流感病毒神经氨酸酶双表达DNA疫苗的构建及在小鼠中的免疫保护研究

目的研究对多亚型流感病毒流行提供预防的DNA疫苗。方法以pIRES为双表达载体,在其两个多克隆位点中分别插入H1N1（A/PR/8/34）和H3N2（A/Guizhou/54/89）的NA DNA片段,构建双表达质粒pN1-IRES-N2及pN2-IRES-N1。以BALB/c小鼠为模型,采用电穿孔法进行免疫,并用致死量流感病毒感染以检测双表达载体的保护力。结果实验表明pN1-IRES-N2能完全保护小鼠抵御致死量H1N1同源病毒的攻击,部分保护小鼠抵御致死量H3N2同源病毒攻击；pN2-IRES-N1能完全保护小鼠抵御致死量H3N2同源病毒攻击,对致死量H1N1同源病毒的攻击提供部分保护。结论双表达DNA疫苗可望开发为一种提供广泛保护的新型流感疫苗。

Construction of co-expression DNA vaccine containing influenza neuraminidases and the immune response in mice

Objective In order to efficiendy deliver multiple antigens with the use of one plasmid, new antigen delivery systems must be assessed. Methods A bicistronic vector construct was utilized, containing an internal ribosome entry site （IRES） ,expressing a combination of neuraminidase （NA） from influenza virus A/PR/8/34 （H1N1） and A/Guizhou/ 54/89 （H3N2）. Female BALB/c mice aged 6-8 weeks were immunized twice with 60μg pN1-IRES-N2 or pN2-IRES-N1 at a 3-week interval by electroporation. The mice were challenged with a lethal dose of influenza H1N1 or H3N2 virus one week after the second immunization. The protections of DNA vaccines were evaluated by the serum antibody titers, residual lung virus titers, and survival rates of the mice. Results It was shown in our study that pN1-IRES-N2 provided complete protection against H1 N1 influenza virus challenge, but only partial protecion against H3 N2 influenza virus infection was provided. Mice immunized with pN2-IRES-N1 survived 100% after H3N2 influenza virus challenge ,but only 60% after HI N1 influenza virus infection. Conclusion These results suggested that IRES vectors can produce polycistronic mRNAs to produce two foreign proteins.