We have previously reported that exogenous nicotinamide adenine dinucleotide hydrate (NAD) decreases basal O-GlcNAc levels as well as attenuates the increase in O-GlcNAc that occurs in response to glucose deprivation (GD); however, the mechanism by which NAD regulates O-GlcNAcylation is unknown. Recent studies have shown that a major pathway for extracellular NAD metabolism is via the cell surface glycohydrolases, CD38 and the resulting formation of cADPR. Therefore, the goal of this study was to determine whether the effects of NAD on the cellular responses to GD were mediated via CD38 and cADPR. Consistent with earlier studies AC16 cardiomyocytes subjected to GD for up to 24hrs, demonstrated significant increases in O-GlcNAcylation as well as with increased protein levels of CHOP, BIP, P-PERK/T-PERK and LC3-II/LC3-I ratio; all of which were markedly attenuated with the addition of NAD (250µM). The addition of the CD38 inhibitor,Luteolinidin chloride (100µM), significantly blunted the effects of NAD on the cellular responses to GD. Conversely, the addition of cADPR (100µM) mimicked the effects of NAD, attenuating the GD-induced increase in O-GlcNAc and CHOP, BIP, P-PERK/T-PERK and LC3-II/LC3-I. These results demonstrate for the first time that NAD inhibits multiple cell survival pathways including increased protein O-GlcNAcylation, ER stress and autophagy through a CD38 and cADPR mediated pathway.
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