Angiopoietin Mechanisms in Sepsis

描述(申请人提供)：败血症在美国每年影响30多万人，每年治疗费用超过160亿美元，与30%的急性死亡率有关，预计发病率将增加，使这种疾病成为一个主要的公共卫生问题。尽管进行了密集的基础和临床研究工作，但对这些患者的护理仍然主要是支持性的。对其发病机制的新见解可以应用于患者护理是迫切需要的。我们的实验室正在研究作为一种细胞类型的内皮细胞，它在脓毒症中受到损害，进而介导对宿主的进一步损害。具体地说，我们正在研究血管生成素，这是一种分泌蛋白，通过Tie-2受体作用于内皮细胞，作为这种疾病中血管通透性的调节因素。我们发现，在患有严重脓毒症的人类受试者中，循环血管生成素-2(Ang-2)升高，过量的Ang-2足以促进内皮细胞和其他健康成年小鼠的渗漏。最近发表的对Ang-2基因缺失小鼠的研究进一步证实了Ang-2是炎症过程中通透性增加的介质。我们还发现，血管生成素-1(Ang-1)在结构上与Ang-2相似，但具有相反的信号作用，可以防止小鼠内毒素血症中的血管渗漏。为了达到这种显著的保护作用，Ang-1必须激活Rho-GTP酶家族中一种名为p190RhoGAP的调节蛋白。在体内耗尽p190RhoGAP足以消除Ang-1对内毒素的保护作用。这些发现使我们假设，血管生成素通过Ang-2过多和/或不足导致的明显的高通透性是脓毒症的原因。为了探索这个中心问题，我们将把啮齿动物败血症、盲肠结扎和穿孔的金标准模型应用于Ang-2缺失的小鼠，以及使用有效的Ang-2中和抗体或表达Ang-1的腺病毒治疗的野生型小鼠，以评估存活率、渗透性和细胞反应(目标1)。考虑到Ang-1通过激活p190RhoGAP来对抗内毒素介导的通透性的意想不到的能力，我们将进一步探索这种屏障防御的机制(目标2)。最后，我们将回到脓毒症患者身上，在我们的急诊科进行一项前瞻性研究，以评估早期循环Ang-2和Ang-1变化对死亡、休克和其他不良结局的预测价值(目标3)。目标1和3是高度翻译的，目标2将实质上阐明脓毒症中的内皮屏障防御机制。试点数据强烈支持每个目标，我们组建的团队在脓毒症领域的合作研究(基础科学、临床研究、病理学和流行病学)方面拥有丰富的经验，测试每个目标都将具有成本效益。这些有针对性的研究将提高我们对脓毒症发病机制的理解，并有可能为严重脓毒症患者提供新的诊断和治疗选择。与公共卫生相关脓毒症是一种发病率不断上升的常见、昂贵和致命的疾病；它可能已经占美国所有死亡人数的10%，使其成为一个主要的公共卫生威胁，其影响只会越来越大。基于大量的初步数据，我们认为血管生成素可能在本病的发病机制中起关键作用。拟议的实验将使我们能够评估这些蛋白质在脓毒症中的诊断和治疗潜力。

DESCRIPTION (provided by applicant): Sepsis affects more than 300,000 individuals in the US annually, costs more than $16 billion/yr to treat, is associated with an acute mortality of 30%, and is expected to increase in incidence, making this disease a major public health concern. Care of these patients has remained primarily supportive despite intensive basic and clinical research efforts. Novel insights into its pathogenesis that can be applied to patient care are sorely needed. Our laboratory is studying the endothelium as a cell type that is injured in sepsis and that, in turn, mediates further damage to the host. Specifically, we are studying the angiopoietins a class of secreted proteins that act on endothelial cells through the Tie-2 receptor as regulators of vascular permeability in this disease. We have found that circulating angiopoietin-2 (Ang-2) is elevated in human subjects with severe sepsis, and that excess Ang-2 is sufficient to promote leak across endothelial cells and in otherwise healthy adult mice. More recently published studies of Ang-2 null mice have further validated Ang-2 as a mediator of increased permeability during inflammation. We have also found that angiopoietin-1 (Ang-1), which is similar in structure to Ang-2 but has opposite signaling effects, prevents vascular leak in murine endotoxemia. To achieve this remarkable protective effect, Ang-1 must activate a regulatory protein in the Rho-GTPase family called p190RhoGAP. In vivo depletion of p190RhoGAP is sufficient to abrogate the protection conferred by Ang-1 against endotoxin. These findings have led us to hypothesize that derangements in angiopoietins via excess Ang-2 and/or insufficient Ang-1 contribute to the marked hyperpermeability of sepsis. To explore this central question, we will apply the gold-standard model of rodent sepsis, cecal ligation and perforation, to Ang- 2 null mice, and wild-type mice treated either with a potent Ang-2 neutralizing antibody or with an Ang-1 expressing adenovirus to evaluate survival, permeability, and cellular responses (Aim 1). Given the unexpected ability of Ang-1 to counteract endotoxin-mediated permeability via p190RhoGAP activation, we will explore this mechanism of barrier defense further (Aim 2). Finally, we will return to human subjects with sepsis to perform a prospective study based in our emergency department to evaluate the predictive value of early changes in circulating Ang-2 and Ang-1 for death, shock, and other adverse outcomes (Aim 3). Aims 1 and 3 are highly translational, and Aim 2 will substantially elucidate endothelial barrier defense mechanisms in sepsis. Pilot data are strongly supportive of each aim, the team we have assembled has extensive experience in collaborative research (basic science, clinical research, pathology and epidemiology) in the area of sepsis, and testing each aim will be cost-efficient. These targeted studies will improve our understanding of the pathogenesis of sepsis, and potentially lead to novel diagnostic and therapeutic options for patients with severe sepsis. PUBLIC HEALTH RELEVANCE Sepsis is a common, costly, and lethal disease of increasing incidence; it may already account for 10% of all deaths in the US, making it a major public health threat whose impact is only likely to grow. Based on extensive preliminary data, we believe that angiopoietins may be critically involved in the pathogenesis of this disease. The proposed experiments will enable us to evaluate the diagnostic and therapeutic potential of these proteins in sepsis.