丙型肝炎病毒包膜E2蛋白DNA疫苗诱导小鼠产生中和抗体研究  

Study on the induction of mouse neutralization antibodies to HCVpp by HCV E2 DNA vaccines

作　　者：尹迪[1] 严石[1] 丁惠[2] 华显[2] 童一民[2] 王永智[2] 刘媛[2] 徐凤花[1] 赵平[2] 戚中田[2] 

机构地区：[1]东北农业大学,哈尔滨市150030 [2]第二军医大学微生物学教研室

出　　处：《中华实验和临床感染病杂志（电子版）》2008年第1期24-32,共9页Chinese Journal of Experimental and Clinical Infectious Diseases(Electronic Edition)

基　　金：上海市基础重点项目(06JC14084);国家自然科学基金(30671921;30770094);国家"863"基金(2006AA02Z401)

摘　　要：目的探讨HCV包膜E2蛋白DNA疫苗诱导小鼠产生中和抗体的可行性。方法分别构建HCV E2蛋白全长表达质粒pcDNA3.1-1a746、截除疏水性羧基末端的表达质粒pcDNA3.1-1a661以及同时截除疏水性羧基末端和高变区1(HVR1)的表达质粒pcDNA3.1-1a661Δ,转染293T细胞,以Western blot和ELISA法检测细胞内和培养上清中的HCV E2蛋白,将3种表达质粒以及空载体分别肌注免疫BALB/c小鼠,检测小鼠血清的E2及HVR1抗体,以HCV假病毒模型分析小鼠血清的中和活性。结果3种E2表达质粒均能有效表达HCV E2蛋白,其中pcDNA3.1-1a746质粒的表达产物不能分泌,而pcDNA3.1-1a661和pcDNA3.1-1a661Δ均能分泌表达E2蛋白。分泌表达E2蛋白可显著增强小鼠的抗体应答,pcDNA3.1-1a661免疫血清对HCV假病毒颗粒(HCVpp)的中和活性明显高于pcDNA3.1-1a661Δ免疫血清。pcDNA3.1-1a661免疫血清中的HVR1抗体量仅占总E2抗体的一小部分,却是中和抗体的重要成分。结论表达E2蛋白的DNA疫苗能有效诱导HCV中和抗体的产生,HVR1不仅是重要的中和抗体表位,而且能增强E2蛋白其他抗原表位的抗体应答。Objective To explore the feasibility of induction of neutralization antibodies against HCV infection by HCV envelope 2 protein DNA vaccines.MethodsThree expression plasmids of HCV envelope 2 protein,plasmid pcDNA3.1-1a746 encoding full length of E2,plasmid pcDNA3.1-1a661 encoding hydrophobic carboxyl terminal truncated E2 and pcDNA3.1-1a661Δ encoding E2 with deletion of hypervariable region 1(HVR1)and carboxyl terminal,were constructed,respectively.The 293T cells were transfected with these plasmids,respectively,and the excretion of E2 protein were analyzed by Western blot and ELISA.Then the plasmids were used to immunize BALB/c mouse intramuscularly.The sera antibodies against E2 and HVR1 were detected by ELISA and the neutralization activity of the antibodies were assayed with HCV pseudotype particle(HCVpp).Results All three plasmids could express HCV E2 protein,the expression product of pcDNA3.1-1a746 could not secrete,while that of pcDNA3.1-1a661 and pcDNA3.1-1a661Δ could secrete E2 protein into culture medium.The secretion of HCV E2 protein enhanced antibody response elicited by DNA vaccines in mice significantly.Sera from pcDNA3.1-1a661 immunized mice showed stronger neutralization activity than that from pcDNA3.1-1a661Δ immunized mice.For sera from pcDNA3.1-1a661 immunized mice,the anti-HVR1 antibodies account trivially in total anti-E2 antibodies in quantity,while play a major role in neutralization infectivity of HCVpp.Conclusions DNA vaccines that express and secrete HCV E2 protein could induce neutralization antibodies against HCVpp infection.HVR1 not only acts as a region containing neutralization antibodies epitopes,but also enhance antibody response of other epitopes presented in HCV E2 protein.

关 键 词：丙型肝炎病毒 包膜E2蛋白 DNA疫苗 高变区1 中和抗体 

分 类 号：R392[医药卫生—免疫学]