p38MAP kinase-dependent mechanisms of fever-enhanced acute lung injury

描述（由申请人提供）：这是一项竞争性更新申请，旨在扩展共同主要研究者正在进行的研究，重点是发热和发热抑制对急性肺损伤（ALI）的影响。尽管低潮气量通气被普遍接受用于支持ALI患者，但此类患者的死亡率仍约为40%。我们确定发热/体温过高是另一个潜在的重要贡献者的ARDS发病机制。据报道，大约一半的重症监护室患者和高达90%的ALI/ARDS患者会发生发热。ARDS的特征是富含嗜酸性粒细胞的炎症、内皮屏障丧失和上皮损伤。我们以前的研究表明，暴露于发热范围（FRH，~39.5oC）的高温会大大增强这些致病过程。本申请提出了一种机制研究，其集中于FRH增强的ALI的p38丝裂原活化蛋白激酶（MAPK）依赖性机制的作用。我们已经表明，暴露于FRH增强先天免疫功能的一部分，通过增强中性粒细胞（PMN）的招聘。这种作用加速了病原体清除，但也增加了侧支组织损伤，特别是在肺中。基于新的数据，这一建议的重点是肺血管内皮细胞和应激激活的MAPK，p38的核心作用。初步数据表明：（1）FRH至少部分通过自磷酸化直接激活p38;（2）FRH直接改变p38构象、激酶活性、细胞定位和底物选择性。本提案将检验以下假设： (1)FRH通过多种途径激活p38，包括通过p38的构象变化促进其自身磷酸化和通过激活上游激酶;（2）FRH诱导的p38构象变化将导致p38分子间对接、亚细胞分布和下游信号事件的不同模式，其将独特地改变下游底物磷酸化模式，对肺部炎症和损伤具有重要影响;（3）在伴有发热的ALI患者中，抑制p38或下游信号通路将比抑制发热更好地改善预后。我们将使用原代培养的HMVEC-Ls和ALI小鼠模型：（1）阐明FRH激活p38的分子机制，（2）分析FRH如何改变p38的亚细胞分布和底物磷酸化谱，和（3）测试p38信号通路阻断在不损害病原体清除的情况下减少FRH增强的肺损伤的潜力。我们期望这些研究的结果将阐明FRH诱导的p38信号转导与ALI相关的机制，并提供有关不成比例底物磷酸化的重要信息，以确定在ALI/ARDS中测试的替代治疗靶点。我们希望这些发现在减轻其他组织的炎症和损伤方面也有更广泛的应用。

DESCRIPTION (provided by applicant): This is a competitive renewal application to extend ongoing research by the co-Principal Investigators that is focused on the effects of fever and fever suppression on acute lung injury (ALI). Despite the general acceptance of low tidal volume ventilation for support of patients with ALI, mortality in such patients remains ~40%. We identified fever/hyperthermia as another potentially important contributor to ARDS pathogenesis. Fever is reported to occur in about half of patients admitted to intensive care units and in up to 90% of those with ALI/ARDS. ARDS is characterized by neutrophil-rich inflammation, loss of endothelial barrier, and epithelial injury. Our previous studies show that exposure to hyperthermia in the febrile range (FRH, ~39.5oC) profoundly augments each of these pathogenic processes. The current application proposes a mechanistic study that focuses on the role of p38 mitogen-activated protein kinase (MAPK)-dependent mechanisms of FRH- augmented ALI. We have shown that exposure to FRH augments innate immune function in part by enhancing neutrophil (PMN) recruitment. This effect accelerates pathogen clearance, but also increases collateral tissue injury, especially in the lung. Based on new data, this proposal focuses on the central role of pulmonary vascular endothelium and the stress-activated MAPK, p38. Preliminary data suggests that (1) FRH directly activates p38 at least in part through autophosphorylation and (2) that FRH directly alters p38 conformation, kinase activity, cellular localization, and substrate selectivity. This proposal will test the following hypotheses: (1) FRH activates p38 through multiple pathways including through a conformational change in p38 that facilitates its autophosphorylation and by activating upstream kinases; (2) the FRH-induced conformational changes in p38 will result in distinct patterns of p38 intermolecular docking, subcellular distribution, and downstream signaling events, which will uniquely modify downstream substrate phosphorylation patterns with important consequences for lung inflammation and injury; and (3) in the patient with ALI and fever, inhibition of p38 or downstream signaling pathways will improve outcome better than suppressing fever. We will use primary cultured HMVEC- Ls and mouse models of ALI to: (1) elucidate the molecular mechanisms by which FRH activates p38, (2) analyze how FRH modifies p38 subcellular distribution and substrate phosphorylation profile, and (3) test the potential of p38 signaling pathway blockade to reduce FRH-augmented lung injury without impairing pathogen clearance. We expect that the results of these studies will clarify the mechanisms of FRH-induced p38 signaling relevant to ALI and provide essential information about disproportionate substrate phosphorylation that will identify alternative therapeutic targets to test in ALI/ARDS. We expect that these findings will have wider applications in mitigating inflammation and injury in other tissues as well.