Abstract 655: Paradoxical Reduction in Glycosylation Sensitizes the Diabetic Heart to Mitochondrial Permeability Transition

Abstract

Hexosamine biosynthesis, an accessory pathway for glucose metabolism, culminates in the formation of UDP-GlcNAc, which is the monosaccharide donor for the post-translational modification, O-linked beta-N-acetylglucosamine (O-GlcNAc). Augmented hexosamine biosynthesis has been implicated in the pathogenesis of diabetes in many peripheral tissues. Yet, glucose transporters in cardiac myocytes are predominantly insulin dependent, thereby limiting intracellular glucose availability in diabetic cardiac myocytes. Driven by such paradoxes, we hypothesized that unlike other tissues, intracellular O-GlcNAc signaling would actually be reduced in diabetic hearts. HPLC revealed a 24% reduction in UDP-GlcNAc levels in db/db diabetic (2.2 +/−0.5 nmol) compared to nondiabetic (2.9 +/−0.5 nmol) mouse hearts. Immunoblots showed a reduced expression of O-GlcNAc transferase (61 +/−14% of nondiabetic), which is the enzyme that adds the O-GlcNAc modification to proteins, and reduced O-GlcNAc levels (86 +/−8% of nondiabetic) in diabetic hearts (n = 8/group). We recently reported an interaction between O-GlcNAc signaling and mitochondrial-controlled cell death and, accordingly, evaluated the sensitivity to mitochondrial permeability transition pore (mPTP) formation here. Cardiac mitochondria from diabetic hearts were more sensitive to calcium-induced mitochondrial swelling (138 +/−15% of nondiabetic) compared with mitochondria from nondiabetics (n = 3/group). Because diabetes is multi-faceted, we distilled the system to one variable, reduced O-GlcNAc levels, by injecting additional groups of nondiabetic mice with a drug (TT04) to reduce O-GlcNAc levels, or Vehicle (n =3/group). Cardiac mitochondria from nondiabetic mice with reduced O-GlcNAc levels (TT04) were also more sensitive to calcium-induced swelling (178 +/−12% of Vehicle) than cardiac mitochondria from Vehicle treated mice, thus recapitulating one element of the diabetic phenotype and supporting the idea that reduction of O-GlcNAc levels is sufficient to sensitize cardiac mPTP formation. We conclude that the paradoxical reduction in O-GlcNAc signaling in diabetic hearts may be responsible for heightened sensitivity to cardiac injury, particularly in terms of mPTP formation.