挫伤脑组织对人胚神经干细胞影响的体外研究

目的 建立人胚胎来源的神经干细胞体外培养方法，初步研究挫伤脑组织提取液(TBITE)对人胚神经干细胞(HNSCs)存活、增殖能力和分化的影响。方法 10周龄左右的胎儿大脑皮层细胞体外培养，神经干细胞增殖2～3代后，剔除生长因子，分别加入不同浓度的脑外伤病人的挫伤脑组织提取液，培养2～3周后，免疫细胞化学方法鉴定细胞分化后的类型。用Laemrnli凝胶电泳法分析培养液上清。结果成功获得以神经球形式生长的人胚神经干细胞，Nestin表达阳性；与空白对照相比，TBITE组中的神经干细胞有明显的增殖，并分化为神经细胞、星形胶质细胞和少突胶质细胞。随着TBITE含量的增加，星形胶质细胞的比例增加。结论 脑外伤的局部微环境可促进神经干细胞增殖和分化。

Effect of traumatic brain injury tissue on human embryonic neural stem cells

Objective To establish the methods of culturing human embryonic neural stem cells (NSCs) in vitro and to investigate the effect of traumatic brain injury tissue extraction (TBITE) on the survival, proliferation and differentiation of human embryonic NSCs. Methods The cells were separated from 10-week-old fetal forebrain cortex and were cultured in serum-free DMEM/F12 supplemented with epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and leukemia inhibit factor (LIF). When NSCs were passaged for 2 to 3 times, the growth factors were depleted from culture media. Cells were moved onto coverslips precoated with poly-D-lysine and 10% fetal bovine serum and TBITE were added into the media. Two or three weeks later, immunocytochemistry was used to identify the types of differentiated cells. At the time the remarkable differentiation was observed, culture media were preserved for Laemmli gel electrophoresis and Coomassie light blue R250 dying. Results NSCs were successfully separated and proliferated in vitro. These cells were nestin-positive and could be induced to differentiate into neurons, astrocytes and oligodendrocytes by TBITE. Conclusions TBITE could stimulate the proliferation and differentiation of NSCs.Higher density of TBITE caused more astrocytes production.