体外诱导成人骨髓间充质干细胞转化为神经元细胞的实验研究

目的探讨成人骨髓间充质干细胞(hMSCs)在体外特定的诱导条件下,转化为神经元细胞的规律、效率及长期存活的条件.方法采用β-巯基乙醇为诱导剂,在体外诱导6 h后,改用诱导维持液使诱导后的细胞在诱导维持液中存活6 d.用细胞化学及免疫组织化学、Western blot方法检测神经元细胞、星形胶质细胞标记蛋白的表达.结果诱导6 h,诱导后的细胞尼氏染色胞浆中可见深蓝色的尼氏体形成;细胞表达NSE、NF-M,而不表达GFAP;NSE、NF-M阳性细胞数超过80%以上.Western blot结果显示:诱导6 h,诱导后的细胞表达Nestin,而无MAP-2的表达,随着时间的延长,Nestin的表达逐渐减少,而出现了MAP-2的表达.结论β-巯基乙醇在体外可定向诱导hMSCs经神经干细胞转化为神经元细胞,而且诱导后的神经元细胞逐渐趋于成熟.

Experimental study on neuro-differentiation of human bone marrow mesenchymal stem cells in vitro

Objective To explore the regularity, efficiency, requirements for long-term survival of human bone marrow mesenchymal stem cells (hMSCs) transforming to neuron induced by 2-mercaptoethanol (2-ME) in vitro. Methods At 6 hours after hMSCs induced in vitro by 2-ME in serum-free medium, induction media were removed and transferred to neuronal induction maintenance media for 6 days. The expressions of specific markers of neuron or astrocyte were evaluated by cytochemistry, immunohistochemistry staining and Western blot method. Results After induction for 6 hours, hMSCs displayed neuronal morphologies such as pyramidal cell bodies. The induced hMSCs expressed neuro-specific marker such as neuro-specific enolase (NSE) and neurofilament (NF), but could not express glia fiber acid protein (GFAP). The number of positive cells of NSE and NF-M was more than 80%. The results of Western blot showed that nestin was detectable at 6h, and expression levels of hMSCs-derived neurons decreased with time by Western blot. With the times, there was no detectable nestin expression in any hMSCs-derived neurons, but MAP-2, a marker of mature neuron was detectable. Conclusions hMSCs could differentiate into nerve stem cells in vitro, and then induced neurocyte could be gradually mature.