NF-kB Regulation, ICAM-1 Expression, and Lung Injury

NF-kB 调节、ICAM-1 表达和肺损伤

• 批准号: 8318828
• 负责人: CHINNASWAMY TIRUPPATHI
• 金额: $ 28.44万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8318828
• 关键词: 4-ethoxymethylene-2-phenyl-2-oxazoline-5-one; A20 protein; Acute Lung Injury; Address; Adhesions; Binding; Binding Proteins; Biological Assay; CabP2; Cells; Cloning; Complement; Complementary DNA; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; DNA Binding; Data; Dominant-Negative Mutation; Dreams; EF Hand Motifs; Ectopic Expression; Elements; Endothelial Cells; Esthesia; Gene Expression; Gene Targeting; Genes; Genetic Models; Genetic Transcription; Host Defense; Human; Hydrogen Peroxide; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Response; Injury; Instruction; Intercellular adhesion molecule 1; Intervention; Knock-out; Knockout Mice; Ligation; Link; Lung; Lung Inflammation; Mediating; Modeling; Molecular; Mus; NADPH Oxidase; NF-kappa B; Natural Immunity; Nucleic Acid Regulatory Sequences; Oxidants; Pain; Phenotype; Phosphorylation; Positioning Attribute; Prevention; Promoter Regions; Protein Biosynthesis; Proteins; Puncture procedure; RIPK2 gene; Reactive Oxygen Species; Regulation; Reporter; Repression; Repressor Proteins; Research Proposals; Role; Seminal; Sepsis; Signal Pathway; Signal Transduction; Signaling Pathway Gene; Stimulus; TNF gene; Testing; Therapeutic; Thrombin; Time; Transcription Repressor/Corepressor; Vascular Endothelial Cell; Wild Type Mouse; base; chromatin immunoprecipitation; derepression; gene repression; genetic regulatory protein; glucosylthiazolidine-4-carboxylic acid; in vivo; inhibitor/antagonist; intraperitoneal; knock-down; lung injury; lung vascular injury; mortality; neutrophil; novel; prevent; promoter; research study; response; stem; sulfated glycoprotein 2; transcription factor

PROJECT SUMMARY (See instructions): Transcription factor NF-KB activation-dependent ICAM-1 expression in pulmonary endothelial cells results in neutrophil (PMN) adhesion-mediated lung vascular injury. NF-KB-dependent expression of the NF-KB negative regulatory protein A20 inhibits unrestrained activation of NF-KB and thereby controls the inflammatory response. The pre-transcriptional mechanisms involved in regulating A20 gene expression are poorly understood. This research proposal stems from our seminal discovery of multiple "GTCA" sequences (ORE elements) forthe Ca2+ binding transcriptional repressor protein Dream in the 5'-regulatory region of both human and mouse A20 genes. Thus, in Project 3, we will address the crucial role of Dream in regulating the expression of A20 in pulmonary microvessels and thereby controlling NF-KB activation, ICAM-1 expression, and PMN adhesion-mediated lung vascular injury. We cloned a 1.5 kb human A20 promoter sequence linked to reporter and showed robust promoter activity in response to thrombin and TNF-a which was blocked by co-expression of Dream. In addition, we showed that Ca2+ influx through TRPC4 channels or oxidant-sensitive TRPM2 channels induced the de-repression of A20 transcription in response to thrombin and H2O2 respectively. Importantly, we observed that in Dream knock-out (Dream -/-) mice, basal A20 expression was enhanced and this was associated with prevention of thrombin-TNF-a- or LPS-induced ICAM-1 expression in lungs of Dream''' mice. Further, LPS-induced acute lung injury was markedly reduced in Dream -/- mice. In addition, we observed that NF-KB activation in response to inflammatory stimuli was suppressed in Dream-/- mice. Based on these novel findings, in Aim 1, we will test the hypothesis that repressor Dream regulates the expression of the NF-KB inhibitor A20 by interacting with 5'-regulatory region of the A20 gene. In Aim 2, we will test the hypothesis that H2O2 activation of oxidant-sensitive TRPM2 channels and induces Ca2+ influx leading to de-repression of A20 transcription and its consequent suppression of NF-KB activity. In Aim 3, we will test the hypothesis that Dream in vivo regulates the basal expression of A20 and thereby suppresses unrestrained activation of NF-KB utilizing Dream-/- mice. With this understanding of Dream, we will be in the position to develop therapeutic strategies that can target Dream function and thereby prevent acute lung injury without compromising innate immunity.

项目总结（见说明）： 肺内皮细胞中转录因子NF-κ B活化依赖性ICAM-1表达导致中性粒细胞（PMN）粘附介导的肺血管损伤。NF-κ B负调节蛋白A20的NF-κ B依赖性表达抑制NF-κ B的无限制活化，从而控制炎症反应。参与调节A20基因表达的转录前机制知之甚少。这项研究计划源于我们在人类和小鼠A20基因的5 '调控区发现了多个Ca 2+结合转录抑制蛋白Dream的“GTCA”序列（ORE元件）。因此，在项目3中，我们将讨论Dream在调节肺微血管中A20的表达，从而控制NF-κ B活化、ICAM-1表达和PMN粘附介导的肺血管损伤中的关键作用。我们克隆了一个1.5 kb的人A20启动子序列连接到报告，并显示出强大的启动子活性，凝血酶和TNF-α，这是封闭的梦想的共表达。此外，我们发现，通过TRPC 4通道或氧化剂敏感性TRPM 2通道的Ca 2+内流分别诱导凝血酶和H2 O2对A20转录的去抑制。重要的是，我们观察到在Dream基因敲除（Dream -/-）小鼠中，基础A20表达增强，这与Dream-/-小鼠肺中凝血酶-TNF-α-或LPS-诱导的ICAM-1表达的预防有关。此外，在Dream -/-小鼠中，LPS诱导的急性肺损伤显著减少。此外，我们观察到在Dream-/-小鼠中，响应于炎症刺激的NF-κ B活化被抑制。基于这些新的发现，在目的1中，我们将检验阻遏物Dream通过与A20基因的5 '调控区相互作用来调控NF-κ B抑制剂A20表达的假设。在目标2中，我们将检验H2 O2激活氧化剂敏感性TRPM 2通道并诱导Ca 2+内流导致A20转录去抑制及其随后抑制NF-κ B活性的假设。在目的3中，我们将检验Dream在体内调节A20的基础表达从而抑制利用Dream-/-小鼠的NF-κ B的无限制活化的假设。有了对梦的这种理解，我们将能够开发出针对梦功能的治疗策略，从而在不损害先天免疫的情况下预防急性肺损伤。