基于Tsc/mTORC1和PTEN/PI3K通路研究归肾丸防治卵巢早衰的作用机制

Overactivation of primordial follicles leading to the decrease of remaining primordial follicles is the key point of premature ovarian failure. Hence premature ovarian failure could be fundamentally prevented by suppressing the activation of primordial follicles and maintaining them in a dormant state. Recent research shows that Tsc/mTORC1 and PTEN/PI3K signaling pathways play an important role in regulating the dormancy and activation of primordial follicles. Guishen pill, which represents the method of invigorating the kidney in traditional Chinese medicine, has been widely used in treating POF and menopausal syndrome. In this project, Tsc/mTORC1 and PTEN/PI3K signaling pathways are chosen as the entry point to study the influence of Guishen Pill on activation of primordial follicles from in vivo of mouse, in vitro ovary of new neonate and oocyte. In order to provide the theoretical basis for the application of Guishen Pill in preventing and curing POF, various modern experimental technologies such as radioimmunoassay, immunoprecipitation, immunoblotting, immunohistochemistry, in situ hybridization, Chinese herbs serum pharmacology will be applied to examining the molecular mechanism of Guishen Pill, which would regulate the dormancy and activation of primordial follicles to prolong the length of reproductive life through regulating the Tsc/mTORC1 signaling and PTEN/PI3K signaling.

原始卵泡的提前激活，致使剩余卵泡数量提前减少是卵巢早衰的关键原因，由于静止的原始卵泡一旦发育启动就不会停止，因此，阻止卵巢早衰患者原始卵泡提前激活，才能从根本上阻止卵巢的进一步衰竭。新近研究显示：Tsc/mTORC1和 PTEN/PI3K信号通路在原始卵泡启动、生长中具有重要地位。归肾丸为代表的中医补肾疗法被广泛用于治疗卵巢早衰或更年期综合症，本项目以调节原始卵泡生长启动的Tsc/mTORC1和 PTEN/PI3K信号通路为切入点，从小鼠在体，离体新生大鼠卵巢和卵母细胞三个层面观察归肾丸对原始卵泡启动的影响，采用放免法、免疫沉淀、免疫印迹、免疫组织化学、原位杂交、血清药理学等现代实验技术，从分子水平验证：归肾丸可能调节原始卵泡中的Tsc/mTORC1和 PTEN/PI3K信号通路维持原始卵泡的休眠状态，恢复卵巢储备，延长生育期。为临床归肾丸防治卵巢早衰提供理论依据。

原始卵泡的提前激活，致使剩余卵泡数量提前减少是卵巢早衰的关键原因，由于静止的原始卵泡一旦发育启动就不会停止，因此，阻止卵巢早衰患者原始卵泡提前激活，才能从根本上阻止卵巢的进一步衰竭。Tsc/mTORC1 和 PTEN/PI3K 信号通路在原始卵泡启动、生长中具有重要地位。归肾丸为代表的中医补肾疗法被广泛用于治疗卵巢早衰或更年期综合症，本研究通过小鼠在体，离体新生小鼠卵巢和卵母细胞三个层面观察归肾丸对原始卵泡启动的影响。结果发现（1）归肾丸能明显改善小鼠动情周期，阻止始基卵泡数量的下降，并改善卵巢分泌性激素水平；归肾丸能明显抑制体外培养卵巢的始基卵泡发育启动；归肾丸能明显抑制KL诱导的始基卵泡发育启动。（2）小鼠在体，离体新生小鼠卵巢和卵母细胞实验都显示归肾丸是通过Tsc1/mTORC1和PTEN/PI3K信号通路维持始基卵泡的休眠状态，阻止卵泡消耗。这些研究结果提示，归肾丸通过调节原始卵泡中的Tsc/mTORC1 和 PTEN/PI3K 信号通路维持原始卵泡的休眠状态，恢复卵巢储备，延长生育期，为临床归肾丸防治卵巢早衰提供理论依据。