Background: Using a deuterated tracer (D4-cortisol) to measure cortisol regeneration from cortisone in vivo we have quantified tissue-specific 11βHSD1 activity in health and disease, and acute regulation eg by insulin. These studies relied on tracer enrichment in the total plasma cortisol pool, assuming rapid equilibration between free and protein-bound pools. Slower equilibration would result in underestimation of enzyme activity, and has implications for the contribution of protein-bound cortisol to glucocorticoid action. Here, we investigated cortisol turnover in the free compared with the total cortisol pool at steady state and after acute up-regulation of 11βHSD1 by a high carbohydrate meal.
Methods: As reported (J Clin Endocrinol Metab 2014 99 160), 8 healthy men were infused with D4-cortisol before consuming blinded liquid meals containing minimal calories (placebo), high protein or high carbohydrate in random order. Plasma samples were obtained every 15 min for 3 h after the meals. Free steroids were separated using equilibrium dialysis. Deuterated and endogenous cortisol concentrations were quantified using LCMS/MS. To achieve the sensitivity necessary multiple plasma samples were pooled from each subject across steady state and with each meal type. Data are mean±S.E.M.
Results: At steady state, dilution of D4-cortisol by endogenous cortisol and D3 cortisol were similar in the free vs total pools, yielding similar rates of appearance (Ra) of cortisol and D3-cortisol (Ra cortisol 59.0±7.2 nmol/min vs 54.9±6.6 nmol/min and Ra D3 cortisol 20.1±1.1 nmol/min vs 19.4±1.0 nmol/min). With acute perturbation, similar up-regulation of Ra cortisol (after protein or carbohydrate) and Ra D3-cortisol (after carbohydrate) was observed using enrichments measured in the free and total plasma cortisol pools.
Conclusion: Rapid equilibration occurs between free and bound cortisol pools in plasma, and measurement of cortisol turnover in the total pool does not underestimate 11βHSD1 activity.