Attenuation of <i>Pseudomonas aeruginosa</i> infection by INP0341, a salicylidene acylhydrazide, in a murine model of keratitis

<b>Pseudomonas aeruginosa</b> is an opportunistic pathogen and a major cause of corneal infections worldwide. The bacterium secretes several toxins through its type III secretion system (T3SS) to subvert host immune responses. In addition, it is armed with intrinsic as well as acquired antibiotic resistance mechanisms that make treatment a significant challenge and new therapeutic interventions are needed. Type III secretion inhibitors have been studied as an alternative or in accompaniment to traditional antibiotics to inhibit virulence of bacteria. In this study, INP0341, a T3SS inhibitor, inhibited cytotoxicity by <i>P. aeruginosa</i> toward human corneal epithelial cells (HCEC) at 100 μM without affecting bacterial growth in the liquid media. An increased expression of antimicrobial peptides and reactive oxygen species generation was also observed in cells exposed to <i>P. aeruginosa</i> in the presence of INP0341. Furthermore, INP0341 efficiently attenuated corneal infection by <i>P. aeruginosa</i> in an experimental model of murine keratitis as evident from corneal opacity, clinical score and bacterial load. Thus, INP0341 appears to be a promising candidate to treat corneal infection caused by <i>P. aeruginosa</i> and can be further considered as an alternative therapeutic intervention.铜绿假单胞菌（Pseudomonas aeruginosa）是一种机会致病菌，也是全球范围内角膜炎的主要致病原。该细菌可通过III型分泌系统（type III secretion system, T3SS）分泌多种毒素，以破坏宿主免疫应答。此外，铜绿假单胞菌兼具固有耐药与获得性耐药机制，使其临床治疗面临极大挑战，亟需开发新型治疗干预手段。作为传统抗生素的替代方案或辅助疗法，III型分泌系统抑制剂已被用于研究以抑制细菌毒力。本研究中，III型分泌系统抑制剂INP0341在100 μM浓度下即可抑制铜绿假单胞菌对人角膜上皮细胞（human corneal epithelial cells, HCEC）的细胞毒性，且不会影响该细菌在液体培养基中的生长。在INP0341存在的条件下，暴露于铜绿假单胞菌的细胞中，抗菌肽的表达水平显著上调，活性氧生成量也有所增加。进一步实验显示，在小鼠角膜炎模型中，INP0341可有效减轻铜绿假单胞菌所致的角膜感染，该效果可通过角膜混浊度、临床评分以及细菌载量得到验证。综上，INP0341是治疗铜绿假单胞菌所致角膜炎的极具潜力的候选药物，可进一步作为替代治疗干预手段进行开发。