摘要
目的探讨不同胎龄胎鼠脊髓源性神经干细胞(NSCs)的增殖分化能力。方法 Sprague-Dawley孕鼠分为A组(孕12d)、B组(孕14 d)、C组(孕16 d),酶消化法结合机械分离法提取细胞,检测不同时间各组神经干细胞球的直径、数量,CCK-8法绘制细胞生长曲线。Brd U/Nestin免疫荧光染色鉴定NSCs。10%胎牛血清诱导分化后行β-tubulinⅢ、胶质纤维酸性蛋白(GFAP)免疫荧光染色,比较各组神经元的分化比例。结果各组细胞Brd U、Nestin、β-tubulinⅢ、GFAP免疫荧光染色均为阳性。B组β-tubulinⅢ染色阳性的比例最高;B、C两组各时间点神经球直径、数量及神经元的分化比例高于A组(P<0.05)。第3代NSCs培养第5天,B组细胞吸光度高于C组(P<0.05)。结论孕14 d左右取材培养的脊髓神经干细胞生物学特性稳定,增殖活力强,诱导分化后神经元的分化比例较高。
Objective To study the differentiation and proliferation ability of the spinal neural stem cells（NSCs） at different gestational ages in fetal rats. Methods Sprague-Dawley fetal rats were divided into group A（12 days of pregnancy）, group B（14 days of pregnancy）and group C（16 days of pregnancy）. NSCs were separated with enzyme-assisted microdissection. The diameter and numbers of NSCs balls were measured at different time. The cell growth curve was drawn with CCK8 colorimeter. NSCs were identified with BrdU/Nestin immunohistochemical staining. They were induced with 10% fetal bovine serum for 10 days, and the expression of β-tubulinⅢ and glial fibrillary acidic protein was detected with immunocytochemistry. Results There were cells expressed BrdU, Nestin, β-tubulinⅢ and GFAP in all the group. The most cells（22.74±0.79%） expressed β-tubulinⅢ in the group B, but no significant difference between group B and group C. The cell vitality on the 5th day of third-generation neural stem cells was the most in group B. Conclusion For enzyme-assisted microdissection,it may obtain more neurons to isolate the neural stem cells from 14 days of pregnancy pregnant rats.
出处
《中国康复理论与实践》
CSCD
北大核心
2015年第10期1145-1150,共6页
Chinese Journal of Rehabilitation Theory and Practice
基金
国家自然科学基金项目(No.30772299)
甘肃省自然科学研究基金计划项目(No.1107RJZA140)
