雌激素调控Wnt/β-catenin信号与TGF-β的“串扰”在损伤后子宫内膜病理性纤维化发生中的研究

Intrauterine adhesions (IUA) is a main cause of refractory infertility,as the result of pathological basis of fibrosis and scar from improper endometrial epithelial repair afterr injury. Endometrum epithelial cell proliferation periodical under the estrogen regulate Wnt/beta-catenin signaling on homeostasis. However,there is different influence on cell responses in the setting of homeostasis and injury for the reason of different clinical prognosis.we hypothesis that the vital influence to epithelial pathological fibrosis on acount of abnormal regulation for Wnt/beta - catenin signaling and estrogen not to mention the "crosstalk" with TGF - beta interaction. This proposal aims to explore the mechanism of estrogen-regulated Wnt/ beta-catenin in the pathogenesis of IUA by accomplishing the following 4 specific aims using an air-liquid interface culture model generated with primary human endometrial epithelial cells: (1) analyzing clinical relation of serum estrogen levels and the expression of Wnt/ beta-catenin signaling molecules of endometrial in IUA; (2) dose- and time-dependent effects of estrogen on the Wnt/beta-catenin signaling in endometrial epithelial cells; (3)impacts of interaction of estrogen and Wnt/beta-catenin signaling on the characteristics of endometrial epithelial cell biology and interstitutial property; and (4) investigating the molecular mechanism of estrogen in regulating Wnt/beta-catenin signaling of endometrial epithelial cells using a rabbit IUA model. An accomplishment of this proposal will allow us to guide estrogen treatment for IUA in clinical settings.

宫腔粘连(IUA)可导致难治性不孕,病理表现为子宫内膜上皮创伤后修复障碍导致的纤维化。雌激素调控Wnt/β-catenin促进子宫内膜上皮细胞（EEC）在内稳态的增殖,但是损伤后修复中,EEC中Wnt/β-catenin和体内雌激素信号之间存在交互失衡,我们猜测这种失衡与TGF-β的“串扰”是损伤后子宫内膜纤维化的关键。本项目利用EEC气液相培养模型,通过下述内容研究它们在IUA病理发生中的机制:(1)分析血清雌激素水平和EEC中Wnt/β-catenin信号表达在IUA中的相关性;(2) 雌激素调控EEC中Wnt/β-catenin信号的剂量和时间效应;(3)雌激素调控Wnt/β-catenin信号与TGF-β的“串扰”对EEC生物学特征的影响;(4) 制备兔IUA模型探讨损伤后雌激素作用EEC中Wnt/β-catenin信号的分子机制。本项目的完成将为雌激素治疗IUA提供理论依据。

宫腔粘连(IUA)是子宫内膜创伤后修复异常的结果，纤维化使内膜失去功能导致难治性不孕。雌激素调控Wnt/β-catenin信号是子宫内膜上皮细胞（EEC）增殖的重要机制，我们猜测当两者存在调控失衡并可能与TGF-β发生“串扰”是IUA形成的关键。课题组以此为研究目标展开研究。结果表明：1.子宫内膜腺上皮细胞 Wnt/β-catenin 信号与 TGF-β1 的“串扰” 参与了IUA的形成，其中Wnt3a和TGF-β1两者互相促进纤维化的进程；2.通过二代测序对IUA相关的 lncRNA、miRNA 靶点进行筛选及验证：分析了差异基因的功能学分布，相关性最强的是Wnt信号通路，lncRNA TUG1可能通过靶向DKK1调控Wnt通路参与IUA的病理进程，而miR-543靶向MAPK1抑制Wnt通路活性降低β-catenin的核转位从而降低纤维化ESCs细胞的增殖和迁移能力，而LiCl则通过激活Wnt通路抑制这一作用；3.通过雌激素剂量效应及IUA 动物体内试验，揭示了雌激素通过激活Wnt/β-catenin通路，抑制EMT减弱TGF-β1诱导的IUA；造模后骨髓间充质干细胞联合雌激素通过抑制EMT和激活Wnt通路协同促进受损子宫内膜的修复; 进一步培养了子宫内膜腺上皮来源的类器官，在ENR培养基中添加雌孕激素可使类器官呈现对甾体激素的良好反应，原位移植促进了IUA小鼠子宫内膜创伤后修复；4.我们发现不同雌激素剂量对正常EEC和纤维化EEC具有不同的生理效应，很好的解释了临床应用雌激素产生不同的内膜增殖或抑制的现象，进一步临床观察IUA患者分离粘连术后，大剂量雌激素对IUA重度患者子宫内膜的修复更有效，但远期对小剂量雌激素有反应的子宫内膜意味着更好的功能性修复，可能是影响生殖预后的重要因素。研究结果揭示了雌激素在IUA发病机制中的作用靶点，也为其在IUA患者术后应用雌激素提供了理论依据。