In vivo, gastric infection with Helicobacter pylori leads to substantial production of the inflammatory cytokines IL-1, IL-6, TNF-α, and IL-8. H. pylori strains that contain the cag pathogenicity island (cag+) and are associated with ulceration and gastric carcinoma induce greater cytokine production than cag− strains. Expression of these cytokines is often regulated by the transcription factor complex, nuclear factor-κB (NF-κB) through κB-binding elements in the enhancer/promoter regions of their genes. We report that more virulent cag+ H. pylori strains induce increased NF-κB-DNA binding activity, which elevates IL-8 expression in AGS gastric epithelial cells. The cag+ H. pylori strains induce significant stimulation of IL-8 promoter-driven reporter activity, while cag− strains do not. Furthermore, mutation of specific genes within the cag island (picA1 and picB) ablates enhanced NF-κB activation and IL-8 transcription. Increased IL-8 expression is inhibited by mutation in either the NF-κB or NF-IL-6 binding element. The cag+ strains, compared with the cag− strains, induce enhanced nuclear localization of a RelA-containing NF-κB binding complex, but no increase in NF-IL-6 binding activity. These studies demonstrate that the ability of different types of H. pylori strains to activate NF-κB correlates with their ability to induce IL-8 transcription and indicate a mechanism for the heightened inflammatory response seen in subjects infected with cag+ H. pylori strains.