The Mineralocorticoid Receptor in Salt-Sensitive Hypertension and Renal Injury

Abstract

Hypertension and its comorbidities pose a major public health problem associated with disease-associated factors related to a modern lifestyle, such high salt intake or obesity. Accumulating evidence has demonstrated that aldosterone and its receptor, the mineralocorticoid receptor (MR), have crucial roles in the development of salt-sensitive hypertension and coexisting cardiovascular and renal injuries. Accordingly, clinical trials have repetitively shown the promising effects of MR blockers in these diseases. We and other researchers have identified novel mechanisms of MR activation involved in salt-sensitive hypertension and renal injury, including the obesity-derived overproduction of aldosterone and ligand-independent signaling. Moreover, recent advances in the analysis of cell-specific and context-dependent mechanisms of MR activation in various tissues—including a classic target of aldosterone, aldosterone-sensitive distal nephrons—are now providing new insights. In this review, we summarize recent updates to our understanding of aldosterone-MR signaling, focusing on its role in salt-sensitive hypertension and renal injury.

Aldosterone, secreted from adrenal glands by stimuli including angiotensin II (AngII) or hyperkalemia, binds to the mineralocorticoid receptor (MR) expressed on epithelial cells in the aldosterone-sensitive distal nephron (ASDN) to regulate sodium and potassium flux.^1–3 According to the theory that renal handling of sodium is a key determinant of fluid volume and BP control,^4,5 MR signaling in the connecting tubule and collecting duct is essential for the regulation of BP, especially in the development of salt-sensitive hypertension.⁶ Mutations in aldosterone synthase in the MR or in a major target of MR signaling, the epithelial sodium channel (ENaC), cause genetic disorders with hypertension or hypotension.⁷ Higher aldosterone levels within the physiologic range were associated with hypertension in a community-based study.⁸ Moreover, aldosterone administration in canines shifted the pressure-natriuresis curve, indicating salt-sensitive hypertension.⁹

In addition to the ASDN, MR is expressed in other tissues, including in the renal glomerulus,¹⁰ the myocardium,^11,12 the vasculature,^11,13,14 immune cells,^15,16 and the brain.¹² Accumulating evidence has shown that the aberrant activation of the MR is involved in the development of cardiovascular and renal injuries. Reports of clinical trials findings indicated that treatment with an MR blocker improved the prognosis of heart failure^17–20 and reduced proteinuria in patients with CKD.^21,22 Moreover, the pathogenicity of the aldosterone-MR system in hypertension and its comorbidities is associated with disease-associated conditions related to the modern lifestyle, such as obesity^23,24 and aging.²⁵

Given the benefit of MR blockers, investigation of the mechanism of MR activation may lead to the development of novel therapeutic strategies and targets. Previous studies reported that inappropriate aldosterone secretion^10,26–29 and ligand-independent mechanisms of MR activation^30–32 increased salt sensitivity of hypertension and end organ injuries. Moreover, recent studies demonstrated the tissue- or cell type–specific roles of MR activation in the myocardium,^33,34 in vasculature,^25,35–37 in immune cells,^15,16,38 and in a classic target, the ASDN.^39–43 These findings have updated our knowledge of the aldosterone-MR system and provided a new outlook for the use of MR blockers. Here, we summarize the recent research advances in the mechanism of MR activation in the development of salt-sensitive hypertension and renal injury.

Role of the Aldosterone-MR System in Metabolic Syndrome

Metabolic syndrome, a cluster of comorbidities that includes visceral obesity, hypertension, glucose intolerance, and dyslipidemia, is a major risk factor of cardiovascular and renal injury. Indeed, clinical studies have reported a higher incidence of CKD or proteinuria in individuals with metabolic syndrome than in those without it. Of note, hypertension and renal injury in obese patients are closely associated with high salt intake. Individuals with obesity reportedly showed greater salt-sensitivity of BP with shifted curve of pressure natriuresis, compared with lean individuals. Moreover, the shifted curve was normalized after weight-loss program, and resulted in BP reduction.⁴⁴ The relationship between salt intake and urinary albumin excretion was steeper in obese study participants than in lean ones, suggesting increased salt sensitivity of renal injury in obesity.⁴⁵

Multiple factors commonly observed in obese subjects, including compression of kidneys, hyperinsulinemia, hyperleptinemia, sympathetic overactivation, and activation of the renin-angiotensin-aldosterone system (RAAS), interdependently contribute to the impaired renal sodium handling and the resultant salt-sensitive hypertension in metabolic syndrome.⁴⁶ Clinical studies have shown relatively high plasma aldosterone with low plasma renin activity in individuals of African descent with metabolic syndrome,^47,48 indicating the role of aldosterone even in the presence of a blunted renin-angiotensin system (RAS). Moreover, primary aldosteronism⁴⁹ and a variant in the promoter region of aldosterone synthase (CYP11B2) that leads to hyperaldosteronemia⁵⁰ were associated with incident metabolic syndrome, suggesting an interaction between obesity and hyperaldosteronism.

Because excess aldosterone promotes salt-sensitive hypertension and end organ damage, we and others have explored its role in metabolic syndrome. We found in a study of obese SHR (SHR/NDmcr-cp) rats, a model of metabolic syndrome, that these animals had significantly greater proteinuria with podocyte damage compared with control lean SHR rats, despite both strains showing a similar BP.¹⁰ Obese SHR rats had a higher plasma aldosterone concentration compared with lean SHR rats, and aldosterone level was positively correlated with the amount of proteinuria. Accordingly, the expression of an MR target gene, serum and glucocorticoid-regulated kinase 1 (Sgk1), was increased in the glomerular fraction and a whole-kidney sample from obese SHR rats. Moreover, a selective MR blocker, eplerenone, reversed the proteinuria and podocyte injury in obese SHR rats, demonstrating the role of MR activation induced by aldosterone excess in renal injury in metabolic syndrome.

Of note, proteinuria and podocyte injury in obese SHR rats were markedly aggravated after salt loading.⁵¹ Because salt loading physiologically suppresses renin release in the kidney, plasma renin activity and aldosterone were suppressed in salt-loaded normotensive rats and lean SHR rats. However, plasma aldosterone levels in salt-loaded obese SHR rats were still higher than that in salt-loaded lean SHR rats, suggesting that aldosterone overproduction by escape from physiologic control was sustained in obese SHR rats. Furthermore, inappropriately high levels of aldosterone during salt loading are a key determinant of the pathogenicity of aldosterone. Thus, we hypothesized that the combination of imbalanced excess of aldosterone and salt loading in obese SHR induces the full activation of MR to exacerbate renal injury. Indeed, MR signaling, including the nuclear accumulation of MR protein and expression of Sgk1, was moderately increased in the kidneys of salt-loaded obese SHR rats. Moreover, eplerenone reversed the upregulation of Sgk1, resulting in the attenuation of salt-induced BP elevation and renal injury. Thus, MR activation by inadequate suppression of aldosterone production in the context of a high-salt diet plays a key role in the increased salt sensitivity of BP and renal injury in obese SHR rats.

Previous studies have suggested that obese adipose tissue is involved in the mechanism of aldosterone overproduction in metabolic syndrome. Goodfriend et al.²⁶ demonstrated a positive correlation between plasma aldosterone level and the volume of visceral fat and suggested a role for the epoxy-keto derivates of linoleic acid.²⁷ Ehrhart-Bornstein et al.²⁸ showed that a secretory product from adipocytes isolated from obese subjects stimulated aldosterone production in cultured adrenal cells. We also observed that conditioned medium from obese SHR rat adipocytes, but not from lean SHR adipocytes, stimulated aldosterone production in cultured adrenal cells.⁵¹ Of note, these effects were not affected by the inhibition of the RAS with angiotensin-converting enzyme inhibitor/angiotensin receptor blocker treatment. This implies that these effects are different from physiologic stimuli, including AngII. However, the “aldosterone-releasing factors” from obese adipocytes have not been identified. Some researchers have suggested that complement-C1q TNF-related protein⁵² and leptin²⁹ are candidate aldosterone-releasing factors. Taken together, aldosterone overproduction induced by aldosterone-releasing factors from obese adipocytes causes salt-sensitive hypertension and renal injury in metabolic syndrome (Figure 1).

Figure 1.

Mechanism of MR-mediated hypertension and renal injuries in metabolic syndrome. In metabolic syndrome, aldosterone (Aldo)-releasing factors from obese adipocytes, which are not identified, stimulate Aldo secretion in the adrenal glands, independent of RAS. Subsequently, activation of the MR in the kidney causes salt-sensitive hypertension and renal injury.

Alternative Mechanism of MR Activation by RAC1

In addition to aldosterone excess as a mechanism in activation of the MR, alternative pathways of MR activation have been suggested. The antihypertensive effect of eplerenone in a clinical trial did not depend on plasma renin-aldosterone profiles.⁵³ In contrast to obese SHR rats, despite adequately suppressed plasma aldosterone, Dahl salt-sensitive (Dahl-S) rats, a classic model of salt-sensitive hypertension, develop hypertension and proteinuric renal injury after salt loading, along with increased MR signaling in kidneys that includes the upregulation of Sgk1 or ENaC.^31,54,55

These observations implicate ligand-independent mechanisms of MR activation. The activity of nuclear receptors is affected by signaling molecules that modulate nuclear translocation, epigenetics, and the recruitment of coactivators or corepressors. A Rho family GTPase, Rac1, has multiple roles in intracellular signaling, including organization of the actin cytoskeleton, generation of reactive oxygen species (ROS), cell migration and adhesion, apoptosis, and modulation of the function of nuclear receptors.^56–58 Rac1 functions as a molecular switch by cycling between an inactive GDP-bound and an active GTP-bound state. The transition is controlled positively by guanine nucleotide exchange factors and negatively by GTPase-activating proteins and GDP-dissociation inhibitors (GDIs)^59,60; Rac1 activates downstream effectors, including p21-activated kinase, LIM kinase, and mitogen-activated protein kinases.^56–58 Recently, we found that Rac1 mediates ligand-independent MR activation.

In our study using a stable line of human embryonic kidney cells (HEK293) and podocytes, the transfection of constitutively active Rac1 promoted the nuclear translocation of green fluorescent protein–tagged MR and enhanced MR-dependent transcriptional activity.³⁰ A p21-activated kinase inhibitor reduced the Rac1-mediated activation of MR, indicating that Rac1 acts via activation of p21-activated kinase.³⁰ We then further investigated the role of the Rac1-MR pathway in an in vivo model of Rac1 activation, RhoGDIα knockout mice. RhoGDIα is a negative regulator of Rac1 and expressed in kidney with relatively high abundance in podocytes^61,62 and collecting ducts.⁶³ Accordingly, RhoGDIα knockout mice showed renal activation of Rac1 along with the nuclear accumulation of MR and upregulation of Sgk1 in the kidney, despite normal aldosterone levels. Moreover, these mice developed profound proteinuria with podocyte damage and FSGS. Treatment with a selective Rac1 inhibitor (NSC23766) and eplerenone abolished the MR signaling in RhoGDIα knockout mice, attenuating glomerular injury, which suggests that the Rac1-mediated activation of MR induces proteinuric renal injury. The role of Rac1 activation in proteinuric renal disease was also reported in familial nephrotic syndrome with mutations in genes encoding Arhgap24⁶⁴ and RhoGDIα,^61,62 which induce Rac1 activation in podocytes. Moreover, an Rac1 inhibitor and eplerenone ameliorated the nephrotic phenotype recapitulated in RhoGDIα knockout zebrafish.⁶¹ These results strongly support the important role of the Rac1-MR pathway in glomerular injury, whereas some other Rac1 signaling, including cytoskeleton remodeling^61,62,64 and crosstalk with transient receptor potential channel 5,^65–67 would also be involved in the Rac1-dependent podocyte injuries. We also demonstrated the involvement of the Rac1-MR pathway in the development of proteinuric renal injury in animal models of RAAS activation⁶⁸ and diabetic nephropathy.⁶⁹

Of note, BP and glomerular injury in RhoGDIα knockout mice were salt sensitive, suggesting Rac1 activation has a pivotal role in the development of salt sensitivity.³¹ We assessed the Rac1-MR pathway in Dahl-S rats and their counterparts, Dahl salt-resistant (Dahl-R) rats. After salt loading, both strains showed similarly suppressed levels of plasma aldosterone. However, only the salt-loaded Dahl-S rats showed Rac1 activation. In contrast, the salt-loaded Dahl-R rats showed suppressed Rac1 activity and MR signaling, in accordance with decreased plasma aldosterone, leading to the absence of hypertension and glomerular injury.³¹ The phenotypes in salt-loaded Dahl-S rats were reversed by not only eplerenone but also NSC23766, associated with the significant reduction of Sgk1, a downstream molecule of MR signaling. These results indicated that the Rac1-MR pathway is a key determinant of salt sensitivity of BP and glomerular injury (Figure 2). Given that the knockdown of RhoGDIα in cultured principal cells of cortical collecting ducts caused Rac1 activation and induced ENaC activation,⁶³ the activation MR-ENaC signaling in ASDN would be involved in the development of salt sensitivity of BP, whereas MR activation in non-ASDN might be also involved in the entire phenotype. The downstream effector of Rac1-MR activation in non-ASDN tissues remains unclear, but ROS are known to mediate MR signaling in the cardiovascular system^{25,33,70–72} and glomeruli.^73–75

Figure 2.

Rac1 is a determinant of BP salt sensitivity through the on/off switching of MR activation. In both salt-resistant and salt-sensitive models, high salt intake suppressed plasma aldosterone (Aldo) concentration. In salt-resistant models, the salt loading suppresses renal Rac1 activity and subsequently, suppresses MR activity to maintain normal BP. In contrast, despite decreased plasma Aldo, high salt activates Rac1 in salt-sensitive models, which causes paradoxical activation of MR, leading to salt-induced BP elevation. The contrasting regulation of MR by Rac1 plays a key role in the development of salt-resistant and salt-sensitive phenotypes in each model.

The switching mechanism of Rac1 activity by salt loading in salt-sensitive and salt-resistant phenotypes is still unclear. One possible mechanism is genetic alteration in molecules that are involved in Rac1 activation. The quantitative trait loci involved in salt sensitivity of Dahl-S rats^76–78 include some genes encoding GTPase-activating proteins. Another possibility is the change in the stimuli that activate Rac1. Rac1 is activated by various stimuli, including mechanical stretch,^79,80 shear stress,⁸¹ inflammatory cytokines,⁸² growth factors,⁸³ integrins,⁸⁴ ROS,⁸⁵ high glucose,^86,87 sodium chloride⁸⁸ or osmotic stress,⁸⁹ AngII,^90–92 and aldosterone.^74,93 Tiam1, a guanine nucleotide exchange factor that is activated by some stimuli, including AngII,⁹⁴ was upregulated by salt loading in Dahl-S rats but unchanged in Dahl-R rats.⁹⁵ Given that intrarenal angiotensinogen was elevated by salt loading in Dahl-S rats,⁹⁶ the local increase in AngII could induce Rac1 activation. Based upon the previous findings indicating that MR activation promotes ROS production and that ROS could activate Rac1 in turn, ROS also might play an important role in creating the vicious cycle of Rac1 and MR activation leading to salt-sensitive hypertension and renal injury. In AKI caused by ischemia-reperfusion, it is noteworthy that vascular ROS overproduction—which is attributed to Rac1 activation via the MR in vascular smooth muscle cells—causes endothelial dysfunction, resulting in tubular injury through reduced renal perfusion.⁹⁷ Thus, intricate crosstalk between Rac1 and the MR among various cell types or tissues would also be involved in the MR-dependent renal injuries.

The Eplerenone Combination versus Conventional Agents to Lower Blood Pressure on Urinary Antialbuminuric Treatment Effect study reported that treatment with eplerenone significantly reduced BP and urinary albumin excretion in patients with CKD already receiving angiotensin-converting enzyme inhibitors, angiotensin receptor blockers, or both, independent of the plasma aldosterone level. Moreover, the antihypertensive and antialbuminuric effects of eplerenone were greater in patients with higher dietary salt intake as estimated by urinary sodium excretion.^21,98 These results led us to the plausible hypothesis that patients with CKD have aldosterone-independent MR activation driven by high salt, possibly through Rac1 activation. Thus, the Rac1-MR pathway is a potential therapeutic target, especially for salt-sensitive hypertension and renal injury.

Cell Type–Specific and Context-Dependent Role of Aldosterone and the MR in the ASDN

Recent findings have also provided new insights into our understanding of the ASDN, which consists of distal convoluted tubules (DCTs), connecting tubules, and collecting ducts. In steady state, most of the sodium delivered to the distal convolution is reabsorbed in the DCT and connecting tubule; the late DCT and connecting tubule play a major role in potassium secretion.^99,100 Classically, aldosterone binds to the MR in the principal cells of the connecting tubule and collecting duct to activate the ENaC via signaling pathways, including Sgk1 and neuronal precursor cell expressed developmentally down-regulated protein 4-2 (Nedd4–2).³ The resultant sodium reabsorption by the ENaC generates a lumen-negative potential, which promotes chloride reabsorption and the secretion of potassium.

Of note, both aldosterone and glucocorticoids, which circulate in plasma at concentrations 100–1000 times higher than that of aldosterone, bind to the MR with equivalent high affinity.¹ In principal cells, however, 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2), which converts glucocorticoids to their inactive metabolites, guarantees the sensitivity of principal cells to aldosterone.^101,102 In deficiency of 11β-HSD2, unconverted glucocorticoids should bind to the MR in principal cells and activate ENaC as a result. This mechanism is thought to be one of the causes of salt-sensitive hypertension observed in the syndrome of apparent mineralocorticoid excess, which involves a loss-of-function mutation in the gene encoding 11β-HSD2.¹⁰³ Supporting this, mice with kidney-specific ablation of the gene encoding 11β-HSD2 develop salt-sensitive hypertension in association with ENaC activation in the kidney, and an ENaC inhibitor amiloride attenuates the hypertension.¹⁰⁴

DCT cells express sodium chloride cotransporter (NCC), a target of thiazide diuretics. It has been reported that the activity of NCC is modulated differently in some conditions with aldosterone excess, such as volume depletion, hyperkalemia, and primary aldosteronism. During volume depletion that activates RAAS, AngII directly activates NCC via with-no-lysine kinase (WNK)–dependent pathways, enhancing sodium chloride reabsorption in the DCTs.^105,106 Conversely, during high potassium intake, hyperkalemia directly suppresses NCC activity and increases urinary sodium excretion in the DCTs, despite the presence of hyperaldosteronism.¹⁰⁷

It has been suggested that the NCC activation in volume depletion would reduce distal sodium delivery and flow rate, which may limit the ENaC-dependent generation of lumen-negative potential for potassium secretion and flow-dependent potassium excretion, and also that the NCC suppression on a high-potassium diet may exert the opposite effect to enhance potassium excretion.^108,109 Supporting this hypothesis, acute hyperkalemia induced by intravenous infusion of potassium in mice was associated with both natriuresis and kaliuresis, along with significant suppression of NCC and redistribution of ENaC and the renal outer medullary potassium channel to the luminal membrane of principal cells.¹⁰⁷ However, some recent studies have indicated that the influence of NCC activity upon potassium secretion is not solely explained by the alteration in distal sodium delivery and flow but would depend on remodeling of ASDN in some conditions.^109,110 A recent study showed that the inhibition of NCC by acute administration of hydrochlorothiazide in mice induced natriuresis but did not caused kaliuresis.¹¹¹ In another study, mice with DCT-specific expression of a constitutively active mutant of STE20/SPS1-related proline-alanine–rich protein kinase, which induced NCC activation, exhibited hyperkalemia in association with remodeling of ASDN; this remodeling involved a reduction of connecting tubule mass and a decrease in expression and apical localization of ENaC and renal outer medullary potassium.¹¹² Hydrochlorothiazide treatment in these mutant mice immediately induced natriuresis but gradually caused kaliuresis to attenuate the hyperkalemia in association with reversal of the ASDN remodeling.¹¹²

Regarding the mechanism of NCC regulation by potassium, a recent study showed that changes in plasma potassium levels altered the polarization of DCT cells, and the resultant change in intracellular chloride level modulated WNK kinases to regulate NCC.¹¹³ In contrast to hyperkalemia, hypokalemia induces NCC activation. In primary aldosteronism, hypokalemia-induced NCC activation at the DCT, in cooperation with MR-ENaC activation at the connecting tubule and collecting duct, contributes to the aberrant sodium reabsorption and the resultant salt-sensitive hypertension. Supporting this, aldosterone infusion in rodents caused hypokalemia and NCC activation, but the correction of hypokalemia by an ENaC inhibitor or potassium supplementation reversed the NCC activation.¹¹⁴ Moreover, NCC inhibitors ameliorated salt-sensitive hypertension in mineralocorticoid-infused rats¹¹⁵ and in mice with the kidney-specific deletion of 11β-HSD2; in both cases, NCC is activated via hypokalemia induced by activation of MR-ENaC signals in principal cells.¹⁰⁴ Of note, some studies offer evidence indicating a direct role of aldosterone in NCC regulation, although the modulation of NCC by aldosterone largely depends on change in plasma potassium. Late DCT, the marginal zone expressing both NCC and ENaC, expresses 11β-HSD2, although the expression level and area vary among species.^116–118 Ackermann et al.² have shown that suppression of aldosterone by dietary salt repletion in rats eliminated the nuclear localization of the MR in late DCT, demonstrating that the MR in late DCT is sensitive to the change in aldosterone. Some researchers have suggested that the aldosterone-MR cascade in DCT activates NCC by modulating interaction among Sgk1, Nedd4–2, and WNK1.¹¹⁹ In addition, a recent study showed that rapid aldosterone-mediated signaling activates NCC through an MR-independent, nongenomic mechanism via GPR30, a membrane-associated receptor.¹²⁰

Emerging evidence has also shown that a chloride/bicarbonate exchanger, pendrin, expressed on β-intercalated cells (β-ICs) in the connecting tubule and cortical collecting duct, mediates chloride flux and is involved in sodium chloride reabsorption in coordination with a sodium-dependent chloride/bicarbonate exchanger.^121–123 Pendrin knockout mice showed slightly reduced arterial pressure detectable by telemetric analysis under basal conditions,^124,125 and they exhibited more severe hypotension with apparent fluid loss after salt depletion.¹²¹ In addition, mice with IC-specific pendrin overexpression developed salt-sensitive hypertension,¹²⁶ demonstrating the important role of pendrin in fluid homeostasis and salt sensitivity of BP. Of note, it has been suggested that pendrin complements NCC. Some studies have indicated that NCC knockout mice showed compensatory activation of pendrin.^127,128 Moreover, pendin/NCC double-knockout mice exhibited visibly enhanced fluid loss and hypotension with normal salt intake compared with mice that have a single knockout of pendrin or NCC.¹²⁸ Similar to NCC, pendrin is upregulated by AngII during volume depletion^129,130 and hypokalemic status during primary aldosterone excess.¹³¹ Interestingly, these upregulations of pendrin were suppressed by an MR blocker, suggesting the involvement of the MR.^130,131

It is still unclear whether aldosterone is a sole ligand of the MR in ICs because in immunohistologic studies^2,102 and single-cell transcriptome analysis,¹³² 11β-HSD2 expression was absent or minimal in ICs, which would permit glucocorticoids to occupy the MR. Notably, recent studies suggest that ICs have a specific mechanism to regulate the ligand-binding affinity of MRs, which are associated with pendrin regulation. Shibata et al.⁹⁵ reported that the MR in ICs was phosphorylated at S843 in the ligand-binding domain and that its phosphorylation prevented ligand binding in the steady state. AngII and hypokalemia dephosphorylate this site and increase the MR’s ligand-binding affinity, leading to the nuclear accumulation of MR in ICs and the upregulation of pendrin only when the ligand was present.^95,130,131 These data indicate that the switching mechanism of MR affinity by phosphorylation in ICs is involved in the regulation of pendrin. Indeed, we and others demonstrated that mice with an IC-specific MR deletion (MR^IC-KO) displayed reduced expression of pendrin after AngII infusion or endogenous AngII elevation by dietary salt depletion.^42,43 Salt-depleted MR^IC-KO mice showed normal BP related to the compensatory activation of NCC to maintain BP; they also exhibited fluid loss and hypotension after an additional treatment with an NCC inhibitor.⁴² Importantly, NCC inhibitor treatment of salt-depleted MR^IC-KO mice resulted in hypokalemia, which is consistent with a recent finding indicating that pendrin has a potassium-sparing effect.¹³³ Thus, the AngII-NCC and AngII-MR-pendrin cascades might coordinate to maintain fluid levels and BP and simultaneously maintain potassium balance during salt depletion.

We also observed that aldosterone administration to salt-loaded mice induced pendrin upregulation and NCC activation; this occurred in association with hypokalemia and metabolic alkalosis induced by activation of the MR-ENaC cascade in principal cells.⁴² In addition, immunohistologic analysis showed that aldosterone significantly increased the relative abundance of pendrin label in the cells’ apical membrane region.^42,43,134 Correction of hypokalemic alkalosis by an ENaC inhibitor or potassium supplementation reversed the changes in pendrin and NCC. However, the aldosterone-induced upregulation of pendrin was only partially suppressed in MR^IC-KO mice.^42,43 A report described significant attenuation of apical membrane accumulation of pendrin label in MR^IC-KO mice in immunohistochemical analysis⁴³; however, another report found that aldosterone-induced change in subcellular distribution of pendrin is still observed in MR^IC-KO mice.⁴² Despite this inconsistency, the correction of hypokalemic alkalosis by potassium supplementation in aldosterone-treated MR^IC-KO mice significantly reduced pendrin expression.⁴² Thus, a substantial portion of aldosterone-induced regulation of pendrin is dependent on hypokalemic alkalosis but is independent of the MR in ICs.

We also examined whether hypokalemia or metabolic alkalosis was responsible for pendrin activation related to primary aldosterone excess. Because pendrin was originally reported as a mediator of bicarbonate excretion responding to alkali loading,¹³⁵ we hypothesized that alkalosis drives the aldosterone-induced pendrin activation. Accordingly, we found that treating aldosterone-infused mice with a carbonic anhydrase inhibitor, acetazolamide, corrected alkalosis without a change in hypokalemia and reversed the upregulation of pendrin with no change in NCC activation.⁴² Thus, during aldosterone excess, hypokalemia and alkalosis—consequences of MR-ENaC pathway activation—induced the parallel activation of NCC and pendrin, respectively, which was independent of the MR.

On the basis of a previous finding that pendrin knockout attenuated hypertension induced by mineralocorticoid excess,¹³⁴ pendrin upregulation induced by hypokalemic alkalosis contributed to aldosterone-induced hypertension. Furthermore, we observed that the correction of hypokalemic alkalosis by potassium supplementation suppressed pendrin expression and salt-sensitive hypertension in NCC knockout mice,⁴² indicating the role of pendrin as a complement pathway in NCC inhibition. As is well known, the clinical use of thiazide diuretics occasionally activates RAAS and hypokalemic alkalosis, resulting in thiazide resistance. Moreover, MR blockers effectively reduced the BP in a patient with resistant hypertension, defined as BP that remains above the target level despite treatment with at least three antihypertensive drugs, including a diuretic.¹³⁶ Thus, the two pathways of pendrin upregulation—mediated through either the MR in ICs activated by RAS stimulation during volume depletion or hypokalemic alkalosis driven by the MR-ENaC cascade in principal cells during aldosterone excess (Figure 3)—may be therapeutic targets for thiazide-resistant hypertension.

Figure 3.

Putative mechanism of salt-sensitive hypertension in primary aldosteronism. Cell type–specific mechanisms of sodium reabsorption are mediated by MR in the ASDN. Classically, aldosterone activates the MR-ENaC pathway in principal cells (PCs) of the connecting tubules (CNTs) and cortical collecting ducts (CCDs) to reabsorb sodium and secrete potassium. The interaction between aldosterone and MR in PCs is guaranteed by 11β-HSD2 that converts glucocorticoids to inactive metabolites. NCC expressed in DCT is directly activated by AngII and hypokalemia, which is induced by the aldosterone-induced activation of MR-ENaC in PCs. In β-ICs in the CNTs and CCDs, AngII induces dephosphorylation of MR, which increases affinity of ligands, and resultantly causes MR activation, leading to upregulation of pendrin. In addition, metabolic alkalosis, which is induced by activation of MR-ENaC in PCs, directly induces pendrin upregulation in β-ICs. The contrasting activation of NCC and pendrin induced by aldosterone excess, in a coordinated manner, mediates sodium chloride reabsorption, leading to the development of salt-sensitive hypertension. P, phosphorylation of MR; ROMK, renal outer medullary potassium.

Conclusions

Updated information about the renal activation of the aldosterone-MR system in the development of salt-sensitive hypertension and renal injury will provide new insights into the use of MR antagonists for the treatment of resistant hypertension and renal injury. Recent studies have also revealed that the aldosterone-MR system in various tissues participates in the pathogenesis of cardiorenal diseases by both BP-dependent and BP-independent mechanims.^{15,16,33,34,137–140} Of note, clinical trials have demonstrated the efficacy and improved safety profiles of novel nonsteroidal MR antagonists, such as finerenone^{20,22,141,142} and esaxerenone,¹⁴³ in cardiovascular and renal injuries in diabetic individuals. Given the promising effect of MR antagonists, further investigation of the aldosterone-MR system may lead to the discovery of therapeutic strategies and targets for hypertension and end organ injury.

Disclosures

T. Fujita reports research funding from Bayer Japan and Omron; honoraria from Daiichi-Sankyo; and scientific advisor or membership with Hypertension, Hypertension Research, the Japanese Society of Hypertension, and the Japanese Society of Nephrology. The remaining author has nothing to disclose.

Funding

This work was supported by Japan Society for the Promotion of Science grants KAKENHI JP20K08585 and KAKENHI JP15H05788.