高龄妊娠对子鼠海马神经干细胞发育的影响

Effect of advanced maternal age on development of hippocampal neural stem cells in offspring rats

摘要:

目的 探讨高龄妊娠对子鼠海马神经干细胞发育的影响。方法 将3月龄（n=10）与12月龄（n=10）的雌性大鼠分别与3月龄雄性大鼠（n=20）合笼，以其子鼠为研究对象，分为适龄子代组和高龄子代组，每组40只。分别采用免疫荧光及Western blot法定位及定量检测生后第7天两组海马组织巢蛋白（Nestin）、双皮质素（DCX）及生后第28天成熟神经元标志物NeuN、胶质纤维酸性蛋白（GFAP）表达水平；采用免疫荧光法定位检测生后第14天两组海马组织多聚唾液酸神经细胞黏附分子（PSA-NCAM）表达情况；每组各检测指标随机取8只大鼠。结果 Western blot检测结果发现高龄子代组海马DCX蛋白的表达明显低于适龄子代组（P < 0.05），Nestin、NeuN及GFAP蛋白表达在两组间比较差异均无统计学意义（P > 0.05）。免疫荧光检测结果显示，高龄子代组海马齿状回（DG）区Nestin、DCX及PSA-NCAM的表达均明显低于适龄子代组（P < 0.05），而上述指标分别在两组海马CA1区、CA3区的表达比较差异均无统计学意义（P > 0.05）。高龄子代组海马CA1区NeuN的表达高于适龄子代组（P < 0.01），两组CA3区和DG区NeuN的表达比较差异无统计学意义（P > 0.05）。高龄子代组海马CA1区、CA3区、DG区GFAP的表达均低于适龄子代组（P < 0.05）。结论 高龄妊娠可能引起子鼠海马神经干细胞的增殖、存活及迁移受到抑制，并影响其向神经元和星形胶质细胞的分化，最终导致子鼠海马神经干细胞发育障碍。

Abstract:

Objective To study the effect of advanced maternal age (AMA) on the development of hippocampal neural stem cells in offspring rats. Methods Ten 3-month-old and ten 12-month-old female Sprague-Dawley rats were housed individually with 3-month-old male rats (1:1, n=20), whose offspring rats were assigned to a control group and an AMA group. A total of 40 rats were randomly selected from each group. Immunofluorescence assay and Western blot were used to localize and determine the levels of protein expression of Nestin and doublecortin (DCX) on day 7 as well as neuronal nuclear antigen (NeuN) and glial fibrillary acidic protein (GFAP) on day 28 (n=8 for each marker). Immunofluorescence assay was also used to localize the hippocampal expression of polysialylated isoforms of neural cell adhesion molecule (PSA-NCAM) on day 14 (n=8 for each marker). Results According to the Western blot results, the AMA group had significantly lower protein expression of DCX than the control group (P < 0.05), while there were no significant differences in the protein expression of Nestin, NeuN, and GFAP between the two groups (P > 0.05). According to the results of immunofluorescence assay, the AMA group had significantly lower protein expression of Nestin, DCX, and PSA-NCAM in the hippocampal dentate gyrus (DG) region than the control group (P < 0.05), while there were no significant differences in the above indices in the hippocampal CA1 and CA3 regions between the two groups (P > 0.05). The AMA group had significantly higher expression of NeuN in the hippocampal CA1 region than the control group (P < 0.01), while there were no significant differences in the expression of NeuN in the hippocampal DG and CA3 regions between the two groups (P > 0.05). The AMA group had significantly lower expression of GFAP in the hippocampal CA1, CA3, and DG regions than the control group (P < 0.05). Conclusions AMA may cause inhibition of proliferation, survival, and migration of hippocampal neural stem cells. AMA may also affect their differentiation into neurons and astrocytes, which will eventually lead to developmental disorders of hippocampal neural stem cells in offspring rats.