Neutrophil interactions with apical ICAM-1 regulate intestinal epithelial homeost

中性粒细胞与顶端 ICAM-1 的相互作用调节肠上皮稳态

• 批准号: 8985321
• 负责人: Ronen Sumagin
• 金额: $ 9.2万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-19 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8985321
• 关键词: Neutrophil; interactions; apical; ICAM; regulate

DESCRIPTION (provided by applicant): It is well appreciated that neutrophils (PMN) can act as a "double edged sword" in promoting both tissue injury and healing. PMN transepithelial migration (TEM), and accumulation within colonic epithelial crypts, is a hallmark of mucosal inflammation, and particularly, diseases of the gastrointestinal tract. These include inflammatory bowel diseases, ulcerative colitis and Crohn's disease, where PMN TEM is often associated with epithelial injury and barrier dysfunction. However, there is also abundant evidence for PMN function in resolution of inflammation, including release of lipid mediators, such as lipoxins, resolvins and protectins that facilitate healing. PMN TEM has also been shown to induce β-catenin dependent proliferative responses promoting mucosal wound repair. Histological analysis of inflamed intestinal tissue from patients with active IBD show increased numbers of PMN that remain in intimate contact with apical IEC surface after completing TEM, thus continuously engaging intestinal epithelial cell (IEC) apical ligands. Extensive efforts were dedicated to identifying key players that mediate PMN TEM, and establishing the link between PMN TEM and epithelial barrier function, however, our understanding of this process is still limited. Even less is known about PMN interactions with apical IEC ligands and the consequent effects of these interactions on epithelial homeostasis, and specifically epithelial wound repair. One such ligand is intracellular adhesion molecule 1 (ICAM-1). ICAM-1 is expressed at low levels in healthy tissue, and is highly upregulated during inflammation. Unlike in endothelium, where the role of ICAM-1 in mediating PMN transendothelial migration and in regulating barrier function is well established, in epithelium the role for ICAM-1 is still not clear. In inflamed tisue its expression is restricted to apical epithelial surface, suggesting that ICAM-1 on the luminal surface may play a role in mediating PMN-epithelial cell interactions, promoting PMN retention and triggering signaling events to alter epithelial function. Our preliminary data suggest that PMN after completion of TEM are retained on the luminal epithelial membrane through binging to ICAM-1. PMN binding to ICAM-1 significantly inhibits PMN apoptosis, resulting in prolonged life span of PMN adherent to the apical epithelial membrane. Moreover, we found that engagement of ICAM-1 leads to decreased epithelial integrity and an induction of proliferative signaling facilitating epithelial wound repair. Thus the overall goal of this proposal is to determine how PMN engagement of ICAM-1 on the apical epithelial surface triggers signaling events to increase epithelial permeability, promote PMN transmigration and stimulate epithelial repair. In Aim 1 we will define the specific signaling events downstream of ICAM-1 ligation that increase epithelial permeability to facilitate enhanced PMN TEM, using in- vitro and in-vivo models of PMN TEM, supplemented with molecular and protein approaches, and advanced multi-photon intravital imaging. In Aim 2 we will define the role for specific engagement of ICAM-1 in regulation of epithelial homeostasis and wound repair using in-vitro scratch wound assays and novel in-vivo models of acute injury and inflammation, including colonoscopic biopsy-wound and DSS-induced colonic mucosal injury. Experiments outlined in the current proposal will shed new light on mechanisms regulating PMN TEM and retention at the mucosal surfaces, and aid in identification of specific molecules that link PMN-epithelial cell interactions with epithelial barrier function and wound repair. This is imperative for the development of new and improved therapeutic approaches facilitating enhanced host defense function and resolution of mucosal inflammation, and reestablishing IEC homeostasis.

描述(由申请人提供)：中性粒细胞(PMN)在促进组织损伤和愈合方面可以起到“双刃剑”的作用，这是很好的理解。中性粒细胞在结肠上皮隐窝内的跨上皮迁移和积聚是粘膜炎症，尤其是胃肠道疾病的标志。这些疾病包括炎症性肠病、溃疡性结肠炎和克罗恩病，其中PMN的透射电子显微镜通常与上皮损伤和屏障功能障碍有关。然而，也有大量的证据表明PMN在消解炎症方面的作用，包括释放脂类介质，如促进愈合的脂氧素、溶血素和保护素。PMN的透射电子显微镜也被证明能够诱导β-连环蛋白依赖的增殖反应，促进粘膜伤口修复。对活动期IBD患者炎性肠组织的组织学分析显示，在完成透射电子显微镜检查后，PMN数量增加，这些PMN仍与IEC根尖表面密切接触，从而持续与肠上皮细胞(IEC)根尖配体结合。人们致力于寻找PMN-TEMs的关键调节因子，并建立PMN-TEMs与上皮屏障功能之间的联系，然而，我们对这一过程的了解仍然有限。中性粒细胞与顶端IEC配体的相互作用以及这些相互作用对上皮细胞内稳态的影响，特别是对上皮损伤修复的影响更是鲜为人知。其中一个这样的配体是细胞内黏附分子1(ICAM-1)。ICAM-1在健康组织中低水平表达，在炎症期间高度上调。与内皮细胞不同的是，ICAM-1在介导PMN跨内皮细胞迁移和调节屏障功能方面的作用已经得到证实，而在内皮细胞中，ICAM-1的作用仍然不清楚。在炎症组织中，ICAM-1仅表达于上皮尖表面，提示管腔表面ICAM-1可能在介导PMN-上皮细胞相互作用、促进PMN滞留和触发信号事件改变上皮功能中发挥作用。我们的初步数据表明，在透射电子显微镜完成后，PMN通过与ICAM-1结合而保留在管腔上皮膜上。中性粒细胞与ICAM-1结合可显著抑制中性粒细胞的凋亡，延长中性粒细胞黏附在根尖上皮膜上的寿命。此外，我们发现ICAM-1的参与导致上皮完整性降低，并诱导促进上皮创伤修复的增殖信号。因此，本方案的总体目标是确定PMN如何在顶端上皮表面ICAM-1的参与下触发信号事件，从而增加上皮的通透性，促进PMN的迁移，并刺激上皮修复。在目标1中，我们将使用PMN的体外和体内模型，辅以分子和蛋白质方法，以及先进的多光子活体成像技术，定义ICAM-1连接下游的特定信号事件，这些事件增加上皮的通透性，以促进增强的PMN TEM。在目标2中，我们将使用体外划痕创伤试验和新的体内急性损伤和炎症模型，包括结肠镜活检创伤和DSS诱导的结肠粘膜损伤，确定ICAM-1在调节上皮细胞动态平衡和伤口修复中的特定作用。目前提案中概述的实验将为调节PMN透射电子显微镜和粘膜表面滞留的机制提供新的线索，并有助于识别将PMN-上皮细胞相互作用与上皮屏障功能和伤口修复联系起来的特定分子。这对于开发新的和改进的治疗方法，促进增强宿主防御功能和消除粘膜炎症，以及重建IEC内环境平衡是必要的。