Stéphanie Baron

Resveratrol induces a mitochondrial complex I-dependent increase in NADH oxidation responsible for sirtuin activation in liver cells.

Background: The mechanism of action of resveratrol on sirtuin and mitochondrial metabolism remains elusive. Results: Resveratrol increases the mitochondrial NAD+ level by direct stimulation of complex I, leading to a SIRT3-dependent increase in substrate supplies. Conclusion: Our results link the direct stimulation by resveratrol of NADH oxidation to the SIRT3 activation. Significance: Mitochondrial NAD+/NADH ratio is a critical parameter mediating resveratrol effect on mitochondrial function. Resveratrol (RSV) has been shown to be involved in the regulation of energetic metabolism, generating increasing interest in therapeutic use. SIRT1 has been described as the main target of RSV. However, recent reports have challenged the hypothesis of its direct activation by RSV, and the signaling pathways remain elusive. Here, the effects of RSV on mitochondrial metabolism are detailed both in vivo and in vitro using murine and cellular models and isolated enzymes. We demonstrate that low RSV doses (1–5 μm) directly stimulate NADH dehydrogenases and, more specifically, mitochondrial complex I activity (EC50 ∼1 μm). In HepG2 cells, this complex I activation increases the mitochondrial NAD+/NADH ratio. This higher NAD+ level initiates a SIRT3-dependent increase in the mitochondrial substrate supply pathways (i.e. the tricarboxylic acid cycle and fatty acid oxidation). This effect is also seen in liver mitochondria of RSV-fed animals (50 mg/kg/day). We conclude that the increase in NADH oxidation by complex I is a crucial event for SIRT3 activation by RSV. Our results open up new perspectives in the understanding of the RSV signaling pathway and highlight the critical importance of RSV doses used for future clinical trials.

Assessment of hydration status in a large population

Both acute and chronic dehydration can have important implications for human behaviour and health. Young children, non-autonomous individuals and the elderly are at a greater risk of dehydration. Mild hypertonic dehydration could be related to less efficient cognitive and physical performance and has been reported to be associated with frequently occurring pathological conditions, especially nephrolithiasis. The assessment of hydration status in a large sample appears to be of interest for conducting epidemiological and large clinical studies aimed at improving preventive and curative care. Especially in large-population studies, methods that are used have to be accurate, cheap, quick and require no technical expertise. Body weight change is widely used to determine acute hydration changes, but seems to be insufficiently accurate in longitudinal studies. Bioimpedance analysis methods enable the assessment of total body water content, but their use is still under debate. Because plasma osmolality directly reflects intracellular osmolality, it constitutes a good marker to assess acute hydration changes, but not chronic hydration status because it changes constantly. Moreover, venepuncture is considered to be invasive and is not suitable for a large-sample study, especially in children. Urinary markers appear to be good alternatives for assessing hydration status in large populations. Collection of urine samples is non-invasive and cheap. High technical expertise is not required to perform urinary marker measurements and these measurements can be carried out quickly. Thus, methods based on urinary markers are very well suited for field studies. Urine colour is probably the least sensitive marker despite its high specificity. Urine osmolality and especially urine specific gravity could be easily used for determining hydration status in large-sample studies.

Plasma Soluble (Pro)Renin Receptor Is Independent of Plasma Renin, Prorenin, and Aldosterone Concentrations But Is Affected by Ethnicity

A soluble (pro)renin receptor (sPRR) circulates in plasma and is able to bind renin and prorenin. It is not known whether plasma sPRR concentrations vary with the activity of the renin–angiotensin system. We measured plasma sPRR, renin, prorenin, and aldosterone concentrations in 121 white and 9 black healthy subjects, 40 patients with diabetes mellitus, 41 hypertensive patients with or without renin–angiotensin system blockers, 9 patients with primary aldosteronism, and 10 patients with Gitelman syndrome. Median physiological plasma sPRR concentration was 23.5 ng/mL (interquartile range, 20.9–26.5) under usual uncontrolled sodium diet. sPRR concentration in healthy subjects, unlike renin and prorenin, did not display circadian variation or dependence on age, sex, posture, or hormonal status. sPRR concentrations were ≈25% lower in black than in white subjects, whereas renin concentrations were ≈40% lower. Patients with diabetes mellitus (average renin–high prorenin levels) and with hypertension only (average renin–average prorenin levels) had sPRR concentrations similar to healthy subjects. Renin–angiotensin system blockade was associated with increase of sPRR concentration by ≈12%. sPRR in patients with primary aldosteronism (low renin–low prorenin) and Gitelman syndrome (high renin–high prorenin) were similar and ≈10% higher than in healthy subjects. There was no correlation between sPRR and renin or prorenin. In conclusion, our results show that plasma sPRR concentrations are dependent on ethnicity and independent of renin, prorenin, and aldosterone concentrations in healthy subjects and in patients with contrasted degrees of renin–angiotensin system activity.

Familial Hypocalciuric Hypercalcemia Types 1 and 3 and Primary Hyperparathyroidism: Similarities and Differences

CONTEXT Familial hypocalciuric hypercalcemia (FHH) is a genetically heterogeneous condition resembling primary hyperparathyroidism (PHPT) but not curable by surgery; FHH types 1, 2, and 3 are due to loss-of-function mutations of the CASR, GNA11, or AP2S1 genes, respectively. OBJECTIVE This study aimed to compare the phenotypes of patients with genetically proven FHH types 1 or 3 or PHPT. DESIGN, SETTING, AND PATIENTS This was a mutation analysis in a large cohort, a cross-sectional comparison of 52 patients with FHH type 1, 22 patients with FHH type 3, 60 with PHPT, and 24 normal adults. INTERVENTION There were no interventions. MAIN OUTCOME MEASURES Abnormalities of the CASR, GNA11, and AP2S1 genes, blood calcium, phosphate, and PTH concentrations, urinary calcium excretion were measured. RESULTS In 133 families, we detected 101 mutations in the CASR gene, 68 of which were previously unknown, and in 19 families, the three recurrent AP2S1 mutations. No mutation was detected in the GNA11 gene. Patients with FHH type 3 had higher plasma calcium concentrations than patients with FHH type 1, despite having similar PTH concentrations and urinary calcium excretion. Renal tubular calcium reabsorption levels were higher in patients with FHH type 3 than in those with FHH type 1. Plasma calcium concentration was higher whereas PTH concentration and urinary calcium excretion were lower in FHH patients than in PHPT patients. In patients with FHH or PHPT, all data groups partially overlapped. CONCLUSION In our population, AP2S1 mutations affect calcium homeostasis more severely than CASR mutations. Due to overlap, the risk of confusion between FHH and PHPT is high.

Indomethacin, Amiloride, or Eplerenone for Treating Hypokalemia in Gitelman Syndrome

Patients with Gitelman syndrome (GS), an inherited salt-losing tubulopathy, are usually treated with potassium-sparing diuretics or nonsteroidal anti-inflammatory drugs and oral potassium and magnesium supplementations. However, evidence supporting these treatment options is limited to case series studies. We designed an open-label, randomized, crossover study with blind end point evaluation to compare the efficacy and safety of 6-week treatments with one time daily 75 mg slow-release indomethacin, 150 mg eplerenone, or 20 mg amiloride added to constant potassium and magnesium supplementation in 30 patients with GS (individual participation: 48 weeks). Baseline plasma potassium concentration was 2.8±0.4 mmol/L and increased by 0.38 mmol/L (95% confidence interval [95% CI], 0.23 to 0.53; P<0.001) with indomethacin, 0.15 mmol/L (95% CI, 0.02 to 0.29; P=0.03) with eplerenone, and 0.19 mmol/L (95% CI, 0.05 to 0.33; P<0.01) with amiloride. Fifteen patients became normokalemic: six with indomethacin, three with eplerenone, and six with amiloride. Indomethacin significantly reduced eGFR and plasma renin concentration. Eplerenone and amiloride each increased plasma aldosterone by 3-fold and renin concentration slightly but did not significantly change eGFR. BP did not significantly change. Eight patients discontinued treatment early because of gastrointestinal intolerance to indomethacin (six patients) and hypotension with eplerenone (two patients). In conclusion, each drug increases plasma potassium concentration in patients with GS. Indomethacin was the most effective but can cause gastrointestinal intolerance and decreased eGFR. Amiloride and eplerenone have similar but lower efficacies and increase sodium depletion. The benefit/risk ratio of each drug should be carefully evaluated for each patient.

Double Knockout of the Na+-Driven Cl−/HCO3− Exchanger and Na+/Cl− Cotransporter Induces Hypokalemia and Volume Depletion

We recently described a novel thiazide-sensitive electroneutral NaCl transport mechanism resulting from the parallel operation of the Cl-/HCO3- exchanger pendrin and the Na+-driven Cl-/2HCO3- exchanger (NDCBE) in β-intercalated cells of the collecting duct. Although a role for pendrin in maintaining Na+ balance, intravascular volume, and BP is well supported, there is no in vivo evidence for the role of NDCBE in maintaining Na+ balance. Here, we show that deletion of NDCBE in mice caused only subtle perturbations of Na+ homeostasis and provide evidence that the Na+/Cl- cotransporter (NCC) compensated for the inactivation of NDCBE. To unmask the role of NDCBE, we generated Ndcbe/Ncc double-knockout (dKO) mice. On a normal salt diet, dKO and single-knockout mice exhibited similar activation of the renin-angiotensin-aldosterone system, whereas only dKO mice displayed a lower blood K+ concentration. Furthermore, dKO mice displayed upregulation of the epithelial sodium channel (ENaC) and the Ca2+-activated K+ channel BKCa. During NaCl depletion, only dKO mice developed marked intravascular volume contraction, despite dramatically increased renin activity. Notably, the increase in aldosterone levels expected on NaCl depletion was attenuated in dKO mice, and single-knockout and dKO mice had similar blood K+ concentrations under this condition. In conclusion, NDCBE is necessary for maintaining sodium balance and intravascular volume during salt depletion or NCC inactivation in mice. Furthermore, NDCBE has an important role in the prevention of hypokalemia. Because NCC and NDCBE are both thiazide targets, the combined inhibition of NCC and the NDCBE/pendrin system may explain thiazide-induced hypokalemia in some patients.

Resveratrol Directly Binds to Mitochondrial Complex I and Increases Oxidative Stress in Brain Mitochondria of Aged Mice.

Resveratrol is often described as a promising therapeutic molecule for numerous diseases, especially in metabolic and neurodegenerative disorders. While the mechanism of action is still debated, an increasing literature reports that resveratrol regulates the mitochondrial respiratory chain function. In a recent study we have identified mitochondrial complex I as a direct target of this molecule. Nevertheless, the mechanisms and consequences of such an interaction still require further investigation. In this study, we identified in silico by docking study a binding site for resveratrol at the nucleotide pocket of complex I. In vitro, using solubilized complex I, we demonstrated a competition between NAD+ and resveratrol. At low doses (<5μM), resveratrol stimulated complex I activity, whereas at high dose (50 μM) it rather decreased it. In vivo, in brain mitochondria from resveratrol treated young mice, we showed that complex I activity was increased, whereas the respiration rate was not improved. Moreover, in old mice with low antioxidant defenses, we demonstrated that complex I activation by resveratrol led to oxidative stress. These results bring new insights into the mechanism of action of resveratrol on mitochondria and highlight the importance of the balance between pro- and antioxidant effects of resveratrol depending on its dose and age. These parameters should be taken into account when clinical trials using resveratrol or analogues have to be designed.

Dual Effects of Resveratrol on Arterial Damage Induced By Insulin Resistance in Aged Mice

Aging leads to increased insulin resistance and arterial dysfunction, with oxidative stress playing an important role. This study explored the metabolic and arterial effects of a chronic treatment with resveratrol, an antioxidant polyphenol compound that has been shown to restore insulin sensitivity and decrease oxidative stress, in old mice with or without a high-protein diet renutrition care. High-protein diet tended to increase insulin resistance and atheromatous risk. Resveratrol improved insulin sensitivity in old mice fed standard diet by decreasing homeostasis model of assessment-insulin resistance and resistin levels. However, resveratrol did not improve insulin resistance status in old mice receiving the high-protein diet. In contrast, resveratrol exhibited deleterious effects by increasing inflammation state and superoxide production and diminishing aortic distensibility. In conclusion, we demonstrate that resveratrol has beneficial or deleterious effects on insulin sensitivity and arterial function, depending on nutritional status in our models.

Estimated or Measured GFR in Living Kidney Donors Work-up?

The value of estimated glomerular filtration rate (eGFR) in living kidney donors screening is unclear. A recently published web‐based application derived from large cohorts, but not living donors, calculates the probability of a measured GFR (mGFR) lower than a determined threshold. Our objectives were to validate the clinical utility of this tool in a cohort of living donors and to test two other strategies based on chronic kidney disease epidemiology collaboration (CKD‐EPI) and on MDRD‐eGFR. GFR was measured using 51Cr‐ ethylene‐diamine tetraacetic acid urinary clearance in 311 potential living kidney donors (178 women, mean age 50 ± 11.6 years). The web‐based tool was used to predict those with mGFR < 80 mL/min/1.73 m2. Inputs to the application were sex, age, ethnicity, and plasma creatinine. In our cohort, a web‐based probability of mGFR <90 mL/min/1.73 m2 higher than 2% had 100% sensitivity for detection of actual mGFR <80 mL/min/1.73 m2. The positive predictive value was 0.19. A CKD‐EPI‐eGFR threshold of 104 mL/min/1.73 m2 and an MDRD‐eGFR threshold of 100 mL/min/1.73 m2 had 100% sensitivity to detect donors with actual mGFR <80 mL/min/1.73 m2. We obtained similar results in an external cohort of 354 living donors. We confirm the usefulness of the web‐based application to identify potential donors who should benefit from GFR measurement.

Resveratrol Decreases TXNIP mRNA and Protein Nuclear Expressions With an Arterial Function Improvement in Old Mice

Aging leads to a high prevalence of glucose intolerance and cardiovascular diseases, with oxidative stress playing a potential role. Resveratrol has shown promising effects on glucose tolerance and tends to improve endothelial function in elderly patients. Thioredoxin-interacting protein (TXNIP) was recently proposed as a potential link connecting glucose metabolism to oxidative stress. Here, we investigated the resveratrol-induced improvement of arterial aging phenotype in old mice and the expression of aortic TXNIP. Using an in vivo model of old mice with or without 3-month resveratrol treatment, we investigated the effects of resveratrol on age-related impairments from a cardiovascular Doppler analysis, to a molecular level, by studying inflammation and oxidative stress factors. We found a dual effect of resveratrol, with a decrease of age-related glucose intolerance and oxidative stress imbalance leading to reduced matrix remodeling that forestalls arterial aging phenotype in terms of intima-media thickness and arterial distensibility. These results provide the first evidence that aortic TXNIP mRNA and protein nuclear expressions are increased in the arterial aging and decreased by resveratrol treatment. In conclusion, we demonstrated that resveratrol helped to restore several aging impaired processes in old mice, with a decrease of aortic TXNIP mRNA and protein nuclear expressions.