In the endo/sarcoplasmic reticulum (ER/SR), hexose-6-phosphate dehydrogenase (H6PDH) generates an NADPH/ NADP ratio sufficient to drive 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1)-mediated glucocorticoid (GC) activation (11-DHC to corticosterone). In H6PDH knockout (H6PDKO) mice a reduced NADPH/NADP ratio leads to reversed 11β-HSD1 activity (corticosterone to 11-DHC), and GC insensitivity. They also display skeletal myopathy driven by activation of the unfolded protein response (UPR), which is 11β-HSD1 independent. We have begun to evaluate a novel H6PDH allele (euH6PDKO purchased from EUCOMM), generated to carry out conditional mutagenesis, in which initial recombination of a LacZ reporter cassette into intron 2 is intended to completely knockout H6PDH. We have now characterised 11β-HSD1 activity and muscle function in euH6PDKO and previously generated H6PDKO mice. We can demonstrate low-level H6PDH mRNA in euH6PDKO liver (21-fold decrease) and muscle (5-fold decrease) compared to control which is absent in H6PDHKOs. 11β-HSD1 activity in hepatic microsomes and muscle explants in both euH6PDKOs and H6PDKOs is switched as previously described, meaning GCs produced by the adrenal glands are inactivated in the peripheral tissues. This is reflected in urinary steroid analysis which shows that approximately 90% of GC metabolites are inactive 11-DHC, compared to 10% in control mice. However, histological analysis of quadriceps from 6 week old euH6PDKOs shows no evidence of the myopathy present in H6PDKOs, and no upregulation of the UPR. Finally, while NADPH production is absent in H6PDKO microsomes, we detected ≤10% vs. control levels in the euH6PDKO, potentially explaining the absence of myopathy. These data suggest that the euH6PDKO allele is hypomorphic, and that low level NADPH production is sufficient to rescue myopathy, but insufficient to rescue the defects in 11β-HSD1-mediated GC metabolism. This allele represents a useful tool to further assess SR function and 11β-HSD1 mediated GC metabolism in metabolic disease.
Declaration of interest: There is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.
Funding: No specific grant from any funding agency in the public, commercial or not-for-profit sector.