Mechanisms of flagellin-induced protection against microbial keratitis

鞭毛蛋白诱导的微生物性角膜炎保护机制

• 批准号: 8840592
• 负责人: Fu-Shin X Yu
• 金额: $ 37.24万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-05-01 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8840592
• 关键词: Acetylation; Adjuvant; Anti-Inflammatory Agents; Anti-inflammatory; Antibiotic Resistance; Aspergillus; Biological Assay; Blindness; Bone Marrow; Candida; Candida albicans; Cells; Chromatin; Cornea; Data; Dendritic Cells; Development; Epithelial; Epithelial Cells; Epithelium; Flagellin; Gene Expression; Genes; Genomics; Goals; Grant; Histone Deacetylase Inhibitor; Homeostasis; Hour; Immune; Immune response; Infection; Inflammation; Inflammation Mediators; Inflammatory; Invaded; Keratitis; Knock-out; Knockout Mice; Knowledge; Lead; Ligands; Light; Mediating; Mediator of activation protein; Methods; Modality; Modeling; Molecular Structure; Mucosal Immunity; Mus; Myeloid Cells; Natural Immunity; Neutrophil Infiltration; Play; Production; Pseudomonas aeruginosa; Recovery of Function; Recruitment Activity; Regulation; Resistance; Resistance to infection; Role; Severities; Signal Transduction; Small Interfering RNA; TLR2 gene; Testing; Therapeutic; Toll-Like Receptor 5; Toll-like receptors; Topical application; Transgenic Mice; activating transcription factor 3; antimicrobial peptide; base; cathelicidin; chemokine; chromatin immunoprecipitation; chromatin modification; chromatin remodeling; corneal epithelium; cytokine; innovation; macrophage; microbial; mouse model; neutrophil; ocular surface; pathogen; prevent; promoter; prophylactic; reconstitution; response; transcription factor

Microbial keratitis is a common cause of vision loss. The cornea is exquisitely sensitive to inflammation- mediated damage and therefore has strong innate defenses. However, when the epithelial barrier function is breached, opportunistic bacterial and fungal pathogens can gain access to the epithelial cell layers and colonize in the cornea, leading to infection. During the initial grant period, flagellin, the ligand of Toll-like receptor 5 (TLR5), was used to fortify corneal innate immunity and render resistance to a broad spectrum of keratitis causing pathogens, including Pseudomonas aeruginosa, Candida albicans and Aspergillus fumigates. This highly inducible and robust innate mucosal protection can be attributed to flagellin-induced genomic reprogramming in the epithelium which, in turn, interacts with professional immune cells to control inflammation and to eradicate invading pathogens. The hypothesis in this application is that flagellin, through TLR5, stimulates protective corneal innate mucosal immunity that involves epithelial cells, infiltrated PMNs, and activated dendritic cells, collectively conferring robust resistance to microbial keratitis. Three specific aims are proposed to test this hypothesis: 1) To determine how flagellin-induced reprogramming is regulated in corneal epithelial cells and in the cornea. This can be assessed by using the chromatin immunoprecipitation assay and histone deacetylase inhibitors for gene-specific chromatin modification and by using siRNA and knockout mice for delineating the role of activating transcription factor-3, a transcription factor induced by fg, in regulating the innate immune response in the cornea. 2) To determine how the epithelium participates in and coordinates flagellin-induced mucosal innate immune defense against microbial keratitis. The role of the epithelium and its interaction with dendritic cells in innate defense will be determined by using various transgenic and knockout mice, bone marrow reconstitution, and cell depletion. 3) To determine how flagellin induces protection against non-flagellated pathogens in the cornea. The therapeutic potential of flagellin and its mechanisms of action will be characterized by using mouse models of fungal keratitis (Candida and Aspergillus as pathogens and post-infection topical flagellin application) and double TLR knockout mice. The results of the proposed study should shed light on corneal innate immunity and its induction, and may lead to the development of new prophylactic/therapeutic modalities for preventing and treating microbial keratitis.

微生物性角膜炎是视力丧失的常见原因。角膜对炎症极其敏感 介导的损害，因此具有强大的先天防御。然而，当上皮屏障功能减弱时 破损的机会性细菌和真菌病原体可以进入上皮细胞层并 定植于角膜，导致感染。在初始资助期间，鞭毛蛋白（Toll-like 的配体） 受体 5 (TLR5)，用于增强角膜先天免疫并提供对广谱病原体的抵抗力 引起角膜炎的病原体，包括铜绿假单胞菌、白色念珠菌和曲霉菌 熏蒸。这种高度诱导性和强大的先天粘膜保护可归因于鞭毛蛋白诱导的 上皮细胞中的基因组重编程，进而与专业免疫细胞相互作用以控制 炎症并消灭入侵的病原体。本申请中的假设是鞭毛蛋白通过 TLR5，刺激保护性角膜先天粘膜免疫，涉及上皮细胞、浸润的中性粒细胞、 和激活的树突状细胞，共同赋予对微生物角膜炎的强大抵抗力。三具体 提出的目标是检验这一假设：1）确定鞭毛蛋白诱导的重编程是如何调节的 在角膜上皮细胞和角膜中。这可以通过使用染色质来评估 免疫沉淀试验和组蛋白脱乙酰酶抑制剂用于基因特异性染色质修饰，并通过 使用 siRNA 和基因敲除小鼠来描述激活转录因子 3（一种转录因子）的作用 fg 诱导的因子，调节角膜的先天免疫反应。 2) 确定如何 上皮参与并协调鞭毛蛋白诱导的粘膜针对微生物的先天免疫防御 角膜炎。上皮细胞的作用及其与树突状细胞在先天防御中的相互作用将被确定 通过使用各种转基因和基因敲除小鼠、骨髓重建和细胞耗竭。 3) 至 确定鞭毛蛋白如何诱导针对角膜中非鞭毛病原体的保护。治疗性的 将通过使用真菌小鼠模型来表征鞭毛蛋白的潜力及其作用机制 角膜炎（念珠菌和曲霉作为病原体以及感染后局部鞭毛蛋白应用）和双重 TLR 基因敲除小鼠。拟议研究的结果应有助于揭示角膜先天免疫及其影响 诱导，并可能导致开发新的预防/治疗方式来预防和治疗 治疗微生物性角膜炎。