Distinguishing the activities of 11beta-hydroxysteroid dehydrogenases in vivo using isotopically labeled cortisol.
The isozymes of 11beta-hydroxysteroid dehydrogenase (11betaHSDs) catalyze the interconversion of cortisol and cortisone. The type 2 dehydrogenase inactivates cortisol to cortisone, whereas the type 1 catalyzes predominantly the reverse reductive reaction. These reactions take place in different tissues, where they are subject to distinct regulation, and may be important in common pathologies. Current methods to determine the activities of these enzymes in vivo rely only on the balance between cortisol and cortisone, do not measure turnover, and cannot distinguish between the two reactions. We have investigated the use of [9,11,12,12-2H4]cortisol (d4F) to distinguish the dehydrogenase and reductase activities. On metabolism by dehydrogenation, d4F loses 11alpha- deuterium, forming trideuterated cortisone (d3E) and is regenerated by reduction to trideuterated cortisol (d3F). Healthy men (n = 6) participated in a randomized, double blind, cross-over study comparing oral placebo and the 11betaHSD inhibitor, carbenoxolone (100 mg every 8 h for 7 d). d4F and its metabolites were measured in plasma and urine during a steady state infusion. Inhibition of 11betaHSDs by carbenoxolone was measured by increased steady state concentrations of d4F (41 +/- 5.1 vs. 48 +/- 7.7 nM; P < 0.05) and a fall in the rate of appearance of d3F (P < 0.05). 11betaHSD1 reductase activity could be measured specifically as conversion of d3E to d3F (28 +/- 4.2 vs. 17 +/- 3.1 nM; P < 0.05), whereas 11betaHSD2 could be measured by initial rates of appearance of d3E or from urinary ratios of d4F/(d3E + d3F) (0.73 +/- 0.06 vs. 1.02 +/- 0.03; P < 0.05). This technique offers a significant advance in the methods available to measure turnover in 11betaHSDs and isozymes of 11betaHSDs in vivo in human studies, and this study confirms that carbenoxolone inhibits both isozymes of 11betaHSD.
Andrew
Ruth
R
Endocrinology Unit, Department of Medical Sciences, University of Edinburgh, Western General Hospital, Edinburgh, Scotland EH4 2XU, United Kingdom. ruth.andrew@ed.ac.uk
Smith
Kenneth
K
Jones
Gregory C
GC
Walker
Brian R
BR
D042842:11-beta-Hydroxysteroid Dehydrogenases; D000328:Adult; D002229:Carbenoxolone / Q000494:pharmacology; D003348:Cortisone / Q000097:blood / Q000378:metabolism; D018592:Cross-Over Studies; D003903:Deuterium; D004311:Double-Blind Method; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006854:Hydrocortisone / Q000097:blood / Q000378:metabolism*; D006913:Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007262:Infusions, Intravenous; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D007700:Kinetics; D008297:Male; D013058:Mass Spectrometry
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D003903:Deuterium; D006913:Hydroxysteroid Dehydrogenases; D042842:11-beta-Hydroxysteroid Dehydrogenases; D002229:Carbenoxolone; D003348:Cortisone; D006854:Hydrocortisone