Thrombospondin-1 inhibits ossification of tissue engineered cartilage constructed by ADSCs

Cartilage tissue engineering provides a new method in the treatment of cartilage defects, and adipose derived stem cells seem to be an ideal seed cell in cartilage tissue engineering because of its characteristics. However, ossification after in vivo implantation of tissue engineered cartilage remains a challenge. Thrombospondin-1 which has been reported to have an inhibitory effect on angiogenesis, may play an important role in inhibiting the ossification of tissue engineered cartilage constructed by adipose derived stem cells. Therefore, the effect of thrombospondin-1 in inhibiting the ossification of tissue engineered cartilage was evaluated in this study. Lentivirus vectors carrying thrombospondin-1 cDNA were transfected into adipose derived stem cells, and the transfected cells were used in the experiments. The expression of thrombospondin-1 was evaluated by quantitative reverse transcriptase-polymerase chain reaction and western blot, and the effects of thrombospondin-1 over-expression on angiogenesis were analyzed by angiogenesis assays. The quality of tissue engineered cartilage and the degree of ossification were assessed by biomechanical and molecular biology methods. The results showed that thrombospondin-1 infected cells have a high expression of thrombospondin-1 in mRNA and protein level, which inhibited the tube formation of endothelial cells, indicating the anti-angiogenic effects. Gene expression analyses in vitro showed that thrombospondin-1 inhibits the osteogenic differentiation of adipose derived stem cells significantly, and the results of in vivo study revealed that thrombospondin-1 significantly inhibits the expression of osteogenic genes. Compared to that in the control group, tissue engineered cartilage constructed by thrombospondin-1 transfected adipose derived stem cells in vivo showed a higher GAG content and lower compressive modulus, which indicating lower level of ossification. In conclusion, the current study indicated that the anti-angiogenic factor thrombospondin-1 suppresses the osteogenic differentiation of adipose derived stem cells in vitro, and inhibits ossification of tissue engineered cartilage constructed by adipose derived stem cells in vivo.