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病理生物学的见解是逐步有效的靶向药物治疗方法 尚未实现。项目＃2解决了编码烟酰胺的基因NAMPT的新作用 在ARDS和呼吸机诱导的肺损伤（VILI）的病理生物学中，磷酸贝糖基转移酶（VILI）。我们确定了 使用过量机械的细胞和临床前模型的基因组密集型方法的NAMPT 压力和vili。我们证明，超级机械应力会诱导稳健的NAMPT表达和 分泌（细胞外NAMPT或ENAMPT）是一种新型的ARDS生物标志物。我们已经证明了nampt 展示5'启动子单核苷酸多态性（SNP），显着改变NAMPT启动子活性 会议大大提高了ARDS的敏感性和ARDS严重性（无通风机的日子减少， 增加了ARDS死亡率）。我们确定ENAMPT是VILI病理生物学的重要参与者 产生嗜中性肺泡炎和肺损伤，而enampt供应量的减少（中和） 抗体，siRNA，NAMPT +/-小鼠）急剧减轻了临床前vili /ards中肺损伤的严重程度 型号。最后，我们证明了NAMPT表达通过可靠的表达在空间上定位，并且 肺内皮细胞（EC）的分泌，用ENAMPT诱导了Toll -like受体4（TLR4）的新型配体 NFκB转录活性和炎症性肺损伤。虽然Enampt显然是一个有吸引力的 ARDS/VILI目标，关键差距在理解NAMPT介导的肺病理生物学的理解中仍然存在问题 需要解决重大转化为ICU疗法的解决方案。项目＃2将解决这些关键差距 专注于机械应力挑战的肺EC（分泌的ENAMPT的主要来源）， 有助于增加血管通透性（ARDS中的主要热目标）以及关键影响 在VILI发育中，ENAMPT与肺EC TLR4结合。项目＃2特定目标（SAS）旨在 通过SA＃1阐明机械应力介导的遗传和表观遗传调节来解决这些差距 NAMPT表达（转录因子，CpG脱甲基化，3'UTR miRNA结合，NAMPT SNP）。基于 在令人兴奋的初步数据上，SA＃2将通过caspase介导的裂解来定义对ENAMPT分泌的调节 和ABC转运蛋白。与核心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将定义参与TLR4的NAMPT结合的结构/功能机制和TLR4的影响 在配体 - 受体相互作用上编码SNP的NAMPT编码SNP。最后，使用临床前vili/ards模型， 包括一种新型的条件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数据转化为可行的信息，以减轻Vili/ards和 定义降低NAMPT表达和分泌的影响（STAT5/HIF2α抑制剂），ENAMPT发射 （中和抗体）和TLR4拮抗作用（肽抑制剂）。项目2将提高对 NAMPT参与VILI/ARDS，并促进针对重病的个性化疗法的应用。