Histone modifications play a critical role in chromatin remodelling and regulate gene expression in health and disease. Histone methyltransferases EZH1, EZH2, and demethylases UTX, JMJD3, and UTY catalyse trimethylation of lysine 27 on histone H3 (H3K27me3). This study was designed to investigate whether H3K27me3 triggers hyperglycemia-induced oxidative and inflammatory transcriptional programs in the endothelium. We studied human aortic endothelial cells exposed to high glucose (HAEC) or isolated from individuals with diabetes (D-HAEC). RT-qPCR, immunoblotting, chromatin immunoprecipitation (ChIP-qPCR), and confocal microscopy were performed to investigate the role of H3K27me3. We determined superoxide anion (O2−) production by ESR spectroscopy, NF-κB binding activity, and monocyte adhesion. Silencing/overexpression and pharmacological inhibition of chromatin modifying enzymes were used to modulate H3K27me3 levels. Furthermore, isometric tension studies and immunohistochemistry were performed in aorta from wild-type and db/db mice. Incubation of HAEC to high glucose showed that upregulation of EZH2 coupled to reduced demethylase UTX and JMJD3 was responsible for the increased H3K27me3. ChIP-qPCR revealed that repressive H3K27me3 binding to superoxide dismutase and transcription factor JunD promoters is involved in glucose-induced O2− generation. Indeed, loss of JunD transcriptional inhibition favours NOX4 expression. Furthermore, H3K27me3-driven oxidative stress increased NF-κB p65 activity and downstream inflammatory genes. Interestingly, EZH2 inhibitor GSK126 rescued these endothelial derangements by reducing H3K27me3. We also found that H3K27me3 epigenetic signature alters transcriptional programs in D-HAEC and aortas from db/db mice. EZH2-mediated H3K27me3 represents a key epigenetic driver of hyperglycemia-induced endothelial dysfunction. Targeting EZH2 may attenuate oxidative stress and inflammation and, hence, prevent vascular disease in diabetes.

中文翻译：

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抑制性 H3K27me3 驱动高血糖诱导的人内皮氧化和炎症转录程序

组蛋白修饰在染色质重塑中发挥着关键作用，并调节健康和疾病中的基因表达。组蛋白甲基转移酶 EZH1、EZH2 和去甲基化酶 UTX、JMJD3 和 UTY 催化组蛋白 H3 (H3K27me3) 上赖氨酸 27 的三甲基化。本研究旨在调查 H3K27me3 是否触发高血糖诱导的内皮细胞氧化和炎症转录程序。我们研究了暴露于高葡萄糖（HAEC）或从糖尿病患者中分离的人主动脉内皮细胞（D-HAEC）。采用 RT-qPCR、免疫印迹、染色质免疫沉淀 (ChIP-qPCR) 和共聚焦显微镜来研究 H3K27me3 的作用。我们通过 ESR 光谱、NF-κB 结合活性和单核细胞粘附测定了超氧阴离子 (O2−) 的产生。使用染色质修饰酶的沉默/过度表达和药理学抑制来调节 H3K27me3 水平。此外，在野生型和 db/db 小鼠的主动脉中进行了等长张力研究和免疫组织化学。 HAEC 与高葡萄糖的孵育表明，EZH2 的上调与还原的去甲基化酶 UTX 和 JMJD3 结合是 H3K27me3 增加的原因。 ChIP-qPCR 显示，抑制性 H3K27me3 与超氧化物歧化酶和转录因子 JunD 启动子的结合参与了葡萄糖诱导的 O2− 生成。事实上，JunD 转录抑制的丧失有利于 NOX4 的表达。此外，H3K27me3 驱动的氧化应激增加了 NF-κB p65 活性和下游炎症基因。有趣的是，EZH2 抑制剂 GSK126 通过减少 H3K27me3 来挽救这些内皮紊乱。我们还发现 H3K27me3 表观遗传特征改变了 db/db 小鼠 D-HAEC 和主动脉的转录程序。 EZH2 介导的 H3K27me3 是高血糖诱导的内皮功能障碍的关键表观遗传驱动因素。靶向 EZH2 可能会减轻氧化应激和炎症，从而预防糖尿病中的血管疾病。