Neutrophils (PMNs) play a critical role in clearing invading microbes and promoting tissue repair following infection/injury. However, dysregulated PMN trafficking and associated tissue damage is pathognomonic of numerous inflammatory mucosal diseases. The final step in PMN influx into mucosal lined organs (including the lungs, kidneys, skin and gut) involves transepithelial migration (TEpM). The b2-integrin CD11b/CD18 plays an important role in mediating PMN intestinal trafficking with recent studies highlighting that terminal Fucose and GlcNAc glycans on CD11b/CD18 can be targeted to reduce TEpM. However, the role of the most abundant terminal glycan, Sialic acid (Sia), in regulating PMN epithelial influx and mucosal inflammatory function is not well understood. Here we demonstrate that inhibiting sialidase mediated removal of a2-3 linked Sia from CD11b/CD18 inhibits PMN migration across intestinal epithelium in vitro and in vivo. Sialylation was also found to regulate critical PMN inflammatory effector functions including degranulation and superoxide release. Finally, we demonstrate that sialidase inhibition reduces bacterial peptide mediated CD11b/CD18 activation in PMN and blocks downstream intracellular signaling mediated by Spleen tyrosine kinase (Syk) and p38 MAP kinase. These findings suggest that sialylated glycans on CD11b/CD18 represent novel targets for ameliorating PMN mediated tissue destruction in inflammatory mucosal diseases.
Background Incidences of inflammatory bowel disease (IBD), including Crohn’s disease and ulcerative colitis, are escalating worldwide and can be considered a global public health problem. Given that the gold standard approach to IBD therapeutics focuses on reducing the severity of symptoms, there is an urgent unmet need to develop alternative therapies that halt not only inflammatory processes but also promote mucosal repair. Previous studies have identified increased stem cell factor (SCF) expression in inflamed intestinal mucosal tissues. However, the role that SCF plays in mediating intestinal inflammation and repair has not been explored. Methods Changes in the expression of SCF were evaluated in the colonic tissue of healthy mice and during dextran sodium sulfate (DSS)-induced colitis. Furthermore, mucosal wound healing and colitis severity were analyzed in mice subjected to either mechanical biopsy or DSS treatment, respectively, following intestinal epithelial cell–specific deletion of SCF or anti-SCF antibody administration. Results We report robust expression of SCF by intestinal epithelial cells during intestinal homeostasis with a switch to immune cell-produced SCF during colitis. Data from mice with intestinal epithelial cell–specific deletion of SCF highlight the importance of immune cell–produced SCF in driving the pathogenesis of colitis. Importantly, antibody-mediated neutralization of total SCF or the specific SCF248 isoform decreased immune cell infiltration and enhanced mucosal wound repair following biopsy-induced colonic injury or DSS-induced colitis. Conclusions These data demonstrate that SCF functions as a pro-inflammatory mediator in mucosal tissues and that specific neutralization of SCF248 could be a viable therapeutic option to reduce intestinal inflammation and promote mucosal wound repair in individuals with IBD.
Dysregulated transepithelial migration (TEM) of neutrophils (PMN) is pathognomonic of numerous inflammatory disorders including inflammatory bowel disease (IBD), yet mechanisms controlling PMN TEM remain poorly defined. The glycan determinant Lewis X (Galβ1‐4 (Fucα1‐3) GlcNAc‐β‐R), expressed by PMN surface glycoproteins (including MAC‐1 and LFA‐1), has previously been implicated in adhesive interactions between PMN and endothelium. However, little is known about the wider role of Lex in PMN function following extravasation. Studies revealed that engagement of terminal Lex glycans (but not the related glycan sLex) increased PMN adhesive interactions, both with other PMNs and with intestinal epithelial cells, blocking PMN chemotaxis and TEM. Further, flow cytometry analyses revealed increased surface expression of Lex on human PMN following TEM. In addition to blocking PMN trafficking, targeting of Lex also altered post‐migratory PMN functions, increasing both PMN phagocytosis and the surface mobilization of azurophilic granule (CD63 and MPO) and specific granule (CD66b and lactoferrin) markers. Finally, immunohistochemistry analyses revealed robust expression of Lex by PMN in the crypt abscesses of individuals with active IBD. Therefore, Lex represents a potential target for regulating PMN trafficking into the intestinal lumen and PMN luminal function. Further, given its abundant expression on migrating PMN and during inflammation, Lex may be a rational target for modulating inflammation in IBD.
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