新生鼠和胎鼠海马神经干细胞分离和培养方法的比较

Comparison of the methods on isolation culture of neural stem cells from postnatal and embryonic rat hippocampus

目的：探索大鼠海马神经于细胞离体培养的实验条件和方法，并建立一种简便且获得率较高的实验方法。方法：分别随机选取新生1—3dsD大鼠和孕18—19dSD胎鼠，并提取海马组织，采用机械吹打结合胰蛋白酶消化的方法获得单细胞悬液，然后对培养出的细胞进行鉴定和诱导分化，最后测定并分别比较两组原代细胞获得率和增殖率以及凋亡率之间的差异。结果：来源于两种不同发育阶段大鼠的海马组织培养出的细胞均呈Nestin阳性和BrdU阳性的神经球样结构，诱导分化后的细胞高表达βⅢ-tublin和GFAP。其中，胎鼠组原代细胞获得率较高、增殖能力较强，并有统计学差异（P〈0．05）；但两组神经干细胞的凋亡率无明显差异（P〉0．05）。结论：胎鼠和新生鼠海马组织均能培养出神经干细胞，而且胎鼠组细胞产出率更高。

Objective： To establish a simple and productive methods of culturing rat hippocampal neural stem cells （NSCs） . Methods： Hippocampus of postnatal rats （P1-3） and E18-19 SD embryonic rat were taken out, and mechani- cal pipetting combined with chemical digestion were used to obtain monoplast suspension. NSCs was identified by immu- nocytochemistry and immunofluorescence from cultured cells, then differentiation of NSCs were induced. Finally prolifera- tion and apoptosis difference and cell obtained rate were measured and compared. Results： Both the primary cells from two different age of rat hippoeampus expressed Nestin immunoreactivity and BrdU immunoreactivity, and the differentiated cells expressed 13 llI-tublin and GFAP immunoreactivity. Meanwhile NSCs from embryonic rat hippocampus had better pro- liferation ability and obtained monoplast suspension more than NSCs from newborn rat hippocampus （ P 〈 0. 05 ）. But there was no apparent difference in apoptosis rate between the two groups （ P 〉 0. 05 ）. Conclusion ： The hippocampal neural stem cells could be cultured from the postnatal and embryonic rat. Embryonic rat bippocampus is suitable for large- quantity cultivation of neural stem cells because of its high output rate.