雌激素对GT1-7细胞GnRH分泌及相关基因表达的影响

The effects of 17β-estradiol on GnRH serection and GnRH-related gene expressions by GT1-7 cell line

【目的】研究低浓度雌激素对下丘脑GnRH分泌以及ERα、GnRH等6个生殖相关基因表达的影响.【方法】采用体外培养GT1-7细胞,对照组添加溶剂(无水乙醇),试验组添加雌激素(100pmol/L),培养1、3、6、12、18、24、30、36h收集细胞和培养上清液,利用酶联免疫法(ELISA)测定收集上清液中GnRH浓度,并利用相对荧光定量法测定目的基因的表达.【结果】与对照组相比,试验组GnRH分泌呈增加趋势,培养12h左右GnRH分泌达到峰值(P<0.05);ERα、GnRH、Kiss-1、GPR54mRNA相对表达量先增加后降低,4种基因mRNA依次在18h(P<0.01)和24h(P<0.05)、6h(P<0.01)、3h(P<0.01)、6h和12h(P<0.05)表达显著增加.nNOS和c-fos mRNA相对表达量较对照组呈降低趋势,这2种基因mRNA依次在30h和36h(P<0.05)、3h(P<0.01)表达显著降低.【结论】100pmol/L雌激素能够促进GT1-7细胞分泌GnRH,这种作用可能是通过调控ERα、GnRH、Kiss-1、GPR54、nNOS以及c-fos mRNA的表达实现的.

[Objective] To investigate the effect of low concentration of 17/3-estradiol on GnRH secretion and 6 reproduction-related gene expressions including ERa,GnRH,Kiss- 1,GPR54, nNOS and c-los on hy- pothalamus. [Method] Taken GT1-7 cells as a model in vitro, test groups supplementation with 17]3-estra- diol （100 pmol/L）,while control groups adminstration only solvent （ethanol）, cells and cultured superna- tant were collected at 1,3,6,12,18,24,30 and 36 h, respectively, GnRH level of supernatants were deter- mined by enzyme-linked immunoassay （ELISA） and genes expression were determined by relative fluores- cence quantitative method. [Result] Compared with control groups, the GnRH secretion increased in test groups and reached secretion peak at 12 h （P〈0. 05）. ERa,GnRH,Kiss-1 and GPR54 mRNA relative ex- pression levels were firstly increased then decreased. The four genes exhibited significantly increase at 18 h （P〈0.01） and 24 h （P〈0. 05）,6 h （P〈0.01）,3 h （P〈0. 01）,6 h and 12 h （P〈0. 05）,respectively. Compared with the controls,nNOS and c-fos mRNA relative expression levels were decreased. The two genes were significantly decreased at 30 h and 36 h （P〈0. 05） ,3 h （P〈0. 01） ,respectively. [Conclusion] The 100 pmol/L 17β-estradiol can promote GnRH secretion of GT1-7 cells in vitro, which may be regulated mRNA expressions of ERa,GnRH,Kiss-l,GPR54,nNOS and c-fos genes.