Bioactive polymer/extracellular matrix scaffolds fabricated with a flow perfusion bioreactor for cartilage tissue engineering.

In this study, electrospun poly(ɛ-caprolactone) (PCL) microfiber scaffolds, coated with cartilaginous extracellular matrix (ECM), were fabricated by first culturing chondrocytes under dynamic conditions in a flow perfusion bioreactor and then decellularizing the cellular constructs. The decellularization procedure yielded acellular PCL/ECM composite scaffolds containing glycosaminoglycan and collagen. PCL/ECM composite scaffolds were evaluated for their ability to support the chondrogenic differentiation of mesenchymal stem cells (MSCs) in vitro using serum-free medium with or without the addition of transforming growth factor-β1 (TGF-β1). PCL/ECM composite scaffolds supported chondrogenic differentiation induced by TGF-β1 exposure, as evidenced in the up-regulation of aggrecan (11.6 ± 3.8 fold) and collagen type II (668.4 ± 317.7 fold) gene expression. The presence of cartilaginous matrix alone reduced collagen type I gene expression to levels observed with TGF-β1 treatment. Cartilaginous matrix further enhanced the effects of growth factor treatment on MSC chondrogenesis as evidenced in the higher glycosaminoglycan synthetic activity for cells cultured on PCL/ECM composite scaffolds. Therefore, flow perfusion culture of chondrocytes on electrospun microfiber scaffolds is a promising method to fabricate polymer/extracellular matrix composite scaffolds that incorporate both natural and synthetic components to provide biological signals for cartilage tissue engineering applications.

[1]  Magnus Lundberg,et al.  Treatment of osteochondral defects with autologous bone marrow in a hyaluronan-based delivery vehicle. , 2002, Tissue engineering.

[2]  Jerry C. Hu,et al.  Low-density cultures of bovine chondrocytes: effects of scaffold material and culture system. , 2005, Biomaterials.

[3]  Antonios G Mikos,et al.  Design of a flow perfusion bioreactor system for bone tissue-engineering applications. , 2003, Tissue engineering.

[4]  L. Bonassar,et al.  Comparison of Chondrogensis in Static and Perfused Bioreactor Culture , 2000, Biotechnology progress.

[5]  H. Stegemann,et al.  Determination of hydroxyproline. , 1967, Clinica chimica acta; international journal of clinical chemistry.

[6]  Antonios G Mikos,et al.  In vitro generated extracellular matrix and fluid shear stress synergistically enhance 3D osteoblastic differentiation. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[7]  V. Hascall,et al.  Turnover of proteoglycans in cultures of bovine articular cartilage. , 1984, Archives of biochemistry and biophysics.

[8]  R. Tuan,et al.  Biological response of chondrocytes cultured in three-dimensional nanofibrous poly(epsilon-caprolactone) scaffolds. , 2003, Journal of biomedical materials research. Part A.

[9]  Margherita Cioffi,et al.  The effect of hydrodynamic shear on 3D engineered chondrocyte systems subject to direct perfusion. , 2006, Biorheology.

[10]  M. Weiss,et al.  Effect of initial seeding density on human umbilical cord mesenchymal stromal cells for fibrocartilage tissue engineering. , 2009, Tissue engineering. Part A.

[11]  H. Kleinman,et al.  Complex Extracellular Matrices Promote Tissue‐Specific Stem Cell Differentiation , 2005, Stem cells.

[12]  A. Mikos,et al.  Electrospun poly(epsilon-caprolactone) microfiber and multilayer nanofiber/microfiber scaffolds: characterization of scaffolds and measurement of cellular infiltration. , 2006, Biomacromolecules.

[13]  B. Obradovic,et al.  Bioreactor cultivation conditions modulate the composition and mechanical properties of tissue‐engineered cartilage , 1999, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[14]  Antonios G. Mikos,et al.  Injectable biodegradable hydrogel composites for rabbit marrow mesenchymal stem cell and growth factor delivery for cartilage tissue engineering. , 2007, Biomaterials.

[15]  Antonios G Mikos,et al.  The influence of an in vitro generated bone-like extracellular matrix on osteoblastic gene expression of marrow stromal cells. , 2008, Biomaterials.

[16]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[17]  M. Akens,et al.  Changes in subchondral bone in cartilage resurfacing--an experimental study in sheep using different types of osteochondral grafts. , 2003, Osteoarthritis and cartilage.

[18]  David A. Bader,et al.  Facial Expression Recognition System using Statistical Feature and Neural Network , 2012 .

[19]  T. Simon,et al.  Articular Cartilage: Injury Pathways and Treatment Options , 2006, Sports medicine and arthroscopy review.

[20]  D Herbage,et al.  Optimization of dynamic culture conditions: effects on biosynthetic activities of chondrocytes grown in collagen sponges. , 2005, Tissue engineering.

[21]  A. Yarin,et al.  Chondrogenic differentiation of human mesenchymal stem cells on oriented nanofibrous scaffolds: engineering the superficial zone of articular cartilage. , 2009, Tissue engineering. Part A.

[22]  G. Dickson,et al.  Fabrication and repair of cartilage defects with a novel acellular cartilage matrix scaffold. , 2010, Tissue engineering. Part C, Methods.

[23]  M. Brittberg.  Autologous chondrocyte implantation--technique and long-term follow-up. , 2008, Injury.

[24]  Wan-Ju Li,et al.  Biological response of chondrocytes cultured in three-dimensional nanofibrous poly(ϵ-caprolactone) scaffolds† , 2003 .

[25]  V. Goldberg,et al.  High Variability in Rabbit Bone Marrow-Derived Mesenchymal Cell Preparations , 1999, Cell transplantation.

[26]  W. Comper,et al.  Passive loss of proteoglycan from articular cartilage explants. , 1989, Biochimica et biophysica acta.

[27]  R. Tompkins,et al.  Differences in dermal analogs influence subsequent pigmentation, epidermal differentiation, basement membrane, and rete ridge formation of transplanted composite skin grafts. , 1997, Transplantation.

[28]  V. Sikavitsas,et al.  Effect of bone extracellular matrix synthesized in vitro on the osteoblastic differentiation of marrow stromal cells. , 2005, Biomaterials.

[29]  Boon Chin Heng,et al.  Directing Stem Cell Differentiation into the Chondrogenic Lineage In Vitro , 2004, Stem cells.

[30]  F. Guilak,et al.  Chondrogenic differentiation of adipose-derived adult stem cells by a porous scaffold derived from native articular cartilage extracellular matrix. , 2009, Tissue engineering. Part A.

[31]  James D. Kang,et al.  Quantitative analysis of gene expression in a rabbit model of intervertebral disc degeneration by real-time polymerase chain reaction. , 2005, The spine journal : official journal of the North American Spine Society.

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

[33]  K. Athanasiou,et al.  Extraction techniques for the decellularization of tissue engineered articular cartilage constructs. , 2009, Biomaterials.

[34]  A. Mikos,et al.  Modulation of osteogenic properties of biodegradable polymer/extracellular matrix scaffolds generated with a flow perfusion bioreactor. , 2010, Acta biomaterialia.

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

[36]  Brian J. Cole,et al.  Autologous Chondrocyte Implantation , 2001 .

[37]  John J. Keeling,et al.  A comparison of open versus arthroscopic harvesting of osteochondral autografts. , 2009, The Knee.

[38]  R. Langer,et al.  Tissue engineering of bovine articular cartilage within porous poly(ether ester) copolymer scaffolds with different structures. , 2005, Tissue engineering.

[39]  Antonios G Mikos,et al.  In vitro localization of bone growth factors in constructs of biodegradable scaffolds seeded with marrow stromal cells and cultured in a flow perfusion bioreactor. , 2006, Tissue engineering.