Effect of NaCl and Helicobacter pylori vacuolating cytotoxin on cytokine expression and viability

AIM: To determine whether Helicobacter pylori (H pylori) vacuolating cytotoxin (VacA) regulates release of proinflammatory cytokines (IL-1 beta, IL-8, TNF-alpha, and IL-6) or alters gastric epithelial cell viability and to determine whether NaCl affects these VacA-incluced changes. METHODS: Vacuolating activity was determined by measuring the uptake of neutral red Into vacuoles of VacA-treated human gastric epithelial (AGS) cells. AGS cell viability was assessed by direct cell counting. Specific enzyme-linked immunosorbent assays (ELISA) and reverse transcriptase-polymerase chain reaction(RT-PCR) were performed to examine the effects of H pylori (VacA and NaCl on cell pro-inflammatory cytokine production in AGS cells. Immunohistochemical staining of gastric tissue from Mongolian gerbils was used to confirm VacA-induced pro-inflammatory cytokine production and the effects of NaCl on this VacA-incluced response. RESULTS: Addition of VacA alone reduced AGS cell viability (P < 0.05), and this reduction was enhanced by high doses of NaCl (P < 0.05). VacA alone induced expression of TNF-alpha, IL-8 and IL-1 beta, while NaCl alone induced expression of TNF-alpha and IL-1 beta, Changes in mRNA levels in the presence of both VacA and NaCl were more complicated. For the case of TNF-alpha, expression was close-dependent on NaCl. IL-6 mRNA was not detected. However, low levels of IL-6 were detected by ELISA. Positive immunohistochemical staining of IL-1, IL-6, and TNF-alpha was found in gastric tissue of H pylori-infected gerbils fed with either a normal diet or a high salt diet. However, the staining of these three cytokines was stronger in H pylori-infected animals fed with a 5g/kg NaCl diet. CONCLUSION: VacA decreases the viability of AGS cells, and this effect can be enhanced by NaCl. NaCl also affects the production of pro-inflammatory cytokines induced by VacA, suggesting that NaCl plays an important role in Hpylori-induced gastric epithelial cell cytotoxicity. (c) 2006 The WJG Press. All rights reserved.