大鼠嗅鞘细胞培养和纯化方法

Method of and on Purification of Rat Olfactory Ensheasing Cells

目的:建立一种可行的培养和纯化嗅鞘细胞的方法。方法:利用显微外科技术,从新生大鼠脑中取出嗅球的嗅神经组织,去除脑膜和微血管,消化后细胞接种在含有10%的DMEM/F12(1:1)培养液中进行体外培养,利用阿糖胞苷Arac和Thy1.1单抗去除成纤维细胞,胰酶传代时通过仔细的监测将星形胶质细胞去掉,通过纯化获得大量的嗅鞘细胞。结果:嗅鞘细胞较容易培养,其增殖速度较快,污染的星形胶质细胞和成纤维细胞可有效地被去除,显微镜下嗅鞘细胞有卵圆形的胞体和长的、细的突触,为双级或多级的细胞,透射电镜显示细胞具有锯齿状的核,伴有大块染色体和电子密度的胞浆及散在的纤维,形态学和超微结构显示已获得了较纯的细胞。结论:所建立的培养方法可行,适宜从新生大鼠中获得嗅鞘细胞。

Objective：To set up an applicable method of culturing the ensheasing cells （ECs）. Methods： The olfactory nerve layer of olfactory bulbs was removed from the neonatal rat head using microsurgical techniques. The meningeal coverings and minute capillaries was pialed. The cells were seeded into DMEM/F12 （1：1） containing 10% fetal bovine serum. Cytosine arabi- noside （Ara-c） and antiThy1.1 monoclonal antibody were used to remove fibroblasts. The astrocytes were separated from the ECs through careful monitoring of trypsination. Results：The ensheasing cells could be cultured easily, contaminating cells such as fibroblasts and astrocytes were eliminated from the cultures effectively. The cells were bipolar or multipolar, transmission e lectron microscopy revealed the ECs possessed an irregular indented nucleous with pachy chromatin. Ultrastructure analysis indicated the cells were ensheasting cells. Conclusion：The culturing method is applicable and is fit for isolating ensheasing cells from neonatal rats.