摘要
目的评价抗原85B-6kDa早期分泌靶抗原融合蛋白(AE)亚单位疫苗经黏膜免疫小鼠诱导的免疫应答及其对结核分枝杆菌(MTB)感染的保护力。方法分别用AE、AE联合环二腺苷酸(c-di-AMP)亚单位疫苗经鼻黏膜免疫小鼠,ELISA检测抗体水平以及1型辅助T(Th1)细胞的细胞因子γ干扰素(IFN-γ)、白细胞介素2(IL-2)和Th2细胞的IL-10分泌水平,实时荧光定量PCR检测IFN-γ、IL-2、IL-10和肿瘤坏死因子α(TNF-α)的mRNA水平。MTB静脉感染免疫小鼠,ELISA检测感染小鼠血清抗体及脾细胞细胞因子分泌水平,HE染色分析肺病理改变,平板法计数菌落形成单位(CFU)检测脾和肺荷菌数。结果AE和AE联合c-di-AMP经鼻黏膜免疫可诱导小鼠产生高水平的特异性抗体,促进脾细胞增殖、脾和肺脏Th1/Th2型细胞因子和TNF-α转录增加、脾Th1/Th2型细胞因子分泌增加。小鼠MTB感染后,与对照组相比,AE和AE联合c-di-AMP免疫小鼠特异性抗体水平仍升高,Th1/Th2型细胞免疫应答增强,肺组织呈炎症反应,肺、脾荷菌数显著降低,且AE联合c-di-AMP免疫小鼠肺、脾荷菌数进一步降低。结论AE亚单位疫苗经黏膜接种可诱导小鼠产生体液与细胞免疫应答,并提供对MTB感染的保护力;c-di-AMP为佐剂可在一定程度上提高AE的免疫原性。
Objective To identify the immune responses induced by subunit vaccine of Ag85B-ESAT-6(AE)fusion protein by mucosal route and the protection against Mycobacterium tuberculosis(MTB)infection in mice.Methods AE and AE with c-di-AMP as adjuvant were inoculated intranasally in mice.The generation of specific IgG,cytokines secreted by Th1 cells(IFN-γ,IL-2)and Th2 cells(IL-10)were detected by ELISA.The transcriptional levels of IFN-γ,IL-2,IL-10,and TNF-αwere determined using real-time quantitative PCR.After MTB infection by vein,the antibodies level in mice sera and cytokines secretion of splenocytes were detected by ELISA.Histopathological changes in mice lung was illustrated by HE staining,and bacteria burdens of spleen and lung were counted by colony-forming units(CFUs)on plate.Results AE and AE combined with c-di-AMP via nasal mucosal immunization could induce high level specific antibodies in sera,promote splenocyte proliferation,and lead to increased Th1/Th2 cytokines and TNF-αtranscription in spleen and lung,and secret more Th1/Th2 cytokines in spleen.After MTB infection,compared with the control group,the specific antibody levels of AE and AE combined with c-di-AMPimmunized mice still increased,with enhanced the Th1/Th2 cellular immune responses,inflammatory response in the lung tissues,and reduced bacteria loads in spleen and lung,especially in mice immunized with AE combined with c-di-AMP.ConclusionIntranasal mucosal vaccination of AE subunit vaccine can induce humoral and cellular immune responses,and provide protection against MTB infection in mice,c-di-AMP as an adjuvant can improve the immunogenicity of AE to a certain extent.
作者
宁唤唤
张芳琳
康健
王立飞
路延之
任瑞
白鹭
梁璇
谢燕玲
柏银兰
NING Huanhuan;ZHANG Fanglin;KANG Jian;WANG Lifei;LU Yanzhi;REN Run;BAI Lu;LIANG Xuan;XIE Yanling;BAI Yinlan(School of Basic Medicine,Department of Microbiology and Pathogen Biology,Air Force Medical University,Xi'an 710032;School of Life Sciences,Yan'an University,Yan'an 716000;Key Laboratory of Resources Biology and Biotechnology in Westerm China,Ministry of Education,College of Life Sciences,Northwest University,Xi'an 710069,China)
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2022年第10期886-892,共7页
Chinese Journal of Cellular and Molecular Immunology
基金
国家“十三五”重大传染病专项课题(2018ZX10302302002004)
国家自然科学基金(81971560,81671638,81371774)
陕西省重点研发计划(2022ZDLSF01-07,2019ZDLSF02-04)。
