猪IL-2真核表达质粒构建及对PRRSV-ORF5基因疫苗免疫佐剂作用的研究

Construction of Pig IL-2 Eukaryotic Expression Plasmid pcDNA-IL-2 and Its Adjuvant Effect on pcDNA-PRRSV-SC2-ORF5 Gene Vaccine

本研究应用真核表达载体pcDNA3.1(+)和猪白细胞介素2(IL-2)基因成功构建了IL-2的真核表达质粒(pcDNA-IL-2),探讨了pcDNA-IL-2作为分子免疫佐剂对猪繁殖与呼吸综合征(PRRS)ORF5基因疫苗(pcDNA-PRRSV-SC2-ORF5)免疫猪的增强作用。经间接ELISA法、MTT比色法及流式细胞仪分别对pcDNA-IL-2与pcDNA-PRRSV-SC2-ORF5共同免疫猪、pcDNA-PRRSV-SC2-ORF5单独免疫猪、pcDNA3.1(+)空载体和灭菌水免疫对照猪的血清抗PRRSV抗体IgG、外周血T淋巴细胞的增殖活性、CD4+和CD8+细胞比例进行检测,结果表明pcDNA-IL-2与pcDNA-PRRSV-SC2-ORF5共同免疫猪的IgG含量、外周血T淋巴细胞的增殖活性、CD4+和CD8+细胞比例与pcDNA-PRRSV-SC2-ORF5单独免疫猪相比有显著差异(P<0.05)。说明pcDNA-IL-2能显著增强pcDNA-PRRSV-SC2-ORF5基因疫苗免疫猪的体液和细胞免疫应答,可作为PRRSV基因疫苗的良好佐剂。

In this study, pig IL-2 eukaryotic expression plasmid pcDNA-IL-2 was constructed and its adjuvant effect was investigated. Four groups of pigs were immunized with sterilized water, empty vector pcDNA3. 1 （＋）, PRRSV-SC2-OFR5 gene eukaryotic expression plasmid pcDNA- PRRSV-SC2-ORF5 and a mixed solution of pcDNA-IL-2 and pcDNA-PRRSV-SC2-ORF5 respectively. Serum of pig in each group was collected at different time post inoculation, and antibody to PRRSV was measured by ELISA. Peripheral T lymphocytes were acquired from a small quantity of blood of each pig, and the activity of lymphocyte proliferation was detected by MTT methods after being cultured with RPMI1640 culture medium. Content of CD4^＋ and CD8^＋ of peripheral T lymphocytes in each groups of pigs were tested by cytospectrometry. Results showed that the antibody and peripheral T lymphocytes level, content of CD4^＋,CD8^＋ cell of the pigs immunized with both pcDNA-IL-2 and pcDNA-PRRSV-SC2-ORF5 were significantly higher than that of the pigs immunized with pcDNA-PRRSV-SC2-ORF5 alone, indicate that plasmid pcDNA-IL-2 can be used as a adjuvant to enhance PRRS gene vaccine.