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目的探讨含非甲基化胞嘧啶和鸟嘌呤核苷酸序列的寡脱氧核糖核苷酸(CpG-ODN)对卡介苗(BCG)的免疫增强作用,为提高BCG的免疫效果提供实验依据。方法 (1)细胞实验:分空白组、低浓度组和高浓度组,均接种小鼠巨噬细胞株Ana-1预培养。分别加入培养液、低浓度CpG-ODN和高浓度CpG-ODN,培养后测定上清液中IL-12、IL-18和IFN-γ含量。(2)动物实验:分对照组(A组)、感染组(B组)、BCG预防组(C组)和BCG+CpG-ODN预防组(D组)。A组常规饲养,B、C、D组分别注射无菌PBS、BCG、BCG和CpG-ODN混合液,2周后B、C、D组注射结核分枝杆菌,再2周后处死所有小鼠,取肺组织病检,收集血清测定IL-12、IL-18和IFN-γ含量。结果 (1)细胞实验:CpG-ODN刺激培养下,Ana-1分泌IL-12、IL-18和IFN-γ均呈CpG-ODN剂量依赖性,加入CpG-ODN含量越高,Ana-1分泌上述三种Th1型细胞因子含量越多。(2)动物实验:A组肺组织未见炎症细胞浸润;B组肺组织可见大量炎症细胞浸润;C组和D组肺组织可见少量炎症细胞浸润。B组小鼠血清中IL-12、IL-18和IFN-γ含量均显著低于A组小鼠(均P<0.05)。C组和D组小鼠血清中IL-12、IL-18和IFN-γ含量均显著高于A组(均P<0.05)。D组小鼠血清中IL-12、IL-18和IFN-γ含量均显著高于C组(均P<0.05)。结论 CpG-ODN能够增强BCG的免疫预防效果,该免疫增强作用可能与CpG-ODN上调Th1型细胞因子含量有关。
OBJECTIVE: To investigate the immune enhancement effect of CpG-ODN containing non-methylated cytosine and guanine nucleotide sequences on BCG and to provide evidence for improving the immunogenicity of BCG. Methods (1) Cell experiments: The blank control group, low concentration group and high concentration group were inoculated with the mouse macrophage cell line Ana-1 pre-culture. Culture medium, low concentration of CpG-ODN and high concentration of CpG-ODN were added respectively. The contents of IL-12, IL-18 and IFN-γ in the supernatant were measured after culture. (2) Animal experiments: The control group (group A), infection group (group B), BCG prevention group (group C) and BCG + CpG-ODN prevention group (group D). Groups A, B, C and D were injected with sterile PBS, BCG, BCG and CpG-ODN mixture respectively. After 2 weeks, mice in groups B, C and D were injected with Mycobacterium tuberculosis. Two weeks later, all mice were sacrificed The lung tissue was taken for pathological examination. Serum levels of IL-12, IL-18 and IFN-γ were measured. Results: (1) Cell experiment: Under the stimulation of CpG-ODN, IL-12, IL-18 and IFN-γ secreted by Ana-1 were all in a CpG-ODN dose-dependent manner. The above three Th1-type cytokines content more. (2) Animal experiments: There was no infiltration of inflammatory cells in group A lungs; Group B showed a large number of infiltration of inflammatory cells; Group C and Group D lung tissue infiltration of a small amount of inflammatory cells. The levels of IL-12, IL-18 and IFN-γ in serum of group B were significantly lower than those of group A (all P <0.05). The serum levels of IL-12, IL-18 and IFN-γ in group C and group D were significantly higher than those in group A (all P <0.05). The serum levels of IL-12, IL-18 and IFN-γ in group D were significantly higher than those in group C (all P <0.05). Conclusion CpG-ODN can enhance the immunoprecipitation effect of BCG, which may be related to the up-regulation of Th1-type cytokines by CpG-ODN.