糖尿病患者脂肪间充质干细胞的分离培养与鉴定

Culture and identification of adipose mesenchymal stem cells from diabetic patients in vitro

目的探讨体外分离培养糖尿病患者脂肪间充质干细胞(ADSCs)的方法并分析其生物学特性。方法采用胶原酶消化法从糖尿病患者的脂肪组织中分离出ADSCs。对ADSCs进行体外培养,倒置相差显微镜下观察细胞形态;绘制第3、6、8代传代细胞的生长曲线;流式细胞仪检测第3代细胞表面标记物(CD34、CD45、CD29、CD44、CD105、CD90);通过成脂诱导实验鉴定细胞的分化潜能(油红染色阳性判定为脂肪细胞,证实细胞可向脂肪细胞分化)。结果分离出的ADSCs,能稳定传代;其细胞表面标记物CD34、CD45呈阴性表达,CD29、CD90、CD44、CD105呈阳性表达。成脂诱导后油红染色阳性。结论在糖尿病患者脂肪组织中成功分离到ADSCs,在体外适宜条件下ADSCs可向脂肪细胞分化,具有分化潜能。

Objective To establish the isolation and culture method of adipose mesenchymal stem cells derived from diabetic patients in vitro,so as to observe biological properties. Methods The ADSCs were isolated by digestion with collagenase from adipose tissue of diabetic patients,then cultured and purified in vitro. The cells were observed by inverted microscopeand. The P3,P6,P8 generation cells were collected and analyzed the cell growth curve. Then the cell surface markers and karyotype analysis of the P3 generation cells were detected. Their differentiation potential was proved by adipogenic differentiation. Oil red staining positive proved they were fat cells. Results After isolation and culture,we obtained the ADSCs,which could passage stably and proliferate highly. The flow cytometry examination showed that the expression of CD34 and CD45were negative,CD29,CD90,CD44 and CD105were positive. Oil red staining was positive. Conclusion The ADSCs could be isolated from diabetic patients adipose tissue and be functionally induced into adipocytes in the presence of appropriate conditioned media.