神经干细胞球和单层贴壁神经干细胞的培养及鉴定

Journal of Materials Sciences & Technology››2011,Vol. 1››Issue (1): 51-55.

神经干细胞球和单层贴壁神经干细胞的培养及鉴定

郝军荣, 张丽, 秦正红^*

苏州大学基础医学与生物科学学院衰老与神经疾病实验室，江苏苏州 215123

出版日期:2011-02-26发布日期:2011-10-20

Cultivation and identification of neural stem cells cultivated by the methods of neurospheres and adherent monolayer cells

HAO Jun-rong, ZHANG Li, QIN Zheng-hong^*

Laboratory of Aging and Neurological Diseases，Department of Pharmacology，Suzhou University School of Medicine，Suzhou 215123，China

Online:2011-02-26Published:2011-10-20

摘要/Abstract

摘要：目的：探讨小鼠大脑皮层源性神经干细胞(NSCs)体外培养的两种方法并对培养的细胞进行鉴定。方法：分别取孕14~16d的昆明小鼠胚胎脑皮质并用细胞球悬浮培养和单层贴壁培养两种方法进行体外培养，用光学显微镜观察NSCs的生长情况，并用免疫细胞化学鉴定NSCs特异性抗原的表达，经过血清对NSCs分化诱导后进行胶质纤维酸性蛋白及微管相关蛋白-2的检测。结果：①培养出来的细胞可以扩增生长；②这两种方法分别培养的神经干细胞球和单层神经干细胞经抗巢蛋白免疫细胞染色均呈阳性；③两种培养方法培养出的NSCs经诱导分化后，免疫细胞化学染色显示微管相关蛋白-2、神经胶质酸性蛋白染色阳性。结论：采用无血清培养基中加入特定生长因子的神经干细胞球培养和单层贴壁两种培养技术，可培养出在体外稳定增殖并有多向分化潜能的NSCs。

关键词:大脑皮层,神经干细胞,神经干细胞球,单层贴壁培养

Abstract:Objective:AIM To explore two methods of culturing mouse embryo neural stem cells in vitro and identification of their differentiation．Methods:The neural stem cells were derived from the cortex tissue of mouse embryo at gestation day of 14～16 with two methods: culturing of neurosphere in floating and culturing of monolayer adherent cells. To observe the development of the neural stem cell，the expression of the stem cell marker Nestin and the neural marker MAP-2 and the glial marker GFAP was examined with immunofluorescence.Results:The cultured neural stem cells proliferated．The maintenance of stem cell property was preserved with both methods as revealed by Nestin immunofluorescence. These stem cells are able to differentiate into glial (GFAP positive) and neuronal lineage (MAP-2positive).Conclusion:With the help of growth factors and serum-free technique，the mouse neural stem cells cultivated by methods of neural stem cell spheres and adherent monolayer cells，have potential of proliferation and differentiation.

Key words:cerebral cortex,neural stem cells,neurosphere,monolayer adherent cells

郝军荣, 张丽, 秦正红. 神经干细胞球和单层贴壁神经干细胞的培养及鉴定[J]. Journal of Materials Sciences & Technology, 2011, 1(1): 51-55.

HAO Jun-Rong, ZHANG Li, QIN Zheng-Hong. Cultivation and identification of neural stem cells cultivated by the methods of neurospheres and adherent monolayer cells[J]. Journal of Materials Sciences & Technology, 2011, 1(1): 51-55.

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