A novel selective 11b-hydroxysteroid dehydrogenase type 1 inhibitor prevents human adipogenesis.

Glucocorticoid excess increases fat mass, preferentially within omental depots; yet circulating cortisol concentrations are normal in most patients with metabolic syndrome (MS). At a pre-receptor level, 11b-hydroxysteroid dehydrogenase type 1 (11b-HSD1) activates cortisol from cortisone locally within adipose tissue, and inhibition of 11b-HSD1 in liver and adipose tissue has been proposed as a novel therapy to treat MS by reducing hepatic glucose output and adiposity. Using a transformed human subcutaneous preadipocyte cell line (Chub-S7) and human primary preadipocytes, we have defined the role of glucocorticoids and 11b-HSD1 in regulating adipose tissue differentiation. Human cells were differentiated with 1.0 mM cortisol (F), or cortisone (E) with or without 100 nM of a highly selective 11b-HSD1 inhibitor PF-877423. 11b-HSD1 mRNA expression increased across adipocyte differentiation (P!0.001, nZ4), which was paralleled by an increase in 11b-HSD1 oxo-reductase activity (from nil on day 0 to 5.9G1.9 pmol/mg per h on day 16,P!0.01, nZ7). Cortisone enhanced adipocyte differentiation; fatty acid-binding protein 4 expression increased 312-fold (P!0.001) and glycerol-3-phosphate dehydrogenase 47-fold (P!0.001) versus controls. This was abolished by co-incubation with PF-877423. In addition, cellular lipid content decreased significantly. These findings were confirmed in the primary cultures of human subcutaneous preadipocytes. The increase in 11b-HSD1 mRNA expression and activity is essential for the induction of human adipogenesis. Blocking adipogenesis with a novel and specific 11b-HSD1 inhibitor may represent a novel approach to treat obesity in patients with MS

11 beta-hydroxysteroid dehydrogenase type 1 regulates glucocorticoid-induced insulin resistance in skeletal muscle

OBJECTIVE: Glucocorticoid excess is characterized by increased adiposity, skeletal myopathy, and insulin resistance, but the precise molecular mechanisms are unknown. Within skeletal muscle, 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) converts cortisone (11-dehydrocorticosterone in rodents) to active cortisol (corticosterone in rodents). We aimed to determine the mechanisms underpinning glucocorticoid-induced insulin resistance in skeletal muscle and indentify how 11beta-HSD1 inhibitors improve insulin sensitivity. \ud RESEARCH DESIGN AND METHODS: Rodent and human cell cultures, whole-tissue explants, and animal models were used to determine the impact of glucocorticoids and selective 11beta-HSD1 inhibition upon insulin signaling and action. \ud RESULTS: Dexamethasone decreased insulin-stimulated glucose uptake, decreased IRS1 mRNA and protein expression, and increased inactivating pSer$^{307}$ insulin receptor substrate (IRS)-1. 11beta-HSD1 activity and expression were observed in human and rodent myotubes and muscle explants. Activity was predominantly oxo-reductase, generating active glucocorticoid. A1 (selective 11beta-HSD1 inhibitor) abolished enzyme activity and blocked the increase in pSer$^{307}$ IRS1 and reduction in total IRS1 protein after treatment with 11DHC but not corticosterone. In C57Bl6/J mice, the selective 11beta-HSD1 inhibitor, A2, decreased fasting blood glucose levels and improved insulin sensitivity. In KK mice treated with A2, skeletal muscle pSer$^{307}$ IRS1 decreased and pThr$^{308}$ Akt/PKB increased. In addition, A2 decreased both lipogenic and lipolytic gene expression.\ud CONCLUSIONS: Prereceptor facilitation of glucocorticoid action via 11beta-HSD1 increases pSer$^{307}$ IRS1 and may be crucial in mediating insulin resistance in skeletal muscle. Selective 11beta-HSD1 inhibition decreases pSer$^{307}$ IRS1, increases pThr$^{308}$ Akt/PKB, and decreases lipogenic and lipolytic gene expression that may represent an important mechanism underpinning their insulin-sensitizing action

Short- and long-term glucocorticoid treatment enhances insulin signalling in human subcutaneous adipose tissue

BACKGROUND: Endogenous or exogenous glucocorticoid (GC) excess (Cushing's syndrome) is characterized by increased adiposity and insulin resistance. Although GCs cause global insulin resistance in vivo, we have previously shown that GCs are able to augment insulin action in human adipose tissue, contrasting with their action in skeletal muscle. Cushing's syndrome develops following chronic GC exposure and, in addition, is a state of hyperinsulinemia. OBJECTIVES: We have therefore compared the impact of short- (24 h) and long-term (7 days) GC administration on insulin signalling in differentiated human adipocytes in the presence of low or high concentrations of insulin. RESULTS: Both short- (24 h) and long-term (7 days) treatment of chub-s7 cells with dexamethasone (Dex) (0.5 μM) increased insulin-stimulated pTyr612IRS1 and pSer473akt/PKB, consistent with insulin sensitization. Chronic high-dose insulin treatment induced insulin resistance in chub-s7 cells. However, treatment with both high-dose insulin and Dex in combination still caused insulin sensitization. CONCLUSIONS: In this human subcutaneous adipocyte cell line, prolonged GC exposure, even in the presence of high insulin concentrations, is able to cause insulin sensitization. We suggest that this is an important mechanism driving adipogenesis and contributes to the obese phenotype of patients with Cushing's syndrome

11 beta-hydroxysteroid dehydrogenase type 1 regulates glucocorticoid-induced insulin resistance in skeletal muscle

OBJECTIVE: Glucocorticoid excess is characterized by increased adiposity, skeletal myopathy, and insulin resistance, but the precise molecular mechanisms are unknown. Within skeletal muscle, 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) converts cortisone (11-dehydrocorticosterone in rodents) to active cortisol (corticosterone in rodents). We aimed to determine the mechanisms underpinning glucocorticoid-induced insulin resistance in skeletal muscle and indentify how 11beta-HSD1 inhibitors improve insulin sensitivity. \ud RESEARCH DESIGN AND METHODS: Rodent and human cell cultures, whole-tissue explants, and animal models were used to determine the impact of glucocorticoids and selective 11beta-HSD1 inhibition upon insulin signaling and action. \ud RESULTS: Dexamethasone decreased insulin-stimulated glucose uptake, decreased IRS1 mRNA and protein expression, and increased inactivating pSer$^{307}$ insulin receptor substrate (IRS)-1. 11beta-HSD1 activity and expression were observed in human and rodent myotubes and muscle explants. Activity was predominantly oxo-reductase, generating active glucocorticoid. A1 (selective 11beta-HSD1 inhibitor) abolished enzyme activity and blocked the increase in pSer$^{307}$ IRS1 and reduction in total IRS1 protein after treatment with 11DHC but not corticosterone. In C57Bl6/J mice, the selective 11beta-HSD1 inhibitor, A2, decreased fasting blood glucose levels and improved insulin sensitivity. In KK mice treated with A2, skeletal muscle pSer$^{307}$ IRS1 decreased and pThr$^{308}$ Akt/PKB increased. In addition, A2 decreased both lipogenic and lipolytic gene expression.\ud CONCLUSIONS: Prereceptor facilitation of glucocorticoid action via 11beta-HSD1 increases pSer$^{307}$ IRS1 and may be crucial in mediating insulin resistance in skeletal muscle. Selective 11beta-HSD1 inhibition decreases pSer$^{307}$ IRS1, increases pThr$^{308}$ Akt/PKB, and decreases lipogenic and lipolytic gene expression that may represent an important mechanism underpinning their insulin-sensitizing action

A novel selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor prevents human adipogenesis

Glucocorticoid excess increases fat mass, preferentially within omental depots; yet circulating cortisol concentrations are normal in most patients with metabolic syndrome (MS). At a pre-receptor level, 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) activates cortisol from cortisone locally within adipose tissue, and inhibition of 11β-HSD1 in liver and adipose tissue has been proposed as a novel therapy to treat MS by reducing hepatic glucose output and adiposity. Using a transformed human subcutaneous preadipocyte cell line (Chub-S7) and human primary preadipocytes, we have defined the role of glucocorticoids and 11β-HSD1 in regulating adipose tissue differentiation. Human cells were differentiated with 1·0 μM cortisol (F), or cortisone (E) with or without 100 nM of a highly selective 11β-HSD1 inhibitor PF-877423. 11β-HSD1 mRNA expression increased across adipocyte differentiation (P<0·001, n=4), which was paralleled by an increase in 11β-HSD1 oxo-reductase activity (from nil on day 0 to 5·9±1.9 pmol/mg per h on day 16, P<0·01, n=7). Cortisone enhanced adipocyte differentiation; fatty acid-binding protein 4 expression increased 312-fold (P<0·001) and glycerol-3-phosphate dehydrogenase 47-fold (P<0·001) versus controls. This was abolished by co-incubation with PF-877423. In addition, cellular lipid content decreased significantly. These findings were confirmed in the primary cultures of human subcutaneous preadipocytes. The increase in 11β-HSD1 mRNA expression and activity is essential for the induction of human adipogenesis. Blocking adipogenesis with a novel and specific 11β-HSD1 inhibitor may represent a novel approach to treat obesity in patients with MS

Characterisation of 11β-hydroxysteroid dehydrogenase 1 in human orbital adipose tissue: a comparison with subcutaneous and omental fat

Glucocorticoids (GCs) have a profound effect on adipose biology increasing tissue mass causing central obesity. The pre-receptor regulation of GCs by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) that activates cortisol from cortisone has been postulated as a fundamental mechanism underlying the metabolic syndrome mediating adipocyte hyperplasia and hypertrophy in the omental (OM) depot. Orbital adipose tissue (OF) is the site of intense inflammation and tissue remodelling in several orbital inflammatory disease states. In this study, we describe features of the GC metabolic pathways in normal human OF depot and compare it with subcutaneous (SC) and OM depots. Using an automated histological characterisation technique, OF adipocytes were found to be significantly smaller (parameters: area, maximum diameter and perimeter) than OM and SC adipocytes (P<0·001). Although immunohistochemical analyses demonstrated resident CD68+ cells in all three whole tissue adipose depots, OF CD68 mRNA and protein expression exceeded that of OM and SC (mRNA, P<0·05; protein, P<0·001). In addition, there was higher expression of glucocorticoid receptor (GR)α mRNA in the OF whole tissue depot (P<0·05). Conversely, 11β-HSD1 mRNA together with the markers of late adipocyte differentiation (FABP4 and G3PDH) were significantly lower in OF. Primary cultures of OF preadipocytes demonstrated predominant 11β-HSD1 oxo-reductase activity with minimal dehydrogenase activity. Orbital adipocytes are smaller, less differentiated, and express low levels of 11β-HSD1 but abundant GRα compared with SC and OM. OF harbours a large CD68+ population. These characteristics define an orbital microenvironment that has the potential to respond to sight-threatening orbital inflammatory disease

Regulation of Lipogenesis by Glucocorticoids and Insulin in Human Adipose Tissue

Patients with glucocorticoid (GC) excess, Cushing's syndrome, develop a classic phenotype characterized by central obesity and insulin resistance. GCs are known to increase the release of fatty acids from adipose, by stimulating lipolysis, however, the impact of GCs on the processes that regulate lipid accumulation has not been explored. Intracellular levels of active GC are dependent upon the activity of 11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) and we have hypothesized that 11β-HSD1 activity can regulate lipid homeostasis in human adipose tissue (Chub-S7 cell line and primary cultures of human subcutaneous (sc) and omental (om) adipocytes. Across adipocyte differentiation, lipogenesis increased whilst β-oxidation decreased. GC treatment decreased lipogenesis but did not alter rates of β-oxidation in Chub-S7 cells, whilst insulin increased lipogenesis in all adipocyte cell models. Low dose Dexamethasone pre-treatment (5 nM) of Chub-S7 cells augmented the ability of insulin to stimulate lipogenesis and there was no evidence of adipose tissue insulin resistance in primary sc cells. Both cortisol and cortisone decreased lipogenesis; selective 11β-HSD1 inhibition completely abolished cortisone-mediated repression of lipogenesis. GCs have potent actions upon lipid homeostasis and these effects are dependent upon interactions with insulin. These in vitro data suggest that manipulation of GC availability through selective 11β-HSD1 inhibition modifies lipid homeostasis in human adipocytes

Advantage of rare infanticide strategies in an invasion experiment of behavioural polymorphism

Killing conspecific infants (infanticide) is among the most puzzling phenomena in nature. Stable polymorphism in such behaviour could be maintained by negative frequency-dependent selection (benefit of rare types). However, it is currently unknown whether there is genetic polymorphism in infanticidal behaviour or whether infanticide may have any fitness advantages when rare. Here we show genetic polymorphism in non-parental infanticide. Our novel invasion experiment confirms negative frequency-dependent selection in wild bank vole populations, where resource benefits allow an infanticidal strategy to invade a population of non-infanticidal individuals. The results show that infanticidal behaviour is highly heritable with genetic correlation across the sexes. Thus, a positive correlative response in male behaviour is expected when selection operates on females only and vice versa. Our results, on one hand, demonstrate potential benefits of infanticide, and on the other, they open a new perspective of correlative evolution of infanticide in females and males

A Switch in Hepatic Cortisol Metabolism across the Spectrum of Non Alcoholic Fatty Liver Disease

Context: Non alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome. NAFLD represents a spectrum of liver disease ranging from reversible hepatic steatosis, to non alcoholic steato-hepatitis (NASH) and cirrhosis. The potential role of glucocorticoids (GC) in the pathogenesis of NAFLD is highlighted in patients with GC excess, Cushing's syndrome, who develop central adiposity, insulin resistance and in 20% of cases, NAFLD. Although in most cases of NAFLD, circulating cortisol levels are normal, hepatic cortisol availability is controlled by enzymes that regenerate cortisol (F) from inactive cortisone (E) (11β-hydroxysteroid dehydrogenase type 1, 11β-HSD1), or inactivate cortisol through A-ring metabolism (5α- and 5β-reductase, 5αR and 5βR). Objective and Methods: In vitro studies defined 11β-HSD1 expression in normal and NASH liver samples. We then characterised hepatic cortisol metabolism in 16 patients with histologically proven NAFLD compared to 32 obese controls using gas chromatographic analysis of 24 hour urine collection and plasma cortisol generation profile following oral cortisone. Results: In patients with steatosis 5αR activity was increased, with a decrease in hepatic 11β-HSD1 activity. Total cortisol metabolites were increased in this group consistent with increased GC production rate. In contrast, in patients with NASH, 11β-HSD1 activity was increased both in comparison to patients with steatosis, and controls. Endorsing these findings, 11β-HSD1 mRNA and immunostaining was markedly increased in NASH patients in peri septal hepatocytes and within CD68 positive macrophages within inflamed cirrhotic septa. Conclusion: Patients with hepatic steatosis have increased clearance and decreased hepatic regeneration of cortisol and we propose that this may represent a protective mechanism to decrease local GC availability to preserve hepatic metabolic phenotype. With progression to NASH, increased 11β-HSD1 activity and consequent cortisol regeneration may serve to limit hepatic inflammation

Hexose-6-phosphate Dehydrogenase Modulates 11β-Hydroxysteroid Dehydrogenase Type 1-Dependent Metabolism of 7-keto- and 7β-hydroxy-neurosteroids

BACKGROUND: The role of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) in the regulation of energy metabolism and immune system by locally reactivating glucocorticoids has been extensively studied. Experiments determining initial rates of enzyme activity revealed that 11beta-HSD1 can catalyze both the reductase and the dehydrogenase reaction in cell lysates, whereas it predominantly catalyzes the reduction of cortisone to cortisol in intact cells that also express hexose-6-phosphate dehydrogenase (H6PDH), which provides cofactor NADPH. Besides its role in glucocorticoid metabolism, there is evidence that 11beta-HSD1 is involved in the metabolism of 7-keto- and 7-hydroxy-steroids; however the impact of H6PDH on this alternative function of 11beta-HSD1 has not been assessed. METHODOLOGY: We investigated the 11beta-HSD1-dependent metabolism of the neurosteroids 7-keto-, 7alpha-hydroxy- and 7beta-hydroxy-dehydroepiandrosterone (DHEA) and 7-keto- and 7beta-hydroxy-pregnenolone, respectively, in the absence or presence of H6PDH in intact cells. 3D-structural modeling was applied to study the binding of ligands in 11beta-HSD1. PRINCIPAL FINDINGS: We demonstrated that 11beta-HSD1 functions in a reversible way and efficiently catalyzed the interconversion of these 7-keto- and 7-hydroxy-neurosteroids in intact cells. In the presence of H6PDH, 11beta-HSD1 predominantly converted 7-keto-DHEA and 7-ketopregnenolone into their corresponding 7beta-hydroxy metabolites, indicating a role for H6PDH and 11beta-HSD1 in the local generation of 7beta-hydroxy-neurosteroids. 3D-structural modeling offered an explanation for the preferred formation of 7beta-hydroxy-neurosteroids. CONCLUSIONS: Our results from experiments determining the steady state concentrations of glucocorticoids or 7-oxygenated neurosteroids suggested that the equilibrium between cortisone and cortisol and between 7-keto- and 7-hydroxy-neurosteroids is regulated by 11beta-HSD1 and greatly depends on the coexpression with H6PDH. Thus, the impact of H6PDH on 11beta-HSD1 activity has to be considered for understanding both glucocorticoid and neurosteroid action in different tissues