Single-cell transcriptomics and chromatin accessibility profiling elucidate the kidney-protective mechanism of mineralocorticoid receptor antagonists.

Mineralocorticoid excess commonly leads to hypertension (HTN) and kidney disease. In our study, we used single-cell expression and chromatin accessibility tools to characterize the mineralocorticoid target genes and cell types. We demonstrated that mineralocorticoid effects were established through open chromatin and target gene expression, primarily in principal and connecting tubule cells and, to a lesser extent, in segments of the distal convoluted tubule cells. We examined the kidney-protective effects of steroidal and nonsteroidal mineralocorticoid antagonists (MRAs), as well as of amiloride, an epithelial sodium channel inhibitor, in a rat model of deoxycorticosterone acetate, unilateral nephrectomy, and high-salt consumption-induced HTN and cardiorenal damage. All antihypertensive therapies protected against cardiorenal damage. However, finerenone was particularly effective in reducing albuminuria and improving gene expression changes in podocytes and proximal tubule cells, even with an equivalent reduction in blood pressure. We noted a strong correlation between the accumulation of injured/profibrotic tubule cells expressing secreted posphoprotein 1 (Spp1), Il34, and platelet-derived growth factor subunit b (Pdgfb) and the degree of fibrosis in rat kidneys. This gene signature also showed a potential for classifying human kidney samples. Our multiomics approach provides fresh insights into the possible mechanisms underlying HTN-associated kidney disease, the target cell types, the protective effects of steroidal and nonsteroidal MRAs, and amiloride.

The influence of pharmacological mineralocorticoid and glucocorticoid receptor blockade on the cortisol response to psychological stress.

The glucocorticoid cortisol is the end product of the hypothalamic-pituitary-adrenal (HPA) axis and crucial for the stress response in humans. Cortisol regulates numerous biological functions by binding to two different types of receptors: the mineralocorticoid receptor (MR) and the glucocorticoid receptor (GR). Both receptors are found in the brain where they are crucially involved in various mental functions and in feedback inhibition of cortisol release. The precise role of both receptors in the human stress response is not completely understood. In this study, we examined the effects of pharmacological blockade of the MR or the GR on stress-induced cortisol release in a sample of 318 healthy young men (M = 25.42, SD = 5.01). Participants received the MR antagonist spironolactone (300 mg), the GR antagonist mifepristone (600 mg), or a placebo and were subjected 90 min later to a social-evaluative stressor (Trier Social Stress Test) or a non-stressful control condition. We found significantly higher stress-induced cortisol release in the spironolactone group, whereas participants after mifepristone administration did not differ from the control groups. These results suggest that MR blockade results in attenuated fast negative feedback processes and emphasize the important role of the MR during the early phase of the stress response.

Mineralocorticoid receptor agonist aldosterone rescues hippocampal neural stem cell proliferation defects and improves postoperative cognitive function in aged mice.

OBJECTIVES: Hippocampal neurogenesis is closely related to learning and memory, and hippocampal neurogenesis disorders are involved in the development of many neurodegenerative diseases. Mineralocorticoid receptor (MR) plays a vital role in regulating stress response, neuroendocrine and cognitive functions, and is involved in regulating the integrity and stability of neural networks. However, the potential role of MR in the pathogenesis of postoperative cognitive dysfunction (POCD) is unclear. Therefore, this study evaluated the effect and mechanism of MR activation on postoperative hippocampal neurogenesis and cognitive function in aged mice. METHODS: 18-month-old male Kunming mice were randomly divided into Control group (C group), Surgery group (S group), Surgery+ Aldosterone group (S+Aldo group), Surgery + Wortmannin group (S+Wort group), Surgery + Aldosterone + Wortmannin group (S+Aldo+Wort group). Laparotomy was used to establish an animal model of postoperative cognitive dysfunction. After surgery, mice were intraperitoneally injected with aldosterone (100 ug/kg,150 ug/kg,200 ug/kg) and / or wortmannin (1 mg/kg); One day before the sacrifice, mice were injected intraperitoneally with BrdU (100 mg / kg / time, 3 times in total). Mice were subjected to Morris water maze and field tests at 1, 3, 7, and 14 days after surgery. Immunofluorescence was used to detect the number of BrdU +, Nestin +, BrdU/Nestin + positive cells in the hippocampal dentate gyrus of mice at 1, 3, 7 and 14 days after surgery. Western-blot was used to detect PI3K/Akt/GSK-3ß signaling pathway related proteins Akt, p-Akt, GSK-3ß, P-GSK-3ß expression. RESULTS: Stress impairs the performance of aged mice in water maze and open field tests, reduces the number of BrdU/Nestin+ cells in the hippocampal dentate gyrus, and inhibits the phosphorylation of Akt and GSK-3ß proteins in the hippocampus. Aldosterone treatment promotes P-Akt, P-GSK-3ß protein expression and hippocampal neural stem cell proliferation, and improves postoperative cognitive dysfunction. However, wortmannin treatment significantly reversed these effects of aldosterone. CONCLUSIONS: The mineralocorticoid receptor agonist aldosterone promotes the proliferation of hippocampal neural stem cells and improves cognitive dysfunction in aged mice after surgery, and the mechanism may be related to activation of PI3K/Akt/GSK-3ß signaling.

Corticotropin-stimulated steroid profiles to predict shock development and mortality in sepsis: From the HYPRESS study.

RATIONALE: Steroid profiles in combination with a corticotropin stimulation test provide information about steroidogenesis and its functional reserves in critically ill patients. OBJECTIVES: We investigated whether steroid profiles before and after corticotropin stimulation can predict the risk of in-hospital death in sepsis. METHODS: An exploratory data analysis of a double blind, randomized trial in sepsis (HYPRESS [HYdrocortisone for PRevention of Septic Shock]) was performed. The trial included adult patients with sepsis who were not in shock and were randomly assigned to placebo or hydrocortisone treatment. Corticotropin tests were performed in patients prior to randomization and in healthy subjects. Cortisol and precursors of glucocorticoids (17-OH-progesterone, 11-desoxycortisol) and mineralocorticoids (11-desoxycorticosterone, corticosterone) were analyzed using the multi-analyte stable isotope dilution method (LC-MS/MS). Measurement results from healthy subjects were used to determine reference ranges, and those from placebo patients to predict in-hospital mortality. MEASUREMENTS AND MAIN RESULTS: Corticotropin tests from 180 patients and 20 volunteers were included. Compared to healthy subjects, patients with sepsis had elevated levels of 11-desoxycorticosterone and 11-desoxycortisol, consistent with activation of both glucocorticoid and mineralocorticoid pathways. After stimulation with corticotropin, the cortisol response was subnormal in 12% and the corticosterone response in 50% of sepsis patients. In placebo patients (n = 90), a corticotropin-stimulated cortisol-to-corticosterone ratio > 32.2 predicted in-hospital mortality (AUC 0.8 CI 0.70-0.88; sensitivity 83%; and specificity 78%). This ratio also predicted risk of shock development and 90-day mortality. CONCLUSIONS: In this exploratory analysis, we found that in sepsis mineralocorticoid steroidogenesis was more frequently impaired than glucocorticoid steroidogenesis. The corticotropin-stimulated cortisol-to-corticosterone ratio predicts the risk of in-hospital death. Trial registration Clinical trial registered with www. CLINICALTRIALS: gov Identifier: NCT00670254. Registered 1 May 2008, https://clinicaltrials.gov/ct2/show/NCT00670254 .

Effects of steroidal mineralocorticoid receptor antagonists on acute and chronic estimated glomerular filtration rate slopes in patients with chronic heart failure.

AIMS: Steroidal mineralocorticoid receptor antagonists (MRAs) form a cornerstone of the management of heart failure (HF), but little is known about the long-term effects of MRA therapy on kidney function. We evaluated acute and chronic estimated glomerular function (eGFR) slopes in the two largest completed trials testing steroidal MRAs in chronic HF. METHODS AND RESULTS: We conducted parallel post hoc eGFR slope analyses in two multinational, double-blind randomized, placebo-controlled trials of steroidal MRAs in chronic HF with reduced ejection fraction (EMPHASIS-HF) and preserved ejection fraction (TOPCAT Americas region). GFR was estimated using the Chronic Kidney Disease Epidemiology Collaboration equation. Annual slopes of eGFR were assessed by generalized random coefficient models. Least square mean differences in eGFR slopes between steroidal MRA and placebo arms were assessed. Median follow-up was 1.8 years (EMPHASIS-HF) and 3.3 years (TOPCAT Americas). From baseline to month 4-6 ('acute eGFR slope'), compared to placebo, MRA treatment led to an acute decline in eGFR of -2.4 ml/min/1.73 m2 (95% confidence interval [CI] -3.4 to -1.4; p < 0.001) and -2.0 ml/min/1.73 m2 (95% CI -3.0 to -1.8; p < 0.001) in EMPHASIS-HF and TOPCAT Americas, respectively. From month 4-6 to end of study, there was no difference in 'chronic eGFR slope' between MRA and placebo arms (-0.3 ml/min/1.73 m2 /year [95% CI -1.3 to 0.7; p = 0.53] and 0.1 ml/min/1.73 m2 /year [95% CI -1.4 to 1.7; p = 0.86]) in EMPHASIS-HF and TOPCAT Americas, respectively. CONCLUSIONS: Steroidal MRAs result in acute declines in eGFR but do not modify long-term kidney disease trajectories in chronic HF with reduced or preserved ejection fraction. CLINICAL TRIAL REGISTRATION: EMPHASIS-HF (ClinicalTrials.gov NCT00232180) and TOPCAT (ClinicalTrials.gov NCT00094302).

No influence of mineralocorticoid and glutamatergic NMDA receptor stimulation on spatial learning and memory in individuals with major depression.

BACKGROUND: Major depressive disorder (MDD) is associated with impairments in spatial learning and memory and with altered functioning of central mineralocorticoid receptors (MR) and glutamatergic N-methyl-D-aspartate receptors (NMDA-R). Both receptors are highly expressed in the hippocampus and prefrontal cortex - brain areas that are critical for spatial learning and memory. Here, we examined the effects of separate and combined MR and NMDA-R stimulation on spatial learning and memory in individuals with MDD and healthy controls. METHODS: We used a randomized, double-blind, placebo-controlled between-group study design to examine the effects of separate and combined stimulation of the MR (with 0.4 mg fludrocortisone) and NMDA-R (with 250 mg D-cycloserine) in 116 unmedicated individuals with MDD (mean age: 34.7 ± 13.3 years; 78.4% women) and 116 age-, sex-, and education-matched healthy controls. Participants were randomly assigned to one of four conditions: 1) placebo; 2) MR stimulation; 3) NMDA-R stimulation; and 4) combined MR/NMDA-R stimulation. Three hours after drug administration, spatial learning and memory were assessed using a virtual Morris Water Maze task. RESULTS: Individuals with MDD and healthy controls did not differ in spatial learning and memory performance. Neither separate nor combined MR or NMDA-R stimulation altered measures of spatial performance. CONCLUSION: In this study of relatively young, predominantly female, and unmedicated individuals, we found no effect of MDD and no effect of separate or combined MR and NMDA-R stimulation on spatial learning and memory.

Secreted protein acidic and rich in cysteine (SPARC) and a disintegrin and metalloproteinase with thrombospondin type 1 motif (ADAMTS1) increments by the renin-angiotensin system induce renal fibrosis in deoxycorticosterone acetate-salt hypertensive rats.

Secreted protein acidic and rich in cysteine (SPARC), an extracellular matrix (ECM) protein, was recently shown to induce collagen deposition through the production of a disintegrin and metalloproteinase with thrombospondin type 1 motif (ADAMTS1) in the aging heart. ADAMTS1 regulates ECM turnover by degrading ECM components, and its excessive activation contributes to various pathological states, including fibrosis. The present study investigated the pathophysiological regulation and role of SPARC and ADAMTS1 in renal fibrosis using uninephrectomized rats treated with deoxycorticosterone acetate (DOCA, 40 mg/kg/week, subcutaneously) and salt (1% in drinking water). The administration of DOCA and salt gradually and significantly elevated systolic blood pressure during the 3-week treatment period, induced proteinuria, decreased creatinine clearance, and increased NADPH oxidase-derived superoxide production, malondialdehyde concentrations, and monocyte chemoattractant protein-1 and osteopontin expression in the kidneys. Glomerulosclerosis, fibrillar collagen deposition, and transforming growth factor-ß expression increased in a time-dependent manner, and SPARC and ADAMTS1 expression showed a similar pattern to these changes. The angiotensin II type-1 receptor blocker losartan suppressed the overexpression of SPARC and ADAMTS1, and an in vitro exposure to angiotensin II induced the production of both SPARC and ADAMTS1 in renal fibroblast NRK-49F cells. Knockdown of the SPARC gene with small interfering RNA reduced all forms (the 110-kDa latent and 87- and 65-kDa bioactive forms) of ADAMTS1 expression as well as collagen production. These results suggest that SPARC is induced by the renin-angiotensin system and may be a fibrogenic factor, at least in part, by producing ADAMTS1 in hypertensive renal disease.

Further evidence against the role renal medullary perfusion in short-term control of arterial pressure in normotensive and mildly or overtly hypertensive rats.

Earlier evidence from studies of rat hypertension models undermines the widespread view that the rate of renal medullary blood flow (MBF) is critical in control of arterial pressure (MAP). Here, we examined the role of MBF in rats that were normotensive, with modest short-lasting pressure elevation, or with overt established hypertension. The groups studied were anaesthetised Sprague-Dawley rats: (1) normotensive, (2) with acute i.v. norepinephrine-induced MAP elevation, and (3) with hypertension induced by unilateral nephrectomy followed by administration of deoxycorticosterone-acetate (DOCA) and 1% NaCl drinking fluid for 3 weeks. MBF was measured (laser-Doppler probe) and selectively increased using 4-h renal medullary infusion of bradykinin. MAP, renal excretion parameters and post-experiment medullary tissue osmolality and sodium concentration were determined. In the three experimental groups, baseline MAP was 117, 151 and 171 mmHg, respectively. Intramedullary bradykinin increased MBF by 45%, 65% and 70%, respectively, but this was not associated with a change in MAP. In normotensive rats a significant decrease in medullary tissue sodium was seen. The intramedullary bradykinin specifically increased renal excretion of water, sodium and total solutes in norepinephrine-treated rats but not in the two other groups. As previously shown in models of rat hypertension, in the normotensive rats and those with acute mild pressure elevation (resembling labile borderline human hypertension), 4-h renal medullary hyperperfusion failed to decrease MAP. Nor did it decrease in DOCA-salt model mimicking low-renin human hypertension. Evidently, within the 4-h observation, medullary perfusion was not a critical determinant of MAP in normotensive and hypertensive rats.

Increased SGK1 activity potentiates mineralocorticoid/NaCl-induced kidney injury.

Serum and glucocorticoid-regulated kinase 1 (SGK1) stimulates aldosterone-dependent renal Na+ reabsorption and modulates blood pressure. In addition, genetic ablation or pharmacological inhibition of SGK1 limits the development of kidney inflammation and fibrosis in response to excess mineralocorticoid signaling. In this work, we tested the hypothesis that a systemic increase in SGK1 activity would potentiate mineralocorticoid/salt-induced hypertension and kidney injury. To that end, we used a transgenic mouse model with increased SGK1 activity. Mineralocorticoid/salt-induced hypertension and kidney damage was induced by unilateral nephrectomy and treatment with deoxycorticosterone acetate and NaCl in the drinking water for 6 wk. Our results show that although SGK1 activation did not induce significantly higher blood pressure, it produced a mild increase in glomerular filtration rate, increased albuminuria, and exacerbated glomerular hypertrophy and fibrosis. Transcriptomic analysis showed that extracellular matrix- and immune response-related terms were enriched in the downregulated and upregulated genes, respectively, in transgenic mice. In conclusion, we propose that systemically increased SGK1 activity is a risk factor for the development of mineralocorticoid-dependent kidney injury in the context of low renal mass and independently of blood pressure.NEW & NOTEWORTHY Increased activity of the protein kinase serum and glucocorticoid-regulated kinase 1 may be a risk factor for accelerated renal damage. Serum and glucocorticoid-regulated kinase 1 expression could be a marker for the rapid progression toward chronic kidney disease and a potential therapeutic target to slow down the process.

Greater T Regulatory Cells in Females Attenuate DOCA-Salt-Induced Increases in Blood Pressure Versus Males.

Hypertension is the most common risk factor for cardiovascular disease, causing over 18 million deaths a year. Although the mechanisms controlling blood pressure (BP) in either sex remain largely unknown, T cells play a critical role in the development of hypertension. Further evidence supports a role for the immune system in contributing to sex differences in hypertension. The goal of the current study was to first, determine the impact of sex on the renal T-cell profiles in DOCA-salt hypertensive males and females and second, test the hypothesis that greater numbers of T regulatory cells (Tregs) in females protect against DOCA-salt-induced increases in BP and kidney injury. Male rats displayed greater increases in BP than females following 3 weeks of DOCA-salt treatment, although increases in renal injury were comparable between the sexes. DOCA-salt treatment resulted in an increase in proinflammatory T cells in both sexes; however, females had more anti-inflammatory Tregs than males. Additional male and female DOCA-salt rats were treated with anti-CD25 to decrease Tregs. Decreasing Tregs significantly increased BP only in females, thereby abolishing the sex difference in the BP response to DOCA-salt. This data supports the hypothesis that Tregs protect against the development of hypertension and are particularly important for the control of BP in females.

Molecular mechanisms of posaconazole- and itraconazole-induced pseudohyperaldosteronism and assessment of other systemically used azole antifungals.

Recent reports described cases of severe hypertension and hypokalemia accompanied by low renin and aldosterone levels during antifungal therapy with posaconazole and itraconazole. These conditions represent characteristics of secondary endocrine hypertension caused by mineralocorticoid excess. Different mechanisms can cause mineralocorticoid excess, including inhibition of the adrenal steroidogenic enzymes CYP17A1 and CYP11B1, inhibition of the peripheral cortisol oxidizing enzyme 11ß-hydroxysteroid dehydrogenase type 2 (11ß-HSD2) or direct activation of the mineralocorticoid receptor (MR). Compared to previous experiments revealing a threefold more potent inhibition of 11ß-HSD2 by itraconazole than with posaconazole, the current study found sevenfold stronger CYP11B1 inhibition by posaconazole over itraconazole. Both compounds most potently inhibited CYP11B2. The major pharmacologically active itraconazole metabolite hydroxyitraconazole (OHI) resembled the effects of itraconazole but was considerably less active. Molecular modeling calculations assessed the binding of posaconazole, itraconazole and OHI to 11ß-HSD2 and the relevant CYP enzymes, and predicted important interactions not formed by the other systemically used azole antifungals, thus providing an initial explanation for the observed inhibitory activities. Together with available clinical observations, the presented data suggest that itraconazole primarily causes pseudohyperaldosteronism through cortisol-induced MR activation due to 11ß-HSD2 inhibition, and posaconazole by CYP11B1 inhibition and accumulation of the mineralocorticoids 11-deoxycorticosterone and 11-deoxycortisol because of hypothalamus-pituitary-adrenal axis (HPA) feedback activation. Therapeutic drug monitoring and introduction of upper plasma target levels may help preventing the occurrence of drug-induced hypertension and hypokalemia. Furthermore, the systemically used azole antifungals voriconazole, isavuconazole and fluconazole did not affect any of the mineralocorticoid excess targets, offering alternative therapeutic options.

Plasma Renin Measurements are Unrelated to Mineralocorticoid Replacement Dose in Patients With Primary Adrenal Insufficiency.

CONTEXT: No consensus exists for optimization of mineralocorticoid therapy in patients with primary adrenal insufficiency. OBJECTIVE: To explore the relationship between mineralocorticoid (MC) replacement dose, plasma renin concentration (PRC), and clinically important variables to determine which are most helpful in guiding MC dose titration in primary adrenal insufficiency. DESIGN: Observational, retrospective, longitudinal analysis. PATIENTS: A total of 280 patients (with 984 clinical visits and plasma renin measurements) with primary adrenal insufficiency were recruited from local databases and the international congenital adrenal hyperplasia (CAH) registry (www.i-cah.org). Thirty-seven patients were excluded from the final analysis due to incomplete assessment. Data from 204 patients with salt-wasting CAH (149 adults and 55 children) and 39 adult patients with Addison disease (AD) were analysed. MAIN OUTCOME MEASURES: PRC, electrolytes, blood pressure (BP), and anthropometric parameters were used to predict their utility in optimizing MC replacement dose. RESULTS: PRC was low, normal, or high in 19%, 36%, and 44% of patients, respectively, with wide variability in MC dose and PRC. Univariate analysis demonstrated a direct positive relationship between MC dose and PRC in adults and children. There was no relationship between MC dose and BP in adults, while BP increased with increasing MC dose in children. Using multiple regression modeling, sodium was the only measurement that predicted PRC in adults. Longitudinally, the change in MC dose was able to predict potassium, but not BP or PRC. CONCLUSIONS: The relationship between MC dose and PRC is complex and this may reflect variability in sampling with respect to posture, timing of last MC dose, adherence, and concomitant medications. Our data suggest that MC titration should not primarily be based only on PRC normalization, but also on clinical parameters such as BP and electrolyte concentration.

The selective mineralocorticoid receptor modulator AZD9977 reveals differences in mineralocorticoid effects of aldosterone and fludrocortisone.

INTRODUCTION:: AZD9977 is a novel mineralocorticoid receptor (MR) modulator, which in preclinical studies demonstrated organ protection without affecting aldosterone-regulated urinary electrolyte excretion. However, when tested in humans, using fludrocortisone as an MR agonist, AZD9977 exhibited similar effects on urinary Na+/K+ ratio as eplerenone. The aim of this study is to understand whether the contradictory results seen in rats and humans are due to the mineralocorticoid used. MATERIALS AND METHODS:: Rats were treated with single doses of AZD9977 or eplerenone in combination with either aldosterone or fludrocortisone. Urine was collected for five to six hours and total amounts excreted Na+ and K+ were assessed. RESULTS:: AZD9977 dose-dependently increased urinary Na+/K+ ratio in rats when tested against fludrocortisone, but not when tested against aldosterone. Eplerenone dose-dependently increased urinary Na+/K+ ratio when tested against fludrocortisone as well as aldosterone. CONCLUSIONS:: The data suggest that the contrasting effects of AZD9977 on urinary electrolyte excretion observed in rats and humans are due to the use of the synthetic mineralocorticoid fludrocortisone. Future clinical studies are required to confirm the reduced electrolyte effects of AZD9977 and the subsequent lower predicted hyperkalemia risk.

Influence of glucocorticoid and mineralocorticoid receptor stimulation on task switching.

The influence of stress on executive functions has been demonstrated in numerous studies and is potentially mediated by the stress-induced cortisol release. Yet, the impact of cortisol on cognitive flexibility and task switching in particular remains equivocal. In this study, we investigated the influence of pharmacological glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) stimulation, two corticosteroid receptor types known to be responsible for cortisol effects on the brain. We conducted two experiments, each with 80 healthy participants (40 women and 40 men), and tested the effect of the unspecific MR/GR agonist hydrocortisone (Experiment I) and the more specific MR agonist fludrocortisone (Experiment II) on switch costs and task rule congruency in a bivalent, cued task switching paradigm. The results did not confirm our hypotheses; we found no significant effects of our manipulations on task switching capacity, although general switching and congruency effects were observed. We discuss the absence of MR/GR-mediated effects and propose alternative mechanisms that could explain stress induced effects on task switching.

Mineralocorticoid antagonism enhances brown adipose tissue function in humans: A randomized placebo-controlled cross-over study.

AIM: To investigate whether mineralocorticoid (MC) antagonism enhances brown adipose tissue (BAT) function in humans. MATERIALS AND METHODS: In a randomized double-blind, cross-over designed trial, 10 healthy adults (two men, eight women) underwent 2 weeks of spironolactone (100 mg/d) treatment and placebo, with an intervening 2-week wash-out period. BAT function was assessed in response to cooling and to a mixed meal. Metabolic activity was measured by fluoro-deoxyglucose (FDG) uptake (maximal standardized uptake value, SUVmax ) using PET-CT. Thermogenic activity was measured by skin temperatures overlying supraclavicular (SCL) BAT depots using infrared thermography. Postprandial metabolism was measured by energy production rate (EPR) and lipid synthesis using indirect calorimetry. RESULTS: During cooling, BAT metabolic activity (SUV 6.30 ± 2.16 vs 3.98 ± 1.34; P < 0.05) and volume (54.9 ± 22.8 vs 21.6 ± 11.8 cm3 ; P < 0.05) were significantly higher, and mean SCL temperature decreased by a smaller degree (-0.3°C°± 0.2°C vs -0.9°C ± 0.2°C; P = 0.05) with spironolactone treatment. A mixed meal increased SCL temperature and EPR. The postprandial rise in SCL temperature (+0.4°C ± 0.1°C vs +0.1°C ± 0.1°C; P < 0.05) but not in EPR was greater during spironolactone treatment. Postprandial lipid synthesis occurred in three participants with placebo but in none with spironolactone treatment (P = 0.06). CONCLUSION: MC antagonism enhanced human BAT function in response to cooling and to a meal during which lipid synthesis was suppressed. As postprandial EPR comprises energy dissipated as heat and energy required to store nutrients, the reduction in lipid synthesis during MC antagonism is a probable consequence of concurrent stimulation of BAT thermogenesis. The shift in energy usage from storage to heat dissipation indicates that MC antagonists may have therapeutic benefit for obesity.

Ecdysone Elicits Chronic Renal Impairment via Mineralocorticoid-Like Pathogenic Activities.

BACKGROUND/AIMS: Ecdysteroids are steroidal insect molting hormones that also exist in herbs. Ecdysteroid-containing adaptogens have been popularly used to improve well-being and by bodybuilders for muscle growth. However, the use of ecdysone in mammals is also associated with kidney growth and enlargement, indications of disturbed kidney homeostasis. The underlying pathogenic mechanism remains to be clarified. METHODS: Virtual screening tools were employed to identify compounds that are homologous to ecdysone and to predict putative ecdysone-interacting proteins. The kidney effect of ecdysone was examined in vitro and in vivo and compared with that of aldosterone. Cellular apoptosis was estimated by terminal deoxynucleotidyl transferase dUTP nick end labeling. Cell motility was assessed by scratch-wound cell migration assay. Blood urea nitrogen was measured to evaluate renal function. Western immunblot analysis was employed to determine the expression profile of interested proteins. RESULTS: Computational molecular structure analysis revealed that ecdysone is highly homologous to aldosterone. Moreover, virtual screening based on compound-protein interaction profiles identified the Mineralocorticoid Receptor (MR) to potentially interact with ecdysone. Accordingly, to assess potential biological functions of ecdysone in mammals, ecdysone was applied to mineralocorticoid-sensitive inner medullar collecting duct cells. Ecdysone induced mesenchymal accumulation of extracellular matrix and epithelial dedifferentiation characterized by de novo expression of α-smooth muscle actin. In addition, ecdysone elicited cellular apoptosis and retarded cell motility, akin to the effect of aldosterone. In vivo, daily treatment of mice with ecdysone increased cell apoptosis in the kidney, impaired renal function and elicited early signs of renal fibrogenesis, marked by deposition of collagen and fibronectin in tubulointerstitium, reminiscent of the action of aldosterone. The MR signaling pathway is likely responsible for the cellular and pathobiological effects of ecdysone, as evidenced by strong ecdysone-induced MR nuclear translocation in renal tubular cells both in vitro and in vivo, while blockade of MR by concomitant spironolactone treatment largely abolished the detrimental effects of ecdysone. CONCLUSION: Our findings suggest that ecdysone induces mineralocorticoid-dependent activities that impair renal function and elicit renal injury.

Immune Effects of Corticosteroids in Sepsis.

Sepsis, a life-threatening organ dysfunction, results from a dysregulated host response to invading pathogens that may be characterized by overwhelming systemic inflammation or some sort of immune paralysis. Sepsis remains a major cause of morbidity and mortality. Treatment is nonspecific and relies on source control and organ support. Septic shock, the most severe form of sepsis is associated with the highest rate of mortality. Two large multicentre trials, undertaken 15 years apart, found that the combination of hydrocortisone and fludrocortisone significantly reduces mortality in septic shock. The corticosteroids family is composed of several molecules that are usually characterized according to their glucocorticoid and mineralocorticoid power, relative to hydrocortisone. While the immune effects of glucocorticoids whether mediated or not by the intracellular glucocorticoid receptor have been investigated for several decades, it is only very recently that potential immune effects of mineralocorticoids via non-renal mineralocorticoid receptors have gained popularity. We reviewed the respective role of glucocorticoids and mineralocorticoids in counteracting sepsis-associated dysregulated immune systems.

Desoxycorticosterone Pivalate Duration of Action and Individualized Dosing Intervals in Dogs with Primary Hypoadrenocorticism.

BACKGROUND: Clinicians alter dosing for desoxycorticosterone pivalate (DOCP) to mitigate costs, but this practice has not been critically evaluated in a prospective clinical trial. HYPOTHESIS/OBJECTIVES: The duration of action of DOCP is longer than 30 days in dogs with primary hypoadrenocorticism (PH). ANIMALS: A total of 53 client-owned dogs with PH. Twenty-four dogs with newly diagnosed PH (Group 1) and 29 dogs with treated PH (Group 2). METHODS: Prospective, multicenter, clinical trial. For phase I, DOCP was administered and plasma sodium and potassium concentrations were measured until the dog developed hyponatremia or hyperkalemia at a planned evaluation, or displayed clinical signs with plasma electrolyte concentrations outside of the reference interval independent of a planned evaluation, thus defining DOCP duration of action. Plasma electrolyte concentrations then were assessed at the end of the individualized dosing interval (IDI; i.e., DOCP duration of action minus 7 days, phase II and at least 3 months after concluding phase II, phase III). RESULTS: The duration of action of DOCP in dogs in phase I with naïve PH (n = 24) ranged from 32 to 94 days (median, 62 days; 95% confidence interval [CI], 57, 65) and previously treated PH (n = 29) from 41 to 124 days (median, 67 days; CI, 56, 72). Overall, the final DOCP dosing interval for all dogs that completed phase II (n = 36) ranged from 38 days to 90 days (median, 58 days; CI, 53, 61). No dog that completed phase III (n = 15) required reduction in the IDI. The DOCP duration of action, independent of group, was not significantly associated with several baseline variables. The median drug cost reduction using IDI was approximately 57.5% per year. CONCLUSION AND CLINICAL IMPORTANCE: The duration of action of DOCP in dogs with PH is >30 days, and plasma sodium and potassium concentrations can be maintained with an IDI >30 days long term.

Mineralocorticoids modulate the expression of the ß-3 subunit of the Na+, K+-ATPase in the renal collecting duct.

Renal sodium reabsorption depends on the activity of the Na+,K+-ATPase α/ß heterodimer. Four α (α1-4) and 3 ß (ß1-3) subunit isoforms have been described. It is accepted that renal tubule cells express α1/ß1 dimers. Aldosterone stimulates Na+,K+-ATPase activity and may modulate α1/ß1 expression. However, some studies suggest the presence of ß3 in the kidney. We hypothesized that the ß3 isoform of the Na+,K+-ATPase is expressed in tubular cells of the distal nephron, and modulated by mineralocorticoids. We found that ß3 is highly expressed in collecting duct of rodents, and that mineralocorticoids decreased the expression of ß3. Thus, we describe a novel molecular mechanism of sodium pump modulation that may contribute to the effects of mineralocorticoids on sodium reabsorption.

30 YEARS OF THE MINERALOCORTICOID RECEPTOR: Evolution of the mineralocorticoid receptor: sequence, structure and function.

The mineralocorticoid receptor (MR) is descended from a corticoid receptor (CR), which has descendants in lamprey and hagfish, cyclostomes (jawless fish), a taxon that evolved at the base of the vertebrate line. A distinct MR and GR first appear in cartilaginous fishes (Chondrichthyes), such as sharks, skates, rays and chimeras. Skate MR has a strong response to corticosteroids that are mineralocorticoids and glucocorticoids in humans. The half-maximal responses (EC50s) for skate MR for the mineralocorticoids aldosterone and 11-deoxycorticosterone are 0.07 nM and 0.03 nM, respectively. EC50s for the glucocorticoids cortisol and corticosterone are 1 nM and 0.09 nM, respectively. The physiological mineralocorticoid in ray-finned fish, which do not synthesize aldosterone, is not fully understood because several 3-ketosteroids, including cortisol, 11-deoxycortisol, corticosterone, 11-deoxycorticosterone and progesterone are transcriptional activators of fish MR. Further divergence of the MR and GR in terrestrial vertebrates, which synthesize aldosterone, led to emergence of aldosterone as a selective ligand for the MR. Here, we combine sequence analysis of the CR and vertebrate MRs and GRs, analysis of crystal structures of human MR and GR and data on transcriptional activation by 3-ketosteroids of wild-type and mutant MRs and GRs to investigate the evolution of selectivity for 3-ketosteroids by the MR in terrestrial vertebrates and ray-finned fish, as well as the basis for binding of some glucocorticoids by human MR and other vertebrate MRs.