大鼠神经干细胞的培养和鉴定

Identification and culture of neural stem cells from rat

目的 :自新生第 1天 (P1期 )和成年SD大鼠海马和纹状体分离培养神经干细胞。方法 :分别分离P1期和成年SD大鼠海马和纹状体组织 ,制成单细胞悬液 ,利用无血清培养技术 ,在添加碱性成纤维细胞生长因子(bFGF)、表皮生长因子 (EGF)和B2 7的DMEM F12培养液中进行培养 ,形成神经球 (neurosphere)后每 8～ 10d传代 1次 ,以免疫组织化学方法对原代和传代培养形成的神经球以及原代和传代培养的神经球分化后的细胞进行鉴定。结果 :在上述条件下培养及传代的细胞不断分裂增殖 ,形成悬浮生长的呈巢蛋白 (nestin)阳性的神经球 ;神经球贴壁后分化的细胞表现为神经元和星形胶质细胞的形态 ,且分别呈神经元特异性烯醇化酶 (NSE)阳性和胶质纤维酸性蛋白 (GFAP)阳性。结论

Objective:To culture and identify neural stem cells of hippocampus and striatum respectively from P1 rat brain and adult rat brain. Methods:Hippocampus and striatum tissue were respectively dissected out from P1 rat and adult rat brain and made into single cell suspension, which was cultured in DMEM/F12 with the presence of bFGF, EGF and B27.After neurospheres formed they were repeatedly passaged every 8 10 days. Neurospheres and differentiated cells were identified by immunocytochemisty.Results:Cells under above culture conditions continuously proliferated and formed neurospheres which were nestin positive. Differentiated cells took on the shapes of neuron and astrocyte cell and were NSE and GFAP positive respectively. Conclusion:There are neural stem cells in hippocampus and striatum of P1 and adult rat.