Monochloramine-induced toxicity and dysregulation of intracellular Zn ²⁺ in parietal cells of rabbit gastric glands

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 ([Zn ²⁺]_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⁺]_i were 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⁺]_i is increased in gastric glands exposed to NH₂Cl and that elevated [Zn²⁺]_i decreases 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⁺]_i early in the process and that zinc chelation can protect, but not rescue, gastric glands exposed to toxic doses of NH₂Cl.