细胞因子联合纹状体条件培养液定向诱导间充质干细胞体外分化为多巴胺能神经元的研究(英文)  

Cytokines Combined with Striatal Conditioned Medium Promote Directional Differentiation of Mesenchymal Stem Cells into Dopaminergic Neurons in Vitro

作　　者：李府[1] 马丽霞[1] 张乐玲[1] 郑立波[1] 陈颖杰[2] 吴镇[2] 王世富[1] 

机构地区：[1]山东大学齐鲁儿童医院儿科研究所,济南250022 [2]山东大学第二医院血液学实验室,济南250033

出　　处：《实用儿科临床杂志》2009年第24期1922-1925,共4页Journal of Applied Clinical Pediatrics

摘　　要：目的探讨联合应用碱性成纤维细胞生长因子(bFGF)、表皮生长因子(EGF)和纹状体条件培养液定向诱导大鼠骨髓间充质干细胞(BMMSCs)分化为多巴胺能神经元的可能性。方法1.BMMSCs的分离培养:分离纯化健康成年Wistar大鼠BMMSCs进行传代培养。2.纹状体条件培养液的制备:出生24h内的新生Wistar大鼠,分离纹状体组织用于制备纹状体条件培养液。3.BMMSCs的诱导分化:取体外培养的第5代间充质干细胞,用含bFGF和EGF的预诱导液进行预诱导,24h后去除预诱导液,换用纹状体条件培养液进行诱导。4.倒置显微镜下观察细胞形态变化,并用细胞免疫化学技术检测细胞内神经元特异烯醇化酶(NSE)和酪氨酸羟化酶(TH)的表达。结果大鼠BMMSCs经bFGF、EGF和纹状体条件培养液诱导后细胞胞体逐渐回缩成团,形成梭形,部分细胞可见突起伸出,类似神经元。细胞免疫化学检测,联合诱导后NSE阳性表达为(72.70±14.81)%,TH阳性表达为(34.50±15.93)%。结论bFGF、EGF联合纹状体条件培养液体外可定向诱导大鼠BMMSCs分化为多巴胺能神经元。Objective To explore the possibility of basic fibroblast growth factor（bFGF）and epidermal growth factor（EGF）combined with st-riatal conditioned medium promoting the directional differentiation of rat bone marrow mesenchymal stem cells（BMMSCs）into dopaminergic neurons.Methods 1.Separation and culture of BMMSCs：BMMSCs were harvested from healthy adult Wistar rats for serial subcultivation.2.Preparation of st-riatal conditioned medium：newborn Wistar rats within 24 hours were selected,and their brain tissues were removed to prepare striatal conditioned me-dium.3.Induced differentiation of BMMSCs：the 5th passage BMMSCs were collected and pre-induced in low glucose-Dulbecco＇s modified eagle medium（L-DMEM）containing bFGF and EGF.Twenty-four hours later,pre-induction liquor was replaced with striatal conditioned medium for further induced differentiation.4.Result assessment：the morphological changes of stem cells were observed under inverted phase microscope.The expression of neuron specific enolase（NSE）and tyrosine hydroxylase（TH）were identified by immunocytochemical technique.Results The cell body of rat BMMSCs contracted into round and spindle shape after induction by bFGF and EGF combined with striatal conditioned medium.Partial neuron-like cells with prominence could be found.Immunocytochemical detection showed that the percentages of NSE and TH positive cells were（72.70±14.81）% and（34.50±15.93）%,respectively.Conclusion BMMSCs can be induced directionally into dopaminergic neurons by bFGF and EGF combined with striatal conditioned medium in vitro.

关 键 词：间充质干细胞 多巴胺能神经元 条件培养液 纹状体 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]