细菌影载体负载防龋DNA疫苗免疫小鼠的效果研究

摘要

目的 研制鼠伤寒沙门菌细菌影,负载防龋DNA疫苗,探寻增强防龋DNA疫苗黏膜免疫效能的方法.方法将pREP4质粒和噬菌体PhiX基因E表达质粒同时转入鼠伤寒沙门菌减毒株J357中,加入异丙基硫代半乳糖苷(isopropyl β-D-1-thiogalactopyranoside,IPTG)诱导,收集洗涤,负载防龋DNA疫苗,分4组经鼻免疫小鼠,分别为:细菌影+pGJGLU/VAX组,细菌影负载5μg防龋DNA疫苗pGJGLU/VAX;细菌影+pVAX1组,细菌影负载5μg空载体pVAX1;pGJGLU/VAX组,布比卡因包裹5 μg pGJGLU/VAX; pVAX1组,布比卡因包裹5μg pVAX1.酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)法检测唾液抗体产生结果.结果 ELISA结果显示经鼻黏膜免疫鼠伤寒沙门菌J357细菌影负载的防龋DNA疫苗pGJGLU/VAX(细菌影+pGJGLU/VAX组)后,诱导了显著的唾液特异性抗葡聚糖结合区的IgA抗体,(x)±sx为(0.367 ±0.086) A/μg,与细菌影+pVAX1组[(0.122±0.077)A/μg]、pGJGLU/VAX组[(0.068±0.068)A/μg]和pVAX1组[(0.089±0.089) A/μg]相比,差异均有统计学意义(P值分别为0.028、0.012和0.030).结论 成功制备了鼠伤寒沙门菌细菌影,负载防龋DNA疫苗后经鼻黏膜途径免疫小鼠能有效提高免疫效能.%Objective To develop an anti-caries DNA vaccine-loaded Salmonella typhimurium(St) ghost and enhance the efficacy of immune responses induced by anti-caries DNA vaccine via mucosal route.Methods Both pREP4 and PhiX gene E expression plasmids were transformed into StJ357 and then induced with isopropyl β-D-1-thiogalactopyranoside (IPTG).The bacterial ghosts (BG) were collected after wash and loaded with anti-caries DNA vaccine pGJGLU/VAX.Mice were divided into four groups and immunized through the nasal route with pGJGLU/VAX-loaded BG(Group Ghost + pGJGLU/VAX),pVAX1-loaded BG (Group Ghost + pVAX1),pGJGLU/VAX-Bupivacaine complex (Group pGJGLU/VAX) and pVAX1-Bupivacaine complex (Group pVAX1),respectively.Enzyme-linked immunosorbent assay (ELISA) was used to evaluate the immune responses.Results ELISA results showed that group Ghost + pGJGLU/ VAX had significantly higher level of specific anti-GLU SIgA antibody [(0.367 ± 0.086) A/μg] compared with group Ghost + pVAX1 [(0.122 ± 0.077) A/μg],Group pGJGLU/VAX [(0.068 ± 0.068) A/μg] or Group pVAX1 [(0.089 ±0.089) A/μg] (P =0.028,0.012 or 0.030,respectively).Conclusions St ghost was developed successfully,which enhanced the efficacy of immune responses induced by anti-caries DNA vaccine pGJGLU/VAX via the nasal route.