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目的: 体外诱导骨髓间充质干细胞(MSC)向软骨细胞定向分化, 并对分化后的细胞进行鉴定。方法: 由健康成人骨髓中分离出MSC, 取第 3代细胞进行实验。鉴定后, 将微小细胞团在TGF -β1、地塞米松(Dex)及维生素C(VitC)等诱导下分化。14d后, 细胞团经石蜡包埋、切片及HE染色后, 进行甲苯胺蓝染色及II型胶原(ColII)的免疫组化染色。采用Westernblot和RT- PCR, 分别检测诱导前后MSC中ColII及前ColIImRNA的表达。结果: HE染色显示, 诱导后细胞呈软骨细胞样形态; 甲苯胺蓝及ColII染色的细胞外基质呈阳性。Westernblot和RT PCR的结果显示, 诱导分化后的MSC可表达ColII和前ColII的mRNA。结论: MSC在TGF- β1、Dex及VitC等诱导后, 可分化为软骨细胞。
OBJECTIVE: To induce the differentiation of bone marrow mesenchymal stem cells (MSC) into chondrocytes in vitro and to identify the differentiated cells. METHODS: MSC were isolated from healthy adult bone marrow and the third generation of cells were harvested for experiments. After identification, tiny cell clusters were differentiated under the induction of TGF-β1, Dex and VitC. After 14 days, the cell mass was paraffin-embedded, sectioned and HE stained for toluidine blue staining and type II collagen (ColII) immunohistochemical staining. Western Blot and RT-PCR were used to detect the expression of ColII and procollagen mRNA in MSC before and after induction. Results: The HE staining showed that the cells showed chondrocyte-like morphology after induction, and the extracellular matrix of toluidine blue and ColII staining was positive. The results of Western blot and RT PCR showed that differentiated MSCs can express ColII and pre-ColII mRNA. Conclusion: MSC can differentiate into chondrocytes after induced by TGF-β1, Dex and VitC.