基于甲病毒复制子的PRRSV DNA疫苗在小鼠体内诱导的免疫应答  

Immune Responses Induced by the DNA Vaccines Based on the Alphavirus Replicon against Porcine Reproductive and Respiratory Syndrome in Mice

作　　者：梁欠欠[1] 侯绍华[1] 贾红[1] 袁维峰[1] 郭晓宇[1] 朱鸿飞[1] 

机构地区：[1]中国农业科学院北京畜牧兽医研究所动物医学研究室,北京100193

出　　处：《畜牧兽医学报》2013年第11期1805-1811,共7页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家高技术研究发展计划(863)项目(2012AA101302);农业部动物疫情监测与防治项目;国家科技支撑计划重点项目(2010BAD04B02);公益性行业(农业)科研专项经费项目(200903027)

摘　　要：本研究旨在构建含PRRSV FZ06A株GP3、GP5、M蛋白修饰后基因的复制子疫苗pSCA-Km5、pSCAVPm3、pSCA-VPm5、pSCA-VP6和pSCA-V56,研究其特异性表达目的蛋白的能力和在小鼠体内诱导免疫应答的能力。将构建的复制子质粒和空载体以100μg·只-1的剂量免疫小鼠,免疫3次,监测血清中特异性抗体和中和抗体水平及淋巴细胞经特异性抗原刺激后的增殖情况和释放IFN-γ、IL-4的水平。结果显示,各疫苗均能在转染细胞中高效表达目的蛋白。免疫小鼠,pSCA-VPm3、pSCA-VP6能引起较好的细胞免疫应答,但血清中未检测到特异性抗体。pSCA-Km5、pSCA-VPm5血清中能检测到低水平特异性抗体,但引起的细胞免疫较pSCA-VPm3、pSCAVP6低。pSCA-V56和pSCA-VPm5-PEI均能诱导细胞免疫反应和体液免疫反应,但体液免疫水平低于pSCAVPm5。pSCA-VPm5-PEI抗体水平在首免后7周突然降低,至首免后10周,无特异性抗体再次产生。所有免疫小鼠中未能检测到中和抗体。结果表明,构建的基于甲病毒复制子的PRRS DNA疫苗能诱导动物机体产生一定水平的细胞免疫和体液免疫,有望成为具有开发价值的PRRS标记疫苗。但各目的基因所能诱导机体产生体液免疫和细胞免疫的能力不同。Abstract： We have constructed five recombinant plasmids contain modified GP3, GP5 and M genes of PRRSV based on the alphavirus replicon vector, pSCA1, named pSCA-Km5, pSCA- VPm3, pSCA-VPm5, pSCA-VP6 and pSCA-V56. And we studied the expression characteristics in vitro. Expression of proteins was confirmed by western-blotting. In order to evaluate the im- munogenicity of the five plasmids in BALB/c mouse model, we detected the antibody, IL-4 and IFN-y level by ILISA, valued the lymphocyte proliferation activity by MTS staining assay. For the group of pSCA-VPm3, pSCA-VP6, specific lymphoproliferative responses to the PRRSV stimulation were induced in the splenocytes of the immunized mice as demonstrated by MTS stai- ning assay, and antigen specific IFN-y was detected in the splenocytes by cytokine ELSIA. For the group of pSCA-Km5, pSCA-VPm5 and pSCA-VPm5-PEI, low-level of specific antibody was detected by ELSIA, and antigen specific IL-4 was detected in the splenocytes by cytokine ELSIA. While, the antibody and IL-4 level of the pSCA-Km5 is lower than that of the pSCA-VPmS, which indicated that VP22 gene have transduction capability. The antibody level of pSCA-VPmS- PEI was reduced after 7 weeks, which may because of its toxicity to cells. No neutralizing anti- body was detected in the mice. Analysis of the data suggests that the recombined plasmids have well immunity and the capability inducing humoral and cellular immune responses in the mice, in- dicating that alphavirus replicon-vectored DNA-based vaccine can be potential marker vaccine against PRRS. And the statistics indicate the genes have different ability in inducing immune re- sponse.

关 键 词：甲病毒复制子 PRRSV 小鼠 免疫应答 

分 类 号：S852.659.6[农业科学—基础兽医学]