Hypermethylation of the Nrf2 Promoter Induces Ferroptosis by Inhibiting the Nrf2-GPX4 Axis in COPD.
Hypermethylation of the Nrf2 Promoter Induces Ferroptosis by Inhibiting the Nrf2-GPX4 Axis in COPD.
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DOI：
10.2147/copd.s340113
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发表时间：
2021
影响因子：
2.8
通讯作者：
Bian T
中科院分区：
文献类型：
--
作者：
Zhang Z;Fu C;Liu J;Sai X;Qin C;Di T;Yang Y;Wu Y;Bian T
核因子E2相关因子2（Nrf 2）参与氧化应激和肺部炎症，并调节慢性阻塞性肺疾病（COPD）的病因。铁凋亡的特征在于通过亚铁离子依赖性芬顿反应的脂质活性氧（ROS）的积累，并且涉及COPD。然而，Nrf 2在铁凋亡中的作用及其在COPD发病机制中的表观遗传调节仍不清楚。采用4-HNE、MDA、C11 BODIPY、DCFH-DA、Peals染色和CCK-8法检测血清铁蛋白含量。进行qPCR和蛋白质印迹以检查外周肺组织、从COPD患者和具有正常肺功能的受试者（从不吸烟者[对照-NS];吸烟者[对照-S]）收集的原代上皮细胞和香烟烟雾提取物（CSE）处理的人支气管上皮（HBE）细胞中的Nrf 2水平。ELISA用于定量IL-8和IL-1β水平。通过亚硫酸氢盐测序和焦磷酸测序分析Nrf 2启动子的甲基化。COPD患者存在铁蛋白沉积，谷胱甘肽过氧化物酶4（GPX 4）表达下调。CSE处理的HBE细胞活性氧、脂质过氧化物和MDA含量增加，GPX 4和SOD含量减少。IL-1β和IL-8的产生在HBE细胞中响应于CSE而被促进，但可以被铁凋亡抑制剂fer-1逆转。COPD组Nrf 2水平显著低于对照组和NS组。增加Nrf 2表达增强GPX 4和SOD水平，抑制上清液中的铁凋亡和促炎细胞因子。GPX 4的抑制逆转了Nrf 2过表达的作用，并促进了铁凋亡。在COPD组中，Nrf 2启动子内的两个特异性CpG位点高甲基化。类似地，CSE处理的HBE细胞表现出Nrf 2基因的高甲基化。COPD患者肺组织中Nrf 2基因表达下调是由于Nrf 2基因启动子甲基化，抑制Nrf 2/GPX 4和铁凋亡，与COPD的发生和发展有关。靶向Nrf 2/GPX 4可以抑制铁凋亡，这可以提供延迟或治疗COPD的策略。
Nuclear factor E2-related factor 2 (Nrf2) is involved in oxidative stress and lung inflammation and regulates the etiology of chronic obstructive pulmonary disease (COPD). Ferroptosis is characterized by the accumulation of lipid reactive oxygen species (ROS) via ferrous ion-dependent Fenton reactions and is involved in COPD. However, the role of Nrf2 in ferroptosis and its epigenetic regulation in the pathogenesis of COPD remain unclear. Ferroptosis was detected by 4-HNE, MDA, C11BODIPY, DCFH-DA, Peals’ staining and CCK-8 assays. qPCR and Western blotting were performed to examine the Nrf2 levels in peripheral lung tissues, primary epithelial cells collected from patients with COPD and subjects with normal pulmonary function (never-smoker [control-NS]; smoker [control-S]), and cigarette smoke extract (CSE)-treated human bronchial epithelial (HBE) cells. ELISA was used to quantify IL-8 and IL-1β levels. Methylation of the Nrf2 promoter was analyzed by bisulfite sequencing and pyrosequencing. Ferroptosis was involved in COPD and glutathione peroxidase 4 (GPX4) expression was downregulated in the COPD group. Reactive oxygen species (ROS), lipid peroxides and MDA were increased, but GPX4 and SOD were exhausted in CSE-treated HBE cells. The production of IL-1β and IL-8 was promoted in HBE cells in response to CSE but could be reversed by the ferroptosis inhibitor fer-1. The Nrf2 level was significantly decreased in the COPD group compared with the control-S and control-NS groups. Increased Nrf2 expression enhanced GPX4 and SOD levels and inhibited ferroptosis and proinflammatory cytokines in the supernatant. Inhibition of GPX4 reversed the effect of Nrf2 overexpression and promoted ferroptosis. Two specific CpG sites within the Nrf2 promoter were hypermethylated in the COPD group. Similarly, CSE-treated HBE cells exhibited hypermethylation of the Nrf2 gene. Nrf2 expression was downregulated in the lungs of COPD patients due to hypermethylation of the Nrf2 promoter, inhibiting Nrf2/GPX4 and ferroptosis, which is related to the initiation and progression of COPD. Targeting Nrf2/GPX4 may inhibit ferroptosis, which could provide strategies to delay or treat COPD.