大鼠胚胎脑皮层神经干细胞的分离和培养

目的探讨分离、培养、纯化大鼠胚胎神经干细胞(neuralstem cell,NSCs)的最佳条件,以获得充足的神经干细胞来源,用于中枢神经系统疾病的治疗.方法分离孕13～15 d胎鼠大脑皮层,在无血清含神经生长因子N2培养液中培养,利用有限稀释法单克隆培养和改良法连续传代纯化并扩增NSCs,免疫组织化学法对NSCs及分化细胞进行鉴定.结果可以通过体外培养获得大量神经源性干细胞,在体外经多次传代后仍具有很强的增殖能力和多向分化潜能.结论NSCs的存活和分裂依赖于神经生长因子和N2添加剂的浓度,胚胎脑组织和神经球分离方法影响NSCs的形成速度和数量.掌握NSCs的体外纯化培养和鉴定手段可为进一步研究NSCs生物学特性及神经系统损伤的治疗提供新方法.

Isolation and cultivation of neural stem cells from the embryonic rat brain

Objective To study the isolation, cultivation and purification of neural stem cells from the embryonic rat in order to get large quantity of neural stem cells(NSCs), which were used to treat the central nerve system diseases.Methods The cells were derived from the 13 to 15 d embryonic brain cortex expanded in blood-serum-free N_ 2 supplemented culture medium which contained nerve growth factors. The NSCs were purified by limited dilution and expanded numerously in consecutive passages. NSCs and differentiated cells were detected by immunohistochemical techniques.Results Neural stem cells could be obtained in large quantity by the culture in vitro and retained the capacity after several passages to proliferate and differentiate into all three major types of cells in central nervous system.Conclusion The survival and proliferation of NSCs relies on the concentration of nerve growth factors and N_ 2 supplement. The process of asunder embryonic brain tissue and neurospheres influence the speed and quantity in NSCs' expanding. The technologies for the purification and proliferation of NSCs in vitro provide a new way for further research of the biologic specificity and the treatment of the brain trauma.