Modulation of Lung Inflammation and Cell Death by VIP

VIP 对肺部炎症和细胞死亡的调节

• 批准号: 6752407
• 负责人: SAMI I SAID
• 金额: $ 18.9万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6752407
• 关键词: CD95 molecule; NMDA receptors; adult respiratory distress syndrome; apoptosis; cysteine endopeptidases; cytokine; endotoxins; genetically modified animals; glutamate receptor; immunofluorescence technique; inflammation; laboratory mouse; lipopolysaccharides; lung injury; neuropeptides; nuclear factor kappa beta; oxidative stress; paraquat; pathologic process; pentosyltransferase; protein structure function; terminal nick end labeling; vasoactive intestinal peptide; xanthine oxidase; xanthines

DESCRIPTION (provided by applicant): The broad, long-term objective is to achieve a deeper understanding of the pathogenesis of acute lung injury (ALI) causing the Acute Respiratory Distress Syndrome (ARDS), and to develop more effective means of its treatment and prevention. Hypotheses to be tested: 1) ALI due to oxidant stress is attributable in large measure to two fundamental and interrelates processes: inflammation and cell death by apoptosis or necrosis; 2) Excitotic mechanisms are a major pathogenic factor in cell death due to oxidant stress; and 3) Vasoactive intestinal peptide (VIP) and the related pituitary adenylate cyclase activating peptide (PACAP) can effectively reduce or prevent at least some forms of ALI, by blocking these critical processes, and by promoting cell survival pathways. Specific Objectives fall into two broad categories: a) Establish the role of inflammatory, apoptotic and excitotoxic mechanisms in the pathogenesis of ALI. This goal is justified by the need to establish criteria for evaluating the second objective; b) Determine the efficacy, potency, and specificity of VIP & PACAP in the protection against ALI, with special reference to the above pathogenic mechanisms, and the receptors and pathways mediating this protection. 1) Document the importance of key inflammatory mechanisms in ALI, including NF- KB activation, an the expression and production of major pro- and anti-inflammatory cytokines. NF-KB activation, will be assessed by evaluating evidence for such activation (nuclear displacement of p50 and p65 subunits) in models of ALI, and correlating the activation with the degree of injury, and the inhibition of activation with protection from injury. Transgenic mice with targeted deletion of NF-KB subunit p50 will be examined for possible resistance to injury. 2) Assess the contribution of apoptotic and necrotic cell death to ALI by: morphologiv evidence, TUNEL-testing, caspase activation, poly (ADP-ribose) polymerase PARP) activation (which often leads to necrosis) or its degradation (a marker of apoptosis), and the degree of protection afforded by anti-apoptotic measures, including caspase inhibitors and upregulation of anti-apoptosis protein bcl2, relative to pro-apoptotic BAX. 3) Validate the concept that endogenous glutamate receptor activation is an important mechanism of oxidant-induced ALI. 4) Determine the mechanisms, receptors, and pathways of lung protection by VIP and PACAP in these experimental models of injury: a) Excitotoxic lung injury due to excessive glutamate receptors due to excessive activation of glutamate receptors, a model that we recently characterized and is uniquely suited to a critical analysis of cell death mechanisms in lung injury; b) oxidant injury due to paraquat and xanthine + xanthine oxidase; and c) endotoxin lipopolysaccharide (LPS)-and Fas- induced apoptotic cell death. Injury will be induced in isolated lungs, cell preparations (alveolar macrophages, alveolar epithelial & pulmonary endothelial cells), and in vivo model of endotoxemia. Whenever available, we will study selected strains of transgenic mice either lacking or overexpressing key factors regulating the inflammatory response, death pathways and survival signals. In searching for the pathways of injury and its modulation by VIP/PACAP, we will use the newly introduced gene microarrays.

描述（由申请人提供）：广泛的长期目标是 更深入地了解急性肺损伤（ALI）的发病机制 导致急性呼吸窘迫综合症（ARDS），并发展更多 其治疗和预防的有效手段。待检验的假设：1) 氧化应激引起的 ALI 在很大程度上可归因于两个基本因素 并相互关联的过程：炎症和细胞凋亡或细胞死亡 坏死； 2）兴奋机制是细胞死亡的主要致病因素 由于氧化应激； 3) 血管活性肠肽 (VIP) 和 相关垂体腺苷酸环化酶激活肽（PACAP）可以有效 通过阻止这些关键因素，减少或预防至少某些形式的 ALI 过程，并通过促进细胞存活途径。具体目标下降 分为两大类： a) 确定炎症、细胞凋亡和 ALI 发病机制中的兴奋性毒性机制。这一目标的合理性在于 需要制定评估第二个目标的标准； b) 确定 VIP 和 PACAP 的功效、效力和特异性 预防 ALI，特别是上述致病菌 机制，以及介导这种保护的受体和途径。 1） 记录 ALI 中关键炎症机制的重要性，包括 NF-KB 激活，主要亲和物质的表达和产生 抗炎细胞因子。 NF-KB 激活，将通过评估来评估 这种激活的证据（p50 和 p65 亚基的核位移） ALI 模型，并将激活与损伤程度相关联，以及 抑制激活并保护免受伤害。转基因小鼠 将检查 NF-KB 亚基 p50 的靶向删除是否可能产生耐药性 以致受伤。 2) 评估细胞凋亡和坏死细胞死亡对细胞凋亡的贡献 ALI 通过：形态学证据、TUNEL 测试、半胱天冬酶激活、多聚 （ADP-核糖）聚合酶 PARP）激活（通常导致坏死）或其 降解（细胞凋亡的标志），以及所提供的保护程度 抗凋亡措施，包括半胱天冬酶抑制剂和上调 抗凋亡蛋白 bcl2，相对于促凋亡 BAX。 3) 验证 内源性谷氨酸受体激活是一个重要机制的概念 氧化剂诱发的 ALI。 4) 确定其机制、受体和途径 在这些实验损伤模型中通过 VIP 和 PACAP 进行肺保护：a) 过量谷氨酸受体导致的兴奋性中毒性肺损伤 谷氨酸受体的激活，这是我们最近描述的一个模型 非常适合对肺细胞死亡机制进行严格分析 受伤; b) 百草枯和黄嘌呤+黄嘌呤氧化酶造成的氧化损伤；和 c)内毒素脂多糖(LPS)和Fas诱导的细胞凋亡。 将在离体肺、细胞制品（肺泡）中诱导损伤 巨噬细胞、肺泡上皮细胞和肺内皮细胞）以及体内 内毒素血症模型。只要有可能，我们将研究选定的菌株 转基因小鼠缺乏或过度表达调节基因的关键因子 炎症反应、死亡途径和生存信号。在寻找中 损伤途径及其 VIP/PACAP 调节，我们将使用新的 引入基因微阵列。