THE MOLECULAR REGULATION OF NEUTROPHIL CALCIUM SIGNALING IN ACUTE LUNG INJURY

 描述（由申请人提供）：急性肺损伤（ALI）是一种破坏性疾病，其特征是肺泡上皮屏障破坏，并导致肺水肿和低氧血症。目前没有有效的疾病修饰疗法，支持性护理仍然是医疗管理的支柱。最终，ALI导致多达30%的患者发生多器官功能障碍和/或死亡。中性粒细胞对于宿主防御细菌显然是重要的，然而毒性中性粒细胞介质如活性氧自由基、颗粒酶和中性粒细胞胞外陷阱也通过引起内皮、实质和肺泡损伤而促成肺损伤的发病机制。中性粒细胞是操纵炎症反应的主要靶点，因此了解引导中性粒细胞反应的分子机制至关重要。克莱门斯博士的长期研究目标是了解细胞内信号通路如何在急性肺损伤和其他炎症性疾病期间调节中性粒细胞活化。钙是一种进化上保守的信号信使，是多种信号通路的中心组分。在免疫细胞中，细胞质钙的增加通过钙库操纵的钙进入（SOCE）来控制，其中钙释放激活的钙（CRAC）通道允许当内质网（ER）库耗尽时细胞外钙的流入。STIM蛋白是ER钙“传感器”，其感测钙消耗并直接门控CRAC通道的孔亚基奥赖。关于STIM、奥赖或中性粒细胞中钙信号传导的分子调节知之甚少。本提案的目的是确定调节中性粒细胞中SOCE的分子途径，并确定这些途径如何调节ALI期间中性粒细胞的活化。核心假设是：1）STIM钙传感器和奥赖钙通道协同调节中性粒细胞钙信号传导，2）钙依赖性中性粒细胞活化对于ALI的诱导至关重要。目的1和2中拟定的研究将使用一组独特的中性粒细胞特异性Stim或奥赖基因缺失小鼠，确定中性粒细胞中SOCE所需的分子机制，并确定中性粒细胞钙信号传导在急性肺损伤中的作用。目的 3将利用一种新的方法与人源化小鼠来测试这些假设在人类中性粒细胞。该项目与NIH和NIAID的任务相关，因为这些研究将 阐明以前未探索的中性粒细胞钙信号传导途径，可能导致新的治疗方法来治疗急性肺损伤和其他炎症过程。

 DESCRIPTION (provided by applicant): Acute lung injury (ALI) is a devastating disease characterized by alveolar-epithelial barrier disruption and resultant pulmonary edema and hypoxemia. There are currently no effective disease-modifying therapies and supportive care remains the mainstay of medical management. Ultimately, ALI leads to multi-organ dysfunction and/or death in up to 30% of patients. Neutrophils are clearly important for host defense against bacteria, however toxic neutrophil mediators such as reactive oxygen radicals, granule enzymes and neutrophil extracellular traps also contribute to the pathogenesis of lung injury by causing endothelial, parenchymal and alveolar injury. Neutrophils are prime targets for manipulating the inflammatory response and therefore it is critical to understand the molecular mechanisms that guide neutrophil responses. Dr. Clemens' long-term research goal is to understand how intracellular signaling pathways regulate neutrophil activation during acute lung injury and other inflammatory diseases. Calcium is an evolutionarily conserved signaling messenger that is a central component of multiple signaling pathways. In immune cells, increases in cytoplasmic calcium are controlled via store operated calcium entry (SOCE), where calcium release-activated calcium (CRAC) channels allow influx of extracellular calcium when endoplasmic reticulum (ER) stores are depleted. STIM proteins are ER calcium "sensors" which sense calcium depletion and directly gate ORAI, a pore subunit of the CRAC channel. Little is known about STIM, ORAI or the molecular regulation of calcium signaling in neutrophils. The objectives of this proposal are to define the molecular pathways that regulate SOCE in neutrophils and to determine how these pathways modulate neutrophil activation during ALI. The central hypotheses are: 1) STIM calcium sensors and ORAI calcium channels cooperate to regulate neutrophil calcium signaling, and 2) calcium-dependent neutrophil activation is critical for induction of ALI. The studies proposed in Aims 1 and 2 will identify the molecular machinery required for SOCE in neutrophils and determine the role of neutrophil calcium signaling in acute lung injury using a unique set of mice with neutrophil specific deletion in Stim or Orai genes. Aim 3 will utilize a novel approach with humanized mice to test these hypotheses in human neutrophils. This project is relevant to the missions of the NIH and NIAID since these studies will elucidate previously unexplored pathways of calcium signaling in neutrophils that may lead to new therapeutic approaches to treat acute lung injury and other inflammatory processes.