[Potential of chondrogenesis of bone marrow stromal cells co-cultured with chondrocytes on biodegradable scaffold: in vivo experiment with pigs and mice].
OBJECTIVE
To explore the feasibility of in vivo chondrogenesis of bone marrow stromal cells (BMSCs) co-cultured with chondrocytes on biodegradable scaffold.


METHODS
Porcine BMSCs were isolated, expanded and labeled with enhanced green fluorescent protein (EGFP), and then were mixed with articular chondrocytes isolated from porcine knee joint at the ratio of 1:1. The mixed cells were seeded onto polyglycolic acid (PGA) scaffold at the ultimate concentration of 5.0 x 10(7)/ml (co-culture group). Pure chondrocytes and BMSCs of the same ultimate concentration were seeded respectively onto the scaffold as positive control group and negative control group. After two weeks' culture in vitro, they were planted subcutaneously into nude mice respectively. These specimens were collected after in vivo implantation for 8 weeks to undergo microscopy. Laser confocal microscopy was used to observe the distribution of EGFP-labeled cells in the tissue. RT-PCR was used to examine the expression of collagen type II and aggrecan. Immunohistochemistry was used to observe the protein expression of collagen type II.


RESULTS
The cell-scaffold constructs of the co-culture group and positive control group, could maintain the original size and shape no matter in vitro or in vivo. After 8 weeks' in vivo implantation, the constructs in both co-culture group and positive control group formed cartilage-like tissue with typical histological structure and extracellular matrix staining similar to those of the normal cartilage. The GAG content and compressive modulus of the co-culture group reached over 80% of those of the positive control group. Confocal microscopy revealed the presence of EGFP-labeled cells in the engineered cartilage lacuna. Histological examination showed that the constructs of the negative control group shrunk gradually after in vivo implantation with no typical cartilage-like tissue formation.


CONCLUSIONS
In vitro co-cultured BMSC-chondrocyte-PGA constructs have the potential to form mature cartilage-like tissue in subcutaneous non-chondrogenesis environment, indicating that chondrocytes still provide enough signals for BMSC chondrogenic differentiation.
Liu
Xia
X
Department of Plastic and Reconstructive Surgery, Shanghai 9th People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai Key Laboratory of Tissue Engineering, Shanghai 200011, China.
Zhou
Guang-dong
GD
Lü
Xiao-jie
XJ
Liu
Tian-yi
TY
Zhang
Wen-jie
WJ
Liu
Wei
W
Cao
Yi-lin
YL
D053674:Aggrecans / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D017690:Cell Transplantation / Q000379:methods*; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D018920:Coculture Techniques; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D005240:Feasibility Studies; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008819:Mice, Nude; D011100:Polyglycolic Acid / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017154:Stromal Cells / Q000166:cytology* / Q000378:metabolism; D013552:Swine; D054457:Tissue Scaffolds / Q000737:chemistry; D014162:Transfection
D053674:Aggrecans; D001672:Biocompatible Materials; D024043:Collagen Type II; D049452:Green Fluorescent Proteins; D011100:Polyglycolic Acid