title: NO-dependent and NO-independent IL-1 production by a human colonic epithelial cell line under inflammatory stress.
abstract: 1. The present study was designed to investigate, in an in vitro model of the human intestinal barrier, the ability of epithelial cells to produce interleukin-1 (IL-1), the cellular mechanisms involved in IL-1 release, and the intracellular signalling pathways involved in IL-1 up-regulation during inflammatory stress. 2. This study was based on the human colonic epithelial cell line HT29-Cl.16E, maintained as polarized monolayers on filters mounted in culture chambers and treated with various proinflammatory cytokines (interferon gamma (IFN gamma), tumour necrosis factor alpha (TNF alpha), IL-1 beta) alone or in combination. 3. IL-1 production, restricted to IL-1 alpha, was induced by the combination of IFN gamma/TNF alpha. When IL-1 beta was added to IFN gamma/TNF alpha, it led to an additional production of IL-1 alpha. IL-1 alpha release was associated with cell damage, as shown by the correlation between lactate dehydrogenase (LDH) release and extracellular IL-1 production, and was not accounted for by a secretory mechanism. 4. Both IFN gamma/TNF alpha and IFN gamma/TNF alpha/IL-1 beta induced inducible nitric oxide synthase (iNOS) expression as shown by quantitation of NO2-/NO3- by use of the Griess reagent, quantitation of cells scoring positive with an anti-iNOS antibody and detection of mRNAs coding for iNOS by RT-PCR. The use of NG-monomethyl-L-arginine (L-NMMA), an inhibitor of NOS, led to the demonstration of two distinct signalling pathways in IL-1 production by HT29-Cl.16E cells, one dependent on NO (L-NMMA-sensitive) under treatment with IFN gamma/TNF alpha/IL-1 beta, and the other independent of NO (L-NMMA-insensitive) under treatment with IFN gamma/TNF alpha. 5. Moreover, we examined whether a redox-based mechanism could be responsible for the apparent discrepancy between NO production and NO implication in IL-1 production under IFN gamma/TNF alpha and IFN gamma/TNF alpha/IL-1 beta treatments. Experiments with cysteine, which acts as a powerful reductant, suggest that the nitrosonium character of NO is involved in the NO-dependent pathway in IL-1 production.
authors.affiliation: Groupe de Recherche Fonctions Sécrétoires des Epithéliums Digestifs, CJF INSERM 94-04, Faculté de Médecine, Nantes, France.
authors.forename: G
authors.initials: G
authors.lastname: Vallette
authors.forename: A
authors.initials: A
authors.lastname: Jarry
authors.forename: P
authors.initials: P
authors.lastname: Lemarre
authors.forename: J E
authors.initials: JE
authors.lastname: Branka
authors.forename: C L
authors.initials: CL
authors.lastname: Laboisse
mesh_terms: D002478:Cells, Cultured / Q000378:metabolism; D003106:Colon / Q000378:metabolism*; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D007375:Interleukin-1 / Q000096:biosynthesis*; D009569:Nitric Oxide / Q000096:biosynthesis* / Q000502:physiology; D013312:Stress, Physiological / Q000378:metabolism*
chemical_list: D007375:Interleukin-1; D009569:Nitric Oxide