PPARγ通过调控MAT2A基因转录影响子宫内膜异位症发病的分子机制研究

Endometriosis(EMs) is one of the most commonly encountered benign gynecological disorders. Up to now, the etiology and pathogenesis of endometriosis are not clear. In previous study, we found that overexpression of MAT2A in eutopic endometrium of endometriosis was associated with enhanced cell proliferation, invasion, and reduced apoptosis. It provided the initial evidence that overexpression of MAT2A facilitated the formation of ectopic lesions and played an important role in the development of endometriosis. Peroxisome proliferator-activated receptor typeγ(PPARγ) is an essential transcription factor. It was reported that PPARγ exerted a strong, negative regulatory control on MAT2A transcription by binding to PPAR response elements (PPREs) in the MAT2A promoter of quiescent hepatic stellate cells (HSCs). Our previous data showed that silencing PPARγ increased MAT2A transcription in normal endometrium whereas over-expressing it had the opposite effect in the eutopic endometrium of endometriosis, which demonstrated that PPARγ may exert its biological effects in the pathogenesis of endometriosis by negatively controlling MAT2A gene. Our present studies plan to detect mRNA and protein levels of MAT2A and PPARγ between the eutopic endometrium of endometriosis and controls and analyze the correlation of their expression. Meanwhile, we identify PPREs in the MAT2A promoter of endometrial stromal cells, examine the binding of PPARγ to each PPRE sites on the MAT2A promoter, and discuss that whether PPARγ effects the biological characteristics of the endometrium by regulating the MAT2A transcriptional activity.We first investigate the the molecular mechanism responsible for the transcriptional regulation of the MAT2A gene in endometriosis, which provide a wider perspective for understanding the pathogenesis of endometriosis and offer the basis for molecular-targeted diagnosis and therapy of endometriosis.

子宫内膜异位症（内异症）是妇科领域的常见病及难治病，目前发病机制尚不明确。前期研究发现内异症在位内膜MAT2A基因高表达促进原代子宫内膜细胞增殖、侵袭及转移，抑制细胞凋亡，提示MAT2A可能是导致子宫内膜细胞异位种植生长的重要基因。已知大鼠MAT2A基因启动子区中存在PPREs与过氧化物酶体增殖物激活受体γ（PPARγ）相互作用，调节细胞静止/激活状态。预实验研究表明内异症在位内膜组织与细胞中，PPARγ与MAT2A表达呈负相关，据此我们推测PPARγ在内异症中的作用也是通过调节MAT2A基因的表达来实现的。本研究旨在鉴定子宫内膜细胞MAT2A基因启动子区中是否存在PPREs及PPARγ与每个PPRE区域的结合效率，检测PPARγ是否通过调控MAT2A转录活性进而影响子宫内膜细胞功能，探讨内异症在位内膜MAT2A基因表达增高的分子机制，为寻找新的内异症诊断标志物及基因治疗靶点提供理论依据。

本项目紧紧围绕MAT2A基因分子调控机制展开，在前期研究筛查发现内异症在位内膜MAT2A基因较正常子宫内膜表达升高的基础上，探讨PPARγ通过调控MAT2A基因转录影响子宫内膜异位症发病的分子机制。研究发现，与正常内膜组织相比，异位内膜组织和在位内膜组织的MAT2A表达明显升高，PPARγ表达明显下降，差异具有统计学意义（P<0.05）；异位内膜组织和在位内膜组织比较PPARγ和 MAT2A表达无差异，进一步佐证了“在位内膜决定论”。本项目成功分离子宫内膜间质细胞并筛选出合适的PPARγ激动剂罗格列酮（RSG）的干预浓度（10μmol/L）。通过构建干扰和过表达的PPARγ载体，发现过表达内异症在位内膜细胞（ESC）中PPARγ表达或给予RSG干预后，MAT2A的表达明显下降，细胞增殖、迁移和侵袭能力明显下降，细胞质内可见大量空泡形成，出现典型的凋亡细胞形态学改变。相反，沉默正常子宫内膜细胞（NSC）中PPARγ表达后，MAT2A的表达明显升高，细胞增殖、迁移和侵袭能力明显升高，凋亡能力下降。双荧光素酶检测证实，PPARγ与MAT2A 基因启动子的PPREs全长、PPREs1、PPREs4和PPREs5区域具有结合作用。本项目完成了PPARγ 可通过靶定MAT2A 基因调节子宫内膜细胞生物学行为的研究。另外，本项目通过构建子宫内膜异位症大鼠模型并应用RSG进行干预，发现加入RSG干预的各组大鼠的子宫内膜异位病灶重量明显减少，异位病灶组织中PPARγ表达明显升高，MAT2A的表达明显下降，VEGF表达明显下降，Caspase-3表达明显升高，细胞活力降低，异位内膜组织内可见大量空泡形成，出现典型的凋亡细胞形态学改变，表明PPARγ激动剂类药物可通过改变 PPARγ的表达来调节MAT2A基因的转录，进而影响子宫内膜细胞的异位种植及生长。