Critical role of mac-1 sialyl lewis x moieties in regulating neutrophil degranulation and transmigration |
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Authors: | Zen Ke Cui Lun-Biao Zhang Chen-Yu Liu Yuan |
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Affiliation: | 1 Jiangsu CDC-Nanjing University Joint Institute of Virology, Nanjing University, Nanjing, Jiangsu 210093, China 2 School of Life Sciences, Nanjing University, Nanjing, Jiangsu 210093, China 3 Cellular and Molecular Biology Program, Department of Biology, Georgia State University, Atlanta, GA 30303, USA |
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Abstract: | Leukocyte cell surface sialyl Lewis x (sLex) and related epitopes play an important role in cell rolling and adhesion during diapedesis via interaction with E-selectin. Here, we present evidence that Mac-1 (CD11b/CD18, CR-3) is a major neutrophil glycoprotein decorated with sLex and ligation of these carbohydrate moieties by anti-sLex antibody significantly impairs neutrophil functions. First, Western blot analysis shows that both CD11b and CD18 subunit of purified Mac-1 are decorated with sLex moieties. A significant co-localization of CD11b and sLex moieties is observed at neutrophil secondary granules. With stimulation of formyl-Met-Leu-Phe (fMLP), neutrophil surface labeling with anti-sLex antibody follows an identical up-regulation pattern of Mac-1. Second, protein-binding assays indicate that sLex moieties on Mac-1 are critical for binding interaction of Mac-1 to E-selectin. Removal of sLex moieties completely abolishes Mac-1-E-selectin binding. Finally, ligation of Mac-1 sLex by anti-sLex antibody induces a significant degranulation of neutrophil secondary granules at the absence of chemoattractant stimulation. This “dysregulated” degranulation induced by anti-sLex antibody strongly inhibits neutrophil transmigration in response to fMLP. In summary, Mac-1 sLex moieties play a critical role in regulating β2 integrin functions during neutrophil transmigration and degranulation. |
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Keywords: | sLex, sialyl Lewis x fMLP, formyl-Met-Leu-Phe PMN, polymorphonuclear leukocyte FACS, fluorescence-activated cell scanning mAb, monoclonal antibody MPO, meyloperoxidase HBSS, Hank's balanced salt solution |
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