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.
In the case of bacterial infection and mucosal injury neutrophil transepithelial migration is essential for host defense and tissue homeostasis; however; it may also lead to detrimental inflammation; which is a hallmark of inflammatory disorders of mucosal surfaces; particularly diseases of the gastrointestinal tract; including ulcerative colitis and Cohn's disease. This project focuses on interactions of innate immune cells; specifically neutrophils with luminally expressed epithelial protein; ICAM-1; and the contribution of these interactions to regulation of epithelial barrier and mucosal wound healing; under the conditions of intestinal inflammation. A better understanding of the mechanisms regulating PMN recruitment and retention at the mucosal surfaces; and identification of specific molecules that may link PMN-epithelial cell interactions with epithelial barrier function and wound repair are imperative for the development of new and improved therapeutic approaches aiding in the resolution of mucosal inflammation; and reestablishing epithelial homeostasis.
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