Department of Basic Sciences-Microbiology Division, Ponce Health Sciences University, Ponce, Puerto Rico, USA.
Department of Biomedical Sciences, Division of Neurosciences, University of Texas at Rio Grande Valley-School of Medicine, Texas, USA.
Biol Reprod. 2018 Aug 1;99(2):293-307. doi: 10.1093/biolre/ioy030.
Although the histone methyltransferase EZH2 and its product H3K27me3 are well studied in cancer, little is known about their role and potential as therapeutic targets in endometriosis. We have previously reported that endometriotic lesions are characterized by global enrichment of H3K27me3. Therefore, we aimed to (1) characterize the expression levels of EZH2 in endometriotic tissues; (2) assess H3K27me3 enrichment in candidate genes promoter regions; and (3) determine if pharmacological inhibition of EZH2 impacts migration, proliferation, and invasion of endometriotic cells. Immunohistochemistry of an endometriosis-focused tissue microarray was used to assess the EZH2 protein levels in tissues. Chromatin immunoprecipitation-qPCR was conducted to assess enrichment of H3K27me3 in candidate gene promoter regions in tissues. Immunofluorescence was performed to assess the effect of an EZH2-specific pharmacological inhibitor on H3K27me3 global enrichment in cell lines. To measure effects of the inhibitor in migration, proliferation, and invasion in vitro we used Scratch, BrdU, and Matrigel assays, respectively. Endometriotic lesions had significantly higher EZH2α nuclear immunostaining levels compared to eutopic endometrium from patients (glands, stroma) and controls (glands). H3K27me3 was enriched within promoter regions of candidate genes in some but not all of the endometriotic lesions. Inhibition of EZH2 reduced H3K27me3 levels in the endometriotic cells specifically, and also reduced migration, proliferation but not invasion of endometriotic epithelial cells (12Z). These findings support future preclinical studies to determine in vivo efficacy of EZH2 inhibitors as promising nonhormonal treatments for endometriosis, still an incurable gynecological disease.
尽管组蛋白甲基转移酶 EZH2 及其产物 H3K27me3 在癌症中研究得很好，但它们在子宫内膜异位症中的作用和作为治疗靶点的潜力知之甚少。我们之前报道过，子宫内膜异位症病变的特征是 H3K27me3 的全局富集。因此，我们旨在：(1) 描述 EZH2 在子宫内膜异位症组织中的表达水平；(2) 评估候选基因启动子区域 H3K27me3 的富集情况；(3) 确定 EZH2 的药理学抑制是否影响子宫内膜异位症细胞的迁移、增殖和侵袭。通过子宫内膜异位症组织微阵列的免疫组织化学评估组织中 EZH2 蛋白水平。进行染色质免疫沉淀-qPCR 以评估候选基因启动子区域 H3K27me3 的富集情况。免疫荧光用于评估 EZH2 特异性药理学抑制剂对细胞系中 H3K27me3 全局富集的影响。为了测量抑制剂在体外迁移、增殖和侵袭中的作用，我们分别使用划痕、BrdU 和 Matrigel 测定法。与患者（腺体、基质）和对照（腺体）的在位子宫内膜相比，子宫内膜异位症病变的 EZH2α 核免疫染色水平显著升高。候选基因启动子区域内 H3K27me3 在一些但不是所有的子宫内膜异位症病变中都有富集。EZH2 的抑制作用降低了子宫内膜异位症细胞中 H3K27me3 的水平，同时也降低了子宫内膜异位症上皮细胞的迁移、增殖（12Z），但不降低侵袭。这些发现支持未来的临床前研究，以确定 EZH2 抑制剂作为治疗子宫内膜异位症的有前途的非激素治疗方法的体内疗效，子宫内膜异位症仍然是一种无法治愈的妇科疾病。
