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MUCOSAL BIOLOGY

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

¹Department of Surgery and ²Veterans Affairs Medical Center, Oregon Health Sciences University, Portland, Oregon, 97201; ³Department of Medicine, Howard University, Washington, District of Columbia 20060; and ⁴Departments of Medicine, Microbiology, Immunology, and Veterans Affairs Medical Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232

Submitted 1 July 2002 ; accepted in final form 15 February 2003

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.

vacA; cagA; picB/cagE; bacteria; signal transduction; fura 2; fluo 4; cell culture; immunofluorescence; thapsigargin; genistein; herbimycin; G protein; stomach