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Instrument	Year	Title	Journal	Author	Institution	Keywords	Abstract
BioTester	2009	Fabrication Of Nanofiber Reinforced Protein Structures For Tissue Engineering	Materials Science and Engineering	V. Beachley, X. Wen	Clemson University, Medical University of South Carolina		Extracellular matrices and degradable nanofibers are two very promising materials in the field of tissue engineering; however both of these structures face limitations as tissue engineering scaffolds. Extracellular matrices, such as collagen, gelatin, and laminin, have excellent biocompatibility and allow cell in growth and survival, but structural weakness makes them difficult to handle and greatly limits their uses. Degradable nanofibers support cell attachment and can provide structural support and directional guidance, but individual degradable nanofibers are fragile and have a tendency to form dense fiber bundles which limit cell penetration into the spaces between the nanofibers, especially in the case of aligned nanofibers. To overcome these difficulties, degradable loose nanofibers were embedded in protein matrix in an attempt to fabricate a hybrid scaffold with improved properties, such as improved strength, guidance, spacing among nanofibers, etc. Polycaprolactone (PCL) was used as a model material for degradable nanofibers. Gelatin was employed as a model protein for matrix structure formation. Thin hybrid films (average thickness = 2.78 um) were fabricated by wetting the loose aligned undirectional nanofiber arrays or loose aligned bi-directional nanofiber grids with a gelatin aqueous solution, which also allows for live cell loading into the nanofiber-protein composite if cell are premixed with protein solution or on the surface of the films. Gelatin film alone without nanofiber reinforcement is difficult to handle due to the weakness of the thin membrane. Gelatin films with a fiber density as low as 3% v/v were structurally robust enough for handling, and manipulation into complex shapes. Mechanical testing confirmed that the addition of nanofibers enhanced the strength of gelatin films, in both dry and hydrated state. In vitro testing confirmed that nanofiber reinforced films were biocompatible and provided cells with directional guidance. Results demonstrate the promise of gelatin/PCL nanofiber composites as a tissue scaffolding material. Cell/Tissues: collagen gelatin laminin
BioTester	2009	Strain Uniformity In Biaxial Specimens Is Highly Sensitive To Attachment Details	Journal of Biomechanical Engineering	A. Eilaghi, J. Flanagan, G.W. Brodland, R. Ethier	University of Toronto		Biaxial testing has been used widely to characterize the mechanical properties of soft tissues and other flexible materials, but fundamental issues related to specimen design and attachment have remained. Finite element models and experiments were used to investigate how specimen geometry and attachment details affect uniformity of the strain field inside the attachment points. The computational studies confirm that increasing the number of attachment points increases the size of the area that experiences sensibly uniform strain (defined here as the central sample region where the ratio of principal strains E11/E22<1.10E11/E22<1.10), and that the strains experienced in this region are less than nominal strains based on attachment point movement. Uniformity of the strain field improves substantially when the attachment points span a wide zone along each edge. Subtle irregularities in attachment point positioning can significantly degrade strain field uniformity. In contrast, details of the apron, the region outside of the attachment points, have little effect on the interior strain field. When nonlinear properties consistent with those found in human sclera are used, similar results are found. Experiments were conducted on 6×6 mm6×6 mm talc-sprinkled rubber specimens loaded using wire “rakes.” Points on a grid having 12×12 bays12×12 bays were tracked, and a detailed strain map was constructed. A finite element model based on the actual geometry of an experiment having an off-pattern rake tine gave strain patterns that matched to within 4.4%. Finally, simulations using nonequibiaxial strains indicated that the strain field uniformity was more sensitive to sample attachment details for the nonequibiaxial case as compared to the equibiaxial case. Specimen design and attachment were found to significantly affect the uniformity of the strain field produced in biaxial tests. Practical guidelines were offered for design and mounting of biaxial test specimens. The issues addressed here are particularly relevant as specimens become smaller in size. Cell/Tissue: soft tissue
BioTester	2009	The Influence Of The Tensile Material Properties Of Single Annulus Fibrosus Lamellae And The Interlamellar Matrix Strength On Disc Herniation And Progression	PhD Thesis	D. Gregory	University of Waterloo		Low back pain is highly prevalent in the developed world, with 80% of the population being affected at some point in their lives. Herniation, a common injury to the intervertebral disc, is characterized as the posterior migration of the nucleus pulposus through the layers of the annulus fibrosus. Various risk factors have been associated with the development of disc herniation, but the mechanisms are largely not understood. For example, exposure to vibration has been linked to the occurrence of herniation, yet our understanding of this association is not clear. It is hypothesized that vibration cyclically loads the tissues of the intervertebral disc until failure occurs as a result of fatigue. Tissues at risk of fatigue failure may include the intra-lamellar matrix, the connective tissue found between collagen fibres within a single lamella, and the inter-lamellar matrix, the connective tissue found between adjacent lamellae. In order to determine the mechanistic link between vibration and herniation, a firm understanding of the properties of the intervertebral disc as well as the intra and inter-lamellar matrices are of utmost importance. Further, it is important to determine these properties under physiological loading scenarios. This thesis consists of five studies, which have each provided a unique piece to the intervertebral disc herniation puzzle in order to better understand this mechanistic link. First, it was discovered that annular tissue is subject to significantly higher stresses and is stiffer under biaxial strain as compared to uniaxial strain. Biaxial strain is more representative of the in vivo loading scenario and provides more accurate information regarding scenarios that the annulus can tolerate and those that can result in injury. It was also revealed that, when strained at physiological strain rates (up to 4%/sec), these mechanical properties do not change such that they are independent of iv strain rate. Therefore, when strained at varying rates akin to voluntary movement, the annulus is not subject to higher stresses or altered stiffness. Second, the effect of vibration, an acknowledged risk factor for herniation, was examined on the mechanical properties of the intra and inter-lamellar matrices. It was discovered that vibration altered these matrices such that they were more extensible and strained to greater magnitudes, yet did not reach higher stresses before failing. It was hypothesized that this increased extensibility was due to damage to elastin, as elastin assists in minimizing tissue deformation and helps tissues recover from tensile strain. The final study assessed the effect of exposure to vibration on the development of disc herniation. The initiation of herniation was observed in a significantly greater number of intervertebral discs exposed to vibration as compared to a control condition. Although epidemiological studies had documented a correlation between exposure to vibration and herniation, this was the first study to conclude that exposure to vibration is in fact a mechanical risk factor for the development of herniation and increases the incidence of herniation. Further, based on the findings of the mechanical properties of the intra and inter-lamellar matrices, and in particular the observed 15-20 times greater failure strength of the intra as compared to inter-lamellar matrix, it would appear that the inter-lamellar matrix, and thus delamination, may be the weakest link in the herniation pathway. This thesis has uncovered new information regarding physiological mechanical properties of the annulus. Further, new information regarding the intra and inter-lamellar matrices was obtained, improving our understanding of the healthy disc. Last, by subjecting the disc to a known risk factor for herniation, hypotheses were generated v regarding the initiation and progression of disc herniation, specifically related to the roles of the intra and inter-lamellar matrices. Cell/Tissues: intra-lamellar matrix inter-lamellar matrix annulus fibrosus
BioTester	2010	An Examination Of The Influence Of Strain Rate On Subfailure Mechanical Properties Of The Annulus Fibrosus	Journal of Biomechanical Engineering	D. Gregory, J. Callaghan	University of Waterloo		Disk herniation is often considered a cumulative injury in that repetitive stress on the posterior annulus can result in the nucleus pulposus penetrating the annulus fibrosus and eventually extruding posteriorly. Further, it has been documented that the nucleus pulposus works its way through the annulus through clefts, which form as a result of repetitive tensile strain. The annulus fibrosus is viscoelastic in nature and therefore could express different mechanical responses to applied strain at varying rates. Other viscoelastic tissues, including tendons and ligaments, have shown altered mechanical responses to different rates of applied strain, but the response of the annulus to varying rates of strain is largely unknown. The present study examined the mechanical properties of 20 two-layered samples of porcine annulus fibrosus tissue at three distinct rates of applied 20% biaxial strain (20% strain over 20 s (slow), over 10 s (medium), and over 5 s (fast)); these three rates are considered applicable to nontraumatic loading. No differences in the stiffness or maximum stress in each of the two directions of applied strain were observed between the three strain rates. Specifically, the average (standard deviation) moduli calculated at the fast, medium, and slow rates, respectively, in the axial direction were 7.42 MPa (6.06), 7.77 MPa (6.61), and 7.63 MPa (6.67) and 8.22 MPa (8.4), 8.63 MPa (9.00), and 8.49 MPa (8.69) in the circumferential direction. The maximum stress values reached during the fast, medium, and slow rates, respectively, in the axial direction were 0.40 (0.36) MPa, 0.40 (0.36) MPa, and 0.39 (0.35) MPa and 0.45 (0.47) MPa, 0.44 (0.46) MPa, and 0.43 (0.46) MPa in the circumferential direction. At submaximal strain magnitudes over a range of nontraumatic rates likely to result in clefts in the annulus and potentially leading to disk herniation, any strain rate dependence is not significant. Cell/Tissue: porcine annulus fibrosus
BioTester	2010	Biaxial Mechanical Testing Of Human Sclera	Journal of Biomechanics	A. Eilaghi, J. Flanagan, I. Tertinegg, C. Simmons, W. Brodland, R. Ethier	University of Toronto, Imperial College		The biomechanical environment of the optic nerve head (ONH), of interest in glaucoma, is strongly affected by the biomechanical properties of sclera. However, there is a paucity of information about the variation of scleral mechanical properties within eyes and between individuals. We thus used biaxial testing to measure scleral stiffness in human eyes. Ten eyes from 5 human donors (age 55.4±3.5 years; mean±SD) were obtained within 24 h of death. Square scleral samples (6 mm on a side) were cut from each ocular quadrant 3–9 mm from the ONH centre and were mechanically tested using a biaxial extensional tissue tester (BioTester 5000, CellScale Biomaterials Testing, Waterloo). Stress–strain data in the latitudinal (toward the poles) and longitudinal (circumferential) directions, here referred to as directions 1 and 2, were fit to the four-parameter Fung constitutive equation W=c(eQ−1), where and W, c’s and Eij are the strain energy function, material parameters and Green strains, respectively. Fitted material parameters were compared between samples. The parameter c3 ranged from 10−7 to 10−8, but did not contribute significantly to the accuracy of the fitting and was thus fixed at 10−7. The products cc1 and cc2, measures of stiffness in the 1 and 2 directions, were 2.9±2.0 and 2.8±1.9 MPa, respectively, and were not significantly different (two-sided t-test; p=0.795). The level of anisotropy (ratio of stiffness in orthogonal directions) was 1.065±0.33. No statistically significant correlations between sample thickness and stiffness were found (correlation coefficients=−0.026 and −0.058 in directions 1 and 2, respectively). Human sclera showed heterogeneous, near-isotropic, nonlinear mechanical properties over the scale of our samples. Cell/Tissue: scleral tissue
BioTester	2010	Effects Of Sclera Stiffness On Biomechanics Of The Optic Nerve Head In Glaucoma	Doctoral Theses	A. Eilaghi	University of Toronto		Glaucoma is a common cause of blindness worldwide, yet the etiology of the disease is unclear. A leading hypothesis is that elevated intraocular pressure (IOP) affects the biomechanical environment within the tissues of the optic nerve head (ONH), and that the altered biomechanical environment contributes to optic nerve damage and consequent loss of vision. The biomechanical environment of the ONH is strongly dependent on the biomechanical properties of sclera, particularly scleral stiffness. However there is significant variability in reported stiffness data for human sclera. Therefore, our research goal was to measure the stiffness of human sclera and incorporate this information into finite element models of the human eye to characterize and quantify the biomechanical environment within and around the optic nerve head region at different IOP levels. Human sclera adjacent to the optic nerve head showed highly nonlinear, nearly isotropic and heterogeneous stiffness which was found to be substantially lower than that previously assumed, particularly at lower levels of IOP. The products cc1 and cc2, measures of stiffness in the latitudinal and longitudinal directions from the Fung constitutive model, were 2.9 ± 2.0 MPa and 2.8 ± 1.9 MPa, respectively, and were not significantly different (two-sided t-test; p = 0.795). Scleral stiffness was not statistically different between left and right eyes of an individual (p = 0.952) and amongst the quadrants of an eye (p = 0.412 and p = 0.456 in latitudinal and longitudinal directions, respectively). Three stress-strain relationships consistent with the 5th, 50th and 95th percentiles of the measured scleral stiffness distribution were selected as representatives of compliant, median and stiff scleral properties and were implemented in a generic finite element model of the eye using a hyperelastic five-parameter Mooney-Rivlin material model. Models were solved for IOPs of 15, 25 and 50 mmHg. The magnitudes of strains at the optic nerve head region were substantial at even the lowest applied IOP (15 mmHg) and increased at elevated IOPs (e.g. the third principal strain in the compliant model reached as much as 5.25% in the lamina cribrosa at 15mmHg and 8.84% in the lamina cribrosa at 50 mmHg). Scleras that are “weak”, but still within the physiologic range, are predicted to lead to appreciably increased optic nerve head strains and could represent a risk factor for glaucomatous optic neuropathy. As IOP increased from 15 to 50 mmHg, principal strains in the model with a compliant sclera increased at a lower rate than in the model with a stiff sclera. We quantified the biomechanical environment within and around the optic nerve head region using a range of experimentally measured mechanical properties of sclera and at different IOPs. We showed that IOP-related strains within optic nerve head tissues can reach potentially biologically significant levels (capable of inducing a range of effects in glial cells) even at average levels of IOP and for typical human scleral biomechanical properties. Cell/Tissue: scleral tissue optic nerve head tissues
BioTester	2010	Effects Of Sclera Stiffness Properties On Optic Nerve Head Biomechanics	Annals of Biomedical Engineering	A. Eilaghi, J. Flanagan, C. Simmons, R. Ethier	University of Toronto		The biomechanical environment within the optic nerve head, important in glaucoma, depends strongly on scleral biomechanical properties. Here we use a range of measured nonlinear scleral stress–strain relationships in a finite element (FE) model of the eye to compute the biomechanical environment in the optic nerve head at three levels of intraocular pressure (IOP). Three stress–strain relationships consistent with the 5th, 50th and 95th percentiles of measured human scleral stiffness were selected from a pool of 30 scleral samples taken from 10 eyes and implemented in a generic FE model of the eye using a hyperelastic five-parameter Mooney-Rivlin material model. Computed strains within optic nerve head tissues depended strongly on scleral properties, with most of this difference occurring between the compliant and median scenarios. Also, the magnitudes of strains were found to be substantial even at normal IOP (up to 5.25% in the lamina cribrosa at 15 mmHg), being larger than previously reported values even at normal levels of IOP. We conclude that scleras that are “weak”, but still within the physiologic range, will result in appreciably increased optic nerve head strains and could represent a risk factor for glaucomatous optic neuropathy. Estimations of the deformation at the optic nerve head region, particularly at elevated IOP, should take into account the nonlinear nature of scleral stiffness. Cell/Type: scleral tissue optic nerve head tissues
BioTester	2011	A Comparison Of Uniaxial And Biaxial Mechanical Properties Of The Annulus Fibrosus: A Porcine Model	Journal of Biomechanical Engineering	Diane Gregory, J. Callaghan	University of Waterloo		The annulus fibrosus of the intervertebral disk experiences multidirectional tension in vivo, yet the majority of mechanical property testing has been uniaxial. Therefore, our understanding of how this complex multilayered tissue responds to loading may be deficient. This study aimed to determine the mechanical properties of porcine annular samples under uniaxial and biaxial tensile loading. Two-layer annulus samples were isolated from porcine disks from four locations: anterior superficial, anterior deep, posterior superficial, and posterior deep. These tissues were then subjected to three deformation conditions each to a maximal stretch ratio of 1.23: uniaxial, constrained uniaxial, and biaxial. Uniaxial deformation was applied in the circumferential direction, while biaxial deformation was applied simultaneously in the circumferential and compressive directions. Constrained uniaxial consisted of a stretch ratio of 1.23 in the circumferential direction while holding the tissue stationary in the axial direction. The maximal stress and stress-stretch ratio (S-S) moduli determined from the biaxial tests were significantly higher than those observed during both the uniaxial tests (maximal stress, 97.1% higher during biaxial; p=0.002p=0.002; S-S moduli, 117.9% higher during biaxial; p=0.0004p=0.0004) and the constrained uniaxial tests (maximal stress, 46.8% higher during biaxial; S-S moduli, 82.9% higher during biaxial). These findings suggest that the annulus is subjected to higher stresses in vivo when under multidirectional tension. Cell/Tissue: annulus fibrosus
BioTester	2011	Does Vibration Influence The Initiation Of Intervertebral Disc Herniation	The Spine Journal	Diane Gregory, J. Callaghan	University of Gothenburg		Study Design. Pain behavior assessment in rats following disc puncture (DP) and simultaneous tumor necrosis factor (TNF) inhibition. Objective. To assess if treatment with TNF inhibition could reduce the pain behavior changes induced by DP in the rat. Summary of Background Data. Anular tears with leakage of nucleus pulposus have been suggested to be one possible cause of low back pain (LBP). In an experimental model, it was recently shown that DP might induce specific pain behavior changes. The aim of the present study was to a study if inhibition of TNF might reduce such pain behavior changes. Methods. Sixty rats underwent facetectomy and puncture of the fourth lumbar disc. The rats were simultaneously treated with doxycycline locally at 0.3 and 3.0 mg/kg and systemically at 3.0 mg/kg, or infliximab locally at 0.5 and 5.0 mg/kg, and systemically at 5.0 mg/kg, (n ∇ 10 for each subseries). The rats were videotaped at 1, 3, 7, 14, and 21 days after surgery. The videos were analyzed regarding presence of wet-dog shakes (WDS). Data from a previous study with sham surgery and DP without treatment were included for comparison. Results. All groups treated with doxycycline resulted in a statistically significant reduction of WDS compared to the group without treatment (DP). In infliximab treated animals, WDS decreased with statistically significance compared to the nontreated DP group at all analyzed days except for the group with high dose local treatment where a statistically significant reduction was obtained only at days 14 and 21. Conclusion. The present study showed that TNF inhibition induced a marked reduction of wet dog shakes. It is not fully understood if wet-dog shakes may relate to LBP, but in view of recent clinical findings one may consider clinical studies of TNF inhibition for the treatment of LBP. Cell/Tissue: lumbar tissue annular tissue nucleus pulposus
BioTester	2011	Novel Lap Test Determines The Mechanics Of Delamination Between Annular Lamellae Of The Intervertebral Disc	Journal of Biomechanics	D. Gregory, J. Veldhuis, C. Horst, W. Brodland, J. Callaghan	University of Waterloo		Delamination between lamellae of the annulus fibrosus is a crucial stage of intervertebral disc herniation, and to better understand the mechanics of the delamination process, a novel lap test was devised. Specimens consisting of two adjacent, naturally bonded lamellae were obtained from the cervical region of frozen porcine spines. They were cut into specimens nominally 3.5 mm wide by 7 mm long and tabs of the deep and superficial layers were removed from opposite ends of the specimens so that a 4.5–5.0 mm long intact interface remained between the lamellae. Specimens were mounted in a BioTester tensile instrument using BioRake attachments having 5 sharpened points side-by-side, and they were strained at 2%/s. Force–time curves were obtained and, using tracking software, a detailed map was made of the time course of the displacements within the specimens. Extensibility of the lamellae themselves was found to substantially complicate interpretation of the data. The experiments, together with mathematical analyses and finite element models, show that much of the shear load is transferred between lamellae at the ends of the bonded region, a finding of clinical importance. The inter-lamellae bond was found to have a peak strength of 0.30±0.05 N/mm of specimen width (not to be confused with lap length), and the remarkable ability to carry substantial load even when lamellae had displaced up to 10 mm relative to each other. Cell/Tissue: cervical lamellae annulus fibrosus
BioTester	2011	Surfactant Assisted Incorporation Of Single-Walled Carbon Nanotubes Into A Chitosan-Polyvinylpyrrolidone Polymer	Journal of Nanoengineering and Nanomanufacturing	T. Davis, J. Zhang, J.E. Herrera	Western University		A novel method for the incorporation of highly dispersed single-walled carbon nanotubes (SWNTs) into a chitosan matrix is reported. This was achieved through the use of a precursor SWNT dispersion in a biocompatible surfactant solution containing cetyl trimethylammonium bromide (CTAB) and polyvinylpyrrolidone (PVP). Once a desirable degree of dispersion was attained this precursor SWNT dispersion was mixed with a chitosan solution followed by chemical crosslinking of the chitosan. Vis/NIR and Raman spectra of several sections of the resulting solid nanocomposite material suggested uniform dispersion of SWNTs throughout the solid matrix. The mechanical properties of the nanocomposite were tested and compared to a material obtained using a similar protocol without the use of SWNTs. The results indicate that the presence of a relatively low amount (0.32 wt%) of SWNTs is enough to significantly increase the stiffness and elastic modulus of uncrosslinked chitosan by over 20 times. Cell/Tissue: chitosan - polyvinylpyrrolidone polymer matrix single-walled carbon nanotubes
BioTester	2012	A Murine Experimental Model For The Mechanical Behavior Of Viable Right-Ventricular Myocardium	Journal of Physiology	D. Valdez-Jasso, M.A. Simon, H.C. Champion, M.S. Sacks	University of Pittsburgh		Although right-ventricular function is an important determinant of cardio-pulmonary performance in health and disease, right ventricular myocardium mechanical behaviour has received relatively little attention. We present a novel experimental method for quantifying the mechanical behaviour of transmurally intact, viable right-ventricular myocardium. Seven murine right ventricular free wall (RVFW) specimens were isolated and biaxial mechanical behaviour measured, along with quantification of the local transmural myofibre and collagen fibre architecture. We developed a complementary strain energy function based method to capture the average biomechanical response. Overall, murine RVFW revealed distinct mechanical anisotropy. The preferential alignment of the myofibres and collagen fibres to the apex-to-outflow-tract direction was consistent with this also being the mechanically stiffer axis. We also observed that the myofibre and collagen fibre orientations were remarkably uniform throughout the entire RVFW thickness. Thus, our findings indicate a close correspondence between the tissue microstructure and biomechanical behaviour of the RVFW myocardium, and are a first step towards elucidating the structure–function of non-contracted murine RVFW myocardium in health and disease. Cell/Tissue: right ventricular myocardium
BioTester	2012	An Examination Of The Mechanical Properties Of The Annulus Fibrosus: The Effect Of Vibration On The Intra-Lamellar Matrix Strength	Medical Engineering & Physics	Diane Gregory, J. Callaghan	University of Waterloo		Vibration has been associated with low back pain and disc herniation; however the mechanism responsible for this association is unclear. It is theorized that herniation propagates through annular layers via clefts that form in the extracellular matrix between collagen fibres (intra-lamellar matrix) within each lamella. The effect of cyclic compressive loading at 5 Hz, applied to porcine functional spine units, on the mechanical properties of excised single annular lamellae was examined. These lamellae were tested under tension applied perpendicular to the collagen fibre orientation, effectively isolating the intra-lamellar matrix. Vibration affected the deformation magnitude at the end of the toe region of the stress–stretch ratio curve; specifically vibrated tissues showed larger toe regions (stretch ratio of 1.50 as compared to 1.31 observed in the control tissues, p = 0.027). It is hypothesized that this result may be due to damaged elastin—a protein responsible for minimizing deformation and assisting with returning tissues to normal length following tension. Cell/Tissue: lamella
BioTester	2012	Application Of Simple Biomechanical And Biochemical Tests To Heart Valve Leaflets: Implications For Heart Valve Characterization And Tissue Engineering	Journal of Engineering in Medicine	H-Y. Huang, B.N. Balhouse, S. Huang	North Carolina State University		A simple biomechanical test with real-time displacement and strain mapping is reported, which provides displacement vectors and principal strain directions during the mechanical characterization of heart valve tissues. The maps reported in the current study allow us to quickly identify the approximate strain imposed on a location in the samples. The biomechanical results show that the aortic valves exhibit stronger anisotropic mechanical behavior than that of the pulmonary valves before 18% strain equibiaxial stretching. In contrast, the pulmonary valves exhibit stronger anisotropic mechanical behavior than aortic valves beyond 28% strain equibiaxial stretching. Simple biochemical tests are also conducted. Collagens are extracted at different time points (24, 48, 72, and 120 h) at different locations in the samples. The results show that extraction time plays an important role in determining collagen concentration, in which a minimum of 72 h of extraction is required to obtain saturated collagen concentration. This work provides an easy approach for quantifying biomechanical and biochemical properties of semilunar heart valve tissues, and potentially facilitates the development of tissue engineered heart valves. Cell/Tissue: cardiovascular tissue collagen
BioTester	2012	Biomechanical Properties Of The Transverse Carpal Ligament Under Biaxial Strain	Journal of Orthopaedic Research	Michael Holmes, S. Howarth, J. Callaghan, P. Keir	McMaster University		The transverse carpal ligament (TCL) influences carpal stability and carpal tunnel mechanics, yet little is known about its mechanical properties. We investigated the tissue properties of TCLs extracted from eight cadaver arms and divided into six tissue samples from the distal radial, distal middle, distal ulnar, proximal radial, proximal middle, and proximal ulnar regions. The 5% and 15% strains were applied biaxially to each sample at rates of 0.1, 0.25, 0.5, and 1%/s. Ligament thickness ranged from 1.22 to 2.90 mm. Samples from the middle of the TCL were thicker proximally than distally (p < 0.013). Tissue location significantly affected elastic modulus (p < 0.001). Modulus was greatest in the proximal radial samples (mean 2.8 MPa), which were 64% and 44% greater than the distal radial and proximal ulnar samples, respectively. Samples from the middle had a modulus that was 20–39% greater in the proximal versus more distal samples. The TCL exhibited different properties within different locations and in particular greater moduli were found near the carpal bone attachments. These properties contribute to the understanding of carpal tunnel mechanics that is critical to understanding disorders of the wrist. Cell/Tissue: transverse carpal ligament
BioTester	2012	Multi-Scale Mechanical Characterization Of Scaffolds For Heart Valve Tissue Engineering	Journal of Biomechanics	G. Argento, M. Simonet, C.W.J. Oomens, F.P.T. Baaijens	Eindhoven University of Technology		Electrospinning is a promising technology to produce scaffolds for cardiovascular tissue engineering. Each electrospun scaffold is characterized by a complex micro-scale structure that is responsible for its macroscopic mechanical behavior. In this study, we focus on the development and the validation of a computational micro-scale model that takes into account the structural features of the electrospun material, and is suitable for studying the multi-scale scaffold mechanics. We show that the computational tool developed is able to describe and predict the mechanical behavior of electrospun scaffolds characterized by different microstructures. Moreover, we explore the global mechanical properties of valve-shaped scaffolds with different microstructural features, and compare the deformation of these scaffolds when submitted to diastolic pressures with a tissue engineered and a native valve. It is shown that a pronounced degree of anisotropy is necessary to reproduce the deformation patterns observed in the native heart valve. Cell/Tissue: cardivascular tissue
BioTester	2013	A Nanocomposite Contact Lens For The Delivery Of Hydrophilic Protein Drugs	Journal of Materials Chemistry B	J. Zhang, R. Bi, W. Hodge, P. Yin, W.H. Tse	University of Western Ontario		To improve the efficiency of topical ocular drug administration, we developed a nanocomposite contact lens to deliver hydrophilic protein drugs over a prolonged period of time. Here, an in situ route was used to encapsulate the hydrophilic protein drug bovine serum albumin (BSA) within gelatin nanoparticles (NPs), 180 ± 20 nm in diameter, which were then grafted onto the lens material, a copolymer of 2-hydroxyethyl methacrylate and 2-aminoethyl methacrylate p(HEMA-co-AEMA), through photopolymerization. The thickness of the nanocomposite lens was controlled at 150 μm. The release kinetics of BSA from plain p(HEMA-co-AEMA), gelatin NPs, and gelatin NP-grafted p(HEMA-co-AEMA) in phosphate buffer saline (PBS) at pH = 7.4 were studied. The release profile of BSA encapsulated within gelatin NPs could be monitored for 7 days, three times longer than that of BSA soaked in p(HEMA-co-AEMA). Our findings indicate that use of the nanocomposite contact lens, i.e. BSA-loaded gelatin NPs incorporated into p(HEMA-co-AMEA), can prolong the release profile of BSA to 12 days. The swelling behavior and interior strain of p(HEMA-co-AEMA) with and without grafted NPs (1000 : 1 w/w) were further investigated. The nanocomposite lens shows higher swelling behavior than the plain p(HEMA-co-AEMA) lens does. The addition of gelatin NPs to hydrogels leads to a relatively uniform interior strain with lower stiffness. Thus, the prolonged release might be due to a combination of effects. Internal diffusion of the nanocomposite lens materials may significantly contribute to the prolonged release of protein drugs. Furthermore, the nanocomposite lens materials had no cytotoxicity. This new biocompatible nanocomposite might be further developed as an alternative tool for continuous topical ocular drug delivery over a prolonged period of time. Cell/Tissue: nanocomposite ocular lens
BioTester	2013	Comparison Of Methods Used To Measure The Thickness Of Soft Tissues And Their Influence On The Evaluation Of Tensile Stress	Journal of Biomechanics	S.A. O'Leary, B.J. Doyle, T.M. McGloughlin	University of Limerick		Measuring the physical dimensions of soft tissue is difficult due to its deformable nature. Such measurements are used to evaluate the tissue's mechanical properties. Imprecise measurements of the tissue's thickness can alter the assessment of tensile stress which may have significant clinical relevance when used as a diagnostic tool. The performance of routinely used measurement methods including a (i) vernier calipers, (ii) micrometer, (iii) thickness gauge, (iv) glass slide technique coupled with (i) and (ii) and a (v) laser displacement sensor were assessed by comparing them to a photogrammetric technique which was considered to be the measurement standard. All measurements were performed on two tissue types: porcine aorta and human intraluminal thrombus from an abdominal aortic aneurysm (AAA) and results were compared against predetermined criteria whose limits represented a 10% change in experimentally derived tensile stress. The inter-rater and retest reliability of the vernier calipers, micrometer and thickness gauge were also investigated. The thickness gauge was shown to be the most reliable and could accurately measure the thickness of aortic tissue. The conditions of the criteria were not met by any instrument used to measure the thickness of the AAA intraluminal thrombus, however, the micrometer, which proved highly reliable, was considered the most suitable (effects on tensile stress: +14.7%). For both tissues the glass slide and laser techniques significantly over estimated the thickness measurement altering the tensile stress by up to −29.6%. This study highlights the effects of inaccurate measurements on the assessment of tensile stress and recommends a thickness gauge be used to measure structured tissue (aorta) and a micrometer for unstructured tissue (AAA intraluminal thrombus). Cell/Type: cardiavascular tissue, porcine aorta intraluminal thrombus
BioTester	2013	Corneal Resistance To Shear Force After Uva-Riboflavin Cross-Linking	Investigative ophthalmology & Visual Science Journal	A.P. Sondergaard, A. Ivarsen, J. Hjortdal	Aarhus University Hospital		Purpose.: We evaluated whether UVA-riboflavin collagen cross-linking (CXL) increases transverse stromal shear moduli ex vivo, whether the shear moduli are greater in the anterior compared to the posterior stroma, and whether the shear moduli are affected by CXL. Methods.: The resistance to unidirectional transverse shear of human (n = 18) and porcine (n = 42) corneas was measured in a custom engineered biaxial biomechanical setup at different hydrations. The corneas were separated into untreated, riboflavin solution–treated, and CXL–treated groups. The depth dependency of shear moduli within groups was assessed in femtosecond laser cut sheets. Dry weights were obtained for solids correction. Results.: In porcine full-thickness buttons and 300 μm anterior sheets, a significantly increased unidirectional transverse shear modulus was detected in riboflavin-treated and CXL-treated groups compared to the respective untreated groups. There was no significant difference in shear modulus between riboflavin- and CXL-treated groups. In all groups, the shear moduli were greater in the anterior sheets compared to posterior sheets. Similar results were detected in human corneas. Conclusions.: A method for unidirectional transverse shear resistance measurement was developed. The shear moduli were greater in the anterior compared to the posterior sheets. Increase in shear moduli was observed in the riboflavin and CXL groups compared to the untreated group, indicating that the immediate effects of the riboflavin or CXL treatment may be due partly to ground substance/dextran-5-phosphate interaction. Cell/Type: corneal scleral
BioTester	2013	In Vitro Fabrication Of Autologous Living Tissue-Engineered Vascular Grafts Based On Prenatally Harvested Ovine Amniotic Fluid-Derived Stem Cells	Journal of Tissue Engineering and Regenerative Medicine	B. Weber, D. Kehl, U. Bleul, S. Sammut, L. Frese, A. Ksiazek, L Behr, J. Achermann, G. Stranzinger, J. Robert, B. Sanders, M. Sidler, C.E. Brokopp, S.T. Proulx, T. Frauenfelder, R. Schoenauer, M.Y.Emmert, V. Falk, S.P.Herstrup	University of Zurich		Amniotic fluid cells (AFCs) have been proposed as a valuable source for tissue engineering and regenerative medicine. However, before clinical implementation, rigorous evaluation of this cell source in clinically relevant animal models accepted by regulatory authorities is indispensable. Today, the ovine model represents one of the most accepted preclinical animal models, in particular for cardiovascular applications. Here, we investigate the isolation and use of autologous ovine AFCs as cell source for cardiovascular tissue engineering applications. Fetal fluids were aspirated in vivo from pregnant ewes (n = 9) and from explanted uteri post mortem at different gestational ages (n = 91). Amniotic non-allantoic fluid nature was evaluated biochemically and in vivo samples were compared with post mortem reference samples. Isolated cells revealed an immunohistochemical phenotype similar to ovine bone marrow-derived mesenchymal stem cells (MSCs) and showed expression of stem cell factors described for embryonic stem cells, such as NANOG and STAT-3. Isolated ovine amniotic fluid-derived MSCs were screened for numeric chromosomal aberrations and successfully differentiated into several mesodermal phenotypes. Myofibroblastic ovine AFC lineages were then successfully used for the in vitro fabrication of small- and large-diameter tissue-engineered vascular grafts (n = 10) and cardiovascular patches (n = 34), laying the foundation for the use of this relevant pre-clinical in vivo assessment model for future amniotic fluid cell-based therapeutic applications. Cell/Tissue: cardiovascular tissue amniotic fluid cells
BioTester	2013	Mechanical Analysis Of Ovine And Pediatric Pulmonary Artery For Heart Valve Stent Design	Journal of Biomechanics	M.S. Cabrera, C.W.J. Oomens, C.V.C. Bouten, A.J.J.C. Bogers, S.P. Hoerstrup, F.P.T. Baaijens	Eindhoven University of Technology		Transcatheter heart valve replacement is an attractive and promising technique for congenital as well as acquired heart valve disease. In this procedure, the replacement valve is mounted in a stent that is expanded at the aimed valve position and fixated by clamping. However, for this technique to be appropriate for pediatric patients, the material properties of the host tissue need to be determined to design stents that can be optimized for this particular application. In this study we performed equibiaxial tensile tests on four adult ovine pulmonary artery walls and compared the outcomes with one pediatric pulmonary artery. Results show that the pediatric pulmonary artery was significantly thinner (1.06±0.36 mm (mean±SD)) than ovine tissue (2.85±0.40 mm), considerably stiffer for strain values that exceed the physiological conditions (beyond 50% strain in the circumferential and 60% in the longitudinal direction), more anisotropic (with a significant difference in stiffness between the longitudinal and circumferential directions beyond 60% strain) and presented stronger non-linear stress–strain behavior at equivalent strains (beyond 26% strain) compared to ovine tissue. These discrepancies suggest that stents validated and optimized using the ovine pre-clinical model might not perform satisfactorily in pediatric patients. The material parameters derived from this study may be used to develop stent designs for both applications using computational models. Cell/Tissue: pulmonary cardiovascular tissue
BioTester	2013	Mechanics Of The Pulmonary Valve In The Aortic Position	Journal of the Mechanical Behavior of Biomedical Materials	A.L.F. Soares, D. van Geeman, A.J. van den Bogaerdt, C.W.J. Oomens, C.V.C. Bouten, F.P.T. Baaijens	Eindhoven University of Technology		Mathematical models can provide valuable information to assess and evaluate the mechanical behavior and remodeling of native tissue. A relevant example when studying collagen remodeling is the Ross procedure because it involves placing the pulmonary autograft in the more demanding aortic valve mechanical environment. The objective of this study was therefore to assess and evaluate the mechanical differences between the aortic valve and pulmonary valve and the remodeling that may occur in the pulmonary valve when placed in the aortic position. The results from biaxial tensile tests of pairs of human aortic and pulmonary valves were compared and used to determine the parameters of a structurally based constitutive model. Finite element analyzes were then performed to simulate the mechanical response of both valves to the aortic diastolic load. Additionally, remodeling laws were applied to assess the remodeling of the pulmonary valve leaflet to the new environment. The pulmonary valve showed to be more extensible and less anisotropic than the aortic valve. When exposed to aortic pressure, the pulmonary leaflet appeared to remodel by increasing its thickness and reorganizing its collagen fibers, rotating them toward the circumferential direction. Cell/Tissue: cardivascular tissue, human aorta and pulmonary valves
BioTester	2013	Reduction Of Stromal Swelling Pressure After Uva-Riboflavin Cross-Linking	Investigative ophthalmology & Visual Science	A.P. Sondergaard, A. Ivarsen, J. Hjortdal	Aarhus University Hospital		Purpose.: We evaluated whether UVA-riboflavin collagen cross-linking (CXL) reduces stromal swelling pressure (SP) in porcine and human corneas ex vivo. Methods.: Porcine corneas (n = 35) were divided into five groups: Full-thickness buttons, riboflavin-treated buttons, CXL-treated buttons, riboflavin-treated anterior and posterior lamellae, and CXL-treated anterior and posterior lamellae. Riboflavin- or CXL-treated human corneas (n = 6) were cut into anterior and posterior lamellae. The force exerted by the specimens during swelling in saline was recorded and SP was calculated. Dry weights were obtained for solids correction. Results.: In full-thickness porcine buttons, a significantly reduced SP was observed after CXL (40.07 ± 3.86 mm Hg) compared to riboflavin treatment (68.13 ± 11.39 mm Hg, P = 0.02) at 5% compression. Also, a trend toward reduced SP in the CXL-treated anterior human lamellae (9.9 mm Hg) was found compared to the riboflavin group (25 mm Hg) at 5% compression. In the anterior porcine lamellae a significant SP reduction was observed in the CXL group versus the riboflavin group (P < 0.001, two-way ANOVA). Likewise, in the posterior porcine lamellae, a significant SP reduction was observed in the CXL group versus riboflavin alone (P < 0.05, two-way ANOVA). Posterior human lamellae did not differ in SP when comparing CXL and riboflavin groups. Conclusions.: Our study demonstrated a significant reduction in anterior stromal SP in porcine and human corneas after CXL. This finding suggested that CXL may reduce corneal SP in vivo, and thereby reduce edema and improve vision. Thus, in the clinical setting, patients suffering from corneal edema may benefit from CXL treatment. Cell/Tissue: corneal
BioTester	2014	Cardiac Function Of The Naked Mole-Rat: Echophysiological Responses To Working Underground	American Journal of Physiology: Heart and Circulatory Physiology	K.M. Grimes, A. Voorhees, Y.A. Chiao, H-C. Han, M.L. Lindsey, R. Buffenstein	University of Texas		The naked mole-rat (NMR) is a strictly subterranean rodent with a low resting metabolic rate. Nevertheless, it can greatly increase its metabolic activity to meet the high energetic demands associated with digging through compacted soils in its xeric natural habitat where food is patchily distributed. We hypothesized that the NMR heart would naturally have low basal function and exhibit a large cardiac reserve, thereby mirroring the species' low basal metabolism and large metabolic scope. Echocardiography showed that young (2–4 yr old) healthy NMRs have low fractional shortening (28 ± 2%), ejection fraction (43 ± 2%), and cardiac output (6.5 ± 0.4 ml/min), indicating low basal cardiac function. Histology revealed large NMR cardiomyocyte cross-sectional area (216 ± 10 μm2) and cardiac collagen deposition of 2.2 ± 0.4%. Neither of these histomorphometric traits was considered pathological, since biaxial tensile testing showed no increase in passive ventricular stiffness. NMR cardiomyocyte fibers showed a low degree of rotation, contributing to the observed low NMR cardiac contractility. Interestingly, when the exercise mimetic dobutamine (3 μg/g ip) was administered, NMRs showed pronounced increases in fractional shortening, ejection fraction, cardiac output, and stroke volume, indicating an increased cardiac reserve. The relatively low basal cardiac function and enhanced cardiac reserve of NMRs are likely to be ecophysiological adaptations to life in an energetically taxing environment. Cell/Tissue: cardiac tissue, cardiovascular tissue myocardium cardiomyocytes
BioTester	2014	Directional Biomechanical Properties Of Porcine Skin Tissue	Journal of Mechanics in Medicine and Biology	H-Y. Huang, S. Huang, C.P. Frazier, P.M. Prim, O. Harrysson	North Carolina State University		Skin is a multilayered composite material and composed principally of the proteins collagen, elastic fibers and fibroblasts. The direction-dependent material properties of skin tissue is important for physiological functions like skin expansion. The current study has developed methods to characterize the directional biomechanical properties of porcine skin tissues as studies have shown that pigs represent a useful animal model due to similarities between porcine and human skin. It is observed that skin tissue has a nonlinear anisotropy biomechanical behavior, where the parameters of material modulus is 378 ± 160 kPa in the preferred-fiber direction and 65.96 ± 40.49 kPa in the cross-fiber direction when stretching above 30% strain equibiaxially. The result from the study provides methods of characterizing biaxial mechanical properties of skin tissue, as the collagen fiber direction appears to be one of the primary determinants of tissue anisotropy. Cell/Tissue: porcine and human skin tissue, epithelial tissue
BioTester	2014	Modeling The Impact Of Scaffold Architecture And Mechanical Loading On Collagen Turnover In Engineered Cardiovascular Tissues	Biomechanics and Modeling in MechanoBiology	G. Argento, N. de Jonge, S.H.M. Sontjens, C.W.J. Oomens, C.V.C. Bouten, F.P.T. Baaijens	Eindhoven University of Technology		The anisotropic collagen architecture of an engineered cardiovascular tissue has a major impact on its in vivo mechanical performance. This evolving collagen architecture is determined by initial scaffold microstructure and mechanical loading. Here, we developed and validated a theoretical and computational microscale model to quantitatively understand the interplay between scaffold architecture and mechanical loading on collagen synthesis and degradation. Using input from experimental studies, we hypothesize that both the microstructure of the scaffold and the loading conditions influence collagen turnover. The evaluation of the mechanical and topological properties of in vitro engineered constructs reveals that the formation of extracellular matrix layers on top of the scaffold surface influences the mechanical anisotropy on the construct. Results show that the microscale model can successfully capture the collagen arrangement between the fibers of an electrospun scaffold under static and cyclic loading conditions. Contact guidance by the scaffold, and not applied load, dominates the collagen architecture. Therefore, when the collagen grows inside the pores of the scaffold, pronounced scaffold anisotropy guarantees the development of a construct that mimics the mechanical anisotropy of the native cardiovascular tissue. Cell/Tissue: cardiovascuclar tissue
BioTester	2014	Should A Native Depth-Dependent Distribution Of Human Meniscus Constitutive Components Be Considered In Fea-Models Of The Knee Joint?	Journal of the Mechanical Behavior of Biomedical Materials	J.M. Parraga Quiroga, P. Emmans, W. Wilson, K. Ito, C.C. van Donkelaar	Eindhoven University of Technology		The depth-dependent matrix composition of articular cartilage is important for its mechanical behavior. It is unknown whether the depth-dependent matrix composition of a meniscus is similarly important for its load-bearing function. The present objective was to determine whether it is necessary to account for the native distribution of matrix components in the cross-sectional plane of the meniscus, when studying its mechanical behavior in numerical models. To address this objective, measured depth-dependent distribution of matrix contents in the human meniscus, and fitted visco-elastic mechanical properties of the collagen were used as input in FEA simulations of a knee joint. The importance of including the depth-dependent matrix component constitution in the meniscus was determined by comparing simulations with an axisymmetric representation of the knee joint, which incorporated either the depth-dependent matrix composition or homogenized matrix. Depth-dependent differences in water, collagen and proteoglycan contents were observed, but these were not significantly different. The anterior region, with significantly higher collagen content, was statistically stiffer than the posterior region. However, depth wise, stiffness did not correlate to the constitution of the tissue. GAG content was significantly higher in the posterior than in the anterior region. Visco-elastic properties of meniscus collagen were fitted against tensile test data. Simulations show that the distribution of stresses and strains in the cartilage is slightly low when the meniscus contains a depth-dependent constitution, but this difference is only modest. Therefore, this study suggests that knee joint mechanics is rather insensitive to the distribution of constitutive components in the cross section of the meniscus, and that the depth-dependent matrix distribution of the meniscus is not essential to be included in axisymmetric computational models of the knee joint. Cell/Tissue: meniscus articular cartilage
BioTester	2014	Strain-Dependent Modulation Of Macrophage Polarization Within Scaffolds	Biomaterials	V. Ballotta, A. Dressen-Mol, C.V.C. Bouten, F.P.T. Baaijens	Eindhoven University of Technology		Implanted synthetic substrates for the regeneration of cardiovascular tissues are exposed to mechanical forces that induce local deformation. Circulating inflammatory cells, actively participating in the healing process, will be subjected to strain once recruited. We investigated the effect of deformation on human peripheral blood mononuclear cells (hPBMCs) adherent onto a scaffold, with respect to macrophage polarization towards an inflammatory (M1) and reparative (M2) phenotype and to early tissue formation. HPBMCs were seeded onto poly-ε-caprolactone bisurea strips and subjected to 0%, 7% and 12% cyclic strain for up to one week. After 1 day, cells subjected to 7% deformation showed upregulated expression of pro and anti-inflammatory chemokines, such as MCP-1 and IL10. Immunostaining revealed presence of inflammatory macrophages in all groups, while immunoregulatory macrophages were detected mainly in the 0 and 7% groups and increased significantly over time. Biochemical assays indicated deposition of sulphated glycosaminoglycans and collagen after 7 days in both strained and unstrained samples. These results suggest that 7% cyclic strain applied to hPBMCs adherent on a scaffold modulates their polarization towards reparative macrophages and allows for early synthesis of extracellular matrix components, required to promote further cell adhesion and proliferation and to bind immunoregulatory cytokines. Cell/Tissue: cardiovascular tissue, arterial tissue human peripheral blood mononuclear cells macrophages
BioTester	2014	Structural And Mechanical Adaptations Of Right Ventrical Free Wall Myocardium To Pressure Overload	Annals of Biomedical Engineering	M.R. Hill, M.A. Simon, D. Valdez-Jasso, H.C. Champion, M.S. Sacks	University of Texas		Right ventricular (RV) failure in response to pulmonary hypertension (PH) is a severe disease that remains poorly understood. PH-induced pressure overload leads to changes in the RV free wall (RVFW) that eventually results in RV failure. While the development of computational models can benefit our understanding of the onset and progression of PH-induced pressure overload, detailed knowledge of the underlying structural and biomechanical events remains limited. The goal of the present study was to elucidate the structural and biomechanical adaptations of RV myocardium subjected to sustained pressure overload in a rat model. Hemodynamically confirmed severe chronic RV pressure overload was induced in Sprague–Dawley rats via pulmonary artery banding. Extensive tissue-level biaxial mechanical and histomorphological analyses were conducted to assess the remodeling response in the RV free wall. Simultaneous myofiber hypertrophy and longitudinal re-orientation of myo- and collagen fibers were observed, with both fiber types becoming more highly aligned. Transmural myo- and collagen fiber orientations were co-aligned in both the normal and diseased state. The overall tissue stiffness increased, with larger increases in longitudinal vs. circumferential stiffness. The latter was attributed to longitudinal fiber re-orientation, which increased the degree of anisotropy. Increased mechanical coupling between the two axes was attributed to the increased fiber alignment. Interestingly, estimated myofiber stiffness increased while the collagen fiber stiffness remained unchanged. The increased myofiber stiffness was consistent with clinical results showing titin-associated increased sarcomeric stiffening observed in PH patients. These results further our understanding of the underlying adaptive and maladaptive remodeling mechanisms and may lead to improved techniques for prognosis, diagnosis, and treatment for PH. Cell/Tissue: cardiovascular tissue, myocardium, cardiac tissue
BioTester	2014	The Biaxial Biomechanical Behavior Of Adominal Aortic Aneurysm Tissue	Annals of Biomedical Engineering	S.A. O'Leary, D.A. Healey, E.G. Kavanagh, M.T. Walsh, T.M. McGloughling, B.J. Doyle	University of Western Australia		Rupture of the abdominal aortic aneurysm (AAA) occurs when the local wall stress exceeds the local wall strength. Knowledge of AAA wall mechanics plays a fundamental role in the development and advancement of AAA rupture risk assessment tools. Therefore, the aim of this study is to evaluate the biaxial mechanical properties of AAA tissue. Multiple biaxial test protocols were performed on AAA samples harvested from 28 patients undergoing open surgical repair. Both the Tangential Modulus (TM) and stretch ratio (λ) were recorded and compared in both the circumferential (ϴ) and longitudinal (L) directions at physiologically relevant stress levels, the influence of patient specific factors such as sex, age AAA diameter and status were examined. The biomechanical response was also fit to a hyperplastic material model. The AAA tissue was found to be anisotropic with a greater tendency to stiffen in the circumferential direction compared to the longitudinal direction. An anisotropic hyperelastic constitutive model represented the data well and the properties were not influenced by the investigated patient specific factors however, a future study utilizing a larger cohort of patients is warranted to confirm these findings. This work provides further insights on the biomechanical behavior of AAA and may be useful in the development of more reliable rupture risk assessment tools. Cell/Tissue: cardiovascular tissue
BioTester	2014	The Biaxial Mechanical Behaviour Of Abdominal Aortic Aneurysm Intraluminal Thrombus: Classification Of Morphology And The Determination Of Layer And Region Specific Properties	Journal of Biomechanics	S.A. O'Leary, E.G. Kavanagh, P.A. Grace, T.M. McGloughlin, B.J. Doyle	University of Western Australia		Intraluminal thrombus (ILT) is present in 75% of clinically-relevant abdominal aortic aneurysms (AAAs) yet, despite much research effort, its role in AAA biomechanics remains unclear. The aim of this work is to further evaluate the biomechanics of ILT and determine if different ILT morphologies have varying mechanical properties. Biaxial mechanical tests were performed on ILT samples harvested from 19 patients undergoing open surgical repair. ILT were separated into luminal, medial and medial/abluminal layers. A total of 356 tests were performed and the Cauchy stress (σ) and tangential modulus (TM) at a stretch ratio (λ) of 1.14 were recorded for each test in both the circumferential (θ) and longitudinal (L) directions. Our data revealed three distinct types of ILT morphologies, each with a unique set of mechanical properties. All ILT layers were found to be isotropic and inhomogeneous. Type 1 (n=10) was a multi-layered ILT (thick medial/abluminal layer) whose strength and stiffness decreased gradually from the luminal to the medial/abluminal layer. Type 2 (n=6) was a multi-layered ILT (thin/highly degraded medial/abluminal layer) whose strength and stiffness decreased abruptly between the luminal and medial/abluminal layer and Type 3 (n=3) is a single layered ILT with a lower strength and stiffness than Types 1 and 2. In a sub-study, we found the luminal layer to be stronger and stiffer in the posterior than the anterior region. This work provides further insights to the biomechanical behaviour of ILT and the use of our ILT classification may be useful in future studies. Cell/Tissue: luminal intraluminal thrombus, cardiovascular tissue medial intraluminal thrombus abluminal intraluminal thrombus
BioTester	2014	The Impact Of Long Term Freezing On The Mechanical Properties Of Porcine Aortic Tissue	Journal of the Mechanical Behavior of Biomedical Materials	S. O'Leary, B. Doyle, T. McGloughlin	University of Limerick		Background Preservation of the native artery׳s functionality can be important in both clinical and experimental applications. Although, simple cryopreservation techniques offer an attractive solution to this problem, the extent to which freezing affects the tissue׳s properties is widely debated. Earlier assessments of the mechanical properties post-freezing have been limited by one or more of the following: small sample numbers, uncontrolled inter-specimen/animal variability, failure to account for the impact of potential errors in thickness measurements, short storage times and uniaxial test methods. Material and methods Biaxial mechanical tests were performed on porcine aortic samples (n=89) extracted from superior, middle and inferior regions of five aortas, stored in isotonic saline at −20°C for 1 day, 1 week, 1, 6 and 12 months, thawed and retested. The sample׳s weight and thickness were also measured pre and post-freezing. A total of 178 tests were performed and elastic modulus was assessed by calculating the slope of the Cauchy stress–stretch curve at the low and high stretch regions in both the circumferential (θ) and longitudinal (L) directions. Results The weight of the samples increased post-freezing. However, in general, no significant difference was found between the elastic modulus of porcine aortic tissue before and after freezing at −20°C and was unaffected by storage time. Although more accurate measuring instruments are warranted to confirm this finding, minor changes to the elastic modulus as a result of freezing were negatively correlated with regional variances i.e. changes in the elastic modulus decreased from the superior to the inferior region. Conclusions These results indicate that for applications which require preservation of the gross mechanical properties, storing the tissue at −20 °C in isotonic saline, for an extended period of time, is acceptable. Cell/Tissue: porcine aortic tissue, cardiovascular tissue
BioTester	2014	Vascular Elastography: A Validation Study	Ultrasound in Medicine & Biology	R.G.P. Lopata, M.F.J. Peters, J. Nijs, C.W. Oomens, M.C.M. Rutten, F.N. van de Vosse	Eindhoven University of Technology		Vascular elastography techniques are promising tools for mechanical characterization of diseased arteries. These techniques are usually validated with simulations or phantoms or by comparing results with histology or other imaging modalities. In the study described here, vascular elastography was applied to porcine aortas in vitro during inflation testing (n = 10) and results were compared with those of standard bi-axial tensile testing, a technique that directly measures the force applied to the tissue. A neo-Hookean model was fit to the stress-strain data, valid for large deformations. Results indicated good correspondence between the two techniques, with GUS = 110 ± 11 kPa and GTT = 108 ± 10 kPa for ultrasound and tensile testing, respectively. Bland-Altman analysis revealed little bias (GUS−GTT = 2 ± 20 kPa). The next step will be the application of a non-linear material model that is also adaptable for in vivo measurements. Cell/Tissue: cardiovascular tissue, arterial tissue human peripheral blood mononuclear cells macrophages
BioTester	2015	Adipose Derived Tissue Engineered Heart Valve	Journal of Tissue Science & Engineering	L. Frese, B. Sanders, G.M. Beer, B. Weber, A. Driessen-Mol, F.P.T. Baaijens, S.P. Hoerstrup	University Hospital of Zurich		Introduction: A major challenge associated with heart valve tissue engineering is the in vitro creation of mature tissue structures compliant with native valve functionality. Various cell types have been investigated for heart valve tissue engineering. In addition to prenatal, umbilical cord- and vascular-derived cells, mesenchymal stem cells (MSCs) have gained large interest for tissue engineering purposes, because of their broad differentiation potential. However, bone marrow derived MSCs require a highly invasive harvesting procedure and decline in both cell number and differentiation potential proportionally with the donor’s age. In contrast, adipose derived stem cells (ADSCs) represent an interesting alternative. The ease of repeated access to subcutaneous adipose tissue as well as the less invasive donation procedures provide clear advantages. Therefore, this study investigated the suitability of ADSCs as alternative cell source for tissue engineered heart valves (TEHVs). Methods: Human ADSCs were seeded on TEHV-scaffolds (n=11) made of nonwoven polyglycolic acid coated with poly-4-hydroxybutyrate. TEHVs were cultivated in diastolic-pulse-duplicator-bioreactor systems and subsequently seeded with a superficial layer of ADSC-derived endothelial cells. Quantitative assessment of extracellular matrix composition of the TEHV-leaflets was performed with biochemical analyses for sulphated glycosaminoglycans, hydroxyproline and DNA content. Microstructural evaluation was performed on representative samples of the TEHVleaflets by (immuno-)histochemistry and scanning electron microscopy. The mechanical properties of the ADSC derived TEHV-leaflets were characterized by biaxial tensile tests. Results: ADSC-derived TEHV-leaflets showed a homogenous vital cell distribution throughout the whole leaflet structure that consisted of large amounts of glycosaminoglycans and collagen and was endothelialized. Furthermore, the mechanically stable matrix of the ADSC-derived TEHVs showed a stiffness range in the right order of magnitude for heart valve applications. Conclusion: Human ADSCs represent a promising alternative autologous mesenchymal cell source for TEHVs that is of large clinical relevance due to their easy accessibility, efficient proliferation and excellent tissue formation capacities Cell/Tissue: heart valve tissue cardiac tissue adipose derived stem cellsendothelial cells
BioTester	2015	Biaxial Stress Relaxation Of Semilunar Heart Valve Leaflets During Simulated Collagen Catabolism: Effects Of Collagenase Concentration And Equibiaxial Strain State	Journal of Engineering in Medicine	S. Huang, H.-Y. S. Huang	North Carolina State University		Heart valve leaflet collagen turnover and remodeling are innate to physiological homeostasis; valvular interstitial cells routinely catabolize damaged collagen and affect repair. Moreover, evidence indicates that leaflets can adapt to altered physiological (e.g. pregnancy) and pathological (e.g. hypertension) mechanical load states, tuning collagen structure and composition to changes in pressure and flow. However, while valvular interstitial cell-secreted matrix metalloproteinases are considered the primary effectors of collagen catabolism, the mechanisms by which damaged collagen fibers are selectively degraded remain unclear. Growing evidence suggests that the collagen fiber strain state plays a key role, with the strain-dependent configuration of the collagen molecules either masking or presenting proteolytic sites, thereby protecting or accelerating collagen proteolysis. In this study, the effects of equibiaxial strain state on collagen catabolism were investigated in porcine aortic valve and pulmonary valve tissues. Bacterial collagenase (0.2 and 0.5 mg/mL) was utilized to simulate endogenous matrix metalloproteinases, and biaxial stress relaxation and biochemical collagen concentration served as functional and compositional measures of collagen catabolism, respectively. At a collagenase concentration of 0.5 mg/mL, increasing the equibiaxial strain imposed during stress relaxation (0%, 37.5%, and 50%) yielded significantly lower median collagen concentrations in the aortic valve (p = 0.0231) and pulmonary valve (p = 0.0183), suggesting that relatively large strain magnitudes may enhance collagen catabolism. Collagen concentration decreases were paralleled by trends of accelerated normalized stress relaxation rate with equibiaxial strain in aortic valve tissues. Collectively, these in vitro results indicate that biaxial strain state is capable of affecting the susceptibility of valvular collagens to catabolism, providing a basis for further investigation of how such phenomena may manifest at different strain magnitudes or in vivo. Cell/Tissue: porcine aortic valve tissue porcine pulmonary valve tissue cardivascular tissue
BioTester	2015	Building A Better Infarct: Modulation Of Collagen Cross-Linking To Increase Infarct Stiffness And Reduce Left Ventricular Dilation Post-Myocardial Infarction	Journal of Molecular and Cellular Cardiology	A.P. Voorhees, K.Y. DeLeon-Pennell, Y. Ma, G. V. Halade, A. Yabluchanskiy, R. Padmanabhan, E. Flynn, C.A. Cates, M.L. Lindsey, H.-C. Han	University of Texas		Matrix metalloproteinase-9 (MMP-9) deletion attenuates collagen accumulation and dilation of the left ventricle (LV) post-myocardial infarction (MI); however the biomechanical mechanisms underlying the improved outcome are poorly understood. The aim of this study was to determine the mechanisms whereby MMP-9 deletion alters collagen network composition and assembly in the LV post-MI to modulate the mechanical properties of myocardial scar tissue. Adult C57BL/6J wild-type (WT; n = 88) and MMP-9 null (MMP-9−/−; n = 92) mice of both sexes underwent permanent coronary artery ligation and were compared to day 0 controls (n = 42). At day 7 post-MI, WT LVs displayed a 3-fold increase in end-diastolic volume, while MMP-9−/− showed only a 2-fold increase (p < 0.05). Biaxial mechanical testing revealed that MMP-9−/− infarcts were stiffer than WT infarcts, as indicated by a 1.3-fold reduction in predicted in vivo circumferential stretch (p < 0.05). Paradoxically, MMP-9−/− infarcts had a 1.8-fold reduction in collagen deposition (p < 0.05). This apparent contradiction was explained by a 3.1-fold increase in lysyl oxidase (p < 0.05) in MMP-9−/− infarcts, indicating that MMP-9 deletion increased collagen cross-linking activity. Furthermore, MMP-9 deletion led to a 3.0-fold increase in bone morphogenetic protein-1, the metalloproteinase that cleaves pro-collagen and pro-lysyl oxidase (p < 0.05) and reduced fibronectin fragmentation by 49% (p < 0.05) to enhance lysyl oxidase activity. We conclude that MMP-9 deletion increases infarct stiffness and prevents LV dilation by reducing collagen degradation and facilitating collagen assembly and cross-linking through preservation of the fibronectin network and activation of lysyl oxidase. Cell/Tissue: myocardial tissue cardiac tissue
BioTester	2015	Corneal Collagen Cross-Linking Combined With Simulation Of Femtosecond Laser-Assisted Refractive Lens Extraction: An Ex Vivo Biomechanical Effect Evaluation	The Journal of Cornea and External Disease	A.J. Kanellopoulos, M.A. Kontos, S. Chen, G. Asimellis	New York University		PURPOSE: To evaluate biomechanical changes induced by in situ corneal cross-linking (CXL) with stromal pocket delivered enhanced concentration riboflavin and high-fluence, high-energy UV-A irradiation. METHODS: Eight human donor corneas were subjected to intrastromal lamellar corneal tissue removal of anterior 140-μm deep, 80-μm thick × 5-mm diameter central stromal buttons, extracted through a 3.5-mm width tunnel, surfacing in the superior cornea periphery. Enhanced concentration riboflavin solution (0.25%) was instilled in the pocket. In study group A (CXL), superficial high-fluence UV-A irradiation was applied, whereas in control group B (no CXL), none. To comparatively assess changes in corneal rigidity, corneal specimens were subjected to transverse biaxial resistance measurements by application of a unidirectional tangential shear force. Biomechanical differences were evaluated through stress and Young shear modulus. RESULTS: Stress at 10% strain was 305 ± 24 kPa in study group A versus 157 ± 11 kPa in control group B (relative difference Δ = 107%, P = 0.021). Stress at 20% strain was 1284 ± 34 kPa in study group A versus 874 ± 29 kPa in control group B (Δ = 47%, P = 0.043). Average shear modulus in study group A at 10% strain was 6.98 ± 1.12 MPa versus 4.04 ± 0.85 MPa in control group B (Δ = 73%, P = 0.036). Average shear modulus in study group A at 20% strain was 11.46 ± 0.75 MPa versus 8.80 ± 0.72 MPa in group B (Δ = 30%, P = 0.047). CONCLUSIONS: Adjunct CXL in this ex vivo simulation refractive lens extraction procedure seems to provide significant increase in corneal rigidity, up to +107%. These findings also support our previous reported work on laser in situ keratomileusis combined with CXL. Cell/Tissue: lamellar corneal tissue
BioTester	2015	Development Of Non-Cell Adhesive Vascular Grafts Using Supramolecular Building Blocks	Macromolecular Bioscience Journal	G. C. van Almen, H. Talacua, B.D. Ippel, B.B. Mollet, M. Ramaekers, M. Simonet, A.I.P.M. Smits, C.V.C. Bouten, J. Kluin, P.Y.W. Dankers	Eindhoven University of Technology		Cell-free approaches to in situ tissue engineering require materials that are mechanically stable and are able to control cell-adhesive behavior upon implantation. Here, the development of mechanically stable grafts with non-cell adhesive properties via a mix-and-match approach using ureido-pyrimidinone (UPy)-modified supramolecular polymers is reported. Cell adhesion is prevented in vitro through mixing of end-functionalized or chain-extended UPy-polycaprolactone (UPy-PCL or CE-UPy-PCL, respectively) with end-functionalized UPy-poly(ethylene glycol) (UPy-PEG) at a ratio of 90:10. Further characterization reveals intimate mixing behavior of UPy-PCL with UPy-PEG, but poor mechanical properties, whereas CE-UPy-PCL scaffolds are mechanically stable. As a proof-of-concept for the use of non-cell adhesive supramolecular materials in vivo, electrospun vascular scaffolds are applied in an aortic interposition rat model, showing reduced cell infiltration in the presence of only 10% of UPy-PEG. Together, these results provide the first steps toward advanced supramolecular biomaterials for in situ vascular tissue engineering with control over selective cell capturing. Cell/Tissue: cardiovascular, vascular, aortic
BioTester	2015	High-Irradiance Cxl Combined With Myopic Lasik: Flap And Residual Stroma Biomechanical Properties Studied Ex-Vivo	British Journal of ophthalmology	A.J. Kanellopoulos, G. Asimellis, B.Salvador-Culla, J. Chodosh, J.B. Ciolino	New York University		Background/aims To evaluate ex vivo biomechanical and enzymatic digestion resistance differences between standard myopic laser in-situ keratomileusis (LASIK) compared with LASIK+CXL, in which high-irradiance cross-linking (CXL) is added. Methods Eight human donor corneas were subjected to femtosecond-assisted myopic LASIK. Group A (n=4) served as a control group (no CXL). The corneas in LASIK+CXL group B were subjected to concurrent prophylactic high-irradiance CXL (n=4). Saline-diluted (0.10%) riboflavin was instilled on the stroma, subsequently irradiated with UV-A through the repositioned flap. The cornea stroma and flap specimens were separately subjected to transverse biaxial resistance measurements; biomechanical differences were assessed via stress and Young's shear modulus. Subsequently, the specimens were subjected to enzymatic degradation. Results For the corneal stroma specimen, stress at 10% strain was 128±11 kPa for control group A versus 293±20 kPa for the LASIK+CXL group B (relative difference Δ=+129%, p<0.05). The stress in group B was also increased at 20% strain by +68% (p<0.05). Shear modulus in group B was increased at 10% strain by +79%, and at 20% strain by +48% (both statistically significant, p<0.05). The enzymatic degradation time to dissolution was 157.5±15.0 min in group A versus 186.25±7.5 min in group B (Δ=+18%, p=0.014). For the flaps, both biomechanical, as well as enzymatic degradation tests showed no significant differences. Conclusions LASIK+CXL appears to provide significant increase in underlying corneal stromal rigidity, up to +130%. Additionally, there is significant relevant enzymatic digestion resistance confirmatory to the above. LASIK flaps appear unaffected biomechanically by the LASIK+CXL procedure, suggesting effective CXL just under the flap. Cell/Tissue: cornea, corneal stroma
BioTester	2015	Leaflet Stress And Strain Distributions Following Incomplete Transcatheter Aortic Valve Expansion	Journal of Biomechanics	M. Abbasi, A.N. Azadani	University of Denver		Transcatheter aortic valve replacement (TAVR) is an established treatment alternative to surgical valve replacement in high-risk patients with severe symptomatic aortic stenosis. The current guidelines for TAVR are to upsize transcatheter aortic valve (TAV) relative to the native annulus to secure the device and minimize paravalvular leakage. Unlike surgical stented bioprosthetic valves where leaflets are attached to a rigid frame, TAVs must expand to fit within the native annulus. Fully-expanded circular TAVs have consistent leaflet kinematics; however, subtle variations in the degree of stent expansion may affect leaflet coaptation. The objective of this study was to determine the impact of incomplete TAV expansion on leaflet stress and strain distributions. In this study, we developed finite element models of a 23 mm homemade TAV expanded to diameters ranging from 18 to 23 mm in 1 mm increments. Through dynamic finite element simulations, we found that leaflet stress and strain distributions were dependent on the diameter of the inflated TAV. After complete expansion of the TAV to 23 mm, high stress and strain regions were observed primarily in the commissures during diastole. However, 2–3 mm incomplete TAV stent expansion induced localized high stress regions within the TAV commissures, while 4–5 mm incomplete stent expansion induced localized high stress regions within the belly of the TAV leaflets during the diastolic phase of the cardiac cycle. Increased mechanical stress and flexural deformation on TAV leaflets due to incomplete stent expansion may lead to accelerated tissue degeneration and diminished long-term valve durability. Cell/Tissue: transcatheter aortic valve cardiovascular tissue
BioTester	2015	Methods For Using 3-D Ultrasound Speckle Tracking In Biaxial Mechanical Testing Of Biological Tissue Samples	Ultrasound in Medicine & Biology Journal	C.H. Yap, D.W. Park, D. Dutta, M. Simon, K. Kim	University of Pittsburgh		Being multilayered and anisotropic, biological tissues such as cardiac and arterial walls are structurally complex, making the full assessment and understanding of their mechanical behavior challenging. Current standard mechanical testing uses surface markers to track tissue deformations and does not provide deformation data below the surface. In the study described here, we found that combining mechanical testing with 3-D ultrasound speckle tracking could overcome this limitation. Rat myocardium was tested with a biaxial tester and was concurrently scanned with high-frequency ultrasound in three dimensions. The strain energy function was computed from stresses and strains using an iterative non-linear curve-fitting algorithm. Because the strain energy function consists of terms for the base matrix and for embedded fibers, spatially varying fiber orientation was also computed by curve fitting. Using finite-element simulations, we first validated the accuracy of the non-linear curve-fitting algorithm. Next, we compared experimentally measured rat myocardium strain energy function values with those in the literature and found a matching order of magnitude. Finally, we retained samples after the experiments for fiber orientation quantification using histology and found that the results satisfactorily matched those computed in the experiments. We conclude that 3-D ultrasound speckle tracking can be a useful addition to traditional mechanical testing of biological tissues and may provide the benefit of enabling fiber orientation computation. Cell/Tissue: cardiac tissue, myocardium arterial tissue cardiovascular tissue
BioTester	2015	Nanocomposite Silicone Hydrogels With A Laser-Assisted Surface Modification For Inhibiting The Growth Of Baterial Biofilm	Journal of Materials Chemistry B	P. Yin, G.B. Huang, W.H. Tse, Y.G. Bao, J. Denstedt, J. Zhang	University of Western Ontario		Surface and interface modifications of synthetic silicone hydrogels used for wearable and implantable medical devices, e.g. catheters and contact lenses, are critical to overcome their poor mechanical properties and biofouling. In this paper, silica nanoparticles (SiO2 NPs) were incorporated within silicone hydrogels through photo-polymerization. As compared to the silicone hydrogel, the nanocomposited silicone hydrogel shows highly textured microstructures, increased swelling behaviour and improved stiffness. Meanwhile, a hydrophilic surface of silicone hydrogel is important to minimize protein fouling which forms a conditioning layer for the growth of bacterial biofilm. Here, we applied matrix-assisted pulsed laser evaporation (MAPLE) with a pulsed Nd:YAG laser at 532 nm to deposit polyethylene glycol (PEG) on the surface of the nanocomposited silicone hydrogels. The PEG deposited on the nanocomposited silicone hydrogels forms islands at the submicron-scale, which increase with increasing irradiation time (t). The protein adsorption on nanocomposited silicone hydrogel with PEG deposition decreases over 40 ± 2% when t = 2 h. Compared to the commercial silicone catheters, the nanocomposited silicone hydrogel with PEG deposition can reduce the growth of bacteria from 1.20 × 106 CFU cm−2 to 3.69 × 105 CFU cm−2. In addition, the relative cell viabilities of NIH/3T3 mouse fibroblast cells treated using the nanocomposited silicone hydrogels coated with/without PEG were studied. No toxic effect is imposed on the cells. Consequently, the MAPLE process is a controllable, contamination-free technique to modify the surface of silicone hydrogels. We expect that the nanocomposited silicone hydrogels with appropriate surface treatment can be applied in various wearable and implantable medical devices. Cell/Tissue: syntehtic silicone hydrogels mouse fibroblast cells
BioTester	2015	Regional And Depth Variability Of Porcine Meniscal Mechanical Properties Through Biaxial Testing	Journal of the Mechanical Behavior of Biomedical Materials	A. Kahlon, M.B. Hurtig, K.D. Gordon	University of Guelph		The menisci in the knee joint undergo complex loading in-vivo resulting in a multidirectional stress distribution. Extensive mechanical testing has been conducted to investigate the tissue properties of the knee meniscus, but the testing conditions do not replicate this complex loading regime. Biaxial testing involves loading tissue along two different directions simultaneously, which more accurately simulates physiologic loading conditions. The purpose of this study was to report mechanical properties of meniscal tissue resulting from biaxial testing, while simultaneously investigating regional variations in properties. Ten left, fresh porcine joints were obtained, and the medial and lateral menisci were harvested from each joint (twenty menisci total). Each menisci was divided into an anterior, middle and posterior region; and three slices (femoral, deep and tibial layers) were obtained from each region. Biaxial and constrained uniaxial testing was performed on each specimen, and Young׳s moduli were calculated from the resulting stress strain curves. Results illustrated significant differences in regional mechanical properties, with the medial anterior (Young׳s modulus (E)=11.14±1.10 MPa), lateral anterior (E=11.54±1.10 MPa) and lateral posterior (E=9.0±1.2 MPa) regions exhibiting the highest properties compared to the medial central (E=5.0±1.22 MPa), medial posterior (E=4.16±1.13 MPa) and lateral central (E=5.6±1.20 MPa) regions. Differences with depth were also significant on the lateral meniscus, with the femoral (E=12.7±1.22 MPa) and tibial (E=8.6±1.22 MPa) layers exhibiting the highest Young׳s moduli. This data may form the basis for future modeling of meniscal tissue, or may aid in the design of synthetic replacement alternatives. Cell/Tissue: meniscus medial meniscilateral menisci
BioTester	2015	Superior Tissue Evolution In Slow Degrading Scaffolds For Valvular Tissue Engineering	Tissue Engineering Part A	M. Brugmans, S. Soekhradj-Soechit, D. van Geemen, M.A.J. Cox, C. Bouten, F.P.T. Baaijens, A. Driessen-Mol	Eindhoven University of Technology		Synthetic polymers are widely used to fabricate porous scaffolds for the regeneration of cardiovascular tissues. To ensure mechanical integrity, a balance between the rate of scaffold absorption and tissue formation is of high importance. A higher rate of tissue formation is expected in fast-degrading materials than in slow-degrading materials. This could be a result of synthetic cells, which aim to compensate for the fast loss of mechanical integrity of the scaffold by deposition of collagen fibers. Here, we studied the effect of fast-degrading polyglycolic acid scaffolds coated with poly-4-hydroxybutyrate (PGA-P4HB) and slow-degrading poly-ɛ-caprolactone (PCL) scaffolds on amount of tissue, composition, and mechanical characteristics in time, and compared these engineered values with values for native human heart valves. Electrospun PGA-P4HB and PCL scaffolds were either kept unseeded in culture or were seeded with human vascular-derived cells. Tissue formation, extracellular matrix (ECM) composition, remaining scaffold weight, tissue-to-scaffold weight ratio, and mechanical properties were analyzed every week up to 6 weeks. Mass of unseeded PCL scaffolds remained stable during culture, whereas PGA-P4HB scaffolds degraded rapidly. When seeded with cells, both scaffold types demonstrated increasing amounts of tissue with time, which was more pronounced for PGA-P4HB-based tissues during the first 2 weeks; however, PCL-based tissues resulted in the highest amount of tissue after 6 weeks. This study is the first to provide insight into the tissue-to-scaffold weight ratio, therewith allowing for a fair comparison between engineered tissues cultured on scaffolds as well as between native heart valve tissues. Although the absolute amount of ECM components differed between the engineered tissues, the ratio between ECM components was similar after 6 weeks. PCL-based tissues maintained their shape, whereas the PGA-P4HB-based tissues deformed during culture. After 6 weeks, PCL-based engineered tissues showed amounts of cells and ECM that were comparable to the number of human native heart valve leaflets, whereas values were lower in the PGA-P4HB-based tissues. Although increasing in time, the number of collagen crosslinks were below native values in all engineered tissues. In conclusion, this study indicates that slow-degrading scaffold materials are favored over fast-degrading materials to create organized ECM-rich tissues in vitro, which keep their three-dimensional structure before implantation. Cell/Tissue: cardiovascular tissue
BioTester	2015	The Biomechanics Of Eyelid Tarsus Tissue	Journal of Biomechanics	M.T. Sun, D. Pham, A.J. O'Conner, J. Wood, R. Casson, D. Selva, J.J. Costi	South Australian Institute of Ophthalmology and Royal Adelaide Hospital		Reconstruction of the eyelid remains challenging due to the unique properties of the tarsal plate, which is a fibrocartilagenous structure within the eyelid providing structural support and physical form. There are no previous studies investigating the biomechanical properties of tarsus tissue, which is vital to the success of bioengineered tarsal substitutes. We therefore aimed to determine the biomechanical properties of human tarsus tissue, and used a CellScale BioTester 5000 (CellScale, Waterloo, Canada) to perform uniaxial tensile tests on ten samples of healthy eyelid tarsus. All samples were tested ‘fresh’ within two hours of harvest. A tensile preload of 50 mN was applied for 10 min before the sample was subjected to uniaxial tension under linear ramp displacement control. Maximum strain was 30% of the original tissue length and thirty dynamic cycles were performed at a strain rate of 1%/s using a triangular waveform. Of the samples tested, the mean (SD) width was 5.51 mm (1.45 mm) whilst mean thickness was 1.6 mm (0.51 mm). The mean toe modulus was 0.14 (0.10) MPa, elastic modulus was 1.73 (0.61) MPa, with an extensibility of 15.8 (2.1)%, and phase angle of 6.4° (2.4)°. After adjusting for the initial tissue slack, the maximum strain ranged from 23.8% to 30.0%. At maximum strain, it was observed that the linear region of the stress–strain curve was reached without the sample slipping out of the clamps. Our results establish a benchmark for native tarsus tissue, which can be used when evaluating tissue engineered tarsal substitutes in the future. Cell/Tissue: tarsus tissue
BioTester	2016	A Comparison Between Porcine, Ovine, And Bovine Intervertebral Disc Anatomy And Single Lamella Annulus Fibrosis Tensile Properties	Journal of Morphology	L.A. Monaco, S.J. DeWitte-Orr, D.E. Gregory	Wilfrid Laurier University		This project aimed to compare gross anatomical measures and biomechanical properties of single lamellae from the annulus fibrosus of ovine and porcine lumbar vertebrae, and bovine tail vertebrae. The morphology of the vertebrae of these species differ significantly both from each other and from human, yet how these differences alter biomechanical properties is unknown. Geometric parameters measured in this study included: 1) absolute and relative intervertebral (IVD) and vertebral body height and 2) absolute and relative intervertebral disc (IVD) anterior-posterior (AP) and medial-lateral (ML) widths. Single lamella tensile properties included toe-region stress and stretch ratio, stiffness, and tensile strength. As expected, the bovine tail IVD revealed a more circular shape compared with both the ovine and porcine lumbar IVD. The bovine tail also had the largest IVD to vertebral body height ratio (due to having the highest absolute IVD height). Bovine tail lamellae were also found to be strongest and stiffest (in tension) while ovine lumbar lamellae were weakest and most compliant. Histological analysis revealed the greatest proportion of collagen in the bovine corroborating findings of increased strength and stiffness. The observed differences in anatomical shape, connective tissue composition, and tensile properties need to be considered when choosing an appropriate model for IVD research. J. Morphol. 277:244–251, 2016. Cell/Tissue: porcine aortic valve tissue porcine pulmonary valve tissue cardivascular tissue
BioTester	2016	Acute Pergolide Exposure Stiffens Engineered Valve Interstitial Cell Tissues And Reduces Contractility In Vitro	Cardiovascular Pathology	A.K. Capulli, L.A. MacQueen, B.B. O'Conner, S. Dauth, K.K. Parker	Harvard University		Medications based on ergoline-derived dopamine and serotonin agonists are associated with off-target toxicities that include valvular heart disease (VHD). Reports of drug-induced VHD resulted in the withdrawal of appetite suppressants containing fenfluramine and phentermine from the US market in 1997 and pergolide, a Parkinson's disease medication, in 2007. Recent evidence suggests that serotonin receptor activity affected by these medications modulates cardiac valve interstitial cell activation and subsequent valvular remodeling, which can lead to cardiac valve fibrosis and dysfunction similar to that seen in carcinoid heart disease. Failure to identify these risks prior to market and continued use of similar drugs reaffirm the need to improve preclinical evaluation of drug-induced VHD. Here, we present two complimentary assays to measure stiffness and contractile stresses generated by engineered valvular tissues in vitro. As a case study, we measured the effects of acute (24 h) pergolide exposure to engineered porcine aortic valve interstitial cell (AVIC) tissues. Pergolide exposure led to increased tissue stiffness, but it decreased both basal and active contractile tone stresses generated by AVIC tissues. Pergolide exposure also disrupted AVIC tissue organization (i.e., tissue anisotropy), suggesting that the mechanical properties and contractile functionality of these tissues are governed by their ability to maintain their structure. We expect further use of these assays to identify off-target drug effects that alter the phenotypic balance of AVICs, disrupt their ability to maintain mechanical homeostasis, and lead to VHD. Cell/Tissue: cardiac aortic valve interstitial cell tissues cardiovascular tissue
BioTester	2016	Age-Dependent Changes In Geometry, Tissue Composition And Mechanical Properties Of Fetal To Adult Cryopreserved Human Heart Valves	PLOS One (Public Library of Science One)	D. van Geeman, A.L.F. Soares, P.J.M. Oomen, A. Dressen-Mol, M.W.J.T. Janssen-van den Broek, A.J. van den Bogaerdt, J.J.C. Bogers, M-J. T.H. Goumans, F.P.T. Baaijens, C.V.C. Bouten	Eindhoven University of Technology		There is limited information about age-specific structural and functional properties of human heart valves, while this information is key to the development and evaluation of living valve replacements for pediatric and adolescent patients. Here, we present an extended data set of structure-function properties of cryopreserved human pulmonary and aortic heart valves, providing age-specific information for living valve replacements. Tissue composition, morphology, mechanical properties, and maturation of leaflets from 16 pairs of structurally unaffected aortic and pulmonary valves of human donors (fetal-53 years) were analyzed. Interestingly, no major differences were observed between the aortic and pulmonary valves. Valve annulus and leaflet dimensions increase throughout life. The typical three-layered leaflet structure is present before birth, but becomes more distinct with age. After birth, cell numbers decrease rapidly, while remaining cells obtain a quiescent phenotype and reside in the ventricularis and spongiosa. With age and maturation–but more pronounced in aortic valves–the matrix shows an increasing amount of collagen and collagen cross-links and a reduction in glycosaminoglycans. These matrix changes correlate with increasing leaflet stiffness with age. Our data provide a new and comprehensive overview of the changes of structure-function properties of fetal to adult human semilunar heart valves that can be used to evaluate and optimize future therapies, such as tissue engineering of heart valves. Changing hemodynamic conditions with age can explain initial changes in matrix composition and consequent mechanical properties, but cannot explain the ongoing changes in valve dimensions and matrix composition at older age. Cell/Tissue: cardiac valve cardiovascular tissue aortic pulmonary
BioTester	2016	Are Adipose-Derived Stem Cells Cultivated In Human Platelet Lysate Suitable For Heart Valve Tissue Engineering	Journal of Tissue Engineering and Regenerative Medicine	L. Frese, T. Sasse, B. Sanders, F.P.T. Baaijens, G.M. Beer, S.P. Hoerstrup	University Hospital of Zurich		Tissue-engineered heart valves represent a promising strategy for the growing need for valve replacements in cardiovascular medicine. Recent studies have shown that adipose-derived stem cells (ADSC) are a viable cell source, as they are readily available in both the young and the elderly, show diverse differentiation potential and adapt their extracellular matrix (ECM) to a varying mechanical load. In vitro culture medium is usually enriched with fetal calf serum (FCS). However, a promising substitute has recently been found in human platelet lysate (HPL), which is superior in terms of proliferation speed and allogenicity. This study sought to elucidate the suitability of ADSC and HPL for heart valve tissue engineering (TE). ADSC harvested from five healthy individuals were cultured in both FCS and HPL. The cells were observed for differentiation potential, proliferation speed and immunophenotype, using immunohistochemistry and FACS analysis. Neotissue was assessed for ECM composition, human collagen I (hColl1) formation, histomorphology and mechanical stiffness under uniaxial tensile stress. Neotissue cultured in HPL was found to be significantly inferior in mechanical rigidity; it showed a three-fold higher proliferation rate and a more dense ECM, but also a more heterogeneous hColl1 distribution. ECM analysis showed significantly higher amounts of DNA and glycosaminoglycans (GAG) in HPL-cultured tissue. No significant differences were observed for differentiation potential and immunophenotype, apart from a lower CD166 expression in HPL. The mechanical inferiority of neotissue cultured in HPL represents a limitation to the use of HPL-enriched media for heart valve TE with ADSC. This result concurs with data published about HPL and myofibroblasts derived from the venous wall. Similarly, the mechanical inferiority is not rooted in a difference in ECM composition, but rather in hColl1 architecture. Stem cell properties, as documented in the literature, are retained with HPL. A possible connection between the mechanical inferiority and the observed decrease in CD166 needs further investigation. Cell/Tissue: cardiovascular tissue adipose derived stem cells
BioTester	2016	Biaxial Quantification Of Deep Layer Transverse Carpal Ligament Elastic Properties By Sex And Region	Clinical Biomechanics	B. Mathers, A. Agur, M. Oliver, K. Gordon	University of Guelph		Background The transverse carpal ligament is a major component of the carpal tunnel and is an important structure in the etiology of carpal tunnel syndrome. The current study aimed to quantify biaxial elastic moduli of the transverse carpal ligament and compare differences between sex and region (Radial and Ulnar). Methods Biaxial testing of radial and ulnar samples from twenty-two (thirteen male, nine female) human fresh frozen cadaveric transverse carpal ligaments was performed. Elastic moduli and stiffness were calculated and compared. Findings Biaxial elastic moduli of the transverse carpal ligament ranged from 0.76 MPa to 3.38 MPa, varying based on region (radial and ulnar), testing direction (medial-lateral and proximal-distal) and sex. Biaxial elastic moduli were significantly larger in the medial-lateral direction than the proximal-distal direction (P < 0.001). Moduli were significantly larger ulnarly than radially (P = 0.001). No significant differences due to gender were noted. Interpretation The regional variations in biaxial elastic moduli of the transverse carpal ligament may help improve non-invasive treatment methods for carpal tunnel syndrome, specifically manipulative therapy. The smaller biaxial elastic moduli found in the radial region suggests that manipulative therapy should be focused on the radial aspect of the transverse carpal ligament. The trend where female transverse carpal ligaments had larger stiffness in the ulnar location than males suggests that that the increased prevalence of carpal tunnel syndrome in women may be related to an increased stiffness of the transverse carpal ligament, however further work is warranted to evaluate this trend. Cell/Tissue: transverse carpal ligament radial ulnar
BioTester	2016	Biomechanical Characterization Of Ascending Aortic Aneurysms	Biomechanics and Modeling in MechanoBiology	M. Smoljkic, H. Fehervary, P. Van den Bergh, A. Jorge-Penas, L. Kluyskens, S. Dymarkowski, P. Verbrugghe, B. Meuris, J. Vander Sloten, N. Famaey	Katholieke Universiteit Leuven		Ascending thoracic aortic aneurysms (ATAAs) are a silent disease, ultimately leading to dissection or rupture of the arterial wall. There is a growing consensus that diameter information is insufficient to assess rupture risk, whereas wall stress and strength provide a more reliable estimate. The latter parameters cannot be measured directly and must be inferred through biomechanical assessment, requiring a thorough knowledge of the mechanical behaviour of the tissue. However, for healthy and aneurysmal ascending aortic tissues, this knowledge remains scarce. This study provides the geometrical and mechanical properties of the ATAA of six patients with unprecedented detail. Prior to their ATAA repair, pressure and diameter were acquired non-invasively, from which the distensibility coefficient, pressure–strain modulus and wall stress were calculated. Uniaxial tensile tests on the resected tissue yielded ultimate stress and stretch values. Parameters for the Holzapfel–Gasser–Ogden material model were estimated based on the pre-operative pressure–diameter data and the post-operative stress–stretch curves from planar biaxial tensile tests. Our results confirmed that mechanical or geometrical information alone cannot provide sufficient rupture risk estimation. The ratio of physiological to ultimate wall stress seems a more promising parameter. However, wall stress estimation suffers from uncertainties in wall thickness measurement, for which our results show large variability, between patients but also between measurement methods. Our results also show a large strength variability, a value which cannot be measured non-invasively. Future work should therefore be directed towards improved accuracy of wall thickness estimation, but also towards the large-scale collection of ATAA wall strength data. Cell/Tissue: thoracic aortic cadiovascular cardiac tissue
BioTester	2016	Biomechanical Properties And Microstructure Of Heart Chambers: A Paired Comparison Study In An Ovine Model	Annals of Biomedical Engineering	S. Javani, M. Gordon, A.N. Azadani	University of Denver		Mechanical properties of the cardiac tissue play an important role in normal heart function. The goal of this study was to determine the passive mechanical properties of all heart chambers through a paired comparison study in an ovine model. Ovine heart was used due its physiological and anatomical similarities to human heart. A total of 189 specimens from anterior and posterior portions of the left and right ventricles, atria, and appendages underwent biaxial mechanical testing. A Fung-type strain energy function was used to fit the experimental data. Tissue behavior was quantified based on the magnitude of strain energy, as indicator of tissue stiffness, at equibiaxial strains of 0.10, 0.15, and 0.20. Statistical analysis revealed no significant difference in strain energy storage between anterior and posterior portions of each chamber, except for the right ventricle where strain energy storage in the posterior specimens were higher than the anterior specimens. Additionally, all chambers from the left side of the heart had significantly higher strain energy storage than the corresponding chambers on the right side. Furthermore, the highest to lowest stored strain energy were associated with ventricles, appendages, and atria, respectively. Microstructure of tissue specimens from different chambers was also compared using histology.Cell/Tissue: cardiac tissue ventricular atrial
BioTester	2016	Characterization Of Three-Dimensional Anisotropic Heart Valve Tissue Mechanical Properties Using Inverse Finite Element Analysis	Journal of the Mechanical Behavior of Biomedical Materials	M. Abbasi, M.S. Barakat, K. Vanhidkhah, A.N. Azadani	University of Denver		Computational modeling has an important role in design and assessment of medical devices. In computational simulations, considering accurate constitutive models is of the utmost importance to capture mechanical response of soft tissue and biomedical materials under physiological loading conditions. Lack of comprehensive three-dimensional constitutive models for soft tissue limits the effectiveness of computational modeling in research and development of medical devices. The aim of this study was to use inverse finite element (FE) analysis to determine three-dimensional mechanical properties of bovine pericardial leaflets of a surgical bioprosthesis under dynamic loading condition. Using inverse parameter estimation, 3D anisotropic Fung model parameters were estimated for the leaflets. The FE simulations were validated using experimental in-vitro measurements, and the impact of different constitutive material models was investigated on leaflet stress distribution. The results of this study showed that the anisotropic Fung model accurately simulated the leaflet deformation and coaptation during valve opening and closing. During systole, the peak stress reached to 3.17 MPa at the leaflet boundary while during diastole high stress regions were primarily observed in the commissures with the peak stress of 1.17 MPa. In addition, the Rayleigh damping coefficient that was introduced to FE simulations to simulate viscous damping effects of surrounding fluid was determined. Cell/Tissue: pericardium, pericardial leaflets cardiovascular tissue
BioTester	2016	Constitutive Description Of Human Femoropopliteal Artery Aging	Biomechanics and Modeling in MechanoBiology	A. Kamenskiy, A. Seas, P. Deegan, W. Poulson, E. Anttila, S. Sim, A. Desyatova, J. MacTaggart	University of Nebraska		Femoropopliteal artery (FPA) mechanics play a paramount role in pathophysiology and the artery’s response to therapeutic interventions, but data on FPA mechanical properties are scarce. Our goal was to characterize human FPAs over a wide population to derive a constitutive description of FPA aging to be used for computational modeling. Fresh human FPA specimens ( n=579n=579) were obtained from n=351n=351 predominantly male (80 %) donors 54±15 years old (range 13–82 years). Morphometric characteristics including radius, wall thickness, opening angle, and longitudinal pre-stretch were recorded. Arteries were subjected to multi-ratio planar biaxial extension to determine constitutive parameters for an invariant-based model accounting for the passive contributions of ground substance, elastin, collagen, and smooth muscle. Nonparametric bootstrapping was used to determine unique sets of material parameters that were used to derive age-group-specific characteristics. Physiologic stress–stretch state was calculated to capture changes with aging. Morphometric and constitutive parameters were derived for seven age groups. Vessel radius, wall thickness, and circumferential opening angle increased with aging, while longitudinal pre-stretch decreased ( p<0.01p<0.01). Age-group-specific constitutive parameters portrayed orthotropic FPA stiffening, especially in the longitudinal direction. Structural changes in artery wall elastin were associated with reduction of physiologic longitudinal and circumferential stretches and stresses with age. These data and the constitutive description of FPA aging shed new light on our understanding of peripheral arterial disease pathophysiology and arterial aging. Application of this knowledge might improve patient selection for specific treatment modalities in personalized, precision medicine algorithms and could assist in device development for treatment of peripheral artery disease. Cell/Tissue: femoropopliteal artery cardiovascular tissue arterial
BioTester	2016	Deposition Of A Hydrophilic Nanocomposite-Based Coating On A Silicone Hydrogel Through A Laser Process To Minimize Uv Exposure And Bacterial Contamination	RSC Advances (Royal Society of Chemistry Advances)	G. Huang, W. H. Tse, J. Zhang	University of Western Ontario		To prevent contact lens from biofouling and to minimize UV exposure to human eyes, a nanocomposite-based coating made of silver (Ag) nanoparticles and polyvinylpyrrolidone (PVP) is deposited on synthetic silicone hydrogels through matrix assisted pulsed laser evaporation (MAPLE) with a pulsed Nd:YAG laser at 532 nm. The average diameter of Ag NPs that have undergone the MAPLE process for 60 min is 11.61 ± 3.58 nm. The thickness of the Ag–PVP nanocomposite coating with a deposition time of 60 min is around 930 ± 15 nm. Our results demonstrate that the oxygen permeability of silicone hydrogel with a nanocomposite coating is similar to that of commercialized contact lenses; over 60% of UV light in the range of 300–450 nm can be blocked. Moreover, the silicone hydrogel with the nanocomposite coating can reduce over 65.4 ± 1.6% of protein (human lysozyme) absorption as compared to silicone hydrogel-based contact lens, and kill all cultured bacteria in 8 hours. This research work demonstrates a new way to deposit biocompatible nanocomposite coatings on silicone hydrogels used as contact lens to efficiently minimize UV exposure and biofouling. Cell/Tissue: synthetic silicone hydrogels
BioTester	2016	Improved Geometry Of Decellularized Tissue Engineered Heart Valves To Prevent Leaflet Retraction	Annals of Biomedical Engineering	B. Sanders, S. Loerakker, E.S. Fioretta, D.J.P. Bax, A. Driessen-Mol, S.P. Hoerstrup, F.P.T. Baaijens	Eindhoven University of Technology		Recent studies on decellularized tissue engineered heart valves (DTEHVs) showed rapid host cell repopulation and increased valvular insufficiency developing over time, associated with leaflet shortening. A possible explanation for this result was found using computational simulations, which revealed radial leaflet compression in the original valvular geometry when subjected to physiological pressure conditions. Therefore, an improved geometry was suggested to enable radial leaflet extension to counteract for host cell mediated retraction. In this study, we propose a solution to impose this new geometry by using a constraining bioreactor insert during culture. Human cell based DTEHVs (n = 5) were produced as such, resulting in an enlarged coaptation area and profound belly curvature. Extracellular matrix was homogeneously distributed, with circumferential collagen alignment in the coaptation region and global tissue anisotropy. Based on in vitro functionality experiments, these DTEHVs showed competent hydrodynamic functionality under physiological pulmonary conditions and were fatigue resistant, with stable functionality up to 16 weeks in vivo simulation. Based on implemented mechanical data, our computational models revealed a considerable decrease in radial tissue compression with the obtained geometrical adjustments. Therefore, these improved DTEHV are expected to be less prone to host cell mediated leaflet retraction and will remain competent after implantation. Cell/Tissue: heart valve tissue cardiac tissue cardiovascular tissue
BioTester	2016	Inflation And Bi-Axial Tensile Testing Of Healthy Porcine Carotid Arteries	Ultrasound in Medicine & Biology	R.W. Boekhoven, M.F.J. Peters, M.C.M. Rutten, M.R. van Sambeek, F.N. van de Vosse, R.G.P Lopata	Eindhoven University of Technology		Knowledge of the intrinsic material properties of healthy and diseased arterial tissue components is of great importance in diagnostics. This study describes an in vitro comparison of 13 porcine carotid arteries using inflation testing combined with functional ultrasound and bi-axial tensile testing. The measured tissue behavior was described using both a linear, but geometrically non-linear, one-parameter (neo-Hookean) model and a two-parameter non-linear (Demiray) model. The shear modulus estimated using the linear model resulted in good agreement between the ultrasound and tensile testing methods, GUS = 25 ± 5.7 kPa and GTT = 23 ± 5.4 kPa. No significant correspondence was observed for the non-linear model aUS = 1.0 ± 2.7 kPa vs. aTT = 17 ± 8.8 kPa, p ∼ 0); however, the exponential parameters were in correspondence (bUS = 12 ± 4.2 vs. bTT = 10 ± 1.7, p > 0.05). Estimation of more complex models in vivo is cumbersome considering the sensitivity of the model parameters to small changes in measurement data and the absence of intraluminal pressure data, endorsing the use of a simple, linear model in vivo. Cell/Tissue: arterial tissue cardiovascular tissue
BioTester	2016	Modulation Of Collaen Fiber Orientation By Strain-Controlled Enzymatic Degradation	Acta Biomaterialia	S. Ghazanfari, A. Dressen-Mol, C.V.C. Bouten, F.P.T. Baaijens	Eindhoven University of Technology		Collagen fiber anisotropy has a significant influence on the function and mechanical properties of cardiovascular tissues. We investigated if strain-dependent collagen degradation can explain collagen orientation in response to uniaxial and biaxial mechanical loads. First, decellularized pericardial samples were stretched to a fixed uniaxial strain and after adding a collagen degrading enzyme (collagenase), force relaxation was measured to calculate the degradation rate. This data was used to identify the strain-dependent degradation rate. A minimum was observed in the degradation rate curve. It was then demonstrated, for the first time, that biaxial strain in combination with collagenase alters the collagen fiber alignment from an initially isotropic distribution to an anisotropic distribution with a mean alignment corresponding with the strain at the minimum degradation rate, which may be in between the principal strain directions. When both strains were smaller than the minimum degradation point, fibers tended to align in the direction of the larger strain and when both strains were larger than the minimum degradation, fibers mainly aligned in the direction of the smaller strain. However, when one strain was larger and one was smaller than the minimum degradation point, the observed fiber alignment was in between the principal strain directions. In the absence of collagenase, uniaxial and biaxial strains only had a slight effect on the collagen (re)orientation of the decellularized samples. Statement of Significance Collagen fiber orientation is a significant determinant of the mechanical properties of native tissues. To mimic the native-like collagen alignment in vitro, we need to understand the underlying mechanisms that direct this alignment. In the current study, we aimed to control collagen fiber orientation by applying biaxial strains in the presence of collagenase. We hypothesized that strain‐dependent collagen degradation can describe specific collagen orientation when biaxial mechanical strains are applied. Based on this hypothesis, collagen fibers align in the direction where the degradation is minimal. Pericardial tissues, as isotropic collagen matrices, were decellularized and subjected to a fixed uniaxial strain. Then, collagenase was added to initiate the collagen degradation and the relaxation of force was measured to indicate the degradation rate. The V‐shaped relationship between degradation rate and strain was obtained to identify the minimum degradation rate point. It was then demonstrated, for the first time, that biaxial strain in combination with collagenase alters the collagen fiber alignment from almost isotropic to a direction corresponding with the strain at the minimum degradation rate. Cell/Tissue: cardiovascular tissue pericardial
BioTester	2016	Nanocomposited Coatings Produced By Laser-Assisted Process To Prevent Silicone Hydrogels From Protein Fouling And Bacterial Contamination	Applied Surface Science	G. Huang, Y. Chen, J. Zhang	University of Western Ontario		Zinc oxide (ZnO) nanoparticles incorporating with polyethylene glycol (PEG) were deposited together on the surface of silicone hydrogel through matrix-assisted pulsed laser evaporation (MAPLE). In this process, frozen nanocomposites (ZnO–PEG) in isopropanol were irradiated under a pulsed Nd:YAG laser at 532 nm for 1 h. Our results indicate that the MAPLE process is able to maintain the chemical backbone of polymer and prevent the nanocomposite coating from contamination. The ZnO–PEG nanocomposited coating reduces over 50% protein absorption on silicone hydrogel. The cytotoxicity study shows that the ZnO–PEG nanocomposites deposited on silicone hydrogels do not impose the toxic effect on mouse NIH/3T3 cells. In addition, MAPLE-deposited ZnO–PEG nanocomposites can inhibit the bacterial growth significantly.
BioTester	2016	Patient Specific Vascular Benchtop Models For Development And Validation Of Medical Devices For Minimally Invasive Procedures	Journal of Medical Robotics Research	M. Kvasnytsia, N. Famaey, M. Bohm, E. Verhoelst	Katholieke Universiteit Leuven		Using realistic benchtop models in early stages of device development can reduce time and efforts necessary to move the device to further testing. In this study, we propose several patient specific vascular benchtop models for the development and validation of a robotic catheter for transcatheter aortic valve implantation. The design and manufacturing of these models, and their properties are presented. Additionally, it is demonstrated that the described design process provides virtual models that are accurately linked to the physical models. Cell/Tissue: cardiovascular tissue aortic valve
BioTester	2016	Peak Stress In The Annulus Fibrosis Under Cyclic Biaxial Tensile Loading	Journal of Biomechanical Engineering	C.E. Gooyers, J.P. Callaghan	University of Waterloo		Numerous in vitro studies have examined the initiation and propagation of fatigue injury pathways in the annulus fibrosus (AF) using isolated motion segments; however, the cycle-varying changes to the AF under cyclic biaxial tensile loading conditions have yet to be examined. Therefore, the primary objective of this study was to characterize the cycle-varying changes in peak tensile stress in multilayer AF tissue samples within a range of physiologically relevant loading conditions at subacute magnitudes of tissue stretch up to 100 loading cycles. A secondary aim was to examine whether the stress-relaxation response would be different across loading axes (axial and circumferential) and whether this response would vary across regions of the intervertebral disk (IVD) (anterior and posterior–lateral). The results from the study demonstrate that several significant interactions emerged between independent factors that were examined in the study. Specifically, a three-way interaction between the radial location, magnitude of peak tissue stretch, and cycle rate (p = 0.0053) emerged. Significant two-way interactions between the magnitude of tissue stretch and cycle number (p < 0.0001) and the magnitude of tissue stretch and loading axis (p < 0.0001) were also observed. These findings are discussed in the context of known mechanisms for structural damage, which have been linked to fatigue loading in the IVD (e.g., cleft formation, radial tearing, increased neutral zone, disk bulging, and loss of intradiscal pressure). Cell/Tissue: annulus fibrosus
BioTester	2016	Phototactic Guidance Of A Tissue-Engineered Soft-Robotic Ray	Science Journal	S-J. Park, M. Gazzola, K.S. Park, S. Park, V. Di Santo, E.L Blevins, J.U. Lind, P.H. Campbell, S. Dauth, K.K. Parker	Harvard University		Inspired by the relatively simple morphological blueprint provided by batoid fish such as stingrays and skates, we created a biohybrid system that enables an artificial animal—a tissue-engineered ray—to swim and phototactically follow a light cue. By patterning dissociated rat cardiomyocytes on an elastomeric body enclosing a microfabricated gold skeleton, we replicated fish morphology at Embedded Image scale and captured basic fin deflection patterns of batoid fish. Optogenetics allows for phototactic guidance, steering, and turning maneuvers. Optical stimulation induced sequential muscle activation via serpentine-patterned muscle circuits, leading to coordinated undulatory swimming. The speed and direction of the ray was controlled by modulating light frequency and by independently eliciting right and left fins, allowing the biohybrid machine to maneuver through an obstacle course. Cell/Tissue: cardiomyocytes
BioTester	2016	Planar Biaxial Testing Of Heart Valve Cusp Replacement Biomaterials: Experiments, Theory And Material Constants	Acta Biomaterialia	M.R. Labrosse, R. Jafar, J.Ngu, M. Boodhwani	University of Ottawa		Objectives Aortic valve (AV) repair has become an attractive option to correct aortic insufficiency. Yet, cusp reconstruction with various cusp replacement materials has been associated with greater long-term repair failures, and it is still unknown how such materials mechanically compare with native leaflets. We used planar biaxial testing to characterize six clinically relevant cusp replacement materials, along with native porcine AV leaflets, to ascertain which materials would be best suited for valve repair. Methods We tested at least six samples of: 1) fresh autologous porcine pericardium (APP), 2) glutaraldehyde fixed porcine pericardium (GPP), 3) St Jude Medical pericardial patch (SJM), 4) CardioCel patch (CC), 5) PeriGuard (PG), 6) Supple PeriGuard (SPG) and 7) fresh porcine AV leaflets (PC). We introduced efficient displacement-controlled testing protocols and processing, as well as advanced convexity requirements on the strain energy functions used to describe the mechanical response of the materials under loading. Results The proposed experimental and data processing pipeline allowed for a robust in-plane characterization of all the materials tested, with constants determined for two Fung-like hyperelastic, anisotropic strain energy models. Conclusions Overall, CC and SPG (respectively PG) patches ranked as the closest mechanical equivalents to young (respectively aged) AV leaflets. Because the native leaflets as well as CC, PG and SPG patches exhibit significant anisotropic behaviors, it is suggested that the fiber and cross-fiber directions of these replacement biomaterials be matched with those of the host AV leaflets. Cell/Tissue: pericardium cardiac tissue aortic valve cardiovascular tissue
BioTester	2016	Planar Biaxial Testing Of Soft Biological Tissue Using Rakes: A Critical Analysis Of Protocol And Fitting Process	Journal of the Mechanical Behavior of Biomedical Materials	H. Fehervary, M. Smoljkic, J.Vander Sloten, N. Famaey	KU Leuven		Mechanical characterizationofsoftbiologicaltissueisbecomingmoreandmoreprevalent. Despite thegrowinguseofplanarbiaxialtestingforsofttissuecharacterization,testing conditions andsubsequentdataanalysishavenotbeenstandardizedandvarywidely.This also influences thequalityoftheresultoftheparameter fitting. Moreover,thetesting conditions anddataanalysisareoftennotorincompletelyreported,whichimpedesthe proper comparisonofparametersobtainedfromdifferentstudies. With afocusonplanarbiaxialtestsusingrakes,thispaperinvestigatesvaryingtesting conditions andvaryingdataanalysismethodsandtheireffectonthequalityofthe parameter fitting results.Bymeansofaseriesof finite elementsimulations,aspectssuch as numberofrakes,rakes'width,loadingprotocol,constitutivemodel,materialstiffness and anisotropyareevaluatedbasedonthedegreeofhomogeneityofthestress field, and on thecorrelationbetweentheexperimentallyobtainedstressandthestressderivedfrom the constitutivemodel.Whencalculatingtheaforementionedstresses,differentdefinitions ofthesectionwidthanddeformationgradientareusedinliterature,eachofwhich are lookedinto.Apartfromthisdegreeofhomogeneityandcorrelation,alsotheeffecton the qualityoftheparameter fitting resultisevaluated. The resultsshowthatinhomogeneitiescanbereducedtoaminimumforwisechoices of testingconditionsandanalysismethods,butnevercompletelyeliminated.Therefore,a new parameteroptimizationprocedureisproposedthatcorrectsfortheinhomogeneities in thestress field andinducessignificant improvementstothe fitting results.Recommen- dations aremadeforbestpracticeinrake-basedplanarbiaxialtestingofsoftbiological tissues andsubsequentparameter fitting, andguidelinesareformulatedforreporting thereof inpublications.
BioTester	2016	Production Of Synthetic, Para-Aramid And Biopolymer Nanofibers By Immersion Rotary Jet-Spinning	Macromolecular Materials and Engineering	G.M. Gonzalez, L.A. MacQueen, J.U. Lind, S.A. Fitzgibbons, C. O. Chantre, I. Huggler, H.M. Golecki, J.A. Goss, K.K. Parker	Harvard University		Nanofiber production platforms commonly rely on volatile carrier solvents or high voltages. Production of nanofibers comprised of charged polymers or polymers requiring nonvolatile solvents thus typically requires customization of spinning setup and polymer dope. In severe cases, these challenges can hinder fiber formation entirely. Here, a versatile system is presented which addresses these challenges by employing centrifugal force to extrude polymer dope jet through an air gap, into a flowing precipitation bath. This voltage-free approach ensures that nanofiber solidification occurs in liquid, minimizing surface tension instability that results in jet breakup and fiber defects. In addition, nanofibers of controlled size and morphology can be fabricated by tuning spinning parameters including air gap length, spinning speed, polymer concentration, and bath composition. To demonstrate the versatility of our platform, para-aramid (e.g., Kevlar) and biopolymer (e.g., DNA, alginate) nanofibers are produced that cannot be readily produced using standard nanofiber production methods. Cell/Tissue: nanofibers
BioTester	2016	Quantification Of Coupled Stiffness And Fiber Orientation Remodeling In Hypertensive Rat Right-Ventricular Myocardium Using 3d Ultrasound Speckle Tracking With Biaxial Testing	PLOS One (Public Library of Science One)	D.W. Park, A. Sebastiani, C.H. Yap, M.A. Simon, K. Kim	University of Pittsburgh		Mechanical and structural changes of right ventricular (RV) in response to pulmonary hypertension (PH) are inadequately understood. While current standard biaxial testing provides information on the mechanical behavior of RV tissues using surface markers, it is unable to fully assess structural and mechanical properties across the full tissue thickness. In this study, the mechanical and structural properties of normotensive and pulmonary hypertension right ventricular (PHRV) myocardium through its full thickness were examined using mechanical testing combined with 3D ultrasound speckle tracking (3D-UST). RV pressure overload was induced in Sprague–Dawley rats by pulmonary artery (PA) banding. The second Piola–Kirchhoff stress tensors and Green-Lagrangian strain tensors were computed in the RV myocardium using the biaxial testing combined with 3D-UST. A previously established non-linear curve-fitting algorithm was applied to fit experimental data to a Strain Energy Function (SEF) for computation of myofiber orientation. The fiber orientations obtained by the biaxial testing with 3D-UST compared well with the fiber orientations computed from the histology. In addition, the re-orientation of myofiber in the right ventricular free wall (RVFW) along longitudinal direction (apex-to-outflow-tract direction) was noticeable in response to PH. For normotensive RVFW samples, the average fiber orientation angles obtained by 3D-UST with biaxial test spiraled from 20° at the endo-cardium to -42° at the epi-cardium (Δ = 62°). For PHRV samples, the average fiber orientation angles obtained by 3D-UST with biaxial test had much less spiral across tissue thickness: 3° at endo-cardium to -7° at epi-cardium (Δ = 10°, P<0.005 compared to normotensive). Cell/Tissue: right ventricular myocardium cardiac cardiovascular endo-cardium epi-cardium
BioTester	2016	The Aging Disc: Using An Ovine Model To Examine Age-Related Differences In The Biomechanical Properties Of The Intralamellar Matrix Of Single Lamellae	European Spine Journal	D.M. Stewart, L.A. Monaco, D.E. Gregory	Wilfrid Laurier University		Purpose To determine the effect of age on the biomechanical properties of the intralamellar matrix of single annulus fibrosus (AF) lamellae. Methods One intervertebral disc (IVD) was excised from five young (<12 months), five middle-aged (2–4 years) and five older (5–7 years) ovine lumbar spines. From each IVD, a maximum of four single AF lamellae samples were harvested: two from the anterior region and two from the posterior region. Tissues were mounted in a tensile testing apparatus such that tension was applied perpendicular to the orientation of the collagen fibers to isolate the intralamellar matrix. Variables of interest from the stress–strain relationship were: end of toe-region strain and corresponding stress, initial failure stress and strain, and elastic stiffness. Results When compared to the middle-aged and old samples, the intralamellar matrix of young AF samples displayed significantly higher stress values at the end of the end of toe-region (p = 0.008) and at initial failure (p = 0.002). Further, the young samples were stiffer than both middle-aged and old samples (p = 0.04). Conclusions This study was the first to show that the intralamellar matrix of single AF lamellae is weaker and more compliant in middle-aged and old ovine IVDs compared to young IVDs. These findings are likely a result of the remarkable age-related changes that occur that ultimately weaken the IVD as a whole. Cell/Tissue: annulus fibrosus lamellae
BioTester	2016	The Choice Of A Constitutive Formulation For Modeling Limb Flexion-Induced Deformations And Stresses In The Human Femoropopliteal Arteries Of Different Ages	Biomechanics and Modeling in MechanoBiology	A. Desyatova, J. MacTaggart, W. Poulson, P. Deegan, C. Lomneth, A. Sandip, A. Kamenskiy	University of Nebraska		Open and endovascular treatments for peripheral arterial disease are notorious for high failure rates. Severe mechanical deformations experienced by the femoropopliteal artery (FPA) during limb flexion and interactions between the artery and repair materials play important roles and may contribute to poor clinical outcomes. Computational modeling can help optimize FPA repair, but these simulations heavily depend on the choice of constitutive model describing the arterial behavior. In this study finite element model of the FPA in the standing (straight) and gardening (acutely bent) postures was built using computed tomography data, longitudinal pre-stretch and biaxially determined mechanical properties. Springs and dashpots were used to represent surrounding tissue forces associated with limb flexion-induced deformations. These forces were then used with age-specific longitudinal pre-stretch and mechanical properties to obtain deformed FPA configurations for seven age groups. Four commonly used invariant-based constitutive models were compared to determine the accuracy of capturing deformations and stresses in each age group. The four-fiber FPA model most accurately portrayed arterial behavior in all ages, but in subjects younger than 40 years, the performance of all constitutive formulations was similar. In older subjects, Demiray (Delfino) and classic two-fiber Holzapfel–Gasser–Ogden formulations were better than the Neo-Hookean model for predicting deformations due to limb flexion, but both significantly overestimated principal stresses compared to the FPA or Neo-Hookean models. Cell/Tissue: femoropopliteal artery cardiovascular tissue arterial
BioTester	2016	The Effect Of Local Hydration Environment On The Mechanical Properties And Unloaded Temporal Changes Of Isolated Porcine Annular Samples	Journal of Biomedical Engineering	K.M. Gruevski, C.E. Gooyers, T. Karakolis, J.P. Callaghan	University of Waterloo		Preventing dehydration during in vitro testing of isolated layers of annulus fibrosus tissue may require different test conditions than functional spine units. The purpose of the study was twofold: (A) to quantify changes in mass and thickness of multilayer annulus samples in four hydration environments over 120 min; and (B) to quantify cycle-varying biaxial tensile properties of annulus samples in the four environments. The environments included a saline bath, air, relative humidity control, and misting combined with controlled humidity. The loading protocol implemented 24 cycles of biaxial tensile loading to 20% strain at a rate of 2%/s with 3-, 8-, and 13-min of intermittent rest. Specimen mass increased an average (standard deviation) 72% (11) when immersed for 120 min (p < 0.0001). The air condition and the combined mist and relative humidity conditions reduced mass by 45% (15) and 25% (23), respectively, after 120 min (p < 0.0014). Stress at 16% stretch in the air condition was higher at cycle 18 (18 min of exposure) and cycle 24 (33 min of exposure) compared to all other environments in both the axial and circumferential directions (p < 0.0460). There was no significant change in mass or thickness over time in the relative humidity condition and the change in circumferential stress at 16% stretch between cycles 6 and 24 was a maximum of 0.099 MPa and not statistically significant. Implementation of a controlled relative humidity environment is recommended to maintain hydration of isolated annulus layers during cyclic tensile testing. Cell/Tissue: annulus fibrosus tissue
BioTester	2017	Biaxial Experimental And Analytical Characterization Of A Dielectric Elastomer	Applied Physics A	A. Helal, M. Doumit, R. Shaheen	University of Ottawa		Electroactive polymers (EAPs) have emerged as a strong contender for use in low-cost efficient actuators in multiple applications especially related to biomimetic and mobile-assistive devices. Dielectric elastomers (DE), a subcategory of these smart materials, have been of particular interest due to their large achievable deformation and favourable mechanical and electro-mechanical properties. Previous work has been completed to understand the behaviour of these materials; however, their properties require further investigation to properly integrate them into real-world applications. In this study, a biaxial tensile experimental evaluation of 3M™ VHB 4905 and VHB 4910 is presented with the purpose of illustrating the elastomers’ transversely isotropic mechanical behaviours. These tests were applied to both tapes for equibiaxial stretch rates ranging between 0.025 and 0.300 s−1. Subsequently, a dynamic planar biaxial visco-hyperelastic constitutive relationship was derived from a Kelvin–Voigt rheological model and the general Hooke’s law for transversely isotropic materials. The model was then fitted to the experimental data to obtain three general material parameters for either tapes. The model’s ability to predict tensile stress response and internal energy dissipation, with respect to experimental data, is evaluated with good agreement. The model’s ability to predict variations in mechanical behaviour due to changes in kinematic variables is then illustrated for different conditions. Cell/Tissue: electroactive polymers elastomers
BioTester	2017	Biaxial Mechanical Properties Of Bovine Jugular Venous Valve Leaflet Tissues	Biomechanics and Modeling in MechanoBiology	H-Y. S. Huang, J. Lu	North Carolina State University		Venous valve incompetence has been implicated in diseases ranging from chronic venous insufficiency (CVI) to intracranial venous hypertension. However, while the mechanical properties of venous valve leaflet tissues are central to CVI biomechanics and mechanobiology, neither stress–strain curves nor tangent moduli have been reported. Here, equibiaxial tensile mechanical tests were conducted to assess the tangent modulus, strength and anisotropy of venous valve leaflet tissues from bovine jugular veins. Valvular tissues were stretched to 60% strain in both the circumferential and radial directions, and leaflet tissue stress–strain curves were generated for proximal and distal valves (i.e., valves closest and furthest from the right heart, respectively). Toward linking mechanical properties to leaflet microstructure and composition, Masson’s trichrome and Verhoeff–Van Gieson staining and collagen assays were conducted. Results showed: (1) Proximal bovine jugular vein venous valves tended to be bicuspid (i.e., have two leaflets), while distal valves tended to be tricuspid; (2) leaflet tissues from proximal valves exhibited approximately threefold higher peak tangent moduli in the circumferential direction than in the orthogonal radial direction (i.e., proximal valve leaflet tissues were anisotropic; p<0.01p<0.01 ); (3) individual leaflets excised from the same valve apparatus appeared to exhibit different mechanical properties (i.e., intra-valve variability); and (4) leaflets from distal valves exhibited a trend of higher soluble collagen concentrations than proximal ones (i.e., inter-valve variability). To the best of the authors’ knowledge, this is the first study reporting biaxial mechanical properties of venous valve leaflet tissues. These results provide a baseline for studying venous valve incompetence at the tissue level and a quantitative basis for prosthetic venous valve design. Cell/Tissue: venous valve leaflets tissues valvular tissues cardiovascular tissue
BioTester	2017	Comparison Of Femoropopliteal Artery Stents Under Axial And Radial Compression, Axial Tension, Bending, And Torsion Deformations	Journal of the Mechanical Behavior of Biomedical Materials	K. Maleckis, P. Deegan, W. Poulson, C. Sievers, A. Desyatova, J. MacTaggart, A. Kamenskiy	University of Nebraska		High failure rates of Peripheral Arterial Disease (PAD) stenting appear to be associated with the inability of certain stent designs to accommodate severe biomechanical environment of the femoropopliteal artery (FPA) that bends, twists, and axially compresses during limb flexion. Twelve Nitinol stents (Absolute Pro, Supera, Lifestent, Innova, Zilver, Smart Control, Smart Flex, EverFlex, Viabahn, Tigris, Misago, and Complete SE) were quasi-statically tested under bench-top axial and radial compression, axial tension, bending, and torsional deformations. Stents were compared in terms of force-strain behavior, stiffness, and geometrical shape under each deformation mode. Tigris was the least stiff stent under axial compression (6.6 N/m axial stiffness) and bending (0.1 N/m) deformations, while Smart Control was the stiffest (575.3 N/m and 105.4 N/m, respectively). Under radial compression Complete SE was the stiffest (892.8 N/m), while Smart Control had the lowest radial stiffness (211.0 N/m). Viabahn and Supera had the lowest and highest torsional stiffness (2.2 μN m/° and 959.2 μN m/°), respectively. None of the 12 PAD stents demonstrated superior characteristics under all deformation modes and many experienced global buckling and diameter pinching. Though it is yet to be determined which of these deformation modes might have greater clinical impact, results of the current analysis may help guide development of new stents with improved mechanical characteristics. Cell/Tissue: femoropopliteal artery cardiovascular tissue
BioTester	2017	Constitutive Modeling Of Human Femoropopliteal Artery Biaxial Stiffening Due To Aging And Diabetes	Acta Biomaterialia	A. Desyatova, J. MacTaggart, A. Kamenskiy	University of Nebraska		Atherosclerotic obstructive disease of the femoropopliteal artery (Peripheral Arterial Disease, PAD) is notorious for high treatment failure rates. Older age and diabetes mellitus (DM) are among the major risk factors for PAD, and both are associated with increased arterial stiffness. Our goal was to develop a constitutive model describing multiaxial arterial stiffening, and use it to portray aging of normal and diabetic human femoropopliteal arteries (FPA). Fresh human FPAs (n = 744) were obtained from 13–82-year-old donors. Arteries were tested using planar biaxial extension, and their behavior was modeled with a constitutive relation that included stiffening functions of age. FPA diameter, wall thickness, circumferential, and longitudinal opening angles increased with age, while longitudinal pre-stretch decreased. Diameter and circumferential opening angle did not change with age in subjects with DM. Younger FPAs were more compliant longitudinally but became more isotropic with age. Arteries with DM stiffened significantly faster in the circumferential direction than arteries without DM. Constitutive model accurately portrayed orthotropic stiffening with age of both normal and diabetic arteries. Constitutive description of FPA aging contributes to understanding of arterial pathophysiology and can help improve fidelity of computational models investigating device-artery interaction in PAD repair by providing more personalized arterial properties. Cell/Tissue: femoropopliteal artery cardiovascular tissue arterial
BioTester	2017	Constitutive Modeling Of Jugular Vein-Derived Venous Valve Leaflet Tissues	Journal of the Mechanical Behavior of Biomedical Materials	N. Kaul, H-Y. S. Huang	North Carolina State University		Venous valve tissues, though used in vein reconstruction surgeries and bioprosthetic valves with moderate success, have not been extensively studied with respect to their structure. Their inherent anisotropic, non-linear behavior combined with severe diseases which affect veins, such as chronic venous insufficiency, warrant understanding the structure and material behavior of these tissues. Hence, before any bioprosthetic grafts may be used in place of tissues, it is of the utmost importance to understand the mechanical and structural properties of these tissues as this may lead to higher success rates for valve replacement surgeries. The longevity of the bioprosthetics may also increase if the manufactured grafts behave the same as native valves. Building on the scant information about the uniaxial and biaxial mechanical properties of jugular venous valves and wall tissues from previous studies, the current focus of our investigation lies in understanding the material behavior by establishing a phenomenological strain energy-based constitutive relation for the tissues. We used bovine veins to study the behavior of valve leaflet tissue and adjoining wall tissue (from the proximal and distal ends of the veins) under different biaxial testing protocols. We looked at the behavior of numerical partial derivatives of the strain energy to select a suitable functional form for the strain energy for wall and valve tissues. Using this strain energy descriptor, we determined the Cauchy stress and compared it with experimental results under additional sets of displacement-controlled biaxial testing protocols to find material specific model parameters by the Powell's method algorithm. Results show that whereas wall tissue strain energy can be explained using a polynomial non-linear function, the valve tissue, due to higher non-linearities, requires an exponential function. This study may provide useful information for the primary stages of bioprosthetic designs and replacement surgeries and may support future studies investigating structural models. It may also support the study of valvular diseases by providing a way to understand material properties and behavior and to form a continuum model when required for numerical analyses and computational simulations. Cell/Tissue: venous valve leaflets tissues valvular tissues cardiovascular tissue
BioTester	2017	Design And Characterization Of A Hyperelastic Tubular Soft Composite	Journal of the mechanical behavior of biomedical materials	R. Shaheen, M. Doumit, A. Helal	University of Ottawa		Research in the field of human mobility assistive devices, aiming to reduce the metabolic cost of daily activities, is seeing the benefits of the exclusive use of passive actuators to store and release energy during the gait cycle. Current devices commonly employ either mechanical springs or Pneumatic Artificial Muscles as the primary method of passive actuation. The Pneumatic Artificial Muscle has proven to be a superior actuation choice for these devices, when compared to its alternatives. However, challenges regarding muscle pressure loss and limited elongation potential have been identified. This paper presents a hyperelastic tubular Soft Composite that replicates the distinctive mechanical behaviour of the Pneumatic Artificial Muscle without the need for internal pressurization. The proposed Soft Composite solution is achieved by impregnating a prefabricated polyethylene terephthalate braided sleeve, held at a high initial fibre angle, with a silicone prepolymer. A comprehensive experimental evaluation is achieved on numerous prototypes for a variety of customizable design parameters including: the initial fibre angle, the silicone stiffness, and the braided sleeve style. This research has successfully developed, tested, and validated a novel Soft Composite that can achieve the desired nonlinear stiffness and elongation potential for optimal use as passive actuation in human mobility assistive devices. Cell/Type: muscular hyperelastic tubular soft composite
BioTester	2017	In Situ Heart Valve Tissue Engineering Using A Bioresorbable Elastomeric Implant – From Material Design To 12 Months Follow-Up In Sheep	Biomaterials Journal	J. Kluin, H. Talacua, A.I.P.M. Smits, M. Y. Emmert, M.C.P. Brugmans, E.S. Fioretta, P.E. Dijkman, S.H.M. Sontjens, R.Duijvelshoff, S. Dekker, M.W.J.T. Janssen-van den Broek, V. Lintas, A. Vink, S.P. Hoerstrup, H.M. Janssen, P.Y.W. Dankers, F.P.T. Baaijens, C.V.C. Bouten	Eindhoven University of Technology		The creation of a living heart valve is a much-wanted alternative for current valve prostheses that suffer from limited durability and thromboembolic complications. Current strategies to create such valves, however, require the use of cells for in vitro culture, or decellularized human- or animal-derived donor tissue for in situ engineering. Here, we propose and demonstrate proof-of-concept of in situ heart valve tissue engineering using a synthetic approach, in which a cell-free, slow degrading elastomeric valvular implant is populated by endogenous cells to form new valvular tissue inside the heart. We designed a fibrous valvular scaffold, fabricated from a novel supramolecular elastomer, that enables endogenous cells to enter and produce matrix. Orthotopic implantations as pulmonary valve in sheep demonstrated sustained functionality up to 12 months, while the implant was gradually replaced by a layered collagen and elastic matrix in pace with cell-driven polymer resorption. Our results offer new perspectives for endogenous heart valve replacement starting from a readily-available synthetic graft that is compatible with surgical and transcatheter implantation procedures. Cell/Tissue: valvular tissue cardiac cardiovascular tissue
BioTester	2017	Jetvalve: Rapid Manufacturing Of Biohybrid Scaffolds For Biomimetic Heart Valve Replacement	Biomaterials Journal	A.K. Capulli, M.Y. Emmert, F.S. Pasqualini, D. Kehl, E. Caliskan, J.U. Lind, S. P. Sheehy, S.J. Park, S. Ahn, B. Weber, J.A. Goss, S.P Hoerstrup, K.K. Parker	Harvard University		Tissue engineered scaffolds have emerged as a promising solution for heart valve replacement because of their potential for regeneration. However, traditional heart valve tissue engineering has relied on resource-intensive, cell-based manufacturing, which increases cost and hinders clinical translation. To overcome these limitations, in situ tissue engineering approaches aim to develop scaffold materials and manufacturing processes that elicit endogenous tissue remodeling and repair. Yet despite recent advances in synthetic materials manufacturing, there remains a lack of cell-free, automated approaches for rapidly producing biomimetic heart valve scaffolds. Here, we designed a jet spinning process for the rapid and automated fabrication of fibrous heart valve scaffolds. The composition, multiscale architecture, and mechanical properties of the scaffolds were tailored to mimic that of the native leaflet fibrosa and assembled into three dimensional, semilunar valve structures. We demonstrated controlled modulation of these scaffold parameters and show initial biocompatibility and functionality in vitro. Valves were minimally-invasively deployed via transapical access to the pulmonary valve position in an ovine model and shown to be functional for 15 h. Cell/Tissue: heart valve tissue cardiac cardiovascular tissue
BioTester	2017	Limb Flexion-Induced Twist And Associated Intramural Stresses In The Human Femoropopliteal Artery	Journal of the Royal Society Interface	A. Desyatova, W. Poulson, P. Deegan, C. Lomneth, A. Seas, K. Maleckis, J. MacTaggart, A. Kamenskiy	University of Nebraska		High failure rates of femoropopliteal artery (FPA) interventions are often attributed to severe mechanical deformations that occur with limb movement. Torsion of the FPA likely plays a significant role, but is poorly characterized and the associated intramural stresses are currently unknown. FPA torsion in the walking, sitting and gardening postures was characterized in n = 28 in situ FPAs using intra-arterial markers. Principal mechanical stresses and strains were quantified in the superficial femoral artery (SFA), adductor hiatus segment (AH) and the popliteal artery (PA) using analytical modelling. The FPA experienced significant torsion during limb flexion that was most severe in the gardening posture. The associated mechanical stresses were non-uniformly distributed along the length of the artery, increasing distally and achieving maximum values in the PA. Maximum twist in the SFA ranged 10–13° cm−1, at the AH 8–16° cm−1, and in the PA 14–26° cm−1 in the walking, sitting and gardening postures. Maximum principal stresses were 30–35 kPa in the SFA, 27–37 kPa at the AH and 39–43 kPa in the PA. Understanding torsional deformations and intramural stresses in the FPA can assist with device selection for peripheral arterial disease interventions and may help guide the development of devices with improved characteristics. Cell/Tissue: femoropopliteal artery cardiovascular tissue arterial
BioTester	2017	Mechanically Robust Electrospun Hydrogel Scaffolds Crosslinked Via Supramolecular Interactions	Macromolecular Bioscience Journal	B.B. Mollet, S. Spaans, P.G. Fard, N.A.M. Bax, C.V.C. Bouten, P.Y.W. Dankers	Eindhoven University of Technology		One of the major challenges in the processing of hydrogels based on poly(ethylene glycol) (PEG) is to create mechanically robust electrospun hydrogel scaffolds without chemical crosslinking postprocessing. In this study, this is achieved by the introduction of physical crosslinks in the form of supramolecular hydrogen bonding ureido-pyrimidinone (UPy) moieties, resulting in chain-extended UPy-PEG polymers (CE-UPy-PEG) that can be electrospun from organic solvent. The resultant fibrous meshes are swollen in contact with water and form mechanically stable, elastic hydrogels, while the fibrous morphology remains intact. Mixing up to 30 wt% gelatin with these CE-UPy-PEG polymers introduce bioactivity into these scaffolds, without affecting the mechanical properties. Manipulating the electrospinning parameters results in meshes with either small or large fiber diameters, i.e., 0.63 ± 0.36 and 2.14 ± 0.63 µm, respectively. In that order, these meshes provide support for renal epithelial monolayer formation or a niche for the culture of cardiac progenitor cells. Cell/Tissue: synthetic hydrogrel Upy-PEG polymers renal epithelial monolayer
BioTester	2017	Nondestructive Mechanical Characterization Of Developing Biological Tissues Using Inflation Testing	Journal of the mechanical behavior of biomedical materials	P.J.A. Oomen, M.A.J. van Kelle, C.W.J. Oomens, C.V.C. Bouten, S. Loerakker	Eindhoven University of Technology		One of the hallmarks of biological soft tissues is their capacity to grow and remodel in response to changes in their environment. Although it is well-accepted that these processes occur at least partly to maintain a mechanical homeostasis, it remains unclear which mechanical constituent(s) determine(s) mechanical homeostasis. In the current study a nondestructive mechanical test and a two-step inverse analysis method were developed and validated to nondestructively estimate the mechanical properties of biological tissue during tissue culture. Nondestructive mechanical testing was achieved by performing an inflation test on tissues that were cultured inside a bioreactor, while the tissue displacement and thickness were nondestructively measured using ultrasound. The material parameters were estimated by an inverse finite element scheme, which was preceded by an analytical estimation step to rapidly obtain an initial estimate that already approximated the final solution. The efficiency and accuracy of the two-step inverse method was demonstrated on virtual experiments of several material types with known parameters. PDMS samples were used to demonstrate the method's feasibility, where it was shown that the proposed method yielded similar results to tensile testing. Finally, the method was applied to estimate the material properties of tissue-engineered constructs. Via this method, the evolution of mechanical properties during tissue growth and remodeling can now be monitored in a well-controlled system. The outcomes can be used to determine various mechanical constituents and to assess their contribution to mechanical homeostasis. Cell/Tissue: Polydimethylsiloxane PDMS
BioTester	2017	On The Influence Of Wall Calcification And Intraluminal Thrombus On Prediction Of Abdominal Aortic Aneurysm Rupture	Journal of Vascular Surgery	H.E. Barrett, E.M. Cunnane, H. Hidayat, J.M. O'Brien, M.A. Moloney, E.G. Kavanagh, M.T. Walsh	University of Limerick		Objective Parameters other than maximum diameter that predict rupture of abdominal aortic aneurysms (AAAs) may be helpful for risk-benefit analysis in individual patients. The aim of this study was to characterize the biomechanical-structural characteristics associated with AAA walls to better identify the related mechanistic variables required for an accurate prediction of rupture risk. Methods Anterior AAA wall (n = 40) and intraluminal thrombus (ILT; n = 114) samples were acquired from 18 patients undergoing open surgical repair. Biomechanical characterization was performed using controlled circumferential stretching tests combined with a speckle-strain tracking technique to quantify the spatial heterogeneity in deformation and localized strains in the AAA walls containing calcification. After mechanical testing, the accompanying microstructural characteristics of the AAA wall and ILT types were examined using electron microscopy. Results No significant correlation was found between the AAA diameter and the wall mechanical properties in terms of Cauchy stress (rs = −0.139; P = .596) or stiffness (rs = −0.451; P = .069). Quantification of significant localized peak strains, which were concentrated in the tissue regions surrounding calcification, reveals that peak strains increased by a mean of 174% as a result of calcification and corresponding peak stresses by 18.2%. Four ILT types characteristic of diverse stages in the evolving tissue microstructure were directly associated with distinct mechanical stiffness properties of the ILT and underlying AAA wall. ILT types were independent of geometric factors, including ILT volume and AAA diameter measures (ILT stiffness and AAA diameter [rs = −0.511; P = .074]; ILT stiffness and ILT volume [rs = −0.245; P = .467]). Conclusions AAA wall stiffness properties are controlled by the load-bearing capacity of the noncalcified tissue portion, and low stiffness properties represent a highly degraded vulnerable wall. The presence of calcification that is contiguous with the inner wall causes severe tissue overstretching in surrounding tissue areas. The results highlight the use of additional biomechanical measures, detailing the biomechanical-structural characteristics of AAA tissue, that may be a helpful adjunct to improve the accuracy of rupture prediction. Cell/Tissue: abdominal aortic intraluminal thrombus cardiovascular tissue
BioTester	2017	Paraspinal Muscle Passive Stiffness Remodels In Direct Response To Spine Stiffness: A Study Using The Ent1-Deficient Mouse	Spine Journal	K. Gsell, D. Zwambag, D.E. Fournier, C.A. Seguin, S.H.M. Brown	University of Guelph		Study Design. Basic science study of the relationship between the structural properties of the spine and its surrounding musculature. Objective. To determine whether an increase in spine stiffness causes an inverse compensatory change in the passive stiffness of the adjacent paraspinal muscles. Summary of Background Data. Intervertebral disc degeneration causes an increase in multifidus passive stiffness; this was hypothesized to compensate for a decrease in spine stiffness associated with disc degeneration. Mice lacking equilibrative nucleoside transporter 1 (ENT1) develop progressive ectopic calcification of the fibrous connective tissues of the spine, which affects the lumbar spine by 6 months of age and likely creates a mechanically stiffer spine. Methods. Experiments were conducted on four groups of mice (n = 8 mice/group): wild-type (WT) and ENT1 knockout (KO) at 2 or 8 months of age. Lumbar spines were removed and tested in cyclic axial compression to determine neutral zone length and stiffness. Single muscle fibers and bundles of fibers were isolated from lumbar multifidus and erector spinae, as well as tibialis anterior (a non–spine-related control) and tested to determine elastic modulus (passive stiffness). Results. At 2 months of age, neither spine nor muscle stiffness was different between KO and WT. At 8 months of age, compared with WT the lumbar spines of ENT1 KO mice had a stiffer and shorter neutral zone, and the paraspinal muscle fibers were less stiff; however, fiber bundles were not different. In addition, tibialis anterior was not different between KO and WT. Conclusion. This work has confirmed that calcification of spinal connective tissues in the ENT1 KO mouse results in a stiffened spine, whereas the concurrent decrease in muscle fiber elastic modulus in the adjacent paraspinal muscles suggests a direct compensatory relationship between the stiffness of the spine and the muscles that are attached to it. Cell/Tissue: lumbar multifiduserector spinaetibialis anterior and paraspinal muscles
BioTester	2017	Tensile Behaviour Of Individual Fibre Bundles In The Human Lumbar Anulus Fibrosus	Journal of Biomechanics	D.T. Pham, J.G. Shapter, J.J. Costi	Flinders University		Disc degeneration is a common medical affliction whose origins are not fully understood. An improved understanding of its underlying mechanisms could lead to the development of more effective treatments. The aim of this paper was to investigate the effect of (1) degeneration, (2) circumferential region and (3) strain rate on the microscale mechanical properties (toe region modulus, linear modulus, extensibility, phase angle) of individual fibre bundles in the anulus fibrosus lamellae of the human intervertebral disc. Healthy and degenerate fibre bundles excised from different circumferential regions in the outer anulus (posterolateral, lateral, anterolateral, anterior) were tensile tested at slow (0.1%/s), medium (1%/s) and fast (10%/s) strain rates using a micromechanical testing system. Our preliminary results showed that neither degeneration nor circumferential region significantly affected the fibre bundles’ mechanical behaviour. However, when the fibre bundles were tested at higher strain rates, this resulted in significantly higher linear moduli and lower phase angles. These findings, compared with data from other studies investigating single and multiple lamellae sections, suggest that degeneration has minimal effect on outer anulus mechanics irrespective of structural level, and the inter- and intra-lamellar arrangement and continuity of the fibre bundles may influence the lamellae’s regional behaviour and viscoelasticity. Cell/Tissue: anulus fibrosus lamellae intervertebral lumbar spine
BioTester	2017	Urinary Bladder Versus Gastrointestinal Tissue: A Comparative Study Of Their Biomechanical Properties For Urinary Tract Reconstruction	Urology the Gold Journal	N.F. Davis, J.J.E. Mulvihill, S. Mulay, E.M. Cunnane, D.M. Bolton, M.T. Walsh	University of Limerick		Objective To evaluate the mechanical properties of gastrointestinal (GI) tissue segments and to compare them with the urinary bladder for urinary tract reconstruction. Methods Urinary bladders and GI tissue segments were sourced from porcine models (n = 6, 7 months old [5 male; 1 female]). Uniaxial planar tension tests were performed on bladder tissue, and Cauchy stress-stretch ratio responses were compared with stomach, jejunum, ileum, and colonic GI tissue. Results The biomechanical properties of the bladder differed significantly from jejunum, ileum, and colonic GI tissue. Young modulus (kPa—measure of stiffness) of the GI tissue segments was on average 3.07-fold (±0.21 standard error) higher than bladder tissue (P < .01), and the strain at Cauchy stress of 50 kPa for bladder tissues was on average 2.27-fold (±0.20) higher than GI tissues. There were no significant differences between the averaged stretch ratio and Young modulus of the horizontal and vertical directions of bladder tissue (315.05 ± 49.64 kPa and 283.62 ± 57.04, respectively, P = .42). However, stomach tissues were 1.09- (±0.17) and 0.85- (±0.03) fold greater than bladder tissues for Young modulus and strain at 50 kPa, respectively. Conclusion An ideal urinary bladder replacement biomaterial should demonstrate mechanical equivalence to native tissue. Our findings demonstrate that GI tissue does not meet these mechanical requirements. Knowledge on the biomechanical properties of bladder and GI tissue may improve development opportunities for more suitable urologic reconstructive biomaterials. Cell/Tissue: gastrointestinal jejunum ileum colonicurinary bladder
BioTester	2018	Biaxial Mechanical Behavior Of Bovine Saphenous Venous Valve Leaflets	Journal of the mechanical behavior of biomedical materials	J. Lu, H-Y S. Huang	North Carolina State University		Chronic venous disease is caused by chronic venous insufficiency (CVI), which results in significant symptoms such as venous ulcers, ankle eczema, leg swelling, etc. Venous valve incompetence is a major cause of CVI. When the valves of veins in the leg become incompetent (i.e., do not close properly), blood is able to flow backwards (i.e., reflux), which results in blood pooling in the lower extremities, distal venous hypertension, and CVI. Current clinical therapies, such as surgical venous valve reconstruction and bioprosthetic venous valve replacement, are highly invasive and only moderately successful. This is due, in part, to the scanty information available about venous valve leaflet structure and mechanical properties. To date, only one previous study by our research group has reported on the mechanical properties of venous valve leaflet tissue, and specifically in the case of jugular vein valves. In this study, we conducted equibiaxial tensile tests on bovine saphenous vein valve leaflet tissues to better understand their nonlinear, anisotropic mechanical behavior. By stretching the valvular tissues to 60% strain in both the circumferential and radial directions, we generated stress-strain curves for proximal (i.e., those closest to the heart) and distal (i.e., those furthest from the heart) valve leaflets. Histology and collagen assays were also conducted to study corresponding leaflet microstructures and the biochemical properties of the tissues. Results showed: (1) saphenous venous valve tissues possessed overall anisotropic properties. The tissues were stiffer in the circumferential direction than in the radial direction (p<0.01), and (2) saphenous venous valve tissues from the proximal end showed nonlinear isotropic mechanical properties, while those from the distal end showed nonlinear anisotropic mechanical properties. (3) Distal saphenous venous valve tissues appeared to be stiffer than proximal ones in the circumferential direction, p=0.04 (i.e., inter-valvular variability), and (4) the collagen concentration showed a decreasing trend from the proximal to the distal end. This study focuses on highly relevant animal (bovine) tissues to develop test protocols, establish biomechanical structure-function correlations, and to provide data critical to the design of clinical prosthetic venous valves. To the best of the author's knowledge, this is the first study reporting the biaxial mechanical properties of saphenous venous valve leaflet tissues and thus contributes toward refining our collective understanding of valvular tissue biomechanics. Cell/Tissue: saphenous vein valve leaflet valvular tisssues cardiovascular tissue
BioTester	2018	Prevalence Of Calcification In Human Femoropopliteal Arteries And Its Association With Demographics, Risk Factors, And Arterial Stiffness	Vasciular Biology	A. Kamenskiy, W. Poulson, S. Sim, A. Reilly, J. Luo, J. MacTaggart	University of Nebraska		Objective—Arterial calcification and stiffening increase the risk of reconstruction failure, amputation, and mortality in patients with peripheral arterial disease, but underlying mechanisms and prevalence are unclear. Approach and Results—Fresh human femoropopliteal arteries were obtained from n=431 tissue donors aged 13 to 82 years (mean age, 53±16 years) recording the in situ longitudinal prestretch. Arterial diameter, wall thickness, and opening angles were measured optically, and stiffness was assessed using planar biaxial extension and constitutive modeling. Histological features were determined using transverse and longitudinal Verhoeff–Van Gieson and Alizarin stains. Medial calcification was quantified using a 7-stage grading scale and was correlated with structural and mechanical properties and clinical characteristics. Almost half (46%) of the femoropopliteal arteries had identifiable medial calcification. Older arteries were more calcified, but small calcium deposits were observed in arteries as young as 18 years old. After controlling for age, positive correlations were observed between calcification, diabetes mellitus, dyslipidemia, and body mass index. Tobacco use demonstrated a negative correlation. Calcified arteries were larger in diameter but had smaller circumferential opening angles. They were also stiffer longitudinally and circumferentially and had thinner tunica media and external elastic lamina with more discontinuous elastic fibers. Conclusions—Although aging is the dominant risk factor for femoropopliteal artery calcification and stiffening, these processes seem to be linked and can begin at a young age. Calcification is associated with the presence of certain risk factors and with elastic fiber degradation, suggesting overlapping molecular pathways that require further investigation.
BioTester	2018	Comparison Of Corneal Biomechanics After Myopic Small-Incision Lenticule Extraction Compared To Lasik: An Ex Vivo Study	Clinical Opthamology	A.J. Kanellopoulos			Small-incision lenticule extraction (SMILE) procedure1,2 is a new method of intrastromal keratomileusis that seems to be a viable alternative for refractive correction, since numerous recent studies validate its safety and efficacy, even when compared to LASIK procedure the current laser refractive procedure of choice to treat myopia.3,4 SMILE is a new single femtosecond laser “flapless” procedure that involves the creation of an intrastromal lenticule between two photo disruption planes that is mechanically removed through a small corneal incision tunnel of 2–3 mm diameter typically created superotemporally.5 On the other hand, LASIK procedure involves two distinct steps: the creation of the flap (corresponding to a lamellar cut typically 110–140 µm within the cornea) by either a mechanical microkeratome or a femtosecond laser (usually operating at 1,056 nm),6 followed by the refractive part, the ablative removal of stromal tissue from the exposed bed under the lifted flap using an excimer laser (usually operating at 193 nm).7 The removal of corneal tissue by either procedure inevitably leads to reduction of corneal tensile strength.10,11 Both procedures alter corneal biomechanical properties that are thought to play an important role in the development of post-LASIK ectasia, but the nature of each procedure may produce different biomechanical effects. First, it is known that vertical cuts have more biomechanical impact than horizontal cuts. In SMILE significantly less anterior cornea is subjected to transverse separation, since side cut diameter is 2–3 mm (50°) compared to LASIK where flap diameter is of 300°, that is, 360° minus only the hinge. Additionally, it is also known that anterior stromal lamellae are stronger than posterior stromal lamellae, and the anterior 40% of the central corneal stroma constitutes the strongest region of the cornea, whereas the posterior 60% of the stroma is at least 50% weaker. In SMILE, since the anterior stroma remains uncut and the tissue is removed from deeper stromal layers than in LASIK, the strongest part of the stroma continues to contribute to the strength of the cornea postoperatively. In this study, we attempted to comparatively evaluate the changes in corneal biomechanical properties ex vivo following either SMILE (Visumax Laser; Carl Zeiss Meditec AG, Jena, Germany) or Femtosecond LASIK (FS200 & EX500 Lasers; Alcon Surgical, Fort Worth, TX, USA) for high or low myopic correction.
BioTester	2018	Dehiscence Of Patch Augmentation Of A Left-Sided Atrioventricular Valve Related To Strenuous Isometric Exercise: Case Report And Failure Analysis	The Journal of Thoracic and Cardiovascular Surgery	P.E. Hammer, C.W. Baird, P.J. del Nido, G.R. Marx	Boston Children's Hospital
BioTester	2018	Puncturing Of Lyophilized Tissue Engineered Vascular Matrices Enhances The Efficiency Of Their Recellularization	Acta Biomaterialia	A.A. Ksiazek, L. Frese, P.E. Dijkman, B. Sanders, S. E. Motta, B. Weber, S.P. Hoerstrup	Eindhoven		Data on in vitro engineered “off the shelf” matrices support the concept of endogenous cellular repopulation driving the graft’s remodeling via immune-mediated response. This seems important to further accelerate the cell reconstitution and may play a crucial role when mononuclear cells are used. Nevertheless, studies on decellularized xenogeneic grafts showed only limited host cell repopulation post-implantation. This study aims at a systematic comparison of reseeding methods (dripping, injection, bathing in a cell suspension and combined puncturing-dripping method) to define the most efficient technique enhancing recellularization of tissue engineered vascular matrices (patches, vessels, small diameter and standard size valves) prior implantation. The constructs were analyzed histologically, biochemically and biomechanically. Various preconditioning treatments (wet, lyophilized and air-dried) combined with reseeding methods demonstrated the highest cell loading efficiency, despite applied crimping and flow stress, of lyophilization followed by puncturing-dripping technique. This novel seeding method allows for an efficient, time-saving graft reseeding that can be used within a one-step cardiovascular clinical intervention.
BioTester	2018	Biomechanical Evaluation Of A Personalized External Aortic Root Support Applied In The Ross Procedure	Journal of the Mechanical Behavior of Biomedical Materials	J. Vastmans, H. Fehervary, P. Verbrugghe, T. Verbelen, E. Vanderveken, J. Vander Sloten, T. Treasure, F. Rega, N. Famaey	Leuven		A commonly heard concern in the Ross procedure, where a diseased aortic valve is replaced by the patient's own pulmonary valve, is the possibility of pulmonary autograft dilatation. We performed a biomechanical investigation of the use of a personalized external aortic root support or exostent as a possibility for supporting the autograft. In ten sheep a short length of pulmonary artery was interposed in the descending aorta, serving as a simplified version of the Ross procedure. In seven of these cases, the autograft was supported by an external mesh or so-called exostent. Three sheep served as control, of which one was excluded from the mechanical testing. The sheep were sacrificed six months after the procedure. Samples of the relevant tissues were obtained for subsequent mechanical testing: normal aorta, normal pulmonary artery, aorta with exostent, pulmonary artery with exostent, and pulmonary artery in aortic position for six months. After mechanical testing, the material parameters of the Gasser-Ogden-Holzapfel model were determined for the different tissue types. Stress-strain curves of the different tissue types show significantly different mechanical behavior. At baseline, stress-strain curves of the pulmonary artery are lower than aortic stress-strain curves, but at the strain levels at which the collagen fibers are recruited, the pulmonary artery behaves stiffer than the aorta. After being in aortic position for six months, the pulmonary artery tends towards aorta-like behavior, indicating that growth and remodeling processes have taken place. When adding an exostent around the pulmonary autograft, the mechanical behavior of the composite artery (exostent + artery) differs from the artery alone, the non-linearity being more evident in the former.
BioTester	2018	Polydopamine As Sizing On Carbon Fiber Surfaces For Enhancement Of Epoxy Laminated Composites	Composites Part A: Applied Science and Manufacturing	W. Han, H-P. Zhang, J. Tavakoli, J. Campbell, Y. Tang	Flinders University		Carbon fiber reinforced polymer (CFRP) laminate normally has plastic dominant crack propagation behavior, inducing potential insecurity in the safety and reliability of structures in practical applications. In this study, we report a simple process to increase the stability of crack growth by using polydopamine (PDA) as sizing on the surface of carbon fiber (CF) fabric. The crack propagation behavior changes from a saw-tooth-shaped curve in neat CFRP laminate to a relatively smooth trending curve in PDA coated CFRP laminate with increased Mode I interlaminar fracture toughness. Enhanced impact strength and interlaminar shear strength of PDA coated CFRP laminates is also observed. A single fiber pull-out experiment and morphological study reveal that, with PDA coating on CF fabrics, cracks tend to fracture through the epoxy matrix rather than between fiber and matrix interfaces. The use of PDA as sizing on the CF contributes to improving the load transfer between the CF and the polymer matrix by enhancing the interfaces between the epoxy and the CF, increasing the friction of the fractured interface, reducing unstable crack growth, and thereby enhancing interfacial fracture toughness and impact performance.
BioTester	2018	Comparison Of In Vivo Vs. Ex Situ Obtained Material Properties Of Sheep Common Carotid Artery	Medical Engineering & Physics	M. Smoljkic, P. Verbrugghe, M. Larsson, E. Widman, H. Fehervary, J. D'hooge, J. Vander Sloten, N. Famaey	Leuven		Patient-specific biomechanical modelling can improve preoperative surgical planning. This requires patient-specific geometry as well as patient-specific material properties as input. The latter are, however, still quite challenging to estimate in vivo. This study focuses on the estimation of the mechanical properties of the arterial wall. Firstly, in vivo pressure, diameter and thickness of the arterial wall were acquired for sheep common carotid arteries. Next, the animals were sacrificed and the tissue was stored for mechanical testing. Planar biaxial tests were performed to obtain experimental stress-stretch curves. Finally, parameters for the hyperelastic Mooney–Rivlin and Gasser–Ogden–Holzapfel (GOH) material model were estimated based on the in vivo obtained pressure-diameter data as well as on the ex situ experimental stress-stretch curves. Both material models were able to capture the in vivo behaviour of the tissue. However, in the ex situ case only the GOH model provided satisfactory results. When comparing different fitting approaches, in vivo vs. ex situ, each of them showed its own advantages and disadvantages. The in vivo approach estimates the properties of the tissue in its physiological state while the ex situ approach allows to apply different loadings to properly capture the anisotropy of the tissue. Both of them could be further enhanced by improving the estimation of the stress-free state, i.e. by adding residual circumferential stresses in vivo and by accounting for the flattening effect of the tested samples ex vivo.
BioTester	2018	Soy Protein/Cellulose Nanofiber Scaffolds Mimicking Skin Extracellular Matrix For Enhanced Wound Healing	Advanced Healthcare Materials	S. Ahn, C.O. Chantre, A.R. Gannon, J.U. Lind, P.H. Campbell, T. Grevesse, B.B. O'Conner, K.K. Parker	Harvard University		Historically, soy protein and extracts have been used extensively in foods due to their high protein and mineral content. More recently, soy protein has received attention for a variety of its potential health benefits, including enhanced skin regeneration. It has been reported that soy protein possesses bioactive molecules similar to extracellular matrix (ECM) proteins and estrogen. In wound healing, oral and topical soy has been heralded as a safe and cost‐effective alternative to animal protein and endogenous estrogen. However, engineering soy protein‐based fibrous dressings, while recapitulating ECM microenvironment and maintaining a moist environment, remains a challenge. Here, the development of an entirely plant‐based nanofibrous dressing comprised of cellulose acetate (CA) and soy protein hydrolysate (SPH) using rotary jet spinning is described. The spun nanofibers successfully mimic physicochemical properties of the native skin ECM and exhibit a high water retaining capability. In vitro, CA/SPH nanofibers promote fibroblast proliferation, migration, infiltration, and integrin β1 expression. In vivo, CA/SPH scaffolds accelerate re‐epithelialization and epidermal thinning as well as reduce scar formation and collagen anisotropy in a similar fashion to other fibrous scaffolds, but without the use of animal proteins or synthetic polymers. These results affirm the potential of CA/SPH nanofibers as a novel wound dressing.
BioTester	2018	New Findings Confirm The Viscoelastic Behaviour Of The Inter-Lamellar Matrix Of The Disc Annulus Fibrosus In Radial And Circumferential Directions Of Loading	Acta Biomaterialia	J. Tavakoli, J.J. Costi	Flinders University		While few studies have improved our understanding of composition and organization of elastic fibres in the inter-lamellar matrix (ILM), its clinical relevance is not fully understood. Moreover, no studies have measured the direct tensile and shear failure and viscoelastic properties of the ILM. Therefore, the aim of this study was, for the first time, to measure the viscoelastic and failure properties of the ILM in both the tension and shear directions of loading. Using an ovine model, isolated ILM samples were stretched to 40% of their initial length at three strain rates of 0.1%s−1 (slow), 1%s−1 (medium) and 10%s−1 (fast) and a ramp test to failure was performed at a strain rate of 10%s−1. The findings from this study identified that the stiffness of the ILM was significantly larger at faster strain rates, and energy absorption significantly smaller, compared to slower strain rates, and the viscoelastic and failure properties were not significantly different under tension and shear loading. We found a strain rate dependent response of the ILM during dynamic loading, particularly at the fastest rate. The ILM demonstrated a significantly higher capability for energy absorption at slow strain rates compared to medium and fast strain rates. A significant increase in modulus was found in both loading directions and all strain rates, having a trend of larger modulus in tension and at faster strain rates. The finding of no significant difference in failure properties in both loading directions, was consistent with our previous ultra-structural studies that revealed a well-organized (±45°) elastic fibre orientation in the ILM. The results from this study can be used to develop and validate finite element models of the AF at the tissue scale, as well as providing new strategies for fabricating tissue engineered scaffolds.
BioTester	2018	3d Printed, Controlled Release, Tritherapeutic Tablet Matrix For Advanced Anti-Hiv-1 Drug Delivery	European Journal of Pharmaceutics and Biopharmaceutics	M. Siyawamwaya, L.C. du Toit, P. Kumar, Y.E. Choonara, P.P.P.D Kondiah, V. Pillay	Witwatersrand		A 3D-Bioplotter® was employed to 3D print (3DP) a humic acid-polyquaternium 10 (HA-PQ10) controlled release fixed dose combination (FDC) tablet comprising of the anti-HIV-1 drugs, efavirenz (EFV), tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC).MethodsChemical interactions, surface morphology and mechanical strength of the FDC were ascertained. In vitro drug release studies were conducted in biorelevant media followed by in vivo study in the large white pigs, in comparison with a market formulation, Atripla®. In vitro-in vivo correlation of results was undertaken.EFV, TDF and FTC were successfully entrapped in the 24-layered rectangular prism-shaped 3DP FDC with a loading of ~12.5?mg/6.3?mg/4?mg of EFV/TDF/FTC respectively per printed layer. Hydrogen bonding between the EFV/TDF/FTC and HA-PQ10 was detected which was indicative of possible drug solubility enhancement. The overall surface of the tablet exhibited a fibrilla structure and the 90° inner pattern was determined to be optimal for 3DP of the FDC. In vitro and in vivo drug release profiles from the 3DP FDC demonstrated that intestinal-targeted and controlled drug release was achieved.A 3DP FDC was successfully manufactured with the aid of a 3D-Bioplotter in a single step process. The versatile HA-PQ10 entrapped all drugs and achieved an enhanced relative bioavailability of EFV, TDF, and FTC compared to the market formulation for potentially enhanced HIV treatment.
BioTester	2018	Computational Modeling Guides Tissue-Engineered Heart Valve Design For Long-Term In Vivo Performance In A Translational Sheep Model	Science Translational Medicine	M.Y. Emmert, B.A. Schmitt, S. Loerakker, B. Sanders, H. Spriestersbach,, E.S. Fioretta, L. Bruder, K. Brakmann, S.E. Motta, V. Lintas, P.E. Dijkman, L. Frese, F. Berger, F.P.T. Baaijens, S.P. Hoerstrup	Eindhoven University of Technology		Valvular heart disease is a major cause of morbidity and mortality worldwide. Current heart valve prostheses have considerable clinical limitations due to their artificial, nonliving nature without regenerative capacity. To overcome these limitations, heart valve tissue engineering (TE) aiming to develop living, native-like heart valves with self-repair, remodeling, and regeneration capacity has been suggested as next-generation technology. A major roadblock to clinically relevant, safe, and robust TE solutions has been the high complexity and variability inherent to bioengineering approaches that rely on cell-driven tissue remodeling. For heart valve TE, this has limited long-term performance in vivo because of uncontrolled tissue remodeling phenomena, such as valve leaflet shortening, which often translates into valve failure regardless of the bioengineering methodology used to develop the implant. We tested the hypothesis that integration of a computationally inspired heart valve design into our TE methodologies could guide tissue remodeling toward long-term functionality in tissue-engineered heart valves (TEHVs). In a clinically and regulatory relevant sheep model, TEHVs implanted as pulmonary valve replacements using minimally invasive techniques were monitored for 1 year via multimodal in vivo imaging and comprehensive tissue remodeling assessments. TEHVs exhibited good preserved long-term in vivo performance and remodeling comparable to native heart valves, as predicted by and consistent with computational modeling. TEHV failure could be predicted for nonphysiological pressure loading. Beyond previous studies, this work suggests the relevance of an integrated in silico, in vitro, and in vivo bioengineering approach as a basis for the safe and efficient clinical translation of TEHVs.
BioTester	2018	Pressure-Induced End-Plate Fracture In The Porcine Spine: Is The Annulus Fibrosus Susceptible To Damage?	European Spine Journal	C.R. Snow, M.Harvey-Burgess, B. Laird, S.H.M Brown, D.E. Gregory	Wilfrid Laurier University		To determine if the mechanical properties of the annulus fibrosus (AF) are altered following end-plate fracture. Vertebral fractures, particularly those in the growth plate, are relatively common among adolescents. What is unclear is whether or not these fractures are also associated with concomitant damage to the intervertebral disc (IVD), in particular the AF.
BioTester	2018	Dual Electrospun Supramolecular Polymer Systems For Selective Cell Migration	Macromolecular Bioscience Journal	S.H. Thakker, A. Di Luca, S. Zaccaria, F.P.T Baaijens, C.V.C. Bouten, P.Y.W. Dankers	TU Eindhoven		Dual electrospinning can be used to make multifunctional scaffolds for regenerative medicine applications. Here, two supramolecular polymers with different material properties are electrospun simultaneously to create a multifibrous mesh. Bisurea (BU)‐based polycaprolactone, an elastomer providing strength to the mesh, and ureido‐pyrimidinone (UPy) modified poly(ethylene glycol) (PEG), a hydrogelator, introducing the capacity to deliver compounds upon swelling. The dual spun scaffolds are modularly tuned by mixing UPyPEG hydrogelators with different polymer lengths, to control swelling of the hydrogel fiber, while maintaining the mechanical properties of the scaffold. Stromal cell derived factor 1 alpha (SDF1α) peptides are embedded in the UPyPEG fibers. The swelling and erosion of UPyPEG increase void spaces and released the SDF1α peptide. The functionalized scaffolds demonstrate preferential lymphocyte recruitment proposed to be created by a gradient formed by the released SDF1α peptide. This delivery approach offers the potential to develop multifibrous scaffolds with various functions.
BioTester	2018	Regional Mechanical And Biochemical Properties Of The Porcine Cortical Meninges	Acta Biomaterialia	D.R. Walsh, A.M. Ross, S. Malijauskaite, B.D. Flanagan, D.T. Newport, K.D. McGourty, J.J.E. Mulvihill	Limerick		The meninges are pivotal in protecting the brain against traumatic brain injury (TBI), an ongoing issue in most mainstream sports. Improved understanding of TBI biomechanics and pathophysiology is desirable to improve preventative measures, such as protective helmets, and advance our TBI diagnostic/prognostic capabilities. This study mechanically characterised the porcine meninges by performing uniaxial tensile testing on the dura mater (DM) tissue adjacent to the frontal, parietal, temporal, and occipital lobes of the cerebellum and superior sagittal sinus region of the DM. Mechanical characterisation revealed a significantly higher elastic modulus for the superior sagittal sinus region when compared to other regions in the DM. The superior sagittal sinus and parietal regions of the DM also displayed local mechanical anisotropy. Further, fatigue was noted in the DM following ten preconditioning cycles, which could have important implications in the context of repetitive TBI. To further understand differences in regional mechanical properties, regional variations in protein content (collagen I, collagen III, fibronectin and elastin) were examined by immunoblot analysis. The superior sagittal sinus was found to have significantly higher collagen I, elastin, and fibronectin content. The frontal region was also identified to have significantly higher collagen I and fibronectin content while the temporal region had increased elastin and fibronectin content. Regional differences in the mechanical and biochemical properties along with regional tissue thickness differences within the DM reveal that the tissue is a non-homogeneous structure. In particular, the potentially influential role of the superior sagittal sinus in TBI biomechanics warrants further investigation.
BioTester	2018	New Insights Into The Viscoelastic And Failure Mechanical Properties Of The Elastic Fiber Network Of The Inter-Lamellar Matrix In The Annulus Fibrosus Of The Disc	Acta Biomaterialia	J. Tavakoli, J.J. Costi	Flinders University		The mechanical role of elastic fibers in the inter-lamellar matrix (ILM) is unknown; however, it has been suggested that they play a role in providing structural integrity to the annulus fibrosus (AF). Therefore, the aim of this study was to measure the viscoelastic and failure properties of the elastic fiber network in the ILM of ovine discs under both tension and shear directions of loading. Utilizing a technique, isolated elastic fibers within the ILM from ovine discs were stretched to 40% of their initial length at three strain rates of 0.1% s−1 (slow), 1% s−1 (medium) and 10% s−1 (fast), followed by a ramp test to failure at 10% s−1. A significant strain-rate dependent response was found, particularly at the fastest rate for phase angle and normalized stiffness (p < 0.001). The elastic fibers in the ILM demonstrated a significantly higher capability for energy absorption at slow compared to medium and fast strain rates (p < 0.001). These finding suggests that the elastic fiber network of the ILM exhibits nonlinear elastic behavior. When tested to failure, a significantly higher normalized failure force was found in tension compared to shear loading (p = 0.011), which is consistent with the orthotropic structure of elastic fibers in the ILM. The results of this study confirmed the mechanical contribution of the elastic fiber network to the ILM and the structural integrity of the AF. This research serves as a foundation for future studies to investigate the relationship between degeneration and ILM mechanical properties.
BioTester	2018	Initial Scaffold Thickness Affects The Emergence Of A Geometrical And Mechanical Equilibrium In Engineered Cardiovascular Tissues	Journal of the Royal Society Interface	M.A.J. van Kelle, P.J.A. Oomen, W.J.T. Janssen-van den Broek, R.G.P. Lopata, S. Loerakker, C.V.C. Bouten	TU Eindhoven		In situ cardiovascular tissue-engineering can potentially address the shortcomings of the current replacement therapies, in particular, their inability to grow and remodel. In native tissues, it is widely accepted that physiological growth and remodelling occur to maintain a homeostatic mechanical state to conserve its function, regardless of changes in the mechanical environment. A similar homeostatic state should be reached for tissue-engineered (TE) prostheses to ensure proper functioning. For in situ tissue-engineering approaches obtaining such a state greatly relies on the initial scaffold design parameters. In this study, it is investigated if the simple scaffold design parameter initial thickness, influences the emergence of a mechanical and geometrical equilibrium state in in vitro TE constructs, which resemble thin cardiovascular tissues such as heart valves and arteries. Towards this end, two sample groups with different initial thicknesses of myofibroblast-seeded polycaprolactone-bisurea constructs were cultured for three weeks under dynamic loading conditions, while tracking geometrical and mechanical changes temporally using non-destructive ultrasound imaging. A mechanical equilibrium was reached in both groups, although at different magnitudes of the investigated mechanical quantities. Interestingly, a geometrically stable state was only established in the thicker constructs, while the thinner constructs’ length continuously increased. This demonstrates that reaching geometrical and mechanical stability in TE constructs is highly dependent on functional scaffold design.
BioTester	2018	Development Of An Improved Parameter Fitting Method For Planar Biaxial Testing Using Rakes	Numerical Methods in Biomedical Engineering	H. Fehervary, J. Vander Sloten, N. Famaey	KU Leuven		A correct estimation of the material parameters from a planar biaxial test is crucial since they will affect the outcome of the finite element model in which they are used. In a virtual planar biaxial experiment, a difference can be noticed in the stress calculated from the force measured experimentally at the rakes and the actual stress at the center of the sample. As a consequence, a classic parameter fitting does not result in a correct estimation of the material parameters. This difference is caused by the boundary conditions of the set‐up and is among others dependent on the sample material. To overcome this problem, a new parameter fitting procedure is proposed that takes this difference into account by calculating a finite element‐based correction vector.
BioTester	2018	The Biomechanics Of The Inter-Lamellar Matrix And The Lamellae During Progression To Lumbar Disc Herniation: Which Is The Weakest Structure?	Annals of Biomedical Engineering	J. Tavakoli, D.B. Amin, B.J.C. Freeman, J.J. Costi	Flinders University		While microstructural observations have improved our understanding of possible pathways of herniation progression, no studies have measured the mechanical failure properties of the inter-lamellar matrix (ILM), nor of the adjacent lamellae during progression to herniation. The aim of this study was to employ multiscale, biomechanical and microstructural techniques to evaluate the effects of progressive induced herniation on the ILM and lamellae in control, pre-herniated and herniated discs (N = 7), using 2 year-old ovine spines. Pre-herniated and herniated (experimental) groups were subjected to macroscopic compression while held in flexion (13°), before micro-mechanical testing. Micro-tensile testing of the ILM and the lamella from anterior and posterolateral regions was performed in radial and circumferential directions to measure failure stress, modulus, and toughness in all three groups. The failure stress of the ILM was significantly lower for both experimental groups compared to control in each of radial and circumferential loading directions in the posterolateral region (p < 0.032). Within each experimental group in both loading directions, the ILM failure stress was significantly lower by 36% (pre-herniation), and 59% (herniation), compared to the lamella (p < 0.029). In pre-herniated compared to control discs, microstructural imaging revealed significant tissue stretching and change in orientation (p < 0.003), resulting in a loss of distinction between respective lamellae and ILM boundaries.
BioTester	2018	Mechanically Robust Cryogels With Injectability And Bioprinting Supportability For Adipose Tissue Engineering	Acta Biomaterialia	D. Qi, S. Wu, M.A.Kuss, W.Shi, S. Chung, P.T. Deegan, A. Kamenskiy, Y. He, B. Duan	Nebraska		Bioengineered adipose tissues have gained increased interest as a promising alternative to autologous tissue flaps and synthetic adipose fillers for soft tissue augmentation and defect reconstruction in clinic. Although many scaffolding materials and biofabrication methods have been investigated for adipose tissue engineering in the last decades, there are still challenges to recapitulate the appropriate adipose tissue microenvironment, maintain volume stability, and induce vascularization to achieve long-term function and integration. In the present research, we fabricated cryogels consisting of methacrylated gelatin, methacrylated hyaluronic acid, and 4arm poly(ethylene glycol) acrylate (PEG-4A) by using cryopolymerization. The cryogels were repeatedly injectable and stretchable, and the addition of PEG-4A improved the robustness and mechanical properties. The cryogels supported human adipose progenitor cell (HWA) and adipose derived mesenchymal stromal cell adhesion, proliferation, and adipogenic differentiation and maturation, regardless of the addition of PEG-4A. The HWA laden cryogels facilitated the co-culture of human umbilical vein endothelial cells (HUVEC) and capillary-like network formation, which in return also promoted adipogenesis. We further combined cryogels with 3D bioprinting to generate handleable adipose constructs with clinically relevant size. 3D bioprinting enabled the deposition of multiple bioinks onto the cryogels. The bioprinted flap-like constructs had an integrated structure without delamination and supported vascularization.
BioTester	2018	Biomechanical Characterization Of Human Dura Mater	JMBBM	D. DeKegel, J. Vastmans, H. Fehervary, B. Depreitere, J. Vander Sloten, N. Famaey	KU Leuven		A reliable computational model of the human head is necessary for better understanding of the physical mechanisms of traumatic brain injury (TBI), car-crash investigation, development of protective head gear and advancement of dural replacement materials. The performance and biofidelity of these models depend largely on the material description of the different structures present in the head. One of these structures is the dura mater, the protective layer around the brain.
BioTester	2018	3d Printed, Pva–Paa Hydrogel Loaded-Polycaprolactone Scaffold For The Delivery Of Hydrophilic In-Situ Formed Sodium Indomethacin	Materials	M. Govender, S. Indermum, P. Kumar, Y.E. Choonara, V. Pillay	Wits		3D printed polycaprolactone (PCL)-blended scaffolds have been designed, prepared, and evaluated in vitro in this study prior to the incorporation of a polyvinyl alcohol–polyacrylic acid (PVA–PAA) hydrogel for the delivery of in situ-formed sodium indomethacin. The prepared PCL–PVA–PAA scaffold is proposed as a potential structural support system for load-bearing tissue damage where inflammation is prevalent. Uniaxial strain testing of the PCL-blended scaffolds were undertaken to determine the scaffold’s resistance to strain in addition to its thermal, structural, and porosimetric properties. The viscoelastic properties of the incorporated PVA–PAA hydrogel has also been determined, as well as the drug release profile of the PCL–PVA–PAA scaffold. Results of these analyses noted the structural strength, thermal stability, and porosimetric properties of the scaffold, as well as the ability of the PCL–PVA–PAA scaffold to deliver sodium indomethacin in simulated physiological conditions of pH and temperature. The results of this study therefore highlight the successful design, fabrication, and in vitro evaluation of a 3D printed polymeric strain-resistant supportive platform for the delivery of sodium indomethacin.
BioTester	2018	A Transverse Isotropic Constitutive Model For The Aortic Valve Tissue Incorporating Rate-Dependency And Fibre Dispersion: Application To Biaxial Deformation	JMBBM	A. Anssari-Benam, Y-T. Tseng, A. Bucchi			This paper presents a continuum-based transverse isotropic model incorporating rate-dependency and fibre dispersion, applied to the planar biaxial deformation of aortic valve (AV) specimens under various stretch rates. The rate dependency of the mechanical behaviour of the AV tissue under biaxial deformation, the (pseudo-) invariants of the right Cauchy-Green deformation-rate tensor associated with fibre dispersion, and a new fibre orientation density function motivated by fibre kinematics are presented for the first time. It is shown that the model captures the experimentally observed deformation of the specimens, and characterises a shear-thinning behaviour associated with the dissipative (viscous) kinematics of the matrix and the fibres. The application of the model for predicting the deformation behaviour of the AV under physiological rates is illustrated and an example of the predicted curves is presented. While the development of the model was principally motivated by the AV biomechanics requisites, the comprehensive theoretical approach employed in the study renders the model suitable for application to other fibrous soft tissues that possess similar rate-dependent and structural attributes.
BioTester	2018	Decoupling The Effect Of Shear Stress And Stretch On Tissue Growth And Remodeling In A Vascular Graft	Tissue Engineering Part C: Methods	E.E. van Haaften, et. al.	TU Eindhoven		The success of cardiovascular tissue engineering (TE) strategies largely depends on the mechanical environment in which cells develop a neotissue through growth and remodeling processes. This mechanical environment is defined by the local scaffold architecture to which cells adhere, that is, the microenvironment, and by external mechanical cues to which cells respond, that is, hemodynamic loading. The hemodynamic environment of early developing blood vessels consists of both shear stress (due to blood flow) and circumferential stretch (due to blood pressure). Experimental platforms that recapitulate this mechanical environment in a controlled and tunable manner are thus critical for investigating cardiovascular TE. In traditional perfusion bioreactors, however, shear stress and stretch are coupled, hampering a clear delineation of their effects on cell and tissue response. In this study, we uniquely designed a bioreactor that independently combines these two types of mechanical cues in eight parallel vascular grafts. The system is computationally and experimentally validated, through finite element analysis and culture of tissue constructs, respectively, to distinguish various levels of shear stress (up to 5 Pa) and cyclic stretch (up to 1.10). To illustrate the usefulness of the system, we investigated the relative contribution of cyclic stretch (1.05 at 0.5 Hz) and shear stress (1 Pa) to tissue development. Both types of hemodynamic loading contributed to cell alignment, but the contribution of shear stress overruled stretch-induced cell proliferation and matrix (i.e., collagen and glycosaminoglycan) production. At a macroscopic level, cyclic stretching led to the most linear stress–stretch response, which was not related to the presence of shear stress. In conclusion, we have developed a bioreactor that is particularly suited to further unravel the interplay between hemodynamics and in situ TE processes. Using the new system, this work highlights the importance of hemodynamic loading to the study of developing vascular tissues.
BioTester	2018	An Investigation Of The Anisotropic Mechanical Properties And Anatomical Structure Of Porcine Atrioventricular Heart Valves	Journal of the Mechanical Behavior of Biomedical Materials	S. Jett, D. Laurence, R. Kunkel, A. R. Babu, K. Kramer, R. Baumwart, R. Towner, Y. Wu, C-H. Lee	University of Oklahoma		Valvular heart diseases are complex disorders, varying in pathophysiological mechanism and affected valve components. Understanding the effects of these diseases on valve functionality requires a thorough characterization of the mechanics and structure of the healthy heart valves. In this study, we performed biaxial mechanical experiments with extensive testing protocols to examine the mechanical behaviors of the mitral valve and tricuspid valve leaflets. We also investigated the effect of loading rate, testing temperatures, species (porcine versus ovine hearts), and age (juvenile vs adult ovine hearts) on the mechanical responses of the leaflet tissues. In addition, we evaluated the structure of chordae tendineae within each valve and performed histological analysis on each atrioventricular leaflet. We found all tissues displayed a characteristic nonlinear anisotropicmechanical response, with radial stretches on average 30.7% higher than circumferential stretches under equibiaxial physiological loading. Tissue mechanical responses showed consistent mechanical stiffening in response to increased loading rate and minor temperature dependence in all five atrioventricular heart valve leaflets. Moreover, our anatomical study revealed similar chordae quantities in the porcine mitral (30.5 ± 1.43 chords) and tricuspid valves (35.3 ± 2.45 chords) but significantly more chordae in the porcine than the ovine valves (p < 0.010). Our histological analyses quantified the relative thicknesses of the four distinct morphological layers in each leaflet. This study provides a comprehensive database of the mechanics and structure of the atrioventricular valves, which will be beneficial to development of subject-specific atrioventricular valve constitutive models and toward multi-scale biomechanical investigations of heart valve function to improve valvular disease treatments.
BioTester	2018	Biaxial Mechanical Data of Porcine Atrioventricular Valve Leaflets	Data in Brief	S. Jett, D. Laurence, R. Kunkel, A. Babu, K. Kramer, R. Baumwart, R. Towner, Y. Wu, C. H. Lee	University of Oklahoma		This dataset contains the anisotropic tissue responses of porcine atrioventricular valve leaflets to force-controlled biaxial mechanical testing. The set includes the first Piola-Kirchhoff Stress and the specimen stretches (λ) in both circumferential and radial tissue directions (C and R, respectively) for the mitral valve anterior and posterior leaflets (MVAL and MVPL), and the tricuspid valve anterior, posterior, and septal leaflets (TVAL, TVPL, and TVSL) from six porcine hearts at five separate force-controlled biaxial loading protocols. This dataset is associated with a companion journal article, which can be consulted for further information about the methodology, results, and discussion of this biaxial mechanical testing (Jett et al., in press) [1].
BioTester	2018	Biaxial Mechanical Characterization of Atrioventricular Heart Valves	Journal of Visualized Experiments	C. Ross, D. Laurence, Y. Wu, C. H. Lee	University of Oklahoma		Extensive biaxial mechanical testing of the atrioventricular heart valve leaflets can be utilized to derive optimal parameters used in constitutive models, which provide a mathematical representation of the mechanical function of those structures. This presented biaxial mechanical testing protocol involves (i) tissue acquisition, (ii) the preparation of tissue specimens, (iii) biaxial mechanical testing, and (iv) postprocessing of the acquired data. First, tissue acquisition requires obtaining porcine or ovine hearts from a local Food and Drug Administration-approved abattoir for later dissection to retrieve the valve leaflets. Second, tissue preparation requires using tissue specimen cutters on the leaflet tissue to extract a clear zone for testing. Third, biaxial mechanical testing of the leaflet specimen requires the use of a commercial biaxial mechanical tester, which consists of force-controlled, displacement-controlled, and stress-relaxation testing protocols to characterize the leaflet tissue's mechanical properties. Finally, post-processing requires the use of data image correlation techniques and force and displacement readings to summarize the tissue's mechanical behaviors in response to external loading. In general, results from biaxial testing demonstrate that the leaflet tissues yield a nonlinear, anisotropic mechanical response. The presented biaxial testing procedure is advantageous to other methods since the method presented here allows for a more comprehensive characterization of the valve leaflet tissue under one unified testing scheme, as opposed to separate testing protocols on different tissue specimens. The proposed testing method has its limitations in that shear stress is potentially present in the tissue sample. However, any potential shear is presumed negligible.
BioTester	2019	An Investigation Of Regional Variations In The Biaxial Mechanical Properties And Stress Relaxation Behaviors Of Porcine Atrioventricular Heart Valve Leaflets	Journal of Biomechanics	D. Laurence, C. Ross, S. Jett, C. Johns, A. Echols, R. Baumwart, R. Towner, J. Liao, P. Bajona, Y. Wu, C. H. Lee	University of Oklahoma		The facilitation of proper blood flow through the heart depends on proper function of heart valve components, and alterations to any component can lead to heart disease or failure. Comprehension of these valvular diseases is reliant on thorough characterization of healthy heart valve structures for use in computational models. Previously, computational models have treated these leaflet structures as a structurally and mechanically homogenous material, which may not be an accurate description of leaflet mechanical response. In this study, we aimed to characterize the mechanics of the heart valve leaflet as a structurally heterogenous material. Specifically, porcine mitral valve and tricuspid valve anterior leaflets were sectioned into six regions and biaxial mechanical tests with various loading ratios and stress-relaxation test were performed on each regional tissue sample. Three main findings from this study were summarized as follows: (i) the central regions of the leaflet had a more anisotropic nature than edge regions, (ii) the mitral valve anterior leaflet was more extensible in regions closer to the annulus, and (iii) there was variance in the stress-relaxation behavior among all six regions, with mitral valve leaflet tissue regions exhibiting a greater decay than the tricuspid valve regions. This study presents a novel investigation of the regional variations in the heart valve biomechanics that has not been comprehensively examined. Our results thus allow for a refinement of computational models for more accurately predicting diseased or surgically-intervened condition, where tissue heterogeneity plays an essential role in the heart valve function.
BioTester	2019	Strain Effects On Collagen Proteolysis In Heart Valve Tissues	Mechanics of Time-Depedent Materials	K. Barbour, H. Y. S. Huang	North Carolina State University		Collagen is at the heart of any and all questions concerning semilunar valvular leaflet composition, structure, and function. Whether during development, physiological homeostasis, or pathological degeneration, it is the structural-mechanical state of the heart valve leaflet collagen network that ultimately confers valvular function, and the difference between health and disease. In the current study, the effects of physiologically relevant strain states on collagen catabolism are investigated in porcine aortic and pulmonary valve leaflets. Application of bacterial collagenase to the tissues which acts to simulate collagen degradation by endogenous matrix metalloproteinases, biaxial stress relaxation, and histology are all used to serve as measures of functional and compositional collagen catabolism. Current stress-relaxation results are used in conjunction with previous equibiaxial testing to confirm that a mechanism exists to prevent collagen catabolism when stretched at physiologically relevant strain states. Collectively, these in vitro results indicate that biaxial strain states are capable of impacting the susceptibility of valvular collagens to catabolism, and that at physiological strain states, a protective mechanism exists to effectively block collagen catabolism. The results of the study will be broadly applicable to clarify the roles of tissue microarchitecture and load transmission in a variety of other developmental, homeostatic, or pathogenic tissue processes such as tumor growth, embryogenesis, thrombi formation, and atherogenesis.
BioTester	2019	Comparison Of Biomechanical Properties And Microstructure Of Trabeculae Carneae, Papillary Muscles, And Myocardium In The Human Heart	Journal of Biomechanical Engineering	F. Fatemifar, M. D. Feldman, M. Oglesby, H. C. Han	University of Texas		Trabeculae carneae account for a significant portion of human ventricular mass, despite being considered embryologic remnants. Recent studies have found trabeculae hypertrophy and fibrosis in hypertrophied left ventricles with various pathological conditions. The objective of this study was to investigate the passive mechanical properties and microstructural characteristics of trabeculae carneae and papillary muscles compared to the myocardium in human hearts. Uniaxial tensile tests were performed on samples of trabeculae carneae and myocardium strips, while biaxial tensile tests were performed on samples of papillary muscles and myocardium sheets. The experimental data were fitted with a Fung-type strain energy function and material coefficients were determined. The secant moduli at given diastolic stress and strain levels were determined and compared among the tissues. Following the mechanical testing, histology examinations were performed to investigate the microstructural characteristics of the tissues. Our results demonstrated that the trabeculae carneae were significantly stiffer (Secant modulus SM2 = 80.06 ± 10.04 KPa) and had higher collagen content (16.10 ± 3.80%) than the myocardium (SM2 = 55.14 ± 20.49 KPa, collagen content = 10.06 ± 4.15%) in the left ventricle. The results of this study improve our understanding of the contribution of trabeculae carneae to left ventricular compliance and will be useful for building accurate computational models of the human heart.
BioTester	2019	Rate-Dependency Of The Mechanical Behavior Of Semilunar Heart Valves Under Biaxial Deformation	Acta Biomaterialia	A. Anssari-Benam, Y-T. Tseng, A. Bucchi	University of Portsmouth		This paper presents an experimental investigation and evidence of rate-dependency in the planar mechanical behaviour of semilunar heart valves. Samples of porcine aortic and pulmonary valves were subjected to biaxial deformations across 1000-fold stretch rate, ranging from λ̇=0.001 to 1 s−1. The experimental campaign encompassed protocols covering (i) tests on samples without preconditioning, (ii) preconditioning immediately followed by tensile tests; and (iii) tensile tests at different rates performed on the same preconditioned specimen. Our results indicate that under all employed loading protocols, heart valve samples exhibit a marked rate-dependency in their deformation behaviour. This rate-dependency is reflected in stress-stretch curves and the calculated ensuing gradients, where samples typically show stiffening with increased rate. These results underpin one conclusive outcome: the in-plane mechanical behaviour of semilunar valves is rate-dependent (p<0.05 for Cauchy stress levels ≥50 kPa). This outcome implies that the rate of deformation for characterising the mechanical behaviour of semilunar heart valves may not be chosen arbitrarily low, and models that incorporate rate-effects may be more appropriate for better capturing the mechanical behaviour of heart valves.
BioTester	2019	An Investigation Of Layer-Specific Tissue Biomechanics Of Porcine Atrioventricular Valve Anterior Leaflets	Acta Biomaterialia	K. Kramer, C. Ross, A. Babu, Y. Wu, R. Towner, A. Mir, H. M. Burkhart, G. A. Holzapfel, C. H. Lee	University of Oklahoma		The atrioventricular heart valves (AHVs) are composed of structurally complex and morphologically heterogeneous leaflets. The coaptation of these leaflets during the cardiac cycle facilitates unidirectional blood flow and prevents regurgitation. Valve regurgitation is treated preferably by surgical repair if possible or replacement based on the disease state of the valve tissue. A comprehensive understanding of valvular morphology and mechanical properties is crucial to refining computational models, serving as a patient-specific diagnostic and surgical tool for preoperative planning. Previous studies have modeled the stress distribution throughout the thickness of the heterogeneous leaflet, but validations with layer-specific biaxial mechanical experiments are missing. In this study, we sought to fill this gap in literature by investigating the impact of microstructure constituents on mechanical behavior throughout the thickness of the AHVs’ anterior leaflets. Porcine mitral valve anterior leaflets (MVAL) and tricuspid valve anterior leaflets (TVAL) were micro-dissected into 3 layers (atrialis/spongiosa, fibrosa, and ventricular) and 2 layers (atrialis/spongiosa and fibrosa/ventricularis), respectively, based on their relative distributions of extracellular matrix (ECM) components as quantified by histological analyses: collagen, elastin, and glycosaminoglycans. Our results suggest that: (i) the atrialis/spongiosa layer, for both valves, is the most extensible and anisotropic layer, (ii) the intact TVAL response is stiffer than the atrialis/spongiosa layer but more compliant than the fibrosa/ventricularis layer, and (iii) the MVAL fibrosa and ventricularis layers behave nearly isotropic. These novel findings emphasize the biomechanical variances throughout the AHV leaflets, and our results will be useful to better inform AHV computational models.
BioTester	2019	The Degradation And Performance Of Electrospun Supramolecular Vascular Scaffolds Examined Upon In Vitro Enzymatic Exposure	Acta Biomaterialia	E. E. van Haaften, R. Duijvelshoff, B. D. Ippela, S. H. M. Sontjens, M. H. C. J. van Houtem, H. M. Janssen, A. I. P. M. Smits, N. A. Kurniawan, P. Y. W. Dankers, C. V. C. Bouten	Eindhoven Institute of Technology		To maintain functionality during in situ regeneration of load-bearing tissues, a balance between the rates of implant degradation and neo-tissue formation is required. Strictly segmented thermoplastic elastomers with supramolecularly interacting bis-urea (BU) hard blocks are attractive biomaterials for vascular tissue engineering, as these polymers are soft, tough, and biodegradable. Moreover, these materials possess a sequence-controlled macromolecular structure, so their susceptibility to degradation is tunable by controlling the nature of the polymer backbone. It is unknown, however, how the implant’s functionality is affected by the degradation of the polymers it is composed of. We therefore examined the macro- and microscopic features as well as the mechanical performance of vascular scaffolds upon in vitro enzymatic degradation. Three candidate biomaterials (‘slow-degrading’ polycarbonate-BU (PC-BU), ‘intermediate-degrading’ polycarbonate-ester-BU (PC(e)-BU), and ‘fast-degrading’ polycaprolactone-ester-BU (PCL-BU)) were synthesized and electrospun into microporous scaffolds. The scaffolds were incubated in lipase and monitored for changes in physical, chemical, and mechanical properties. Remarkably, comparing PC-BU to PC(e)-BU, we observed that small changes in macromolecular structure led to significant differences in degradation kinetics. All three scaffold types degraded via surface erosion, which was accompanied by fiber swelling for PC-BU scaffolds, and some bulk degradation and a collapsing network for PCL-BU scaffolds. For the PC-BU and PC(e)-BU scaffolds this resulted in retention of mechanical properties, whereas for the PCL-BU scaffolds this resulted in stiffening. Our in vitro study demonstrates that vascular scaffolds, electrospun from sequence-controlled supramolecular materials with varying ester contents, not only display different susceptibilities to degradation, but also degrade via different mechanisms.
BioTester	2019	Mechanical Damage Characterization In Human Femoropopliteal Arteries Of Different Ages	Acta Biomaterialia	E. Anttila, D. Balzani, A. Desyatova, P. Deegan, J. MacTaggart, A. Kamenskiy	University of Nebraska-Lincoln		Endovascular treatment of Peripheral Arterial Disease (PAD) is notorious for high failure rates, and interaction between the arterial wall and the repair devices plays a significant role. Computational modeling can help improve clinical outcomes of these interventions, but it requires accurate inputs of elastic and damage characteristics of the femoropopliteal artery (FPA) which are currently not available. Fresh human FPAs from n = 104 tissue donors 14–80 years old were tested using planar biaxial extension to capture elastic and damage characteristics. Damage initiation stretches and stresses were determined for both longitudinal and circumferential directions, and their correlations with age and risk factors were assessed. Two and four-fiber-family invariant-based constitutive models augmented with damage functions were used to describe stress softening with accumulating damage. In FPAs younger than 50 years, damage began accumulating after 1.51 ± 0.13 and 1.49 ± 0.11 stretch, or 196 ± 110 kPa and 239 ± 79 kPa Cauchy stress in the longitudinal and circumferential directions, respectively. In FPAs older than 50 years, damage initiation stretches and stresses decreased to 1.27 ± 0.09 (106 ± 52 kPa) and 1.26 ± 0.09 (104 ± 59 kPa), respectively. Damage manifested primarily as tears at the internal and external elastic laminae and within the tunica media layer. Higher body mass index and presence of diabetes were associated with lower damage initiation stretches and higher stresses. The selected constitutive models were able to accurately portray the FPA behavior in both elastic and inelastic domains, and properties were derived for six age groups. Presented data can help improve fidelity of computational models simulating endovascular PAD repairs that involve arterial damage.
BioTester	2019	Cyclic Strain Affects Macrophage Cytokine Secretion And Extracellular Matrix Turnover In Electrospun Scaffolds	Tissue Engineering	V. Bonito, B. J. deKort, C. V. C. Bouten, A. I. P. M. Smits	University of Technology, Eindhoven		Controlling macrophage behavior has become a high-potential target strategy for regenerative therapies, such as in situtissue engineering (TE). In situ TE is an approach, in which acellular resorbable synthetic scaffolds are used, to induce endogenous tissue regeneration. However, little is known regarding the effect of the biomechanical environment on the macrophage response to a scaffold. Therefore, the aim of this study was to assess the effect of cyclic strains (0%, 8%, and 14% strain) on primary human macrophage polarization in electrospun scaffolds with two different fiber diameters in the micrometer range (4 μm or 13 μm). High strains led to a proinflammatory profile in terms of gene expression, expression of surface proteins, and cytokine secretion. These results were consistent for scaffolds with small and large fiber diameters, indicating that the effect of cyclic strain was not affected by the different scaffold microstructures. Notably, macrophages were identified as direct contributors of early secretion of extracellular matrix proteins, including elastin, which was deposited in a strain-dependent manner. These findings are instrumental for the rational design of scaffolds for in situ TE and underline that immunomodulatory scaffolds for biomechanically loaded applications should be mechanically tailored, for example, in terms of stiffness and compliance, to support a desirable proregenerative macrophage phenotype.
BioTester	2019	Image-Based Analysis Of Uniaxial Ring Test For Mechanical Characterization Of Soft Materials And Biological Tissues	Soft Matter	E. E. van Haaften, M. C. van Turnhout, N. A. Kurniawan	Eindhoven University of Technology		Uniaxial ring test is a widely used mechanical characterization method for a variety of materials, from industrial elastomers to biological materials. Here we show that the combination of local material compression, bending, and stretching during uniaxial ring test results in a geometry-dependent deformation profile that can introduce systematic errors in the extraction of mechanical parameters. We identify the stress and strain regimes under which stretching dominates and develop a simple image-based analysis approach that eliminates these systematic errors. We rigorously test this approach computationally and experimentally, and demonstrate that we can accurately estimate the sample mechanical properties for a wide range of ring geometries. As a proof of concept for its application, we use the approach to analyze explanted rat vascular tissues and find a clear temporal change in the mechanical properties of these explants after graft implantation. The image-based approach can therefore offer a straightforward, versatile, and accurate method for mechanically characterizing new classes of soft and biological materials.
BioTester	2019	Mechanical Properties of Autologous Pericardium Change with Fixation Time: Implications for Valve Reconstruction	Seminars in Thoracic and Cardiovascular Surgery	S. C. Hofferberth, C. W. Baird, D. M. Hoganson, L. G. Quinonez, S. M. Emani, P. J. Del Nido, P. E. Hammer	Boston Children's Hospital		Autologous pericardium, fixed in glutaraldehyde, is a common patch material used in heart valve reconstruction. Competence of a reconstructed valve depends on patch dimensions and also on how much the patch deforms when the closed valve is pressurized. In this study, we used biaxial testing to evaluate the deformability of fixed autologous pericardium, and several commercial alternatives, under typical cardiac loads. We found that deformability of fixed autologous pericardium varies predictably with fixation time. This information can be used by surgeons in designing patches for valve repair.
BioTester	2019	Tissue Level Mechanical Properties and Extracellular Matrix Investigation of the Bovine Jugular Venous Valve Tissue	Bioengineering	A. A. Benson, H-Y. S. Huang	North Carolina State University		Jugular venous valve incompetence has no long-term remedy and symptoms of transient global amnesia and/or intracranial hypertension continue to discomfort patients. During this study, we interrogate the synergy of the collagen and elastin microstructure that compose the bi-layer extracellular matrix (ECM) of the jugular venous valve. In this study, we investigate the jugular venous valve and relate it to tissue-level mechanical properties, fibril orientation and fibril composition to improve fundamental knowledge of the jugular venous valves toward the development of bioprosthetic venous valve replacements. Steps include: (1) multi loading biaxial mechanical tests; (2) isolation of the elastin microstructure; (3) imaging of the elastin microstructure; and (4) imaging of the collagen microstructure, including an experimental analysis of crimp. Results from this study show that, during a 3:1 loading ratio (circumferential direction: 900 mN and radial direction: 300 mN), elastin may have the ability to contribute to the circumferential mechanical properties at low strains, for example, shifting the inflection point toward lower strains in comparison to other loading ratios. After isolating the elastin microstructure, light microscopy revealed that the overall elastin orients in the radial direction while forming a crosslinked mesh. Collagen fibers were found undulated, aligning in parallel with neighboring fibers and orienting in the circumferential direction with an interquartile range of −10.38° to 7.58° from the circumferential axis (n = 20). Collagen crimp wavelength and amplitude was found to be 38.46 ± 8.06 µm and 4.51 ± 1.65 µm, respectively (n = 87). Analyzing collagen crimp shows that crimp permits about 12% true strain circumferentially, while straightening of the overall fibers accounts for more. To the best of the authors’ knowledge, this is the first study of the jugular venous valve linking the composition and orientation of the ECM to its mechanical properties and this study will aid in forming a structure-based constitutive model.
BioTester	2019	Regional Biaxial Mechanical Data of the Mitral and Tricuspid Valve Anterior Leaflets	Data in Brief	D. Laurence, C. Ross, S. Jett, C. Johns, A. Echols, R. Baumwart, R. Towner, J. Liao, P. Bajona, Y. Wu, C. H. Lee	University of Oklahoma		The collective data associated with this article presents the biaxial mechanical behavior for six smaller, delimited regions of the mitral valve and tricuspid valve anterior leaflets. Each data set consists of five columns of data, specifically: (i) biaxial testing protocol ID, (ii) circumferential stretch, (iii) radial stretch, (iv) circumferential membrane tension, and (v) radial membrane tension. For further elaboration regarding methodologies or results of the biaxial mechanical characterization please refer to the companion article
BioTester	2019	Mechanics of the Tricuspid Valve-From Clinical Diagnosis/Treatment, In-Vivo and In-Vitro Investigations, to Patient-Specific Biomechanical Modeling.	Bioengineering	C. H. Lee, D. Laurence, C. Ross, K. Kramer, A. Babu, E. Johnson, M. C. Hsu, A. Aggarwal, A. Mir, H. Burkhart, R. Towner, B. Baumwart, Y. Wu	University of Oklahoma		Proper tricuspid valve (TV) function is essential to unidirectional blood flow through the right side of the heart. Alterations to the tricuspid valvular components, such as the TV annulus, may lead to functional tricuspid regurgitation (FTR), where the valve is unable to prevent undesired backflow of blood from the right ventricle into the right atrium during systole. Various treatment options are currently available for FTR; however, research for the tricuspid heart valve, functional tricuspid regurgitation, and the relevant treatment methodologies are limited due to the pervasive expectation among cardiac surgeons and cardiologists that FTR will naturally regress after repair of left-sided heart valve lesions. Recent studies have focused on (i) understanding the function of the TV and the initiation or progression of FTR using both in-vivo and in-vitro methods, (ii) quantifying the biomechanical properties of the tricuspid valve apparatus as well as its surrounding heart tissue, and (iii) performing computational modeling of the TV to provide new insight into its biomechanical and physiological function. This review paper focuses on these advances and summarizes recent research relevant to the TV within the scope of FTR. Moreover, this review also provides future perspectives and extensions critical to enhancing the current understanding of the functioning and remodeling tricuspid valve in both the healthy and pathophysiological states.
BioTester	2019	An Investigation of the Glycosaminoglycan Contribution to Biaxial Mechanical Behaviours of Porcine Atrioventricular Heart Valve Leaflets	Journal of the Royal Society Interface	C. Ross, D. Laurence, Y. Wu, J. Richardson, A. Babu, L. Evans, E. Beyer, R. Childers, Y. Wu, R. Towner, K. M. Fung, A. Mir, H. Burkhart, G. A. Holzapfel, C. H. Lee	University of Oklahoma		The atrioventricular heart valve (AHV) leaflets have a complex microstructure composed of four distinct layers: atrialis, ventricularis, fibrosa and spongiosa. Specifically, the spongiosa layer is primarily proteoglycans and glycosaminoglycans (GAGs). Quantification of the GAGs' mechanical contribution to the overall leaflet function has been of recent focus for aortic valve leaflets, but this characterization has not been reported for the AHV leaflets. This study seeks to expand current GAG literature through novel mechanical characterizations of GAGs in AHV leaflets. For this characterization, mitral and tricuspid valve anterior leaflets (MVAL and TVAL, respectively) were: (i) tested by biaxial mechanical loading at varying loading ratios and by stress-relaxation procedures, (ii) enzymatically treated for removal of the GAGs and (iii) biaxially mechanically tested again under the same protocols as in step (i). Removal of the GAG contents from the leaflet was conducted using a 100 min enzyme treatment to achieve approximate 74.87% and 61.24% reductions of all GAGs from the MVAL and TVAL, respectively. Our main findings demonstrated that biaxial mechanical testing yielded a statistically significant difference in tissue extensibility after GAG removal and that stress-relaxation testing revealed a statistically significant smaller stress decay of the enzyme-treated tissue than untreated tissues. These novel findings illustrate the importance of GAGs in AHV leaflet behaviour, which can be employed to better inform heart valve therapeutics and computational models.
BioTester	2019	Macrophage-Driven Biomaterial Degradation Depends on Scaffold Microarchitecture	Frontiers in Bioengineering and Biotechnology	T. Wissing, V. Bonito, E. Haaften, M. Doeselaar, M. Brugmans, H. Janssen, C. Bouten, A. Smits	Eindhoven University of Technology		In situ tissue engineering is a technology in which non-cellular biomaterial scaffolds are implanted in order to induce local regeneration of replaced or damaged tissues. Degradable synthetic electrospun scaffolds are a versatile and promising class of biomaterials for various in situ tissue engineering applications, such as cardiovascular replacements. Functional in situtissue regeneration depends on the balance between endogenous neo-tissue formation and scaffold degradation. Both these processes are driven by macrophages. Upon invasion into a scaffold, macrophages secrete reactive oxygen species (ROS) and hydrolytic enzymes, contributing to oxidative and enzymatic biomaterial degradation, respectively. This study aims to elucidate the effect of scaffold microarchitecture, i.e., μm-range fiber diameter and fiber alignment, on early macrophage-driven scaffold degradation. Electrospun poly-ε-caprolactone-bisurea (PCL-BU) scaffolds with either 2 or 6 μm (Ø) isotropic or anisotropic fibers were seeded with THP-1 derived human macrophages and cultured in vitro for 4 or 8 days. Our results revealed that macroph age-induced oxidative degradation in particular was dependent on scaffold microarchitecture, with the highest level of ROS-induced lipid peroxidation, NADPH oxidase gene expression and degradation in the 6 μm Ø anisotropic group. Whereas, biochemically polarized macrophages demonstrated a phenotype-specific degradative potential, the observed differences in macrophage degradative potential instigated by the scaffold microarchitecture could not be attributed to either distinct M1 or M2 polarization. This suggests that the scaffold microarchitecture uniquely affects macrophage-driven degradation. These findings emphasize the importance of considering the scaffold microarchitecture in the design of scaffolds for in situ tissue engineering applications and the tailoring of degradation kinetics thereof.
BioTester	2019	The Effect of Axial Torsion on the Mechanical Properties of the Annulus Fibrosus	Biomechanics	M. Harvey-Burgess, D. Gregory	Wilfrid Laurier University		This study exposed bovine intervertebral discs to static axial torsion in order to determine its effect on the inter- and intralamellar matrices. Axial torsion resulted in decreased strength and stiffness of the intralamellar matrix, which may make the intervertebral disc more susceptible to herniation.
BioTester	2019	Integration of Polarized Spatial Frequency Domain Imaging (pSFDI) with a Biaxial Mechanical Testing System for Dynamic Quantification of Collagen Architecture in Soft Collagenous Tissues	SSRN	S. Jett, L. Hudson, R. Baumwart, B. Bohnstedt, A. Mir, H. Burkhart, G. Holzapfel, C-H. Lee, Y. Wu	University of Oklahoma		Collagen fiber networks provide the structural strength of tissues such as tendons, skin, and arteries. Quantifying the response of the fiber architecture to mechanical loads is essential towards a better understanding of the tissue-level mechanical behaviors, especially in assessing disease-driven functional changes. To enable novel investigations into these dynamic fiber structures, a polarized spatial frequency domain imaging (pSFDI) device was developed and, for the first time, integrated with a biaxial mechanical testing system. The integrated instrument is capable of a wide-field and dynamic quantification of the fiber orientation and degree of optical anisotropy (DOA), representing the local strength of fiber alignment. The performance of this integrated instrument was assessed through uniaxial testing on tendon tissues with known collagen fiber microstructures. Our results revealed that the fiber orientation of the tendon tissue changed indiscernibly, whereas the fibers became better aligned with the average DOA increasing from 0.126 to 0.215 under 0% and 3% uniaxial strains, respectively. The integrated instrument was further applied to study the mitral valve anterior leaflet (MVAL) tissue subjected to various biaxial loadings. The fiber orientations within the MVAL demonstrated a proclivity towards the tissue's circumferential direction under all loading protocols, while certain fiber groups re-oriented towards the tissue's radial direction under radially-dominant loading. Our results also showed that fibers were generally better aligned under equibiaxial (DOA=0.088) and circumferentially-dominant loading (DOA=0.084) than under the radially-dominant loading (DOA=0.074), indicating circumferential predisposition. These novel findings exemplify a deeper understanding of dynamic collagen fiber microstructures obtained through the integrated opto-mechanical instrument.
BioTester	2019	An Integrated Opto-Mechanical System for Quantification of Dynamic Microstructure and Mechanics of Heart Valve Tissues	SB3C	S. Jett, Z. Schuermann, A. Mir, H. Burkhart, C-H. Lee	University of Oklahoma		Many biomechanical diseases, such as heart valve stenosis, tendon tears, and arterial aneurysms, cause impairment in organ function via alterations and remodeling in tissue microstructure. Consequently, prior studies have examined the microstructure of healthy and diseased tissues [1]. Methodologies for microstructural analysis include histology, small angle light scattering (SALS), polarized light microscopy (PLM), and second-harmonic generation (SHG) imaging, among others. However, these techniques are incapable of capturing the dynamic nature of tissue microstructure and the collagen fibers therein. In collagenous tissues, such as the tendon and heart valves (HVs), the orientations and dispersion of the load-bearing collagen fibers dynamically adjust in response to mechanical loading [2]. In this study, we developed and validated a novel integrated system consisting of a biaxial tester and a polarized spatial frequency domain imaging (pSFDI) device to quantify load-driven changes to collagen microstructural architecture in tissues.
BioTester	2019	Effects of longitudinal pre-stretch on the mechanics of human aorta before and after thoracic endovascular aortic repair (TEVAR) in trauma patients	Biomechanics and Modeling in MechanoBiology	A. Desyatova, J. MacTaggart, A. Kamenskiy	University of Nebraska Omaha		Thoracic endovascular aortic repair (TEVAR) has evolved as a first-line therapy for trauma patients. Most trauma patients are young, and their aortas are compliant and longitudinally pre-stretched. We have developed a method to include longitudinal pre-stretch in computational models of human thoracic aortas of different ages before and after TEVAR. Finite element models were built using computerized tomography angiography data obtained from human subjects in 6 age groups 10–69 years old. Aortic properties were determined with planar biaxial testing, and pre-stretch was simulated using a series of springs. GORE C-Tag stent-graft was computationally deployed in aortas with and without pre-stretch, and the stress–strain fields were compared. Pre-stretch had significant qualitative and quantitative effects on the aortic stress–strain state before and after TEVAR. Before TEVAR, mean intramural aortic stresses with and without pre-stretch decreased with age from 108 kPa and 83 kPa in the youngest age group, to 60 kPa in the oldest age group. TEVAR increased intramural stresses by an average of 73 ± 15 kPa and 48 ± 10 kPa for aortas with and without pre-stretch and produced high stress concentrations near the aortic isthmus. Inclusion of pre-stretch in young aortas increased intramural stresses by 30%, while in > 50-year-old subjects it did not change the results. Computational modeling of aorta-stent-graft interaction that includes pre-stretch can be instrumental for device design and assessment of its long-term performance, and in the future may help more accurately determine the stress–strain characteristics associated with TEVAR complications.
BioTester	2019	Human in vitro model of material-driven vascular regeneration reveals how cyclic stretch and shear stress differentially modulate inflammation and tissue formation	bioRxiv	E. Haaften, T. Wissing, N. Kurniawan, A. Smits, C. Bouten	Eindhoven University of Technology		Resorbable synthetic scaffolds designed to regenerate living tissues and organs inside the body emerge as a clinically attractive technology to replace diseased blood vessels. However, mismatches between scaffold design and in vivo hemodynamic loading (i.e., cyclic stretch and shear stress) can result in aberrant inflammation and adverse tissue remodeling, leading to premature graft failure. Yet, the underlying mechanisms remain elusive. Here, a human in vitro model is presented that mimics the transient local inflammatory and biomechanical environments that drive scaffold-guided tissue regeneration. The model is based on the co-culture of human (myo)fibroblasts and macrophages in a bioreactor platform that decouples cyclic stretch and shear stress. Using a resorbable supramolecular elastomer as the scaffold material, it is revealed that cyclic stretch initially reduces pro-inflammatory cytokine secretion and, especially when combined with shear stress, stimulates IL-10 secretion. Moreover, cyclic stretch stimulates downstream (myo)fibroblast proliferation and neotissue formation. In turn, shear stress attenuates cyclic-stretch-induced tissue growth by enhancing MMP-1/TIMP-1-mediated collagen remodeling, and synergistically alters (myo)fibroblast phenotype when combined with cyclic stretch. The findings suggest that shear stress acts as a stabilizing factor in cyclic stretch-induced tissue formation and highlight the distinct roles of hemodynamic loads in the design of resorbable vascular grafts.
BioTester	2019	Some Effects of Different Constitutive Laws on FSI Simulation for the Mitral Valve	Nature Scientific Reports	L. Cai, Y. Wang, H. Gao, X. Ma, G. Zhu, R. Zhang, X. Shen, X. Luo	Chongqing University		In this paper, three different constitutive laws for mitral leaflets and two laws for chordae tendineae are selected to study their effects on mitral valve dynamics with fluid-structure interaction. We first fit these three mitral leaflet constitutive laws and two chordae tendineae laws with experimental data. The fluid-structure interaction is implemented in an immersed boundary framework with finite element extension for solid, that is the hybrid immersed boundary/finite element(IB/FE) method. We specifically compare the fluid-structure results of different constitutive laws since fluid-structure interaction is the physiological loading environment. This allows us to look at the peak jet velocity, the closure regurgitation volume, and the orifice area. Our numerical results show that different constitutive laws can affect mitral valve dynamics, such as the transvalvular flow rate, closure regurgitation and the orifice area, while the differences in fiber strain and stress are insignificant because all leaflet constitutive laws are fitted to the same set of experimental data. In addition, when an exponential constitutive law of chordae tendineae is used, a lower closure regurgitation flow is observed compared to that of a linear material model. In conclusion, combining numerical dynamic simulations and static experimental tests, we are able to identify suitable constitutive laws for dynamic behaviour of mitral leaflets and chordae under physiological conditions.
BioTester	2019	Biomechanical Restoration Potential of Pentagalloyl Glucose after Arterial Extracellular Matrix Degeneration	Bioengineering	S. Patnaik, S. Piskin, N. Pillalamarri, G. Romero, G. Escobar, E. Sprague, E. Finol	University of Texas at San Antonio		The objective of this study was to quantify pentagalloyl glucose (PGG) mediated biomechanical restoration of degenerated extracellular matrix (ECM). Planar biaxial tensile testing was performed for native (N), enzyme-treated (collagenase and elastase) (E), and PGG (P) treated porcine abdominal aorta specimens (n = 6 per group). An Ogden material model was fitted to the stress–strain data and finite element computational analyses of simulated native aorta and aneurysmal abdominal aorta were performed. The maximum tensile stress of the N group was higher than that in both E and P groups for both circumferential (43.78 ± 14.18 kPa vs. 10.03 ± 2.68 kPa vs. 13.85 ± 3.02 kPa; p = 0.0226) and longitudinal directions (33.89 ± 8.98 kPa vs. 9.04 ± 2.68 kPa vs. 14.69 ± 5.88 kPa; p = 0.0441). Tensile moduli in the circumferential direction was found to be in descending order as N > P > E (195.6 ± 58.72 kPa > 81.8 ± 22.76 kPa > 46.51 ± 15.04 kPa; p = 0.0314), whereas no significant differences were found in the longitudinal direction (p = 0.1607). PGG binds to the hydrophobic core of arterial tissues and the crosslinking of ECM fibers is one of the possible explanations for the recovery of biomechanical properties observed in this study. PGG is a beneficial polyphenol that can be potentially translated to clinical practice for preventing rupture of the aneurysmal arterial wall.
BioTester	2019	An investigation of the effect of freezing storage on the biaxial mechanical properties of excised porcine tricuspid valve anterior leaflets	Journal of the Mechanical Behavior of Biomedical Materials	G. Duginski, C. Ross, D. Laurence, C. Johns, C-H. Lee	University of Oklahoma		The atrioventricular heart valve (AHV) leaflets are critical to the facilitation of proper unidirectional blood flow through the heart. Previously, studies have been conducted to understand the tissue mechanics of healthy AHV leaflets to inform the development of valve-specific computational models and replacement materials for use in diagnosing and treating valvular heart disease. Generally, these studies involved biaxial mechanical testing of the AHV leaflet tissue specimens to extract relevant mechanical properties. Most of those studies considered freezing-based storage systems based on previous findings for other connective tissues such as aortic tissue or skin. However, there remains no study that specifically examines the effects of freezing storage on the characterized mechanical properties of the AHV leaflets. In this study, we aimed to address this gap in knowledge by performing biaxial mechanical characterizations of the tricuspid valve anterior leaflet (TVAL) tissue both before and after a 48-h freezing period. Primary findings of this study include: (i) a statistically insignificant change in the tissue extensibilities, with the frozen tissues being slightly stiffer and more anisotropic than the fresh tissues; and (ii) minimal variations in the stress relaxation behaviors between the fresh and frozen tissues, with the frozen tissues demonstrating slightly lessened relaxation. The findings from this study suggested that freezing-based storage does not significantly impact the observed mechanical properties of one of the five AHV leaflets—the TVAL. The results from this study are useful for reaffirming the experimental methodologies in the previous studies, as well as informing the tissue preservation methods of future investigations of AHV leaflet mechanics.
BioTester	2019	A Detailed Mechanical and Microstructural Analysis of the Ovine Tricuspid Valve Leaflets	Acta Biomaterialia	W. Meador, M. Mathur, T. Jazwiec, M. Malinowski, M. Bersi, T. Timek, M. Rausch	University of Texas at Austin		The tricuspid valve ensures unidirectional blood flow from the right atrium to the right ventricle. The three tricuspid leaflets operate within a dynamic stress environment of shear, bending, tensile, and compressive forces, which is cyclically repeated nearly two billion times in a lifetime. Ostensibly, the microstructural and mechanical properties of the tricuspid leaflets have mechanobiologically evolved to optimally support their function under those forces. Yet, how the tricuspid leaflet microstructure determines its mechanical properties and whether this relationship differs between the three leaflets is unknown. Here we perform a microstructural and mechanical analysis in matched ovine tricuspid leaflet samples. We found that the microstructure and mechanical properties vary among the three tricuspid leaflets in sheep. Specifically, we found that tricuspid leaflet composition, collagen orientation, and valve cell nuclear morphology are spatially heterogeneous and vary across leaflet type. Furthermore, under biaxial tension the leaflets' mechanical behaviors exhibited unequal degrees of mechanical anisotropy. Most importantly, we found that the septal leaflet was stiffer in the radial direction and not the circumferential direction as with the other two leaflets. The differences we observed in leaflet microstructure coincide with the varying biaxial mechanics among leaflets. Our results demonstrate the structure-function relationship for each leaflet in the tricuspid valve. We anticipate our results to be vital toward developing more accurate, leaflet-specific tricuspid valve computational models. Furthermore, our results may be clinically important, informing differential surgical treatments of the tricuspid valve leaflets. Finally, the identified structure-function relationships may provide insight into the homeostatic and remodeling potential of valvular cells in altered mechanical environments, such as in diseased or repaired tricuspid valves.
BioTester	2019	The regional-dependent biaxial behavior of young and aged mouse skin: A detailed histomechanical characterization, residual strain analysis, and constitutive model	Acta Biomaterialia	W. Meador, G. Sugerman, H. Story, A. Seifert, M. Bersi, A. Tepole, M. Rausch	University of Texas at Austin		Skin fulfills several vital functions, many of which are dependent on its mechanical properties. Therefore, as mice have become an invaluable model for skin research, determining murine skin's mechanical properties is important. Specifically, skin's mechanical properties are important for functional tests as well as for prognostic and diagnostic purposes. Additionally, computational simulations of skin behavior are becoming commonplace, rendering accurate models of murine skin's constitutive behavior necessary. To date, our knowledge of mouse skin mechanics shows significant gaps. For example, there are no comprehensive reports correlating skin's mechanical properties with region, age, and direction. Moreover, mouse skin's residual strain behavior has not been reported on. In our current work, we set out to fill these gaps. Based on histology, 2-photon microscopy, and planar biaxial testing, while accurately tracking various reference configurations, we report on differences in gross structure, microstructural organization, and constitutive response of skin, and cast those properties into a versatile Fung-type hyperelastic constitutive law for three reference configurations. Our data is the most comprehensive report contrasting the mechanical properties of young (12 weeks) and aged (52 weeks) mouse skin and will, thus, be valuable to basic science, as control data, and provide accurate constitutive laws for mouse skin modeling.
BioTester	2019	Integration of polarized spatial frequency domain imaging (pSFDI) with a biaxial mechanical testing system for quantification of load-dependent collagen architecture in soft collagenous tissues	Acta Biomaterialia	S. Jett, L. Hudson, R. Baumwart, B. Bohnstedt, A. Mir, H. Burkhart, G. Holzapfel, C-H. Lee, Y. Wu	University of Oklahoma		Collagen fiber networks provide the structural strength of tissues, such as tendons, skin and arteries. Quantifying the fiber architecture in response to mechanical loads is essential towards a better understanding of the tissue-level mechanical behaviors, especially in assessing disease-driven functional changes. To enable novel investigations into these load-dependent fiber structures, a polarized spatial frequency domain imaging (pSFDI) device was developed and, for the first time, integrated with a biaxial mechanical testing system. The integrated instrument is capable of a wide-field quantification of the fiber orientation and the degree of optical anisotropy (DOA), representing the local degree of fiber alignment. The opto-mechanical instrument''s performance was assessed through uniaxial loading on tendon tissues with known collagen fiber microstructures. Our results revealed that the bulk fiber orientation angle of the tendon tissue changed minimally with loading (median ± 0.5*IQR of 52.7° ± 3.3° and 51.9° ± 3.3° under 0 and 3% longitudinal strains, respectively), whereas on a micro-scale, the fibers became better aligned with the direction of loading: the DOA (mean ± SD) increased from 0.149 ± 0.032 to 0.198 ± 0.056 under 0 and 3% longitudinal strains, respectively, p < 0.001. The integrated instrument was further applied to study two representative mitral valve anterior leaflet (MVAL) tissues subjected to various biaxial loads. The fiber orientations within these representative MVAL tissue specimens demonstrated noticeable heterogeneity, with the local fiber orientations dependent upon the sample, the spatial and transmural locations, and the applied loading. Our results also showed that fibers were generally better aligned under equibiaxial (DOA = 0.089 ± 0.036) and circumferentially-dominant loading (DOA = 0.086 ± 0.037) than under the radially-dominant loading (DOA = 0.077 ± 0.034), indicating circumferential predisposition. These novel findings exemplify a deeper understanding of the load-dependent collagen fiber microstructures obtained through the use of the integrated opto-mechanical instrument.
BioTester	2019	Methods of Delivering Mechanical Stimuli to Organ-on-a-Chip	Micromachines	K. Kaarj, J-Y., Yoon	University of Arizona		Recent advances in integrating microengineering and tissue engineering have enabled the creation of promising microengineered physiological models, known as organ-on-a-chip (OOC), for experimental medicine and pharmaceutical research. OOCs have been used to recapitulate the physiologically critical features of specific human tissues and organs and their interactions. Application of chemical and mechanical stimuli is critical for tissue development and behavior, and they were also applied to OOC systems. Mechanical stimuli applied to tissues and organs are quite complex in vivo, which have not adequately recapitulated in OOCs. Due to the recent advancement of microengineering, more complicated and physiologically relevant mechanical stimuli are being introduced to OOC systems, and this is the right time to assess the published literature on this topic, especially focusing on the technical details of device design and equipment used. We first discuss the different types of mechanical stimuli applied to OOC systems: shear flow, compression, and stretch/strain. This is followed by the examples of mechanical stimuli-incorporated OOC systems. Finally, we discuss the potential OOC systems where various types of mechanical stimuli can be applied to a single OOC device, as a better, physiologically relevant recapitulation model, towards studying and evaluating experimental medicine, human disease modeling, drug development, and toxicology.
BioTester	2019	Optimization of Oxygen Dynamics, UV-A Delivery, and Drug Formulation for Accelerated Epi-On Corneal Crosslinking	Current Eye Research	J. Hill, C. Liu, P. Deardorff, B. Tavakol, W. Eddington, V. Thompson, D. Gore, M. Raizman, D. Adler	Avedro Inc		Purpose: Corneal collagen crosslinking (CXL) through an intact epithelium (epi-on) at high irradiance could potentially improve patient comfort, visual recovery, and clinical workflow compared to conventional epi-off CXL. However, intact epithelium limits stromal delivery of the oxygen, photosensitizer, and ultraviolet-A (UV-A) radiation needed to drive CXL. This ex vivo study evaluated three different epi-on CXL protocols compared to positive and negative controls, specifically focusing on the impact of supplemental oxygen. Endpoints included stromal oxygen levels, stiffness of crosslinked tissue, and acute flattening of whole eyes.
BioTester	2019	Reproducibility Assessment of Ultrasound-based Aortic Stiffness Quantification and Verification using Bi-axial Tensile Testing	Journal of the Mechanical Behavior of Biomedical Materials	E. Disseldorp, M. van den Hoven, F. van de Vosse, M. van Sambeek, R. Lopata	Eindhoven University of Technology		Current guidelines for abdominal aortic aneurysm (AAA) repair are primarily based on the maximum diameter. Since these methods lack robustness in decision making, new image-based methods for mechanical characterization have been proposed. Recently, time-resolved 3D ultrasound (4D US) in combination with finite element analysis was shown to provide additional risk estimators such as patient-specific peak wall stresses and wall stiffness in a non-invasive way. The aim of this study is to: 1) assess the reproducibility of this US-based stiffness measurement in vitro and in vivo, and 2) verify this 4D US stiffness using the gold standard: bi-axial tensile testing of the excised aortic tissue.
BioTester	2019	Mechanical and Structural Changes in Human Thoracic Aortas with Age	Acta Biomaterialia	M. Jadidi, M. Habibnezhad, E. Anttila, K. Maleckis, A. Desyatova, J. MacTaggart, A. Kamenskiy	University of Nebraska-Lincoln		Aortic mechanical and structural characteristics have profound effects on pathophysiology, but many aspects of physiologic stress-stretch state and intramural changes due to aging remain poorly understood in human tissues. While difficult to assess in vivo due to residual stresses and pre-stretch, physiologic stress-stretch characteristics can be calculated using experimentally-measured mechanical properties and constitutive modeling. Mechanical properties of 76 human descending thoracic aortas (TA) from 13 to 78-year-old donors (mean age 51±18 years) were measured using multi-ratio planar biaxial extension. Constitutive parameters were derived for aortas in 7 age groups, and the physiologic stress-stretch state was calculated. Intramural characteristics were quantified from histological images and related to aortic morphometry and mechanics. TA stiffness increased with age, and aortas became more nonlinear and anisotropic. Systolic and diastolic elastic energy available for pulsation decreased with age from 30 to 8 kPa and from 18 to 5 kPa, respectively. Cardiac cycle circumferential stretch dropped from 1.14 to 1.04, and circumferential and longitudinal physiologic stresses decreased with age from 90 to 72 kPa and from 90 to 17 kPa, respectively. Aortic wall thickness and radii increased with age, while the density of elastin in the tunica media decreased. The number of elastic lamellae and circumferential physiologic stress per lamellae unit remained constant with age at 102±10 and 0.85±0.04 kPa, respectively. Characterization of mechanical, physiological, and structural features in human aortas of different ages can help understand aortic pathology, inform the development of animal models that simulate human aging, and assist with designing devices for open and endovascular aortic repairs.
BioTester	2020	Load-dependent Collagen Fiber Architecture Data of Representative Bovine Tendon and Mitral Valve Anterior Leaflet Tissues as Quantified by an Integrated Opto-mechanical System	Data in Brief	S. Jett, L. Hudson, R. Baumwart, B. Bohnstedt, A. Mir, H. Burkhart, G. Holzapfel, C-H. Lee, Y. Wu	University of Oklahoma		The data presented in this article provide load-dependent collagen fiber architecture (CFA) of one representative bovine tendon tissue sample and two representative porcine mitral valve anterior leaflet tissues, and they are stored in a MATLAB MAT-file format. Each dataset contains: (i) the number of pixel points, (ii) the array of pixel's x- and y-coordinates, (iii) the three acquired pixel intensity arrays, and (iv) the Delaunay triangulation for visualization purpose. This dataset is associated with a companion journal article, which can be consulted for further information about the methodology, results, and discussion of the opto-mechanical characterization of the tissue's CFA's (Jett et al. [1]).
BioTester	2020	Mechanics of Porcine Heart Valves’ Strut Chordae Tendineae Investigated as a Leaflet–Chordae–Papillary Muscle Entity	Annals of Biomedical Engineering	C. Ross, D. Laurence, M-C. Hsu, R. Baumwart, Y. Zhao, A. Mir, H. Burkhart, G. Holzapfel, Y. Wu, C-H. Lee	University of Oklahoma		Proper blood flow through the atrioventricular heart valves (AHVs) relies on the holistic function of the valve and subvalvular structures, and a failure of any component can lead to life-threatening heart disease. A comprehension of the mechanical characteristics of healthy valvular components is necessary for the refinement of heart valve computational models. In previous studies, the chordae tendineae have been mechanically characterized as individual structures, usually in a clamping-based approach, which may not accurately reflect the in vivo chordal interactions with the leaflet insertion and papillary muscles. In this study, we performed uniaxial mechanical testing of strut chordae tendineae of the AHVs under a unique tine-based leaflet–chordae–papillary muscle testing to observe the chordae mechanics while preserving the subvalvular component interactions. Results of this study provided insight to the disparity of chordae tissue stress-stretch responses between the mitral valve (MV) and the tricuspid valve (TV) under their respective emulated physiological loading. Specifically, strut chordae tendineae of the MV anterior leaflet had peak stretches of 1.09–1.16, while peak stretches of 1.08–1.11 were found for the TV anterior leaflet strut chordae. Constitutive parameters were also derived for the chordae tissue specimens using an Ogden model, which is useful for AHV computational model refinement. Results of this study are beneficial to the eventual improvement of treatment methods for valvular disease.
BioTester	2020	Pre-mounted dry TAVI valve with improved endothelialization potential using REDV-loaded PEGMA hydrogel hybrid pericardium	Journal of Materials Chemistry B	L. Yang, X. Huang, L. Deng, X. Ma, H. Jiang, Q. Ning, Z. Liang, Y. Lei, Y. Wang	Sichuan University		The pre-mounted dry transcatheter aortic valve implantation (TAVI) valve is a new technology in the development of biological heart valves. Dry valves do not need to be placed in special preservation solution and can be opened and used immediately, meeting the needs of clinical emergency valve implantation. However, current biological valves obtained by simple air drying cannot be unfolded quickly. In addition, the crimping process leads to structural damage to the valve fiber microstructure, reducing the service life of biological valves. Furthermore, current biological valves still have problems such as calcification, endothelialization difficulty, and immune rejection. In this study, a poly(ethylene glycol)methacrylate (PEGMA) hydrogel hybrid pericardium loaded with REDV was developed. The PEGMA monomer solution can penetrate the space of the pericardium. REDV was loaded into the PEGMA hydrogel, which was hybridized with pericardium via in situ polymerization. The results showed improved unfolding properties, less mechanical damage after crimping, and improved endothelialization potential of the biological valve. Thus, REDV-loaded PEGMA hydrogel hybrid pericardium is a promising approach for obtaining pre-mounted dry TAVI valves with enhanced endothelialization properties.
BioTester	2020	Radiological, dosimetric and mechanical properties of a deformable breast phantom for radiation therapy and surgical applications	Biomedical Physics and Engineering Express	G. Aldosary, T. Tse, A. Arnaout, J-M.Caudrelier, C. Czyrnyj, R. Romain, L. McLean, C. Foottit, E. Vandervoort, J. Belec	University of Ottawa		The displacement of tumor bed walls during oncoplastic breast surgery (OBS) decreases the accuracy of using clips as the sole surrogate for tumor bed location. This highlights the need for better communication of OBS techniques to radiation oncologists. To facilitate OBS practice and investigate clip placement reliability, a realistic silicone-based breast phantom was constructed with components emulating a breast parenchyma, epidermis, areola, nipple, chest wall, and a tumor. OBS was performed on the phantom and surgical clips were placed to mark the tumor bed. The phantom was imaged with CT, MRI, and ultrasound (US). The parenchyma's signal-to-noise ratio (SNR) and clips to parenchyma's contrast-to-noise ratio (CNR) were measured. The phantom's CT Hounsfield Unit (HU), relative electron density (RED), and mass density were determined. 6 and 10 MV photon beam attenuation measurements were performed in phantom material. The Young's Modulus and ultimate tensile strength (UTS) of the phantom parenchyma and epidermis were measured. Results showed that the breast phantom components were visible on all imaging modalities with adequate CNR and SNR. The phantom's HU is 130 ± 10. The RED is 0.983. Its mass density is 1.01 ± 0.01 g/cm3. Photon attenuation measurements in phantom material were within 1% of those in water. The Young's Moduli were 13.4 ± 4.2 kPa (mechanical) and 30.2 ± 4.1 kPa (US elastography) for the phantom parenchyma. The UTS' were 0.05 ± 0.01 MPa (parenchyma) and 0.23 ± 0.12 MPa (epidermis). We conclude that the phantom's imaging characteristics resemble a fibroglandular breast's and allow clear visualization of high-density markers used in radiation therapy. The phantom material is suitable for dose measurements in MV photon beams. Mechanical results confirmed the phantom's similarity to breast tissue. The phantom enables investigation of surgical clip displacements pre and post-OBS, and is useful for radiation therapy quality assurance applications.
BioTester	2020	A combined in vivo and in vitro approach to assess supraspinatus activation and tissue responses to arm elevation demands	UWSpace	A. Cudlip	University of Waterloo		Rotator cuff degeneration affects a large portion of the human population, yet knowledge surrounding which loading scenarios allow transition from healthy to diseased states remains largely unresolved. Mechanistic progression of rotator cuff pathology often originates in the supraspinatus before cascading to other tissues, leading to substantial degeneration. Posture, loading and repetitive motions are known risk factors that exacerbate shoulder injury progression. This suggests a causal relationship between specific upper extremity task scenarios and degenerative rotator cuff loading. This thesis intentionally explored regional activations of the supraspinatus and accompanying tendon loading across a range of postures. The global objective was to evaluate how postural and task intensity differences alter tissue-level mechanical parameters in both in vivo muscular activation and in vitro tangent stiffness, hysteresis and optical stretch ratios.
Custom Solutions	2019	Functional Arrays Of Human Pluripotent Stem Cell-Derived Cardiac Microtissues	bioRxiv	N. Thavandiran, C. Hale, P. Blit, M. L. Sandberg, M. E. McElvain, M. Gagliardi, B. Sun, A. Witty, G. Graham, M. Mcintosh, M. A. Bakooshli, H. Le, J. Ostblom, S. McEwen, E. Chau, A. Prowse, I. Fernandes, P. M. Gilbert, G. Keller, P. Tagari, H. Xu, M. Radisic, P. W. Zandstra, D. Nojima, H. Vargas, Y. Qu, A. Motani, J. Reagan	University of Toronto		To accelerate the cardiac drug discovery pipeline, we set out to develop a platform that would be amenable to standard multiwell-plate manipulations and be capable of quantifying tissue-level functions such as contractile force. We report a 96-well-based array of 3D human pluripotent stem cell (hPSC)-derived cardiac microtissues - termed Cardiac MicroRings (CaMiRi) - in custom printed multiwell plates capable of contractile force measurement. Within each well, two elastomeric microcantilevers are situated above a ramp. The wells are seeded with cell-laden collagen which, in response to the slope of the ramp, self-organizes around tip-gated microcantilevers to form contracting CaMiRi. The contractile force exerted by the CaMiRi is measured and calculated using the deflection of the cantilevers. Platform responses were robust and comparable across wells and we used it to determine an optimal tissue formulation. We validated contractile force response of CaMiRi using selected cardiotropic compounds with known effects. Additionally, we developed automated protocols for CaMiRi seeding, image acquisition, and analysis to enable measurement of contractile force with increased throughput. The unique tissue fabrication properties of the platform, and the consequent effects on tissue function, were demonstrated upon adding hPSC-derived epicardial cells to the system. This platform represents an open-source contractile force screening system useful for drug screening and tissue engineering applications.
MechanoCulture B1	2018	Role Of Boundary Conditions In Determining Cell Alignment In Response To Stretch	PNAS	K. Chen, A. Vigliotti, M. Bacca, R. M. McMeeking, V.S. Dashpande, J.W. Holmes	University of VIrginia		The ability of cells to orient in response to mechanical stimuli is essential to embryonic development, cell migration, mechanotransduction, and other critical physiologic functions in a range of organs. Endothelial cells, fibroblasts, mesenchymal stem cells, and osteoblasts all orient perpendicular to an applied cyclic stretch when plated on stretchable elastic substrates, suggesting a common underlying mechanism. However, many of these same cells orient parallel to stretch in vivo and in 3D culture, and a compelling explanation for the different orientation responses in 2D and 3D has remained elusive. Here, we conducted a series of experiments designed specifically to test the hypothesis that differences in strains transverse to the primary loading direction give rise to the different alignment patterns observed in 2D and 3D cyclic stretch experiments (“strain avoidance”). We found that, in static or low-frequency stretch conditions, cell alignment in fibroblast-populated collagen gels correlated with the presence or absence of a restraining boundary condition rather than with compaction strains. Cyclic stretch could induce perpendicular alignment in 3D culture but only at frequencies an order of magnitude greater than reported to induce perpendicular alignment in 2D. We modified a published model of stress fiber dynamics and were able to reproduce our experimental findings across all conditions tested as well as published data from 2D cyclic stretch experiments. These experimental and model results suggest an explanation for the apparently contradictory alignment responses of cells subjected to cyclic stretch on 2D membranes and in 3D gels.
MechanoCulture B1	2019	Stretch In Focus: 2d Inplane Cell Stretch Systems For Studies Of Cardiac Mechano-Signaling	Frontiers in Bioengineering and Biotechnology	O. Friedrich, A-L. Merten, D. Schneidereit, Y. Guo, S. Schurmann, N. Martinac	Friedrich-Alexander-University Erlangen-Nürnberg		Mechanobiology is a rapidly growing interdisciplinary research field, involving biophysics, molecular and cell biology, biomedical engineering, and medicine. Rapid progress has been possible due to emerging devices and tools engineered for studies of the effect of mechanical forces, such as stretch or shear force, impacting on biological cells and tissues. In response to such mechanical stimuli, cells possess various mechanosensors among which mechanosensitive ion channels are molecular transducers designed to convert mechanical stimuli into electrical and/or biochemical intracellular signals on millisecond time scales. To study their role in cellular signaling pathways, devices have been engineered that enable application of different strain protocols to cells allowing for determination of the stress-strain relationship or other, preferably optical, readouts. Frequently, these devices are mounted on fluorescence microscopes, allowing simultaneous investigation of cellular mechanotransduction processes combined with live-cell imaging. Mechanical stress in organs/tissues can be complex and multiaxial, e.g., in hollow organs, like lung alveoli, bladder, or the heart. Therefore, biomedical engineers have, in recent years, developed devices based on elastomeric membranes for application of biaxial or multiaxial stretch to 2D substrate-adhered or even 3D-embedded cells. Here, we review application of stretch technologies to cellular mechanotransduction with a focus on cardiovascular systems. We also present new results obtained by our IsoStretcher device to examine mechanosensitivity of adult ventricular cardiomyocytes. We show that sudden isotropic stretch of cardiomyocytes can already trigger arrhythmic Ca2+ transients on the single cell level.
MechanoCulture B1	2019	Methods of Delivering Mechanical Stimuli to Organ-on-a-Chip	Micromachines	K. Kaarj, J-Y., Yoon	University of Arizona		Recent advances in integrating microengineering and tissue engineering have enabled the creation of promising microengineered physiological models, known as organ-on-a-chip (OOC), for experimental medicine and pharmaceutical research. OOCs have been used to recapitulate the physiologically critical features of specific human tissues and organs and their interactions. Application of chemical and mechanical stimuli is critical for tissue development and behavior, and they were also applied to OOC systems. Mechanical stimuli applied to tissues and organs are quite complex in vivo, which have not adequately recapitulated in OOCs. Due to the recent advancement of microengineering, more complicated and physiologically relevant mechanical stimuli are being introduced to OOC systems, and this is the right time to assess the published literature on this topic, especially focusing on the technical details of device design and equipment used. We first discuss the different types of mechanical stimuli applied to OOC systems: shear flow, compression, and stretch/strain. This is followed by the examples of mechanical stimuli-incorporated OOC systems. Finally, we discuss the potential OOC systems where various types of mechanical stimuli can be applied to a single OOC device, as a better, physiologically relevant recapitulation model, towards studying and evaluating experimental medicine, human disease modeling, drug development, and toxicology.
MechanoCulture FX	2017	Mechanical And Signaling Roles For Keratin Intermediate Filaments In The Assembly And Morphogenesis Of Mesendoderm Tissue At Gastrulation	Development Journal	P.R. Sonavane, C. Wang, B. Dzamba, G.F. Weber, A. Periasamy, D.W. DeSimone	University of Virginia		Coordination of individual cell behaviors is a critical step in the assembly and morphogenesis of tissues. Xenopus mesendoderm cells migrate collectively along a fibronectin (FN) substrate at gastrulation but how the adhesive and mechanical forces required for these movements are generated and transmitted is unclear. Traction force microscopy (TFM) was used to establish that traction stresses are limited primarily to leading edge cells in mesendoderm explants and that these forces are balanced by intercellular stresses in follower rows. This is further reflected in the morphology of these cells, with broad lamellipodial protrusions, mature focal adhesions and a gradient of activated Rac1 evident at the leading edge; while small protrusions, rapid turnover of immature focal adhesions, and lack of a Rac1 activity gradient characterize cells in following rows. Depletion of keratin (8) with antisense morpholinos results in high traction stresses in follower row cells, misdirected protrusions, and the formation of actin stress fibers anchored in streak-like focal adhesions. We propose that maintenance of mechanical integrity in the mesendoderm by keratin intermediate filaments is required to balance stresses within the tissue to regulate collective cell movements. Cell/Tissue: xenopus mesendoderm cells
MechanoCulture FX	2017	Tenogenic Phenotype Maintenance And Differentiation Using Macromolecular Crowding And Mechanical Loading	Orthopaedic Proceedings	D. Gaspar, A. Pandit, D. Zeugolis	National University of Ireland		Cell-based tissue engineering strategies for tendon repair have limited clinical applicability due to delayed extracellular matrix (ECM) deposition and subsequent prolonged culture periods, which lead to tenogenic phenotypic drift. Deposition of ECM in vitrocan be enhanced by macromolecular crowding (MMC), a biophysical phenomenon that governs the intra- and extra-cellular milieu of multicellular organisms2, which has been described to accelerate ECM deposition in human tenocytes1. A variety of cell sources have been studied for tendon repair including tenocytes, dermal fibroblasts and mesenchymal stem cells (MSCs)3and various biophysical, biochemical and biological tools have been used to mimic tendon microenvironment and induce phenotype maintenance in long term cultures or differentiation4. Therefore, we propose to assess the combined effect of macromolecular crowding and mechanical loading on different cell sources to determine their suitability for the in vitro fabrication of tendon-like tissue. Human dermal fibroblasts, tenocytes and bone marrow mesenchymal stem cells were cultured for 3 days with 100 µg/ml of carrageenan (MMC) under static and dynamic culture conditions. Cyclic uniaxial strain was applied using a MechanoCulture FX (CellScale) at 1 Hz and 10% strain for 12 hours a day. Cell morphology and alignment were evaluated by fluorescein isothiocyanate (FITC) labelled phalloidin and 4’,6-diamidino-2-phenylindole (DAPI) staining. Extracellular matrix composition was evaluated by immunocytochemistry. Cell phenotype maintenance/differentiation (tenogenic, chondrogenic and osteogenic lineages) were assessed by gene and protein analysis. After 12 hours of exposure to the uniaxial load, permanently differentiated cells are strictly aligned in the direction perpendicular to the load while the MSCs do not show preferential alignment. ECM deposition (e.g. collagens type I, III, V, VI) is increased in the presence of MMC and this effect is maintained under mechanical loading. ECM deposited under mechanical loading is also aligned in the direction perpendicular to the load. Tenogenic, osteogenic and chondrogenic markers are being tested to assess cell phenotype. Mechanical loading and macromolecular crowding can induce cell and ECM alignment and increased ECM deposition without affecting cell metabolic activity or viability. Cell and ECM alignment alongside ECM composition and tenogenic marker expression suggest this approach might be suitable to maintain or differentiate towards tenogenic lineage. Cell/Tissue: tenocytesbone marrow mesenchymal stem cells dermal fibroblasts
MechanoCulture FX	2018	Stretching Vascular Smooth Muscle Cells on Micropatterned Surfaces	Clemson University Thesis	S. N. Bradley	Clemson University Thesis		Vascular smooth muscle cells regulate blood flow by contracting and relaxing blood vessels. Vascular smooth muscle cells display one of two distinct phenotypes: contractile or synthetic. The contractile phenotype enables cells to contract, and the majority of cells in healthy blood vessels exhibit this phenotype. Transition to the synthetic phenotype is associated with abnormal mechanical forces and the development of vascular diseases, including hypertension and atherosclerosis. Cell-extracellular matrix interactions and mechanical stimulation are factors that affect phenotypic modulation. Micropatterning techniques create a microenvironment used to control cell morphology, adhesion, migration, proliferation, and differentiation. Vascular smooth muscle cells are circumferentially arranged in blood vessels, so parallel cell alignment mimics the native environment. Micropatterning vascular smooth muscle cells, either onto topographical features or extracellular matrix protein patterns, has been shown to promote a more contractile, in vivo-like phenotype. Vascular smooth muscle cells are exposed to continuous mechanical signaling from pulsatile blood flow, which enforces the contractile phenotype. Applying a physiologically relevant, cyclic stretching regimen to vascular smooth muscle cells also promotes contractile functioning. The purpose of this study was to determine the effects of micropatterning and mechanical stimulation on vascular smooth muscle cell phenotype. Three micropatterning techniques were implemented to determine the best method for aligning vascular smooth muscle cells. Microstamping was effective at patterning collagen but did not promote sufficient cell alignment. Stencil patterning vascular smooth muscle cells on a collagen layer was found to promote cell alignment with minimal cell spreading. Stenciled cells were exposed to physiological cyclic stretch using the MechanoCulture FX mechanical stimulation system (CellScale). The combination of stencil patterning and stretching reduced cell proliferation and promoted a more contractile phenotype.
MechanoCulture FX	2019	The Effect of Cyclic Strain on Human Fibroblasts with Lamin A/C Mutations and Its Relation to Heart Disease	Journal of Biomechanical Engineering	R. Tran, M. Siemens, C. Nguyen, A. Ochs, M. Zaragoza, A. Grosberg	University of California, Irvine		Although mutations in the Lamin A/C gene (LMNA) cause a variety of devastating diseases, the pathological mechanism is often unknown. Lamin A/C proteins play a crucial role in forming a meshwork under the nuclear membrane, providing the nucleus with mechanical integrity and interacting with other proteins for gene regulation. Most LMNA mutations result in heart diseases, including some types that primarily have heart disease as the main pathology. In this study, we used cells from patients with different LMNA mutations that primarily lead to heart disease. Indeed, it is a mystery why a mutation to the protein in every nucleus of the body manifests as a disease of primarily the heart in these patients. Here, we aimed to investigate if strains mimicking those within the myocardial environment are sufficient to cause differences in cells with and without the LMNA mutation. To test this, a stretcher device was used to induce cyclic strain upon cells, and viability/proliferation, cytoskeleton and extracellular matrix organization, and nuclear morphology were quantified. The properties of cells with HutchinsonGilford progeria syndrome were found to be significantly different from all other cell lines and were mostly in line with previous findings. However, the properties of cells from patients that primarily had heart diseases were not drastically different when compared to individuals without the LMNA mutation. Our results indicated that cyclic strain alone was insufficient to cause any significant differences that could explain the mechanisms that lead to heart diseases in these patients with LMNA mutations.
MechanoCulture FX	2019	Dynamic Stretching of Fibrillar Collagen Enhances Cross-linking by Transglutaminas	AOFAS	N. Shealy, J. Rex, A. Bradshaw, C. Gross	Medical U of South Carolina		New approaches to improve tendon repair after injury are an active area of research. Critical properties of tendons are governed by the production and assembly of fibrillar collagens. Cross-linking of fibrillar collagen is a primary factor in determining the function and mechanical properties of the collagen fibers comprising Enzymatic cross-linking by lysyl oxidase in the telopeptide domain of collagen I and III is one determinant of collagen fibril assembly and is the best characterized biochemical cross-link. Transglutaminase catalyzes the modification of lysine residues that in turn form an n-e-glutamyl lysine bond between proteins in the extracellular space. We hypothesize that transglutaminase-dependent modification of collagen in tendons is also a principal determinant of tendon strength and function and is dependent upon tension.
MechanoCulture T6	2016	Additive Manufacturing Of A Photo-Cross-Linkable Polymer Via Direct Meld Electrospinning Writing For Producing High Strength Structures	Biomacromolecules	F. Chen, G. Hochleitner, T. Woodfield, J. Groll, P. Dalton, B.G. Amsden	Queen's University		Melt electrospinning writing (MEW) is an emerging additive manufacturing technique that enables the design and fabrication of micrometer-thin fibrous scaffolds made of biocompatible and biodegradable polymers. By using a computer-aided deposition process, a unique control over pore size and interconnectivity of the resulting scaffolds is achieved, features highly interesting for tissue engineering applications. However, MEW has been mainly used to process low melting point thermoplastics such as poly(ε-caprolactone). Since this polymer exhibits creep and a reduction in modulus upon hydration, we manufactured scaffolds of poly(l-lactide-co-ε-caprolactone-co-acryloyl carbonate) (poly(LLA-ε-CL-AC)), a photo-cross-linkable and biodegradable polymer, for the first time. We show that the stiffness of the scaffolds increases significantly (up to ∼10-fold) after cross-linking by UV irradiation at room temperature, compared with un-cross-linked microfiber scaffolds. The preservation of stiffness and high average fiber modulus (370 ± 166 MPa) within the cross-linked hydrated scaffolds upon repetitive loading (10% strain at 1 Hz up to 200,000 cycles) suggests that the prepared scaffolds may be of potential interest for soft connective tissue engineering applications. Moreover, the approach can be readily adapted through manipulation of polymer properties and scaffold geometry to prepare structures with mechanical properties suitable for other tissue engineering applications. Cell/Tissue: poly(l-lactide-co-ε-caprolactone-co-acryloyl carbonate) (poly(LLA-ε-CL-AC)) polymerpoly(ε-caprolactone)
MechanoCulture T6	2017	Living Nanofiber Yarn-Based Woven Biotextiles For Tendon Tissue Engineering Using Cell Tri-Culture And Mechanical Stimulation	Acta Biomaterialia	S. Wu, Y. Wang, P.N. Streubel, B. Duan	University of Nebraska		Non-woven nanofibrous scaffolds have been developed for tendon graft application by using electrospinning strategies. However, electrospun nanofibrous scaffolds face some obstacles and limitations, including suboptimal scaffold structure, weak tensile and suture-retention strengths, and compact structure for cell infiltration. In this work, a novel nanofibrous, woven biotextile, fabricated based on electrospun nanofiber yarns, was implemented as a tissue engineered tendon scaffold. Based on our modified electrospinning setup, polycaprolactone (PCL) nanofiber yarns were fabricated with reproducible quality, and were further processed into plain-weaving fabrics interlaced with polylactic acid (PLA) multifilaments. Nonwoven nanofibrous PCL meshes with random or aligned fiber structures were generated using typical electrospinning as comparative counterparts. The woven fabrics contained 3 D aligned microstructures with significantly larger pore size and obviously enhanced tensile mechanical properties than their nonwoven counterparts. The biological results revealed that cell proliferation and infiltration, along with the expression of tendon-specific genes by human adipose derived mesenchymal stem cells (HADMSC) and human tenocytes (HT), were significantly enhanced on the woven fabrics compared with those on randomly-oriented or aligned nanofiber meshes. Co-cultures of HADMSC with HT or human umbilical vein endothelial cells (HUVEC) on woven fabrics significantly upregulated the functional expression of most tenogenic markers. HADMSC/HT/HUVEC tri-culture on woven fabrics showed the highest upregulation of most tendon-associated markers than all the other mono- and co-culture groups. Furthermore, we conditioned the tri-cultured constructs with dynamic conditioning and demonstrated that dynamic stretch promoted total collagen secretion and tenogenic differentiation. Our nanofiber yarn-based biotextiles have significant potential to be used as engineered scaffolds to synergize the multiple cell interaction and mechanical stimulation for promoting tendon regeneration. Cell/Tissue: adipose derived mesenchymal stem cell stenocytesumbilical vein endothelial cells
MechanoCulture T6	2018	Mechanotransduction And Control Of Valvular Cell Phenotype As Tools To Inform Valvular Pathophysiology	Dissertation	Mir Ali	Texas Tech Uni
MechanoCulture T6	2019	Multifactorial Bottom-Up Bioengineering Approaches For The Development Of Living Tissues Substitutes	FASEB	D. Gaspar, Christina N. M. Ryan, Dimitrios I. Zeugolis	National University of Ireland		Bottom-up bioengineering utilizes the inherent capacity of cells to build highly sophisticated structures with high levels of biomimicry. Despite the significant advancements in the field, monodomain approaches require prolonged culture time to develop an implantable device, usually associated with cell phenotypic drift in culture. Herein, we assessed the simultaneous effect of macromolecular crowding (MMC) and mechanical loading in enhancing extracellular matrix (ECM) deposition while maintaining tenocyte (TC) phenotype and differentiating bone marrow stem cells (BMSCs) or transdifferentiating neonatal and adult dermal fibroblasts toward tenogenic lineage. At d 7, all cell types presented cytoskeleton alignment perpendicular to the applied load independently of the use of MMC. MMC enhanced ECM deposition in all cell types. Gene expression analysis indicated that MMC and mechanical loading maintained TC phenotype, whereas tenogenic differentiation of BMSCs or transdifferentiation of dermal fibroblasts was not achieved. Our data suggest that multifactorial bottom-up bioengineering approaches significantly accelerate the development of biomimetic tissue equivalents.—Gaspar, D., Ryan, C. N. M., Zeugolis, D. I. Multifactorial bottom-up bioengineering approaches for the development of living tissue substitutes.
MechanoCulture T6	2019	The Effect Of Physiological Stretch And The Valvular Endothelium On Mitral Valve Proteomes	Experimental Biology and Medicine	M. S. Ali, X. Wang, C. M. R. Lacerda	Texas Tech Uni		Heart valves reside in a dynamic mechanical environment experiencing a multitude of forces. These forces have been shown to play a key role in valvular pathophysiology. However, knowledge of proteins involved in valvular mechanobiology is limited to only a few proteins of interest, namely, α-smooth muscle actin, transforming growth factor β, etc. Valvular endothelium mediates valvular homeostasis and controls valvular interstitial cell phenotype transformation. But, how endothelium mediates valvular response to dynamic forces is also unknown. In this study, proteomic analysis of mitral valve anterior leaflets under 10% cyclic radial strain was performed. Endothelium from these samples was removed to test how endothelium mediates mitral valve response to stretch. Results show that stretch downregulated cytoskeletal proteins and proteins involved in energy metabolism such as glycolysis and oxireductase activity. Endothelium removal resulted in downregulation of extracellular matrix and cell-matrix adhesion proteins. Removal of endothelium also resulted in upregulation of translation-related and chaperone proteins. Overall, this high throughput study provides insights into new protein groups that may be involved in mitral valve response to mechanical stretch and loss of endothelium.
MechanoCulture T6	2019	Effect Of Topography And Physical Stimulus On Hmsc Phenotype Using A 3d In Vitro Model	Nanomaterials	D. Kumar, S. A. Cain, L. A. Bosworth	University of Manchester		This communication reports the first comparative study addressing the effects of both structural architecture and mechanical loading on human mesenchymal stem cells (hMSC) positioned at the interface of a 3D in vitro model composed of a nanofibre/hydrogel laminate composite. hMSC phenotype was affected by both stimuli over a seven-day period. Cells were orientated parallel to the underlying fibre direction irrespective of environment (electrospun 2D fibre sheet or laminate 2D sheet with collagen gel layer). Application of cyclical tensile force (5% strain, 1 Hz, 1 h per day) encouraged hMSCs to remain at the fibre/gel interface, whereas cells cultured in static conditions migrated from the interface to the upper hydrogel layer. Depending on the stimulus applied, hMSCs presented an up-regulation in gene expression, indicative of several cell lineages, with those cultured at the interface and physically stimulated expressing markers indicative of angiogenesis, osteogenesis, and tenogenesis. This study highlights the importance of developing biomaterial scaffolds with environmental cues to specifically drive cells towards the tissue intended for bioengineering.
MechanoCulture T6	2019	Methods of Delivering Mechanical Stimuli to Organ-on-a-Chip	Micromachines	K. Kaarj, J-Y., Yoon	University of Arizona		Recent advances in integrating microengineering and tissue engineering have enabled the creation of promising microengineered physiological models, known as organ-on-a-chip (OOC), for experimental medicine and pharmaceutical research. OOCs have been used to recapitulate the physiologically critical features of specific human tissues and organs and their interactions. Application of chemical and mechanical stimuli is critical for tissue development and behavior, and they were also applied to OOC systems. Mechanical stimuli applied to tissues and organs are quite complex in vivo, which have not adequately recapitulated in OOCs. Due to the recent advancement of microengineering, more complicated and physiologically relevant mechanical stimuli are being introduced to OOC systems, and this is the right time to assess the published literature on this topic, especially focusing on the technical details of device design and equipment used. We first discuss the different types of mechanical stimuli applied to OOC systems: shear flow, compression, and stretch/strain. This is followed by the examples of mechanical stimuli-incorporated OOC systems. Finally, we discuss the potential OOC systems where various types of mechanical stimuli can be applied to a single OOC device, as a better, physiologically relevant recapitulation model, towards studying and evaluating experimental medicine, human disease modeling, drug development, and toxicology.
MechanoCulture T6	2019	In vitro degradation of electrospun polycaprolactone tissue engineering scaffolds under cyclical dynamic loading	Poster	J. Bracamonte, S. Saunders, S. Cole, G. Annohene, G. Tepper, J. Soares	Virginia Commonwealth U
MechanoCulture TR	2019	Hierarchical Electrospun Tendon-ligament Bioinspired Scaffolds Induce Changes in Fibroblasts Morphology under Static and Dynamic Conditions	Microscopy	A. Sensini, L. Christofolini, A. Zucchelli, M. L. Focarete, C. Gualandi, A. De Mori, A. P. Kao, M. Roldo, G. Blunn, G. Tozzi	University of Bologna		The regeneration of injured tendons and ligaments is challenging because the scaffolds needs proper mechanical properties and a biomimetic morphology. In particular, the morphological arrangement of scaffolds is a key point to drive the cells growth to properly regenerate the collagen extracellular matrix. Electrospinning is a promising technique to produce hierarchically structured nanofibrous scaffolds able to guide cells in the regeneration of the injured tissue. Moreover, the dynamic stretching in bioreactors of electrospun scaffolds had demonstrated to speed up cell shape modifications in vitro. The aim of the present study was to combine different imaging techniques such as high-resolution X-ray tomography (XCT), scanning electron microscopy (SEM), fluorescence microscopy and histology to investigate if hierarchically structured poly (L-lactic acid) and collagen electrospun scaffolds can induce morphological modifications in human fibroblasts, while cultured in static and dynamic conditions. After 7 days of parallel cultures, the results assessed that fibroblasts had proliferated on the external nanofibrous sheath of the static scaffolds, elongating themselves circumferentially. The dynamic cultures revealed a preferential axial orientation of fibroblasts growth on the external sheath. The aligned nanofibre bundles inside the hierarchical scaffolds instead, allowed a physiological distribution of the fibroblasts along the nanofibre direction. Inside the dynamic scaffolds, cells appeared thinner compared with the static counterpart. This study had demonstrated that hierarchically structured electrospun scaffolds can induce different fibroblasts morphological modifications during static and dynamic conditions, modifying their shape in the direction of the applied loads. LAY DESCRIPTION: To enhance the regeneration of injured tendons and ligaments cells need to growth on dedicated structures (scaffolds) with mechanical properties and a fibrous morphology similar to the natural tissue. In particular, the morphological organisation of scaffolds is fundamental in leading cells to colonise them, regenerating the collagen extracellular matrix. Electrospinning is a promising technique to produce fibres with a similar to the human collagen fibres, suitable to design complex scaffolds able to guide cells in the reconstruction of the natural tissue. Moreover, it is well established that the cyclic stretching of these scaffolds inside dedicated systems called bioreactors, can speed up cells growth and their shape modification. The aim of the present study was to investigate how hierarchically structured electrospun scaffolds, made of resorbable material such as poly(L-lactic acid) and collagen, could induce morphological changes in human fibroblasts, while cultured during static and dynamic conditions. These scaffolds were composed by an external electrospun membrane that grouped inside it a ring-shaped bundle, made of axially aligned nanofibres, resembling the morphological arrangement of tendon and ligament tissue. After 7 days of parallel cultures, the scaffolds were investigated using the following imaging techniques: (i) high-resolution X-ray tomography (XCT); (ii) scanning electron microscopy (SEM); (iii) fluorescence microscopy and (iv) histology. The results showed that fibroblasts were able to grow on the external nanofibrous sheath of the static scaffolds, by elongating themselves along their circumference. The dynamic cultures revealed instead a preferential axial orientation of fibroblasts grown on the external sheath. The aligned nanofibre bundles inside the hierarchical scaffolds allowed an axial distribution of the fibroblasts along the nanofibres direction. This study has demonstrated that the electrospun hierarchically structured scaffolds investigated can modify the fibroblasts morphology both in static and dynamic conditions, in relation with the direction of the applied loads.
MechanoCulture TR	2019	Advanced In-vitro Models with Integrated Sensing for Real-time Monitoring of Electrical and Mechanical Properties of Cellular Constructs	University of Pisa	C. Ludovica	University of Pisa
MechanoCulture TR	2020	A New Load-controlled Testing Method for Viscoelastic Characterisation Through Stress-rate Measurements	Materialia	C. Ludovica, G. Mattei, A. Ahluwalia	University of Pisa		Viscoelastic or time-dependent behaviour is typical of several materials, including biological tissues. Thus, several testing methods have been developed to characterise material viscoelastic properties. In this study, we propose a new load-controlled testing approach, called the sigma-dot method (σ˙M), based on measurements at different constant stress-rates. The method can be implemented with either standard load-controlled testing devices or bioreactors with load-controlled mechano-actuation to perform in-situ viscoelastic characterisation during cell culture. Both polydimethylsiloxane and hydroxyapatite/gelatin composites were tested using the σ˙M, by performing measurements with the MechanoCulture TR- a load-controlled bioreactor - and a universal testing machine as control. The results show that the proposed method is effective for deriving sample viscoelastic parameters, independent of the testing device. The main advantage of the σ˙M with respect to classical step-response viscoelastic tests, such as creep and stress-relaxation, is its ramp stress input which is physically implementable and does not need any a priori determination of the sample linear viscoelastic region.
MicroTester/MicroSquisher	2009	Cellular Interfacial And Surface Tensions Determined From Aggregate Compression Tests Using A Finite Element Model. Hfsp Journal	Frontiers in Life Science	Brodland, G.W., Yang, J., Sweny, J.	University of Waterloo		Although previous studies suggested that the interfacial tension γ cc acting along cell‐cell boundaries and the effective viscosity μ of the cell cytoplasm could be measured by compressing a spherical aggregate of cells between parallel plates, the mechanical understanding necessary to extract this information from these tests—tests that have provided the surface tension σcm acting along cell‐medium interfaces—has been lacking. These tensions can produce net forces at the subcellular level and give rise to cell motions and tissue reorganization, the rates of which are regulated by μ. Here, a three‐dimensional (3D) cell‐based finite element model provides insight into the mechanics of the compression test, where these same forces are at work, and leads to quantitative relationships from which the effective viscosity μ of the cell cytoplasm, the tension γ cc that acts along internal cell‐cell interfaces and the surface tension σcp along the cell‐platen boundaries can be determined from force‐time curves and aggregate profiles. Tests on 5‐day embryonic chick mesencephalon, neural retina, liver, and heart aggregates show that all of these properties vary significantly with cell type, except γcc, which is remarkably constant. These properties are crucial for understanding cell rearrangement and tissue self‐organization in contexts that include embryogenesis, cancer metastases, and tissue engineering. Cell/Tissue: embryonic mesencephalon embryonic neural retina embryonic liver and heart
MicroTester/MicroSquisher	2012	Stiffening Of Human Mesenchymal Stem Cell Spheroid Mircoenvironments Induced By Incorporation Of Gelatin Microparticles	Journal of the mechanical behavior of biomedical materials	P. R. Baraniak, M.T. Cooke, R. Saeed, M.A. Kinney, K.M. Fridley, T.C. mcDevitt	Georgia Institute of Technology and Emory University		Culturing multipotent adult mesenchymal stem cells as 3D aggregates augments their differentiation potential and paracrine activity. One caveat of stem cell spheroids, though, can be the limited diffusional transport barriers posed by the inherent 3D structure of the multicellular aggregates. In order to circumvent such limitations, polymeric microparticles have been incorporated into stem cell aggregates as a means to locally control the biochemical and physical properties of the 3D microenvironment. However, the introduction of biomaterials to the 3D stem cell microenvironment could alter the mechanical forces sensed by cells within aggregates, which in turn could impact various cell behaviors and overall spheroid mechanics. Therefore, the objective of this study was to determine the acute effects of biomaterial incorporation within mesenchymal stem cell spheroids on aggregate structure and mechanical properties. The results of this study demonstrate that although gelatin microparticle incorporation results in similar multi-cellular organization within human mesenchymal stem cell spheroids, the introduction of gelatin materials significantly impacts spheroid mechanical properties. The marked differences in spheroid mechanics induced by microparticle incorporation may hold major implications for in vitro directed differentiation strategies and offer a novel route to engineer the mechanical properties of tissue constructs ex vivo. Cell/Tissue: mesenchymal stem cell spheroids
MicroTester/MicroSquisher	2013	Alginate Encapsulation Parameters Influence The Differentiation Of Microencapsulated Embryonic Stem Cell Aggregates	Biotechnology and Bioengineering	Wilson, J.L., Ali Naijia, M., Saeed, R., McDevitt, T.C.	Georgia Institute of Technology and Emory University		Pluripotent embryonic stem cells (ESCs) have tremendous potential as tools for regenerative medicine and drug discovery, yet the lack of processes to manufacture viable and homogenous cell populations of sufficient numbers limits the clinical translation of current and future cell therapies. Microencapsulation of ESCs within microbeads can shield cells from hydrodynamic shear forces found in bioreactor environments while allowing for sufficient diffusion of nutrients and oxygen through the encapsulation material. Despite initial studies examining alginate microbeads as a platform for stem cell expansion and directed differentiation, the impact of alginate encapsulation parameters on stem cell phenotype has not been thoroughly investigated. Therefore, the objective of this study was to systematically examine the effects of varying alginate compositions on microencapsulated ESC expansion and phenotype. Pre-formed aggregates of murine ESCs were encapsulated in alginate microbeads composed of a high or low ratio of guluronic to mannuronic acid residues (High G and High M, respectively), with and without a poly-L-lysine (PLL) coating, thereby providing four distinct alginate bead compositions for analysis. Encapsulation in all alginate compositions was found to delay differentiation, with encapsulation within High G alginate yielding the least differentiated cell population. The addition of a PLL coating to the High G alginate prevented cell escape from beads for up to 14 days. Furthermore, encapsulation within High M alginate promoted differentiation toward a primitive endoderm phenotype. Taken together, the findings of this study suggest that distinct ESC expansion capacities and differentiation trajectories emerge depending on the alginate composition employed, indicating that encapsulation material physical properties can be used to control stem cell fate. Cell/Tissue: murine embryonic stem cells
MicroTester/MicroSquisher	2014	Mesenchymal Morphogenesis Of Embryonic Stem Cells Dynamically Modulates The Biophysical Microtissue Niche	Scientific Reports	M.A. Kinney, R.Saeed, T.C. McDevitt	Georgia Institute of Technology and Emory University		Stem cell fate and function are dynamically modulated by the interdependent relationships between biochemical and biophysical signals constituting the local 3D microenvironment. While approaches to recapitulate the stem cell niche have been explored for directing stem cell differentiation, a quantitative relationship between embryonic stem cell (ESC) morphogenesis and intrinsic biophysical cues within three-dimensional microtissues has not been established. In this study, we demonstrate that mesenchymal embryonic microtissues induced by BMP4 exhibited increased stiffness and viscosity accompanying differentiation, with cytoskeletal tension significantly contributing to multicellular stiffness. Perturbation of the cytoskeleton during ESC differentiation led to modulation of the biomechanical and gene expression profiles, with the resulting cell phenotype and biophysical properties being highly correlated by multivariate analyses. Together, this study elucidates the dynamics of biophysical and biochemical signatures within embryonic microenvironments, with broad implications for monitoring tissue dynamics, modeling pathophysiological and embryonic morphogenesis and directing stem cell patterning and differentiation. Cell/Tissue: mesenchymal embryonic microtissues embryonic stem cells
MicroTester/MicroSquisher	2015	Burr-Like, Laser-Made 3d Microscaffolds For Tissue Spheroid Encagement	Biointerphases	P. Danilevicius, R.A. Rezende, F.D.A.S. Pereira, A. Selimis, V. Kasyanov, P.Y. Noritomi, J.V.L. da Silva, M.Chatzinikolaidou, M. Farsari, V. Mironov	University of Crete		The modeling, fabrication, cell loading, and mechanical and in vitro biological testing of biomimetic, interlockable, laser-made, concentric 3D scaffolds are presented. The scaffolds are made by multiphoton polymerization of an organic–inorganic zirconium silicate. Their mechanical properties are theoretically modeled using finite elements analysis and experimentally measured using a MicroTester/MicroSquisher®. They are subsequently loaded with preosteoblastic cells, which remain live after 24 and 72 h. The interlockable scaffolds have maintained their ability to fuse with tissue spheroids. This work represents a novel technological platform, enabling the rapid, laser-based, in situ 3D tissue biofabrication. Cell/Tissue: tissue spheroids preosteaoblastic cells
MicroTester/MicroSquisher	2016	An Injectable Hydrogel Prepared Using A Peg/Vitamin E Copolymer Facilitating Aqueous-Driven Gelation	Biomacromolecules	J. Zhang, B. Muirhead, M. Dodd, L. Liu, N. Mangiacotte, T. Hoare, S. Sheardown	McMaster University		Hydrogels have been widely explored for biomedical applications, with injectable hydrogels being of particular interest for their ability to precisely deliver drugs and cells to targets. Although these hydrogels have demonstrated satisfactory properties in many cases, challenges still remain for commercialization. In this paper, we describe a simple injectable hydrogel based on poly(ethylene glycol) (PEG) and a vitamin E (Ve) methacrylate copolymer prepared via simple free radical polymerization and delivered in a solution of low molecular weight PEG and Ve as the solvent instead of water. The hydrogel formed immediately in an aqueous environment with a controllable gelation time. The driving force for gelation is attributed to the self-assembly of hydrophobic Ve residues upon exposure to water to form a physically cross-linked polymer network via polymer chain rearrangement and subsequent phase separation, a spontaneous process with water uptake. The hydrogels can be customized to give the desired water content, mechanical strength, and drug release kinetics simply by formulating the PEGMA-co-Ve polymer with an appropriate solvent mixture or by varying the molecular weight of the polymer. The hydrogels exhibited no significant cytotoxicity in vitro using fibroblasts and good tissue compatibility in the eye and when injected subcutaneously. These polymers thus have the potential to be used in a variety of applications where injection of a drug or cell containing depot would be desirable. Cell/Tissue: poly(ethylene glycol) hydrogelvitamin E methacrylate copolymer hydrogeleye tissue fibroblast
MicroTester/MicroSquisher	2016	Delivery Of Human Adipose Stem Cells Spheroids Into Lockyballs	PLOS One (Public Library of Science One)	K.R. Silva, R.A. Rezende, F.D.A.S. Pereira, P. Gruber, M.P. Stuart, A. Ovsianikov, K. Brakke, V. Kasyanov, J.V.L. da Silva, J.M. Granjeiro, L.S. Baptista, V. Mironov	Federal University of Rio de Janeiro-Xerém		Adipose stem cells (ASCs) spheroids show enhanced regenerative effects compared to single cells. Also, spheroids have been recently introduced as building blocks in directed self-assembly strategy. Recent efforts aim to improve long-term cell retention and integration by the use of microencapsulation delivery systems that can rapidly integrate in the implantation site. Interlockable solid synthetic microscaffolds, so called lockyballs, were recently designed with hooks and loops to enhance cell retention and integration at the implantation site as well as to support spheroids aggregation after transplantation. Here we present an efficient methodology for human ASCs spheroids biofabrication and lockyballs cellularization using micro-molded non-adhesive agarose hydrogel. Lockyballs were produced using two-photon polymerization with an estimated mechanical strength. The Young’s modulus was calculated at level 0.1362 +/-0.009 MPa. Interlocking in vitro test demonstrates high level of loading induced interlockability of fabricated lockyballs. Diameter measurements and elongation coefficient calculation revealed that human ASCs spheroids biofabricated in resections of micro-molded non-adhesive hydrogel had a more regular size distribution and shape than spheroids biofabricated in hanging drops. Cellularization of lockyballs using human ASCs spheroids did not alter the level of cells viability (p › 0,999) and gene fold expression for SOX-9 and RUNX2 (p › 0,195). The biofabrication of ASCs spheroids into lockyballs represents an innovative strategy in regenerative medicine, which combines solid scaffold-based and directed self-assembly approaches, fostering opportunities for rapid in situ biofabrication of 3D building-blocks. Cell/Tissue: adipose stem cell spheroids lockyballs
MicroTester/MicroSquisher	2016	Direct Production Of Human Cardiac Tissues By Pluripotent Stem Cell Encapsulation In Gelatin Methacryloyl	ACS Biomaterials Science and Engineering (American Chemical Society Biomaterials Science and Engineering)	P. Kerscher, J.A. Kaczmarek, S.E. Head, M. Brazel, W. Seeto, S. Bhattacharya, J. Kim, V. Suppiramaniam, E.A. Lipke	Auburn University		Direct stem cell encapsulation and cardiac differentiation within supporting biomaterial scaffolds are critical for reproducible and scalable production of the functional human tissues needed in regenerative medicine and drug-testing applications. Producing cardiac tissues directly from pluripotent stem cells rather than assembling tissues using pre-differentiated cells can eliminate multiple cell-handling steps that otherwise limit the potential for process automation and production scale-up. Here we asked whether our process for forming 3D developing human engineered cardiac tissues using poly(ethylene glycol)-fibrinogen hydrogels can be extended to widely used and printable gelatin methacryloyl (GelMA) hydrogels. We demonstrate that low-density GelMA hydrogels can be formed rapidly using visible light (<1 min) and successfully employed to encapsulate human induced pluripotent stem cells while maintaining high cell viability. Resulting constructs had an initial stiffness of approximately 220 Pa, supported tissue growth and dynamic remodeling, and facilitated high-efficiency cardiac differentiation (>70%) to produce spontaneously contracting GelMA human engineered cardiac tissues (GEhECTs). GEhECTs initiated spontaneous contractions on day 8 of differentiation, with synchronicity, frequency, and velocity of contraction increasing over time, and displayed developmentally appropriate temporal changes in cardiac gene expression. GEhECT-dissociated cardiomyocytes displayed well-defined and aligned sarcomeres spaced at 1.85 ± 0.1 μm and responded appropriately to drug treatments, including the β-adrenergic agonist isoproterenol and antagonist propranolol, as well as to outside pacing up to 3.0 Hz. Overall results demonstrate that GelMA is a suitable biomaterial for the production of developing cardiac tissues and has the potential to be employed in scale-up production and bioprinting of GEhECTs. Cell/Tissue: cardiac tissues induced pluripotent stem cells
MicroTester/MicroSquisher	2016	Mineral Particles Modulate Osteo-Chondrogenic Differentiation Of Embryonic Stem Cell Aggregates	Acta Biomaterialia	Y. Wang, X. Yu, C. Baker, W.L. Murphy, T.C. McDevitt	University of California		Pluripotent stem cell aggregates offer an attractive approach to emulate embryonic morphogenesis and skeletal development. Calcium phosphate (CaP) based biomaterials have been shown to promote bone healing due to their osteoconductive and potential osteoinductive properties. In this study, we hypothesized that incorporation of CaP-coated hydroxyapatite mineral particles (MPs) within murine embryonic stem cell (ESC) aggregates could promote osteo-chondrogenic differentiation. Our results demonstrated that MP alone dose-dependently promoted the gene expression of chondrogenic and early osteogenic markers. In combination with soluble osteoinductive cues, MPs enhanced the hypertrophic and osteogenic phenotype, and mineralization of ESC aggregates. Additionally, MPs dose-dependently reduced ESC pluripotency and thereby decreased the size of teratomas derived from MP-incorporated ESC aggregates in vivo. Our data suggested a novel yet simple means of using mineral particles to control stem cell fate and create an osteochondral niche for skeletal tissue engineering applications. Cell/Tissue: murine embryonic stem cells
MicroTester/MicroSquisher	2016	Peg-Fibrinogen Hydrogels For Three-Dimensional Breast Cancer Cell Culture	Journal of Biomedical Materials Research	S. Pradhan, I. Hassani, W.J. Seeto, E. A. Lipke	Auburn University		Tissue-engineered three-dimensional (3D) cancer models employing biomimetic hydrogels as cellular scaffolds provide contextual in vitro recapitulation of the native tumor microenvironment, thereby improving their relevance for use in cancer research. This study reports the use of poly(ethylene glycol)-fibrinogen (PF) as a suitable biosynthetic hydrogel for the 3D culture of three breast cancer cell lines: MCF7, SK-BR-3, and MDA-MB-231. Modification of the matrix characteristics of PF hydrogels was achieved by addition of excess poly(ethylene glycol) diacrylate, which resulted in differences in Young's moduli, degradation behavior, release kinetics, and ultrastructural variations in scaffold microarchitecture. Cancer cells were maintained in 3D culture with high viability within these hydrogels and resulted in cell-type dependent morphological changes over time. Cell proliferation and 3D morphology within the hydrogels were visualized through immunofluorescence staining. Finally, spatial heterogeneity of colony area within the hydrogels was quantified, with peripheral cells forming colonies of higher area compared to those in the interior regions. Overall, PF-based hydrogels facilitate 3D culture of breast cancer cells and investigation of cellular behavior in response to varying matrix characteristics. PF-based cancer models could be potentially used in future investigations of cancer biology and in anti-cancer drug-testing applications. Cell/Tissue: poly(ethylene glycol)-fibrinogen (PF)biosynthetic hydrogel breast cancer cells
MicroTester/MicroSquisher	2016	Reactive Eletrospinning Of Degradable Poly(Oligoethylene Glycol Methacrylate)-Based Nanofibrous Hydrogel Networks	Chemical Communications	F. Xu, H. Sheardown, T. Hoare	McMaster University		A direct, all-aqueous electrospinning method for fabricating degradable nanofibrous hydrogel networks is reported in which hydrazide and aldehyde-functionalized poly(oligoethylene glycol methacrylate) (POEGMA) polymers are simultaneously electrospun and cross-linked. The resulting networks are spatially well-defined, mechanically stable (both dry and wet), and offer extremely fast swelling responses, suggesting potential utility as smart hydrogels and tunable tissue engineering matrices.Cell/Tissue: nanofibrous hydrogelpoly(oligoethylene glycol methacrylate) (POEGMA) polymers
MicroTester/MicroSquisher	2017	A Three-Dimensional Spheroidal Cancer Model Based On Peg-Fibrinogen Hydrogel Microspheres	Biomaterials	S.Pradhan, J. M. Clary, D. Seliktar, E. A. Lipke	Auburn University		Three-dimensional (3D) in vitro cancer models offer an attractive approach towards the investigation of tumorigenic phenomena and other cancer studies by providing dimensional context and higher degree of physiological relevance than that offered by conventional two-dimensional (2D) models. The multicellular tumor spheroid model, formed by cell aggregation, is considered to be the “gold standard” for 3D cancer models, due to its ease and simplicity of use. Although better than 2D models, tumor spheroids are unable to replicate key features of the native tumor microenvironment, particularly due to a lack of surrounding extracellular matrix components and heterogeneity in shape, size and aggregate forming tendencies. In order to address this issue, we have developed a 3D “tumor microsphere” model, formed by a dual-photoinitiator, aqueous-oil emulsion technique, for the encapsulation of cancer cells within PEG-fibrinogen hydrogel microspheres and for subsequent long-term 3D culture. In comparison to self-aggregated tumor spheroids, the tumor microspheres displayed a higher degree of size and shape homogeneity throughout long-term culture. In sharp contrast to cells in tumor spheroids, cells within tumor microspheres demonstrated significant loss in apico-basal polarity and cellular architecture, cellular and nuclear atypia, increased disorganization, elevated nuclear cytoplasmic ratio and nuclear volume density and reduction in cell-cell junction length, all of which are hallmarks of malignant transformation and tumorigenic progression. Additionally, the tumor microsphere model was extended for the 3D encapsulation and maintenance of a wide range of other cancer cell (metastatic and non-metastatic) types. Taken together, our results reinforce the importance of incorporating a biomimetic matrix in the cellular microenvironment of 3D tumor models and the influential effects of the matrix on the tumorigenic morphology of 3D cultured cells. The tumor microsphere system established in this study has the potential to be used in future investigations of 3D cancer cell-cell and cell-ECM interactions and in drug-testing applications. Cell/Tissue: tumorigenic cancer cells tumor microsphere
MicroTester/MicroSquisher	2017	Decellularized Adipose Tissue Microcarriers As A Dynamic Culture Platform For Human Adipose-Derived Stem/Stromal Cell Expansion	Biomaterials	C. Yu, A. Kornmuller, C. Brown, T. Hoare, L.E. Flynn	University of Western Ontario		With the goal of designing a clinically-relevant expansion strategy for human adipose-derived stem/stromal cells (ASCs), methods were developed to synthesize porous microcarriers derived purely from human decellularized adipose tissue (DAT). An electrospraying approach was applied to generate spherical DAT microcarriers with an average diameter of 428 ± 41 μm, which were soft, compliant, and stable in long-term culture without chemical crosslinking. Human ASCs demonstrated enhanced proliferation on the DAT microcarriers relative to commercially-sourced Cultispher-S microcarriers within a spinner culture system over 1 month. ASC immunophenotype was maintained post expansion, with a trend for reduced expression of the cell adhesion receptors CD73, CD105, and CD29 under dynamic conditions. Upregulation of the early lineage-specific genes PPARγ, LPL, and COMP was observed in the ASCs expanded on the DAT microcarriers, but the cells retained their multilineage differentiation capacity. Comparison of adipogenic and osteogenic differentiation in 2-D cultures prepared with ASCs pre-expanded on the DAT microcarriers or Cultispher-S microcarriers revealed similar adipogenic and enhanced osteogenic marker expression in the DAT microcarrier group, which had undergone a higher population fold change. Further, histological staining results suggested a more homogeneous differentiation response in the ASCs expanded on the DAT microcarriers as compared to either Cultispher-S microcarriers or tissue culture polystyrene. A pilot chondrogenesis study revealed higher levels of chondrogenic gene and protein expression in the ASCs expanded on the DAT microcarriers relative to all other groups, including the baseline controls. Overall, this study demonstrates the promise of applying dynamic culture with tissue-specific DAT microcarriers as a means of deriving regenerative cell populations. Cell/Tissue: adipose tissue adipose derived stem stromal
MicroTester/MicroSquisher	2017	Encapsulation Of Equine Endothelial Colony Forming Cells In Highly Uniform, Injectable Hydrogel Microspheres For Local Cell Delivery	Tissue Engineering Part C: Methods	W.J. Seeto, Y. Tian, R.L. Winter, F.J. Caldwell, A.A. Wooldridge, E.A. Lipke	Auburn University		A common challenge in cell therapy is the inability to routinely maintain survival and localization of injected therapeutic cells. Delivering cells by direct injection increases the flexibility of clinical applications, but may cause low cell viability and retention rates due to the high shear forces in the needle and mechanical wash out. In this study, we encapsulated endothelial colony forming cells (ECFCs) in poly(ethylene glycol)-fibrinogen (PF) hydrogel microspheres using a custom-built microfluidic device; this system supports rapid encapsulation of high cell concentrations (10 million cells per mL) and resulting cell-laden microspheres are highly uniform in shape and size. The encapsulated ECFCs were shown to have >95% viability and continued to rapidly proliferate. Expression of cell markers (von Willebrand factor, CD105, and CD14), the ability to form tubules on basement membrane matrix, and the ability to take up low-density lipoprotein were similar between pre- and post-encapsulated cells. Viability of encapsulated ECFCs was maintained after shear through 18–23-gauge needles. Ex vivo and in vivo cell delivery studies were performed by encapsulating and injecting autologous equine ECFCs subcutaneously into distal limb full-thickness wounds of adult horses. Injected ECFCs were visualized by labeling with fluorescent nanodots before encapsulation. One week after injection, confocal microscopy analysis of biopsies of the leading edges of the wounds showed that the encapsulated ECFCs migrated into the surrounding host tissue indicating successful retention and survival of the delivered ECFCs. Rapid, scalable cell encapsulation into PF microspheres was demonstrated to be practical for use in large animal cell therapy and is a clinically relevant method to maintain cell retention and survival after local injection. Cell/Tissue: endothelial colony forming cells
MicroTester/MicroSquisher	2017	Microfluidic Production Of Degradable Thermoresponsive Poly(N-Isopropylacrylamide)-Based Microgels	Soft Matter	D. Sivakumaran, E. Mueller, T. Hoare	McMaster University		Highly monodisperse and hydrolytically degradable thermoresponsive microgels on the tens-to-hundreds of micron size scale have been fabricated based on simultaneous on-chip mixing and emulsification of aldehyde and hydrazide-functionalized poly(N-isopropylacrylamide) precursor polymers. The microfluidic chip can run for extended periods without upstream gelation and can produce monodisperse (<10% particle size variability) microgels on the size range of ∼30–90 μm, with size tunable according to the flow rate of the oil continuous phase. Fluorescence analysis indicates a uniform distribution of each reactive pre-polymer inside the microgels while micromechanical testing suggests that smaller microfluidic-produced microgels exhibit significantly higher compressive moduli compared to bulk hydrogels of the same composition, an effect we attribute to improved mixing (and thus crosslinking) of the precursor polymer solutions within the microfluidic device. The microgels retain the reversible volume phase transition behavior of conventional microgels but can be hydrolytically degraded back into their oligomeric precursor polymer fragments, offering potential for microgel clearance following use in vivo. Cell/Tissue: aldehyde and hydrazide-functionalized poly(N-isopropylacrylamide) precursor polymers
MicroTester/MicroSquisher	2017	Polysaccharide Hydrogels Support The Long-Term Viability Of Encapsulated Human Mesenchymal Stem Cells And Their Ability To Secrete Immunomodulatory Factors	Stem Cells International	F. Hached, C. Vinatier, P-G. Pinta, P. Hulin, C. Le Visage, P. Weiss, J. Guicheux, A. Billon-Chabaud, G. Grimandi	Institut national de la santé et de la recherche médicale		While therapeutically interesting, the injection of MSCs suffers major limitations including cell death upon injection and a massive leakage outside the injection site. We proposed to entrap MSCs within spherical particles derived from alginate, as a control, or from silanized hydroxypropyl methylcellulose (Si-HPMC). We developed water in an oil dispersion method to produce small Si-HPMC particles with an average size of about 68 μm. We evidenced a faster diffusion of fluorescein isothiocyanate-dextran in Si-HPMC particles than in alginate ones. Human adipose-derived MSCs (hASC) were encapsulated either in alginate or in Si-HPMC, and the cellularized particles were cultured for up to 1 month. Both alginate and Si-HPMC particles supported cell survival, and the average number of encapsulated hASC per alginate and Si-HPMC particle (7102 and 5100, resp.) did not significantly change. The stimulation of encapsulated hASC with proinflammatory cytokines resulted in the production of IDO, PGE2, and HGF whose concentration was always higher when cells were encapsulated in Si-HPMC particles than in alginate ones. We have demonstrated that Si-HPMC and alginate particles support hASC viability and the maintenance of their ability to secrete therapeutic factors. Cell/Tissue: adipose-derived mesenchymal stem cells
MicroTester/MicroSquisher	2017	Pullulan Microbeads/Si-Hpmc Hydrogel Injectable System For The Sustained Delivery Of Gdf-5 And Tgf-Β1: New Insight Into Intervertebral Disc Regenerative Medicine	Drug Delivery	N. Henry, J. Clouet, A. Fragale, L.Griveau, C. Chedevile, J. Veziers, P. Weiss, J. Le Bideau, J. Guicheaux, C. Le Visage	Université de Nantes		Discogenic low back pain is considered a major health concern and no etiological treatments are today available to tackle this disease. To clinically address this issue at early stages, there is a rising interest in the stimulation of local cells by in situ injection of growth factors targeting intervertebral disc (IVD) degenerative process. Despite encouraging safety and tolerability results in clinic, growth factors efficacy may be further improved. To this end, the use of a delivery system allowing a sustained release, while protecting growth factors from degradation appears of particular interest. We propose herein the design of a new injectable biphasic system, based on the association of pullulan microbeads (PMBs) into a cellulose-based hydrogel (Si-HPMC), for the TGF-β1 and GDF-5 growth factors sustained delivery. We present for the first time the design and mechanical characterization of both the PMBs and the called biphasic system (PMBs/Si-HPMC). Their loading and release capacities were also studied and we were able to demonstrate a sustained release of both growth factors, for up to 28 days. Noteworthy, the growth factors biological activity on human cells was maintained. Altogether, these data suggest that this PMBs/Si-HPMC biphasic system may be a promising candidate for the development of an innovative bioactive delivery system for IVD regenerative medicine. Cell/Tissue: intervertebral disc cells
MicroTester/MicroSquisher	2017	Real-Time And Non-Invasive Measurements Of Cell Mechanical Behaviour With Optical Coherence Phase Microscopy	Methods Journal	D. Gillies, W. Gamal, A. Downes, Y. Reinwald, Y. Yang, A.J. El Haj, P.O. Bagnaninchi	University of Edinburgh		Cell mechanical behaviour is increasingly recognised as a central biophysical parameter in cancer and stem cell research, and methods of investigating their mechanical behaviour are therefore needed. We have developed a novel qualitative method based on quantitative phase imaging which is capable of investigating cell mechanical behaviour in real-time at cellular resolution using optical coherence phase microscopy (OCPM), and stimulating the cells non-invasively using hydrostatic pressure. The method was exemplified to distinguish between cells with distinct mechanical properties, and transient change induced by Cytochalasin D. We showed the potential of quantitative phase imaging to detect nanoscale intracellular displacement induced by varying hydrostatic pressure in microfluidic channels, reflecting cell mechanical behaviour. Further physical modelling is required to yield quantitative mechanical properties. Cell/Tissue: stem cells
MicroTester/MicroSquisher	2017	The Physical Basis Of Coordinated Tissue Spreading In Zebrafish Gastrulation	Developmental Cell	H. Morita, S. Grigolon, M. Bock, S.F.G. Krens, G. Salbreux, C-P. Heisenberg	University of Yamanashi		Embryo morphogenesis relies on highly coordinated movements of different tissues. However, remarkably little is known about how tissues coordinate their movements to shape the embryo. In zebrafish embryogenesis, coordinated tissue movements first become apparent during “doming,” when the blastoderm begins to spread over the yolk sac, a process involving coordinated epithelial surface cell layer expansion and mesenchymal deep cell intercalations. Here, we find that active surface cell expansion represents the key process coordinating tissue movements during doming. By using a combination of theory and experiments, we show that epithelial surface cells not only trigger blastoderm expansion by reducing tissue surface tension, but also drive blastoderm thinning by inducing tissue contraction through radial deep cell intercalations. Thus, coordinated tissue expansion and thinning during doming relies on surface cells simultaneously controlling tissue surface tension and radial tissue contraction. Cell/Tissue: blastodermepithelial surface cells
MicroTester/MicroSquisher	2017	The Role Of Adhesion Junctions In The Biomechanical Behaviour And Osteogenic Differentiation Of 3d Mesenchymal Stem Cell Spheroids	Journal of biomechanics	F.E. Griffin, J. Schiavi, T.C. McDevitt, J.P. McGarry, L.M. McNamara	National University of Ireland		Osteogenesis of mesenchymal stem cells (MSC) can be regulated by the mechanical environment. MSCs grown in 3D spheroids (mesenspheres) have preserved multi-lineage potential, improved differentiation efficiency, and exhibit enhanced osteogenic gene expression and matrix composition in comparison to MSCs grown in 2D culture. Within 3D mesenspheres, mechanical cues are primarily in the form of cell-cell contraction, mediated by adhesion junctions, and as such adhesion junctions are likely to play an important role in the osteogenic differentiation of mesenspheres. However the precise role of N- and OB-cadherin on the biomechanical behaviour of mesenspheres remains unknown. Here we have mechanically tested mesenspheres cultured in suspension using parallel plate compression to assess the influence of N-cadherin and OB-cadherin adhesion junctions on the viscoelastic properties of the mesenspheres during osteogenesis. Our results demonstrate that N-cadherin and OB-cadherin have different effects on mesensphere viscoelastic behaviour and osteogenesis. When OB-cadherin was silenced, the viscosity, initial and long term Young's moduli and actin stress fibre formation of the mesenspheres increased in comparison to N-cadherin silenced mesenspheres and mesenspheres treated with a scrambled siRNA (Scram) at day 2. Additionally, the increased viscoelastic material properties correlate with evidence of calcification at an earlier time point (day 7) of OB-cadherin silenced mesenspheres but not Scram. Interestingly, both N-cadherin and OB-cadherin silenced mesenspheres had higher BSP2 expression than Scram at day 14. Taken together, these results indicate that N-cadherin and OB-cadherin both influence mesensphere biomechanics and osteogenesis, but play different roles. Cell/Tissue: mesenchymal stem cells
MicroTester/MicroSquisher	2018	Acrylate-Based Materials For Heart Valve Scaffold Engineering	Royal Society of Chemistry	R. Santoro, S. Venkateswaran, F. Amandeo, R. Zhang, M. Brioschi, A. Callanan, M. Agrifoglio, C. Banfi, M. Bradley, M. Pesce	Edinburgh		Calcific aortic valve disease (CAVD) is the most frequent cardiac valve pathology. Its standard treatment consists of surgical replacement either with mechanical (metal made) or biological (animal tissue made) valve prostheses, both of which have glaring deficiencies. In the search for novel materials to manufacture artificial valve tissue, we have conducted a high-throughput screening with subsequent up-scaling to identify non-degradable polymer substrates that promote valve interstitial cells (VICs) adherence/growth and, at the same time, prevent their evolution toward a pro-calcific phenotype. Here, we provide evidence that one of the two identified ‘hit’ polymers, poly(methoxyethylmethacrylate-co-diethylaminoethylmethacrylate), provided robust VICs adhesion and maintained the healthy VICs phenotype without inducing pro-osteogenic differentiation. This ability was also maintained when the polymer was used to coat a non-woven poly-caprolactone (PCL) scaffold using a novel solvent coating procedure, followed by bioreactor-assisted VICs seeding. Since we observed that VICs had an increased secretion of the elastin-maturing component MFAP4 in addition to other valve-specific extracellular matrix components, we conclude that valve implants constructed with this polyacrylate will drive the biological response of human valve-specific cells.
MicroTester/MicroSquisher	2018	A Microvascularized Tumor-Mimetic Platform For Assessing Anti-Cancer Drug Efficacy	Scientific Reports	S. Pradhan, A.M. Smith, C.J. Garson, I. Hassani, W.J. Seeto, K. Pant, R.D. Arnold, B. Prabhakarpandian, E.A. Lipke	Auburn University		Assessment of anti-cancer drug efficacy in in vitro three-dimensional (3D) bioengineered cancer models provides important contextual and relevant information towards pre-clinical translation of potential drug candidates. However, currently established models fail to sufficiently recapitulate complex tumor heterogeneity. Here we present a chip-based tumor-mimetic platform incorporating a 3D in vitro breast cancer model with a tumor-mimetic microvascular network, replicating the pathophysiological architecture of native vascularized breast tumors. The microfluidic platform facilitated formation of mature, lumenized and flow-aligned endothelium under physiological flow recapitulating both high and low perfused tumor regions. Metastatic and non-metastatic breast cancer cells were maintained in long-term 3D co-culture with stromal fibroblasts in a poly(ethylene glycol)-fibrinogen hydrogel matrix within adjoining tissue chambers. The interstitial space between the chambers and endothelium contained pores to mimic the “leaky” vasculature found in vivo and facilitate cancer cell-endothelial cell communication. Microvascular pattern-dependent flow variations induced concentration gradients within the 3D tumor mass, leading to morphological tumor heterogeneity. Anti-cancer drugs displayed cell type- and flow pattern-dependent effects on cancer cell viability, viable tumor area and associated endothelial cytotoxicity. Overall, the developed microfluidic tumor-mimetic platform facilitates investigation of cancer-stromal-endothelial interactions and highlights the role of a fluidic, tumor-mimetic vascular network on anti-cancer drug delivery and efficacy for improved translation towards pre-clinical studies.
MicroTester/MicroSquisher	2018	Nanoscale Dysregulation Of Collagen Structure-Function Disrupts Mechano-Homeostasis And Mediates Pulmonary Fibrosis	eLIFE	M.G. Jones, O.G. Andriotis, J.J.W. Roberts, K. Lunn, V.J. Tear, L.Cao, K. Ask, D.E. Smart, A. Bonfanti, P.Johnson, A. Alzetani, D.E. Davies	Southampton
MicroTester/MicroSquisher	2018	Extracellular Matrix Surface Regulates Self-Assembly Of Three-Dimensional Placental Trophoblast Spheroids	PLOS One	M.K. Wong, S.A. Shawky, A. Aryasomayajula, M.A. Green, T. Ewart, P.R. Selvanganapathy, S. Raha	McMaster University		The incorporation of the extracellular matrix (ECM) is essential for generating in vitro models that truly represent the microarchitecture found in human tissues. However, the cell-cell and cell-ECM interactions in vitro remains poorly understood in placental trophoblast biology. We investigated the effects of varying the surface properties (surface thickness and stiffness) of two ECMs, collagen I and Matrigel, on placental trophoblast cell morphology, viability, proliferation, and expression of markers involved in differentiation/syncytial fusion. Most notably, thicker Matrigel surfaces were found to induce the self-assembly of trophoblast cells into 3D spheroids that exhibited thickness-dependent changes in viability, proliferation, syncytial fusion, and gene expression profiles compared to two-dimensional cultures. Changes in F-actin organization, cell spread morphologies, and integrin and matrix metalloproteinase gene expression profiles, further reveal that the response to surface thickness may be mediated in part through cellular stiffness-sensing mechanisms. Our derivation of self-assembling trophoblast spheroid cultures through regulation of ECM surface alone contributes to a deeper understanding of cell-ECM interactions, and may be important for the advancement of in vitro platforms for research or diagnostics.
MicroTester/MicroSquisher	2018	3d Skeletal Muscle Fascicle Engineering Is Improved With Tgf-Β1 Treatment Of Myogenic Cells And Their Co-Culture With Myofibroblasts		J. Krieger, B-W. Park, C.R. Lambert, C. Malcuit	Kent State		Skeletal muscle wound healing is dependent on complex interactions between fibroblasts, myofibroblasts, myogenic cells, and cytokines, such as TGF-β1. This study sought to clarify the impact of TGF-β1 signaling on skeletal muscle cells and discern between the individual contributions of fibroblasts and myofibroblasts to myogenesis when in co-culture with myogenic cells. 3D tissue-engineered models were compared to equivalent 2D culture conditions to assess the efficacy of each culture model to predictively recapitulate the in vivo muscle environment.
MicroTester/MicroSquisher	2018	Combined Use Of Parallel-Plate Compression And Finite Element Modeling To Analyze The Mechanical Properties Of Intact Porcine Lens	Journal of Mechanics in Medicine and Biology	K. Wang, D.T. Venetsanos, J. Wang, B.K. Pierscionek	Nottingham		The objective of this study is to explore the feasibility of a compression test for measuring mechanical properties of intact eye lenses using novel parallel plate compression equipment to compare the accuracy of implementing a classical Hertzian model and a newly proposed adjusted Hertzian model to calculate Young’s modulus from compression test results using finite element (FE) analysis. Parallel-plate compression tests were performed on porcine lenses. An axisymmetric FE model was developed to simulate the experimental process to evaluate the accuracy of using the classical Hertzian theory of contact mechanics as well as a newly proposed adjusted Hertzian theory model for calculating the equivalent Young’s modulus. By fitting the force-displacement relation obtained from FE simulations to both the classical and adjusted Hertzian theory model and comparing the calculated modulus to the input modulus of the FE model, the results demonstrated that the classical Hertzian theory model overestimated the Young’s modulus with a proportional error of over 10%. The adjusted Hertzian theory model produced results that are closer to original input values with error ratios all lower than 1.29%. Measurements of three porcine lenses from the parallel plate compression experiments were analyzed with resulting values of Young’s modulus of between 3.2kPa and 4.3kPa calculated. This study demonstrates that the adjusted Hertzian theory of contact mechanics can be applied in conjunction with the parallel-plate compression system to investigate the overall mechanical behavior of intact lenses.
MicroTester/MicroSquisher	2018	Mammary Fibroblasts Remodel Fibrillar Collagen Microstructure In A Biomimetic Nanocomposite Hydrogel	Acta Biomaterialia	C. Liu, B. Chiang, D.L. Mejia, K.E. Luker, G.D. Luker, A. Lee	U of Michigan		Architecture and microstructure of type I collagen fibers constitute central regulators of tumor invasion with aligned fibers providing a route for migration of stromal and cancer cells. Several different aspects of fibrillar collagen, such as stiffness, density, thickness, and pore size, may regulate migration of cancer cells, but determining effects of any one parameter requires clear decoupling of physical properties of collagen networks. The objective of this work is to develop and apply an in vitro three-dimensional (3D) tumor-extra cellular matrix (ECM) model with tunable physical parameters to define how stromal fibroblasts modulate collagen microstructure to control migration of breast cancer cells. We incorporated two different types of polyhedral oligomeric silsesquioxane (POSS) nano-molecules into a collagen/alginate matrix to induce different mechanisms of gelling. The resultant biomimetic, nanocomposite hydrogels show different collagen fibrillar microstructures while maintaining constant overall matrix stiffness, density, and porosimetry. Spheroids of human mammary fibroblasts embedded in these 3D matrices remodel the collagen network to varying extents based on differences in underlying matrix microstructures. The remodeled collagen matrix shows oriented, thicker fibrillar tracks, facilitating invasion of tumor cells. By decoupling effects of matrix stiffness and architecture, our nanocomposite hydrogels serve as robust platforms to investigate how biophysical properties of tumor environments control key processes regulating tumor progression in breast cancer and other malignancies.
MicroTester/MicroSquisher	2018	Hybrid Collagen Alginate Hydrogel As A Platform For 3d Tumor Spheroid Invasion	Acta Biomaterialia	C. Liu, D.L. Mejia, B. Chiang, K.E. Luker, G.D. Luker	U of Michigan		Extracellular matrix regulates hallmark features of cancer through biochemical and mechanical signals, although mechanistic understanding of these processes remains limited by lack of models that recreate physiology of tumors. To tissue-engineer models that recapitulate three-dimensional architecture and signaling in tumors, there is a pressing need for new materials permitting flexible control of mechanical and biophysical features. We developed a hybrid hydrogel system composed of collagen and alginate to model tumor environments in breast cancer and other malignancies. Material properties of the hydrogel, including stiffness, microstructure and porosimetry, encompass parameters present in normal organs and tumors. The hydrogel possesses a well-organized, homogenous microstructure with adjustable mechanical stiffness and excellent permeability. Upon embedding multicellular tumor spheroids, we constructed a 3D tumor invasion model showing follow-the-leader migration with fibroblasts leading invasion of cancer cells similar to in vivo. We also demonstrated effects of CXCL12-CXCR4 signaling, a pathway implicated in tumor progression and metastasis, in a dual-tumor spheroid invasion model in 3D hydrogels. These studies establish a new hydrogel platform with material properties that can be tuned to investigate effects of environmental conditions on tumor progression, which will advance future studies of cancer cell invasion and response to therapy.
MicroTester/MicroSquisher	2018	3d Bioprinting Of Hydrogels For Retina Cell Culturing	Bioprinting	P. Wang, X. Li, W. Zhu, Z. Zhong, A. Moran, W. Wang, K. Zhang, S. Chen	UCSD		Recapitulating native retina environment is crucial in isolation and culturing of retina photoreceptors (PRs). To date, maturation of PRs remains incomprehensible in vitro. Here we present a strategy of integrating the physical and chemical signals through 3D-bioprinting of hyaluronic acid (HA) hydrogels and co-differentiation of retinal progenitor cells (RPCs) into PRs with the support of retinal-pigment epithelium (RPEs). To mimic the native environment during retinal development, we chemically altered the functionalization of HA hydrogels to match the compressive modulus of HA hydrogels with native retina. RPEs were incorporated in the culturing system to support the differentiation due to their regeneration capabilities. We found that HA with a specific functionalization can yield hydrogels with compressive modulus similar to native retina. This hydrogel is also suitable for 3D bioprinting of retina structure. The results from cell study indicated that derivation of PRs from RPCs was improved in the presence of RPEs.
MicroTester/MicroSquisher	2018	Matrix Composition In 3-D Collagenous Bioscaffolds Modulates The Survival And Angiogenic Phenotype Of Human Chronic Wound Dermal Fibroblasts	Acta Biomaterialia	P.M. Martin, A. Grant, D.W. Hamilton, L.E. Flynn	UWO		There is a substantial need for new strategies to stimulate cutaneous tissue repair in the treatment of chronic wounds. To address this challenge, our team is developing modular biomaterials termed “bead foams”, comprised of porous beads synthesized exclusively of extracellular matrix (ECM) and assembled into a cohesive three-dimensional (3-D) network. In the current study, bead foams were fabricated from human decellularized adipose tissue (DAT) or commercially-sourced bovine tendon collagen (COL) to explore the effects of ECM composition on human wound edge dermal fibroblasts (weDF) sourced from chronic wound tissues. The DAT and COL bead foams were shown to be structurally similar, but compositionally distinct, containing different levels of glycosaminoglycan content and collagen types IV, V, and VI. In vitro testing under conditions simulating stresses within the chronic wound microenvironment indicated that weDF survival and angiogenic marker expression were significantly enhanced in the DAT bead foams as compared to the COL bead foams. These findings were corroborated through in vivo assessment in a subcutaneous athymic mouse model. Taken together, the results demonstrate that weDF survival and paracrine function can be modulated by the matrix source applied in the design of ECM-derived scaffolds and that the DAT bead foams hold promise as cell-instructive biological wound dressings.
MicroTester/MicroSquisher	2018	Understanding Interfacial Interactions Of Polydopamine And Glass Fiber And Their Enhancement Mechanisms In Epoxy-Based Laminates	Composites Part A: Applied Science and Manufacturing	H. Zhang, W. Han, J. Tavakoli, Y. Zhang, X. Lin, X. Lu, Y. Ma, Y. Tang	Flinders University		Interfacial behavior greatly affects the bulk mechanical performance of fiber reinforced polymer laminates. In this study, polydopamine modified glass fiber was used to fabricate short glass fiber reinforced polymer (GFRP) laminates. The interactions between glass fiber and polydopamine were studied experimentally and theoretically by X-ray photoelectron spectroscopy (XPS) and density functional calculation respectively. Theoretical study clearly demonstrated that electronic interactions existed between polydopamine and glass fiber, indicating the hydrogen bonds/chemical interactions between them that were also demonstrated by XPS results. The enhanced interfacial interaction significantly benefited GFRP laminates, as demonstrated by various mechanical characterizations such as single fiber pull-out and Mode I interlaminar fracture toughness tests. Combining the theoretical and experimental studies indicated that polydopamine modification of glass fiber could be an easy and effective way to significantly improve the interfacial interactions between glass fiber and matrix and enhance the mechanical properties of GFRP laminates.
MicroTester/MicroSquisher	2018	Colloidal Gelatin Microgels With Tunable Elasticity Support The Viability And Differentiation Of Mesenchymal Stem Cells Under Pro‐Inflammatory Conditions	Journal of Biomedical Materials Research	B. Sung, J. Krieger, B. Yu, M-H. Kim	Kent State		Despite a promising potential for mesenchymal stem cells (MSCs) in tissue regeneration, a major challenge in MSC‐based therapy has been associated with poor cell survival and low levels of cell integration into host tissue following transplantation. The objective of this study was to develop a gelatin‐based colloidal microgel platform that enables the encapsulation of viable MSCs as well as confer fine‐tuning of mechanical stiffness and low cytotoxicity. In this study, we report a facile method of fabricating gelatin‐based microgel spheres for the encapsulation of MSCs using a water‐in‐oil mini‐emulsification method, which is covalently crosslinked by genipin. At a given seeding cell number, there was a positive correlation between the size of the microsphere and the number of encapsulated MSCs. Controlling the crosslinking degree of gelatin matrix enabled a fine‐tuning of mechanical stiffness of gel microsphere. MSCs within softer microgel exhibit more spread morphology than the cells in the stiffer matrix, while cells within stiffer matrix become more elongated morphology. Importantly, we show that the colloidal gelatin microgel could support the viability and differentiation of encapsulated MSCs in a pro‐inflammatory environment. This study demonstrates the feasibility of using genipin‐crosslinked gelatin gel microspheres as an injectable carrier of MSCs for tissue engineering applications, which can be further explored for MSC‐based cell therapy for tissue repair.
MicroTester/MicroSquisher	2018	Toward Morphologically Relevant Extracellular Matrix In Vitro Models: 3d Fiber Reinforced Hydrogels	Frontiers in Physiology	A. Williams, J.F. Nowak, R. Dass, J. Samuel, K.L. Mills	Rensselaer		The extracellular matrix (ECM) is known to play an important role in the health of cells and tissues. Not only are chemical signals transmitted via bonds and tightly controlled diffusion, but the structure of the ECM also provides important physical signaling for the cells attached to it. The structure is composed of a mesh of fibrous proteins, such as collagen, embedded in a hydrated gel matrix of glycosaminoglycans. To study cell behavior with respect to the combined morphology and mechanics of such matrices is not currently possible with the types of 3D cell culture matrices available. Most of the cell culture matrices are single-phase bio- or polymeric hydrogels. Therefore, here we developed a continuous hybrid manufacturing process to make fiber-reinforced composite hydrogels. A far field electrospinning process was used to deposit the fibrous component with the aid of guiding electrodes; and a gravity-assisted, droplet-based system controlled the rate of addition of the cell-laden hydrogel component. The addition of the fibrous component slightly increased the elastic modulus of the pure hydrogel. The cells that were embedded into the fiber-reinforced hydrogels were viable for 8 days. The cells were randomly placed in the matrix such that some had no contact to the fibers and others were initially in proximity to fibers. The cells with no contact to fibers grew into spheroidal clusters within the hydrogel, and those in proximity to the fibers spread out and grew along the fibers showing that the fiber-reinforced hydrogels are able to control cell behavior with morphological cues.
MicroTester/MicroSquisher	2018	Scaffold-Free, Label-Free And Nozzle-Free Biofabrication Technology Using Magnetic Levitational Assembly	Biofabrication	V. A. Parfenov, E.V. Koudan, E.A. Bulanova, A.D. Knyazeva, A.A. Gryadunova, O.F. Petrov, V.A. Mironov	Laboratory for Biotechnological Research, Russia		Tissue spheroids have been proposed as building blocks in 3D biofabrication. Conventional magnetic force-driven 2D patterning of tissue spheroids requires prior cell labeling by magnetic nanoparticles, meanwhile a label-free approach for 3D magnetic levitational assembly has been introduced. Here we present first time report on rapid assembly of 3D tissue construct using scaffold-free, nozzle-free and label-free magnetic levitation of tissue spheroids. Chondrospheres of standard size, shape and capable to fusion have been biofabricated from primary sheep chondrocytes using non-adhesive technology. Label-free magnetic levitation was performed using a prototype device equipped with permanent magnets in presence of gadolinium (Gd3+) in culture media, which enables magnetic levitation. Mathematical modeling and computer simulations were used for prediction of magnetic field and kinetics of tissue spheroids assembly into 3D tissue constructs. First, we used polystyrene beads to simulate the assembly of tissue spheroids and to determine the optimal settings for magnetic levitation in presence of Gd3+. Second, we proved the ability of chondrospheres to assemble rapidly into 3D tissue construct in the permanent magnetic field in the presence of Gd3+. Thus, scaffold- and label-free magnetic levitation of tissue spheroids is a promising approach for rapid 3D biofabrication and attractive alternative to label-based magnetic force-driven tissue engineering.
MicroTester/MicroSquisher	2018	Rapid 3d Bioprinting Of Decellularized Extracellular Matrix With Regionally Varied Mechanical Properties And Biomimetic Microarchitecture	Biomaterials	X. Ma, C. Yu, P. Wang, W. Xu, X. Wan, C.S.E. Lai, J. Liu, A. K-Maharajh, S. Chen	UCSD		Hepatocellular carcinoma (HCC), as the fifth most common malignant cancer, develops and progresses mostly in a cirrhotic liver where stiff nodules are separated by fibrous bands. Scaffolds that can provide a 3D cirrhotic mechanical environment with complex native composition and biomimetic architecture are necessary for the development of better predictive tissue models. Here, we developed photocrosslinkable liver decellularized extracellular matrix (dECM) and a rapid light-based 3D bioprinting process to pattern liver dECM with tailorable mechanical properties to serve as a platform for HCC progression study. 3D bioprinted liver dECM scaffolds were able to stably recapitulate the clinically relevant mechanical properties of cirrhotic liver tissue. When encapsulated in dECM scaffolds with cirrhotic stiffness, HepG2 cells demonstrated reduced growth along with an upregulation of invasion markers compared to healthy controls. Moreover, an engineered cancer tissue platform possessing tissue-scale organization and distinct regional stiffness enabled the visualization of HepG2 stromal invasion from the nodule with cirrhotic stiffness. This work demonstrates a significant advancement in rapid 3D patterning of complex ECM biomaterials with biomimetic architecture and tunable mechanical properties for in vitro disease modeling.
MicroTester/MicroSquisher	2018	Trans-Differentiation Of Human Adipose-Derived Mesenchymal Stem Cells Into Cardiomyocyte-Like Cells On Decellularized Bovine Myocardial Extracellular Matrix-Based Films	Journal of Materials Science	Y.E. Arslan, Y.F. Galata, T.S. Arslan, B. Derkus	Regenerative Biomaterials Laboratory, Turkey		In this study, we aimed at fabricating decellularized bovine myocardial extracellular matrix-based films (dMEbF) for cardiac tissue engineering (CTE). The decellularization process was carried out utilizing four consecutive stages including hypotonic treatment, detergent treatment, enzymatic digestion and decontamination, respectively. In order to fabricate the dMEbF, dBM were digested with pepsin and gelation process was conducted. dMEbF were then crosslinked with N-hydroxysuccinimide/1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide (NHS/EDC) to increase their durability. Nuclear contents of native BM and decellularized BM (dBM) tissues were determined with DNA content analysis and agarose-gel electrophoresis. Cell viability on dMEbF for 3rd, 7th, and 14th days was assessed by MTT assay. Cell attachment on dMEbF was also studied by scanning electron microscopy. Trans-differentiation capacity of human adipose-derived mesenchymal stem cells (hAMSCs) into cardiomyocyte-like cells on dMEbF were also evaluated by histochemical and immunohistochemical analyses. DNA contents for native and dBM were, respectively, found as 886.11 ± 164.85 and 47.66 ± 0.09 ng/mg dry weight, indicating a successful decellularization process. The results of glycosaminoglycan and hydroxyproline assay, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), performed in order to characterize the extracellular matrix (ECM) composition of native and dBM tissue, showed that the BM matrix was not damaged during the proposed method. Lastly, regarding the histological study, dMEbF not only mimics native ECM, but also induces the stem cells into cardiomyocyte-like cells phenotype which brings it the potential of use in CTE.
MicroTester/MicroSquisher	2018	Extracellular Matrix Determines Biomechanical Properties Of Chondrospheres During Their Maturation In Vitro	Cartilage	N.P. Omelyanenko, P.A. Karalkin, E.A. Bulanova	Priorov National Medical Research Center of Traumatology and Orthopedics		Chondrospheres represent a variant of tissue spheroids biofabricated from chondrocytes. They are already being used in clinical trials for cartilage repair; however, their biomechanical properties have not been systematically investigated yet. The aim of our study was to characterize chondrospheres in long-term in vitro culture conditions for morphometric changes, biomechanical integrity, and their fusion and spreading kinetics.
MicroTester/MicroSquisher	2018	Influence Of Hydrophobic Cross-Linkers On Carboxybetaine Copolymer Stimuli Response And Hydrogel Biological Properties	Langmuir	V. Huynh, A. H. Jesmer, M. M. Shoaib, R. G. Wylie	McMaster University		Poly(carboxybetaine) (pCB) hydrogels do not elicit a foreign body response due to their low-fouling properties, making them ideal implantable materials for in vivo drug and cell delivery. Current reported pCB hydrogels are cross-linked using cytotoxic UV-initiated radical polymerization limiting clinical and in vivo translation. For clinical translation, we require in situ and biorthogonal cross-linking of pCB hydrogels that are both low-fouling and low-swelling to limit nonspecific interactions and minimize tissue damage, respectively. To this end, we synthesized carboxybetaine (CB) random copolymers (molecular weight (MW): ∼7–33 kDa; Đ: 1.1–1.36) containing azide (pCB-azide) or strained alkyne (Dibenzocyclooctyne (DBCO); pCB-DBCO) that rapidly cross-link upon mixing. Unlike CB homopolymers and other CB copolymers studied, high DBCO content pCB-DBCO30 (30% DBCO mole fraction) is thermoresponsive with a upper critical solution temperature (UCST; cloud point of ∼20 °C at 50 g/L) in water due to electrostatic associations. Due to the antipolyelectrolyte effect, pCB-DBCO30 is salt-responsive and is soluble even at low temperatures in 5 M NaCl, which prevents zwitterion electrostatic associations. pCB-azide and pCB-DBCO with 0.05 to 0.16 cross-linker mole fractions rapidly formed 10 wt % hydrogels upon mixing that were low-swelling (increase of ∼10% in wet weight) while remaining low-fouling to proteins (∼10–20 μg cm–2) and cells, making them suitable for in vivo applications. pCB-X31 hydrogels composed of pCB-azide32 and pCB-DBCO30 formed opaque gels in water and physiological conditions that shrunk to ∼70% of their original wet weight due to pCB-DBCO30’s greater hydrophobicity and interchain electrostatic interactions, which promotes nonspecific protein adsorption (∼35 μg cm–2) and cell binding. Once formed, the electrostatic interactions in pCB-X31 hydrogels are not fully reversible with heat or salt. Although, pCB-X31 hydrogels are transparent when initially prepared in 5 M NaCl. This is the first demonstration of a thermo- and salt-responsive CB copolymer that can tune hydrogel protein and cell fouling properties.
MicroTester/MicroSquisher	2018	Microfluidics Device for Fabrication of Large, Uniform, Injectable Hydrogel Microparticles for Cell Encapsulation	US Patent Application Publication	E. Lipke, W. Seeto, Y. Tian	Auburn University		The devices, methods, and compositions disclosed herein accomplish robust cell encapsulation in polymer microparticles using a vertically oriented microfluidic device. A hydrophilic polymer precursor solution is flowed into a first inlet channel, which extends inward from an upper surface of the device housing. A hydrophobic fluid is flowed into a second inlet channel, which extends inward from a lower surface of the device housing. The two inlet channels meet at a junction, and an outlet channel extends away from the two inlet channels. When the two inwardly flowing streams meet at the junction, the polymer precursor solution disperses into the hydrophobic fluid. The dispersed precursor droplets are photopolymerized into microparticles as they travel through the outlet channel. The resulting microparticles are highly uniform, and are larger than conventionally formed microparticles. Cells of varying types can be encapsulated with high viability and spatial uniformity.
MicroTester/MicroSquisher	2019	Rapid 3d Bioprinting Of In-Vitro Cardiac Tissue Models Using Human Embryonic Stem Cell-Derived Cardiomyocytes	Bioprinting	J. Liu, J. He, J. Liu, X. Ma, Q. Chen, N. Lawrence, W. Zhu, Y. Xu, S. Chen	UCSD		Cardiovascular disease (CVD) is well recognized as the leading cause of death worldwide, attributing 32% of global deaths to CVD [1]. The majority of deaths are attributed to ischemic heart disease (IHD). Current treatments of IHD can only delay the progression of the disease. Thus, there is a significant need to develop new strategies to replace injured or damaged myocardium. Over the past decades, human embryonic stem cells (hESCs) have offered opportunities of repairing damaged organ such as heart [2], especially for the patients suffering loss of functional cardiomyocytes. hESCs have been extensively studied and their robust differentiation towards cardiomyocyte lineages have been well established. The strategies for generating cardiomyocytes from stem cells, maturation eliciting physiological response, and how to improve the survival of the engraftments remains a concern [3]. Potential strategies include alignment [4] and co-culture with endothelial cellsand fibroblasts [5]. However, stem-cell derived cardiomyocytes are still in state of research where obtaining quality cells is both time consuming and resource intensive to differentiate with limited ability to proliferate once differentiated, a means of creating 3D micro-tissues that elicit physiological responses is necessary.
MicroTester/MicroSquisher	2019	Scanningless And Continuous 3d Bioprinting Of Human Tissues With Decellularized Extracellular Matrix	Biomaterials	C. Yu, X. Ma, W. Zhu, P. Wang, Kathleen L. Miller, J. Stupin, A. Koroleva-Maharajh, A. Hairabedian, S. Chen	UCSD	3	Decellularized extracellular matrices (dECMs) have demonstrated excellent utility as bioscaffolds in recapitulating the complex biochemical microenvironment, however, their use as bioinks in 3D bioprinting to generate functional biomimetic tissues has been limited by their printability and lack of tunable physical properties. Here, we describe a method to produce photocrosslinkable tissue-specific dECM bioinks for fabricating patient-specifictissues with high control over complex microarchitecture and mechanical properties using a digital light processing (DLP)-based scanningless and continuous 3D bioprinter. We demonstrated that tissue-matched dECM bioinks provided a conducive environment for maintaining high viability and maturation of human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes and hepatocytes. Microscale patterning also guided spontaneous cellular reorganization into predesigned striated heart and lobular liver structures through biophysical cues. Our methodology enables a light-based approach to rapidly bioprint dECM bioinks with accurate tissue-scale design to engineer physiologically-relevant functional human tissues for applications in biology, regenerative medicine, and diagnostics.
MicroTester/MicroSquisher	2019	Cell Force-Mediated Matrix Reorganization Underlies Multicellular Network Assembly	Nature.com	C. D. Davidson, W. Y. Wang, I. Zaimi, D. K. P. Jayco, B. M. Baker	University of Michigan		Vasculogenesis is the de novo formation of a vascular network from individual endothelial progenitor cells occurring during embryonic development, organogenesis, and adult neovascularization. Vasculogenesis can be mimicked and studied in vitro using network formation assays, in which endothelial cells (ECs) spontaneously form capillary-like structures when seeded in the appropriate microenvironment. While the biochemical regulators of network formation have been well studied using these assays, the role of mechanical and topographical properties of the extracellular matrix (ECM) is less understood. Here, we utilized both natural and synthetic fibrous materials to better understand how physical attributes of the ECM influence the assembly of EC networks. Our results reveal that active cell-mediated matrix recruitment through actomyosin force generation occurs concurrently with network formation on Matrigel, a reconstituted basement membrane matrix regularly used to promote EC networks, and on synthetic matrices composed of electrospun dextran methacrylate (DexMA) fibers. Furthermore, modulating physical attributes of DexMA matrices that impair matrix recruitment consequently inhibited the formation of cellular networks. These results suggest an iterative process in which dynamic cell-induced changes to the physical microenvironment reciprocally modulate cell behavior to guide the formation and stabilization of multicellular networks.
MicroTester/MicroSquisher	2019	A Multimaterial Microphysiological Platform Enabled By Rapid Casting Of Elastic Microwires	Advanced Healthcare Materials	Y. Zhao, E. Y. Wang, L. H. Davenport, Y. Liao, K. Yeager, G. Vunjak-Novakovic, M. Radisic, B. Zhang	McMaster University		Due to escalating drug developmental costs and limitations of cardiotoxicity screening, there is an urgent need to develop robust in vitro 3D tissue culture platforms that can both facilitate the culture of human cardiac tissues and provide noninvasive functional readouts predictive of cardiotoxicity in clinical settings. However, such platforms commonly require complex fabrication procedures that are difficult to scale up to high‐throughput testing platforms. Here, innovative multimaterial processing into a scalable and functional platform is proposed in the format of a 96‐well plate. Three classes of materials are integrated into the platform. An array of soft elastic microwires is used both as anchors for tissue formation as well as sensors for recording tissue contraction. Conductive carbon electrodes are embedded into the plate to drive electrical stimulation for tissue maturation and pace tissue contraction during drug testing. The bulk of the device is made of rigid polystyrene plastic to eliminate drug‐absorbing polydimethylsiloxane (PDMS). The platform has higher throughput than the current state‐of‐the‐art devices, at a significantly reduced cost of manufacturing and tissue production.
MicroTester/MicroSquisher	2019	Significant Adhesion Enhancement Of Bioinspired Dry Adhesives By Simple Thermal Treatment	International Journal of Precision Engineering and Manufacturing - Green Technology	M. Seong, J. Lee, I. Hwang, H. E. Jeong	Ulsan National Institute of Science and Technology		Bioinspired dry adhesives with micropillar arrays can be harnessed for precise and environment-friendly manufacturing. This study presents a simple and robust approach for developing synthetic dry adhesives with significantly enhanced adhesion strength without sophisticated structural modification or chemical surface treatment. We show that when dry adhesives with micropillar arrays are annealed at slightly elevated temperatures of 150–200 °C, their adhesion strengths are remarkably enhanced (maximum normal adhesion: 50.0 N cm−2) compared to those that are not treated thermally (normal adhesion: 17.6 N cm−2). The enhanced adhesion levels obtained by simple annealing surpass those of previously reported dry adhesives having nanoscale hairs with high aspect ratios or mushroom-like pillars with large tips. Experimental investigations regarding the chemical structure, surface roughness, surface energy, and elastic modulus of the dry adhesive samples indicate that the enhanced adhesion originates from the annealing-induced enhancement of the adhesive’s elastic modulus
MicroTester/MicroSquisher	2019	A Scaffold- And Serum-Free Method To Mimic Human Stable Cartilage Validated By Secretome	Tissue Engineering	I. Cortes, R. A. M. Matsui, M. S. Azevedo, A. Beatrici, K. L. A. Souza, G. Launay, F. Delolme, J. M. Granjeiro, C. Moali, L. S. Baptista	National Institute of Metrology, Quality and Technology		A stabilized cartilage construct without signs of hypertrophy in chondrocytes is still a challenge. Suspensions of ASCs and cartilage progenitor cells (CPCs) were seeded into micro-molded non-adhesive hydrogel to produce spheroids (scaffold- and serum-free method) characterized by size, immunohistochemistry, fusion and biomechanical properties. After cell dissociation, they were characterized for mesenchymal cell surface markers, cell viability, and quantitative real-time PCR. Both targeted and non-targeted (shotgun mass-spectrometry) analysis were conducted on the culture supernatants. Induced-ASC spheroids (ø = 350m) showed high cell viability and CD73 downregulation contrasting to CD90. The TGF-β3/TGF-β1 ratio and SOX-9 increased (p<0.05) while IL-6, IL-8, RUNX2 and ALPL decreased. Induced-ASC spheroids were able to completely fuse and showed a higher force required to compression at day 14 (p < 0.0001). Strong collagen type II in situ was associated with gradual decrease of collagen type X and a lower COLXA1 gene expression at day 14 compared to 7 (p = 0,0352). The comparison of the secretome content of induced, non-induced ASCs and CPCs identified 138 proteins directly relevant to chondrogenesis out of 704 proteins in total. While collagen X was absent, TSP-1, described as anti-angiogenic and anti-hypertrophic, and COMP, a biomarker of chondrogenesis were upregulated in induced ASC spheroids. Our scaffold- and serum-free method mimics stable cartilage acting as a tool for biomarker discovery and for regenerative medicine protocols.
MicroTester/MicroSquisher	2019	A Platform For Generation Of Chamber-Specific Cardiac Tissues And Disease Modeling	Cell	Y. Zhao, N. Rafatian, N. T. Feric, B. J. Cox, R. Aschar-Sobbi, E. Y. Wang, P. Aggarwal, B. Zhang, G. Conant, K. Ronaldson-Bouchard, A. Pahnke, S. Protze, J. H. Lee, L. D. Huyer, D. Jekic, A. Wickeler, H. E. Naguib, G. M. Keller, G. Vunjak-Novakovic, U. Broeckel, P. H. Backx, M. Radisic	University of Toronto		Tissue engineering using cardiomyocytes derived from human pluripotent stem cells holds a promise to revolutionize drug discovery, but only if limitations related to cardiac chamber specification and platform versatility can be overcome. We describe here a scalable tissue-cultivation platform that is cell source agnostic and enables drug testing under electrical pacing. The plastic platform enabled on-line noninvasive recording of passive tension, active force, contractile dynamics, and Ca2+ transients, as well as endpoint assessments of action potentials and conduction velocity. By combining directed cell differentiation with electrical field conditioning, we engineered electrophysiologically distinct atrial and ventricular tissues with chamber-specific drug responses and gene expression. We report, for the first time, engineering of heteropolar cardiac tissues containing distinct atrial and ventricular ends, and we demonstrate their spatially confined responses to serotonin and ranolazine. Uniquely, electrical conditioning for up to 8 months enabled modeling of polygenic left ventricular hypertrophy starting from patient cells.
MicroTester/MicroSquisher	2019	Actomyosin Contractility-dependent Matrix Stretch and Recoil Induces Rapid Cell Migration	Nature Communications	W. Y. Wang, C. D. Davidson, D. Lin, B. M. Baker	University of Michigan		Cells select from a diverse repertoire of migration strategies. Recent developments in tunable biomaterials have helped identify how extracellular matrix properties influence migration, however, many settings lack the fibrous architecture characteristic of native tissues. To investigate migration in fibrous contexts, we independently varied the alignment and stiffness of synthetic 3D fiber matrices and identified two phenotypically distinct migration modes. In contrast to stiff matrices where cells migrated continuously in a traditional mesenchymal fashion, cells in deformable matrices stretched matrix fibers to store elastic energy; subsequent adhesion failure triggered sudden matrix recoil and rapid cell translocation. Across a variety of cell types, traction force measurements revealed a relationship between cell contractility and the matrix stiffness where this migration mode occurred optimally. Given the prevalence of fibrous tissues, an understanding of how matrix structure and mechanics influences migration could improve strategies to recruit repair cells to wound sites or inhibit cancer metastasis.
MicroTester/MicroSquisher	2019	A 96-Well Culture Platform Enables Longitudinal Analyses Of Engineered Human Skeletal Muscle Microtissue Strength	bioRxiv	M. E. Afshar, H. Y. Abraha, M. A. Bakooshli, S. Davoudi, N. Thavandiran, K. Tung, H. Ahn, H. Ginsberg, P. W. Zandstra, P. M. Gilbert	University of Toronto		Three-dimensional (3D) in vitro models of human skeletal muscle mimic aspects of native tissue structure and function, thereby providing a promising system for disease modeling, drug discovery or pre-clinical validation, and toxicity testing. Widespread adoption of this research approach is hindered by the lack of an easy-to-use platform that is simple to fabricate and yields arrays of human skeletal muscle micro-tissues (hMMTs) in culture with reproducible physiological responses that can be assayed non-invasively. Here, we describe a design and methods to generate a reusable mold to fabricate a 96-well platform, referred to as MyoTACTIC, that enables bulk production of 3D hMMTs. All 96-wells and all well features are cast in a single step from the reusable mold. Non-invasive calcium transient and contractile force measurements are performed on hMMTs directly in MyoTACTIC, and unbiased force analysis occurs by a custom automated algorithm, allowing for longitudinal studies of function. Characterizations of MyoTACTIC and resulting hMMTs confirms the reproducibility of device fabrication and biological responses. We show that hMMT contractile force mirrors expected responses to compounds shown by others to decrease (dexamethasone, cerivistatin) or increase (IGF-1) skeletal muscle strength. Since MyoTACTIC supports hMMT long-term culture, we evaluated direct influences of pancreatic cancer chemotherapeutics agents on contraction competent human skeletal muscle fibers. A single application of a clinically relevant dose of Irinotecan decreased hMMT contractile force generation, while clear effects on fiber atrophy were observed histologically only at a higher dose. This suggests an off-target effect that may contribute to cancer associated muscle wasting, and highlights the value of the MyoTACTIC platform to non-invasively predict modulators of human skeletal muscle function.
MicroTester/MicroSquisher	2019	Biomechanical Impact of Localized Corneal Cross-linking Beyond the Irradiated Treatment Area	Journal of Refractive Surgery	J. N. Webb, E. Langille, F. Hafezi, J. B. Randleman, G. Scarcelli	University of Maryland		To investigate the stiffening effect of localized corneal cross-linking (L-CXL) within and beyond the irradiated region in three dimensions.
MicroTester/MicroSquisher	2019	Molecularly-ordered Hydrogels with Controllable, Anisotropic Stimulus Response	Soft Matter	J. M. Boothby, J. Samuel, T. H. Ware	University of Texas at Dallas		Hydrogels which morph between programmed shapes in response to aqueous stimuli are of significant interest for biosensors and artificial muscles, among other applications. However, programming hydrogel shape change at small size scales is a significant challenge. Here we use the inherent ordering capabilities of liquid crystals to create a mechanically anisotropic hydrogel; when coupled with responsive comonomers, the mechanical anisotropy in the network guides shape change in response to the desired aqueous condition. Our synthetic strategy hinges on the use of a methacrylic chromonic liquid crystal monomer which can be combined with a non-polymerizable chromonic of similar structure to vary the magnitude of shape change while retaining liquid crystalline order. This shape change is directional due to the mechanical anisotropy of the gel, which is up to 50% stiffer along the chromonic stack direction than perpendicular. Additionally, we show that the type of stimulus to which these anisotropic gels respond can be switched by incorporating responsive, hydrophilic comonomers without destroying the nematic phase or alignment. The utility of these properties is demonstrated in polymerized microstructures which exhibit Gaussian curvature in response to high pH due to emergent ordering in a micron-sized capillary.
MicroTester/MicroSquisher	2019	Tunable Degradation of Low-Fouling Carboxybetaine-Hyaluronic Acid Hydrogels for Applications in Cell Encapsulation	Biomedical Materials	V. Huynh, A. D'Angelo, R. Wylie	McMaster University		Low-fouling hydrogels with tunable degradation rates and biochemical environments have the potential to improve adoptive cell therapies for cancer immunotherapy and regenerative medicine. To this end, we developed in situ gelling hydrogels from low-fouling poly(carboxybetaine-co-maleimide) (pCBM) random copolymers and thiolated hyaluronic acid (HA-SH). pCBM-HA hydrogel enzymatic degradation rates were tuned 5 fold by altering pCBM composition (4, 11, and 16 maleimide mol%) and 2.3 fold by HA-SH concentration (1–2 wt%). pCBM-HA gels were low-fouling towards bovine serum albumin (BSA; adsorbed ~20 μg cm−2) and resisted fibroblast adhesion. To control pCBM-HA bioactivity, the cell adhesive peptide CGRGDS was immobilized on pCBM to promote fibroblast adhesion (39% decrease in circularity), which increased metabolic activity by ~50%. pCBM-HA modified with CGRGDS enhanced the metabolic activity of encapsulated T cells by ~21% compared to gels without HA, indicating their potential for immunotherapies. Low-fouling pCBM-HA hydrogels provide a vehicle with tunable degradation rates and biochemical environments for encapsulation applications in cell adoptive therapies.
MicroTester/MicroSquisher	2019	Biowire Model of Interstitial and Focal Cardiac Fibrosis	ACS Central Science	E. Wang, N. Rafatian, Y. Zhao, A. Lee, B. Lai, R. Lu, D. Jekic, L. Huyer, E. Knee-Walden, S. Bhattacharya, P. Backx, M. Radisic	University of Toronto		Myocardial fibrosis is a severe global health problem due to its prevalence in all forms of cardiac diseases and direct role in causing heart failure. The discovery of efficient antifibrotic compounds has been hampered due to the lack of a physiologically relevant disease model. Herein, we present a disease model of human myocardial fibrosis and use it to establish a compound screening system. In the Biowire II platform, cardiac tissues are suspended between a pair of poly(octamethylene maleate (anhydride) citrate) (POMaC) wires. Noninvasive functional readouts are realized on the basis of the deflection of the intrinsically fluorescent polymer. The disease model is constructed to recapitulate contractile, biomechanical, and electrophysiological complexities of fibrotic myocardium. Additionally, we constructed a heteropolar integrated model with fibrotic and healthy cardiac tissues coupled together. The integrated model captures the regional heterogeneity of scar lesion, border zone, and adjacent healthy myocardium. Finally, we demonstrate the utility of the system for the evaluation of antifibrotic compounds. The high-fidelity in vitro model system combined with convenient functional readouts could potentially facilitate the development of precision medicine strategies for cardiac fibrosis modeling and establish a pipeline for preclinical compound screening.
MicroTester/MicroSquisher	2019	Mechanical considerations for design and implementation of peripheral intraneural devices	Journal of Neural Engineering	A. Stiller, M. Gonzalez-Gonzalez, J. Boothby, S. Sherman, J. Benavides, M. Romero-Ortega, J. Pancrazio, B. Black	University of Texas at Dallas		Objective. Neural interfaces designed to stimulate or record electrical activity from peripheral nerves have applications ranging from the electrical modulation of nerve activity as a therapeutic option (e.g. epilepsy and untreatable depression) to the design of prosthetics. Currently, most peripheral nerve interfaces are either cuff-style devices that wrap around the target nerve or intraneural devices that are implanted within the nerve. While the latter option offers higher specificity and signal-to-noise ratio, penetrating devices can cause significant damage to the nerve due to the high degree of mechanical mismatch. Because of this, there is interest in developing penetrating devices fabricated from soft or softening materials (materials having a low elastic modulus). However, there is currently a lack of understanding regarding implantation forces required for successful insertion, which is a constraint for soft device design. Softer devices require robust designs to achieve a critical buckling force that is larger than forces experienced during device insertion. Approach. This study comprehensively assesses insertion force under different implantation conditions, with three variations for implantation speed, angle, and device tip angle, during insertion of silicon shanks in rat sciatic nerve. Additionally, we report compression modulus for rat sciatic nerve at different compression rates to inform computational modeling. Main results. We found that insertion speed and angle had significant effects on peak insertion force. We observed lower insertion forces (10-60 mN) when the device was implanted at higher angles relative to the perpendicular insertion (80-125 mN). We also demonstrate use of a nerve-stabilizing device to keep the nerve immobile during implantation. Additionally, we found that compression modulus was significantly different in small and large strain regions of the stress-strain curve with values between 1500-4500 Pa depending on compression rate. Significance. This study provides information imperative to the design and successful implementation of soft penetrating peripheral nerve interfaces.
MicroTester/MicroSquisher	2019	Mesenchymal stem cell encapsulation in alginate micro-particles for intra-articular injection in osteoarthritis	Osteoarthritis and Cartilage	A. Smith, J. Boulestreau, M. Marquis, D. Renard, B. Legoff, F. Blanchard, C. Vinatier, J. Guicheux, A. des Rieux, C. Le Visage	Inserm		Osteoarthritis (OA) is a degenerative and inflammatory joint disease that affects cartilage, subchondral bone and joint tissues. Mesenchymal Stromal Cells (MSC) ability to secrete anti-inflammatory and immuno-modulatory factors represents an attractive tool in the treatment of OA. Considering cell death and the risk of cell leakage upon intra-articular (IA) injection, MSC encapsulation could therefore protect cells from death, avoid cell effusion outside the articular space, and provide a suitable 3D microenvironment supporting the biological activity of MSC.
MicroTester/MicroSquisher	2019	Compressive mechanical properties of rat and pig optic nerve head	Journal of Biomechanics	E. Boazak, J. d'Humieres, A. Read, C. Ethier	Georgia Institute of Technology and Emory University		Glaucoma is the leading cause of irreversible blindness worldwide. Elevated intraocular pressure (IOP), the primary risk factor for glaucoma, is thought to induce abnormally high strains in optic nerve head (ONH) tissues, which ultimately result in retinal ganglion cell damage and vision loss. The mechanisms by which excessive deformations result in vision loss remain incompletely understood. The ability of computational and in vitro models of the ONH to provide insight into these mechanisms, in many cases, depends on our ability to replicate the physiological environment, which in turn requires knowledge of tissue biomechanical properties. The majority of mechanical data published to date regarding the ONH has been obtained from tensile testing, yet compression has been shown to be the main mode of deformation in the ONH under elevated IOP. We have thus tested pig and rat ONH tissue using unconfined cyclic compression. The material constants C1, obtained from fitting the stress vs. strain data with a neo-Hookean material model, were 428 [367, 488] Pa and 64 [53, 76] Pa (mean [95% Confidence Interval]) for pig and rat optic nerve head, respectively. Additionally, we investigated the effects of strain rate and tissue storage on C1 values. These data will inform future efforts to understand and replicate the in vivo biomechanical environment of the ONH.
MicroTester/MicroSquisher	2019	3D printed micro-scale force gauge arrays to improve human cardiac tissue maturation and enable high throughput drug testing	Acta Biomaterialia	X. Ma, S. Dewan, J. Liu, M. Tang, K. Miller, C. Yu, N. Lawrence, A. McCulloch, S. Chen	University of California San Diego		Human induced pluripotent stem cell – derived cardiomyocytes (iPSC-CMs) are regarded as a promising cell source for establishing in-vitro personalized cardiac tissue models and developing therapeutics. However, analyzing cardiac force and drug response using mature human iPSC-CMs in a high-throughput format still remains a great challenge. Here we describe a rapid light-based 3D printing system for fabricating micro-scale force gauge arrays suitable for 24-well and 96-well plates that enable scalable tissue formation and measurement of cardiac force generation in human iPSC-CMs. We demonstrate consistent tissue band formation around the force gauge pillars with aligned sarcomeres. Among the different maturation treatment protocols we explored, 3D aligned cultures on force gauge arrays with in-culture pacing produced the highest expression of mature cardiac marker genes. We further demonstrated the utility of these micro-tissues to develop significantly increased contractile forces in response to treatment with isoproterenol, levosimendan, and omecamtiv mecarbil. Overall, this new 3D printing system allows for high flexibility in force gauge design and can be optimized to achieve miniaturization and promote cardiac tissue maturation with great potential for high-throughput in-vitro drug screening applications.
MicroTester/MicroSquisher	2019	Rapid pacing by circulating traveling waves improves maturation of hiPSC-derived cardiomyocytes in self-organized tissue ring	bioRxiv	J. Li, L. Zhang, L. Yu, I. Minami, M. Horning, J. Dong, J. Qiao, N. Fujimoto, Y. Shiba, Y. Zhao, F. Tang, S. Miyagawa, Y. Chen, Y. Sawa, C. Tang, L. Liu	Osaka University Graduate School of Medicine		Directed differentiation methods allow acquisition of high-purity cardiomyocytes (CMs) differentiated from human induced pluripotent stem cells (hiPSCs); however, their immaturity characteristic limits their application for drug screening and regenerative therapy. The rapid electrical pacing of cardiomyocytes have been used for efficiently promoting the maturation of cardiomyocytes, here we describe a simple device in modified culture plate on which hiPSC-derived CMs (hiPSC-CMs) can form three-dimensional self-organized tissue rings (SOTRs). Using calcium imaging, we show that within the ring, traveling waves (TWs) of action potential spontaneously originated and ran robustly at a frequency up to 4 Hz. After 2 weeks, SOTRs with TW training showed matured features including structural organization, increased cardiac-specific gene expression, enhanced Ca2+-handling properties, an increased oxygen-consumption rate, and enhanced contractile force. We subsequently used a mathematical model to interpret the origination, propagation, and long-term behavior of the TWs within the SOTRs. This new idea for spontaneous hiPSC-CM maturation also has potential for pacing the electrical excitable cells such as neuron and retina cells for various applications.
MicroTester/MicroSquisher	2019	Biomechanical Characterization of Mouse Sclera in Myopia	Molecular & Cellular Biomechanics	C. Ethier, D. Brown, E. Landis, M. Pardue	Georgia Institute of Technology and Emory University		Myopia, or near-sightedness, is a common ocular condition in which the eye elongates excessively. Development of myopia is associated with, and thought to be facilitated by, changes in the biomechanical properties of the sclera (the white part of the eye). We characterized scleral biomechanics in a mouse model of myopia using unconfirmed compression testing and biphasic theory to extract scleral permeability, in- plane scleral tensile modulus, and through-plane scleral compressive modulus. We find that myopia reduces in-plane tensile modulus and permeability, consistent with scleral tissue remodeling. Such biomechanical outcome measures may offer advantages over more traditional assessments of myopia-associated changes in the small mouse eye.
MicroTester/MicroSquisher	2019	Rapid fabrication of collagen bundles mimicking tumor-associated collagen signatures	bioRxiv	X. Gong, J. Kulwatno, K. Mills	Rensselaer Polytechnic Institute		Stromal collagen surrounding a solid tumor tends to present as dense, thick bundles. The collagen bundles are remodeled during tumor progression: first tangential to the tumor boundary (indicating growth) and later perpendicular to the tumor boundary (indicating likely metastasis). Current reconstituted-collagen in vitro tumor models are unable to recapitulate the in vivo structural features of collagen bundling and alignment. Here, we present a rapid yet simple procedure to fabricate collagen bundles with an average thickness of 9 µm, compared to the reticular dense collagen nanofiber (~900 nm-diameter, on average) prepared using common protocols. The versatility of the collagen bundles was demonstrated with their incorporation into two in vitro models where the thickness and alignment of the collagen bundles resembled the various in vivo arrangements. First, collagen bundles aligned by a microfluidic device elicited cancer cell contact guidance and enhanced their directional migration. Second, the presence of the collagen bundles in a bio-inert agarose hydrogel was shown to provide a highway for cancer cell invasion. The unique structural features of the collagen bundles advance the physiological relevance of in vitro collagen-based tumor models for accurately capturing cancer cell-stroma interactions.
MicroTester/MicroSquisher	2019	Chitosan Microbeads Produced by One-Step Scalable Stirred Emulsification: A Promising Process for Cell Therapy Applications	ACS Publications	Y. Alinejad, C. Bitar, K. Villegas, S. Perignon, C. Hoesli, S. Lerouge	École de technologie supérieure (ETS)		Cell microencapsulation is a promising approach to improve cell therapy outcomes by protecting injected cells from rapid dispersion and allowing bidirectional diffusion of nutrients, oxygen, and waste that promote cell survival in the target tissues. Here, we describe a simple and scalable emulsification method to encapsulate animal cells in chitosan microbeads using thermosensitive gel formulations without any chemical modification and cross-linker. The process consists of a water-in-oil emulsion where the aqueous phase droplets contain cells (L929 fibroblasts or human mesenchymal stromal cells), chitosan acidic solution and gelling agents (sodium hydrogen carbonate and phosphate buffer or beta-glycerophosphate). The oil temperature is maintained at 37 °C, allowing rapid physical gelation of the microbeads. Alginate beads prepared with the same method were used as a control. Microbeads with a diameter of 300–450 μm were successfully produced. Chitosan and alginate (2% w/v) microbeads presented similar rigidity in compression, but chitosan microbeads endured >80% strain without rupture, while alginate microbeads presented fragile breakage at <50% strain. High cell viability and metabolic activity were observed after up to 7 days in culture for encapsulated cells. Mesenchymal stromal cells encapsulated in chitosan microbeads released higher amounts of the vascular endothelial growth factor after 24 h compared to the cells encapsulated in manually cast macrogels. Moreover, microbeads were injectable through 23G needles without significant deformation or rupture. The emulsion-generated chitosan microbeads are a promising delivery vehicle for therapeutic cells because of their cytocompatibility, biodegradation, mechanical strength, and injectability. Clinical-scale encapsulation of therapeutic cells such as mesenchymal stromal cells in chitosan microbeads can readily be achieved using this simple and scalable emulsion-based process.
MicroTester/MicroSquisher	2019	Rapid Production of Cell-Laden Microspheres Using a Flexible Microfluidic Encapsulation Platform	Small Journal	W. Seeto, Y. Tian S. Pradhan, P. Kerscher, E. Lipke	Auburn University		This study establishes a novel microfluidic platform for rapid encapsulation of cells at high densities in photocrosslinkable microspherical hydrogels including poly(ethylene glycol)-diacrylate, poly(ethylene glycol)-fibrinogen, and gelatin methacrylate. Cell-laden hydrogel microspheres are advantageous for many applications from drug screening to regenerative medicine. Employing microfluidic systems is considered the most efficient method for scale-up production of uniform microspheres. However, existing platforms have been constrained by traditional microfabrication techniques for device fabrication, restricting microsphere diameter to below 200 μm and making iterative design changes time-consuming and costly. Using a new molding technique, the microfluidic device employs a modified T-junction design with readily adjustable channel sizes, enabling production of highly uniform microspheres with cell densities (10–60 million cells mL−1) and a wide range of diameters (300–1100 μm), which are critical for realizing downstream applications, through rapid photocrosslinking (≈1 s per microsphere). Multiple cell types are encapsulated at rates of up to 1 million cells per min, are evenly distributed throughout the microspheres, and maintain high viability and appropriate cellular activities in long-term culture. This microfluidic encapsulation platform is a valuable and readily adoptable tool for numerous applications, including supporting injectable cell therapy, bioreactor-based cell expansion and differentiation, and high throughput tissue sphere-based drug testing assays.
MicroTester/MicroSquisher	2020	Nanostructured Degradable Macroporous Hydrogel Scaffolds with Controllable Internal Morphologies via Reactive Electrospinning	Acta Biomaterialia	F. Xu, I. Gough, J. Dorogin, H. Sheardown, T. Hoare	McMaster University		Creating micro/nanostructured hydrogels with tunable morphologies under cell-friendly processing conditions would enable rational engineering of hydrogel scaffolds for targeted biomedical applications. Herein, an all-aqueous single-step reactive electrospinning method is applied to prepare hydrogel networks with controlled morphologies on both the nanoscale and the microscale. Hydrazide and aldehyde-functionalized poly(oligo ethylene glycol methacrylate) (POEGMA) are co-spun from a double barrel syringe together with poly(ethylene oxide) (PEO) as an electrospinning aid. By varying the concentrations and molecular weights of PEO and/or POEGMA, various morphologies from pure fibers to beaded fibers to bead network morphologies with tunable bead sizes can be fabricated, all of which remain monolithically stable in water due to the dynamic covalent crosslinks formed within the gel structure. The rates and magnitudes of swelling, degradation, and mechanics of the resulting scaffolds can be tuned by independently controlling gel morphologies on the nanoscale (i.e. crosslink density within the gel) and the microscale (i.e. the network structure formed), with an atypical independence of swelling relative to the mechanics and degradation rate observed. Furthermore, the internal morphology of the networks is demonstrated to systematically alter both the cell responses within the scaffolds and the rate of protein release from the scaffolds, with small fibers showing optimal cell proliferation, bead networks exhibiting the slowest protein release kinetics and very high maintained cell viabilities post-electrospinning, and beaded fibers showing intermediate properties.
MicroTester/MicroSquisher	2020	Myofibroblast activation in synthetic fibrous matrices composed of dextran vinyl sulfone	Acta Biomaterialia	C. Davidson, D. Jayco, D. Matera, S. DePalma, H. Hiraki, W. Wang, B. Baker	University of Michigan		Mechanical interactions between fibroblasts and their surrounding extracellular matrix (ECM) guide fundamental behaviors such as spreading, migration, and proliferation that underlie disease pathogenesis. The challenges of studying ECM mechanics in vivo have motivated the development of in vitro models of the fibrous ECM in which fibroblasts reside. Natural materials such as collagen hydrogels bear structural and biochemical resemblance to stromal ECM, but mechanistic studies in these settings are often confounded by cell-mediated material degradation and the lack of structural and mechanical tunability. Here, we established a new material system composed of electrospun dextran vinyl sulfone (DexVS) polymeric fibers. These fibrous matrices exhibit mechanical tunability at both the single fiber (80–340 MPa) and bulk matrix (0.77–11.03 kPa) level, as well as long-term stability in mechanical properties over a two-week period. Cell adhesion to these matrices can be either user-defined by functionalizing synthetic fibers with thiolated adhesive peptides or methacrylated heparin to sequester cell-derived ECM proteins. We utilized DexVS fibrous matrices to investigate the role of matrix mechanics on the activation of fibroblasts into myofibroblasts, a key step of the fibrotic progression. In contrast to previous findings with non-fibrous hydrogel substrates, we find that fibroblasts in soft and deformable matrices exhibit increased spreading, focal adhesion formation, proliferation, and myofibroblast activation as compared to cells on stiffer matrices with equivalent starting architecture.
MicroTester/MicroSquisher	2020	Multiparametric Analysis of Tissue Spheroids Fabricated from Different Types of Cells	Biotechnology Journal	E. Koudan, A. Gryadunova, P. Karalkin, J. Korneva, N. Meteleva, I. Babichenko, A. Volkov, S. Rodionov, V. Parfenov, F. Pereira, Y. Khesuani, V. Mironov, E. Bulanova	Laboratory for Biotechnological Research 3D Bioprinting Solutions		Reproducible, scalable, and cost effective fabrication and versatile characterization of tissue spheroids (TS) is highly demanded by 3D bioprinting and drug discovery. Consistent geometry, defined mechanical properties, optimal viability, appropriate extracellular matrix/cell organization are required for cell aggregates aimed for application in these fields. A straightforward procedure for fabrication and systematic multiparametric characterization of TS with defined properties and uniform predictable geometry employing non‐adhesive technology is suggested. Applying immortalized and primary cells, the reproducibility of spheroid generation, the strong correlation of ultimate spheroid diameter, and growth pattern with cell type and initial seeding concentration are demonstrated. Spheroids viability and mechanical properties are governed by cell derivation. In this study, a new decision procedure to apply for any cell type one starts to work with to prepare and typify TS meeting high quality standards in biofabrication and drug discovery is suggested.
MicroTester/MicroSquisher	2020	Shape-morphing Living Composites	Science Advances	L. Rivera-Tarazona, V. Bhat, H. Kim, Z. Campbell, T. Ware	University of Texas at Dallas		This work establishes a means to exploit genetic networks to create living synthetic composites that change shape in response to specific biochemical or physical stimuli. Baker’s yeast embedded in a hydrogel forms a responsive material where cellular proliferation leads to a controllable increase in the composite volume of up to 400%. Genetic manipulation of the yeast enables composites where volume change on exposure to L-histidine is 14× higher than volume change when exposed to D-histidine or other amino acids. By encoding an optogenetic switch into the yeast, spatiotemporally controlled shape change is induced with pulses of dim blue light (2.7 mW/cm2). These living, shape-changing materials may enable sensors or medical devices that respond to highly specific cues found within a biological milieu.
MicroTester/MicroSquisher	2020	Development of Scaffolds with Adjusted Stiffness for Mimicking Disease-Related Alterations of Liver Rigidity	Journal of Functional Biomaterials	M. Ruoß, S. Rebholz, M. Weimer, C. Grom-Baumgarten, K. Anthanasopulu, R. Kemkemer, H. Käß, S. Ehnert, A. Nussler	Eberhard Karls University		Drug-induced liver toxicity is one of the most common reasons for the failure of drugs in clinical trials and frequent withdrawal from the market. Reasons for such failures include the low predictive power of in vivo studies, that is mainly caused by metabolic differences between humans and animals, and intraspecific variances. In addition to factors such as age and genetic background, changes in drug metabolism can also be caused by disease-related changes in the liver. Such metabolic changes have also been observed in clinical settings, for example, in association with a change in liver stiffness, a major characteristic of an altered fibrotic liver. For mimicking these changes in an in vitro model, this study aimed to develop scaffolds that represent the rigidity of healthy and fibrotic liver tissue. We observed that liver cells plated on scaffolds representing the stiffness of healthy livers showed a higher metabolic activity compared to cells plated on stiffer scaffolds. Additionally, we detected a positive effect of a scaffold pre-coated with fetal calf serum (FCS)-containing media. This pre-incubation resulted in increased cell adherence during cell seeding onto the scaffolds. In summary, we developed a scaffold-based 3D model that mimics liver stiffness-dependent changes in drug metabolism that may more easily predict drug interaction in diseased livers.
MicroTester/MicroSquisher	2020	Shape recoverable and mechanically robust cellulose aerogel beads for efficient removal of copper ions	Chemical Engineering Journal	C. Tang, P. Brodie, Y. Li, N. Grishkewich, M. Brunsting, K. Tam	University of Waterloo		Developing bio-based absorbents with high removal capacity towards heavy metals is one of the key strategies to manage water pollution worldwide. Robust cellulose aerogel beads (CGP, diameter of 3–4 mm) with high amine content were prepared by cross-linking polyethylenimine (PEI) to cellulose nanofibrils (CNF) using 3-glycidyloxypropyl) trimethoxysilane (GPTMS). The mass ratio between PEI and GPTMS (1:1 or 3:1), crosslinking mechanism, morphologies, elemental compositions, total amine contents, mechanical strengths as well as the adsorption capacity and kinetic of the aerogel beads were investigated. Small CGP beads possessed several desirable characteristics, such as high amine group content (5.74 mmol/g for CGP3 beads), large maximum Cu (II) adsorption capacity (163.40 mg/g for CGP3 beads), very fast adsorption rate (<10 h to reach equilibrium), high shape recovery (2.00% plastic deformation for CGP3 beads at 50% strain), robust mechanical strength (stress around 10.5 KPa at 50% strain), and they could be regenerated and reused.
UniVert	2016	Development Of An Infusion Bioreactor For The Accelerated Preparation Of Decellularized Skeletal Muscle Scaffolds	Cell Culture and Tissue Engineering	B. M. Kasukonis, J.T. Kim, T.A. Washington, J.C. Wolchok	University of Arkansas		The implantation of decellularized tissue has shown effectiveness as a strategy for the treatment of volumetric muscle loss (VML) injuries. The preparation of decellularized tissue typically relies on the diffusion driven removal of cellular debris. For bulky tissues like muscle, the process can be lengthy, which introduces opportunities for both tissue contamination and degradation of key ECM molecules. In this study we report on the accelerated preparation of decellularized skeletal muscle (DSM) scaffolds using a infusion system and examine scaffold performance for the repair of VML injuries. The preparation of DSM scaffolds using infusion was dramatically accelerated. As the infusion rate (1% SDS) was increased from 0.1 to 1 and 10ml/hr, the time needed to remove intracellular myoglobin and actin decreased from a maximum of 140 ± 3hrs to 45 ± 3hrs and 10 ± 2hrs respectively. Although infusion appeared to remove cellular debris more aggressively, it did not significantly decrease the collagen or glycosaminoglycan composition of DSM samples when compared to un-infused controls. Infusion prepared DSM samples retained the aligned network structure and mechanical integrity of control samples. Infusion prepared DSM samples supported the attachment and in-vitro proliferation of myoblast cells and was well tolerated by the host when examined in-vivo. Cell/Tissue: decellularized skeletal muscle
UniVert	2016	The Global Mechanical Properties And Multi-Scale Failure Mechanics Of Heterogeneous Human Stratum Corneum	Acta Biomaterialia	X. Liu, J. Cleary, G.K. German	Binghamton University		The outermost layer of skin, or stratum corneum, regulates water loss and protects underlying living tissue from environmental pathogens and insults. With cracking, chapping or the formation of exudative lesions, this functionality is lost. While stratum corneum exhibits well defined global mechanical properties, macroscopic mechanical testing techniques used to measure them ignore the structural heterogeneity of the tissue and cannot provide any mechanistic insight into tissue fracture. As such, a mechanistic understanding of failure in this soft tissue is lacking. This insight is critical to predicting fracture risk associated with age or disease. In this study, we first quantify previously unreported global mechanical properties of isolated stratum corneum including the Poisson’s ratio and mechanical toughness. African American breast stratum corneum is used for all assessments. We show these parameters are highly dependent on the ambient humidity to which samples are equilibrated. A multi-scale investigation assessing the influence of structural heterogeneities on the microscale nucleation and propagation of cracks is then performed. At the mesoscale, spatially resolved equivalent strain fields within uniaxially stretched stratum corneum samples exhibit a striking heterogeneity, with localized peaks correlating closely with crack nucleation sites. Subsequent crack propagation pathways follow inherent topographical features in the tissue and lengthen with increased tissue hydration. At the microscale, intact corneocytes and polygonal shaped voids at crack interfaces highlight that cracks propagate in superficial cell layers primarily along intercellular junctions. Cellular fracture does occur however, but is uncommon. Cell/Tissue: breast stratum corneum
UniVert	2017	A Highly Deformable Conducting Traces For Printed Antennas And Interconnects: Silver/Fluoropolymer Composite Amalgamated By Triethanolamine	Flexible and Printed Electronics	A. Kumar, H. Saghlatoon, T-G. La, M. M. Honari, H. Charaya, H.A. Damis, R. Mirzavand, P. Mousavi, H-J. Chung	University of Alberta		Stretchable conducting traces are the key component to realize wearable healthcare electronics; a conductor material that can withstand high strain conditions can be crucial. Here, we describe a simple fabrication pathway to achieve stretchable conductive ink for printing. Specifically, silver flakes and fluorine rubber are amalgamated by the aid of triethanolamine (TEA), which enhanced the compatibility of the components in solution state where methylisobutylketone is the co-solvent. Moreover, TEA plasticizes the composites after the printing and drying of the solvent, causing the composite to deform freely without losing conductivity. The composite exhibits a conductivity value of 8.49 × 104 S m−1 at rest. The printed composite itself is not mechanically resilient after plastic deformation, but it has remarkable adhesion on elastomeric substrates. Thus, the printed ink on elastomers allows stretchable trace that can accommodate repeated stretching/releasing cycles. We fabricate and characterize stretchable printed antenna with three different designs (loop, patch, and bowtie) for the application of skin-adhesive electronics. Cell/Tissue: silver flakes fluorine rubber elastomers
UniVert	2017	Self-Reinforcing Graphene Coatings On 3d Printed Elastomers For Flexible Radio Frequency Antennas And Strain Sensors	Flexible and Printed Electronics	X. Li, M.M. Honari, Y. Fu, A. Kumar, H. Saghlatoon	University of Alberta		The emergence of the Internet of Things (IoT) necessitates the development of electronic components with various form factors and mechanical properties. 3D printing is an effective tool to realize objects with arbitrary form factors. Various 3D printable materials have recently been commercialized; among them, stretchable materials are particularly useful in the IoT because they enable adaptability in the dimensional change of the electronics. Most of these stretchable materials are, however, not electrically conductive; conductive coating can enable the functionality. Here, we propose a self-reinforcing conductive coating strategy, which reduced graphene oxide (RGO) self-assembles to wrap graphene nanoflakes (GNF) as a conductive binder that can also achieve mechanical integrity. The conductivity of the GNF-RGO coating reaches 4.47 × 104 S m−1. To demonstrate the potential applications of the GNF-RGO coating, applying the coating on 3D printed porous elastomers enabled flexible radio frequency (RF) antennas and strain sensors. The RF antenna shows high radiation efficiency and maintains excellent performance under bending conditions. The coating also produces a strain sensor with a gauge factor of ~13 up to 40% of strain. We foresee that the electrically conductive GNF-RGO composite coating can provide versatile functionalization strategy in flexible electronics and in wearable biomedical devices. Cell/Tissue: graphene nanoflakes reduced graphene oxide
UniVert	2018	Mechanical Strength Of Nafion®/Zro2 Nano-Composite Membrane	International Journal of Manufacturing, Materials and Mechanical Engineering (IJMMME)	R. Sigwadi, F. Nemavhola, M. Dhlamini, T. Mokrani	University of South Africa		The mechanical stability of modified membranes has become a priority for fuel cell applications as the membranes must endure all the fuel cell operations (to prevent crossover of the fuel while still conducting). Their mechanical stress and yielding stress in the recast and impregnation methods compared with the commercial Nafion® membrane were observed under tensile tests. The modulus of elasticity of wet commercial Nafion117 membrane, Nafion®/ Zr-0, Nafion®/Zr-50 and Nafion®/ Zr-80 membranes and Nafion®/ Zr-100 nano-composite membrane using impregnation methods in the region between 0 and 0.23 strain were determined to be 4817.5 kPa, 2434.7 kPa, 1872.4 kPa, 2092.1 kPa and 2661.4 kPa respectively. The tensile strength of the dry nano-composite membrane prepared using the recast method is higher than the wet nano-composite membrane prepared using the recast methods. It was found that the impregnation method plays an important role in strengthening the nan-composite membranes. Cell/Tissue: proton exchange membrane fuel cells
UniVert	2018	Three-Dimensional Printing Of Bisphenol A-Free Polycarbonates	Applied Materials and Interfaces	W. Zhu, S-H. Pyo, S. You, C. Yu, J. Alido, J. Liu, Y. Leong, S. Chen	University of California San Diego		Polycarbonates are widely used in food packages, drink bottles, and various healthcare products such as dental sealants and tooth coatings. However, bisphenol A (BPA) and phosgene used in the production of commercial polycarbonates pose major concerns to public health safety. Here, we report a green pathway to prepare BPA-free polycarbonates (BFPs) by thermal ring-opening polymerization and photopolymerization. Polycarbonates prepared from two cyclic carbonates in different mole ratios demonstrated tunable mechanical stiffness, excellent thermal stability, and high optical transparency. Three-dimensional (3D) printing of the new BFPs was demonstrated using a two-photon laser direct writing system and a rapid 3D optical projection printer to produce structures possessing complex high-resolution geometries. Seeded C3H10T1/2 cells also showed over 95% viability with potential applications in biological studies. By combining biocompatible BFPs with 3D printing, novel safe and high-performance biomedical devices and healthcare products could be developed with broad long-term benefits to society.
UniVert	2018	Fabrication Of Functionalized Citrus Pectin/Silk Fibroin Scaffolds For Skin Tissue Engineering.	Journal of Biomedical Materials Research	S. Tukkan, D. Atila, A. Akdaq, A. Tezcaner	Middle Eastern Technical U		In this study, novel porous three-dimensional (3D) scaffolds from silk fibroin (SF) and functionalized (amidated and oxidized) citrus pectin (PEC) were developed for skin tissue engineering applications. Crosslinking was achieved by Schiff's reaction in borax presence as crosslinking coordinating agent and CaCl2 addition. After freeze-drying and methanol treatment, plasma treatment (10 W, 3 min) was applied to remove surface skin layer formed on scaffolds. 3D matrices had high porosity (83%) and interconnectivity with pore size about 120 µm that providing suitable microenvironment for cells. Modifications on PEC chain and crosslinking of scaffolds were verified by fourier-transform infrared spectroscopy (FTIR) analysis and spectrophotometric assay. Scaffolds showed low weight loss (21.3% in 40 days) and high water uptake ability in phosphate-buffered saline (800% in 24 h). Mechanical properties of 3D matrices satisfied the stability of scaffolds under compressive stress and supported adhesion, proliferation and penetration of fibroblast cells. Our results suggested that modified PEC-SF scaffolds would be proposed for use in tissue engineered skin dermal substitutes.
UniVert	2018	Rapid Continuous 3d Printing Of Customizable Peripheral Nerve Guidance Conduits	Materials Today	W. Zhu, K.R. Tringale, S.A. Woller, S. You, S. Johnson, H. Shen, J. Schimelman, M. Whitney, J. Steinauer, W. Xu, T.L. Yaksh, Q.T. Nguyen, S.Chen	University of California San Diego		Engineered nerve guidance conduits (NGCs) have been demonstrated for repairing peripheral nerve injuries. However, there remains a need for an advanced biofabrication system to build NGCs with complex architectures, tunable material properties, and customizable geometrical control. Here, a rapid continuous 3D-printing platform was developed to print customizable NGCs with unprecedented resolution, speed, flexibility, and scalability. A variety of NGC designs varying in complexity and size were created including a life-size biomimetic branched human facial NGC. In vivo implantation of NGCs with microchannels into complete sciatic nerve transections of mouse models demonstrated the effective directional guidance of regenerating sciatic nerves via branching into the microchannels and extending toward the distal end of the injury site. Histological staining and immunostaining further confirmed the progressive directional nerve regeneration and branching behavior across the entire NGC length. Observational and functional tests, including the von Frey threshold test and thermal test, showed promising recovery of motor function and sensation in the ipsilateral limbs grafted with the 3D-printed NGCs.
UniVert	2018	Functionalization Of Silk Fibroin Through Anionic Fibroin Derived Polypeptides	Biomedical Materials	G. Griffanti, M. James-Bhasin, I. Donelli, G. Freddi, S.N. Nazhat	McGill		While silk fibroin (SF)-based fibrous matrices are often considered as templates to mimic the native biomineralization process, their limited ability to induce apatite deposition hinders their potential applications in bone tissue engineering. In this study, it was hypothesized that the incorporation of anionic fibroin derived polypeptides (Cs), generated through the α-chymotrypsin digestion of SF, into SF would induce apatite deposition. The effect of Cs incorporation and content on the mineralization of fibrous, electrospun (ES) SF matrices, was assessed in simulated body fluid (SBF). Moreover, the potential role of Cs in mediating the proliferation and osteoblastic differentiation of seeded mesenchymal stem cells (MSCs), in vitro, was also investigated. Methylene blue staining indicated that the ES SF matrices became increasingly negatively charged with an increase in Cs content. Furthermore, the mechanical properties of the ES SF matrices were modulated through variations in Cs content. Their subsequent immersion in SBF demonstrated rapid mineralization, attributable to the carboxyl groups provided by the negatively charged Cs polypeptides, which served as nucleation sites for apatite deposition. Seeded MSCs attached on all scaffold types with differences observed in metabolic activities when cultured in osteogenic medium. Relative to basal medium, there was an up-regulation of alkaline phosphatase, runt related transcription factor 2 and osteocalcin in osteogenic medium (at days 14 and 21). Cell-induced mineralized matrix deposition appeared to be accelerated on Cs incorporated ES SF suggesting an osteoinductive potential of these polypeptides. In sum, the ability to incorporate Cs into SF scaffolds offers promise in bone tissue engineering applications.
UniVert	2018	A Novel Gelma-Phema Hydrogel Nerve Guide For The Treatment Of Peripheral Nerve Damages	International Journal of Biological Macromolecules	T.D. Usal, D. Yucel, V. Hasirci	METU		Damage to the nervous system due to age, diseases or trauma may inhibit signal transfer along the nervous system. Nerve guides are used to treat these injuries by bridging the proximal and the distal end together. The design of the guide is very important for the reconnection of the severed axons. Methacrylated gelatin-poly(2-hydroxyethylmethacrylate) (GelMA-pHEMA) hydrogel was produced as the outer part of the nerve guide. pHEMA was added in various amounts into GelMA and increased the mechanical strength which is needed for the suturability of the guide. Porosity (15–70%), pore size (10–35 μm), water content (42–92%), and mechanical strength (65–710 kPa) of GelMA-pHEMA hydrogels were found to be suitable for nerve tissue engineering applications. Schwann cells attached and proliferated on GelMA, GelMA-pHEMA (5:5), and pHEMA hydrogels. Providing guidance is very important in the development of a nerve guide due to the anisotropic nature of the nerve tissue. Therefore, gelatin-poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) aligned fiber mats were used inside of the nerve guide. High degree of alignment with low deviation (7°) of this mats provided PC12 cell alignment throughout the fibers. Combination of GelMA-pHEMA (5:5) hydrogel and gelatin-PHBV aligned mat would provide an ideal nerve guide for the treatment of peripheral nerve damages.
UniVert	2018	Microstructure And Mechanical Properties Of Stainless Steel 316l Vertical Struts Manufactured By Laser Powder Bed Fusion Process	Materials Science and Engineering	X. Wang, J.A. Muniz-Lerma, O. Sanchez-Mata, M.A. Shandiz, M. Brochu	McGill		Stainless steel 316L (SS316L) vertical struts with various diameters ranging from 0.25 mm to 5 mm were manufactured by laser powder bed fusion (LPBF) process. A systematic investigation was conducted on the microstructure and mechanical properties of the produced struts. The struts possessed hierarchical microstructures consisting of cellular sub-grain structures inside columnar grains. The primary dendrite arm spacing (PDAS) of the cellular sub-grains decreased monotonically with increasing strut diameter until reaching a plateau after 1 mm. In contrast, the columnar grain width did not show a clear relationship with respect to the variation in the strut diameter. A <110> to <100> texture transition along the building direction (BD) of the struts was observed as the strut diameter decreased from 5 mm to 0.25 mm, which was attributed to the change of the heat extraction direction. Microstructure-property relations were established via Hall-Petch type correlations between the PDAS and the microhardness as well as the PDAS and the strengths of the struts, suggesting the importance of the role played by the cellular sub-grain structures in the strengthening of LPBF manufactured SS316L. Electron backscatter diffraction (EBSD) analysis confirmed that the strong <110> texture within the thicker struts promoted the twinning-induced plasticity, and thus resulted in a better strength-ductility combination compared with that of the thinner struts with <100> texture or weak <110> texture.
UniVert	2018	One-Step Fabrication Of Apatite-Chitosan Scaffold As A Potential Injectable Construct For Bone Tissue Engineering	Carbohydrate Polymers	K. Jahan, M. Mekhail, M. Tabrizian	McGill		Biomineralization of soft scaffolds is a new venture in bone tissue engineering. This work aimed to develop a new injectable and in-situ gelling soft scaffold with chitosan and apatites through a rapid purine-crosslinking reaction. The scaffolds were fabricated by mixing chitosan, adenosine diphosphate and biominerals in an ‘all-in-one-step’ procedure. The gelling of chitosan via the crosslinker occurs in <4 s as measured by impedance spectroscopy. These soft gels could retain up to 4 times their weight in water. Spectroscopy showed the formation of ionic bonds between chitosan and the apatites. Morphological analyses revealed an interconnected, highly porous structure, with pore size ranging from 200 nm-200 μm that was maintained even with the biominerals. Rheology showed a viscoelastic behavior of the solutions and the elastic behavior of the sponges, and therefore their injectability potential. in vitro studies showed good cell adhesion and morphology, as well as potential use for bone tissue engineering applications.
UniVert	2018	Skin Stiffness In Ray‐Finned Fishes: Contrasting Material Properties Between Species And Body Regions	Journal of Morphology	C.P. Kenaley, A. Sanin, J. Ackerman, J. Yoo, A. Alberts	Boston College		The skin of aquatic vertebrates surrounds all the mechanical lineages of the body and must, therefore, play an important role in locomotion. A cross‐woven collagenous dermal design has converged across several clades of vertebrates. Despite this intriguing pattern, the biomechanical role of skin in swimming fishes remains largely unknown. A direct force transmission role for fish skin has been proposed, a hypothesis that is supported by the arrangement of the connective tissues linking the skin to the axial musculature. To evaluate this direct force‐transmission hypothesis, we undertook hundreds of uniaxial tensile tests on skin samples from coho salmon (Oncorhynchus kisutch), Florida pompano (Trachinotus carolinus), and red snapper (Lutjanus campechanus). To do this, we developed highly precise, low‐cost, custom‐built material testing units. To augment our data, we also assembled a data set of skin stiffness of four additional species of actinopterygians fishes from previously published studies. We found that stiffness varies significantly between species and that the skin of our study species was increasingly stiff along a rostrocaudal gradient. Placing our results in the context of the limited body of previous work, we found that species with lower skin stiffness exhibit shorter propulsive wavelengths and low thrust production at the caudal fin and species with higher skin stiffness possess longer propulsive wavelengths and high thrust production at the caudal fin. In addition, we found that mean collagen fiber angle was close to 50° and that fiber angle was lower in posterior samples than in anterior and midlateral samples. Taken as a whole, our mechanical and morphological results support the hypothesis that the skin functions as an important direct force‐transmission device in actinopterygians whereby muscular force generated in anterior myotomes is transmitted to the posterior of the body through the increasingly stiff skin.
UniVert	2019	Preparation And Characterization Of Poly (Ε-Caprolactone) Scaffolds Modified With Cell-Loaded Fibrin Gel	International Journal of Biological Macromolecules	E. Malikmammadov, T. E. Tanir, A. Kiziltay, N. Hasirci	Middle Eastern Technical U		Poly(ε-caprolactone) (PCL) is one of the most commonly used polymers in the production of tissue engineered scaffolds for hard tissue treatments. Incorporation of cells into these scaffolds significantly enhances the healing rate of the tissue. In this study, PCL scaffolds were prepared by wet spinning technique and modified by addition of fibrinogen in order to form a fibrin network between the PCL fibers. By this way, scaffolds would have micro- and nanofibers in their structures. Drying of the wet spun constructs was achieved by application of ethanol dehydration or freeze drying techniques. Fibrinogen solutions (as low: 2 mg/mL; or high: 10 mg/mL concentrations) were added onto the scaffolds and fibrin formation was achieved via fibrinogen crosslinking. Results showed that ethanol dehydration led to film-like coating on the fibers while freeze-drying led to nanofiber bridges between PCL fibers establishing an interconnected web in the structure. Mechanical properties of the scaffolds were improved in the presence of the fibrin net. After the seeding of Saos-2 cells, higher attachment and homogeneous distribution of the cells was achieved on the samples modified with high concentration of fibrinogen. These scaffolds can be good candidates for the treatment of problematic bone defects.
UniVert	2019	Characterization Of Single Crystalline Austenitic Stainless Steel Thin Struts Processed By Laser Powder Bed Fusion	Scripta Materialia	X. Wang, J.A. Muniz-Lerma, O. Sanchez-Mata, M.A. Shandiz, N. Brodusch, R. Gauvin, M. Brochu	McGill		Single crystalline austenitic stainless steel struts with a diameter of 250 μm were successfully fabricated using laser powder bed fusion (LPBF). The struts exhibited a strong 〈110〉 texture parallel to the building direction (BD). Excellent strength-ductility combination was achieved for the struts, which is attributed to the strengthening of the fine cellular sub-grains and the twinning-induce plasticity (TWIP) effect. After deformation, dynamic recrystallization (DRX) was observed and its presence is closely related to the unique microstructure of the single crystalline struts.
UniVert	2019	Poly(Ester Amide) Particles For Controlled Delivery Of Celecoxib	Journal of Biomedical Materials Research	I. J. Villamagna, T. N. Gordon, M. B. Hurtig, F. Beier, E. R. Gillies	Western University		Many potential pharmacological treatments for osteoarthritis can result in undesirable side effects due to the systemic administration of drugs, making the direct delivery of drugs to joints an attractive alternative. Poly(ester amide)s (PEAs) have been shown to exhibit promising properties for the development of particle‐based intra‐articular delivery vehicles. However, a limited range of PEA structures has been investigated. In this study, we prepared and characterized the properties of two different PEA particles composed of l‐phenylalanine, sebacic acid, and either 1,4‐butanediol or 1,8‐octanediol (PBSe and POSe, respectively). The anti‐inflammatory drug celecoxib (CXB) was encapsulated into the particles. Despite minor structural differences, PBSe and POSe exhibited different thermal and mechanical properties, and encapsulation of CXB influenced these properties. PBSe‐CXB particles provided a slower release of drug in vitro relative to POSe‐CXB. Toxicity studies showed that particles without drug exhibited low toxicity to ATDC5 and C2C12 cells, while the PBSe‐CXB particles exhibited concentration‐dependent toxicity. Host response to the particles was evaluated in an ovine model. No adverse effects were observed following intra‐articular injection and it was observed that the particles diffused into the surrounding tissues. This work shows the importance of structural tuning in PEA delivery vehicles and demonstrates their potential for further development. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2019.
UniVert	2019	Effect Of Drug Loading On The Properties Of Temperature‐Responsive Polyester‐Poly(Ethylene Glycol)‐Polyester Hydrogels	Polymer International	D. A. Prince, I. J. Villamagna, C. C. Hopkins, J. R. de Bruyn, E. R. Gillies	Western University		Hydrogels that can undergo gelation upon injection in vivo are promising systems for the site‐specific delivery of drugs. In particular, some thermo‐responsive gels require no chemical additives but simply gel in response to a change from a lower temperature to physiological temperature (37 °C). The gelation mechanism does not involve covalent bonds, and it is possible that incorporation of drugs into the hydrogel could disrupt gelation. We investigated the incorporation of drugs into thermo‐responsive hydrogels based on poly(ε‐caprolactone‐co‐lactide)‐block‐poly(ethylene glycol)‐b‐poly(ε‐caprolactone‐co‐lactide) (PCLA‐PEG‐PCLA). Significant differences in properties and in the response to incorporation of the anti‐inflammatory drug celecoxib (CXB) were observed as the PEG block length was varied from 1500 to 3000 g mol‐1. Linear viscoelastic moduli of a PCLA‐PEG‐PCLA hydrogel containing a 2000 g mol‐1 PEG block were least affected by the incorporation of CXB and this gel also exhibited the slowest release of CXB, so the incorporation of phenylbutazone, methotrexate, ibuprofen, diclofenac, and etodolac were also investigated for this hydrogel. Different drugs resulted in varying degrees of syneresis of the hydrogels, suggesting that they interact with the polymer networks in different ways. In addition, the drugs had varying effects on the viscoelastic and compressive moduli of the gels. The results showed that the effects of drug loading on the properties of thermo‐responsive hydrogels can be substantial and depend on the drug. For applications such as intra‐articular drug delivery, in which the mechanical properties of the hydrogel are important, these effects should thus be studied on a case by case basis.
UniVert	2019	A 3d Printed Pcl/Hydrogel Construct With Zone-Specific Biochemical Composition Mimicking That Of The Meniscus	Biofabrication	G. Bahcecioglu, N. Hasirci, B. Bilgen, V. Hasirci	Middle East Technical University		Engineering the meniscus is challenging due to its bizonal structure; the tissue is cartilaginous at the inner portion and fibrous at the outer portion. Here, we constructed an artificial meniscus mimicking the biochemical organization of the native tissue by 3D printing a meniscus shaped PCL scaffold and then impregnating it with agarose (Ag) and gelatin methacrylate (GelMA) hydrogels in the inner and outer regions, respectively. After incubating the constructs loaded with porcine fibrochondrocytes for 8 weeks, we demonstrated that presence of Ag enhanced glycosaminoglycan (GAG) production by about 4 fold (p < 0.001), while GelMA enhanced collagen production by about 50 fold (p < 0.001). In order to mimic the physiological loading environment, meniscus shaped PCL/hydrogel constructs were dynamically stimulated at strain levels gradually increasing from the outer region (2% of initial thickness) towards the inner region (10%). Incorporation of hydrogels protected the cells from the mechanical damage caused by dynamic stress. Dynamic stimulation resulted in increased ratio of collagen type II (COL 2) in the Ag-impregnated inner region (from 50% to 60% of total collagen), and increased ratio of collagen type I (COL 1) in the GelMA-impregnated outer region (from 60% to 70%). We were able to engineer a meniscus, which is cartilage-like at the inner portion and fibrocartilage-like at the outer portion. Our construct has a potential for use as a substitute for total meniscus replacement.
UniVert	2019	Evaluation Of Human Dental Pulp Stem Cells Behavior On A Novel Nanobiocomposite Scaffold Prepared For Regenerative Endodontics	Material Science and Engineering	R. M. Rad, D. Atila, E. E. Akgun, Z. Evis, D. Keskin, A. Tezcaner	Middle East Technical University		Dental caries is a dental disease affecting public health, which results in many socio-economic consequences. This disease causes loss of tooth hard tissue and subsequent inflammation and loss of the dental pulp. In this study, it was aimed to develop and characterize boron (B) modified bioactive glass nanoparticles (BG-NPs) containing cellulose acetate/oxidized pullulan/gelatin (CA/ox-PULL/GEL) three dimensional scaffolds with tubular morphology for dentin regeneration. 3D nanobiocomposite structures were prepared by thermally induced phase separation and porogen leaching methods and characterized by in vitro degradation analysis, water absorption (WA) capacity measurement, SEM, in vitro biomineralizationanalysis, porosity measurement and mechanical tests. Scaffolds lost about (30–40)% of their weight during one month and WA capacity decreased with increase in immersion time in phosphate buffer solution (PBS) during one month. According to SEM, aligned and tubular structures were formed with mean diameter of about 11 μm, and BG-NPS were distributed evenly in all parts of the scaffolds. Scaffolds (without BG-NPs) possessed the highest porosity percentage. Addition of 10% BG-NPs improved the mechanical properties of scaffolds. Scaffolds surfaces were fully covered by calcium phosphate (Ca-P) deposits after conditioning in simulated body fluid for 14 days with higher quantity of deposition in groups with inclusion of B-BG-NPs. Human dental pulp stem cells (hDPSCs) were isolated from third molar teeth and used in cell culture studies. In all groups, cells adhered well 1 day after culture. Group B14–10 showed a slight increase of proliferation than group (without BG-NPs) after 7 days of incubation. Alkaline phosphatase activity (ALP) and intracellular calcium amounts increased significantly 14 days after incubation with highest values in B14–10 and B14–20 groups. Confocal laser scanning microscopy(CSLM) analysis, showed that cells on B14–10 and B21–10 scaffold groups, spread more 14 days after culture, and they also possessed extended processes specific to odontoblasts. Alizarin Red quantification showed that the highest calcium deposition was observed on B14–10 scaffolds. Immunohistochemical and Von Kossa stainings showed that scaffolds positively affected the odontoblastic differentiation of the hDPSCs. In this work, results showed that boron modified BG-NPs (B-BG-NPs) incorporated dentin-like constructs bring a new approach for dental tissue engineering applications.
UniVert	2019	A Comparative Study Of Monoaxial And Coaxial Pcl/Gelatin/Poloxamer 188 Scaffolds For Bone Tissue Engineering	International Journal of Polymeric Materials and Polymeric Biomaterials	H. A. Alam, A. D. Dalgic, A. Tezcaner, C. Ozen, D. Keskin	Middle East Technical University		Structure of fibrous scaffolds varies depending on the electrospinning method and parameters which will define cell fate. In this study, monoaxial and coaxial electrospun PCL/gelatin/P-188 scaffolds were characterized in terms of morphology, chemistry, mechanical property, water retention, weight loss, porosity and release properties. Coaxial fibers supported duel release of model hydrophilic protein (Beta-lactoglobulin) and hydrophobic agent vitamin K2. Novel PCL/P-188(Shell):Gelatin/P-188(Core) coaxial scaffold had superior stability, average weight loss, no swelling behavior, controlled release of Beta-lactoglobulin and vitamin K2, enhanced Saos-2 cell viability and ALP activity and could be a good candidate as a bone tissue scaffold.
UniVert	2019	Development of in-situ Forming Hydrogels for Intra-articular Drug Delivery	Western Libraries	A. Prince	Western University		Hydrogels are 3-dimensional crosslinked polymer networks that can absorb significant amounts of water. The physical properties associated with hydrogels affords them resemblance to biological tissues making them good candidates for biomedical applications. Many pharmaceuticals, specifically non-steroidal anti-inflammatory drugs (NSAIDs), have poor aqueous solubility, which limits their bioavailability and efficacy. People suffering from chronic osteoarthritis (OA) are required to frequently take large doses to mitigate pain, which can lead to serious side effects. Hydrogels are good strategies to deliver NSAIDs via articular injection because they can form solid gels in situ. This thesis describes the synthesis, formulation, mechanical testing, in vitro and in vivo trials of triblock copolymer (PCLA-PEG-PCLA) hydrogels. We observed that drug-loading can have negatively impact the gelation behaviour. Block lengths were tuned, and we found that using PEG2000 was better to maintain gelation integrity upon drug-loading. We further looked to improve mechanical properties by investigating a redox initiated crosslinking system with methacrylated end-caps. These gels were able to maintain or improve gelation upon loading of various drugs. The optimized chemically crosslinked formulation was able to provide a sustained release of celecoxib in horses for up to 32 d. These results are significant because currently, there is no curative or pain relief option for OA patients beyond 16 d while limiting systemic drug concentration.
UniVert	2019	Development of a Novel Functionally Graded Membrane Containing Boron Modified Bioactive Glass Nanoparticles for Guided Bone Regeneration	Journal of Tissue Engineering and Regenerative Medicine	R. M. Rad, D. Atila, Z. Evis, D. Keskin, A. Tezcaner	Middle East Technical University		Barrier membranes are used in periodontal tissue engineering for successful neo‐bone tissue formation and prevention of bacterial colonization. We aimed to prepare and characterize novel 7% boron modified bioactive glass (7B‐BG) containing bilayered membrane for this end. We hypothesized that presence of 7B‐BG could promote structural and biological properties of GBR membrane. Cellulose acetate (CA) layer was prepared by solvent casting and functionally graded layer of CA/gelatin/BG nanoparticles was prepared by electrospinning. 0B‐BG, and 7B‐BG were synthesized by quick alkali‐mediated sol‐gel method and were characterized by SEM and FT‐Raman spectroscopy. Membranes were cross‐linked with glutaraldehyde to preserve their stability. SEM analysis showed the asymmetric nature of membranes consisting of a smooth membrane layer and a rough surface composed of (0B‐BG, 7B‐BG) containing nanofibers. 7B‐BG addition increased surface wettability (from 110.5° ± 0.8 to 73.46° ± 7.6) and biodegradability of the membranes. Additionally, a significant increase in Ca‐P layer formation was observed in 7B‐BG containing group after 1‐week incubation in stimulated body fluid. 7B‐BG incorporation resulted in a decrease in tensile strength and Young's modulus values. Human Dental Pulp Stem Cells (hDPSCs) showed better attachment, spreading and proliferation on 7B‐BG containing bilayered membranes. Osteogenic differentiation analysis revealed higher ALP enzyme activity of cells (~1.5 fold), higher intracellular calcium deposition (~2 fold), and higher calcium deposition revealed by Alizarin red staining on 7B‐BG containing bilayered membranes. Overall, results suggested that functionally graded bilayered membranes hold potential for GBR applications in regenerative dentistry.
UniVert	2019	Anatomical Meniscus Construct with Zone Specific Biochemical Composition and Structural Organization	bioRxiv	G. Bahcecioglu, N. Hasirci, B. Bilgen, V. Hasirci	Middle East Technical University		A PCL/hydrogel construct that would mimic the structural organization, biochemistry and anatomy of meniscus was engineered. The compressive (380 ± 40 kPa) and tensile modulus (18.2 ± 0.9 MPa) of the PCL scaffolds were increased significantly when constructs were printed with a shifted design and circumferential strands mimicking the collagen organization in native tissue (p<0.05). Gene expression of human fibrochondrocytes in agarose (Ag), gelatin methacrylate (GelMA), and GelMA-Ag hydrogels was significantly higher than that of cells on the PCL scaffolds after a 21-day culture. GelMA exhibited the highest level of collagen type I (COL1A2) mRNA expression, while GelMA-Ag exhibited the highest level of aggrecan (AGG) expression (p<0.001, compared to PCL). GelMA and GelMA-Ag exhibited a high level of collagen type II (COL2A1) expression (p<0.05, compared to PCL). Anatomical scaffolds with circumferential PCL strands were impregnated with cell-loaded GelMA in the periphery and GelMA-Ag in the inner region. GelMA and GelMA-Ag hydrogels enhanced the production of COL 1 and COL 2 proteins after a 6-week culture (p<0.05). COL 1 expression increased gradually towards the outer periphery, while COL 2 expression decreased. We were thus able to engineer an anatomical meniscus with a cartilage-like inner region and fibrocartilage-like outer region.
UniVert	2019	Niobium Pentoxide and Hydroxyapatite Particle Loaded Electrospun Polycaprolactone/Gelatin Membranes for Bone Tissue Engineering	Colloids and Surfaces B: Biointerfaces	N. Marins, B. Lee, R. Silva, A. Raghavan, N. Carreno, K. Grandfield	Federal University of Pelotas		Effective methods of accelerating the bone regeneration healing process are in demand for a number of bone-related diseases and trauma. This work developed scaffolds with improved properties for bone tissue engineering by electrospinning composite polycaprolactone-gelatin-hydroxyapatite-niobium pentoxide (PGHANb) membranes. Composite membranes, with average fiber diameters ranging from 123 to 156 nm, were produced by adding hydroxyapatite (HA) and varying concentrations of niobium pentoxide (Nb2O5) particles (0, 3, 7, and 10 wt%) to a polycaprolactone (PCL) and gelatin (GL) matrix prior to electrospinning. The morphology, mechanical, chemical and biological properties of resultant membranes were evaluated. Bioactivity was assessed using simulated body fluid (SBF) and it confirmed that the presence of particles induced the formation of hydroxyapatite crystals on the surface of the membranes. Samples were hydrophilic and cell metabolism results showed that the niobium-containing membranes were non-toxic while improving cell proliferation and differentiation compared to controls. This study demonstrated that electrospun membranes containing HA and Nb2O5 particles have potential to promote cell adhesion and proliferation while exhibiting bioactive properties. PGHANb membranes are promising candidates for bone tissue engineering applications.
UniVert	2019	Printability of Methacrylated Gelatin upon Inclusion of a Chloride Salt and Hydroxyapatite Nano‐Particles	Macromolecular Materials and Engineering	P. Comeau, T. Willett	University of Waterloo		3D biomaterial printing requires an ink to have suitable printability characteristics, as well as creating a final construct of controllable swelling and stiffness. To tune such properties, the impact of adding different levels of chloride salts (NaCl and CaCl2) and hydroxyapatite nano‐particles (nHA) to a highly concentrated and photo‐crosslinkable methacrylated gelatin (GelMA) is investigated. By adding up to 100 mm CaCl2 or 1.11 mNaCl, the GelMA viscosity decreases from that of control GelMA (no salt). Interestingly, a 25G needle and strong photo‐polymerization kinetics are able to overcome the low viscosity of the 50CaG ink during printing. Adding further CaCl2 increases GelMA viscosity, while decreasing both the swelling and dynamic modulus of the UV‐cured construct observed in water. As all UV‐cured constructs have a dynamic modulus greater than 1 MPa, this novel system is able to match the dynamic modulus of articular cartilage—a feat not previously reported for a GelMA‐based system. Lastly, nHA inclusion improves ink printability, as well as decreases swelling and increases dynamic modulus of the final construct. Overall, this study leads to the successful development of a new advanced functional ink which will be beneficial in the 3D printing of biomaterials toward tissue engineering applications.
UniVert	2019	A Smart Bandage Equipped with Miniaturized Needle Arrays and Wirelessly-Controlled Drug Delivery System for the Treatment of Chronic Wounds	University of Nebraska-Lincoln DigitalCommons	H. Derakhshandeh	University of Nebraska-Lincoln		Chronic wounds are one of the most challenging complications of diabetes and are the leading cause of non-traumatic limb amputation. Despite the progress in identifying factors and promising in vitro results for the treatment of chronic wounds, their clinical translation has been limited. Given the range of disruptive processes necessary for wound healing, different pharmacological agents are needed at different stages of tissue regeneration. This requires the capability of wearable devices to deliver agents to critical layers of the epidermis with minimum invasiveness. Here, for the first time, I have engineered a programmable platform which is capable of actively delivering a variety of drugs with independent temporal profiles through miniaturized needle arrays (MNAs) into deeper layers of the wound bed. The delivery of vascular endothelial growth factor (VEGF) through the MNAs demonstrated that, in addition to the selection of suitable therapeutics, the delivery method and their spatial distribution within the wound bed is equally important. Administration of VEGF to chronic dermal wounds of diabetic mice using the programmable platform showed a significant increase in wound closure, re-epithelialization, angiogenesis, and hair growth when compared to standard topical delivery of therapeutics.
UniVert	2019	The Effect of Halloysite Addition on the Material Properties of Chitosan–Halloysite Hydrogel Composites	MDPI	Y. Luo, D. Mills	Louisiana Tech University		Chitosan-based hydrogels are being widely used in biomedical applications due to their eco-friendly, biodegradable, and biocompatible properties, and their ability to mimic the extracellular matrix of many tissues. However, the application of chitosan hydrogels has been limited due to their inherent mechanical weakness. Halloysite nanotubes (HNTs) are naturally occurring aluminosilicate clay minerals and are widely used as a bulk filler to improve the performance characteristics of many polymeric materials. HNTs have also been shown to be a viable nanocontainer able to provide the sustained release of antibiotics, chemicals, and growth factors. This study’s objective was to develop a stable drug delivery chitosan/HNT nanocomposite hydrogel that is biocompatible, biodegradable, and provides sustained drug release. In this study, chitosan/HNTs hydrogels containing undoped or gentamicin-doped HNTs were combined in different wt./wt. ratios and cross-linked with tripolyphosphate. The effects of chitosan and HNTs concentration and combination ratios on the hydrogel surface morphology, degradability, and mechanical properties, as well as its drug release capability, were analyzed. The results clearly showed that the addition of HNTs improved chitosan mechanical properties, but only within a narrow range. The nanocomposite hydrogels provided a sustained pattern of drug release and inhibited bacterial growth, and the live/dead assay showed excellent cytocompatibility
UniVert	2019	Fabrication of 3D Conjugated Polymer Structures via Vat Polymerization Additive Manufacturing	Semantic Scholar	A. Cullen, A. Price	University of Western Ontario		Conjugated polymers are a class of electroactive active material that have been studied for their potential in advanced sensing and actuation technologies. Conventional methods to fabricate conjugated polymer transducers produce devices with planar morphologies that limit fabricated devices to simplistic linear or bending modes of operation. To overcome this limitation, this work develops a photosensitive polypyrrole formulation and light-based additive manufacturing method capable of realizing three-dimensional conjugated polymer structures with micro-scale features. The electroactivity and strain sensing properties of the photosensitive polymer formulation are reported to show the early development of this method towards new sensing and actuation technologies.
UniVert	2019	Design and fabrication of a low-cost pilot-scale melt-processing system	Polymer	D. Wirth, J. Pokorski	University of California		Melt processing of polymeric materials is a ubiquitous technique for forming, shaping, refining and homogenizing polymers and polymer composites. Melt-processing techniques are the primary manufacturing method of consumer and industrial thermoplastic parts, especially when using commodity polymers with high-throughput production. Melt-processing, however, is underutilized in academic laboratories when developing high value-added materials due to the capital expense of the equipment and relatively large-scale required to carry out such processing. These concerns make pilot-scale melt-processing challenging, particularly for new polymers or polymer composites where materials can only be generated in small-scale at reasonable costs. The current study designs and evaluates a bench-top, sub-milliliter volume extrusion and injection-molding device, which sources parts from current 3D print technology at minimal expense. The plans presented will open this convenient technique to academic research laboratories interested in pilot-scale experiments. A systematic approach to melt processing of PLA, PLGA, and PCL polymer composites is demonstrated. Characterization of the dispersion of pharmaceuticals, small molecules and nanoparticles in melt processed polymers is presented as a demonstration of potential utility.
UniVert	2019	Materials and Processing for Wearable Healthcare Electronics Systems: Flexible Circuit Boards and Stretchable e-Textile Patches for Surface Electromyography	University of Alberta ERA	S. Qiu	University of Alberta		Skin-adhesive electrodes – more preferably, integrated electronic systems with embedded signal processing and transmittance – that can be worn comfortably on a patient have irreplaceable importance in healthcare. For example, surface electromyography (sEMG) is a non-invasive electrodiagnostic medical technique to detect, record, and interpret electric activity of groups of muscles from skin surface right above them. If skin-adhesive sEMG electrodes can be integrated with wireless data processing and transmittance system in a compact and wearable format, these devices can provide unprecedented opportunities for applications such as posture monitoring for rehab medicine, swallowing self-coaching, and muscle fatigue detector. However, there are stringent demands for form factors that are specific for the target locations in body, such as required level of flexibility, foldability and stretchability. The conversion of electronics into such advanced form factors has been arising the current challenges in the skin-adhesive electronic systems, such as poor reusability, complicated fabrication methods, and poor connection between soft and hard components in the electronics system. In this thesis, we have explored the potential solutions to solve these problems.Overall, this thesis is divided into two themes. In the first theme, we developed a versatile technology with capability of rapid, simple, customizable fabrication of flexible printed circuit boards (Flex-PCBs) and sensors. A simple and cost-effective wax-printing technique to pattern flexible interconnects allowed the system-level integration of sensing, processing, power, and wireless communication units. The flexible circuitry with reusable adhesive can follow the contour of curvilinear skin topology, thus allow enhanced wearability for skin-adhesive electronic systems. In the second theme, we developed textile-based electronics, also known as 'e-textiles', which offer stretchability and comfort for patients. A stretchable conductive silver ink was developed for ink-jet printing to pattern stretchable circuits on nano-textiles. The deep permeation of ink into nano-textiles resulted in stable adhesion, mechanically and electrically strengthen. With our custom-developed electronic circuits, we demonstrated a sEMG system with wireless data emission.
UniVert	2019	Surface Representation of the Urinary Bladder and Bio-mechanical Analysis by Linear Dynamic Model	Latin American Conference on Biomedical Engineering	K. Guzman-Diaz, A. Alvarado-Gonzalez, D. Herrera, A. Hernandez-Reynoso, S. Rodriguez-Reynoso, A. Garcia-Gonzalez	Tecnologico de Monterrey		The biomechanical characterization of Urinary Bladder is essential to create new therapeutic tools as artificial tissue replacement or electrical stimulators. In this paper, we applied a linear model, the so-called viscoelastic Voigt’s model to calculate the viscosity and elasticity in three regions of a bladder under the experimental condition of tensile stresses. Additionally, we acquired the tridimensional surface of urinary bladders by a cloud of points that were processed in digital format. Viscosity and elasticity are represented on the surface reconstruction. Three bovine bladders were processed with an average sample viscosity of 89.33 × 10−6 Ns/mm2 and average elasticity of 1.3 × 10−3 N/mm, the isotropic condition was evaluated. The digital representation includes a cloud of points of 9 × 106 elements on average. These results are the base of a future computational simulation platform.
UniVert	2019	Hyperelastic characterization of synthetic mesh for abdominal wall hernia repair	MRS Advances	J. Ortiz Ortiz, G. Carbajal de la Torre, M. Villagomez Galindo, M. Antonio Espinosa Medina, H. Aguilar Rodriguez	Universidad Michoacana de San Nicolas de Hidalgo		Hernia is defined as the protrusion of one or several internal organs through an opening in the cavity that contains them due to a tissue defect, abdominal wall surgery by means of synthetic meshes is the most common method used for hernia repair, however, postsurgical effects can range from some discomfort, to chronic pain and even the reappearance of the hernia due to a poor mechanical adaptability between the synthetic tissue and the host tissue. The knowledge of the mechanical properties of the materials involved in hernia repair is fundamental in the understanding and subsequent solution of this type of problems. In this work, experimental data were obtained by means of uniaxial tensile tests in two perpendicular directions of commercial meshes used in hernia repair. The tests were carried out on the UniVert® machine of the CellScale® brand. Anisotropic mechanical behavior is observed due to the structure of the mesh and the interaction between each of the yarns that make it up. The data found vary with respect to the direction of traction and also has non-linear hyperelastic behavior, so the adjustment of curves was made through a hyperelastic model in the COMSOL Multiphysics® software through the Levenberg-Marquardt Algorithm for the characterization of these materials.
UniVert	2019	An Alternative Approach to the Surface Methacrylation of Non-stoichiometric Hydroxyapatite Nanoparticles for Use in Bone-Inspired Composites	Frontiers in Materials	P. Comeau, T. Willett	University of Waterloo		Modification of the inorganic particle surface is one approach toward enhancing the mechanical and swelling properties of a bone-inspired composite. In this study we sought to manipulate the surface of non-stoichiometric nHA particles by utilizing a gentler methacrylation approach with two different methacrylates—methacrylic anhydride (MAh) and glycidyl methacrylate (GMA). While silanization of nHA (Si-nHA) is a recognized functionalization approach, one notable disadvantage is the need for relatively harsh reaction conditions. Here, ATR-FTIR and liquid state 13C-NMR provided evidence for a stronger affinity of the nHA surface for methacrylic anhydride compared to glycidyl methacrylate under the same reaction conditions; fewer GMA molecules reacted with the nHA surface. However, the affinity of MAh is more electrostatic in nature, whereas GMA attachment involves some covalent bond formation. In addition, while in water, there was no detectable dependence of particle settling on methacrylate choice, in ethanol, native nHA and nHA methacrylated with GMA each had detectably greater stability in suspension than the other sample groups. Lastly, the addition of GMA-nHA detectably increased the dynamic stiffness and did not impact the swelling in 37°C water of a GelMA-based composite compared to that observed upon adding native nHA. Overall, the GMA-based methacrylation approach was found to be a viable alternative to silanization and offers possibilities for future fabrication of bone-inspired composites.
UniVert	2019	Hybrid Cornea: Cell Laden Hydrogel Incorporated Decellularized Matrix	ACS Biomaterials	M. Uyaniklar, G. Günal, A. Tevlek, P. Hosseinian, H. Aydin	Hacettepe University		The decellularization protocols applied on the corneal stromal constructs in the literature usually fail to provide a corneal matrix with sufficient mechanical and optical properties since they alter the extracellular matrix (ECM) microstructure. In this study, to overcome these limitations, a hybrid cornea stromal construct was engineered by combining gelatin methacrylate (GelMA) and decellularized ECM. Photo-cross-linking of impregnated cell laden GelMA in situ using different UV cross-linking energies (3200, 6210, and 6900 μJ/cm2) and impregnation times (up to 24 h) within a decellularized bovine cornea enhanced light transmission and restored lost mechanical features following the harsh decellularization protocol. The light transmittance value for optimized hybrid constructs (53.6%) was increased nearly 10 fold compared to that of decellularized cornea (5.84%). The compressive modulus was also restored up to 6 fold with calculated values of 5040 and 870 kPa for the hybrid and decellularized samples, respectively. These values were found to be quite close to that of native cornea (48.5%, 9790 kPa). ATR-FTIR analyses were carried out to confirm the final chemical structure. The degradation profiles showed similar decomposition behaviors to that of native cornea. In vitro culture studies showed a high level of cell viability and cell proliferation rate was found remarkable up to the 14th day of the culture period regardless of selected UV energy level. The methodology used in the preparation of the hybrid cornea stromal constructs in this study is a promising approach toward the development of successful corneal transplants.
UniVert	2019	Synthesis of Crosslinkable Poly(ester amide)s for Cell Encapsulation and Delivery	Western Electronic Thesis and Dissertation Repository	Y. Liang	Western University		Tissue engineering using adipose-derived stromal cells (ASCs) shows promise for soft tissue regeneration. Biodegradable polymers are potential biomaterials as they support the growth and delivery of cells. Specifically, poly(ester amide)s (PEAs) are a class of biodegradable polymers with tunable structures that have been shown to exhibit low cytotoxicity and support the growth of various cell types. This thesis involved the development of new water soluble amino acid-based PEAs with crosslinkable moieties to enable formation of hydrogel scaffolds for ASC encapsulation. These hydrophilic phenylalanine-based and alanine-based PEAs were synthesized by solution polycondensation and photo-crosslinked into a series of hydrogels with and without poly(ethylene glycol) dimethacrylate, methacrylated chondroitin sulphate or decellularized adipose tissue, and hydrogel properties including gel content, equilibrium water content, and swelling were examined. ASC viability and adipogenesis were studied in selected PEA hydrogels, and the overall results demonstrated the potential of these new water soluble PEAs as biomaterials for adipose tissue engineering, as they provided a supportive environment for ASCs survival and adipogenic differentiation in culture.
UniVert	2019	Design of cell-instructiv Design of cell-instructive biomaterial scaffolds for inter e biomaterial scaffolds for intervertebral disc regeneration	Western Electronic Thesis and Dissertation Repository	N. Sharma	Western University		Biomaterials-based therapies targeting the nucleus pulposus (NP) have the potential to promote regeneration and restore mechanical function to the intervertebral disc. This study developed composite hydrogels incorporating decellularized NP (DNP) and assessed its effects on viability, retention and differentiation of U-CH1 cells, an NP progenitor-like cell line. A minimal protocol was developed to decellularize bovine NP that reduced nuclear content while preserving key extracellular matrix components predicted to be favourable for bioactivity. The resulting DNP demonstrated cell-instructive effects, supporting U-CH1 viability and retention within the hydrogels, and promoted the differentiation of the progenitor-like cells towards an NP-like phenotype. These studies established a 3-D platform that mimics the native NP microenvironment and holds promise for applications in cell culture and delivery. Further in vitro studies using this system will provide valuable insight into the effects of tissue-specific extracellular matrix on NP progenitor cell fate.
UniVert	2020	Process–Structure–Quality Relationships of Three-Dimensional Printed Poly(Caprolactone)-Hydroxyapatite Scaffolds	Tissue Engineering	S. Gerdes, A. Mostafavi, S. Ramesh, A. Memic, I. Rivero, P. Rao, A. Tamayol	University of Nebraska-Lincoln		Bone defects are common and, in many cases, challenging to treat. Tissue engineering is an interdisciplinary approach with promising potential for treating bone defects. Within tissue engineering, three-dimensional (3D) printing strategies have emerged as potent tools for scaffold fabrication. However, reproducibility and quality control are critical aspects limiting the translation of 3D printed scaffolds to clinical use, which remain to be addressed. To elucidate the factors that yield to the generation of defects in bioprinting and to achieve reproducible biomaterial printing, the objective of this article is to frame a systematic approach for optimizing and validating 3D printing of poly(caprolactone) (PCL)-hydroxyapatite (HAp) composite scaffolds. We delineate the effect of PCL-to-HAp ratio, print velocity, print temperature, and extrusion pressure on the architectural and mechanical properties of the 3D printed scaffold. Furthermore, we present an in situ image-based monitoring approach to quantify key quality-related aspects of constructs, such as the ability to deposit material consistently and print elementary shapes with fewer flaws. Our results show that small defects generated during the printing process have a significant role in lowering the mechanical properties of 3D printed polymeric scaffolds. In addition, the in vitro osteoinductivity of the fabricated scaffolds is demonstrated.
UniVert	2020	A Wirelessly Controlled Smart Bandage with 3D-Printed Miniaturized Needle Arrays	Advanced Functional Materials	H. Derakhshandeh, D. Aghabaglou, A. McCarthy, A. Mostafavi, C. Wiseman, Z. Bonick, I. Ghanavati, S. Harris, C. Kreikemeier-Bower, S. Basri, J. Rosenbohm, R. Yang, P. Mostafalu, D. Orgill, A. Tamayol	University of Nebraska-Lincoln		Chronic wounds are one of the most devastating complications of diabetes and are the leading cause of nontraumatic limb amputation. Despite the progress in identifying factors and promising in vitro results for the treatment of chronic wounds, their clinical translation is limited. Given the range of disruptive processes necessary for wound healing, different pharmacological agents are needed at different stages of tissue regeneration. This requires the development of wearable devices that can deliver agents to critical layers of the wound bed in a minimally invasive fashion. Here, for the first time, a programmable platform is engineered that is capable of actively delivering a variety of drugs with independent temporal profiles through miniaturized needles into deeper layers of the wound bed. The delivery of vascular endothelial growth factor (VEGF) through the miniaturized needle arrays demonstrates that, in addition to the selection of suitable therapeutics, the delivery method and their spatial distribution within the wound bed is equally important. Administration of VEGF to chronic dermal wounds of diabetic mice using the programmable platform shows a significant increase in wound closure, re-epithelialization, angiogenesis, and hair growth when compared to standard topical delivery of therapeutics.
UniVert	2020	Single-crystalline-like stainless steel 316L with different geometries fabricated by laser powder bed fusion	Progress in Additive Manufacturing	X. Wang, J. Muniz-Lerma, O. Sanchez-Mata, S. Atabay, M. Shandiz, M. Brochu	McGill University		In this study, we explored the feasibility of fabricating single-crystalline or single-crystalline-like stainless steel 316L (SS316L) with different geometries (thin struts, cubes, walls and a simulated pump impeller) using laser powder bed fusion (LPBF). The LPBF-fabricated SS316L thin struts possessed a single-crystalline core featuring a 〈110〉 ∥ building direction (BD) crystallographic texture. The cubes, walls and the pump impeller preserved this 〈110〉 ∥ BD texture and also exhibited a well-defined single-crystalline-like {110}〈001〉 Goss texture. Cellular sub-grain structures with their primary dendrite arm spacing (PDAS) values smaller than 1 μm were discovered in all the samples with their growth directions showing a 45° angular deviation from the BD. Nanoscale precipitates and dislocations were also found in the cellular sub-grain structures of the thin struts. The mechanical properties of different geometries (the thin struts, the walls, and the simulated pump impeller) were studied and compared. The anisotropic mechanical responses of the walls and the simulated pump impeller were correlated with their crystallographic textures.
UniVert	2020	Development and Characterization of a Benchtop Corneal Puncture Injury Model	Nature	E. Snider, L. Cornell, J. Acevedo, B. Gross, P. Edsall, B. Lund, D. Zamora	US Army Institute of Surgical Research		During recent military operations, eye-related injuries have risen in frequency due to increased use of explosive weaponry which often result in corneal puncture injuries. These have one of the poorest visual outcomes for wounded soldiers, often resulting in blindness due to the large variations in injury shape, size, and severity. As a result, improved therapeutics are needed which can stabilize the injury site and promote wound healing. Unfortunately, current corneal puncture injury models are not capable of producing irregularly shaped, large, high-speed injuries as seen on the battlefield, making relevant therapeutic development challenging. Here, we present a benchtop corneal puncture injury model for use with enucleated eyes that utilizes a high-speed solenoid device suitable for creating military-relevant injuries. We first established system baselines and ocular performance metrics, standardizing the different aspects of the benchtop model to ensure consistent results and properly account for tissue variability. The benchtop model was evaluated with corneal puncture injury objects up to 4.2 mm in diameter which generated intraocular pressure levels exceeding 1500 mmHg. Overall, the created benchtop model provides an initial platform for better characterizing corneal puncture injuries as seen in a military relevant clinical setting and a realistic approach for assessing potential therapeutics.
UStretch	2014	Intrastromal Application Of Riboflavin For Corneal Crosslinking	Investigative ophthalmology & Visual Science	T.G. Seiler, I. Fischinger, T. Senfft, G. Schmidinger, T. Seiler	The Institut für Refraktive und Ophthalmo-Chirurgie (IROC)		Purpose.: To experimentally evaluate the efficacy of corneal crosslinking (CXL) by injecting the photomediator riboflavin into the corneal stroma via intrastromal channels. Methods.: Five groups of pig corneas, nine each, were compared regarding stress–strain relationship and UV-absorption. Group 1 had intrastromal channels floated with riboflavin 0.5%-solution followed by UVA-irradiation (3 mW/cm2 for 30 minutes); group 2 was handled like group 1, but were irradiated with 9 mW/cm2 for 10 minutes; group 3 was treated according to the Dresden protocol (epi-off, 9 mW/cm2 for 10 minutes); group 4 had the identical channel system, no riboflavin but identical irradiation; group 5 with native corneas served as a control group. The intrastromal channels were created with a femtosecond laser. The stress–strain relations were measured in corneal strips using a uniaxial material tester at strains up to 12%. The UV-transmission of the corneas was measured in groups 1, 3, and 5. Results.: The stress needed for a 10% strain was significantly increased by 82% in the corneas treated with the Dresden protocol compared with native cornea (P = 0.0005). With intrastromal application of riboflavin the significant increase was 87% (P = 0.0005) in group 1 and 64% (P = 0.007) in group 2. The channel formation alone did not alter biomechanics (P = 0.923). The corneal UVA-transmission was 2.4% after intrastromal riboflavin application, 8.9% after the treatment according to the Dresden protocol, and 57.9% in native corneas. Conclusions.: The experiments demonstrate the intrastromal application of riboflavin by means of intrastromal channels a feasible “epi-on” approach for CXL. More experimental data are needed before clinical testing. Cell/Tissue: pig corneas corneal stroma
UStretch	2015	Development Of A Biological Scaffold Engineered Using The Extracellular Matrix Secreted By Skeletal Muscle Cells	Biomaterials Journal	S.A. Hurd, N.M. Bhatti, A.M. Walker, B.M. Kasukonis, J.C. Wolchok	University of Arkansas		The performance of implantable biomaterials derived from decellularized tissue, including encouraging results with skeletal muscle, suggests that the extracellular matrix (ECM) derived from native tissue has promising regenerative potential. Yet, the supply of biomaterials derived from donated tissue will always be limited, which is why the in-vitro fabrication of ECM biomaterials that mimic the properties of tissue is an attractive alternative. Towards this end, our group has utilized a novel method to collect the ECM that skeletal muscle myoblasts secrete and form it into implantable scaffolds. The cell derived ECM contained several matrix constituents, including collagen and fibronectin that were also identified within skeletal muscle samples. The ECM was organized into a porous network that could be formed with the elongated and aligned architecture observed within muscle samples. The ECM material supported the attachment and in-vitro proliferation of cells, suggesting effectiveness for cell transplantation, and was well tolerated by the host when examined in-vivo. The results suggest that the ECM collection approach can be used to produce biomaterials with compositions and structures that are similar to muscle samples, and while the physical properties may not yet match muscle values, the in-vitro and in-vivo results indicate it may be a suitable first generation alternative to tissue derived biomaterials. Cell/Tissue: cardiac tissue, myocardium arterial tissue cardiovascular tissue
UStretch	2016	Material Properties From Air Puff Corneal Deformation By Numerical Simulations On Model Corneas	PLOS One (Public Library of Science One)	N. Bekesi, C. Dorronsoro, A. de la Hoz, S. Marcos	Consejo Superior de Investigaciones Científicas (Spanish National Research Council)		Objective To validate a new method for reconstructing corneal biomechanical properties from air puff corneal deformation images using hydrogel polymer model corneas and porcine corneas. Methods Air puff deformation imaging was performed on model eyes with artificial corneas made out of three different hydrogel materials with three different thicknesses and on porcine eyes, at constant intraocular pressure of 15 mmHg. The cornea air puff deformation was modeled using finite elements, and hyperelastic material parameters were determined through inverse modeling, minimizing the difference between the simulated and the measured central deformation amplitude and central-peripheral deformation ratio parameters. Uniaxial tensile tests were performed on the model cornea materials as well as on corneal strips, and the results were compared to stress-strain simulations assuming the reconstructed material parameters. Results The measured and simulated spatial and temporal profiles of the air puff deformation tests were in good agreement (< 7% average discrepancy). The simulated stress-strain curves of the studied hydrogel corneal materials fitted well the experimental stress-strain curves from uniaxial extensiometry, particularly in the 0–0.4 range. Equivalent Young´s moduli of the reconstructed material properties from air-puff were 0.31, 0.58 and 0.48 MPa for the three polymer materials respectively which differed < 1% from those obtained from extensiometry. The simulations of the same material but different thickness resulted in similar reconstructed material properties. The air-puff reconstructed average equivalent Young´s modulus of the porcine corneas was 1.3 MPa, within 18% of that obtained from extensiometry. Conclusions Air puff corneal deformation imaging with inverse finite element modeling can retrieve material properties of model hydrogel polymer corneas and real corneas, which are in good correspondence with those obtained from uniaxial extensiometry, suggesting that this is a promising technique to retrieve quantitative corneal biomechanical properties. Cell/Tissue: porcine corneal tissue
UStretch	2018	Self-Assembled Collagen-Fibrin Hydrogel Reinforces Tissue Engineered Adventitia Vessels Seeded With Human Fibroblasts	Scientific Reports	B. Patel, Z. Xu, C. Pinnock, L. S. Kabbani, M. T. Lam	Wayne State		Efforts for tissue engineering vascular grafts focuses on the tunica media and intima, although the tunica adventitia serves as the primary structural support for blood vessels. In surgery, during endarterectomies, surgeons can strip the vessel, leaving the adventitia as the main strength layer to close the vessel. Here, we adapted our recently developed technique of forming vascular tissue rings then stacking the rings into a tubular structure, to accommodate human fibroblasts to create adventitia vessels in 8 days. Collagen production and fibril cross-linking was augmented with TGF-β and ascorbic acid, significantly increasing tensile strength to 57.8 ± 3.07 kPa (p = 0.008). Collagen type I gel was added to the base fibrin hydrogel to further increase strength. Groups were: Fibrin only; 0.7 mg/ml COL; 1.7 mg/ml COL; and 2.2 mg/ml COL. The 0.7 mg/ml collagen rings resulted in the highest tensile strength at 77.0 ± 18.1 kPa (p = 0.015). Culture periods of 1–2 weeks resulted in an increase in extracellular matrix deposition and significantly higher failure strength but not ultimate tensile strength. Histological analysis showed the 0.7 mg/ml COL group had significantly more, mature collagen. Thus, a hydrogel of 0.7 mg/ml collagen in fibrin was ideal for creating and strengthening engineered adventitia vessels.
UStretch	2018	Intraocular Photobonding To Enable Accommodating Intraocular Lens Function	TVST	N. Alejandre-Alba, R. Cutierrez-Contreras, Carlos Dorronsoro, S. Marcos			Accommodating intraocular lenses (A-IOLs) require capturing the ciliary muscle forces. Prior work demonstrated strong photo-initiated bonding between strips of capsular bag and poly(2-hydroxyethyl methacrylate); (pHEMA) polymer in an extraocular setting. We demonstrate that photobonding can be achieved intraocularly.
UStretch	2019	Anti-Fatigue-Fracture Hydrogels	Science Advances	S. Lin, X. Liu, J. Liu, H. Yuk, H. C. Loh, G. A. Parada, C. Settens, J. Song, A. Masic, G. H. McKinley, X. Zhao	MIT		The emerging applications of hydrogels in devices and machines require hydrogels to maintain robustness under cyclic mechanical loads. Whereas hydrogels have been made tough to resist fracture under a single cycle of mechanical load, these toughened gels still suffer from fatigue fracture under multiple cycles of loads. The reported fatigue threshold for synthetic hydrogels is on the order of 1 to 100 J/m2. We propose that designing anti-fatigue-fracture hydrogels requires making the fatigue crack encounter and fracture objects with energies per unit area much higher than that for fracturing a single layer of polymer chains. We demonstrate that the controlled introduction of crystallinity in hydrogels can substantially enhance their anti-fatigue-fracture properties. The fatigue threshold of polyvinyl alcohol (PVA) with a crystallinity of 18.9 weight % in the swollen state can exceed 1000 J/m2.
UStretch	2019	Repeated In Vitro Impact Conditioning Of Astrocytes Decreases The Expression And Accumulation Of Extracellular Matrix	Annals of Biomedical Engineering	A. Walker, J. Kim, J. Wyatt, A. Terlouw, K. Balachandran, J. Wolchok	University of Arkansas		Pathological changes to the physical and chemical properties of brain extracellular matrix (ECM) occur following injury. It is generally assumed that astrocytes play an important role in these changes. What remain unclear are the triggers that lead to changes in the regulation of ECM by astrocytes following injury. We hypothesize that mechanical stimulation triggers genotypic and phenotypic changes to astrocytes that could ultimately reshape the ECM composition of the central nervous system following injury. To explore astrocyte mechanobiology, an in vitro drop test bioreactor was employed to condition primary rat astrocytes using short duration (10 ms), high deceleration (150G) and strain (20%) impact stimuli. Experiments were designed to explore the effect of single and repeated impact (single vs. double) on mechano-sensitive behavior including cell viability; ECM gene (collagens I and IV, fibronectin, neurocan, versican) and reactivity gene [glial fibrillary acidic protein (GFAP), S100B, vimentin] expression; matrix regulatory cytokine secretion [matrix metalloproteinase 2 (MMP-2), tissue inhibitor of metalloproteinases 1 (TIMP1), transforming growth factor beta 1 (TGFβ1)]; and matrix accumulation [collagen and glycosaminoglycan (GAG)]. Experiments revealed that both ECM and reactive gliosis gene expression was significantly decreased in response to impact conditioning. The decreases for several genes, including collagen, versican, and GFAP were sensitive to impact number, suggesting mechano-sensitivity to repeated impact conditioning. The measured decreases in ECM gene expression were supported by longer-term in vitro experiments that demonstrated significant decreases in chondroitin sulfate proteoglycan (CSPG) and collagen accumulation within impact conditioned 3-D scaffolds accompanied by a 25% decrease in elastic modulus. Overall, the general trend across all samples was towards altered ECM and reactive gliosis gene expression in response to impact. These results suggest that the regulation of ECM production by astrocytes is sensitive to mechanical stimuli, and that repeated impact conditioning may increase this sensitivity.
UStretch	2019	Ingestible Hydrogel Device	Nature Communications	X. Liu, C. Steiger, S. Lin, G. A. Parada, J. Liu, H. F. Chan, H. Yuk, N. V. Phan, J. Collins, S. Tamang, G. Traverso, X. Zhao	MIT		Devices that interact with living organisms are typically made of metals, silicon, ceramics, and plastics. Implantation of such devices for long-term monitoring or treatment generally requires invasive procedures. Hydrogels offer new opportunities for human-machine interactions due to their superior mechanical compliance and biocompatibility. Additionally, oral administration, coupled with gastric residency, serves as a non-invasive alternative to implantation. Achieving gastric residency with hydrogels requires the hydrogels to swell very rapidly and to withstand gastric mechanical forces over time. However, high swelling ratio, high swelling speed, and long-term robustness do not coexist in existing hydrogels. Here, we introduce a hydrogel device that can be ingested as a standard-sized pill, swell rapidly into a large soft sphere, and maintain robustness under repeated mechanical loads in the stomach for up to one month. Large animal tests support the exceptional performance of the ingestible hydrogel device for long-term gastric retention and physiological monitoring.
UStretch	2019	Pure PEDOT: PSS Hydrogels	Nature Communications	B. Lu, H. Yuk, S. Lin, N. Jian, K. Qu, J. Xu, X. Zhao	MIT		Hydrogels of conducting polymers, particularly poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS), provide a promising electrical interface with biological tissues for sensing and stimulation, owing to their favorable electrical and mechanical properties. While existing methods mostly blend PEDOT:PSS with other compositions such as non-conductive polymers, the blending can compromise resultant hydrogels’ mechanical and/or electrical properties. Here, we show that designing interconnected networks of PEDOT:PSS nanofibrils via a simple method can yield high-performance pure PEDOT:PSS hydrogels. The method involves mixing volatile additive dimethyl sulfoxide (DMSO) into aqueous PEDOT:PSS solutions followed by controlled dry-annealing and rehydration. The resultant hydrogels exhibit a set of properties highly desirable for bioelectronic applications, including high electrical conductivity (~20 S cm−1 in PBS, ~40 S cm−1 in deionized water), high stretchability (> 35% strain), low Young’s modulus (~2 MPa), superior mechanical, electrical and electrochemical stability, and tunable isotropic/anisotropic swelling in wet physiological environments.
UStretch	2019	Muscle-like Fatigue-Resistant Hydrogels by Mechanical Training	National Academy of Sciences	S. Lin, J. Liu, X. Liu, X. Zhao	MIT		Skeletal muscles possess the combinational properties of high fatigue resistance (1,000 J/m2), high strength (1 MPa), low Young’s modulus (100 kPa), and high water content (70 to 80 wt %), which have not been achieved in synthetic hydrogels. The muscle-like properties are highly desirable for hydrogels’ nascent applications in load-bearing artificial tissues and soft devices. Here, we propose a strategy of mechanical training to achieve the aligned nanofibrillar architectures of skeletal muscles in synthetic hydrogels, resulting in the combinational muscle-like properties. These properties are obtained through the training-induced alignment of nanofibrils, without additional chemical modifications or additives. In situ confocal microscopy of the hydrogels’ fracturing processes reveals that the fatigue resistance results from the crack pinning by the aligned nanofibrils, which require much higher energy to fracture than the corresponding amorphous polymer chains. This strategy is particularly applicable for 3D-printed microstructures of hydrogels, in which we can achieve isotropically fatigue-resistant, strong yet compliant properties.
UStretch	2019	Prediction of Hyperelastic Material Properties of Nafion117 and Nafion/ZrO2 Nano-Composite Membrane	International Journal of Automotive and Mechanical Engineering	F. Nemavhola, R. Sigwadi	University of South Africa		This paper presents constitutive laws suitable for the prediction of mechanical behaviour of nano-composite membrane compared with the commercial membrane Nafion®117. The uniaxial tensile data of commercial Nafion®117 and Nafion®/ Zr-150 nano-composite membrane utilised for fitting hyperelastic models was determined experimentally. Several material models on mechanical behaviour of nano-composite and commercial Nafion® 117 membrane material was fitted to determined accuracy. In order to observe yield and fracture behaviour, the com-mercial Nafion®117 and Nafion®/ Zr-150 nano-composite membranes were loaded in uniaxial direction at a constant strain rate. To obtain the optimal material constants form six different material models considered in this study, the OriginLab® version 9 was used and the Leven-berg-Marquardt (M) optimization logarithm. Hyperplastic material models including Mooney-Rivlin, Yeoh, Ogden, Humphrey, Martins and Veronda-Westmann were selected to use in an inverse method to fit the experimental uniaxial data of nano-composite material. The hyper-plastic material parameters could then be used to simulate material behaviour of nano mem-brane using finite element analysis (FEA) technique. The procedure discussed in this paper could be used to accurately determine the constitutive parameters of various constitutive models of Polymer Nafion presented.
UStretch	2019	Ultraviolet Light Degrades the Mechanical and Structural Properties of Human Stratum Corneum	bioRxiv	Z. Lipsky, G. German	Binghamton University		Prolonged exposure of human skin to sunlight causes photodamage, which induces the early onset of wrinkles and increased tissue fragility. While solar ultraviolet (UV) light is considered to have the most damaging effect, the UV range that is most harmful remains a topic of significant debate. In this study, we take a first step towards elucidating biomechanical photoageing effects by quantifying how exposure to different UV ranges and dosages impacts the mechanical and structural properties of human stratum corneum (SC), the most superficial skin layer. Mechanical testing reveals that irradiation of isolated human SC to UVA (365 nm), UVB (302 nm), or UVC (265 nm) light with dosages of up to 4000 J/cm2 notably alters the elastic modulus, fracture stress, fracture strain, and work of fracture. For equivalent incident dosages, UVC degrades SC the greatest. However, upon discounting reflected and transmitted components of the incident light, a generalized scaling law relating the photonic energy absorbed by the SC to the energy cost of tissue fracture emerges. This relationship indicates that no one UV range is more damaging than another. Rather, the magnitude of absorbed UV energy governs the degradation of tissue mechanical integrity. Subsequent structural studies are performed to elucidate the cause of this mechanical degradation. UV absorption scales with the spatial dispersion of desmoglein 1 (Dsg 1), a component of corneocyte cell-cell junctions, away from intercellular sites. Combining both scaling laws, we establish a mechanical-structural model capable of predicting UV induced tissue mechanical integrity from Dsg 1 dispersion.
UStretch	2019	Enhancing the Mechanical Properties of Zirconia/Nafion® Nanocomposite Membrane through Carbon Nanotubes for Fuel Cell Application	Heliyon	R. Sigwadi, F. Nemavhola, M. Dhlamini, T. Mokrani	University of South Africa		Membranes are widely used daily, such as for filtration in reverse osmosis, or in the form of electrolyte membrane fuel cells. Modified Nafion® membranes were synthesised by impregnation and their mechanical properties were observed. The effect of the incorporation of a ZrO2-CNT nano-filler within Nafion® membrane on the thermal stability and crystallinity was investigated by TGA and XRD. Tensile test results show the increases in the mechanical properties of Nafion® 117 membranes impregnated with ZrO2-CNT when compared with that of commercial Nafion® 117 membranes. The results also show that adding ZrO2-CNT in Nafion® 117 membranes improves the water contact angle and water uptake, as it enhances water retention within the membrane. The SEM results indicated that ZrO2-CNT was well distributed in the Nafion® 117 membrane pores through the impregnation method.
UStretch	2019	Polyvinyl Alcohol-Poly Acrylic Acid Bilayer Oral Drug Delivery Systems: A Comparison Between Thin Films and Inverse Double Network Bilayers	Journal of Biomaterials Applications	S. Miar, C. Perez, J. Ong, T. Guda	University of Texas at San Antonio		Polymer network structures in the design of bilayer hydrogels for oral drug delivery systems contribute to the mechanical stability as well as the stimulus responsive drug release. We report a strategy to improve the stability of polyelectrolyte layer interfaces in bilayer hydrogels by employing concepts from tough interpenetrating hydrogel network design. Polyvinyl alcohol as the neutral substrate and Polyacrylic acid as the pH-sensitive layer were used to prepare bilayers capable of pH-sensitive bending behavior, and inverse double network structures were compared to conventional thin film coated bilayers in this study. The morphology of the bilayers observed after exposure to stimulus revealed that the polyacrylic acid layer was fragmented in conventional thin films. The delamination of the coating also had an adverse impact on bending behavior. Additionally, bilayers with inverse double network structure had a better pH-sensitive swelling behavior which was lacking in regular thin films. Controllable, pH-specific drug release behavior was a direct benefit of using the inverse double network structure, and was demonstrated by controlled vancomycin release under different pHs chosen to simulate the physiological milieu of exposure corresponding to hydrogel passage along the gastrointestinal tract. Additionally, the inverse double networks showed improved mucoadhesive properties and mechanical stability compared to the thin film bilayers. The use of inverse double network structured hydrogels has many potential applications in improving bilayer hydrogel stability; specifically, for drug delivery systems, designing hydrogel origamis, and to prepare soft material actuators.
UStretch	2019	A Bioink Blend for Rotary 3D Bioprinting Tissue Engineered Small-Diameter Vascular Constructs	Acta Biomaterialia	S. Freeman, R. Ramos, P. Chando, L. Zhou, K. Reeser, S. Jin, P. Soman, K. Ye	State University of New York		3D bioprinted vascular constructs have gained increased interest due to their significant potential for creating customizable alternatives to autologous vessel grafts. In this study, we developed a new approach for biofabricating fibrin-based vascular constructs using a novel rotary 3D bioprinter developed in our lab. We formulated a new bioink by incorporating fibrinogen with gelatin to achieve a desired shear-thinning property for rotary bioprinting. The blending of heat-treated gelatin with fibrinogen turned unprintable fibrinogen into a printable biomaterial for vessel bioprinting by leveraging the favorable rheological properties of gelatin. We discovered that the heat-treatment of gelatin remarkably affects the rheological properties of a gelatin-fibrinogen blended bioink, which in turn influences the printability of the ink. Further characterizations revealed that not only concentration of the gelatin but the heat treatment also affects cell viability during printing. Notably, the density of cells included in the bioinks also influenced printability and tissue volumetric changes of the printed vessel constructs during cultures. We observed increased collagen deposition and construct mechanical strength during two months of the cultures. The burst pressure of the vessel constructs reached 1110 mmHg, which is about 52% of the value of the human saphenous vein. An analysis of the tensile mechanical properties of the printed vessel constructs unveiled an increase in both the circumferential and axial elastic moduli during cultures. This study highlights important considerations for bioink formulation when bioprinting vessel constructs.
UStretch	2019	The Effect of Axial Torsion on the Mechanical Properties of the Annulus Fibrosus	Biomechanics	M. Harvey-Burgess, D. Gregory	Wilfrid Laurier University		This study exposed bovine intervertebral discs to static axial torsion in order to determine its effect on the inter- and intralamellar matrices. Axial torsion resulted in decreased strength and stiffness of the intralamellar matrix, which may make the intervertebral disc more susceptible to herniation.
UStretch	2019	The proton conductivity and mechanical properties of Nafion®/ ZrP nanocomposite membrane	Heliyon	R. Sigwadi, M. Dhlamini, T. Mokrani, F. Nemovhola, P. Nonjola, P. Msomi	University of South Africa		Zirconium phosphates (ZrP) were incorporated into Nafion® 117 membrane by impregnating method to obtain a reduced methanol permeation and improved proton conductivity for fuel cell application. The mechanical properties and water uptake of Nafion® membrane incorporated with zirconium phosphates nanoparticles was more improvement when compared to the commercial Nafion® 117, due to the presence of phosphoric acid within the nanoparticles. The effect of ZrP nano filler on the membrane structural morphology and thermal properties were investigated by Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), Thermal gravimetric analysis (TGA) and Scanning Electron Microscopy (SEM). The improved ion conductivity and decreased methanol permeability on the nanocomposite membranes showed a great potential for fuel cell applications. The nanocomposite membrane with high tensile strength was obtained due to the well dispersed zirconium phosphates within the Nafion® matrix.
UStretch	2019	Tissue-like Hydrogels by Design	MIT Libraries	S. Lin	MIT		Human bodies are mostly made of soft, wet yet robust biological hydrogels such as cartilages, ligaments, and muscles. The biological hydrogels commonly possess mechanical properties such as high toughness, resilience and fatigue resistance to guarantee the bodies' reliable functions and activities. While hydrogels with tissue-like mechanical properties are highly desirable in applications as diverse as tissue engineering, drug delivery and soft machines, these properties are rarely achieved in synthetic hydrogels. The first part of this dissertation is aimed to design synthetic hydrogels that possess tissue-like mechanical properties, including high toughness, resilience and fatigue threshold, through combined theory, modeling and experiments. First, we develop a coupled cohesive-zone and Mullins effect model to predict the fracture energies of tough hydrogels. Based on the model, we further provide a toughening diagram that can guide the design of new tough hydrogels.
UStretch	2020	Endothelial Safety and Efficacy of ex-vivo Collagen Cross Linking (CXL) of Human Corneal Transplants	American Journal of Ophthalmology	L. Jan, L. Maria, P. Niklas, F. Isaak, H. Christina, S. Gerald	Medical University of Vienna		Ex-vivo CXL of human donor tissue is an effective and safe procedure with no difference regarding efficacy between 2 commercially available deswelling media. Biochemical and -mechanical resistance were significantly increased after CXL. Patients requiring keratoplasty due to corneal melting might benefit from the strengthening effect of pre-operative CXL of donor tissue.
UStretch	2020	Strong adhesion of wet conducting polymers on diverse substrates	Science Advances	A. Inoue, H. Yuk, B. Lu, X. Zhao	MIT		Conducting polymers such as poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS), polypyrrole (PPy), and polyaniline (PAni) have attracted great attention as promising electrodes that interface with biological organisms. However, weak and unstable adhesion of conducting polymers to substrates and devices in wet physiological environment has greatly limited their utility and reliability. Here, we report a general yet simple method to achieve strong adhesion of various conducting polymers on diverse insulating and conductive substrates in wet physiological environment. The method is based on introducing a hydrophilic polymer adhesive layer with a thickness of a few nanometers, which forms strong adhesion with the substrate and an interpenetrating polymer network with the conducting polymer. The method is compatible with various fabrication approaches for conducting polymers without compromising their electrical or mechanical properties. We further demonstrate adhesion of wet conducting polymers on representative bioelectronic devices with high adhesion strength, conductivity, and mechanical and electrochemical stability.
BioTester	2020	Vortex fluidic mediated one-step fabrication of polyvinyl alcohol hydrogel films with tunable surface morphologies and enhanced self-healing properties	Science China Materials	J. Tavakoli, C. Raston, Y. Ma, Y. Tang	Flinders University		Previous strategies for controlling the surface morphologies of polyvinyl alcohol (PVA)-based hydrogels, including freeze-drying and electrospinning, require a posttreatment process, which can affect the final textures and properties of the hydrogels. Of particular interest, it is almost impossible to control the surface morphology during the formation of PVA hydrogels using these approaches. The strategy reported in this study used the novel vortex fluidic device (VFD) technology, which for the first time provided an opportunity for one-step fabrication of PVA hydrogel films. PVA hydrogels with different surface morphologies could be readily fabricated using a VFD. By also reducing the crosslinking agent concentration, a self-healing gel with enhanced fracture stress (60% greater than that of traditionally made hydrogel) was achieved. Interestingly, the associated selfhealing property remained unchanged during the 260-s mechanical testing performed with the strain rate of 5% s−1. The VFD can effectively tune the surface morphologies of the PVA-based hydrogels and their associated properties, particularly the self-healing property
BioTester	2020	An advanced 3D monofilament biosuture	South African Pharmaceutical Journal	K. de la Harpe, P. Kondiah, T. Marimuthu, L. du Toit, P. Kumar, Y. Choonara, V. Pillay	University of the Witwarersrand, SA		Sutures are one of the most widely used medical devices with employment in over 12 million procedures per year globally.1 Yet, the ideal suture material does not exist. Over the years scientists and surgeons alike have set out to find a suture material that is biocompatible, easy to handle, does not cause unnecessary tissue damage and creates an optimal environment for wound healing.2 This has led to the discovery of numerous suture materials ranging from silk and catgut in the early 1800s to synthetic polymers such as polylactic acid and polyglycolide that are currently in use.3 Sutures on the market today are associated with distinct disadvantages that threaten the success of the procedures they are used in. For example, sutures consisting of polyglactin have been found to cause severe tissue damage and necrosis through the release of acidic degradation products that result in strong inflammatory responses.4 Additionally, most suture materials are multifilament or braided materials that tend to cut through tissues causing cellular damage and extending the time of wound healing.5 Suture-related complications have been implicated in numerous clinical problems such as hematoma formation, pulmonary embolisms and false aneurysms. A retrospective study done by Starr et al., ound that 26 of 39 false aneurysm cases could be directly ascribed to insufficiencies in the suture material.6 There is, therefore, a clear yet understated need for a superior suture material that can limit suture-related complications and improve the success rate of surgical procedures.
UniVert	2020	Triethyleneglycol dimethacrylate addition improves the 3D-printability and construct properties of a GelMA-nHA composite system towards tissue engineering applications	Material Science and Engineering	P. Comeau, T. Willett	University of Waterloo		In tissue engineering, there is a growing interest in the development of 3D printable bone tissue-inspired nanocomposites. However, most such nanocomposites have poor mechanical properties, owing to poor dispersion of the mineral phase (e.g. nano-hydroxyapatite, nHA) within the organic phase (e.g. methacrylated gelatin, GelMA) and low volume fractions of each phase. Triethyleneglycol dimethacrylate (TEGDMA) is commonly added to dental resin-based composites to improve the properties of the dental resin. Here, the effects of substituting a portion of the water phase in a GelMA-nHA composite with TEGDMA were evaluated. TEGDMA improved the dispersion of nHA within the highly-concentrated GelMA-based composite ink, as well as increased the ink's shear yield strength and reduced the critical energy for ink cure. As a result, the printability of the composite ink was greatly improved upon TEGDMA inclusion. Lastly, while the swelling of the cast composite in 37 °C water increased slightly, the mechanical properties (tensile strength, toughness, and stiffness) of the cast composite increased by at least an order of magnitude upon TEGDMA addition, and all composites demonstrated MSC cytocompatibility after 24 h. Overall, TEGDMA shows promise as an additive to tune properties of the GelMA-nHA system towards use in tissue engineering applications.
UniVert	2020	Multifunctional periodontal membrane for treatment and regeneration purposes	Journal of Bioactive and Compatible Polymers	G. Isik, N. Hasirci, A. Tezcaner, A. Kiziltay	Middle East Technical University		Periodontitis is a chronic inflammatory disease that causes gum tissue degeneration and alveolar bone and tooth loss. The aim of this study is to develop a multifunctional matrix for the treatment of periodontitis and enhancement of regeneration of the periodontal tissue. The matrix was prepared from vitamin E containing hydrogel made of alginate and gelatin, and doxycycline HCl containing methoxy poly(ethylene glycol)-block-polycaprolactone micelles. Methoxy poly(ethylene glycol)-block-polycaprolactone was synthesized with ring-opening polymerization technique and characterized by proton nuclear magnetic resonance (1H NMR), Fourier-transform infrared spectroscopy, differential scanning calorimetry, and gel permeation chromatography. Micelles were characterized by measuring zeta potential, hydrodynamic diameter, drug encapsulation efficiency, drug loading capacity, and in vitro drug-release kinetics. Micelles were obtained with an average size of 164 nm and drug loading amount of 5.8%. The activity of doxycycline HCl–loaded micelles and vitamin E containing hydrogels was determined against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) with disk diffusion method. Bio-efficacy of micelle-loaded alginate–gelatin hydrogels were tested in vitro using L929 fibroblasts and dental pulp stem cells. Doxycycline HCl–loaded micelles and vitamin E containing hydrogels showed a sustained release and exhibited inhibition zone against E. coli and S. aureus. Hydrogels with vitamin E and doxycycline HCl–loaded micelles promoted osteogenic differentiation of dental pulp stem cells. Results suggest that alginate–gelatin hydrogels containing doxycycline HCl–loaded micelles and vitamin E can be good candidates for the treatment of periodontitis and tissue regeneration.
MicroTester/MicroSquisher	2020	Bionic 3D Printed Corals	Nature Communications	D. Wangpraseurt, S. You, F. Azam, G. Jacucci, O. Gaidarenko, M. Hildebrand, M. Kuhl, A. Smith, M. Davey, A. Smith, D. Deheyn, S. Chen, S. Vignolini	University of Cambridge		Corals have evolved as optimized photon augmentation systems, leading to space-efficient microalgal growth and outstanding photosynthetic quantum efficiencies. Light attenuation due to algal self-shading is a key limiting factor for the upscaling of microalgal cultivation. Coral-inspired light management systems could overcome this limitation and facilitate scalable bioenergy and bioproduct generation. Here, we develop 3D printed bionic corals capable of growing microalgae with high spatial cell densities of up to 109 cells mL−1. The hybrid photosynthetic biomaterials are produced with a 3D bioprinting platform which mimics morphological features of living coral tissue and the underlying skeleton with micron resolution, including their optical and mechanical properties. The programmable synthetic microenvironment thus allows for replicating both structural and functional traits of the coral-algal symbiosis. Our work defines a class of bionic materials that is capable of interacting with living organisms and can be exploited for applied coral reef research and photobioreactor design.
MicroTester/MicroSquisher	2020	High throughput direct 3D bioprinting in multiwell plates	Biofabrication	H. Hwang, S. You, X. Ma, L. Kwe, G. Victorine, N. Lawrence, X. Wan, H. Shen, W. Zhu, S. Chen	University of California San Diego		Advances in three dimensional (3D) bioprinting have enabled the fabrication of sophisticated 3D tissue scaffolds for biological and medical applications, where high speed, high throughput production in well plates is a critical need. Here, we present an integrated 3D bioprinting platform based on microscale continuous optical printing, capable of high throughput in situ rapid fabrication of complex 3D biomedical samples in multiwell plate formats for subsequent culture and analysis. Our high throughput 3D bioprinter (HT-3DP) was used to showcase constructs of varying spatial geometries of biomimetic significance, tunable mechanical properties, as well as reproducibility. Live hepatocellular carcinoma 3D tissue scaffolds were fabricated in situ in multiwell plates, after which a functional drug response assay against the chemotherapy drug doxorubicin was performed. Dual cell-type populations involving both live hepatocellular carcinoma as well as human umbilical vein endothelial cells were also printed to demonstrate dual-tissue fabrication capability. This work demonstrates a significant advancement in that the production rate of 3D bioprinted tissue scaffolds with controllable spatial architectures and mechanical properties can now be done on a high throughput scale, enabling rapid generation of in vitro 3D tissue models within conventional multiwell cell culture plates for high throughput preclinical drug screening and disease modeling.