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In peripheral target tissues, levels of active glucocorticoid hormones are controlled by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), a dimeric enzyme that catalyzes the reduction of cortisone to cortisol within the endoplasmic reticulum. Loss of this activity results in a disorder termed cortisone reductase deficiency (CRD), typified by increased cortisol clearance and androgen excess. To date, only mutations in H6PD, which encodes an enzyme supplying cofactor for the reaction, have been identified as the cause of disease. Here we examined the HSD11B1 gene in two cases presenting with biochemical features indicative of a milder form of CRD in whom the H6PD gene was normal. Novel heterozygous mutations (R137C or K187N) were found in the coding sequence of HSD11B1. The R137C mutation disrupts salt bridges at the subunit interface of the 11β-HSD1 dimer, whereas K187N affects a key active site residue. On expression of the mutants in bacterial and mammalian cells, activity was either abolished (K187N) or greatly reduced (R137C). Expression of either mutant in a bacterial system greatly reduced the yield of soluble protein, suggesting that both mutations interfere with subunit folding or dimer assembly. Simultaneous expression of mutant and WT 11β-HSD1 in bacterial or mammalian cells, to simulate the heterozygous condition, indicated a marked suppressive effect of the mutants on both the yield and activity of 11β-HSD1 dimers. Thus, these heterozygous mutations in the HSD11B1 gene have a dominant negative effect on the formation of functional dimers and explain the genetic cause of CRD in these patients.

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.

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.

Background: Toll-like receptor-4 (TLR4) has been implicated in the pathogenesis of different renal diseases in rodent models. However, in human kidney disease, TLR4 expression and regulation is not well understood. We hypothesized that renal TLR4 expression plays a role in chronic kidney disease (CKD) and is associated with proteinuria, kidney function, histological diagnosis, and inflammatory mediators., Methods: We quantified TLR4 mRNA as well as expression of macrophage chemoattractant protein-1 (MCP-1), transforming growth factor-β₁ (TGF-β₁) and interleukin-6 (IL-6) in human kidney biopsies from 70 patients with CKD. We measured kidney function, urinary MCP-1 protein excretion, and the amount of chronic damage. Macrophage load was quantified by CD68 and vascularization by CD34 immunostaining., Results: TLR4 expression correlated significantly with MCP-1 and TGF-β₁ expression. High TLR4 expression was associated with high IL-6 expression. TLR4 expression was significantly upregulated in patients with severe proteinuria, and in patients with chronic ischemic renal damage and IgA nephropathy, when compared to patients with thin glomerular basement membrane disease. TLR4 expression did not correlate with creatinine clearance, renal outcome, macrophage load or chronic damage., Conclusions: We show for the first time that renal TLR4 expression was significantly associated with the pro-inflammatory marker MCP-1 and the profibrotic molecule TGF-β₁ in kidney biopsies from patients with CKD, suggesting that increased expression of TLR4 is an important feature of CKD., (2011 S. Karger AG, Basel.)

Objective: The combined thickness of the intima and media of the carotid artery (carotid intima-medial thickness, CIMT) is associated with cardiovascular disease and stroke. Previous studies indicate that carotid intima-medial thickness is a significantly heritable phenotype, but the responsible genes are largely unknown. Hexose-6 phosphate dehydrogenase (H6PDH) is a microsomal enzyme whose activity regulates corticosteroid metabolism in the liver and adipose tissue; variability in measures of corticosteroid metabolism within the normal range have been associated with risk factors for cardiovascular disease. We performed a genetic association study in 854 members of 224 families to assess the relationship between polymorphisms in the gene coding for hexose-6 phosphate dehydrogenase (H6PD) and carotid intima-medial thickness., Methods: Families were ascertained via a hypertensive proband. CIMT was measured using B-mode ultrasound. Single nucleotide polymorphisms (SNPs) tagging common variation in the H6PD gene were genotyped. Association was assessed following adjustment for significant covariates including "classical" cardiovascular risk factors. Functional studies to determine the effect of particular SNPs on H6PDH were performed., Results: There was evidence of association between the single nucleotide polymorphism rs17368528 in exon five of the H6PD gene, which encodes an amino-acid change from proline to leucine in the H6PDH protein, and mean carotid intima-medial thickness (p = 0.00065). Genotype was associated with a 5% (or 0.04 mm) higher mean carotid intima-medial thickness measurement per allele, and determined 2% of the population variability in the phenotype., Conclusions: Our results suggest a novel role for the H6PD gene in atherosclerosis susceptibility.

Glucose-6-phosphate (G6P) metabolism by the enzyme hexose-6-phosphate dehydrogenase (H6PDH) within the sarcoplasmic reticulum lumen generates nicotinamide adenine dinucleotide phosphate (reduced) to provide the redox potential for the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) to activate glucocorticoid (GC). H6PDH knockout (KO) mice have a switch in 11β-HSD1 activity, resulting in GC inactivation and hypothalamic-pituitary-adrenal axis activation. Importantly, H6PDHKO mice develop a type II fiber myopathy with abnormalities in glucose metabolism and activation of the unfolded protein response (UPR). GCs play important roles in muscle physiology, and therefore, we have examined the importance of 11β-HSD1 and GC metabolism in mediating aspects of the H6PDHKO myopathy. To achieve this, we examined 11β-HSD1/H6PDH double-KO (DKO) mice, in which 11β-HSD1 mediated GC inactivation is negated. In contrast to H6PDHKO mice, DKO mice GC metabolism and hypothalamic-pituitary-adrenal axis set point is similar to that observed in 11β-HSD1KO mice. Critically, in contrast to 11β-HSD1KO mice, DKO mice phenocopy the salient features of the H6PDHKO, displaying reduced body mass, muscle atrophy, and vacuolation of type II fiber-rich muscle, fasting hypoglycemia, increased muscle glycogen deposition, and elevated expression of UPR genes. We propose that muscle G6P metabolism through H6PDH may be as important as changes in the redox environment when considering the mechanism underlying the activation of the UPR and the ensuing myopathy in H6PDHKO and DKO mice. These data are consistent with an 11β-HSD1-independent function for H6PDH in which sarcoplasmic reticulum G6P metabolism and nicotinamide adenine dinucleotide phosphate-(oxidized)/nicotinamide adenine dinucleotide phosphate (reduced) redox status are important for maintaining muscle homeostasis.

Glucocorticoids (GCs) are highly detrimental to skin integrity and function both when applied topically for anti-inflammatory treatments and during conditions of circulating excess, e.g., Cushing's syndrome. Within target tissues, GC availability is regulated at a prereceptor level, independently of systemic levels, by isozymes of 11β-hydroxysteroid dehydrogenase (11β-HSD) that interconvert active cortisol and inactive cortisone. Many of the adverse effects of GCs on skin are also reminiscent of the natural aging process. 11β-HSD1 (which activates cortisol), but not 11β-HSD2 (which inactivates cortisol), was expressed in epidermal keratinocytes and dermal fibroblasts in human skin and also in outer hair follicle root sheath cells in murine skin. 11β-HSD1 activity was present ex vivo in both species and increased with age in human skin tissue explants. In primary human dermal fibroblasts (HDF) from both photoprotected and photoexposed sites, 11β-HSD1 also increased with donor age. Additionally, photoexposed HDF displayed higher 11β-HSD1 mRNA expression than donor-matched photoprotected HDF. GC treatment of HDF caused upregulation of 11β-HSD1 mRNA levels independent of donor age or site. The age- and site-associated increase in dermal 11β-HSD1, and the ensuing increased local GC activation, may contribute to the adverse changes in skin morphology and function associated with chronological aging and photoaging.

The prevalence of primary hyperaldosteronism approaches 10% of all hypertensive patients, and besides efficient diagnostic procedures, effective treatment is of increasing importance to reverse increased morbidity and mortality. Aldosterone-producing adenoma and unilateral adrenal hyperplasia are amenable to cure by endoscopic adrenalectomy. Bilateral adrenal hyperplasia (micro- or macronodular), which comprises two-thirds of primary hyperaldosteronism, is treated primarily by mineralocorticoid receptor antagonists (starting dose 12.5-25mg/day spironolactone with titration up to 100mg/day, alternatively 50-100mg/day eplerenone). If blood pressure is not normalised by this first-line treatment, additional treatment with potassium-sparing diuretics (amiloride or triamterene) or calcium channel antagonists is necessary. The start of medication should be closely monitored by serum electrolyte and creatinine controls., (2010 Elsevier Ltd. All rights reserved.)

Context: The etiology of idiopathic intracranial hypertension (IIH) is unknown. We hypothesized that obesity and elevated intracranial pressure may be linked through increased 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) activity., Objective: The aim was to characterize 11β-HSD1 in human cerebrospinal fluid (CSF) secretory [choroid plexus (CP)] and drainage [arachnoid granulation tissue (AGT)] structures, and to evaluate 11β-HSD1 activity after therapeutic weight loss in IIH., Design and Setting: We conducted in vitro analysis of CP and AGT and a prospective in vivo cohort study set in two tertiary care centers., Patients or Other Participants: Twenty-five obese adult female patients with active IIH were studied, and 22 completed the study., Intervention: Fasted serum, CSF, and 24-h urine samples were collected at baseline, after 3-month observation, and after a 3-month diet., Main Outcome Measures: Changes in urine, serum, and CSF glucocorticoids (measured by gas chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry) after weight loss were measured., Results: 11β-HSD1 and key elements of the glucocorticoid signaling pathway were expressed in CP and AGT. After weight loss (14.2±7.8 kg; P<0.001), global 11β-HSD1 activity decreased (P=0.001) and correlated with reduction in intracranial pressure (r=0.504; P=0.028). CSF and serum glucocorticoids remained stable, although the change in CSF cortisone levels correlated with weight loss (r=-0.512; P=0.018)., Conclusions: Therapeutic weight loss in IIH is associated with a reduction in global 11β-HSD1 activity. Elevated 11β-HSD1 may represent a pathogenic mechanism in IIH, potentially via manipulation of CSF dynamics at the CP and AGT. Although further clarification of the functional role of 11β-HSD1 in IIH is needed, our results suggest that 11β-HSD1 inhibition may have therapeutic potential in IIH.

Patients with early chronic kidney disease (CKD) have an increased risk for cardiovascular disease. Aldosterone levels are elevated and might impair ventricular function through adverse myocardial and vascular proinflammatory and fibrotic effects. In the Chronic Renal Impairment in Birmingham II (CRIB II) study, it was hypothesized that mineralocorticoid receptor blockade with spironolactone in addition to angiotensin-converting enzyme inhibitors or angiotensin receptor blockers would improve left ventricular (LV) function and markers of inflammation, ventricular stretch, and collagen turnover in early CKD. A total of 112 patients with early CKD were randomized to spironolactone 25 mg/day or placebo for 40 weeks. Ventricular function was assessed by echocardiography and cardiac magnetic resonance imaging tagging. High-sensitivity C-reactive protein, N-terminal-pro-B-type natriuretic peptide, and aminoterminal propeptide of type III procollagen were measured. Spironolactone improved LV long-axis systolic function (Sm 8.2 ± 1.4 vs 7.7 ± 1.3 cm/s, p <0.05), torsion (7.77 ± 1.61° vs 6.77 ± 1.48°, p <0.05), and myocardial deformation (strain rate -1.14 ± 0.24 vs -1.09 ± 0.20 s(-1), p <0.05) compared to placebo, without a change in the ejection fraction. Markers of LV relaxation (E/e' ratio 7.2 ± 2.3 vs 8.5 ± 2.3, p <0.05) and suction (M-mode propagation velocity 56 ± 12 vs 50 ± 12 cm/s, p <0.05) were also improved. Spironolactone reduced N-terminal-pro-B-type natriuretic peptide (24.8 pmol/L [range 0.4 to 122.4] vs 39.4 pmol/L [range 10.8 to 102.4], p <0.01) and attenuated an increase in aminoterminal propeptide of type III procollagen observed with placebo. In conclusion, spironolactone improves markers of regional LV systolic and diastolic function in early CKD., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Context: Aldosterone has emerged as an important mediator of disease progression and mortality in patients with chronic heart and kidney disease (CKD). Despite the increasing use of mineralocorticoid receptor antagonists in these patients, little is known about the effects on corticosteroid hormone secretion and metabolism., Objective: To assess corticosteroid hormone secretion and metabolism in patients with early stage CKD before and after spironolactone (Spiro)., Design: Randomized, double-blind, placebo-controlled interventional study., Setting: Single tertiary referral centre., Patients: A total of 112 patients with stable stage 2/3 CKD., Interventions: Patients were randomized to receive either Spiro 25 mg once daily or placebo for 36 weeks., Main Outcome Measures: Plasma renin activity (PRA), angiotensin II (AngII) and steroid hormones were analysed by standard assays; urinary corticosteroid hormone metabolites (5α+5β-tetrahydro-cortisol (5α+5β-THF), TH-cortisone (THE), 3α5β-TH-aldosterone (TH-Aldo), 5α+5β-TH-deoxycorticosterone (5α+5β-TH-DOC), TH-11-desoxycortisol (THS)) were analysed by gas chromatography/mass spectrometry., Results: Plasma aldosterone concentration (PAC) was inversely correlated with eGFR (r = -0·331, P < 0·001). Urinary 24-h excretion of TH-Aldo was correlated with PAC (r = 0·214, P < 0·05) and diastolic blood pressure (BP) (r = 0·212, P = <0·05), whereas total 24-h urinary cortisol metabolite excretion was correlated with systolic BP (r = 0·316, P < 0·01). In addition, 11β-hydroxysteroid dehydrogenase (11β-HSD) type 1 activity (urinary 5α+5β-THF)/THE) ratio) was correlated with PRA (r = 0·277, P < 0·01). Spiro treatment significantly reduced BP (123 ± 11/76 ± 7 vs 119 ± 11/73 ± 8 mmHg, P < 0·01) despite renin-angiotensin-aldosterone system induction, reflected by increased urinary 24-h TH-Aldo excretion (17·6 (12, 86) vs 26 (18, 80) μg/24 h, P < 0·05). By contrast, Spiro had no effect on total urinary cortisol metabolite excretion, 11β-hydroxylase, 11β-HSD type 1 and 2 activity., Conclusions: Aldo and cortisol are positively associated with BP suggesting that adrenal hyperactivity may in part explain the increased cardiovascular risk in patients with early end-stage CKD. Addition of Spiro had no effect on glucocorticoid metabolism or total 24-h corticosteroid production., (© 2010 Blackwell Publishing Ltd.)

Aim: Peroxisome proliferator-activated receptor gamma (PPARγ) is generally accepted as renoprotective factor in type 2 diabetes mellitus, and PPARγ agonists have been reported to reduce albuminuria. However, little is known about renal PPARγ expression in chronic kidney disease, and especially human data are scarce. We hypothesized that renal PPARγ expression is associated with extent of proteinuria, kidney function, histological diagnosis and inflammatory mediators. Therefore, we investigated PPARγ mRNA expression in human kidney biopsies., Methods: We quantified PPARγ mRNA as well as the expression of macrophage chemoattractant protein-1, transforming growth factor beta-1 and interleukin-6 in 64 human kidney biopsies from patients with chronic kidney disease and mild-to-marked proteinuria of diverse aetiology. We measured renal function, and macrophage invasion was quantified by CD68 and vascularization by CD34 immunostaining., Results: PPARγ mRNA expression correlated inversely with renal function. Higher blood pressure levels were associated with higher PPARγ expression levels. PPARγ mRNA expression correlated significantly (P<0.001) with macrophage chemoattractant protein-1 mRNA expression and showed a negative trend with transforming growth factor beta-1 mRNA expression. No differences in PPARγ expression were detected with regard to extent of proteinuria, histological diagnosis, macrophage invasion, interleukin-6 expression, and age or body mass index., Conclusions: PPARγ expression increases with loss of renal function and may be an important factor in maintaining normal renal function serving as a key protective mechanism to renal injury., (© 2010 The Authors. Nephrology © 2010 Asian Pacific Society of Nephrology.)

Due to its impact upon health and the economy, the mechanisms that contribute to the pathogenesis of obesity and the metabolic syndrome are under intense scrutiny. In addition to understanding the pathogenesis of disease it is important to design and trial novel therapies. Patients with cortisol excess, Cushing's syndrome, have a phenotype similar to that of the metabolic syndrome and as a result there is much interest the manipulation of glucocorticoid (GC) action as a therapeutic strategy. Intracellular GC levels are regulated by 11β-hydroxysteroid dehydrogenase (11β-HSD1) which converts inactive cortisone to cortisol, thereby increasing local GC action. There is an abundance of data implicating 11β-HSD1 in the pathogenesis of obesity, type 2 diabetes and the metabolic syndrome and 11β-HSD1 is an attractive therapeutic target. Selective 11β-HSD1 inhibitors, which do not act upon 11β-HSD2 (which inactivates cortisol to cortisone) are in development. So far studies have primarily been carried out in rodents, with results showing improvements in metabolic profile. Data are now beginning to emerge from human studies and the results are promising., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Liquid chromatography tandem mass spectrometry (LC/MS/MS) is replacing classical methods for steroid hormone analysis. It requires small sample volumes and has given rise to improved specificity and short analysis times. Its growth has been fueled by criticism of the validity of steroid analysis by older techniques, testosterone measurements being a prime example. While this approach is the gold-standard for measurement of individual steroids, and panels of such compounds, LC/MS/MS is of limited use in defining novel metabolomes. GC/MS, in contrast, is unsuited to rapid high-sensitivity analysis of specific compounds, but remains the most powerful discovery tool for defining steroid disorder metabolomes. Since the 1930s almost all inborn errors in steroidogenesis have been first defined through their urinary steroid excretion. In the last 30 years, this has been exclusively carried out by GC/MS and has defined conditions such as AME syndrome, glucocorticoid remediable aldosteronism (GRA) and Smith-Lemli-Opitz syndrome. Our recent foci have been on P450 oxidoreductase deficiency (ORD) and apparent cortisone reductase deficiency (ACRD). In contrast to LC/MS/MS methodology, a particular benefit of GC/MS is its non-selective nature; a scanned run will contain every steroid excreted, providing an integrated picture of an individual's metabolome. The "Achilles heel" of clinical GC/MS profiling may be data presentation. There is lack of familiarity with the multiple hormone metabolites excreted and diagnostic data are difficult for endocrinologists to comprehend. While several conditions are defined by the absolute concentration of steroid metabolites, many are readily diagnosed by ratios between steroid metabolites (precursor metabolite/product metabolite). Our work has led us to develop a simplified graphical representation of quantitative urinary steroid hormone profiles and diagnostic ratios., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Objective: To observe intracranial pressure in women with idiopathic intracranial hypertension who follow a low energy diet., Design: Prospective cohort study., Setting: Outpatient department and the clinical research facility based at two separate hospitals within the United Kingdom., Participants: 25 women with body mass index (BMI) >25, with active (papilloedema and intracranial pressure >25 cm H(2)O), chronic (over three months) idiopathic intracranial hypertension. Women who had undergone surgery to treat idiopathic intracranial hypertension were excluded., Intervention: Stage 1: no new intervention; stage 2: nutritionally complete low energy (calorie) diet (1777 kJ/day (425 kcal/day)); stage 3: follow-up period after the diet. Each stage lasted three months., Main Outcome Measure: The primary outcome was reduction in intracranial pressure after the diet. Secondary measures included score on headache impact test-6, papilloedema (as measured by ultrasonography of the elevation of the optic disc and diameter of the nerve sheath, together with thickness of the peripapillary retina measured by optical coherence tomography), mean deviation of Humphrey visual field, LogMAR visual acuity, and symptoms. Outcome measures were assessed at baseline and three, six, and nine months. Lumbar puncture, to quantify intracranial pressure, was measured at baseline and three and six months., Results: All variables remained stable over stage 1. During stage 2, there were significant reductions in weight (mean 15.7 (SD 8.0) kg, P<0.001), intracranial pressure (mean 8.0 (SD 4.2) cm H(2)O, P<0.001), score on headache impact test (7.6 (SD 10.1), P=0.004), and papilloedema (optic disc elevation (mean 0.15 (SD 0.23) mm, P=0.002), diameter of the nerve sheath (mean 0.7 (SD 0.8) mm, P=0.004), and thickness of the peripapillary retina (mean 25.7 (SD 36.1) micro, P=0.001)). Mean deviation of the Humphrey visual field remained stable, and in only five patients, the LogMAR visual acuity improved by one line. Fewer women reported symptoms including tinnitus, diplopia, and obscurations (10 v 4, P=0.004; 7 v 0, P=0.008; and 4 v 0, P=0.025, respectively). Re-evaluation at three months after the diet showed no significant change in weight (0.21 (SD 6.8) kg), and all outcome measures were maintained., Conclusion: Women with idiopathic intracranial hypertension who followed a low energy diet for three months had significantly reduced intracranial pressure compared with pressure measured in the three months before the diet, as well as improved symptoms and reduced papilloedema. These reductions persisted for three months after they stopped the diet.

Pituitary disease is associated with increased mortality predominantly due to vascular disease. Control of cortisol secretion and GH hypersecretion (and cardiovascular risk factor reduction) is key in the reduction of mortality in patients with Cushing's disease and acromegaly, retrospectively. For patients with acromegaly, the role of IGF-I is less clear-cut. Confounding pituitary hormone deficiencies such as gonadotropins and particularly ACTH deficiency (with higher doses of hydrocortisone replacement) may have a detrimental effect on outcome in patients with pituitary disease. Pituitary radiotherapy is a further factor that has been associated with increased mortality (particularly cerebrovascular). Although standardized mortality ratios in pituitary disease are falling due to improved treatment, mortality for many conditions are still elevated above that of the general population, and therefore further measures are needed. Craniopharyngioma patients have a particularly increased risk of mortality as a result of the tumor itself and treatment to control tumor growth; this is a key area for future research in order to optimize the outcome for these patients.

Background: Insulin-like growth factor I (IGF-1) is for the most part bound in a ternary complex with IGF-binding protein-3 (IGFBP-3) and acid-labile subunit (ALS). This ternary complex is a storage form of IGF-1 in blood and passes not through the renal glomerulus. Little information is available in regard to the components of the ternary complex in adult renal disease., Objective: To investigate levels of serum IGF-1, IGFBP-3 and ALS in relation to renal function and extent of proteinuria., Design and Patients: We measured IGF-1, IGFBP-3 and ALS concentrations in 137 patients who were investigated due to proteinuria and/or haematuria and/or renal impairment. The patients received renal biopsies and the histological diagnosis was documented. Urinary albumin excretion and relevant clinical parameter were evaluated., Results: IGF-1 showed a highly positive correlation to IGFBP-3 and ALS, and the latter to IGFBP-3. IGF-1, IGFBP-3 and ALS decreased with increasing age. IGF-1 and IGFBP-3 showed no significant change depending on the creatinine clearance. However, ALS decreased with decreasing renal function. In patients with heavy proteinuria ALS levels, but not IGF-1 and IGFBP-3 levels, decreased significantly. Patients with chronic ischaemic renal damage and diabetic glomerulopathy showed higher IGF-1 and IGFBP-3 levels compared to patients with thin glomerular basement membrane disease despite their older age., Conclusions: IGF-1 and IGFBP-3 levels seem to be independent of renal function and severity of proteinuria. However, ALS levels are altered in renal failure and nephrotic syndrome, which may be due to increased renal loss or diminished hepatic production or both., (Copyright (c) 2009 Elsevier Ltd. All rights reserved.)

We sought to evaluate the diagnostic accuracy of metabolomic biomarker profiles in neurological conditions (idiopathic intracranial hypertension (IIH), multiple sclerosis (MS) and cerebrovascular disease (CVD) compared to controls with either no neurological disease or mixed neurological diseases). Spectra of CSF (n = 87) and serum (n = 72) were acquired using (1)H NMR spectroscopy. Multivariate pattern recognition analysis was used to identify disease-specific metabolite biomarker profiles. The metabolite profiles were then used to predict the diagnosis of a second cohort of patients (n = 25). CSF metabolite profiles were able to predict diagnosis with a sensitivity and specificity of 80% for both IIH and MS. The CVD serum metabolite profile was 75% sensitive and specific. On analysing the second patient cohort, the established metabolite biomarker profiles generated from the first cohort showed moderate ability to segregate patients with IIH and MS (sensitivity:specificity of 63:75% and 67:75%, respectively). These findings suggest that NMR spectroscopic metabolic profiling of CSF and serum can identify differences between IIH, MS, CVD and mixed neurological diseases. Metabolomics may, therefore, have the potential to be developed into a clinically useful diagnostic tool. The identification of disease-unique metabolites may also impart information on disease pathology., ((c) 2009 John Wiley & Sons, Ltd.)

Context: Thyroid-associated ophthalmopathy (TAO) is a sight-threatening autoimmune disease in which de novo adipogenesis has been identified as a fundamental pathogenic mechanism. 11beta-Hydroxysteroid dehydrogenase 1 (11beta-HSD1) increases cortisol bioavailability and is pivotal in mediating glucocorticoid responses in adipose tissue and inflammation., Objective: In this study we characterize 11beta-HSD1 as a determinant of the adipogenic and inflammatory pathways in TAO orbital fat (OF) compared with normal OF., Patients and Methods: OF was harvested from 46 TAO and 44 control patients undergoing orbital surgery. Samples were examined by a combination of immunohistochemistry, real-time RT-PCR, primary cell culture, specific enzyme assays, colorimetric proliferation assays, and bead-based ELISA., Results: Glucocorticoid (glucocorticoid receptor-alpha,11beta-HSD1, hexose-6-phosphate dehydrogenase) and inflammatory cytokines (IL-1beta, IL-1 receptor, IL-6, TNF-alpha, TNF-alpha inductible protein, TGF-beta2) target genes together with markers of late adipocyte differentiation (fatty-acid-binding-protein-4, glycerol-6-phosphate-dehydrogenase) were highly expressed in TAO whole OF (P < 0.05) compared with controls. Primary cultures of TAO OF stromal cells demonstrated greater 11beta-HSD1 oxoreductase activity (P < 0.05), which was regulated by cytokines, most notably TNF-alpha (P < 0.01), compared with controls. Activity increased across differentiation, and this was most marked in TAO cells (P < 0.01). Similarly, stromal cell proliferation was limited by incubation with cortisol in TAO cells only. Furthermore, cortisone decreased IL-6 (P < 0.005), IL-8 (P < 0.05), and macrophage chemoattractant protein-1 (P < 0.05) production by cultured TAO cells only, an effect that was abrogated by inhibition of 11beta-HSD1., Conclusions: Induction of 11beta-HSD1 activity and expression by inflammatory cytokines (TNF-alpha, IL-6) may enhance orbital adipocyte differentiation (adipogenesis) and limit proliferation in TAO. 11beta-HSD1 may also have a role in regulating the local orbital inflammatory response.

Cushing's disease results from prolonged overexposure of tissues to endogenous glucocorticoids, secondary to adrenocorticotrophin excess from the pituitary. Common clinical signs and symptoms include weight gain, hypertension, and osteoporosis. Effective treatment of Cushing's disease can normalize biochemical levels, reverse comorbidities, and improve overall survival and quality of life. Treatment options include pituitary or adrenal surgery, radiotherapy, and various medical therapies, but each has important limitations. Medical therapies that effectively target the pituitary tumour are urgently needed., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Context: 11Beta-hydroxysteroid dehydrogenase (11beta-HSD) enzymes are now appreciated to be important regulators of hormone action at a tissue level. 11Beta-HSD1 is widely expressed and increases glucocorticoid action through its unique ability to convert inactive glucocorticoids (cortisone in man, 11-dehydrocorticosterone in rodents) to their active forms (cortisol and corticosterone, respectively). The enzyme has roles in the normal hypothalamus-pituitary-adrenal (HPA) axis, has been implicated in metabolic syndrome, and may modulate various aspects of the immune response., Evidence Acquisition: A review of published, peer-reviewed medical literature (1990 to June 2009) on the physiology and pathophysiology of 11beta-HSD1 was performed with an emphasis on HPA axis consequences, the metabolic syndrome, and the inflammatory response., Evidence Synthesis: Studies of patients with genetic defects in 11beta-HSD1 action show abnormal HPA axis responses with hyperandrogenism being a major consequence. The mechanisms underlying these abnormalities have been explored in mouse models with targeted deletion of components of the 11beta-HSD1 system. A range of experimental studies emphasize the role of 11beta-HSD1 in the metabolic syndrome and the potential for treatment with chemical inhibitors. An emerging area is the role of 11beta-HSD1 in the inflammatory response., Conclusions: 11Beta-HSD1 activity is an important component of the HPA axis and contributes to the metabolic syndrome and the normal immune response. Ongoing clinical observations and the development of selective inhibitors will further clarify the role of 11beta-HSD1 in these areas.

Context: A number of retrospective studies report that patients with acromegaly have increased morbidity and premature mortality, with standardized mortality ratios (SMR) of 1.3-3. Many patients with acromegaly develop hypopituitarism as a result of the pituitary adenoma itself or therapies such as surgery and radiotherapy. Pituitary radiotherapy and hypopituitarism have also been associated with an increased SMR., Methods: Using the West MIDLANDS: Acromegaly database (n = 501; 275 female), we assessed the influence of prior radiotherapy and hypopituitarism (and replacement therapy) on mortality in patients with acromegaly. Median duration of follow-up was 14.0 yr (interquartile range, 7.9-21 yr)., Results: All-cause mortality was elevated [SMR, 1.7 (1.4, 2.0); P < 0.001]. On external analysis, prior radiotherapy, ACTH, and gonadotropin deficiency were associated with an elevated SMR [radiotherapy SMR, 2.1 (1.7-2.6); P = 0.006; ACTH deficiency SMR, 2.5 (1.9-3.2); P < 0.0005; and gonadotropin deficiency SMR, 2.1 (1.6-2.7); P = 0.037]. On internal analysis, the relative risk (RR) of mortality was increased in the radiotherapy [RR, 1.8 (1.2-2.8); P = 0.008] and ACTH-deficiency groups [RR, 1.7 (1.2-2.5); P = 0.004], but not in the gonadotropin- or TSH-deficiency groups. In the ACTH-deficient group, increased replacement doses of hydrocortisone greater than 25 mg/d were associated with increased mortality compared to lower doses., Conclusions: Radiotherapy and ACTH deficiency are significantly associated with increased mortality in patients with acromegaly. In ACTH-deficient patients, a daily dose of more than 25 mg hydrocortisone is associated with increased mortality compared to lower doses. These results have important implications for the treatment of patients with acromegaly and also raise issues as to the optimum hydrocortisone treatment regimens for ACTH-deficient patients.

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., 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., 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., 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.

Context: Polycystic ovary syndrome (PCOS) is characterized by hyperandrogenism, anovulation, and susceptibility to the metabolic syndrome. Altered peripheral cortisol metabolism has been reported in PCOS, but also in simple obesity., Objective: The aim of the study was to describe cortisol metabolism and metabolic characteristics of a large PCOS cohort and to delineate the effect of obesity by comparison to body mass index (BMI)-matched controls., Design and Setting: We conducted an observational, cross-sectional study at outpatient clinics of two secondary/tertiary care centers., Patients or Other Participants: A total of 178 PCOS patients fulfilling Rotterdam criteria and 100 BMI-matched controls participated in the study., Intervention: The study included 24-h urine collection for steroid metabolite excretion and fasting blood samples, followed by an oral glucose tolerance test., Main Outcome Measures: We measured urinary steroid metabolites including glucocorticoids and androgens and the ratios reflecting enzymatic activities involved in peripheral cortisol and androgen metabolism, 5 alpha-reductase, and 11 beta-hydroxysteroid dehydrogenase types 1 and 2. We also measured circulating levels of glucose, insulin, dehydroepiandrosterone, dehydroepiandrosterone sulfate, and testosterone and calculated homeostasis model assessment., Results: Total androgen metabolites were higher in PCOS patients compared to BMI-matched controls (4,105 +/- 2,047 vs. 2,532 +/- 1,610 microg/24 h for the nonobese; 5,547 +/- 2,911 vs. 2,468 +/- 1,794 microg/24 h for the obese; both P < 0.001). Total glucocorticoid metabolites were higher in obese PCOS vs. controls (10,786 +/- 3,852 vs. 8,834 +/- 4,487 microg/24 h; P = 0.001). 5 alpha-Reductase activity correlated with BMI, insulin levels, and homeostasis model assessment. Both obese and nonobese PCOS patients had higher 5 alpha-reductase activity than controls (all P < 0.05). 11 beta-Hydroxysteroid dehydrogenase activities did not differ between PCOS and controls., Conclusions: PCOS is associated with enhanced androgen and cortisol metabolite excretion and increased 5 alpha-reductase activity that cannot be explained by obesity alone. Increased adrenal corticosteroid production represents an important pathogenic pathway in PCOS.

Objectives: We sought to determine whether the addition of spironolactone to angiotensin-converting enzyme (ACE) inhibitors and angiotensin receptor blockers (ARBs) improves left ventricular mass and arterial stiffness in early-stage chronic kidney disease (CKD)., Background: Chronic kidney disease is associated with a high risk of cardiovascular disease and a high prevalence of left ventricular hypertrophy and arterial stiffness that confer an adverse prognosis. It is believed that these abnormalities are in part a result of activation of the renin-angiotensin-aldosterone system., Methods: After an active run-in phase with spironolactone 25 mg once daily, 112 patients with stage 2 and 3 CKD with good blood pressure control (mean daytime ambulatory blood pressure <130/85 mm Hg) on established treatment with ACE inhibitors or ARBs were randomized to continue spironolactone or to receive a matching placebo. Left ventricular mass (cardiac magnetic resonance) and arterial stiffness (pulse wave velocity/analysis, aortic distensibility) were measured before run in and after 40 weeks of treatment., Results: Compared with placebo, the use of spironolactone resulted in significant improvements in left ventricular mass (-14 +/- 13 g vs. +3 +/- 11 g, p < 0.01), pulse wave velocity (-0.8 +/- 1.0 m/s vs. -0.1 +/- 0.9 m/s, p < 0.01), augmentation index (-5.2 +/- 6.1% vs. -1.4 +/- 5.9%, p < 0.05), and aortic distensibility (0.69 +/- 0.86 x 10(-3) mm Hg vs. 0.04 +/- 1.04 x 10(-3) mm Hg, p < 0.01)., Conclusions: The use of spironolactone reduces left ventricular mass and improves arterial stiffness in early-stage CKD. These effects suggest that aldosterone exerts adverse cardiovascular effects in CKD and that spironolactone is worthy of further study as a treatment that could reduce adverse cardiovascular events. (Is Spironolactone Safe and Effective in the Treatment of Cardiovascular Disease in Mild Chronic Renal Failure; NCT00291720).

11 beta-Hydroxysteroid dehydrogenase type 1 (11 beta-HSD1) is a key enzyme in the conversion of cortisone to the functional glucocorticoid hormone cortisol. This activation has been implicated in several human disorders, notably the metabolic syndrome where 11 beta-HSD1 has been identified as a novel target for potential therapeutic drugs. Recent crystal structures have revealed the presence of a pronounced hydrophobic surface patch lying on two helices at the C-terminus. The physiological significance of this region has been attributed to facilitating substrate access by allowing interactions with the endoplasmic reticulum membrane. Here, we report that single mutations that alter the hydrophobicity of this patch (I275E, L266E, F278E, and L279E in the human enzyme and I275E, Y266E, F278E, and L279E in the guinea pig enzyme) result in greatly increased yields of soluble protein on expression in E. coli. Kinetic analyses of both reductase and dehydrogenase reactions indicate that the F278E mutant has unaltered K(m) values for steroids and an unaltered or increased k(cat). Analytical ultracentrifugation shows that this mutation also decreases aggregation of both the human and guinea pig enzymes, resulting in greater monodispersity. One of the mutants (guinea pig F278E) has proven easy to crystallize and has been shown to have a virtually identical structure to that previously reported for the wild-type enzyme. The human F278E enzyme is shown to be a suitable background for analyzing the effects of naturally occurring mutations (R137C, K187N) on enzyme activity and stability. Hence, the F278E mutants should be useful for many future biochemical and biophysical studies of the enzyme.

Context: The aims of treatment in patients with acromegaly are to achieve serum GH/IGF-I concentrations associated with cure or normalization of mortality and alleviation of symptoms., Objective and Methods: Using the West Midlands Acromegaly database (n = 501) we investigated the reliability of basal fasting GH in predicting nadir or mean GH during oral glucose tolerance test (OGTT) or GH day curve (GHDC), respectively, the degree of discordance between disease activity measured by GH and IGF-I values and the effect of radiotherapy on the above relationships. In total 773 OGTT and 507 GHDC were performed., Results: Basal fasting GH was strongly correlated with nadir/mean GH on OGTT/GHDC (r = +0.87, P < 0.0001, r = +0.93, P < 0.0001, respectively). A basal GH < 2.5 microg/l was associated with a nadir/mean GH during OGTT/GHDC < 2.5 microg/l in 98.6% and 88.2% of cases, respectively. Elevated IGF-I was seen in 32.4% and 46.4% of patients with GH nadir values during OGTT < 1 and < 2.5 microg/l, respectively, and in 21.2% and 45.9% of GHDC with mean GH < 1 and < 2.5 microg/l, respectively. Radiotherapy increased the discordance in GH and IGF-I as markers of disease activity at GH < 2.5 microg/l (elevated IGF-I-values when OGTT nadir GH < 2.5 microg/l: radiotherapy 55.5%vs. no radiotherapy 36.9%, P = 0.002)., Conclusions: There is a close relationship between a basal fasting GH < 2.5 microg/l and nadir/mean GH < 2.5 microg/l during OGTT/GHDC. There is a large discordance between disease activity when assessed by GH and IGF-I which is further increased by radiotherapy. These observations illustrate the challenge of defining appropriate biochemical end-points to achieve control of disease and normalization of mortality in acromegaly.

19-nor-progesterone (19-nor-P) has the characteristics of a potent mineralocorticoid in adrenalectomized or salt-loaded rats and is capable of causing hypertension. In human placenta, progesterone is converted to 19-hydroxy-progesterone, a precursor of 19-nor-P. In some states of pregnancy hypertension, 19-nor-P may inhibit renal 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2), thus allowing cortisol to bind to the mineralocorticoid receptor (MR). Therefore, we investigated the ability of 19-nor-P to inhibit human 11beta-HSD2. Fetal kidney cells (HEK 293) were transfected with human 11beta-HSD2 and incubated with increasing concentrations of 19-nor-P, labelled and unlabelled cortisol. Steroids were extracted, separated by TLC, and radioactivity was measured using a TLC scanner. 19-nor-P treatment did not significantly reduce 11beta-HSD2 activity (430 to 300 pmol/mg protein/h) in the range of tested concentrations. In conclusion, 19-nor-P did not inhibit human 11beta-HSD2 and seems not to be involved in human hypertension. Nevertheless, 19-nor-P may be converted by extra-adrenal tissues into 19-nor-deoxycorticosterone (DOC) or 19-nor-corticosterone, which are potent mineralocorticoids and may be involved in the pathogenesis of hypertension during pregnancy.

Objective: The aetiology of idiopathic intracranial hypertension (IIH) is not known, but its association with obesity is well-recognized. Recent studies have linked obesity with abnormalities in circulating inflammatory and adiposity related cytokines. The aim of this study was to characterize adipokine and inflammatory cytokine profiles in IIH., Design: Paired serum and cerebrospinal fluid (CSF) specimens were collected from 26 patients with IIH and compared to 62 control subjects. Samples were analysed for leptin, resistin, adiponectin, insulin, IL-1beta, IL-6, IL-8 (CXCL8), TNFalpha, MCP-1 (CCL2), hepatocyte growth factor, nerve growth factor and PAI-1 using multiplex bead immunoassays., Results: CSF leptin was significantly higher in patients with IIH (P = 0.001) compared to controls after correction for age, gender and body mass index (BMI). In the control population, BMI correlated with serum leptin (r = 0.34; P = 0.007) and CSF leptin (r = 0.51; P < 0.0001), but this was not the case for the IIH population. Profiles of other inflammatory cytokines and adipokines did not differ between IIH patients and controls once anthropometric factors had been accounted for., Conclusions: IIH was characterized by significantly elevated CSF leptin levels which did not correlate with BMI. We suggest that CSF leptin may be important in the pathophysiology of IIH and that obesity in IIH may occur as a result of hypothalamic leptin resistance.

Intramyocellular lipids (IMCL) are stored as discrete lipid droplets which are associated with a number of proteins. The lipid droplet-associated protein adipophilin (the human orthologue of adipose differentiation-related protein) is ubiquitously expressed and is one of the predominant lipid droplet-proteins in skeletal muscle. The aim of this study was to investigate the subcellular distribution of adipophilin in human muscle fibres and to measure the colocalization of adipophilin with IMCL. Muscle biopsies from six lean male cyclists (BMI 23.4 +/- 0.4, aged 31 +/- 2 years, W (max) 346 +/- 8) were stained for myosin heavy chain type 1, IMCL, adipophilin and mitochondria using immunofluorescence and viewed with widefield and confocal fluorescence microscopy. The present study shows that like IMCL, the adipophilin content is ~twofold greater in type I skeletal muscle fibres and is situated in the areas between the mitochondrial network. Colocalization analysis demonstrated that 61 +/- 2% of IMCL contain adipophilin. Although the majority of adipophilin is contained within IMCL, 36 +/- 4% of adipophilin is not associated with IMCL. In conclusion, this study indicates that the IMCL pool is heterogeneous, as the majority but not all IMCL contain adipophilin.

COX-2 promotes colon cancer. While both nonselective NSAIDs and selective COX-2 inhibitors reduce disease burden, their adverse gastrointestinal and cardiovascular side effects limit their therapeutic use. In this issue of the JCI, Zhang et al. used gene silencing and a derivative of licorice root to show that inhibition of the enzyme 11beta-hydroxysteroid dehydrogenase type II(11betaHSD2) reduces tumor COX-2 activity, tumor growth, and metastasis by increasing the tonic glucocorticoid-mediated suppression of the COX-2 signaling pathway without the adverse effects associated with NSAIDs and selective COX-2 inhibitors (see the related article beginning on page 876). Their findings suggest that 11betaHSD2 inhibition may be a potential therapeutic option in colon cancer, warranting further investigation.

Context: Acromegaly is associated with increased morbidity and mortality. Treatment options include surgery, radiotherapy, and medical therapy., Aims: The objective of the study was to examine the role of prolactin status, prior surgery, and radiotherapy on the response to medical therapy in patients with acromegaly and assess the relative efficacy of dopamine agonist therapy compared with somatostatin analog therapy., Materials and Methods: A total of 276 patients with acromegaly received either dopamine agonists (DA) and/or somatostatin analogs (SSA). One hundred seventy-two patients had received surgery and 73 radiotherapy prior to receiving medical therapy. One hundred ninety-eight of 276 received DA, and 143 of 276 received SSA. GH and IGF-I values at baseline and after 12 months on therapy were analyzed., Results: In the DA group, basal prolactin concentration did not predict response to therapy, GH percent reduction: hyperprolactinemia, 26.7% (-10.4 to 48) vs. normoprolactinemia, 34.8% (0.2-53.2), P = 0.58; IGF-I percent reduction: hyperprolactinemia 30.0% (9.2-43.1) vs. normoprolactinemia 16.8% (4-37), P = 0.45. Prior surgery was not associated with any difference in response to DA: GH percent reduction (P = 0.1) and IGF-I percent reduction (P = 0.08). By contrast, prior radiotherapy was associated with an enhanced efficacy of GH response to DA, P = 0.02. In the SSA group, there was no effect of prior surgery or radiotherapy on response of GH, but radiotherapy was associated with less marked IGF-I percent reduction (P = 0.05). SSA were more potent than DA at decreasing both GH [62.8% (20.7-85%) vs. 42.4% (-6.5 to 68.6), P < 0.008] and IGF-I [SSA 40.4% (0-64.3) vs. 8% (0-40.8), P = 0.05]., Conclusions: The effects of DA are irrespective of baseline prolactin concentrations. Prior radiotherapy is associated with differences in GH and IGF-I response to DA and SSA therapy.

Context: Nonalcoholic fatty liver disease represents the hepatic manifestation of the metabolic syndrome. Nonalcoholic steatohepatitis (NASH) is the progressive form of liver injury. The pathophysiology that leads to NASH is not well understood., Objective: We hypothesize that an altered cortisol metabolism in the liver may be a pathogenetic factor., Design and Patients: 75 patients (28 men, 47 women) underwent liver biopsy for elevation in liver enzymes. Histological diagnosis identified normal liver in eight, fatty liver in 20, NASH grade 1 in 22, grade 2 in nine, grade 3 in three patients, and other forms of hepatitis or cirrhosis in 13 patients. We quantified hepatic 11beta-hydroxysteroid dehydrogenase type1 (11beta-HSD1) and hexose-6-phosphate-dehydrogenase (H6PDH) mRNA expression by real-time PCR. In addition, analysis of 24 h urinary excretion of cortisol metabolites using GCMS was performed and compared with healthy controls., Results: 11beta-HSD1 mRNA expression correlated significantly (R2= 0.809; P < 0.001) with H6PDH mRNA expression, negatively with waist-to-hip ratio in women (R2= 0.394; P= 0.005), but not with urinary (THF + 5alpha-THF)/THE ratio, total cortisol metabolite excretion, age, BMI, degree of fatty liver or NASH stages. Total cortisol metabolite excretion was increased in patients with fatty liver or NASH compared with healthy controls., Conclusions: Our data suggest that expression of hepatic 11beta-HSD1 and H6PDH are closely interlinked. 11beta-HSD1 gene expression does not seem to be involved in the pathogenesis of fatty liver or NASH. However, those patients showed an increased 5alpha- and 5beta-reduction of cortisol leading to an increased cortisol turnover rate and an activation of the HPA axis.

External beam radiotherapy has been used in the management of pituitary adenomas for nearly a century, preventing tumor regrowth following surgery for non-functioning pituitary adenomas and suppressing functional hypersecretion in those which are hormonally active. However, it has been linked with a number of potentially significant complications including formation of secondary intracranial tumors, cognitive impairment, hypopituitarism and cerebrovascular disease, as well as increased mortality. Radiation may cause a variety of vascular injuries and hemodynamic changes to the cerebral vasculature, and several authors have reported cerebrovascular complications and an increase in cerebrovascular mortality in patients receiving radiotherapy for pituitary and other central nervous system tumors. Ten years following pituitary radiotherapy, over 50% of patients develop deficiencies in one or more anterior pituitary hormones. A number of studies have demonstrated increased mortality in patients with hypopituitarism, predominantly due to cerebrovascular and cardiovascular disease. However, no clear answer has emerged with regards to causation, and pituitary radiotherapy has only been linked directly to mortality in one of these studies. Questions remain unanswered, and the use of conventional external beam radiotherapy in the management of pituitary disease must involve a critical risk-benefit analysis in each case.

The activity of the pituitary-adrenal axis can profoundly impact on body composition. This is dramatically seen in Cushing's syndrome (CS) but changes in body composition are also implicated in depression and alcoholic pseudocushing's. The pathophysiological mechanisms underlying these changes remain poorly understood. Changes to body composition in CS include increased fat mass, decreased bone mass, thinning of the skin and reduced lean mass. Why these tissues are affected so dramatically is unclear. Additionally, the change in body composition between individuals varies considerably for reasons which are only now becoming evident. This paper reviews the phenotypic changes with altered pituitary-adrenal axis activity and discusses the mechanisms involved. The primary focus is on adipose, bone, muscle and skin since the most dramatic changes are seen in these tissues.

Aims/hypothesis: Exposure to excess glucocorticoid is associated with pancreatic beta cell damage and decreased glucose-stimulated insulin secretion (GSIS). Inactive glucocorticoids (cortisone, 11-dehydrocorticosterone) are converted to active cortisol and corticosterone by 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), which requires NADPH as cofactor, which is generated by hexose-6-phosphate dehydrogenase (H6PDH). We investigated the localisation and activity of 11beta-HSD1 within pancreatic islets, and determined its functional role in the regulation of insulin and glucagon secretion., Methods: mRNA expression of 11beta-HSD1 (also known as HSD11B1), glucocorticoid receptor and H6PDH (also known as H6PD) in human pancreas and murine islets was examined by real-time PCR. 11beta-HSD1 protein levels were examined by immunohistochemistry and immunofluorescence. 11beta-HSD1 activity was assessed in intact tissue and isolated islets of wild-type (WT) and both 11beta-Hsd1- and H6pdh-null mice. Glucagon secretion and insulin secretion were analysed by RIA and ELISA respectively in isolated murine islets incubated with dexamethasone., Results: 11beta-HSD1 co-localised with glucagon in the periphery of murine and human islets, but not with insulin or somatostatin. Dexamethasone, 11-dehydrocorticosterone and corticosterone induced a dose-dependent decrease in GSIS and glucagon secretion following low glucose stimulation. Reduction of 11beta-HSD1 activity with specific inhibitors or in experiments carried out in H6pdh-null mice reversed the effects of 11-dehydrocorticosterone, but had no effect following treatment with corticosterone., Conclusions/interpretation: Local regeneration of glucocorticoid via 11beta-HSD1 within alpha cells regulates glucagon secretion and in addition may act in a paracrine manner to limit insulin secretion from beta cells.

To examine any potential role for 1,25-dihydroxyvitamin D (1,25(OH)2D) in inflammation associated with chronic kidney disease we measured vitamin D metabolites, markers of inflammation and gene expression in 174 patients with a variety of kidney diseases. Urinary MCP-1 protein and renal macrophage infiltration were each significantly but inversely correlated with serum 1,25(OH)2D levels. Logistic regression analysis with urinary MCP-1 as binary outcome showed that a 10-unit increase in serum 1,25(OH)2D or 25OHD resulted in lower renal inflammation. Analysis of 111 renal biopsies found that renal injury was not associated with a compensatory increase in mRNA for the vitamin D-activating enzyme 25-hydroxyvitamin D-1alpha-hydroxylase (CYP27B1), its catabolic counterpart 24-hydroxylase, or the vitamin D receptor. There was, however, a significant association between tissue MCP-1 and CYP27B1. Patients with acute renal inflammation had a significant increase in urinary and tissue MCP-1, macrophage infiltration, and macrophage and renal epithelial CYP27B1 expression but significantly lower levels of serum 1,25(OH)2D in comparison to patients with chronic ischemic disease despite similar levels of renal damage. In vitro, 1,25(OH)2D attenuated TNFalpha-induced MCP-1 expression by human proximal tubule cells. Our study indicates that renal inflammation is associated with decreased serum vitamin D metabolites and involves activation of the paracrine/autocrine vitamin D system.

Context: Cortisone reductase deficiency (CRD) is characterized by a failure to regenerate cortisol from cortisone via 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), resulting in increased cortisol clearance, activation of the hypothalamic-pituitary-axis (HPA) and ACTH-mediated adrenal androgen excess. 11beta-HSD1 oxoreductase activity requires the reduced nicotinamide adenine dinucleotide phosphate-generating enzyme hexose-6-phosphate dehydrogenase (H6PDH) within the endoplasmic reticulum. CRD manifests with hyperandrogenism resulting in hirsutism, oligo-amenorrhea, and infertility in females and premature pseudopuberty in males. Recent association studies have failed to corroborate findings that polymorphisms in the genes encoding H6PDH (R453Q) and 11beta-HSD1 (Intron 3 inserted adenine) interact to cause CRD., Objective: Our objective was to reevaluate the genetics and steroid biochemistry of patients with CRD., Design: We analyzed 24-h urine collection for steroid biomarkers by gas chromatography/mass spectrometry and sequenced the HSD11B1 and H6PD genes in our CRD cohort., Patients: Patients included four cases presenting with hyperandrogenism and biochemical features clearly indicative of CRD., Results: Gas chromatography/mass spectrometry identified steroid biomarkers that correlated with CRD in each case. Three cases were identified as homozygous (R109AfsX3, Y316X, and G359D) and one case identified as compound heterozygous (c.960G-->A and D620fsX3) for mutations in H6PD. No mutations affecting enzyme activity were identified in the HSD11B1 gene. Expression and activity assays demonstrate loss of function for all reported H6PDH mutations., Conclusions: CRD is caused by inactivating mutations in the H6PD gene, rendering the 11beta-HSD1 enzyme unable to operate as an oxoreductase, preventing local glucocorticoid regeneration. These data highlight the importance of the redox control of cortisol metabolism and the 11beta-HSD1-H6PDH pathway in regulating hypothalamic-pituitary-adrenal axis activity.

Data are presented on the urinary corticosteroid metabolic profile of the mouse strain 129/svJ. Through the use of GC/MS we have characterized, or tentatively identified corticosterone (Kendall's compound B) metabolites of both the 11beta-hydroxy and 11-carbonyl (compound A) series in urine. Full mass spectra of the methyloxime-trimethylether derivatives of 15 metabolites are included in the paper as an aid to other researchers in the field. Metabolites ranged in polarity from tetrahydrocorticosterone (THB) to dihydroxy-corticosterone with dominance of highly polar steroids. We found that prior to excretion corticosterone can undergo oxidation at position 11beta, reduction at position 20 and A-ring reduction. Metabolites retaining the 3-oxo-4-ene structure can be hydroxylated at position 6beta- as well as at an unidentified position, probably 16alpha-. Saturated steroids can be hydroxylated at positions 1beta-, 6alpha-, 15alpha- and 16alpha. A pair of hydroxy-20-dihydro-corticosterone metabolites (OH-DHB) were the most important excretory products accounting for about 40% of the total. One metabolite of this type was identified as 6beta-hydroxy-DHB; the other, of similar quantitative importance was probably 16alpha-hydroxy-DHB. The ratio of metabolites of corticosterone (B) to those of 11-dehydro-corticosterone (A) was greater than 9:1, considerably higher than that for the equivalent "human" ratio of 1:1 for cortisol to cortisone metabolites. Results from this study allowed the evaluation of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) activity in mice with deleted glucose-6-phosphate transporter (G6PT). These mice had attenuated back-conversion of A to B resulting in an increased ratio of A-metabolites to B-metabolites [Walker EA, Ahmed A, Lavery GG, Tomlinson JW, Kim SY, Cooper MS, Stewart PM, 11beta-Hydroxysteroid dehydrogenase type 1 regulation by intracellular glucose-6-phosphate, provides evidence for a novel link between glucose metabolism and HPA axis function. J Biol Chem 2007;282:27030-6]. We believe this study is currently the most comprehensive on the urinary steroid metabolic profile of the mouse. Quantitatively less steroid is excreted in urine than in feces by this species but urine analysis is more straightforward and the hepatic metabolites are less subject to microbial degradation than if feces was analyzed.

The effect of progestogens on bone is controversial with some studies suggesting an anabolic action while others show no effect. Prereceptor metabolism via localized expression of specific enzymes may have major impact on progesterone action in bone and may explain some of the discrepancies between studies. We therefore investigated the metabolism of progesterone in primary cultures of human osteoblasts and MG-63 osteoblastic cells. Osteoblasts and MG-63 cells were incubated with 4- (14)C-progesterone tracer and 50 nM unlabeled progesterone, and magnitude and pattern of progesterone metabolism were determined by two-dimensional thin-layer chromatography. Conventional and Taqman real-time PCR analysis were used to assess expression of progesterone metabolizing enzymes. In both types of cells the two major metabolic products of progesterone were 20 alpha-dihydroprogesterone and 5 alpha-dihydroprogesterone, but conversion to 3 alpha, 5 alpha- and 3 beta, 5 alpha-tetrahydroprogesterone was also detected. This activity was concomitant with expression of mRNAs for the enzymes AKR1C1, 5 alpha-reductase type 1 and AKR1C2, and 3 beta-HSD type 1 and 3-hydroxysteroid epimerase. In MG-63 cells progesterone metabolism was largely mediated via 5 alpha-reductase. In primary osteoblasts progesterone metabolism was unaffected by treatment with dexamethasone or estradiol, but in MG-63 cells dexamethasone pretreatment increased 5 alpha-reductase activity. Progesterone is subject to extensive intracellular inactivation in human osteoblasts, with potential attenuation of local progesterone receptor responses. Conversely, osteoblasts have the capacity to convert progestogens to metabolites reported to have anabolic actions through the estrogen receptor.

The present study identified three patients with bronchiectasis receiving inhaled corticosteroids (ICSs) who had symptomatic adrenal suppression secondary to ICS. The prevalence of adrenal suppression is unknown in bronchiectasis. The frequency of adrenal suppression and the impact of ICS use in bronchiectasis patients were examined. In total, 50 outpatients (33 receiving ICSs) underwent a short Synacthen test and completed a St George's Respiratory Questionnaire (SGRQ). Symptoms of adrenal suppression, steroid use and lung function were compared between subjects who were suppressed and those who were not. Adrenal suppression was evident in 23.5% of subjects who did not receive ICSs and 48.5% of those who did. Basal cortisol and the increments by which cortisol increased 30 min after Synacthen were lower in suppressed than in nonsuppressed subjects. The incremental cortisol rise was negatively correlated with SGRQ impacts and total score, suggesting a worse quality of life in those who had an impaired adrenal response. The greatest frequency of generalised symptoms was seen in the suppressed group. A significant proportion of subjects with bronchiectasis have evidence of adrenal suppression, and this is increased when inhaled corticosteroids are also used. Impairment of the cortisol response to stimulation is associated with poorer health status.

Idiopathic intracranial hypertension (IIH) is a common blinding condition amongst the young obese female population (20 per 100,000) characterised by elevated intracranial pressure (ICP). The aetiology of IIH is not known. In this review we explore the literature investigating the pathogenesis of IIH and suggest additional hypotheses. Chronic inflammation is emerging as an aetiological factor in the pathogenesis of obesity and we propose that this may be a feature of IIH. Obesity is also related to dysregulation of cortisol production by the pre-receptor enzyme, 11beta-hydroxysteroid dehydrogenase, and we speculate that this may have a role in the pathogenesis of obesity and raised ICP seen in IIH.

The question addressed in this study was the nature of the enzyme required to remove the side-chain of 17-hydroxycorticosteroids, leading in the case of cortisol to the excretion of 11beta-hydroxyandrosterone, 11-oxo-androsterone and the corresponding etiocholanolones. We questioned whether it could be CYP17, the 17-hydroxylase/17,20-lyase utilized in androgen synthesis. The conversion of exogenous cortisol to C(19) steroids in patients with complete 17-hydroxylase deficiency (17HD) was studied rationalizing that if CYP17 was involved no C(19) steroids would be formed. The urinary excretion of the four 11-oxy-C(19) steroids as well as many of the major C(21) cortisol metabolites were measured by GC/MS. Our results showed that the conversion of cortisol to C(19) steroids was normal in 17HD indicating that a currently unidentified enzyme must be responsible for this transformation. A secondary goal was to determine to what extent 11-oxy-C(19) steroids were metabolites of cortisol or adrenal synthesized 11beta-hydroxyandrostenedione. Since cortisol-treated 17HD patients cannot produce androstenedione, all C(19) 11-oxy-metabolites excreted must be derived from exogenous cortisol. The extent to which 17HD patients have lower relative excretion of C(19) steroids should reflect the absence of 11beta-hydroxyandrostenedione metabolites. Our results showed almost all of 11-oxo-etiocholanolone and 11beta-hydroxyetiocholanolone were cortisol metabolites, but in contrast the excretion of 11beta-hydroxyandrosterone was less than 10% that of normal individuals, indicating that in excess of 90% must be a metabolite of 11beta-hydroxyandrostenedione.

Objective: Our objective was to evaluate the published literature and reach a consensus on the treatment of patients with ACTH-dependent Cushing's syndrome, because there is no recent consensus on the management of this rare disorder., Participants: Thirty-two leading endocrinologists, clinicians, and neurosurgeons with specific expertise in the management of ACTH-dependent Cushing's syndrome representing nine countries were chosen to address 1) criteria for cure and remission of this disorder, 2) surgical treatment of Cushing's disease, 3) therapeutic options in the event of persistent disease after transsphenoidal surgery, 4) medical therapy of Cushing's disease, and 5) management of ectopic ACTH syndrome, Nelson's syndrome, and special patient populations., Evidence: Participants presented published scientific data, which formed the basis of the recommendations. Opinion shared by a majority of experts was used where strong evidence was lacking., Consensus Process: Participants met for 2 d, during which there were four chaired sessions of presentations, followed by general discussion where a consensus was reached. The consensus statement was prepared by a steering committee and was then reviewed by all authors, with suggestions incorporated if agreed upon by the majority., Conclusions: ACTH-dependent Cushing's syndrome is a heterogeneous disorder requiring a multidisciplinary and individualized approach to patient management. Generally, the treatment of choice for ACTH-dependent Cushing's syndrome is curative surgery with selective pituitary or ectopic corticotroph tumor resection. Second-line treatments include more radical surgery, radiation therapy (for Cushing's disease), medical therapy, and bilateral adrenalectomy. Because of the significant morbidity of Cushing's syndrome, early diagnosis and prompt therapy are warranted.

Background and Aim: The alcohol-induced pseudo-Cushing's syndrome is an important differential diagnosis of hypercortisolism that is poorly understood. Two isozymes of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) interconvert hormonally active cortisol (F) and inactive cortisone (E). Previously we have shown higher urinary F:E metabolite ratios (a reflection of global 11beta-HSD activity) in patients with alcoholic liver disease (ALD) compared to patients with chronic liver disease (CLD) of other aetiologies, suggesting that the phenotype of alcoholic pseudo-Cushing's may relate to altered metabolism of F., Subjects and Methods: We performed selective venous sampling of the hepatic, renal and peripheral veins measuring F and E concentrations (using in-house radioimmunoassay) in 20 patients with histologically confirmed ALD and 19 patients with CLD. Six patients who also had selective venous sampling for investigation of suspected hyperaldosteronism were used as 'normal' controls., Results: There was a significant difference in the hepatic F gradient (mean +/- SEM) between groups, indicating increased F production in the liver in patients with ALD (34.5 +/- 21.7 nmol/l) compared to those with CLD (-21.0 +/- 18.5 nmol/l) (P < 0.05) and normals (-19.7 +/- 17.2 nmol/l) (P < 0.05). 11beta-HSD1 mRNA expression was increased fivefold in the ALD group compared with normal controls (P < 0.01)., Conclusions: These results indicate significant induction of HSD11B1 gene expression and activity in patients with ALD during short- and long-term abstinence from alcohol. The mechanism is unknown but might be explained on the basis of alcohol-induced changes in intracellular redox potential or as a protective mechanism to limit liver inflammation and injury. Selective 11beta-HSD1 inhibitors may offer a novel therapeutic approach to treat alcoholic pseudo-Cushing's.

In adipose tissue, glucocorticoids regulate lipogenesis and lipolysis. Hexose-6-phosphate dehydrogenase (H6PDH) is an enzyme located in the endoplasmic reticulum that provides a cofactor for the enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), regulating the set point of its activity and allowing for tissue-specific activation of glucocorticoids. The aim of this study was to examine the adipose tissue biology of the H6PDH null (H6PDH/KO) mouse. Real-time PCR analysis confirmed similar mRNA levels of 11beta-HSD1 and glucocorticoid receptor-alpha in wild-type (WT) and H6PDH/KO mice in liver and gonadal fat depots. Microsomal 11beta-HSD1 protein levels shown by Western blot analysis corresponded well with mRNA expression in gonadal fat of WT and H6PDH/KO mice. Despite this, the enzyme directionality in these tissues changed from predominately oxoreductase in WT to exclusively dehydrogenase activity in the H6PDH/KO mice. In the fed state, H6PDH/KO mice had reduced adipose tissue mass, but histological examination revealed no difference in average adipocyte size between genotypes. mRNA expression levels of the key lipogenic enzymes, acetyl CoA carboxylase, adiponutrin, and stearoyl-coenzyme A desaturase-2, were decreased in H6PDH/KO mice, indicative of impaired lipogenesis. In addition, lipolysis rates were also impaired in the H6PDH/KO as determined by lack of mobilization of fat and no change in serum free fatty acid concentrations upon fasting. In conclusion, in the absence of H6PDH, the set point of 11beta-HSD1 enzyme activity is switched from predominantly oxoreductase to dehydrogenase activity in adipose tissue; as a consequence, this leads to impairment of fat storage and mobilization.