Abstract
Two major research topics were discussed in this thesis. One is evaluating osteochondral defect repair potential of autologous rabbit bone marrow cells on type II collagen scaffold (COL II scaffold); the other is to discuss the effects of chondrogenesis by culture of mesenchymal stem cells with various types of chondroitin sulfate C (CSC). Moreover, the defect repair potential of CSC-COL II scaffold was also evaluated this thesis. In the first section, after 3-week of in vitro induction, chondrocytic behavior, including marker genes expression and specific extracellular matrix (ECM) secretion, was observed. In the in vivo evaluation experiment, the scaffolds containing RBMSCs without prior induction were autologous implanted into the articular cartilage defects made by subchondral drilling. The rabbits were sacrificed after eight and twenty four weeks. Eight weeks later, chondrocyte-like cells with lacuna structure and corresponding ECM were found in the repaired sites with¬out apparent inflammation. After twenty four weeks, we could easily find cartilage structure the same with normal cartilage in the repair site. In conclusion, it was shown that the scaffolds in combination of in vivo condi¬tions can induce RBMSCs into chondrocytes in repaired area and would be a possible method for articular cartilage repair in clinic and cartilage tissue engineering. In the second section, we studied the effects of chondroitin sulfate C (CSC) on the differentiation of human mesenchymal stem cells (MSCs) toward the chondrocyte lineage. The MSCs were either cultured on type II collagen scaffolds with CSC addition in the medium (free CSC) or with free oligosaccharide CSC. Special attention was given to the effects of MSCs cultured on CSC crosslinked type II scaffolds (crosslinked CSC). According to the analyses of histology stain, gene expression and ECM secretion, our results showed that MSCs cultured with free CSC, free oligosaccharides CSC and on the crosslinked CSC scaffolds all would be induced into chondrocytes. We also found crosslinked CSC scaffold had the good response for chondrogenesis when 35 ug of CSC was crosslinked to COL II scaffold (T+CCR,H35). Moreover, free oligosaccharide CSC presented in the microenvironment could significantly up-regulate MSC chondrogenesis gene expression and stimulate cartilage ECM accumulation more than free CSC with high molecular weight after 3-week induction. Thus, we believed that crosslinked CSC in the scaffold would play the similar roles with free oligosaccharide CSC in the medium. The defect repair potential of CSC-COL II scaffold (T+CCR,H35) was also evaluated by rabbit model. Our results showed that T+CCR,H35 scaffold showed better repair ability on cartilage defect than T+ scaffold by histological and immunohistological staining and the recovered cells in the T+CCR,H35 scaffold had lacuna structure. At the meanwhile, T+CCR,H35 scaffold revealed the type II collagen fibers in the defect site. Furthermore, the generated cells in the T+CCR,H35 scaffold on the repair site showed the higher COL II and aggrecan gene expression and lower type I collagen gene expression compared to the cells in the T+ scaffold after twenty-four week transplantation. Thus, we believed that T+CCR,H35 scaffold would be a potential candidate for cartilage defect repair by tissue engineering approach.