Interleukin-18 and Intestinal Barrier Function Following Alcohol and Burn Injury

描述(申请人提供)：我们实验室最近的实验结果表明，烧伤前的急性酒精中毒会恶化肠道屏障功能，增加细菌易位。我们建议的研究旨在调查IL-18在乙醇中毒和烧伤复合损伤后肠道屏障功能改变中的作用。在我们的初步研究中，我们观察到乙醇中毒和烧伤的联合损伤上调了脾和肠道淋巴器官(肠系膜淋巴结和Peyer‘s Patch)中IL-18的产生。IL-18的升高伴随着肠道髓过氧化物酶(MPO，中性粒细胞聚集的一个指标)活性的增加和肠道水肿。损伤时给予caspase-1的抑制剂Ac-YVAD-CHO(5 mg/kg)治疗，可阻止乙醇中毒烧伤大鼠肠道中IL-18的生成增加以及肠道MPO活性的增加和肠道水肿。 这些初步发现提示IL-18在乙醇中毒和烧伤后肠组织中性粒细胞聚集和水肿中起作用。由于已知中性粒细胞会导致组织损伤，IL-18很可能会增强中性粒细胞释放氧化剂(O2-)和蛋白酶(例如弹性蛋白酶)的能力，这反过来又会导致肠道通透性增加和水肿。因此，这项拟议研究的总体目标是确定IL-18是否影响中性粒细胞的组织破坏行为。我们假设，乙醇中毒和烧伤复合损伤后IL-18上调，通过增强O2-和弹性蛋白酶的释放，以及它们在肠道组织中的募集，增加了中性粒细胞的组织破坏作用。提出的假设将在一个已建立的急性乙醇中毒和烧伤大鼠模型中进行验证。针对特定目标1的研究将确定，通过中和抗IL-18抗体或用caspase-1抑制剂Ac-YVAD-CHO治疗动物来阻断IL-18水平是否可以防止中性粒细胞02-产生、弹性酶释放及其在无水乙醇和烧伤大鼠肠道中的隔离。在特定的目标2中，我们将确定a)IL-18介导的中性粒细胞在乙醇中毒和烧伤后肠道中的隔离是IL-18的直接作用还是通过IL-18依赖的中性粒细胞趋化因子CINC的上调来介导的，以及b)中性粒细胞是否是IL-18介导的乙醇中毒和烧伤后肠道通透性增加和水肿的关键决定因素。我们希望本申请中描述IL-18在肠屏障功能受损中的作用的拟议研究将有助于设计特定的治疗方法来治疗在乙醇中毒影响下发生损伤的烧伤患者。

DESCRIPTION (provided by applicant): Recent experimental findings from our laboratory suggest that acute alcohol (EtOH) intoxication prior to bum injury deteriorates intestinal barrier function and increases bacterial translocation. Our proposed studies are designed to investigate the role of IL-18 in altered intestinal barrier functions following a combined insult of EtOH intoxication and burn injury. In our preliminary studies, we observed that a combined insult of EtOH intoxication and burn injury up-regulates IL-18 production in spleen and intestinal lymphoid organs (Mesenteric lymph nodes and Peyer's patches). The increase in IL-18 is accompanied by an increase in intestinal myeloperoxidase (MPO, an index of neutrophil accumulation) activity and intestinal edema. The treatment of rats at the time of injury with Ac-YVAD-CHO (5 mg/kg), an inhibitor of caspase-1 (enzyme that converts pro-IL-18, an inactive form of IL-18, to mature IL-18), prevented the increase in IL-18 production, as well as the increase in intestinal MPO activity and intestinal edema in EtOH intoxicated, burn-injured rats. These preliminary findings suggest a role for IL-18 in neutrophil accumulation and edema in intestinal tissue following EtOH intoxication and burn injury. Since neutrophils are known to cause tissue injury, it is likely that IL-18 potentiates neutrophils' ability to release oxidants (O2-) and proteases (e.g., elastase), which in turn can contribute to increased intestinal permeability and edema. Thus the overall goal of the proposed studies is to determine whether or not IL-18 influences neutrophils' tissue-damaging actions. We hypothesize that IL-18 up-regulation following a combined insult of EtOH intoxication and burn injury increases neutrophils' tissue-damaging actions via potentiating the release of O2- and elastase, and their recruitment to the intestinal tissue. The proposed hypothesis will be tested in a well-established rat model of acute EtOH intoxication and burn injury. Studies in Specific aim 1 will determine whether blocking IL-18 levels by treating animals with neutralizing anti-IL-18 antibodies or with caspase-1 inhibitor Ac-YVAD-CHO prevents neutrophils' 02- production, elastase release, and their sequestration in the intestine of rats given EtOH and burn injury. In Specific aim 2, we will determine a) whether IL-18-mediated neutrophil sequestration in the intestine following EtOH intoxication and burn injury is a direct effect of IL-18 or is mediated via IL-18-dependent up-regulation of neutrophil chemotactic factor, CINC, and b) whether neutrophils are critical determinant of IL-18-mediated increase in intestinal permeability and edema following EtOH intoxication and burn injury. We hope that the proposed studies as outlined in this application delineating the role of IL-18 in impaired intestinal barrier function would help in designing specific therapeutic approaches for treatment of burn patients who sustained injury under the influence of EtOH intoxication.