ACS DIVISION OF BIOLOGICAL CHEMISTRY

July 3rd, 2014 by Samad, A., James, A., Wong, J., Mankad, P., Whitehouse, J., Patel, W., Alves-Simoes, M., Siriwardena, A. K., Bruce, J. I. E.

Acute pancreatitis is a serious and sometimes fatal inflammatory disease where the pancreas digests itself. The non-oxidative ethanol metabolites palmitoleic acid (POA) and POA-ethylester (POAEE), are reported to induce pancreatitis due to impaired mitochondrial metabolism, cytosolic Ca2+ ([Ca2+]i) overload and necrosis of pancreatic acinar cells. Metabolism and [Ca2+]i are linked critically by the ATP-driven plasma membrane Ca2+-ATPase (PMCA) important for maintaining low resting [Ca2+]i. The aim of the current study was to test the protective effects of insulin on cellular injury induced by the pancreatitis-inducing agents, ethanol, POA and POAEE. Rat pancreatic acinar cells were isolated by collagenase digestion and [Ca2+]i was measured by fura-2 imaging. An in situ [Ca2+]i clearance assay was used to assess PMCA activity. Magnesium green (MgGreen) and a luciferase-based ATP kit was used to assess celular ATP depletion. Ethanol (100 mM) and POAEE (100 µM) induced a small but irreversible Ca2+ overload response but had no significant effect on PMCA activity. POA (50-100 µM) induced a robust Ca2+ overload, ATP depletion, inhibited PMCA activity and consequently induced necrosis. Insulin pretreatment (100 nM for 30 minutes) prevented the POA-induced Ca2+ overload, ATP depletion, inhibition of the PMCA and necrosis. Moreover, the insulin-mediated protection of the POA-induced Ca2+ overload was partially prevented by the phosphoinositide-3-kinase (PI3K) inhibitor, LY294002. These data provide the first evidence that insulin directly protects pancreatic acinar cell injury induced by bona fide pancreatitis-inducing agents, such as POA. This may have important therapeutic implications for the treatment of pancreatitis.