大鼠脂肪干细胞的分离培养及向软骨表型分化的实验研究

The study of isolation, cultivation and differentiation to chondrocytic type of adiposederived stem cells

目的探讨从大鼠的脂肪组织中分离、培养脂肪干细胞（ADSCs）并定向诱导分化为软骨细胞的可行性。方法取Wistar雄性大鼠脂肪组织培养，传代。并用免疫荧光法检测传代ADSCs表面抗原CD29表达。取第3代ADSCs用含有TGF－β4，的培养基定向诱导培养，行Ⅱ型胶原免疫组化染色进行检测鉴定。结果显微镜下可见脂肪干细胞的形态特点。并经免疫荧光法进行检测证实。定向诱导ADSCs向软骨细胞分化，观察细胞形态变化，并通过Ⅱ型胶原免疫组化检测，证实向表达II型胶原的软骨细胞表型分化的可行性。结论证实大鼠脂肪组织中存在干细胞来源细胞。并且细胞在体外培养条件下生物学性状保持稳定。通过外源性转化生长因子TGF－β定向分化、诱导、培养可表现出软骨细胞的部分特性。证实脂肪干细胞具有向软骨细胞定向分化的能力。

Objective To investigate the methods of isolation, cultivation of adipose derived stem cells （ADSCs） from the rats＇ adipose tissue and the feasibility of inducting differentiation to chondrocyte. Methods The adipose tissue was obtained from the wistar＇s male rats, and it was isolated, cultivated, and subcultured. The expression of CD29 in the subcultured ADSCs was detected by immunofluorescence. The 3rd generation ADSCs was cultured in the medium contained TGF-β1 , and the presence was identified by type II collagen immunohistochemistry. Results The character of ADSCs＇morphology could be observed by microscope, and it can be identified by immunofluorescence. Through the course of inducting the ADSCs to chondrocyte, the feasibility of inducting ADSCs to chondrocyte showed that type II collagen was expressed. Conclusion We demonstrated the presence of stem cells in the adipose tissue, and the stability of biologi- cal feature in vitro. The chondrocyte could be obtained from the adipose tissue through the committed differentiation, induction, cultivation by exogenous transforming growth factor （TGF-β）. The ability of the adi- pose stem cell differentiating to chondrocyte was also proved.