Chronic gastritis induced by Helicobacter pylori is a strong risk factor for the development of distal gastric adenocarcinoma. A specific host response to H. pylori that may contribute to gastric carcinogenesis is epithelial cell apoptosis. The aim of this study was to investigate the capacity of H. pylori vacuolating toxin (VacA) to induce gastric epithelial cell apoptosis. When cocultured with AGS gastric epithelial cells, H. pylori strain 60190, which expresses a type s1/m1 VacA toxin, induced significantly higher levels of apoptosis than did an isogenic vacA null mutant strain. VacA purified from strain 60190 induced apoptosis in a dose-dependent manner, which required acid activation of the purified toxin and the presence of ammonium chloride. In contrast, apoptosis was not induced after incubation with a chimeric s2/m1 toxin (in which the s1 sequence at the NH(2) terminus of VacA from strain 60190 was replaced with the s2 sequence from the nontoxigenic strain Tx30a) or a VacA mutant protein (VacA Delta 6-27) that lacks a unique strongly hydrophobic region near the VacA NH(2) terminus. Moreover, when an equimolar mixture of purified VacA Delta 6-27 and purified wild-type VacA were added simultaneously to AGS cells, the mutant toxin exhibited a dominant negative effect, completely inhibiting the apoptosis-inducing activity of wild-type VacA. These results indicate that VacA induces gastric epithelial cell apoptosis and suggest that differences in levels of gastric mucosal epithelial apoptosis among H. pylori-infected persons may result from strain-dependent variations in VacA structure.