Inhibition of 11β-HSD1 activity in intact cells.

<p>(A), Inhibition of 11β-HSD1-dependent cortisol formation in intact HEK-293 cells. HEK-293 cells stably co-expressing human 11β-HSD1 and H6PDH were incubated for 30 min in the presence of 200 nM radiolabeled cortisone and either 1 μM of the positive control inhibitor glycyrrhetinic acid (CTRL 1) or 100 nM and 1 μM of the respective test compounds. Formation of cortisol was determined by separation of steroids by TLC and scintillation counting. Data represent mean ± SD of three independent experiments, each performed in triplicate. (B), Inhibition of 11β-HSD1-dependent cortisol formation in intact human keratinocytes. Primary human keratinocytes were grown for two days, followed by incubation for 24 h with 1 μM of cortisone and various concentrations of inhibitor. Compound CAS 1009373-58-3 (Merck, CTRL 4) was used as positive control. An additional vehicle control was measured in the absence of exogenous cortisone (black bar). Formation of cortisol was determined by an enzyme immune assay. Data represent mean ± SD from three experiments. Shapiro-Wilk test was used to assess the normality of data. One-way analysis of variance (ANOVA) and Dunnett's multiple-comparison test were performed to evaluate differences between inhibitor treatments compared to the solvent control. All values were significantly different (p<0.01) compared to vehicle control in the presence of cortisone, except treatment of keratinocytes with 10 nM <b>2b</b>.