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Background Amiloride is a competitive blocker of the epithelial sodium (Na) channel in the renal collecting duct. It is a less potent diuretic than thiazides or loop diuretics, but is often used in association with its potassium (K)-sparing profile. Whether amiloride has a hypocalciuric effect similar to thiazides remains unclear. Animal studies and experiments on cell lines suggested that amiloride increases calcium (Ca) reabsorption in the distal nephron, but human studies are scarce. Methods We performed a post hoc analysis of a study with 48 healthy males (mean ± standard deviation age, 23.2 ± 3.9 years) who were assigned to a high-Na/low-K diet for 7 days before receiving 20 mg of amiloride orally. Urinary excretions of electrolytes were measured at 3 and 6 h afterwards; we calculated the relative changes in urinary excretion rates after amiloride administration. Results The high-Na/low-K diet led to an expected suppression of plasma renin and aldosterone. Amiloride showed a mild natriuretic effect associated with a decreased kaliuresis. Urinary Ca excretion dropped substantially (by 80%) 3 h after amiloride administration and remained low at the sixth hour. At the same time, fractional excretion of lithium decreased by a third, reflecting an increased proximal tubular reabsorption. Conclusions During a high-Na/low-K diet, amiloride had a strong acute hypocalciuric effect, most probably mediated by increased proximal Ca reabsorption, even though a distal effect cannot be excluded. Further studies should establish if chronic amiloride or combined amiloride/thiazide treatment may decrease calciuria more efficiently and be useful in preventing kidney stones.

OBJECTIVE There is increasing evidence that sodium can be stored in the skin and muscles without being osmotically active, yet whether acute changes in dietary sodium intake alter sweat and muscle sodium content has not been investigated previously. METHODS In a cross-over design, we assessed muscle sodium content by Na-MRI in 38 healthy normotensive volunteers (aged 33.5 ± 11.1 years, 76.3% women) after 5 days of high-sodium diet (6 g of salt added to their normal diet) and 5 days of a low-sodium diet. In a subgroup of 18 participants (72.2% women) we conducted quantitative pilocarpine iontophoretic sweat collections and measured the sodium concentration in sweat. Plasma aldosterone and plasma renin activity levels were measured in all participants. RESULTS Under high-sodium diet conditions urinary sodium excretion, muscle sodium content and sweat sodium concentration all increased significantly. Muscle sodium content (rm = 0.47, P = 0.03) and sodium sweat concentration (rm = 0.72, P

Glomerular filtration rate (GFR), or the rate of primary urine formation, is the key indicator of renal function. Studies have demonstrated that GFR exhibits significant circadian rhythmicity and, that these rhythms are disrupted in a number of pathologies. Here, we tested a hypothesis that the circadian rhythm of GFR is driven by intrinsic glomerular circadian clocks. We used mice lacking the circadian clock protein BMAL1 specifically in podocytes, highly specialized glomerular cells critically involved in the process of glomerular filtration (Bmal1lox/lox/Nphs2-rtTA/LC1 or, cKO mice). Circadian transcriptome profiling performed on isolated glomeruli from control and cKO mice revealed that the circadian clock controls expression of multiple genes encoding proteins essential for normal podocyte function. Direct assessment of glomerular filtration by inulin clearance demonstrated that circadian rhythmicity in GFR was lost in cKO mice that displayed an ultradian rhythm of GFR with 12-h periodicity. The disruption of circadian rhythmicity in GFR was paralleled by significant changes in circadian patterns of urinary creatinine, sodium, potassium and water excretion and by alteration in the diurnal pattern of plasma aldosterone levels. Collectively, these results indicate that the intrinsic circadian clock in podocytes participate in circadian rhythmicity of GFR.

BACKGROUND Renal sodium handling could be a potential mediator linking adipokines to hypertension. The aim of the study was to assess the relationship of leptin with urinary sodium excretion and proximal sodium reabsorption in humans. METHODS This cross-sectional study was conducted on participants of hypertensive families from the Seychelles Island. A split urine (daytime and nighttime) collection and plasma leptin were measured. Endogenous lithium clearance was used to assess proximal sodium reabsorption. Mixed multiple linear regression tests adjusted for confounding factors were used. RESULTS Three hundred and sixty-five participants (57% women) were included in this analysis. Leptin and adiponectin were higher in women (P < 0.001). Leptin was associated positively with daytime (coefficient [c]: 0.16, standard deviation (SD): 0.03, P < 0.001), nighttime urinary sodium excretion (c: 0.17, SD: 0.04), P < 0.01), daytime lithium clearance (c: 0.40, SD: 0.08, P < 0.001), and nighttime lithium clearance (c: 0.39, SD: 0.10, P < 0.001) after adjusting for sex. The association was lost or mitigated only when BMI was introduced in the model. When BMI was categorized in normal vs. overweight participant, leptin was associated with daytime and nighttime sodium excretion rates (c: 0.14, SD: 0.05, P = 0.011 and c: 0.22, SD: 0.07, P = 0.002, respectively) only in overweight participants. CONCLUSION Leptin is associated positively with daytime and nighttime sodium excretion and lithium clearance suggesting a natriuretic rather than a sodium retaining effect of leptin. Sex and body mass index (BMI) are major confounders in this association. These results highlight the importance of sex and obesity in our understanding of the relationships between leptin, blood pressure, and renal sodium handling.

Aprocitentan is a novel, oral, dual endothelin receptor antagonist (ERA) in development in difficult-to-control hypertension. As fluid retention and edema are concerns with ERAs, we investigated whether aprocitentan causes weight gain in healthy subjects on a high sodium diet and explored potential mechanisms if occurring. This double-blind, randomized, placebo-controlled, crossover study enrolled 28 subjects. Three doses of aprocitentan (10, 25, or 50 mg/day for 9 days) were compared with placebo. Increases in body weight were observed with aprocitentan (placebo-corrected mean weight gains [90% confidence interval]) of 0.43 [0.05-0.80], 0.77 [0.03-1.51], and 0.83 [0.33-1.32] kg at 10 mg, 25 mg, and 50 mg, respectively. Decreases in hemoglobin and uric acid were observed. Plasma volume increased at most by 5.5% without dose-response relationship. Urinary sodium excretion decreased at 10 mg and 25 mg but not at 50 mg. Therefore, aprocitentan produced moderate weight increases in healthy subjects on high sodium diet, without obvious sodium retention.

Background The effects of sodium (Na+) intakes on renal handling of potassium (K+) are insufficiently studied. Methods We assessed the effect of Na+ on renal K+ handling in 16 healthy males assigned to three 7-day periods on low salt diet [LSD, 3 g sodium chloride (NaCl)/day], normal salt diet (NSD, 6 g NaCl/day) and high salt diet (HSD, 15 g NaCl/day), with constant K+ intake. Contributions of distal NaCl co-transporter and epithelial Na+ channel in the collecting system on K+ and Na+ handling were assessed at steady state by acute response to 100 mg oral hydrochlorothiazide and with addition of 10 mg of amiloride to hydrochlorothiazide, respectively. Results Diurnal blood pressure slightly increased from 119.30 ± 7.95 mmHg under LSD to 123.00 ± 7.50 mmHg (P = 0.02) under HSD, while estimated glomerular filtration rate increased from 133.20 ± 34.68 mL/min under LSD to 187.00 ± 49.10 under HSD (P = 0.005). The 24-h K+ excretion remained stable on all Na+ intakes (66.28 ± 19.12 mmol/24 h under LSD; 55.91 ± 21.17 mmol/24 h under NSD; and 66.81 ± 20.72 under HSD, P = 0.9). The hydrochlorothiazide-induced natriuresis was the highest under HSD (30.22 ± 12.53 mmol/h) and the lowest under LSD (15.38 ± 8.94 mmol/h, P = 0.02). Hydrochlorothiazide increased kaliuresis and amiloride decreased kaliuresis similarly on all three diets. Conclusions Neither spontaneous nor diuretic-induced K+ excretion was influenced by Na+ intake in healthy male subjects. However, the respective contribution of the distal convoluted tubule and the collecting duct to renal Na+ handling was dependent on dietary Na+ intake.

Background and ObjectivesThe pathophysiological mechanisms linking weight loss to blood pressure (BP) reduction are not completely understood. The objective of this study was to compare the effect of weight loss after Roux-en-Y gastric bypass (RYGB) on BP, renin-angiotensin-aldosterone system (RAAS), and urinary electrolytes excretion to those of dietary advice.MethodsThis was a case-control prospective study including obese patients referred for RYGB (cases) and obese receiving diet advice only (controls). Ambulatory BP, plasma renin activity (PRA), plasma aldosterone concentration (PAC), and urinary electrolytes were measured before (M0) and after intervention (M3: 3 months and M12: 12 months).ResultsTwenty-five patients were included in the RYGB group and twelve patients in the control group. After 12 months, weight loss (-42 ± 11.5 vs -12.3 ± 6.3 kg in the control group, p=0.001) and decrease in PAC were more pronounced in the RYGB group (-34 ± 76 vs +14 ± 45 pg/ml in the control group, p=0.002). There was no difference in PRA between both groups (-0.08 ± 1.68 vs 0.01 ± 0.37 ng/ml/h, p=0.31). Sodium excretion was more marked in the RYGB group after 3 months only (-89 ± 14.9 vs -9.9 ± 27.9 mmol/day, p=0.009). The decrease in SBP was similar between both groups (-6.9 ± 9.9 vs -7.1 ± 11.9 mmHg in the control group, p=0.96).ConclusionsBariatric-induced weight loss induces a progressive decrease in PAC independently of PRA and sodium excretion. Whether this decrease in PAC affects target organ damage in the long term remains to be determined.Clinical Trial RegistrationClinicalTrials.gov, identifier NCT02218112.

Key pointsAcclimatization to hypoxia leads to a reduction in plasma volume (PV) that restores arterial O-2 content.Findings from studies investigating the mechanisms underlying this PV contraction have been controversial, possibly as experimental conditions were inadequately controlled.We examined the mechanisms underlying the PV contraction evoked by 4 days of exposure to hypobaric hypoxia (HH) in 11 healthy lowlanders, while strictly controlling water intake, diet, temperature and physical activity.Exposure to HH-induced an similar to 10% PV contraction that was accompanied by a reduction in total circulating protein mass, whereas diuretic fluid loss and total body water remained unchanged.Our data support an oncotically driven fluid redistribution from the intra- to the extravascular space, rather than fluid loss, as the mechanism underlying HH-induced PV contraction.Extended hypoxic exposure reduces plasma volume (PV). The mechanisms underlying this effect are controversial, possibly as previous studies have been confounded by inconsistent experimental conditions. Here, we investigated the effect of hypobaric hypoxia (HH) on PV in a cross-over study that strictly controlled for diet, water intake, physical activity and temperature. Eleven males completed two 4-day sojourns in a hypobaric chamber, one in normoxia (NX) and one in HH equivalent to 3500 m altitude. PV, urine output, volume-regulating hormones and plasma protein concentration were determined daily. Total body water (TBW) was determined at the end of both sojourns by deuterium dilution. Although PV was 8.1 +/- 5.8% lower in HH than in NX after 24 h and remained similar to 10% lower thereafter (all P 0.23). Plasma renin activity and circulating aldosterone were suppressed in HH during the first half of the sojourn (all P 0.05). Markers for atrial natriuretic peptide were higher in HH than NX after 30 min (P = 0.001) but lower during the last 2 days (P

Background The sodium/glucose cotransporter 2 inhibitor empagliflozin has cardiorenal protective properties through mechanisms beyond glucose control. In this study we assessed whether empagliflozin modifies renal oxygenation as a possible mechanism of renal protection, and determined the metabolic, renal, and hemodynamic effects of empagliflozin in nondiabetic subjects. Methods and Results In this double‐blind, randomized, placebo‐controlled study, 45 healthy volunteers underwent blood and urine sampling, renal ultrasound, and blood‐oxygenation‐level–dependent magnetic resonance imaging before and 180 minutes after administration of 10 mg empagliflozin (n=30) or placebo (n=15). These examinations were repeated after 1 month of daily intake. Cortical and medullary renal oxygenation were not affected by the acute or chronic administration of empagliflozin, as determined by 148 renal blood‐oxygenation‐level–dependent magnetic resonance imaging examinations. Empagliflozin increased glucosuria (24‐hour glucosuria at 1 month: +50.1±16.3 g). The acute decrease in proximal sodium reabsorption, as determined by endogenous fractional excretion of lithium (−34.6% versus placebo), was compensated at 1 month by a rise in plasma renin activity (+28.6%) and aldosterone (+55.7%). The 24‐hour systolic and diastolic ambulatory blood pressures decreased significantly after 1 month of empagliflozin administration (−5.1 and −2.0 mm Hg, respectively). Serum uric acid levels decreased (−28.4%), hemoglobin increased (+1.7%), and erythropoietin remained the same. Conclusions Empagliflozin has a rapid and significant effect on tubular function, with sustained glucosuria and transient natriuresis in nondiabetic normotensive subjects. These effects favor blood pressure reduction. No acute or sustained changes were found in renal cortical or medullary tissue oxygenation. It remains to be determined whether this is the case in nondiabetic or diabetic patients with congestive heart failure or kidney disease. REGISTRATION : URL: https://www.clini​caltr​ials.gov ; Unique identifier: NCT03093103 .

Myo-inositol hexakisphosphate (IP6) is a natural product known to inhibit vascular calcification (VC), but with limited potency and low plasma exposure following bolus administration. Here we report the design of a series of inositol phosphate analogs as crystallization inhibitors, among which 4,6-di-O-(methoxy-diethyleneglycol)-myo-inositol-1,2,3,5-tetrakis(phosphate), (OEG2)2-IP4, displays increased in vitro activity, as well as more favorable pharmacokinetic and safety profiles than IP6 after subcutaneous injection. (OEG2)2-IP4 potently stabilizes calciprotein particle (CPP) growth, consistently demonstrates low micromolar activity in different in vitro models of VC (i.e., human serum, primary cell cultures, and tissue explants), and largely abolishes the development of VC in rodent models, while not causing toxicity related to serum calcium chelation. The data suggest a mechanism of action independent of the etiology of VC, whereby (OEG2)2-IP4 disrupts the nucleation and growth of pathological calcification., Cardiovascular calcification is a serious pathology for which effective pharmacological treatments are lacking. Here the authors show that an optimized oligo(ethylene glycol) derivative of inositol phosphate interferes with calcium phosphate crystallization and inhibits soft tissue calcification in vivo following subcutaneous injection.

Sodium and water handling by the kidney and the sympathetic nervous system have been implicated in the development of obesity-related hypertension and kidney disease. They have seldom been studied together during stress conditions. The objective of this study was to compare the systemic, renal and hormonal responses to lower body negative pressure (LBNP) in adult healthy participants (H), obese normotensive (OBN) and obese hypertensive patients (OBH). This was a prospective case-control study. Participants from the three groups were exposed to one hour of LBNP. Systemic and renal haemodynamics, sodium and water excretion and hormones were measured before and after LBNP. Intergroup LBNP responses were tested using a Student t-test or a Wilcoxon rank-sum test. An extension of the Wilcoxon rank-sum test was used to test for a trend across the three groups. The study included 54 participants (H: 25, OBN: 16, OBH: 13). LBNP induced a stepwise increase in systolic blood pressure (+2.7 ± 4.7 mmHg (H) vs. +4.7 ± 8.8 mmHg (OBN) vs. +8.0 ± 8.6 mmHg (OBH, p = .028)) and heart rate (−1.3 ± 4.9 bpm (H) vs. 2.2 ± 6.1 bpm (OBN) vs. 1.9 ± 4.1 bpm (OBH, p = .041). Urinary output (−2.8 ± 2.1 ml/min vs. −1.4 ± 1.7 ml/min, p = .028) and free water clearance (−1.9 ± 1.7 mOsm/kg vs. −0.7 ± 1.3 mOsm/kg, p = .016) responses were more marked in OBN compared to H. These results show that the systemic and the renal response to LBNP differ according to weight and to BP categories. Systolic BP and heart show a progressive increased response form healthy volunteers to OBN and then to obese hypertensive participants while urinary output and free water clearance responses are increased in OBN only, suggesting that the occurrence of hypertension in obese individuals modifies the early kidney responses to stress. NCT01734096

The mechanism of sodium retention and its location in kidney tubules may vary with time in nephrotic syndrome (NS). We studied the mechanisms of sodium retention in transgenic POD-ATTAC mice, which display an inducible podocyte-specific apoptosis. At day 2 after the induction of NS, the increased abundance of NHE3 and phosphorylated NCC in nephrotic mice compared with controls suggest that early sodium retention occurs mainly in the proximal and distal tubules. At day 3, the abundance of NHE3 normalized, phosphorylated NCC levels decreased, and cleavage and apical localization of γ-ENaC increased in nephrotic mice. These findings indicate that sodium retention shifted from the proximal and distal tubules to the collecting system. Increased cleavage and apical localization of γ-ENaC persisted at day 5 in nephrotic mice when hypovolemia resolved and steady-state was reached. Sodium retention and γ-ENaC cleavage were independent of the increased plasma levels of aldosterone. Nephrotic mice displayed decreased glomerular filtration rate and urinary potassium excretion associated with hyperkaliemia at day 3. Feeding nephrotic mice with a low potassium diet prevented hyperkaliemia, γ-ENaC cleavage, and led to persistent increased phosphorylation of NCC. These results suggest that potassium homeostasis is a major determinant of the tubular site of sodium retention in nephrotic mice.

The amiloride-sensitive epithelial sodium channel (ENaC) and the thiazide-sensitive sodium chloride cotransporter (NCC) are key regulators of sodium and potassium and colocalize in the late distal convoluted tubule of the kidney. Loss of the αENaC subunit leads to a perinatal lethal phenotype characterized by sodium loss and hyperkalemia resembling the human syndrome pseudohypoaldosteronism type 1 (PHA-I). In adulthood, inducible nephron-specific deletion of αENaC in mice mimics the lethal phenotype observed in neonates, and as in humans, this phenotype is prevented by a high sodium (HNa+)/low potassium (LK+) rescue diet. Rescue reflects activation of NCC, which is suppressed at baseline by elevated plasma potassium concentration. In this study, we investigated the role of the γENaC subunit in the PHA-I phenotype. Nephron-specific γENaC knockout mice also presented with salt-wasting syndrome and severe hyperkalemia. Unlike mice lacking αENaC or βEΝaC, an HNa+/LK+ diet did not normalize plasma potassium (K+) concentration or increase NCC activation. However, when K+ was eliminated from the diet at the time that γENaC was deleted, plasma K+ concentration and NCC activity remained normal, and progressive weight loss was prevented. Loss of the late distal convoluted tubule, as well as overall reduced βENaC subunit expression, may be responsible for the more severe hyperkalemia. We conclude that plasma K+ concentration becomes the determining and limiting factor in regulating NCC activity, regardless of Na+ balance in γENaC-deficient mice.

Na+ excretion by the kidney varies according to dietary Na+ intake. We undertook a systematic study on the effects of dietary salt intake on glomerular filtration rate (GFR) and tubular Na+ reabsorption. We examined the renal adaptive response in mice subjected to 7 days of a low sodium diet (LSD) containing 0.01% Na+, a normal sodium diet (NSD) containing 0.18% Na+, as well as a moderately high sodium diet (HSD) containing 1.25% Na+. As expected, LSD did not alter measured GFR and increased the abundance of total and cell-surface NHE3, NKCC2, NCC, α-ENaC, and cleaved γ-ENaC compared to NSD. Mathematical modelling predicted that tubular Na+ reabsorption increased in the proximal tubule but decreased in the distal nephron because of diminished Na+ delivery. This prediction was confirmed by the natriuretic response to diuretics targeting the thick ascending limb, the distal convoluted tubule or the collecting system. On the other hand, HSD did not alter measured GFR but decreased the abundance of the aforementioned transporters compared to NSD. Mathematical modelling predicted that tubular Na+ reabsorption decreased in the proximal tubule but increased in distal segments with lower transport efficiency with respect to O2 consumption. This prediction was confirmed by the natriuretic response to diuretics. The activity of the metabolic sensor AMPK was related to the changes in tubular Na+ reabsorption. Our data show that fractional Na+ reabsorption is distributed differently according to dietary Na+ intake and induces changes in tubular O2 consumption and sodium transport efficiency. This article is protected by copyright. All rights reserved

Aims Clinical studies suggest beneficial effects of renin–angiotensin system blockade for prevention of left ventricular (LV) dysfunction after chemotherapy. However, the efficacy of this strategy as primary prevention has been poorly studied. This study aimed at identifying the pathophysiological mechanisms by which mineralocorticoid receptor antagonism (MRA) or angiotensin converting enzyme inhibition (ACEi) provide protection against doxorubicin-induced cardiotoxicity (DIC) in mouse models of acute and chronic toxicity. Methods and results Acute DIC was induced by a single injection of Dox at 15 mg/kg and chronic DIC applied 5 injections of Dox at 4 mg/kg/week. MRA was achieved using eplerenone or cardiomyocyte-specific ablation of the MR gene in transgenic mice and ACEi using enalapril. Drugs were provided with the first dose of Dox and applied until the end of the study. In both model of DIC, Dox induced cardiac atrophy with decreased LV volume, reduced cardiomyocyte cell size, and cardiac dysfunction. In the acute model, neither MRA nor ACEi protected against these manifestations of DIC. In the chronic model, concomitant treatment with eplerenone did not protect against DIC and drastically increased plasma aldosterone levels and cardiac levels of angiotensin II type 1 receptor and of connective tissue growth factor (CTGF), as observed in acute DIC. Enalapril treatment in the chronic model, however, protected against cardiac dysfunction and cardiomyocyte atrophy and was associated with increased activation of the PI3K/AKT/mTOR pathway along with normal levels of CTGF. Conclusion Enalapril and eplerenone disparately impact on cellular signalling in DIC. Eplerenone, on top of Dox treatment was not protective and associated with increased levels of plasma aldosterone and of cardiac CTGF. In contrast, we show that primary prevention with enalapril preserves LV morphology and function in a clinically relevant model of chronic DIC, with increased stimulation of the PI3K/AKT/mTOR axis and normal CTGF levels suggesting potential therapeutic implications.

The mineralocorticoid hormone aldosterone plays a crucial role in the control of Na + and K + balance, blood volume, and arterial blood pressure, by acting in the aldosterone-sensitive distal nephron (ASDN) and stimulating a complex transcriptional, translational, and cellular program. Because the complexity of the aldosterone response is still not fully appreciated, we aimed at identifying new elements in this pathway. Here, we demonstrate that the expression of the proto-oncogene PIM3 (Proviral Integration Site of Moloney Murine Leukemia Virus 3), a serine/threonine kinase belonging to the calcium/calmodulin-regulated group of kinases, is stimulated by aldosterone in vitro (mCCD cl1 cells), ex vivo (mouse kidney slices), and in vivo in mice. Characterizing a germline Pim3 - / - mouse model, we found that these mice have an upregulated Renin-Angiotensin-Aldosterone System (RAAS), with high circulating aldosterone and plasma renin activity levels on both standard or Na + -deficient diet. Surprisingly, we did not observe any obvious salt-losing phenotype in Pim3 KO mice as shown by normal blood pressure, plasma and urinary electrolytes, as well as unchanged expression levels of the major Na + transport proteins. These observations suggest that the potential effects of the loss of the Pim3 gene are physiologically compensated. Indeed, the 2 other family members of the PIM kinase family, PIM1 and PIM2 are upregulated in the kidney of Pim3 - / - mice, and may therefore be involved in such compensation. In conclusion, our data demonstrate that the PIM3 kinase is a novel aldosterone-induced protein, but its precise role in aldosterone-dependent renal homeostasis remains to be determined.

International audience; Chronic glucocorticoid infusion impairs NCC activity and induces a non-dipping profile in mice, suggesting that glucocorticoids are essential for daily blood pressure variations. In this paper, we studied mice lacking the renal tubular glucocorticoid receptor (GR) in adulthood (GR knockouts, Nr3c1Pax8/LC1). Upon standard salt diet, Nr3c1Pax8/LC1 mice grow normally, but show reduced NCC activity despite normal plasma aldosterone levels. Following diet switch to low sodium, Nr3c1Pax8/LC1 mice exhibit a transient but significant reduction in the activity of NCC and expression of NHE3 and NKCC2 accompanied by significant increased Spak activity. This is followed by transiently increased urinary sodium excretion and higher plasma aldosterone concentrations. Plasma corticosterone levels and 11βHSD2 mRNA expression and activity in the whole kidney remain unchanged. High salt diet does not affect whole body Na+ and/or K+ balance and NCC activity is not reduced, but leads to a significant increase in diastolic blood pressure dipping in Nr3c1Pax8/LC1 mice. When high sodium treatment is followed by 48 h of darkness, NCC abundance is reduced in knockout mice although activity is not different. Our data show that upon Na+ restriction renal tubular GR-deficiency transiently affects Na+ handling and transport pathways. Overall, upon standard, low Na+ and high Na+ diet exposure Na+ and K+ balance is maintained as evidenced by normal plasma and urinary Na+ and K+ and aldosterone concentrations.

The circadian clock controls a wide variety of metabolic and homeostatic processes in a number of tissues, including the kidney. However, the role of the renal circadian clocks remains largely unknown. To address this question, we performed a combined functional, transcriptomic, and metabolomic analysis in mice with inducible conditional knockout (cKO) of BMAL1, which is critically involved in the circadian clock system, in renal tubular cells (Bmal1(lox/lox)/Pax8-rtTA/LC1 mice). Induction of cKO in adult mice did not produce obvious abnormalities in renal sodium, potassium, or water handling. Deep sequencing of the renal transcriptome revealed significant changes in the expression of genes related to metabolic pathways and organic anion transport in cKO mice compared with control littermates. Furthermore, kidneys from cKO mice exhibited a significant decrease in the NAD(+)-to-NADH ratio, which reflects the oxidative phosphorylation-to-glycolysis ratio and/or the status of mitochondrial function. Metabolome profiling showed significant changes in plasma levels of amino acids, biogenic amines, acylcarnitines, and lipids. In-depth analysis of two selected pathways revealed a significant increase in plasma urea level correlating with increased renal Arginase II activity, hyperargininemia, and increased kidney arginine content as well as a significant increase in plasma creatinine concentration and a reduced capacity of the kidney to secrete anionic drugs (furosemide) paralleled by an approximate 80% decrease in the expression level of organic anion transporter 3 (SLC22a8). Collectively, these results indicate that the renal circadian clocks control a variety of metabolic/homeostatic processes at the intrarenal and systemic levels and are involved in drug disposition.

Background/Aims In patients with resistant hypertension, renal denervation (RDN) studies have mainly focused their outcomes on blood pressure (BP). The aim of this study was to evaluate the long-term effect of RDN on neurohormonal profiles, renal hemodynamics and sodium excretion in a resting state and during stress induced by lower body negative pressure (LBNP). Materials and methods This was a single center prospective observational study. Norepinephrine, plasma renin activity (PRA), glomerular filtration rate (GFR), renal plasma flow (RPF) and sodium excretion were measured in unstimulated conditions (rest) and after one hour of LBNP at three different time points: before (M0), one (M1) and twelve months (M12) after RDN. Results Thirteen patients with resistant hypertension were included. In the resting state, no differences were observed in norepinephrine, PRA, sodium excretion and mean BP levels after RDN. GFR (78 ± 32 ml/min at M0 vs 66 ± 26 ml/min at M12 (p = 0.012) and filtration fraction (22.6 ±5.4% at M0 vs 15.1 ±5.3% at M12 (p = 0.002)) both decreased after RDN. During LBNP, the magnitude of the mean BP increase was reduced from +6.8 ± 6.6 mm Hg at M0 to +2.3 ± 1.3 mm Hg at M12 (p = 0.005). The LBNP-induced increase in norepinephrine and decrease in GFR and sodium excretion observed before RDN were blunted after the procedure. Conclusion A decrease in GFR and filtration fraction was observed one year after RDN. In addition, our results suggest that RDN blunts not only the norepinephrine but also the mean BP, the GFR and the sodium excretion responses to an orthostatic stress one year after the intervention. Registry number NCT01734096

Cirrhosis is a frequent and severe disease, complicated by renal sodium retention leading to ascites and oedema. A better understanding of the complex mechanisms responsible for renal sodium handling could improve clinical management of sodium retention. Our aim was to determine the importance of the amiloride-sensitive epithelial sodium channel (ENaC) in collecting ducts in compensate and decompensate cirrhosis. Bile duct ligation was performed in control mice (CTL) and collecting duct-specific αENaC knockout (KO) mice, and ascites development, aldosterone plasma concentration, urinary sodium/potassium ratio and sodium transporter expression were compared. Disruption of ENaC in collecting ducts (CDs) did not alter ascites development, urinary sodium/potassium ratio, plasma aldosterone concentrations or Na,K-ATPase abundance in CCDs. Total αENaC abundance in whole kidney increased in cirrhotic mice of both genotypes and cleaved forms of α and γ ENaC increased only in ascitic mice of both genotypes. The sodium chloride cotransporter (NCC) abundance was lower in non-ascitic KO, compared to non-ascitic CTL, and increased when ascites appeared. In ascitic mice, the lack of αENaC in CDs induced an upregulation of total ENaC and NCC and correlated with the cleavage of ENaC subunits. This revealed compensatory mechanisms which could also take place when treating the patients with diuretics. These compensatory mechanisms should be considered for future development of therapeutic strategies.

Background/Aims: The purpose of the present study was to compare the direct renin inhibitor aliskiren to the diuretic hydrochlorothiazide (HCTZ) in their ability to modulate renal tissue oxygenation in hypertensive patients. Methods: 24 patients were enrolled in this randomized prospective study and 20 completed the protocol. Patients were randomly assigned to receive either aliskiren 150-300 mg/d or HCTZ 12.5 - 25 mg/d for 8 weeks. Renal oxygenation was measured by BOLD-MRI at weeks 0 and 8. BOLD-MRI was also performed before and after an i.v. injection of 20 mg furosemide at week 0 and at week 8. BOLD-MRI data were analyzed by measuring the oxygenation in 12 computed layers of the kidney enabling to asses renal oxygenation according to the depth within the kidney and by the classical method of regions of interest (ROI). Results: The classical ROI analysis of the data showed no difference between the groups at week 8. The analysis of renal oxygenation according to the 12 layers method shows no significant difference between aliskiren and HCTZ at week 8 before administration of furosemide. However, within group analyses show that aliskiren slightly but not significantly increased oxygenation in the cortex and decreased medullary oxygenation whereas HCTZ induced a significant overall decrease in renal tissue oxygenation. With the same method of analysis we observed that the response to furosemide was unchanged in the HCTZ group at week 8 but was characterized by an increase in both cortical and medullary oxygenation in aliskiren-treated patients. Patients responding to aliskiren and HCTZ by a fall in systolic blood pressure of >10 mmHg improved their renal tissue oxygenation when compared to non-responders. Conclusion: With the classical method of evaluation using regions no difference were found between aliskiren and HCTZ on renal tissue oxygenation after 8 weeks. In contrast, with our new method that takes into account the entire kidney, within group analyses show that aliskiren slightly increases cortical and medullary renal tissue oxygenation in hypertensive patients whereas HCTZ decreases significantly renal oxygenation at trough.

Elevated serum phosphorus, calcium, and fibroblast growth factor 23 (FGF23) levels are associated with cardiovascular disease in chronic renal disease. This study evaluated the effects of sucroferric oxyhydroxide (PA21), a new iron-based phosphate binder, versus lanthanum carbonate (La) and sevelamer carbonate (Se), on serum FGF23, phosphorus, calcium, and intact parathyroid hormone (iPTH) concentrations, and the development of vascular calcification in adenine-induced chronic renal failure (CRF) rats. After induction of CRF, renal function was significantly impaired in all groups: uremic rats developed severe hyperphosphatemia, and serum iPTH increased significantly. All uremic rats (except controls) then received phosphate binders for 4 weeks. Hyperphosphatemia and increased serum iPTH were controlled to a similar extent in all phosphate binder-treatment groups. Only sucroferric oxyhydroxide was associated with significantly decreased FGF23. Vascular calcifications of the thoracic aorta were decreased by all three phosphate binders. Calcifications were better prevented at the superior part of the thoracic and abdominal aorta in the PA21 treated rats. In adenine-induced CRF rats, sucroferric oxyhydroxide was as effective as La and Se in controlling hyperphosphatemia, secondary hyperparathyroidism, and vascular calcifications. The role of FGF23 in calcification remains to be confirmed.

BACKGROUND. Glomerular hyperfiltration (GHF) is a well-recognized early renal alteration in diabetic patients. As the prevalence of GHF is largely unknown in populations in the African region with respect to normal fasting glucose (NFG), impaired fasting glucose (IFG) and type 2 diabetes [diabetes mellitus (DM)], we conducted a cross-sectional study in the Seychelles islands among families including at least one member with hypertension. METHODS. The glomerular filtration rate (GFR), effective renal plasma flow (ERPF) and proximal tubular sodium reabsorption were measured using inulin, p-aminohippurate (PAH) and endogenous lithium clearance, respectively. Twenty-four-hour urine was collected on the preceding day. RESULTS. Of the 363 participants (mean age 44.7 years), 6.6% had IFG, 9.9% had DM and 63.3% had hypertension. The prevalence of GHF, defined as a GFR >140 ml/min, was 17.2%, 29.2% and 52.8% in NFG, IFG and DM, respectively (P trend

BACKGROUND: Adipokines are a group of fat-secreted hormones and cytokines, including leptin and adiponectin, with important functions in humans. Peritoneal dialysis (PD) is associated with markedly raised plasma adipokines, suggesting increased production in this setting. We have shown that low pH down-regulates leptin production. The current study was designed to test if novel pH-neutral PD fluids may regulate leptin and adiponectin secretion in vitro. METHODS: We exposed 3T3-L1 adipocytes to a 50 : 50 mixture of dialysate and M199 containing 10% serum for upto 48 h. Dialysates were commercial PD fluids, i.e. conventional acidic, lactate-buffered solutions (PD-acid) and pH-neutral lactate-buffered (PD-Bal) or bicarbonate-buffered solutions (PD-Bic). Leptin and adiponectin concentrations in culture-cell media were measured by ELISA. RESULTS: Compared with PD-acid, PD-Bal and PD-Bic produced a 25 and 43% increase, respectively, in leptin secretion at 48 h (P < 0.05). In contrast, adiponectin secretion was not affected. High glucose PD fluids (4.25%) specifically inhibited leptin secretion vs 1.5% glucose, buffer-matched solutions (P < 0.05). However, differences in leptin secretion due to pH and type of buffer remained significant. In further experiments, the pH of test media were extensively varied without the presence of dialysates. Leptin secretion was shown to increase in a parallel to pH, whereas large changes in pH did not affect adiponectin secretion. CONCLUSION: The pH-neutral PD solutions specifically induce leptin, but not adiponectin secretion from 3T3-L1 adipocytes. PD-Bic produced a greater leptin stimulation than PD-Bal, but this difference was attributable to pH per se, rather than the type of buffer.

BACKGROUND In this study, we assessed whether the white-coat effect (difference between office and daytime blood pressure (BP)) is associated with nondipping (absence of BP decrease at night). METHODS Data were available in 371 individuals of African descent from 74 families selected from a population-based hypertension register in the Seychelles Islands and in 295 Caucasian individuals randomly selected from a population-based study in Switzerland. We used standard multiple linear regression in the Swiss data and generalized estimating equations to account for familial correlations in the Seychelles data. RESULTS The prevalence of systolic and diastolic nondipping (

In adulthood, an induced nephron-specific deficiency of αENaC (Scnn1a) resulted in pseudohypoaldosteronism type 1 (PHA-1) with sodium loss, hyperkalemia, and metabolic acidosis that is rescued through high-sodium/low-potassium (HNa + /LK + ) diet. In the present study, we addressed whether renal βENaC expression is required for sodium and potassium balance or can be compensated by remaining (α and γ) ENaC subunits using adult nephron-specific knockout (Scnn1b Pax8/LC1 ) mice. Upon induction, these mice present a severe PHA-1 phenotype with weight loss, hyperkalemia, and dehydration, but unlike the Scnn1a Pax8/LC1 mice without persistent salt wasting. This is followed by a marked downregulation of STE20/SPS1-related proline-alanine-rich protein kinase (SPAK) and Na + /Cl - co-transporter (NCC) protein expression and activity. Most of the experimental Scnn1b Pax8/LC1 mice survived with a HNa + /LK + diet that partly normalized NCC phosphorylation, but not total NCC expression. Since salt loss was minor, we applied a standard-sodium/LK + diet that efficiently rescued these mice resulting in normokalemia and normalization of NCC phosphorylation, but not total NCC expression. A further switch to LNa + /standard-K + diet induced again a severe PHA-1-like phenotype, but with only transient salt wasting indicating that low-K + intake is critical to decrease hyperkalemia in a NCC-dependent manner. In conclusion, while the βENaC subunit plays only a minor role in sodium balance, severe hyperkalemia results in downregulation of NCC expression and activity. Our data demonstrate the importance to primarily correct the hyperkalemia with a low-potassium diet that normalizes NCC activity.

Introduction Angiotensin II AT 1-receptor antagonists are highly bound to plasma proteins (≥ 99%). With some antagonists, such as DuP-532, the protein binding was such that no efficacy of the drug could be demonstrated clinically. Whether protein binding interferes with the efficacy of other antagonists is not known. We have therefore investigated in vitro how plasma proteins may affect the antagonistic effect of different AT1-receptor antagonists. Methods A radio-receptor binding assay was used to analyse the interaction between proteins and the ability of various angiotensin II (Ang II) antagonists to block AT1-receptors. In addition, the Biacore technology, a new technique which enables the real-time monitoring of binding events between two molecules, was used to evaluate the dissociation rate constants of five AT1-receptor antagonists from human serum albumin. Results The in vitro AT 1-antagonistic effects of different Ang II receptor antagonists were differentially affected by the presence of human plasma, with rightward shifts of the IC50 ranging from one to several orders of magnitude. The importance of the shift correlates with the dissociation rate constants of these drugs from albumin. Our experiments also show that the way that AT1-receptor antagonists bind to proteins differs from one compound to another. These results suggest that the interaction with plasma proteins appears to modulate the efficacy of some Ang II antagonists. Conclusion Although the high binding level of Ang II receptor antagonist to plasma proteins appears to be a feature common to this class of compounds, the kinetics and characteristics of this binding is of great importance. With some antagonists, protein binding interferes markedly with their efficacy to block AT1-receptors.

Chronic renal failure (CRF) is associated with the development of secondary hyperparathyroidism and vascular calcifications. We evaluated the efficacy of PA21, a new iron-based noncalcium phosphate binder, in controlling phosphocalcic disorders and preventing vascular calcifications in uremic rats. Rats with adenine-diet-induced CRF were randomized to receive either PA21 0.5, 1.5, or 5% or CaCO3 3% in the diet for 4 weeks, and were compared with uremic and nonuremic control groups. After 4 weeks of phosphate binder treatment, serum calcium, creatinine, and body weight were similar between all CRF groups. Serum phosphorus was reduced with CaCO3 3% (2.06 mM; P ≤ 0.001), PA21 1.5% (2.29 mM; P < 0.05), and PA21 5% (2.21 mM; P ≤ 0.001) versus CRF controls (2.91 mM). Intact parathyroid hormone was strongly reduced in the PA21 5% and CaCO3 3% CRF groups to a similar extent (1138 and 1299 pg/ml, respectively) versus CRF controls (3261 pg/ml; both P ≤ 0.001). A lower serum fibroblast growth factor 23 concentration was observed in the PA21 5%, compared with CaCO3 3% and CRF, control groups. PA21 5% CRF rats had a lower vascular calcification score compared with CaCO3 3% CRF rats and CRF controls. In conclusion, PA21 was as effective as CaCO3 at controlling phosphocalcic disorders but superior in preventing the development of vascular calcifications in uremic rats. Thus, PA21 represents a possible alternative to calcium-based phosphate binders in CRF patients.

Aim To assess whether blockade of the renin–angiotensin system (RAS), a recognized strategy to prevent the progression of diabetic nephropathy, affects renal tissue oxygenation in type 2 diabetes mellitus (T2DM) patients. Methods Prospective randomized 2-way cross over study; T2DM patients with (micro)albuminuria and/or hypertension underwent blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI) at baseline, after one month of enalapril (20mgqd), and after one month of candesartan (16mgqd). Each BOLD-MRI was performed before and after the administration of furosemide. The mean R 2 * (=1/ T 2 *) values in the medulla and cortex were calculated, a low R 2 * indicating high tissue oxygenation. Results Twelve patients (mean age: 60±11 years, eGFR: 62±22ml/min/1.73m 2 ) completed the study. Neither chronic enalapril nor candesartan intake modified renal cortical or medullary R 2 * levels. Furosemide significantly decreased cortical and medullary R 2 * levels suggesting a transient increase in renal oxygenation. Medullary R 2 * levels correlated positively with urinary sodium excretion and systemic blood pressure, suggesting lower renal oxygenation at higher dietary sodium intake and blood pressure; cortical R 2 * levels correlated positively with glycemia and HbA1c. Conclusion RAS blockade does not seem to increase renal tissue oxygenation in T2DM hypertensive patients. The response to furosemide and the association with 24h urinary sodium excretion emphasize the crucial role of renal sodium handling as one of the main determinants of renal tissue oxygenation.

Ubiquitylation plays an important role in the control of Na+ homeostasis by the kidney. It is well established that the epithelial Na+ channel ENaC is regulated by the ubiquitin-protein ligase NEDD4-2, limiting ENaC cell surface expression and activity. Ubiquitylation can be reversed by the action of deubiquitylating enzymes (DUBs). One such DUB, USP2-45, was identified previously as an aldosterone-induced protein in the kidney and is also a circadian output gene. In heterologous expression systems, USP2-45 binds to ENaC, deubiquitylates it, and enhances channel density and activity at the cell surface. Because the role of USP2-45 in renal Na+ transport had not been studied in vivo, we investigated here the effect of Usp2 gene inactivation in this process. We demonstrate first that USP2-45 protein has a rhythmic expression with a peak at ZT12. Usp2-KO mice did not show any differences from wild-type littermates with respect to the diurnal control of Na+ or K+ urinary excretion and plasma levels either on a standard diet or after acute and chronic changes to low- and high-Na+ diets, respectively. Moreover, they had similar aldosterone levels on either a low- or high-Na+ diet. Blood pressure measurements using telemetry did not reveal variations compared with control mice. Usp2-KO mice did not display alterations in expression of genes involved in sodium homeostasis or the ubiquitin system, as evidenced by transcriptome analysis in the kidney. Our data suggest that USP2 does not play a primary role in the control of Na+ balance or blood pressure.

BACKGROUND Cocoa is rich in flavonoids, has anti-oxidative properties and increases the bioavailability of nitric oxide (NO). Adequate renal tissue oxygenation is crucial for the maintenance of renal function. The goal of this study was to investigate the effect of cocoa-rich dark chocolate (DC) on renal tissue oxygenation in humans, as compared to flavonoid-poor white chocolate (WC). METHODS Ten healthy volunteers with preserved kidney function (mean age ± SD 35 ± 12 years, 70% women, BMI 21 ± 3 kg/m2) underwent blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI) before and 2 hours after the ingestion of 1 g/kg of DC (70% cocoa). Renal tissue oxygenation was determined by the measurement of R2* maps on 4 coronal slices covering both kidneys. The mean R2* (= 1/T2*) values in the medulla and cortex were calculated, a low R2* indicating high tissue oxygenation. Eight participants also underwent BOLD-MRI at least 1 week later, before and 2 hours after the intake of 1 g/kg WC. RESULTS The mean medullary R2* was lower after DC intake compared to baseline (28.2 ± 1.3 s-1 vs. 29.6 ± 1.3 s-1, p = 0.04), whereas cortical and medullary R2* values did not change after WC intake. The change in medullary R2* correlated with the level of circulating (epi)catechines, metabolites of flavonoids (r = 0.74, p = 0.037), and was independent of plasma renin activity. CONCLUSION This study suggests for the first time an increase of renal medullary oxygenation after intake of dark chocolate. Whether this is linked to flavonoid-induced changes in renal perfusion or oxygen consumption, and whether cocoa has potentially renoprotective properties, merits further study.

The stimulation of postprandial K+ clearance involves aldosterone-independent and -dependent mechanisms. In this context, serum- and glucocorticoid-induced kinase (SGK)1, a ubiquitously expressed kinase, is one of the primary aldosterone-induced proteins in the aldosterone-sensitive distal nephron. Germline inactivation of SGK1 suggests that this kinase is fundamental for K+ excretion under conditions of K+ load, but the specific role of renal SGK1 remains elusive. To avoid compensatory mechanisms that may occur during nephrogenesis, we used inducible, nephron-specific Sgk1 Pax8/ LC1 mice to assess the role of renal tubular SGK1 in K+ regulation. Under a standard diet, these animals exhibited normal K+ handling. When challenged by a high-K+ diet, they developed severe hyperkalemia accompanied by a defect in K+ excretion. Molecular analysis revealed reduced neural precursor cell expressed developmentally downregulated protein (NEDD)4-2 phosphorylation and total expression. γ-Epithelial Na+ channel (ENaC) expression and α/γENaC proteolytic processing were also decreased in mutant mice. Moreover, with no lysine kinase (WNK)1, which displayed in control mice punctuate staining in the distal convoluted tubule and diffuse distribution in the connecting tubule/cortical colleting duct, was diffused in the distal convoluted tubule and less expressed in the connecting tubule/collecting duct of Sgk Pax8/ LC1 mice. Moreover, Ste20-related proline/alanine-rich kinase phosphorylation, and Na+-Cl− cotransporter phosphorylation/apical localization were reduced in mutant mice. Consistent with the altered WNK1 expression, increased renal outer medullary K+ channel apical localization was observed. In conclusion, our data suggest that renal tubular SGK1 is important in the regulation of K+ excretion via the control of NEDD4-2, WNK1, and ENaC.

The circadian clock contributes to the control of BP, but the underlying mechanisms remain unclear. We analyzed circadian rhythms in kidneys of wild-type mice and mice lacking the circadian transcriptional activator clock gene. Mice deficient in clock exhibited dramatic changes in the circadian rhythm of renal sodium excretion. In parallel, these mice lost the normal circadian rhythm of plasma aldosterone levels. Analysis of renal circadian transcriptomes demonstrated changes in multiple mechanisms involved in maintaining sodium balance. Pathway analysis revealed the strongest effect on the enzymatic system involved in the formation of 20-HETE, a powerful regulator of renal sodium excretion, renal vascular tone, and BP. This correlated with a significant decrease in the renal and urinary content of 20-HETE in clock-deficient mice. In summary, this study demonstrates that the circadian clock modulates renal function and identifies the 20-HETE synthesis pathway as one of its principal renal targets. It also suggests that the circadian clock affects BP, at least in part, by exerting dynamic control over renal sodium handling.

The glucocorticoid-induced leucine zipper (Tsc22d3-2) is a widely expressed dexamethasone-induced transcript that has been proposed to be important in immunity, adipogenesis, and renal sodium handling based on in vitro studies. To address its function in vivo, we have used Cre/loxP technology to generate mice deficient for Tsc22d3-2. Male knockout mice were viable but surprisingly did not show any major deficiencies in immunological processes or inflammatory responses. Tsc22d3-2 knockout mice adapted to a sodium-deprived diet and to water deprivation conditions but developed a subtle deficiency in renal sodium and water handling. Moreover, the affected animals developed a mild metabolic phenotype evident by a reduction in weight from 6 months of age, mild hyperinsulinemia, and resistance to a high-fat diet. Tsc22d3-2-deficient males were infertile and exhibited severe testis dysplasia from postnatal d 10 onward with increases in apoptotic cells within seminiferous tubules, an increased number of Leydig cells, and significantly elevated FSH and testosterone levels. Thus, our analysis of the Tsc22d3-2-deficient mice demonstrated a previously uncharacterized function of glucocorticoid-induced leucine zipper protein in testis development.

Background Acute kidney injury (AKI) is a common and important complication of critical illness associated with increased morbidity, mortality and costs. Alterations in renal perfusion are thought to play a central role in its pathogenesis. This causal relationship remains, however, largely speculative as data on renal perfusion in AKI and critical illness are scarce. Indeed, methods enabling renal perfusion quantification are either invasive, expensive or inapplicable to critically ill patients. Contrast-enhanced ultrasonography (CEUS) is a recent imaging modality which provides a unique means of visualizing tissue perfusion. Studies have suggested that CEUS could enable blood flow quantification. CEUS would be an ideal tool in the intensive care unit (ICU) where knowledge of renal perfusion could help prevent or manage AKI. However, CEUS was never used nor validated in the ICU and its clinical use remains to be established. Hypothesis and specific aims We hypothesized that evaluation of renal microcirculation with CEUS was feasible and could change medical management in the ICU. To test this hypothesis, we have designed four studies with the following specific aims: 1. Validate preliminary results and improve technical measurements in a sheep model. 2. Demonstrate CEUS’s feasibility and safety in the ICU. Obtain preliminary results of renal microcirculation modifications associated with elective cardiac surgery. 3. Evaluate the impact of CEUS measurements in two clinical situations: a. Circulatory failure requiring noradrenaline infusion b. Type-1 hepato-renal syndrome requiring terlipressin treatment Overall methods and main results 1. In a sheep model in which renal blood flow could be modified pharmacologically and mechanically, we have compared CEUS-derived parameters to measurements of a an implanted flow probe. We found that CEUS derived parameters were highly heterogeneous and that they did not parallel changes in renal blood flow. Technical measurements were improved and simpler protocols designed for further studies. 2. We have performed CEUS studies before and after an elective cardiac surgery in twelve patients. This study was the first attempt to quantify renal microcirculation in critically ill patients and has demonstrated CEUS’s safety and feasibility in ICU. 3. We have compared changes in CEUS-derived parameters under two levels of blood pressure (BP) in twelve patients with circulatory shock. We have established that microcirculatory response to an increase in BP as assessed by CEUS was highly heterogeneous and unpredictable. This finding is consistent with data from large trials and suggest that a CEUS-guided strategy to determine ideal target BP could be tested. 4. We have compared changes in CEUS-derived parameters before and after the administration of terlipressin in four patients with type-1 hepato-renal syndrome. We found dramatic changes in CEUS-derived parameters after terlipressin therapy and perfusion responses appeared variable. Larger studies are necessary to establish the sensitivity and specificity of CEUS to predict terlipressin responsiveness. Conclusions CEUS is safe and feasible in the intensive care unit. CEUS-derived perfusion indices might not fully correlate with renal blood flow as they are indicative of renal microcirculation changes. Further studies are required to establish the significance of CEUS-derived indices changes in clinical practice.

Background Obesity and African American ethnicity are established independent risk factors for the development of chronic kidney disease. No data exist about the association between obesity and renal hemodynamics in the African region. Study Design Cross-sectional study. Setting & Participants 301 nondiabetic participants (97 lean, 108 overweight, and 96 obese) of African descent with a positive family history of hypertension from the Seychelles islands. Predictor Body mass index (BMI). Outcomes Glomerular hyperfiltration, glomerular filtration rate (GFR), effective renal plasma flow (ERPF), and filtration fraction. Measurements GFR and ERPF were measured using inulin and para-aminohippurate clearances, respectively. Participants' baseline demographics, laboratory data, and blood pressure were measured using standard techniques. Results The prevalence of glomerular hyperfiltration (defined as GFR ≥140 mL/min) increased across BMI categories (7.2%, 14.8%, and 27.1% for lean, overweight, and obese participants, respectively; P 25 kg/m2) and increased GFR, ERPF, and filtration fraction. No association between BMI categories and GFR was found with adjustment for body surface area. Limitations Participants had a positive family history of hypertension. Conclusion Overweight and obesity are associated with increased GFR, ERPF, and filtration fraction and a high prevalence of glomerular hyperfiltration in nondiabetic individuals of African descent. The absence of associations between BMI categories and GFR indexed for body surface area raises questions regarding the appropriateness of indexing GFR for body surface area in overweight populations.

The cardiovascular system is under the control of the circadian clock, and disturbed circadian rhythms can induce cardiovascular pathologies. This cyclic regulation is probably brought about by the circadian expression of genes encoding enzymes and regulators involved in cardiovascular functions. We have previously shown that the rhythmic transcription of output genes is, in part, regulated by the clock-controlled PAR bZip transcription factors DBP (albumin D-site binding protein), HLF (hepatic leukemia factor), and TEF (thyrotroph embryonic factor). The simultaneous deletion of all three PAR bZip transcription factors leads to increased morbidity and shortened life span. In the present study, we demonstrate that Dbp/ Tef/ Hlf triple knockout mice develop cardiac hypertrophy and left ventricular dysfunction associated with a low blood pressure. These dysfunctions are exacerbated by an abnormal response to this low blood pressure characterized by low aldosterone levels. The phenotype of PAR bZip knockout mice highlights the importance of circadian regulators in the modulation of cardiovascular functions.

Endothelin (ET) is an important modulator of renal function through its binding to ETA and ETB receptors in renal tissue. Various renal cells have the ability to synthesize and release endothelin-1 and elevated plasma and urinary endothelin levels have been measured in patients with chronic kidney diseases. Within the last 5 year, several studies have demonstrated that ET plays a role in the pathogenesis and progression of chronic kidney diseases and associated cardiovascular diseases. With this increasing evidence, several ET receptor antagonists have been developed, some of them being specifically investigated for their ability to provide renal protection in diabetic nephropathy. For this indication, a selective blockade of ETA receptors appears to be the preferred approach. Thus, recent clinical phase II and phase III studies have shown that ETA receptor blockers such as avosentan are able to lower proteinuria significantly in type 2 diabetic patients even on top of a full treatment with angiotensin converting enzyme inhibitors or angiotensin II receptor blockers. However, today, the clinical benefits of ET receptor antagonists appear to be limited by the development of fluid retention and peripheral edema which have been reported to occur with all antagonists, but more so with non-selective ET antagonists. Fluid retention, like headache, nausea and nasal congestion probably represent class side-effects. Nevertheless, provided a good equilibrium can be obtained between their clinical benefits and their tolerability profile, ET receptor blockers remain promising for the management of patients with chronic kidney diseases.

The endothelin receptor antagonist avosentan may cause fluid overload at doses of 25 and 50 mg, but the actual mechanisms of this effect are unclear. We conducted a placebo-controlled study in 23 healthy subjects to assess the renal effects of avosentan and the dose dependency of these effects. Oral avosentan was administered once daily for 8 days at doses of 0.5, 1.5, 5, and 50 mg. The drug induced a dose-dependent median increase in body weight, most pronounced at 50 mg (0.8 kg on day 8). Avosentan did not affect renal hemodynamics or plasma electrolytes. A dose-dependent median reduction in the fractional renal excretion of sodium was found (up to 8.7% at avosentan 50 mg); this reduction was paralleled by a dose-related increase in proximal sodium reabsorption. It is suggested that avosentan dose-dependently induces sodium retention by the kidney, mainly through proximal tubular effects. The potential clinical benefits of avosentan should therefore be investigated at doses of ≤ 5 mg. Clinical Pharmacology & Therapeutics (2009); 85, 6, 628–634 doi:10.1038/clpt.2009.15

Previous studies reported on the association of left ventricular mass index (LVMI) with urinary sodium or with circulating or urinary aldosterone. We investigated the independent associations of LVMI with the urinary excretion of both sodium and aldosterone. We randomly recruited 317 untreated subjects from a white population (45.1% women; mean age 48.2 years). Measurements included echocardiographic left ventricular (LV) properties, the 24-hour urinary excretion of sodium and aldosterone, plasma renin activity (PRA), and proximal (RNa prox ) and distal (RNa dist ) renal sodium reabsorption, assessed from the endogenous lithium clearance. In multivariable-adjusted models, we expressed changes in LVMI per 1-SD increase in the explanatory variables, while accounting for sex, age, systolic blood pressure, and the waist-to-hip ratio. LVMI increased independently with the urinary excretion of both sodium (+2.48 g/m 2 ; P =0.005) and aldosterone (+2.63 g/m 2 ; P =0.004). Higher sodium excretion was associated with increased mean wall thickness (MWT: +0.126 mm, P =0.054), but with no change in LV end-diastolic diameter (LVID: +0.12 mm, P =0.64). In contrast, higher aldosterone excretion was associated with higher LVID (+0.54 mm; P =0.017), but with no change in MWT (+0.070 mm; P =0.28). Higher RNa dist was associated with lower relative wall thickness (−0.81×10 −2 , P =0.017), because of opposite trends in LVID (+0.33 mm; P =0.13) and MWT (−0.130 mm; P =0.040). LVMI was not associated with PRA or RNa prox. In conclusion, LVMI independently increased with both urinary sodium and aldosterone excretion. Increased MWT explained the association of LVMI with urinary sodium and increased LVID the association of LVMI with urinary aldosterone.

BACKGROUND: Segmental handling of sodium along the proximal and distal nephron might be heritable and different between black and white participants. METHODS: We randomly recruited 95 nuclear families of black South African ancestry and 103 nuclear families of white Belgian ancestry. We measured the (FENa) and estimated the fractional renal sodium reabsorption in the proximal (RNaprox) and distal (RNadist) tubules from the clearances of endogenous lithium and creatinine. In multivariable analyses, we studied the relation of RNaprox and RNadist with FENa and estimated the heritability (h) of RNaprox and RNadist. RESULTS: Independent of urinary sodium excretion, South Africans (n = 240) had higher RNaprox (unadjusted median, 93.9% vs. 81.0%; P < 0.001) than Belgians (n = 737), but lower RNadist (91.2% vs. 95.1%; P < 0.001). The slope of RNaprox on FENa was steeper in Belgians than in South Africans (-5.40 +/- 0.58 vs. -0.78 +/- 0.58 units; P < 0.001), whereas the opposite was true for the slope of RNadist on FENa (-3.84 +/- 0.19 vs. -13.71 +/- 1.30 units; P < 0.001). h of RNaprox and RNadist was high and significant (P < 0.001) in both countries. h was higher in South Africans than in Belgians for RNaprox (0.82 vs. 0.56; P < 0.001), but was similar for RNadist (0.68 vs. 0.50; P = 0.17). Of the filtered sodium load, black participants reabsorb more than white participants in the proximal nephron and less postproximally. CONCLUSION: Segmental sodium reabsorption along the nephron is highly heritable, but the capacity for regulation in the proximal and postproximal tubules differs between whites and blacks.

Activation of type-1 dopamine receptors ( DRD1 ) reduces renal sodium reabsorption. In a family-based random sample of 611 untreated whites (women, 45.0%; mean age, 38.6 years), we measured blood pressure (BP). We used the endogenous lithium clearance to assess fractional sodium excretion (FE Na ) and proximal (RNa prox ) and distal (RNa dist ) tubular sodium reabsorption. We investigated multivariate-adjusted associations with the DRD1 promoter ( A − 48G , G − 94A , and C − 800T ) and GRK4 (Ala142Val ). The frequent DRD1 haplotypes were AGC (48.2%), GGT (34.4%), and AAC (14.3%). While standardizing to mean sodium excretion (8.7 mmol/h) and adjusting for covariates and relatedness, RNa dist was lower in DRD1 − 94GG homozygotes than − 94A allele carriers (effect size, −0.94%; P =0.005) with opposite findings for FE Na (+0.084%; P =0.014). AGC carriers (−0.88%; P =0.012) and AAC carriers (+1.00%; P =0.004) had different RNa dist compared to corresponding noncarriers. Furthermore, FE Na was lower in AAC carriers than in noncarriers (−0.082%; P =0.019). The family-based analyses identified a significant between-family component in the variance of the renal phenotypes associated with the DRD1 polymorphisms. Transmission of the DRD1 AGC haplotype was also associated with lower systolic (−3.54 mm Hg; P =0.016) and diastolic (−2.80 mm Hg; P =0.0064) BPs without significant between-family variance component. Plasma renin activity and urinary aldosterone excretion were not associated with DRD1 variation. The GRK4 Ala142Val polymorphism did not contribute to the phenotypes under study. In conclusion, renal sodium handling and BP were associated with genetic variation in the DRD1 promoter. The between-family variance component excluded population stratification for BP, but not for the renal phenotypes.

The basolateral Na pump drives renotubular reabsorption. In cultured renal cells, mutant adducins, as well as sub-nanomolar ouabain concentrations, stimulate the Na-K pump.To determine whether these factors interact and affect Na handling and blood pressure (BP) in vivo, we studied 155 untreated hypertensive patients subdivided on the basis of their plasma endogenous ouabain or alpha-adducin genotype (ADD1 Gly460Trp-rs4961).Under basal conditions, proximal tubular reabsorption and plasma Na were higher in patients with mutated Trp ADD1 or increased endogenous ouabain (P = 0.002 and 0.05, respectively). BPs were higher in the high plasma endogenous ouabain group (P = 0.001). Following volume loading, the increment in BP (7.73 vs. 4.81 mmHg) and the slopes of the relationship between BP and Na excretion were greater [0.017 +/- 0.002 vs. 0.009 +/- 0.003 mmHg/(muEq min)] in ADD1 Trp vs. ADD1 Gly carriers (P0.05). BP changes were similar, whereas the slopes of the relationship between BP and Na excretion were lower [0.016 +/- 0.003 vs. 0.008 +/- 0.002 mmHg/(muEq min)] in patients with low vs. high endogenous ouabain (P0.05). In patients with high endogenous ouabain, volume loading increased the BP in the ADD1 Trp group but not in the Gly group (P0.05). Thus, patients with ADD1 Trp alleles are sensitive to salt and tubular Na reabsorption remains elevated after volume expansion.With saline loading, BP changes are similar in high and low endogenous ouabain patients, whereas tubular Na reabsorption increases in the high endogenous ouabain group. Saline loading unmasks differences in renal Na handling in patients with mutant adducin or high endogenous ouabain and exposes an interaction of endogenous ouabain and Trp alleles on BP.

Leptin and adiponectin are adipokines with respective pro-atherogenic and anti-atherogenic properties, defining the plasma leptin/adiponectin ratio as a novel marker for atherosclerosis. In non-renal patients, both hyperleptinemia and hypoadiponectinemia are associated with cardiovascular complications. In peritoneal dialysis (PD) patients, the leptin/adiponectin ratio is markedly elevated, which is consistent with their increased cardiovascular risk. As glucose metabolism regulates adipokines, we hypothesized that glucose and/or other PD fluid components may affect adipokine production balance. This review summarizes the available data arising from research in this area. In 3T3-L1 adipocytes, glucose-based PD4 1.36% significantly increased leptin secretion vs amino-acid-based (AA) and icodextrin (ICOD)-based PD fluids. In contrast, adiponectin secretion was significantly reduced by PD4 1.36% vs glucose-free dialysates. Glucose concentration in PD fluids was shown to determine leptin secretion. Preliminary data from PD patients showed that a single 6-h dwell with PD4 3.86% glucose acutely increased plasma leptin vs AA (P0.05). The reduction in glucose load in a standard PD regimen was associated with an improvement in the plasma leptin/adiponectin ratio at 6 months. pH-neutral PD fluids increased leptin secretion in vitro vs acidic PD fluids, without effect on adiponectin. Whether this effect may have an impact on plasma leptin levels in PD patients is unknown. In conclusion, glucose-based PD fluids worsen the adipokine production balance in vitro while glucose-free solutions improve it. In PD patients, hypertonic glucose-based PD fluids may increase plasma leptin levels. Glucose-sparing PD regimens appear to improve the leptin/adiponectin ratio. However, their potential to reduce cardiovascular complications needs to be demonstrated.

Interactions between sodium and calcium regulating systems are poorly characterized but clinically important. Parathyroid hormone (PTH) levels are increased shortly after furosemide treatment by an unknown mechanism, and this effect is blunted by the previous administration of a calcimimetic in animal studies. Here, we explored further the possible underlying mechanisms of this observation in a randomized crossover placebo-controlled study performed in 18 human males. Volunteers took either cinacalcet (60 mg) or placebo and received a 20 mg furosemide injection 3 h later. Plasma samples were collected at 15-min intervals and analyzed for intact PTH, calcium, sodium, potassium, magnesium, phosphate, plasma renin activity (PRA), and aldosterone up to 6 h after furosemide injection. Urinary electrolyte excretion was also monitored. Subjects under placebo presented a sharp increase in PTH levels after furosemide injection. In the presence of cinacalcet, PTH levels were suppressed and marginal increase of PTH was observed. No significant changes in electrolytes and urinary excretion were identified that could explain the furosemide-induced increase in PTH levels. PRA and aldosterone were stimulated by furosemide injection but were not affected by previous cinacalcet ingestion. Expression of NKCC1, but not NKCC2, was found in parathyroid tissue. In conclusion, our results indicate that furosemide acutely stimulates PTH secretion in the absence of any detectable electrolyte changes in healthy adults. A possible direct effect of furosemide on parathyroid gland needs further studies.

Aldosterone is the main mineralocorticoid hormone controlling sodium balance, fluid homeostasis, and blood pressure by regulating sodium reabsorption in the aldosterone-sensitive distal nephron (ASDN). Germline loss-of-function mutations of the mineralocorticoid receptor (MR) in humans and in mice lead to the "renal" form of type 1 pseudohypoaldosteronism (PHA-1), a case of aldosterone resistance characterized by salt wasting, dehydration, failure to thrive, hyperkalemia, and metabolic acidosis. To investigate the importance of MR in adult epithelial cells, we generated nephron-specific MR knockout mice (MR(Pax8/LC1)) using a doxycycline-inducible system. Under standard diet, MR(Pax8/LC1) mice exhibit inability to gain weight and significant weight loss compared to control mice. Interestingly, despite failure to thrive, MR(Pax8/LC1) mice survive but develop a severe PHA-1 phenotype with higher urinary Na(+) levels, decreased plasma Na(+), hyperkalemia, and higher levels of plasma aldosterone. This phenotype further worsens and becomes lethal under a sodium-deficient diet. Na(+)/Cl(-) co-transporter (NCC) protein expression and its phosphorylated form are downregulated in the MR(Pax8/LC1) knockouts, as well as the αENaC protein expression level, whereas the expression of glucocorticoid receptor (GR) is increased. A diet rich in Na(+) and low in K(+) does not restore plasma aldosterone to control levels but is sufficient to restore body weight, plasma, and urinary electrolytes. In conclusion, MR deletion along the nephron fully recapitulates the features of severe human PHA-1. ENaC protein expression is dependent on MR activity. Suppression of NCC under hyperkalemia predominates in a hypovolemic state.