[大鼠胎脑神经干细胞的分离培养及电穿孔转染]
[The isolating culture of neural stem cells from rat's fetus brain and transfection by electroporation].
作者信息
Liu Chun-jiang, Yin Fei, Zheng Xiang-rong, Zhang Shan-shan, Tan Jie-lu
机构信息
Department of Pediatrics, Xiangya Hospital, Central South University, Changsha, China.
出版信息
Zhonghua Yi Xue Za Zhi. 2009 Nov 17;89(42):3007-9.
OBJECTIVE
To isolate and culture the neural stem cells (NSC) from rat's fetus brain, to study the transfection efficacy of NSC using electroporation.
METHODS
We isolated, cultured and amplified NSC from the brain of SD fetal rats. NSC and differentiated cells were identified using immunofluorescent histochemical methods. Using green fluorescence protein (GFP) as the marker, pEGFP-N1 was transfected into NSC using electroporation, and the transfection rate was calculated by counting the NSCs with green fluorescent.
RESULTS
The cells isolated from brain tissue of fetal rats can proliferate for long time, both primary and passage culture of NSC can express specific antigen of NSC-Nestin, and after induced differentiation, the cells can express specific antigen of astrocytes-Glial fibrillary acidic protein (GFAP) and specific antigen of neurons-Neuron-specific enolase (NSE). The transfection rates of electroporation were 17.9% - 69.1%, the average was 30.5%.
CONCLUSION
Isolated NSC which had the features of self-proliferation and differentiation potential from brain tissue of SD fetal rats, confirmed that electroporation was an efficient transfection method for NSC, provided experimental basis for gene therapy in the future.
目的
从大鼠胎脑分离培养神经干细胞（NSC），研究电穿孔法对NSC的转染效率。
方法
从SD胎鼠脑内分离、培养并扩增NSC。采用免疫荧光组织化学方法鉴定NSC及其分化细胞。以绿色荧光蛋白（GFP）为标记，用电穿孔法将pEGFP-N1转染入NSC，通过计数发绿色荧光的NSC计算转染率。
结果
从胎鼠脑组织分离的细胞能长期增殖，NSC原代及传代培养均能表达NSC特异性抗原巢蛋白（Nestin），诱导分化后细胞能表达星形胶质细胞特异性抗原胶质纤维酸性蛋白（GFAP）和神经元特异性抗原神经元特异性烯醇化酶（NSE）。电穿孔法转染率为17.9% - 69.1%，平均为30.5%。
结论
从SD胎鼠脑组织分离出具有自我增殖和分化潜能特性的NSC，证实电穿孔法是NSC的一种高效转染方法，为今后的基因治疗提供了实验依据。
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