丹参诱导大鼠骨髓间充质干细胞向神经样细胞分化

目的:探讨骨髓间充质干细胞(marrow stromal stem cells,MSCs)向神经细胞分化中神经蛋白分子及相关基因的表达情况。方法:分离提取大鼠的MSCs,体外培养扩增。用含丹参的无血清的L-DMEM培养基进行诱导,并以不含丹参的无血清L-DMEM培养基作为对照。提取两组的MSCs总RNA,RT-PCR检测ngn-1、mash-1的表达。结果:丹参诱导90 min后,细胞发生了明显的形态学变化,大多数细胞转变为类似双极或多极神经元样形态,伸出轴突或树突样突起。免疫细胞化学显示诱导后的MSCs表现为nestin、NSE、GFAP染色阳性,对照组为阴性。未经诱导的MSCs ngn-1,mash-1 mRNA为阴性,诱导后有表达。结论:丹参可诱导大鼠MSCs向神经前体细胞和神经细胞分化。MSCs向神经元样的分化可能与ngn-1,mash-1有关。

Differentiation of rat bone marrow stromal stem cells into neuron-like cells induced by salvia mitiorrhiza

Objective:To investigate the expression of the neural markers and the gene in differentiation of SD rat bone marrow derived mesenchymal stem cells (MSCs)into neuron-like cells induced by salvia mitiorrhiza.Methods:The nu- cleated cells were isolated from male Sprague-Dawley rat's bone marrow through gradient eentrifugation and cultured, and the unadherent cells were removed to purify MSCs.Salvia mitiorrhiza was used as inducer to differentiate MSCs, and then morphologic changes of differentiated cells were observed and identified.The nestin,neuron specific enolase (NSE) and glial fibrillary acidic protein (GFAP) were detected by immunoeytoehemical staining.Results:The primary cells grew well in adherent way when cultured in L-DMEM containing 10% fetal calf serum (FCS),and were in fusi- form shape,forming cell clones;subeultured cells,which were fibroblast-like,had higher reproductive activity than the primary cells.Morphology of MSCs changed when cultured with salvia mitiorrhiza.Initially,cytoplasm in the flat MSCs retracted towards the nucleus,forming a similar bipolar or multipolar structures,and cell bodies became increas- ingly spherical and refractile,exhibiting a typical neuronal perikaryal appearance.Processes continued to elaborate, displaying primary and secondary branches.The differentiated cells expressed nestin,NSE and GFAP at 90 min, which was negative in the control group.The expression of mash-1 and ngn-1 without induction was negative and could be found after induction.Conclusion:MSCs can differentiate into neuron-like cells while in the existance of salvia mitiorrhi- za,and mash-1 and ngn-1 may play an important role in the MSCs differentiation into neuron-like cells.