Pathogenesis and Treatment of Experimental Peritonitis

描述(申请人提供)：肠道上皮是一种高度组织化的选择性屏障，可防止肠腔细菌从胃肠道进入。腹膜炎可导致肠道屏障受损，导致病原体跨肠粘膜移位，导致全身炎症加剧，并建立炎症引起的上皮损伤的恶性循环。了解腹膜炎、脓毒症等肠屏障功能障碍的发病机制是外科研究的重要目标之一。炎症性肠道屏障衰竭的机制尚不清楚。环氧合酶-2(COX-2)的表达及其产物前列腺素E_2(PGE_2)的积聚是肠道炎症的关键事件。PGE2通过其同源受体(EP1-EP4)发出信号，引发大量反应，包括局部血液供应增加，发烧和疼痛。PGE2通过EP1传递信号的结果之一是从细胞内的钙离子储存库中动员钙离子。已知钙离子激活肌球蛋白轻链激酶(MLCK)，导致MLC磷酸化和肌动球蛋白收缩。在多种上皮细胞中，MLC的磷酸化导致封闭细胞边界的紧密连接(TJ)的破坏。我们已经证实，PGE2能显著降低Caco-2单层的跨皮细胞电阻(TEER)，这种作用是由EP1、磷脂酶C(PLC)、钙离子和MLCK介导的。我们推测腹膜炎时肠屏障的破坏包括COX-2在肠上皮的表达，PGE2的积聚，以及PGE2通过EP1-PLC-Ca~(2+)-MLCK诱导的TJ的破坏。我们提出了三个特定的目标：1.明确COX-2、PGE2和EP1信号在腹膜炎时肠屏障衰竭中的作用。2.阐明前列腺素E_2诱导COX-2的作用机制。3.检测实验性腹膜炎期间保护肠屏障的COX-2靶向治疗。这项研究将深入了解COX-2和PGE2在肠道屏障衰竭发病机制中的作用，并有助于设计新的治疗方法，以在全身炎症期间维持肠道屏障的完整性。 与公共卫生相关：肠道屏障功能障碍影响着数十万患有各种急性和慢性炎症性疾病的患者。如果不了解炎症性肠道屏障失效的机制，就不能开发屏障支持疗法。我们建议阐明炎性前列腺素E2(PGE2)在实验性腹膜炎时屏障功能障碍中的作用。这项研究的结果将有助于在腹膜炎、休克和脓毒症的情况下设计新的屏障维护。

DESCRIPTION (provided by applicant): The intestinal epithelium is a highly organized selective barrier that prevents the entry of luminal bacteria from the GI tract. Peritonitis can lead to impairment of the gut barrier resulting in translocation of pathogens across the intestinal mucosa, leading exacerbation of the systemic inflammation and establishment a vicious cycle of inflammation-inflicted epithelial damage. Understanding the pathogenesis of gut barrier dysfunction in peritonitis, sepsis, etc, is one of the most important goals in surgical research. The mechanisms of inflammatory gut barrier failure are poorly understood. Expression of cyclooxygenase-2 (COX-2) and accumulation of its product prostaglandin E2 (PGE2) are critical events in the intestinal inflammation. PGE2 signals through its cognate receptors (EP1-EP4) to elicit a plethora of responses including increased local blood supply, fever and pain. One of the consequences of PGE2 signaling via EP1 is Ca2+ mobilization from the intracellular stores. It is known that Ca2+ activates myosin light chain kinase (MLCK), leading to MLC phosphorylation and actomyosin contractility. In a variety of epithelial cells, MLC phosphorylation causes disruption of tight junctions (TJ) that seal the cell borders. We have established that PGE2 causes dramatic decrease of transepithelial electric resistance (TEER) in Caco-2 monolayers, and that this effect is mediated by EP1, phospholipase C (PLC), Ca2+, and MLCK. We hypothesize that gut barrier breakdown during peritonitis involves expression of COX-2 in the intestinal epithelium, accumulation of PGE2, and PGE2-induced disruption of TJ via EP1-PLC-Ca2+-MLCK. We propose 3 specific aims: 1. To define the roles of COX-2, PGE2, and EP1 signaling in gut barrier failure during peritonitis. 2. To elucidate the mechanism of COX-2 induction by PGE2. 3. To test COX-2-targeting therapies aimed at protecting the gut barrier during experimental peritonitis. This study will yield considerable insight into the role of COX-2 and PGE2 in the pathogenesis of gut barrier failure and help design novel therapies to maintain gut barrier integrity during systemic inflammation. PUBLIC HEALTH RELEVANCE: Gut barrier failure affects hundreds of thousands of patients with a variety of acute and chronic inflammatory disorders. Barrier support treatments cannot be developed without understanding the mechanisms of inflammatory gut barrier failure. We propose to elucidate the role of inflammatory prostaglandin E2 (PGE2) in barrier dysfunction during experimental peritonitis. Results of this study will help to design new barrier maintenance under conditions of peritonitis, shock, and sepsis.