Glucagon-like peptide-1 increases beta-cell glucose competence and proliferation by translational induction of insulin-like growth factor-1 receptor expression.

Glucagon-like peptide-1 (GLP-1) protects beta-cells against apoptosis, increases their glucose competence, and induces their proliferation. We previously demonstrated that the anti-apoptotic effect was mediated by an increase in insulin-like growth factor-1 receptor (IGF-1R) expression and signaling, which was dependent on autocrine secretion of insulin-like growth factor 2 (IGF-2). Here, we further investigated how GLP-1 induces IGF-1R expression and whether the IGF-2/IGF-1R autocrine loop is also involved in mediating GLP-1-increase in glucose competence and proliferation. We show that GLP-1 up-regulated IGF-1R expression by a protein kinase A-dependent translational control mechanism, whereas isobutylmethylxanthine, which led to higher intracellular accumulation of cAMP than GLP-1, increased both IGF-1R transcription and translation. We then demonstrated, using MIN6 cells and primary islets, that the glucose competence of these cells was dependent on the level of IGF-1R expression and on IGF-2 secretion. We showed that GLP-1-induced primary beta-cell proliferation was suppressed by Igf-1r gene inactivation and by IGF-2 immunoneutralization or knockdown. Together our data show that regulation of beta-cell number and function by GLP-1 depends on the cAMP/protein kinase A mediated-induction of IGF-1R expression and the increased activity of an IGF-2/IGF-1R autocrine loop.

Intracellular trafficking of interleukin-1 receptor I requires Tollip.

Interleukin-1 receptor (IL-1RI) is a master regulator of inflammation and innate immunity. When triggered by IL-1beta, IL-1RI aggregates with IL-1R-associated protein (IL-1RAcP) and forms a membrane proximal signalosome that potently activates downstream signaling cascades. IL-1beta also rapidly triggers endocytosis of IL-1RI. Although internalization of IL-1RI significantly impacts signaling, very little is known about trafficking of IL-1RI and therefore about precisely how endocytosis modulates the overall cellular response to IL-1beta. Upon internalization, activated receptors are often sorted through endosomes and delivered to lysosomes for degradation. This is a highly regulated process that requires ubiquitination of cargo proteins as well as protein-sorting complexes that specifically recognize ubiquitinated cargo. Here, we show that IL-1beta induces ubiquitination of IL-1RI and that via these attached ubiquitin groups, IL-1RI interacts with the ubiquitin-binding protein Tollip. By using an assay to follow trafficking of IL-1RI from the cell surface to late endosomes and lysosomes, we demonstrate that Tollip is required for sorting of IL-1RI at late endosomes. In Tollip-deficient cells and cells expressing only mutated Tollip (incapable of binding IL-1RI and ubiquitin), IL-1RI accumulates on late endosomes and is not efficiently degraded. Furthermore, we show that IL-1RI interacts with Tom1, an ubiquitin-, clathrin-, and Tollip-binding protein, and that Tom1 knockdown also results in the accumulation of IL-1RI at late endosomes. Our findings suggest that Tollip functions as an endosomal adaptor linking IL-1RI, via Tom1, to the endosomal degradation machinery.

Translation and assembly of HLA-DR antigens in Xenopus oocytes injected with mRNA from a human B-cell line.

HLA-DR antigens are polymorphic cell surface glycoproteins, expressed primarily in B lymphocytes and macrophages, which are thought to play an important role in the immune response. Two polypeptide chains, alpha and beta, are associated at the cell surface, and a third chain associates with alpha and beta intracellularly. RNA isolated from the human B-cell line Raji was injected in Xenopus laevis oocytes. Immunoprecipitates of translation products with several monoclonal antibodies revealed the presence of HLA-DR antigens similar to those synthesized in Raji cells. One monoclonal antibody was able to bind the beta chain after dissociation of the three polypeptide chains with detergent. The presence of all three chains was confirmed by two-dimensional gel electrophoresis. The glycosylation pattern of the three chains was identical to that observed in vivo, as evidenced in studies using tunicamycin, an inhibitor of N-linked glycosylation. The presence of alpha chains assembled with beta chains in equimolar ratio was further demonstrated by amino-terminal sequencing. An RNA fraction enriched for the three mRNAs, encoding alpha, beta, and intracellular chains, was isolated. This translation-assembly system and the availability of monoclonal antibodies make it possible to assay for mRNA encoding specific molecules among the multiple human Ia-like antigens.

Cryo-negative staining: advantages and applications for three-dimensional electron microscopy of biological macromolecules

Les échantillons biologiques ne s?arrangent pas toujours en objets ordonnés (cristaux 2D ou hélices) nécessaires pour la microscopie électronique ni en cristaux 3D parfaitement ordonnés pour la cristallographie rayons X alors que de nombreux spécimens sont tout simplement trop << gros D pour la spectroscopie NMR. C?est pour ces raisons que l?analyse de particules isolées par la cryo-microscopie électronique est devenue une technique de plus en plus importante pour déterminer la structure de macromolécules. Néanmoins, le faible rapport signal-sur-bruit ainsi que la forte sensibilité des échantillons biologiques natifs face au faisceau électronique restent deux parmi les facteurs limitant la résolution. La cryo-coloration négative est une technique récemment développée permettant l?observation des échantillons biologiques avec le microscope électronique. Ils sont observés à l?état vitrifié et à basse température, en présence d?un colorant (molybdate d?ammonium). Les avantages de la cryo-coloration négative sont étudiés dans ce travail. Les résultats obtenus révèlent que les problèmes majeurs peuvent êtres évités par l?utilisation de cette nouvelle technique. Les échantillons sont représentés fidèlement avec un SNR 10 fois plus important que dans le cas des échantillons dans l?eau. De plus, la comparaison de données obtenues après de multiples expositions montre que les dégâts liés au faisceau électronique sont réduits considérablement. D?autre part, les résultats exposés mettent en évidence que la technique est idéale pour l?analyse à haute résolution de macromolécules biologiques. La solution vitrifiée de molybdate d?ammonium entourant l?échantillon n?empêche pas l?accès à la structure interne de la protéine. Finalement, plusieurs exemples d?application démontrent les avantages de cette technique nouvellement développée.<br/><br/>Many biological specimens do not arrange themselves in ordered assemblies (tubular or flat 2D crystals) suitable for electron crystallography, nor in perfectly ordered 3D crystals for X-ray diffraction; many other are simply too large to be approached by NMR spectroscopy. Therefore, single-particles analysis has become a progressively more important technique for structural determination of large isolated macromolecules by cryo-electron microscopy. Nevertheless, the low signal-to-noise ratio and the high electron-beam sensitivity of biological samples remain two main resolution-limiting factors, when the specimens are observed in their native state. Cryo-negative staining is a recently developed technique that allows the study of biological samples with the electron microscope. The samples are observed at low temperature, in the vitrified state, but in presence of a stain (ammonium molybdate). In the present work, the advantages of this novel technique are investigated: it is shown that cryo-negative staining can generally overcome most of the problems encountered with cryo-electron microscopy of vitrified native suspension of biological particles. The specimens are faithfully represented with a 10-times higher SNR than in the case of unstained samples. Beam-damage is found to be considerably reduced by comparison of multiple-exposure series of both stained and unstained samples. The present report also demonstrates that cryo-negative staining is capable of high- resolution analysis of biological macromolecules. The vitrified stain solution surrounding the sample does not forbid the access to the interna1 features (ie. the secondary structure) of a protein. This finding is of direct interest for the structural biologist trying to combine electron microscopy and X-ray data. developed electron microscopy technique. Finally, several application examples demonstrate the advantages of this newly

Mutations in NDUFB11, Encoding a Complex I Component of the Mitochondrial Respiratory Chain, Cause Microphthalmia with Linear Skin Defects Syndrome.

Microphthalmia with linear skin defects (MLS) syndrome is an X-linked male-lethal disorder also known as MIDAS (microphthalmia, dermal aplasia, and sclerocornea). Additional clinical features include neurological and cardiac abnormalities. MLS syndrome is genetically heterogeneous given that heterozygous mutations in HCCS or COX7B have been identified in MLS-affected females. Both genes encode proteins involved in the structure and function of complexes III and IV, which form the terminal segment of the mitochondrial respiratory chain (MRC). However, not all individuals with MLS syndrome carry a mutation in either HCCS or COX7B. The majority of MLS-affected females have severe skewing of X chromosome inactivation, suggesting that mutations in HCCS, COX7B, and other as-yet-unidentified X-linked gene(s) cause selective loss of cells in which the mutated X chromosome is active. By applying whole-exome sequencing and filtering for X-chromosomal variants, we identified a de novo nonsense mutation in NDUFB11 (Xp11.23) in one female individual and a heterozygous 1-bp deletion in a second individual, her asymptomatic mother, and an affected aborted fetus of the subject's mother. NDUFB11 encodes one of 30 poorly characterized supernumerary subunits of NADH:ubiquinone oxidoreductase, known as complex I (cI), the first and largest enzyme of the MRC. By shRNA-mediated NDUFB11 knockdown in HeLa cells, we demonstrate that NDUFB11 is essential for cI assembly and activity as well as cell growth and survival. These results demonstrate that X-linked genetic defects leading to the complete inactivation of complex I, III, or IV underlie MLS syndrome. Our data reveal an unexpected role of cI dysfunction in a developmental phenotype, further underscoring the existence of a group of mitochondrial diseases associated with neurocutaneous manifestations.

Patterns of practice of regional nodal irradiation in breast cancer: results of the European Organization for Research and Treatment of Cancer (EORTC) NOdal Radiotherapy (NORA) survey.

BACKGROUND: Predicting outcome of breast cancer (BC) patients based on sentinel lymph node (SLN) status without axillary lymph node dissection (ALND) is an area of uncertainty. It influences the decision-making for regional nodal irradiation (RNI). The aim of the NORA (NOdal RAdiotherapy) survey was to examine the patterns of RNI. METHODS: A web-questionnaire, including several clinical scenarios, was distributed to 88 EORTC-affiliated centers. Responses were received between July 2013 and January 2014. RESULTS: A total of 84 responses were analyzed. While three-dimensional (3D) radiotherapy (RT) planning is carried out in 81 (96%) centers, nodal areas are delineated in only 51 (61%) centers. Only 14 (17%) centers routinely link internal mammary chain (IMC) and supraclavicular node (SCN) RT indications. In patients undergoing total mastectomy (TM) with ALND, SCN-RT is recommend by 5 (6%), 53 (63%) and 51 (61%) centers for patients with pN0(i+), pN(mi) and pN1, respectively. Extra-capsular extension (ECE) is the main factor influencing decision-making RNI after breast conserving surgery (BCS) and TM. After primary systemic therapy (PST), 49 (58%) centers take into account nodal fibrotic changes in ypN0 patients for RNI indications. In ypN0 patients with inner/central tumors, 23 (27%) centers indicate SCN-RT and IMC-RT. In ypN1 patients, SCN-RT is delivered by less than half of the centers in patients with ypN(i+) and ypN(mi). Twenty-one (25%) of the centers recommend ALN-RT in patients with ypN(mi) or 1-2N+ after ALND. Seventy-five (90%) centers state that age is not considered a limiting factor for RNI. CONCLUSION: The NORA survey is unique in evaluating the impact of SLNB/ALND status on adjuvant RNI decision-making and volumes after BCS/TM with or without PST. ALN-RT is often indicated in pN1 patients, particularly in the case of ECE. Besides the ongoing NSABP-B51/RTOG and ALLIANCE trials, NORA could help to design future specific RNI trials in the SLNB era without ALND in patients receiving or not PST.

Nonenhanced arterial spin labeled carotid MR angiography using three-dimensional radial balanced steady-state free precession imaging.

PURPOSE: To optimize and preliminarily evaluate a three-dimensional (3D) radial balanced steady-state free precession (bSSFP) arterial spin labeled (ASL) sequence for nonenhanced MR angiography (MRA) of the extracranial carotid arteries. MATERIALS AND METHODS: The carotid arteries of 13 healthy subjects and 2 patients were imaged on a 1.5 Tesla MRI system using an undersampled 3D radial bSSFP sequence providing a scan time of ∼4 min and 1 mm(3) isotropic resolution. A hybridized scheme that combined pseudocontinuous and pulsed ASL was used to maximize arterial coverage. The impact of a post label delay period, the sequence repetition time, and radiofrequency (RF) energy configuration of pseudocontinuous labeling on the display of the carotid arteries was assessed with contrast-to-noise ratio (CNR) measurements. Faster, higher undersampled 2 and 1 min scans were tested. RESULTS: Using hybridized ASL MRA and a 3D radial bSSFP trajectory, arterial CNR was maximized with a post label delay of 0.2 s, repetition times ≥ 2.5 s (P < 0.05), and by eliminating RF energy during the pseudocontinuous control phase (P < 0.001). With higher levels of undersampling, the carotid arteries were displayed in ≤ 2 min. CONCLUSION: Nonenhanced MRA using hybridized ASL with a 3D radial bSSFP trajectory can display long lengths of the carotid arteries with 1 mm(3) isotropic resolution. J. Magn. Reson. Imaging 2015;41:1150-1156. © 2014 Wiley Periodicals, Inc.

Self-navigated isotropic three-dimensional cardiac T2 mapping.

PURPOSE: To implement and characterize an isotropic three-dimensional cardiac T2 mapping technique. METHODS: A self-navigated three-dimensional radial segmented balanced steady-state free precession pulse sequence with an isotropic 1.7-mm spatial resolution was implemented at 3T with a variable T2 preparation module. Bloch equation and Monte Carlo simulations were performed to determine the influence of the heart rate, B1 inhomogeneity and noise on the T2 fitting accuracy. In a phantom study, the accuracy of the pulse sequence was studied through comparison with a gold-standard spin-echo T2 mapping method. The robustness and homogeneity of the technique were ascertained in a study of 10 healthy adult human volunteers, while first results obtained in patients are reported. RESULTS: The numerical simulations demonstrated that the heart rate and B1 inhomogeneity cause only minor deviations in the T2 fitting, whereas the phantom study showed good agreement of the technique with the gold standard. The volunteer study demonstrated an average myocardial T2 of 40.5 ± 3.3 ms and a <15% T2 gradient in the base-apex and anterior-inferior direction. In three patients, elevated T2 values were measured in regions with expected edema. CONCLUSION: This respiratory self-navigated isotropic three-dimensional technique allows for accurate and robust in vitro and in vivo T2 quantification. Magn Reson Med 73:1549-1554, 2015. © 2014 Wiley Periodicals, Inc.

Two-dimensional probabilistic inversion of plane-wave electromagnetic data: Methodology, model constraints and joint inversion with electrical resistivity data

Probabilistic inversion methods based on Markov chain Monte Carlo (MCMC) simulation are well suited to quantify parameter and model uncertainty of nonlinear inverse problems. Yet, application of such methods to CPU-intensive forward models can be a daunting task, particularly if the parameter space is high dimensional. Here, we present a 2-D pixel-based MCMC inversion of plane-wave electromagnetic (EM) data. Using synthetic data, we investigate how model parameter uncertainty depends on model structure constraints using different norms of the likelihood function and the model constraints, and study the added benefits of joint inversion of EM and electrical resistivity tomography (ERT) data. Our results demonstrate that model structure constraints are necessary to stabilize the MCMC inversion results of a highly discretized model. These constraints decrease model parameter uncertainty and facilitate model interpretation. A drawback is that these constraints may lead to posterior distributions that do not fully include the true underlying model, because some of its features exhibit a low sensitivity to the EM data, and hence are difficult to resolve. This problem can be partly mitigated if the plane-wave EM data is augmented with ERT observations. The hierarchical Bayesian inverse formulation introduced and used herein is able to successfully recover the probabilistic properties of the measurement data errors and a model regularization weight. Application of the proposed inversion methodology to field data from an aquifer demonstrates that the posterior mean model realization is very similar to that derived from a deterministic inversion with similar model constraints.

Three-dimensional hydrodynamic numerical modelling of fold nappe formation, basement-cover deformation and slab detachment with applications to the helvetic nappe system (W Switzerland)

Many three-dimensional (3-D) structures in rock, which formed during the deformation of the Earth's crust and lithosphere, are controlled by a difference in mechanical strength between rock units and are often the result of a geometrical instability. Such structures are, for example, folds, pinch-and-swell structures (due to necking) or cuspate-lobate structures (mullions). These struc-tures occur from the centimeter to the kilometer scale and the related deformation processes con-trol the formation of, for example, fold-and-thrust belts and extensional sedimentary basins or the deformation of the basement-cover interface. The 2-D deformation processes causing these structures are relatively well studied, however, several processes during large-strain 3-D defor-mation are still incompletely understood. One of these 3-D processes is the lateral propagation of these structures, such as fold and cusp propagation in a direction orthogonal to the shortening direction or neck propagation in direction orthogonal to the extension direction. Especially, we are interested in fold nappes which are recumbent folds with amplitudes usually exceeding 10 km and they have been presumably formed by ductile shearing. They often exhibit a constant sense of shearing and a non-linear increase of shear strain towards their overturned limb. The fold axes of the Morcles fold nappe in western Switzerland plunges to the ENE whereas the fold axes in the more eastern Doldenhorn nappe plunges to the WSW. These opposite plunge direc-tions characterize the Rawil depression (Wildstrubel depression). The Morcles nappe is mainly the result of layer parallel contraction and shearing. During the compression the massive lime-stones were more competent than the surrounding marls and shales, which led to the buckling characteristics of the Morcles nappe, especially in the north-dipping normal limb. The Dolden-horn nappe exhibits only a minor overturned fold limb. There are still no 3-D numerical studies which investigate the fundamental dynamics of the formation of the large-scale 3-D structure including the Morcles and Doldenhorn nappes and the related Rawil depression. We study the 3-D evolution of geometrical instabilities and fold nappe formation with numerical simulations based on the finite element method (FEM). Simulating geometrical instabilities caused by sharp variations of mechanical strength between rock units requires a numerical algorithm that can accurately resolve material interfaces for large differences in material properties (e.g. between limestone and shale) and for large deformations. Therefore, our FE algorithm combines a nu-merical contour-line technique and a deformable Lagrangian mesh with re-meshing. With this combined method it is possible to accurately follow the initial material contours with the FE mesh and to accurately resolve the geometrical instabilities. The algorithm can simulate 3-D de-formation for a visco-elastic rheology. The viscous rheology is described by a power-law flow law. The code is used to study the 3-D fold nappe formation, the lateral propagation of folding and also the lateral propagation of cusps due to initial half graben geometry. Thereby, the small initial geometrical perturbations for folding and necking are exactly followed by the FE mesh, whereas the initial large perturbation describing a half graben is defined by a contour line inter-secting the finite elements. Further, the 3-D algorithm is applied to 3-D viscous nacking during slab detachment. The results from various simulations are compared with 2-D resulats and a 1-D analytical solution. -- On retrouve beaucoup de structures en 3 dimensions (3-D) dans les roches qui ont pour origines une déformation de la lithosphère terrestre. Ces structures sont par exemple des plis, des boudins (pinch-and-swell) ou des mullions (cuspate-lobate) et sont présentés de l'échelle centimétrique à kilométrique. Mécaniquement, ces structures peuvent être expliquées par une différence de résistance entre les différentes unités de roches et sont généralement le fruit d'une instabilité géométrique. Ces différences mécaniques entre les unités contrôlent non seulement les types de structures rencontrées, mais également le type de déformation (thick skin, thin skin) et le style tectonique (bassin d'avant pays, chaîne d'avant pays). Les processus de la déformation en deux dimensions (2-D) formant ces structures sont relativement bien compris. Cependant, lorsque l'on ajoute la troisiéme dimension, plusieurs processus ne sont pas complètement compris lors de la déformation à large échelle. L'un de ces processus est la propagation latérale des structures, par exemple la propagation de plis ou de mullions dans la direction perpendiculaire à l'axe de com-pression, ou la propagation des zones d'amincissement des boudins perpendiculairement à la direction d'extension. Nous sommes particulièrement intéressés les nappes de plis qui sont des nappes de charriage en forme de plis couché d'une amplitude plurikilométrique et étant formées par cisaillement ductile. La plupart du temps, elles exposent un sens de cisaillement constant et une augmentation non linéaire de la déformation vers la base du flanc inverse. Un exemple connu de nappes de plis est le domaine Helvétique dans les Alpes de l'ouest. Une de ces nap-pes est la Nappe de Morcles dont l'axe de pli plonge E-NE tandis que de l'autre côté de la dépression du Rawil (ou dépression du Wildstrubel), la nappe du Doldenhorn (équivalent de la nappe de Morcles) possède un axe de pli plongeant O-SO. La forme particulière de ces nappes est due à l'alternance de couches calcaires mécaniquement résistantes et de couches mécanique-ment faibles constituées de schistes et de marnes. Ces différences mécaniques dans les couches permettent d'expliquer les plissements internes à la nappe, particulièrement dans le flanc inver-se de la nappe de Morcles. Il faut également noter que le développement du flanc inverse des nappes n'est pas le même des deux côtés de la dépression de Rawil. Ainsi la nappe de Morcles possède un important flanc inverse alors que la nappe du Doldenhorn en est presque dépour-vue. A l'heure actuelle, aucune étude numérique en 3-D n'a été menée afin de comprendre la dynamique fondamentale de la formation des nappes de Morcles et du Doldenhorn ainsi que la formation de la dépression de Rawil. Ce travail propose la première analyse de l'évolution 3-D des instabilités géométriques et de la formation des nappes de plis en utilisant des simulations numériques. Notre modèle est basé sur la méthode des éléments finis (FEM) qui permet de ré-soudre avec précision les interfaces entre deux matériaux ayant des propriétés mécaniques très différentes (par exemple entre les couches calcaires et les couches marneuses). De plus nous utilisons un maillage lagrangien déformable avec une fonction de re-meshing (production d'un nouveau maillage). Grâce à cette méthode combinée il nous est possible de suivre avec précisi-on les interfaces matérielles et de résoudre avec précision les instabilités géométriques lors de la déformation de matériaux visco-élastiques décrit par une rhéologie non linéaire (n>1). Nous uti-lisons cet algorithme afin de comprendre la formation des nappes de plis, la propagation latérale du plissement ainsi que la propagation latérale des structures de type mullions causé par une va-riation latérale de la géométrie (p.ex graben). De plus l'algorithme est utilisé pour comprendre la dynamique 3-D de l'amincissement visqueux et de la rupture de la plaque descendante en zone de subduction. Les résultats obtenus sont comparés à des modèles 2-D et à la solution analytique 1-D. -- Viele drei dimensionale (3-D) Strukturen, die in Gesteinen vorkommen und durch die Verfor-mung der Erdkruste und Litosphäre entstanden sind werden von den unterschiedlichen mechani-schen Eigenschaften der Gesteinseinheiten kontrolliert und sind häufig das Resulat von geome-trischen Istabilitäten. Zu diesen strukturen zählen zum Beispiel Falten, Pich-and-swell Struktu-ren oder sogenannte Cusbate-Lobate Strukturen (auch Mullions). Diese Strukturen kommen in verschiedenen Grössenordungen vor und können Masse von einigen Zentimeter bis zu einigen Kilometer aufweisen. Die mit der Entstehung dieser Strukturen verbundenen Prozesse kontrol-lieren die Entstehung von Gerbirgen und Sediment-Becken sowie die Verformung des Kontaktes zwischen Grundgebirge und Stedimenten. Die zwei dimensionalen (2-D) Verformungs-Prozesse die zu den genannten Strukturen führen sind bereits sehr gut untersucht. Einige Prozesse wäh-rend starker 3-D Verformung sind hingegen noch unvollständig verstanden. Einer dieser 3-D Prozesse ist die seitliche Fortpflanzung der beschriebenen Strukturen, so wie die seitliche Fort-pflanzung von Falten und Cusbate-Lobate Strukturen senkrecht zur Verkürzungsrichtung und die seitliche Fortpflanzung von Pinch-and-Swell Strukturen othogonal zur Streckungsrichtung. Insbesondere interessieren wir uns für Faltendecken, liegende Falten mit Amplituden von mehr als 10 km. Faltendecken entstehen vermutlich durch duktile Verscherung. Sie zeigen oft einen konstanten Scherungssinn und eine nicht-lineare zunahme der Scherverformung am überkipp-ten Schenkel. Die Faltenachsen der Morcles Decke in der Westschweiz fallen Richtung ONO während die Faltenachsen der östicher gelegenen Doldenhorn Decke gegen WSW einfallen. Diese entgegengesetzten Einfallrichtungen charakterisieren die Rawil Depression (Wildstrubel Depression). Die Morcles Decke ist überwiegend das Resultat von Verkürzung und Scherung parallel zu den Sedimentlagen. Während der Verkürzung verhielt sich der massive Kalkstein kompetenter als der Umliegende Mergel und Schiefer, was zur Verfaltetung Morcles Decke führ-te, vorallem in gegen Norden eifallenden überkippten Schenkel. Die Doldenhorn Decke weist dagegen einen viel kleineren überkippten Schenkel und eine stärkere Lokalisierung der Verfor-mung auf. Bis heute gibt es keine 3-D numerischen Studien, die die fundamentale Dynamik der Entstehung von grossen stark verformten 3-D Strukturen wie den Morcles und Doldenhorn Decken sowie der damit verbudenen Rawil Depression untersuchen. Wir betrachten die 3-D Ent-wicklung von geometrischen Instabilitäten sowie die Entstehung fon Faltendecken mit Hilfe von numerischen Simulationen basiert auf der Finite Elemente Methode (FEM). Die Simulation von geometrischen Instabilitäten, die aufgrund von Änderungen der Materialeigenschaften zwischen verschiedenen Gesteinseinheiten entstehen, erfortert einen numerischen Algorithmus, der in der Lage ist die Materialgrenzen mit starkem Kontrast der Materialeigenschaften (zum Beispiel zwi-schen Kalksteineinheiten und Mergel) für starke Verfomung genau aufzulösen. Um dem gerecht zu werden kombiniert unser FE Algorithmus eine numerische Contour-Linien-Technik und ein deformierbares Lagranges Netz mit Re-meshing. Mit dieser kombinierten Methode ist es mög-lich den anfänglichen Materialgrenzen mit dem FE Netz genau zu folgen und die geometrischen Instabilitäten genügend aufzulösen. Der Algorithmus ist in der Lage visko-elastische 3-D Ver-formung zu rechnen, wobei die viskose Rheologie mit Hilfe eines power-law Fliessgesetzes beschrieben wird. Mit dem numerischen Algorithmus untersuchen wir die Entstehung von 3-D Faltendecken, die seitliche Fortpflanzung der Faltung sowie der Cusbate-Lobate Strukturen die sich durch die Verkürzung eines mit Sediment gefüllten Halbgraben bilden. Dabei werden die anfänglichen geometrischen Instabilitäten der Faltung exakt mit dem FE Netz aufgelöst wäh-rend die Materialgranzen des Halbgrabens die Finiten Elemente durchschneidet. Desweiteren wird der 3-D Algorithmus auf die Einschnürung während der 3-D viskosen Plattenablösung und Subduktion angewandt. Die 3-D Resultate werden mit 2-D Ergebnissen und einer 1-D analyti-schen Lösung verglichen.

Characterization of hepatic fatty acids in mice with reduced liver fat by ultra-short echo time (1)H-MRS at 14.1 T in vivo.

Alterations in the hepatic lipid content (HLC) and fatty acid composition are associated with disruptions in whole body metabolism, both in humans and in rodent models, and can be non-invasively assessed by (1)H-MRS in vivo. We used (1)H-MRS to characterize the hepatic fatty-acyl chains of healthy mice and to follow changes caused by streptozotocin (STZ) injection. Using STEAM at 14.1 T with an ultra-short TE of 2.8 ms, confounding effects from T2 relaxation and J-coupling were avoided, allowing for accurate estimations of the contribution of unsaturated (UFA), saturated (SFA), mono-unsaturated (MUFA) and poly-unsaturated (PUFA) fatty-acyl chains, number of double bonds, PU bonds and mean chain length. Compared with in vivo (1) H-MRS, high resolution NMR performed in vitro in hepatic lipid extracts reported longer fatty-acyl chains (18 versus 15 carbons) with a lower contribution from UFA (61 ± 1% versus 80 ± 5%) but a higher number of PU bonds per UFA (1.39 ± 0.03 versus 0.58 ± 0.08), driven by the presence of membrane species in the extracts. STZ injection caused a decrease of HLC (from 1.7 ± 0.3% to 0.7 ± 0.1%), an increase in the contribution of SFA (from 21 ± 2% to 45 ± 6%) and a reduction of the mean length (from 15 to 13 carbons) of cytosolic fatty-acyl chains. In addition, SFAs were also likely to have increased in membrane lipids of STZ-induced diabetic mice, along with a decrease of the mean chain length. These studies show the applicability of (1)H-MRS in vivo to monitor changes in the composition of the hepatic fatty-acyl chains in mice even when they exhibit reduced HLC, pointing to the value of this methodology to evaluate lipid-lowering interventions in the scope of metabolic disorders.

Error models in hydrogeology applications

Notre consommation en eau souterraine, en particulier comme eau potable ou pour l'irrigation, a considérablement augmenté au cours des années. De nombreux problèmes font alors leur apparition, allant de la prospection de nouvelles ressources à la remédiation des aquifères pollués. Indépendamment du problème hydrogéologique considéré, le principal défi reste la caractérisation des propriétés du sous-sol. Une approche stochastique est alors nécessaire afin de représenter cette incertitude en considérant de multiples scénarios géologiques et en générant un grand nombre de réalisations géostatistiques. Nous rencontrons alors la principale limitation de ces approches qui est le coût de calcul dû à la simulation des processus d'écoulements complexes pour chacune de ces réalisations. Dans la première partie de la thèse, ce problème est investigué dans le contexte de propagation de l'incertitude, oú un ensemble de réalisations est identifié comme représentant les propriétés du sous-sol. Afin de propager cette incertitude à la quantité d'intérêt tout en limitant le coût de calcul, les méthodes actuelles font appel à des modèles d'écoulement approximés. Cela permet l'identification d'un sous-ensemble de réalisations représentant la variabilité de l'ensemble initial. Le modèle complexe d'écoulement est alors évalué uniquement pour ce sousensemble, et, sur la base de ces réponses complexes, l'inférence est faite. Notre objectif est d'améliorer la performance de cette approche en utilisant toute l'information à disposition. Pour cela, le sous-ensemble de réponses approximées et exactes est utilisé afin de construire un modèle d'erreur, qui sert ensuite à corriger le reste des réponses approximées et prédire la réponse du modèle complexe. Cette méthode permet de maximiser l'utilisation de l'information à disposition sans augmentation perceptible du temps de calcul. La propagation de l'incertitude est alors plus précise et plus robuste. La stratégie explorée dans le premier chapitre consiste à apprendre d'un sous-ensemble de réalisations la relation entre les modèles d'écoulement approximé et complexe. Dans la seconde partie de la thèse, cette méthodologie est formalisée mathématiquement en introduisant un modèle de régression entre les réponses fonctionnelles. Comme ce problème est mal posé, il est nécessaire d'en réduire la dimensionnalité. Dans cette optique, l'innovation du travail présenté provient de l'utilisation de l'analyse en composantes principales fonctionnelles (ACPF), qui non seulement effectue la réduction de dimensionnalités tout en maximisant l'information retenue, mais permet aussi de diagnostiquer la qualité du modèle d'erreur dans cet espace fonctionnel. La méthodologie proposée est appliquée à un problème de pollution par une phase liquide nonaqueuse et les résultats obtenus montrent que le modèle d'erreur permet une forte réduction du temps de calcul tout en estimant correctement l'incertitude. De plus, pour chaque réponse approximée, une prédiction de la réponse complexe est fournie par le modèle d'erreur. Le concept de modèle d'erreur fonctionnel est donc pertinent pour la propagation de l'incertitude, mais aussi pour les problèmes d'inférence bayésienne. Les méthodes de Monte Carlo par chaîne de Markov (MCMC) sont les algorithmes les plus communément utilisés afin de générer des réalisations géostatistiques en accord avec les observations. Cependant, ces méthodes souffrent d'un taux d'acceptation très bas pour les problèmes de grande dimensionnalité, résultant en un grand nombre de simulations d'écoulement gaspillées. Une approche en deux temps, le "MCMC en deux étapes", a été introduite afin d'éviter les simulations du modèle complexe inutiles par une évaluation préliminaire de la réalisation. Dans la troisième partie de la thèse, le modèle d'écoulement approximé couplé à un modèle d'erreur sert d'évaluation préliminaire pour le "MCMC en deux étapes". Nous démontrons une augmentation du taux d'acceptation par un facteur de 1.5 à 3 en comparaison avec une implémentation classique de MCMC. Une question reste sans réponse : comment choisir la taille de l'ensemble d'entrainement et comment identifier les réalisations permettant d'optimiser la construction du modèle d'erreur. Cela requiert une stratégie itérative afin que, à chaque nouvelle simulation d'écoulement, le modèle d'erreur soit amélioré en incorporant les nouvelles informations. Ceci est développé dans la quatrième partie de la thèse, oú cette méthodologie est appliquée à un problème d'intrusion saline dans un aquifère côtier. -- Our consumption of groundwater, in particular as drinking water and for irrigation, has considerably increased over the years and groundwater is becoming an increasingly scarce and endangered resource. Nofadays, we are facing many problems ranging from water prospection to sustainable management and remediation of polluted aquifers. Independently of the hydrogeological problem, the main challenge remains dealing with the incomplete knofledge of the underground properties. Stochastic approaches have been developed to represent this uncertainty by considering multiple geological scenarios and generating a large number of realizations. The main limitation of this approach is the computational cost associated with performing complex of simulations in each realization. In the first part of the thesis, we explore this issue in the context of uncertainty propagation, where an ensemble of geostatistical realizations is identified as representative of the subsurface uncertainty. To propagate this lack of knofledge to the quantity of interest (e.g., the concentration of pollutant in extracted water), it is necessary to evaluate the of response of each realization. Due to computational constraints, state-of-the-art methods make use of approximate of simulation, to identify a subset of realizations that represents the variability of the ensemble. The complex and computationally heavy of model is then run for this subset based on which inference is made. Our objective is to increase the performance of this approach by using all of the available information and not solely the subset of exact responses. Two error models are proposed to correct the approximate responses follofing a machine learning approach. For the subset identified by a classical approach (here the distance kernel method) both the approximate and the exact responses are knofn. This information is used to construct an error model and correct the ensemble of approximate responses to predict the "expected" responses of the exact model. The proposed methodology makes use of all the available information without perceptible additional computational costs and leads to an increase in accuracy and robustness of the uncertainty propagation. The strategy explored in the first chapter consists in learning from a subset of realizations the relationship between proxy and exact curves. In the second part of this thesis, the strategy is formalized in a rigorous mathematical framework by defining a regression model between functions. As this problem is ill-posed, it is necessary to reduce its dimensionality. The novelty of the work comes from the use of functional principal component analysis (FPCA), which not only performs the dimensionality reduction while maximizing the retained information, but also allofs a diagnostic of the quality of the error model in the functional space. The proposed methodology is applied to a pollution problem by a non-aqueous phase-liquid. The error model allofs a strong reduction of the computational cost while providing a good estimate of the uncertainty. The individual correction of the proxy response by the error model leads to an excellent prediction of the exact response, opening the door to many applications. The concept of functional error model is useful not only in the context of uncertainty propagation, but also, and maybe even more so, to perform Bayesian inference. Monte Carlo Markov Chain (MCMC) algorithms are the most common choice to ensure that the generated realizations are sampled in accordance with the observations. Hofever, this approach suffers from lof acceptance rate in high dimensional problems, resulting in a large number of wasted of simulations. This led to the introduction of two-stage MCMC, where the computational cost is decreased by avoiding unnecessary simulation of the exact of thanks to a preliminary evaluation of the proposal. In the third part of the thesis, a proxy is coupled to an error model to provide an approximate response for the two-stage MCMC set-up. We demonstrate an increase in acceptance rate by a factor three with respect to one-stage MCMC results. An open question remains: hof do we choose the size of the learning set and identify the realizations to optimize the construction of the error model. This requires devising an iterative strategy to construct the error model, such that, as new of simulations are performed, the error model is iteratively improved by incorporating the new information. This is discussed in the fourth part of the thesis, in which we apply this methodology to a problem of saline intrusion in a coastal aquifer.

Mechanisms of OGT-mediated HCF-1 protein maturation

Post-translational protein modifications are crucial for many fundamental cellular and extracellular processes and greatly contribute to the complexity of organisms. Human HCF-1 is a transcriptional co-regulator that undergoes complex protein maturation involving reversible and irreversible post-translational modifications. Upon synthesis as a large precursor protein, HCF-1 undergoes extensive reversible glycosylation with β-N-acetylglucosamine giving rise to O-linked-β-N-acetylglucosamine (O-GlcNAc) modified serines and threonines. HCF-1 also undergoes irreversible site-specific proteolysis, which is important for one of HCF-1's major functions - the regulation of the cell-division cycle. HCF-1 O-GlcNAcylation and site-specific proteolysis are both catalyzed by a single enzyme with an unusual dual enzymatic activity, the O-GlcNAc transferase (OGT). HCF-1 is cleaved by OGT at any of six highly conserved 26 amino acid repeated sequences (HCF-1PRO repeats), but the mechanisms and the substrate requirements for OGT-mediated cleavage are not understood. In the present work, I characterized substrate requirements for OGT-mediated cleavage and O-GlcNAcylation of HCF-1. I identified key elements within the HCF-1PRO-repeat sequence that are important for proteolysis. Remarkably, an invariant single amino acid side-chain within the HCF-1PRO-repeat sequence displays particular OGT-binding properties and is essential for proteolysis. Additionally, I characterized substrate requirements for proteolysis outside of the HCF-1PRO repeat and identified a novel, highly O-GlcNAcylated OGT-binding sequence that enhances cleavage of the first HCF-1PRO repeat. These results link OGT association and its O-GlcNAcylation activities to HCF-1PRO-repeat proteolysis.

Information transfer between large and small two-dimensional polyacrylamide gel electrophoresis.

To determine the feasibility of data transfer, an interlaboratory comparison was conducted on colon carcinoma cell line (DLD-1) proteins resolved by two-dimensional polyacrylamide gel electrophoresis either on small (6 x 7 cm) or large (16x18 cm) gels. The gels were silver-stained and scanned by laser densitometry, and the image obtained was analyzed using Melanie software. The number of spots detected was 1337+/-161 vs. 2382+/-176 for small vs. large format gels, respectively. After gel calibration using landmarks determined using pl and Mr markers, large- and small-format gels were matched and 712+/-36 proteins were found on both types of gels. Having performed accurate gel matching it was possible to acquire additional information after accessing a 2-D PAGE reference database (http://www.expasy.ch/ cgibin/map2/def?DLD1_HUMAN). Thus, the difference in gel size is not an obstacle for data transfer. This will facilitate exchanges between laboratories or consultation concerning existing databases.

Trabecular bone score in type 1 diabetes--a cross-sectional study.

UNLABELLED: Trabecular bone score (TBS) seems to provide additive value on BMD to identify individuals with prevalent fractures in T1D. TBS did not significantly differ between T1D patients and healthy controls, but TBS and HbA1c were independently associated with prevalent fractures in T1D. A TBS cutoff <1.42 reflected prevalent fractures with 91.7 % sensitivity and 43.2 % specificity. INTRODUCTION: Type 1 diabetes (T1D) increases the risk of osteoporotic fractures. TBS was recently proposed as an indirect measure of bone microarchitecture. This study aimed at investigating the TBS in T1D patients and healthy controls. Associations with prevalent fractures were tested. METHODS: One hundred nineteen T1D patients (59 males, 60 premenopausal females; mean age 43.4 ± 8.9 years) and 68 healthy controls matched for gender, age, and body mass index (BMI) were analyzed. The TBS was calculated in the lumbar region, based on two-dimensional (2D) projections of DXA assessments. RESULTS: TBS was 1.357 ± 0.129 in T1D patients and 1.389 ± 0.085 in controls (p = 0.075). T1D patients with prevalent fractures (n = 24) had a significantly lower TBS than T1D patients without fractures (1.309 ± 0.125 versus 1.370 ± 0.127, p = 0.04). The presence of fractures in T1D was associated with lower TBS (odds ratio = 0.024, 95 % confidence interval (CI) = 0.001-0.875; p = 0.042) but not with age or BMI. TBS and HbA1c were independently associated with fractures. The area-under-the curve (AUC) of TBS was similar to that of total hip BMD in discriminating T1D patients with or without prevalent fractures. In this set-up, a TBS cutoff <1.42 discriminated the presence of fractures with a sensitivity of 91.7 % and a specificity of 43.2 %. CONCLUSIONS: TBS values are lower in T1D patients with prevalent fractures, suggesting an alteration of bone strength in this subgroup of patients. Reliable TBS cutoffs for the prediction of fracture risk in T1D need to be determined in larger prospective studies.