Mechanisms of essential calcium signaling during gastric epithelial wound healing

 描述(申请人提供)：消化性溃疡疾病和上消化道刺激是美国医疗保健系统和世界范围内的重大负担，在很大程度上是由摄入的非类固醇抗炎药(NSAIDs)和致病菌H.Pylori引起的上消化道毒性。我们工作的基本原理是，改进的或替代的治疗策略可以建立在理解胃防御机制的基础上，以应对这些挑战。初步研究表明，细胞内和细胞外钙离子动员在胃表面上皮细胞修复中起重要作用。我们假设这种钙动员是修复的中央调节因子，它是多种激动剂调节修复过程的基础，并且这一途径可以被非类固醇抗炎药和幽门螺杆菌破坏。这项应用的目的是利用新发现的钙信号作为先兆，以确定介导胃修复的潜在上游和下游机制，并作为一种工具来研究非类固醇抗炎药和/或幽门螺杆菌是否通过这些途径影响修复。我们开创了光学技术的先河，可以实时产生局部损伤并持续量化修复。我们专注于体内局部病变修复的工作将被外推到评估溃疡的实验，以增强与临床观察到的胃病理的相关性，并研究胃有机物质作为一种潜在的强大的胃上皮功能的新模型。我们的第一个目标是确定在受损组织中启动细胞外刺激钙动员和胃修复的信号。我们将重点关注TFF2和前列腺素，它们被证明是胃修复所必需的，但它们在钙信号转导中的作用尚未得到测试。第二个目的是建立钙依赖的细胞内信号通路的层次结构，这些信号通路对于修复是必不可少的。实验基于初步发现，磷脂酶C和蛋白激酶C是完整胃有效修复所必需的。我们的第三个目标是评估执行胃修复工作的钙依赖信号的下游靶点。我们将评估NHE2Na/H交换器作为TFF2下游效应因子在胃修复中的作用。我们还将评估钙离子对肌动蛋白细胞骨架、紧密连接以及在修复过程中调节这些细胞结构的小GTP酶的调节。所有的目的都集中在环氧合酶(COX)/前列腺素途径被非甾体抗炎药抑制的作用上，第一和第三个目的另外考察了幽门螺杆菌感染引起的所研究途径的变化。这些结果将为了解胃发病机制的早期因素提供一个独特的窗口，这些因素有可能导致新的靶点和新的干预策略，以最大限度地减少胃损伤和加速愈合。

 DESCRIPTION (provided by applicant): Peptic ulcer disease and upper GI irritation are a significant burden in the US healthcare system and worldwide, caused in large part by the upper GI toxicity of ingested non-steroidal anti-inflammatory drugs (NSAIDs) and the pathogen H. pylori. The rationale for our work is that improved or alternative therapeutic strategies can be based on understanding the mechanisms of gastric defense rallied against such challenges. Preliminary studies show an essential role for both intracellular and extracellular Ca2+ mobilization in gastric repair of the surface epithelium. We hypothesize that this Ca2+ mobilization is a central regulator of repair that underlies the effects of diverse agonists shown to modulate the repair process, and that this pathway can be disrupted by NSAIDs and H. pylori. The objective of this application is to use the newly identified Ca2+ signals as heralds to identif the underlying upstream and downstream mechanisms mediating gastric repair, and as a tool to investigate if NSAIDs and/or H. pylori compromise repair via these pathways. We have pioneered optical technologies that allow real-time creation of focal damage and continual quantification of repair. Our work focusing on the repair of focal lesions in vivo will be extrapolated to experiments evaluating ulceration to enhance relevance to gastric pathologies observed clinically, and to study of gastric organoids as a potentially powerful new model of gastric epithelial function. Our first aim is to identify the initiating extracellular signals in damaged tissue that stimulate Ca2+ mobilization and gastric repair. We will focus on TFF2 and prostaglandins, shown to be essential to gastric repair, but whose role in Ca2+ signaling is untested. The second aim establishes the hierarchy of Ca2+-dependent intracellular signaling pathways that are essential to allow repair. Experiments are based on preliminary findings that show phospholipase C, and protein kinase C are required for efficient gastric repair in the intact stomach. Our third aim evaluates the downstream targets of Ca2+- dependent signaling that perform the work of gastric repair. We will evaluate the role of the NHE2 Na/H exchanger, shown to be essential for gastric repair as a downstream effector of TFF2. We will also evaluate Ca2+ -dependent regulation of the actin cytoskeleton, tight junctions, and the small GTPases that regulate these cellular structures during repair. All aims focus on the role of the cyclooxygenase (COX)/prostaglandin pathway inhibited by NSAIDs, and the first and third aims additionally examine changes in the studied pathways caused by infection with H. pylori. The outcomes will provide a unique window into understanding early factors in gastric pathogenesis, which have potential to lead to new targets and new strategies for interventions that can minimize gastric damage and speed healing.