Large strain stimulation promotes extracellular matrix production and stiffness in an elastomeric scaffold model.

Mechanical conditioning of engineered tissue constructs is widely recognized as one of the most relevant methods to enhance tissue accretion and microstructure, leading to improved mechanical behaviors. The understanding of the underlying mechanisms remains rather limited, restricting the development of in silico models of these phenomena, and the translation of engineered tissues into clinical application. In the present study, we examined the role of large strip-biaxial strains (up to 50%) on ECM synthesis by vascular smooth muscle cells (VSMCs) micro-integrated into electrospun polyester urethane urea (PEUU) constructs over the course of 3 weeks. Experimental results indicated that VSMC biosynthetic behavior was quite sensitive to tissue strain maximum level, and that collagen was the primary ECM component synthesized. Moreover, we found that while a 30% peak strain level achieved maximum ECM synthesis rate, further increases in strain level lead to a reduction in ECM biosynthesis. Subsequent mechanical analysis of the formed collagen fiber network was performed by removing the scaffold mechanical responses using a strain-energy based approach, showing that the denovo collagen also demonstrated mechanical behaviors substantially better than previously obtained with small strain training and comparable to mature collagenous tissues. We conclude that the application of large deformations can play a critical role not only in the quantity of ECM synthesis (i.e. the rate of mass production), but also on the modulation of the stiffness of the newly formed ECM constituents. The improved understanding of the process of growth and development of ECM in these mechano-sensitive cell-scaffold systems will lead to more rational design and manufacturing of engineered tissues operating under highly demanding mechanical environments.

[1]  Kyriacos A. Athanasiou,et al.  Tension-Compression Loading with Chemical Stimulation Results in Additive Increases to Functional Properties of Anatomic Meniscal Constructs , 2011, PloS one.

[2]  M. Sacks,et al.  A meso-scale layer-specific structural constitutive model of the mitral heart valve leaflets. , 2016, Acta biomaterialia.

[3]  William R Wagner,et al.  Fabrication of biodegradable elastomeric scaffolds with sub-micron morphologies. , 2004, Journal of biomedical materials research. Part A.

[4]  Ankush Aggarwal,et al.  An inverse modeling approach for semilunar heart valve leaflet mechanics: exploitation of tissue structure , 2016, Biomechanics and modeling in mechanobiology.

[5]  K. Liao,et al.  Bovine pericardium versus porcine aortic valve: comparison of tissue biological properties as prosthetic valves. , 2008, Artificial organs.

[6]  R. Panettieri,et al.  Cyclic mechanical strain‐induced proliferation and migration of human airway smooth muscle cells: role of EMMPRIN and MMPs , 2005, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[7]  Jianjun Guan,et al.  Microintegrating smooth muscle cells into a biodegradable, elastomeric fiber matrix. , 2006, Biomaterials.

[8]  Marcel C M Rutten,et al.  Autologous Human Tissue-Engineered Heart Valves: Prospects for Systemic Application , 2006, Circulation.

[9]  Stephen F Badylak,et al.  The extracellular matrix as a biologic scaffold material. , 2007, Biomaterials.

[10]  R. Chernecky,et al.  Biaxial mechanical/structural effects of equibiaxial strain during crosslinking of bovine pericardial xenograft materials. , 1999, Biomaterials.

[11]  Michael S Sacks,et al.  Synergistic effects of cyclic tension and transforming growth factor-beta1 on the aortic valve myofibroblast. , 2007, Cardiovascular pathology : the official journal of the Society for Cardiovascular Pathology.

[12]  F J Schoen,et al.  Functional Living Trileaflet Heart Valves Grown In Vitro , 2000, Circulation.

[13]  V. Barocas,et al.  Multiscale, structure-based modeling for the elastic mechanical behavior of arterial walls. , 2007, Journal of biomechanical engineering.

[14]  Victor H Barocas,et al.  Affine versus non-affine fibril kinematics in collagen networks: theoretical studies of network behavior. , 2006, Journal of biomechanical engineering.

[15]  Michael S Sacks,et al.  Insights into regional adaptations in the growing pulmonary artery using a meso-scale structural model: effects of ascending aorta impingement. , 2014, Journal of biomechanical engineering.

[16]  Michael S Sacks,et al.  Design and analysis of tissue engineering scaffolds that mimic soft tissue mechanical anisotropy. , 2006, Biomaterials.

[17]  Victor H Barocas,et al.  Image-based multiscale modeling predicts tissue-level and network-level fiber reorganization in stretched cell-compacted collagen gels , 2009, Proceedings of the National Academy of Sciences.

[18]  D. Vorp,et al.  Development of a tissue-engineered vascular graft combining a biodegradable scaffold, muscle-derived stem cells and a rotational vacuum seeding technique. , 2008, Biomaterials.

[19]  A. Grodzinsky,et al.  Fluorometric assay of DNA in cartilage explants using Hoechst 33258. , 1988, Analytical biochemistry.

[20]  M. Sacks,et al.  Biaxial mechanical properties of the natural and glutaraldehyde treated aortic valve cusp--Part I: Experimental results. , 2000, Journal of biomechanical engineering.

[21]  V. Mow,et al.  Chondrocyte deformation and local tissue strain in articular cartilage: A confocal microscopy study , 1995, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[22]  Ross A. Marklein,et al.  The potential to improve cell infiltration in composite fiber-aligned electrospun scaffolds by the selective removal of sacrificial fibers. , 2008, Biomaterials.

[23]  D. Mooney,et al.  Combining chondrocytes and smooth muscle cells to engineer hybrid soft tissue constructs. , 2000, Tissue engineering.

[24]  Victor H. Barocas,et al.  Volume-averaging theory for the study of the mechanics of collagen networks , 2007 .

[25]  Michael S Sacks,et al.  Prediction of extracellular matrix stiffness in engineered heart valve tissues based on nonwoven scaffolds , 2008, Biomechanics and modeling in mechanobiology.

[26]  John M. Gosline,et al.  Elastin as a random‐network elastomer: A mechanical and optical analysis of single elastin fibers , 1981 .

[27]  Frank P T Baaijens,et al.  Intermittent straining accelerates the development of tissue properties in engineered heart valve tissue. , 2009, Tissue engineering. Part A.

[28]  Jennifer L. West,et al.  Physiologic Pulsatile Flow Bioreactor Conditioning of Poly(ethylene glycol)-based Tissue Engineered Vascular Grafts , 2007, Annals of Biomedical Engineering.

[29]  Brendon M. Baker,et al.  Dynamic culture enhances stem cell infiltration and modulates extracellular matrix production on aligned electrospun nanofibrous scaffolds. , 2011, Acta biomaterialia.

[30]  M. Sacks,et al.  A triphasic constrained mixture model of engineered tissue formation under in vitro dynamic mechanical conditioning , 2015, Biomechanics and Modeling in Mechanobiology.

[31]  S. Badylak,et al.  Abdominal wall reconstruction by a regionally distinct biocomposite of extracellular matrix digest and a biodegradable elastomer , 2016, Journal of tissue engineering and regenerative medicine.

[32]  Michael S Sacks,et al.  Microstructural manipulation of electrospun scaffolds for specific bending stiffness for heart valve tissue engineering. , 2012, Acta biomaterialia.

[33]  Victor H Barocas,et al.  Functional tissue-engineered valves from cell-remodeled fibrin with commissural alignment of cell-produced collagen. , 2008, Tissue engineering. Part A.

[34]  Joseph H. Gorman,et al.  Patient-Specific Modeling of Heart Valves: From Image to Simulation , 2013, FIMH.

[35]  H. Vogel Influence of maturation and aging on mechanical and biochemical properties of connective tissue in rats , 1980, Mechanisms of Ageing and Development.

[36]  M. Sacks,et al.  Biaxial mechanical properties of the native and glutaraldehyde-treated aortic valve cusp: Part II--A structural constitutive model. , 2000, Journal of biomechanical engineering.

[37]  Ankush Aggarwal,et al.  A Framework for Determination of Heart Valves' Mechanical Properties Using Inverse-Modeling Approach , 2015, FIMH.

[38]  Michael S Sacks,et al.  The independent role of cyclic flexure in the early in vitro development of an engineered heart valve tissue. , 2005, Biomaterials.

[39]  Marcel C. M. Rutten,et al.  Tissue Engineering of Human Heart Valve Leaflets: A Novel Bioreactor for a Strain-Based Conditioning Approach , 2005, Annals of Biomedical Engineering.

[40]  Christopher A. Carruthers,et al.  Quantification and simulation of layer-specific mitral valve interstitial cells deformation under physiological loading. , 2015, Journal of theoretical biology.

[41]  D. Elliott,et al.  Homologous structure-function relationships between native fibrocartilage and tissue engineered from MSC-seeded nanofibrous scaffolds. , 2011, Biomaterials.

[42]  Marcel C M Rutten,et al.  Deformation-controlled load application in heart valve tissue engineering. , 2009, Tissue engineering. Part C, Methods.

[43]  V. Barocas,et al.  Multiscale mechanical simulations of cell compacted collagen gels. , 2013, Journal of biomechanical engineering.

[44]  Frederick J. Schoen,et al.  Early In Vivo Experience With Tissue-Engineered Trileaflet Heart Valves , 2000, Circulation.

[45]  Michael S Sacks,et al.  From single fiber to macro-level mechanics: A structural finite-element model for elastomeric fibrous biomaterials. , 2014, Journal of the mechanical behavior of biomedical materials.

[46]  Michael S Sacks,et al.  Bioengineering challenges for heart valve tissue engineering. , 2009, Annual review of biomedical engineering.

[47]  David P. Martin,et al.  Fabrication of a trileaflet heart valve scaffold from a polyhydroxyalkanoate biopolyester for use in tissue engineering. , 2000, Tissue engineering.

[48]  F P T Baaijens,et al.  The relevance of large strains in functional tissue engineering of heart valves. , 2003, The Thoracic and cardiovascular surgeon.

[49]  V. Barocas,et al.  Comparison of 2D fiber network orientation measurement methods. , 2009, Journal of biomedical materials research. Part A.

[50]  Cwj Cees Oomens,et al.  Multi-scale mechanical characterization of scaffolds for heart valve tissue engineering. , 2012, Journal of biomechanics.

[51]  Chung-Hao Lee,et al.  A generalized method for the analysis of planar biaxial mechanical data using tethered testing configurations. , 2015, Journal of biomechanical engineering.

[52]  Michael S Sacks,et al.  Optimal elastomeric scaffold leaflet shape for pulmonary heart valve leaflet replacement. , 2013, Journal of biomechanics.

[53]  Y. Lanir Mechanistic micro-structural theory of soft tissues growth and remodeling: tissues with unidirectional fibers , 2015, Biomechanics and modeling in mechanobiology.

[54]  P. Cahill,et al.  Cyclic strain-mediated regulation of endothelial matrix metalloproteinase-2 expression and activity. , 2004, Cardiovascular research.

[55]  Kevin E. Healy,et al.  Engineering gene expression and protein synthesis by modulation of nuclear shape , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[56]  Cees W J Oomens,et al.  Predicting local cell deformations in engineered tissue constructs: a multilevel finite element approach. , 2002, Journal of biomechanical engineering.

[57]  Robert T. Tranquillo,et al.  Controlled cyclic stretch bioreactor for tissue-engineered heart valves. , 2009, Biomaterials.

[58]  Michael S Sacks,et al.  An elastic, biodegradable cardiac patch induces contractile smooth muscle and improves cardiac remodeling and function in subacute myocardial infarction. , 2007, Journal of the American College of Cardiology.

[59]  F. Baaijens,et al.  Matrix Production and Organization by Endothelial Colony Forming Cells in Mechanically Strained Engineered Tissue Constructs , 2013, PloS one.

[60]  Michael S. Sacks,et al.  Orthotropic Mechanical Properties of Chemically Treated Bovine Pericardium , 1998, Annals of Biomedical Engineering.

[61]  M. Sacks,et al.  Biodegradable poly(ether ester urethane)urea elastomers based on poly(ether ester) triblock copolymers and putrescine: synthesis, characterization and cytocompatibility. , 2004, Biomaterials.

[62]  Robert M. Nerem,et al.  Dynamic Mechanical Conditioning of Collagen-Gel Blood Vessel Constructs Induces Remodeling In Vitro , 2000, Annals of Biomedical Engineering.

[63]  R. J. Pawluk,et al.  ACCELERATION OF FRACTURE REPAIR BY ELECTROMAGNETIC FIELDS. A SURGICALLY NONINVASIVE METHOD , 1974, Annals of the New York Academy of Sciences.

[64]  Michael S Sacks,et al.  Tissue-to-cellular level deformation coupling in cell micro-integrated elastomeric scaffolds. , 2008, Biomaterials.

[65]  S. Zucker,et al.  Mechanical stretch induces MMP-2 release and activation in lung endothelium: role of EMMPRIN. , 2003, American journal of physiology. Lung cellular and molecular physiology.

[66]  Boris Martinac,et al.  The ion channels to cytoskeleton connection as potential mechanism of mechanosensitivity. , 2014, Biochimica et biophysica acta.

[67]  Frank P. T. Baaijens,et al.  Strain-induced Collagen Organization at the Micro-level in Fibrin-based Engineered Tissue Constructs , 2012, Annals of Biomedical Engineering.

[68]  Michael S Sacks,et al.  Scale-dependent fiber kinematics of elastomeric electrospun scaffolds for soft tissue engineering. , 2009, Journal of biomedical materials research. Part A.

[69]  Michael S Sacks,et al.  Elastomeric Electrospun Polyurethane Scaffolds: The Interrelationship Between Fabrication Conditions, Fiber Topology, and Mechanical Properties , 2011, Advanced materials.

[70]  David E. Schmidt,et al.  On the biomechanics of heart valve function. , 2009, Journal of biomechanics.

[71]  Robert T Tranquillo,et al.  Cyclic distension of fibrin-based tissue constructs: Evidence of adaptation during growth of engineered connective tissue , 2008, Proceedings of the National Academy of Sciences.

[72]  L. R. Duarte The stimulation of bone growth by ultrasound , 2004, Archives of orthopaedic and traumatic surgery.

[73]  Michael S Sacks,et al.  Geometric characterization and simulation of planar layered elastomeric fibrous biomaterials. , 2015, Acta biomaterialia.

[74]  Young-Mi Kang,et al.  Nanofiber alignment and direction of mechanical strain affect the ECM production of human ACL fibroblast. , 2005, Biomaterials.

[75]  M. Sacks,et al.  Simulation of planar soft tissues using a structural constitutive model: Finite element implementation and validation. , 2014, Journal of biomechanics.

[76]  E. A. Trowbridge,et al.  The tensile strength of natural and chemically modified bovine pericardium. , 1988 .

[77]  Michael S Sacks,et al.  Cyclic flexure and laminar flow synergistically accelerate mesenchymal stem cell-mediated engineered tissue formation: Implications for engineered heart valve tissues. , 2006, Biomaterials.

[78]  D. Elliott,et al.  Modeling interlamellar interactions in angle-ply biologic laminates for annulus fibrosus tissue engineering , 2011, Biomechanics and modeling in mechanobiology.

[79]  Laurent Bozec,et al.  Mechanical properties of collagen fibrils. , 2007, Biophysical journal.

[80]  David J. Mooney,et al.  Cyclic mechanical strain regulates the development of engineered smooth muscle tissue , 1999, Nature Biotechnology.