Effects of<i>Helicobacter pylori</i>on intracellular Ca²⁺signaling in normal human gastric mucous epithelial cells

Abstract

In stomach, Helicobacter pylori (Hp) adheres to gastric mucous epithelial cells (GMEC) and initiates several different signal transduction events. Alteration of intracellular Ca2+ concentration ([Ca2+]i) is an important signaling mechanism in numerous bacteria-host model systems. Changes in [Ca2+]i induced by Hp in normal human GMEC have not yet been described; therefore, we examined effects of Hp on [Ca2+]i in normal human GMEC and a nontransformed GMEC line (HFE-145). Cultured cells were grown on glass slides, porous filters, or 96-well plates and loaded with fura 2 or fluo 4. Hp wild-type strain 60190 and vacA-, cagA-, and picB-/cagE- isogenic mutants were incubated with cells. Changes in [Ca2+]i were recorded with a fluorimeter or fluorescence plate reader. Wild-type Hp produced dose-dependent biphasic transient [Ca2+]i peak and plateau changes in both cell lines. Hp vacA- isogenic mutant produced changes in [Ca2+]i similar to those produced by wild type. Compared with wild type, cagA- and picB-/cagE- isogenic mutants produced lower peak changes and did not generate a plateau change. Preloading cultures with intracellular Ca2+ chelator BAPTA blocked all Hp-induced [Ca2+]i changes. Thapsigargin pretreatment of cultures to release Ca2+ from internal stores reduced peak change. Extracellular Ca2+ removal reduced plateau response. Hp-induced peak response was sensitive to G proteins and PLC inhibitors. Hp-induced plateau change was sensitive to G protein inhibitors, src kinases, and PLA2. These findings are the first to show that H. pylori alters [Ca2+]i in normal GMEC through a Ca2+ release/influx mechanism that depends on expression of cagA and picB/cagE genes.