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目的探讨日本血吸虫疫苗候选分子—钙离子激活的中性蛋白激酶(Calpain)的保护性免疫力和免疫保护性机制。方法用重组纯化的Calpain抗原免疫BALB/c小鼠,制备抗Calpain血清,抗Calpain血清与机械转化的日本血吸虫童虫和来自于同种鼠激活的嗜酸性粒细胞共同培养,观察细胞对血吸虫童虫的粘附及对童虫的杀伤效果;RT-PCR分析Calpain抗原免疫BALB/c小鼠一氧化氮激酶(iNos.eNos.nNos)mRNA的表达;ELISA分析免疫小鼠体外培养脾细胞产生的细胞因子IFN-γ和IL-4。结果重组Calpain抗原免疫小鼠后,产生了一个极高的抗Calpain特异性抗体,此抗体介导了同种鼠的嗜酸性粒细胞对日本血吸虫童虫的粘附,37℃、5%CO2培养48h后,33.8%的日本血吸虫童虫被杀死,与控制组14.5%的自然死亡率相比有一个显著性的增高(P<0.01);RT-PCR测定重组Calpain抗原免疫1周小鼠iNosmRNA的表达显著性的增强,培养24h脾细胞上清中IFN-γ浓度高于对照组。结论Calpain特异性抗体介导了对日本血吸虫童虫的细胞毒作用,Calpain抗原能够诱导IFN-γ的产生,提示Calpain诱导Th1的免疫应答反应来抗日本血吸虫的感染。
Objective To investigate the protective immunity and immunoprotective mechanism of Schistosoma japonicum vaccine candidate molecule Calpain activated by calcium ion. Methods BALB / c mice were immunized with recombinant purified calpain antigen, anti-Calpain serum was prepared, anti-Calpain serum and mechanically transformed Schistosoma japonicum Schistosoma japonicum were co-cultured with eosinophils activated by the same kind of rat, And the killing effect on the parasites. The expression of iNos.NNos mRNA in BALB / c mice immunized with Calpain antigen was analyzed by RT-PCR, and the splenocytes Cytokines IFN-γ and IL-4. Results After immunization of mice with recombinant Calpain antigen, a very high anti-Calpain-specific antibody was produced and this antibody mediated the adhesion of eosinophils of the same kind to Schistosoma japonicum, 5% CO2 at 37 ℃ After 48 h, 33.8% of Schistosoma japonicum schistosomulas were killed, which was significantly higher than that of control group (14.5%) (P <0.01). The iNos mRNA of mice was immunized with recombinant Calpain antigen by RT-PCR The expression of IFN-γ in supernatant of cultured spleen cells was higher than that of control group. Conclusion Calpain-specific antibodies mediate the cytotoxic effect on Schistosoma japonicum, and Calpain antigen can induce the production of IFN-γ, suggesting that Calpain induces Th1 immune response to resist Schistosoma japonicum infection.