大鼠骨髓间充质干细胞向神经元样细胞定向诱导分化及免疫组化鉴定

目的建立体外分离、培养大鼠骨髓间充质干细胞(MSCs)的方法,体外诱导其向神经元样细胞方向分化。方法应用细胞培养技术从大鼠股骨、胫骨中分离、纯化骨髓间充质干细胞并在体外进行培养,以形态学方法鉴定间充质干细胞,在倒置显微镜下观察细胞的形态特征,利用含10 ng/ml bFGF的LG-DMEM及含200μmol/L BHA、2%DMSO的无血清DMEM诱导其向神经元样细胞分化,并通过免疫组化方法鉴定。结果经原代及传代培养的骨髓间充质干细胞呈梭形,类似于成纤维细胞;成神经元诱导24 h后,多数MSCs变为典型的神经元样细胞,胞体向胞核收缩并有较多的长突起,免疫组化显示神经元特异性标志NSE、神经巢蛋白Nestin染色阳性。结论体外成功的进行了MSCs原代及传代培养,细胞生长稳定、增殖迅速并可多次传代,经诱导后具有向神经元样细胞分化潜能。

neuron cell formation of mesenchymal stem cells of rats by induction differentiation and immunohistochemistry identification

Objective To establish method of isolation and culture of rat bone marrow mesenchymal stem cells in vitro and to induce it differentiate to neuro cell.Methods MSCs were isolated from the femurs and tibias bones and purified and cultured in vitro,MSCs＇,quality was evaluated with morphology.States of MSCs growing in culture were observed with inverted microscope.MSCs were induced to neuro cell by LG-DMEM including 10 ng/ml bFGF and DMEM including 200μmol/L BHA、2%DMSO.They are identified by immunohistochemistry.Results The morphous of MCSs was fusiform shape and fibroblast cell-shape after cultivation of primary and subcultured.Typical neuron cell appear after 24 hours nerve cell induction,NSE and Nestin are expressed positively by immunohistochemistry staining.Conclusion The method of isolation and culture of rat bone marrow mesenchymal stem cells in vitro is ideal and MCSs have the potency of neuron cell differentiation.