Role of ENaC in skin wound healing and phenotypic analysis of GILZ-deficient mice

In my first project, I analyzed the role of the amiloride-sensitive epithelial sodium channel ENaC) in the skin during wound healing. ENaC is present in the skin and a function in keratinocyte differentiation and barrier formation has been demonstrated. Previous findings suggested, that ENaC might be implicated in keratinocyte migration, although its role in wound healing was not analyzed yet. Using skin-specific (K14-Cre) conditional ENaC knockout and overexpressing mice, I determined the wound closure kinetic and performed morphometric measurements. The time course of wound repair was not significantly different in knockouts or transgenics when compared to control mice and the morphology of the closing wound was not altered. In my second project, I studied the glucocorticoid-induced leucine zipper (GILZ, Tsc22d3). GILZ is widely expressed and an important role has been predicted in immunity, adipogenesis and renal sodium handling. Mice were generated that constitutively lack all the functional domains of the Gilz gene. In these mice, the expression of GILZ mRNA transcripts and protein were completely abolished in all tissues tested. Surprisingly, knockout mice survived. To test whether GILZ mimicks glucocorticoid action, we studied its implication in T- and B- cell development and in a model of sepsis. We measured cytokine secretion in different inflammatory models, like in peritoneal and bone marrow-derived macrophages, in splenocytes and a model of sepsis. In all our experiments, cytokine secretion from GILZ- deficient cells was not different from controls. From 6 months onwards, knockout mice contained significantly less body fat and were lighter. Following sodium and water deprivation experiments, water and salt homeostasis was preserved. Sterility of knockout males was associated with a severe testis dysplasia, smaller seminiferous tubules, the number of Sertoli and germ cell was reduced while increased apoptosis, but not cell proliferation, was evidenced. The interstitial Leydig cell population was augmented, and higher plasma FSH and testosterone levels were found. Interestingly, the expression of the target gene Ppar2 was diminished in the testis and in the liver, but not in the skin, kidney or fat. Tsc22d1 mRNA transcript level was found to be upregulated in testis, but not in the kidney or fat tissue. In most tissue, excepted the testis, GILZ-deficient mice reveal functional redundancy amongst members of the Tsc22d family or genes involved in the same regulatory pathways. In summary, contrarily to the published in vitro data, GILZ does not play a crucial role attributed in immunology or inflammation, but we identified a novel function in spermatogenesis. -- Dans mon premier projet, j'ai analysé le rôle du canal épithélial sodique sensible à l'amiloride (ENaC) dans la cicatrisation de la peau. ENaC est présent dans la peau et il a une fonction dans la différenciation des kératinocytes et dans la formation de la barrière. Des études suggèrent qu'ENaC pourrait être impliqué dans la migration des kératinocytes, cependant, son rôle dans la cicatrisation n'a pas encore été étudié. A l'aide de souris qui surexpriment ou qui sont knockout pour ENaC, spécifiquement dans la peau (K14-Cre), j'ai analysé le temps de clôture de la cicatrice et j'ai aussi étudié la morphologie de la plaie guérissant. Chez les souris qui surexpriment ou chez les knockouts, la vitesse de fermeture et la morphologie de la cicatrice étaient identiques aux souris contrôles. Dans mon second projet, j'ai étudié le glucocorticoid-induced leucine zipper (GILZ, Tsc22d3). GILZ est largement exprimé et un rôle important a été prédit dans l'immunité, l'adipogénèse et le transport sodique rénal. Des souris ont été générées dont les domaines fonctionnels du gène Gilz sont éliminés. L'expression de GILZ en ARNm et protéine a été complètement abolie dans tous les tissus testés. Étonnamment, ces souris knockout survivent. Afin de tester si GILZ imite les effets des glucocorticoïdes, nous avons étudié son implication dans le développement des cellules T et B ainsi qu'un modèle de septicémie. Nous avons mesuré la sécrétion de cytokines à partir de différents modèles d'inflammation tels que des macrophages péritonéaux ou de moelle, de splénocytes ou encore d'un modèle de septicémie. Dans toutes nos expériences, la sécrétion de cytokines de cellules GILZ-déficientes était semblable. Dès 6 mois, les knockouts contenaient significativement moins de graisses et étaient plus légères. Suite à une privation sodique et aqueuse, l'homéostasie du sel et de l'eau était préservée. Les mâles knockouts présentaient une stérilité accompagnée d'une dysplasie testiculaire sévère, de tubules séminifères étaient plus petits et contenaient un nombre réduit de cellules de Sertoli et de cellules germinales. L'apoptose était augmentée dans ces cellules mais pas la prolifération cellulaire. Le nombre de cellules de Leydig était aussi plus élevé, ainsi que la FSH et la testostérone. L'expression du gène cible Pparγ2 était diminuée dans le testicule et le foie, mais pas dans la peau, le rein ou le tissu adipeux. L'ARNm de Tsc22d1 était plus exprimé dans le testicule, mais pas dans le rein ou le tissu adipeux. Dans la plupart des tissus, sauf le testicule, les souris knockouts révélaient une redondance fonctionnelle des autres membres de la famille Tsc22d ou de gènes impliqués dans les mêmes voies de régulation. En résumé, contrairement aux données in vitro, GILZ ne joue pas un rôle essentiel en immunologie, mais nous avons identifié une nouvelle fonction dans la spermatogénèse.

The neuroretina is a novel mineralocorticoid target: aldosterone up-regulates ion and water channels in Müller glial cells.

Glucocorticoids reduce diabetic macular edema, but the mechanisms underlying glucocorticoid effects are imperfectly elucidated. Glucocorticoids may bind to glucocorticoid (GR) and mineralocorticoid (MR) receptors. We hypothesize that MR activation may influence retinal hydration. The effect of the MR agonist aldosterone (24 h) on ion/water channel expression (real-time PCR, Western blot, immunofluorescence) was investigated on cultured retinal Müller glial cells (RMGs, which contribute to fluid homeostasis in the retina), in Lewis rat retinal explants, and in retinas from aldosterone-injected eyes. We evidenced cell-specific expression of MR, GR, and 11-beta-hydroxysteroid dehydrogenase type II. Aldosterone significantly enhances expression of sodium and potassium channels ENaC-alpha (6.5-fold) and Kir4.1 (1.9-fold) through MR and GR occupancy, whereas aquaporin 4 (AQP4, 2.9-fold) up-regulation is MR-selective. Aldosterone intravitreous injection induces retinal swelling (24% increase compared to sham-injected eyes) and activation of RMGs. It promotes additional localization of Kir4.1 and AQP4 toward apical microvilli of RMGs. Our results highlight the mineralocorticoid-sensitivity of the neuroretina and show that aldosterone controls hydration of the healthy retina through regulation of ion/water channels expression in RMGs. These results provide a rationale for future investigations of abnormal MR signaling in the pathological retina.

BMI group-related differences in physical fitness and physical activity in preschool-age children: a cross-sectional analysis.

In the Ballabeina study, we investigated age- and BMI-group-related differences in aerobic fitness (20 m shuttle run), agility (obstacle course), dynamic (balance beam) and static balance (balance platform), and physical activity (PA, accelerometers) in 613 children (M age = 5.1 years, SD = 0.6). Normal weight (NW) children performed better than overweight (OW) children in aerobic fitness, agility, and dynamic balance (all p <.001), while OWchildren had a better static balance (p < .001). BMI-group-related differences in aerobic fitness and agility were larger in older children (p for interaction with age = .01) in favor of the NW children. PA did not differ between NW and OW (p > or = .1), but did differ between NW and obese children (p < .05). BMI-group-related differences in physical fitness can already be present in preschool-age children.

Explaining Fiscal Balances with a Simultaneous Equation Model of Revenue and Expenditure : A Case Study of Swiss Cantons Using Panel Data

Empirical literature on the analysis of the efficiency of measures for reducing persistent government deficits has mainly focused on the direct explanation of deficit. By contrast, this paper aims at modeling government revenue and expenditure within a simultaneous framework and deriving the fiscal balance (surplus or deficit) equation as the difference between the two variables. This setting enables one to not only judge how relevant the explanatory variables are in explaining the fiscal balance but also understand their impact on revenue and/or expenditure. Our empirical results, obtained by using a panel data set on Swiss Cantons for the period 1980-2002, confirm the relevance of the approach followed here, by providing unambiguous evidence of a simultaneous relationship between revenue and expenditure. They also reveal strong dynamic components in revenue, expenditure, and fiscal balance. Among the significant determinants of public fiscal balance we not only find the usual business cycle elements, but also and more importantly institutional factors such as the number of administrative units, and the ease with which people can resort to political (direct democracy) instruments, such as public initiatives and referendum.

Roles of PPARβ and PPARγ in mouse placental development

RESUME Les gènes des PPARs jouent des rôles importants dans la régulation du métabolisme énergétique, lipidique et glucidique. Le présent travail, caractérise et analyse les défauts placentaires responsables de la mort embryonnaire des souris mutantes pour PPARβ et pour PPARγ, entre le jour E9.5 et E10.5. Les placentas issus d'embryons PPARP présentent un sévère retard de croissance, alors que les placentas mutants PPARγ montrent de graves défauts vasculaires. Nous montrons que les placentas issus d'embryons PPARβ-/-, au jour E9.5 présentent une réduction prononcée de la couche de cellules géantes, associée à une diminution des niveaux de protéines exprimées par les cellules géantes, tel que le placenta lactogène-I et la « proliferin ». Par ailleurs, nous montrons que le traitement d'un lignée trophoblastique par un ligand spécifique de PPARP augmente considérablement leur différentiation en cellules géantes. Cette différentiation dépendante de la voie de signalisation P13-kinase, s'accompagne d'une élévation de l'expression de l'ADRP, une protéine de structure associée aux vésicules lipidiques. Ainsi nous démontrons que PPAR5 est un régulateur majeur de la différentiation des cellules géantes, lesquelles sont primordiales aussi bien pour l'établissement de la structure placentaire, que pour la fonction endocrine. Par contre, les placentas PPARγ-/- présentent un défaut de vascularisation. Le niveau d'une protéine anti-angiogénique, la « proliferin-related protein », est très basse et ne peut pas contre-balancer l'élévation normale de la protéine pro-angiogénique « proliferin ». La formation des vaisseaux se trouve alors altérée. Ainsi, PPARγ constitue un régulateur majeur de l'activité anti-angiogénique. En conclusion, ce travail fournit de nouveaux éléments sur le rôle complémentaires de PPARβet PPARγ dans les événements complexes qui régissent le développement placentaire. SUMMARY Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors involved in energy homeostasis and growth. Herein, we characterize the placental defects that cause embryonic lethality around E9.5/E10.5 in PPARβ- and in PPARγ-deficient mouse lines. Most but not all PPARβ-null mutants die around E9.5/E10.5 with severe growth retardation. The placentas from PPARβ-/- embryos at E9.5 exhibit a strongly reduced giant cell layer, associated with reduced levels of proteins expressed by giant cells such as Placental lactogen-I and Proliferin. Ectopic treatment of a rat trophoblast cell line with PPARβ ligand markedly accelerated PI3 kinase-dependent giant cell differentiation. In addition, we demonstrate that ADRP, a pen-related lipid droplet-bound protein, is up-regulated by PPARβ in differentiated Rcho-1 cells. These results indicate that PPARβ is a crucial regulator of the differentiation secondary giant cells, which play a major role in the establishment of the placental structure as well as an important endocrine function. In contrast, the main alteration of the PPARγ-null placentas concerns the vasculogenesis. We show that in these placentas, the level of the anti-angiogenic proliferin-related protein is very low, and cannot balance the normal elevation of the pro-angiogenic proliferin expression, leading to the defective placental vessel formation. Consistently, the dramatic increase of PPARγ expression in late stage of gestation in wild-type mice is likely a major regulator of the anti-angiogenic activity, particularly important at the end of the pregnancy. This work emphasizes the important and complementary roles of PPARβ and PPARγ in mouse placental development and provides new tools for understanding the complex regulatory events that governs placental development and function. Understanding the function of PPARβ and PPARγ are of crucial interest with respect to human placental development and associated pathologies.

The cataract and glucosuria associated monocarboxylate transporter MCT12 is a new creatine transporter.

Creatine transport has been assigned to creatine transporter 1 (CRT1), encoded by mental retardation associated SLC6A8. Here, we identified a second creatine transporter (CRT2) known as monocarboxylate transporter 12 (MCT12), encoded by the cataract and glucosuria associated gene SLC16A12. A non-synonymous alteration in MCT12 (p.G407S) found in a patient with age-related cataract (ARC) leads to a significant reduction of creatine transport. Furthermore, Slc16a12 knockout (KO) rats have elevated creatine levels in urine. Transport activity and expression characteristics of the two creatine transporters are distinct. CRT2 (MCT12)-mediated uptake of creatine was not sensitive to sodium and chloride ions or creatine biosynthesis precursors, breakdown product creatinine or creatine phosphate. Increasing pH correlated with increased creatine uptake. Michaelis-Menten kinetics yielded a Vmax of 838.8 pmol/h/oocyte and a Km of 567.4 µm. Relative expression in various human tissues supports the distinct mutation-associated phenotypes of the two transporters. SLC6A8 was predominantly found in brain, heart and muscle, while SLC16A12 was more abundant in kidney and retina. In the lens, the two transcripts were found at comparable levels. We discuss the distinct, but possibly synergistic functions of the two creatine transporters. Our findings infer potential preventive power of creatine supplementation against the most prominent age-related vision impaired condition.

Health at work and risk factors related to ergonomics: some results from the Swiss sample of the 4th European working conditions survey

Introduction The European Foundation for the improvement of living and working conditions conducts a survey every 5 years since 1990. The foundation also offers the possibility to non-EU countries to be included in the survey: in 2005, Switzerland took part for the first time in the fourth edition of this survey. The Institute for Work and Health (IST) has been associated to the Swiss project conducted under the leadership of the SECO and the Fachhochschule Nordwestschweiz. The survey covers different aspects of work like job characteristics and employment conditions, health and safety, work organization, learning and development opportunities, and the balance between working and non-working life (Parent-Thirion, Fernandez Macias, Hurley, & Vermeylen, 2007). More particularly, one question assesses the worker's self-perception of the effects of work on health. We identified (for the Swiss sample) several factors affecting the risk to report health problems caused by work. The Swiss sample includes 1040 respondents. Selection of participants was based on a random multi-stage sampling and was carried out by M.I.S Trend S.A. (Lausanne). Participation rate was 59%. The database was weighted by household size, gender, age, region of domicile, occupational group, and economic sector. Specially trained interviewers carried out the interviews at the respondents home. The survey was carriedout between the 19th of September 2005 and the 30th of November 2005. As detailed in (Graf et al., 2007), 31% of the Swiss respondents identify work as the cause of health problems they experience. Most frequently reported health problems include back pain (18%), stress (17%), muscle pain (13%), and overall fatigue (11%). Ergonomic aspects associated with higher risk of reporting health problems caused by work include frequent awkward postures (odds ratio [OR] 4.7, 95% confidence interval [CI] 3.1 to 5.4), tasks involving lifting heavy loads (OR 2.7, 95% CI 2.0 to 3.6) or lifting people (OR 2.2, 95% CI 1.4 to 3.5), standing or walking (OR 1.4, 95% CI 1.1 to 1.9), as well as repetitive movements (OR 1.7, 95% CI 1.3 to 2.3). These results highlight the need to continue and intensify the prevention of work related health problems in occupations characterized by risk factors related to ergonomics.

A comprehensive analysis of gene expression profiles in distal parts of the mouse renal tubule.

The distal parts of the renal tubule play a critical role in maintaining homeostasis of extracellular fluids. In this review, we present an in-depth analysis of microarray-based gene expression profiles available for microdissected mouse distal nephron segments, i.e., the distal convoluted tubule (DCT) and the connecting tubule (CNT), and for the cortical portion of the collecting duct (CCD; Zuber et al., Proc Natl Acad Sci USA 106:16523-16528, 2009). Classification of expressed transcripts in 14 major functional gene categories demonstrated that all principal proteins involved in maintaining the salt and water balance are represented by highly abundant transcripts. However, a significant number of transcripts belonging, for instance, to categories of G-protein-coupled receptors or serine/threonine kinases exhibit high expression levels but remain unassigned to a specific renal function. We also established a list of genes differentially expressed between the DCT/CNT and the CCD. This list is enriched by genes related to segment-specific transport functions and by transcription factors directing the development of the distal nephron or collecting ducts. Collectively, this in silico analysis provides comprehensive information about relative abundance and tissue specificity of the DCT/CNT and the CCD expressed transcripts and identifies new candidate genes for renal homeostasis.

Chronic potassium depletion increases adrenal progesterone production that is necessary for efficient renal retention of potassium.

Modern dietary habits are characterized by high-sodium and low-potassium intakes, each of which was correlated with a higher risk for hypertension. In this study, we examined whether long-term variations in the intake of sodium and potassium induce lasting changes in the plasma concentration of circulating steroids by developing a mathematical model of steroidogenesis in mice. One finding of this model was that mice increase their plasma progesterone levels specifically in response to potassium depletion. This prediction was confirmed by measurements in both male mice and men. Further investigation showed that progesterone regulates renal potassium handling both in males and females under potassium restriction, independent of its role in reproduction. The increase in progesterone production by male mice was time dependent and correlated with decreased urinary potassium content. The progesterone-dependent ability to efficiently retain potassium was because of an RU486 (a progesterone receptor antagonist)-sensitive stimulation of the colonic hydrogen, potassium-ATPase (known as the non-gastric or hydrogen, potassium-ATPase type 2) in the kidney. Thus, in males, a specific progesterone concentration profile induced by chronic potassium restriction regulates potassium balance.

Prevalences of masked and white-coat hypertension in the Lausanne population: the Hercules Study

Background and Objectives: Few population-based data on the prevalences of masked and white-coat hypertension exist. We collected 24-hour ambulatory blood pressure (BP) and urine in a random subset of participants to the population-based CoLaus study. Methods: Clinic BP was measured using an Omron HEM 907 device and ambulatory BP (ABP) using a Diasys Integra device. Masked hypertension (MH) was defined as clinic BP < 140/90mm Hg and 24-hour ABP >¼135/85mmHg. White coat hypertension (WCH) was defined as clinic BP >¼ 140/90mm Hg and ABP <135/85mm Hg. Microalbuminuria was defined as present if urinary albumin excretion was > 20mg/min. Results: The 198 men and 213 women were aged (mean_SD) 56.2_10.7 and 57.2_10.3 years and had mean urinary excretion of 148_65 and 122_52 mmol/24 h for sodium and 70_24 and 5721 mmol/24 h for potassium, respectively. In men and women, the prevalences were 34.9% and 31.0% for clinic hypertension, 42.9% and 32.9% for ambulatory hypertension, 12.6% and 5.6% for MH, and 4.5% and 3.8% for WCH, respectively. The higher prevalence of MH in men was explained, in part, by higher alcohol consumption and smoking. Participants with MH tended to have higher microalbuminuria (13.5% vs 5.8%, P¼0.067). Participants with WCH had no microalbuminuria. Conclusions: In the Lausanne population aged 38 to 78 years, the prevalence of hypertension based on ABP was high, despite moderate dietary salt intake. Men had higher prevalence of MH then women. The prevalence of WCH was low and similar in men and women. MH tended to be associated with early kidney damage.

Influence of a lifestyle intervention in preschool children on physiological and psychological parameters (Ballabeina): study design of a cluster randomized controlled trial.

Childhood obesity and physical inactivity are increasing dramatically worldwide. Children of low socioeconomic status and/or children of migrant background are especially at risk. In general, the overall effectiveness of school-based programs on health-related outcomes has been disappointing. A special gap exists for younger children and in high risk groups. This paper describes the rationale, design, curriculum, and evaluation of a multicenter preschool randomized intervention study conducted in areas with a high migrant population in two out of 26 Swiss cantons. Twenty preschool classes in the German (canton St. Gallen) and another 20 in the French (canton Vaud) part of Switzerland were separately selected and randomized to an intervention and a control arm by the use of opaque envelopes. The multidisciplinary lifestyle intervention aimed to increase physical activity and sleep duration, to reinforce healthy nutrition and eating behaviour, and to reduce media use. According to the ecological model, it included children, their parents and the teachers. The regular teachers performed the majority of the intervention and were supported by a local health promoter. The intervention included physical activity lessons, adaptation of the built infrastructure; promotion of regional extracurricular physical activity; playful lessons about nutrition, media use and sleep, funny homework cards and information materials for teachers and parents. It lasted one school year. Baseline and post-intervention evaluations were performed in both arms. Primary outcome measures included BMI and aerobic fitness (20 m shuttle run test). Secondary outcomes included total (skinfolds, bioelectrical impedance) and central (waist circumference) body fat, motor abilities (obstacle course, static and dynamic balance), physical activity and sleep duration (accelerometry and questionnaires), nutritional behaviour and food intake, media use, quality of life and signs of hyperactivity (questionnaires), attention and spatial working memory ability (two validated tests). Researchers were blinded to group allocation. The purpose of this paper is to outline the design of a school-based multicenter cluster randomized, controlled trial aiming to reduce body mass index and to increase aerobic fitness in preschool children in culturally different parts of Switzerland with a high migrant population. Trial Registration: (clinicaltrials.gov) NCT00674544.

A novel vasopressin-induced transcript promotes MAP kinase activation and ENaC downregulation.

In the principal cell of the renal collecting duct, vasopressin regulates the expression of a gene network responsible for sodium and water reabsorption through the regulation of the water channel and the epithelial sodium channel (ENaC). We have recently identified a novel vasopressin-induced transcript (VIT32) that encodes for a 142 amino acid vasopressin-induced protein (VIP32), which has no homology with any protein of known function. The Xenopus oocyte expression system revealed two functions: (i) when injected alone, VIT32 cRNA rapidly induces oocyte meiotic maturation through the activation of the maturation promoting factor, the amphibian homolog of the universal M phase trigger Cdc2/cyclin; and (ii) when co-injected with the ENaC, VIT32 cRNA selectively downregulates channel activity, but not channel cell surface expression. In the kidney principal cell, VIP32 may be involved in the downregulation of transepithelial sodium transport observed within a few hours after vasopressin treatment. VIP32 belongs to a novel gene family ubiquitously expressed in oocyte and somatic cells that may be involved in G to M transition and cell cycling.

Relationship of aerobic fitness and motor skills with memory and attention in preschoolers (Ballabeina): A cross-sectional and longitudinal study.

Background: The debate about a possible relationship between aerobic fitness and motor skills with cognitive development in children has recently re-emerged, because of the decrease in children's aerobic fitness and the concomitant pressure of schools to enhance cognitive performance. As the literature in young children is scarce, we examined the cross-sectional and longitudinal relationship of aerobic fitness and motor skills with spatial working memory and attention in preschool children.Methods: Data from 245 ethnically diverse preschool children (mean age: 5.2 (0.6) years, girls: 49.4%) analyzed at baseline and 9 months later. Assessments included aerobic fitness (20 m shuttle run) and motor skills with agility (obstacle course) and dynamic balance (balance beam). Cognitive parameters included spatial working memory (IDS) and attention (KHV-VK). All analyses were adjusted for age, sex, BMI, migration status, parental education, native language and linguistic region. Longitudinal analyses were additionally adjusted for the respective baseline value.Results: In the cross-sectional analysis, aerobic fitness was associated with better attention (r = 0.16, p = 0.03). A shorter time in the agility test was independently associated with a better performance both in working memory (r = -0.17, p = 0.01) and in attention (r = -0.20, p = 0.01). In the longitudinal analyses, baseline aerobic fitness was independently related to improvements in attention (r = 0.16, p = 0.03), while baseline dynamic balance was associated with improvements in working memory (r = 0.15, p = 0.04).Conclusions: In young children, higher baseline aerobic fitness and motor skills were related to a better spatial working memory and/or attention at baseline, and to some extent also to their future improvements over the following 9 months.

Postgraduate Symposium: Positive influence of nutritional alkalinity on bone health.

There is growing evidence that consumption of a Western diet is a risk factor for osteoporosis through excess acid supply, while fruits and vegetables balance the excess acidity, mostly by providing K-rich bicarbonate-rich foods. Western diets consumed by adults generate approximately 50-100 mEq acid/d; therefore, healthy adults consuming such a diet are at risk of chronic low-grade metabolic acidosis, which worsens with age as a result of declining kidney function. Bone buffers the excess acid by delivering cations and it is considered that with time an overstimulation of this process will lead to the dissolution of the bone mineral content and hence to reduced bone mass. Intakes of K, Mg and fruit and vegetables have been associated with a higher alkaline status and a subsequent beneficial effect on bone health. In healthy male volunteers an acid-forming diet increases urinary Ca excretion by 74% and urinary C-terminal telopeptide of type I collagen (C-telopeptide) excretion by 19% when compared with an alkali (base-forming) diet. Cross-sectional studies have shown that there is a correlation between the nutritional acid load and bone health measured by bone ultrasound or dual-energy X-ray absorptiometry. Few studies have been undertaken in very elderly women (>75 years), whose osteoporosis risk is very pertinent. The EVAluation of Nutrients Intakes and Bone Ultra Sound Study has developed and validated (n 51) an FFQ for use in a very elderly Swiss population (mean age 80.4 (sd 2.99) years), which has shown intakes of key nutrients (energy, fat, carbohydrate, Ca, Mg, vitamin C, D and E) to be low in 401 subjects. A subsequent study to assess net endogenous acid production (NEAP) and bone ultrasound results in 256 women aged > or = 75 years has shown that lower NEAP (P=0.023) and higher K intake (P=0.033) are correlated with higher bone ultrasound results. High acid load may be an important additional risk factor that may be particularly relevant in very elderly patients with an already-high fracture risk. The latter study adds to knowledge by confirming a positive link between dietary alkalinity and bone health indices in the very elderly. In a further study to complement these findings it has also been shown in a group of thirty young women that in Ca sufficiency an acid Ca-rich water has no effect on bone resorption, while an alkaline bicarbonate-rich water leads to a decrease in both serum parathyroid hormone and serum C-telopeptide. Further investigations need to be undertaken to study whether these positive effects on bone loss are maintained over long-term treatment. Mineral-water consumption could be an easy and inexpensive way of helping to prevent osteoporosis and could be of major interest for long-term prevention of bone loss.

Deubiquitylation regulates activation and proteolytic cleavage of ENaC

The epithelial sodium channel (ENaC) is critical for sodium and BP homeostasis. ENaC is regulated by Nedd4-2-mediated ubiquitylation, which leads to its internalization; this process can be reversed by deubiquitylation, which is regulated by the aldosterone-induced enzyme Usp2-45. In a second regulatory pathway, ENaC can be activated by luminal serine protease-mediated cleavage of its extracellular loops. Whether these two regulatory processes interact, however, is unknown. Here, in HEK293 cells stably transfected with ENaC, Usp2-45 interacted with ENaC, leading to deubiquitylation of the channel and stimulation of ENaC activity >20-fold. This was accompanied by a modest increase in cell surface expression of ENaC and by proteolytic cleavage of alphaENaC and gammaENaC at their extracellular loops. When endocytosis was inhibited with dominant negative dynamin (DynK44R), channel density and gammaENaC cleavage were increased, but alphaENaC cleavage and ENaC activity were not augmented. When Usp2-45 was coexpressed with DynK44R, both alphaENaC cleavage and activity were recovered. In summary, these data suggest that Usp2-45 deubiquitylation of ENaC enhances the proteolytic activation of both alphaENaC and gammaENaC, possibly by inducing a conformational change and by interfering with endocytosis, respectively