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Lung epithelial cell regulation of immunity to inhaled fungi

肺上皮细胞对吸入真菌免疫的调节

基本信息

批准号：
9975090
负责人：
BRUCE Steven KLEIN
金额：
$ 46.17万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-01 至 2022-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9975090
关键词：
Address Adult Air Alveolar Antifungal Agents Behavior Blastomyces C Type Lectin Receptors CCL20 gene CRISPR/Cas technology Cell Line Cell Lineage Cells Confocal Microscopy Data Defense Mechanisms Ecosystem Elements Enterobacteria phage P1 Cre recombinase Epithelial Epithelial Cells Epithelium Event Fostering Fungal Spores Genes Granulocyte-Macrophage Colony-Stimulating Factor Hematopoietic Histoplasma Host Defense Human Image Immunity Individual Infection Inflammatory Inhalation Interleukin-1 Interleukin-17 Interleukin-18 Interleukin-6 Knowledge Learning Leukocytes Life Style Link Location Lung Lung Inflammation Lymphocyte Mammalian Cell Molecular Mucosal Immunity Mucous Membrane Mus Myelogenous Myeloid Cells Natural Immunity Neurosecretory Systems Outcome Pathogenicity Persons Production Public Health Regulation Reporter Reproduction spores Resistance Respiratory Mucosa Role Signal Pathway Signal Transduction Source T-Lymphocyte Testing Transgenic Mice Work climate change cytokine design extracellular fungus genome editing improved in vivo insight interleukin-22 interleukin-23 lymphocyte product particle pathogen pathogenic fungus promoter receptor recruit response restraint therapeutic target tissue repair tool γδ T cells

项目摘要

PROJECT SUMMARY We propose to define how lung epithelial cells (EC) sense and respond to inhaled fungi. A person inhales a billion spores/day, but the modes by which fungi are sensed and restrained in lung are unclear. Our preliminary data reveal that EC regulate mucosal immunity to two primary pathogens, Blastomyces and Histoplasma. We find that EC NF-κB activation is essential, as are upstream signals of IL-1R, MyD88 and CARD9 and mobiliza- tion of innate lymphocytes that make IL-17 and GM-CSF. We posit that EC subsets, notably Club cells, sense fungi and regulate innate immunity, and that EC intrinsic signals via IL-1R and MyD88 drive NFkB-regulated products to trigger IL-17- and GM-CSF-producing innate lymphocytes, which recruit/activate myeloid effectors. The lung contains 6 EC subsets: alveolar, club (formerly clara), goblet, neuroendocrine, basal and ciliated columnar. We'll test the role of Club cells in regulating mucosal immunity by using transgenic mice that display a lineage-specific GFP reporter to track and analyze the behavior of this subset and, since our reporter mouse harbors cre recombinase, test its role in resistance by crossing the mouse with IKK2fl/fl or CRISPR/Cas9 mice to block NF-κB signaling. To learn how EC mobilize innate immunity to fungi, we will use two fungal pathogens that represent paradigmatic extracellular (Blastomyces) and intracellular (Histoplasma) pathogenic lifestyles that confront the epithelium of mammalian lungs. Our aims are to: 1. Delineate the role and action of Club cells in sensing inhaled fungi. By using NFκB-eGFP reporter mice, advanced imaging and FACS analysis, we will track the timing and location of activation of EC and hemato- poietic cells in response to inhaled fungi. Transgenic mice will be used to dissect the roles of Club cells and their signaling pathways and downstream products in resistance, and CRISPR/Cas9 used to GFP tag, iso- late, profile and functionally interrogate specific functions of Club cells in regulating innate immunity. 2. Define how Club cells and products regulate γδ T cells and nTh17 cells to restrain inhaled fungi. We will define the regulatory role of Club cells and their signals on anti-fungal TCR γδ and nTh17 cells – e.g. activa- tion or recruitment and cytokine production. The role of innate T cell IL-17A, GM-CSF and IL-22 will be dis- sected in promoting killing of fungi and fostering EC integrity. Soluble signals such as CCL20, IL-1, IL-6, IL- 18 & IL-23 that activate innate T cells in mice and humans will be defined, and the cell source(s) identified. Our work will reveal how lung Club cells regulate innate mucosal immunity to fungi offering conceptually new insight and powerful gene editing tools that will advance the field. By divining antifungal defense mechanisms, we will improve prospects for therapeutically targeting early events designed to optimize mucosal immunity to fungi and lung inflammation.

项目总结