自噬与乙型肝炎病毒复制的关系  

Research on relationship between autophagy and hepatitis B virus replication

作　　者：王安娜[1] 封晓昆 闫晓东[2] 石英[1] 陈德喜[1] 

机构地区：[1]首都医科大学附属北京佑安医院北京市肝病研究所,北京100069 [2]山东省肿瘤医院普外科,山东济南250117

出　　处：《临床荟萃》2015年第2期210-213,共4页Clinical Focus

基　　金：国家自然科学基金面上项目(81272266);国家自然科学基金青年基金资助项目(81100288);"十二五"国家科技支撑计划项目(2012BAI15B08)

摘　　要：目的探究自噬与乙型肝炎病毒(hepatitis B virus,HBV)复制之间的相互关系。方法利用HepG2细胞,转染绿色荧光标记的微管相关蛋白轻链3(GFP-LC3),分别在正常和饥饿条件下培养,另共转染HBV WT和GFP-LC3,正常条件培养,免疫荧光观察细胞自噬;分别转染1.3mer HBV野生型质粒(HBV WT)与HBV突变型(HBV X-)质粒,正常和饥饿两种条件下培养,蛋白印迹检测自噬相关蛋白LC3和乙型肝炎病毒核心蛋白(HBc)的表达;细胞转染HBV WT后,分别正常条件培养(对照组),无血清饥饿培养(饥饿组),加入自噬抑制剂3-甲基腺嘌呤(3-MA)培养(3-MA组),采用酶联免疫吸附测定法(ELISA)检测细胞上清乙型肝炎表面抗原(HBsAg)和乙型肝炎e抗原(HBeAg)的含量。结果转染HBV WT质粒的细胞内自噬荧光颗粒较对照组明显增加发生自噬的细胞比例分别为13%和2%;饥饿组HBsAg含量(1.856±1.415)较对照组(1.531±0.944)升高,HBeAg含量(0.533±0.391)较对照组(0.334±0.110)升高,差异具有统计学意义(P<0.05);自噬抑制剂组HBsAg含量(1.184±0.759)较饥饿组(1.856±1.415)降低,HBeAg含量(0.306±0.130)较饥饿组(0.533±0.391)降低(P<0.05);蛋白印迹显示,转染HBV WT质粒的细胞中LC3蛋白表达较HBV X-组增高,且在饥饿条件下,HBc蛋白的表达增高。结论乙型肝炎病毒可诱导细胞自噬的发生,且自噬可促进乙型肝炎病毒的复制。Objective To investigate the interaction between autophagy and hepatitis B virus replication.Methods HepG2 cells were transfected with GFP-LC3,cultured under normal conditions or starvation.Another cotransfected with wild-type 1.3 mer HBV（HBV WT）and GFP-LC3,normal culture conditions.Autophagy was observed by immunofluorescence.The cells were transfected with HBV WT plasmid and mutant HBV（HBV X-）plasmid,then treated with nutrient starvation.Western blotting was used to analyze the expression of LC3 and HBc.The cells were transfected with HBV WT,cultured with normal conditions（control）,no serum（starvation）,autophagy inhibitor（3-MA）,ELISA was used to detect HBsAg and HBeAg of cell supernatants.Results A significant increase of autophagy particles in HBV WT transfected cells（13%）was found compared with that in control group（2%）;Titres of HBsAg（1.856±1.415）and HBeAg（0.533±0.391）of nutrient starvation group were higher than those of control group（1.531±0.944;0.334±0.110）（P〈0.05）;Titres of HBsAg（1.184±0.759）and HBeAg（0.306±0.130）of 3-MA group were lower than those of starvation group（1.856±1.415;0.533±0.391）（P〈0.05）;Western blotting analysis showed that the expression of LC3 in the HBV WT-transfected cells was higher than in mutant group.HBc expression increased in starvation condition.Conclusion Hepatitis B virus can induce cell autophagy.Further more,autophage can enhance HBV DNA replication.

关 键 词：肝炎病毒 乙型 自噬 病毒复制 

分 类 号：R512.62[医药卫生—内科学]