摘要
目的:基于脂质多聚复合物(lipopolyplex,LPP)递送平台和保守抗原开发的新型流感病毒mRNA疫苗在小鼠体内的免疫原性评价。方法:将4个拷贝的甲型流感病毒基质蛋白2胞外域(extracellular domain of matrix 2 protein,M2e)与核蛋白(nucleoprotein,NP)编码基因按照真核密码子优化,串联构建融合抗原并体外转录为4M2eNP-mRNA,利用LPP技术平台制备甲型流感病毒mRNA疫苗(LPP-4M2eNP)。转染真核细胞后通过免疫荧光体外验证NP和M2e表达;BALB/c小鼠经肌肉注射10μg、30μg mRNA疫苗,间隔4周加强免疫一次,酶联免疫法检测免疫后血清抗体滴度,免疫斑点法检测细胞免疫应答。结果:间接免疫荧光检测表明4M2eNP-mRNA转染真核细胞后NP和M2e正确表达。单针免疫可明显诱导抗原(NP和M2e)特异性细胞免疫应答,加强免疫后在小鼠中诱发了抗原特异性体液免疫应答呈Th1型偏向;NP细胞免疫应答显著提高,但M2e细胞免疫应答强度无明显改变。结论:本研究制备的LPP-4M2eNP疫苗在小鼠中单针免疫即可诱导较强的细胞免疫应答,加强免疫后可诱导明显的体液和细胞免疫应答,表明该疫苗具有较好的研发和应用前景。
Objective To evaluate the immunogenicity of a novel influenza virus mRNA vaccine based on conserved antigens delivered by lipopolyplex(LPP)platform in a mouse model.Methods Four copies of genes coding for extracellular domain of matrix 2 protein(M2e)and nucleoprotein(NP)of influenza A virus were synthetized after codon optimization.The fusion antigens were transcribed in vitro and delivered by LPP platform,named as LPP-4M2eNP.Expression of M2e and NP in eukaryotic cells was detected by immunofluorescence assay(IFA).BALB/c mice were inoculated intramuscularly twice with 10μg or 30μg LPP-4M2eNP vaccine at an interval of four weeks.Antibody response was detected by ELISA and cellular-mediated immunity(CMI)was detected by enzyme-linked immunospot assay(ELISPOT).Results IFA showed that NP and M2e were expressed correctly in eukaryotic cells.Single dose immunization could induce significant antigen(NP,M2e)-specific CMI and antigen(NP,M2e)-specific antibody response was induced in mice with Th1 type bias after boost immunization.Moreover,NP-specific CMI was increased significantly after the second immunization,while no significant change in M2e-specific CMI was observed.Conclusions Stronger CMI was triggered in mice by single dose of LPP-4M2eNP vaccine.Furthermore,robust humoral and cellular immune responses were induced after boost immunization.This study suggested that LPP-4M2eNP vaccine,which based on conserved antigen of influenza A and delivered by LPP platform,had great potential for development and application.
作者
郭俊佳
王文玲
邓瑶
黄保英
叶飞
阿茹罕
王娜
孙新蕾
谭文杰
Guo Junjia;Wang Wenling;Deng Yao;Huang Baoying;Ye Fei;A Ruhan;Wang Na;Sun Xinlei;Tan Wenjie(Zhejiang Provincial Key Laboratory of Medical Genetics,School of Laboratory Medicine and Life Science,Wenzhou Medical University,Wenzhou 325035,China;National Health Commission Key Laboratory of Biosafety,National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China)
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2022年第3期209-215,共7页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金(82041041)
国家重点研发计划项目(2020YFC0842200,2020YFC0860300)。
