大鼠骨髓基质干细胞体外分化为神经元样细胞并表达肠神经相关神经营养因子

目的探讨维甲酸、锌联合胎肠培养基诱导大鼠骨髓基质干细胞向神经元细胞分化的可能性以及分化前后肠神经相关神经营养因子表达的变化。方法体外培养大鼠骨髓基质干细胞（BMSC），传代至第6代，进行神经诱导分化。先以bFGF（10ng／ml）预诱导24h，然后以维甲酸（Retinoic acid，RA）、锌在胎肠培养基（FGCM）条件下诱导10d。免疫荧光法检测神经元标志神经特异性烯醇化酶（NSE）和神经丝蛋白（NF）的表达。RT-PCR法检测胶质源神经营养因子（GDNF）和神经生长因子（NGF）mRNA的表达。结果流式细胞检测BMSC高表达CD90（99，7％），而CD45表达阴性。诱导10d后，部分细胞在形态上表现出神经元样改变，免疫荧光染色神经元标志NSE和NF阳性，而胶质细胞标志GFAP阴性。RT-PCR检测发现，BMSC诱导前低表达神经营养因子NGF和GDNF mRNA，而诱导后表达水平显著增加。结论维甲酸、锌联合胎肠培养基不仅能在体外诱导BMSC分化为神经元样细胞，而且能促进肠神经相关神经营养因子表达显著上调。

Rat bone marrow stromal cells differentiate into neurons and express enteric nerve related neurotrophic factors invitro

Objective To investigate the possibility of the differentiation from bone marrow stromal cells(BMSC) induced by retinoic acid,Zn and fetal gut condition medium into neurons and the expression of enteric nerve related neurotrophic factors,glial cell line-derived neurotrophic factor(GDNF) and nerve growth factor(NGF) in vitro.Methods BMSC were harvested from male rats and cultured in DMEM supplemented with 20% fetal bovine serum,and characterized by flow cytometry. At passage 6,BMSC were pre-induced by bFGF (10ng/ml) for 24h,then induced with retinoic acid,Zn in fetal gut condition medium (FGCM) for 10 days. The expressions of neuronal markers NSE (neural specific enolase),NF(neurofilament),and glial cell marker GFAP (glial fibrillary acidic protein) were detected by fluorescent immunohistochemistry method. The mRNA expressions of GDNF and NGF were detected by RT-PCR method. Results The cultured BMSCs were CD90 (99.7%) positive and CD45 negative on flow cytometry. After 10 days of induction,a certain number of cells adopted neuron-like morphological changes and showed the expressions of NSE and NF without the expression of GFAP by fluorescent immunohistochemistry. BMSC expressed low level NGF and GDNF mRNA. After induction of ratinoic acid,Zn in FGCM,the expression of NGF and GDNF mRNA were highly increased.Conclusion Retinoic acid,Zn and fetal gut condition medium can not only induce neuronal differentiation of BMSC,but also increase the expression of NGF and GDNF significantly.