Analysis of apoptotic and antiapoptotic signalling pathways induced by Helicobacter pylori.

Background and aims: Although it is reported that Helicobacter pylori induces apoptosis on gastric epithelial cells, the mechanism remains unknown. Antiapoptotic effects generated by H pylori have not yet been evaluated.

Methods: (1) H pylori strains (type 1 wild, TN2-deltacagE, TN2-deltavacA) were cocultured with MKN45, TMK1, and HeLa cells, and cell viability and apoptosis were assessed by trypan blue exclusion and DNA laddering, respectively. (2) Activation of caspases-3, 7, and 8, cytochrome c release from the mitochondria, and Fas, Fas associated death domain protein (FADD), Bax, Bak, and Bcl-X expression were evaluated by immunoblot analysis. (3) To investigate whether nuclear factor kappa B (NFkappaB) activation induced by cag pathogenicity island (PAI) positive H pylori affects antiapoptosis, MKN45 cells stably expressing super-repressor IkappaBalpha were cocultured with H pylori, and cell viability and caspase activation were evaluated. NFkappaB regulated gene expression was also evaluated by ribonuclease protection assay.

Results: (1) Wild-type and deltavacA mutant H pylori induced apoptosis more potently than the deltacagE mutant. Inhibition of cell contact between H pylori and cancer cells and heat killing H pylori diminished cell death. (2) Caspases-3, 7, and 8 were activated time dependently by H pylori as well as by the agonist anti-Fas. Cytochrome c release from mitochondria was observed and was not inhibited by caspase-8 inhibitor. Although protein expression of Fas, FADD, Bax, Bak, and Bcl-X in the whole cell lysates was not changed by H pylori, Bax was decreased from mitochondria free cytosol suggesting that Bax was translocated into mitochondria. (3) Cell death and the activities of caspases-3 and 8 were promoted in MKN45 cells stably expressing super-repressor IkappaBalpha that inhibits NFkappaB activation. Antiapoptotic proteins c-IAP1 and c-IAP2 were upregulated by the wild-type strains.

Conclusion: cag PAI positive H pylori is capable of inducing apoptotic effects mainly through the mitochondrial pathway. Antiapoptotic effects mediated by NFkappaB activation were also observed.