RA和SHH体外联合诱导骨髓基质干细胞转化为神经细胞

Research of BMSCs differentiating into neural cells induced by SHH and RA in vitro

目的研究体外使用全反式维甲酸(RA)和音猬因子(SHH)联合诱导人骨髓基质干细胞(BMSCs)转化为神经细胞的可行性。方法从健康志愿者髂骨处抽取骨髓5ml,体外分离、纯化、扩增到第三代后分别接种于A组[RA0.5μg/ml]、B组[SHH500ng/ml+碱性成纤维生长因子-8(FGF-8)100ng/ml]和C组[RA0.5μg/ml+SHH500ng/ml+FGF-8100ng/ml]诱导液中,诱导BMSCs分化为神经样细胞。使用倒置相差显微镜及免疫荧光技术观察诱导前后细胞改变。结果诱导7d后,各组均有部分BMSCs胞体收缩,折光性增强,突起伸出,表现出神经细胞的形态。免疫荧光表现神经元烯醇化酶(NSE)、微管相关蛋白(MAP-2)和β微管蛋白(β-tubulin)阳性。C组的各种分化细胞数均高于A组和B组,差异有统计学意义(P<0.01)。结论SHH联合RA可以在体外有效诱导人BMSCs转化为神经细胞,两者之间具有协同作用。

Objective To investigate the possibility of inducing human bone marrow stromal cells （BMSCs） into neural cells by sonic hedgehog （SHH） and all-trans-retinoic acid （RA）. Methods BMSCs were harvested from normal human marrow, then were passaged and purifed in culture medium. BMSCs were passaged three times before experiment, then purifed BMSCs were induced by SHH and/or RA with different concentrations and detected with immunocytochemisty staining for microtubule associated protein-2 （MAP-2）, neuron specific enolase （NSE） and β-tubolin. Results After 7 days of induction, some BMSCs exhibited immunoreactivity. MAP-2, NSE and β-tubulin positive cells existed in all groups, while NSE, MAP-2 and β-tubulin positive cells increased markedly in BMSCs treated with SHH and RA. There were significant differences （P 〈 0. 01 ）. Conclusion SHH plus RA can effectively induce human BMSCs to differentiate into neural cells in vitro. There is an interaction between RA and SHH.