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Reactive oxygen species (ROS) generated by NADPH oxidases (NOX) provide antimicrobial defense, redox signaling and gut barrier maintenance. NADPH oxidase mutations have been identified in IBD patients, and mucus layer disruption, a critical aspect in IBD pathogenesis, was connected to NOX inactivation. To gain insight into ROS-dependent modification of epithelial glycosylation the colonic and ileal mucin <jats:italic>O<\/jats:italic>-glycome of mice with genetic NOX inactivation (<jats:italic>Cyba<\/jats:italic> mutant) was analyzed. <jats:italic>O<\/jats:italic>-glycans were released from purified murine mucins and analyzed by hydrophilic interaction ultra-performance liquid chromatography in combination with exoglycosidase digestion and mass spectrometry. We identified five novel glycans in ileum and found minor changes in <jats:italic>O<\/jats:italic>-glycans in the colon and ileum of <jats:italic>Cyba<\/jats:italic> mutant mice. Changes included an increase in glycans with terminal HexNAc and in core 2 glycans with Fuc-Gal- on C3 branch, and a decrease in core 3 glycans in the colon, while the ileum showed increased sialylation and a decrease in sulfated glycans. Our data suggest that NADPH oxidase activity alters the intestinal mucin <jats:italic>O<\/jats:italic>-glycans that may contribute to intestinal dysbiosis and chronic inflammation.<\/jats:p>","DOI":"10.1371\/journal.pone.0297292","type":"journal-article","created":{"date-parts":[[2024,3,14]],"date-time":"2024-03-14T17:38:09Z","timestamp":1710437889000},"page":"e0297292","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":5,"title":["Characterization of intestinal O-glycome in reactive oxygen species deficiency"],"prefix":"10.1371","volume":"19","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5085-5080","authenticated-orcid":true,"given":"Radka","family":"Saldova","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kristina 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