Nguyen T D, Canada A T
Department of Medicine, Duke University, Durham, NC.
Biochem Pharmacol. 1994 Jan 20;47(2):403-10. doi: 10.1016/0006-2952(94)90032-9.
Hydrogen peroxide (H2O2) is a reactive oxygen species that can be produced in the digestive tract by inflammatory cells or during reperfusion following ischemia. To evaluate a possible direct effect of H2O2 on epithelial secretory cells, well-differentiated colonic T84 cells were grown to confluence on permeable membranes and studied in Ussing chambers. In this model, where the measured short-circuit current (Isc) reflects electrogenic secretion, we observed that H2O2 stimulated a concentration-dependent and transient secretory response: 5.5 mM H2O2 produced a peak Isc of 12.4 microA/cm2 after 4 min, 2.2 mM H2O2 a peak Isc of 7.9 microA/cm2 after 4 min, and 1.1 mM H2O2 a peak Isc of 5.5 microA/cm2 after 16 min (N = 5). When 97 experiments using 5.5 mM H2O2 were reviewed, the mean peak Isc response was 8.9 +/- 0.5 microA/cm2. A similar secretory response was elicited whether H2O2 was added to the serosal, to the mucosal, or simultaneously to both sides of the T84 cell monolayer. This secretory response reflected transcellular chloride secretion because it was inhibited by the depletion of chloride in the medium and by the suppression of the Na+,K+,2Cl- co-transporter activity necessary for the chloride gradient driving chloride secretion. When T84 cell monolayer resistance was studied, 5.5 mM H2O2 produced a transient decrease in resistance, reflecting transcellular chloride secretion, and a gradual decline in resistance (75% of the initial value after 55 min). The secretory response to H2O2 was increased 2-fold in T84 cells maximally stimulated with 10 nM vasoactive intestinal peptide (VIP), a neuropeptide which acts via cAMP, demonstrating synergism between the two agents. In contrast, the secretory responses produced by H2O2 and carbachol, which acts through the Ca2+ pathway, were additive. A late inhibitory effect of H2O2 was also observed: in cells previously treated with 5.5 mM H2O2, the subsequent secretory responses to either VIP or carbachol were partially inhibited. These secretory effects were specific for the oxidant properties of H2O2 because they were inhibited by 450 U/mL catalase and by 5 mM dithiothreitol, but were unaffected by 50 microM deferoxamine B or Fe3+. H2O2 may be a potential modulator of intestinal or colonic secretion in certain pathologic conditions such as inflammation or ischemia-reperfusion.
过氧化氢（H₂O₂）是一种活性氧物质，可由炎症细胞在消化道中产生，或在缺血后的再灌注过程中产生。为了评估H₂O₂对上皮分泌细胞的可能直接作用，将分化良好的结肠T84细胞在可渗透膜上培养至汇合，并在尤斯灌流小室中进行研究。在该模型中，所测量的短路电流（Isc）反映了电生性分泌，我们观察到H₂O₂刺激了浓度依赖性和短暂的分泌反应：5.5 mM H₂O₂在4分钟后产生的Isc峰值为12.4微安/平方厘米，2.2 mM H₂O₂在4分钟后产生的Isc峰值为7.9微安/平方厘米，1.1 mM H₂O₂在16分钟后产生的Isc峰值为5.5微安/平方厘米（N = 5）。当回顾使用5.5 mM H₂O₂进行的97次实验时，平均峰值Isc反应为8.9±0.5微安/平方厘米。无论将H₂O₂添加到T84细胞单层的浆膜侧、粘膜侧还是同时添加到两侧，都会引发类似的分泌反应。这种分泌反应反映了跨细胞氯分泌，因为它受到培养基中氯的耗尽以及抑制驱动氯分泌的氯梯度所需的Na⁺、K⁺、2Cl⁻共转运体活性的抑制。当研究T84细胞单层电阻时，5.5 mM H₂O₂导致电阻短暂下降，反映跨细胞氯分泌，随后电阻逐渐下降（55分钟后降至初始值的75%）。在用10 nM血管活性肠肽（VIP）最大程度刺激的T84细胞中，对H₂O₂的分泌反应增加了2倍，VIP是一种通过cAMP起作用的神经肽，表明这两种物质之间存在协同作用。相比之下，通过Ca²⁺途径起作用的H₂O₂和卡巴胆碱产生的分泌反应是相加的。还观察到H₂O₂的后期抑制作用：在先前用5.5 mM H₂O₂处理的细胞中，随后对VIP或卡巴胆碱的分泌反应受到部分抑制。这些分泌效应是H₂O₂氧化特性所特有的，因为它们受到450 U/mL过氧化氢酶和5 mM二硫苏糖醇的抑制，但不受50 microM去铁胺B或Fe³⁺的影响。在某些病理状况如炎症或缺血 - 再灌注中，H₂O₂可能是肠道或结肠分泌的潜在调节因子。
