流感疫苗株为载体的重组HAdV疫苗候选株构建及免疫效果  

Influenza virus vaccine expressing human adenovirus epitopes induce protective antibodies in mice

作　　者：高雅倩[1] 高雅倩[2] 段跃强[2] 邢丽[2] 杨鹏辉[2] 王希良 GAO Yaqian DUAN Yueqiang XING Li YANG Penghui WANG Xiliang(Anhui Medical University, Hefei 230032, China Beijing Institute of Microbiology and Epidemiology, State Key Laboratory of Pathogen and Biosecurity, Academy of Military Medical Sciences, Beijing 100071, China)

机构地区：[1]安徽医科大学,230032 [2]军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室,100071

出　　处：《免疫学杂志》2016年第12期1073-1077,共5页Immunological Journal

基　　金：国家自然科学基金(30972614)

摘　　要：目的以冷适应、减毒流感活疫苗株为载体,构建并拯救喷鼻重组HAd V疫苗候选株,并评价其免疫效果,为腺病毒候选疫苗研制提供实验数据。方法应用反向遗传学技术,以冷适应、减毒流感活疫苗A/Ann Arbor/6/60ca(H2N2)株为骨架,与季节性流感病毒A/California/07/209疫苗株的血凝素(haemagglutination,HA)及插入HAd V六邻体蛋白抗优势原表位的2个重复序列经改造的神经氨酸酶(neuraminidase,NA)基因构建流感病毒八质粒系统,转染COS-1/MDCK细胞筛选疫苗候选株并鉴定;制备的候选疫苗株滴鼻免疫Balb/c小鼠,通过中和抗体、s Ig A黏膜抗体及细胞因子的测定评价其免疫原性,并用野生HAd V-7病毒株攻击评价免疫保护效果。结果成功拯救获得重组HAd V疫苗候选株,命名为rg Flu/HAd V-Ca,其形态符合流感病毒典型特征,可有效表达HAd V病毒Hexon优势表位,且具有冷适应、减毒表型。小鼠免疫后可产生针对HAd V-7及H1N1流感病毒的特异性中和抗体,肺鼻灌洗液中可检测到针对HAd V-7中和抗体s Ig A抗体,并可检测到脾淋巴细胞分泌HAd V-7特异性细胞因子IFN-γ、IL-4;攻毒实验显示,rg Flu/HAd V-Ca疫苗候选株可有效降低小鼠肺病毒载量。结论重组冷适应、减毒HAd V活疫苗株rg Flu/HAd V-Ca具有良好的免疫效果,为腺病毒候选疫苗的研制提供了新思路。Human adenovirus（HAd V） is an important pathogen of respiratory diseases that cause seriousinfections and even death in children and the immunocompromised people. At present, there is no specific drug ofadenovirus, and vaccine is still the most effective measure to prevent it. In this study, we used reverse geneticstechnique, inserted two copies of adenovirus hexon epitope into the influenza NA protein gene and successfullyrescued a cold adapted recombinant virus, termed as rg Flu/HAd V-Ca. Western blotting and transmission electronmicroscope analysis showed that the recombinant virus expressed the specific protein of adenovirus hexon and hadtypical morphology of influenza virus. rg Flu/HAd V-Ca also had the identical cold adapted phenotype with H1N1,the cold adapted vaccine strain. After immunization with the purified rg Flu/HAd V-Ca intranasally, Balb/c micedemonstrated neutralizing antibodies against HAd V-7 and influenza in serum, high titer antibodies（s Ig A） againstHAd V-7 viruses in lavage fluid of lung or nasal, and specific cytokines（IFN-γ and IL-4） in spleen lymphocyte.These data implied that the candidate vaccine strain rg Flu/HAd V-Ca elicited nice immune response includinghumoral immunity, cellular immunity and mucosal immunity. Furthermore, the HAd V-7 viral loads in lung weresignificantly reduced, which demonstrated that the candidate vaccine rg Flu/HAd V-Ca provided effective protectionagainst HAd V-7 infection. The research provides a novel method for the study of adenovirus vaccine.

关 键 词：腺病毒 冷适应、减毒活疫苗 六邻体抗原表位 免疫效果 

分 类 号：R392-33[医药卫生—免疫学]