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目的研究结核分枝杆菌能量代谢对吡嗪酰胺抗结核作用的机制和作用靶点。方法将饥饿3、5和10 d 的结核分枝杆菌及未经过营养饥饿的结核分枝杆菌分别加入100μg/ml 吡嗪酰胺和脂肪酸、苯甲酸和水杨酸进行处理,观察营养饥饿处理过程对于吡嗪酰胺抗结核分枝杆菌作用的影响,以及酸性物质对吡嗪酰胺抗菌活性的影响。同时用流式细胞仪检测结核分枝杆菌膜电位和平均荧光强度,以探讨吡嗪酰胺的作用机制。结果未经过营养饥饿的结核分枝杆菌和饥饿3、5和10 d的结核分枝杆菌经吡嗪酰胺处理后,菌落形成单位(CFU)分别减少23.08%、37.75%、82.32%和81.03%;营养饥饿5 d 后,结核分枝杆菌的膜电位明显降低,经吡嗪酰胺处理后的膜电位则大幅降低,加入能量物质葡萄糖后,因吡嗪酰胺作用而降低的膜电位可得到恢复。脂肪酸、苯甲酸和水杨酸对于正常生长的和营养饥饿后的结核分枝杆菌均有促进吡嗪酰胺抗菌效果的作用,对营养饥饿后的结核分枝杆菌作用更明显。结论饥饿状态和弱酸能增强吡嗪酰胺对结核分枝杆菌的抗菌作用,吡嗪酰胺可能是通过干扰结核分枝杆菌的膜电位及其能量代谢而产生作用。
Objective To study the mechanisms and targets of Mycobacterium tuberculosis (BCG) energy metabolism on pyrazinamide anti-TB activity. Methods Mycobacterium tuberculosis on 3, 5, and 10 days of starvation and Mycobacterium tuberculosis not on nutrient-starvation were respectively treated with 100μg / ml pyrazinamide and fatty acid, benzoic acid and salicylic acid to observe the effect of nutrient starvation treatment on Pyrazinamide against Mycobacterium tuberculosis and the effect of acidic substances on the antibacterial activity of pyrazinamide. At the same time by flow cytometry Mycobacterium tuberculosis membrane potential and average fluorescence intensity to explore the mechanism of pyrazinamide. Results After treatment with pyrazinamide, mycobacterium tuberculosis (M. tuberculosis) without starvation and starved 3, 5 and 10 d after treatment with pyrazinamide reduced the colony forming units (CFUs) by 23.08%, 37.75%, 82.32% and 81.03%, respectively. After 5 days of nutrition starvation, the membrane potential of Mycobacterium tuberculosis decreased significantly, and the membrane potential decreased significantly after treated with pyrazinamide. After the addition of energy substance glucose, the membrane potential decreased due to pyrazinamide recovery. Fatty acids, benzoic acid and salicylic acid have the effect of promoting the antibacterial effect of pyrazinamide on the normal growth and the nutritionally hungry Mycobacterium tuberculosis, and have a more obvious effect on the post-nutrition-hungry Mycobacterium tuberculosis. Conclusion Starvation and weak acid can enhance the antimicrobial effect of pyrazinamide on M. tuberculosis. Pyrazinamide may play an important role by interfering with the membrane potential of M. tuberculosis and its energy metabolism.