Charakterisierung der Rolle des Leukozyten-Adhäsionsmoleküls JAML während der Interaktion von Leukozyten und Epithelzellen in entzündlichen Darmerkrankungen und Bestimmung der funktionellen Konsequenz der Leukozyten-Adhäsion mittels JAML an die epithelialen Liganden CAR und CLMP
Final Report Abstract
Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the gastrointestinal tract in which leukocytes play a major role by altering epithelial barrier function and triggering aberrant inflammatory responses. The communication between leukocytes and epithelial cells is mediated by receptor ligand pairs that induce pathophysiological processes. The characterization of receptor-ligand pairs improves the understanding of intestinal inflammation and may provide new ideas for IBD treatment strategies. Junctional adhesion molecule-like (JAML) was originally discovered In the promyelocytic cell lines HL60 and NB4 where it was described to mediate cell adhesion and neutrophil transepithelial migration via binding to Its receptor CAR. In this project, I identified the ability of JAML to form homodimers for which the free cysteine residues in the membrane-distal D1-dohiain are important. By contrast, JAML binding to CAR is mediated by a conserved salt-bridge motif in the Dl-domain. Additionally, I focussed on the function of JAML in intestinal epithelial cells, since I unexpectedly detected strong mRNA signals for JAML only in intestinal epithelial but not in other epithelial cell lines. A gene silencing approach using JAML-specific sIRNA revealed that knock-down of JAML leads to stronger barriers in SK-C015 cells. Transepithelial resistance increased after loss of JAML whereas paracellular flux decreased. By contrast, leukocyte transmigration was increased after loss of epithelial JAML. This discrepancy may be explained by increased levels of CAR. Immunofluorescence analysis of ectopically overexpressed recombinant JAML in the intestinal epithelial cell line SK-C015 showed strong staining of JAML at the lateral membrane at cell contacts. These results point to an important role of JAML in the maintenance of epithelial barrier integrity. However, during this project, I identified a crossreactivity of our JAML antibody to the large GTPase guanylate binding protein (GBP)-I. Following on this result, I discovered that GBP-1 is an important mediator of epithelial barrier function during inflammation. GBP-1 is not only induced in intestinal epithelial cells but also translocated to tight junctions in response to IFN-y treatment. Moreover, GBP-1 is also upregulated in the colon of individuals with IBD. Loss of GBP-1 leads to impaired epithelial barrier function which is due to an increased rate of caspase-mediated apoptosis. These data suggest that GBP-1 is a novel marker of intestinal mucosal inflammation that protects against epithelial apoptosis induced by inflammatory cytokines and subsequent loss of barrier function.
Publications
- Disassembly of endothelial and epithelial junctions during leukocyte transmigration. Frontiers in Bioscience, 2008, 13: 6638-6652
Schnoor, M., Parkos, C. A.
- Guanylate binding protein-1 is expressed at tight junctions of intestinal epithelial cells in response to interferon-γ and regulates barrier function through effects on apoptosis. Mucosal Immunology, 2009, 2: 33-42
Schnoor, M., Betanzos, A., Weber, D. A., Parkos, C. A.
