代谢综合征合并慢性肾病中醛固酮的激活作用

REVIEW ARTICLEActivation of the aldosterone/mineralocorticoid receptor system in chronic kidney disease and metabolic syndromeMiki NagaseReceived:3January 2010/Accepted:13May 2010/Published online:9June 2010ÓJapanese Society of Nephrology 2010Abstract Recent clinical and experimental studies have shown that aldosterone is a potent inducer of proteinuria and that mineralocorticoid receptor (MR)antagonists confer efficient antiproteinuric effects.We identified glo-merular epithelial cells (podocytes)as novel targets of aldosterone;activation of MR injures podocytes possibly via oxidative stress,resulting in disruption of glomerular filtration barrier,proteinuria,and progression of chronic kidney disease.We also demonstrated that SHR/cp,a rat model of metabolic syndrome,was susceptible to podocyte injury and proteinuria.Aldosterone excess caused by adipocyte-derived aldosterone-releasing factors was sug-gested to underlie the nephropathy.High salt intake aug-mented MR activation in the kidney and exacerbated the nephropathy.Furthermore,we identified an alternative pathway of MR activation by small GTPase Rac1.Rho-GDI a knockout mice,a model with Rac1activation in the kidney,showed albuminuria,podocyte injury,and glo-merulosclerosis.Renal injury in the knockout mice was accompanied by enhanced MR signaling in the kidney despite normoaldosteronemia,and was ameliorated by an MR antagonist,eplerenone.Moreover,Rac-specific inhib-itor significantly reduced the nephropathy,concomitantly with repression of MR activation.In vitro transfection studies provided direct evidence of Rac1-mediated MR activation.In conclusion,our findings suggest that MRactivation plays a pivotal role in the pathogenesis of chronic kidney disease in metabolic syndrome,and that MR may be activated both aldosterone dependently (via aldosterone-releasing factors)and independently (via Rac1).MR antagonists are promising antiproteinuric drugs in metabolic syndrome,although long-term effects on renal outcomes,mortality,and safety need to be established.Keywords Proteinuria ÁPodocyte injury ÁSalt ÁAldosterone-releasing factors ÁRac1IntroductionSince its isolation in 1953,aldosterone (formerly called ‘‘electrocortin’’)has been recognized as a hormone that regulates electrolyte,fluid volume,and blood pressure (BP)homeostasis.Aldosterone acts on its receptor,mineralo-corticoid receptor (MR)[1],in the epithelial cells of the aldosterone-sensitive distal nephron in the kidney (late distal convoluted tubules,the connecting tubules,and the principal cells of the collecting ducts),thereby controlling sodium reabsorption and potassium excretion.This classic mode of action has been substantiated by many studies of genetic disorders and knockout mice.Recently,however,aldosterone has emerged as a dele-terious hormone in the cardiovascular and renal system [2–4].Accumulating clinical evidence suggests the role of aldosterone in the pathogenesis of hypertension,meta-bolic syndrome,cardiac outcomes,and renal munity-based prospective studies revealed that higher serum aldosterone levels within the physiologic range confer increased risk of development of hypertension and metabolic syndrome [5,6].Higher circulating con-centrations of aldosterone were associated with increasedThis article was presented as the Oshima Award memorial lecture at the 52nd annual meeting of the Japanese Society of Nephrology,held at Yokohama,Japan,in 2009.M.Nagase (&)Division of Chronic Kidney Disease,Department of Nephrology and Endocrinology,University of Tokyo Graduate Schoolof Medicine,7-3-1Hongo,Bunkyo-ku,Tokyo 113-8655,Japan e-mail:mnagase-tky@umin.ac.jpClin Exp Nephrol (2010)14:303–314DOI 10.1007/s10157-010-0298-8mortality among patients with chronic heart failure[7]and with ST-elevation myocardial infarction[8].As for renal dysfunction,plasma aldosterone concentrations were in most instances elevated in chronic renal disease(creatinine clearance below50%of normal)with normal serum potassium and renin levels[9].Quinkler et al.[10]reported that serum aldosterone concentrations were negatively correlated with creatinine clearance and positively associ-ated with renal scarring in chronic kidney disease(CKD) patients.We recently found that the glomerular epithelial cell (podocyte),finalfiltration barrier in the glomerulus,is a novel target of aldosterone[11],and activation of MR in podocyte causes proteinuria and glomerulosclerosis,pos-sibly via oxidative stress,in metabolic syndrome model [12].We also identified a novel mechanism of MR acti-vation by small GTPase Rac1and its contribution to CKD [13].Herein,we summarize recent advances in aldosterone research and introduce our data on the roles and regulation of the aldosterone/MR system in CKD and metabolic syndrome.Aldosterone and target organ injuryHistorically,it was already known in the1940s(even before isolation of aldosterone and MR)that MR activation causes inflammation andfibrosis in target organs;for example,Hans Selye[14],an advocate of‘‘stress theory(non-specific adaptation response to stressor by glucocorticoids),’’reported that administration of desoxy(11-deoxy)corticosterone acetate(DOCA),thefirst synthetic steroid which stimulates MR,to rats induces inflammatory andfibrotic changes in the heart and kidney.Uninephrectomy and salt loading were performed as conditioning factors.He considered that these phenotypes were due to mineralocorticoid actions of DOCA.Furthermore,he proposed that glucocorticoids have anti-inflammatory actions in adaptation to stress, whereas mineralocorticoids may have pro-inflammatory effects.Half a century later,aldosterone,the physiological mineralocorticoid in our body,was shown to cause myo-cardial and vascularfibrosis[15,16].MR was found to be expressed not only in the epithelia of the kidney and colon but also in various nonepithelial cells.Based on these experimentalfindings,together with aldosterone break-through phenomenon(see below),Pitt et al.[17,18]con-ducted two milestone randomized clinical trials,RALES and EPHESUS,in which addition of low-dose MR antag-onist,spironolactone or eplerenone,dramatically improved outcomes of heart failure patients already taking standard medications including angiotensin-converting enzyme inhibitors(ACEIs),angiotensin II(Ang II)type1receptor blockers(ARBs),diuretics,and b-blockers.The organ-protective efficacy of MR antagonists was further con-firmed by subsequent clinical studies[19,20].The aldosterone/MR system plays a pathogenic role also in the progression of CKD[2,3].MR is present in various nonepithelial cells,where aldosterone causes inflammation andfibrosis through genomic and nongenomic actions. Multiple factors are proposed to be involved:plasminogen activator inhibitor(PAI)-1,transforming growth factor (TGF)-b1,connective tissue growth factor,proinflammatory cytokines such as osteopontin and monocyte chemoattrac-tant protein(MCP)-1,and reactive oxygen species(ROS).MR binds both mineralocorticoids and glucocorticoids with high affinity[1].Because plasma glucocorticoid con-centrations are100–1000times higher than those of aldo-sterone,aldosterone target cells have to equip machinery for preventing occupation of MR by glucocorticoids.In the Na?-transporting epithelia,such as the renal distal nephron, the enzyme11b-hydroxysteroid dehydrogenase type2 (11b HSD2)confers specificity on aldosterone to activate MR,by converting cortisol and corticosterone into inactive 11-keto derivatives,which have low affinity for MR[21].In nonepithelial tissues without11b HSD2,glucocorticoids activate MR or,in some cases,glucocorticoids just occupy and do not activate MR.In the kidney,11b HSD2is abun-dantly expressed in the distal tubules and collecting ducts, but is also found in the glomeruli,proximal tubules,cultured podocytes,and mesangial cells[22–24].The enzyme is also expressed in vascular endothelial and smooth muscle cells, while its expression is minimal or absent in the myocardium, hippocampus,adipocytes,and macrophages. Aldosterone and proteinuriaCumulative evidence suggests that aldosterone is a potent inducer of rge-scale,randomized clinical trials have proved the efficacy of renin–angiotensin–aldo-sterone system(RAAS)blockade by ACEIs or ARBs in reducing proteinuria and retarding progression of kidney disease in diabetic and nondiabetic patients[25–30]. Conventionally,Ang II has been regarded as the primary factor responsible for injurious actions of the RAAS in the kidney.Recent studies,however,indicated aldosterone as an additional pathogenetic factor.According to the report by Greene et al.[31],Ang II blockade by losartan and enalapril markedly attenuated proteinuria in rat remnant kidney,a model with enhanced RAAS activity.However, these drugs not only inhibited the action of Ang II but also blocked aldosterone synthesis,and reversal of plasma aldosterone levels by exogenous aldosterone infusion recapitulated the nephropathy.Conversely,adrenalecto-mized rats with glucocorticoid replacement were protectedfrom renal-ablation-induced proteinuria[32].Similarfind-ings were observed in stroke-prone spontaneously hyper-tensive rats(SHR)[33,34],in which renal MR contents as well as plasma aldosterone levels are elevated[35].Indeed, Ang II excess models,such as Ang II/salt rats and double transgenic rats carrying both human renin and human angi-otensinogen genes,develop severe albuminuria and kidney injury,which were abrogated by aldosterone synthase inhibitor FAD286,adrenalectomy or MR antagonists [36,37].The pivotal role of aldosterone was directly proved by creating mineralocorticoid excess models,such as uni-nephrectomized,high-salt-fed rats loaded with aldosterone or DOCA,which were quite prone to proteinuria and glo-merulosclerosis,despite suppressed plasma renin activity and Ang II concentrations[38,39].Serum-and glucocorti-coid-inducible kinase1(Sgk1),an aldosterone effector kinase,might mediate the proteinuric effect of aldosterone, because mice lacking Sgk1were protected against DOCA/ salt-induced albuminuria[40].As for clinical evidence supporting the association between aldosterone and proteinuria,Conn et al.[41] described that85%of patients with primary aldosteronism (PA),a high-aldosterone state,were positive for protein-uria.More recent studies documented that urinary protein excretion was higher in patients with PA than in those with essential hypertension[42,43].Conversely,MR antago-nists were shown to effectively ameliorate proteinuria in patients with hypertension,diabetes mellitus,and CKD. For example,Williams et al.[20]compared the efficacy of eplerenone and enalapril in patients with stage1or2 hypertension.After12months,the extent of BP reduction was similar between the two groups,whereas eplerenone was superior to enalapril in reducing albuminuria(-62% versus-26%;P=0.01).A recent double-blind,placebo-controlled trial by Mehdi et al.[44]showed that addition of spironolactone,but not losartan or placebo,afforded greater antiproteinuric effect in diabetic hypertensive patients already under maximal ACE inhibition.They carefully controlled factors known to affect protein excretion in diabetes including BP,sodium and protein intake,and glycemia.After48weeks,albuminuria decreased by34.0%in the spironolactone group and by 16.8%in the losartan group compared with the placebo group.Regarding safety,however,incidence of asymp-tomatic hyperkalemia(C6.0mEq/L on at least one occasion)was highest in the spironolactone group(14out of27patients).A recent systematic review and meta-analysis by Navaneethan et al.[45]revealed that MR antagonists could significantly reduce proteinuria in CKD patients receiving ACEI and/or ARB,but increase the risk of hyperkalemia.It should be noted,however,that long-term effects on renal outcomes,mortality,and safety need to be established.Aldosterone breakthroughThe role of aldosterone as a mediator of proteinuria was also suggested by the‘‘aldosterone breakthrough’’phenomenon [46].Staessen et al.[47]first described in1981that treat-ment of hypertensive patients with high dose of ACEI captopril lowered plasma aldosterone from74to21pg/mL after1month,but plasma aldosterone began to rise there-after,and after1year reached a level of165pg/mL.Plasma Ang II levels remained suppressed throughout the long-term therapy.This phenomenon was originally known as ‘‘aldosterone escape’’but is now termed‘‘aldosterone breakthrough,’’because‘‘aldosterone escape’’also means loss of salt-retaining ability of excess aldosterone.Although the renin–angiotensin system(RAS)is the principal regu-lator of aldosterone synthesis and secretion in the adrenal glands,aldosterone synthesis is also regulated by various non-RAS stimuli including plasma potassium levels and adrenocorticotropic hormone.These non-RAS secreta-gogues may stimulate aldosterone release under long-term RAS blockage.Sato et al.[48]reported that aldosterone breakthrough was observed in40%of patients with type2diabetes and early nephropathy during40weeks of trandolapril therapy (plasma aldosterone concentration:112.0±18.7and 53.2±15.1pg/mL in patients with and without aldoste-rone breakthrough,respectively).Trandolapril reduced urinary albumin excretion in patients without aldosterone breakthrough(119±95mg/g Cre),whereas the antialbu-minuric effect of trandolapril was lost in patients with aldosterone breakthrough(368±142mg/g Cre).The contribution of unsuppressed aldosterone was further sup-ported by the observation that addition of low-dose spiro-nolactone on top of trandolapril for24weeks significantly reduced albuminuria without BP change in patients with aldosterone breakthrough.Bianchi et al.[49]reported that spironolactone(25mg/day)is useful in reducing protein-uria and retarding the decrease in estimated glomerular filtration rate(GFR)after1year of therapy in nondiabetic CKD patients already treated with ACEIs and/or ARBs. They found that baseline aldosterone levels under Ang II blockade were positively correlated with protein-uria,and predicted the degree of reduction in proteinuria with spironolactone.These studies indicate that aldoste-rone breakthrough indeed contributes to albuminuria and decline in GFR.Aldosterone evokes podocyte injury:role of oxidative stressThe glomerularfiltration barrier comprises three layers:the fenestrated glomerular endothelium,the glomerularbasement membrane,and the podocytes.Podocytes line the outer aspect of the glomerular basement membrane,and serve as thefinal barrier against urinary protein loss[50, 51].Podocytes are injured in many types of proteinuric renal diseases,including nephrotic syndrome,lupus nephritis,diabetic and hypertensive nephropathy,and obesity-related glomerulopathy[39,52–55].The intimate relationship between aldosterone and pro-teinuria prompted us to investigate the effects of aldoste-rone on podocytes.Wefirst analyzed proteinuria and podocyte injury in uninephrectomized,high-salt-fed rats continuously infused with aldosterone(0.75l g/h via osmotic minipump)[11].This is an established chronic mineralocorticoid excess model in which plasma aldoste-rone levels rise to those seen in human congestive heart failure,whereas plasma renin activity and circulating Ang II are quickly suppressed.After4weeks,aldosterone-infused rats developed hypertension and massive protein-uria.Glomerular expressions of slit diaphragm-associated molecules nephrin and podocin were markedly decreased, whereas expression of a damaged podocyte marker desmin was upregulated in aldosterone-infused rats.Electron microscopic analysis revealed podocyte foot process effacement.Podocytes were already impaired at2weeks of aldosterone infusion,when proteinuria was modestly increased.Proteinuria and podocyte damage were com-pletely reversed by selective MR antagonist eplerenone. Podocyte injury was also reported in another mineralo-corticoid excess model,uninephrectomized DOCA/salt rats [39].Thesefindings suggest that podocyte injury underlies the pathogenesis of proteinuria caused by aldosterone administration.So,how did aldosterone evoke podocyte injury?First, aldosterone-induced hypertension should be an important contributor to the podocyte damage.Our uninephrectom-ized aldosterone/salt rats developed hypertension.Systemic BP load,especially in the presence of impaired autoregu-latory vasoconstriction of the preglomerular arterioles, enhances transmission of systemic pressure to the glome-ruli,and the resultant glomerular hypertension/hyperfiltra-tion predisposes to hypertensive glomerular damage[56]. Indeed,a micropuncture study proved these hemodynamic factors as the basis for glomerular injury in uninephrec-tomized DOCA/salt rats[57].Mechanical stress is postu-lated as a key mediator of podocyte damage[51]. Podocytes,whose foot processes overlie glomerular capil-lary tufts,are sensitive to mechanical stress.Cultured podocytes exposed to mechanical strain were reported to enhance expression of proinflammatory cytokines,TGF-b, and Ang II type1receptor,resulting in podocyte damage [58,59].Alternatively,aldosterone may modulate podocyte function directly by acting on MR within podocytes.MR transcripts and proteins were actually detected in in vitro cultured podocytes as well as in vivo glomerular podo-cytes.Exposure of cultured podocytes to aldosterone resulted in nuclear translocation of MR,activation of reduced nicotinamide adenine dinucleotide phosphate (NADPH)oxidase,and ROS increment.In addition,aldo-sterone upregulated expression of an aldosterone effector kinase,Sgk1,in cultured podocytes as well as in the kidney of aldosterone-infused rats,which was prevented by MR antagonist.Taken together,our results suggest that at least some of the proteinuric effects of aldosterone are attribut-able to direct actions on podocytes.We next examined the role of oxidative stress,because ROS are proposed as important mediators of aldosterone-induced target organ injury.Aldosterone-infused rats exhibited enhanced oxidative stress markers such as increased urinary excretion of thiobarbituric acid reactive substances,elevated renal NADPH oxidase activity deter-mined using lucigenin chemiluminescence assay,and stimulation of membrane translocation of NADPH oxidase cytosolic components p67phox and Rac1.Indeed,an anti-oxidant,tempol,significantly reduced oxidative stress markers and corrected podocyte damage and proteinuria. Oxidative stress markers were also suppressed by eplere-none.These results suggest that aldosterone causes podo-cyte injury and proteinuria,possibly through induction of oxidant stress.Metabolic syndrome,CKD,and aldosterone Metabolic syndrome,a constellation of comorbidities that include visceral obesity,hypertension,glucose intolerance, and dyslipidemia,has become a major public health con-cern.Metabolic syndrome is known as a highly predis-posing condition for cardiovascular disease(CVD).Recent epidemiological studies revealed that metabolic syndrome is an important modifiable risk factor for proteinuria and CKD as well.For instance,a cross-sectional study based on the data of NHANES III showed that metabolic syndrome subjects have higher incidence of microalbuminuria (12.3%versus4.7%)and CKD(6.0%versus1.2%)[60]. According to a prospective longitudinal study,based on the data of community-living nondiabetic adults with normal kidney function,having metabolic syndrome increased the risk of developing CKD by approximately50%over 9years,and the risk increased with more traits of the syndrome[61].However,the mechanisms linking meta-bolic syndrome to CKD have not been clearly elucidated.It cannot be solely attributed to the additive effects of indi-vidual component such as hypertension and diabetes mel-litus[61],and some unifying underlying mechanism has been suggested.It is widely accepted that the RAAS plays a crucial role in the pathogenesis of metabolic syndrome.Recent clinical evidence suggests a role for aldosterone per se.Two cross-sectional clinical studies of patients of African descents demonstrated that plasma aldosterone concentration is independently associated with metabolic syndrome,while plasma renin activity was rather suppressed [62,63].In another study,the C allele of -344C/T variant in the promoter of the aldosterone synthase (CYP11B2)gene,which is associated with hyperaldosteronemia,was shown to increase susceptibility to metabolic syndrome in European men [64].Fallo et al.[65]observed that patients with PA had higher incidence of metabolic syndrome than those with essential hypertension (41.1%versus 29.6%;P \0.05).Podocyte injury and proteinuria in metabolic syndrome rats:contribution of aldosterone/MR activation Considering the interrelationships among metabolic syn-drome,CKD,and aldosterone,we postulated a working hypothesis that aldosterone/MR activation plays a critical role in progression of CKD in metabolic syndrome.We used SHR/NDmcr-cp (SHR/cp,obese SHR)as a rat model of metabolic syndrome (Fig.1)[12].This rat is a derivative of SHR with introduction of leptin receptor gene cpmutation,and manifests a clustering of obesity,hyperten-sion,hyperinsulinemia,and hypertriglyceridemia.Obese SHR/cp developed marked proteinuria in an age-dependent manner,while urinary protein excretion remained low in nonobese SHR despite similar BP level.Proteinuria in SHR/cp was accompanied by podocyte injury,as indicated by attenuation of nephrin,induction of desmin,and foot process effacement under electron microscopic analysis.Notably,serum aldosterone levels were higher in obese SHR/cp than in nonobese SHR,and there was positive correlation between circulating aldosterone concentration and proteinuria.Expression of Sgk1was upregulated in the glomerular fraction as well as in the whole kidney of SHR/cp,supporting the causative role of aldosterone/MR acti-vation.Indeed,selective MR antagonist eplerenone effec-tively reduced proteinuria and podocyte damage in SHR/cp.These data suggest that aldosterone-provoked podocyte injury plays a pivotal role in pathogenesis of proteinuria in SHR/cp.We further explored the role of oxidative stress.Similar to the case of aldosterone-infused rats,oxidative stress markers were upregulated in SHR/cp.Importantly,eplerenone reversed the increased oxidative stress,suggesting partici-pation of endogenous aldosterone.Treatment with antioxi-dant tempol also significantly alleviatedproteinuriaFig.1Hyperaldosteronemia in SHR/cp (metabolic syndrome rats),and antiproteinuric effects of MR antagonist eplerenone in SHR/cp and SHR/cp fed a high salt (8%NaCl)diet.a Serum aldosterone concentrations in SHR at 17weeks old and SHR/cp at 17and 24weeks old.b Proteinuria in SHR,SHR/cp,and SHR/cp treated with Epl (SHR/cp ?Epl).Epl wasadministered from 13weeks old for 4weeks.c Proteinuria and nephrin staining in SHR/cp fed a normal diet (SHR/cp),SHR/cp fed a high salt diet (SHR/cp ?HS),and SHR/cp ?HS treated with Epl (SHR/cp ?HS ?Epl).*P \0.05,**P \0.01versus SHR;##P \0.01versus SHR/cp;$$P \0.01versus SHR/cp ?HS.Epl,eplerenoneand podocyte injury in SHR/cp.These data suggest that oxidative stress is an important mediator of aldosterone-induced podocyte injury in this model.Mechanisms of aldosterone excess in SHR/cpWe further investigated the mechanisms of high-aldoste-rone state in SHR/cp.Expression of CYP11B2was enhanced in the adrenal glands of SHR/cp compared with nonobese SHR,but was below the detection level in the kidney,suggesting that aldosterone production in the adrenals is responsible for high circulating aldosterone. However,aldosterone excess in this model was not attrib-utable to conventional aldosterone secretagogues,such as Ang II or hyperkalemia.Recent evidence suggests the presence of nonclassical pathways of adrenal aldosterone production in obesity and metabolic syndrome.Goodfriend et al.[66,67]demon-strated correlation between plasma aldosterone concentra-tion and the amount of visceral fat,and proposed the role of oxidized products of linoleic acid.Ehrhart-Bornstein et al.[68]reported that adipocytes from obese subjects secrete potent aldosterone-releasing factors(ARFs).Although ARFs themselves are as yet unidentified,we compared the aldosterone-releasing activity of adipocytes between SHR/ cp and SHR,according to their method[68].Interestingly, aldosterone production in H295R adrenocortical cells was markedly increased by adipocyte conditioned medium from SHR/cp but not that from nonobese SHR.In parallel, adipocyte conditioned medium from SHR/cp but not from SHR stimulated expression of CYP11B2and steroidogenic acute regulatory protein,another key factor in aldosterone synthesis that mediates transfer of cholesterol to mito-chondria in H295R cells.The activity was not recapitulated by known adipocytokines,including angiotensinogen. Thesefindings suggest involvement of adipocyte-derived substances other than Ang II in hyperaldosteronism in SHR/cp.Aldosterone excess has been reported in other animal models of metabolic syndrome.For example,plasma aldosterone as well as renal Sgk1expression were increased in obese db/db mice,which have a nonfunctional leptin receptor[69,70].Complement-C1q TNF-related protein1(CTRP1)was suggested to participate in hyper-aldosteronemia in db/db mice.This32-kDa adipokine is expressed at high level in adipose tissues of obese db/db mice and Zucker diabetic fatty(fa/fa)rats and has ability to stimulate aldosterone secretion in adrenal cortical cell line H295R[71,72].It should be noted that ARF-mediated hyperaldosteron-emia is not inhibitable by ACEIs or ARBs.Thus,eplere-none should have benefit over Ang II blockade in situations where such factors are overproduced.We expect that ARFs can be a novel target of therapy in metabolic syndrome.Taken together,our data indicate that adipocytes pro-duce ARFs in SHR/cp,which might contribute to elevated circulating aldosterone,podocyte injury,and proteinuria in this model.Salt accelerates renal and cardiac damagein SHR/cp via MR activationRecent clinical studies showed increased salt sensitivity of target organ injury in obese subjects.For example,Verhave et al.[73]demonstrated that higher sodium intake increa-ses urinary albumin excretion in overweight subjects but not in nonoverweight people.However,the mechanisms have not been clearly elucidated.The deleterious effect of aldosterone is known to be augmented under high salt intake.So we examined the effects of salt loading in SHR/cp.High-salt feeding for4weeks markedly enhanced pro-teinuria in SHR/cp.Notably,MR antagonist eplerenone perfectly inhibited the salt-induced exacerbation,suggest-ing involvement of aldosterone/MR cascade.Although salt loading suppressed circulating renin and aldosterone,it paradoxically activated renal MR signaling,as revealed by increased MR in the nuclear fraction,induction of aldo-sterone effector kinase Sgk1,and upregulation of putative mediators of aldosterone-evoked organ damage,such as PAI-1,MCP-1,and TGF-b1in kidney of salt-loaded SHR/ cp.Eplerenone completely inhibited these MR-dependent cascades.The paradoxical MR activation might be attributable in part to adipocyte-derived ARFs.While RAS-regulated aldosterone generation is counterbalanced by salt,our preliminary data suggest that aldosterone production by ARFs lacks negative feedback regulation in response to high salt intake.As a result,suppression of circulating aldosterone level might be less than expected,causing inappropriately high aldosterone for the amount of salt intake.Cross-talk between the kidney and the heart has recently become a major topic.We examined whether the same mechanism can be extrapolated to the pathogenesis of cardiac injury in our rats[74].Left ventricular diastolic function,as revealed by cardiac catheterization study and Doppler echocardiographic analysis,was impaired in salt-loaded SHR/cp,which was accompanied by increased ROS and perivascularfibrosis in the heart.Again,cardiac abnormalities were fully recovered by eplerenone.Thesefindings corroborate our hypothesis that obesity and salt,two cardinal features of modern society,cause MR activation,leading ultimately to CKD and CVD.。