title: Increased iNOS activity is essential for intestinal epithelial tight junction dysfunction in endotoxemic mice.
abstract: We tested the hypothesis that increased production of nitric oxide (NO.) associated with lipopolysaccharide (LPS)-induced systemic inflammation leads to functionally significant alterations in the expression and/or targeting of key tight junction (TJ) proteins in ileal and colonic epithelium. Wild-type or inducible NO. synthase (iNOS) knockout male C57B1/6J mice were injected intraperitoneally with 2 mg/kg Escherichia coli O111:B4 LPS. iNOS was inhibited using intraperitoneal L-N(6)-(1-iminoethyl)lysine (L-NIL; 5 mg/kg). Immunoblotting of total protein and NP-40 insoluble proteins revealed decreased expression and decreased TJ localization, respectively, of the TJ proteins, zonula occludens (ZO)-1, ZO-2, ZO-3, and/or occludin in ileal mucosa and colonic mucosa (total protein only) after injection of C57B1/6J mice with LPS. Immunohistochemistry showed deranged distribution of ZO-1 and occludin in both tissues from endotoxemic mice. Endotoxemia was associated with evidence of gut epithelial barrier dysfunction evidenced by increased ileal mucosal permeability to fluorescein isothiocyanate-dextran (Mr=4 kDa) and increased bacterial translocation to mesenteric lymph nodes. Pharmacologic inhibition of iNOS activity using L-NIL or genetic ablation of the iNOS gene ameliorated LPS-induced changes in TJ protein expression and gut mucosal barrier function. These results support the view that at least one mechanism contributing to the pathogenesis of gastrointestinal epithelial dysfunction secondary to systemic inflammation is increased iNOS-dependent NO. production leading to altered expression and localization of key TJ proteins.
authors.affiliation: Department of Critical Care Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, 15261, USA.
authors.forename: Xiaonan
authors.initials: X
authors.lastname: Han
authors.forename: Mitchell P
authors.initials: MP
authors.lastname: Fink
authors.forename: Runkuan
authors.initials: R
authors.lastname: Yang
authors.forename: Russell L
authors.initials: RL
authors.lastname: Delude
mesh_terms: D000818:Animals; D003106:Colon / Q000378:metabolism / Q000473:pathology; D003911:Dextrans / Q000494:pharmacology; D004848:Epithelium / Q000201:enzymology*; D004926:Escherichia coli / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives* / Q000494:pharmacology; D019084:Fluorescent Antibody Technique, Indirect; D007082:Ileum / Q000378:metabolism; D015151:Immunoblotting; D007150:Immunohistochemistry; D007413:Intestinal Mucosa / Q000473:pathology; D008070:Lipopolysaccharides / Q000378:metabolism; D008196:Lymph / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000096:biosynthesis / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D009092:Mucous Membrane / Q000473:pathology; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D062793:Occludin; D017830:Octoxynol; D010750:Phosphoproteins / Q000096:biosynthesis; D011092:Polyethylene Glycols / Q000378:metabolism; D011233:Precipitin Tests; D019108:Tight Junctions / Q000201:enzymology*; D013997:Time Factors; D062826:Zonula Occludens-1 Protein
chemical_list: D003911:Dextrans; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D062793:Occludin; C568986:Ocln protein, mouse; D010750:Phosphoproteins; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein; C015219:fluorescein isothiocyanate dextran; D009569:Nitric Oxide; D011092:Polyethylene Glycols; D017830:Octoxynol; C010615:Nonidet P-40; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D016650:Fluorescein-5-isothiocyanate