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Understanding Severe Asthma Using an Experimental Model

使用实验模型了解严重哮喘

基本信息

批准号：
10215597
负责人：
Anuradha Ray
金额：
$ 49.49万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-01-07 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10215597
关键词：
Adoptive Transfer Adrenal Cortex Hormones Antigens Asthma Biological Assay Bronchoalveolar Lavage CD4 Positive T Lymphocytes CXCL10 gene Cells Characteristics Chymase Complex DNA Data Dendritic Cells Disease Dose Down-Regulation Experimental Models Follow-Up Studies Frequencies GTP-Binding Proteins Gene Expression Genes Glucocorticoid Receptor Goals Human Immune System Diseases Inflammation Inhalation Interferon Type II Interleukin-10 Interleukin-12 Lead Lung Mediating Messenger RNA Molecular Mucous body substance Mus Myosin Heavy Chains Oral Outcome Measure Pathogenesis Pathway interactions Patients Peptide Hydrolases Phenocopy Phenotype Play Production Proteins Publishing Refractory Reporter Reporting Resolution Role SLPI gene STAT1 gene Sampling Signal Transduction Smooth Muscle Myocytes Structure of parenchyma of lung T cell response T-Lymphocyte TBK1 gene Testing Th1 Cells Time Up-Regulation airway hyperresponsiveness airway remodeling asthma exacerbation asthma model asthmatic c-myc Genes chemokine chromatin immunoprecipitation cysteinyl leukotriene receptor cysteinyl-leukotriene draining lymph node effector T cell inhibitor/antagonist insight mRNA Expression macrophage mast cell monocyte mouse model novel peripheral blood promoter pulmonary function recruit response standard of care transcription factor transcriptome sequencing

项目摘要

Severe asthma is a complex heterogeneous disease, which poorly responds to corticosteroids (CS), the current standard of care for asthma. In a previous study, we reported that bronchoalveolar lavage (BAL) cells in >50% of subjects with severe asthma (SA) secrete high levels of IFN-γ from CD4+ T cells despite treatment of the subjects with high doses of inhaled CS, often combined with oral CS. In a recent follow-up study to understand the molecular mechanisms that promote a persistent Th1high phenotype in SA, we have shown an essential role for the transcription factor IRF5 in promoting IL-12 production from lung dendritic cells (DCs) and macrophages (Mφs) to elicit high levels of IFN-γ production from T cells. In another study published recently, we show that SA subjects with a Th1high profile harbor higher levels of expression of the Th1-recruiting chemokine CXCL10 in their airways compared to mild asthma subjects, strongly associated with IFN-γ expression, frequency of oral CS use, and expression of mast cell-specific proteases. Chromatin immunoprecipitation (ChIP) assay used to determine high CXCL10 levels in SA revealed recruitment of both STAT1 to and GR to the CXCL10 promoter with no accompanying suppression of CXCL10 mRNA expression. While CS were unable to suppress IFN-γ-induced CXCL10 mRNA and protein levels in monocytes, IL-10 completely inhibited CXCL10 gene expression in the context of IFN-γ and CS, an effect not realized in many severe asthmatics. In new data generated using RNA-seq approaches, we show downregulation of the negative regulator of TGF-β1-mediated signaling, Smad7, and upregulation of c-Myc, in lungs of mice subjected to the SA model, which may impact mast cells and airway remodeling. Indeed in our recent report we have shown a strong correlation between CXCL10 mRNA and mast cell signatures in humans and mice. Mast cells produce cysteinyl leukotrienes (CysLTs), and our RNA-seq data show that IFN-γ selectively upregulates expression of the CysLT receptor, CySLTR2, in the lung. Collectively, these studies lead us to hypothesize that the IRF-5-IFN-γ-CXCL10 axis in the context of CS induces a feed-forward loop promoting a Th1high phenotype that impacts lung function and airway remodeling involving mast cells. Although refractory to CS, the Th1high state is sensitive to IL-10. To test this hypothesis we will: Aim 1. Establish IRF5 in lung APCs as a sustainer of Th1 (IFN-γ) effector function and identify IRF5 SNPs in humans. Aim 2. Determine the role of specific IFN-γ-regulated molecules and mast cells in disease pathogenesis in the SA model. Aim 3. Investigate defective IL-10 production in T cells in humans with SA and study inhibition by IL-10 of CS- refractory CXCL10 gene expression induced by IFN-γ.

重症哮喘是一种复杂的异质性疾病，对皮质类固醇(CS)的反应很差。目前哮喘的护理标准。在之前的研究中，我们报告了支气管肺泡灌洗(BAL)细胞尽管接受治疗，50%的重症哮喘患者仍能从CD+T细胞分泌高水平的干扰素-γ 大剂量吸入CS的受试者，常合并口服CS。在最近一项关于了解促进SA持续Th1高表型的分子机制，我们已经展示了转录因子IRF5在促进肺树突状细胞(DC)产生IL-12中的重要作用巨噬细胞(MφS)诱导T细胞产生高水平的干扰素-γ。在最近发表的另一项研究中，我们发现Th1水平较高的SA患者体内Th1-Recruiting的表达水平较高呼吸道趋化因子CXCL10与轻度哮喘患者的比较，与干扰素-γ密切相关肥大细胞特异性蛋白水解酶的表达、口服CS的使用频率和表达。染色质免疫沉淀(CHIP)法检测SA中高水平的CXCL10发现两者都有募集 STAT1至CXCL10启动子，GR至CXCL10启动子，但不伴随抑制CXCL10 mRNA的表达。而CS不能抑制干扰素-γ诱导的单核细胞CXCL10mRNA和蛋白水平，IL-10 在干扰素-γ和CS的背景下，完全抑制CXCL10基因的表达，这是许多人没有意识到的效果严重的哮喘患者。在使用rna-seq方法生成的新数据中，我们显示了对转化生长因子-β1介导的信号负调控因子Smad7与小鼠肺组织c-Myc的上调受SA模型影响，可能影响肥大细胞和气道重塑。事实上，在我们最近的报告中我们已经在人类和小鼠中证明了CXCL10mRNA和肥大细胞特征之间的强烈相关性。肥大细胞产生半胱氨酰白三烯(CysLts)，我们的rna-seq数据表明，干扰素-γ选择性地上调肺组织CysLT受体CySLTR2的表达。总而言之，这些研究引导我们假设，在CS的背景下，IRF-5-干扰素-γ-CXCL10轴诱导前馈循环，促进Th1高表型，影响肺功能和气道重塑涉及肥大细胞。虽然对CS不敏感，但Th1高状态对IL-10敏感。为了检验这一假设，我们将：目的1.在肺泡巨噬细胞中建立IRF5作为Th1型(干扰素-γ)效应功能的维持者，并鉴定IRF5SNP。人类。目的2.确定特异性干扰素-γ调节分子和肥大细胞在慢性粒细胞白血病发病机制中的作用。 SA模型。目的3.研究SA患者T细胞产生IL-10缺陷及IL-10对CS-10的抑制作用。干扰素-γ诱导难治性CXCL10基因表达。