Comparison between computational alanine scanning and per-residue binding free energy decomposition for protein-protein association using MM-GBSA: application to the TCR-p-MHC complex.

Recognition by the T-cell receptor (TCR) of immunogenic peptides (p) presented by Class I major histocompatibility complexes (MHC) is the key event in the immune response against virus-infected cells or tumor cells. A study of the 2C TCR/SIYR/H-2K(b) system using a computational alanine scanning and a much faster binding free energy decomposition based on the Molecular Mechanics-Generalized Born Surface Area (MM-GBSA) method is presented. The results show that the TCR-p-MHC binding free energy decomposition using this approach and including entropic terms provides a detailed and reliable description of the interactions between the molecules at an atomistic level. Comparison of the decomposition results with experimentally determined activity differences for alanine mutants yields a correlation of 0.67 when the entropy is neglected and 0.72 when the entropy is taken into account. Similarly, comparison of experimental activities with variations in binding free energies determined by computational alanine scanning yields correlations of 0.72 and 0.74 when the entropy is neglected or taken into account, respectively. Some key interactions for the TCR-p-MHC binding are analyzed and some possible side chains replacements are proposed in the context of TCR protein engineering. In addition, a comparison of the two theoretical approaches for estimating the role of each side chain in the complexation is given, and a new ad hoc approach to decompose the vibrational entropy term into atomic contributions, the linear decomposition of the vibrational entropy (LDVE), is introduced. The latter allows the rapid calculation of the entropic contribution of interesting side chains to the binding. This new method is based on the idea that the most important contributions to the vibrational entropy of a molecule originate from residues that contribute most to the vibrational amplitude of the normal modes. The LDVE approach is shown to provide results very similar to those of the exact but highly computationally demanding method.

GLUT4, AMP kinase, but not the insulin receptor, are required for hepatoportal glucose sensor-stimulated muscle glucose utilization.

Recent evidence suggests the existence of a hepatoportal vein glucose sensor, whose activation leads to enhanced glucose use in skeletal muscle, heart, and brown adipose tissue. The mechanism leading to this increase in whole body glucose clearance is not known, but previous data suggest that it is insulin independent. Here, we sought to further determine the portal sensor signaling pathway by selectively evaluating its dependence on muscle GLUT4, insulin receptor, and the evolutionarily conserved sensor of metabolic stress, AMP-activated protein kinase (AMPK). We demonstrate that the increase in muscle glucose use was suppressed in mice lacking the expression of GLUT4 in the organ muscle. In contrast, glucose use was stimulated normally in mice with muscle-specific inactivation of the insulin receptor gene, confirming independence from insulin-signaling pathways. Most importantly, the muscle glucose use in response to activation of the hepatoportal vein glucose sensor was completely dependent on the activity of AMPK, because enhanced hexose disposal was prevented by expression of a dominant negative AMPK in muscle. These data demonstrate that the portal sensor induces glucose use and development of hypoglycemia independently of insulin action, but by a mechanism that requires activation of the AMPK and the presence of GLUT4.

Design of a Control Slide for Cyanoacrylate Polymerization : Application to the CA-Bluestar Sequence

Casework expercience has shown that, in some cases, long exposures of surfaces subjected to cyanoacrylate (CA) fuming had detrimental effects on the subsequent application of Bluestar. This study aimed to develop a control mechanism to monitor the amount of CA deposited prior to the subsequent treatment. A control slide bearing spots of sodium hydroxide (NaOH) of known concentrations and volume was designed and validated against both scanning electron microscopy (SEM) observations and latent print examiners' assessments of the quality of the developed marks. The control slide allows one to define three levels of development that were used to monitor the Bluestar reaction on depleting footwear marks left in diluted blood. The appropriate conditions for a successful application of both CA and Bluestar were determined.

Application de la modélisation moléculaire à de nouvelles approches thérapeutiques du cancer [Application of molecular modeling to new therapeutic cancer approaches].

Recent progress in the experimental determination of protein structures allow to understand, at a very detailed level, the molecular recognition mechanisms that are at the basis of the living matter. This level of understanding makes it possible to design rational therapeutic approaches, in which effectors molecules are adapted or created de novo to perform a given function. An example of such an approach is drug design, were small inhibitory molecules are designed using in silico simulations and tested in vitro. In this article, we present a similar approach to rationally optimize the sequence of killer T lymphocytes receptors to make them more efficient against melanoma cells. The architecture of this translational research project is presented together with its implications both at the level of basic research as well as in the clinics.

Phase Contrast X-Ray Tomographic Microscopy for Biological and Materials Science Applications

The application of two approaches for high-throughput, high-resolution X-ray phase contrast tomographic imaging being used at the tomographic microscopy and coherent radiology experiments (TOMCAT) beamline of the SLS is discussed and illustrated. Differential phase contrast (DPC) imaging, using a grating interferometer and a phase-stepping technique, is integrated into the beamline environment at TOMCAT in terms of the fast acquisition and reconstruction of data and the availability to scan samples within an aqueous environment. A second phase contrast method is a modified transfer of intensity approach that can yield the 3D distribution of the decrement of the refractive index of a weakly absorbing object from a single tomographic dataset. The two methods are complementary to one another: the DPC method is characterised by a higher sensitivity and by moderate resolution with larger samples; the modified transfer of intensity approach is particularly suited for small specimens when high resolution (around 1 mu m) is required. Both are being applied to investigations in the biological and materials science fields.

A viscous bioerodible poly(ortho ester) as a new biomaterial for intraocular application.

The biocompatibility of a viscous, hydrophobic, bioerodible poly(ortho ester) (POE) intended for intraocular application was investigated. POE was evaluated as a blank carrier and as containing modulators of degradation. Each formulation was injected intracamerally and intravitreally in rabbit eyes, and clinical and histological examinations were performed postoperatively for 2 weeks. In the case of intracameral injections, polymer biocompatibility appeared to depend on the amount injected in the anterior chamber. When 50 microL was administered, the polymer degraded within 2 weeks, and clinical observations showed good biocompatibility of POE with no toxicity to the ocular tissues or increase in intraocular pressure. The injection of a larger volume, 100 microL, of POE, appeared inappropriate because of direct contact of polymeric material with the corneal endothelium, and triggered reversible edema and inflammation in the anterior chamber of the eye that regressed after a few days. After intravitreal administration, POE was well tolerated and no inflammatory reaction developed during the observation period. The polymer degraded slowly, appearing as a round whitish bubble in the vitreous cavity. The presence of modulators of degradation both improved POE biocompatibility and prolonged polymer lifetime in the eye. POE appears to be a promising biomaterial for clinical intraocular application.

Améliorations et optimisation globale de la méthode des analogues pour la prévision statistique des précipitations. Développement d'un outil de prévision et application opérationnelle au bassin du Rhône à l'amont du Léman.

Le bassin du Rhône à l'amont du Léman peut être sujet à de fortes précipitations en mesure de provoquer des crues significatives. L'objectif du projet MINERVE dans lequel s'inscrit le présent travail consiste à fournir des outils pour la prévision et la gestion des crues par des actions préventives sur les aménagements hydroélectriques à accumulation. Pour satisfaire ce dernier, il est nécessaire de prévoir au mieux les cumuls de précipitations pour les jours suivants. Ceci est actuellement effectué par le modèle numérique de prévision de MétéoSuisse ; mais, en raison des grandes incertitudes liées à la quantification des événements extrêmes, il a été décidé qu'une approche parallèle de nature statistique pourrait compléter l'information disponible. Ainsi, nous avons adapté la méthode des analogues, qui est une technique de prévision statistique des précipitations, au contexte alpin du bassin d'étude. Pour ce faire, plusieurs paramétrisations de la méthode ont été documentées et calibrées. Afin de prendre en main la méthode, nous avons effectué de multiples analyses paramétriques sur les variables synoptiques, mais également sur la constitution de groupements pluviométriques. Une partie conséquente de cette étude a été consacrée à la programmation d'un logiciel de prévision automatique par la méthode des analogues, ainsi qu'à un outil de visualisation des résultats sous forme de cartes et graphiques. Ce logiciel, nommé Atmoswing, permet d'implémenter un grand nombre de méthodes différentes de prévision par analogie. L'outil est opérationnel depuis mi-2011 et nous a permis de confirmer l'intérêt de la prévision par analogie. La méthode étant ici appliquée à un nouveau contexte, un grand nombre de variables synoptiques ont été évaluées. Nous avons alors confirmé l'intérêt des deux niveaux d'analogie sur la circulation atmosphérique et sur le flux d'humidité, tout en apportant des améliorations à celles-ci. Il en résulte des paramétrisations présentant des scores de performance supérieurs aux méthodes de référence considérées. Nous avons également évalué d'autres améliorations, comme l'introduction d'une fenêtre temporelle glissante afin de rechercher de meilleures analogies synoptiques à d'autres heures de la journée, ce qui s'est avéré intéressant, tout comme une prévision infrajournalière à pas de temps de 6 h. Finalement, nous avons introduit une technique d'optimisation globale, les algorithmes génétiques, capable de calibrer la méthode des analogues en considérant tous les paramètres des différents niveaux d'analogie de manière conjointe. Avec cette technique, nous pouvons nous approcher objectivement d'une paramétrisation optimale. Le choix des niveaux atmosphériques et des fenêtres temporelles et spatiales étant automatisé, cette technique peut engendrer un gain de temps, même si elle est relativement exigeante en calculs. Nous avons ainsi pu améliorer la méthode des analogues, et y ajouter de nouveaux degrés de liberté, notamment des fenêtres spatiales et des pondérations différenciées selon les niveaux atmosphériques retenus.

A deterministic model to assess the impact of AIDS interventions in countries with generalized HIV epidemic. An application to Botswana

General introductionThe Human Immunodeficiency/Acquired Immunodeficiency Syndrome (HIV/AIDS) epidemic, despite recent encouraging announcements by the World Health Organization (WHO) is still today one of the world's major health care challenges.The present work lies in the field of health care management, in particular, we aim to evaluate the behavioural and non-behavioural interventions against HIV/AIDS in developing countries through a deterministic simulation model, both in human and economic terms. We will focus on assessing the effectiveness of the antiretroviral therapies (ART) in heterosexual populations living in lesser developed countries where the epidemic has generalized (formerly defined by the WHO as type II countries). The model is calibrated using Botswana as a case study, however our model can be adapted to other countries with similar transmission dynamics.The first part of this thesis consists of reviewing the main mathematical concepts describing the transmission of infectious agents in general but with a focus on human immunodeficiency virus (HIV) transmission. We also review deterministic models assessing HIV interventions with a focus on models aimed at African countries. This review helps us to recognize the need for a generic model and allows us to define a typical structure of such a generic deterministic model.The second part describes the main feed-back loops underlying the dynamics of HIV transmission. These loops represent the foundation of our model. This part also provides a detailed description of the model, including the various infected and non-infected population groups, the type of sexual relationships, the infection matrices, important factors impacting HIV transmission such as condom use, other sexually transmitted diseases (STD) and male circumcision. We also included in the model a dynamic life expectancy calculator which, to our knowledge, is a unique feature allowing more realistic cost-efficiency calculations. Various intervention scenarios are evaluated using the model, each of them including ART in combination with other interventions, namely: circumcision, campaigns aimed at behavioral change (Abstain, Be faithful or use Condoms also named ABC campaigns), and treatment of other STD. A cost efficiency analysis (CEA) is performed for each scenario. The CEA consists of measuring the cost per disability-adjusted life year (DALY) averted. This part also describes the model calibration and validation, including a sensitivity analysis.The third part reports the results and discusses the model limitations. In particular, we argue that the combination of ART and ABC campaigns and ART and treatment of other STDs are the most cost-efficient interventions through 2020. The main model limitations include modeling the complexity of sexual relationships, omission of international migration and ignoring variability in infectiousness according to the AIDS stage.The fourth part reviews the major contributions of the thesis and discusses model generalizability and flexibility. Finally, we conclude that by selecting the adequate interventions mix, policy makers can significantly reduce the adult prevalence in Botswana in the coming twenty years providing the country and its donors can bear the cost involved.Part I: Context and literature reviewIn this section, after a brief introduction to the general literature we focus in section two on the key mathematical concepts describing the transmission of infectious agents in general with a focus on HIV transmission. Section three provides a description of HIV policy models, with a focus on deterministic models. This leads us in section four to envision the need for a generic deterministic HIV policy model and briefly describe the structure of such a generic model applicable to countries with generalized HIV/AIDS epidemic, also defined as pattern II countries by the WHO.