【摘要】 目的 探讨灭活的新型结核病疫苗菌株（B/R菌株）对小鼠T淋巴细胞免疫记忆的影响。方法 以PBS、BCG菌株、B/R菌株与灭活B/R菌株为分组方式，单次皮下接种免疫C57BL/6小鼠。于免疫后第9周、第12周和第15周，分别取各组小鼠脾脏，提取脾脏淋巴细胞。各组脾脏淋巴细胞半数直接检测，其余部分进行体外培养，并按实验设计以结核菌素纯蛋白衍生物（PPD）进行刺激，而后进行检测。应用流式细胞技术分别检测未经PPD刺激和经刺激的小鼠脾脏淋巴细胞内中央型记忆性T细胞(T_CM)和效应型记忆性T细胞(T_EM)数量。结果 免疫后未经PPD刺激，第9周时，灭活B/R组、B/R组、BCG组诱导产生的CD₄⁺ T_CM（F=13.20,P<0.05）与CD₄⁺ T_EM（F=28.15,P<0.05）高于PBS组；诱导产生的CD₈⁺ T_CM（F=8.92,P<0.05）与CD₈⁺ T_EM（F=6.13,P<0.05）高于PBS组。第12周时，各组诱导产生的CD₄⁺ T_CM（F=15.97,P<0.05）与CD₄⁺ T_EM（F=13.60,P<0.05）高于PBS组；诱导产生的CD₈⁺ T_CM（F=5.52,P<0.05）与CD₈⁺ T_EM（F=20.15,P<0.05）高于PBS组。第15周时，各组诱导产生的CD₄⁺ T_CM（F=15.40,P<0.05）与CD₄⁺ T_EM（F=7.43,P<0.05）高于PBS组；诱导产生的CD₈⁺ T_CM（F=6.57,P<0.05）与CD₈⁺ T_EM（F=9.27,P<0.05）高于PBS组。免疫后经结核特异性抗原PPD刺激，在第9周时，灭活B/R组、B/R组、BCG组诱导产生的CD₄⁺ T_CM（F=9.66,P<0.05）与CD₄⁺T_EM（F=11.20,P<0.05）高于PBS组；诱导产生的CD₈⁺ T_CM（F=7.24,P<0.05）与CD₈⁺ T_EM（F=9.30,P<0.05）高于PBS组。第12周时，各组诱导产生的CD₄⁺ T_CM（F=9.33,P<0.05）与CD₄⁺T_EM（F=6.94,P<0.05）高于PBS组；诱导产生的CD₈⁺ T_CM（F=67.71,P<0.05）与CD₈⁺ T_EM（F=10.86,P<0.05）高于PBS组。第15周时，各组诱导产生的CD₄⁺ T_CM（F=39.88,P<0.05）与CD₄⁺T_EM（F=11.93,P<0.05）高于PBS组；诱导产生的CD₈⁺ T_CM（F=138.80,P<0.05）与CD₈⁺ T_EM（F=38.47,P<0.05）高于PBS组。值得注意的是，在第15周时，灭活B/R组与B/R组诱导产生的CD₈⁺ T_CM(q_灭活B/R=12.24,q_B/R=12.61,P<0.05)与CD₈⁺ T_EM(q_灭活B/R=7.19,q_B/R=5.00,P<0.05)均高于BCG组。结论 灭活B/R菌株免疫小鼠后，所诱导对小鼠T淋巴细胞免疫记忆的能力与B/R菌株相当，热灭活未影响B/R菌株诱导小鼠免疫记忆的能力。更多还原

【Abstract】 To investigate the effect of inactivated new tuberculosis vaccine strain（B/R strain） on immune memory of T lymphocytes in mice, C57BL/6 mice were immunized with PBS, BCG strain, B/R strain and inactivated B/R strain. At week 9, 12 and 15 after immunization, the spleens of each group were taken, and the spleen lymphocytes were extracted. Half of the spleen lymphocytes in each group were directly detected, and the rest were cultured in vitro and stimulated with PPD according to the experimental design. Flow cytometry was used to detect the number of central memory T cells(T_CM) and effector memory T cells(T_EM) in spleen lymphocytes of unstimulated and stimulated mice.Without PPD stimulation after immunization, the CD₄⁺ T_CM（F=13.20,P<0.05） and CD₄⁺ T_EM（F=28.15, P<0.05） induced by inactivated B/R group, B/R group and BCG group was higher than that of PBS group at week 9.The induced CD₈⁺ T_CM（F=8.92, P<0.05） and CD₈⁺ T_EM（F=6.13, P<0.05） was higher than that of PBS group.At week 12, the CD₄⁺ T_CM（F=15.97, P<0.05） and CD₄⁺ T_EM（F=13.60, P<0.05） induced by each group was higher than that of PBS group.The induced CD₈⁺ T_CM（F=5.52, P<0.05） and CD₈⁺ T_EM（F=20.15, P<0.05） was higher than that of PBS group. At week 15, the CD₄⁺ T_CM（F=15.40, P<0.05） and CD₄⁺ T_EM（F=7.43, P<0.05） induced by each group was higher than that of PBS group. The induced CD₈⁺ T_CM（F=6.57, P<0.05） and CD₈⁺ T_EM（F=9.27, P<0.05） was higher than that of PBS group. At week 9, the CD₄⁺ T_CM（F=9.66, P<0.05） and CD₄⁺ T_EM（F=11.20, P<0.05）induced by inactivated B/R group, B/R group and BCG group was higher than that of PBS group. The induced CD₈⁺ T_CM（F=7.24, P<0.05） and CD₈⁺ T_EM（F=9.30, P<0.05） was higher than that of PBS group. At week 12, the CD₄⁺ T_CM（F=9.33, P<0.05） and CD₄⁺ T_EM（F=6.94, P<0.05） induced by each group was higher than that of PBS group. The induced CD₈⁺ T_CM（F=67.71, P<0.05） and CD₈⁺ T_EM（F=10.86, P<0.05） was higher than that of PBS group.At week 15, the CD₄⁺ T_CM（F=39.88, P<0.05） and CD₄⁺ T_EM（F=11.93, P<0.05） induced by each group was higher than that of PBS group. The induced CD₈⁺ T_CM and CD₈⁺ T_EM（F=38.47, P<0.05） was higher than that of PBS group（F=138.80, P<0.05）. It is worth noting that at week 15, the CD₈⁺ T_CM(q_{inactivated B/R}=12.24, q_B/R=12.61, P<0.05) and CD₈⁺ T_EM(q_{inactivated B/R}=7.19, q_B/R=5.00, P<0.05) induced by inactivated B/R group and B/R group were higher than those of BCG group. The immunizing mice with inactivated B/R strain, the ability of inducing immune memory of T lymphocytes in mice was equivalent to that of B/R strain. Heat-inactivation did not affect the ability of B/R strain to induce immune memory in mice.更多还原