Rao R, Baker R D, Baker S S
Department of Pediatrics, Medical University of South Carolina, Charleston 29425, USA.
Biochem Pharmacol. 1999 Mar 15;57(6):685-95. doi: 10.1016/s0006-2952(98)00333-5.
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.
评估表皮生长因子（EGF）对H2O2诱导的Caco-2和T-84细胞单层细胞旁通透性增加的影响，以研究EGF在肠道黏膜抗氧化应激保护中的作用。通过将细胞单层暴露于H2O2或黄嘌呤氧化酶+黄嘌呤（XO+X）混合物中来诱导氧化应激。通过测量跨上皮电阻（TER）、氯化钠稀释电位和[3H]甘露醇的单向通量来评估细胞旁通透性。H2O2（0.1至5.0 mM）降低了TER和稀释电位，并增加了甘露醇通量。给予EGF可延迟H2O2和XO+X诱导的TER、稀释电位和[3H]甘露醇通量的变化。顶端或基底给予EGF的这种保护作用与浓度相关，A50（95%置信限）值分别为2.1（1.17至4.34）和6.0（4.37至8.34）nM。用酪氨酸激酶抑制剂染料木黄酮（10 microM）处理细胞单层可阻止EGF介导的保护作用。H2O2和XO+X还诱导了Caco-2和T-84细胞单层中多种蛋白质的酪氨酸磷酸化。EGF处理可抑制氧化剂诱导的蛋白质酪氨酸磷酸化，特别是分子量为110-220 kDa的蛋白质。用抗转化生长因子-α抗体处理Caco-2细胞可增强H2O2诱导的TER、稀释电位和甘露醇通量的变化。这些研究表明，EGF受体介导的机制可能通过抑制氧化剂诱导的蛋白质酪氨酸磷酸化来延迟氧化剂诱导的上皮屏障功能破坏。
