HSV-gD-IL-21 DNA疫苗免疫效应及抗鼠眼角膜HSV-1感染的初步研究

Preliminary study on the HSV-gD-IL-21 DNA vaccine and its immunological effects and immunoprotection against HSV-1 challenging on cornea in mouse model

构建共表达HSV-1型gD和鼠源性IL-21DNA疫苗,探讨其是否能诱导动物免疫应答以及抵抗眼角膜HSV-1的感染。PCR扩增HSV-1gD和IL-21基因,分别定向插入载体pRSC中,构建DNA疫苗pRSC-gD-IL-21,对其进行鉴定后再分别以pRSC-gD-IL-21、pRSC-gD、pRSC免疫小鼠3次,间隔2周,末次免疫2周后检测血清抗体、IL-4及IFN-γ水平,特异性脾细胞增殖反应以及CTL、NK细胞杀伤活性。同时评价小鼠对病毒攻击角膜的免疫保护作用。经测序、酶切和RT-PCR鉴定后表明,pRSC-gD-IL-21中目的基因在细胞内成功转录表达。与对照鼠相比,该疫苗在鼠体内产生了较强的特异性抗体、IL-4及IFN-γ、特异性脾细胞增殖反应、CTL及NK细胞杀伤活性均增强,同时疫苗使小鼠产生了较强的针对眼角膜HSV-1感染免疫保护作用。结果显示,pRSC-gD-IL-21疫苗能诱导小鼠产生较强的免疫应答及针对HSV-1眼部感染的免疫保护作用。

The DNA vaccine expressing herpes simplex virus type 1 （HSV-1） glycoprotein D（gD） and mouse interleukin-21（IL-21） was constructed,and their ability to induce immune response and to protect against the HSV-1 infection of cornea in mice was investigated. The genes of gD and IL-21 were respectively amplified with PCR and then the amplified DNA fragments were inserted into the plasmid pRSC to form the DNA vaccine pRSC-gD-IL-21. After being identified by molecular methods,the vaccine pRSC-gD-IL-21,the pRSC-gD and the blank pRSC were respectively inoculated into BALB/c mice for 3 times with 2 weeks interval. The levels of specific antibody,IL-4 and IFN-γ in serum were detected 2 weeks after the last immunization. At the same time,the proliferative response of the special splenic lymphocytes,the activities of CTL and NK were assayed,respectively. The immunoprotective against HSV-1 challenging on cornea in mice was also evaluated. The presented data showed that the targeted genes of gD and IL-21 were successfully expressed in the transfected SP2/0 cells after the DNA vaccine was identified by DNA sequencing,the restricted endonuclease analysis,and RT-PCR,respectively. Compared with the control mice,the DNA vaccine pRSC-gD-IL-21 induced mice to generate higher levels of antibody,IL-4 and IFN-γ,and enhanced the splenic cell proliferation response as well as the activities of CTL and NK. Meanwhile,the pRSC-gD-IL-21 could elicit a stronger immunoprotective effect against the corneal infection of HSV-1 in mice. These findings indicate that the DNA vaccine pRSC-gD-IL-21 may induce an effective immune response in immunized mice. More importantly,the efficacy against the HSV-1 challenge in mouse cornea was enhanced markedly.