BRCA-1基因调控大鼠神经干细胞增殖的实验研究

A study on the influence of BRCA-1 on proliferation of neural stem cells

目的探讨BRCA-1基因在17β-雌二醇刺激下从大鼠分离培养的神经干细胞(NSCs)增殖中的作用。方法成体SD大鼠海马NSCs的分离、纯化和培养。分为对照组及加入不同浓度的17β-雌二醇溶液共6组。于1d、7d、14d、21d分别将各组细胞进行传代培养及Nestin、BrdU免疫荧光检测,激光共聚焦显微镜对其表达进行定量分析。收集以上各时间点的NSCs以荧光定量RT-PCR法检测BRCA-1基因的表达,Western Blot法检测BRCA-1蛋白表达。相关性分析研究神经干细胞增殖与BRCA-1基因表达的关系。结果与对照组相比,Nestin表达及BrdU荧光强度在10-5、10-6、10-717β-雌二醇组逐渐加强,各实验组BRCA-1 mRNA及蛋白表达增加,统计学差异均有显著意义。NSCs增殖量大时BRCA-1基因及蛋白的表达量也增高。相关性分析说明神经干细胞的增殖与BRCA-1基因表达存在正相关趋势(相关系数r=0.46)。结论17β-雌二醇作为一种辅助因子可以促进神经干细胞的增殖。其作用与其浓度有关。BRCA-l基因能够调控成体神经干细胞的增殖。成体神经干细胞的增殖与BRCA-l基因表达蛋白量成正相关。BRCA-l基因的表达受到雌激素的调控,为正向调控。

Objective To study the possible role of BRCA-1 on proliferation of neural stem cells （NSCs） of rats under the simulating of 17β-estradiol in vitro. Methods NSCs from adult SD rats hippocampus were isolated,purified and cultured. NSCs were divided into six groups, including control group and groups adding different concentration of 17β-estradiol. At day 1,7,14,21, each group of NSCs were serially subcuhivated,immunofluorescence （IMF） of Nestin and BrdU were detected by Laser confocal microscopy. The expression of BRCA-1 mRNA and protein were detected by fluorescent quantitation RT-PCR and western blot. The relationship of NSCs proliferation and BRCA-1 expression were analyzed by correlative analysis. Results Comparing with control group,IMF of Nestin and BrdU were gradually intense in 10^-5,10^-6, 10^-7 groups, and the BRCA-1 mRNA and protein increased in experimental groups. The expression of mRNA and protein of BRCA-1 increased according to the quantity of NSCs. Correlative analysis shows that NSCs proliferation and BRCA-1 expression was positive correlation （ r = 0.46）. Condusion 17β-estradiol is a helpful factor in the proliferation of NSCs and its effect is related to its concentration. BRCA-1 can regulate adult NSCs proliferation. NSCs proliferation and BRCA-1 expression show positive correlation. The expression of BRCA-1 is positively regulated by estrogen.