MiR-214 Promotes the Alcohol-Induced Oxidative Stress via Down-Regulation of Glutathione Reductase and Cytochrome P450 Oxidoreductase in Liver Cells
MiR-214 Promotes the Alcohol-Induced Oxidative Stress via Down-Regulation of Glutathione Reductase and Cytochrome P450 Oxidoreductase in Liver Cells
复制标题
MiR-214 通过下调肝细胞中谷胱甘肽还原酶和细胞色素 P450 氧化还原酶促进酒精诱导的氧化应激
DOI：
10.1111/acer.12209
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发表时间：
2014-01-01
影响因子：
3.2
通讯作者：
Zhao, Yanyan
中科院分区：
文献类型：
--
作者：
Dong, Xiaolong;Liu, Hong;Zhao, Yanyan
背景氧化应激在酒精性肝损伤的病理生理过程中的作用已被研究了几十年。然而，针对氧化应激基因的microRNAs(MiRNAs)在酒精性肝损伤发病机制中的作用尚未确定。本研究的目的是确定miR-214对谷胱甘肽还原酶(GSR)和细胞色素P450氧化还原酶(POR)基因的靶向性，并阐明其对酒精诱导的肝细胞氧化应激的影响。人肝癌细胞(Bel7402)、人胚胎肾293细胞(HEK293)和大鼠正常肝细胞(BRL)分别用乙醇(EtoH)转染和刺激。用乙醇喂养Wistar大鼠4周。用免疫印迹法检测GSR和POR蛋白水平，用分光光度法检测其活性。实时定量聚合酶链式反应检测miR-214的表达。用商品试剂盒检测氧化应激指标，包括总抗氧化能力(T-AOC)和丙二醛(MDA)水平。结果smiR-214与GSR和POR 3-UTR特异性结合，抑制GSR和POR的表达和活性。乙醇可上调人和大鼠肝细胞miR-214的表达，下调GSR和POR的蛋白水平和活性，并诱导肝细胞氧化应激。进一步证实酒精可上调肝脏miR-214的表达，抑制GSR和POR的表达。结论酒精通过上调miR-214的表达，进而诱导肝细胞氧化应激，从而抑制GSR和POR的表达，这是一种新的机制。
BackgroundThe involvement of oxidative stress in the pathophysiological process of alcohol-induced liver injury has been studied for decades. However, the role of microRNAs (miRNAs) targeting to oxidative stress genes in the pathogenesis of alcohol-induced liver injury has not yet been determined. The aim of this study was to identify the targeting of miR-214 to both glutathione reductase (GSR) and cytochrome P450 oxidoreductase (POR) genes and elucidate their impact on alcohol-induced oxidative stress in liver cells.MethodsThe miR-214 expression vector and reporter vectors of GSR and POR 3-UTR were constructed. Human hepatoma cell (Bel7402), human embryonic kidney 293 cell (HEK293), and rat normal hepatocyte (BRL) were transfected and stimulated with ethanol (EtOH). Wistar rats were fed with EtOH for 4weeks. The GSR and POR protein levels were detected by Western blot, and their activities were measured using the spectrophotometric method. The miR-214 expression was detected by real-time PCR. The index of oxidative stress including the total antioxidant capacity (T-AOC) and malondialdehyde (MDA) level was detected by commercial kits.ResultsmiR-214 bound specifically to the GSR and POR 3-UTR and repressed the expressions and activities of both GSR and POR. EtOH up-regulated the miR-214 expression, down-regulated the GSR and POR protein levels and activities, and induced the oxidative stress in human and rat liver cells. EtOH-fed Wistar rats further confirmed that alcohol up-regulates the miR-214 expression in liver and repressed both GSR and POR in vivo.ConclusionsThese findings demonstrated a new mechanism by which the alcohol repressed the GSR and POR expression via up-regulation of miR-214 and in turn induced oxidative stress in liver cells.