摘要
流感病毒的核蛋白(Nucleoprotein,NP)高度保守,具有型特异性,能够诱导产生细胞介导的免疫应答,从而起到保护作用,抵抗不同亚型流感病毒的攻击。本研究利用杆状病毒表面展示技术,将杆状病毒GP64蛋白的信号肽(Signal peptide,SP)和胞质尾(Cytoplasmic tail,CT)与甲型流感病毒(A/Hubei/1/2010(H5H1))的NP蛋白融合表达,从而将NP蛋白展示在杆状病毒的表面,并对该病毒NP蛋白的免疫原性和保护效果进行了研究。Westernblot实验证明NP蛋白可以在Sf9细胞中有效表达。免疫电镜试验显示NP蛋白已成功展示在杆状病毒表面。通过小鼠感染实验,ELISA结果证明NP疫苗可以诱导产生高水平的IgG血清抗体。ELISPOT结果表明NP蛋白的2个细胞免疫抗原表位(NP57-66IQNSITIERM和NP441-450RTEIIKMMES)可以诱导小鼠脾淋巴细胞产生IFN-γ,利用NP57-66、NP441-450和NP蛋白均可刺激CD4+和CD8+T细胞产生IFN-γ,但主要以CD8+诱导为主,表明重组病毒可以产生有效的细胞免疫反应。利用小鼠鼠肺适应株A/PR/8/34(H1N1)进行攻毒实验,结果表明该重组病毒感染小鼠后可以降低小鼠肺病毒载量,减轻肺组织损伤,减少体重下降,并可以保护50%小鼠存活。
Nucleoprotein（NP） of influenza virus is highly conserved and type-specific. NP can trigger strong cell-mediated immune responses in host and is involved in the protection against the challenges with differ- ent subtype influenza viruses. Here, NP of an avian H5N1 （A/Hubei/1/2010, HB） was expressed by bac- ulovirus surface-display technology and its immunogenicity as well as protective mechanism was investiga- ted in mice infection model. Western blot and immunolabeled electron microscopy assay showed NP was displayed on baculovirus surface. ELISA results showed NP could induce high level of anti-NP IgG in the sera from NP-Bac-inoculated mice. Two cellular immune peptides （NP57-74 IQNSITIERMVLSAFDER and NP441-458 RTEIIKMMESARPEDLSF）were identified by IFN-r ELISPOT assay. NP57-66 and NP441-450 and NP protein could be able to trigger the activation of CD4＋ and CD8＋ T cells , and the re- sponse of CD8＋ T was more predominant. The challenge study of mice-adapted virus A/PR/8/34 （HIN1） showed that NP-Bac could reduce viral load and attenuate the damage to lung tissue. 50% protection ratio against the virus could be detected.
出处
《病毒学报》
CAS
CSCD
北大核心
2013年第3期265-272,共8页
Chinese Journal of Virology
基金
国家863课题(2010AA022901)
国家科技重大专项(2009ZX10004-016)
