Computationally Optimizing the Compliance of Multilayered Biomimetic Tissue Engineered Vascular Grafts.

Coronary artery bypass grafts used to treat coronary artery disease (CAD) often fail due to compliance mismatch. In this study, we have developed an experimental/computational approach to fabricate an acellular biomimetic hybrid tissue engineered vascular graft (TEVG) composed of alternating layers of electrospun porcine gelatin/polycaprolactone (PCL) and human tropoelastin/PCL blends with the goal of compliance-matching to rat abdominal aorta, while maintaining specific geometrical constraints. Polymeric blends at three different gelatin:PCL (G:PCL) and tropoelastin:PCL (T:PCL) ratios (80:20, 50:50, and 20:80) were mechanically characterized. The stress-strain data were used to develop predictive models, which were used as part of an optimization scheme that was implemented to determine the ratios of G:PCL and T:PCL and the thickness of the individual layers within a TEVG that would compliance match a target compliance value. The hypocompliant, isocompliant, and hypercompliant grafts had target compliance values of 0.000256, 0.000568, and 0.000880 mmHg-1, respectively. Experimental validation of the optimization demonstrated that the hypercompliant and isocompliant grafts were not statistically significant from their respective target compliance values (p-value = 0.37 and 0.89, respectively). The experimental compliance values of the hypocompliant graft were statistically significant than their target compliance value (p-value = 0.047). We have successfully demonstrated a design optimization scheme that can be used to fabricate multilayered and biomimetic vascular grafts with targeted geometry and compliance.

Computationally Optimizing the Compliance of a Biopolymer Based Tissue Engineered Vascular Graft.

Coronary heart disease is a leading cause of death among Americans for which coronary artery bypass graft (CABG) surgery is a standard surgical treatment. The success of CABG surgery is impaired by a compliance mismatch between vascular grafts and native vessels. Tissue engineered vascular grafts (TEVGs) have the potential to be compliance matched and thereby reduce the risk of graft failure. Glutaraldehyde (GLUT) vapor-crosslinked gelatin/fibrinogen constructs were fabricated and mechanically tested in a previous study by our research group at 2, 8, and 24 hrs of GLUT vapor exposure. The current study details a computational method that was developed to predict the material properties of our constructs for crosslinking times between 2 and 24 hrs by interpolating the 2, 8, and 24 hrs crosslinking time data. matlab and abaqus were used to determine the optimal combination of fabrication parameters to produce a compliance matched construct. The validity of the method was tested by creating a 16-hr crosslinked construct of 130 µm thickness and comparing its compliance to that predicted by the optimization algorithm. The predicted compliance of the 16-hr construct was 0.00059 mm Hg-1 while the experimentally determined compliance was 0.00065 mm Hg-1, a relative difference of 9.2%. Prior data in our laboratory has shown the compliance of the left anterior descending porcine coronary (LADC) artery to be 0.00071 ± 0.0003 mm Hg-1. Our optimization algorithm predicts that a 258-µm-thick construct that is GLUT vapor crosslinked for 8.1 hrs would match LADC compliance. This result is consistent with our previous work demonstrating that an 8-hr GLUT vapor crosslinked construct produces a compliance that is not significantly different from a porcine coronary LADC.

2-Photon Characterization of Optical Proteolytic Beacons for Imaging Changes in Matrix-Metalloprotease Activity in a Mouse Model of Aneurysm.

Abdominal aortic aneurysm is a multifactorial disease that is a leading cause of death in developed countries. Matrix-metalloproteases (MMPs) are part of the disease process, however, assessing their role in disease initiation and progression has been difficult and animal models have become essential. Combining Förster resonance energy transfer (FRET) proteolytic beacons activated in the presence of MMPs with 2-photon microscopy allows for a novel method of evaluating MMP activity within the extracellular matrix (ECM). Single and 2-photon spectra for proteolytic beacons were determined in vitro. Ex vivo experiments using the apolipoprotein E knockout angiotensin II-infused mouse model of aneurysm imaged ECM architecture simultaneously with the MMP-activated FRET beacons. 2-photon spectra of the two-color proteolytic beacons showed peaks for the individual fluorophores that enable imaging of MMP activity through proteolytic cleavage. Ex vivo imaging of the beacons within the ECM revealed both microstructure and MMP activity. 2-photon imaging of the beacons in aneurysmal tissue showed an increase in proteolytic cleavage within the ECM (p<0.001), thus indicating an increase in MMP activity. Our data suggest that FRET-based proteolytic beacons show promise in assessing MMP activity within the ECM and will therefore allow future studies to identify the heterogeneous distribution of simultaneous ECM remodeling and protease activity in aneurysmal disease.

Sequential-digital image correlation for mapping human posterior sclera and optic nerve head deformation.

Optic nerve head (ONH) deformations may be involved in the onset or further development of glaucoma, including in patients with relatively normal intraocular pressures (IOPs). Characterizing posterior scleral deformations over physiological pressures may provide a better understanding of how changes in IOP lead to changes in the mechanical environment of the ONH and possibly retinal ganglion cell death. Pressure inflation measurement test protocols are commonly used to measure deformation of the peripapillary sclera with full-field noncontact optical methods. The purpose of this work was to develop and validate a new sequential 3D digital image correlation (S-DIC) approach for quantification of posterior scleral pressure induced deformation that improves z (in-depth) resolution of the DIC measurement without losing in-plane sensitivity, while also being able to contour and map deformations of the complex-shaped ONH. Our approach combines two orthogonal axes of parallax with standard 3D DIC methods using a single high-resolution camera. The enhanced capabilities of S-DIC with respect to standard 3D DIC has been demonstrated by carrying out a complete benchmark for shape, deformation, and strain measurement on an object of known complex geometry. Our S-DIC method provided a reconstruction accuracy of 0.17% and an uncertainty in z-position measurement of 8 µm. The developed methodology has also been applied to a human posterior scleral shell, including the full peripapillary sclera and optic nerve. The relatively inexpensive S-DIC approach may provide new information on the biomechanical deformations of the optic nerve head and, thus, the death of retinal ganglion cells in primary open angle glaucoma.

Differences in the microstructure and biomechanical properties of the recurrent laryngeal nerve as a function of age and location.

Idiopathic onset of unilateral vocal fold paralysis (UVP) is caused by damage to the recurrent laryngeal nerve (RLN) and results in difficulty speaking, breathing, and swallowing. This damage may occur in this nerve as it loops around the aortic arch, which is in a dynamic biomechanical environment. The goal of this study is to determine if the location-dependent biomechanical and microstructural properties of the RLN are different in piglets versus adolescent pigs. The neck/distal and thoracic/proximal (near the aortic arch) regions of the RLN from eight adolescent pigs and six piglets were isolated and mechanically assessed in uni-axial tension. Two-photon imaging (second harmonic) data were collected at 5%, 10%, and 15% strain during the mechanical test. The tangential modulus (TM) and the strain energy density (W) were determined at each level of strain. The mean mode of the preferred fiber angle and the full width at half maximum (FWHM, a measure of fiber splay) were calculated from the imaging data. We found significantly larger values of TM, W, and FWHM in the proximal segments of the left RLN when compared to the distal segments (18.51 MPa ± 1.22 versus 10.78 MPa ± 1.22, p < 0.001 for TM, 0.046 MPa ± 0.01 versus 0.026 MPa ± 0.01, p < 0.003 for W, 15.52 deg ± 1.00 versus 12.98 deg ± 1.00, p < 0.001 for FWHM). TM and W were larger in the left segments than the right (15.32 MPa ± 1.20 versus 11.80 MPa ± 1.20, p < 0.002 for TM, 0.038 MPa ± 0.01 versus 0.028 MPa ± 0.01, p < 0.0001 for W). W was larger in piglets when compared to adolescent pigs (0.042 MPa ± 0.01 versus 0.025 MPa ± 0.01, p < 0.04). The proximal region of the left porcine RLN is more stiff than the distal region and has a higher degree of fiber splay. The left RLN of the adolescent pigs also displayed a higher degree of strain stiffening than the right. These differences may develop as a result of the more dynamic environment the left RLN is in as it loops around the aortic arch.

Location-dependent coronary artery diffusive and convective mass transport properties of a lipophilic drug surrogate measured using nonlinear microscopy.

PURPOSE: Arterial wall mass transport properties dictate local distribution of biomolecules or locally delivered dugs. Knowing how these properties vary between coronary artery locations could provide insight into how therapy efficacy is altered between arterial locations. METHODS: We introduced an indocarbocyanine drug surrogate to the lumens of left anterior descending and right coronary (LADC; RC) arteries from pigs with or without a pressure gradient. Interstitial fluorescent intensity was measured on live samples with multiphoton microscopy. We also measured binding to porcine coronary SMCs in monoculture. RESULTS: Diffusive transport constants peaked in the middle sections of the LADC and RC arteries by 2.09 and 2.04 times, respectively, compared to the proximal and distal segments. There was no statistical difference between the average diffusivity value between LADC and RC arteries. The convection coefficients had an upward trend down each artery, with the RC being higher than the LADC by 3.89 times. CONCLUSIONS: This study demonstrates that the convective and diffusive transport of lipophilic molecules changes between the LADC and the RC arteries as well as along their length. These results may have important implications in optimizing drug delivery for the treatment of coronary artery disease.

A finite element study on variations in mass transport in stented porcine coronary arteries based on location in the coronary arterial tree.

Drug-eluting stents have a significant clinical advantage in late-stage restenosis due to the antiproliferative drug release. Understanding how drug transport occurs between coronary arterial locations can better help guide localized drug treatment options. Finite element models with properties from specific porcine coronary artery sections (left anterior descending (LAD), right (RCA); proximal, middle, distal regions) were created for stent deployment and drug delivery simulations. Stress, strain, pore fluid velocity, and drug concentrations were exported at different time points of simulation (0-180 days). Tests indicated that the highest stresses occurred in LAD sections. Higher-than-resting homeostatic levels of stress and strain existed at upwards of 3.0 mm away from the stented region, whereas concentration of species only reached 2.7 mm away from the stented region. Region-specific concentration showed 2.2 times higher concentrations in RCA artery sections at times corresponding to vascular remodeling (peak in the middle segment) compared to all other segments. These results suggest that wall transport can occur differently based on coronary artery location. Awareness of peak growth stimulators and where drug accumulation occurs in the vasculature can better help guide local drug delivery therapies.

Mechanical alterations of electrospun poly(ϵ-caprolactone) in response to convective thermobonding.

Vascular graft failure has persisted as a major clinical problem. Mechanical, structural, and transport properties of vascular grafts are critical factors that substantially affect their function and thus the outcome of implantation. The manufacturing method, post-processing technique, and material of choice have a significant impact on these properties. The goal of this work is to use thermal treatment to modulate the transport properties of PCL-based vascular engineered constructs. To this end, we electrospun PCL tubular constructs and thermally bonded the electrospun fibers in a convective oven at various temperatures (54, 57, and 60°C) and durations of treatment (15, 30, and 45 s). The effects of fiber thermal bonding (thermobonding) on the transport, mechanical, and structural properties of PCL tubular constructs were characterized. Increasing the temperature and treatment duration enhanced the degree of thermobonding by removing the interconnected void and fusing the fibers. Thermobonding at 57°C and 60°C for longer than 30 s increased the median tangential modulus (E = 126.1 MPa, [IQR = 20.7]), mean suture retention (F = 193.8 g, [SD = 18.5]), and degradation rate while it decreased the median permeability (kA  = 0 m/s), and median thickness (t = 60 µm, [IQR = 2.5]). In particular, the thermobonding at 57°C allowed a finer modulation of permeability via treatment duration. We believe that the thermobonding method can be utilized to modulate the properties of vascular engineered constructs which can be useful in designing functional vascular grafts.

Altered tissue behavior of a non-aneurysmal descending thoracic aorta in the mouse model of Marfan syndrome.

Aortic aneurysm is predominantly found in the ascending aorta in patients with Marfan syndrome (MFS). However, descending aortic disease has emerged as a problem since people are living longer because of improved medical and surgical management of the ascending aorta. Diagnostic procedures before disease onset and the mechanisms involved in the transition of normal aortic tissue to aneurysm remain unclear. We determined signs of descending aortic disease before disease onset in mice with a mutation in the fibrillin 1 gene (Fbn1(+/C1039G)), a validated mouse model of disease susceptibility and progression of aortic aneurysm of MFS. We analyzed a tubular unfixed non-aneurysmal descending thoracic aorta from 8-month-old wild-type and Fbn1(+/C1039G) mice by a tubular biaxial tester that works in conjunction with a two-photon nonlinear microscope. Fbn1(+/C1039G) mouse aorta was more compliant in the circumferential direction. Two-photon imaging showed defective organization of adventitial collagen fibers in the pressurized aortas of Fbn1(+/C1039G) mice. Moreover, disruption in the elastic lamina was noted in the absence of aneurysms in pressurized aortas but not unpressurized aortas of Fbn1(+/C1039G) mice. At the molecular level, this altered tissue behavior in non-aneurysmal descending aortas of Fbn1(+/C1039G) mice was accompanied by an increasing trend of canonical but not noncanonical, transforming growth factor-ß (TGFß) signaling. Finally, assays of in vitro collagen lattice formation in mouse wild-type and TGFß1-deficient embryonic fibroblasts indicate that TGFß1 can regulate collagen organization. The ability to reveal the presence of altered biomechanics and microstructure coupled with subtle changes in TGFß signaling provides a novel surrogate measure of tissue susceptibility to aneurysm before disease onset.

Acute Elution of TGFß2 Affects the Smooth Muscle Cells in a Compliance-Matched Vascular Graft.

Transforming growth factor beta 2 (TGFß2) is a pleiotropic growth factor that plays a vital role in smooth muscle cell (SMC) function. Our prior in vitro work has shown that SMC response can be modulated with TGFß2 stimulation in a dose dependent manner. In particular, we have shown that increasing concentrations of TGFß2 shift SMCs from a migratory to a synthetic behavior. In this work, electrospun compliance-matched and hypocompliant TGFß2-eluting tissue engineered vascular grafts (TEVGs) were implanted into Sprague Dawley rats for 5 days to observe SMC population and collagen production. TEVGs were fabricated using a combined computational and experimental approach that varied the ratio of gelatin:polycaprolactone to be either compliance matched or twice as stiff as rat aorta (hypocompliant). TGFß2 concentrations of 0, 10, 100 ng/mg were added to both graft types (n = 3 in each group) and imaged in vivo using ultrasound. Histological markers (SMC, macrophage, collagen, and elastin) were evaluated following explanation at 5 days. In vivo ultrasound showed that compliance-matched TEVGs became stiffer as TGFß2 increased (100 ng/mg TEVGs compared to rat aorta, p < 0.01), while all hypocompliant grafts remained stiffer than control rat aorta. In vivo velocity and diameter were also not significantly different than control vessels. The compliance-matched 10 ng/mg group had an elevated SMC signal (myosin heavy chain) compared to the 0 and 100 ng/mg grafts (p = 0.0009 and 0.0006). Compliance-matched TEVGs containing 100 ng/mg TGFß2 had an increase in collagen production (p < 0.01), general immune response (p < 0.05), and a decrease in SMC population to the 0 and 10 ng/mg groups. All hypocompliant groups were found to be similar, suggesting a lower rate of TGFß2 release in these TEVGs. Our results suggest that TGFß2 can modulate in vivo SMC phenotype over an acute implantation period, which is consistent with our prior in vitro work. To the author's knowledge, this is the first in vivo rat study that evaluates a TGFß2-eluting TEVG. Impact statement TGFß2 affects the SMCs in a vascular graft.

Microstructural Deformations Within the Depth of the Lamina Cribrosa in Response to Acute In Vivo Intraocular Pressure Modulation.

Purpose: The lamina cribrosa (LC) is a leading target for initial glaucomatous damage. We investigated the in vivo microstructural deformation within the LC volume in response to acute IOP modulation while maintaining fixed intracranial pressure (ICP). Methods: In vivo optic nerve head (ONH) spectral-domain optical coherence tomography (OCT) scans (Leica, Chicago, IL, USA) were obtained from eight eyes of healthy adult rhesus macaques (7 animals; ages = 7.9-14.4 years) in different IOP settings and fixed ICP (8-12 mm Hg). IOP and ICP were controlled by cannulation of the anterior chamber and the lateral ventricle of the brain, respectively, connected to a gravity-controlled reservoir. ONH images were acquired at baseline IOP, 30 mm Hg (H1-IOP), and 40 to 50 mm Hg (H2-IOP). Scans were registered in 3D, and LC microstructure measurements were obtained from shared regions and depths. Results: Only half of the eyes exhibited LC beam-to-pore ratio (BPR) and microstructure deformations. The maximal BPR change location within the LC volume varied between eyes. BPR deformer eyes had a significantly higher baseline connective tissue volume fraction (CTVF) and lower pore aspect ratio (P = 0.03 and P = 0.04, respectively) compared to BPR non-deformer. In all eyes, the magnitude of BPR changes in the anterior surface was significantly different (either larger or smaller) from the maximal change within the LC (H1-IOP: P = 0.02 and H2-IOP: P = 0.004). Conclusions: The LC deforms unevenly throughout its depth in response to IOP modulation at fixed ICP. Therefore, analysis of merely the anterior LC surface microstructure will not fully capture the microstructure deformations within the LC. BPR deformer eyes have higher CTVF than BPR non-deformer eyes.

Extracellular matrix deformations of the porcine recurrent laryngeal nerve in response to hydrostatic pressure.

Damage to the recurrent laryngeal nerve (RLN) caused by supraphysiological compression or tension imposed by adjacent tissue structures, such as the aorta, may contribute to onset of idiopathic unilateral vocal fold paralysis (iUVP) resulting in difficulty speaking, breathing, and swallowing. We previously demonstrated in adolescent pigs that the right RLN epineurium exhibits uniform composition of adipose tissue, with larger quantities along its length within the neck region in contrast to the left RLN that shows greater collagen composition in the thoracic region and greater quantities of adipose tissue in the neck region. In contrast, the epineurium in piglets was primarily composed of collagen tissue that remained uniform along the length of the left and right RLNs. Tensile testing of the left and right RLN in piglets and pigs showed associated differences in strain by RLN side and segment by age. The goal of this study was to investigate how external hydrostatic compression of the RLN affects the nerve's connective tissue and microstructure. RLN segments were harvested from the distal (cervical/neck) regions and proximal (subclavian for the right RLN, thoracic for the left RLN) regions from eight adolescent pigs and nine piglets. RLN segments were isolated and assessed under fluid compression to test hypotheses regarding epineurium composition and response to applied forces. Second harmonic generation (SHG) imaging of epineurial collagen was conducted at 0, 40, and 80 mmHg of compression. The cartesian strain tensor, principal strain (Eps1), and principal direction of the RLN collagen fibers were determined at each pressure step. Significantly larger values of the 1st principal strain occurred in the proximal segments of the pig left RLN when compared to the same segment in piglets (p = 0.001, pig = 0.0287 [IQR = 0.0161 - 0.0428], piglet = 0.0061 [IQR = 0.0033 - 0.0156]). Additionally, the median transverse strain Eyy) for the second pressure increment was larger in the right proximal segment of pigs compared to piglets (p < 0.001, pig = 0.0122 [IQR = 0.0033 - 0.0171], piglet = 0.0013 [IQR = 0.00001 - 0.0028]). Eyy values were significantly larger in the right proximal RLN versus the left proximal RLNs in pigs but not in piglets (p < 0.001). In contrast to piglets, histological analysis of pig RLN demonstrated increased axial alignment of epineurial and endoneurial collagen in response to compressive pressure. These findings support the hypothesis that the biomechanical response of the RLN to compressive pressure changed from being similar to being different between the right and left RLNs during development in the porcine model. Further investigation of these findings associated with age-related onset of idiopathic UVP may illuminate underlying etiologic mechanisms. STATEMENT OF SIGNIFICANCE: Damage to the recurrent laryngeal nerve (RLN) caused by compression imposed by the aorta may contribute to the onset of left-sided idiopathic unilateral vocal fold paralysis resulting in difficulty speaking, breathing, and swallowing. The goal of this study was to investigate how compression affects the connective tissue and microstructure of the RLN. We quantified the pressure induced deformation of the RLN using multiphoton imaging as a function of both location (proximal versus distal) and age (piglets, adolescent pigs). Our results demonstrate that the biomechanical response of the RLN to compression changes in the right versus left RLN throughout development, providing further evidence that the the left RLN is exposed to increasing dynamic loads with age.

Adaptation of a planar microbiaxial optomechanical device for the tubular biaxial microstructural and macroscopic characterization of small vascular tissues.

Murine models of disease are a powerful tool for researchers to gain insight into disease formation, progression, and therapies. The biomechanical indicators of diseased tissue provide a unique insight into some of these murine models, since the biomechanical properties in scenarios such as aneurysm and Marfan syndrome can dictate tissue failure and mortality. Understanding the properties of the tissue on the macroscopic scale has been shown to be important, as one can then understand the tissue's ability to withstand the high stresses seen in the cardiac pulsatile cycle. Alterations in the biomechanical response can foreshadow prospective mechanical failure of the tissue. These alterations are often seen on the microstructural level, and obtaining detailed information on such changes can offer a better understanding of the phenomena seen on the macroscopic level. Unfortunately, mouse models present problems due to the size and delicate features in the mechanical testing of such tissues. In addition, some smaller arteries in large-animal studies (e.g., coronary and cerebral arteries) can present the same issues, and are sometimes unsuitable for planar biaxial testing. The purpose of this paper is to present a robust method for the investigation of the mechanical properties of small arteries and the classification of the microstructural orientation and degree of fiber alignment. This occurs through the cost-efficient modification of a planar biaxial tester that works in conjunction with a two-photon nonlinear microscope. This system provides a means to further investigate how microstructure and mechanical properties are modified in diseased transgenic animals where the tissue is in small tube form. Several other hard-to-test tubular specimens such as cerebral aneurysm arteries and atherosclerotic coronary arteries can also be tested using the described modular device.

A gimbal-mounted pressurization chamber for macroscopic and microscopic assessment of ocular tissues.

The biomechanical model of glaucoma considers intraocular pressure-related stress and resultant strain on load bearing connective tissues of the optic nerve and surrounding peripapillary sclera as one major causative influence that effects cellular, vascular, and axonal components of the optic nerve. By this reasoning, the quantification of variations in the microstructural architecture and macromechanical response of scleral shells in glaucomatous compared to healthy populations provides an insight into any variations that exist between patient populations. While scleral shells have been tested mechanically in planar and pressure-inflation scenarios the link between the macroscopic biomechanical response and the underlying microstructure has not been determined to date. A potential roadblock to determining how the microstructure changes based on pressure is the ability to mount the spherical scleral shells in a method that does not induce unwanted stresses to the samples (for instance, in the flattening of the spherical specimens), and then capturing macroscopic and microscopic changes under pressure. Often what is done is a macroscopic test followed by sample fixation and then imaging to determine microstructural organization. We introduce a novel device and method, which allows spherical samples to be pressurized and macroscopic and microstructural behavior quantified on fully hydrated ocular specimens. The samples are pressurized and a series of markers on the surface of the sclera imaged from several different perspectives and reconstructed between pressure points to allow for mapping of nonhomogenous strain. Pictures are taken from different perspectives through the use of mounting the pressurization scheme in a gimbal that allows for positioning the sample in several different spherical coordinate system configurations. This ability to move the sclera in space about the center of the globe, coupled with an upright multiphoton microscope, allows for collecting collagen, and elastin signal in a rapid automated fashion so the entire globe can be imaged.

Coupled porohyperelastic mass transport (PHEXPT) finite element models for soft tissues using ABAQUS.

Finite element models (FEMs) including characteristic large deformations in highly nonlinear materials (hyperelasticity and coupled diffusive/convective transport of neutral mobile species) will allow quantitative study of in vivo tissues. Such FEMs will provide basic understanding of normal and pathological tissue responses and lead to optimization of local drug delivery strategies. We present a coupled porohyperelastic mass transport (PHEXPT) finite element approach developed using a commercially available ABAQUS finite element software. The PHEXPT transient simulations are based on sequential solution of the porohyperelastic (PHE) and mass transport (XPT) problems where an Eulerian PHE FEM is coupled to a Lagrangian XPT FEM using a custom-written FORTRAN program. The PHEXPT theoretical background is derived in the context of porous media transport theory and extended to ABAQUS finite element formulations. The essential assumptions needed in order to use ABAQUS are clearly identified in the derivation. Representative benchmark finite element simulations are provided along with analytical solutions (when appropriate). These simulations demonstrate the differences in transient and steady state responses including finite deformations, total stress, fluid pressure, relative fluid, and mobile species flux. A detailed description of important model considerations (e.g., material property functions and jump discontinuities at material interfaces) is also presented in the context of finite deformations. The ABAQUS-based PHEXPT approach enables the use of the available ABAQUS capabilities (interactive FEM mesh generation, finite element libraries, nonlinear material laws, pre- and postprocessing, etc.). PHEXPT FEMs can be used to simulate the transport of a relatively large neutral species (negligible osmotic fluid flux) in highly deformable hydrated soft tissues and tissue-engineered materials.

Design and demonstration of a microbiaxial optomechanical device for multiscale characterization of soft biological tissues with two-photon microscopy.

The biomechanical response of tissues serves as a valuable marker in the prediction of disease and in understanding the related behavior of the body under various disease and age states. Alterations in the macroscopic biomechanical response of diseased tissues are well documented; however, a thorough understanding of the microstructural events that lead to these changes is poorly understood. In this article we introduce a novel microbiaxial optomechanical device that allows two-photon imaging techniques to be coupled with macromechanical stimulation in hydrated planar tissue specimens. This allows that the mechanical response of the microstructure can be quantified and related to the macroscopic response of the same tissue sample. This occurs without the need to fix tissue in strain states that could introduce a change in the microstructural configuration. We demonstrate the passive realignment of fibrous proteins under various types of loading, which demonstrates the ability of tissue microstructure to reinforce itself in periods of high stress. In addition, the collagen and elastin response of tissue during viscoelastic behavior is reported showing interstitial fluid movement and fiber realignment potentially responsible for the temporal behavior. We also demonstrate that nonhomogeneities in fiber strain exist over biaxial regions of assumed homogeneity.

In-vivo assessment of a tissue engineered vascular graft computationally optimized for target vessel compliance.

Tissue engineered vascular grafts (TEVGs) have the ability to be tuned to match a target vessel's compliance, diameter, wall thickness, and thereby prevent compliance mismatch. In this work, TEVG compliance was manipulated by computationally tuning its layered composition or by manipulating a crosslinking agent (genipin). In particular, these three acelluluar TEVGs were compared: a compliance matched graft (CMgel - high gelatin content); a hypocompliant PCL graft (HYPOpcl - high polycaprolactone content); and a hypocompliant genipin graft (HYPOgen - equivalent composition as CMgel but hypocompliant via increased genipin crosslinking). All constructs were implanted interpositionally into the abdominal aorta of 21 Sprague Dawley rats (n=7, males=11, females=10) for 28 days, imaged in-vivo using ultrasound, explanted, and assessed for remodeling using immunofluorescence and two photon excitation fluorescence imaging. Compliance matched grafts remained compliance-matched in-vivo compared to the hypocompliant grafts through 4 weeks (p<0.05). Construct degradation and cellular infiltration was increased in the CMgel and HYPOgen TEVGs. Contractile smooth muscle cell markers in the proximal anastomosis of the graft were increased in the CMgel group compared to the HYPOpcl (p=0.007) and HYPOgen grafts (p=0.04). Both hypocompliant grafts also had an increased pro-inflammatory response (increased ratio of CD163 to CD86 in the mid-axial location) compared to the CMgel group. Our results suggest that compliance matching using a computational optimization approach leads to the improved acute (28 day) remodeling of TEVGs. To the authors' knowledge, this is the first in-vivo rat study investigating TEVGs that have been computationally optimized for target vessel compliance.

Porohyperelastic finite element modeling of abdominal aortic aneurysms.

Abdominal aortic aneurysm (AAA) is the gradual weakening and dilation of the infrarenal aorta. This disease is progressive, asymptomatic, and can eventually lead to rupture--a catastrophic event leading to massive internal bleeding and possibly death. The mechanical environment present in AAA is currently thought to be important in disease initiation, progression, and diagnosis. In this study, we utilize porohyperelastic (PHE) finite element models (FEMs) to investigate how such modeling can be used to better understand the local biomechanical environment in AAA. A 3D hypothetical AAA was constructed with a preferential anterior bulge assuming both the intraluminal thrombus (ILT) and the AAA wall act as porous materials. A parametric study was performed to investigate how physiologically meaningful variations in AAA wall and ILT hydraulic permeabilities affect luminal interstitial fluid velocities and wall stresses within an AAA. A corresponding hyperelastic (HE) simulation was also run in order to be able to compare stress values between PHE and HE simulations. The effect of AAA size on local interstitial fluid velocity was also investigated by simulating maximum diameters (5.5 cm, 4.5 cm, and 3.5 cm) at the baseline values of ILT and AAA wall permeability. Finally, a cyclic PHE simulation was utilized to study the variation in local fluid velocities as a result of a physiologic pulsatile blood pressure. While the ILT hydraulic permeability was found to have minimal affect on interstitial velocities, our simulations demonstrated a 28% increase and a 20% decrease in luminal interstitial fluid velocity as a result of a 1 standard deviation increase and decrease in AAA wall hydraulic permeability, respectively. Peak interstitial velocities in all simulations occurred on the luminal surface adjacent to the region of maximum diameter. These values increased with increasing AAA size. PHE simulations resulted in 19.4%, 40.1%, and 81.0% increases in peak maximum principal wall stresses in comparison to HE simulations for maximum diameters of 35 mm, 45 mm, and 55 mm, respectively. The pulsatile AAA PHE FEM demonstrated a complex interstitial fluid velocity field the direction of which alternated in to and out of the luminal layer of the ILT. The biomechanical environment within both the aneurysmal wall and the ILT is involved in AAA pathogenesis and rupture. Assuming these tissues to be porohyperelastic materials may provide additional insight into the complex solid and fluid forces acting on the cells responsible for aneurysmal remodeling and weakening.

Determination of coefficient of friction for self-expanding stent-grafts.

Migration of stent-grafts (SGs) after endovascular aneurysm repair of abdominal aortic aneurysms is a serious complication that may require secondary intervention. Experimental, analytical, and computational studies have been carried out in the past to understand the factors responsible for migration. In an experimental setting, it can be very challenging to correctly capture and understand the interaction between a SG and an artery. Quantities such as coefficient of friction (COF) and contact pressures that characterize this interaction are difficult to measure using an experimental approach. This behavior can be investigated with good accuracy using finite element modeling. Although finite element models are able to incorporate frictional behavior of SGs, the absence of reliable values of coefficient of friction make these simulations unreliable. The aim of this paper is to demonstrate a method for determining the coefficients of friction of a self-expanding endovascular stent-graft. The methodology is demonstrated by considering three commercially available self-expanding SGs, labeled as A, B, and C. The SGs were compressed, expanded, and pulled out of polymeric cylinders of varying diameters and the pullout force was recorded in each case. The SG geometries were recreated using computer-aided design modeling and the entire experiment was simulated in ABAQUS 6.8/STANDARD. An optimization procedure was carried out for each SG oversize configuration to determine the COF that generated a frictional force corresponding to that measured in the experiment. The experimental pullout force and analytically determined COF for SGs A, B, and C were in the range of 6-9 N, 3-12 N, and 3-9 N and 0.08-0.16, 0.22-0.46, and 0.012-0.018, respectively. The computational model predicted COFs in the range of 0.00025-0.0055, 0.025-0.07, and 0.00025-0.006 for SGs A, B, and C, respectively. Our results suggest that for SGs A and B, which are exoskeleton based devices, the pullout forces increase upto a particular oversize beyond which they plateau, while pullout forces showed a continuous increase with oversize for SG C, which is an endoskeleton based device. The COF decreased with oversizing for both types of SGs. The proposed methodology will be useful for determining the COF between self-expanding stent-grafts from pullout tests on human arterial tissue.

Decellularizing the Porcine Optic Nerve Head: Toward a Model to Study the Mechanobiology of Glaucoma.

Purpose: Studying the extracellular matrix (ECM) remodeling of the lamina cribrosa in vivo can be extremely challenging and costly. There exist very few options for studying optic nerve head (ONH) mechanobiology in vitro that are able to reproduce the complex anatomic and biomechanical environment of the ONH. Herein, we have developed a decellularization procedure that will enable more anatomically relevant and cost-efficient future studies of ECM remodeling of the ONH. Methods: Porcine posterior poles were decellularized using a detergent and enzyme-based decellularization protocol. DNA quantification and histology were used to investigate the effectiveness of the protocol. We subsequently investigated the ability of a polyethylene glycol (PEG)-based hydrogel to restore the ONH's ability to hold pressure following decellularization. Anterior-posterior displacement of the decellularized and PEG treated ONH in a pressure bioreactor was used to evaluate the biomechanical response of the ONH. Results: DNA quantification and histology confirmed decellularization using Triton X-100 at low concentration for 48 hours successfully reduced the cellular content of the tissue by 94.9% compared with native tissue while preserving the ECM microstructure and basal lamina of the matrix. Infiltrating the decellularized tissues with PEG 6000 and PEG 10,000 hydrogel restored their ability to hold pressure, producing displacements similar to those measured for the non-decellularized control samples. Conclusions: Our decellularized ONH model is capable of producing scaffolds that are cell-free and maintain the native ECM microstructure. Translational Relevance: This model represents a platform to study the mechanobiology in the ONH and potentially for glaucoma drug testing.