Blockade of mineralocorticoid receptor has been shown to improve the clinical outcomes of proteinuric kidney diseases. However, little is known about the regulation of mineralocorticoid receptor-dependent transcriptional activity in renal disease. Here we identify a new role for Rac1, a member of the Rho family GTPases, as a potent activator of mineralocorticoid receptor signal transduction both in vitro and in vivo. Transient transfection assays in HEK 293 cells revealed that constitutively active Rac1 (CA-Rac1) enhanced mineralocorticoid receptor-dependent reporter activity, which was accompanied by increased nuclear translocation of mineralocorticoid receptor. CA-Rac1 facilitated mineralocorticoid receptor nuclear accumulation also in podocytes via p21-activated kinase phosphorylation. In mice lacking Rho GDP-dissociation inhibitor-alpha (Arhgdia(-/-) mice), renal abnormalities, including heavy albuminuria and podocyte damage, were associated with increased Rac1 (but not RhoA) and mineralocorticoid receptor signaling in the kidney, without alteration in systemic aldosterone status. Pharmacological intervention with a Rac-specific small-molecule inhibitor diminished mineralocorticoid receptor overactivity and renal damage in this model. Furthermore, albuminuria and histological changes in Arhgdia(-/-) mice were suppressed by mineralocorticoid receptor blockade, confirming the pathological role of Rac1-mineralocorticoid receptor interaction. Our results provide evidence that signaling cross-talk between Rac1 and mineralocorticoid receptor modulates mineralocorticoid receptor activity and identify Rac1 as a therapeutic target for chronic kidney disease.

Abstract-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.

Metabolic syndrome is an important risk factor for proteinuria and chronic kidney disease independent of diabetes and hypertension; however, the underlying mechanisms have not been elucidated. Aldosterone is implicated in target organ injury of obesity-related disorders. This study investigated the role of aldosterone in the early nephropathy of 17-wk-old SHR/ NDmcr-cp, a rat model of metabolic syndrome. Proteinuria was prominent in SHR/NDmcr-cp compared with nonobese SHR, which was accompanied by podocyte injury as evidenced by foot process effacement, induction of desmin and attenuation of nephrin. Serum aldosterone level, renal and glomerular expressions of aldosterone effector kinase Sgk1, and oxidative stress markers all were elevated in SHR/NDmcr-cp. Mineralocorticoid receptors were expressed in glomerular podocytes. Eplerenone, a selective aldosterone blocker, effectively improved podocyte damage, proteinuria, Sgk1, and oxidant stress. An antioxidant tempol also alleviated podocyte impairment and proteinuria, along with inhibition of Sgk1. As for the mechanisms of aldosterone excess, visceral adipocytes that were isolated from SHR/NDmcr-cp secreted substances that stimulate aldosterone production in adrenocortical cells. The aldosterone-releasing activity of adipocytes was not inhibited by candesartan. Adipocytes from nonobese SHR did not show such activity. In conclusion, SHR/NDmcr-cp exhibit enhanced aldosterone signaling, podocyte injury, and proteinuria, which are ameliorated by eplerenone or tempol. The data also suggest that adipocyte-derived factors other than angiotensin II might contribute to the aldosterone excess of this model.

Hypertension is a leading contributor to cardiovascular mortality worldwide. Despite this, its underlying mechanism(s) and the role of excess salt in cardiorenal dysfunction are unclear. Previously, we have identified cross-talk between mineralocorticoid receptor (MR), a nuclear transcription factor regulated by the steroid aldosterone, and the small GTPase Rac1, which is implicated in proteinuric kidney disease. We here show that high-salt loading activates Rac1 in the kidneys in rodent models of salt-sensitive hypertension, leading to blood pressure elevation and renal injury via an MR-dependent pathway. We found that a high-salt diet caused renal Rac1 upregulation in salt-sensitive Dahl (Dahl-S) rats and downregulation in salt-insensitive Dahl (Dahl-R) rats. Despite a reduction of serum aldosterone levels, salt-loaded Dahl-S rats showed increased MR signaling in the kidneys, and Rac1 inhibition prevented hypertension and renal damage with MR repression. We further demonstrated in aldosterone-infused rats as well as adrenalectomized Dahl-S rats with aldosterone supplementation that salt-induced Rac1 and aldosterone acted interdependently to cause MR overactivity and hypertension. Finally, we confirmed the key role of Rac1 in modulating salt susceptibility in mice lacking Rho GDP-dissociation inhibitor α. Therefore, our data identify Rac1 as a determinant of salt sensitivity and provide insights into the mechanism of salt-induced hypertension and kidney injury.

Abstract-Accumulating evidence suggests that mineralocorticoid receptor blockade effectively reduces proteinuria in hypertensive patients. However, the mechanism of the antiproteinuric effect remains elusive. In this study, we investigated the effects of aldosterone on podocyte, a key player of the glomerular filtration barrier. Uninephrectomized rats were continuously infused with aldosterone and fed a high-salt diet. Aldosterone induced proteinuria progressively, associated with blood pressure elevation. Notably, gene expressions of podocyte-associated molecules nephrin and podocin were markedly decreased in aldosterone-infused rats at 2 weeks, with a gradual decrease thereafter. Immunohistochemical studies and electron microscopy confirmed the podocyte damage. Podocyte injury was accompanied by renal reduced nicotinamide-adenine dinucleotide phosphate oxidase activation, increased oxidative stress, and enhanced expression of aldosterone effector kinase Sgk1. Treatment with eplerenone, a selective aldosterone receptor blocker, almost completely prevented podocyte damage and proteinuria, with normalization of elevated reduced nicotinamide-adenine dinucleotide phosphate oxidase activity. In addition, proteinuria, podocyte damage, and Sgk1 upregulation were significantly alleviated by tempol, a membrane-permeable superoxide dismutase, suggesting the pathogenic role of oxidative stress. Although hydralazine treatment almost normalized blood pressure, it failed to improve proteinuria and podocyte damage. In cultured podocytes with consistent expression of mineralocorticoid receptor, aldosterone stimulated membrane translocation of reduced nicotinamide-adenine dinucleotide phosphate oxidase cytosolic components and oxidative stress generation in podocytes. Furthermore, aldosterone enhanced the expression of Sgk1, which was inhibited by mineralocorticoid receptor antagonist and tempol. In conclusion, podocytes are injured at the early stage in aldosterone-infused rats, resulting in the occurrence of proteinuria. Aldosterone can directly modulate podocyte function, possibly through the induction of oxidative stress and Sgk1.

No abstract

Abstract-Aldosterone is implicated in the pathogenesis of proteinuria and chronic kidney disease. We previously demonstrated the contribution of elevated serum aldosterone in the early nephropathy of SHR/NDmcr-cp (SHR/cp), a rat model of metabolic syndrome. In the present study, we investigated the effect of salt loading on renal damage in SHR/cps and explored the underlying mechanisms. SHR/cps fed a high-sodium diet for 4 weeks developed severe hypertension, massive proteinuria, and advanced renal lesions. High salt also worsened glomerular podocyte impairment. Surprisingly, selective mineralocorticoid receptor (MR) antagonist eplerenone dramatically ameliorated the salt-induced proteinuria and renal injury in SHR/cps. Although salt loading reduced circulating aldosterone, it increased nuclear MR and expression of aldosterone effector kinase Sgk1 in the kidney. Gene expressions of transforming growth factor-␤1 and plasminogen activator inhibitor-1 were also enhanced in the kidneys of salt-loaded SHR/cps, and eplerenone completely inhibited these injury markers. To clarify the discrepancy between decreased aldosterone and enhanced MR signaling by salt, we further investigated the role of oxidative stress, a putative key factor mediating salt-induced tissue damage. Interestingly, antioxidant Tempol attenuated the salt-evoked MR upregulation and Sgk1 induction and alleviated proteinuria and renal histological abnormalities, suggesting the involvement of oxidative stress in salt-induced MR activation. MR activation by salt was not attributed to increased serum corticosterone or reduced 11␤-hydroxysteroid dehydrogenase type 2 activity. In conclusion, sodium loading exacerbated proteinuria and renal injury in metabolic syndrome rats. Salt reduced circulating aldosterone but caused renal MR activation at least partially via induction of oxidative stress, and eplerenone effectively improved the nephropathy. Key Words: aldosterone Ⅲ mineralocorticoids Ⅲ metabolic syndrome Ⅲ sodium Ⅲ dietary Ⅲ oxidative stress (kidney) Ⅲ chronic kidney disease Ⅲ proteinuria Ⅲ eplerenone S alt plays an important role in the progression of target organ injury, as well as in the pathogenesis of hypertension. Epidemiologic studies revealed that higher sodium intake increases the risk of cardiovascular disease, independent of other risk factors, 1 and causes poor prognosis of renal disease. 2 Experimental animal studies indicated that increased dietary salt promotes renal fibrosis and progression of kidney disease. 3,4 The deleterious effects of salt are, at least partially, independent of blood pressure (BP). 1,2,4 Susceptibility to salt is reported to be augmented in certain disease conditions. For example, high salt exacerbates proteinuria in overweight but not nonoverweight subjects. 5 However, the underlying mechanisms have not been clearly elucidated.Accumulating evidence suggests that aldosterone is an important pathogenetic factor in the progression of renal disease and heart failure. 6 In the kidney, aldosterone was demonstrated to ...

No abstract