CCCP Facilitates Aminoglycoside to Kill Late Stationary-Phase Escherichia coli by Elevating Hydroxyl Radical.
CCCP Facilitates Aminoglycoside to Kill Late Stationary-Phase Escherichia coli by Elevating Hydroxyl Radical.
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DOI：
10.1021/acsinfecdis.2c00522
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发表时间：
2023-03
影响因子：
5.3
通讯作者：
Zhongyan Li;Linchao Wu;Zhijie Huang;Boyan Lv;Yajuan Fu;Lunjiang Zhou;Xinmiao Fu
中科院分区：
文献类型：
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作者：
Zhongyan Li;Linchao Wu;Zhijie Huang;Boyan Lv;Yajuan Fu;Lunjiang Zhou;Xinmiao Fu
提高现有抗生素的功效对于对抗对人类健康构成重大威胁的抗生素耐药性具有重要意义。羰基氰化物间氯苯肼（CCCP）是一种众所周知的消散质子动力（PMF）的质子载体，被广泛用于阻断PMF依赖性氨基糖苷类抗生素的摄取，从而抑制氨基糖苷类抗生素的致死率。在这里，我们报告了CCCP及其功能类似物FCCP，而不是其他类型的原载体，史无前例地增强氨基糖苷类（例如，tobramycin和gentamicin），对固定期的大肠杆菌、金黄色葡萄球菌、福氏志贺氏菌和溶藻弧菌细胞具有3-4个数量级的杀伤作用，但对指数期的这些细胞和我们研究的其他12种细菌都没有杀伤作用。总的来说，CCCP对氨基糖苷致死率的影响经历了一个逐渐的转变，从对大肠杆菌指数期细胞的抑制到对晚静止期细胞的增强，细胞生长状态和培养基是至关重要的。同样，通过改变跨膜质子梯度（ΔpH）或电势（ΔΨ）对PMF的干扰也会增强妥布霉素。然而，CCCP既没有增加妥布霉素的细胞内浓度，也没有降低抗生素的MIC，因此排除了CCCP作为外排泵抑制剂增强氨基糖苷的可能性。相反，我们表明，在有氧和厌氧培养条件下，联合处理显著提高了羟基自由基的细胞水平，在这种条件下，抗氧化剂n -乙酰半胱氨酸完全抑制羟基自由基的积累和细胞死亡。总之，这些发现为开发某些原载体作为氨基糖苷佐剂对抗固定期病原体开辟了新的途径，也说明了羟基自由基在氨基糖苷致死性中的重要作用，而不管有氧呼吸。
Improving the efficacy of existing antibiotics is significant for combatting antibiotic resistance that poses a major threat to human health. Carbonyl cyanide m-chlorophenylhydrazine (CCCP), a well-known protonophore for dissipating proton motive force (PMF), has been widely used to block the PMF-dependent uptake of aminoglycoside antibiotics and thus suppress aminoglycoside lethality. Here, we report that CCCP and its functional analog FCCP, but not other types of protonophores, unprecedently potentiate aminoglycosides (e.g., tobramycin and gentamicin) by 3-4 orders of magnitude killing of Escherichia coli, Staphylococcus aureus, Shigella flexneri, and Vibrio alginolyticus cells in stationary phase but not these cells in exponential phase nor other 12 bacterial species we examined. Overall, the effect of CCCP on aminoglycoside lethality undergoes a gradual transition from suppression against E. coli exponential-phase cells to potentiation against late stationary-phase cells, with the cell growth status and culture medium being crucial. Consistently, disturbance of the PMF by changing transmembrane proton gradient (ΔpH) or electric potential (ΔΨ) also potentiates tobramycin. Nevertheless, CCCP neither increases the intracellular concentration of tobramycin nor decreases the MIC of the antibiotic, thus excluding that CCCP acts as an efflux pump inhibitor to potentiate aminoglycosides. Rather, we show that the combined treatment dramatically enhances the cellular level of hydroxyl radical under both aerobic and anaerobic culturing conditions, under which the antioxidant N-acetyl cysteine fully suppresses both hydroxyl radical accumulation and cell death. Together, these findings open a new avenue to develop certain protonophores as aminoglycoside adjuvants against pathogens in stationary phase and also illustrate an essential role of hydroxyl radical in aminoglycoside lethality regardless of aerobic respiration.