title: Glutamine regulates Caco-2 cell tight junction proteins.
abstract: Intestinal epithelial tight junction (TJ) barrier dysfunction may lead to inflammation and mucosal injury. Glutamine (GLN) plays a role in maintenance of intestinal barrier function in various animal models and critically ill humans. Recent evidence from intestinal cell monolayers indicates that GLN maintains transepithelial resistance and decreases permeability. The mechanisms of these effects remain undefined. We hypothesized that GLN affects proteins involved in the intercellular junctional complex. GLN availability was controlled in Caco-2 monolayers by addition to the medium and treatment with methionine sulfoximine (MSO) to inhibit glutamine synthetase (GS). Expression of TJ proteins, claudin-1, occludin, and zonula occluden (ZO)-1 was measured by immunoblotting. Localization of TJ proteins was evaluated by immunofluorescence light microscopy. Structure of TJ was determined by transmission electron microscopy (TEM). Deprivation of GLN decreased claudin-1, occludin, and ZO-1 protein expression and caused a disappearance of perijunctional claudin-1 and a reduction of occludin but had no effect on ZO-1. TEM revealed that MSO-treated cells in the absence of GLN formed irregular junctional complexes between the apical lateral margins of adjoining cells. These findings indicate that TJ protein expression and cellular localization in Caco-2 cell monolayers rely on GLN. This mechanism may similarly relate to GLN-mediated modulation of intestinal barrier function in stressed animals and humans.
authors.affiliation: Division of Neonatology, Department of Pediatrics, University of Florida College of Medicine, Gainesville, Florida 32610, USA.
authors.forename: Nan
authors.initials: N
authors.lastname: Li
authors.forename: Patricia
authors.initials: P
authors.lastname: Lewis
authors.forename: Don
authors.initials: D
authors.lastname: Samuelson
authors.forename: Kellym
authors.initials: K
authors.lastname: Liboni
authors.forename: Josef
authors.initials: J
authors.lastname: Neu
mesh_terms: D015153:Blotting, Western; D018938:Caco-2 Cells; D062445:Claudin-1; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D005973:Glutamine / Q000494:pharmacology*; D005981:Glutathione Synthase / Q000037:antagonists & inhibitors; D006801:Humans; D015151:Immunoblotting; D008565:Membrane Proteins / Q000096:biosynthesis; D008854:Microscopy, Electron; D062793:Occludin; D010750:Phosphoproteins / Q000096:biosynthesis; D014176:Protein Biosynthesis; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D062826:Zonula Occludens-1 Protein
chemical_list: C568108:CLDN1 protein, human; D062445:Claudin-1; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D005973:Glutamine; D005981:Glutathione Synthase