The use of poly(l-lactide) and RGD modified microspheres as cell carriers in a flow intermittency bioreactor for tissue engineering cartilage.

The use of biodegradable microcarriers as initial supports for tissue engineering has been demonstrated to be advantageous for maintaining a differentiated cell phenotype; the high surface area also allows rapid cell expansion. Poly l-lactide (PLLA) is a significant member of a group of polymers regarded as bioresorbable and has been widely used for manufacturing 3D scaffolds for tissue engineering. In this study, the hypothesis that PLLA microspheres could be surface modified using RGD peptide sequences to improve the cell adhesion and function of those cells in contact with PLLA was tested. Using this type of approach it may be possible to generate larger structures that contain a high cell number relative to the amount of polymer, whilst remaining free from mass transport limitations. PLLA microspheres were prepared using an oil-in-water solvent-evaporation technique and then an RGD-motif was incorporated onto the microspheres surface by conjugation to improve cell attachment and function. Both PLLA and GRGDSPK modified PLLA microspheres were used as cell microcarriers for chondrocytes cultured in a flow intermittency bioreactor. At the same time, the degradation of the microspheres has been studied after 7, 14, 21, 28, 35, 49 and 56 days. The molecular weight of the PLLA microspheres was determined by Gel Permeation Chromatography. The morphology was assessed by scanning electron microscopy, and the thermal properties determined by Differential Scanning Calorimetry. It was demonstrated that the RGD modified and pure PLLA microspheres degraded gradually at a steady rate over the experimental period, which would provide a controlled degradation profile, both could serve as cell microcarriers because of their thermal and mechanical stabilities. The microspheres with RGD surface modification enhanced cell adhesion and increased the cell numbers in the microspheres aggregates.

[1]  Y. Ikada,et al.  Degradation of high molecular weight poly(L-lactide) in alkaline medium. , 1995, Biomaterials.

[2]  T. Park,et al.  Biodegradable PLGA Microcarriers for Injectable Delivery of Chondrocytes: Effect of Surface Modification on Cell Attachment and Function , 2004, Biotechnology progress.

[3]  Antonios G Mikos,et al.  Biomimetic materials for tissue engineering. , 2003, Biomaterials.

[4]  Judith M Curran,et al.  Expansion of human chondrocytes in an intermittent stirred flow bioreactor, using modified biodegradable microspheres. , 2005, Tissue engineering.

[5]  T. Chung,et al.  Effects of solvent evaporation rate on the properties of protein-loaded PLLA and PDLLA microspheres fabricated by emulsion-solvent evaporation process , 2002, Journal of microencapsulation.

[6]  Jia-cong Shen,et al.  Cartilage tissue engineering PLLA scaffold with surface immobilized collagen and basic fibroblast growth factor. , 2005, Biomaterials.

[7]  T. Park,et al.  Microencapsulation of dissociable human growth hormone aggregates within poly(D,L-lactic-co-glycolic acid) microparticles for sustained release. , 2001, International journal of pharmaceutics.

[8]  Fred Leonard,et al.  Polylactic Acid for Surgical Implants , 1966 .

[9]  W C de Bruijn,et al.  In vivo degradation and biocompatibility study of in vitro pre-degraded as-polymerized polyactide particles. , 1995, Biomaterials.

[10]  J Tramper,et al.  Expansion of bovine chondrocytes on microcarriers enhances redifferentiation. , 2003, Tissue engineering.

[11]  C. Satriano,et al.  The effect of irradiation modification and RGD sequence adsorption on the response of human osteoblasts to polycaprolactone. , 2005, Biomaterials.

[12]  M. Dame,et al.  Growth of three established cell lines on glass microcarriers , 1983, Biotechnology and bioengineering.

[13]  K. Shakesheff,et al.  Poly(L-lysine)-GRGDS as a biomimetic surface modifier for poly(lactic acid). , 2001, Biomaterials.

[14]  Makarand V Risbud,et al.  Tissue engineering: advances in in vitro cartilage generation. , 2002, Trends in biotechnology.

[15]  Dai Kato,et al.  The design of polymer microcarrier surfaces for enhanced cell growth. , 2003, Biomaterials.

[16]  J. West,et al.  Modification of surfaces with cell adhesion peptides alters extracellular matrix deposition. , 1999, Biomaterials.

[17]  D. Williams,et al.  Mechanisms of polymer degradation in implantable devices. 2. Poly(DL-lactic acid). , 1993, Journal of biomedical materials research.

[18]  K A Athanasiou,et al.  Basic science of articular cartilage repair. , 2001, Clinics in sports medicine.

[19]  E Ruoslahti,et al.  RGD and other recognition sequences for integrins. , 1996, Annual review of cell and developmental biology.

[20]  C. M. Agrawal,et al.  Sterilization, toxicity, biocompatibility and clinical applications of polylactic acid/polyglycolic acid copolymers. , 1996, Biomaterials.

[21]  G. Hofmann Biodegradable implants in orthopaedic surgery--a review on the state-of-the-art. , 1992, Clinical materials.

[22]  C. Heath,et al.  Influence of intermittent pressure, fluid flow, and mixing on the regenerative properties of articular chondrocytes. , 1999, Biotechnology and bioengineering.

[23]  Horst Kessler,et al.  RGD modified polymers: biomaterials for stimulated cell adhesion and beyond. , 2003, Biomaterials.

[24]  G. Wegner,et al.  Investigation of the structure of solution grown crystals of lactide copolymers by means of chemical reactions , 1973 .

[25]  A. Mikos,et al.  Modulation of marrow stromal osteoblast adhesion on biomimetic oligo[poly(ethylene glycol) fumarate] hydrogels modified with Arg-Gly-Asp peptides and a poly(ethyleneglycol) spacer. , 2002, Journal of biomedical materials research.

[26]  Daniel I. C. Wang,et al.  Optimization of growth surface parameters in microcarrier cell culture , 1979 .

[27]  S. Li,et al.  New insights on the degradation of bioresorbable polymeric devices based on lactic and glycolic acids. , 1992, Clinical materials.

[28]  W. S. Hu,et al.  Cultivation of mammalian cells on macroporous microcarriers. , 1992, Enzyme and microbial technology.

[29]  Microencapsulation of Dissociable Human Growth Hormone Aggregates within PLGA Microparticles for Sustained Release , 1999 .

[30]  C. Zavaglia,et al.  In vitro study of poly(lactic acid) pin degradation , 1999 .

[31]  R. Cameron,et al.  Polyglycolide: degradation and drug release. Part I: Changes in morphology during degradation , 2001, Journal of materials science. Materials in medicine.