含有H5N1 NA基因重组腺病毒疫苗在小鼠体内的免疫效果研究

目的 考察含有禽流感病毒A/Anhui/1/2005(H5N1)优化型NA基因的重组腺病毒在BALB/c小鼠体内的免疫效果,并筛选合适的免疫剂量.方法 重组病毒以肌内注射方式在第0周和第4周免疫BALB/c小鼠两次,低、中、高组的免疫剂量分别是10~5、10~7和10~9 TCID_(50)/次,第5周时分别使用神经氨酸酶活性抑制试验和EHSpot实验来检测和比较疫苗的体液和细胞免疫效果.结果 重组病毒免疫后的小鼠均可检测出针对NA抗原的特异性体液和细胞免疫反应,并且免疫效果与免疫剂量呈正相关,10~7 TCID_(50)/只/次是合适的免疫剂量.另外,从包含NA全长氨基酸残基的合成肽库中筛选到两个能够刺激BALB/c小鼠T淋巴细胞分泌IFN-γ的表位,即NA_(109-124):CRTFFLTQGALLNDKH和NA_(182-199):AVAVLKYNGIITIKSW.结论 含有优化型NA基因的重组腺病毒疫苗能够诱导BALB/c小鼠同时产生NA抗原特异性的体液和细胞免疫反应,是较好的H5N1流感病毒候选疫苗株,值得进一步深入研究.

Study on immunogenicity of a recombinant adenovirus vaccine containing neuraminidase gene of H5N1 influenza virus(A/Anhui/1/2005)in mice

Objective To investigate immunity of a recombinant adenovirus vaccine(rAdV)containing codon-modified neuraminidase(Mod.NA)gene of H5N1 influenza virus in BALB/c mice and to screen for appropriate dose.Methods BALB/c mice were immunized with the rAdV-Mod.NA vaccine intramuscularly twice (double injection at 0 and 4~(th) week)in three groups,low dosage(10~5 TCI_(50) per dose),medium dosage(10~7 TCID_(50) per dose)and high dosage(10~9 TCID_(50) per dose).The effect of humoral and cell-mediated immunity were analysod at 5~th week.Results ①The rAdV-Mod.NA vaccine eould elicit both humoral and cell-mediated robust NA specific immunity in mice by neuraminidase inhibitor assay and IFN-γ ELISpot assay;②10~7 TCID_(50) per dose Was the appropriate dose;③ Peptide NA_(109-124):CRTFFLTQGALLNDKH and peptide NA_(182-199):AVAVLKYNGIITDTIKSW were the dominant epitopes for neuraminidase-immunized BALB/c mice,which was screened out from the whole length of neuraminidase of an H5N1 virus,A/Anhui/1/2005.Conclusion The recombinant adenovirus NA could induce specific humoral and cellular immune responses in BALB/c after immunization,which suggest rAdV-Mod.NA vaccine was a potential vaccine candidate against H5N1 influenza and worthy of further investigation.