Endocrine Abstracts

11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) is an NADPH-dependant oxo-reductase located in the sarcoplasmic reticulum (SR) lumen of skeletal muscle. Here it generates active glucocorticoids to regulate permissive and adaptive metabolism, and can mediate the pathological effects of glucocorticoid excess. Hexose-6-phosphate dehydrogenase (H6PD) in the SR interacts with 11β-HSD1 to generate an appropriate NADPH/NADP+ ratio to support activity. H6PD depletion impairs SR NADPH generation causing 11β-HSD1 to assume glucocorticoid inactivating dehydrogenase activity. We tested whether modulating cellular nicotinamide adenine dinucleotide (NAD+) availability (as the parent molecule of NAD(P)(H)) influenced 11β-HSD1 activity in the SR. We used FK866 to inhibit nicotinamide phospho-ribosyltransferase (NAMPT, rate-limiting enzyme in NAD+ biosynthesis) to deplete NAD(P)(H) in mouse C2C12 myotubes. 48 h FK866 treatment impaired cellular energetic status, reducing NAD+(>90%), NADP+(>50%) and ATP (>30%) levels without inducing apoptosis or affecting cell viability. 11β-HSD1 reductase activity was 30% that of untreated cells (152±18 vs. 512±44 pmol steroid/mg protein/h respectively, P<0.005). Despite impaired reductase activity, FK866 treatment did not induce 11β-HSD1 dehydrogenase activity, likely reflecting an intact SR 11β-HSD1-H6PD system. To examine the mechanisms of FK866 mediated suppression of 11β-HSD1 activity, cells were co-treated with the NAD+ precursor nicotinamide riboside (NR, 0.5 mM) to bypass NAMPT inhibition. NR supplemented to 48 h FK866 treated myotubes for the last 24 h fully restored NAD+ levels and fully rescued 11β-HSD1 activity. We next ascertained the time course of the NR rescue effect. Intriguingly, 11β-HSD1 activity normalised in proportion to the duration of NR supplementation, with as little as 30 min inducing a significant increase in 11β-HSD1 activity (202±22 pmol steroid/mg protein/h, P<0.05). These data suggest a novel level of regulated glucocorticoid metabolism in skeletal muscle whereby 11β-HSD1 activity can be influenced by cellular redox status and NAD+ levels beyond the SR.