Nouveaux traitements de l'hépatite C: aspects pharmacologiques et potentiel d'interaction [New hepatitis C treatments: pharmacological considerations and potential for drug interactions].

Progress in the understanding of the hepatitis C virus life cycle allowed the development of new, very promising antiviral therapies. Although these new drugs have a favourable profile in terms of efficacy, tolerance and interaction potential, their prescription in the setting of comedication and impaired renal or hepatic function remains a challenge. Here, we provide a summary of pharmacological considerations, focusing on sofosbuvir, simeprevir and daclatasvir. A better understanding of their metabolic pathways and transporters may help the prescriber to identify and manage drug interactions especially in patients under immunosuppressive or anti-HIV therapy. Recommendations for the prescription of these drugs in specific situations are also discussed.

Grabbing your ear: rapid auditory-somatosensory multisensory interactions in low-level sensory cortices are not constrained by stimulus alignment.

Multisensory interactions are observed in species from single-cell organisms to humans. Important early work was primarily carried out in the cat superior colliculus and a set of critical parameters for their occurrence were defined. Primary among these were temporal synchrony and spatial alignment of bisensory inputs. Here, we assessed whether spatial alignment was also a critical parameter for the temporally earliest multisensory interactions that are observed in lower-level sensory cortices of the human. While multisensory interactions in humans have been shown behaviorally for spatially disparate stimuli (e.g. the ventriloquist effect), it is not clear if such effects are due to early sensory level integration or later perceptual level processing. In the present study, we used psychophysical and electrophysiological indices to show that auditory-somatosensory interactions in humans occur via the same early sensory mechanism both when stimuli are in and out of spatial register. Subjects more rapidly detected multisensory than unisensory events. At just 50 ms post-stimulus, neural responses to the multisensory 'whole' were greater than the summed responses from the constituent unisensory 'parts'. For all spatial configurations, this effect followed from a modulation of the strength of brain responses, rather than the activation of regions specifically responsive to multisensory pairs. Using the local auto-regressive average source estimation, we localized the initial auditory-somatosensory interactions to auditory association areas contralateral to the side of somatosensory stimulation. Thus, multisensory interactions can occur across wide peripersonal spatial separations remarkably early in sensory processing and in cortical regions traditionally considered unisensory.

Expression analyses of three members of the AtPHO1 family reveal differential interactions between signaling pathways involved in phosphate deficiency and the responses to auxin, cytokinin, and abscisic acid.

The PHO1 protein is involved in loading inorganic phosphate (Pi) to the root xylem. Ten genes homologous to AtPHO1 are present in the Arabidopsis thaliana (L.) Heyn genome. From this gene family, transcript levels of only AtPHO1, AtPHO1;H1 and AtPHO1;H10 were increased by Pi-deficiency. While the up-regulation of AtPHO1;H1 and AtPHO1;H10 by Pi deficiency followed the same rapid kinetics and was dependent on the PHR1 transcription factor, phosphite only strongly suppressed the expression of AtPHO1;H1 and had a minor effect on AtPHO1;H10. Addition of sucrose was found to increase transcript levels of both AtPHO1 and AtPHO1;H1 in Pi-sufficient or Pi-deficient plants, but to suppress AtPHO1:H10 under the same conditions. Treatments of plants with auxin or cytokinin had contrasting effect depending on the gene and on the Pi status of the plants. Thus, while both hormones down-regulated expression of AtPHO1 independently of the plant Pi status, auxin and cytokinin up-regulated AtPHO1;H1 and AtPHO1;H10 expression in Pi-sufficient plants and down-regulated expression in Pi-deficient plants. Treatments with abscisic acid inhibited AtPHO1 and AtPHO1;H1 expression in both Pi-sufficient and Pi-deficient plants, but increased AtPHO1;H10 expression under the same conditions. The inhibition of expression by abscisic acid of AtPHO1 and AtPHO1;H1, and of the Pi-starvation responsive genes AtPHT1;1 and AtIPS1, was dependant on the ABI1 type 2C protein phosphatase. These results reveal that various levels of cross talk between the signal transduction pathways to Pi, sucrose and phytohormones are involved in the regulation of expression of the three AtPHO1 homologues.

Neutrophils in infectious diseases

Host defense to intracellular pathogens depends upon both innate and adaptive cell-mediated immune responses. Polymorphonuclear neutrophil leukocytes which belong to the innate immune system are the first cells that are recruited massively within hours of microbial infection. Neutrophils are the main players in the killing of microorganisms and recently new methods of killing including nets formation have been described. Neutrophils mediate tissue damage at infected sites. By promoting tissue injury neutrophils contribute to the initiation of inflammation, which is now recognized as an essential step in launching immunity. The importance of neutrophils as decision shaper in the development of an immune response is only emerging as they have long been considered by immunologists as short lived, non-dividing cells, of poor interest. Now, neutrophils are emerging as key components of the inflammatory response, and are shown to have immunoregulatory roles in microbial infections. In addition, neutrophils were also reported to contribute to the recruitment and activation of antigen presenting cells. Thus early interactions between neutrophils and surrounding cells may influence the development/resolution of both inflammatory lesion and pathogen-specific immune response. The impact of neutrophils on cells present at the site of infection are only beginning to be studied and deserves more attention.In this e-book the reader will find updated information about the role of neutrophils in the pathogenesis of 1) bacterial diseases including sepsis, mycobacteria and Chlamydia infections, and of 2) parasitic diseases including leishmaniasis and toxoplasmosis. The role of neutrophils in the protection against microorganisms has largely been underestimated and, until recently, their role was mostly thought to limited to a "kill and die" response. New neutrophil mode of killing, such as their release of extracellular traps to kill extracellular bacterial pathogens, together with several microbial strategies designed to escape NETs are presented in Chapter 1. We will emphasize standard and advanced light microscopy techniques that allowed major advances in the understanding of neutrophil biology, through the visualization of the interaction of selected pathogens with neutrophils in living animals (Chapter 2).The aim of this e-book is to provide an overview of the recent advances made in the field of neutrophil biology. It will provide a basis for understanding future development that will occur in this area, and provide the reader with a short overview of some of the exciting new directions in which neutrophil research is moving.

"In vitro" study of the molecular mechanism of the "E.Coli" Hsp 70/Hsp40/NEF chaperone system and its interactions with ?-sinuclein

SUMMARY Under stressful conditions, mutant or post-translationally modified proteins may spontaneously misfold and form toxie species, which may further assemble into a continuum of increasingly large and insoluble toxic oligomers that may further condense into less toxic, compact amyloids in the cell Intracellular accumulation of aggregated proteins is a common denominator of several neurodegenerative diseases. To cope with the cytotoxicity induced by abnormal, aggregated proteins, cells have evolved various defence mechanisms among which, the molecular chaperones Hsp70. Hsp70 (DnaK in E. coii) is an ATPase chaperone involved in many physiological processes in the cell, such as assisting de novo protein folding, dissociating native protein oligomers and serving as pulling motors in the import of polypeptides into organelles. In addition, Hsp70 chaperones can actively solubilize and reactivate stable protein aggregates, such as heat- or mutation-induced aggregates. Hsp70 requires the cooperation of two other co-chaperones: Hsp40 and NEF (Nucleotide exchange factor) to fulfil its unfolding activity. In the first experimental section of this thesis (Chapter II), we studied by biochemical analysis the in vitro interaction between recombinant human aggregated α-synuclein (a-Syn oligomers) mimicking toxic a-Syn oligomers species in PD brains, with a model Hsp70/Hsp40 chaperone system (the E. coii DnaK/DnaJ/GrpE). We found that chaperone-mediated unfolding of two denatured model enzymes were strongly affected by α-Syn oligomers but, remarkably, not by monomers. This in vitro observed dysfunction of the Hsp70 chaperone system resulted from the sequestration of the Hsp40 proteins by the oligomeric α-synuclein species. In the second experimental part (Chapter III), we performed in vitro biochemical analysis of the co-chaperone function of three E. coii Hsp40s proteins (DnaJ, CbpA and DjlA) in the ATP-fuelled DnaK-mediated refolding of a model DnaK chaperone substrate into its native state. Hsp40s activities were compared using dose-response approaches in two types of in vitro assays: refolding of heat-denatured G6PDH and DnaK-mediated ATPase activity. We also observed that the disaggregation efficiency of Hsp70 does not directly correlate with Hsp40 binding affinity. Besides, we found that these E. coii Hsp40s confer substrate specificity to DnaK, CbpA being more effective in the DnaK-mediated disaggregation of large G6PDH aggregates than DnaJ under certain conditions. Sensibilisées par différents stress ou mutations, certaines protéines fonctionnelles de la cellule peuvent spontanément se convertir en formes inactives, mal pliées, enrichies en feuillets bêta, et exposant des surfaces hydrophobes favorisant l'agrégation. Cherchant à se stabiliser, les surfaces hydrophobes peuvent s'associer aux régions hydrophobes d'autres protéines mal pliées, formant des agrégats protéiques stables: les amyloïdes. Le dépôt intracellulaire de protéines agrégées est un dénominateur commun à de nombreuses maladies neurodégénératives. Afin de contrer la cytotoxicité induite par les protéines agrégées, les cellules ont développé plusieurs mécanismes de défense, parmi lesquels, les chaperonnes moléculaires Hsp70. Hsp70 nécessite la collaboration de deux autres co-chaperonnes : Hsp40 et NEF pour accomplir son activité de désagrégation. Hsp70 (DnaK, chez E. coli) est impliquée par ailleurs dans d'autres fonctions physiologiques telles que l'assistanat de protéines néosynthétisées à la sortie du ribosome, ou le transport transmembranaire de polypeptides. Par ailleurs, les chaperonnes Hsp70 peuvent également solubiliser et réactiver des protéines agrégées à la suite d'un stress ou d'une mutation. Dans la première partie expérimentale de cette thèse (Chapter II), nous avons étudié in vitro l'interaction entre les oligomères d'a-synucleine, responsables entre autres, de la maladie de Parkinson, et le système chaperon Hsp70/Hsp40 (système Escherichia coli DnaK/DnaJ/GrpE). Nous avons démontré que contrairement aux monomères, les oligomères d'a-synucleine inhibaient le système chaperon lors du repliement de protéines agrégées. Cette dysfonction du système chaperon résulte de la séquestration des chaperonnes Hsp40 par les oligomères d'a-synucleine. La deuxième partie expérimentale (Chapitre III) est consacrée à une étude in vitro de la fonction co-chaperonne de trois Hsp40 d'is. coli (DnaJ, CbpA, et DjlA) lors de la désagrégation par DnaK d'une protéine pré-agrégée. Leurs activités ont été comparées par le biais d'une approche dose-réponse au niveau de deux analyses enzymatiques: le repliement de la protéine agrégée et l'activité ATPase de DnaK. Par ailleurs, nous avons mis en évidence que l'efficacité de désagrégation d'Hsp70 et l'affinité des chaperonnes Hsp40 vis-à-vis de leur substrat n'étaient pas corrélées positivement. Nous avons également montré que ces trois chaperonnes Hsp40 étaient directement impliquées dans la spécificité des fonctions accomplies par les chaperonnes Hsp70. En effet, DnaK en présence de CbpA assure la désagrégation de large agrégats protéiques avec une efficacité nettement plus accrue qu'en présence de DnaJ.

Cytomegalovirus (CMV)-specific cellular immune responses.

A large percentage of healthy individuals (50-90%) is chronically infected with Cytomegalovirus (CMV). Over the past few years, several techniques were developed in order to monitor CMV-specific T-cell responses. In addition to the identification of antigen-specific T cells with peptide-loaded MHC complexes, most of the current strategies to identify CMV-specific T cells are centered on the assessment of the functions of memory T cells including their ability to mediate effector function, to proliferate or to secrete cytokines following antigen-specific stimulation. The investigation of these functions has allowed the characterization of the CMV-specific T-cell responses that are present during different phases of the infection. Furthermore, it has also been shown that the combination of virus-specific CD4 and CD8 T-cell responses are critical components of the immune response in the control of virus replication.

Histone deacetylase inhibitors impair innate immune responses to Toll-like receptor agonists and to infection.

Regulated by histone acetyltransferases and deacetylases (HDACs), histone acetylation is a key epigenetic mechanism controlling chromatin structure, DNA accessibility, and gene expression. HDAC inhibitors induce growth arrest, differentiation, and apoptosis of tumor cells and are used as anticancer agents. Here we describe the effects of HDAC inhibitors on microbial sensing by macrophages and dendritic cells in vitro and host defenses against infection in vivo. HDAC inhibitors down-regulated the expression of numerous host defense genes, including pattern recognition receptors, kinases, transcription regulators, cytokines, chemokines, growth factors, and costimulatory molecules as assessed by genome-wide microarray analyses or innate immune responses of macrophages and dendritic cells stimulated with Toll-like receptor agonists. HDAC inhibitors induced the expression of Mi-2β and enhanced the DNA-binding activity of the Mi-2/NuRD complex that acts as a transcriptional repressor of macrophage cytokine production. In vivo, HDAC inhibitors increased the susceptibility to bacterial and fungal infections but conferred protection against toxic and septic shock. Thus, these data identify an essential role for HDAC inhibitors in the regulation of the expression of innate immune genes and host defenses against microbial pathogens.

Public health expenditure and spatial interactions in a decentralized national health system

One of the limitations of cross-country health expenditure analysis refers to the fact that the financing, the internal organization and political restraints of health care decision-making are country-specific and heterogeneous. Yet, a potential solution is to examine the influence of such effects in those countries that have undertaken decentralization processes. In such a setting, it is possible to examine potential expenditure spillovers across the geography of a country as well as the influence of the political ideology of regional incumbents on public health expenditure. This paper examines the determinants of public health expenditure within Spanish region-states (Autonomous Communities, ACs), most of them subject to similar financing structures although exhibiting significant heterogeneity as a result of the increasing decentralization, region-specific political factors along with different use of health care inputs, economic dimension and spatial interactions

SBML qualitative models: a model representation format and infrastructure to foster interactions between qualitative modelling formalisms and tools.

BACKGROUND: Qualitative frameworks, especially those based on the logical discrete formalism, are increasingly used to model regulatory and signalling networks. A major advantage of these frameworks is that they do not require precise quantitative data, and that they are well-suited for studies of large networks. While numerous groups have developed specific computational tools that provide original methods to analyse qualitative models, a standard format to exchange qualitative models has been missing. RESULTS: We present the Systems Biology Markup Language (SBML) Qualitative Models Package ("qual"), an extension of the SBML Level 3 standard designed for computer representation of qualitative models of biological networks. We demonstrate the interoperability of models via SBML qual through the analysis of a specific signalling network by three independent software tools. Furthermore, the collective effort to define the SBML qual format paved the way for the development of LogicalModel, an open-source model library, which will facilitate the adoption of the format as well as the collaborative development of algorithms to analyse qualitative models. CONCLUSIONS: SBML qual allows the exchange of qualitative models among a number of complementary software tools. SBML qual has the potential to promote collaborative work on the development of novel computational approaches, as well as on the specification and the analysis of comprehensive qualitative models of regulatory and signalling networks.

Regulation of inflammatory responses by the commensal microbiota.

It is well established that dysregulation of the interactions between the immune system and commensal bacteria is one factor that underpins the development and chronicity of a number of inflammatory diseases. Certain phyla of bacteria within the microbiota have been associated with 'health', but the mechanisms by which the presence of these bacteria supports a healthy environment are still being unravelled. Recent evidence indicates that one such mechanism involves the anti-inflammatory properties of fermentation products of fibre, short-chain fatty acids and their signalling through the G-protein coupled receptor GPR43. Recent findings also indicate that, even in health, bacterial communities harbour in the airways, indicating that direct exposure to bacterial products at this site may provide a further explanation for how commensal bacteria can regulate chronic airway inflammation.

Vaccinia immune globulin: current policies, preparedness, and product safety and efficacy.

In 1980 the World Health Organization declared that smallpox was eradicated from the world, and routine smallpox vaccination was discontinued. Nevertheless, samples of the smallpox virus (variola virus) were retained for research purposes, not least because of fears that terrorist groups or rogue states might also have kept samples in order to develop a bioweapon. Variola virus represents an effective bioweapon because it is associated with high morbidity and mortality and is highly contagious. Since September 11, 2001, countries around the world have begun to develop policies and preparedness programs to deal with a bioterror attack, including stockpiling of smallpox vaccine. Smallpox vaccine itself may be associated with a number of serious adverse events, which can often be managed with vaccinia immune globulin (VIG). VIG may also be needed as prophylaxis in patients for whom pre-exposure smallpox vaccine is contraindicated (such as those with eczema or pregnant women), although it is currently not licensed in these cases. Two intravenous formulations of VIG (VIGIV Cangene and VIGIV Dynport) have been licensed by the FDA for the management of patients with progressive vaccinia, eczema vaccinatum, severe generalized vaccinia, and extensive body surface involvement or periocular implantation following inadvertent inoculation.

Genetic polymorphisms and drug interactions modulating CYP2D6 and CYP3A activities have a major effect on oxycodone analgesic efficacy and safety

Background and purpose: The major drug-metabolizing enzymes for the oxidation of oxycodone are CYP2D6 and CYP3A. A high interindividual variability in the activity of these enzymes because of genetic polymorphisms and/or drug-drug interactions is well established. The possible role of an active metabolite in the pharmacodynamics of oxycodone has been questioned and the importance of CYP3A-mediated effects on the pharmacokinetics and pharmacodynamics of oxycodone has been poorly explored. Experimental approach: We conducted a randomized crossover (five arms) double-blind placebo-controlled study in 10 healthy volunteers genotyped for CYP2D6. Oral oxycodone (0.2 mg·kg−1) was given alone or after inhibition of CYP2D6 (with quinidine) and/or of CYP3A (with ketoconazole). Experimental pain (cold pressor test, electrical stimulation, thermode), pupil size, psychomotor effects and toxicity were assessed. Key results: CYP2D6 activity was correlated with oxycodone experimental pain assessment. CYP2D6 ultra-rapid metabolizers experienced increased pharmacodynamic effects, whereas cold pressor test and pupil size were unchanged in CYP2D6 poor metabolizers, relative to extensive metabolizers. CYP2D6 blockade reduced subjective pain threshold (SPT) for oxycodone by 30% and the response was similar to placebo. CYP3A4 blockade had a major effect on all pharmacodynamic assessments and SPT increased by 15%. Oxymorphone Cmax was correlated with SPT assessment (ρS= 0.7) and the only independent positive predictor of SPT. Side-effects were observed after CYP3A4 blockade and/or in CYP2D6 ultra-rapid metabolizers. Conclusions and implications: The modulation of CYP2D6 and CYP3A activities had clear effects on oxycodone pharmacodynamics and these effects were dependent on CYP2D6 genetic polymorphism.