Emodin, an 11β-hydroxysteroid dehydrogenase type 1 inhibitor, regulates adipocyte function in vitro and exerts anti-diabetic effect in ob/ob mice

Abstract

Aim: Emodin (1,3,8-trihydroxy-6-methylanthraquinone) is a potent and selective inhibitor of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) with the ability to ameliorate metabolic disorders in diet-induced obese mice. In the present study, we investigated the effects of emodin on adipocyte function and the underlying mechanisms in vitro, and its anti-diabetic effects in ob/ob mice.

Methods: 3T3-L1 adipocytes were used for in vitro studies. 11β-HSD1A activity was evaluated with a scintillation proximity assay. The adipogenesis, glucose uptake, lipolysis and adiponectin secretion were investigated in 3T3-L1 adipocytes treated with emodin in the presence of active (corticosterone) or inactive glucocorticoid (11-dehydrocorticosterone). For in vivo studies, ob/ob mice were administered emodin (25 and 50 mg·kg⁻¹·d⁻¹, ip) for 26 d. On the last day of administration, the serum was collected and the mesenteric and perirenal fat were dissected for analyses.

Results: Emodin inhibited the 11β-HSD1 activity in 3T3-L1 adipocytes in concentration- and time-dependent manners (the IC₅₀ values were 7.237 and 4.204 μmol/L, respectively, after 1 and 24 h treatment. In 3T3-L1 adipocytes, emodin (30 μmol/L) suppressed 11-dehydrocorticosterone-induced adipogenesis without affecting corticosterone-induced adipogenesis; emodin (3 μmol/L) reduced 11-dehydrocorticosterone-stimulated lipolysis, but had no effect on corticosterone-induced lipolysis. Moreover, emodin (3 μmol/L) partly reversed the impaired insulin-stimulated glucose uptake and adiponectin secretion induced by 11-dehydrocorticosterone but not those induced by corticosterone. In ob/ob mice, long-term emodin administration decreased 11β-HSD1 activity in mesenteric adipose tissues, lowered non-fasting and fasting blood glucose levels, and improved glucose tolerance.

Conclusion: Emodin improves the inactive glucocorticoid-induced adipose tissue dysfunction by selective inhibition on 11β-HSD1 in adipocyte in vitro and improves glycemic control in ob/ob mice.

Figure 1

Emodin inhibited 11β-HSD1 activity in 3T3-L1 adipocytes after 1 h (A) or 24 h (B) of treatment. Differentiated 3T3-L1 adipocytes were incubated with the indicated concentrations of emodin or 0.1% DMSO for 1 h or 24 h. 11β-HSD1 enzyme activity was determined by scintillation proximity assay. Each point represents the mean±SEM. n=3.

Figure 2

Emodin regulated adipogenesis and energy metabolism in 3T3-L1 adipocytes. (A) Emodin inhibited the adipogenesis induced by 11-DHC (an inactive glucocorticoid) but not by corticosterone (an active glucocorticoid). The values are expressed as the fold increase over the values for the dexamethasone-treated DMSO group. ^cP<0.01 vs DMSO-treated 11-DHC group; n=4. (B) Emodin inhibited the glycerol release induced by 10 nmol/L 11-DHC but not by 10 nmol/L corticosterone. The values are expressed as the fold increase over the values for the DMSO-treated control group. ^cP<0.01 vs DMSO-treated control group, ^fP<0.01 vs DMSO-treated 11-DHC group; n=3. (C) Emodin significantly reversed the impaired insulin-stimulated glucose uptake induced by 11-DHC but not by corticosterone. ^bP<0.05, ^cP<0.01 vs insulin-stimulated control group, ^fP<0.01 vs insulin stimulated 11-DHC group; n=6. (D) 11-DHC- but not corticosterone-impaired adiponectin release was reversed by 3 μmol/L of emodin in 3T3-L1 adipocytes. Values are expressed as the fold increase over the values for the DMSO-treated control group. ^cP<0.01 vs DMSO-treated control group, ^fP<0.01 vsDMSO-treated 11-DHC group; n=3.

Figure 3

Emodin suppressed 11β-HSD1 activity in the mesenteric fat of ob/obmice. Ob/ob mice were subjected to intraperitoneal injection treatment twice daily with vehicle (0.5% Tween 80), 25 mg/kg emodin or 50 mg/kg emodin for 26 d. 11β-HSD1 activity in the mesenteric fat was measured by SPA at the end of the treatment period. Values are expressed as fold of the values for the vehicle group. ^bP<0.05 vsvehicle group; n=6.

Figure 4

Emodin lowered blood glucose and improved the glucose tolerance of ob/ob mice. ob/ob mice were subjected to intraperitoneal injection twice daily with vehicle (0.5% Tween 80), 25 mg/kg emodin or 50 mg/kg emodin for 26 d. A pair-fed group was set by providing the ob/obmice each day with the amount of food eaten by freely fed 50-mg/kg emodin-treated mice. Random-fed blood glucose concentrations (A) and fasting blood glucose concentrations (B) were measured on day 16 and day 20. Glucose tolerance (C and D) was determined on day 20 of the treatment. Values are expressed as the mean±SEM. ^bP<0.05, ^cP<0.01 vs vehicle group; ^eP<0.05, ^fP<0.01 vs 50-mg/kg emodin pair-fed group; n=8.

Figure 5

Effects of emodin on serum lipids of ob/ob mice. ob/ob mice were subjected to intraperitoneal injection twice daily with vehicle (0.5% Tween 80), 25 mg/kg emodin or 50 mg/kg emodin for 26 d. A pair-fed group was set by providing the ob/obmice each day with the amount of food eaten by freely fed 50-mg/kg emodin-treated mice. Serum triacylglycerol (A), cholesterols (B) and NEFA (C) concentrations were evaluated at the end of the treatment period. Data are expressed as the mean±SEM. ^bP<0.05, ^cP<0.01 vs vehicle group; n=8.

Figure 6

Emodin decreased the food intake (A), body weight gain (B), mesenteric fat weight (C) and perirenal fat weight (D) of ob/ob mice. ob/ob mice were subjected to intraperitoneal injection twice daily with vehicle (0.5% Tween 80), 25 mg/kg emodin or 50 mg/kg emodin for 26 d. A pair-fed group was set by providing the ob/obmice each day with the amount of food eaten by freely fed 50-mg/kg emodin-treated mice. Values are expressed as the mean±SEM. ^cP<0.01 vs vehicle group; n=8.

Figure 7

Emodin increased the mRNA levels of adiponectin (A) and PPARγ (B) in the mesenteric fat of ob/ob mice. ob/ob mice were subjected to intraperitoneal injection twice daily with vehicle (0.5% Tween 80) or emodin (50 mg/kg) for 26 d. The relative mRNA levels of adiponectin and PPARγ in the mesenteric fat were determined by real-time PCR at the end of the treatment period. Values are expressed as the fold increase over the values for the vehicle group. ^cP<0.01 vs vehicle group; n=7.