Role of Calcium in Pancreatic Islet Cell Death by IFN-γ/TNF-α

Abstract

We studied the intracellular events associated with pancreatic β cell apoptosis by IFN-γ/TNF-α synergism. IFN-γ/TNF-α treatment of MIN6N8 insulinoma cells increased the amplitude of high voltage-activated Ca2+ currents, while treatment with IFN-γ or TNF-α alone did not. Cytosolic Ca2+ concentration ([Ca2+]c) was also increased by IFN-γ/TNF-α treatment. Blockade of L-type Ca2+ channel by nifedipine abrogated death of insulinoma cells by IFN-γ/TNF-α. Diazoxide that attenuates voltage-activated Ca2+ currents inhibited MIN6N8 cell death by IFN-γ/TNF-α, 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-γ/TNF-α. Following the increase in [Ca2+]c, calpain was activated, and calpain inhibitors decreased insulinoma cell death by IFN-γ/TNF-α. As a downstream of calpain, calcineurin was activated and the inhibition of calcineurin activation by FK506 diminished insulinoma cell death by IFN-γ/TNF-α. BAD phosphorylation was decreased by IFN-γ/TNF-α because of the increased calcineurin activity, which was reversed by FK506. IFN-γ/TNF-α 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-γ/TNF-α treatment. These results indicate that IFN-γ/TNF-α synergism induces pancreatic β 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.

Cytokines Tumor Necrosis Factor-α and Interferon-γ Induce Pancreatic β-Cell Apoptosis through STAT1-mediated Bim Protein Activation

“…The group of BH3-only sensitizer proteins is composed of DP5, Bad, Noxa, Bik, and other Bcl-2 members (9, 11). Bad was described to be activated by calcineurin in TNF-␣ ϩ IFN-␥-treated MIN6N8 cells (39). We also evaluated the putative response of Noxa and Bik in ␤-cells, but these proteins were not induced by TNF-␣ ϩ IFN-␥ (data not shown).…”

Novel Mechanisms of Target Cell Death and Survival and of Therapeutic Action of IVIg in Pemphigus

“…71 TNF-␣ has been shown to activate calpains because of a rise in the concentration of intracellular free Ca 2ϩ . 72 It is well established that TNF-␣ is elevated in the serum of PV patients, [73][74][75][76] deposited in the lesional skin, 77,78 present in the blister fluid, 74 and up-regulated in keratinocytes due to PV antibody binding 78,79 and that anti-TNF-␣ therapy with etanercept (a fusion protein of TNF-␣ receptor) can ameliorate recalcitrant PV. 80,81 During cell injury, calpain functions as a signal to mediate the influx of extracellular Ca 2ϩ after ATP depletion and endoplasmic reticulum Ca 2ϩ release.…”

Autophagy deficiency in beta cells leads to compromised unfolded protein response and progression from obesity to diabetes in mice

“…Islet cell death Primary pancreatic islets were isolated from fasted mice using the collagenase digestion technique [15]. Islets were placed on Millicell culture plate inserts (Millipore, Carrigtwohill, Ireland) and treated with thapsigargin (Tg; Sigma, St Louis, MO, USA), palmitic acid (PA; Sigma) or oleic acid (OA; Sigma) as described [16].…”

“…cDNA was synthesised using Superscript II (Invitrogen) and oligo(dT) [12][13][14][15][16][17][18] primer. Expression of UPR-related genes, antioxidant genes and p85α and p85β subunits of phosphoinositide 3-kinase (PI3K) was examined by real-time RT-PCR analysis using specific primers (electronic supplementary material [ESM] Table 1) [13].…”