Menadione-induced oxidative stress inhibits cholecystokinin-stimulated secretion of pancreatic acini by cell dehydration

The present study evaluated the effects of free radicals generated by menad ione on morphology and function of pancreatic acinar cells focusing on enzy me secretion, stimulus-secretion coupling, and cell hydration. Various expe riments evaluated morphology and function of isolated rat pancreatic acinar cells exposed to menadione. Menadione instantaneously generated free radic als (luminol and deoxyribose assays) followed by a time-dependent cell inju ry (uptake of trypan blue). Early ultrastructural changes included vacuoliz ation and alterations of mitochondria, endoplasmic reticulum, and nucleus. Menadione caused a rapid glutathione oxidation followed by a depletion in r educed glutathione. An increase in lipid peroxides and a depletion of adeno sine triphosphate were seen only after 30-60 minutes. Menadione markedly in hibited amylase release stimulated by cholecystokinin (CCK) and carbachol a nd simultaneously caused cell shrinkage after a few minutes. Similar degree s of cell shrinkage induced by hyperosmolar incubation and by menadione inh ibited amylase secretion to a similar extent. CCK binding and its effect on calcium and inositol 1,4,5-trisphosphate (IP3) were not affected by menadi one. Menadione (without CCK) induced an instantaneous increase of intracell ular calcium followed by a slow constant increase. In single cells, menadio ne induced calcium oscillations with a frequency lower than that seen after CCK stimulation. Some morphologic and functional alterations owing to mena dione-induced oxidative stress may be caused by adenosine triphosphate and glutathione depletion, lipid peroxidation, and changes in cytosolic calcium . The marked inhibition of secretagogue-stimulated enzyme secretion owing t o menadione may be mediated to a large part by cell dehydration, whereas cl assical steps of stimulus-secretion coupling like receptor binding, calcium release, and IP3 generation remained unchanged.