为建立小鼠结核分枝杆菌持续感染模型并研究其免疫应答特征，选取雌性C57BL/6小鼠，经尾静脉感染结核分枝杆菌H37Rv株；并以异烟肼和利福平联合治疗。分别于感染后4、8、12周处死小鼠，用平板法计数肺和脾荷菌数，酶联免疫吸附试验（ELISA）检测血清中特异性抗体水平及亚类，流式细胞术检测CD4⁺脾淋巴细胞经结核分枝杆菌抗原—纯化蛋白衍生物（PPD）刺激后分泌细胞因子的细胞比例。结果显示，感染4 周后小鼠肺和脾荷菌数lg cfu分别达3.67±0.25和3.54±0.24，至少持续8 周；药物治疗可有效降低脏器荷菌数。感染12周后，感染组血清中抗结核分枝杆菌特异性抗体水平显著升高（P<0.01），且以IgG1为主；化疗组总IgG抗体水平显著低于感染组（P<0.01），且IgG2a相对较高（P<0.05）。感染组CD4⁺脾淋巴细胞中γ干扰素（IFN-γ）和白细胞介素2（IL-2）分泌细胞比例显著增加（P<0.01和P<0.001），而IL-4分泌细胞比例显著降低（P<0.01）；化疗组IL-2和IL-4分泌细胞比例显著低于正常组（P<0.05和P<0.01）。本研究建立的小鼠结核分枝杆菌持续感染模型有望用于结核病治疗性疫苗及药物的研发和筛选。

To establish a persistent infection mouse model and study its immunological characteristics, female C57BL/6 mice were infected with Mycobacterium tuberculosis (M. tuberculosis) H37Rv at a dose of 1×10⁵ cfu by tail vein injection. Four weeks post infection, one group of mice was treated with isoniazid and rifampicin in drinking water for 8 weeks. At 4, 8 and 12 weeks after infection, 6 mice were euthanized to examine bacterial burden in the lungs and spleens. The homogenate of each organ was serially diluted and plated on plates for bacterial count after 21-day incubation. Mice sera were separated and anti-M. tuberculosis IgG and subclasses were analyzed by enzyme-linked immunosorbent assay (ELISA). CD4⁺ splenocytes were cultured with purified protein derivative (PPD) stimulation, and the percentage of interferon γ (IFN-γ), interleukin 2 (IL-2), IL-4, and tumor necrosis factor α (TNF-α) expressing cells were then identified by intracellular cytokine staining and flowcytometry. The results showed that the lg CFU of bacterial loads in lungs and spleens reached to 3.67±0.25 and 3.54±0.24, respectively 4 weeks after infection and the level retained for at least 8 weeks. The chemotherapy reduced the bacterial load of organs significantly. 12 weeks after infection, M. tuberculosis specific total IgG level increased significantly in the untreated group (P<0.01), especially IgG1 (P<0.01). The IgG level in the chemotherapy-treated group also increased（P<0.05）but lower than that in the untreated group (P <0.01). Compared with the untreated group, the chemotherapy-treated group had a lower IgG1 level (P <0.01) but a higher IgG2a level (P <0.05). The untreated group had a significantly higher percentage of IFN-γ and IL-2 expressing CD4⁺ splenocytes (P<0.01, P<0.001) and lower percentage of IL-4 expressing CD4⁺ splenocytes compared with the control group (P <0.01). The percentages of IL-2 and IL-4 expressing CD4⁺ splenocytes were both lower after chemotherapy compared with untreated group (P<0.05, P<0.01). A persistent infection mouse model of M. tuberculosis was established, and its immunological characteristics were identified in the study. It may provide certain applications in new vaccine and drug screening study against tuberculosis.