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: 9242020
• 负责人: Ronen Sumagin
• 金额: $ 14.96万
• 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/9242020
• 关键词: Actins; Actomyosin; Acute; Adhesions; Adhesives; Apical; Apoptosis; Bacterial Infections; Binding; Biochemical; Biological Assay; Biopsy; Blood; CD59 Antigen; Cell Adhesion Molecules; Cell Communication; Cell Line; Cell Proliferation; Cell membrane; Cell surface; Cells; Chemotactic Factors; Complex; Crohn' s disease; Data; Development; Dextrans; Disease; E-Cadherin; Endothelial Cells; Endothelium; Epithelial; Epithelial Cell Junction; Epithelial Cell Proliferation; Epithelial Cells; Epithelium; Event; Fluorescence; Fluorescence Microscopy; Fluorescence Recovery After Photobleaching; Focal Adhesion Kinase 1; Functional disorder; Gastrointestinal Diseases; Gastrointestinal tract structure; Goals; Histologic; Homeostasis; Host Defense; Image; Immune; In Vitro; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Inflammatory disease of the intestine; Injury; Integrins; Interferons; Intestines; Knockout Mice; Lateral; Lead; Ligands; Ligation; Light; Link; Lipoxins; Longevity; Luciferases; Mediating; Membrane; Modeling; Molecular; Mucositis; Mus; Patients; Permeability; Play; Process; Proteins; Quantitative Reverse Transcriptase PCR; R-cadherin; Receptor Signaling; Recruitment Activity; Regulation; Reporter; Resistance; Resolution; Role; Salmonella; Signal Transduction; Surface; Testing; Therapeutic; Time; Tissues; Ulcerative Colitis; Up-Regulation; Wound Healing; cell motility; crosslink; experimental study; healing; improved; in vivo; in vivo Model; injury and repair; intravital imaging; intravital microscopy; knock-down; lipid mediator; migration; multi-photon; neutrophil; novel; occludin; overexpression; prevent; public health relevance; repaired; response; two-photon; wound

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 �-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（由适用提供）：非常感谢中性粒细胞（PMN）可以作为促进组织损伤和愈合的“双边剑”。 PMN transepthelial迁移（TEM）和结肠上皮加密中的积累是粘膜感染的标志，尤其是胃肠道的疾病。其中包括炎症性肠病，溃疡性结肠炎和克罗恩病，PMN TEM通常与上皮损伤和屏障功能障碍有关。但是，也有大量证据证明PMN在炎症的分辨率方面功能，包括释放脂质介质的释放，例如脂肪毒素，分辨率和促进愈合的脂质介质。 PMN TEM也已显示出诱导``catenin依赖性增生剂''，促进粘膜伤口修复。活跃IBD患者发炎的肠道组织的组织学分析显示，PMN的数量增加，在完成TEM后仍与顶端IEC表面保持亲密接触，从而继续引起肠上皮细胞（IEC）顶端配体。广泛的努力致力于确定调解PMN TEM的关键参与者，并在PMN TEM和上皮屏障功能之间建立联系，但是，我们对这一过程的理解仍然有限。关于PMN与顶端IEC配体的相互作用以及这些相互作用对上皮稳态的影响，尤其是上皮伤口修复的影响。一种这样的配体是细胞内粘合分子1（ICAM-1）。 ICAM-1在健康组织中以低水平表达，并且在炎症期间高度更新。与原始植物不同，ICAM-1在介导PMN跨内皮迁移和调节屏障功能方面的作用已经很好地确定，在上皮中，ICAM-1的作用仍然不清楚。在发炎的tisue中，其表达仅限于上皮表面，这表明腔表面上的ICAM-1可能在介导PMN上皮细胞相互作用，促进PMN保留和触发信号事件以改变上皮功能。我们的初步数据表明，完成后TEM后的PMN通过弯曲与ICAM-1保留在腔上皮膜上。 PMN与ICAM-1的结合显着抑制PMN的凋亡，从而导致PMN遵循的长期寿命与顶上上皮膜。此外，我们发现ICAM-1的参与会导致上皮完整性的提高，并诱导支持上皮伤口修复的增殖信号传导。该提案的总体目标是确定ICAM-1在上皮表面上的PMN接合如何触发信号事件，以提高上皮渗透性，促进PMN传播并刺激上皮修复。在AIM 1中，我们将定义ICAM-1连接下游的特定信号事件，从而增加上皮渗透性，以促进增强的PMN TEM，并使用PMN TEM的体内和体内模型，并补充了分子和蛋白质方法，以及先进的多光子插入式Intervital Intravital Impravital Imasting。在AIM 2中，我们将使用维特罗刮伤伤口测定法和急性损伤和感染的新型体内模型，包括结肠镜检查和DSS诱导的结肠粘膜损伤，定义ICAM-1在调节上皮稳态和伤口修复中的特定作用。当前提案中概述的实验将为预防PMN TEM和在粘膜表面保留的机制提供新的启示，并有助于鉴定将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必须支持增强的宿主防御功能和粘膜感染的解决方案，并重新建立IEC稳态。