CEBPD-Mediated Mechanisms of Glucocorticoid Insensitivity in Severe Asthma

CEBPD 介导的严重哮喘糖皮质激素不敏感机制

• 批准号: 9914313
• 负责人: Blanca E Himes
• 金额: $ 63.59万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9914313
• 关键词: Affect; Agonist; Airway Disease; American; Asthma; Binding; Binding Sites; Biological Assay; Biological Markers; Biology; Bronchodilator Agents; Budesonide; CCAAT-enhancer-binding protein-delta; Cell Differentiation process; ChIP-seq; Characteristics; Clinical; Complex; Cyclic AMP; DNA Binding; Data; Disease; Enzyme-Linked Immunosorbent Assay; Epigenetic Process; Event; Exposure to; Gene Expression; Gene Targeting; Genes; Genetic Transcription; Glucocorticoid Receptor; Glucocorticoids; Health Expenditures; Histones; Immune; Individual; Inflammatory; Inflammatory Response; Interleukin-1 beta; Life; Luciferases; Lung diseases; Measures; Mediating; Messenger RNA; Modification; Morbidity - disease rate; Patients; Pattern; Pharmaceutical Preparations; Physiological; Physiological Processes; Play; Post-Translational Protein Processing; Proteins; Proteomics; RNA analysis; Relaxation; Reporter; Research Personnel; Resources; Role; Signal Transduction; Small Interfering RNA; Stimulus; Structure; Testing; Therapeutic; Tissues; Transcript; Transcription Alteration; Validation; Work; airway hyperresponsiveness; asthmatic patient; base; biomarker development; cytokine; differential expression; health care service utilization; insight; knock-down; mRNA Expression; novel therapeutics; overexpression; respiratory smooth muscle; response; time use; transcription factor; transcriptome; transcriptome sequencing; transcriptomics

PROJECT SUMMARY Asthma is an episodic inflammatory disease that affects over 25 million Americans and manifests as airway hyperresponsiveness to specific environmental stimuli. Despite effective medications that control asthma in most individuals, 15% respond inadequately and suffer life-threatening exacerbations. A feature shared by most patients with severe disease is glucocorticoid insensitivity, a poorly understood physiological process. The airway smooth muscle (ASM), which plays an important role in asthma, is a target of glucocorticoids that act in part via modulation of gene transcription, alteration of histone post-translational modifications, and inhibition of transcription factors. CCAAT/Enhancer Binding Protein D (CEBPD) is a pleiotropic glucocorticoid- responsive transcription factor that regulates inflammatory responses, cell differentiation and tissue remodeling. Based on our data showing that CEBPD expression is (1) lower in fatal asthma vs. non-asthma ASM, (2) induced with glucocorticoid treatment in non-asthma ASM but unchanged in fatal asthma ASM, and that (3) decreasing CEBPD via knockdown resulted in increased IL1β-induced NFkB-luciferase expression with glucocorticoid stimulation, our central hypothesis states that low CEBPD levels in ASM elicits transcriptomic and epigenetic modifications that decrease glucocorticoid sensitivity. We will test this hypothesis by using the following unbiased and complementary `omic approaches to study the effect of CEBPD on glucocorticoid response in ASM from fatal asthma and non-asthma donors: (1) RNA-Seq to identify transcriptomic effects, (2) proteomics to determine global histone post-translational modification effects, and (3) ChIP-Seq to measure transcription factor binding of NFkB and the glucocorticoid receptor. An integrated analysis of RNA-Seq, proteomics and ChIP-Seq results will identify major targets of CEBPD that influence glucocorticoid response, whose role will be confirmed via NFkB-luciferase assays. Our project will offer insights into transcriptional and epigenetic signatures that are characteristic of severe asthma, enable glucocorticoid sensitivity biomarker development, and offer therapeutic insights that benefit the most vulnerable individuals with asthma.

项目概要 哮喘是一种间歇性炎症性疾病，影响超过 2500 万美国人，表现为气道炎症 对特定环境刺激的过度反应。尽管有有效的药物可以控制哮喘 对于大多数人来说，15% 的人反应不充分，病情恶化，危及生命。共享的一个功能 大多数重症患者对糖皮质激素不敏感，这是一个人们知之甚少的生理过程。 气道平滑肌（ASM）在哮喘中起重要作用，是糖皮质激素的靶标， 部分通过基因转录的调节、组蛋白翻译后修饰的改变以及 转录因子的抑制。 CCAAT/增强子结合蛋白 D (CEBPD) 是一种多效性糖皮质激素 调节炎症反应、细胞分化和组织的反应性转录因子 重塑。根据我们的数据显示，与非哮喘相比，致死性哮喘中的 CEBPD 表达 (1) 较低 ASM，(2) 在非哮喘 ASM 中用糖皮质激素治疗诱导，但在致死性哮喘 ASM 中没有变化，以及 (3) 通过敲低减少 CEBPD 导致 IL1β 诱导的 NFkB 荧光素酶表达增加 糖皮质激素刺激，我们的中心假设指出 ASM 中低 CEBPD 水平引发转录组学 以及降低糖皮质激素敏感性的表观遗传修饰。我们将使用以下方法来检验这个假设 遵循公正和互补的组学方法来研究 CEBPD 对糖皮质激素的影响 来自致死性哮喘和非哮喘捐赠者的 ASM 反应：(1) RNA-Seq 鉴定转录组效应，(2) 蛋白质组学来确定全局组蛋白翻译后修饰效应，以及 (3) ChIP-Seq 来测量 NFkB 和糖皮质激素受体的转录因子结合。 RNA-Seq 的综合分析， 蛋白质组学和 ChIP-Seq 结果将确定影响糖皮质激素反应的 CEBPD 主要靶标， 其作用将通过 NFkB-荧光素酶测定得到证实。我们的项目将提供对转录和 严重哮喘特征的表观遗传特征使糖皮质激素敏感性生物标志物成为可能 发展，并提供有利于最脆弱的哮喘患者的治疗见解。