11β-HSD1 is the major regulator of the tissue-specific effects of circulating glucocorticoid excess

Abstract

The adverse metabolic effects of prescribed and endogenous glucocorticoid (GC) excess, Cushing syndrome, create a significant health burden. We found that tissue regeneration of GCs by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), rather than circulating delivery, is critical to developing the phenotype of GC excess; 11β-HSD1 KO mice with circulating GC excess are protected from the glucose intolerance, hyperinsulinemia, hepatic steatosis, adiposity, hypertension, myopathy, and dermal atrophy of Cushing syndrome. Whereas liver-specific 11β-HSD1 KO mice developed a full Cushingoid phenotype, adipose-specific 11β-HSD1 KO mice were protected from hepatic steatosis and circulating fatty acid excess. These data challenge our current view of GC action, demonstrating 11β-HSD1, particularly in adipose tissue, is key to the development of the adverse metabolic profile associated with circulating GC excess, offering 11β-HSD1 inhibition as a previously unidentified approach to treat Cushing syndrome.

Keywords: HSD11b1; cortisol; endocrinology; hypercortisolemia; steroids.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

WT and 11β-HSD1 KO mice administered CORT via the drinking water results in elevated circulating CORT levels. C57BL/6 WT (white bars) and 11β-HSD1 KO mice (GKO, black bars) were treated with CORT (100 μg/mL) or vehicle via the drinking water for 5 wk (n = 7–9 in each group). Elevated circulating CORT (A) and suppressed adrenal weights (B) were observed in both CORT-treated WT and GKO mice. Data were analyzed using two-way ANOVA; see Fig. S1 for the complete dataset used in the analysis. ***P < 0.001 vs. WT vehicle; $$$P < 0.001 vs. GKO vehicle. (C) Schematic diagram summarizing the three mechanisms by which circulating CORT results in GR activation: (1) direct binding of circulating CORT with the GR in peripheral tissues; (2) inactivation of circulating CORT by 11β-HSD2, then reactivation in peripheral tissues by 11β-HSD1; and (3) GR activation increases 11β-HSD1 expression and activity, further amplifying intracellular CORT availability.

Fig. 2.

Deletion of 11β-HSD1 ameliorates the adverse metabolic side-effect profile induced by CORT. C57BL/6 WT (white bars) and 11β-HSD1 KO mice (GKO, black bars) were treated with CORT (100 μg/mL) or vehicle via the drinking water for 5 wk (n = 7–9 in each group). Glucose tolerance (A), calculated area under the curve (AUC) for glucose tolerance data (B), fasting insulin levels (C), fasting glucose (D), HOMA-IR index (E), and systolic blood pressure (BP) (F) were improved in CORT-treated GKO mice compared with CORT-treated WTs. Data were analyzed using two-way ANOVA; see Fig. S1 for the complete dataset used in the analysis. **P < 0.01, ***P < 0.001 vs. WT vehicle; ∅P < 0.05, ∅∅P < 0.01, ∅∅∅P < 0.001 vs. WT CORT.

Fig. 3.

The 11β-HSD1 KO mice are protected from the development of CORT-induced hepatic steatosis. Frozen liver sections stained with oil red O (A) and a quantitative TAG assay (B) revealed 11β-HSD1 KO (GKO) mice were protected from CORT-induced hepatic steatosis. GKO mice were also protected from increased serum free fatty acids (C) and increased haptic mRNA expression of CD36 (D) induced by CORT. The expression of the key lipogenic modulators in the liver, ACC1, FAS, DGAT1, PPARG, and CREB, was unaffected by CORT in both WT and GKO mice (D). Data were analyzed using two-way ANOVA; see Fig. S2 for the complete dataset used in the analysis. Hepatic 11β-HSD1 mRNA expression (E) and oxo-reductase activity (F) were increased following CORT treatment in WT mice but not GKO mice. Data were analyzed using Student t tests (n = 7–9 in each group). **P < 0.01, ***P < 0.001 vs. WT vehicle; ∅P < 0.05, ∅∅∅P < 0.001 vs. WT CORT. C, CORT; N.D, not detected; V, vehicle.

Fig. 4.

The 11β-HSD1 KO mice are protected from increased adiposity induced by CORT. The 11β-HSD1 KO mice (GKO, black bars) were protected from CORT-mediated increases in gonadal (A), s.c. (B), retroperitoneal (C), and mesenteric (D) adiposity compared with CORT-treated WT mice (white bars). H&E staining of paraffin-embedded gonadal fat sections revealed increased adipocyte size in both CORT-treated WT and GKO mice (E and G). A similar increase in adipocyte size was observed in the s.c. adipose tissue depot (F). The mRNA expression of the key lipogenic mediatorsACC1, FAS, PPARG, and CREB was unaffected by CORT treatment in both WT and GKO mice, whereas IRS2 and DGAT1 were increased by CORT in WT mice only (H). The mRNA expression of key lipolytic genes was increased in gonadal adipose tissue of CORT-treated WT mice, but not CORT- treated GKOs (I). Data were analyzed using two-way ANOVA; see Fig. S3 for the complete dataset used in the analysis. The 11β-HSD1 mRNA expression (J) and oxo-reductase activity (K) in adipose tissue were increased in CORT-treated WT mice but not GKO mice. Data were analyzed using Student t tests (n = 7–9 in each group). *P < 0.05, **P < 0.01 vs. WT vehicle; ∅P < 0.05, ∅∅P < 0.01 vs. WT CORT; $P < 0.05, $$P < 0.01 vs. GKO vehicle. C, CORT; N.D, not detected; V, vehicle. (Scale bars, 100 μm.)

Fig. 5.

The 11β-HSD1 KO mice are protected from skeletal myopathy induced by CORT. Grip strength (A), quadriceps muscle bed weights (B), and tibialis anterior muscle bed weights (C) were reduced in CORT-treated WT mice (white bars), whereas 11β-HSD1 KO mice (GKO, black bars) were protected from the effects of CORT. Paraffin-embedded quadriceps muscle sections stained with H&E revealed a smaller myofiber diameter in CORT-treated WT mice, whereas CORT was without effect on myofiber diameter in GKO animals (D) (Scale bars, 200 μm.) Similarly, GKO mice were protected from the induction of several muscle atrophy markers including MuRF1 (E), atrogin-1 (F), myostatin (G), and FoxO1 (H) following CORT treatment. Data were analyzed using two-way ANOVA; see Fig. S4 for the complete dataset used in the analysis. The 11β-HSD1 mRNA expression (I) and oxo-reductase activity (J) in quadriceps muscle was increased in CORT-treated WT mice but not GKO mice. Data were analyzed using Student t tests (n = 7–9 in each group). *P < 0.05, **P < 0.01, ***P < 0.001 vs. WT vehicle; ∅P < 0.05 vs. WT CORT. N.D, not detected.

Fig. 6.

The 11β-HSD1 KO mice are protected from skin thinning induced by CORT. Paraffin-embedded skin sections stained with H&E revealed dramatically reduced skin thickness in WT mice following CORT treatment (A). By contrast, CORT was without effect on skin thickness in 11β-HSD1 KO mice (GKO) mice (A). (Scale bars, 200 μm.) Dermal thickness was quantified using ImageJ software (B). In agreement with the histology, GKO mice were shielded from decreased expression of several genes involved in collagen biosynthesis and processing in skin induced by CORT, including Col1A1 (C), Leprel1 (D), and PLOD1 (E), but not LOX (F). n = 3 for skin histology and n = 7–9 for gene expression and 11β-HSD1 activity data. Data were analyzed using two-way ANOVA; see Fig. S5 for the complete dataset used in the analysis. The 11β-HSD1 mRNA expression (G) and oxo-reductase activity (H) in skin were increased in CORT-treated WT mice but not GKO mice. Data were analyzed using Student t tests. *P < 0.05, **P < 0.01 vs. WT vehicle; ∅P < 0.05 vs. WT CORT. N.D, not detected.

Fig. 7.

Fat-specific 11β-HSD1 KO mice, and not liver-specific 11β-HSD1 KO mice, are protected from the development of CORT-induced hepatic steatosis. Frozen liver sections stained with oil red O (A) and hepatic TAG quantification (B) revealed adipose-specific 11β-HSD1 KO mice (FKO, black bars), and not liver-specific 11β-HSD1 KO mice (LKO, gray bars), were protected from CORT-induced hepatic steatosis, compared with CORT-treated WT controls (white bars). Similarly, FKO mice (and not LKO mice) were shielded from increased serum free fatty acids (C) and increased mRNA expression of key lipolytic mediators in adipose tissue (D) following CORT treatment. FKO mice were not protected from increased expression of the hepatic free fatty acid transporter CD36 induced by CORT in WT mice (E). Data were analyzed using two-way ANOVA; see Figs. S9 and S10 for the complete datasets used in the analysis (n = 6–7 in each group). *P < 0.05, ***P < 0.001 vs. WT vehicle; ∅P < 0.05, ∅∅P < 0.01 vs. WT CORT; $P < 0.05, $$P < 0.01, $$$P < 0.001 vs. LKO vehicle. C, CORT; V, vehicle.