Wnt信号分子诱导骨髓间充质干细胞向神经元样细胞分化

Wnt signaling molecules induce the differentiation of bone marrow mesenchymal stem cells into neuron-like cells

背景:间充质干细胞可向神经方向转化,但分子机制目前不清楚。目的:观察Wnt3a和Wnt5a在骨髓间充质干细胞向神经细胞分化过程中的作用。方法:体外分离培养大鼠骨髓间充质干细胞,传代后通过形态学和流式细胞仪检测细胞表面标志物CD14,CD44,CD9,CD34,CD45表达。采用碱性成纤维细胞生长因子分别联合Wnt3a和Wnt5a的诱导方案,免疫组织化学法和RT-PCR检测Wnt3a、Wnt5a在骨髓间充质干细胞向神经样细胞分化过程中的作用。结果与结论:骨髓间充质干细胞为长梭形,表面标志物CD9,CD44高表达,CD34,CD45低表达。诱导后细胞Wnt3a诱导组巢蛋白,神经元特异性烯醇化酶呈阳性,而胶质纤维酸性蛋白无明显表达,细胞活力良好。Wnt5a诱导组巢蛋白呈弱阳性表达,而神经元特异性烯醇化酶及胶质纤维酸性蛋白阴性。RT-PCR结果显示Wnt3a诱导组巢蛋白在诱导前后均有表达,神经元特异性烯醇化酶在诱导后5d可见明显的扩增条带,10d后更加明显。胶质纤维酸性蛋白在诱导10d后出现较弱的扩增条带。提示Wnt3a分子能够促进体外培养的间充质干细胞向神经元样细胞分化。

BACKGROUND：Mesenchymal stem cells（MSCs） could differentiate into nerve cells,but the molecular mechanism is still unknown.OBJECTIVE：To observe effects of Wnt3a and Wnt5a on differentiation of bone marrow mesenchymal stem cells（BMSCs） into nerve cells.METHODS：The rat BMSCs were separated and cultured in vitro.Following passage,expression of CD14,CD44,CD9,CD34 and CD45 was detected by morphology and flow cytometry.Wnt3a,Wnt5a contained with basic fibroblast growth factor（bFGF） were used to induce the differentiation of BMSCs into neuron-like cells.Wnt3a and Wnt5a effects were identified by immunohistochemistry and RT-PCR.RESULTS AND CONCLUSION：BMSCs were fibroblast-like.CD9 and CD44 were highly expressed,but CD34 and CD45 low expressed.Nestin and neuron specific enolase（NSE） were positive and glial fibrillary acidic protein（GFAP） was not positive when they was cultured with Wnt3a.In Wnt5a group,Nestin was weak positive,but NSE and GFAP were negative.The RT-PCR results revealed Nestin expressed before and after induction;NSE was significant at 5 days following induction,especially at 10 days;weak GFAP expression was detectable at 10 days in Wnt3a group.These indicated that Wnt3a can promote the differentiation of BMSCs into neuron-like cells.