DNA疫苗诱导健康小鼠细胞免疫及HBV转基因小鼠抗-HBs产生

DNA vaccine induced cellular immunity in healthy mice and HBsAg serum conversion in HBV transgenic mice

目的 :在HBVDNA疫苗成功地诱导健康小鼠体液免疫应答的基础上 ,探讨其作为抗 HBV治疗的可行性及作用机理。方法 :应用基因重组技术 ,构建编码HBV中蛋白 (preS2 +S)及人白细胞介素融合蛋白基因的真核表达质粒pS2 .S及pFP ,继经肌注免疫健康Balb C及HBV转基因 (Tg)小鼠并观察健康小鼠细胞免疫应答及HBVTg小鼠HBsAg的血清转换。结果 :1 体外HBsAg对DNA疫苗免疫后的T细胞的刺激呈浓度相关 ,HBsAg 30 μg ml时刺激pS2 .S免疫小鼠脾细胞增殖指数(5 6± 0 9)明显较pcDNA3 1组 (2 0± 0 5 )高。细胞培养上清液细胞因子水平检测结果 :免疫实验组IL 2及IFN γ的分泌水平明显较对照组高 ,而IL 4水平于各组影响不明显。 2 pS2 .S免疫小鼠局部引流淋巴结DCs诱导HBsAg致敏的T 细胞增殖指数(4 2 0 )较pcDNA3.1组 (2 5 5高 )。 3 高剂量的pS2 .S组与联合pFP组各有一只Tg小鼠分别于 2、4w开始发生HBsAg血清转换 ,且其抗体水平随时间而增长。结论 :结果表明HBVDNA疫苗能有效诱导HBsAg特异性的细胞免疫应答 ;HBV Tg小鼠初步实验结果为治疗型HBVDNA疫苗的深入研制提供了实验依据。

Objective:To investigate the feasibility and its action of the therapeutic DNA vaccine for treatment of HBV infection.Methods:By genetic recombinant technique,have constructed 2 eukaryotic expressing plasmids namely pS2.S and pFP,encoding HBV envelope middle protein(preS2+HBsAg) and human leukocyte cytokines fusion protein(IL 2/IFN γ) genes respectively and evaluated its efficacy for inducing cellular immunity in healthy BALB/c mice and HBsAg serum conversion in HBV transgenic(Tg) mice after immunization of the plasmids by intramuscular administration.Results:1.The T cell proliferation by in vitro HBsAg stimulation closely correlated with HBsAg concentration,with its stimulation index(SI=5.6±0.9) in pS2.S immunized group,being significantly higher than that (2.0±0.5) of the control.The IL 2 and IFN γ secretion level in supernatant of the DNA vaccine immunized spleen cell culture were significantly higher than that of the control,while the IL 4 secretion level was not affected.2.The dendritic cells(DCs) extracted from the local draining lymph node(LN) after DNA vaccination induced a HBsAg specific T cell proliferation with its SI(4.20) being higher than that(2.55) of the control.3.In each group of high dose pS2.S and the middle dose combined with pFP inoculation,the HBsAg serum conversion occurred in one HBV Tg mouse,with the serum anti HBs level increasing as the time prolonged,while the serum HBsAg level of the rest mice were significantly lower than that of the control.Conclusion:The results suggest that HBV DNA vaccine can effectively induced cellular immunity in healthy mice;and the preliminary results of the immunized HBV Tg mice may provide an experimental evidence for further investigation of DNA vaccine for its use in treatment of HBV infection.