Abstract
ObjectiveThe hexosamine biosynthesis pathway (HBP) flux and protein O-linked N-acetyl-glucosamine (O-GlcNAc) levels have been implicated in mediating the adverse effects of diabetes in the cardiovascular system. Activation of these pathways with glucosamine has been shown to mimic some of the diabetes-induced functional and structural changes in the heart; however, the effect on cardiac metabolism is not known. Therefore, the primary goal of this study was to determine the effects of glucosamine on cardiac substrate utilization.MethodsIsolated rat hearts were perfused with glucosamine (0–10 mM) to increase HBP flux under normoxic conditions. Metabolic fluxes were determined by 13C-NMR isotopomer analysis; UDP-GlcNAc a precursor of O-GlcNAc synthesis was assessed by HPLC and immunoblot analysis was used to determine O-GlcNAc levels, phospho- and total levels of AMPK and ACC, and membrane levels of FAT/CD36.ResultsGlucosamine caused a dose dependent increase in both UDP-GlcNAc and O-GlcNAc levels, which was associated with a significant increase in palmitate oxidation with a concomitant decrease in lactate and pyruvate oxidation. There was no effect of glucosamine on AMPK or ACC phosphorylation; however, membrane levels of the fatty acid transport protein FAT/CD36 were increased and preliminary studies suggest that FAT/CD36 is a potential target for O-GlcNAcylation.Conclusion/InterpretationThese data demonstrate that acute modulation of HBP and protein O-GlcNAcylation in the heart stimulates fatty acid oxidation, possibly by increasing plasma membrane levels of FAT/CD36, raising the intriguing possibility that the HBP and O-GlcNAc turnover represent a novel, glucose dependent mechanism for regulating cardiac metabolism.
Highlights
Cardiovascular complications, including diabetic cardiomyopathy are the leading cause of excessive premature morbidity and mortality in diabetic patients
Flux through the hexosamine biosynthesis pathway (HBP) and the synthesis of UDP-GlcNAc is regulated in large part by the metabolism of glucose; this is regulated by L-glutamine-D-fructose 6-phosphate amidotransferase (GFAT), which converts fructose6-phosphate to glucosamine-6-phosphate with glutamine as the amine donor [9]
Most of our understanding of the role of OGlcNAcylation on cellular function is in the context of chronic diseases, including diabetes and increased O-GlcNAc levels have been associated with the adverse effects of hyperglycemia and diabetes on the heart [7,13,14,35,36]
Summary
Cardiovascular complications, including diabetic cardiomyopathy are the leading cause of excessive premature morbidity and mortality in diabetic patients. In contrast to classical protein glycosylation in the ER and Golgi, characterized by stable and complex elongated oligosaccharide structures, O-GlcNAcylation is a dynamic process involving the reversible addition of a single O-GlcNAc moiety to serine and threonine residues of nuclear and cytosolic proteins [8]. This process is regulated by the activities of two key enzymes, O-GlcNAc transferase (OGT), which catalyzes the attachment of O-GlcNAc and N-acetylglucosaminidase (OGlcNAcase), which catalyzes its removal [8]. Flux through the HBP and the synthesis of UDP-GlcNAc is regulated in large part by the metabolism of glucose; this is regulated by L-glutamine-D-fructose 6-phosphate amidotransferase (GFAT), which converts fructose6-phosphate to glucosamine-6-phosphate with glutamine as the amine donor [9]
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