microRNA-126通过转录后调控Crk和BCAR3对PI3K信号通路的影响在子宫内膜异位症发生中的作用机制研究

Endometriosis(EMs) is one of the most commonly encountered benign gynecological disorders. The pathogenesis of the establishment of EMs is the focal point of the clinical and scientific studies in gynecology. Considerable circumstantial and laboratory evidence suggests that EMs is an estrogen-dependent disease with cell functional disorders. microRNAs regulate the stability and expression of target mRNAs as essential post-transcriptional regulators that determine cell identity and fate. Since each microRNA has been predicted to have a broad range of target mRNA based on degree of sequence homology, it is estimated that changes in the expression of even a single microRNA could have a signi?cant impact on the outcome of diverse cellular activities regulated by the product of target genes..In our previous study, we found decreased microRNA-126(miR-126) expression was signi?cantly correlated with development of EMs. Our previous evidences further validate that the rate of apoptosis is up-regulated, while the ability of proliferation, migration and angiogenesis is down-regulated along with the up-regulation of miR-126. It may provide initial evidence that down-expression of miR-126 may facilitate the formation of ectopic lesions and play an important role in the development of EMs.On the other hand, in our present study, the expression of Crk protein negatively correlated with miR-126, whereas the expression of Crk mRNA was not correlated with miR-126. These results further indicate that Crk may be a target gene of miR-126, and miR-126 may play biological function in endometrium through down-regulation of Crk protein expression post-transcriptionally. It can be concluded that miR-126 are involved in molecular pathways implicated in EMs through post-transcriptional regulation on the targets, and finally control the cytolergy. .Our present studies plan to investigate that miR-126 regulate of biological behaviours of endometrial cells and the sensitivity to estrogen through PI3K signal pathway by post-transcriptional regulation of Crk and BCAR3, and first investigate the role and molecular mechanism of miR-126 in pathogenesis of EMs with the method of biochemistry, immunology and molecular biology. We may finally demonstrate that miR-126 play a primal role on development of endometriosis, and explore the origin of pathogenesis of EMs, which provide the theoretical evidence to therapeutic targets.

子宫内膜异位症（内异症）是妇科常见病及难治病，其发生、发展的机制一直是妇科基础与临床研究探索的重点。microRNA作为转录后调节因子，在许多生命过程中起到关键作用。本课题组在前期相关研究中发现内异症的发生与microRNA-126的低表达有关，并且发现随着microRNA-126表达的升高，子宫内膜细胞凋亡率增加，增殖、迁移及血管生成能力均下降，其原因可能与microRNA-126通过转录后调控靶基因而参与一系列信号通路从而调控细胞活动有关。本研究拟采用生物化学、免疫学及分子生物学等方法，研究microRNA-126可能通过转录后调控靶基因Crk和BCAR3影响PI3K信号通路，调节子宫内膜细胞的功能及对雌激素敏感性影响，探讨microRNA-126在内异症中作用的分子机制，证明microRNA-126在内异症发生、发展中起关键作用，为寻找内异症发生的源头分子机制及治疗靶点提供理论依据。

本课题旨在前期已经进行的miR-126在子宫内膜异位症中的作用及功能鉴定研究基础上，进一步研究miR-126通过对Crk、BCAR3的靶向调控参与PI3K信号通路及雌激素调控作用，寻找子宫内膜异位症源头分子机制及治疗靶点提供理论依据，同时紧紧围绕内异症在位内膜决定论这一研究主线，进行扩展及延伸研究。通过组织、细胞等多层次分子生物学技术，进行的主要研究内容包括:（1） miR-126-5p调控BCAR3基因在内异症在位内膜上皮间质转化及间质细胞侵袭性方面具有重要影响，为后需进一步认识miR-126-5p调控BCAR3基因在内异症中的作用机制提供了新思路。（2）子宫内膜异位症在位内膜其他分子标志物相关研究：进一步延伸了前期在位内膜靶基因cofilin作用机制研究，即LIMK1基因表达增加促进子宫内膜细胞增殖、侵袭以及ICAM-1、MMP-9、VEGF表达，促进子宫内膜异位症的发生发展，而LIMK1调控cofilin1的作用与雌激素代谢相关，作为雌激素依赖性疾病，这一发现具有重要意义；利用LncRNA芯片技术，筛查子宫内膜异位症患者在位内膜和正常子宫内膜差异表达的lncRNAs和mRNAs，并对相关差异lncRNAs进行初步验证，为内异症在位内膜源头分子机制研究提供了新线索。（3）子宫内膜异位症恶变在位内膜生物学特征研究：发现在位内膜的生物学特性不但与内异症发生发展相关，而且可能是内异症恶变的源头。尤其在动物模型实验中发现,在位内膜与异位内膜具有相似的恶变风险，而异位内膜对高危因素刺激更敏感，内异症恶变问题是值得关注的热点问题，而针对在位内膜的候选基因的检测有可能成为预测内异症恶变的分子靶标，对指导临床具有重要价值。同时，还利用分子生物学技术检测了RUNX3基因的表观失活是卵巢子宫内膜异位症恶变的早期事件，并在其发展进程中起作用。在位内膜中RUNX3基因启动子区异常甲基化及蛋白表达失活可作为恶变风险的指标。由此提出了“在位内膜决定新论”，具有重要创新意义，也为下一个研究方向奠定了良好研究基础。