Endocrine Abstracts (2016) 44 P189 | DOI: 10.1530/endoabs.44.P189

Mechanisms of ageing metabolic decline revealed by targeted metabolomics and energy metabolism in NAD+ depleted skeletal muscle

Rachel Fletcher1,2, Lucy Oldacre-Bartley1,2, Craig Doig1,2, Charles Brenner3 & Gareth Lavery1,2


1Institute of Metabolism and Systems Research, University of Birmingham, Birmingham, B15 2TT, UK; 2Centre for Endocrinology, Diabetes and Metabolism, Birmingham Health Partners, Birmingham, B15 2TT, UK; 3University of Iowa, Iowa City, Iowa, USA.


Nicotinamide adenine dinucleotide (NAD+) levels increase during metabolic stress, which acts as a consumed substrate by, amongst other proteins, the sirtuins, which adapt transcriptional programmes to increase energy availability and regulate insulin sensitivity. Thus, maintaining appropriate skeletal muscle NAD+ availability is critical for regulating systemic energy homeostasis. In order to gain better insight into ageing muscle NAD+ dynamics we used targeted LC/MS-based metabolomics and assessed NAD+ associated metabolome in young (3-months) and aged (30-months) mouse quadriceps (n=8). In aged muscle we identified significantly reduced NAD+(~20%). ADPribose was also significantly reduced (~30%). ADPribose is a product of NAD+ consumption and signalling molecule, implying reduced sirtuin mediated NAD+ turnover and impaired energy harvesting. Thus, low NAD+ and ADPribose levels corroborate the significantly reduced ATP levels (~40%). Following NAD+ consumption, nicotinamide is also released. Nicotinamide is recycled by the enzyme nicotinamide phosphoribosyltransferase (NAMPT) and critical to maintaining intracellular NAD+ levels. To model aged-related NAD+ decline, we exposed primary muscle myotubes (C57BL/6J quadriceps) to the NAMPT inhibitor FK866 (100 nM, 48–72 h) to deplete NAD+. After 48–72 h basal oxygen consumption was significantly reduced by 50% implying severely impaired mitochondrial function. Furthermore we also observed increased caspase 3 activity after 72 h indicative of cellular apoptosis initiation. 72 h FK866 treated myotubes were supplemented with the NAD+ precursor nicotinamide riboside (NR) for the final 24 h, which fully restored NAD+ levels, mitochondrial function and cell viability. Nicotinamide was unable to rescue these effects and supports the notion that muscle has a limited NAD+ salvage system which comprises NAMPT, and NR kinases (NMRK) 1/2 to maintain NAD+ availability from extracellular sources. Overall these data identify perturbed NAD+ dynamics in aged muscle and its potential impact on energy homeostasis that may underpin age-related metabolic decline. Enhancing NAD+ by NR supplementation may prove a useful nutraceutical approach to combat age-related muscle decline.

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