血性脑脊液对胚胎神经干细胞增殖和分化的影响

Proliferation and differentiation of neural stem cells in hemorrhagic cerebrospinal fluid

目的观察体外血性脑脊液培养对神经干细胞增殖和分化的影响,以期为临床治疗这类患者提供依据。方法提取终止妊娠的16周人胚胎脑细胞,冻存于液氮中。组织复苏后,在DMEM/F12培养基(含EGF、bFGF、B27和N2)中培养14 d可获得形态完好的神经球(神经干细胞)。从颅脑外伤患者和非外伤患者分别留取血性脑脊液和正常脑脊液。将制备的胚胎神经干细胞分为两组,分别用血性脑脊液和正常脑脊液培养。动态观察神经干细胞在两组脑脊液中生长、增殖和分化的情况。用免疫细胞化学技术对两组脑脊液中神经干细胞的分化进行标记和鉴定。结果神经干细胞在两组脑脊液中均能存活、增殖和分化。但神经干细胞在血性脑脊液中分化速度较快,分化比例也较高。在血性脑脊液中,神经干细胞更倾向于向胶质细胞分化;而在正常脑脊液中,神经干细胞更倾向于向神经元分化。结论血性脑脊液可能会影响神经干细胞的分化速度和分化方向。这一结果对采用神经干细胞治疗颅脑外伤和蛛网膜下腔出血等疾病有一定提示作用。

Objective To observe proliferation and differentiation of neural stem cells in hemorrhagic eerebrospinal fluid in vitro. Methods Human fetal brain cells of gestational 16 weeks were obtained from those who were induced abortion and frozen in liquid nitrogen. Tissue was then resuscitated and incubated in DMEM/FI2 medium with epithium growth factor, basic fibroblast growth factor, B27 and N2. The neural stem cells （NSCs） were obtained after brain tissue cultured for 14 days. Obtained NSCs were randomized and cultured in hemorrhagic CSF from brain injurypatient and normal CSF from other patients. The growth, proliferation and differentiation of NSCs in two groups were observed dynamically. The cell differentiation was identified by immunocytochemistry method. Results NSCs could survive, proliferate and differentiate in both of hemorrhagic and normal CSF. In hemorrhagic CSF, differentiation of NSCs was faster and attachment ratio was higher compared with that of in normal CSF. NSCs were prone to differentiate into astrocyte in hemorrhagic CSF, but were to differentiate into neurons in normal CSF. Conclusions Hemorrhagic CSF may influence proliferation and differentiation of NSCs. This data is referential for treating traumatic brain injury with NSCs transplantation.