Pathogenesis of P. aeruginosa corneal infection

铜绿假单胞菌角膜感染的发病机制

• 批准号: 6854861
• 负责人: Gerald B Pier
• 金额: $ 40.78万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-01-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6854861
• 关键词: Pseudomonas aeruginosa; alginates; bacterial cytopathogenic effect; cell line; chloride channels; corneal epithelium; disease /disorder model; eye infections; genetically modified animals; host organism interaction; immunotherapy; interferon gamma; interleukin 1; interleukin 18; interleukin 6; keratitis; laboratory mouse; microorganism disease chemotherapy; monoclonal antibody; nonhuman therapy evaluation; therapy design /development

DESCRIPTION: The long-term goal of this application is to understand the molecular and cellular responses of corneal epithelial cells to Pseudomonas aeruginosa and develop chemotherapeutic and immunotherapeutic interventions for treatment of this serious eye infection. P. aeruginosa enters corneal epithelial cells via the cystic fibrosis transmembrane conductance regulator (CFTR), and these intracellular bacteria are able to persist in the tissue and avoid host defenses, leading to serious corneal pathology. Goals of aim 1 include exploring the role of the CFTR-P. aeruginosa interactions in corneal pathology as manifest by host chemokine and cytokine responses. Using isogenic human corneal epithelial cell lines expressing either mutant or wild-type (WT) CFTR, P. aeruginosa induced and CFTR-dependent release of cytokines implicated in pathogenesis and resistance to infection, notably IL-1, IL-6, IL-18 and IFN-gamma, will be examined. The role of the identified factors in eye infection and pathology will be tested in appropriate WT and transgenic mice using a corneal scratch-injured eye infection model or in mice in which the factor is neutralized with antibody. P. aeruginosa- CFTR interactions depend on formation of lipid rafts, and preliminary data indicate disruption of rafts prevents corneal pathology and promotes bacterial clearance from infected eyes. In aim 2, the role of these rafts in pathogenesis will be further evaluated using the WT and CF corneal cell lines as well as mice unable to form lipid rafts due to disruption of the acid sphingomyelinase gene, and in mice treated with cyclodextrin, which disrupts lipid rafts. Cyclodextrin treatment of P. aeruginosa eye infections has the potential to be highly efficacious and become an important component of therapy. Finally, a fully human monoclonal antibody (MAb) has been developed to P. aeruginosa alginate a conserved, cell surface polysaccharide that is expressed at low levels by corneal isolates. Importantly, alginate expression is detected n the corneas of P. aeruginosa infected mice and the MAb is highly protective against P. aeruginosa infection. In aim 3, the Mab will be evaluated for therapeutic efficacy against 6 different P. aeruginosa strains in the murine corneal infection model with the goal of having a single immunotherapeutic reagent useful against most clinical isolates as an adjunct to current treatment modalities for P. aeruginosa ulcerative keratitis. Overall at the conclusion of these aims, significant advances in the development of therapies for P. aeruginosa keratitis should become apparent based on use of cellular and molecular studies on host-pathogen interactions.

描述：该应用的长期目标是了解角膜上皮细胞对铜绿假单胞菌的分子和细胞反应，并开发化学治疗和免疫治疗干预措施，以治疗这种严重的眼部感染。 铜绿假单胞菌通过囊性纤维化跨膜电导调节剂（CFTR）进入角膜上皮细胞，这些细胞内细菌能够在组织中持续存在并避免宿主防御，从而导致严重的角膜病理学。目标1的目标包括探索CFTR-P的作用。角膜病理学中的铜质相互作用由宿主趋化因子和细胞因子反应表现出来。使用表达突变体或野生型CFTR，P。铜绿假单胞菌诱导的和抗CFTR依赖性的细胞因子释放的异源性人角膜上皮细胞系，与感染性的抗性和抗性有关，尤其是IL-1，IL-6，IL-6，IL-18和IFN-GAMMA，将被检查。鉴定因子在眼感染和病理学中的作用将在适当的WT和转基因小鼠中使用角膜刮擦感染的眼感染模型或在用抗体中和该因子中和的小鼠中进行测试。 铜绿假单胞菌CFTR相互作用取决于脂质筏的形成，初步数据表明筏的破坏可防止角膜病理学并促进受感染眼睛的细菌清除率。 在AIM 2中，这些筏在发病机理中的作用将通过WT和CF角膜细胞系进一步评估，以及由于酸鞘磷脂酶基因的破坏而无法形成脂质筏的小鼠，以及在用环糊精处理的小鼠中，破坏了脂质筏。铜绿假单胞菌眼感染的环糊精治疗具有高效的潜力并成为治疗的重要组成部分。最后，已经开发出一种完全人类的单克隆抗体（MAB）为铜绿假单胞菌藻酸酯一种保守的细胞表面多糖，该多糖通过角膜分离株在低水平表达。重要的是，检测到藻酸盐的表达n N的铜绿假单胞菌感染小鼠的角膜，MAB对铜绿假单胞菌感染具有高度保护性。在AIM 3中，将评估MAB在鼠角膜感染模型中对6种不同的铜绿假单胞菌菌株的治疗功效，其目的是具有对大多数临床分离株有用的单个免疫治疗试剂，作为大多数临床分离株，作为铜绿假单胞菌溃疡性角膜炎的当前治疗方法的辅助方法。总体而言，在这些目标的结论中，基于使用细胞和分子研究对宿主 - 病原体相互作用的使用，应显而易见铜绿假单胞菌角膜炎疗法的显着进步。