兔胚胎神经干细胞的分离、培养和鉴别

目的：研究兔胎脑神经干细胞体外生长特性，为探讨神经干细胞的临床应用及神经系统的发育奠定基础。方法：采用含碱性成纤维细胞生长因子（bFGF）和表皮细胞生长因子（EGF）的N2无血清培养技术，取18天龄兔胚胎脑组织，分离神经干细胞，并观察分离的细胞体外培养、增殖、分化潜能，免疫组化鉴定。结果：从18天龄兔胎脑皮质和纹状体中成功分离出具有自我更新和多分化潜能的神经干细胞，在无血清培养时细胞呈半贴壁状态生长，形成神经球，可传代。细胞呈Nestin免疫反应阳性；在含血清培养基中培养时则分化，分化后的细胞表达神经元细胞、星形胶质细胞和少突胶质细胞的特异性抗原。结论：来自兔胎脑神经干细胞能在体外培养、增殖并保持传代能力。无血清N2EGF、bFGF培养基有利于兔胎脑神经干细胞的存活和增殖，含血清培养基能诱导兔胎脑神经干细胞分化。

Isolation, Culturing and Identification of Neural Stem Cells from Embryonic Rabbit

Objective To investigate the growth characterization, differentiation and the cultivation of neural stem cells from rabbit embryonic cerebral in vitro for related research. Method Neural stem cells (NSCs) were isolated from embryonic brain of rabbit about 18d, collected and cultured with serum free N2 medium containing basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Cultured and differentiated cells were identified with immunocytochemistry staining. The apoptosis cells' percentage was measared by cytorheological methodResults Rabbit neural stem cells having the abilities of reproduction and multipotency were successfully isolated at embryonic age 18d from rabbit fetal cerebral and cultured from rabbit fetal striatum with serum free medium. They formed typical neuraospheres in suspension. The cells could be passed continuously, expressing Nestin antigen. Serum medium induced neural stem cells to differentiate and express specific antigens of neuron, astrocyte and oligodendrocyte. Conclusion The results suggest that cortex remain to be cultured in vitro, remain to be reprodcted and passed; Neural stem cells at embryonic from rabbit fetal cerebral can be growth and reproduction on cultured with serum free N2 medium containing basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF), and were capable of continuous passage, which have the abilities of multipotencypotential. on cultured with serum free N2 medium.