Interplay between wheat cultivars, biocontrol pseudomonads, and soil.

There is a significant potential to improve the plant-beneficial effects of root-colonizing pseudomonads by breeding wheat genotypes with a greater capacity to sustain interactions with these bacteria. However, the interaction between pseudomonads and crop plants at the cultivar level, as well as the conditions which favor the accumulation of beneficial microorganisms in the wheat rhizosphere, is largely unknown. Therefore, we characterized the three Swiss winter wheat (Triticum aestivum) cultivars Arina, Zinal, and Cimetta for their ability to accumulate naturally occurring plant-beneficial pseudomonads in the rhizosphere. Cultivar performance was measured also by the ability to select for specific genotypes of 2,4-diacetylphloroglucinol (DAPG) producers in two different soils. Cultivar-specific differences were found; however, these were strongly influenced by the soil type. Denaturing gradient gel electrophoresis (DGGE) analysis of fragments of the DAPG biosynthetic gene phlD amplified from natural Pseudomonas rhizosphere populations revealed that phlD diversity substantially varied between the two soils and that there was a cultivar-specific accumulation of certain phlD genotypes in one soil but not in the other. Furthermore, the three cultivars were tested for their ability to benefit from Pseudomonas inoculants. Interestingly, Arina, which was best protected against Pythium ultimum infection by inoculation with Pseudomonas fluorescens biocontrol strain CHA0, was the cultivar which profited the least from the bacterial inoculant in terms of plant growth promotion in the absence of the pathogen. Knowledge gained of the interactions between wheat cultivars, beneficial pseudomonads, and soil types allows us to optimize cultivar-soil combinations for the promotion of growth through beneficial pseudomonads. Additionally, this information can be implemented by breeders into a new and unique breeding strategy for low-input and organic conditions.

Characterization of high osmolarity-induced vacuole fragmentation in "Saccharomyces c."

La majorité des organelles d'une cellule adaptent leur nombre et leur taille pendant les processus de division cellulaire, de trafic vésiculaire ou suite à des changements environnementaux par des processus de fusion et de fragmentation membranaires. Ceci est valable notamment pour le golgi, les mitochondries, les péroxisomes et les lysosomes. La vacuole est le compartiment terminal de la voie endocytaire dans la levure Saccharomyces cerevisiae\ elle correspond aux lysosomes des cellules mammifères. Suite à un choc hyperosmotique, la vacuole se fragmente en plusieurs petites vésicules. Durant ce projet, cette fragmentation a été étudiée en utilisant la technique de microscopie confocale in vivo. J'ai observé que la division de la vacuole se produit d'une façon asymétrique. La première minute après le choc osmotique, les vacuoles rétrécissent et forment des longues invaginations tubulaires. Cette phase est dépendante de la protéine Vps1, un membre de la famille des protéines apparentées à la dynamine, ainsi que d'un gradient transmembranaire de protons. Pendant les 10-15 minutes qui suivent, des vésicules se détachent dans les régions où l'on observe les invaginations pendant la phase initiale. Cette deuxième phase qui mène à la fission des nouveaux compartiments vacuolaires dépend de la production du lipide PI(3,5)P2 par la protéine Fab1. J'ai établi la suite des événements du processus de fragmentation des vacuoles et propose la possibilité d'un rôle régulateur de la protéine kinase cycline-dépendante Pho85.¦En outre, j'ai tenté d'éclaircir plus spécifiquement le rôle de Vps1 pendant la fusion et fission des vacuoles. J'ai trouvé que tous les deux processus sont dépendants de l'activité GTPase de cette protéine. De plus l'association avec la membrane vacuolaire paraît régulée par le cycle d'hydrolyse du GTP. Vps1 peut lier la membrane sans la présence d'un autre facteur protéinique, ce qui permet de conclure à une interaction directe avec des lipides de la membrane. Cette interaction est au moins partiellement effectuée par le domaine GTPase, ce qui est une nouveauté pour un membre de cette famille de protéines. Une deuxième partie de Vps1, nommée insert B, est impliquée dans la liaison à la vacuole, soit par interaction directe avec la membrane, soit par régulation du domaine GTPase. En assumant que Vps1 détienne deux régions capables de liaison aux membranes, je conclus qu'elle pourrait fonctionner comme facteur de « tethering » lors de la fusion des vacuoles.¦-¦La cellule contient plusieurs sous-unités, appelées organelles, possédant chacune une fonction spécifique. Dépendant des processus qui s'y déroulent à l'intérieur, un environnement chimique spécifique est requis. Pour maintenir ces différentes conditions, les organelles sont séparées par des membranes. Lors de la division cellulaire ou en adaptation à des changements de milieu, les organelles doivent être capables de modifier leur morphologie. Cette adaptation a souvent lieu par fusion ou division des organelles. Le même principe est valable pour la vacuole dans la levure. La vacuole est une organelle qui sert principalement au stockage des aliments et à la dégradation des différents composants cellulaires. Alors que la fusion des vacuoles est un processus déjà bien décrit, la fragmentation des vacuoles a jusqu'ici été peu étudiée. Elle peut être induit par un choc osmotique: à cause de la concentration de sel élevé dans le milieu, le cytosol de la levure perd de l'eau. Par un flux d'eau de la vacuole au cytosol, la cellule est capable d'équilibrer celui-ci. Quand la vacuole perd du volume, elle doit réadapter le rapport entre surface membranaire et volume, ce qui se fait efficacement par une fragmentation d'une grande vacuole en plusieurs petites vésicules. Comment ce processus se déroule d'un point de vue morphologique n'a pas été décrit jusqu'à présent. En analysant la fragmentation vacuolaire par microscopie, j'ai trouvé que celle-ci se déroule en deux phases. Pendant la première minute suivant le choc osmotique, les vacuoles rétrécissent et forment des longues invaginations tubulaires. Cette phase dépend de la protéine Vps1, un membre de la famille des protéines apparentées à la dynamine, ainsi que du gradient transmembranaire de protons. Ce gradient s'établit par une pompe membranaire, la V-ATPase, qui transporte des protons dans la vacuole en utilisant l'énergie libérée par hydrolyse d'ATP. Après cette phase initiale, la formation de nouvelles vésicules vacuolaires dépend de la synthèse du lipide PI(3,5)P2.¦Dans la deuxième partie de l'étude, j'ai tenté de décrire comment Vps1 lie la membrane pour effectuer un remodelage de la vacuole. Vps1 est nécessaire pour la fusion et la fragmentation des vacuoles. J'ai découvert que tous les deux processus dépendent de sa capacité d'hydrolyser du GTP. Ainsi l'association avec la membrane est couplée au cycle d'hydrolyse du GTP. Vps1 peut lier la membrane sans la présence d'une autre protéine, et interagit donc très probablement avec les lipides de la membrane. Deux parties différentes de la protéine sont impliquées dans la liaison, dont une, inattendue, le domaine GTPase.¦-¦Numerous organelles undergo membrane fission and fusion events during cell division, vesicular traffic, or in response to changes in environmental conditions. Examples include Golgi (Acharya et al., 1998) mitochondria (Bleazard et al., 1999) peroxisomes (Kuravi et al., 2006) and lysosomes (Ward et al., 1997). In the yeast Saccharomyces cerevisiae the vacuole is the terminal component of the endocytic pathway and corresponds to lysosomes in mammalian cells. Yeast vacuoles fragment into multiple small vesicles in response to a hypertonic shock. This rapid and homogeneous reaction can serve as a model to study the requirements of the fragmentation process. Here, I investigated osmotically induced fragmentation by time-lapse microscopy. I observe that the small fragmentation products originate directly from the large central vacuole by asymmetric scission rather than by consecutive equal divisions and that fragmentation occurs in two distinct phases. During the first minute, vacuoles shrink and generate deep invaginations, leaving behind tubular structures. This phase requires the dynamin-like GTPase Vps1 and the vacuolar proton gradient. In the subsequent 10-15 minutes, vesicles pinch off from the tubular structures in a polarized fashion, directly generating fragmentation products of the final size. This phase depends on the production of phosphatidylinositol- 3,5-bisphosphate by the Fab1 complex. I suggest a possible regulation of vacuole fragmentation by the CDK Pho85. Based on my microscopy study I established a sequential involvement of the different fission factors.¦In addition to the morphological description of vacuole fragmentation I more specifically aimed to shed some light on the role of Vps1 in vacuole fragmentation and fusion. I find that both functions are dependent on the GTPase activity of the protein and that also the membrane association of the dynamin-like protein is coupled to the GTPase cycle. I found that Vps1 has the capacity for direct lipid binding on the vacuole and that this lipid binding is at least partially mediated through residues in the GTPase domain, a complete novelty for a dynamin family member. A second stretch located in the region of insert Β has also membrane-binding activity or regulates the association with the vacuole through the GTPase domain. Under the assumption of two membrane-binding regions I speculate on Vps1 as a possible tethering factor for vacuole fusion.

Digit somatotopy in the human cerebellum: a 7T fMRI study.

The representation of the human body in the human cerebellum is still relatively unknown, compared to the well-studied homunculus in the primary somatosensory cortex. The investigation of the body representation in the cerebellum and its somatotopic organisation is complicated because of the relatively small dimensions of the cerebellum, compared to the cerebrum. Somatotopically organised whole-body homunculi have previously been reported in both humans and rats. However, whether individual digits are represented in the cerebellum in a somatotopically organised way is much less clear. In this study, the high spatial resolution and high sensitivity to the blood oxygenation level dependent (BOLD) signal of 7T fMRI were employed to study the BOLD responses in the human cerebellum to the stroking of the skin of individual digits, the hand and forearm. For the first time, a coarse somatotopic organisation of the digits, ordered from D1-D5, could be visualised in individual human subjects in both the anterior (lobule V) and the posterior (lobule VIII) lobes of the cerebellum using a somatosensory stimulus. The somatotopic gradient in lobule V was found consistently in the posterior to anterior direction, with the thumb most posterior, while the direction of the somatotopic gradient in lobule VIII differed between subjects. No somatotopic organisation was found in Crus I. A comparison of the digit patches with the hand patch revealed that the digit regions are completely covered by the hand region in both the anterior and posterior lobes of the cerebellum, in a non-somatotopic manner. These results demonstrate the promise of ultra-high field, high-resolution fMRI for studies of the cerebellum.

Rare earth elements and stable isotope geochemistry (δ13C and δ18O) of phosphorite deposits in the Gafsa Basin, Tunisia

Rare earth elements (REE) and stable isotope compositions (delta C-13 and delta O-18) of shark teeth and phosphatic coprolites were analyzed from the Lower Maastrichtian layers of the El Haria Formation and two sequences of the Paleocene-Eocene (P/E) Chouabine Formation in the Gafsa Basin (south western of Tunisia) in order to trace the sedimentological, climatic and oceanographic conditions. The REE chemistry and their distribution in the two archives are the same for each of the studied layers indicating that the coprolites and shark teeth experienced the same early diagenetic environments. However major differences occur between the Maastrichtian and the P/E reflecting changes in the depositional conditions. The Early Maastrichtian burial environment tended to be more anoxic with REE derived from reduced FeO. While in the P/E the REE patterns mimic the modern oxic-suboxic seawater, the REE source from remineralisation of organic coating could have more significance. The oxygen isotope compositions of the structural phosphates (delta O-18(PO4)) indicate a stable and warm climate during both studied time intervals. A small offset (-0.4 parts per thousand) in the delta O-18 value between the coprolites and shark teeth show minor thermal gradient between bottom and surface water. The pronounced negative shift of 34%. in delta C-13 values recorded in the upper part of the Chouabine Formation was ascribed to the Paleocene-Eocene boundary. At the same time the lack of negative change in the delta O-18 is explained by the semi-closed situation of the Gafsa Basin, which situation also played an important role in the evolution of the organic matters in the sediment resulting in the exceptional low delta C-13 values. (C) 2008 Elsevier B.V. All rights reserved.

Exogenous selection shapes germination behaviour and seedling traits of populations at different altitudes in a Senecio hybrid zone.

BACKGROUND AND AIMS: The Senecio hybrid zone on Mt Etna, Sicily, is characterized by steep altitudinal clines in quantitative traits and genetic variation. Such clines are thought to be maintained by a combination of 'endogenous' selection arising from genetic incompatibilities and environment-dependent 'exogenous' selection leading to local adaptation. Here, the hypothesis was tested that local adaptation to the altitudinal temperature gradient contributes to maintaining divergence between the parental species, S. chrysanthemifolius and S. aethnensis. METHODS: Intra- and inter-population crosses were performed between five populations from across the hybrid zone and the germination and early seedling growth of the progeny were assessed. KEY RESULTS: Seedlings from higher-altitude populations germinated better under low temperatures (9-13 °C) than those from lower altitude populations. Seedlings from higher-altitude populations had lower survival rates under warm conditions (25/15 °C) than those from lower altitude populations, but also attained greater biomass. There was no altitudinal variation in growth or survival under cold conditions (15/5 °C). Population-level plasticity increased with altitude. Germination, growth and survival of natural hybrids and experimentally generated F(1)s generally exceeded the worse-performing parent. CONCLUSIONS: Limited evidence was found for endogenous selection against hybrids but relatively clear evidence was found for divergence in seed and seedling traits, which is probably adaptive. The combination of low-temperature germination and faster growth in warm conditions might enable high-altitude S. aethnensis to maximize its growth during a shorter growing season, while the slower growth of S. chrysanthemifolius may be an adaptation to drought stress at low altitudes. This study indicates that temperature gradients are likely to be an important environmental factor generating and maintaining adaptive divergence across the Senecio hybrid zone on Mt Etna.

Therapeutic Drug Monitoring of the new targeted anticancer agents imatinib, nilotinib, dasatinib, sunitinib, sorafenib and lapatinib by LC tandem mass spectrometry.

The treatment of some cancer patients has shifted from traditional, non-specific cytotoxic chemotherapy to chronic treatment with molecular targeted therapies. Imatinib mesylate, a selective inhibitor of tyrosine kinases (TKIs) is the most prominent example of this new era and has opened the way to the development of several additional TKIs, including sunitinib, nilotinib, dasatinib, sorafenib and lapatinib, in the treatment of various hematological malignancies and solid tumors. All these agents are characterized by an important inter-individual pharmacokinetic variability, are at risk for drug interactions, and are not devoid of toxicity. Additionally, they are administered for prolonged periods, anticipating the careful monitoring of their plasma exposure via Therapeutic Drug Monitoring (TDM) to be an important component of patients' follow-up. We have developed a liquid chromatography-tandem mass spectrometry method (LC-MS/MS) requiring 100 microL of plasma for the simultaneous determination of the six major TKIs currently in use. Plasma is purified by protein precipitation and the supernatant is diluted in ammonium formate 20 mM (pH 4.0) 1:2. Reverse-phase chromatographic separation of TKIs is obtained using a gradient elution of 20 mM ammonium formate pH 2.2 and acetonitrile containing 1% formic acid, followed by rinsing and re-equilibration to the initial solvent composition up to 20 min. Analyte quantification, using matrix-matched calibration samples, is performed by electro-spray ionization-triple quadrupole mass spectrometry by selected reaction monitoring detection using the positive mode. The method was validated according to FDA recommendations, including assessment of extraction yield, matrix effects variability (<9.6%), overall process efficiency (87.1-104.2%), as well as TKIs short- and long-term stability in plasma. The method is precise (inter-day CV%: 1.3-9.4%), accurate (-9.2 to +9.9%) and sensitive (lower limits of quantification comprised between 1 and 10 ng/mL). This is the first broad-range LC-MS/MS assay covering the major currently in-use TKIs. It is an improvement over previous methods in terms of convenience (a single extraction procedure for six major TKIs, reducing significantly the analytical time), sensitivity, selectivity and throughput. It may contribute to filling the current knowledge gaps in the pharmacokinetics/pharmacodynamics relationships of the latest TKIs developed after imatinib and better define their therapeutic ranges in different patient populations in order to evaluate whether a systematic TDM-guided dose adjustment of these anticancer drugs could contribute to minimize the risk of major adverse reactions and to increase the probability of efficient, long lasting, therapeutic response.

Melanism, body condition and elevational distribution in the asp viper

Alternative morphotypes can confer important selective advantages in different habitats, whereas they can be penalized in other circumstances. In ectotherms, such as reptiles, the body colour can have direct effects on numerous aspects of their existence, such as thermoregulation or prey-predator interactions. Darker melanic individuals show lower skin reflectance and consequently heat up more rapidly and maintain optimal body temperatures more easily than lighter coloured individuals. As a consequence, melanistic individuals of diurnal species in cool areas may exhibit higher body condition, growth rate, survival and fecundity than lighter coloured individuals. Such advantages of dark coloration may be counterbalanced by a lower crypsis to predators and a decreased foraging efficiency. We investigated, in two montane populations of asp vipers Vipera aspis, the relationship between (1) colour polymorphism and body condition and length and (2) the coloration of individuals and their elevational distribution. We showed significant relationships between (1) the coloration, body condition and sex of individuals; (2) sexes and reproductive state and morph frequency; and (3) colour morphs that were distributed following an elevational gradient. Hence, colour polymorphism plays an important role in the ecology and evolution of the asp viper and is maintained through differential selective pressures.

Early white matter alterations in men predisposed to FXTAS revealed by MT imaging.

Introduction: The Fragile X - associated Tremor Ataxia Syndrome (FXTAS) is a recently described, and under-diagnosed, late onset (≈ 60y) neurodegenerative disorder affecting male carriers of a premutation in the Fragile X Mental Retardation 1 (FMR1) gene. The premutation is an CGG (Cytosine-Guanine-Guanine) expansion (55 to 200 CGG repeats) in the proximal region of the FMR1 gene. Patients with FXTAS primarily present with cerebellar ataxia and intention tremor. Neuroradiological features of FXTAS include prominent white matter disease in the periventricular, subcortical, middle cerebellar peduncles and deep white matter of the cerebellum on T2-weighted or FLAIR MR imaging (Jacquemmont 2007, Loesch 2007, Brunberg 2002, Cohen 2006). We hypothesize that a significant white matter alteration is present in younger individuals many years prior to clinical symptoms and/or the presence of visible lesions on conventional MR sequences and might be detectable by magnetization transfer (MT) imaging. Methods: Eleven asymptomatic premutation carriers (mean age = 55 years) and seven intra-familial controls participated to the study. A standardized neurological examination was performed on all participants and a neuropsychological evaluation was carried out before MR scanning performed on a 3T Siemens Trio. The protocol included a sagittal T1-weighted 3D gradient-echo sequence (MPRAGE, 160 slices, 1 mm^3 isotropic voxels) and a gradient-echo MTI (FA 30, TE 15, matrix size 256*256, pixel size 1*1 mm, 36 slices (thickness 2mm), MT pulse duration 7.68 ms, FA 500, frequency offset 1.5 kHz). MTI was performed by acquiring consecutively two set of images; first with and then without the MT saturation pulse. MT images were coregistered to the T1 acquisition. The MTR for every intracranial voxel was calculated as follows: MTR = (M0 - MS)/M0*100%, creating a MTR map for each subject. As first analysis, the whole white matter (WM) was used to mask the MTR image in order to create an histogram of the MTR distribution in the whole tissue class over the two groups examined. Then, for each subject, we performed a segmentation and parcellation of the brain by means of Freesurfer software, starting from the high resolution T1-weighted anatomical acquisition. Cortical parcellations was used to assign a label to the underlying white matter by the construction of a Voronoi diagram in the WM voxels of the MR volume based on distance to the nearest cortical parcellation label. This procedure allowed us to subdivide the cerebral WM in 78 ROIs according to the cortical parcellation (see example in Fig 1). The cerebellum, by the same procedure, was subdivided in 5 ROIs (2 per each hemisphere and one corresponding to the brainstem). For each subject, we calculated the mean value of MTR within each ROI and averaged over controls and patients. Significant differences between the two groups were tested using a two sample T-test (p<0.01). Results: Neurological examination showed that no patient met the clinical criteria of Fragile X Tremor and Ataxia Syndrome yet. Nonetheless, premutation carriers showed some subtle neurological signs of the disorder. In fact, premutation carriers showed a significant increase of tremor (CRST, T-test p=0.007) and increase of ataxia (ICARS, p=0.004) when compared to controls. The neuropsychological evaluation was normal in both groups. To obtain general characterizations of myelination for each subject and premutation carriers, we first computed the distribution of MTR values across the total white matter volume and averaged for each group. We tested the equality of the two distributions with the non parametric Kolmogorov-Smirnov test and we rejected the null-hypothesis at a p=0.03 (fig. 2). As expected, when comparing the asymptomatic permutation carriers with control subjects, the peak value and peak position of the MTR values within the whole WM were decreased and the width of the distribution curve was increased (p<0.01). These three changes point to an alteration of the global myelin status of the premutation carriers. Subsequently, to analyze the regional myelination and white matter integrity of the same group, we performed a ROI analysis of MTR data. The ROI-based analysis showed a decrease of mean MTR value in premutation carriers compared to controls in bilateral orbito-frontal and inferior frontal WM, entorhinal and cingulum regions and cerebellum (Fig 3). The detection of these differences in these regions failed with other conventional MR techniques. Conclusions: These preliminary data confirm that in premutation carriers, there are indeed alterations in "normal appearing white matter" (NAWM) and these alterations are visible with the MT technique. These results indicate that MT imaging may be a relevant approach to detect both global and local alterations within NAWM in "asymptomatic" carriers of premutations in the Fragile X Mental Retardation 1 (FMR1) gene. The sensitivity of MT in the detection of these alterations might point towards a specific physiopathological mechanism linked to an underlying myelin disorder. ROI-based analyses show that the frontal, parahippocampal and cerebellar regions are already significantly affected before the onset of symptoms. A larger sample will allow us to determine the minimum CGG expansion and age associated with these subclinical white matter alterations.

Pulmonary artery pressure and cardiac function in children and adolescents after rapid ascent to 3,450 m.

High-altitude destinations are visited by increasing numbers of children and adolescents. High-altitude hypoxia triggers pulmonary hypertension that in turn may have adverse effects on cardiac function and may induce life-threatening high-altitude pulmonary edema (HAPE), but there are limited data in this young population. We, therefore, assessed in 118 nonacclimatized healthy children and adolescents (mean ± SD; age: 11 ± 2 yr) the effects of rapid ascent to high altitude on pulmonary artery pressure and right and left ventricular function by echocardiography. Pulmonary artery pressure was estimated by measuring the systolic right ventricular to right atrial pressure gradient. The echocardiography was performed at low altitude and 40 h after rapid ascent to 3,450 m. Pulmonary artery pressure was more than twofold higher at high than at low altitude (35 ± 11 vs. 16 ± 3 mmHg; P < 0.0001), and there existed a wide variability of pulmonary artery pressure at high altitude with an estimated upper 95% limit of 52 mmHg. Moreover, pulmonary artery pressure and its altitude-induced increase were inversely related to age, resulting in an almost twofold larger increase in the 6- to 9- than in the 14- to 16-yr-old participants (24 ± 12 vs. 13 ± 8 mmHg; P = 0.004). Even in children with the most severe altitude-induced pulmonary hypertension, right ventricular systolic function did not decrease, but increased, and none of the children developed HAPE. HAPE appears to be a rare event in this young population after rapid ascent to this altitude at which major tourist destinations are located.

Interest of qualitative and quantitative profiling of endocannabinoids for evaluation of their implication in drug addiction process

Introduction: In the middle of the 90's, the discovery of endogenous ligands for cannabinoid receptors opened a new era in this research field. Amides and esters of arachidonic acid have been identified as these endogenous ligands. Arachidonoylethanolamide (anandamide or AEA) and 2-Arachidonoylglycerol (2-AG) seem to be the most important of these lipid messengers. In addition, virodhamine (VA), noladin ether (2-AGE), and N-arachidonoyl dopamine (NADA) have been shown to bind to CB receptors with varying affinities. During recent years, it has become more evident that the EC system is part of fundamental regulatory mechanisms in many physiological processes such as stress and anxiety responses, depression, anorexia and bulimia, schizophrenia disorders, neuroprotection, Parkinson disease, anti-proliferative effects on cancer cells, drug addiction, and atherosclerosis. Aims: This work presents the problematic of EC analysis and the input of Information Dependant Acquisition based on hybrid triple quadrupole linear ion trap (QqQLIT) system for the profiling of these lipid mediators. Methods: The method was developed on a LC Ultimate 3000 series (Dionex, Sunnyvale, CA, USA) coupled to a QTrap 4000 system (Applied biosystems, Concord, ON, Canada). The ECs were separated on an XTerra C18 MS column (50 × 3.0 mm i.d., 3.5 μm) with a 5 min gradient elution. For confirmatory analysis, an information-dependant acquisition experiment was performed with selected reaction monitoring (SRM) as survey scan and enhanced produced ion (EPI) as dependant scan. Results: The assay was found to be linear in the concentration range of 0.1-5 ng/mL for AEA, 0.3-5 ng/mL for VA, 2-AGE, and NADA and 1-20 ng/mL for 2-AG using 0.5 mL of plasma. Repeatability and intermediate precision were found less than 15% over the tested concentration ranges. Under non-pathophysiological conditions, only AEA and 2-AG were actually detected in plasma with concentration ranges going from 104 to 537 pg/mL and from 2160 to 3990 pg/mL respectively. We have particularly focused our scopes on the evaluation of EC level changes in biological matrices through drug addiction and atherosclerosis processes. We will present preliminary data obtained during pilot study after administration of cannabis on human patients. Conclusion: ECs have been shown to play a key role in regulation of many pathophysiological processes. Medical research in these different fields continues to growth in order to understand and to highlight the predominant role of EC in the CNS and peripheral tissues signalisation. The profiling of these lipids needs to develop rapid, highly sensitive and selective analytical methods.

Imaging and quantifying salt-tracer transport in a riparian groundwater system by means of 3D ERT monitoring

Determining groundwater flow paths of infiltrated river water is necessary for studying biochemical processes in the riparian zone, but their characterization is complicated by strong temporal and spatial heterogeneity. We investigated to what extent repeat 3D surface electrical resistance tomography (ERT) can be used to monitor transport of a salt-tracer plume under close to natural gradient conditions. The aim is to estimate groundwater flow velocities and pathways at a site located within a riparian groundwater system adjacent to the perialpine Thur River in northeastern Switzerland. Our ERT time-lapse images provide constraints on the plume's shape, flow direction, and velocity. These images allow the movement of the plume to be followed for 35 m. Although the hydraulic gradient is only 1.43 parts per thousand, the ERT time-lapse images demonstrate that the plume's center of mass and its front propagate with velocities of 2x10(-4) m/s and 5x10(-4) m/s, respectively. These velocities are compatible with groundwater resistivity monitoring data in two observation wells 5 m from the injection well. Five additional sensors in the 5-30 m distance range did not detect the plume. Comparison of the ERT time-lapse images with a groundwater transport model and time-lapse inversions of synthetic ERT data indicate that the movement of the plume can be described for the first 6 h after injection by a uniform transport model. Subsurface heterogeneity causes a change of the plume's direction and velocity at later times. Our results demonstrate the effectiveness of using time-lapse 3D surface ERT to monitor flow pathways in a challenging perialpine environment over larger scales than is practically possible with crosshole 3D ERT.

Vasopressin-stimulated CFTR Cl- currents are increased in the renal collecting duct cells of a mouse model of Liddle's syndrome.

Liddle's syndrome is a genetic form of hypertension linked to Na(+) retention caused by activating mutations in the COOH terminus of the beta or gamma subunit of the epithelial sodium channel (ENaC). In this study, we used the short-circuit current (I(sc)) method to investigate the effects of deamino-8-d-arginine vasopressin (dDAVP) on Na(+) and Cl(-) fluxes in primary cultures of cortical collecting ducts (CCDs) microdissected from the kidneys of mice with Liddle's syndrome carrying a stop codon mutation, corresponding to the beta-ENaC R(566) stop mutation (L) found in the original pedigree. Compared to wild-type (+/+) CCD cells, untreated L/+ and L/L CCD cells exhibited 2.7- and 4.2-fold increases, respectively, in amiloride-sensitive (Ams) I(sc), reflecting ENaC-dependent Na(+) absorption. Short-term incubation with dDAVP caused a rapid and significant increase (approximately 2-fold) in Ams I(sc) in +/+, but not in L/+ or L/L CCD cells. In sharp contrast, dDAVP induced a greater increase in 5-nitro-2-(3-phenylpropamino)benzoate (NPPB)-inhibited apical Cl(-) currents in amiloride-treated L/L and L/+ cells than in their +/+ counterparts. I(sc) recordings performed under apical ion substituted conditions revealed that the dDAVP-stimulated apical secretion of Cl(-), which was absent in cultured CCDs lacking CFTR, was 1.8-fold greater in L/+ and 3.7-fold greater in L/L CCD cells than in their +/+ CCD counterparts. After the basal membrane had been permeabilized with nystatin and a basal-to-apical Cl(-) gradient had been imposed, dDAVP also stimulated larger Cl(-) currents across L/L and L/+ CCD layers than +/+ CCD layers. These findings demonstrate that vasopressin stimulates greater apical CFTR Cl(-) conductance in the renal CCD cells of mice with Liddle's syndrome than in wild-type mice. This effect could contribute to the enhanced NaCl reabsorption observed in the distal nephron of patients with Liddle's syndrome.

Socioeconomic gradients in mortality in the oldest old: a review.

This review aims at identifying gaps in knowledge on socioeconomic gradients in mortality in the oldest old. The authors review literature on oldest old population with a focus on unanswered questions: do socioeconomic status (SES) gradients in mortality persist after 80; does the magnitude of the gradient change as compared with younger populations; which socioeconomic/socio-demographic determinants should be used in this population with specific characteristics (e.g., with respect to sex ratio and household type)? Results are often inconsistent while conclusions drawn by selected studies are generally limited by the difficulty of disentangling the effects of age and cohort, and of generalizing results observed in preponderantly small, selected samples (which typically exclude institutionalized persons). Future research should explore the effects of socio-demographic indicators other than education and social class (e.g., marital status, loss of the partner) and adequately differentiate the social position of oldest old women. The authors recommend that research applies a life-course perspective combined with an interdisciplinary perspective to improve our understanding of the SES gradients in later life. Research is needed to elucidate which causal pathways depending on SES in younger age impact on mortality in higher ages up to oldest old.

Soil fungal communities of grasslands are environmentally structured at a regional scale in the Alps.

Studying patterns of species distributions along elevation gradients is frequently used to identify the primary factors that determine the distribution, diversity and assembly of species. However, despite their crucial role in ecosystem functioning, our understanding of the distribution of below-ground fungi is still limited, calling for more comprehensive studies of fungal biogeography along environmental gradients at various scales (from regional to global). Here, we investigated the richness of taxa of soil fungi and their phylogenetic diversity across a wide range of grassland types along a 2800 m elevation gradient at a large number of sites (213), stratified across a region of the Western Swiss Alps (700 km(2)). We used 454 pyrosequencing to obtain fungal sequences that were clustered into operational taxonomic units (OTUs). The OTU diversity-area relationship revealed uneven distribution of fungal taxa across the study area (i.e. not all taxa are everywhere) and fine-scale spatial clustering. Fungal richness and phylogenetic diversity were found to be higher in lower temperatures and higher moisture conditions. Climatic and soil characteristics as well as plant community composition were related to OTU alpha, beta and phylogenetic diversity, with distinct fungal lineages suggesting distinct ecological tolerances. Soil fungi, thus, show lineage-specific biogeographic patterns, even at a regional scale, and follow environmental determinism, mediated by interactions with plants.

Quantification of nicotine, cotinine, trans-3'-hydroxycotinine and varenicline in human plasma by a sensitive and specific UPLC-tandem mass-spectrometry procedure for a clinical study on smoking cessation.

A sensitive and specific ultra performance liquid chromatography-tandem mass spectrometry method for the simultaneous quantification of nicotine, its metabolites cotinine and trans-3'-hydroxycotinine and varenicline in human plasma was developed and validated. Sample preparation was realized by solid phase extraction of the target compounds and of the internal standards (nicotine-d4, cotinine-d3, trans-3'-hydroxycotinine-d3 and CP-533,633, a structural analog of varenicline) from 0.5mL of plasma, using a mixed-mode cation exchange support. Chromatographic separations were performed on a hydrophilic interaction liquid chromatography column (HILIC BEH 2.1×100mm, 1.7μm). A gradient program was used, with a 10mM ammonium formate buffer pH 3/acetonitrile mobile phase at a flow of 0.4mL/min. The compounds were detected on a triple quadrupole mass spectrometer, operated with an electrospray interface in positive ionization mode and quantification was performed using multiple reaction monitoring. Matrix effects were quantitatively evaluated with success, with coefficients of variation inferior to 8%. The procedure was fully validated according to Food and Drug Administration guidelines and to Société Française des Sciences et Techniques Pharmaceutiques. The concentration range was 2-500ng/mL for nicotine, 1-1000ng/mL for cotinine, 2-1000ng/mL for trans-3'-hydroxycotinine and 1-500ng/mL for varenicline, according to levels usually measured in plasma. Trueness (86.2-113.6%), repeatability (1.9-12.3%) and intermediate precision (4.4-15.9%) were found to be satisfactory, as well as stability in plasma. The procedure was successfully used to quantify nicotine, its metabolites and varenicline in more than 400 plasma samples from participants in a clinical study on smoking cessation.