Kisspeptin通过孕酮介导的Notch1信号通路调控子宫蜕膜化在复发性流产发病中的机制研究

Kisspeptin is a protein synthesized by placental syncytiotrophoblasts. It plays an important role in embryo implantation and pregnancy maintenance, however, the specific regulation mechanism is still unknown. Our preliminary studies found the decreased expression of kisspeptin in placental villa and decidua of patients with recurrent spontaneous abortion(RSA) compared with normal early pregnancy. We found the embryo resorption rate decreased by the rescue of kisspeptin in spontaneous abortion mouse model CBA/J×DBA/2. Simultaneously, we found the condition of decidual dysplasia in spontaneous abortion mouse model improved after the treatment of kisspeptin. Combined with mRNAarray analysis of human decidualization-related molecules, we speculate that kisspeptin can promote synthesis of progesterone in syncytiotrophoblasts, and it plays an important role in the regulation of uterine decidualization through progesterone-mediated Notch1 signaling pathways. We will verify the process that kisspeptin promote progesterone synthesis by the primary culture trophoblast cells and animal model. Co-culturing system of primary trophoblast cells and endometrial stromal cells(ESC) treated by siRNA-kisspeptin and overexpress of kisspeptin will undergo to observe the decidualization of ESC and the multiple role of Notch1 signaling pathway. Foxo1cre/+ mice will be established to evaluate the role of Notch1/Akt/Foxo1 signal pathway in regulating endometrial decidualization during spontaneous pregnancy and pseudopregnancy. We hope the project would well elucidate the mechanism of kisspeptin in decidualization which provides the theoretical evidence to prevention and therapy in RSA.

Kisspeptin是由胎盘合体滋养细胞合成的蛋白，在胚胎植入和妊娠维持中起重要作用，但具体调控机制不明。我们前期研究发现复发性流产(RSA)患者胎盘和蜕膜kisspeptin表达较正常早孕降低，通过对自发流产小鼠模型给予kisspeptin补救能减少流产发生，并能改善蜕膜发育不良情况，结合对人蜕膜化相关分子的mRNAarray分析结果，我们推测kisspeptin通过促进滋养细胞合成孕酮，通过孕酮介导Notch1信号通路在调控子宫蜕膜化中起重要作用。我们应用原代分离培养滋养细胞及动物模型验证kisspeptin促进滋养细胞孕酮合成；原代滋养细胞与子宫基质细胞(ESC)共培养观察ESC蜕膜化转变及Notch1信号通路分子的改变；基因敲除小鼠生育力的评估以及假孕诱导蜕膜化验证Notch1信号通路的调控，希望探究kisspeptin在子宫蜕膜化中的重要作用，为RSA治疗提供理论依据和新的靶标。

本课题组前期研究发现复发性流产患者胎盘和蜕膜Kisspeptin较正常早孕降低，通过对自发流产小鼠模型给予kisspeptin补救能减少流产发生，并能改善蜕膜发育不良情况，但Kisspeptin是否参与和如何介导蜕膜化在流产中的作用机制尚不完全清楚。结合mRNAarray分析结果，我们发现Notch1信号通路在调控子宫内膜蜕膜化中的重要作用。本项目从临床试验、动物实验和细胞实验层面研究了Kisspeptin通过Notch/Akt/Foxo1信号通路调控蜕膜化进而参与复发性流产发病机制。研究发现，复发性流产患者蜕膜组织中，蜕膜化标志物 IGFBP1mRNA 和蛋白表达复发性流产患者组较正常妊娠女性低，KiSS1 mRNA 低表达，Kisspeptin 蛋白表达同样具有统计学差异，Notch1、Akt、Foxo1在复发性流产患者组中表达水平均低于正常妊娠组；动物实验中，正常妊娠模型小鼠注射 Kisspeptin234 与注射生理盐水组比较，可见存活胚胎数目减少，吸收胎增多，差异存在统计学意义，IGFBP1、Notch1、Akt、Foxo1表达均较生理盐水组均有下降；复发性流产模型小鼠注射 Kisspeptin10 后存活胚胎数目增多，吸收胎减少，IGFBP1、Notch1、Akt、Foxo1表达较生理盐水组均有升高； hESC 细胞中发现Notch1、Akt、Foxo1三者关系为上下游关系，Notch1/Akt/Foxo1信号通路是从Notch1向Akt，Akt向Foxo1的顺序依次调控。本研究探究kisspeptin在子宫蜕膜化中的重要作用，为RSA治疗提供理论依据和新的靶标。