Monochloramine-induced toxicity and dysregulation of intracellular Zn2+in parietal cells of rabbit gastric glands

Abstract

Monochloramine (NH2Cl) is a potent, thiol-directed oxidant capable of oxidizing thiol (S-H) residues in a wide variety of proteins. Generated in the stomach by the interaction of bacterial and host products, monochloramine has been shown to dysregulate Ca2+homeostasis and disrupt mucosal integrity. In this report, we show that monochloramine also leads to disturbances in intracellular free zinc concentration ([Zn2+]i) in the gastric gland of the rabbit and that the increased Zn2+within the cell causes an independent decrease in cell viability. Changes in [Zn2+]iwere measured by using the fluorescent reporter FluoZin-3, whereas cell viability was assessed by measuring the conversion of calcein-AM to fluorescent calcein, an assay that is not affected by intracellular oxidation state. Cell death was confirmed using propidium iodide and YO-PRO-1 dye uptake measurements. Our experiments demonstrate that [Zn2+]iis increased in gastric glands exposed to NH2Cl and that elevated [Zn2+]idecreases cell viability. Chelation of Zn2+with tetrakis-(2-pyridylmethyl) ethylenediamine decreases the toxicity of NH2Cl, but only when administered concurrently. These findings suggest that the toxic effect of thiol oxidants present during chronic gastritis is partially due to dysregulation of [Zn2+]iearly in the process and that zinc chelation can protect, but not rescue, gastric glands exposed to toxic doses of NH2Cl.