Role of calcium in pancreatic islet cell death by IFN-γ/TNF-α

We studied the intracellular events associated with pancreatic beta cell apoptosis by IFN-gamma/TNF-alpha synergism. IFN-gamma/TNF-alpha treatment of MIN6N8 insulinoma cells increased the amplitude of high voltage-activated Ca2+ currents, while treatment with IFN-gamma or TNF-alpha alone did not. Cytosolic Ca2+ concentration ([Ca2+](c)) was also increased by IFN-gamma/TNF-alpha treatment. Blockade of L-type Ca2+ channel by nifedipine abrogated death of insulinoma cells by IFN-gamma/TNF-alpha. Diazoxide that attenuates voltage-activated Ca2+ currents inhibited MIN6N8 cell death by IFN-gamma/TNF-alpha, while glibenclamide that accentuates voltage-activated Ca2+ currents augmented insulinoma cell death. A protein kinase C inhibitor attenuated MIN6N8 cell death and the increase in [Ca2+](c) by IFN-gamma/TNF-alpha. Following the increase in [Ca2+](c), calpain was activated, and calpain inhibitors decreased insulinoma cell death by IFN-gamma/TNF-alpha. As a downstream of calpain, calcineurin was activated and the inhibition of calcineurin activation by FK506 diminished insulinoma cell death by IFN-gamma/TNF-alpha. BAD phosphorylation was decreased by IFN-gamma/TNF-alpha because of the increased calcineurin activity, which was reversed by FK506. IFN-gamma/TNF-alpha induced cytochrome c translocation from mitochondria to cytoplasm and activation of caspase-9. Effector caspases such as caspase-3 or -7 were also activated by IFN-gamma/TNF-alpha treatment. These results indicate that IFN-gamma/TNF-alpha synergism induces pancreatic beta cell apoptosis by Ca2+ channel activation followed by downstream intracellular events such as mitochondrial events and caspase activation and also suggest the therapeutic potential of Ca2+ modulation in type 1 diabetes.