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背景:目前关于骨髓间质干细胞能否向神经元方向分化的报道不多,且争论多集中在分化后的神经元是否仅具有神经元形态而不具有神经元功能。目的:探讨海马神经元条件培养液诱导大鼠骨髓间质干细胞向神经元样细胞和神经胶质样细胞分化的可能性。方法:将第5代大鼠骨髓间质干细胞分为4组:条件培养基组加入海马神经元和胶质细胞的培养液;b-FGF组加入含b-FGF的DMEM培养基;无血清培养组加入含Neurobasal和B27的无血清培养基;阴性对照组加入含胎牛血清的DMEM。各组诱导12,24h后,应用免疫细胞化学染色行神经元特异性烯醇化酶、微管相关蛋白2、胶质纤维酸性蛋白的鉴定,Western-blot法检测细胞神经元特异性烯醇化酶、微管相关蛋白2和胶质纤维酸性蛋白的表达。结果与结论:诱导12,24h后,条件培养基组、b-FGF组、无血清培养组骨髓间质充干细胞微管相关蛋白2、胶质纤维酸性蛋白、神经元特异性烯醇化酶均呈阳性表达,阴性对照组未见表达。与阴性对照组比较,诱导后24h,条件培养基组、b-FGF组、无血清培养组微管相关蛋白2表达均明显增强(P<0.05),且条件培养基组增强幅度显著高于另两组(P<0.05);条件培养基组、b-FGF组、无血清培养组神经元特异性烯醇化酶及胶质纤维酸性蛋白表达无明显差异。结果证实海马神经元条件培养液可体外诱导大鼠骨髓间质干细胞分化为神经元样细胞和神经胶质样细胞,与含b-FGF的培养基和无血清培养基相比,海马神经元条件培养基诱导的神经元和神经胶质细胞阳性率最高。
BACKGROUND: Currently, there are few reports on whether bone marrow mesenchymal stem cells can differentiate into neurons, and the controversy focuses more on whether the differentiated neurons have only neuronal morphology but not neuronal function. OBJECTIVE: To investigate the potential of hippocampal neuron conditioned medium in inducing the differentiation of rat bone marrow mesenchymal stem cells into neuron-like cells and glial-like cells. Methods: The 5th generation rat bone marrow mesenchymal stem cells were divided into 4 groups: conditioned medium group was added into the culture medium of hippocampal neurons and glial cells; b-FGF group was added DMEM medium containing b-FGF; serum-free culture Groups were fed with serum-free medium containing Neurobasal and B27; negative controls were added with DMEM containing fetal bovine serum. Immunocytochemical staining was used to identify neuron-specific enolase, microtubule-associated protein 2 and glial fibrillary acidic protein in each group after 12 and 24 hours of induction. Cell-specific enolase, Expression of microtubule-associated protein 2 and glial fibrillary acidic protein. RESULTS AND CONCLUSION: After induced for 12,24 h, microtubule-associated protein 2, glial fibrillary acidic protein and neuron-specific enolase in bone marrow stromal cells were significantly increased in conditioned medium group, b-FGF group and serum-free medium Positive expression, negative control group no expression. Compared with the negative control group, the expression of microtubule-associated protein 2 in the conditioned medium group, the b-FGF group and the serum-free culture group increased significantly (P <0.05) 24 h after induction, and the enhancement extent of the conditioned medium group was significantly higher than that of the negative control group (P <0.05). There was no significant difference in neuron-specific enolase and glial fibrillary acidic protein expression between the conditioned medium group, the b-FGF group and the serum-free culture group. The results confirmed that conditioned medium of hippocampal neurons can induce the differentiation of rat bone marrow mesenchymal stem cells into neuron-like cells and glial-like cells in vitro. Compared with b-FGF-containing medium and serum-free medium, hippocampal neuron conditions The highest positive rate of neurons and glial cells was induced by culture medium.