FIG. 1.

Induction of ROS in gastric epithelial cells after exposure to H. pylori or oxygen metabolites. (A) DCFH₂-DA-treated Kato III cells were exposed to H₂O₂ at concentrations from 0 to 1,000 μM or infected with H. pylori (Hp) at a ratio of bacteria to epithelial cells of 300:1. Intracellular DCF fluorescence was measured by flow cytometry at intervals up to 20 min after stimulation. ROS accumulated in proportion to the concentration of H₂O₂ added to the cells, and bacteria stimulated the production of ROS in epithelial cells in a time-dependent manner. A representative experiment is shown. (B) Superoxide anion was measured by the cytochrome c reduction assay method at 0, 0.5, and 1 h after various concentrations of H. pylori strain 26695 (equivalent to ratios of bacteria to epithelial cells of 0:1, 300:1, 600:1, and 1,000:1) were added to wells containing media alone (dotted line) or media with Kato III gastric epithelial cells (continuous line). A dose-dependent increase in superoxide anion generation was measured with increasing concentrations of bacteria both with and without epithelial cells. Data represent superoxide anion production expressed as means ± SEM (n = 8 to 12). *, P < 0.05 compared to bacteria alone; #, P < 0.05 compared to Kato III cells alone. (C) Kato III and NCI-N87 gastric epithelial cells were exposed to H. pylori (ratio of bacteria to epithelial cells of 300:1), harvested at 6 or 24 h poststimulation, and assayed for levels of intracellular GSH using a colorimetric assay. Oxidative stress generated by H. pylori results in a sustained decrease in GSH levels. The data are means ± SEM, expressed as percentages of control levels. *, P < 0.05 compared to control (n = 5 to 7).