Immunomodulatory role of Pic in EAEC Pathogenesis

Pic 在 EAEC 发病机制中的免疫调节作用

基本信息

批准号：
10427392
负责人：
Fernando Ruiz
金额：
$ 34.4万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-07-01 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10427392
关键词：
Age Anaerobic Bacteria Award Binding Biological Assay CD14 gene Cell Line Cell physiology Cell surface Cells Chemotaxis Chronic Citrobacter Coculture Techniques Collaborations Communicable Diseases Complex Data Developed Countries Developing Countries Development Diarrhea Disease model Epithelial Epithelial Cells Escherichia coli Event Exocytosis Family Flow Cytometry Gastroenterologist Gene Silencing Generations Genes Glycoproteins Goblet Cells Growth Human Immunologist Immunology Impaired cognition Impairment Infection Inflammation Inflammatory Intervention Intestinal Mucosa Intestines Lead Lectin Leukocytes Life MAP3K7 gene Maryland Mediating Microscopy Modeling Mucin 1 protein Mucin-2 Staining Method Mucins Mucous body substance Outcome Pathogenesis Penetration Pharmacology Play Process Productivity Property Protease Inhibitor Research Personnel Role Salmonella Serine Protease Serine Proteinase Inhibitors Shigella Shigella Infections Shigella flexneri Signal Pathway Signal Transduction Specialist System Therapeutic Intervention Thinness Time Traveler&apos s diarrhea Universities Virginia Virulence Factors WFDC2 gene Work base diarrheal disease enteric pathogen enteroaggregative Escherichia coli enterotoxigenic Escherichia coli gut colonization gut inflammation immunoregulation insight knock-down macrophage migration monolayer mouse model multidisciplinary neutrophil pathogen pathogenic Escherichia coli receptor transcriptomics

项目摘要

Abstract Enteroaggregative E. coli (EAEC) is an important pathogen of traveler's diarrhea, diarrhea in industrialized countries and growth faltering in developing countries. In this application, we propose to study the role of highly prevalent serine proteases from the SPATE family in EAEC pathogenesis, and their relationship with chronic inflammatory enteropathy using a newly established human leukocyte-colonoid model. This application comprises a consortium among investigators at Johns Hopkins University (JHU), who have pioneered the development of the enteroid/colonoid model, and researchers at the University of Maryland (UMB) and the University of Virginia (UVA), who are leaders in the study of the pathogenesis of diarrheal disease caused by diarrhegenic E. coli and Shigella. We have successfully used the colonoid model to characterize relevant features of EAEC pathogenesis, highlighting the important role of the lectin-like serine protease Pic in intestinal colonization and inflammation. In this project will leverage the existence of the newly developed leukocyte- colonoid monolayer model to characterize the modulatory activity of Pic and other serine proteases in goblet cells, leukocytes and epithelial cells in EAEC pathogenesis. This project will comprise three Specific Aims. In Aim 1, we will characterize the effect of Pic in goblet cell function by obtaining mechanistic insights into Pic- mediated MUC2 exocytosis and by establishing the role of the goblet-cell secreted serine protease inhibitor WFDC2 in EAEC pathogenesis by using unbiased transcriptomics and pharmacological approaches. In Aim 2, we will characterize the role of SPATEs in EAEC-induced epithelial inflammation by obtaining mechanistic insights into MUC1-mediated epithelial inflammation in EAEC infection, employing gene silencing and pharmacological approaches. In Aim 3, we will characterize the immunomodulatory property of Pic lectin and mucinolytic activity on leukocyte functions in EAEC pathogenesis employing neutrophils- and macrophages- colonoids co-cultures along with state-of-the-art time-lapse microscopy, flow cytometry and Luminex-based assays. This work will continue to advance our understanding of EAEC pathogenesis, generating fundamental insights that will illuminate aspects of pathogenesis relevant to other enteric pathogens, and could open up new avenues for therapeutic intervention. We will work closely and synergistically with investigators in other projects and the Core components.

抽象的肠聚集性大肠杆菌（EAEC）是旅行者腹泻、工业化腹泻的重要病原体国家和发展中国家的增长步履蹒跚。在此应用中，我们建议研究高度的作用 SPATE 家族中常见的丝氨酸蛋白酶在 EAEC 发病机制中及其与慢性疾病的关系使用新建立的人类白细胞-结肠模型来研究炎症性肠病。这个应用程序由约翰·霍普金斯大学 (JHU) 的研究人员组成的联盟组成，他们开创了肠样/结肠样模型的开发以及马里兰大学 (UMB) 和弗吉尼亚大学 (UVA)，该大学在研究由腹泻引起的腹泻疾病的发病机制方面处于领先地位腹泻性大肠杆菌和志贺氏菌。我们已经成功地使用结肠模型来表征相关 EAEC发病机制的特点，强调凝集素样丝氨酸蛋白酶Pic在肠道中的重要作用定植和炎症。在这个项目中将利用新开发的白细胞的存在- 集落单层模型，用于表征杯中 Pic 和其他丝氨酸蛋白酶的调节活性 EAEC 发病机制中的细胞、白细胞和上皮细胞。该项目将包括三个具体目标。在目标 1，我们将通过获得对 Pic 的机制了解来表征 Pic 对杯状细胞功能的影响- 介导 MUC2 胞吐作用并确定杯状细胞分泌的丝氨酸蛋白酶抑制剂的作用通过使用公正的转录组学和药理学方法研究 WFDC2 在 EAEC 发病机制中的作用。在目标 2 中，我们将通过获得机制来描述 SPATE 在 EAEC 诱导的上皮炎症中的作用深入了解 EAEC 感染中 MUC1 介导的上皮炎症，采用基因沉默和药理学方法。在目标 3 中，我们将表征 Pic 凝集素的免疫调节特性和利用中性粒细胞和巨噬细胞在 EAEC 发病机制中对白细胞功能的粘蛋白溶解活性类集落共培养以及最先进的延时显微镜、流式细胞术和基于 Luminex 的技术化验。这项工作将继续推进我们对 EAEC 发病机制的理解，产生基础这些见解将阐明与其他肠道病原体相关的发病机制，并可能开辟新的领域治疗干预的途径。我们将与其他项目的研究人员密切协作和核心组件。