中文摘要：

目的详细了解并比较不同胎龄小鼠全脑和大脑皮层组织中神经干细胞（neural stem cells,NSCs）所占比例,为后期优化分离培养NSCs提供直接量化数据。方法分离不同胎龄小鼠全脑（胎龄12.5、14、16和18d）和大脑皮层（胎龄14、16和18 d）组织,消化成单细胞悬液,贴壁培养3～4 h,细胞免疫荧光标记NSCs特异性标记物Nestin,统计分析各组中NSCs所占比例。另外,通过实时荧光定量PCR技术检测Nestin mRNA在不同胎龄（12.5、14、16和18 d）小鼠大脑皮层中的表达水平。结果细胞免疫荧光结果显示,分离培养的不同胎龄小鼠全脑和大脑皮层组织中均有Nestin阳性细胞。在分离培养的小鼠全脑组织中胎龄12.5 d组NSCs所占比例最高,为53.42%±1.57%;胎龄18 d组NSCs所占比例最低,为25.96%±1.31%,而胎龄14 d组和16 d组位于二者之间。在分离培养的不同胎龄小鼠大脑皮层组织中,胎龄14 d组NSCs所占比例最高,为33.65%±0.29%;胎龄18d组比例最低,为25.29%±0.28%,胎龄16 d组位于二者之间（26.82%±0.30%）。实时荧光定量PCR结果显示,当把胎龄12.5 d组的小鼠大脑皮层组织中Nestin mRNA表达水平设定为1,胎龄14 d组的小鼠大脑皮层组织中Nestin mRNA的相对表达量为0.83±0.04,胎龄16 d组为0.77±0.05,胎龄18 d组为0.44±0.05。因此,随着小鼠胎龄增加,小鼠大脑皮层中的Nestin mRNA的表达水平逐渐下降。结论在小鼠大脑发育过程中,随着胎龄增加,分离培养的小鼠全脑和大脑皮层组织中神经干细胞比例逐渐降低。

英文摘要：

Objective To understand and compare the proportion of neural stem cells （NSCs） in the whole brain and cerebral cortex of mice at different embryonic days, and provide quantitative data for the later optimization of NSCs isolation and culture. Methods The whole brains （at embryonic 12. 5, 14, 16 and 18 days） and cerebral cortex （at embryonicl4, 16 and 18 days） were isolated and digested into single cell suspension, and were adherently cultured for 3 - 4 h. Immunofluoreseence staining of Nestin, a NSCs specific marker, was used to statistically analyze the proportion of NSCs in each group. Expression of Nestin mRNA in the cerebral cortex of mice at El2. 5, El4, El6, and El8 was detected by real-time fluorescence quantitative PCR. Results The result of immunofluorescence assay showed that there were Nestin-positive cells in the whole brain and cerebral cortex of mice at different embryonic days. In the whole brain, the proportion of NSCs was highest at El2.5 （53.42± 1.57% ） and lowest at E18（25.96 ± 1.31% ） , and the proportions at El4 and El6 were placed in the middle among the groups. In the cerebral cortex, the highest proportion of NSCs was at El4 （33.65 ±0. 29% ）, and the lowest at E18（25.29±0. 28% ）, and the middle at El6 （26.82 ±0. 30% ）. The result of real-time PCR showed that when the mRNA expression of Nestin in the cerebral cortex was set to 1, the relative mRNA expression of Nestin was 0.83 ±0. 04 at El4, 0. 77 ±0. 05 at El6, and 0. 44 ±0. 05 at El8. Thus, the mRNA expression level of Nestin in the mouse cerebral cortex was gradually decreasing with the increase of embryonic days. Conclusions During the brain development, the proportion of NSCs is gradually decreasing in the whole brain and cerebral cortex of mice with the increase of embryonic days.