Vascular Mineralocorticoid Receptor Research Articles

Aldosterone Acutely Enhances Coronary Artery Contractile Responsiveness without Inducing Endothelial Dysfunction

Inappropriate activation of the renin‐angiotensin‐aldosterone system (RAAS) contributes to increased contractility of the vasculature in many disease states. Activation of vascular mineralocorticoid receptors (MR) has recently been implicated as an important regulator of vascular function. In vascular smooth muscle, genomic mechanisms of MR activation have been implicated in vascular contractility, however, little is known regarding non‐genomic MR signaling as a modulator of vascular contractility. Based on available evidence, we hypothesized that acute aldosterone incubation of murine coronary arteries ex vivo would enhance coronary contractility via non‐genomic MR activation in smooth muscle cells (SMC). Wire myography experiments were conducted on isolated coronary arteries from C57BL/6J mice. Contractile responses to KCl (10mM–45mM) were determined by repeated measures design before and following one hour aldosterone treatment (10nM or 100nM). KCl induced dose‐dependent coronary vasoconstriction that was enhanced by aldosterone incubation in a dose dependent fashion. Vasoconstriction to 20mM KCl was doubled following incubation with 10nM aldosterone and increased >3‐fold following incubation with 100nM aldosterone. Pretreatment with the selective MR antagonist eplerenone (10μM) did not significantly reduce enhanced KCl vasoconstriction in response to 100nM aldosterone. Furthermore, endothelium‐dependent coronary vasodilation to acetylcholine was unchanged following one hour aldosterone incubation regardless of dose. In conclusion, acute aldosterone incubation enhances coronary vasoconstrictor responses to KCl that, at least in response to high dose aldosterone (100nM), is not dependent on MR activation. Importantly, this effect appears to be smooth muscle‐specific and not secondary to aldosterone‐induced endothelial dysfunction as acute aldosterone incubation did not impair endothelium‐dependent coronary vasodilation. These results highlight important vascular cell‐specific effects of aldosterone that may underlie enhanced coronary contractility in disease states associated with inappropriate RAAS activation.Support or Funding InformationFunded by VA CDA‐2 IK2 BX002030 and the American Physiological Society.

Deletion of mineralocorticoid receptors in smooth muscle cells blunts renal vascular resistance following acute cyclosporine administration

Calcineurin inhibitors such as cyclosporine A (CsA) are still commonly used after renal transplantation, despite CsA--induced nephrotoxicity (CIN), which is partly related to vasoactive mechanisms. The mineralocorticoid receptor (MR) is now recognized as a key player in the control of vascular tone, and both endothelial cell- and vascular smooth muscle cell (SMC)-MR modulate the vasoactive responses to vasodilators and vasoconstrictors. Here we tested whether vascular MR is involved in renal hemodynamic changes induced by CsA. The relative contribution of vascular MR in acute CsA treatment was evaluated using mouse models with targeted deletion of MR in endothelial cell or SMC. Results indicate that MR expressed in SMC, but not in endothelium, contributes to the increase of plasma urea and creatinine, the appearance of isometric tubular vacuolization, and overexpression of a kidney injury biomarker (neutrophil gelatinase--associated lipocalin) after CsA treatment. Inactivation of MR in SMC blunted CsA--induced phosphorylation of contractile proteins. Finally, the in vivo increase of renal vascular resistance induced by CsA was blunted when MR was deleted from SMC cells, and this was associated with decreased L-type Ca2D channel activity. Thus, our study provides new insights into the role of vascular MR in renal hemodynamics during acute CIN, and provides rationale for clinical studies of MR antagonism to manage the side effects of calcineurin inhibitors.

Abstract 133: Role of Vascular and Myeloid Mineralocorticoid Receptor in Renal Ischemia/reperfusion

Introduction: Renal ischemia/reperfusion (IR) is a major cause of acute kidney injury. It is associated with cardiac alterations and chronic kidney disease (CKD) development. We previously showed that mineralocorticoid receptor (MR) antagonism prevents the acute and chronic consequences of renal IR (Barrera-Chimal, Kidney Int, 2013 and JASN, 2015). However, whether the benefit of the MR antagonists is due to the blockade of the MR expressed in the vessels is unclear. Objective: To study the specific contribution of endothelial and smooth muscle cells MR in acute and chronic consequences of renal IR. Methods: To evaluate the contribution of vascular MR we generated two knockout (KO) mouse models. To allow MR inactivation in endothelial cells (MR endoKO mice), floxed MR mice (MR fl/fl ) were crossed with mice expressing the inducible Cre recombinase under the VEcadh promoter. To allow MR inactivation in vascular smooth muscle cells (MR SMCKO mice), MR fl/fl mice were crossed with mice expressing the inducible Cre recombinase under the SMA promoter. In these mice, sham surgery or bilateral renal IR for 20 min was performed in MR fl/fl and KO mice and the animals were studied at short term (24 h) and long term (30 days) after reperfusion. Results: In MR fl/fl mice, IR induced renal dysfunction (plasma creatinine raised from 8.9±0.3 in sham to 33.8±4.8 umol/L in IR), tubular injury and increased mRNA levels of kim-1 (400-fold) and NGAL (220-fold). The MR endoKO mice displayed similar alterations induced by IR as MR fl/fl mice. In contrast, after 24 h of renal IR, the MR SMCKO mice presented normal renal function (plasma creatinine was 9.6±0.7 and 14.0±1.9 umol/L in sham and IR, respectively), absence of histological alterations and reduced kim-1 and NGAL levels. After 30 days, the MR fl/fl mice developed CKD characterized by renal dysfunction (plasma creatinine from 10.5±0.1in sham to 15±0.8 umol/L in IR), tubule-interstitial fibrosis and increased mRNA levels of fibronectin and Galectin-3 (2-fold). The MR SMCKO mice developed similar alterations. Conclusion: We provide evidence that the deficiency of MR in the SMC protects against the development of acute kidney lesions induced by IR, however MR deficiency in SMC did not impact the appearance of CKD induced by IR.

Abstract 112: Endothelial Mineralocorticoid Receptor Knock Out Protects the Endothelium From Aldosterone-Mediated Vascular Stiffness

Background: There is accumulating evidence that increased levels of aldosterone (Aldo) and increased vascular mineralocorticoid receptor (MR) signaling increases vascular inflammation and oxidative stress leading to endothelial dysfunction and associated vascular stiffness. However, the specific role of endothelial cell (EC) MR activation in promotion of end stiffness in female mice has not been explored. Accordingly, we hypothesized that knocking out MR from ECs would attenuate the Aldo-induced endothelial dysfunction and vascular stiffness. Methods and Results: Twenty six week-old female ECMR knockout (ECMR-/-) and wild type (ECMR+/+) mice were infused with 250 μg/Kg/day Aldo for 3 weeks. To assess endothelial-dependent vasodilation, endothelial and aortic stiffness and blood pressure we utilized wire myography, atomic force microscopy (AFM), pulse wave velocity (PWV) (before and after Aldo perfusion) and tail cuff procedures. Aldo infusion did not increase BP or PWV and this was not affected by the presence or absence of ECMR. Aldo impaired endothelial-dependent vasodilation and increased EC stiffness 8.6 fold and these effects were mitigated in ECMR-/-. Aldo did not alter peri-aortic fibrosis by picrosirius red staining as measured by average gray scale intensities, nor did it cause medial thickening or aortic remodeling evaluated by the lumen to aortic wall ratio, in either Aldo-infused group. Moreover, levels of the oxidative marker, 3-nitrotyrosine (3-NT) did not differ in different compartments of the aortic wall in either Aldo treated group. Conclusion: ECMR protects the endothelium from aldo-mediated impaired vasodilation and endothelial cell stiffness, and this protection occurs without changes in BP, total aortic stiffness, or vascular remodeling.

Low-Dose Mineralocorticoid Receptor Blockade Prevents Western Diet-Induced Arterial Stiffening in Female Mice.

Women are especially predisposed to development of arterial stiffening secondary to obesity because of consumption of excessive calories. Enhanced activation of vascular mineralocorticoid receptors impairs insulin signaling, induces oxidative stress, inflammation, and maladaptive immune responses. We tested whether a subpressor dose of mineralocorticoid receptor antagonist, spironolactone (1 mg/kg per day) prevents aortic and femoral artery stiffening in female C57BL/6J mice fed a high-fat/high-sugar western diet (WD) for 4 months (ie, from 4-20 weeks of age). Aortic and femoral artery stiffness were assessed using ultrasound, pressurized vessel preparations, and atomic force microscopy. WD induced weight gain and insulin resistance compared with control diet-fed mice and these abnormalities were unaffected by spironolactone. Blood pressures and heart rates were normal and unaffected by diet or spironolactone. Spironolactone prevented WD-induced stiffening of aorta and femoral artery, as well as endothelial and vascular smooth muscle cells, within aortic explants. Spironolactone prevented WD-induced impaired aortic protein kinase B/endothelial nitric oxide synthase signaling, as well as impaired endothelium-dependent and endothelium-independent vasodilation. Spironolactone ameliorated WD-induced aortic medial thickening and fibrosis and the associated activation of the progrowth extracellular receptor kinase 1/2 pathway. Finally, preservation of normal arterial stiffness with spironolactone in WD-fed mice was associated with attenuated systemic and vascular inflammation and an anti-inflammatory shift in vascular immune cell marker genes. Low-dose spironolactone may represent a novel prevention strategy to attenuate vascular inflammation, oxidative stress, and growth pathway signaling and remodeling to prevent development of arterial stiffening secondary to consumption of a WD.

0227 : Mineralocorticoid receptor deficiency in smooth muscle cells protects against renal injury induced by ischemia/reperfusion

Renal ischemia/reperfusion (IR) is an important cause of acute kidney injury (AKI). AKI has been linked with progression to chronic kidney disease and development of cardiac alterations. Previous studies from our laboratory have shown that mineralocorticoid receptor (MR) antagonism with spironolactone prevents tubular injury and renal dysfunction induced by IR. The protection mechanisms remain unclear and whether the blockade of vascular MR is responsible for the protection conferred by MR antagonists in IR remains unexplored. To evaluate the specific contribution of vascular MR in the development of the kidney injury induced by IR. To investigate the contribution of vascular MR we generated two knockout (KO) mouse models. To allow MR inactivation in endothelial cells (MR endoKO mice), floxed MR mice (MR fl/fl ) were crossed with mice expressing the Cre recombinase under the Tie2cre promoter. To allow MR inactivation in smooth muscle cells (MR SMCKO mice), MRfl/fl mice were crossed with mice expressing the inducible Cre recombinase under the SMA promoter (MR SMCKO ). In these mice, sham surgery or bilateral renal IR for 20min was performed. The animals were studied 24 h after reperfusion. As markers of tubular injury, the mRNA levels of Kim-1 and NGAL were quantified. In MR fl/fl mice, IR induced renal dysfunction (plasma creatinine raised from 8.9±0.3 in sham to 33.8±4.8 μmol/L in IR), tubular injury and an increase in mRNA levels of Kim-1 (400-fold) and NGAL (200-fold). The MR endoKO mice displayed similar alterations induced by IR as MR fl/fl mice. In contrast, after renal IR, the MR SMCKO mice presented normal renal function (plasma creatinine was 9.6±0.7 and 14.0±1.9 μmol/L in sham and IR, respectively), absence of histological alterations and a significant reduction in the mRNA levels of Kim-1 and NGAL. We are now testing the effect of MR deficiency in smooth muscle cells in preventing the development of chronic alterations induced by IR. We show that the deficiency of the MR expressed in smooth muscle cells protects against the renal injury induced by IR. Our data provides further evidence to support the use MR antagonists as a novel therapeutic approach to prevent acute and chronic consequences of renal IR.

Diabetes impairs the vascular effects of aldosterone mediated by G protein-coupled estrogen receptor activation.

Aldosterone promotes non-genomic effects in endothelial and vascular smooth muscle cells via activation of mineralocorticoid receptors (MR) and G protein-coupled estrogen receptors (GPER). GPER activation is associated with beneficial/protective effects in the vasculature. Considering that vascular dysfunction plays a major role in diabetes-associated complications, we hypothesized that the beneficial effects mediated by vascular GPER activation, in response to aldosterone, are decreased in diabetes. Mesenteric resistance arteries from female, 14–16 weeks-old, control and diabetic (db/db) mice were used. Phenylephrine (PhE)-induced contractions were greater in arteries from db/db vs. control mice. Aldosterone (10 nM) increased maximal contractile responses to PhE in arteries from control mice, an effect elicited via activation of GPER. Although aldosterone did not increase PhE responses in arteries from db/db mice, blockade of GPER, and MR decreased PhE-induced contractile responses in db/db mesenteric arteries. Aldosterone also reduced the potency of acetylcholine (ACh)-induced relaxation in arteries from both control and db/db mice via MR-dependent mechanisms. GPER antagonism further decreased ACh-induced relaxation in the control group, but did not affect ACh responses in the diabetic group. Aldosterone increased extracellular signal-regulated kinase 1/2 phosphorylation in arteries from control and db/db mice by a GPER-dependent mechanism. GPER, but not MR, gene, and protein expression, determined by RT-PCR and immunoblotting/immunofluorescence assays, respectively, were increased in arteries from db/db mice vs. control arteries. These findings indicate that aldosterone activates both vascular MR and GPER and that the beneficial effects of GPER activation are decreased in arteries from diabetic animals. Our results further elucidate the mechanisms by which aldosterone influences vascular function and contributes to vascular dysfunction in diabetes. Financial Support: FAPESP, CNPq, and CAPES, Brazil.

Abstract 012: Mineralocorticoid Receptors Mediate Western Diet-Induced Macrophage Polarization and Aortic Stiffness

Mineralocorticoid receptors (MRs) are expressed in vascular smooth muscle cells (VSMCs), macrophages, and endothelial cells (ECs) and signaling through these receptors may play a key role in development of aortic stiffness. Accordingly, we hypothesized that these vascular MRs are involved in the western diet (WD) - induced macrophage polarization and aortic stiffness. The stiffness of primary cultured VSMC and tissue ECs or VSMCs from mice was measured using cell nano-indentation with atomic force microscopy (AFM). Both Ang II (100 nM) and aldosterone (10 nM) increased VSMC stiffness in vitro. Also, B57 mice fed a WD for 16 weeks increased EC and VSMC stiffness as determined by in vivo pulse wave velocity and ex vivo AFM techniques. The elevated aortic stiffness was accompanied by increased mRNA expression of M1 markers MCP-1, CD 86, and CD11b. However, treatment with low dose spironolactone (Sp) (1.0 mg/kg/day), a dose not affecting blood pressure, significantly attenuated WD-induced increases in stiffness in murine ECs and VSMCs. Sp increased M2 marker IL10 and the expression ratio of M2/M1 marker genes in aorta by real time PCR. Interestingly, mice with EC MR knockout did not develop WD- induced aortic vasodilation dysfunction under both Ach and SNP (10-9-10-4 mol/L) stimulation. Furthermore, low dose Sp inhibited WD- caused up-regulation of ERK 1/2 and down-regulation Akt/eNOS signaling pathways in the aorta. These findings support the notion that increased vascular and macrophage MR signaling play a key role in macrophage polarization and associated aortic stiffness that result from consuming a WD high in fructose and fat.

Is there a new dawn for selective mineralocorticoid receptor antagonism?

Aldosterone and the mineralocorticoid receptor contribute to resistant hypertension and cardiovascular mortality, and mineralocorticoid receptor antagonists effectively reduce these complications. Their use is limited in certain populations with a higher risk of hyperkalemia or renal dysfunction. This review will highlight recent developments in extra-renal mineralocorticoid receptor research and the development of novel mineralocorticoid receptor antagonists. Tissue-specific knockout-out models provide definitive evidence that the vascular mineralocorticoid receptor directly contributes to hypertension and vascular remodeling, independent of renal effects. Several nonsteroidal mineralocorticoid receptor antagonists are in preclinical development or early-stage clinical trials. Several nonsteroidal mineralocorticoid receptor antagonists have demonstrated preserved cardiovascular benefit with a reduced incidence of hyperkalemia in preclinical studies. Novel, potent nonsteroidal mineralocorticoid receptor antagonists are in development, although their effect on cardiovascular and adverse drug events requires further investigation.

Estrogen receptor inhibits mineralocorticoid receptor transcriptional regulatory function.

The steroid hormone aldosterone (aldo) contributes to cardiovascular disease in animal models and in humans. Aldo activates the mineralocorticoid receptor (MR), a hormone-activated transcription factor, and indeed, pharmacological MR inhibition improves cardiovascular outcomes. Because the incidence of cardiovascular disease is lower in premenopausal women, we hypothesized that estrogen (E2) signaling through the estrogen receptor (ER) may protect the vasculature by inhibiting the detrimental effects of aldo signaling through the MR. We demonstrate that E2-activated ER inhibits MR-mediated gene transcription from the mouse mammary tumor virus reporter in human embryonic kidney-293 cells. In contrast, aldo-activated MR does not affect ER-mediated gene transcription. The ERα N terminus (amino acids 1-253) containing part of the DNA-binding domain is sufficient to inhibit MR genomic function, although point mutations reveal that DNA binding, ligand-independent activation, and rapid nongenomic ERα signaling are not required for this effect. Furthermore, ERα and MR are part of a complex in cell lysates, with amino acids 1-233 of the ERα N terminus being sufficient to complex with the MR. Overall, the ability of ERα to inhibit MR-mediated gene transcription correlates with the ability of ERα segments to both localize to the nucleus and complex with the MR. In cultured vascular endothelial cells expressing ERα, E2 inhibits aldo induction of the vascular MR target gene intercellular adhesion molecule-1 (ICAM-1). ICAM-1 induction by endothelial MR is known to promote vascular inflammation that could contribute to the mechanism of aldo-induced atherosclerosis. E2 also inhibits aldo induction of ICAM-1 protein and prevents aldo-enhanced leukocyte adhesion to endothelial cells. These studies support a new model in which E2-activated ER in endothelial cells forms a complex with MR in the nucleus to modulate MR regulation of the proinflammatory gene ICAM-1. Estrogen inhibition of MR regulation of genes that contribute to cardiovascular disease may be a new mechanism by which premenopausal women are protected from cardiovascular disease.

0307 Role of vascular mineralocorticoid receptor in renal injury induced by ischemia/reperfusion

Acute kidney injury (AKI) is defined as an abrupt decrease (48h) in kidney function. One of the main causes of AKI is ischemia/reperfusion (I/R). AKI is related with high mortality, chronic kidney disease development and cardiac alterations like heart failure and arrhythmias. Mineralocorticoid receptor (MR) antagonism with spironolactone (Sp) prevents tubular injury and renal dysfunction induced by I/R in the rat. Although there is information supporting a role for aldosterone and MR in kidney injury, it remains unexplored the specific role of the MR expressed in the vasculature in mediating the deleterious effects of aldosterone during renal I/R. To study the effect of inducing I/R in mice lacking the MR specifically in the endothelial cells or in the vascular smooth muscle cells. To test if Sp is also able to prevent renal injury induced by I/R in the mice with the C57BL/6 background (same as MR KO mice) we included three groups of mice: 1) Sham, 2) I/R 20 min and 3) I/R 20 min + Sp pre-treatment. We analyzed the presence of renal dysfunction and inflammatory cytokines. In the MR KO mice, we will analyze the effect of MR deficiency after renal I/R in an acute phase (24h) and in chronic kidney disease development (after 4 weeks). In the acute studies the mechanisms that will be explored include: polarization of macrophages, endothelial injury and oxidative stress. In the chronic studies we will test if the wild type or MR knockout mice develop CKD as a consequence of renal I/R. Mice underwent renal I/R developed injury characterized by increased serum creatinine and urea levels, urinary Hsp72 and elevation in the mRNA of TNF-alpha and MCP-1 pro-inflammatory cytokines. These alterations were prevented by the Sp pre-treatment. The protective effect of Sp against renal I/R that was previously reported in the rat is also observed in the C57BL/6 mice and supports the study of the MR KO mice in the renal I/R setting.

0339: Does mineralocorticoid receptor antoganism could alleviate cyclosporin-induced nephrotoxicity in renal transplant recipients?

Cyclosporine-induced Nephrotoxicity (CIN) is a major adverse event but its pathophysiology remains unclear. Mineralocorticoid Receptor (MR) pharmacological antagonism prevents CIN in rats by modulating the expression of vasoactive factors. Our team has shown that MR is expressed in endothelial and vascular smooth muscle cells (VSMC). Therefore, genetic MR manipulations in endothelium or VSMC modify vascular function. Our working hypothesis is that vascular MR activation plays a key role in CIN. Female mice with low-salt diet were used: 1) Pharmacological approach: Control (Vehicle), Cyclosporine-A (CsA, 100 mg/kg/d) and CsA + Can (CsA + Canrenoate 30 mg/kg/d, MR antagonist); 2) Genetic approach: MR KO in VSMC (MRKO-VSMC) or in Endothelial Cells (MRKO-EC) treated or not with CsA. Body weight loss is greater in Cyclosporine-treated groups (p<0.05). Renal function is impaired (p<0.05) and CsA induces renal histological damages that are prevented by MR antagonism or by MR KO in VSMC but not in endothelial cells. Canrenoate and MR KO in VSMC also prevent Cyclosporine-induced renal expression (mRNA) of NGAL (Neutrophil Gelatinase Associated Lipocalin), a kidney damage marker. CsA induced NGAL expression in proximal tubules (immunohistochemistry); this effect is prevented by MR antagonism and MR KO in VSMC but not in endothelial cells. We show that MR antagonism has beneficial effect on Cyclosporine-induced renal damages that, at least partially, involve VSMC MR. The underlying cellular mechanisms are currently under investigation. A clinical trial testing the safety of MR antagonism (Eplerenone) in renal transplant recipients treated with Cyclosporine is currently ongoing.

Regulation of mineralocorticoid receptor expression by hypoxia and ischemia. (853.6)

Stroke is a leading cause of death and disability in the US. Cerebral ischemia causes a rapid increase in the circulating levels of the mineralocorticoid aldosterone. We have also shown that mineralocorticoid receptor (MR) antagonism at the time of a stroke reduces the damage caused by ischemia. We know little about how ischemia regulates MR expression. We hypothesized that MR expression is up regulated by hypoxia and that this translates to an increase in MR expression in the ischemic arteries after a stroke. To test the effect of hypoxia on MR expression aortic rings from adult Sprague Dawley rats were incubated under control (normoxic) or hypoxic (1% oxygen) conditions for three hours in the presence or absence of glucose (n=7‐8 in each group). qRT‐PCR was performed to measure MR mRNA expression using TAQMAN probes. Results are expressed as fold change from control. Hypoxia caused a marked increase in MR mRNA expression (1.01±0.06 vs 1.72±0.32, normoxic vs hypoxic p&lt;0.05). Concomitant hypoxia and glucose deprivation had similar effects as hypoxia alone (1.09±0.12 vs 1.84±0.31 normoxia‐glucose vs hypoxia‐glucose p&lt;0.03). To tests the effects of ischemia on vascular MR expression rats underwent a middle cerebral artery (MCA) occlusion (MCAO) for 1 hour followed by 47 hours of reperfusion. The MCA from the ischemic hemisphere was compared to the same artery from sham‐operated rats by qRT‐PCR (n=5‐6). Ischemia caused a marked increase in the MR mRNA expression (in fold change from sham: 1.03±0.11 vs 2.01±0.24 sham vs MCAO, P&lt;0.05). These data suggest that after a stroke the expression of the MR in the vasculature may increase and this may exacerbate the effects of the increase in aldosterone in the aftermath of an acute ischemic stroke.Grant Funding Source: AHA ‐ 13GRNT17210000

Aldosterone and vascular mineralocorticoid receptors: regulators of ion channels beyond the kidney.

Elevated levels of the steroid hormone aldosterone have been reported in populations at risk for cardiovascular disease and are associated with adverse cardiovascular events, such as myocardial infarction, stroke, and death.1–3 Clinical trials demonstrate that either inhibition of aldosterone production or antagonism of its receptor, the mineralocorticoid receptor (MR), significantly reduces cardiovascular ischemic events and mortality.4–8 Classically, aldosterone regulates blood pressure by binding to the MR in renal epithelial cells and increasing expression and activity of ion channels in the distal nephron resulting in sodium retention and blood pressure elevation.9 Accordingly, clinical trials of MR antagonists report decreases in blood pressure.4,5 However, the modest reductions in BP do not fully account for the cardioprotective effects of MR antagonism nor do renal electrolyte changes fully explain the antihypertensive effects of MR antagonists.10 For the past 2 decades, it has become clear that aldosterone and MR have extrarenal actions that may contribute to alterations in vascular function leading to the development and progression of cardiovascular disease. Exploration of the precise mechanisms by which aldosterone/MR regulates vascular function and promotes cardiovascular diseases remains a very active area of investigation and a potential avenue for novel pharmacological interventions. Because MR is a critical regulator of renal ion channels,9 and more recently has been implicated in ion channel regulation in the heart,11 we propose that MR may also regulate vascular ion channel expression and function, thereby contributing to aldosterone/MR-induced vascular dysfunction and cardiovascular disease. Vascular ion channels are critical to generation of vascular tone, vessel contraction and relaxation, and vascular stiffness (reviewed elsewhere)12–15 and contribute to vascular dysfunction associated with diseases, including atherosclerosis,16 diabetes mellitus,17–19 and hypertension.20,21 This review focuses on new …

Direct contribution of vascular mineralocorticoid receptors to blood pressure regulation

Hypertension is an extremely prevalent cardiovascular risk factor and current antihypertensive therapies do not adequately treat hypertension in many affected individuals. Thus, a better understanding of mechanisms of hypertension could lead to novel therapies. Mineralocorticoid receptors (MR) are known to regulate blood pressure by responding to aldosterone in the kidney to regulate sodium retention. Recent evidence supports a direct contribution of the vasculature to control of BP and suggests the possibility that MR antagonists may also lower blood pressure by acting on extrarenal MR. This review summarizes existing research considering the role of the vascular MR in regulating vasoreactivity and blood pressure. Multiple studies indicate a role for vascular MR in modulating vasoconstriction and vasorelaxation. Activation of MR in vascular endothelial and smooth muscle cells leads to increased reactive oxygen species production and decreased availability of nitric oxide, important regulators of vascular reactivity. Transgenic mouse models, including an endothelial MR overexpressing mouse and a smooth muscle cell-specific MR-knockout mouse, support a direct role for vascular MR in control of blood pressure. This new evidence demonstrating that vascular MR directly contribute to control of vasoreactivity and blood pressure supports vascular MR and the pathways they control as novel therapeutic targets to treat hypertension.

Vascular Mineralocorticoid Receptors Regulate Blood Pressure Effects on Myogenic Tone and Role in Aging

Effects of aldosterone on blood pressure control have classically been attributed to sodium retention through its action on the kidney. However, increasingly it has been appreciated that aldosterone exerts effects directly on the vasculature. Here, we comment on novel data published recently that sheds light on new mechanisms, whereby aldosterone acting through the mineralocorticoid receptor on blood vessels, in interaction with angiotensin II and via oxidative stress and calcium channels, may affect age-associated changes in myogenic tone and exert effects on blood pressure regulation. Aldosterone is produced by the adrenal glomerulosa in response to angiotensin (Ang) II, elevated serum potassium, and corticotropin. It acts on the kidney to increase sodium reabsorption in the distal nephron interacting with the mineralocorticoid receptor (MR) and by activation of the apical epithelial sodium channel.1 In humans, primary hyperaldosteronism clearly establishes the role that aldosterone has on blood pressure (BP) and hypertension.2 However, in both normotensive subjects and in essential hypertension, contribution of aldosterone to BP has been more difficult to establish. In the Framingham study, quartiles of plasma levels of aldosterone were directly associated with BP.3 A fraction of patients with essential hypertension shows mild elevation of aldosterone plasma concentrations.4 On the contrary, treatment with MR antagonists lowers BP in human patients with essential hypertension,5 and particularly drug-resistant hypertension.6 Because aldosterone synthase inhibitors have similar effects as MR blockade in humans with hypertension,7 it is likely that the effects reported for MR antagonism are mediated by blockade of the action of aldosterone. On which organs does aldosterone act to raise BP? The initial answer is that it may in large measure do so by increasing sodium and water retention …

Targeting activated mineralocorticoid receptor: Occam's razor revisited

Activation of mineralocorticoid receptors (MRs) classically has been associated with electrolyte transport, but we now know that MR activation can also lead to tissue inflammation and fibrosis. Aldosterone consistently activates MR, but under selected circumstances, endogenous glucocorticoids such as cortisol and corticosterone can also trigger MR. Tissue-specific safeguards such as the enzyme 11β-hydroxysteroid dehydrogenase limit glucocorticoid-induced MR activation, while the presence of reactive oxygen species may enhance the ability for glucocorticoid-induced MR activation even in the absence of aldosterone.

Direct regulation of blood pressure by smooth muscle cell mineralocorticoid receptors

Hypertension is a cardiovascular risk factor present in over two-thirds of people over age 60 in North America; elevated blood pressure correlates with increased risk of heart attack, stroke and progression to heart and kidney failure. Current therapies are insufficient to control blood pressure in almost half of these patients. The mineralocorticoid receptor (MR), acting in the kidney, is known to regulate blood pressure through aldosterone binding and stimulation of sodium retention. However, recent studies support the concept that the MR also has extrarenal actions and that defects in sodium handling alone do not fully explain the development of hypertension and associated cardiovascular mortality. We and others have identified functional MR in human vascular smooth muscle cells (SMCs), suggesting that vascular MR might directly regulate blood pressure. Here we show that mice with SMC-specific deficiency of the MR have decreased blood pressure as they age without defects in renal sodium handling or vascular structure. Aged mice lacking MR in SMCs (SMC-MR) have reduced vascular myogenic tone, agonist-dependent contraction and expression and activity of L-type calcium channels. Moreover, SMC-MR contributes to angiotensin II–induced vascular oxidative stress, vascular contraction and hypertension. This study identifies a new role for vascular MR in blood pressure control and in vascular aging and supports the emerging hypothesis that vascular tone contributes directly to systemic blood pressure.

Both estrogen receptor subtypes, ERα and ERβ, prevent aldosterone-induced oxidative stress in VSMC via increased NADPH bioavailability

Activation of vascular mineralocorticoid (MR) or estrogen receptors (ER) exerts opposing effects on vascular remodeling. As we have previously shown, activation of either estrogen receptor subtype, ERα or ERβ, is fully sufficient to attenuate vascular remodeling in aldosterone salt-treated rats. To further elucidate the underlying mechanism(s) we tested the hypothesis that ER and MR activation might differentially modulate vascular reactive oxygen species (ROS) generation. In support of this concept, aldosterone increased ROS generation in vascular smooth muscle cells as determined by quantitative dihydroethidium fluorescence microscopy. Co-treatment with the selective ERα agonist 16α-LE2, the selective ERβ agonist 8β-VE2 or the non-selective ER agonist 17β-estradiol (E2) significantly reduced aldosterone-induced ROS generation. The pure ER antagonist ICI 182,780 completely blocked these salutary effects of E2, 16α-LE2 and 8β-VE2. Activation of ERα or ERβ fully blocked the reduction of intracellular nicotinamide adenine dinucleotide phosphate (NADPH) levels observed in aldosterone treated vascular smooth muscle cells. Intracellular NADPH levels were closely associated with expression and activity of the NADPH generating enzyme glucose-6-phosphate dehydrogenase. In conclusion, estrogens attenuate the detrimental vascular effects of excessive MR activation at least in part by preventing the depletion of intracellular NADPH levels.

Placental growth factor mediates aldosterone-dependent vascular injury in mice

In clinical trials, aldosterone antagonists reduce cardiovascular ischemia and mortality by unknown mechanisms. Aldosterone is a steroid hormone that signals through renal mineralocorticoid receptors (MRs) to regulate blood pressure. MRs are expressed and regulate gene transcription in human vascular cells, suggesting that aldosterone might have direct vascular effects. Using gene expression profiling, we identify the pro-proliferative VEGF family member placental growth factor (PGF) as an aldosterone-regulated vascular MR target gene in mice and humans. Aldosterone-activated vascular MR stimulated Pgf gene transcription and increased PGF protein expression and secretion in the mouse vasculature. In mouse vessels with endothelial damage and human vessels from patients with atherosclerosis, aldosterone enhanced expression of PGF and its receptor, FMS-like tyrosine kinase 1 (Flt1). In atherosclerotic human vessels, MR antagonists inhibited PGF expression. In vivo, aldosterone infusion augmented vascular remodeling in mouse carotids following wire injury, an effect that was lost in Pgf-/- mice. In summary, we have identified PGF as what we believe to be a novel downstream target of vascular MR that mediates aldosterone augmentation of vascular injury. These findings suggest a non-renal mechanism for the vascular protective effects of aldosterone antagonists in humans and support targeting the vascular aldosterone/MR/PGF/Flt1 pathway as a therapeutic strategy for ischemic cardiovascular disease.