The redistribution of cathepsin B, a lysosomal enzyme, from the lysosomal pellet to the zymogen pellet in the subcellular fractionation, the colocalization of cathepsin B with digestive enzyme, and increased cellular, lysosomal, and mitochondrial fragility within acinar cells have been found during the early stages of caerulein-induced acute pancreatitis in rats. In the present study, the authors investigated the protective effects of prostaglandin E1 and E2, a combined therapy of these prostaglandins, and a new, synthetic, low molecular weight protease inhibitor, ONO3307, on the exocrine pancreas in this noninvasive model of experimental pancreatitis in vivo and in vitro. Prostaglandin E2, but not E1, prevented hyperamylasemia, congestion of amylase and trypsinogen in the acinar cells, redistribution of cathepsin B, and amylase and lactate dehydrogenase discharge from the dispersed acini. It also prevented cathepsin B leakage from the lysosomes and malate dehydrogenase leakage from the mitochondria in an almost dose-dependent manner, particularly at the dose of 100 μg/kg/hr continuous infusion. Furthermore, the combined therapy of prostaglandin E2 with ONO3307 strongly inhibited all the parameters tested in this study. This combination therapy seems to be the most effective against secretagogue-induced pancreatic injuries. These results indicate that cellular and sub-cellular organellar fragility seem to be closely involved in the pathogenesis of acute pancreatitis. Prostaglandin E2 seems to have important cytoprotective effects on the biologic membranes, such as a stabilizer of lysosomal or mitochondrial membranes. In addition, these findings also suggest the crucial roles of some unknown proteases in the etiology of acute pancreatitis, and indicate the clinical effectiveness of prostaglandins and this type of low molecular weight protease inhibitor for acute pancreatitis.