Microvascular Permeability and Matrix Fibrinogen Degradation in Trauma

描述（由申请人提供）：增加基质蛋白降解产物，包括纤维蛋白原降解产物（FDPs），与创伤患者全身性炎症和多器官衰竭的发展有关。除了作为止血功能的生物标志物，fdp还与炎症、病理性血管生成和伤口愈合有关。纤维蛋白原及其蛋白水解片段作为一个整体对血管通透性的影响已被报道；然而，关于基于分子构象的特定产物的独特功能的知识有限，对其作用机制的了解甚至更少。我们最近的研究产生了一个有趣的发现，在严重烧伤后，动物血浆和组织中纤维蛋白原-3 （3C）的c端片段（fdp的主要成分）水平升高。这种反应伴随着全身性水肿和死亡率，与其他关于创伤患者循环FDP水平和全身性炎症严重程度相关的报道一致。然后我们检查了3C的微血管作用，发现它直接导致了高通透性。这些数据表明了一种有趣的可能性，即含有3C的基质降解产物在血管炎症中起致病因素的作用。本项目的目的是明确3C诱导的微血管高通透性在烧伤相关炎症条件下的作用和分子机制。中心假设认为，烧伤引起的基质降解和纤维蛋白溶解会升高3C，从而靶向内皮屏障并促进血浆泄漏。3c诱导的高通透性是通过整合素偶联RhoA信号转导导致内皮细胞收缩和连接解离而发生的。本文提出了三个具体目标：1)表征3C在调节微血管通透性中的作用；2)鉴定介导3c诱导的高通透性的内皮受体；3)研究3C增加内皮细胞通透性的分子机制。我们将使用互补的方法，将分子技术结合到生理实验中，深入分析肠系膜作为胃肠道系统的代表性微血管的通透性调节，这是烧伤的主要目标。从这项研究中获得的数据将为胃肠道微血管屏障损伤的分子基础提供新的机制见解。公共卫生相关性：全身性炎症损伤是严重创伤和烧伤患者死亡和发病的主要原因。作为炎症的主要组成部分，微血管屏障功能障碍发生在多器官系统，导致血浆泄漏和组织水肿，这给创伤患者带来了危及生命的问题。开发有效的治疗策略需要深入了解导致损伤过程的终点细胞反应。目前，我们对微血管屏障调节的分子基础的了解相当有限，并且由于细胞生物学转化为系统病理生理学的困难，这一领域的研究受到阻碍。这项工作具有创新性，不仅因为它有可能确定一种介导微血管炎症损伤的新分子途径，而且因为它实现了新的实验模型和技术，可以在临床相关条件下将分子反应与血管功能结合起来。本研究的数据将为微血管屏障功能的病理生理调节提供新的机制见解。识别导致微血管渗漏的关键分子将有助于开发新的抗炎症损伤治疗靶点。

DESCRIPTION (provided by applicant): Increased matrix protein degradation products, including fibrinogen degradation products (FDPs), have been associated with the development of systemic inflammation and multiple organ failure in trauma patients. Besides a well-recognized role as biomarkers of hemostasis function, FDPs have been implicated in inflammation, pathological angiogenesis, and wound healing. A vascular permeability effect of fibrinogen and its proteolytic fragments as a whole has been reported; however, there is limited knowledge regarding specific products which function distinctively based on molecular conformations, and even less is known about their mechanisms of actions. Our recent studies have produced an interesting finding that the plasma and tissue levels of the C-terminal fragment of fibrinogen-3 (3C), a major component of FDPs, are increased in animals after severe burn. The response was coupled with systemic edema and mortality, consistent with other reports regarding correlated circulating FDP levels and severities of systemic inflammation in trauma patients. We then examined the microvascular effects of 3C and found that it directly caused hyperpermeability. These data indicate an intriguing possibility that matrix degradation products containing 3C act as pathogenic factors in vascular inflammation. The objective of this project is to define the role and molecular mechanisms of 3C- induced microvascular hyperpermeability under burn-relevant inflammatory conditions. The central hypothesis states that burn-elicited matrix degradation and fibrinolysis elevate 3C which targets the endothelial barrier and promotes plasma leak. The 3C-induced hyperpermeability occurs via integrin- coupled RhoA signal transduction leading to endothelial cell contraction and junction dissociation. Three specific aims are proposed to 1) characterize the role of 3C in regulating microvascular permeability; 2) identify the endothelial receptor that mediates 3C-induced hyperpermeability; and 3) examine the molecular mechanisms by which 3C increases endothelial permeability. We will use complimentary approaches that incorporate molecular techniques into physiological experiments for an in-depth analysis of permeability regulation in the mesentery as representative microvasculature of the GI system, a major target of burn. Data derived from this study will provide novel, mechanistic insights into the molecular basis of barrier injury in gastrointestinal microvessels. PUBLIC HEALTH RELEVANCE: Systemic inflammatory injury is a major cause of mortality and morbidity in patients with severe trauma and burn. As a cardinal component of inflammation, microvascular barrier dysfunction occurs in multiple organ systems rendering plasma leak and tissue edema, which imposes a life-threatening problem to trauma patients. Development of effective therapeutic strategies requires an in-depth understanding of the end-point cellular reactions responsible for the injurious process. Currently, our knowledge regarding the molecular basis of microvascular barrier regulation is rather limited, and research in this area has been hampered by the difficulties in translating cell biology to systems pathophysiology. The proposed work is innovative not only because it has the potential to identify a new molecular pathway that mediates microvascular inflammatory injury, but because it implements novel experimental models and techniques that can integrate molecular reactions with vascular functions under clinically relevant conditions. Data derived from this study will provide new mechanistic insights into the pathophysiological regulation of microvascular barrier function. Identification of key molecules leading to microvascular leakage will assist in the development of new therapeutic targets against inflammatory injury.