Induction of the nicotinamide riboside kinase NAD⁺ salvage pathway in skeletal myopathy of H6PDH KO mice

Abstract

1Background: Hexose-6-Phosphate Dehydrogenase (H6PDH) is the only known generator 2 of the pyridine nucleotide NADPH from NADP + in the Endoplasmic/Sarcoplasmic Reticulum 3 (ER/SR). However, its ability to influence cell wide pyridine biosynthesis and metabolism is 4 unclear. 5Methods: Skeletal muscle from H6PDH Knockout mice (H6KO) and Wild type controls (WT) 6 was subject to metabolomic assessment and pathway analysis. NAD boosting was 7 performed through intraperitoneal injection of Nicotinamide Riboside (NR) (400mg/kg) twice 8 daily for 4 days. Mice were culled and skeletal muscle tissue collected the next day. Double 9 knockout mice were generated through to breeding of H6KO with NRK2 knockout mice. 10These mice were assessed for NAD levels and ER stress response. 11Results: H6KO skeletal muscle shows significant changes in the pathway regulating NAD+ 12 biosynthesis with the Nicotinamide Riboside Kinase 2 gene (NRK2) being the most elevated 13 gene. Assessment of the metabolic impacts of this dysregulation showed decreases in 14 mitochondrial respiration and Acylcarnitine metabolism. Pharmacologically boosting NAD+ 15 levels with NR had no significant impact over ER stress or Acetyl-CoA metabolism. A duel 16 H6-NRK2 KO mouse exhibited changes in NAD + levels but no overt change in metabolism 17 compared to the H6KO mouse. 18Conclusions: This work shows a relationship between ER/SR status and wider muscle cell 19 metabolism. The utility of NAD + boosting to skeletal muscle is demonstrated however, it is 20 also evident that the extent of cellular stress and/or REDOX status may overcome any 21 beneficial impact. 22 23