Perturbations of NAD + salvage systems impact mitochondrial function and energy homeostasis in mouse myoblasts and intact skeletal muscle

Am J Physiol Endocrinol Metab. 2018 Apr 1;314(4):E377-E395. doi: 10.1152/ajpendo.00213.2017. Epub 2017 Dec 5.

Abstract

Nicotinamide adenine dinucleotide (NAD+) can be synthesized by nicotinamide phosphoribosyltransferase (NAMPT). We aimed to determine the role of NAMPT in maintaining NAD+ levels, mitochondrial function, and metabolic homeostasis in skeletal muscle cells. We generated stable Nampt knockdown (sh Nampt KD) C2C12 cells using a shRNA lentiviral approach. Moreover, we applied gene electrotransfer to express Cre recombinase in tibialis anterior muscle of floxed Nampt mice. In sh Nampt KD C2C12 myoblasts, Nampt and NAD+ levels were reduced by 70% and 50%, respectively, and maximal respiratory capacity was reduced by 25%. Moreover, anaerobic glycolytic flux increased by 55%, and 2-deoxyglucose uptake increased by 25% in sh Nampt KD cells. Treatment with the NAD+ precursor nicotinamide riboside restored NAD+ levels in sh Nampt cells and increased maximal respiratory capacity by 18% and 32% in control and sh Nampt KD cells, respectively. Expression of Cre recombinase in muscle of floxed Nampt mice reduced NAMPT and NAD+ levels by 38% and 43%, respectively. Glucose uptake increased by 40%, and mitochondrial complex IV respiration was compromised by 20%. Hypoxia-inducible factor (HIF)-1α-regulated genes and histone H3 lysine 9 (H3K9) acetylation, a known sirtuin 6 (SIRT6) target, were increased in shNampt KD cells. Thus, we propose that the shift toward glycolytic metabolism observed, at least in part, is mediated by the SIRT6/HIF1α axis. Our findings suggest that NAMPT plays a key role for maintaining NAD+ levels in skeletal muscle and that NAMPT deficiency compromises oxidative phosphorylation capacity and alters energy homeostasis in this tissue.

Keywords: HIF-1α; NAMPT; SIRT6; glycolysis; nicotinamide riboside.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Carbohydrate Metabolism / genetics
  • Cells, Cultured
  • Cytokines / genetics*
  • Cytokines / metabolism
  • Energy Metabolism / genetics*
  • Homeostasis / genetics
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Mitochondria, Muscle / physiology*
  • Muscle, Skeletal / metabolism*
  • Myoblasts / metabolism*
  • NAD / metabolism*
  • Nicotinamide Phosphoribosyltransferase / genetics*
  • Nicotinamide Phosphoribosyltransferase / metabolism
  • Oxidative Phosphorylation
  • Signal Transduction / genetics

Substances

  • Cytokines
  • NAD
  • Nicotinamide Phosphoribosyltransferase
  • nicotinamide phosphoribosyltransferase, mouse