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The function of intact long-type DupA protein in Helicobacter pylori was analyzed using immunoblotting and molecular biology techniques in the study. After cloning, expression and purification, ATPase activity of DupA protein was detected. Antibody was produced for localization and interaction proteins analysis. The dupA -deleted mutant was generated for adhesion and CagA protein translocation assay, susceptibility to different pH, IL-8 secretion assay, cytotoxicity to MKN-45 cells and proteins-involved apoptosis analysis. DupA protein exhibited an ATPase activity (129.5 ± 17.8 U/mgprot) and located in bacterial membrane, while it did not involve the adhesion and CagA protein delivery of H. pylori . DupA protein involved the urease secretion as the interaction proteins. The wild type strain had a stronger growth in low pH than the dupA -deleted mutant ( p < 0.001). IL-8 productions from GES-1 cells infected with the wild type strain were significantly higher than from those with the mutant ( p < 0.001). The amounts of vital MKN-45 cells were decreased and the numbers of apoptotic cells were increased with the wild type strain, compared to those with the mutant after 12 h ( p < 0.05). The increase of cleaved Caspase-3 and Bax was significantly higher and the decrease of Bcl-2 was more obvious in MKN-45 cells exposed to the wild type strain than that exposed to the mutant after 6 h. We demonstrate that intact long-type DupA protein located in membrane as ATPase is a true virulence factor associated with duodenal ulcer development involving the IL-8 induction and urease secretion, while it inhibits gastric cancer cell growth in vitro by activating the mitochondria-mediated apoptotic pathway. [ABSTRACT FROM AUTHOR]

Helicobacter pylori (H. pylori) is one of the most important pathogenic bacteria associated with various gastrointestinal diseases. At present, its apoptotic or antiapoptotic mechanism on gastric epithelial cells remains unknown and needs further illustrated. In this study, acute infection model (H. pylori and GES-1 cells were co-cultured for 24 h at a multiplicity of infection MOI of 100:1) and chronic infection model (GES-1 cells were infected repeatedly every 24 h at a multiplicity of infection MOI of 100:1 for approximately 8 weeks) were established, respectively. the chronic H. pylori infected GES-1 cells underwent a typically morphological change and Western Blot results showed that there was slight decrease in expression of E-cadherin, and obvious increase in expression of Vimentin. Apoptosis of these two models were analyzed by flow cytometry compared with the control cells, meanwhile, apoptosis associated markers (Bcl-xL, Bcl-2, Bax, etc) were detected by Western blot, additional in clinical H. pylori -positive gastric cancer tissues. Results showed that compared with the control cells, acute infection of H. pylori significantly accelerated the apoptosis of GES-1, increased the expression of Bax and Cleaved caspase-3, down-regulated expression of Bcl-xL and Bcl-2. Moreover, an opposite result was found in chronic infection of model and clinical gastric cancer tissues, and enhanced expression of NF–κB p65. Taken together, these findings suggest that H. pylori infection plays differential effects on apoptosis of gastric epithelial cells. • Acute infection with H. pylori promotes gastric epithelial cell apoptosis. • Chronic infection with H. pylori induced the epithelial-to-mesenchymal transition and enhanced the apoptosis tolerance. • The changes of apoptosis of GES-1 cells induced by H. pylori infection were mainly caused by different changes of the mitochondrial apoptosis pathway and NF-κB signaling pathway. [ABSTRACT FROM AUTHOR]