DISTAL EVENTS IN THE HEPATOTOXICITY OF ENDOTOXIN

描述(改编自调查人员摘要)：败血症 由此产生的革兰氏阴性细菌脂多糖(LPS)暴露是一种 主要的临床问题与高死亡率和无数的 病理生理改变，包括肝功能衰竭。在体内，内毒素 触发一系列事件，涉及细胞和可溶性的 最终导致肝功能障碍和肝功能丧失的炎症 肝实质细胞。库普弗细胞、中性粒细胞和血小板 在内毒素诱导的细胞因子中起关键作用 肝脏损伤。研究人员最近在老鼠身上进行的研究表明 体内暴露于内毒素时激活的凝血酶对 发病机制。此外，它的关键作用独立于它的能力 在血管系统中形成阻塞性纤维蛋白凝块，这引起了人们对 其重要作用可能是通过受体介导的 与细胞的相互作用。凝血酶似乎在一连串的 导致肝功能障碍的炎性事件，也就是说， 实质细胞损伤发生的时间。对这种“远端”的认识 事件“很可能在开发有效的临床上有用 干预。 这项提案的总体目标是了解通过哪些机制 凝血酶参与大鼠肝损伤。在目标1中，体内研究 并在隔离的肝脏中执行以定义时间关系 凝血酶的关键作用(S)与显性肝损伤的发病 目标2中的实验将检验凝血酶通过以下途径起作用的假设 提高肝实质细胞对损伤的敏感性 在脂多糖暴露期间。研究将使用隔离的、灌流的 培养物中的肝脏或HCs以确定凝血酶暴露是否增加 HCS对可能造成伤害的侮辱的敏感性。通过哪些机制 凝血酶对HCS的作用将被讨论。凝血酶的假说 通过刺激关键炎症细胞参与肝损伤 释放有毒产品将在目标3中解决。这将是 部分是通过评估凝血酶对人体造成伤害的可能性来实现的 从中性粒细胞、血小板耗尽的大鼠肝脏灌流 或Kupffer细胞，然后用内毒素处理。实验采用 人巨噬细胞与中性粒细胞、血小板或库普弗细胞共培养将 确定凝血酶是否导致受体介导的细胞毒释放 来自关键炎症细胞的因子和/或通过蛋白水解性作用 生物激活一种可溶性炎症介质(目标4)。来自这些的结果 研究将揭示凝血酶和细胞之间的关键相互作用 它们在脂多糖暴露过程中导致肝脏损伤的机制。这个 这些研究的长期目标是发展对 炎症反应将导致有效的干预措施 在降低革兰氏阴性器官衰竭患者死亡率方面的作用 细菌性败血症。

DESCRIPTION (Adapted from the Investigator's Abstract): Septicemia with resultant exposure to gram-negative bacterial lipopolysaccharide (LPS) is a major clinical problem associated with high mortality and a myriad of pathophysiological alterations, including liver failure. In vivo, LPS triggers a cascade of events involving cellular and soluble mediators of inflammation that culminates in liver dysfunction and loss of integrity of hepatic parenchymal cells. Kupffer cells, neutrophils and platelets are among the cellular mediators critical to the development of LPS-induced liver injury. The investigator's recent studies in rats indicate that thrombin activated during LPS exposure in vivo is essential to the pathogenesis. Moreover, its critical role is independent of its ability to form obstructive fibrin clots in the vasculature, which raises interest in the possibility that its important action occurs through receptor-mediated interaction with cells. Thrombin appears to act late in the cascade of inflammatory events that leads to hepatic dysfunction, that is, close in time to the genesis of parenchymal cell injury. Knowledge of such "distal events" is likely to be useful in developing effective clinical intervention. The overall goal of this proposal is to understand the mechanisms by which thrombin participates in liver injury in the rat. In Aim 1, studies in vivo and in isolated livers will be performed to define the temporal relationship between thrombin's critical action(s) and the onset of overt liver injury. Experiments in Aim 2 will test the hypothesis that thrombin acts by increasing the sensitivity of hepatic parenchymal cells (HCs) to injury during LPS exposure. Studies will be performed using isolated, perfused livers or HCs in culture to determine if thrombin exposure increases susceptibility of HCs to potentially injurious insults. Mechanisms by which thrombin acts on HCs will be addressed. The hypothesis that thrombin participates in hepatic injury by stimulating critical inflammatory cells to release toxic products will be addressed in Aim 3. This will be accomplished in part by evaluating thrombin's potential to cause injury to perfused livers from rats which had been depleted of neutrophils, platelets or Kupffer cells and then treated with LPS. Experiments employing cocultures of HCs with neutrophils, platelets or Kupffer cells will determine if thrombin causes the receptor-mediated release of a cytotoxic factor from a critical inflammatory cell and/or acts by proteolytically bioactivating a soluble inflammatory mediator (Aim 4). Results from these studies will reveal critical interactions between thrombin and cells and mechanisms by which they result in liver injury during LPS exposure. The long-term goal of these studies is to develop an understanding of inflammatory responses that will result in interventions that are effective in reducing mortality in people with organ failure from gram-negative bacterial sepsis.