丙型肝炎病毒1b基因型C区氨基酸70替代对干扰素刺激应答元件的影响  

Effects of the Amino Acid 70 Substitution in the Core Region of Hepatitis C Virus Genotype 1b on Interferon- stimulated Response Element

作　　者：胡中杰[1] 刘颖[1] 仇丽霞[1] 范作鹏[1] 聂巍[1] 梁珊[1] 于红卫[1] 金荣华[1] 

机构地区：[1]首都医科大学附属北京佑安医院丙肝与中毒性肝病科,100069

出　　处：《医学研究杂志》2014年第7期75-79,共5页Journal of Medical Research

基　　金：北京市优秀人才培养基金资助项目(2010D003034000009);中国初级卫生保健基金会佑安肝病艾滋病基金资助项目(BJYAH-2011-073);首都医科大学基础-临床科研合作基金资助项目(1000172053-11JL61)

摘　　要：目的探讨丙型肝炎病毒(hepatitis C Virus,HCV)1b基因型(genotype 1b,GT-1b)C区氨基酸(amino acid,aa)70替代对干扰素刺激应答元件(interferon-stimulated response element,ISRE)的影响。方法首先构建HCV GT-1b C区aa70野生型和变异型核心蛋白表达质粒,测序鉴定后瞬时转染肝瘤细胞系Huh7细胞,Western blot法鉴定HCV核心蛋白的成功表达;然后利用ISRE双荧光素酶报告基因实验检测核心蛋白表达对ISRE活化的影响,并以SYBR Green real-time PCR检测3种主要干扰素刺激基因(interferon stimulated genes,ISGs)的表达情况。结果构建了HCV aa70野生型和变异型核心蛋白表达质粒,并在体外培养细胞中成功表达。但野生型和变异型核心蛋白表达对ISRE活化及3种主要ISGs表达的影响无明显差异。结论单纯R70Q/H变异的核心蛋白似乎并不足以直接导致IFNα耐药。Objective To investigate the effects of the amino acid （aa） 70 substitution in the core region （CR） of hepatitis C virus （HCV） genotype 1 b （GT-1 b） on interferon-stimulated response element （ISRE）.Methods The expression plasmids of wild type and mutant aa70 core proteins were constructed and then transfected into human hepatoma cell line Huh7.Western blot was used to conform the expression of HCV core proteins.Next,dual luciferase reporter assays were performed to assess the effects of core proteins on ISRE.Furthermore,expressions of 2＇,5＇-oligoadenylate synthetase 1 （OAS1）,myxovirus resistance protein A （MxA） and protein kinase R （PKR） were tested by SYBR Green real-time PCR.Results The expression plasmids of wild type and mutant aa70 HCV core protein were constructed and expressed in Huh7 cells successfully.However,the effects of wild type and mutant aa70 core proteins on ISRE and 3 major ISGs were not different significantly.Conclusion Substitution of amino acid 70 in HCV GT-1 b core region alone is not sufficient for resistance to IFNα treatment.

关 键 词：丙型肝炎病毒 核心蛋白 干扰素刺激应答元件 干扰素刺激基因 

分 类 号：R5[医药卫生—内科学]