Bacterial Factors that Regulate Ocular Host-Pathogen Interactions and Microbial Keratitis

调节眼部宿主-病原体相互作用和微生物角膜炎的细菌因素

• 批准号: 9910406
• 负责人: ROBERT M SHANKS
• 金额: $ 44.44万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9910406
• 关键词: Address; Animal Model; Antibiotics; Bacteria; Bacterial Antibiotic Resistance; Bacterial Eye Infections; Bacterial Proteins; Biology; Blindness; Bulla; Cell Death; Cell Line; Cells; Centers for Disease Control and Prevention (U.S.); Chemicals; Clinic; Clinical; Communities; Community-Acquired Infections; Complement; Complication; Contact Lenses; Cornea; Corneal Diseases; Cytolysins; Cytolysis; Data; Defect; Dextrans; Disease; Enterobacteriaceae; Epithelial Cells; Escherichia coli; Event; Eye; Eye Infections; Family suidae; Foundations; Genes; Genetic Transcription; Goals; Hospitals; Human; Human Biology; In Vitro; Individual; Infection; Inflammation; Keratitis; Knock-out; Laboratories; Mediating; Membrane; Microscopy; Modeling; Molecular; Morbidity - disease rate; Multi-Drug Resistance; Nosocomial Infections; Oryctolagus cuniculus; Pain; Pathogenesis; Pathogenicity; Pathway interactions; Prevention strategy; Production; Productivity; Protein Inhibition; Proteins; Pseudomonas aeruginosa; Regulation; Reporting; Research; Resolution; Role; Serratia marcescens; Societies; Sterility; Stress; Surface; System; Tertiary Protein Structure; Testing; Therapeutic; Transcriptional Regulation; Type III Secretion System Pathway; United States; Virulence; Virulence Factors; Vision; Work; carbapenem-resistant Enterobacteriaceae; cell injury; corneal epithelium; cost; cytotoxicity; effective therapy; experience; human morbidity; human mortality; improved; in vivo; member; microbial; mortality; mutant; novel; ocular surface; pathogen; prevent; programs; tool; transcriptome sequencing; translational impact; treatment strategy

7. Project Summary / Abstract The overall goal of our research program is to develop better treatment strategies for the prevention of vision loss due to microbial keratitis, as well as increasing our understanding of ocular biology through host-pathogen interaction analysis. This specific study will elucidate a novel host-pathogen interaction important for bacterial infection of the ocular surface. We are focusing on a class of bacteria that frequently cause ocular infections and multidrug resistant hospital infections, the Enterobacteriaceae, that is understudied with respect to ocular infections. Importantly, we will be using recent clinical isolates derived from contact lens associated keratitis event rather than non-ocular laboratory strains that may not share ocular virulence factors. Using the member of the Enterobacteriaceae that most commonly causes keratitis as a model organism, Serratia marcescens, we have uncovered a novel host-pathogen interaction that we will define in this study. Namely, a step in bacteria-mediated ocular cell death is the formation of a distinct type of membrane bleb. Preliminary data indicates the mechanism underlying bleb induction by S. marcescens requires a bacterial type V secretion system, a cytolysin, and a conserved but unstudied stress regulator we are calling GumB. Taking advantage of our state of the art microscopy facility, molecular tools, tunable expression systems, and ocular infection models, we are poised to complete our research goals. Our aims will: 1) address the hypothesis that a type V secretion system involving cytolysin proteins ShlB and ShlA is necessary and sufficient to induce membrane blebs in human ocular cells. This will introduce a completely new mechanism by which bacteria influence corneal cells; 2) determine the mechanism by which GumB regulates bleb formation and cytotoxicity to corneal epithelial cells. This aim will uncover the role of GumB as a virulence factor regulator, which is a conserved bacterial protein found in many species associated with ocular host-pathogen interactions; 3) establish the role of GumB in ocular virulence using a rabbit contact lens keratitis model. This work will show that GumB is a novel master regulator of ocular virulence factors, and will identify those factors. These studies have the potential to discover an entirely new mechanism by which bacteria impact human cells and corneal disease. The long-term implication is that CL-associated keratitis is due, at least in part, to this host-pathogen interaction. The resolution of this mechanism could establish the foundation for more effective infection treatments, such as through chemical inhibition of the proteins studied here.

7. 项目总结/摘要 我们研究计划的总体目标是开发更好的治疗策略来预防视力 微生物角膜炎造成的损失，以及通过宿主病原体增加我们对眼生物学的了解 交互分析。这项具体研究将阐明一种对细菌重要的新型宿主-病原体相互作用 眼表感染。我们关注的是一类经常引起眼部感染的细菌 以及多重耐药性医院感染，即肠杆菌科细菌，在眼部方面尚未得到充分研究 感染。重要的是，我们将使用源自隐形眼镜相关角膜炎的最新临床分离株 事件而不是可能不具有眼部毒力因子的非眼部实验室菌株。 使用最常引起角膜炎的肠杆菌科成员作为模式生物， 粘质沙雷氏菌，我们发现了一种新的宿主-病原体相互作用，我们将在本研究中对其进行定义。 也就是说，细菌介导的眼细胞死亡的一个步骤是形成独特类型的膜泡。 初步数据表明粘质沙门氏菌诱导气泡的机制需要细菌类型 V 分泌系统、溶细胞素和保守但未经研究的应激调节因子，我们称之为 GumB。 利用我们最先进的显微镜设备、分子工具、可调表达系统和 眼部感染模型，我们准备完成我们的研究目标。我们的目标是：1）解决 假设涉及溶细胞素蛋白 ShlB 和 ShlA 的 V 型分泌系统是必要的，并且 足以在人眼细胞中诱导膜泡。这将引入一个全新的 细菌影响角膜细胞的机制； 2）确定GumB的机制 调节泡形成和对角膜上皮细胞的细胞毒性。这一目标将揭示 GumB 作为毒力因子调节剂，是在许多相关物种中发现的保守细菌蛋白 与眼部宿主-病原体相互作用； 3) 使用兔子确定 GumB 在眼毒力中的作用 隐形眼镜角膜炎模型。这项工作将表明 GumB 是一种新型的眼部毒力调节剂 因素，并将识别这些因素。这些研究有可能发现一种全新的机制 细菌影响人体细胞和角膜疾病。长期影响是 CL 相关 角膜炎至少部分归因于这种宿主与病原体的相互作用。这一机制的解决可以 为更有效的感染治疗奠定基础，例如通过化学抑制 蛋白质在这里研究。