Abstract
Inflammatory bowel disease (IBD) affects more than 1 million Americans with more than 30,000 new cases diagnosed each year. IBD increases patient morbidity and susceptibility to colorectal cancer, yet its etiology remains unknown. Current models identify two key determinants of IBD pathogenesis: hyperpermeability of the gut epithelial barrier to bacterial products and an abnormal immune response to these products. Two factors seem critical for hyperpermeability: oxidant-induced stress and proinflammatory cytokines (e.g., tumor necrosis factor-α). The aim of this study was to investigate the role of oxidant stress-mediated transactivation of the epidermal growth factor receptor (EGFR) in intestinal hyperpermeability. This study used the Caco-2 human colonic epithelial cell in vitro model of intestinal epithelium. Cells were grown on inserts for permeability and signaling studies and glass coverslips for microscopy studies. show that oxidant-induced intestinal hyperpermeability can be blocked by specific inhibitors of the EGFR, tumor necrosis factor convertase (TACE) metalloprotease, transforming growth factor (TGF)-α, and mitogen-activated protein kinases, especially extracellular signal-regulated kinase 1/2. We also show that oxidant initiates these signaling events, in part by causing translocation of TACE to cell-cell contact zones. In this study, our data identify a novel mechanism for oxidant-induced intestinal hyperpermeability relevant to IBD. We propose a new intestinal permeability model in which oxidant transactivates EGFR signaling by activation of TACE and cleavage of precursor TGF-α. These data could have a significant effect on our view of IBD pathogenesis and provide new therapeutic targets for IBD treatment.
Footnotes
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This work was supported in part by a grant from the Rush University Medical Center, Department of Internal Medicine, and by National Institutes of Health Grants NIDDK 60511 and NCCAM 01581 (to A.B.) and National Institute on Alcohol Abuse and Alcoholism Grant 13745 (to A.K.).
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.106.113019.
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ABBREVIATIONS: IBD, inflammatory bowel disease; TNF, tumor necrosis factor; TJ, tight junction; IEC, intestinal epithelial cell; EGFR, epidermal growth factor receptor; PKC, protein kinase C; EGF, epidermal growth factor; TGF, transforming growth factor; TACE, tumor necrosis factor-α-converting enzyme; MAPK, mitogen-activated protein kinase; ERK, extracellular signal-regulated kinase; TAPI-2, N-(R)-[2-(hydroxyaminocarbonyl)methyl]-4-methylpentanoyl-l-t-butyl-alanyl-l-alanine, 2-aminoethyl amide, TNF-α protease inhibitor-2; GM6001, Ilomastat or N-[(2R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl]-l-tryptophan methylamide; AG1478, 4-(3-chloroanilino)-6,7-dimethoxyquinazoline; GW2974, N4-(1-benzyl-1H-indazol-5-yl)-N6,N6-dimethyl-pyrido[3,4-d]pyrimidine-4,6-diamine; PD98059, 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; U0126, 1,4-diamino-2,3-dicyano-1,4-bis(o-aminophenylmercapto)butadiene ethanolate; p38, 38-kDa protein (MAPK); SB203580, 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole; JNK, jun kinase; SP600125, 1,9-pyrazoloanthrone anthrapyrazolone; LY29004, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; TAPI, TNF-α protease inhibitor; Ab, antibody; HB-EGF, heparin binding epidermal growth factor; AKT, protein kinase B; DMEM, Dulbecco's modified Eagle's medium; FSA, fluorescein-5-(and-6)-sulfonic acid trisodium salt; FITC, fluorescein isothiocyanate; RT, reverse transcription; PCR, polymerase chain reaction; SiRNA, short inhibitory RNA; MMP, matrix metalloproteinase; DMSO, dimethyl sulfoxide; MLCK, myosin light chain kinase; ADAM, A disintegrin and metalloproteinase; MMPi, matrix metalloprotease inhibitor.
- Received August 28, 2006.
- Accepted January 11, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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