神经干细胞体外长期培养和外源基因表达

The long-term culture in vitro and exogenous gene expression of neural stem cells

目的 探讨胚胎大鼠神经干细胞的体外培养条件及外源基因的表达效率。方法 从胚胎大鼠脑皮质中机械分离细胞,应用N2培养基进行培养和扩增,采用免疫组织化学方法进行鉴定,应用Ad5βgal重组腺病毒载体检测神经干细胞表达外源基因的能力。结果 从胚胎大鼠的大脑皮质中成功分离培养出神经干细胞,在悬浮状态下培养形成典型的神经球并表达波形蛋白和巢蛋白。近100％的细胞可被Ad5βgal基因感染并表达lacZ基因。细胞可用机械方法传代,常规冻融和复苏,体外长期培养可达3个月。结论 神经干细胞可在体外长期稳定培养和传代,能表达βgal外源基因,是细胞治疗和基因治疗的良好载体。

Objective To explore the culture conditions for the neural stem cells from embryonic rat cortices and to test their efficacy of expressing exogenous gene. Methods The cells from the embryonic rat cortices were mechanically dissociated and the N2 medium was adopted to culture and expand the cells. Then the cells were identified by immunocytochemistry method, and the Ad5βgal vector was applied to test their capacity of expressing foreign gene. Results The neural stem cells from embryonic rats were successfully cultured, which formed typical neurospheres in suspension and expressed nestin and vimentin. Most cells were infected by Ad5βgal gene and expressed lacZ gene. The cells could be passaged by mechanical method, frozen and thawed by general procedure, they could be passaged and expanded nearly for up to 3 months. Conclusion The neural stem cells can be cultured and passaged steadily in vitro. They can express βgal foreign gene and are the ideal vectors for cell therapy and gene therapy.