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目的探讨结核分枝杆菌复苏因子D(RpfD)-DNA疫苗在BALB/c小鼠抗H37Rv感染中产生的免疫原性。方法将90只SPF级BALB/c小鼠随机分为3组(每组30只),即RpfD疫苗免疫组、BCG免疫组、生理盐水对照组,分别以RpfD-DNA疫苗、BCG和生理盐水于0、3、6周共3次免疫小鼠,于第9周用结核分枝杆菌气雾感染小鼠。于第17周采集小鼠血清,应用ELISA方法检测RpfD抗体效价、IFNγ水平,ELISA检测脾淋巴细胞培养上清诱导的IFNγ、IL-12水平,采用CCK-8检测淋巴细胞增殖指数,LDH方法检测淋巴细胞CTL杀伤效应,并对小鼠肺组织菌荷量计数。结果 RpfD疫苗免疫组小鼠血清RpfD抗体水平和IFNγ水平均显著高于生理盐水组对照组及BCG组(P<0.05或P<0.01)。RpfD体外刺激培养小鼠脾淋巴细胞释放IFNγ水平RpfD疫苗免疫组显著高于其他组(P<0.05);IL-12水平3组间差异无统计学意义(P>0.05)。RpfD疫苗免疫组淋巴细胞增殖指数和脾特异性CTL杀伤性显著高于其他组(P<0.05)。RpfD疫苗免疫组小鼠肺组织结核分枝杆菌CFU显著低于其他组(P<0.05)。结论 RpfD-DNA疫苗可诱导小鼠产生免疫应答,具有免疫原性。
Objective To investigate the immunogenicity of mycobacterium tuberculosis recovery factor D (RpfD) -DNA vaccine in anti-H37Rv infection in BALB / c mice. Methods Ninety SPF BALB / c mice were randomly divided into 3 groups (30 in each group): RpfD vaccine group, BCG group and saline control group. RpfD-DNA vaccine, BCG and saline The mice were immunized three times at 0, 3 and 6 weeks, and mice were infected with M. tuberculosis aerosol at the 9th week. The serum of mice was collected at the 17th week. The antibody titer and IFNγ level of RpfD antibody were detected by ELISA. The levels of IFNγ and IL-12 induced by splenic lymphocyte culture supernatants were detected by ELISA. The lymphocyte proliferation index and LDH were detected by CCK-8 Lymphocyte CTL killing effect was detected, and the mice lung tissue bacterial count was counted. Results The levels of RpfD antibody and IFNγ in RpfD vaccine group were significantly higher than those in control group and BCG group (P <0.05 or P <0.01). RpfD stimulated the spleen lymphocytes of mice to release IFNγ in vitro. The RpfD vaccine immunized group was significantly higher than other groups (P <0.05). There was no significant difference among the three groups (P> 0.05). RpfD vaccine immunization group lymphocyte proliferation index and spleen-specific CTL cytotoxicity was significantly higher than the other groups (P <0.05). The CFU of Mycobacterium tuberculosis in lungs of mice immunized with RpfD vaccine was significantly lower than that of other groups (P <0.05). Conclusion RpfD-DNA vaccine can induce immune response in mice and is immunogenic.