表达猪瘟病毒E2蛋白的BacMam病毒的构建及其免疫原性分析

Generation of a Recombinant Baculovirus Expressing The E2 Protein of Classical Swine Fever Virus and Its Immunogenicity in a Mouse Model

含具有哺乳动物细胞活性的启动子的重组杆状病毒(BacMam病毒)可有效转导多种哺乳动物细胞,并被广泛用于开发新型非复制型载体疫苗.将水泡性口炎病毒G蛋白(VSV-G)基因插入多角体启动子下游,得到经修饰的杆状病毒转移载体,将对虾白斑综合症病毒(WSSV)ie1启动子控制下的猪瘟病毒E2基因表达盒插入此载体中,构建了BacMam病毒BacMam/G-ie1-E2,以其感染Sf9细胞和转导HeLa细胞,通过间接免疫荧光试验和Western blot分析检测E蛋白的表达,同时用BacMam病毒直接免疫小鼠,用检测猪瘟病毒抗体的间接ELISA方法检测免疫小鼠血清抗体,用基于CFSE和WST-8的淋巴细胞增殖试验评价其细胞免疫应答.结果显示,BacMam/G-ie1-E2能同时在昆虫细胞和哺乳动物细胞中高效表达E2蛋白,免疫小鼠能诱导产生针对猪瘟病毒的特异性抗体,免疫小鼠脾细胞经猪瘟病毒刺激后能诱导特异性的淋巴细胞增殖.这表明,由BacMam病毒介导的基因转移有望用于开发针对猪瘟病毒的非复制型载体疫苗.

Baculovirus-mediated gene transfer into mammalian cells has been used to develop non-replicative vector vaccines against a number of diseases in several animal models. A baculovirus pseudotyped with the glycoprotein of vesicular stomatitis virus was used as vector to construct the recombinant baculovirus expressing classical swine fever virus （CSFV） E2 protein under the control of iel promoter from white spot syndrome virus. The E2 gene was shown to be efficiently expressed in both insect and mammalian cells. Intramuscular injection of mice with the recombinant baculovirus resulted in the production of high-level CSFV-specific antibodies. Specific lymphoproliferative responses to the CSFV stimulation were induced in the splenocytes of the immunized mice as demonstrated by CFSE staining assay and WST-8 assay. The results indicates that the pseudotyped baculovirusdelivered gene can be a potential non-replicative vaccine against CSFV infection.