多种细胞因子诱导大鼠间充质干细胞向多巴胺能神经元分化  

Some cytokines induce rat bone marrow mesenchymal stem cells to differentiate into dopaminergic neurons in vitro

作　　者：陈丹丹[1] 付文玉[1] 庄文欣[2] 吕娥[1] 郭健[3] 李锋杰[2] 

机构地区：[1]潍坊医学院组织学与胚胎学教研室,潍坊261053 [2]潍坊医学院医学研究实验中心,潍坊261053 [3]潍坊医学院药学实验室,潍坊261053

出　　处：《解剖学杂志》2011年第5期584-587,共4页Chinese Journal of Anatomy

基　　金：山东省自然科学基金（Y2003COl;Y2008c129）;留学回国人员科研启动基金（教外司留[20091100l号）

摘　　要：探索音速波状蛋白（sHH）、成纤维细胞生长因子-8（FGF8）、碱性成纤维细胞生长因子（bFGF）、全反式维甲酸（ATRA）和胶质细胞源性神经营养因子（GDNF）多种因子联合诱导大鼠骨髓间充质干细胞（BMSCs）向多巴胺（DA）能神经元分化的可行性及分化后细胞功能的改变。方法：体外分离、扩增和鉴定大鼠BMSCs；分别采用bFGF＋ATRA＋GDNF和bFGF＋SHH＋FGF8细胞因子组合进行诱导，免疫细胞化学显色方法鉴定2组神经元样细胞，计数和比较2组阳性细胞阳性率；荧光分析法测定诱导后细胞培养上清液中儿茶酚胺含量。结果：经过诱导大鼠BMSCs能够分化为神经元样细胞。免疫细胞化学显色呈神经巢蛋白、微管相关蛋白-Ⅱ、酪氨酸羟化酶（TH）阳性；且bFGF＋SHH＋FGF8组诱导的神经元样细胞阳性率高于bFGF＋ATRA＋GDNF组。2组细胞上清液中儿茶酚胺类物质含量分别为（2．731±0．81）μg／L和（2．977±0．16）μg／L。结论：bFGF＋SHH＋FGF8组合能够更有效地诱导大鼠BMSCs分化为DA能神经元样细胞。Objective： To explore the feasibility that various cytokines, including sonic hedgehog （SHH）, basic fibroblast growth factor （bFGF）, all-trans retinoic acid （ATRA）, glial derived neurotrophic factor （GDNF） induce the rat bone marrow mesenchymal stem cells into dopaminergic neurons, and the functions of induced cells. Methods： The BMSCs derived from rat bone marrow were cultured and maintained in vitro. After being induced by basic fibroblast growth factor （bFGF） for 24 h, some BMSCs were incubated with all-trans retinoic acid （ATRA） and glial-derived neurotrophic factor （GDNF） for 4 d, and some BMSCs were incubated with sonic hedgehog （SHH） and fibroblast growth factor-8 （FGF-8） for 12 d. Then the expressions of nestin, microtubule-associated protein-Ⅱ （MAP-Ⅱ） and tyrosine hydroxylase （TH） were detected by immunocytoehemical methods and the content of catecholamine （CA） in supernatant fluid were determined by fluorescent analysis, respectively. Results： Under the inverted phase contrast microscope, the induced BMSCs by two induced methods showed typical neuronal morphological characteristics after induction. The expression rates of nestin, MAP-Ⅱ and TH were （48.1±3.2）%, （63. 2±2. 9）%0, （30. 74±5.9）% induced by BFGF＋ATRA＋GDNF and （54.7±4. 1）%, （75.8±5.2）%, （46. 7±3.3）% and by bFGF＋SHH＋FGF8, respectively. The expression rate of TH of the cells induced by bFGF＋SHH ＋FGF8 was higher than that of the ceils induced by bFGF＋ATRA＋GDNF （P〈0.01）. The content of CA in supernatant fluid was （2. 731±0.81）/lg/L and （2. 977±0. 16）〉g/L, respectively induced by different cytokines. Conclusion： bFGF＋ SHH＋FGF8 eytokines group could induce rat BMSCs to differentiate into dopaminergic neuron-like cells more effectively.

关 键 词：间充质干细胞 神经元 细胞分化 细胞因子 大鼠 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]