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

描述（由申请人提供）：我们实验室最近的实验结果表明，烧伤前急性酒精（EtOH）中毒会恶化肠道屏障功能，增加细菌移位。我们提出的研究旨在研究IL-18在EtOH中毒和烧伤联合损伤后肠屏障功能改变中的作用。在我们的初步研究中，我们观察到EtOH中毒和烧伤的联合损伤上调了脾和肠淋巴器官（肠系膜淋巴结和派尔集合淋巴结）中IL-18的产生。IL-18的增加伴随着肠髓过氧化物酶（MPO，中性粒细胞积聚的指数）活性的增加和肠水肿。在损伤时用Ac-YVAD-CHO（5 mg/kg）（半胱天冬酶-1（将IL-18的非活性形式pro-IL-18转化为成熟IL-18的酶）的抑制剂）治疗大鼠，可防止EtOH中毒、烧伤大鼠中IL-18产生的增加以及肠MPO活性和肠水肿的增加。 这些初步发现表明IL-18在EtOH中毒和烧伤后肠组织中性粒细胞积聚和水肿中的作用。由于已知嗜中性粒细胞引起组织损伤，因此IL-18可能增强嗜中性粒细胞释放氧化剂（O2-）和蛋白酶（例如，弹性蛋白酶），这又会导致肠通透性增加和水肿。因此，所提出的研究的总体目标是确定IL-18是否影响中性粒细胞的组织损伤作用。我们假设，乙醇中毒和烧伤联合损伤后IL-18上调通过增强O2和弹性蛋白酶的释放及其向肠组织的募集来增加中性粒细胞的组织损伤作用。将在完善的急性EtOH中毒和烧伤大鼠模型中检验所提出的假设。具体目标1中的研究将确定通过用中和性抗IL-18抗体或用半胱天冬酶-1抑制剂Ac-YVAD-CHO治疗动物来阻断IL-18水平是否阻止中性粒细胞的O2产生、弹性蛋白酶释放及其在给予EtOH和烧伤的大鼠的肠中的隔离。在具体目标2中，我们将确定a）EtOH中毒和烧伤后肠中IL-18介导的中性粒细胞隔离是否是IL-18的直接作用或通过中性粒细胞趋化因子CINC的IL-18依赖性上调介导，以及B）中性粒细胞是否是EtOH中毒和烧伤后IL-18介导的肠通透性增加和水肿的关键决定因素。我们希望，本申请中概述的描述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用于治疗在EtOH中毒影响下持续损伤的烧伤患者的特定治疗方法。

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.