Disintegrin Metalloprotease and Endothelial Permeability

描述(申请人提供)：ADAM15是一种跨膜糖蛋白，是去整合素金属蛋白酶的特征，具有释放受体分子和调节细胞-细胞/基质黏附的能力。ADAM15表达增加与转移性癌症、动脉粥样硬化和炎症性肠病的发生有关。我们最近的工作确定了该分子的一个新功能，即作为一种能够增加血管内皮细胞通透性的促炎因子。进一步的研究表明，在细菌感染或败血症刺激下，ADAM15在肺部上调，导致中性粒细胞渗透和肺水肿。ADAM15介导的屏障损伤的组织特异性效应和潜在机制尚未确定。该项目的总体目标是了解ADAM15在炎症中的表达和功能的分子调控。目的有两个：1)研究ADAM15活性在脓毒症中的功能和调控途径；2)阐明ADAM15诱导高通透性的分子机制。需要检验的中心假设是脓毒症诱导ADAM15在转录和翻译后水平上调。ADAM15通过胞浆结构域激活Src信号转导通路，增加内皮细胞-细胞连接的通透性，而去整合素调节局部粘连和/或连接结构的金属蛋白酶脱落是另一种途径。这一新概念将通过一系列互补性研究得到验证，这些研究整合了使用动物模型进行的体内生理分析和培养细胞的体外分子分析。创新的实验模型和分子工具将被构建和测试。来自拟议工作的数据不仅将建立蛋白水解酶分子生物学的新理论，而且还将为脓毒症和急性炎症的病理生理学提供新的机制见解。此外，这项研究对确定和开发有效治疗炎症性疾病的新的治疗靶点具有潜在的影响。 公共卫生相关性：全身炎症损伤是败血症患者死亡和发病的主要原因，每年有超过75万美国人受到影响。尽管对其发病机制有先进的理论，但在治疗或预防方面仍未取得重大突破。临床进展有限的部分原因是由于疾病的复杂性和我们对其终点细胞机制的不完全了解，有效的治疗方法发展不足。微血管通透性增加是各种损伤炎症反应的共同终点。内皮屏障功能障碍导致重要器官，特别是肺部的血浆渗漏和白细胞渗入，导致呼吸窘迫和多器官功能衰竭。内皮屏障损伤在急性全身性炎症中的重要作用突出了开发靶向治疗的必要性。这项拟议的工作很重要，因为它将为临床相关脓毒症条件下内皮细胞屏障功能的分子控制和生理调节提供新的机制见解。此外，这项研究对确定和开发新的治疗靶点以改进炎性损伤的治疗和预防具有潜在的影响。

DESCRIPTION (provided by applicant): ADAM15 is a transmembrane glycoprotein characteristic of disintegrin metalloprotease with the ability to shed receptor molecules and regulate cell-cell/matrix adhesions. Increased ADAM15 expression is associated with the development of metastatic cancer, atherosclerosis, and inflammatory bowl disease. Our recent work has identified a novel function of this molecule as a pro-inflammatory factor capable of increasing vascular endothelial permeability. Further studies suggest that ADAM15 is upregulated in the lungs during bacterial infection or septic stimulation, contributing to neutrophil infiltration and pulmonary edema. The tissue-specific effect and underlying mechanisms of ADAM15-mediated barrier injury have not been characterized. The overall goal of this project is to understand the molecular control of ADAM15 expression and function in inflammation. Two specific aims are proposed: 1) to characterize the functions and regulatory pathways of ADAM15 activity in sepsis, and 2) to elucidate the molecular mechanisms by which ADAM15 induces hyperpermeability. The central hypothesis to be tested is that sepsis induces ADAM15 upregulation at the transcriptional and posttranslational levels. ADAM15 increases endothelial cell-cell junction permeability by activating Src signal transduction via its cytoplasmic domain, whereas disintegrin regulation of focal adhesions and/or metalloprotease shedding of junctional structures serve as alternative pathways. This novel concept will be validated through a series of complementary studies that integrate in vivo physiological analyses using animal models and in vitro molecular assays in cultured cells. Innovative experimental models and molecular tools will be constructed and tested. Data derived from the proposed work will not only establish a novel theory in protease molecular biology, but also provide new mechanistic insights into the pathophysiology of sepsis and acute inflammation. In addition, the study has potential impact on the identification and development of novel therapeutic targets for effective treatment of inflammatory disease. PUBLIC HEALTH RELEVANCE: Systemic inflammatory injury is a major cause of mortality and morbidity in patients with sepsis, which affects more than 750,000 Americans annually. Despite the advanced theories regarding its pathogenesis, there has been no major breakthrough in treatment or prevention. The limited clinical progress is partially attributed to the underdevelopment of effective therapies, owing to the complexity of the disease and our incomplete understanding of its endpoint cellular mechanisms. Increased microvascular permeability represents a common endpoint of inflammatory response to various types of injury. Endothelial barrier dysfunction causes plasma leakage and leukocyte infiltration in vital organs, especially in the lungs, leading to respiratory distress and multiple organ failure. The essential role of endothelial barrier injury in acute systemic inflammation highlights the need to develop target-directed treatments. The proposed work is important because it will provide novel mechanistic insights into the molecular control and physiological regulation of endothelial barrier function under clinically relevant septic conditions. In addition, the study has potential impact on the identification and development of new therapeutic targets for improved treatment and prevention of inflammatory injury.