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外源性Ang Ⅱ诱导神经干细胞向多巴胺能神经元分化的研究
引用本文:沈岳飞,罗彦妮,范瑞芳,罗小丹,张维雄,陈梅玲.外源性Ang Ⅱ诱导神经干细胞向多巴胺能神经元分化的研究[J].神经疾病与精神卫生,2010,10(2):129-133.
作者姓名:沈岳飞  罗彦妮  范瑞芳  罗小丹  张维雄  陈梅玲
作者单位:1. 广西医科大学第一附属医院神经内科,530021
2. 桂林医学院附属医院神经内科
摘    要:目的在体外成功分离培养并扩增神经干细胞(NSCs)的基础上,研究外源性血管紧张素Ⅱ(Ang Ⅱ)对NSCs向多巴胺(DA)能神经元分化的影响。方法(1)分离培养新生1d SD大鼠脑组织NSCs,免疫细胞学方法测定NSCs特异性标志物神经上皮干细胞蛋白(Nestin)表达及其分化为神经元、神经胶质细胞的能力;(2)按培养液中Ang Ⅱ的浓度不同,分5个浓度(100、200、400、600、800nmol/L)对第二代NSCs进行诱导分化。10d后采用免疫细胞学方法检测DA能神经元标志物酪氨酸羟化酶(TH)和神经胶质细胞标志物胶质纤维酸性蛋白(GFAP)的表达,半定量RT-PCR(SQ-PCR)测定分化细胞中TH mRNA的相对表达量。结果外源性Ang Ⅱ诱导提高NSCs向TH阳性细胞分化的比率,TH mRNA相对表达量亦增加,以Ang Ⅱ浓度为400nmol/L及600nmol/L的诱导效果最明显,TH阳性细胞率分别为10.77%和11.34%,TH mRNA相对表达量分别为(0.4023±0.0515)和(0.3971±0.0319),两组间比较无统计学意义(P〉0.05);其中400nmol/L Ang Ⅱ组分化细胞中GFAP阳性细胞率高于对照组,有统计学意义(P〈0.05)。结论外源性AngⅡ促进NSCs向DA能神经元分化,在400~600nmol/L浓度范围内AngⅡ的诱导效能更显著;AngⅡ促进NSCs向DA能神经元分化的机制可能与同时促进星型胶质细胞(AS)分化有关。

关 键 词:神经干细胞  分化  多巴胺能神经元  血管紧张素Ⅱ

Effects of exogenous Angiotensin II on differentiation of neural stem cells into dopaminergic neurons
Institution:SHEN Yue-fei, LUO Yan-ni, FAN Rui-fang, et al. Department of Neurology, The First Affili- ated Hospital of Guangxi Medical University, Nanning 530021 , China
Abstract:Objective To investigate the effects of exogenous angiotensin Ⅱ (Ang Ⅱ) on the dif- ferentiation of neural stem cells (NSCs) into dopaminergic (DA) neurons in vitro. Methods (1)The pri- mary NSCs isolated from the whole brain of newborn (within 24h after birth) Sprague Dawley rats were cultured in vitro, the expression of neuroepithelial stem cell protein (Nestin), neuron special enolase (NSE), glial fibrillary acidic protein (GFAP) and cyclic necleotide phosphohydrolase(CNP) was determined by immunohistochemistry. (2)The second generation of NSCs were incubated in medium with dif- ferent doses of exogenous Ang Ⅱ (100--800 nmol/L) for 10 days. After the treatment, tyrosine hydrox- ylase (TH) which is the specific antigen of DA neurons and GFAP which is the speeifc antigen of Astro- cyte (AS) were detected by immunocytochemistry, and the mRNA levels of TH was detected by semi-- quantity RT PCR (SQ--PCR). Results Exogenous Ang II could increase the proportion of NSCs dif- ferentiated into TH--positive cell, both 400 nmol/L and 600 nmol/L Ang II showed elevated expression of TH compared to other Ang II groups, the proportions of TH- positive cell were i0. 77% and 11.34% respectively, the relative expression levels of TH gene were (0. 402 3±0. 051 5) and (0. 397 1± 0. 031 9) respectively; 400 nmot/L Ang Ⅱ displayed an upregulation in GFAP expression compared to control group (P 〈 0.05). Conclusions Exogenous Ang II could promote the differentiaion of NSCs into DA neurons in vitro, the doses of AngⅡ ranged from 400 to 600 nmol/Lmay be more effective on differentiation. The mechanism involved in the Ang Ⅱ--inducible differentiation of NSCs into DA neurons may be associated with promoting the differentiation of astrocytes (AS) simultaneously.
Keywords:Neural stem cells  Differentiation  Dopaminergic neurons  Angiotensin Ⅱ
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