CDK5通过自噬调控子宫内膜异位症上皮——间质转化的分子机制研究

Endometriosis (EMS) is a common benign gynecological disease. We detected the expression of cyclin dependent kinase 5 (CDK5) in the eutopic endometrium of women with endometriosis was significantly higher than in ectopic endometrium as well as in eutopic endometrium from women without endometriosis. High expressed CDK5 contributed to cell proliferation, migration and invasion, whereas decreased the expressions of autophagy-related proteins LC3A, LC3B and Beclin 1, in endometrial epithelial cells. Previous research has domenstrated that autophagy was down-regulated, while the epithelial-mesenchymal transition (EMT) was up-regulated in the eutopic and ectopic endometrium. We speculate that CDK5-inhibit autophagy may promote an EMT in endometrial epithelial cells of women with endometriosis, and finally lead to the pathogenesis and progression of endometriosis. . In this research, we intends to study the relationship between CDK5-mediated autophagy and the EMT in endometrial epithelial cells, using clinical specimens, in vitro model of human endometrial epithelial cells and in vivo endometriosis model of BALB/C mice. We analyse the expressions of CDK5 and its regulators in clinical specimens, and their correlations with the progression of endometriosis. We explore whether autophagy inhibits the CDK5-mediated EMT depending on the activation of Wnt/β-catenin signaling pathway, and clarify its molecular mechanisms, by regulated CDK5, GSK3β and mTOR expressions in endometrial epithelial cells. The BALB/C mice model of endometriosis is also set up to work out the effect of CDK5 in the progresssion of ectopic lesion of endometriosis for in vivo study. Finally, we hope to provide a new target for the precise treatment of endometriosis.

子宫内膜异位症（EMS）是一种妇科良性疾病，我们发现EMS在位子宫内膜中细胞周期素依赖蛋白激酶5（CDK5）表达上调；CDK5高表达促进子宫内膜腺上皮细胞增殖、迁移与侵袭，抑制自噬相关基因LC3A、LC3B及Beclin 1转录。研究已证实EMS子宫内膜中自噬降低、上皮—间质转化（EMT）升高。我们推测CDK5可能抑制子宫内膜腺上皮细胞自噬发生，诱导EMT，促进EMS发生。本项目拟从临床观察、细胞及小鼠实验研究CDK5拮抗的自噬与子宫内膜腺上皮细胞EMT的关联。检测临床标本CDK5及其调节蛋白表达，分析与疾病进程的关联；双向调节子宫内膜腺上皮细胞CDK5、GSK3β和mTOR表达，探讨自噬调控CDK5诱导的EMT途径是否依赖Wnt/β-catenin信号通路激活，阐明其分子机制；抑制EMS小鼠模型CDK5表达，观察病灶变化，验证CDK5对EMS发生发展的影响。为EMS的精细治疗提供新靶点。

子宫内膜异位症（Endometriosis，EMS）是一种常见的妇科良性疾病，指具有生长功能的子宫内膜组织（包括腺体和间质）在子宫腔及宫体肌层以外的部位种植并存活。拥有种植、迁移侵袭及血管生成等“恶性”生物学行为特征。CDK5在中枢神经系统是调节神经元存活与死亡的“开关”，在肿瘤中它促进癌细胞运动、迁移和侵袭，与肿瘤的恶化及不良预后相关。本研究通过免疫组化和Western Blot实验发现CDK5在子宫内膜异位症患者异位子宫内膜腺上皮细胞和间质细胞中的表达均显著高于在位子宫内膜和非子宫内膜异位症患者对照子宫内膜来源的细胞。利用confocal、CCK-8、transwell和western blot等一系列实验，明确了上调CDK5表达，导致子宫内膜腺上皮细胞自噬受到抑制，诱导上皮间充质转化，促进细胞迁移侵袭。阐明了CDK5通过提高p38磷酸化，促进p62表达，抑制Atg5和Atg16L表达，阻遏Atg5-Atg12-Atg16L复合物诱导LC3-I向LC-II的转化，从而抑制自噬体膜的形成。揭示了CDK5介导的自噬调控子宫内膜腺上皮细胞EMT途径，依赖p38 MAPK信号通路的激活。通过子宫内膜异位症裸鼠模型实验，在体内证实下调CDK5表达抑制裸鼠体内子宫内膜异位症病灶的发生与生长，而自噬抑制剂3-methyladenine作用挽救了CDK5对病灶生长的促进作用。通过对该机制的研究，一方面指出了CDK5在子宫内膜异位症中的作用，另一方面为子宫内膜异位症的治疗提供了新思路。