Attenuated Mycobacterium tuberculosis SO2 Vaccine Candidate Is Unable to Induce Cell Death

It has been proposed that Mycobacterium tuberculosis virulent strains inhibit apoptosis and trigger cell death by necrosis of host macrophages to evade innate immunity, while non-virulent strains induce typical apoptosis activating a protective host response. As part of the characterization of a novel tuberculosis vaccine candidate, the M. tuberculosis phoP mutant SO2, we sought to evaluate its potential to induce host cell death. The parental M. tuberculosis MT103 strain and the current vaccine against tuberculosis Bacillus Calmette-Guérin (BCG) were used as comparators in mouse models in vitro and in vivo. Our data reveal that attenuated SO2 was unable to induce apoptotic events neither in mouse macrophages in vitro nor during lung infection in vivo. In contrast, virulent MT103 triggers typical apoptotic events with phosphatidylserine exposure, caspase-3 activation and nuclear condensation and fragmentation. BCG strain behaved like SO2 and did not induce apoptosis. A clonogenic survival assay confirmed that viability of BCG- or SO2-infected macrophages was unaffected. Our results discard apoptosis as the protective mechanism induced by SO2 vaccine and provide evidence for positive correlation between classical apoptosis induction and virulent strains, suggesting apoptosis as a possible virulence determinant during M. tuberculosis infection.

有研究提出，毒力型结核分枝杆菌（Mycobacterium tuberculosis）可通过抑制宿主巨噬细胞凋亡、触发坏死性细胞死亡以逃避先天免疫；而非毒力菌株则诱导典型的细胞凋亡，从而激活宿主的保护性免疫应答。作为新型结核疫苗候选株——结核分枝杆菌phoP突变株SO2的表征工作之一，我们旨在评估其诱导宿主细胞死亡的潜力。以亲本菌株结核分枝杆菌MT103及当前通用的结核疫苗卡介苗（Bacillus Calmette-Guérin, BCG）作为对照，在小鼠体内外模型中开展实验。
研究数据显示，减毒株SO2既无法在体外培养的小鼠巨噬细胞中诱导凋亡事件，也无法在体内肺部感染过程中触发凋亡。与之相反，毒力株MT103可诱导典型的凋亡事件，包括磷脂酰丝氨酸外翻、半胱氨酸天冬氨酸蛋白酶-3（caspase-3）激活以及细胞核固缩与碎裂。卡介苗菌株的表现与SO2类似，均未诱导细胞凋亡。
集落形成存活实验证实，感染BCG或SO2的巨噬细胞存活率未受影响。本研究结果排除了凋亡作为SO2疫苗诱导保护性免疫的机制，并证明经典凋亡诱导与结核分枝杆菌毒力菌株之间存在正相关关系，提示细胞凋亡可能是结核分枝杆菌感染过程中的毒力决定因子之一。