Critical Role of NAMPT and Toll-Like Receptor 4 in Inflammation and Mechanical Ventilator-Induced Lung Injury (VILI)

NAMPT 和 Toll 样受体 4 在炎症和机械呼吸机引起的肺损伤 (VILI) 中的关键作用

• 批准号: 10094248
• 负责人: Joe G. N. Garcia
• 金额: $ 45.99万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-02-05 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10094248
• 关键词: 3' Untranslated Regions; ATP-Binding Cassette Transporters; Acute Lung Injury; Address; Adult Respiratory Distress Syndrome; Alveolitis; Animal Model; Attenuated; Binding; Biological Markers; Blood; Blood Circulation; Caspase; Cell model; Code; Critical Care; Critical Illness; Data; Development; Endothelial Cells; Epigenetic Process; Epithelial; Exhibits; Functional disorder; Gene Targeting; Genes; Genetic; Genetic Polymorphism; Genetic Transcription; Genomics; HMGB1 gene; Immunity; Inflammation; Inflammatory; Knockout Mice; Ligands; Ligation; Lung; Mechanical Stress; Mechanical Ventilators; Mechanical ventilation; Mediating; MicroRNAs; Modeling; Molecular; Molecular Biology; Molecular Genetics; Mus; PBEF Gene; Participant; Patients; Pattern; Peptides; Pharmacotherapy; Post-Translational Protein Processing; Pre-Clinical Model; Predisposition; Protein Chemistry; Regulation; Role; Severities; Signal Transduction; Single Nucleotide Polymorphism; Small Interfering RNA; Source; Stat5 protein; Structure; TLR4 gene; Therapeutic; Tissues; Translating; Translations; United States; Vascular Permeabilities; Ventilator; Ventilator-induced lung injury; base; delta protein; demethylation; design; epigenetic regulation; extracellular; improved; individualized medicine; inflammatory lung disease; inhibitor/antagonist; insight; lung injury; mortality; neutralizing antibody; neutrophil; nicotinamide phosphoribosyltransferase; novel; novel marker; novel therapeutic intervention; pre-B-cell colony-enhancing factor protein; pre-clinical; promoter; receptor; therapeutic target; transcription factor

PROJECT #2 SUMMARY: Insights into ARDS and VILI pathobiology have been incremental and effective targeted pharmacotherapies have not yet been realized. Project #2 addresses the novel role of NAMPT, the gene encoding nicotinamide phosphoribosyltransferase, in the pathobiology of ARDS and ventilator–induced lung injury (VILI). We identified NAMPT by genomic–intensive approaches utilizing cellular and preclinical models of excessive mechanical stress and VILI. We demonstrated that excessive mechanical stress induces robust NAMPT expression and secretion (extracellular NAMPT or eNAMPT) serves as a novel ARDS biomarker. We have shown that NAMPT exhibits 5' promoter single nucleotide polymorphisms (SNPs) that significantly alter NAMPT promoter activity and confer significantly increased ARDS susceptibility and ARDS severity (reduced ventilator-free days, increased ARDS mortality). We determined that eNAMPT is an essential participant in VILI pathobiology directly producing a neutrophilic alveolitis and lung injury whereas reductions in eNAMPT availability (neutralizing antibodies, siRNAs, NAMPT+/- mice) dramatically attenuates the severity of lung injury in preclinical VILI /ARDS models. Finally, we demonstrated that NAMPT expression is spatially-localized with robust expression and secretion by lung endothelial cells (ECs) with eNAMPT a novel ligand for the Toll–like receptor 4 (TLR4) inducing NFκB transcriptional activities and inflammatory lung injury. Although eNAMPT is clearly an attractive ARDS/VILI target, critical gaps remain in the understanding of NAMPT-mediated lung pathobiology, issues which need to be addressed for robust translation to an ICU therapy. Project #2 will address these key gaps focusing on mechanical stress-challenged lung EC (a major source of secreted eNAMPT), on eNAMPT contribution to increases in vascular permeability (a major therapeutic target in ARDS), and on the critical influence of eNAMPT binding to lung EC TLR4 in VILI development. Project #2 Specific Aims (SAs) are designed to address these gaps with SA #1 elucidating mechanical stress-mediated genetic and epigenetic regulation of NAMPT expression (transcription factors, CpG demethylation, 3'UTR miRNA binding, NAMPT SNPs). Based on exciting preliminary data, SA #2 will define regulation of eNAMPT secretion by caspase-mediated cleavage and ABC transporters. With Core B (Molecular Biology & Genetics Core) and Core D (Protein Chemistry Core), SA #3 will define structure/function mechanisms involved in NAMPT binding of TLR4 and the influence of TLR4 and NAMPT coding SNPs on ligand–receptor interactions. Finally, utilizing preclinical VILI/ARDS models, including a novel conditional EC–specific and lung epithelium-specific NAMPT KO mice (Core C: Pre-clinical Animal Model Core), SA #4 will translate SA #1- #3 data into actionable information to attenuate VILI/ARDS and define the impact of reduced NAMPT expression and secretion (STAT5/HIF2α inhibitors), eNAMPT elimination (neutralizing antibodies), and TLR4 antagonism (peptide inhibitors). Project #2 will advance understanding of NAMPT participation in VILI/ARDS and promote the application of individualized therapies for the critically ill.

项目 #2 摘要： 对 ARDS 和 VILI 病理学的深入了解是渐进且有效的靶向药物治疗 尚未实现。项目#2 解决了 NAMPT（编码烟酰胺的基因）的新作用。 我们确定了 ARDS 和呼吸机引起的肺损伤 (VILI) 病理学中的磷酸核糖基转移酶。 NAMPT 通过基因组密集型方法，利用过度机械的细胞和临床前模型 我们证明，过度的机械应力会诱导强大的 NAMPT 表达和 分泌（细胞外 NAMPT 或 eNAMPT）可作为一种新型 ARDS 生物标志物。 表现出 5' 启动子单核苷酸多态性 (SNP)，可显着改变 NAMPT 启动子活性 并显着增加 ARDS 易感性和 ARDS 严重程度（减少不使用呼吸机的天数， ARDS 死亡率增加）我们确定 eNAMPT 是 VILI 病理学的直接重要参与者。 产生中性粒细胞性肺泡炎和肺损伤，同时减少 eNAMPT 的可用性（中和 抗体、siRNA、NAMPT+/- 小鼠）可显着减轻临床前 VILI /ARDS 中肺损伤的严重程度 最后，我们证明了 NAMPT 表达具有稳健的表达和空间定位。 eNAMPT 是 Toll 样受体 4 (TLR4) 的新型配体，可诱导肺内皮细胞 (EC) 分泌 NFκB 转录活性和炎症性肺损伤虽然 eNAMPT 显然很有吸引力。 ARDS/VILI 目标，对 NAMPT 介导的肺部病理学、问题的理解仍存在关键差距 需要解决这些问题才能有力地转化为 ICU 治疗 项目 #2 将解决这些关键差距。 关注机械应力​​挑战的肺 EC（分泌性 eNAMPT 的主要来源），关于 eNAMPT 增加血管通透性（ARDS 的主要治疗目标）的贡献，以及关键影响 VILI 开发中 eNAMPT 与肺 EC TLR4 的结合旨在实现项目 #2 的特定目标 (SA)。 通过 SA #1 阐明了机械应力介导的遗传和表观遗传调控，解决了这些差距 基于 NAMPT 表达（转录因子、CpG 去甲基化、3'UTR miRNA 结合、NAMPT SNP）。 根据令人兴奋的初步数据，SA #2 将定义 caspase 介导的切割对 eNAMPT 分泌的调节 和 ABC 转运蛋白，具有核心 B（分子生物学和遗传学核心）和核心 D（蛋白质化学核心）， SA #3 将定义涉及 TLR4 的 NAMPT 结合的结构/功能机制以及 TLR4 的影响 和 NAMPT 编码配体-受体相互作用的 SNP，最后利用临床前 VILI/ARDS 模型， 包括一种新型条件性 EC 特异性和肺上皮特异性 NAMPT KO 小鼠（核心 C：临床前 动物模型核心），SA #4 将把 SA #1-#3 数据转化为可操作的信息，以减轻 VILI/ARDS 和 定义 NAMPT 表达和分泌减少（STAT5/HIF2α 抑制剂）、eNAMPT 消除的影响 （中和抗体）和 TLR4 拮抗作用（肽抑制剂） 项目 #2 将促进对 TLR4 拮抗作用的理解。 NAMPT参与VILI/ARDS，推动危重症个体化治疗的应用。