藏猪白细胞介素-12基因表达对猪圆环病毒2 疫苗的免疫增强效应

为研究IL-12基因在猪体内表达对PCV2疫苗免疫应答的调控作用,本研究将藏猪IL-12基因克隆至VR1020载体,壳聚糖包裹重组质粒制备成壳聚糖纳米颗粒(VRIL-12-CNP)并与PCV2疫苗共同接种21日龄断奶仔猪.接种后第0d、7d、14d和28d采集猪前腔静脉血进行血细胞分析、PCV2抗体检测和相关免疫基因表达量检测,并记录体重.结果显示:接种VRIL-12-CNP和PCV2疫苗的实验组猪的CD3~+、CD4~+、CD8~+T细胞和PCV2抗体显著增长(P0.05),TLR2/7、IL-12/4/6/15、STAT1/3和Bcl-2基因的表达量显著高于对照组(P0.05);试验期间实验组体重净增长也明显高于对照组(P0.05).结果表明:VRIL12-CNP能增强PCV2疫苗的先天性和获得性体液、细胞免疫应答,是安全、有效的PCV2疫苗佐剂.

Elevation of immunity to PCV2 vaccine by recombinant Tibetan pig interleukin-12 gene in chitosan nanoparticles

To study the regulation effect on vaccination of porcine circovirus-2 (PCV2) by co-delivering interleukin-12 (IL-12) cDNA via chitosan nanoparticles, two subunits of IL-12, P40 and P35 of Tibetan pig, were cloned into mammalian expression plasmid VR1020. VRIL-12 was transfected into HEK293 cells and PCR test was conducted. The results showed that IL-12 gene could express successfully. The recombinant plasmid was packaged into chitosan nanoparticles using ionic crosslinking method. The nanoparticles (VRIL-12-CNP) were confirmed to be nanoparticle size with good dispersion and positive charge. VRIL-12-CNP was then used to inoculate the 21-day-old piglets together with PCV2 vaccine. Blood samples were taken at day 0, 7, 14 and 28 post inoculation for analysis. The results showed that post vaccination the T cells such as CD3+, CD4+ , CD8+, and anti-PCV2 specific antibody, all increased significantly in the VRIL-12-CNP co-delivering pigs (P<0.05), the expression levels of TLR2, TLR7, IL-12, IL-4, IL-6 and IL-15 in the co-delivering group were significantly higher than those in control group (P<0.05), and the expression of STAT1, 3 and Bcl-2 gene was also increased significantly in the co-delivering pigs (P<0.05). Moreover, the net weight gain of the pigs in co-delivery group was significantly increased in the 28 days post inoculation (P<0.05). These results indicated that co-delivery of VRIL-12-CNP elevated swine innate immune response and both humoral and cellular immune response to PCV2, and VRIL-12-CNP is a promising safe and effective adjuvant to PCV2 vaccination.