Transepithelial Migration from Intestinal Epithelium during Polymorphonuclear Leukocyte Detachment Containing Exon 6 Mediates Synthesis of Sialyl Lewis A on CD44 Variant

Polymorphonuclear leukocyte (PMN) migration across the intestinal epithelium closely parallels disease symptoms in patients with inflammatory bowel disease. PMN transepithelial migration (TEM) is a multistep process that terminates with PMN detachment from the apical epithelium into the lumen. Using a unique mAb (GM35), we have previously demonstrated that engagement of the CD44 variant containing exon 6 (CD44v6) blocks both PMN detachment and cleavage of CD44v6. In this article, we report that PMN binding to CD44v6 is mediated by protein-specific O-glycosylation with sialyl Lewis A (sLe a). Analyses of glycosyltransferase expression identified fucosyltransferase 3 (Fut3) as the key enzyme driving sLe a biosynthesis in human intestinal epithelial cells (IECs). Fut3 transfection of sLe a-deficient IECs resulted in robust expression of sLe a. However, this glycan was not expressed on CD44v6 in these transfected IECs; therefore, engagement of sLe a had no effect on PMN TEM across these cells. Analyses of sLe a in human colonic mucosa revealed minimal expression in noninflamed areas, with striking upregulation under colitic conditions that correlated with increased expression of CD44v6. Importantly, intraluminal administration of mAb GM35 blocked PMN TEM and attenuated associated increases in intestinal permeability in a murine intestinal model of inflammation. These findings identify a unique role for protein-specific O-glycosylation in regulating PMN–epithelial interactions at the luminal surface of the intestine. T he migration of polymorphonuclear leukocytes (PMNs) out of the circulation across both endothelial and epithelial cell barriers is critical to the host inflammatory response to infection and injury. When dysregulated, the influx and accumulation of PMN in intestinal crypts is also a hallmark of the symptomatic phase of many intestinal inflammatory processes, including Crohn's disease and ulcerative colitis (UC) (1, 2). Considerable progress has been made in understanding the steps involved in PMN trafficking across vascular endothelium (3–5) and intestinal epithelium (6–9). In addition, the mechanisms governing late events in PMN transmigration across mucosa, including PMN detachment and release from the apical surface of epithelia into the lumen, although incompletely characterized, have become a recent focus of investigation. It has been previously reported that epithelial ICAM-1 is expressed apically and acts as a PMN retention ligand under inflammatory conditions (8). In addition, PMN FcR interactions with apical epithelial proteins have also been implicated in PMN–epithelial retention (10), and it has previously been reported that decay-accelerating factor functions as an antiadhesive epithelial glycoprotein that regulates PMN detachment from the epithelium (11). As described …