Microscale Strategies for Generating Cell-Encapsulating Hydrogels.

Hydrogels in which cells are encapsulated are of great potential interest for tissue engineering applications. These gels provide a structure inside which cells can spread and proliferate. Such structures benefit from controlled microarchitectures that can affect the behavior of the enclosed cells. Microfabrication-based techniques are emerging as powerful approaches to generate such cell-encapsulating hydrogel structures. In this paper we introduce common hydrogels and their crosslinking methods and review the latest microscale approaches for generation of cell containing gel particles. We specifically focus on microfluidics-based methods and on techniques such as micromolding and electrospinning.

[1]  R. Kwapiszewski,et al.  Microfluidic devices as tools for mimicking the in vivo environment , 2011 .

[2]  E. Delamarche,et al.  Patterned delivery of immunoglobulins to surfaces using microfluidic networks. , 1997, Science.

[3]  Jennifer L. West,et al.  Three-dimensional micropatterning of bioactive hydrogels via two-photon laser scanning photolithography for guided 3D cell migration. , 2008, Biomaterials.

[4]  J. Hubbell,et al.  Characterization of permeability and network structure of interfacially photopolymerized poly(ethylene glycol) diacrylate hydrogels. , 1998, Biomaterials.

[5]  A. Khademhosseini,et al.  Modular Tissue Engineering: Engineering Biological Tissues from the Bottom Up. , 2009, Soft matter.

[6]  J. Cooper,et al.  Tumors on chips: oncology meets microfluidics. , 2010, Current opinion in chemical biology.

[7]  Xiaohong Wang,et al.  Recent trends and challenges in complex organ manufacturing. , 2010, Tissue engineering. Part B, Reviews.

[8]  Daniel G. Anderson,et al.  Nanoliter-scale synthesis of arrayed biomaterials and application to human embryonic stem cells , 2004, Nature Biotechnology.

[9]  D. Weitz,et al.  Janus microgels produced from functional precursor polymers. , 2010, Langmuir : the ACS journal of surfaces and colloids.

[10]  Ullrich Scherf,et al.  Micromolding in capillaries and microtransfer printing of silver nanoparticles as soft-lithographic approach for the fabrication of source/drain electrodes in organic field-effect transistors , 2007 .

[11]  Tatiana Kniazeva,et al.  A microfluidic respiratory assist device with high gas permeance for artificial lung applications , 2011, Biomedical microdevices.

[12]  A. Khademhosseini,et al.  Microscale technologies for tissue engineering and biology. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[13]  Sarit B. Bhaduri,et al.  Drop-on-demand printing of cells and materials for designer tissue constructs , 2007 .

[14]  Jennifer L West,et al.  Photocrosslinkable polyvinyl alcohol hydrogels that can be modified with cell adhesion peptides for use in tissue engineering. , 2002, Biomaterials.

[15]  Robert Langer,et al.  Engineering systems for the generation of patterned co-cultures for controlling cell-cell interactions. , 2011, Biochimica et biophysica acta.

[16]  Megan L. McCain,et al.  Ensembles of engineered cardiac tissues for physiological and pharmacological study: heart on a chip. , 2011, Lab on a chip.

[17]  Ying Zhang,et al.  Replica molding of high-aspect-ratio polymeric nanopillar arrays with high fidelity. , 2006, Langmuir : the ACS journal of surfaces and colloids.

[18]  L. Niklason,et al.  Scaffold-free vascular tissue engineering using bioprinting. , 2009, Biomaterials.

[19]  A. Abate,et al.  Microfluidic Assembly of Magnetic Hydrogel Particles with Uniformly Anisotropic Structure , 2009 .

[20]  Chaenyung Cha,et al.  Biodegradable Polymer Crosslinker: Independent Control of Stiffness, Toughness, and Hydrogel Degradation Rate , 2009 .

[21]  D. Mooney,et al.  Controlled degradation of hydrogels using multi-functional cross-linking molecules. , 2004, Biomaterials.

[22]  Adrian Neagu,et al.  Tissue engineering by self-assembly of cells printed into topologically defined structures. , 2008, Tissue engineering. Part A.

[23]  D. Tyler McQuade,et al.  Polymer chemistry in flow: New polymers, beads, capsules, and fibers , 2006 .

[24]  Ali Khademhosseini,et al.  Progress in tissue engineering. , 2009, Scientific American.

[25]  S. Bhatia,et al.  An extracellular matrix microarray for probing cellular differentiation , 2005, Nature Methods.

[26]  Karoly Jakab,et al.  Tissue engineering by self-assembly and bio-printing of living cells , 2010, Biofabrication.

[27]  R. Zengerle,et al.  Alginate bead fabrication and encapsulation of living cells under centrifugally induced artificial gravity conditions. , 2008, Journal of microencapsulation.

[28]  D. Beebe,et al.  Biological implications of polydimethylsiloxane-based microfluidic cell culture. , 2009, Lab on a chip.

[29]  Charles N. Baroud,et al.  Quantitative analysis of the dripping and jetting regimes in co-flowing capillary jets , 2010, 1011.2428.

[30]  D. Beebe,et al.  Cell culture models in microfluidic systems. , 2008, Annual review of analytical chemistry.

[31]  T. Fujii PDMS-based microfluidic devices for biomedical applications , 2002 .

[32]  S. Bhatia,et al.  Three-Dimensional Photopatterning of Hydrogels Containing Living Cells , 2002 .

[33]  Ali Khademhosseini,et al.  Hybrid PGS–PCL microfibrous scaffolds with improved mechanical and biological properties , 2011, Journal of tissue engineering and regenerative medicine.

[34]  Vladimir Mironov,et al.  Review: bioprinting: a beginning. , 2006, Tissue engineering.

[35]  P. Ma,et al.  Ionically crosslinked alginate hydrogels as scaffolds for tissue engineering: part 1. Structure, gelation rate and mechanical properties. , 2001, Biomaterials.

[36]  A. Kasko,et al.  Two-photon lithography in the future of cell-based therapeutics and regenerative medicine: a review of techniques for hydrogel patterning and controlled release. , 2010, Future medicinal chemistry.

[37]  Kristi S Anseth,et al.  Synthesis and characterization of photocrosslinkable, degradable poly(vinyl alcohol)-based tissue engineering scaffolds. , 2002, Biomaterials.

[38]  A. Manz,et al.  Lab-on-a-chip: microfluidics in drug discovery , 2006, Nature Reviews Drug Discovery.

[39]  Catia Bastioli,et al.  Handbook of Biodegradable Polymers , 2005 .

[40]  K. Katti,et al.  Nanoclay Based Composite Scaffolds for Bone Tissue Engineering Applications , 2010 .

[41]  Ali Khademhosseini,et al.  Stimuli-responsive microwells for formation and retrieval of cell aggregates. , 2010, Lab on a chip.

[42]  Loo-Teck Ng,et al.  Synthesis of hydrogel for drug delivery studies utilizing photoinitiator-free photopolymerization based on the donor/acceptor pair, N-vinylpyrrolidinone and hydroxypentyl maleimide , 2002 .

[43]  A. Bigi,et al.  Stabilization of gelatin films by crosslinking with genipin. , 2002, Biomaterials.

[44]  L. Bonassar,et al.  Tissue engineering: the first decade and beyond. , 1998, Journal of cellular biochemistry. Supplement.

[45]  Kytai Truong Nguyen,et al.  Photopolymerizable hydrogels for tissue engineering applications. , 2002, Biomaterials.

[46]  Masanori Fujita,et al.  Controlled release of fibroblast growth factors and heparin from photocrosslinked chitosan hydrogels and subsequent effect on in vivo vascularization. , 2003, Journal of biomedical materials research. Part A.

[47]  I. Morita,et al.  Biocompatible inkjet printing technique for designed seeding of individual living cells. , 2005, Tissue engineering.

[48]  G. Whitesides,et al.  Soft Lithography. , 1998, Angewandte Chemie.

[49]  Leslie Y Yeo,et al.  Microfluidic devices for bioapplications. , 2011, Small.

[50]  Jeffrey A Hubbell,et al.  Photopolymerized hyaluronic acid-based hydrogels and interpenetrating networks. , 2003, Biomaterials.

[51]  Kahp-Yang Suh,et al.  Fabrication of three-dimensional microstructures by soft molding , 2001 .

[52]  D. Beebe,et al.  Fundamentals of microfluidic cell culture in controlled microenvironments. , 2010, Chemical Society reviews.

[53]  G. Moonen,et al.  Poly(D,L-lactide) foams modified by poly(ethylene oxide)-block-poly(D,L-lactide) copolymers and a-FGF: in vitro and in vivo evaluation for spinal cord regeneration. , 2001, Biomaterials.

[54]  Suwan N Jayasinghe,et al.  Bio-electrospraying embryonic stem cells: interrogating cellular viability and pluripotency. , 2009, Integrative biology : quantitative biosciences from nano to macro.

[55]  Vladimir Mironov,et al.  Organ printing: from bioprinter to organ biofabrication line. , 2011, Current opinion in biotechnology.

[56]  T. Lötzbeyer,et al.  Enzymatic cross-linking of proteins with tyrosinase , 2002 .

[57]  Ali Khademhosseini,et al.  Directed assembly of cell-laden microgels for building porous three-dimensional tissue constructs. , 2011, Journal of biomedical materials research. Part A.

[58]  Gwo-Bin Lee,et al.  Microfluidic cell culture systems for drug research. , 2010, Lab on a chip.

[59]  Robert L Sah,et al.  Photo-and Electropatterning of Hydrogel-encapsulated Living Cell Arrays , 2004 .

[60]  S. Takeuchi,et al.  Monodisperse Alginate Hydrogel Microbeads for Cell Encapsulation , 2007 .

[61]  Séverine Le Gac,et al.  Single cells as experimentation units in lab-on-a-chip devices , 2010 .

[62]  Laurent Griscom,et al.  Trends in the development of microfluidic cell biochips for in vitro hepatotoxicity. , 2007, Toxicology in vitro : an international journal published in association with BIBRA.

[63]  Ali Khademhosseini,et al.  Molded polyethylene glycol microstructures for capturing cells within microfluidic channels. , 2004, Lab on a chip.

[64]  Dhananjay Dendukuri,et al.  Stop-flow lithography in a microfluidic device. , 2007, Lab on a chip.

[65]  A. Levchenko,et al.  Lab-on-a-chip devices as an emerging platform for stem cell biology. , 2010, Lab on a chip.

[66]  Hanry Yu,et al.  Stem cells in microfluidics , 2009, Biotechnology progress.

[67]  Shoji Takeuchi,et al.  Monodisperse cell-encapsulating peptide microgel beads for 3D cell culture. , 2010, Langmuir : the ACS journal of surfaces and colloids.

[68]  Suwan N Jayasinghe,et al.  Combining bio-electrospraying with gene therapy: a novel biotechnique for the delivery of genetic material via living cells. , 2010, The Analyst.

[69]  Ali Khademhosseini,et al.  Synthesis and characterization of photocrosslinkable gelatin and silk fibroin interpenetrating polymer network hydrogels. , 2011, Acta biomaterialia.

[70]  Won-Gun Koh,et al.  Fabrication of cell-containing hydrogel microstructures inside microfluidic devices that can be used as cell-based biosensors , 2006, Analytical and bioanalytical chemistry.

[71]  George M. Whitesides,et al.  Replica molding using polymeric materials: A practical step toward nanomanufacturing , 1997 .

[72]  Woo Young Sim,et al.  A pneumatic micro cell chip for the differentiation of human mesenchymal stem cells under mechanical stimulation. , 2007, Lab on a chip.

[73]  Brian Derby,et al.  Bioprinting: Inkjet printing proteins and hybrid cell-containing materials and structures , 2008 .

[74]  Ali Khademhosseini,et al.  A microwell array system for stem cell culture. , 2008, Biomaterials.

[75]  R. Appell,et al.  De Novo Reconstitution of a Functional Mammalian Urinary Bladder by Tissue Engineering , 1999 .

[76]  M. Djabourov,et al.  Water in water emulsions: phase separation and rheology of biopolymer solutions , 2001 .

[77]  A. Giacca,et al.  In vitro and in vivo testing of glucose-responsive insulin-delivery microdevices in diabetic rats. , 2012, Lab on a chip.

[78]  Ali Khademhosseini,et al.  Surface‐directed assembly of cell‐laden microgels , 2010, Biotechnology and Bioengineering.

[79]  Eben Alsberg,et al.  Photocrosslinked alginate hydrogels with tunable biodegradation rates and mechanical properties. , 2009, Biomaterials.

[80]  N. Perrimon,et al.  Droplet microfluidic technology for single-cell high-throughput screening , 2009, Proceedings of the National Academy of Sciences.

[81]  Liang-Yin Chu,et al.  Controllable monodisperse multiple emulsions. , 2007, Angewandte Chemie.

[82]  J. Vorhies,et al.  Synthetic vs. natural/biodegradable polymers for delivery of shRNA-based cancer therapies. , 2009, Methods in molecular biology.

[83]  J L West,et al.  Development and optimization of a dual-photoinitiator, emulsion-based technique for rapid generation of cell-laden hydrogel microspheres. , 2011, Acta biomaterialia.

[84]  Edwin L. Thomas,et al.  Periodic materials and interference lithography , 2008 .

[85]  Christoph A. Merten,et al.  Droplet-based microfluidic platforms for the encapsulation and screening of Mammalian cells and multicellular organisms. , 2008, Chemistry & biology.

[86]  P. Messersmith,et al.  In situ crosslinking of a biomimetic peptide-PEG hydrogel via thermally triggered activation of factor XIII. , 2002, Biomaterials.

[87]  S. Seiffert Functional microgels tailored by droplet-based microfluidics. , 2011, Macromolecular rapid communications.

[88]  Ali Khademhosseini,et al.  Synthesis and characterization of tunable poly(ethylene glycol): gelatin methacrylate composite hydrogels. , 2011, Tissue engineering. Part A.

[89]  Jackie Y Ying,et al.  Hydrodynamic spinning of hydrogel fibers. , 2010, Biomaterials.

[90]  Uwe Sauer,et al.  From good old biochemical analyses to high-throughput omics measurements and back. , 2011, Current opinion in biotechnology.

[91]  Matthew H. M. Lim,et al.  Perfused multiwell plate for 3D liver tissue engineering. , 2010, Lab on a chip.

[92]  Yu-Cheng Lin,et al.  Chitosan microfiber fabrication using a microfluidic chip and its application to cell cultures , 2009 .

[93]  Achim Goepferich,et al.  Rational design of hydrogels for tissue engineering: impact of physical factors on cell behavior. , 2007, Biomaterials.

[94]  G. Whitesides The origins and the future of microfluidics , 2006, Nature.

[95]  A. Bhattacharya,et al.  Studies On The Crosslinking Of Poly (Vinyl Alcohol) , 2006 .

[96]  Roman Stocker,et al.  Microfluidics for bacterial chemotaxis. , 2010, Integrative biology : quantitative biosciences from nano to macro.

[97]  Annelise E Barron,et al.  Modular enzymatically crosslinked protein polymer hydrogels for in situ gelation. , 2010, Biomaterials.

[98]  J. West,et al.  Visible light photoinitiation of mesenchymal stem cell-laden bioresponsive hydrogels. , 2011, European cells & materials.

[99]  Andrew J. deMello,et al.  Rapid cell extraction in aqueous two-phase microdroplet systems , 2010 .

[100]  M. Shoichet,et al.  Synthesis of enzyme-degradable, peptide-cross-linked dextran hydrogels. , 2007, Bioconjugate chemistry.

[101]  G. Whitesides,et al.  Flexible Methods for Microfluidics , 2001 .

[102]  Ali Khademhosseini,et al.  Biomimetic tissues on a chip for drug discovery. , 2012, Drug discovery today.

[103]  Ali Khademhosseini,et al.  Digitally tunable physicochemical coding of material composition and topography in continuous microfibres. , 2011, Nature materials.

[104]  R. Zhuo,et al.  Biodegradable and pH-sensitive hydrogels for cell encapsulation and controlled drug release. , 2008, Biomacromolecules.

[105]  G. Prestwich,et al.  Photocrosslinkable hyaluronan-gelatin hydrogels for two-step bioprinting. , 2010, Tissue engineering. Part A.

[106]  Seeram Ramakrishna,et al.  An Introduction to Electrospinning and Nanofibers (Paperback) , 2005 .

[107]  Liu Ning,et al.  Radiation preparation of PVA-g-NIPAAm in a homogeneous system and its application in controlled release , 2000 .

[108]  Roger D. Kamm,et al.  Microfluidic Platforms for Studies of Angiogenesis, Cell Migration, and Cell–Cell Interactions , 2010, Annals of Biomedical Engineering.

[109]  Chang-Soo Lee,et al.  Synthesis and utilization of E. coli-encapsulated PEG-based microdroplet using a microfluidic chip for biological application. , 2010, Biotechnology and bioengineering.

[110]  Roger D Kamm,et al.  Microfluidic devices for studying heterotypic cell-cell interactions and tissue specimen cultures under controlled microenvironments. , 2011, Biomicrofluidics.

[111]  VLADIMIR MIRONOV,et al.  Bioprinting : A Beginning , 2022 .

[112]  J. A. Hubbell,et al.  Cell‐Responsive Synthetic Hydrogels , 2003 .

[113]  G. Whitesides,et al.  Soft lithography in biology and biochemistry. , 2001, Annual review of biomedical engineering.

[114]  A. Khademhosseini,et al.  A cell-laden microfluidic hydrogel. , 2007, Lab on a chip.

[115]  Hiroyuki Nakamura,et al.  Controllable polymerization of N-carboxy anhydrides in a microreaction system. , 2005, Lab on a chip.

[116]  J. Aizenberg,et al.  Synthesis of photoacid crosslinkable hydrogels for the fabrication of soft, biomimetic microlens arrays , 2005 .

[117]  Christopher Cannizzaro,et al.  Nanofabrication and microfabrication of functional materials for tissue engineering. , 2007, Tissue engineering.

[118]  Robert Langer,et al.  Micromolding of photocrosslinkable hyaluronic acid for cell encapsulation and entrapment. , 2006, Journal of biomedical materials research. Part A.

[119]  Karl R Edminster,et al.  Multi-layered culture of human skin fibroblasts and keratinocytes through three-dimensional freeform fabrication. , 2009, Biomaterials.

[120]  Chien-Chi Lin,et al.  PEG hydrogels formed by thiol-ene photo-click chemistry and their effect on the formation and recovery of insulin-secreting cell spheroids. , 2011, Biomaterials.

[121]  A. Khademhosseini,et al.  Cell-laden microengineered gelatin methacrylate hydrogels. , 2010, Biomaterials.

[122]  George M. Whitesides,et al.  Fabrication of single‐mode polymeric waveguides using micromolding in capillaries , 1996 .

[123]  E. Zussman,et al.  Encapsulation of bacterial cells in electrospun microtubes. , 2009, Biomacromolecules.

[124]  G. Whitesides,et al.  Fabrication of three‐dimensional micro‐structures: Microtransfer molding , 1996 .

[125]  Suming Li,et al.  Hydrolytic and enzymatic degradations of physically crosslinked hydrogels prepared from PLA/PEO/PLA triblock copolymers , 2002, Journal of materials science. Materials in medicine.

[126]  D. Beebe,et al.  Controlled microfluidic interfaces , 2005, Nature.

[127]  Adrian Neagu,et al.  Three-dimensional tissue constructs built by bioprinting. , 2006, Biorheology.

[128]  Suwan N Jayasinghe,et al.  Cell electrospinning: a unique biotechnique for encapsulating living organisms for generating active biological microthreads/scaffolds. , 2006, Biomacromolecules.

[129]  Jun Zhang,et al.  Fabrication of High Aspect Ratio Poly(ethylene glycol)-Containing Microstructures by UV Embossing , 2003 .

[130]  A. Khademhosseini,et al.  Hydrogels in Regenerative Medicine , 2009, Advanced materials.

[131]  Robert Langer,et al.  Thermoresponsive Platforms for Tissue Engineering and Regenerative Medicine. , 2011, AIChE journal. American Institute of Chemical Engineers.

[132]  Suwan N Jayasinghe,et al.  Bio-electrospraying and droplet-based microfluidics: control of cell numbers within living residues. , 2010, Biomedical materials.

[133]  Satoyuki Kawano,et al.  Micro cell encapsulation and its hydrogel-beads production using microfluidic device , 2007 .

[134]  Robert Langer,et al.  Responsive micromolds for sequential patterning of hydrogel microstructures. , 2011, Journal of the American Chemical Society.

[135]  David J Beebe,et al.  From the cellular perspective: exploring differences in the cellular baseline in macroscale and microfluidic cultures. , 2009, Integrative biology : quantitative biosciences from nano to macro.

[136]  F. Horkay,et al.  Nanostructured Hybrid Hydrogels Prepared by a Combination of Atom Transfer Radical Polymerization and Free Radical Polymerization , 2022 .

[137]  Jason A Burdick,et al.  Review: photopolymerizable and degradable biomaterials for tissue engineering applications. , 2007, Tissue engineering.

[138]  E. Kamio,et al.  Monodisperse water-in-water-in-oil emulsion droplets. , 2011, Chemphyschem : a European journal of chemical physics and physical chemistry.

[139]  Mehmet Toner,et al.  Surface engineering with poly(ethylene glycol) photolithography to create high-density cell arrays on glass , 2003 .

[140]  Eric D. Miller,et al.  Inkjet-based biopatterning of bone morphogenetic protein-2 to spatially control calvarial bone formation. , 2010, Tissue engineering. Part A.

[141]  W. Hennink,et al.  The use of aqueous PEG/dextran phase separation for the preparation of dextran microspheres. , 1999, International journal of pharmaceutics.

[142]  Eric D. Miller,et al.  Microenvironments Engineered by Inkjet Bioprinting Spatially Direct Adult Stem Cells Toward Muscle‐ and Bone‐Like Subpopulations , 2008, Stem cells.

[143]  S. Bryant,et al.  Cell encapsulation in biodegradable hydrogels for tissue engineering applications. , 2008, Tissue engineering. Part B, Reviews.

[144]  John A Rogers,et al.  Fabricating three-dimensional nanostructures using two photon lithography in a single exposure step. , 2006, Optics express.

[145]  Adrian Neagu,et al.  Role of physical mechanisms in biological self-organization. , 2005, Physical review letters.

[146]  C. van Nostrum,et al.  Novel crosslinking methods to design hydrogels. , 2002, Advanced drug delivery reviews.

[147]  D. Weitz,et al.  Dripping, Jetting, Drops, and Wetting: The Magic of Microfluidics , 2007 .

[148]  Jiajie Yu,et al.  Microscale 3-D hydrogel scaffold for biomimetic gastrointestinal (GI) tract model. , 2011, Lab on a chip.

[149]  Chang-Soo Lee,et al.  Generation of Monodisperse Inorganic–Organic Janus Microspheres in a Microfluidic Device , 2009 .

[150]  Viness Pillay,et al.  A Review on Composite Liposomal Technologies for Specialized Drug Delivery , 2011, Journal of drug delivery.

[151]  Toru Torii,et al.  Synthesis of Monodisperse Bicolored Janus Particles with Electrical Anisotropy Using a Microfluidic Co‐Flow System , 2006 .

[152]  Vladimir Mironov,et al.  Bioprinting living structures , 2007 .

[153]  George M. Whitesides,et al.  Micromolding in Capillaries: Applications in Materials Science , 1996 .

[154]  A. Abate,et al.  Synthesis of Monodisperse Microparticles from Non‐Newtonian Polymer Solutions with Microfluidic Devices , 2011, Advanced materials.

[155]  S. I. Park,et al.  Involvement of TGF-beta1 signaling in cardiomyocyte differentiation from P19CL6 cells. , 2007, Molecules and cells.

[156]  Niraj K Inamdar,et al.  Microfluidic cell culture models for tissue engineering. , 2011, Current opinion in biotechnology.

[157]  Anthony Atala,et al.  De Novo Reconstitution of a Functional Mammalian Urinary Bladder by Tissue Engineering , 1999 .

[158]  Suwan N Jayasinghe,et al.  Cell electrospinning highly concentrated cellular suspensions containing primary living organisms into cell-bearing threads and scaffolds. , 2007, Nanomedicine.

[159]  S. Quake,et al.  Microfluidics: Fluid physics at the nanoliter scale , 2005 .

[160]  S. Waldman,et al.  Genipin Cross-Linked Fibrin Hydrogels for in vitro Human Articular Cartilage Tissue-Engineered Regeneration , 2009, Cells Tissues Organs.

[161]  J. Fisher,et al.  Photoinitiated Polymerization of Biomaterials , 2001 .

[162]  J. Park,et al.  Continuous generation of hydrogel beads and encapsulation of biological materials using a microfluidic droplet-merging channel , 2008 .

[163]  S. Verma,et al.  Biodegradable Polymers for Emerging Clinical Use in Tissue Engineering , 2011 .