The Trojan Horse Hypothesis: Neutrophil Elastase Reprograms Macrophage Function

特洛伊木马假说：中性粒细胞弹性蛋白酶重新编程巨噬细胞功能

基本信息

批准号：
10191015
负责人：
Judith A Voynow
金额：
$ 58.21万
依托单位：
VIRGINIA COMMONWEALTH UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10191015
关键词：
Acetylation Biological Assay Biological Markers Cell Nucleus Cell Survival Cells Chromatin Chromatin Structure Citrulline Cleaved cell Confocal Microscopy Cystic Fibrosis Cystic Fibrosis Transmembrane Conductance Regulator Cytoplasm Cytoplasmic Organelle Cytoplasmic Tail DNA Data Disease Progression Equus caballus Failure Gender Gene Expression Genetic Transcription Glutathione Disulfide HDAC2 gene HMGB1 Protein Histone Deacetylase Histone H3 Human Infection Control Inflammation Inflammatory Inflammatory Response Interruption Leukocyte Elastase Lung diseases Lysine Maps Mitochondria Modification Molecular Morbidity - disease rate Natural Immunity Nuclear Pathway interactions Pattern Peptide Hydrolases Phagocytosis Phenotype PicoGreen Production Proteins Reactive Oxygen Species SOD2 gene Signal Pathway Sirtuins Small Interfering RNA Source Stimulus Structure Superoxides TNF gene Testing Up-Regulation airway inflammation cystic fibrosis airway cytokine extracellular healthy volunteer histone deacetylase 2 knock-down macrophage monocyte mortality new therapeutic target novel

项目摘要

PROJECT SUMMARY In Cystic Fibrosis (CF), airway innate immunity is breached. Macrophage phagocytosis and efferocytosis fail, and macrophages augment airway inflammation by production of excess cytokines and release of High Mobility Group Box 1 (HMGB1), a damage associated molecular pattern. Although loss of CFTR impacts macrophage function, the CF airway milieu, which is typified by micromolar concentrations of neutrophil elastase (NE), is an overwhelming stimulus, activating a robust pro-inflammatory response in macrophages from both CF and healthy subjects. We propose that NE, a biomarker for lung disease progression in CF subverts macrophage function from protective to pro- inflammatory, yet the mechanisms by which NE reprograms the macrophage are not completely understood. In this application, we present an unprecedented hypothetical mechanism to explain how NE alters macrophage function. Extracellular NE is rapidly endocytosed by the macrophage. But instead of being degraded, proteolytically active NE is localized to both cytoplasmic domains and the nucleus, resulting in increased cytokine expression and release of macrophage extracellular traps (METs). Thus, NE functions like a Trojan Horse to subvert macrophage function. We will test this hypothesis in primary human blood monocyte derived macrophages from healthy volunteers and subjects with CF. The Specific Aims follow: Aim 1. To determine whether NE degrades HDACs, resulting in acetylation of downstream targets, leading to transcriptional upregulation of TNFα and release of HMGB1. Aim 2. To evaluate whether NE protease activity and/or NE-generated reactive oxygen species increase release of mitochondrial and nuclear METs. RELEVANCE: Results from this project will define a novel mechanism utilized by NE to promote sustained airway inflammation, the major cause of morbidity and mortality in CF, and identify targets for new therapies to interrupt the relentless progression of lung disease.

项目概要在囊性纤维化 (CF) 中，气道先天免疫被破坏。失败，巨噬细胞通过产生过量的细胞因子和释放 High Mobility Group Box 1 (HMGB1)，一种与损伤相关的分子模式，尽管 CFTR 丢失。影响巨噬细胞功能，即 CF 气道环境，其特征是微摩尔浓度中性粒细胞弹性蛋白酶 (NE) 是一种压倒性的刺激物，可激活强烈的促炎症反应我们提出 NE 是肺部疾病的生物标志物。 CF 的进展颠覆了巨噬细胞的功能，从保护性转变为促炎性，但 NE 重新编程巨噬细胞的机制尚不完全清楚。在此应用中，我们提出了一种前所未有的假设机制来解释 NE 如何改变巨噬细胞的功能是细胞外的NE被巨噬细胞快速内吞。降解的、具有蛋白水解活性的 NE 定位于细胞质结构域和细胞核，从而导致细胞因子表达和巨噬细胞胞外陷阱（MET）释放增加，因此，NE。像特洛伊木马一样破坏巨噬细胞的功能，我们将在初级阶段测试这个假设。来自健康志愿者和 CF 受试者的人血单核细胞衍生的巨噬细胞。具体目标如下：目标 1. 确定 NE 是否会降解 HDAC，导致下游靶标乙酰化，从而导致 TNFα 的转录上调和 HMGB1 的释放。目标 2. 评估 NE 蛋白酶活性和/或 NE 产生的活性氧是否增加线粒体和核 MET 的释放。相关性：该项目的结果将定义 NE 用来促进的新机制持续气道炎症是 CF 发病和死亡的主要原因，并确定治疗目标阻止肺部疾病持续进展的新疗法。