Helicobacter pylori is a gram negative microaerophilic spiral organism, which is strongly associated with chronic active gastritis and peptic ulcer disease. This bacteria is also postulated to be an independent risk factor for gastric cancer. However, the mechanisms by which H. pylori causes epithelial cell injury, or leads to the development of peptic ulcers is not well understood. This bacterium produces a toxin which causes vacuolation in eukaryotic cells. This toxin appears to act through cell membrane Na+ K+ ATPase's. The bacteria also produces a urease enzyme in great abundance. Ammonia generated by urea breakdown has been shown to protect the organism against hydrochloric acid at the time of initial infection and is postulated to be a pathogenic factor causing gastric cell injury. The hallmark of H. pylori associated gastritis is the presence of both acute and chronic inflammatory cells. H. pylori lipopolysaccharide and surface proteins both stimulate the production of superoxide anion and cytokines by monocytes. The gastric epithelial damage in patients infected with H. pylori is felt to be largely due to the host immune response. Action oxygen, produced by inflammatory cells, is known to be toxic to epithelial cells. Reactive oxygen compounds may also act as carcinogens, and therefore, may be partly responsible for the increased association of gastric cancer in patients infected with H. pylori. This grant proposes to identify mechanisms through which bacterial toxins induce cell injury,k and to determine the extent to which bacterial toxins, such as vacuolizing cytotoxin and ammonia potentiate oxidant and cytokine induced cell injury. Studies will analyze the role of increased cytosolic free calcium in vacuolizing cytotoxin induced cell injury; and investigate activation of the protein kinase C signal transduction pathway by the vacuolizing cytotoxin as one mechanism through which its toxic effects are mediated. Digital imaging fluorescent microscopy will be used in conjunction with fluorescent probes to measure intracellular ionized calcium and mitochondrial respiration. Experiments will be conducted on primary human gastric epithelial cells and a SV40 T-antigen transformed human gastric epithelial cell line. Treatment of peptic ulcers with antibiotics heals ulcers as well as histamine antagonists, and curing H. pylori infection greatly reduces ulcer recurrence. Additional studies elucidating mechanisms by which bacterial toxicity occurs may help to develop safer, more easily tolerated therapies.
