基于炎症与血液高凝状态的关联研究促孕复方改善胚胎着床障碍及早期妊娠丢失的分子机制

In vitro fertilization-embryo transfer (IVF-ET) is a widely used assisted reproductive technology, which has solved the fertility problems of many infertile couples to some extent. But the success rate is still low, and most patients still lose their confidence due to repeated IVF failure. The main reason is that the use of controlled ovarian hyperstimulation (COH) has changed the microenvironment of embryo implantation and reduced endometrial receptivity. The previous work of this research group has confirmed that Promoting Pregnancy Recipe can significantly improve the embryo implantation microenvironment, promote embryo implantation and improve the success rate of pregnancy. But the exact mechanism remains unclear. Based on the correlation between inflammation and prethrombotic state, the possible key role of fgl2 linking inflammation and prethrombosis state, we assume that: fgl2 is the key factor of inflammation and thrombosis formation in mice with embryo implantation dysfunction, Promoting Pregnancy Recipe can improve embryo implantation microenvironment and increase pregnancy rate by promoting M2 macrophage polarization, reducing fgl2 expression. This project will choose fgl2 knockout and wild mice as the research object, make model of COH with gonadotropin, observe blood circulation of early embryo, macrophage activation state, the expression of inflammatory cytokines and fgl2, and further study the intervention effect and the molecular mechanism of Promoting Pregnancy Recipe on the micro environment of embryo implantation. This project will provide reliable experimental basis for Promoting Pregnancy Recipe improving the success rate of pregnancy, lay a good foundation for the popularization and application in the field of reproductive.

体外受精-胚胎移植技术（IVF-ET）是目前广泛应用的辅助生殖技术，解决了许多不孕夫妇的生育问题。但其成功率依然较低，大部分患者仍因反复IVF失败而最终丧失生育信心。其主要原因是控制性超排卵（COH）技术的使用改变了胚胎着床微环境。本课题组前期工作已证实促孕复方能促进胚胎着床，提高妊娠率，但其作用机理仍不清楚。结合炎症与血栓形成的关联以及fgl2在其中的关键纽带作用，我们假设：fgl2是胚胎着床障碍小鼠炎症与血栓形成的关键因子，促孕复方通过促进巨噬细胞向M2极化、降低fgl2的表达而改善胚胎着床微环境提高妊娠率。本项目选用fgl2基因敲除鼠和野生鼠为研究对象，以COH造成胚胎着床障碍模型，观察各组小鼠妊娠早期胚胎血液循环状况、巨噬细胞活化状态、fgl2表达情况以及炎症水平，并在此基础上进一步研究促孕复方改善胚胎着床障碍及早期妊娠丢失的分子机制，为其推广应用提供理论和实验依据。

人类早期妊娠丢失的原因复杂多样，目前研究表明患者年龄、染色体异常、子宫异常、血液高凝状态、内分泌和代谢紊乱、感染、精子质量和生活方式均可影响妊娠的结局。体外受精-胚胎移植技术（IVF-ET）作为目前广泛应用的辅助生殖技术，其成功率仍然较低，主要原因是控制性超促排卵（COH）技术的使用改变了胚胎着床微环境。促孕复方（CYR）是基于中医补肾益气活血法则使用多味中药配伍而成的中药复方，临床用于治疗不孕症或妊娠丢失，已取得较好的临床效果。本课题结合炎症与血栓形成的关联以及fgl2在其中的关键纽带作用，研究促孕复方改善胚胎着床障碍及早期妊娠丢失的分子机制。.本课题通过腹腔注射孕马血清促性腺激素（PMSG）和人绒毛膜促性腺激素（hCG）进行超促排卵造模，成功建立雌性KM小鼠COH模型，通过CYR HPLC指纹图谱分析，确定促孕复方活性成分后，展开实验探究CYR对COH模型小鼠妊娠、卵巢过度刺激状态、子宫蜕膜化的影响，以及CYR对围着床期小鼠炎症和高凝状态的影响。结果表明：与对照组小鼠相比，COH模型组小鼠的妊娠率显著降低，同时平均着床胚泡数降低，卵巢增大，子宫血管发育不良，子宫和脾脏巨噬细胞倾向M1极化，脾脏促炎型单核细胞增多，子宫内膜及血浆中TNF-α、IL1-β、MCP-1、IL-12含量升高，IL-10、TGF-β含量降低，血浆D-二聚体增多，子宫内膜中fgl2表达增加，且子宫内膜巨噬细胞中fgl2表达增加。促孕复方可一定程度逆转上述变化。.本课题研究证明CYR可以通过缓解卵巢过度刺激、促进子宫内膜蜕膜化与血管生成、调节全身炎症状态与子宫局部炎症状态、改善血液高凝状态来改善胚胎着床和妊娠丢失，为临床应用与推广补肾益气活血疗法治疗妊娠丢失提供了理论和实验依据。