title: Metalloprotease-dependent amphiregulin release mediates tumor necrosis factor-alpha-induced IL-8 secretion in the human airway epithelial cell line NCI-H292.
abstract: Tumor necrosis factor-alpha (TNF-alpha) is a potent multifunctional cytokine that plays a central role in the pathogenesis of many inflammatory diseases. Interleukin-8 (IL-8) is a principle neutrophil chemoattractant and activator in humans. The alveolar macrophage-derived TNF-alpha initiates lung inflammation through its ability to stimulate IL-8 synthesis in airway epithelial cells. Since recent studies demonstrated that the stimulation of epidermal growth factor receptor (EGFR) could induce IL-8 secretion, the involvement of EGFR in TNF-alpha-induced IL-8 secretion in airway epithelium-like NCI-H292 cells was investigated in this study. TNF-alpha and epidermal growth factor (EGF) stimulated IL-8 secretion in a time- and concentration-dependent manner. Inhibition of the EGFR by either an anti-EGFR neutralizing antibody or by its specific inhibitor AG1478 (1 microM) blocked TNF-alpha-induced IL-8 secretion. In addition, TNF-alpha stimulated tyrosine phosphorylation of the EGFR within 5 min after stimulation. Further, TNF-alpha-induced IL-8 secretion was completely inhibited by the neutralizing antibody against amphiregulin (AR), an EGFR ligand, suggesting that TNF-alpha-induced IL-8 secretion was mediated by the AR-EGFR pathway. Furthermore, TNF-alpha stimulated the release of AR in a concentration-dependent manner. Finally, both AR and IL-8 release-induced by TNF-alpha were eliminated by pretreatment with either GM6001, a broad-spectrum inhibitor for metalloprotease, or TAPI-1, relatively selective inhibitor for TNF-alpha converting enzyme (TACE). These findings indicate that metalloprotease-mediated AR shedding and subsequent activation of EGFR play a critical role in TNF-alpha-induced IL-8 secretion from the human airway epithelium-like NCI-H292 cells, and that TACE is one of the most possible candidates for metalloprotease responsible for TNF-alpha-induced AR shedding.
authors.affiliation: Bio-medical Evaluation Research Department, Pharmaceutical Discovery Research Laboratories, Teijin Pharma Limited, 4-3-2, Asahigaoka, Hino, Tokyo 191-8512, Japan. m.chiyotsuki@teijin.co.jp
authors.forename: Manabu
authors.initials: M
authors.lastname: Chokki
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Mitsuhashi
authors.forename: Takashi
authors.initials: T
authors.lastname: Kamimura
mesh_terms: D066258:Amphiregulin; D019138:Antibodies, Blocking / Q000494:pharmacology; D002460:Cell Line; D004151:Dipeptides / Q000494:pharmacology; D066255:EGF Family of Proteins; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D015151:Immunoblotting; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D016209:Interleukin-8 / Q000096:biosynthesis*; D007700:Kinetics; D045726:Metalloproteases / Q000037:antagonists & inhibitors / Q000502:physiology*; D011480:Protease Inhibitors / Q000494:pharmacology; D011799:Quinazolines; D015533:Transcriptional Activation; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D020032:Tyrphostins / Q000494:pharmacology
chemical_list: C587848:AREG protein, human; D066258:Amphiregulin; D019138:Antibodies, Blocking; D004151:Dipeptides; D066255:EGF Family of Proteins; D006023:Glycoproteins; D006877:Hydroxamic Acids; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8; C092152:N-((2-(hydroxyaminocarbonyl)methyl)-4-methylpentanoyl)-3-(2'-naphthyl)alanylalanine, 2-aminoethylamide; C078131:N-(2(R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl)-L-tryptophan methylamide; D011480:Protease Inhibitors; D011799:Quinazolines; D014409:Tumor Necrosis Factor-alpha; D020032:Tyrphostins; C101044:RTKI cpd; D066246:ErbB Receptors; D045726:Metalloproteases