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Obesity impairs endothelial-mediated vasodilation, the earliest step in vascular disease and a contributor to hypertension. We previously demonstrated that endothelial cell mineralocorticoid receptor (MR) deletion prevents obesity-induced microvascular dysfunction in females by increasing nitric oxide-mediated vasodilation. Estrogen receptor alpha (ERα) can oppose MR function, therefore we hypothesized that ERα mediates the benefits of endothelial MR deficiency. Females lacking endothelial MR or wildtype littermates were fed control or high fat diet for 20 weeks to cause obesity. MR deletion improved mesenteric artery endothelial-dependent vasodilation in obese females and ERα inhibition ex vivo negated this protective effect. Endothelial MR deletion results in significantly more ERα mRNA and protein. In vitro, estrogen increases endothelial nitric oxide synthase phosphorylation, which is inhibited by aldosterone and dependent on MR. Both proteins co-immunoprecipitate with striatin and a mimetic peptide that disrupts ERα-striatin binding also decreased MR-striatin interaction. Finally, removing endothelial MR in obese females restored endothelial function by increasing the nitric oxide component of vasodilation. Combined deletion of endothelial ERα negated the benefit of endothelial MR deletion. These results indicate that endothelial ERα prevents the detrimental effects of MR in obesity by increasing nitric oxide to rescue vasodilation in females. MR and ERα may compete for striatin binding within endothelial cells to regulate nitric oxide. These data identify a novel mechanism that promotes MR antagonism to prevent obesity-induced microvascular dysfunction in females.

Mineralocorticoid receptors (MRs) are implicated in the pathology of hypertension. MR blockers are recommended for the treatment of salt-sensitive or resistant hypertension. However, use of currently available MR blockers is limited by adverse events. This phase 3 multicenter, randomized, double-blind study compared the efficacy and safety of esaxerenone, a new selective nonsteroidal MR blocker, at 2.5 and 5 mg/day and eplerenone 50 mg/day in Japanese patients with essential hypertension. After a 4-week washout period, 1001 eligible adults with hypertension were randomized evenly to esaxerenone 2.5 or 5 mg/day or eplerenone 50 mg/day treatments, taken orally once daily for 12 weeks. Primary end points were changes in sitting systolic or diastolic blood pressure (BP) from baseline at the end of treatment. Esaxerenone 2.5 mg/day was noninferior to eplerenone for reductions in sitting and 24-hour BP. Reductions in BP with esaxerenone 5 mg/day were significantly greater than those with esaxerenone 2.5 mg/day. Changes in diurnal BP showed persistent 24-hour antihypertensive effects in all treatment groups. The proportions of patients achieving target sitting BP ( Clinical Trial Registration— URL: http://www.clinicaltrials.gov . Unique identifier: NCT02890173.

Objectives: Men are disproportionately affected by the coronavirus disease-2019 (COVID-19) and experience higher mortality as compared to women. Endothelial dysfunction has been proposed as a major inciting factor of pro-inflammatory and thrombotic changes in COVID-19 infection. The vascular effects of androgen signaling and inflammatory cytokines in severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mediated endothelial damage are not defined. Methods: Here, we determined the effects of SARS-CoV-2 spike protein-mediated endothelial damage under conditions of exposure to androgens and TNF-α. We tested the therapeutic effects of approved drugs: angiotensin receptor blockade by valsartan and mineralocorticoid receptor antagonism by spironolactone. Circulating endothelial injury markers VCAM-1 and E-selectin were measured in the plasma from men and women diagnosed with COVID-19. Results: Exposure of endothelial cells (ECs) in vitro to androgen dihydrotestosterone (DHT) exacerbated spike protein S1-mediated endothelial injury transcripts for cell adhesion molecules E-selectin, VCAM-1, and ICAM-1 and anti-fibrinolytic PAI-1 (P Conclusions: Androgen exposure promoted spike protein-mediated endothelial injury by increasing markers of inflammation and thrombosis. A beneficial therapeutic effect of the FDA-approved drug spironolactone was observed, supporting a rationale for further evaluation as an adjunct treatment in COVID-19.

Aldosterone is an important mineralocorticoid responsible for fluid and electrolyte homeostasis produced by aldosterone synthase (CYP11B2). An aldosterone synthase inhibitor (ASI) may be a therapeutic option for primary aldosteronism-related conditions such as resistant hypertension. An ASI with sufficient selectivity for CYP11B2 versus the similar cortisol-producing enzyme CYP11B1 has remained elusive. PB6440 is a novel ASI that is potent and highly selective for CYP11B2. In vitro studies demonstrated 200-300-fold selectivity of PB6440 for human CYP11B2 compared to human CYP11B1. In single and multiple dose cynomolgus monkey studies of orally administered PB6440, dose-and concentration-dependent reduction of plasma aldosterone after ACTH challenge was observed with >90% reduction at higher doses. Consistent with its high selectivity, PB6440 had little effect on the CYP11B1 cortisol pathway. Plasma levels of cortisol, 11-deoxycortisol, and deoxycorticosterone, remained unchanged even at high doses of PB6440. Systolic and diastolic blood pressure was reduced in a dose-dependent manner. Circulating half-life of PB6440 was approximately 17 hours with high oral bioavailability. In summary, PB6440 is a highly selective ASI that demonstrated sustained aldosterone suppression for 14 days with no effect on the CYP11B1 pathway in non-human primates. In single and multiple dose studies, PB6440 appeared well tolerated, demonstrating good oral bioavailability, and a PK profile supportive of once daily dosing. These results suggest that PB6440 may be useful in humans as a novel therapeutic for treating hypertension or other conditions caused by excess aldosterone.

Introduction: Neutrophil Gelatinase-Associated Lipocalin (NGAL) (or lipocalin 2) is a novel mineralocorticoid biotarget in the cardiovascular system. NGAL is also described as an acute renal lesion biomarker and NGAL serum concentration is associated with the severity of renal damages patients with a chronic kidney disease (CKD). Lipocalin2 (Lcn2) gene invalidation in a CKD mouse model protects from proteinuria and renal lesions. Objective: We hypothesized that the NGAL from macrophages promotes expression of chemoattractant molecules involved in renal lesions induced by mineralocorticoid excess. Methods: The role of Lcn2 was analyzed using full Lcn2 knock out mice (NGAL KO) challenged with uni-Nephrectomy, Aldosterone 200μg/kg/day, Salt 1% (NAS model) during 6 weeks. Assessment of CCL5/IL4 in kidney fibrosis were studied using maraviroc administration (50mg/kg/d in chow diet) or by injections of anti-IL4 antibody (600μg/week). Results: NAS induced a significant increase in the expression (relative values, mean±SEM, compared to 1 in the control samples, p&lt0.05) of extracellular matrix proteins such as collagen I (2.35±0.33), α-SMA (2.04±0.44) and fibronectin (3.38±0.42) in the kidney of WT mice associated with interstitial kidney fibrosis (6.49±0.70). These modifications were fully prevented by NGAL deletion. Expression of macrophages markers F4/80 , CD80 and CD86 was increased (5.11±0.46, 4.84±0.19 and 5.22±0.45 respectively) in WT NAS mice and partly prevented in NGAL KO mice. Macrophages isolated from NGAL KO or WT mice were co-treated with aldosterone (10 -8 M) and NaCl (40mM). In WT macrophages, expression of Lcn2 (2.81±0.30) and the CCL5 chemokine (2.48±0.32) was increased. The increase of CCL5 was prevented in NGAL KO macrophages. Such as total deletion of NGAL, CCL5 receptor blockade improved renal fibrosis and high level of Th2 CD4 + cell markers induced by NAS. Neutralization of IL4 Th2 cytokine in NAS mice injected with anti-IL4 antibody blunted kidney fibrosis and overexpression of profibrotic proteins such as collagen I, α-SMA and fibronectin. Conclusion: NGAL produced by macrophages plays a critical role in renal interstitial fibrosis through CCL5/IL4 pathways in mice exposed to mineralocorticoid excess.

We examined whether fingolimod (FTY720), an S1PR (sphingosine-1-phosphate receptor) modulator, has beneficial or harmful effects on mineralocorticoid/salt-induced renal injury. Uninephrectomized rats on 0.9% NaCl/0.3% KCl drinking solution were randomly divided into control, control+FTY720, deoxycorticosterone acetate (DOCA), and DOCA+FTY720 groups and administered vehicle, vehicle+FTY720, DOCA+vehicle, and DOCA+FTY720 for 4 weeks, respectively. Only the DOCA+FTY720 group had reduced survival rates and showed hemolysis because of intravascular mechanical fragmentation of erythrocytes and thrombocytopenia. Both the DOCA+FTY720 and DOCA groups developed malignant hypertension, which was more severe in the DOCA+FTY720 group. In the DOCA+FTY720 group only, thrombotic microangiopathy involving severe renal arteriole endothelial cell injury was observed and was characterized by fibrinoid necrosis and onion-skin lesions in arterioles. There were fewer circulating endothelial progenitor cells in the DOCA+FTY720 group but more in the DOCA group compared with the control group. Expression levels of VEGF (vascular endothelial growth factor), S1PR1, and S1PR3 in renal arteriole endothelial cells were significantly greater in the DOCA+FTY720 and DOCA groups compared with the control group, with levels being similar between the 2 groups. Expression levels of endothelial nitric oxide synthase in renal arteriole endothelial cells were significantly lower in the DOCA+FTY720 group only. The control+FTY720 group showed reduced circulating endothelial progenitor cells but no significant functional or pathological changes in kidneys or changes in blood pressure. Exposure of uninephrectomized rats to DOCA/salt+FTY720 for 4 weeks induced renal arteriolar endothelial cell injury, resulting in the development of thrombotic microangiopathy. Consideration of this possibility is recommended when prescribing FTY720.

Aldosterone, the primary mineralocorticoid, is synthesized in the outer zone of the adrenal cortex called the zona glomerulosa (ZG). The production of aldosterone is tightly regulated by angiotensin II (Ang II) and circulating potassium levels.1 Physiologically, aldosterone plays a key role in the maintenance of intravascular volume and blood pressure through sodium retention in the kidney. Excess aldosterone causes hypertension and induces cardiovascular complications.2–5 The autonomous secretion of aldosterone, independent of Ang II and sodium status, is known as primary aldosteronism (PA). PA is the most common cause of endocrine-related hypertension with a prevalence of 5% to 10% in hypertensive population6–10 and ≈20% in resistant hypertension.11–13 Because of the increased risk for cardiovascular complications in patients with PA, early detection of the disease and targeted treatment is recommended.14 The molecular pathogenesis of PA was largely unknown until recently. The development of specific antibodies against aldosterone synthase (CYP11B2), which is required for the final steps of aldosterone production, has allowed the detection of aldosterone-producing cells in resected adrenals.15,16 Using these antibodies, non-neoplastic foci of CYP11B2-expressing cells called aldosterone-producing cell clusters (APCC)15 have been identified in adrenal tissues adjacent to aldosterone-producing adenomas (APA) and in normal human adrenal glands without tumor or hyperplasia.15–20 Studies have shown that APCC are a common occurrence in normal human adrenals.21–23 Because circulating renin and Ang II levels are suppressed in patients with PA, the observation of APCC adjacent to APA suggested that APCC may represent a source of autonomous and renin-independent aldosterone secretion, perhaps even a precursor to APA.24 Over the past 6 years, somatic and germline mutations that cause inappropriate aldosterone production have been identified in patients with PA. Most of the mutations …

Genome-wide analysis of renal sodium-transporting system has identified specific variations of Mendelian hypertensive disorders, including HSD11B2 gene variants in apparent mineralocorticoid excess. However, these genetic variations in extrarenal tissue can be involved in developing hypertension, as demonstrated in former studies using global and brain-specific Hsd11b2 knockout rodents. To re-examine the importance of renal dysfunction on developing hypertension, we generated kidney-specific Hsd11b2 knockout mice. The knockout mice exhibited systemic hypertension, which was abolished by reducing salt intake, suggesting its salt-dependency. In addition, we detected an increase in renal membrane expressions of cleaved epithelial sodium channel-α and T53-phosphorylated Na + -Cl − cotransporter in the knockout mice. Acute intraperitoneal administration of amiloride-induced natriuresis and increased urinary sodium/potassium ratio more in the knockout mice compared with those in the wild-type control mice. Chronic administration of amiloride and high-KCl diet significantly decreased mean blood pressure in the knockout mice, which was accompanied with the correction of hypokalemia and the resultant decrease in Na + -Cl − cotransporter phosphorylation. Accordingly, a Na + -Cl − cotransporter blocker hydrochlorothiazide significantly decreased mean blood pressure in the knockout mice. Chronic administration of mineralocorticoid receptor antagonist spironolactone significantly decreased mean blood pressure of the knockout mice along with downregulation of cleaved epithelial sodium channel-α and phosphorylated Na + -Cl − cotransporter expression in the knockout kidney. Our data suggest that kidney-specific deficiency of 11β-HSD2 leads to salt-dependent hypertension, which is attributed to mineralocorticoid receptor–epithelial sodium channel–Na + -Cl − cotransporter activation in the kidney, and provides evidence that renal dysfunction is essential for developing the phenotype of apparent mineralocorticoid excess.

Obesity is a risk factor for endothelial dysfunction and hypertension in pregnancy. Our lab has demonstrated that obesity-associated hypertension in female animal models develops via leptin-mediated, aldosterone-induced endothelial dysfunction (ED). We also showed that female mice are more sensitive to leptin-induced aldosterone activation of endothelial mineralocorticoid receptors (ECMR) due to an endogenous sex-specific upregulation of ECMR in females. Our previous data demonstrates that endothelial progesterone receptor activation increases ECMR expression and that ECMR deletion protects female mice from obesity-associated, leptin-mediated ED. However, whether ECMR play a role in ED in pregnancy is unknown. Therefore, we hypothesized that high progesterone levels increase ECMR expression in pregnant mice and that ECMR deletion protects pregnant mice from obesity-associated, leptin-mediated ED. Endothelial cells were isolated from aorta of timed-pregnant Balb/C mice (gestation day (GD) 17) and assessed for ECMR expression via RT-PCR. ECMR mRNA expression increased 9.2±0.2-fold (*P

Introduction: Neutrophil Gelatinase-Associated Lipocalin (NGAL) (or lipocalin 2) is a novel mineralocorticoid biotarget in the cardiovascular system. NGAL is also described as an acute renal lesion biomarker and NGAL serum concentration is associated with the severity of renal damages patients with a chronic kidney disease (CKD). Lipocalin2 (Lcn2) gene invalidation in a CKD mouse model protects from proteinuria and renal lesions. Objective: Since the immune system is involved in renal inflammation and fibrosis process, we hypothesized that the production of NGAL/Lcn2 by immune cells plays an important role in the deleterious mineralocorticoid effects in the kidney. Methods: The role of Lcn2 produced by immune cells was analyzed using full Lcn2 knock out mice (Full KO) and mice depleted for Lcn2 in their immune cells by bone marrow transplantation (BMT lcn2 KO). These mice were challenged with uni-Nephrectomy, Aldosterone 200μg/kg/day, Salt 1% (NAS model) during 6 weeks. Results: NAS induced a significant increase in the expression (relative values, mean±SEM, compared to 1 in the control samples, p-8 M) and NaCl (40mM). In WT macrophages, mRNA expression (relative values, mean±SEM, compared to 1 in the control samples, p Conclusion: NGAL produced by immune cells plays a critical role in renal interstitial fibrosis and inflammation induced by mineralocorticoid excess. The role of RANTES/CCl5 in the renal NGAL-mineralocorticoid pathways is currently analyzed.

We investigated the role of angiotensin type 1a receptors (AGTR1a) in vascular injury induced by aldosterone activation of mineralocorticoid receptors in Agtr1a −/− and wild-type (WT) mice infused with aldosterone for 14 days while receiving 1% NaCl in drinking water. Aldosterone increased systolic blood pressure (BP) by ≈30 mm Hg in WT mice and ≈50 mm Hg in Agtr1a −/− mice. Aldosterone induced aortic and small artery remodeling, impaired endothelium-dependent relaxation in WT mice, and enhanced fibronectin and collagen deposition and vascular inflammation. None of these vascular effects were observed in Agtr1a −/− mice. Aldosterone effects were prevented by the AGTR1 antagonist losartan in WT mice. In contrast to aldosterone, norepinephrine caused similar BP increase and mesenteric artery remodeling in WT and Agtr1a −/− mice. Agtr1a −/− mice infused with aldosterone did not increase sodium excretion in response to a sodium chloride challenge, suggesting that sodium retention could contribute to the exaggerated BP rise induced by aldosterone. Agtr1a −/− mice had decreased mesenteric artery expression of the calcium-activated potassium channel Kcnmb1 , which may enhance myogenic tone and together with sodium retention, exacerbate BP responses to aldosterone/salt in Agtr1a −/− mice. We conclude that although aldosterone activation of mineralocorticoid receptors raises BP more in Agtr1a −/− mice, AGTR1a is required for mineralocorticoid receptor stimulation to induce vascular remodeling and inflammation and endothelial dysfunction.

Primary autonomic failure is characterized by disabling orthostatic hypotension, but at least half of these patients have paradoxical supine hypertension. Renin–angiotensin mechanisms were not initially thought to contribute to this hypertension because plasma renin activity is often undetectable in autonomic failure. Plasma aldosterone levels are normal, however, and we recently showed that plasma angiotensin II is elevated and acts at AT 1 (angiotensin type 1) receptors to contribute to hypertension in these patients. Because aldosterone and angiotensin II can also bind mineralocorticoid receptors to elevate blood pressure, we hypothesized that mineralocorticoid receptor activation plays a role in the hypertension of autonomic failure. To test this hypothesis, we determined the acute effects of the mineralocorticoid receptor antagonist eplerenone (50 mg, oral) versus placebo on supine blood pressure in a randomized, double-blind, crossover study. Medications were given at 8:00 pm with blood pressure recorded every 2 hours for 12 hours. Ten primary autonomic failure patients with supine hypertension completed this study (7 pure autonomic failure, 2 multiple system atrophy, 1 parkinson’s disease; 7 male; 70±2 years of age). Eplerenone maximally reduced supine systolic blood pressure by 32±6 mm Hg at 8 hours after administration (versus 8±10 mm Hg placebo, P =0.016), with no effect on nocturia (12-hour urine volume: 985±134 mL placebo versus 931±94 mL eplerenone, P =0.492; nocturnal weight loss: −1.19±0.15 kg placebo versus −1.18±0.15 kg eplerenone, P =0.766). These findings suggest that inappropriate mineralocorticoid receptor activation contributes to the hypertension of autonomic failure, likely independent of canonical mineralocorticoid effects, and provides rationale for use of eplerenone in these patients.

Loss of mineralocorticoid receptor signaling selectively in cardiomyocytes can ameliorate cardiac fibrotic and inflammatory responses caused by excess mineralocorticoids. The aim of this study was to characterize the role of cardiomyocyte mineralocorticoid receptor signaling in ischemia–reperfusion injury and recovery and to identify a role of mineralocorticoid receptor modulation of cardiac function. Wild-type and cardiomyocyte mineralocorticoid receptor knockout mice (8 weeks) were uninephrectomized and maintained on (1) high salt (0.9% NaCl, 0.4% KCl) or (2) high salt plus deoxycorticosterone pellet (0.3 mg/d, 0.9% NaCl, 0.4% KCl). After 8 weeks of treatment, hearts were isolated and subjected to 20 minutes of global ischemia plus 45 minutes of reperfusion. Mineralocorticoid excess increased peak contracture during ischemia regardless of genotype. Recovery of left ventricular developed pressure and rates of contraction and relaxation post ischemia–reperfusion were greater in knockout versus wild-type hearts. The incidence of arrhythmic activity during early reperfusion was significantly higher in wild-type than in knockout hearts. Levels of autophosphorylated Ca 2+ /calmodulin protein kinase II (Thr287) were elevated in hearts from wild-type versus knockout mice and associated with increased sodium hydrogen exchanger-1 expression. These findings demonstrate that cardiomyocyte-specific mineralocorticoid receptor–dependent signaling contributes to electromechanical vulnerability in acute ischemia–reperfusion via a mechanism involving Ca 2+ /calmodulin protein kinase II activation in association with upstream alteration in expression regulation of the sodium hydrogen exchanger-1.

Cardiac and renal diseases remain major challenges for healthcare systems in developed countries and commonly coexist, with a disorder in the heart or kidney often leading to secondary dysfunction or injury in the other organ. Indeed, the term cardiorenal syndrome has been introduced to describe the broad spectrum of disease involving both the heart and kidneys.1 Aldosterone is a steroid hormone with mineralocorticoid activity, produced primarily in the glomerular zone of the adrenal cortex.2 Aldosterone fulfills its major physiological function of maintaining sodium and potassium balance and blood pressure control by binding to the mineralocorticoid receptor (MR) in the connecting tubule and cortical collecting duct in the kidneys, thereby increasing sodium reabsorption and potassium secretion. There is a growing body of evidence for a broader role of aldosterone and MR activation in the pathophysiology of cardiovascular and renal disease.3,4 This article focuses on our knowledge and understanding of the direct roles of aldosterone and MR activation in the heart and kidneys, including common pathophysiological mechanisms in both organs and implications for clinical use of MR antagonists (MRAs) in the treatment of cardiac and renal diseases. MR expression has been demonstrated in vivo in vascular endothelial cells and vascular smooth muscle cells of interlobar arteries in mouse kidneys and ex vivo in cultured podocytes, mesangial cells, and renal fibroblasts.3 The MR is also expressed in multiple cell types in the heart, including cardiomyocytes, coronary endothelial and vascular smooth muscle cells, fibroblasts, and inflammatory cells, such as macrophages.5 Aldosterone and the glucocorticoid cortisol bind to the MR with similar affinities. Plasma concentrations of glucocorticoids are 100- to 1000-fold higher than those of aldosterone. Overstimulation of the MR is prevented by the coexpression of 11β-hydroxysteroid dehydrogenase type 2 (11-BHSD2). This enzyme converts cortisol into cortisone, which …

The inability of mineralocorticoid receptor (MR) blockade to reduce hypertension associated with high angiotensin (Ang) II suggests direct actions of Ang II to regulate tubular sodium reabsorption via the epithelial Na + channel (ENaC) in the aldosterone-sensitive distal nephron. We used freshly isolated aldosterone-sensitive distal nephron from mice to delineate the synergism and primacy between aldosterone and Ang II in controlling functional ENaC activity. Inhibition of MR specifically prevented the increased number of functionally active ENaC, but not ENaC open probability elicited by a low sodium diet. In contrast, we found no functional role of glucocorticoid receptors in the regulation of ENaC activity by dietary salt intake. Simultaneous inhibition of MR and Ang II type 1 receptors ameliorated the enhanced ENaC activity caused by low dietary salt intake and produced significantly greater natriuresis than either inhibitor alone. Chronic systemic Ang II infusion induced more than 2 times greater increase in ENaC activity than observed during dietary sodium restriction. Importantly, ENaC activity remained greatly above control levels during maximal MR inhibition. We conclude that during variations in dietary salt intake both aldosterone and Ang II contribute complementarily to the regulation of ENaC activity in the aldosterone-sensitive distal nephron. In contrast, in the setting of Ang II–dependent hypertension, ENaC activity is upregulated well above the physiological range and is not effectively suppressed by inhibition of the aldosterone–MR axis. This provides a mechanistic explanation for the resistance to MR inhibition that occurs in hypertensive subjects having elevated intrarenal Ang II levels.

Although elevated renin–angiotensin system activity and angiotensinergic signaling within the brain are required for hypertension, polydipsia, and increased metabolic rate induced by deoxycorticosterone acetate (DOCA)-salt, the contribution of specific receptor subtypes and brain nuclei mediating these responses remains poorly defined. We hypothesized that angiotensin type 1a receptors (AT 1a R) within the subfornical organ (SFO) mediate these responses. Transgenic mice carrying a conditional allele of the endogenous AT 1a R (AT 1a R flox ) were administered an adenovirus encoding Cre-recombinase and enhanced green fluorescent protein (eGFP) or adenovirus encoding eGFP alone into the lateral cerebral ventricle. Adenovirus encoding Cre-recombinase reduced AT 1a R mRNA and induced recombination in AT 1a R flox genomic DNA specifically in the SFO, without significant effect in the paraventricular or arcuate nuclei, and also induced SFO-specific recombination in ROSA TdTomato reporter mice. The effect of SFO-targeted ablation of endogenous AT 1a R was evaluated in AT 1a R flox mice at 3 time points: (1) baseline, (2) 1 week after virus injection but before DOCA-salt, and (3) after 3 weeks of DOCA-salt. DOCA-salt–treated mice with deletion of AT 1a R in SFO exhibited a blunted increase in arterial pressure. Increased sympathetic cardiac modulation and urine copeptin, a marker of vasopressin release, were both significantly reduced in DOCA-salt mice when AT 1a R was deleted in the SFO. Additionally, deletion of AT 1a R in the SFO significantly attenuated the polydipsia, polyuria, and sodium intake in response to DOCA-salt. Together, these data highlight the contribution of AT 1a R in the SFO to arterial pressure regulation potentially through changes on sympathetic cardiac modulation, vasopressin release, and hydromineral balance in the DOCA-salt model of hypertension.

High serum levels of aldosterone have been linked to the development of cardiac disease. In contrast, angiotensin (Ang)-(1-7) was extensively shown to possess cardioprotective effects, including the attenuation of cardiac dysfunction induced by excessive mineralocorticoid activation in vivo, suggesting possible interactions between these 2 molecules. Here, we investigated whether there is cross-talk between aldosterone and Ang-(1-7) and its functional consequences for calcium (Ca 2+ ) signaling in ventricular myocytes. Short-term effects of aldosterone on Ca 2+ transient were assessed in Fluo-4/AM-loaded myocytes. Confocal images showed that Ang-(1-7) had no effect on Ca 2+ transient parameters, whereas aldosterone increased the magnitude of the Ca 2+ transient. Quite unexpectedly, addition of Ang-(1-7) to aldosterone-treated myocytes further enhanced the amplitude of the Ca 2+ transient suggesting a synergistic effect of these molecules. Aldosterone action on Ca 2+ transient amplitude was mediated by protein kinase A, and was related to an increase in Ca 2+ current ( I Ca ) density. Both changes were not altered by Ang-(1-7). When cardiomyocytes were exposed to aldosterone, increased Ca 2+ spark rate was measured. Ang-(1-7) prevented this change. In addition, a NO synthase inhibitor restored the effect of aldosterone on Ca 2+ spark rate in Ang-(1-7)-treated myocytes and attenuated the synergistic effect of these 2 molecules on Ca 2+ transient. These results indicate that NO plays an important role in this cross-talk. Our results bring new perspectives in the understanding of how 2 prominent molecules with supposedly antagonist cardiac actions cross-talk to synergistically amplify Ca 2+ signals in cardiomyocytes.

The circadian clock is important for maintaining rhythms in physiological functions including blood pressure (BP). Circadian disruption leads to increased disease risk. The clock has also been implicated in the maintenance of a normal dip in BP at night. In humans, non-dipping (night/day difference in BP

We have documented recently that bradykinin (BK) directly inhibits activity of the epithelial Na + channel (ENaC) via the bradykinin B2 receptor (B2R)-G q/11 -phospholipase C pathway. In this study, we took advantage of mice genetically engineered to lack bradykinin receptors (B1R, B2R −/− ) to probe a physiological role of BK cascade in regulation of ENaC in native tissue, aldosterone-sensitive distal nephron. Under normal sodium intake (0.32% Na + ), ENaC open probability ( P o ) was modestly elevated in B1R, B2R −/− mice compared with wild-type mice. This difference is augmented during elevated Na + intake (2.00% Na + ) and negated during Na + restriction (+ ). Saturation of systemic mineralocorticoid status with deoxycorticosterone acetate similarly increased ENaC activity in both mouse strains, suggesting that the effect of BK on ENaC is independent of aldosterone. It is accepted that angiotensin-converting enzyme represents the major pathway of BK degradation. Systemic inhibition of angiotensin-converting enzyme with captopril (30 mg/kg of body weight for 7 days) significantly decreases ENaC activity and P o in wild-type mice, but this effect is diminished in B1R, B2R −/− mice. At the cellular level, acute captopril (100 μmol/L) treatment sensitized BK signaling cascade and greatly potentiated the inhibitory effect of 100 nmol/L of BK on ENaC. We concluded that BK cascade has its own specific role in blunting ENaC activity, particularly under conditions of elevated sodium intake. Augmentation of BK signaling in the aldosterone-sensitive distal nephron inhibits ENaC-mediated Na + reabsorption, contributing to the natriuretic and antihypertensive effects of angiotensin-converting enzyme inhibition.

The renin-angiotensin-aldosterone system is involved in the arterial hypertension-associated cardiovascular remodeling. In this context, the development of cardiac fibrosis results from an imbalance between profibrotic and antifibrotic pathways, in which the role of aldosterone is yet not established. To determine the role of intracardiac aldosterone in the development of myocardial fibrosis during hypertension, we used a double transgenic model (AS-Ren) of cardiac hyperaldosteronism (AS) and systemic hypertension (Ren). The 9-month–old hypertensive mice had cardiac fibrosis, and hyperaldosteronism enhanced the fibrotic level. The mRNA levels of connective tissue growth factor and transforming growth factor-β1 were similarly increased in Ren and AS-Ren mice compared with wild-type and AS mice, respectively. Hyperaldosteronism combined with hypertension favored the macrophage infiltration (CD68 + cells) in heart, and enhanced the mRNA level of monocyte chemoattractant protein 1, osteopontin, and galectin 3. Interestingly, in AS-Ren mice the hypertension-induced increase in bone morphogenetic protein 4 mRNA and protein levels was significantly inhibited, and B-type natriuretic peptide expression was blunted. The mineralocorticoid receptor antagonist eplerenone restored B-type natriuretic peptide and bone morphogenetic protein 4 levels and decreased CD68 and galectin 3 levels in AS-Ren mice. Finally, when hypertension was induced by angiotensin II infusion in wild-type and AS mice, the mRNA profiles did not differ from those observed in Ren and AS-Ren mice, respectively. The aldosterone-induced inhibition of B-type natriuretic peptide and bone morphogenetic protein 4 expression was confirmed in vitro in neonatal mouse cardiomyocytes. Altogether, we demonstrate that, at the cardiac level, hyperaldosteronism worsens hypertension-induced fibrosis through 2 mineralocorticoid receptor-dependent mechanisms, activation of inflammation/galectin 3–induced fibrosis and inhibition of antifibrotic factors (B-type natriuretic peptide and bone morphogenetic protein 4).

Mineralocorticoid receptor (MR) activation may be deleterious to the cardiovascular system, and MR antagonists improve morbidity and mortality of patients with heart failure. However, mineralocorticoid signaling in the heart remains largely unknown. Using a pan-genomic transcriptomic analysis, we identified neutrophil gelatinase-associated lipocalin (NGAL or lipocalin 2) as a strongly induced gene in the heart of mice with conditional and targeted MR overexpression in cardiomyocytes (whereas induction was low in glucocorticoid receptor–overexpressing mice). NGAL mRNA levels were enhanced after hormonal stimulation by the MR ligand aldosterone in cultured cardiac cells and in the heart of wild-type mice. Mineralocorticoid pathological challenge induced by nephrectomy/aldosterone/salt treatment upregulated NGAL expression in the heart and aorta and its plasma levels. We show evidence for MR binding to an NGAL promoter, providing a mechanism for NGAL regulation. We propose that NGAL may be a marker of mineralocorticoid-dependent injury in the cardiovascular system in mice.

Overactivation of the mineralocorticoid receptor signaling is implicated in cardiovascular disease, including hypertensive heart disease. Oxidative stress is suggested to augment mineralocorticoid receptor signal transduction, but the precise mechanisms remain unclear. Mineralocorticoid receptor activity is regulated by multiple factors, in addition to plasma ligand levels. We previously identified Rac1 GTPase as a modulator of mineralocorticoid receptor activity. Here we show that oxidative stress induces mineralocorticoid receptor activation in a ligand-independent, Rac1-depenent manner in cardiomyocytes. Oxidant stress was induced in rat cultured cardiomyocytes (H9c2) by l -buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis. BSO depleted intracellular glutathione and concomitantly increased reactive oxygen species (199%; P P N -acetylcysteine. The ligand independency of BSO action was indicated using a mutant mineralocorticoid receptor that does not bind ligands. With this mutant mineralocorticoid receptor, BSO-evoked mineralocorticoid receptor activation remained intact, whereas ligand-induced mineralocorticoid receptor activation was abolished. We next examined the involvement of Rac1. BSO increased active Rac1 in a redox-dependent fashion, and Rac inhibition suppressed the enhancing effect of BSO. Constitutively active Rac1, indeed, potentiated mineralocorticoid receptor transactivation. Furthermore, mineralocorticoid receptor transactivation by BSO was accompanied by enhanced nuclear accumulation of mineralocorticoid receptor. We conclude that alteration of redox state modulates mineralocorticoid receptor–dependent transcriptional activity via Rac1 in the heart. This redox-sensitive, ligand-independent mineralocorticoid receptor activation may contribute to the processes by which oxidant stress promotes cardiac injury.

The purpose of the present article is to provide an overview of plasma membrane steroid hormone receptors and their implications in nongenomic signaling. We especially focus on recent evidence supporting the notion of a possible membrane-associated aldosterone receptor, whether this receptor is different from the classic nuclear receptor, and the possible implications of such a receptor for nongenomic and genomic aldosterone effects in physiological and pathophysiological processes.

Antagonists of the mineralocorticoid receptor improve morbidity and mortality in patients with severe heart failure. However, the cell types involved in these beneficial effects are only partially known. The aim of this work was to evaluate whether genetic deletion of mineralocorticoid receptors in mouse cardiomyocytes or fibroblasts in vivo is cardioprotective after chronic left ventricular pressure overload. After transverse aortic constriction, mice deficient in myocyte mineralocorticoid receptors but not those deficient in fibroblast mineralocorticoid receptors were protected from left ventricular dilatation and dysfunction. After pressure overload, left ventricular ejection fraction was significantly higher in mice lacking myocyte mineralocorticoid receptors (70.2±4.4%) as compared with control mice (54.3±2.5%; P

Salt sensitivity of blood pressure is an independent risk factor for cardiovascular morbidity. Mechanistically, abnormal mineralocorticoid action and subclinical renal impairment may blunt the natriuretic response to high sodium intake, causing blood pressure to rise. 11β-Hydroxysteroid dehydrogenase type 2 (11βHSD2) controls ligand access to the mineralocorticoid receptor, and ablation of the enzyme causes severe hypertension. Polymorphisms in HSD11B2 are associated with salt sensitivity of blood pressure in normotensives. In this study, we used mice heterozygote for a null mutation in Hsd11b2 ( Hsd11b2 +/− ) to define the mechanisms linking reduced enzyme activity to salt sensitivity of blood pressure. A high-sodium diet caused a rapid and sustained increase in blood pressure in Hsd11b2 +/− mice but not in wild-type littermates. During the adaptation to high-sodium diet, heterozygotes displayed impaired sodium excretion, a transient positive sodium balance, and hypokalemia. After 21 days of high-sodium feeding, Hsd11b2 +/− mice had an increased heart weight. Mineralocorticoid receptor antagonism partially prevented the increase in heart weight but not the increase in blood pressure. Glucocorticoid receptor antagonism prevented the rise in blood pressure. In Hsd11b2 +/− mice, high-sodium feeding caused suppression of aldosterone and a moderate but sustained increase in corticosterone. This study demonstrates an inverse relationship among 11βHSD2 activity, heart weight, and blood pressure in a clinically important context. Reduced activity causes salt sensitivity of blood pressure, but this does not reflect illicit activation of mineralocorticoid receptors by glucocorticoids. Instead, we have identified a novel interaction among 11βHSD2, dietary salt, and circulating glucocorticoids.

See related article, pp 1033–1040 Clinically, mineralocorticoid receptor (MR) antagonists are widely prescribed for the treatment of hypertension and heart failure because of their diuretic action in aldosterone-sensitive distal nephron. Clinical trials of MR antagonism in patients with various degrees of heart failure severity have also demonstrated a pronounced reduction of cardiovascular mortality in MR antagonist–treated patients.1 The underlying mechanisms of these cardiovascular benefits are still debated and are probably diverse. Potential beneficial effects of MR antagonism on extracellular matrix remodeling, arrhythmia susceptibility, coronary flow reserve, cardiovascular inflammation, and vascular function have all been suggested. Indeed, the vasculature has been recently highlighted as a primary target of aldosterone and MR antagonists. MR is expressed in human vascular endothelial cells (ECs) and smooth muscle cells as is the 11β-HSD2 enzyme that allows for selective aldosterone versus cortisol activation of MR. The effects of MR activation on vascular reactivity in healthy humans remains controversial because of conflicting results from clinical studies with many demonstrating a constrictive response and some showing vascular relaxation.2 Discrepancies may be due to differences in the vascular health of study participants as well as differences in study design. However, when patients with underlying cardiovascular diseases are studied, the data are consistent with MR activation promoting increased systemic vascular resistance and reduced forearm blood flow and MR antagonism improving endothelium-dependent vasodilatation, independent of changes in blood pressure. The aggregate of data supports that, in healthy vessels, acute MR activation may evoke endothelium-dependent, NO-mediated vasodilatation, whereas, in the presence of endothelial dysfunction, vascular injury, or high vascular oxidative stress (as in patients with cardiovascular risk factors or heart …

International audience; Primary aldosteronism is the most common form of secondary hypertension with hypokalemia and suppressed renin-angiotensin system caused by autonomous aldosterone production. Our aim was to compare zona glomerulosa (ZG) structure and function between control adrenals and the peritumoral tissue from patients operated on for aldosterone-producing adenoma. ZG morphology and CYP11B1, CYP11B2, and disabled 2 expression were studied in 15 control adrenals and 25 adrenals with aldosterone-producing adenoma. A transcriptome analysis was done using publicly available data sets. In control adrenals, ZG was discontinuous, and CYP11B2 expression was focal or partly continuous and localized to 3 structures, foci, megafoci, and aldosterone-producing cell clusters. CYP11B2 expression was restricted to a limited number of ZG cells expressing Dab2 but not CYP11B1; aldosterone-producing cell clusters were composed of cells with an intermediate phenotype expressing CYP11B2 but not disabled 2 or CYP11B1. In peritumoral tissue, large remodeling of the adrenal cortex was observed with increased nodulation and decreased vascularization that were not correlated with CYP11B2 expression. In 17 out of 25 adrenals, hyperplasia of adjacent ZG was observed with persistent expression of CYP11B2 that was extended to the entire ZG. In all of the adrenals from patients with aldosterone-producing adenoma, CYP11B2 expression was present in foci, megafoci, and aldosterone-producing cell clusters. Transcriptome profiling indicates a close relationship between peritumoral and control adrenal cortex. In conclusion, adrenal cortex remodeling, reduced vascularization, and ZG hyperplasia are major features of adrenals with aldosterone-producing adenoma. Transcriptional phenotyping is not in favor of this being an intermediate step toward the formation of aldosterone-producing adenoma.

Obese persons with metabolic syndrome often have associated with salt-sensitive hypertension, microalbuminuria, and cardiac dysfunction, and the plasma aldosterone level in one-third of metabolic syndrome patients is clearly elevated. Hyperaldosteronism, which may be caused at least partially by certain adipocyte-derived factors, contributes to the development of proteinuria in obese hypertensive rats, and salt loading aggravates the proteinuria and induces cardiac diastolic dysfunction because of inadequate suppression of plasma aldosterone level. However, mineralocorticoid receptor (MR) antagonists prevent salt-induced renal and cardiac damage, suggesting that aldosterone excess and a high-salt diet exert an unfavorable synergistic action on the kidney and heart. In Dahl salt-sensitive rats, however, despite appropriate suppression of plasma aldosterone with a high-salt diet, salt loading paradoxically activated renal MR signaling, and the renal injury was markedly prevented by MR antagonists. Accordingly, we discovered an alternative pathway of MR activation in which Rac1, a small GTP-binding protein, activates MRs. Salt loading activates renal Rac1 in Dahl salt-sensitive rats, and Rac1 in turn induces MR activation, which results in renal injury, and the renal injury has been found to be prevented by Rac1 inhibitors. Moreover, several metabolic syndrome-related factors induce Rac1 activation, and one of them, hyperglycemia, activates MRs via Rac1 activation. Consistent with this, Rac1 inhibitors attenuated the proteinuria and renal injury in obese hypertensive animals. Thus, both salt and obesity activate Rac1 and cause MR activation. Abnormal activation of the aldosterone/MR pathway plays a key role in the development of salt-sensitive hypertension and renal injury in metabolic syndrome.

Mineralocorticoid receptor (MR) activation by aldosterone controls salt homeostasis and inflammation in several tissues and cell types. Whether or not a functional MR exists in polymorphonuclear neutrophils is unknown. We investigated the hypothesis that aldosterone modulates inflammatory neutrophil responses via the MR. By flow cytometry, Western blot analysis, and microscopy, we found that neutrophils possess MR. Preincubation with aldosterone (10−11to 10−6M) dose-dependently inhibited nuclear factor κB activation in interleukin (IL)-8– and granulocyte/macrophage colony-stimulating factor–treated neutrophils on fibronectin by IκBα Western blotting, electrophoretic mobility shift assay, and RT-PCR for IκBα mRNA. Aldosterone had no effect on tumor necrosis factor α– and lipopolysaccharide-mediated nuclear factor κB activation or on IL-8– and granulocyte/macrophage colony-stimulating factor–induced extracellular signal-regulated kinase, p38 mitogen-activated protein kinase, or phosphatidylinositol 3-kinase/Akt activation. Spironolactone prevented nuclear factor κB inhibition, indicating an MR-specific aldosterone effect. By RT-PCR, we found that neutrophils have 11β-hydroxysteroid dehydrogenase. Tumor necrosis factor α, which is controlled by nuclear factor κB, increased in the cell supernatant with IL-8 treatment. Aldosterone completely prevented this effect. RT-PCR showed a strong tumor necrosis factor α mRNA increase with IL-8 that was blocked by aldosterone, excluding the possibility that the tumor necrosis factor α increase was merely a consequence of secretion. Finally, conditioned medium from IL-8–treated neutrophils increased intercellular adhesion molecule–1 expression on endothelial cells and subsequently the adhesion of IL-8–treated neutrophils to endothelial cells. These effects were reduced when conditioned medium from aldosterone-pretreated neutrophils was used, and spironolactone blocked the aldosterone effect. Our data indicate that a functional MR exists in neutrophils mediating antiinflammatory effects that are at work when neutrophils interact with endothelial cells. These data could be relevant to MR-blockade treatment protocols.

Myocardial ischemia-reperfusion leads to significant changes in redox state, decreased postischemic functional recovery, and cardiomyocyte apoptosis, with development and progression of heart failure. Ischemia-reperfusion in the isolated perfused rat heart has been used as a model of heart failure. Clinically, mineralocorticoid receptor blockade in heart failure decreases morbidity and mortality versus standard care alone. The effects of corticosteroids on infarct area and apoptosis were determined in rat hearts subjected to 30 minutes of ischemia and 2.5 hours of reperfusion. Both aldosterone and cortisol increased infarct area and apoptotic index, an effect half-maximal between 1 and 10 nM and reversed by spironolactone. Dexamethasone and mifepristone aggravated infarct area and apoptotic index, similarly reversed by spironolactone. Spironolactone alone reduced infarct area and apoptotic index below ischemia-reperfusion alone, in hearts from both intact and adrenalectomized rats. The present study shows that cardiac damage is aggravated by activation of mineralocorticoid receptors by aldosterone or cortisol or of glucocorticoid receptors by dexamethasone. Mifepristone unexpectedly acted as a glucocorticoid receptor agonist, for which there are several precedents. Spironolactone protected cardiomyocytes via inverse agonist activity at mineralocorticoid receptors, an effect near maximal at a relatively low dose (10 nM). Spironolactone acts not merely by excluding corticosteroids from mineralocorticoid receptors but as a protective inverse agonist at low concentration. Mineralocorticoid receptor antagonists may, thus, provide an additional therapeutic advantage in unstable angina and acute myocardial infarction.

Adrenal vein sampling is the gold standard for identification of surgically curable primary aldosteronism, but its accuracy might be hindered by blood dilution from accessory vein blood. We prospectively investigated the presence of accessory veins draining into adrenal veins and their effect on the selectivity index (SI) in 74 consecutive patients undergoing adrenal vein sampling. On the right side, the venous anatomic pattern could be conclusively determined in 91.8% of the cases: we detected hepatic accessory veins in 12.1%, no accessory veins in 42.4%, and renal capsular veins in 45.5%. On the left side there was a phrenico-adrenal trunk in 89.4% and renal capsular accessory veins in 10.6% of the cases. On both sides, renal capsular and phrenic accessory veins did not affect the SI. At variance, on the right side, hepatic accessory veins were associated with SI values ≈3-fold lower than that found when such accessory veins were absent (median: 3.10 [range: 0.80 to 84.2] versus median: 1.10 [range: 0.70 to 2.20]; P =0.01). However, superselective adrenal catheterization resulted into higher SI values (median: 23.88; range: 4.80 to 84.20) in these cases. Thus, hepatic accessory veins sharing egress into the inferior vena cava with the right adrenal vein occurred in ≈12% of the patients and imply a low SI, likely because of adrenal blood dilution by hepatic blood carrying a low cortisol concentration. In the presence of this anatomic variation, superselective catheterization of the right adrenal vein should be undertaken to determine the lateralization of aldosterone secretion.

Previous studies reported on the association of left ventricular mass index (LVMI) with urinary sodium or with circulating or urinary aldosterone. We investigated the independent associations of LVMI with the urinary excretion of both sodium and aldosterone. We randomly recruited 317 untreated subjects from a white population (45.1% women; mean age 48.2 years). Measurements included echocardiographic left ventricular (LV) properties, the 24-hour urinary excretion of sodium and aldosterone, plasma renin activity (PRA), and proximal (RNa prox ) and distal (RNa dist ) renal sodium reabsorption, assessed from the endogenous lithium clearance. In multivariable-adjusted models, we expressed changes in LVMI per 1-SD increase in the explanatory variables, while accounting for sex, age, systolic blood pressure, and the waist-to-hip ratio. LVMI increased independently with the urinary excretion of both sodium (+2.48 g/m 2 ; P =0.005) and aldosterone (+2.63 g/m 2 ; P =0.004). Higher sodium excretion was associated with increased mean wall thickness (MWT: +0.126 mm, P =0.054), but with no change in LV end-diastolic diameter (LVID: +0.12 mm, P =0.64). In contrast, higher aldosterone excretion was associated with higher LVID (+0.54 mm; P =0.017), but with no change in MWT (+0.070 mm; P =0.28). Higher RNa dist was associated with lower relative wall thickness (−0.81×10 −2 , P =0.017), because of opposite trends in LVID (+0.33 mm; P =0.13) and MWT (−0.130 mm; P =0.040). LVMI was not associated with PRA or RNa prox. In conclusion, LVMI independently increased with both urinary sodium and aldosterone excretion. Increased MWT explained the association of LVMI with urinary sodium and increased LVID the association of LVMI with urinary aldosterone.

Mineralocorticoid receptor blockade protects from angiotensin II–induced target-organ damage. 11β-Hydroxysteroid dehydrogenase type 2 protects the mineralocorticoid receptor from activation by glucocorticoids; however, high glucocorticoid concentrations and absent 11β-hydroxysteroid dehydrogenase type 2 in some tissues make glucocorticoids highly relevant mineralocorticoid receptor ligands. We investigated the effects of corticosterone (10 −6 to 10 −12 mol/L) on early vascular mineralocorticoid receptor signaling by Western blotting, confocal microscopy, and myography. Corticosterone initiated extracellular signal–regulated kinase 1/2 phosphorylation in rat vascular smooth muscle cells at ≥10 −11 mol/L doses. Protein synthesis inhibitors had no effect, indicating a nongenomic action. Corticosterone also stimulated c-Jun N-terminal kinase, p38, Src, and Akt phosphorylation at 15 minutes and enhanced angiotensin II–induced signaling at 5 minutes. A specific epidermal growth factor receptor blocker, AG1478, as well as the Src inhibitor PP2, markedly reduced corticosterone-induced extracellular signal–regulated kinase 1/2 phosphorylation, as did preincubation of cells with the mineralocorticoid receptor antagonist spironolactone. Silencing mineralocorticoid receptor with small interfering RNA abolished corticosterone-induced effects. Corticosterone (10 −9 mol/L) enhanced phenylephrine-induced contraction of intact aortic rings. These effects were dependent on the intact endothelium, mineralocorticoid receptor, and mitogen-activated protein kinase kinase 1/extracellular signal-regulated kinase signaling. We conclude that corticosterone induces rapid mineralocorticoid receptor signaling in vascular smooth muscle cells that involves mitogen-activated protein kinase kinase/extracellular signal–regulated kinase–dependent pathways. These new mineralocorticoid receptor–dependent signaling pathways suggest that glucocorticoids may contribute to vascular disease via mineralocorticoid receptor signaling, independent of circulating aldosterone.

The classic role of aldosterone is to regulate water and electrolyte balance and, therefore, blood pressure homeostasis.1 Apart from that, experimental studies have demonstrated that aldosterone induces structural and functional alterations in the heart, kidneys, and vessels with effects such as myocardial fibrosis, nephrosclerosis, vascular inflammation and remodeling, and disturbed fibrinolysis.2,3 This damage seems to be aldosterone mediated, and aldosterone blockade with mineralocorticoid receptor (MR) antagonists, such as spironolactone, may prevent the onset of these effects.4,5 On the other hand, it cannot completely be ruled out that potassium and high blood pressure also play additional key roles in this damage.6,7 This evidence has impressively been supported by clinical studies, such as the Randomized Aldactone Evaluation Study and the Eplerenone Post-Acute Myocardial Infarction Survival and Efficacy Study.8,9 For example, increased mortality in patients with chronic heart failure has been associated with elevated aldosterone plasma levels,10 and high circulating plasma aldosterone levels predict the clinical outcome in patients after myocardial infarction.11 Furthermore, primary aldosteronism (PA) has been demonstrated to enhance the risk of cardiovascular events12 and kidney disease.13 In summary, aldosterone is considered a cardiovascular risk factor, promoting disease processes such as cardiac fibrosis, nephrosclerosis, and arteriosclerosis,2,3,14 all of which are increased in patients with obesity and the metabolic syndrome.15,16 The term “metabolic syndrome” (MSyn) has evolved various definitions in recent times; most of the studies introduced here use slight modifications. Nevertheless, all of the definitions used have a common denominator, which is reflected in a definition by the American Heart Association/National Heart, Lung, and Blood Institute.17 According to this definition, the MSyn is considered as a constellation of interrelated risk factors of metabolic origin, including arterial hypertension, dyslipidemias, alterations in glucose homeostasis with type 2 diabetes mellitus, and abdominal …

Osteopontin (OPN), a proinflammatory cytokine, plays an important role in renal fibrosis. We reported that plasma OPN levels were higher in patients with primary aldosteronism than with essential hypertension. However, the regulatory mechanism of OPN by aldosterone remains unclear. Here, we report the transcriptional regulation of OPN expression by aldosterone and the functional effects of aldosterone-mediated OPN transcription in renal fibroblasts. Aldosterone induced OPN expression in a dose-dependent manner with significant responses at 10 nmol/L (1.6±0.2-fold of controls, P P P cis regulatory element (−2153 to −1758) in the OPN promoter that is responsive to aldosterone. This element contains an activator protein-1 (AP-1) and nuclear factor kappa B (NFκB) site. Electrophoretic mobility shift assays, supershift assays, and chromatin immunoprecipitation assays identified both AP-1 and NFκB as the DNA binding proteins induced by aldosterone with spironolactone inhibiting aldosterone-induced AP-1 or NFκB activity. OPN-siRNA inhibited completely the induction of cell proliferation, type I, III, and IV collagen synthesis by aldosterone. These results indicate that aldosterone induced MR-mediated OPN expression through AP-1 and NFκB activation and suggest that aldosterone plays an important role in renal fibrosis through the induction of OPN.

Experimental data indicate that the cardiorenal effects of aldosterone excess are dependent on concomitant high dietary salt intake. Such an interaction of endogenous aldosterone and dietary salt has not been observed previously in humans. We assessed the hypothesis that excess aldosterone and high dietary sodium intake combine to worsen proteinuria in patients with resistant hypertension. Consecutive subjects with resistant hypertension (n=84) were prospectively evaluated by measurement of 24-hour urinary aldosterone (Ualdo), sodium, and protein (Uprot) excretion. Subjects were analyzed according to aldosterone status (high: Ualdo ≥12 μg/24 hours; or normal: P =0.01). Among subjects with high Ualdo, Uprot increased progressively across urinary sodium groups ( P r =0.47; P =0.003) was observed in subjects with high Ualdo but not in subjects with normal Ualdo ( r =0.18; P value not significant). These results suggest that aldosterone excess and high dietary salt combine to increase urinary protein excretion.

Oxidative stress and inflammation contribute to increased cardiovascular morbidity and mortality associated with activation of the renin-angiotensin (Ang)-aldosterone system (RAAS). Studies in cultured cells in vitro and in rodent models in vivo demonstrate that aldosterone and/or mineralocorticoid receptor (MR) activation cause oxidative stress and vascular inflammation. Translational studies in humans suggest that endogenous aldosterone increases inflammatory biomarkers through an MR-dependent pathway. Clinical studies indicate that the prevalence of hyperaldosteronism may be increased in resistant hypertension,1 that aldosterone concentrations “escape” to pretreatment levels during chronic treatment of congestive heart failure or hypertension with an Ang-converting enzyme (ACE) inhibitor or Ang receptor blocker,2–4 and that MR antagonism decreases mortality in congestive heart failure.5,6 This article reviews the current literature regarding mechanism(s) of aldosterone-induced vascular inflammation and its implications for the prevention of vascular injury in humans. Oxidative stress and inflammation play central roles in the pathogenesis of atherosclerosis.7,8 Reactive oxygen species, including superoxide and hydrogen peroxide, modify vascular low-density lipoproteins to form oxidized low-density lipoproteins. Low-density lipoproteins stimulate endothelial cells to express leukocyte adhesion molecules, such as vascular cell adhesion molecule, leading to the recruitment of monocytes and T lymphocytes, which invade the intima. Monocyte-derived macrophages, rich in nicotinamide-adenine-dinucleotide phosphate (NADPH) oxidase, amplify the generation of reactant oxygen species. Macrophages take up oxidized low-density lipoproteins via scavenger receptors to become foam cells. T cells and macrophages within atherosclerotic lesions produce chemokines and cytokines resulting in the migration and proliferation of vascular smooth muscle cells (VSMCs) within the intima. VSMCs, in turn, promote extracellular matrix formation and secrete matrix metalloproteinases, contributing to the pathogenesis of plaque rupture. Inflammation also triggers a cascade that leads to fibrosis and vascular remodeling.9 During inflammation, the recruitment of monocytes and macrophages promotes a proliferation of fibroblasts in the perivascular space. Cytokines like …

Mineralocorticoid receptor (MR) blockade reduces morbidity and mortality after acute myocardial infarction; however, the underlying mechanisms are still under investigation. This study examined whether MR antagonism promotes healing of the infarcted myocardium. Starting immediately after coronary ligation, male Wistar rats were treated with the selective MR antagonist eplerenone (100 mg/kg per day by gavage) or placebo for 2 to 7 days. At 7 days, eplerenone therapy versus placebo significantly reduced thinning and dilatation of the infarcted wall, improved left ventricular function, and enhanced neovessel formation in the injured myocardium. At 2 days, eplerenone-treated rats displayed lower plasma corticosterone levels, higher circulating blood monocytes, and more macrophages infiltrating the infarcted myocardium. MR blockade led to a transient upregulation (at days 2 and 3 but not at day 7) of monocyte chemoattractant protein-1, tumor necrosis factor-α, interleukin-1β, interleukin-6, interleukin-10, and interleukin-4 and an increase in factor XIIIa protein expression in the healing myocardium. Prevention of macrophage accumulation into the infarct zone by treatment with liposome-encapsulated clodronate almost abrogated the protein expression of factor XIIIa and the beneficial effects of eplerenone on infarct expansion. In conclusion, selective MR blockade immediately after myocardial infarction accelerated macrophage infiltration and transiently increased the expression of healing promoting cytokines and factor XIIIa in the injured myocardium resulting in enhanced infarct neovascularization and reduced early LV dilation and dysfunction.

Aldosterone-producing adenomas (APAs) are a common cause of arterial hypertension, but the underlying molecular mechanisms are unknown, although a transcriptional modulation of aldosterone synthase (CYP11B2) has been suggested. Aldosterone synthesis involves 2 main rate-limiting steps: cholesterol transport into mitochondria and CYP11B2 gene transcription. Evidence supports a role of Ca 2+ /calmodulin-dependent protein kinases (CAMKs) in the regulation of angiotensin II– and potassium-stimulated aldosterone production. CAMK-I mediates CYP11B2 transcription via cAMP response element binding protein and activating transcription factor 1 transcription factors and nuclear receptor Nur-related factor 1. CAMK-II affects cholesterol transport into mitochondria by acting on steroidogenic acute regulatory protein and/or cytoskeleton proteins. We analyzed the whole transcriptome of APAs as compared with a pool of normal human adrenocortical tissues. Based on steroidogenic enzyme gene expression profiles, we identified 2 APA subgroups: 1 featuring overexpression of CYP11B2, CAMK-I, 11-β-hydroxylase, 3-β-hydroxysteroid dehydrogenase, and 21-hydroxylase and the underexpression of CAMK-IIB and the other one with an opposite profile. The low CYP11B2 group exhibited a longer known duration of hypertension and a lower rate of long-term cure. Thus, aldosterone overproduction in APAs involves complex alterations of aldosterone synthesis regulation rather than simply increased aldosterone synthase gene expression. Whether the molecular signature of APA carries prognostic information is worth further investigation.

Peroxisome proliferator-activated receptor-α is widely distributed in the vasculature where it is believed to exert pleiotropic antiatherogenic effects. Its role in the regulation of blood pressure is still unresolved; however, some evidence suggests that it may affect the renin-angiotensin system. We investigated its role in angiotensin II–induced hypertension in the Tsukuba hypertensive mouse (THM). This is a model of hypertension and atherosclerosis because of high angiotensin II and aldosterone levels as a result of the transgenic expression of the entire human renin-angiotensin system. Making the THM animals deficient in Peroxisome proliferator-activated receptor-α (THM/PPARKO) totally abolished hypertension and myocardial hypertrophy. This was accompanied by a reduction in plasma human active renin in THM/PPARKO mice compared with THM animals from 3525±128 mU/L to 1910±750 mU/L ( P P =0.003). In the THM/PPARKO mice, the extent of atherosclerosis at the aortic sinus after a 12-week period on an atherogenic diet was decreased by >80%. In addition, the spontaneous formation of foam cells from peritoneal macrophages, a blood pressure–independent event, was reduced by 92% in the THM/PPARKO mice, suggesting protection from the usual oxidative stress in these animals, possibly because of lower prevailing angiotensin II levels. Finally, chronic fenofibrate treatment further elevated blood pressure in THM animals but not in THM/PPARKO animals. Taken together, these data indicate that peroxisome proliferator-activated receptor-α may regulate the renin-angiotensin system. They raise the possibility that its activation may aggravate hypertension and hasten atherosclerosis in the context of an activated renin-angiotensin system.

Unlike healthy subjects, overt congestive heart failure cannot “escape” the sodium- and water-retaining actions of mineralocorticoid excess. It is undefined whether escape occurs in asymptomatic left ventricular dysfunction (ALVD), which is characterized by preserved sodium homeostasis, natriuretic peptide activation, and normal circulating aldosterone. We hypothesized that, in ALVD, mineralocorticoid excess with exogenous deoxycorticosterone acetate (DOCA) would overwhelm renal compensatory mechanisms, resulting in sodium and water retention, and promote renal and cardiac collagen deposition. ALVD was induced in 2 groups (N=5 each) of dogs by tachypacing at 180 bpm. Urine was collected daily and blood drawn at baseline and days 2, 5, 8, and 11. One group served as control (ALVD), and the other received DOCA (ALVD+DOCA) starting at day 2 of pacing. Urine flow and sodium excretion were unchanged in the ALVD group. In ALVD+DOCA, urine flow and sodium excretion decreased on the first 2 days DOCA was given but normalized starting day 4. Urine flow and urinary cGMP excretion increased in ALVD+DOCA after DOCA escape. Plasma atrial natriuretic peptide, B-type natriuretic peptide, and cGMP increased equally in both groups. There were no differences in cardiorenal and hemodynamic parameters in an acute study on day 11. Although renal collagen area fraction was similar, left ventricular collagen area fraction in ALVD+DOCA was significantly higher than in ALVD (3.3±0.4% versus 2.0±0.2%; P =0.012). We conclude that ALVD can escape the sodium- and water-retaining effects of mineralocorticoid excess. Despite renal escape, increased left ventricular collagen deposition suggests that the heart but not the kidney failed to escape the tissue effects of DOCA.

Aldosterone plays a major role in the maintenance of sodium, potassium, and acid-base balance through its effects on renal electrolyte excretion. This regulation is achieved through aldosterone-induced stimulation of Na+ absorption, K+ secretion, and H+ secretion by the distal nephron, particularly segments of the collecting duct.1–3 These classical actions are mediated through binding of aldosterone to the intracellular mineralocorticoid receptor (MR). The hormone-receptor complex translocates to the nucleus, where it promotes gene transcription and the production of proteins that modulate the expression and activity of the epithelial Na+ channel (ENaC) and other ion transport proteins.1,4–6 Regulatory actions of aldosterone via the MR play an important role in the normal maintenance of blood pressure but also have been implicated in the pathogenesis of hypertension and the progression of renal disease.6–9 In addition to their classical actions, aldosterone and other steroid hormones influence cell processes through nongenomic mechanisms.10,11 Nongenomic effects of aldosterone have been demonstrated in many different epithelial and nonepithelial tissues and are defined by (1) an insensitivity to inhibitors of transcription (actinomycin D) and translation (cycloheximide) and (2) a rapid time course (seconds to a few minutes) that is incompatible with gene regulation and de novo protein synthesis. A rapid onset of action is a sufficient but not necessary criterion for a nongenomic effect. Some nongenomic effects can occur with a slower time course. An additional feature often associated with nongenomic effects is that they are not blocked by spironolactone and/or other MR antagonists, consistent with mediation via a nonclassical aldosterone receptor.10,11 Although compelling evidence exists for rapid effects of aldosterone unrelated to the MR, a novel aldosterone receptor for nongenomic regulation has not been identified, and there is evidence that nongenomic actions of aldosterone can be mediated via the classical …

Aldosterone plays essential roles in body fluid and electrolyte homeostasis and blood pressure. However, the association between polymorphisms in the CYP11B2 gene and hypertension is controversial. We resequenced CYP11B1 and CYP11B2 and identified 35 polymorphisms in this region. We performed association studies between the plasma aldosterone concentration and 13 polymorphisms in this region in 1443 subjects. The subjects were all obtained from the Suita Cohort Study. Multiple regression analysis indicated that aldosterone levels were determined by renin activity, age, total cholesterol, and hematocrit. Residuals of the aldosterone levels after adjusting for these confounding factors were nominally associated with the T(−344)C ( P =0.0026), C(595)T ( P =0.0180), −(4837)C ( P =0.0310), and G(4936)A ( P =0.0498) polymorphisms. Only the T(−344)C polymorphism was significantly associated with the aldosterone level after a correction for multiple testing (Bonferroni). A significant interaction was observed between the T(−344)C polymorphism and renin activity in determining aldosterone levels. Moreover, a significant interaction was observed in 2063 subjects between urinary sodium excretion, which reflects sodium intake, and the T(−344)C polymorphism in determining systolic blood pressure. Only subjects with the TT genotype showed a positive correlation between urinary sodium excretion and systolic blood pressure. In vitro experiments confirmed the functional significance of this T(−344)C polymorphism in terms of angiotensin II reactivity. Thus, the T(−344)C polymorphism in CYP11B2 appears to affect salt sensitivity in Japanese and to have clinical significance.

We have determined that differences in expression of aldosterone synthase (AS) affect responses to a low-salt diet. In AS-null mice (AS −/− ), but not in wild-type, low salt significantly decreased plasma sodium and increased potassium. The increased urine volume (1.5×wild-type) and decreased urine osmolality (0.7×wild-type), present in AS −/− mice on normal salt, became more severe (2.3×wild-type and 0.5×wild-type) on low salt, but neither changed in wild-type. In both genotypes, plasma vasopressin was similar on normal and low salt, and desmopressin injection significantly increased urine osmolality. Renal mRNA levels for aquaporin 1 and 3 were unchanged by genotype or diet and epithelial sodium channel and Na + -K + -2Cl − -cotransporter by genotype. In AS −/− mice, aquaporin 2 mRNA increased on normal salt, whereas Na + Cl − -cotransporter and cortex K + channel mRNAs decreased on both diets. The low blood pressure of AS −/− mice was decreased further by low salt, despite additional increases in renin, intrarenal arterial wall thickness, and macula densa cyclogenase-2 mRNA. In AS +/− mice on normal salt, adrenal AS mRNA was slightly decreased (0.7×wild-type), but blood pressure was normal. On low salt, their blood pressure was less than wild-type (101±2 mm Hg versus 106±2 mm Hg), even though renin mRNA increased to 2×wild-type. We conclude that aldosterone is critical for urine concentration and maintenance of blood pressure and even a mild reduction of AS expression makes blood pressure sensitive to low salt, suggesting that genetic differences of AS levels in humans may influence how blood pressure responds to dietary salt.

Controversial results obtained from human and animal studies on the prevention of heart disease by estrogens and progestins warrant a better understanding of nuclear hormone receptor function and interaction. To address this issue and taking into account that effects of synthetic progestins are not only referable to action through the progesterone receptor but may also be mediated by other steroid receptors, we characterized cardiovascular function and inflammatory gene expression in aldosterone salt-treated rats on long-term administration of 17β-estradiol, medroxyprogesterone acetate, and drospirenone, a new progestogen exhibiting antimineralocorticoid activity. The complex pattern of cardiovascular injury in ovariectomized Wistar rats induced by chronic aldosterone infusion plus a high-salt diet was significantly attenuated in sham-ovariectomized rats and by coadministration of 17β-estradiol in ovariectomized animals after 8 weeks of continuous treatment. The beneficial role of 17β-estradiol on blood pressure, cardiac hypertrophy, vascular osteopontin expression, perivascular fibrosis, and impaired NO-dependent relaxation of isolated aortic rings was completely abrogated by coadministration of medroxyprogesterone acetate. In contrast, drospirenone was either neutral or additive to 17β-estradiol in protecting against aldosterone salt-induced cardiovascular injury and inflammation. The current results support the hypothesis of complex interactions among estrogen, progesterone, glucocorticoid, androgen, and mineralocorticoid receptor signaling in cardiovascular injury and inflammation. Novel progestins, such as drospirenone, confer superior effects compared with medroxyprogesterone acetate in a model of aldosterone-induced heart disease because of its antimineralocorticoid properties.

Aldosterone has been suggested recently to cause vascular injury by directly acting on the vasculature, in addition to causing injury by raising the blood pressure. Bone marrow–derived endothelial progenitor cells (EPCs) have been shown to exert an important role in the repair of the endothelium. In addition, cell-based therapy using EPCs is emerging as a novel therapeutic strategy for myocardial and peripheral vascular diseases. However, impaired formation and function of EPCs has been observed in patients with risk factors for cardiovascular diseases. We evaluated the possible effects of aldosterone on EPCs by examining the progenitor cell formation from bone marrow mononuclear cells ex vivo. Aldosterone (10 to 1000 nmol/L) reduced the formation of progenitor cells in a concentration-dependent manner. This effect of aldosterone was attenuated by cotreatment with spironolactone. Aldosterone reduced the mRNA levels of vascular endothelial growth factor (VEGF) receptor (VEGFR) 2 without having any effect on the production of VEGF or mRNA levels of VEGF and hepatocyte growth factor in the progenitor cells. However, the expression of stromal-derived growth factor 1 mRNA was paradoxically increased. Consistent with the downregulation of VEGFR-2, VEGF-induced phosphorylation of Akt was abolished in the progenitor cells after aldosterone treatment. N-acetylcysteine, an antioxidant, attenuated the inhibitory effects of aldosterone. These data indicate that aldosterone inhibits the formation of bone marrow–derived progenitor cells, at least partly, by attenuating VEGFR-2 expression and the subsequent Akt signaling. Reduction of aldosterone levels, blockade of mineralocorticoid receptor, and/or cotreatment with antioxidants may, therefore, enhance vascular regeneration by EPCs.

The aim of this study was to analyze the associations of plasma aldosterone and plasma renin activity with the metabolic syndrome and each of its components. We analyzed data from a family based study in the Seychelles made up of 356 participants (160 men and 196 women) from 69 families of African descent. In multivariable models, plasma aldosterone was associated positively ( P P P P

Recent clinical studies implicate proteinuria as a key prognostic factor for renal and cardiovascular complications in hypertensives. The pathogenesis of proteinuria in hypertension is, however, poorly elucidated. Podocytes constitute the final filtration barrier in the glomerulus, and their dysfunction may play a pivotal role in proteinuria. In the present study, we examined the involvement of podocyte injury in Dahl salt-hypertensive rats, an animal model prone to hypertensive glomerulosclerosis, and explored the effects of inhibition of aldosterone. Four-week–old Dahl salt-resistant and salt-sensitive rats were fed a 0.3% or 8.0% NaCl diet. Some salt-loaded Dahl salt-sensitive rats were treated with a selective aldosterone blocker eplerenone (1.25 mg/g diet) or hydralazine (0.5 mmol/L). After 6 weeks, salt-loaded Dahl salt-sensitive rats developed severe hypertension, proteinuria, and glomerulosclerosis. Immunostaining for nephrin, a constituent of slit diaphragm, was attenuated, whereas expressions of damaged podocyte markers desmin and B7-1 were upregulated in the glomeruli of salt-loaded Dahl salt-sensitive rats. Electron microscopic analysis revealed podocyte foot process effacement. Podocytes were already impaired at as early as 2 weeks of salt loading in Dahl salt-sensitive rats, when proteinuria was modestly increased. Both eplerenone and hydralazine partially reduced systemic blood pressure as measured by indirect and direct methods in salt-loaded Dahl salt-sensitive rats, but only eplerenone dramatically improved podocyte damage and retarded the progression of proteinuria and glomerulosclerosis. Our findings suggest that podocyte injury underlies the glomerulopathy of Dahl salt-hypertensive rats and that inhibition of aldosterone by eplerenone is protective against podocyte damage, proteinuria, and glomerulosclerosis in this hypertensive model.