Reduced glutathione modulates Ca(2+)-mediated damage to rabbit isolated gastric mucosal cells

Abstract

In the present study, we have examined the role of reduced glutathione (GSH) in the modulation of Ca2+ ionophore A-23187-induced gastric mucosal cellular disruption. Experiments were conducted using cells isolated from rabbit oxyntic mucosa. Cell injury was assessed by release of the lysosomal enzyme acid phosphatase, by trypan blue dye uptake, and by examination of lipid peroxidation. Cellular GSH was assessed spectrophotometrically by measuring total soluble reduced thiol content. Ionophore A-23187 (3-25 microM) induced a concentration-dependent injury to mucosal cells and a concentration-dependent reduction in cellular GSH content. Removal of Ca2+ from the incubation medium abolished both the disruptive action and the GSH-lowering effect of A-23187. Gastric cellular GSH content was reduced significantly by preincubation of cells with an agent that depletes reduced sulfhydryls, diethyl maleate (DEM; 1 mM). Furthermore, DEM significantly augmented cellular disruption in response to A-23187. Similarly, incubation of cells with L-cysteine (1 mM), a stimulant of glutathione synthesis, increased mucosal cellular GSH content and decreased the cellular disruptive effect of A-23187. Exogenous addition of GSH to the cell suspension significantly reduced Ca2+ ionophore-induced cellular disruption and hastened the recovery of cellular free Ca2+ concentration ([Ca2+]) to baseline values. Similarly, exogenous GSH reduced cellular disruption produced by cyclopiazonic acid and thapsigargin, agents that increase intracellular [Ca2+] ([Ca2+]i) by inhibiting Ca2+ sequestration. These data suggest that a sustained increase in [Ca2+]i contributes to the pathogenesis of gastric mucosal cellular injury. This effect appears to be mediated by a reduction in the cellular content of GSH.

Effect of 100 mT homogeneous static magnetic field on [Ca2+]c response to ATP in HL‐60 cells following GSH depletion

“…These findings were in agreement with a number of other studies. Wong and Tepperman [1994] found that DEM significantly increased the susceptibility to cellular damage induced by the calcium ionophore Br-A23187 in rabbit gastric mucosal cells. This increase in susceptibility to cellular damage was attributed to the depletion of intracellular calcium stores.…”

Opposite effects of flurbiprofen and the nitroxybutyl ester of flurbiprofen on apoptosis in cultured guinea‐pig gastric mucous cells

“…Enzyme activity was determined by the breakdown of p‐nitrophenyl phosphate at pH 4.8 as described by Wong & Tepperman (1994) .…”

Real‐time measurement of cytosolic free calcium concentration in DEM‐treated HL‐60 cells during static magnetic field exposure and activation by ATP

“…This line of research has shown that SMF exposure can result in modification of cell proliferation [Aldinucci et al, 2003;Chiu et al, 2007], cell differentiation [Chiu et al, 2007] and cell death by apoptosis or necrosis [Fanelli et al, 1999;Jajte et al, 2002;Tenuzzo et al, 2006]. These results represent a collection of cellular responses that are known to be influenced by the intracellular concentration of ions and free radicals [Wong and Tepperman, 1994]. Several groups have chosen to look at cytosolic free calcium concentration as a measure for the effects of SMFs [Parkinson and Hanks, 1989;Carson et al, 1990;Walleczek and Budinger, 1992].…”