miR-340-5p及PDCD4在过表达乙肝病毒X蛋白的足细胞中的表达及生物学意义  

Expression and Biological Significance of miR-340-5p and PDCD4 in Podocytes Overexpressing Hepatitis B Virus X Protein

作　　者：丁广智 徐佳 刘畅[1] 吴国志 DING Guangzhi;XU Jia;LIU Chang;WU Guozhi(Qingdao Haici Hospital Affiliated to Qingdao University,Qingdao 266033,China)

机构地区：[1]青岛大学附属青岛市海慈医院,青岛266033

出　　处：《病毒学报》2023年第2期419-426,共8页Chinese Journal of Virology

摘　　要：乙肝病毒X蛋白(HBx)引起足细胞损伤与乙型肝炎相关性肾小球肾炎的发病有关,但具体的机制尚不清楚。miR‐340‐5p是受到HBx调控的miR,能够靶向细胞程序性死亡基因4(PDCD4)基因发挥神经元保护作用。本研究观察了过表达HBx的足细胞中miR‐340‐5p及PDCD4表达的变化及生物学意义。培养小鼠足细胞系MPC5后转染HBx质粒、miR‐340‐5p、si‐PDCD4,MTS法检测细胞增殖活力OD490、TUNEL法检测细胞凋亡率、荧光定量PCR检测miR‐340‐5p的表达、Western blot检测PDCD4的表达、双荧光素酶报告基因实验验证miR‐340‐5p靶向PDCD4基因3’UTR。结果显示,与对照组比较,HBx组细胞中miR‐340‐5p的表达、OD490水平降低,PDCD4的表达、凋亡率增加;与HBx组比较,HBx+miR‐340‐5p组细胞中miR‐340‐5p的表达、OD490水平增加,PDCD4的表达、凋亡率降低,HBx+si‐PDCD4组细胞中OD490水平增加,PDCD4的表达、凋亡率降低,miR‐340‐5p的表达无明显改变;PDCD4基因3’UTR中有miR‐340‐5p的结合位点,miR‐340‐5p降低PDCD4基因野生型3’UTR的荧光活力、不影响PDCD4基因突变型3’UTR的荧光活力。以上结果表明,过表达HBx的足细胞中miR‐340‐5p表达减少,进而可能通过增加PDCD4的表达引起足细胞损伤。本研究创新点为探究了HBx引起足细胞损伤的分子机制,国内首次发现miR‐340‐5p靶向PDCD4在HBx引起足细胞损伤中的作用,为今后研究HBV‐GN的发病机制提供了参考。Podocyte injury induced by hepatitis B virus X protein (HBx) is related to the pathogenesis of hepatitis B-associated glomerulonephritis,but the specific mechanism is still unclear.miR-340-5p is a miR regulated by HBx that targets the programmed cell death gene 4 (PDCD4) gene,and plays a neuroprotective role.In this study,we analyzed expression of mir-340-5p and PDCD4 in podocytes overexpressing HBx,and biological significance.The MPC5 mouse podocyte line was transfected with HBx plasmid,miR-340-5p,and si-PDCD4.Cell proliferation was assessed by the MTS method,apoptosis rate was measured by TUNEL assay,mir-340-5p expression was detected by fluorescence quantitative PCR,PDCD4 expression was analyzed by western blotting,and the miR-340-5p-targeted PDCD4 gene 3′-UTR was verified by double luciferase reporter gene assay.The results showed that compared with the control group,expression of miR-340-5p and the OD490value were decreased in the HBx group,while expression of PDCD4 and the apoptosis rate were increased;compared with the HBx group,expression of miR-340-5p and the OD490value were increased in HBx+miR-340-5p group,while expression of PDCD4 and the apoptosis rate were decreased.The OD490value was increased,while expression of PDCD4 and the apoptosis rate were decreased,and expression of miR-340-5p showed no significant change in the HBx+si-PDCD4.A binding site for miR-340-5p was identified in the 3′UTR of the PDCD4 gene.miR-340-5p decreased the fluorescence activity of the wild-type 3′UTR of the PDCD4 gene,but did not affect the fluorescence activity of the mutant type 3′UTR of the PDCD4 gene.These results suggest that expression of miR-340-5p in podocytes overexpressing HBx was decreased,which may lead to podocyte injury by upregulating PDCD4.This study explored the molecular mechanism responsible for podocyte injury induced by HBx,and the results indicate a role for miR-340-5p targeting PDCD4 in podocyte injury induced by HBx,which provides a reference for studying the pathogenesis of HBV-GN in the fu

关 键 词：乙型肝炎相关性肾小球肾炎(HBV-GN) 乙肝病毒X蛋白(HBx) 足细胞 miR-340-5p PDCD4 

分 类 号：R373.9[医药卫生—病原生物学] R373.2[医药卫生—基础医学]