AngⅡ诱导神经干细胞向多巴胺能神经元分化的时间效应

Time effect of angiotensinⅡon differentiation into dopaminergic neurons from neural stem cells in vitro

目的:体外分离培养神经干细胞(NSCs),探讨血管紧张素II(AngII)诱导NSCs向多巴胺(DA)能神经元分化的时间效应。方法:取第二代NSCs在含有AngII的诱导分化液中培养,根据诱导持续时间不同,分为A(10d),B(14d),C(17d)和D(21d)四组。终止诱导后,用免疫学法检测分化细胞中DA能神经元标志物酪氨酸羟化酶(TH)的表达,半定量-PCR测定THmRNA的相对表达量。结果:在AngII作用下,B,C组分化细胞数量均比A组增多,细胞胞体饱满,突起粗长,而D组细胞部分开始老化、脱壁悬浮。其中,C组TH阳性细胞率最高(15.64%),且THmRNA相对表达量(0.5047±0.0212)明显高于其他三组(P<0.05)。结论:AngII诱导NSCs分化成DA能神经元存在分化的时间效应,诱导持续达17d时分化细胞中DA能神经元的比例最高。

Objcective：To investigate the time effect of angiotensin II（Ang II）induced on the differentiation of NSCs to DA neurons in vitro.Methods：The second generation NSCs were underwent in medium with Ang II.Then according to the culture days,NSCs were divided into four groups：group A（incubated 10days）,group B（incubated 14days）,group C（incubated 17days）,group D（incubated 21days）.Tyrosine hydroxylase（TH）which is the specific antigen of DA neurons was detected by immunocytochemistry after the treatment;the relative expression level of TH gene was examined by semi-quantity RT-PCR（SQ-PCR）too.Results：In differentiate solution,the number of differentiated cells in group B and C were more than in group A,the cells were full body and had long synapses,but the cells in group D became aging and suspended from the dish.Immunocytochemistry and SQ-PCR studies revealed that group C showed more immunoractive TH expression than other groups（P〈0.05）,the proportions of TH-positive cell was 15.64%,the relative expression level of TH gene was 0.5047±0.0212.Conclusion：The present results suggest that induction time influences the differentiation of NSCs towards DA neurons by Ang II in vitro.The proportion of DA neurons was the highest at 17 days when NSCs were induced by Ang II.