孕酮回植激活Akt/GSK3β信号通路逆转月经发生

摘要

目的 探讨在孕酮撤退月经模型小鼠月经发生的关键时期前,回植孕酮对蛋白激酶B/糖原合成激酶3β(Akt/GSK3β)信号通路的影响. 方法 30只成熟去势雌性C57BL/6J小鼠在建成小鼠孕酮撤退月经模型的基础上,随机分为3组:孕酮撤退后12 h组(12 h组)、孕酮撤退后16 h组(16 h组),以及在孕酮撤退后12 h回植孕酮皮埋管4 h组(回植孕酮组),每组10只.通过qRT-PCR法检测子宫内膜组织中Akt、GSK3β mRNA表达,Western blotting和免疫组织化学检测总Akt(t-Akt)、磷酸化Akt(p-Akt)(Ser473)和p-GSK3β(Ser9)蛋白的表达. 结果 与12 h组相比,回植孕酮组和16 h组中Akt mRNA表达显著下降(P均<0.05),而回植孕酮组和16 h组无显著性差异(P>0.05);12 h组的GSK3β mRNA表达量显著高于回植孕酮组和16 h组(P均<0.05).与12 h组相比,p-Akt、p-GSK3β蛋白在回植孕酮组表达量显著增高(P均<0.05);而16 h组子宫内膜组织中p-Akt、p-GSK3β蛋白表达量亦显著低于12 h组和回植孕酮组(P均<0.05).免疫组化结果显示,p-Akt和p-GSK3β蛋白在小鼠子宫组织中表达分布呈现相似的模式,主要广泛散在表达于蜕膜化区域的基质细胞及腺上皮和腔上皮细胞中,在蜕膜化区域周边稍有集中分布呈带的趋势. 结论 在月经发生关键时期之前回植孕酮可能通过非转录机制快速激活Akt/GSK3β信号通路及其下游级联反应,从而逆转月经发生.%Objective: To investigate the effects of implanted progesterone at the critical period before menstrual occurrence induced by progesterone withdrawal on the Akt/GSK3 beta signaling pathway in mice menstrual-like model. Methods: Thirty ovariectomized C57BL/6J female mice with menstrual-like model were randomly divided into three groups: 12 hours after progesterone withdrawal group (12 hours group),16 hours after progesterone withdrawal group (16 hours group),12 hours after progesterone withdrawal followed by implantation of progesterone for 4 hours group (implanted progesterone group),ten mice were for each group. The uterus horns of mouse were collected at 12 hours & 16 hours after progesterone withdrawal,and 12 hours after progesterone withdrawal followed by implantation of progesterone for 4 hours. The changes of Akt,p-Akt,GSK3β and p-GSK3β expression in mouse uterine tissue were detected by immunohistochemistry,Western blotting and qRT-PCR respectively. Results: The expression of Akt mRNA of mouse uterine tissue in 12 hours group was significantly higher than implanted progesterone group and 16 hours group (P<0.05),and there was no significantly different between the implanted progesterone group and 16 hours group (P>0.05). The expression of GSK3β mRNA in the 12 hours group was significantly higher than that in the implanted progesterone group and 16 hours group (P<0.05). The expression of p-Akt and p-GSK3β in endometrial tissue in 16 hours group was significantly lower than that in 12 hours group and implanted progesterone group (P<0.05). Immunohistochemical results showed that the expression of p-Akt and p-GSK3β protein presented similar distribution patterns in mouse uterine tissues,and p-Akt and p-GSK3β protein were widely distributed in stromal cells,glandular epithelium and luminal epithelial cells in decidual areas. Conclusions: Implanted progesterone at the critical period,12 hours after progesterone withdrawal,reverses menstruation by rapidly activating Akt/GSK3 beta signaling pathway and its downstream cascade reaction through non-transcriptional mechanisms.