治疗型HBV基因疫苗免疫效果的研究

STUDY ON IMMUNE RESPONSE TO THERAPEUTIC HBV DNA VACCINE

为探索治疗型HBV基因疫苗治疗慢性乙型肝炎的可行性 ,自行构建了HBV包膜蛋白前S2 ·S基因及人IL 2和IFN γ融合蛋白基因真核表达质粒 ,并将双质粒融合蛋白作为佐剂组合成治疗型HBV基因疫苗 ,在健康小鼠、HBV转基因 (Tg)小鼠、新西兰兔和恒河猴体内进行了免疫效果试验。结果发现 :①特异性CTL活性提高 ;②对HBsAg特异性T细胞增殖能力提高 ,HBsAg 3 0 μg /ml对pcS2 ·S免疫的小鼠脾细胞的刺激指数 (SI =5 6± 0 9)明显较空白质粒pcDNA3 1组 (SI =2 0± 0 5 )为高 (P <0 0 1) ,免疫组的IL 2 /IFN γ分泌水平为 (2 2 6 3± 41 0 5pg/ml) /(5 1 1± 7 7pg/ml) ,明显较空白组的 (69 0± 2 2 1pg/ml) /(0 9± 0 7pg/ml)为高 (P <0 0 1) ;③HBV基因疫苗免疫健康小鼠局部引流淋巴结中DCs的诱导HBsAg致敏T细胞增殖指数 (4 2 0 )较pcDNA3 1组 (2 5 5 )为高 ;⑤用在体电脉冲法注射治疗型HBV基因疫苗后 ,检测血清抗 HBs水平 ,无论是小鼠、兔还是猴均有明显提高。提示该治疗型HBV基因疫苗能较好地诱导体液和细胞免疫 。

In this study,we constructed 2 eukaryotic expressing plasmids namely pcS2·S and pFP,encoding HBV preS2+S envelope protein and human IL 2/IFN γ fusion protein, respectively. The double plasmid fusion protein was used as an adjuvant in preparation of a treatment type HBV gene vaccine. To investigate its feasibility for use as a therapeutic DNA vaccine, we've evaluated the immune response after injection into healthy mice, HBV transgenic(Tg) mice, New Zealand rabbits and Rhesus monkeys. The results showed that the therapeutic DNA vaccine can improve: (1)the CTL activity; (2)the HBsAg(30μg/ml) specific T lymphocyte proliferation in which the stimulation index(SI) and cytokines(IL 2/IFN r) release levels of the pS2.S immunized group(SI=5.6±0.9; 226.3±41.1/51.1±7.1pg/ml) were significantly higher( P <0 01) than those of the blank plasmid (pcDNA3.1) group (SI=2.0±0.5; 69.0± 22.1/0.9 ±0.7pg/ml,respectively); (3) the dendric cells(DCs) isolated from the local draining lymph node after vaccination induced higher HBsAg specific T cell proliferation(SI=4.20) than that of the control group(SI=2 55);(4) the serum anti HBsAg level increased in both healthy mice and the higher body weight animals like rabbits and monkeys after we application of in vivo electroporation following intramuscular administration.These results suggest that the therapeutic HBV DNA vaccine contructed by us can successfully induce both humoral and cellular immune response, thus providing an experimental evidence for its use in treatment of HBV infection.