Inhibition of oxidant-induced barrier disruption and protein tyrosine phosphorylation in Caco-2 cell monolayers by epidermal growth factor.
The effect of epidermal growth factor (EGF) on the H202-induced increase in paracellular permeability in Caco-2 and T-84 cell monolayers was evaluated to examine the role of EGF in intestinal mucosal protection from oxidative stress. Oxidative stress was induced by exposing cell monolayers to H2O2 or a mixture of xanthine oxidase + xanthine (XO + X). Paracellular permeability was assessed by measuring transepithelial electrical resistance (TER), sodium chloride dilution potential, and unidirectional flux of [3H]mannitol. H2O2 (0.1 to 5.0 mM) reduced TER and dilution potential and increased mannitol flux. Administration of EGF delayed H2O2 and XO + X-induced changes in TER, dilution potential, and [3H]mannitol flux. This protective effect of apically or basally administered EGF was concentration-related, with A50 (95% confidence limits) values of 2.1 (1.17 to 4.34) and 6.0 (4.37 to 8.34) nM, respectively. The EGF-mediated protection was prevented by treatment of cell monolayers with genistein (10 microM), a tyrosine kinase inhibitor. H2O2 and XO + X also induced tyrosine phosphorylation of a number of proteins in Caco-2 and T-84 cell monolayers. EGF treatment inhibited the oxidant-induced tyrosine phosphorylation of proteins, particularly those with a molecular mass of 110-220 kDa. Treatment of Caco-2 cells with anti-transforming growth factor-alpha antibodies potentiated the H2O2-induced changes in TER, dilution potential, and mannitol flux. These studies demonstrated that an EGF receptor-mediated mechanism delays oxidant-induced disruption of the epithelial barrier function, possibly by suppressing the oxidant-induced tyrosine phosphorylation of proteins.
Rao
R
R
Department of Pediatrics, Medical University of South Carolina, Charleston 29425, USA. raor@musc.edu
Baker
R D
RD
Baker
S S
SS
D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000494:pharmacology; D018938:Caco-2 Cells; D017097:Electric Impedance; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000037:antagonists & inhibitors; D008353:Mannitol / Q000378:metabolism; D016877:Oxidants / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D016211:Transforming Growth Factor alpha / Q000276:immunology; D019820:Xanthine; D014969:Xanthine Oxidase
D000911:Antibodies, Monoclonal; D016877:Oxidants; D016211:Transforming Growth Factor alpha; D019820:Xanthine; D008353:Mannitol; D004815:Epidermal Growth Factor; D006861:Hydrogen Peroxide; D014969:Xanthine Oxidase; D011505:Protein-Tyrosine Kinases