Role of calcium in thromboxane B2-mediated injury to rabbit gastric mucosal cells

A sustained increase in cytosolic Ca²⁺ can damage gastric mucosal cells. The present study has examined the role of Ca²⁺ in thromboxane B₂ (TXB₂)-mediated damage of rabbit isolated gastric mucosal cells. Cells were isolated from rabbit oxyntic mucosa by collagenase-EDTA digestion. Cell metabolic activity and cell damage were estimated by alamar blue dye absorbance and trypan blue uptake, respectively. Cellular Ca²⁺ was monitored by indo-1 dye fluorescence. Addition of TXB₂ (10^–6 and 10^–8 M) to the cell suspension resulted in a decrease in metabolic activity, and this effect was reduced when Ca²⁺ was removed from the incubation Ca²⁺ and incubation of cells with the intracellular Ca²⁺ chelator, BAPTA-AM (20 M), reduced cell injury in response to TXB₂. Incubation of cells with the Ca²⁺ ionophore A23187 (1–25 M) resulted in a dose-dependent increase in trypan blue uptake and a reduction in cell metabolism. Cell unjury in response to A23187 were exacerbated by addition of TXB₂ (10^–8 M) to the cell suspension. TXB₂ treatment reduced cellular content of reduced glutathione (GSH), while exogenous GSH addition (10 mM) reduced TXB₂-mediated cell injury. These data demonstrate that TXB₂ can directly injure gastric mucosal cells. Gastric mucosal cellular damage in response to TXB₂ is mediated in part by a disruption of Ca²⁺ homeostasis as well as a reduction in cellular GSH content.This work was supported by a grant from the Medical Research Council of Canada MT6426.