Pulmonary endothelial glycocalyx degradation causes ARDS during sepsis

描述(由申请人提供)：急性呼吸窘迫综合征(ARDS)是一种严重的(30%死亡率)危重疾病，每年影响美国19万多名患者。尽管进行了40多年的研究，但人们对这种疾病的发展过程知之甚少，也没有具体的治疗方法。虽然脓毒症是ARDS的一个众所周知的原因，但败血症ARDS发生的潜在机制尚不确定。ARDS发展过程中的一个关键步骤是肺内皮屏障功能障碍，该屏障将血液与肺间质分开。对全身(非肺)血管系统的研究表明，正常的内皮屏障功能依赖于完整的糖萼--一层薄薄的蛋白多糖、糖蛋白和糖胺聚糖排列在血管管腔内。新出现的数据表明，脓毒症的炎症介质降低了系统内皮细胞的糖基化反应，导致屏障功能障碍。然而，肺内皮细胞糖基化反应在脓毒症和ARDS中的作用尚不清楚。我们假设脓毒症诱导肺内皮细胞糖基化降解，导致屏障功能障碍和ARDS。由于体外糖萼的结构经常是异常的，这一假说将通过体外和体内模型来探索：隔离的灌流小鼠肺和闭合胸腔活体小鼠肺显微镜。将测定脂多糖(LPS，脓毒症的一种模型)引起的肺糖萼降解的剂量-反应和时间过程。将使用多光子活体显微镜绘制血管节段性(动脉与微血管与静脉)的糖萼丢失模式，并与脓毒症期间内皮细胞高通透性的节段性模式进行比较。肿瘤坏死因子1和乙酰肝素酶的作用将通过药物抑制剂和转基因小鼠模型来探索，作为脂多糖诱导的糖萼丢失的媒介。肝素是一种肝素酶的抑制剂，它的治疗益处将被探索。总而言之，这项研究有望通过生理学(隔离的、灌流的小鼠肺)和专门的显微镜(多光子活体显微镜)相结合的方法，更好地了解与常见临床疾病(脓毒症相关的ARDS)相关的、知之甚少的结构(肺内皮细胞糖萼)。此外，这项研究发现，肝素酶是一种具有吸引力的治疗靶点，用于治疗一种尚未进行特殊治疗的疾病。 公共卫生相关性：败血症是一种常见的严重疾病，由身体对压倒性感染的反应引起。败血症通常会导致肺部血管渗漏，使肺部充满血液和液体(一种通常是致命的疾病，称为急性呼吸窘迫综合征或ARDS)。我们正在研究如何预防ARDS，特别是通过试图保护肺血管内一层薄薄但非常重要的糖层。

DESCRIPTION (provided by applicant): The acute respiratory distress syndrome (ARDS) is a severe (> 30% mortality) critical illness that affects over 190,000 patients in the United States each year. Despite over 40 years of study, little is known about how this disease develops, and no specific treatment exists. While sepsis is a well-known cause of ARDS, the mechanisms underlying the development of septic ARDS are uncertain. A key step in the development of ARDS is dysfunction of the pulmonary endothelial barrier, which separates blood from the lung interstitium. Studies of the systemic (non-pulmonary) vasculature have revealed that proper endothelial barrier function is dependent on an intact glycocalyx-a thin layer of proteoglycans, glycoproteins, and glycosaminoglycans lining the vascular lumen. Emerging data suggest that inflammatory mediators of sepsis degrade the systemic endothelial glycocalyx, causing barrier dysfunction. The role of the pulmonary endothelial glycocalyx in sepsis and ARDS, however, has been unexplored. We hypothesize that sepsis induces pulmonary endothelial glycocalyx degradation, leading to barrier dysfunction and ARDS. As glycocalyx structure is often aberrant in-vitro, this hypothesis will be explored using ex-vivo and in-vivo models: the isolated, perfused mouse lung and closed-chest, intravital mouse lung microscopy. The dose-response and time course of lipopolysaccharide (LPS, a model of sepsis) induced pulmonary glycocalyx degradation will be determined. The vascular segmental (arterial vs. microvascular vs. venous) pattern of glycocalyx loss will be mapped using multiphoton intravital microscopy and compared to the segmental pattern of endothelial hyperpermeability during sepsis. The role of tumor necrosis factor 1 and heparanase will be explored as mediators of LPS-induced glycocalyx loss, using pharmacologic inhibitors and transgenic mouse models. The therapeutic benefit of heparin, an inhibitor of heparanase, will be explored. In summary, this investigation promises a better understanding of a poorly-understood structure (the pulmonary endothelial glycocalyx) with relevance to a common clinical condition (sepsis-associated ARDS) via a combined physiologic (isolated, perfused mouse lung) and specialized microscopic (multiphoton intravital microscopy) approach. Furthermore, this investigation identifies heparanase as an attractive therapeutic target in a disease that yet lacks specific treatment. PUBLIC HEALTH RELEVANCE: Sepsis is a common, severe condition caused by the body's response to an overwhelming infection. Sepsis often causes blood vessels in the lungs to become leaky, flooding the lungs with blood and fluid (an often-fatal condition called the acute respiratory distress syndrome or ARDS). We are studying how to prevent ARDS, specifically by trying to protect a thin, yet very important, layer of sugars lining the blood vessels in the lung.