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Endogenous Ouabain (EO) and Adducin enhance the Na-K pump function and play an important role in sodium homeostasis and blood pressure (BP) regulation. In the general population, plasma EO modulates BP either by inhibiting the prohypertensive effect of an excessive salt intake or counteracting the depressor action of normal-moderate salt intake. Almost 50% of hypertensive patients have increased circulating plasma levels of EO. EO has been associated both to left ventricular dysfunction and hypertrophy. A new antihypertensive agent, PST2238, (17beta-(3-furyl)-5beta-androstan-3beta, 14beta, 17alpha-triol a digitoxigenin derivative) able to selectively antagonize both the EO and adducin prohypertensive and molecular effects, has been developed. In hypertensive rats (MHS strain) carrying both adducin mutations and increased plasma EO and in ouabain-infused rats (OS), PST2238 lowers BP by normalizing the renal Na-K pump function. In OS rats, PST antagonized the cardiac and renal pro-hypertrophic ouabain effect associated to the activation of the Src-EGFr-ERK(1/2) signaling cascade. Phase 1 clinical studies demonstrated a high tolerability of PST2238. In a preliminary phase 2 study on 42 mild never-treated hypertensive patients, PST2238 given for 3 months at 0.5 mg/day, significantly reduced BP in subjects with moderate salt intake, implying that it may be selectively effective in conditions where EO plays a prohypertensive role. In conclusion, PST2238, because of its peculiar action mechanism, represents a new tool to disentangle the complex relationship between salt intake, genetic control of renal sodium handling and EO effect.

High-salt diets elevate circulating Na+ pump inhibitors, vascular resistance, and blood pressure. Ouabain induces a form of hypertension mediated via the α2-Na+ pump isoform and the calcium influx mode of the vascular sodium calcium exchanger (NCX). Whereas elevated levels of an endogenous ouabain (EO) and NCX have been implicated in salt-sensitive hypertension, acute changes in sodium balance do not affect plasma EO. This study investigated the impact of longer-term alterations in sodium balance on the circulating levels and renal clearance of EO in normal humans. Thirteen normal men consumed a normal diet, high-salt diet, and hydrochlorothiazide (HCTZ), each for 5-day periods to alter sodium balance. EO and other humoral and urinary variables were determined daily. On a normal diet, urinary sodium excretion (140 ± 16 meq/day), plasma EO (0.43 ± 0.08 nmol/l) and urinary EO excretion (1.04 ± 0.13 nmol/day) were at steady state. On the 3rd day of a high-salt diet, urine sodium excretion (315 ± 28 meq/day), plasma EO (5.8 ± 2.2 nmol/l), and the urinary EO excretion (1.69 ± 0.27 nmol/day) were significantly increased, while plasma renin activity and aldosterone levels were suppressed. The salt-evoked increase in plasma EO was greater in older individuals, in subjects whose baseline circulating EO was higher, and in those with low renal clearance. During HCTZ, body weight decreased and plasma renin activity, aldosterone, and EO (1.71 ± 0.77 nmol/l) rose, while urinary EO excretion remained within the normal range (1.44 ± 0.31 nmol/day). Blood pressure fell in one subject during HCTZ. HPLC of the plasma extracts showed one primary peak of EO immunoreactivity with a retention time equivalent to ouabain. High-salt diets and HCTZ raise plasma EO by stimulating EO secretion, and a J-shaped curve relates sodium balance and EO in healthy men. Under normal dietary conditions, ∼98% of the filtered load of EO is reabsorbed by the kidney, and differences in the circulating levels of EO are strongly influenced by secretion and urinary excretion of EO. The dramatic impact of high-salt diets on plasma EO is consistent with its proposed role as a humoral vasoconstrictor that links salt intake with vascular function in hypertension.

Digitalis-like steroids and related agents have been a mainstay in the treatment of congestive heart failure ever since the publication, in 1785, of Withering’s seminal monograph on foxglove. Heart failure refers to the clinical syndrome that results when the heart is unable to pump sufficient blood to keep up with the metabolic demands of the body. Congestive heart failure is characterized by excessive neuronal and hormonal-mediated fluid retention, expanded intravascular volume, high pulmonary and systemic venous pressures with consequent dyspnea (shortness of breath) on exertion, reduced exercise tolerance, and fatigue. Most heart failure patients also have impaired ventricular systolic function and depressed cardiac output; these are the patients most often treated with digitalis glycosides. These drugs are positive inotropic agents and enhance cardiac contraction. The “cardiotonic steroids” cause cardiac muscle to lose K+ and gain Na+ because they inhibit the Na+ pump (Na+,K+–ATPase), a plasma membrane protein, present in all cells. The Na+ pump uses the energy from ATP to extrude Na+ and …

Increased circulating levels of endogenous ouabain (EO) have been observed in some heart failure patients, but their long term clinical significance is unknown. This study investigated the prognostic value of EO for worsening heart failure among 140 optimally treated patients (age 50+/-14 years; 104 male; NYHA class 1.9+/-0.7) with idiopathic dilated cardiomyopathy. Plasma EO was determined by RIA and by liquid chromatography mass spectrometry, values were linearly correlated (r = 0.89) in regression analysis. During follow-up (13+/-5 months), heart failure progression was defined as worsening clinical condition leading to one or more of the following: sustained increase in conventional therapies, hospitalization, cardiac transplant, or death. NYHA functional class, age, LVEF, peak VO2 and plasma levels of EO were predictive for heart failure progression. Heart failure worsened 1.5 fold (HR: 1.005; 95% CI: 1.001-1.007; p0.01) for each 100 pmol/L increase in plasma EO. Moreover, those patients with higher plasma EO values had an odds ratio of 5.417 (95% CI: 2.044-14.355; p0.001) for heart failure progression. Following multivariate analysis, LVEF, NYHA class and plasma EO remained significantly linked with clinical events. This study provides the first evidence that circulating EO is a novel, independent and incremental marker that predicts the progression of heart failure.

Experimental evidence and observations in humans strongly support an interactive role of mutated α-adducin, sodium (Na+)/potassium (K+)-adenosine triphosphatase (ATPase) activity and endogenous ouabain in Na+homeostasis and the pathogenesis of hypertension. The Ouabain and Adducin for Specific Intervention on Sodium in HyperTension (OASIS-HT) trial is an early Phase II dose-finding study, which will be conducted across 39 European centers. Following a run-in period of 4 weeks without treatment, eligible patients will be randomized to one of five oral doses of rostafuroxin consisting of 0.05, 0.15, 0.5, 1.5, or 5.0 mg/day. Each dose will be compared to a placebo in a double-blind crossover experiment with balanced randomization. Treatment will be initiated with the active drug and continued with placebo or vice versa. Each double-blind period will last 5 weeks. The primary end point is the reduction in systolic blood pressure defined as the average of three clinic readings with the patient in the sitting position. Secondary end points include the reduction in diastolic blood pressure on clinic measurement, the decrease in the 24-h blood pressure, and the incidence of end points related to safety. Secondary objectives are to investigate the dependence of the blood pressure-lowering activity on the plasma concentration of endogenous ouabain and the genetic variation of the enzymes involved in the metabolism of this hormone, and the adducin cytoskeleton proteins. Eligible patients will have Grade I or II systolic hypertension without associated conditions and no more than two additional risk factors. In conclusion, OASIS-HT is a combination of five concurrent crossover studies, one for each dose of rostafuroxin to be studied. To our knowledge, OASIS-HT is the first Phase II dose-finding study in which a genetic hypothesis is driving primary and secondary end points.

ObjectivesThe goal of our study was to investigate the relationships between atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and type A natriuretic peptide receptor (NPRA) gene polymorphisms and left ventricular structure in human essential hypertension.BackgroundExperimental evidence supports a key role for natriuretic peptides in the modulation of cardiac mass. This relationship has not yet been described in human disease.MethodsA total of 203 hypertensive patients were studied by mono-bidimensional echocardiography. Three markers of the ANP gene (−C664G, G1837A, and T2238C polymorphisms) and a microsatellite marker of both NPRA and BNP genes were characterized.ResultsPatients carrying the ANP gene promoter allelic variant had increased left ventricular mass index (117.4 ± 1.7 g vs. 95.7 ± 1.7 g, p = 0.005), left ventricular posterior wall thickness (1.14 ± 0.07 cm vs. 0.96 ± 0.01 cm, p < 0.0001), left ventricular septal thickness (1.12 ± 0.10 cm vs. 1.04 ± 0.01 cm, p = 0.01), and relative wall thickening (47.5 ± 4.1% vs. 39.4 ± 5.3%, p = 0.001) as compared with the wild-type genotype. These associations were independent from anthropometric factors and major clinical features and were confirmed in a large subgroup of never-treated hypertensive patients (n = 148). Carrier status of the ANP gene promoter allelic variant was associated with significantly lower plasma proANP levels: 1,395 ± 104 fmol/ml versus 3,110 ± 141 fmol/ml in hypertensive patients carrying the wild-type genotype (p < 0.05). A significant association for NPRA gene variants with left ventricular mass index and left ventricular septal thickness was found. The analysis of BNP did not reveal any effect on cardiac phenotypes.ConclusionsOur findings show that the ANP/NPRA system significantly contributes to ventricular remodeling in human essential hypertension.

To the Editor: Sodium-dependent mechanisms play a role in the pathogenesis of hypertension. He et al recently demonstrated that in normotensive and hypertensive subjects an acute reduction in salt intake, from 350 to 10 mmol/d for 5 days, was associated with a decline in serum sodium by ≈3 mmol/L.1 Conversely, a progressive increase in salt intake from 10 to 250 mmol/d by a daily amount of 50 mmol caused an increase in serum sodium in normotensive subjects, but not hypertensive patients.1 These investigators speculated that small changes in serum sodium might directly affect the hypothalamic control of blood pressure through the local pituitary renin-angiotensin system.1 Ouabain is a steroid hormone, which is released from the hypothalamus and the adrenal gland. It is implicated in sodium homeostasis and exerts direct actions on the vasculature, the heart,2 and tubular sodium reabsorption.3 In individuals randomly recruited from a Flemish population, blood pressure increased by 2.2 mm Hg systolic and 1.4 mm Hg diastolic for each 50-mmol/d increment in urinary sodium excretion when the plasma ouabain concentration was below the median (140 pmol/L).4 No association between blood pressure and urinary sodium was found when plasma ouabain exceeded the …

Elevated levels of the endogenous ouabain (EO), a closely related isomer of ouabain, are implicated in rat and human hypertension and in related cardiovascular complications. The pathogenetic mechanisms through which EO affects the cardiovascular system involve the modulation of the renal Na/K-ATPase, implicated in renal tubular sodium reabsorption, and the activation of signal transduction pathways, promoting the transcription of growth-related genes. Experimental and clinical evidence on rats and humans stimulated the pharmacological research for developing novel anti-hypertensive agents able to antagonize the cellular and molecular alterations mediated by EO. Among them, the digitoxigenin derivate, PST 2238, has been selected for its ability to antagonize the ouabain-induced effects on blood pressure and organ hypertrophy at oral doses of microgram/kg/day in vivo. The pharmacological selectivity and safety of PST 2238 suggests that the compound may be effective for the treatment of those forms of hypertension in which renal sodium handling alterations and cardiovascular complications are associated with increased production of EO.

Renin-angiotensin system reactivity and the constitutive capacity of the renal tubule to reabsorb sodium play a role in the individual response to diuretic therapy; therefore we evaluated the blood pressure (BP) response to hydrochlorothiazide in 87 never-treated individuals with mild essential hypertension, according to ACE gene I/D and α-adducin Gly460Trp polymorphism. These genotypes where chosen because previous data showed their interaction in determining the BP response to salt probably was the result of their involvement in the activation of the renin-angiotensin system (ACE) and in the constitutive capacity of the kidney to reabsorb sodium (α-adducin) (treatment for 2 months). BP was measured after 3 run-in visits and after the first and second months of treatment by means of a standardized procedure. Data were analyzed by ANOVA, t test, and multivariate ANOVA for repeated measures (covarying for gender, age, and body mass index). Although basal mean BP (MBP) was similar in the different ACE and α-adducin genotypes, patients carrying at least one I allele of ACE and one 460Trp allele of α-adducin had the largest MBP decrease with treatment (12.7±1.9 mm Hg), the effect of the combination of genotypes being additive but not epistatic. These patients had an odds ratio of 15.75 of being a responder to hydrochlorothiazide compared with patients with Gly460Gly+DD, with the least MBP decrease (3.4±1.7 mm Hg). α-Adducin and ACE I/D polymorphism may be useful to predict the interindividual degree of response to hydrochlorothiazide; the analysis of the combination of the 2 genotypes increases the accuracy of the prediction of response to the drug.

OBJECTIVE: Experimental data show that ouabain is a modulator of the sodium-potassium pump, which plays an important role in sodium homeostasis and blood pressure regulation. We investigated the distribution of plasma ouabain in the general population in relation to blood pressure and other determinants of sodium homeostasis. METHODS: In 379 subjects enrolled in a Belgian population study, we measured plasma ouabain, clinical characteristics including blood pressure, serum and urinary electrolytes, urinary aldosterone excretion, various lifestyle factors, and the Gly460Trp polymorphism of the alpha-adducin gene. Our statistical methods included analysis of covariance and multiple linear regression. RESULTS: Plasma ouabain (median, 140 pmol/l) correlated independently and positively with male gender (n = 182, P = 0.002), smoking (n = 116, P = 0.05), urinary potassium excretion (mean 69 mmol/day, P < 0.0001), and mutation of the alpha-adducin gene (n = 161, P < 0.0001). Both before and after adjustment for covariables, continuous as well as categorical analyses revealed a significant interaction (P < or = 0.02) between plasma ouabain and urinary sodium excretion (mean 194 mmol/day) in relation to blood pressure (mean systolic blood pressure/diastolic blood pressure, 123/76 mmHg). In individuals with plasma ouabain values below the median, blood pressure increased by 2.2 mmHg systolic and 1.4 mmHg diastolic for each 50 mmol/day increment in urinary sodium excretion (P < or = 0.01). No association between blood pressure and urinary sodium excretion was found when plasma ouabain exceeded the median. CONCLUSIONS: Plasma ouabain behaves as a blood pressure modulating factor, possibly released in response to potassium, either inhibiting the pressor effect of an excessive salt intake or counteracting the depressor action of sodium depletion. ispartof: Journal of Hypertension vol:21 issue:8 pages:1475-81 ispartof: location:Netherlands status: published

Human hypertension is a complex, multifactorial disease with a heritability of more than 30-50%. A genetic screening test based on analysis of multiple single-nucleotide polymorphisms (SNPs) to assess the likelihood of developing hypertension would be helpful for disease management.Tailed allele-specific primers were designed to amplify by PCR six biallelic SNP loci [three in G protein-coupled receptor kinase type 4 (GRK4): R65L, A142V, and A486V; two in angiotensinogen: -6G--A and M235T; and one in aldosterone synthase: -344C--T] associated with essential hypertension. PCRs of SNP loci were coupled (via a common sequence of 21 nucleotide tails) to incorporate Universal Amplifluor(TM) primers labeled with fluorescein or sulforhodamine in a homogeneous format. Use of Amplifluors in SNP PCRs produced labeled amplicons, the fluorescence of which was quantified by a microplate reader and then analyzed via an Excel macro to provide genotypes for all six SNP loci. Unique restriction endonucleases were identified for five SNP loci that could independently confirm homogeneous PCR results when needed.We developed six homogeneous PCR assays that were set up, performed, and fluorometrically analyzed in 96-well microplates. Allele frequencies were determined for six SNPs in 60 Italian hypertensive patients and a control group of 60 normotensive persons. A significant correlation (P = 0.034) between one SNP [GRK4 (A486V)] and the hypertensive patients was observed. Genotyping results for five of six SNPs were confirmed by digesting corresponding amplicons with locus-specific restriction endonucleases.We developed a simple and homogeneous fluorescent protocol that has been used to determine the SNP genotype for six loci in a population of hypertensive and normotensive persons. We also observed a significant association (P = 0.034) between one SNP (A486V) and an Italian population of mildly hypertensive patients.

Background Genes involved in cytoskeletal assembly and water/electrolyte balance have been previously linked to hypertension, but recent evidences suggest a role also in the development of inflammation and autoimmunity [1-2]. Systemic Lupus Erythematosus is a prototypic multi-organ autoimmune disease, characterized by polymorphic clinical features and a complex polygenic background. Renal disease in SLE occurs in 40-70% of lupus patients and is responsible of an high burden of morbidity and mortality. Despite substantial efforts, less is known about the role of specific genetic factors in conferring susceptibility to SLE and its complications, including lupus nephritis (LN) [3]. Objectives With these premises we sought if an association between known hypertension-related polymorphisms and development of SLE and LN existed. Methods 106 patients (96 females, 10 males) diagnosed with SLE (n=51) or LN (n=55), 23 patients with Sjoegren9s Syndrome and 62 healthy controls were enrolled. Patients and controls were genotyped for polymorphisms in α-, β- and γ-adducin (ADD1,2,3 respectively), angiotensin converting enzyme, transient receptor potential cation channel-subfamily C, solute carrier family 8 (sodium-calcium exchanger) member 1 (SLC8A1), solute carrier family 24 (sodium/potassium/calcium exchanger) member 3, PKD2 calcium channel and PRKG1 kinase genes. Results Beta-adducin (ADD2) rs4984 CT heterozygosis was significantly more frequent in SLE patients than in LN patients (χ 2 =6.154; p=0.013). No patient had the ADD2 rs4984 TT genotype in accordance with a low prevalence of the ADD2 C1797T homozygosity in the population. Seven patients with SLE (n=3) or LN (n=4), but none of the control groups carried the AA variant of sodium/calcium exchanger 1 (SLC8A1) rs11893826 SNP (χ 2 =11.771; p=0.019). Conclusions Genes involved in the control of electrolyte/water balance and cytoskeletal plasticity could affect the pathogenic background of SLE/LN. Larger studies are required to confirm these data and attempt correlations between genetic data and specific clinical features. References Tintinger GR, et al., Drug Des Devel Ther, 2009 Kleinewietfeld M, et al., Nature, 2013 Ramos PS, et al., Semin Nephrol, 2010 Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.3378

Altered sympathetic nervous system activity has been implicated often in hypertension. We examined short-term potentiation [posttetanic potentiation (PTP)] and long-term potentiation (LTP) in the isolated superior cervical ganglia (SCG) from Sprague-Dawley (SD) rats given vehicle, digoxin, or ouabain by subcutaneous implants as well as in animals with ouabain-induced hypertension (OHR), and inbred Baltimore ouabain-resistant (BOR) and Baltimore ouabain-sensitive (BOS) strains of rats. Postganglionic compound action potentials (CAP) were used to determine PTP and LTP following a tetanic stimulus (20 Hz, 20 s). Baseline CAP magnitude was greater in ganglia from OHR than in vehicle-treated SD rats before tetanus, but the decay time constant of PTP was significantly decreased in OHR and in rats infused with digoxin that were normotensive. In hypertensive BOS and OHR, the time constants for the decay of both PTP and LTP ( t L) were increased and correlated with blood pressure (slope = 0.15 min/mmHg, r = 0.52, P < 0.047 and 6.7 min/mmHg, r = 0.906, P < 0.0001, respectively). In BOS and OHR, t L (minutes) was 492 ± 40 ( n = 7) and 539 ± 41 ( n = 5), respectively, and differed ( P < 0.05) from BOR (257 ± 48, n = 4), SD vehicle rats (240 ± 18, n = 4), and captopril-treated OHR (370 ± 52, n = 5). After the tetanus, the CAP at 90 min in BOS and OHR SCG declined less rapidly vs. SD vehicle rats or BOR. Captopril normalized blood pressure and t L in OHR. We conclude that the duration of ganglionic LTP and blood pressure are tightly linked in ouabain-dependent hypertension. Our results favor the possibility that enhanced duration of LTP in sympathetic neurons contributes to the increase in sympathetic nerve activity in ouabain-dependent hypertension and suggest that a captopril-sensitive step mediates the link of ouabain with LTP.

P20 Elevated levels of an endogenous ouabain (EO) occur in many patients with essential hypertension (e.g., Manunta et al , Hypertension 1999;34:450-456) and the prolonged infusion of ouabain induces hypertension (Manunta et al J. Hypertension 1994;12:549-560). This study used a series of closely related structural analogs of ouabain to test the key hypothesis that sodium pump inhibition underlies ouabain-dependent hypertension. The functional roles of the sugar and lactone ring were tested for their effects on inhibition of the Na,K-ATPase, and on long term blood pressure. Normal male SD rats received infusions of vehicle (VEH), rhamnose (RHA), ouabain (OUA), ouabagenin (OGN), dihydroouabain (DHO), isoouabain (ISO), and ouabain in which the lactone ring was opened (ORO)at 30 μg/Kg/24 hr for 4-5 weeks via s.c. osmotic pumps. Cuff pressures were taken weekly. At the end of the study, blood and tissues were harvested, extracted by C-18 and analyzed by ouabain-RIA using appropriate standards. HPLC showed that plasma and tissues contained one main peak of immunoreactivity corresponding to the infused agent. Systolic pressures (SBP)in VEH, RHA, OUA, OGN, DHO, ISO and ORO were 132±2.5, 133±1.5, 159±2.6*, 154±4*, 167±4*†, 171±2.2*†, 169±2.4*†,respectively (*P

We sought to evaluate the relationships among circulating levels of an endogenous ouabain-like factor (EO) and systemic hemodynamics and left ventricular (LV) geometry in patients with recently diagnosed essential hypertension. We selected 92 never-treated patients with essential hypertension. Blood samples were drawn for estimation of plasma EO (radioimmunoassay) and subjects underwent echocardiographic examination to evaluate LV end-systolic and end-diastolic wall thickness and internal dimensions. LV volumes, stroke volume, cardiac output, total peripheral resistance, LV mass, and relative wall thickness were calculated, and all except the last parameter were indexed by body surface area. LV mass also was indexed by height. On the basis of the values of LV mass index (body surface area or height) and relative wall thickness, subjects were divided into groups with either normal geometry, concentric remodeling, concentric hypertrophy, or eccentric nondilated hypertrophy. In the study population as a whole, circulating EO levels were significantly and directly correlated with mean blood pressure (r = 0.21, P = .048), relative wall thickness (r = 0.34, P = .001), and total peripheral resistance index (r = 0.37, P = .0003). Plasma EO also was significantly and inversely correlated with LV end-diastolic volume index (r = -0.32, P = .002), stroke index (r = -0.34, P = .0009), and cardiac index (r = -0.35, P = .0007). In multiple regression analysis, plasma EO was an independent correlate of total peripheral resistance index, cardiac index, and relative wall thickness. Regardless of the indexation method used for LV mass, plasma EO was higher in patients with concentric remodeling than in those with either normal geometry or concentric hypertrophy. Plasma EO tended to be higher (indexation by body surface area) or was significantly higher (indexation by height) in subjects with concentric remodeling than in those with eccentric nondilated hypertrophy. Patients with concentric remodeling showed the highest total peripheral resistance index and the lowest cardiac index. Our data suggest that EO plays a role in regulating systemic hemodynamics and LV geometry in patients with essential hypertension.

To investigate vascular mechanisms in hypertension, we isolated renal arterial rings from rats with ouabain-dependent hypertension and studied their function. In rats infused with ouabain for 5 weeks, systolic and mean blood pressures (BP) were increased relative to controls. Contractions evoked by high KCl solutions were greater in rings from ouabain-infused rats whereas the threshold concentrations and EC50s for KCl and the peak caffeine contractures were not different. KCl contractures were not affected by 5 microM prazosin. Phenylephrine contractures were increased marginally in ouabain-infused rats, while acetylcholine-induced relaxation was normal. In vitro superfusion of rings with 10 nM ouabain or digoxin did not affect the measured parameters. Plasma ouabain, BP, and all evoked responses were normal one week following interruption of the ouabain infusion. In a second study, BP increased in ouabain (15 microg/kg/day, n= 23), but not digoxin (30 microg/kg/day, n=12), or vehicle-infused (n=16) rats. KCl contractures were greater in rings from ouabain-but decreased in rings from digoxin-infused rats, respectively and correlated with systolic and mean BP (r=0.69, n=30, p

This review focuses on the most recent data published in the field of the sodium-potassium pump inhibitors regarding the hypothetical structure, the secretory stimuli and the pathophysiological implications for particular diseases, such as hypertension. On the basis of the findings published so far, we reconsider and discuss the 'natriuretic hypothesis' for explaining the role of the endogenous sodium-potassium ATPase inhibitor. We propose the ouabain-like factor as a modulator of the renal sodium-potassium pump, that can be considered as a new pharmacological target for hypertension therapy.

Adducin point mutations are associated with genetic hypertension in Milan hypertensive strain (MHS) rats and in humans. In transfected cells, adducin affects actin cytoskeleton organization and increases the Na+-K+-pump rate. The present study has investigated whether rat and human adducin polymorphisms differently modulate rat renal Na+-K+-ATPase in vitro. We report the following. 1) Both rat and human adducins stimulate Na+-K+-ATPase activity, with apparent affinity in tens of nanomolar concentrations. 2) MHS and Milan normotensive strain (MNS) adducins raise the apparent ATP affinity for Na+-K+-ATPase. 3) The mechanism of action of adducin appears to involve a selective acceleration of the rate of the conformational change E2(K) → E1(Na) or E2(K) ⋅ ATP → E1Na ⋅ ATP. 4) Apparent affinities for mutant rat and human adducins are significantly higher than those for wild types. 5) Recombinant human α- and β-adducins stimulate Na+-K+-ATPase activity, as do the COOH-terminal tails, and the mutant proteins display higher affinities than the wild types. 6) The cytoskeletal protein ankyrin, which is known to bind to Na+-K+-ATPase, also stimulates enzyme activity, whereas BSA is without effect; the effects of adducin and ankyrin when acting together are not additive. 7) Pig kidney medulla microsomes appear to contain endogenous adducin; in contrast with purified pig kidney Na+-K+-ATPase, which does not contain adducin, added adducin stimulates the Na+-K+-ATPase activity of microsomes only about one-half as much as that of purified Na+-K+-ATPase. Our findings strongly imply the existence of a direct and specific interaction between adducin and Na+-K+-ATPase in vitro and also suggest the possibility of such an interaction in intact renal membranes.

Abstract —Many patients with essential hypertension (EH) exhibit increased left ventricular mass. Similarly, elevated circulating levels of an endogenous ouabainlike factor (OLF) have been described in some but not all patients with EH. Moreover, ouabain has a hypertrophic influence on isolated cardiac myocytes. Accordingly, we investigated relationships among plasma OLF, left ventricular mass, and cardiac function in patients with EH. Plasma OLF was determined in 110 normotensive subjects and 128 patients with EH. Echocardiographic parameters and humoral determinants were measured in EH. Plasma OLF levels were increased ( P P =0.03). Left ventricular mass index and stroke volume in EH with high OLF were greater than in EH with normal OLF (101.9±3.3 versus 86.1±2.5 g/m 2 , P =0.0003, and 57.10±1.48 versus 52.30±1.14 mL/m 2 , P =0.02, respectively), although heart rate was slower (74.2±1.3 versus 80.5±1.3 bpm, P =0.005). Multiple regression analysis that tested the influence of body mass index, age, gender, 24-hour blood pressure, and OLF on left ventricular mass revealed independent contributions of systolic (13.2%) and diastolic (12.4%) blood pressure and plasma OLF (11.6%) to left ventricular mass. We conclude that ≈50% of patients with uncomplicated EH have elevated-high circulating OLF levels, higher diastolic blood pressure, greater left ventricular mass and stroke volume, and reduced heart rate. We propose that the OLF affects cardiovascular function and structure and should be considered as a factor that contributes to the risk of morbid events.

Summary Background Abnormalities in renal sodium transport may be involved in hypertension. Adducin, an α/β heterodimeric protein found in the renal tubule is thought to regulate ion transport through changes in the actin cytoskeleton. We investigated whether an α-adducin polymorphism (Gly 460 Trp) is involved in essential hypertension in two separate populations. Methods Linkage analysis of three DNA markers at different distances from the α-adducin locus (20-2500 kb) was done in 137 hypertensive sibling-pairs. 477 hypertensive and 322 normotensive individuals were genotyped for the α-adducin polymorphism. The blood-pressure response to acute and chronic changes in sodium balance was studied in hypertensive individuals with and without the 460 Trp α-adducin allele. Findings Significant linkage was found for all three markers in the sibling-pair study. The extra shared alleles (9·1%, 6·5%, and 4·7%) and the significance level for linkage (p=0·0006, p=0·0119, and p=0·0211) both decreased with increasing distance from the α-adducin locus. There was a significant association between the 460 Trp mutation and hypertension (p=0·0003). In the salt-sensitivity test, to assess the acute blood-pressure response to changes in body sodium in 86 hypertensive patients, the decrease in mean arterial pressure was greater in 65 patients who were heterozygous for the mutant allele (Gly/Trp) than in 21 wild-type homozygotes (Gly/Gly) (mean decrease 15·9 [SE 2·0] vs 7·4 [1·3] mm Hg; p=0·001). Similarly, 21 heterozygous hypertensive patients showed a greater fall in mean arterial pressure in response to 2 months' treatment with hydrochlorothiazide than did 37 wild-type homozygous hypertensive patients (mean decrease 14·7 [2·2] vs 6·8 [1·4] mm Hg; p=0·002). Interpretation Our findings of significant linkage of the α-adducin locus to essential hypertension and greater sensitivity to changes in sodium balance among patients with the mutant allele suggest that α-adducin is associated with a salt-sensitive form of essential hypertension. We suggest the α-adducin polymorphism may identify hypertensive patients who will benefit from diuretic treatment or manoeuvres to reduce total body sodium.

increased ( PRA supine 4.2 ng/ml/h with n.v. 0.2‐2.8 ng/ml/h; PRA standing 8.0 ng/ml/h with n.v. 1.5‐5.7 ng/ml/h; serum aldosterone supine 230 pg/ml with n.v. 30‐150 pg/ml; serum aldosterone standing Case Report 393 pg/ml with n.v. 70‐350 pg/ml ), in spite of normal values of both serum and urinary sodium and potassium in two collections (142.2 mEq/l and 4.14 mEq/l A 16-year-old girl was admitted to the Division of with 142.4 mEq/24 h and 50.3 mEq/24 h respectively

To assess possible relationships between endogenous ouabain, sodium balance and blood pressure.This review concerns the structure of endogenous ouabain, circulating levels of this steroid in various disorders of fluid and electrolyte balance, recent evidence for the association of endogenous ouabain with human hypertension, the influence of sodium and volume factors on ouabain-induced hypertension, and possible mechanisms for the hypertensinogenic activity of ouabain.The human circulation contains a closely related isomer of ouabain of putative adrenocortical origin. Elevated circulating levels of this 'endogenous ouabain' are common but not universal in physiologic and pathologic states associated with positive sodium balance or high blood pressure, or both. In the absence of adrenal hyperfunction, elevating circulating levels of endogenous ouabain appear to be secondary to impaired renal clearance. Prolonged elevation of circulating ouabain in the rat induces sustained hypertension. This model exhibits normal plasma renin activity, increased levels of ouabain in the hypothalamus, pituitary, and kidney, and responds to angiotensin converting enzyme inhibitor. In rats with normal kidney function, ouabain-induced hypertension is primarily sodium-insensitive although maneuvers that hinder renal sodium excretion augment the pressor effect of this steroid. Prolonged administration of ouabain into the brain ventricles augments sympathetic nervous system activity and induces sustained hypertension. These observations lead us to propose the following hypothesis. Among Caucasian patients with essential hypertension, a large fraction have elevated circulating levels of endogenous ouabain, possibly caused by an inherited or acquired renal defect in clearance of this steroid. In these patients, and in rats with ouabain-induced hypertension, increased local generation of, or increased target organ sensitivity to, angiotensin II, or both, may contribute critically to heightened vasoconstriction and a sustained increase in blood pressure. Investigations of the efferent pressor mechanisms and the renal handling of endogenous ouabain are novel approaches to the etiology and therapy of several common cardiovascular disorders.

PURPOSE: To evaluate screening with three-dimensional (3D) phase-contrast magnetic resonance (MR) angiography with a phased-array multicoil to detect renal artery stenosis. MATERIALS AND METHODS: Fifty consecutive patients suspected of having renovascular disease were prospectively examined with 3D phase-contrast MR imaging with a phased-array multicoil. Findings were correlated with those at intra-arterial digital subtraction angiography (DSA) as the standard of reference for grade of stenosis. RESULTS: MR angiography depicted 101 of 103 renal arteries depicted at intraarterial DSA; the two missed arteries were an accessory artery outside the imaging volume and an artery with a stent. At intraarterial DSA, a stenosis was found in 31 of 101 arteries. On the basis of findings at 3D phase-contrast MR angiography, the presence of any degree of stenosis was correctly depicted in 29 of 31 cases and the absence of stenosis was correctly depicted in 66 of 70 cases (accuracy, 94%; sensitivity, 94%; negative predictive value, 97%). Overall accuracy was 97% for correct depiction of severe renal artery stenosis (> 50%). CONCLUSION: 3D phase-contrast MR angiography with a phased-array multicoil was an accurate noninvasive screening technique in patients with suspected renal artery stenosis

Genetics of hypertension with its disappointing results may evoke reactions ranging from enthusiasm and hope to suspicion and skepticism. A variety of reasons may account for these contrasting results1 that cannot be discussed here. However, in our opinion, 2 of these reasons are the most important ones: First, there is clear contradiction between the well established notion of the enormous heterogeneity of hypertension (in terms of molecular, biological, and clinical mechanisms) and the demand that a gene variant must be active across different populations, race, age, gender, etc. Second, the probability models of frequentist statistics, based on randomness, trivialize the ordered complexity of biological systems based on natural laws.2 In fact, all the available statistical genetic guidelines for defining causation of a single gene in a complex disease, like hypertension, are unable to fully account for genetic or environmental interactions.3 More than 50 years of pathophysiological, clinical, and epidemiological studies have clearly shown that these interactions are of paramount importance for the elevation of blood pressure. In fact, it is highly unlikely that the same molecular mechanism may account for a disease that affects up to 40% of the adult population of the industrialized countries. Interactions among these factor varieties can only be inferred from empirical observations and not from deductive mathematical logic.3,4 Along this line, studies trying to correlate genotypes with intermediate phenotypes, that are along biologically plausible pathways linking the genes of interest to blood pressure, may reduce this complexity and, therefore, provide more consistent data across the different studies.4 Intermediate phenotypes are also indispensable to dissect the heterogeneity of this human condition …

Knowledge of the air and land surface temperature and their temporal and spatial variations within a city environment is of prime importance to the study of urban climate and human-environment interactions and to monitoring environmental changes due to urbanization. We present a number of air and land surface temperature products that have been produced, archived, evaluated, and analyzed for 10 European cities within the framework of the European Space Agency-funded "Urban Heat Islands and Urban Thermography" project. We evaluate in what way these products are suited to explore the urban thermal dynamics and how products with different temporal and spatial resolution can provide a complementary view, both for thermal patterns as well as heat waves. Level of confidence was evaluated through quantitative, qualitative, and user-based analyses.

Ouabain has recently been identified as an endogenous Na(+)-K+ pump inhibitor. We administered ouabain chronically to normotensive rats with varying degrees of reduced renal mass (RRM) and to normal two-kidney rats to see whether hypertension could be produced. Normal male Wistar rats and rats with 25%, 60%, and 70% RRM received ouabain (13.9 micrograms/kg per day IP) in normal saline for 4 weeks followed by ouabain (27.8 micrograms/kg per day IP) for 3 to 4 more weeks. Respective control animals received vehicle only. Blood pressure was recorded weekly by tail plethysmography. Animals received tap water and standard rat chow, except for 70% RRM rats, which received distilled water and sodium-free chow. After 6 to 8 weeks of treatment, with rats under thiobutabarbital anesthesia, direct blood pressure was determined. Plasma, tissue, and urinary ouabain levels were measured with a specific radioimmunoassay. Animals receiving ouabain developed significant increases in mean blood pressure compared with control animals (70% RRM, 147 +/- 4 vs 116 +/- 4 mm Hg; 60% RRM, 140 +/- 4 vs 107 +/- 3 mm Hg; 25% RRM, 131 +/- 5 vs 100 +/- 2 mm Hg; no RRM, 116 +/- 4 vs 98 +/- 5 mm Hg). Plasma ouabain levels measured 24 hours after the last ouabain dose were not different in animals receiving ouabain vs those receiving vehicle. However, kidney tissue ouabain levels were significantly greater (6.39 +/- 1.17 vs 2.36 +/- 0.52 micrograms/kg, P < .05) in animals receiving ouabain. In conclusion, ouabain, given chronically, is associated with the development of hypertension in RRM rats as well as in normal rats. Blood pressure was greater in animals with greater degrees of RRM for a given ouabain dose.

1. To evaluate whether sodium intake can modulate the action of endogenous kinins on renal function and haemodynamics, a receptor antagonist of bradykinin was infused in conscious normotensive rats maintained on either a normal or a low sodium diet. 2. The antagonist inhibited the hypotensive effect of exogenously administered bradykinin. It did not change the vasodepressor effect of acetylcholine, dopamine or prostaglandin E2. 3. The antagonist did not affect mean blood pressure, glomerular filtration rate, renal blood flow or urinary sodium excretion, in rats on sodium restriction. It did not change mean blood pressure, glomerular filtration rate or urinary sodium excretion, but decreased renal blood flow, in rats on a normal sodium intake. 4. The kallikrein–kinin system has a role in the regulation of renal blood flow in rats on a normal sodium diet.

ObjectivesTo determine the steady-state dose-dependence of blood pressure, plasma and tissue ouabain during continuous infusion of ouabain in the rat, and to evaluate the adrenal dependence and effect of a high salt intake on this form of hypertension.Design and methodsOuabain was administered, via