Addition of hyaluronic acid improves cellular infiltration and promotes early-stage chondrogenesis in a collagen-based scaffold for cartilage tissue engineering.

The response of mesenchymal stem cells (MSCs) to a matrix largely depends on the composition as well as the extrinsic mechanical and morphological properties of the substrate to which they adhere to. Collagen-glycosaminoglycan (CG) scaffolds have been extensively used in a range of tissue engineering applications with great success. This is due in part to the presence of the glycosaminoglycans (GAGs) in complementing the biofunctionality of collagen. In this context, the overall goal of this study was to investigate the effect of two GAG types: chondroitin sulphate (CS) and hyaluronic acid (HyA) on the mechanical and morphological characteristics of collagen-based scaffolds and subsequently on the differentiation of rat MSCs in vitro. Morphological characterisation revealed that the incorporation of HyA resulted in a significant reduction in scaffold mean pore size (93.9 μm) relative to collagen-CS (CCS) scaffolds (136.2 μm). In addition, the collagen-HyA (CHyA) scaffolds exhibited greater levels of MSC infiltration in comparison to the CCS scaffolds. Moreover, these CHyA scaffolds showed significant acceleration of early stage gene expression of SOX-9 (approximately 60-fold higher, p<0.01) and collagen type II (approximately 35-fold higher, p<0.01) as well as cartilage matrix production (7-fold higher sGAG content) in comparison to CCS scaffolds by day 14. Combining their ability to stimulate MSC migration and chondrogenesis in vitro, these CHyA scaffolds show great potential as appropriate matrices for promoting cartilage tissue repair.

[1]  Fergal J O'Brien,et al.  Novel freeze-drying methods to produce a range of collagen-glycosaminoglycan scaffolds with tailored mean pore sizes. , 2010, Tissue engineering. Part C, Methods.

[2]  P. Dieppe,et al.  Pathogenesis and management of pain in osteoarthritis , 2005, The Lancet.

[3]  F. O'Brien,et al.  Osteoblast activity on collagen-GAG scaffolds is affected by collagen and GAG concentrations. , 2009, Journal of biomedical materials research. Part A.

[4]  Fergal J O'Brien,et al.  Influence of freezing rate on pore structure in freeze-dried collagen-GAG scaffolds. , 2004, Biomaterials.

[5]  J. Burdick,et al.  Influence of three-dimensional hyaluronic acid microenvironments on mesenchymal stem cell chondrogenesis. , 2009, Tissue engineering. Part A.

[6]  M. Goldring,et al.  The control of chondrogenesis , 2006, Journal of cellular biochemistry.

[7]  M. Spector,et al.  Cross‐linking affects cellular condensation and chondrogenesis in type II collagen‐GAG scaffolds seeded with bone marrow‐derived mesenchymal stem cells , 2010, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[8]  R. Tuan,et al.  A three-dimensional nanofibrous scaffold for cartilage tissue engineering using human mesenchymal stem cells. , 2005, Biomaterials.

[9]  W. Willems,et al.  Effect of transforming growth factor-beta and growth differentiation factor-5 on proliferation and matrix production by human bone marrow stromal cells cultured on braided poly lactic-co-glycolic acid scaffolds for ligament tissue engineering. , 2007, Tissue engineering.

[10]  Chun-hsien Wu,et al.  Effects of exogenous glycosaminoglycans on human chondrocytes cultivated on type II collagen scaffolds , 2010, Journal of materials science. Materials in medicine.

[11]  P. Janmey,et al.  Tissue Cells Feel and Respond to the Stiffness of Their Substrate , 2005, Science.

[12]  L. Qin,et al.  Gelatin microspheres containing TGF-beta3 enhance the chondrogenesis of mesenchymal stem cells in modified pellet culture. , 2008, Biomacromolecules.

[13]  L. Gibson,et al.  Mechanical characterization of collagen-glycosaminoglycan scaffolds. , 2007, Acta biomaterialia.

[14]  E. Turley,et al.  Ras-transformed cells express both CD44 and RHAMM hyaluronan receptors: only RHAMM is essential for hyaluronan-promoted locomotion. , 1993, Experimental cell research.

[15]  Hsin-Yi Lin,et al.  Chondrogenesis from immortalized human mesenchymal stem cells: comparison between collagen gel and pellet culture methods. , 2008, Artificial organs.

[16]  Amir A. Al-Munajjed,et al.  The healing of bony defects by cell-free collagen-based scaffolds compared to stem cell-seeded tissue engineered constructs. , 2010, Biomaterials.

[17]  M. Spector,et al.  Collagen-GAG Substrate Enhances the Quality of Nerve Regeneration through Collagen Tubes up to Level of Autograft , 1998, Experimental Neurology.

[18]  A J Price,et al.  Temporal trends in hip and knee replacement in the United Kingdom: 1991 to 2006. , 2010, The Journal of bone and joint surgery. British volume.

[19]  W. B. van den Berg,et al.  Linkage of chondroitin-sulfate to type I collagen scaffolds stimulates the bioactivity of seeded chondrocytes in vitro. , 2001, Biomaterials.

[20]  A. U. Daniels,et al.  Effects of scaffold composition and architecture on human nasal chondrocyte redifferentiation and cartilaginous matrix deposition. , 2005, Biomaterials.

[21]  P. Rubin,et al.  Inhibition of conjunctival scarring and contraction by a porous collagen-glycosaminoglycan implant. , 2000, Investigative ophthalmology & visual science.

[22]  Kyoung-Jin Cho,et al.  Functional PLGA scaffolds for chondrogenesis of bone-marrow-derived mesenchymal stem cells. , 2009, Macromolecular bioscience.

[23]  Pieter Buma,et al.  Cross-linked type I and type II collagenous matrices for the repair of full-thickness articular cartilage defects--a study in rabbits. , 2003, Biomaterials.

[24]  Robert A. Brown,et al.  Matrix stiffness and serum concentration effects matrix remodelling and ECM regulatory genes of human bone marrow stem cells , 2008, Journal of tissue engineering and regenerative medicine.

[25]  I. Yannas,et al.  Design of an artificial skin. I. Basic design principles. , 1980, Journal of biomedical materials research.

[26]  Fergal J O'Brien,et al.  The effect of mean pore size on cell attachment, proliferation and migration in collagen-glycosaminoglycan scaffolds for bone tissue engineering. , 2010, Biomaterials.

[27]  T. Fujinaga,et al.  Chondrogenic differentiation of bovine bone marrow mesenchymal stem cells (MSCs) in different hydrogels: Influence of collagen type II extracellular matrix on MSC chondrogenesis , 2006, Biotechnology and bioengineering.

[28]  P. Prendergast,et al.  A collagen-glycosaminoglycan scaffold supports adult rat mesenchymal stem cell differentiation along osteogenic and chondrogenic routes. , 2006, Tissue engineering.

[29]  M. Spector,et al.  Healing of Tendon Defects Implanted with a Porous Collagen-GAG Matrix: Histological Evaluation , 1997 .

[30]  J. Veerkamp,et al.  Attachment of glycosaminoglycans to collagenous matrices modulates the tissue response in rats. , 2000, Biomaterials.

[31]  P. Friedl,et al.  The biology of cell locomotion within three-dimensional extracellular matrix , 2000, Cellular and Molecular Life Sciences CMLS.

[32]  P. Prendergast,et al.  Gene expression by marrow stromal cells in a porous collagen–glycosaminoglycan scaffold is affected by pore size and mechanical stimulation , 2008, Journal of materials science. Materials in medicine.

[33]  N. Rushton,et al.  Current concepts in articular cartilage repair , 2009 .

[34]  S. Gabriel,et al.  Estimates of the prevalence of arthritis and other rheumatic conditions in the United States. Part II. , 2008, Arthritis and rheumatism.

[35]  Maurilio Marcacci,et al.  Patellofemoral Full-Thickness Chondral Defects Treated with Hyalograft-C , 2006, The American journal of sports medicine.

[36]  A. Grodzinsky,et al.  The effects of glycosaminoglycan content on the compressive modulus of cartilage engineered in type II collagen scaffolds. , 2008, Osteoarthritis and cartilage.

[37]  A. P. Newman.  Articular Cartilage Repair , 1998, The American journal of sports medicine.

[38]  M. Pouliot,et al.  Rapid and simple comparison of messenger RNA levels using real-time PCR , 2006, Biological Procedures Online.

[39]  M. Gelinsky,et al.  Pellet culture elicits superior chondrogenic redifferentiation than alginate‐based systems , 2009, Biotechnology progress.

[40]  T. Aigner,et al.  Collagens--major component of the physiological cartilage matrix, major target of cartilage degeneration, major tool in cartilage repair. , 2003, Advanced drug delivery reviews.

[41]  S. Marlovits,et al.  Evaluation of Cartilage Repair Tissue after Matrix-Associated Autologous Chondrocyte Transplantation Using a Hyaluronic-Based or a Collagen-Based Scaffold with Morphological MOCART Scoring and Biochemical T2 Mapping , 2010, The American journal of sports medicine.

[42]  G. Abatangelo,et al.  Functions of hyaluronan in wound repair , 1999, Wound repair and regeneration : official publication of the Wound Healing Society [and] the European Tissue Repair Society.

[43]  F. O'Brien,et al.  Evaluation of early healing events around mesenchymal stem cell-seeded collagen–glycosaminoglycan scaffold. An experimental study in Wistar rats , 2011, Oral and Maxillofacial Surgery.

[44]  A. Mikos,et al.  Review: tissue engineering for regeneration of articular cartilage. , 2000, Biomaterials.

[45]  F. O'Brien,et al.  Towards in vitro vascularisation of collagen-GAG scaffolds. , 2011, European cells & materials.

[46]  W. Knudson,et al.  Hyaluronan and CD44: modulators of chondrocyte metabolism. , 2004, Clinical orthopaedics and related research.

[47]  T. Tsuchiya,et al.  Markedly different effects of hyaluronic acid and chondroitin sulfate-A on the differentiation of human articular chondrocytes in micromass and 3-D honeycomb rotation cultures. , 2007, Journal of biomedical materials research. Part A.

[48]  B. Sharma,et al.  The Future of Cartilage Repair , 2008 .

[49]  S. Nehrer,et al.  Three-year clinical outcome after chondrocyte transplantation using a hyaluronan matrix for cartilage repair. , 2006, European journal of radiology.

[50]  K. Weinberg,et al.  The Role of the Hyaluronan Receptor CD44 in Mesenchymal Stem Cell Migration in the Extracellular Matrix , 2006, Stem cells.

[51]  N. Zini,et al.  Cellular and molecular events during chondrogenesis of human mesenchymal stromal cells grown in a three-dimensional hyaluronan based scaffold. , 2005, Biomaterials.

[52]  Seungju M. Yu,et al.  Enhanced chondrogenesis of mesenchymal stem cells in collagen mimetic peptide-mediated microenvironment. , 2008, Tissue engineering. Part A.

[53]  C. V. van Blitterswijk,et al.  Chitosan scaffolds containing hyaluronic acid for cartilage tissue engineering. , 2011, Tissue engineering. Part C, Methods.

[54]  Adam J. Engler,et al.  Stiffness Gradients Mimicking In Vivo Tissue Variation Regulate Mesenchymal Stem Cell Fate , 2011, PloS one.

[55]  C. Knudson.  Hyaluronan and CD44: strategic players for cell-matrix interactions during chondrogenesis and matrix assembly. , 2003, Birth defects research. Part C, Embryo today : reviews.

[56]  L. Gibson,et al.  The effect of pore size on cell adhesion in collagen-GAG scaffolds. , 2005, Biomaterials.

[57]  J. Buckwalter.  CHONDRAL AND OSTEOCHONDRAL INJURIES: MECHANISMS OF INJURY AND REPAIR RESPONSES , 1997 .

[58]  B. Toole,et al.  Hyaluronan in morphogenesis. , 2001, Journal of internal medicine.

[59]  Weihua Yu,et al.  Chondrogenic differentiation of human mesenchymal stem cells: a comparison between micromass and pellet culture systems , 2010, Biotechnology Letters.