Protection from HIV-1 infection of primary CD4 T cells by CCR5 silencing is effective for the full spectrum of CCR5 expression.

Stable gene silencing by RNA interference (RNAi) can be achieved by expression of small hairpin RNAs (shRNAs) from RNA polymerase III promoters. We have tested lentiviral vectors expressing shRNAs targetting CCR5 in primary CD4 T cells from donors representing various CCR5 and CCR2 genetic backgrounds covering the full spectrum of CCR5 expression levels and permissiveness for HIV-1 infection. A linear decrease in CCR5 expression resulted in a logarithmic decrease in cellular infection, giving up to three logs protection from HIV-1 infection in vitro. Protection was maintained at very high multiplicity of infection. This and other recent reports on RNAi should open a debate about the use of RNAi gene therapy for HIV infection.

Contribution of efflux pumps, porins, and β-lactamases to multidrug resistance in clinical isolates of Acinetobacter baumannii.

We investigated the mechanisms of resistance to carbapenems, aminoglycosides, glycylcyclines, tetracyclines, and quinolones in 90 multiresistant clinical strains of Acinetobacter baumannii isolated from two genetically unrelated A. baumannii clones: clone PFGE-ROC-1 (53 strains producing the OXA-58 β-lactamase enzyme and 18 strains with the OXA-24 β-lactamase) and clone PFGE-HUI-1 (19 strains susceptible to carbapenems). We used real-time reverse transcriptase PCR to correlate antimicrobial resistance (MICs) with expression of genes encoding chromosomal β-lactamases (AmpC and OXA-51), porins (OmpA, CarO, Omp33, Dcap-like, OprB, Omp25, OprC, OprD, and OmpW), and proteins integral to six efflux systems (AdeABC, AdeIJK, AdeFGH, CraA, AbeM, and AmvA). Overexpression of the AdeABC system (level of expression relative to that by A. baumannii ATCC 17978, 30- to 45-fold) was significantly associated with resistance to tigecycline, minocycline, and gentamicin and other biological functions. However, hyperexpression of the AdeIJK efflux pump (level of expression relative to that by A. baumannii ATCC 17978, 8- to 10-fold) was significantly associated only with resistance to tigecycline and minocycline (to which the TetB efflux system also contributed). TetB and TetA(39) efflux pumps were detected in clinical strains and were associated with resistance to tetracyclines and doxycycline. The absence of the AdeABC system and the lack of expression of other mechanisms suggest that tigecycline-resistant strains of the PFGE-HUI-1 clone may be associated with a novel resistance-nodulation-cell efflux pump (decreased MICs in the presence of the inhibitor Phe-Arg β-naphthylamide dihydrochloride) and the TetA(39) system.

Extended family structure in the ant Formica paralugubris: the role of the breeding system

In populations of various ant species, many queens reproduce in the same nest (polygyny), and colony boundaries appear to be absent with individuals able to move fi eely between nests (unicoloniality). Such societies depart strongly from a simple family structure and pose a potential challenge to kin selection theory, because high queen number coupled with unrestricted gene flow among nests should result in levels of relatedness among nestmates close to zero. This study investigated the breeding system and genetic structure of a highly polygynous and largely unicolonial population of the wood ant Formica paralugubris. A microsatellite analysis revealed that nestmate workers, reproductive queens and reproductive males (the queens' mates) are all equally related to each other, with relatedness estimates centring around 0.14. This suggests that most of the queens and males reproducing in the study population had mated within or close to their natal nest, and that the queens did not disperse far after mating. We developed a theoretical model to investigate how the breeding system affects the relatedness structure of polygynous colonies. By combining the model and our empirical data, it was estimated that about 99.8% of the reproducing queens and males originated from within the nest, or from a nearby nest. This high rate of local mating and the rarity of long-distance dispersal maintain significant relatedness among nestmates, and contrast with the common view that unicoloniality is coupled with unrestricted gene flow among nests.

The required beta cell research for improving treatment of type 2 diabetes.

In healthy individuals, insulin resistance is associated with physiological conditions such as pregnancy or body weight gain and triggers an increase in beta cell number and insulin secretion capacity to preserve normoglycaemia. Failure of this beta cell compensation capacity is a fundamental cause of diabetic hyperglycaemia. Incomplete understanding of the molecular mechanisms controlling the plasticity of adult beta cells mechanisms and how these cells fail during the pathogenesis of diabetes strongly limits the ability to develop new beta cell-specific therapies. Here, current knowledge of the signalling pathways controlling beta cell plasticity is reviewed, and possible directions for future research are discussed.

Diffusion spectrum magnetic resonance imaging (DSI) tractography of crossing fibers.

MRI tractography is the mapping of neural fiber pathways based on diffusion MRI of tissue diffusion anisotropy. Tractography based on diffusion tensor imaging (DTI) cannot directly image multiple fiber orientations within a single voxel. To address this limitation, diffusion spectrum MRI (DSI) and related methods were developed to image complex distributions of intravoxel fiber orientation. Here we demonstrate that tractography based on DSI has the capacity to image crossing fibers in neural tissue. DSI was performed in formalin-fixed brains of adult macaque and in the brains of healthy human subjects. Fiber tract solutions were constructed by a streamline procedure, following directions of maximum diffusion at every point, and analyzed in an interactive visualization environment (TrackVis). We report that DSI tractography accurately shows the known anatomic fiber crossings in optic chiasm, centrum semiovale, and brainstem; fiber intersections in gray matter, including cerebellar folia and the caudate nucleus; and radial fiber architecture in cerebral cortex. In contrast, none of these examples of fiber crossing and complex structure was identified by DTI analysis of the same data sets. These findings indicate that DSI tractography is able to image crossing fibers in neural tissue, an essential step toward non-invasive imaging of connectional neuroanatomy.

Transforming growth factor beta 1 production by CD4+ CD25+ regulatory T cells in peripheral blood mononuclear cells from healthy subjects stimulated with Leishmania guyanensis.

Transforming growth factor beta (TGF-beta) has been shown to be a central immunomodulator used by leishmaniae to escape effective mechanisms of protection in human and murine infections with these parasites. However, all the information is derived from studies of established infection, while little is known about TGF-beta production in response to Leishmania stimulation in healthy subjects. In this study, TGF-beta1 production was demonstrated in peripheral blood mononuclear cells from healthy subjects never exposed to leishmaniae in response to live Leishmania guyanensis, and the TGF-beta1-producing cells were described as a distinct subpopulation of CD4(+) CD25(+) regulatory T cells. The suppressive properties of CD4(+) CD25(+) T cells were demonstrated in vitro by their inhibition of production of interleukin 2 (IL-2) and IL-10 by CD4(+) CD25(-) T cells in the presence of either anti-CD3 or L. guyanensis. Although neutralization of TGF-beta1 did not reverse the suppressive activity of CD4(+) CD25(+) T cells activated by anti-CD3, it reversed the suppressive activity of CD4(+) CD25(+) T cells activated by L. guyanensis. Altogether our data demonstrated that TGF-beta1 is involved in the suppressive activity of L. guyanensis-stimulated CD4(+) CD25(+) T cells from healthy controls.

Role for inducible cAMP early repressor in promoting pancreatic beta cell dysfunction evoked by oxidative stress in human and rat islets.

AIMS/HYPOTHESIS: Pro-atherogenic and pro-oxidant, oxidised LDL trigger adverse effects on pancreatic beta cells, possibly contributing to diabetes progression. Because oxidised LDL diminish the expression of genes regulated by the inducible cAMP early repressor (ICER), we investigated the involvement of this transcription factor and of oxidative stress in beta cell failure elicited by oxidised LDL. METHODS: Isolated human and rat islets, and insulin-secreting cells were cultured with human native or oxidised LDL or with hydrogen peroxide. The expression of genes was determined by quantitative real-time PCR and western blotting. Insulin secretion was monitored by EIA kit. Cell apoptosis was determined by scoring cells displaying pycnotic nuclei. RESULTS: Exposure of beta cell lines and islets to oxidised LDL, but not to native LDL raised the abundance of ICER. Induction of this repressor by the modified LDL compromised the expression of important beta cell genes, including insulin and anti-apoptotic islet brain 1, as well as of genes coding for key components of the secretory machinery. This led to hampering of insulin production and secretion, and of cell survival. Silencing of this transcription factor by RNA interference restored the expression of its target genes and alleviated beta cell dysfunction and death triggered by oxidised LDL. Induction of ICER was stimulated by oxidative stress, whereas antioxidant treatment with N-acetylcysteine or HDL prevented the rise of ICER elicited by oxidised LDL and restored beta cell functions. CONCLUSIONS/INTERPRETATION: Induction of ICER links oxidative stress to beta cell failure caused by oxidised LDL and can be effectively abrogated by antioxidant treatment.

Identification of key amino acids of the mouse mammary tumor virus superantigen involved in the specific interaction with T-cell receptor V(beta) domains.

Mouse mammary tumor virus (MMTV) is a retrovirus encoding a superantigen that is recognized in association with major histocompatibility complex class II by the variable region of the beta chain (V(beta)) of the T-cell receptor. The C-terminal 30 to 40 amino acids of the superantigen of different MMTVs display high sequence variability that correlates with the recognition of particular T-cell receptor V(beta) chains. Interestingly, MMTV(SIM) and mtv-8 superantigens are highly homologous but have nonoverlapping T-cell receptor V(beta) specificities. To determine the importance of these few differences for specific V(beta) interaction, we studied superantigen responses in mice to chimeric and mutant MMTV(SIM) and mtv-8 superantigens expressed by recombinant vaccinia viruses. We show that only a few changes (two to six residues) within the C terminus are necessary to modify superantigen recognition by specific V(beta)s. Thus, the introduction of the MMTV(SIM) residues 314-315 into the mtv-8 superantigen greatly decreased its V(beta)12 reactivity without gain of MMTV(SIM)-specific function. The introduction of MMTV(SIM)-specific residues 289 to 295, however, induced a recognition pattern that was a mixture of MMTV(SIM)- and mtv-8-specific V(beta) reactivities: both weak MMTV(SIM)-specific V(beta)4 and full mtv-8-specific V(beta)11 recognition were observed while V(beta)12 interaction was lost. The combination of the two MMTV(SIM)-specific regions in the mtv-8 superantigen established normal MMTV(SIM)-specific V(beta)4 reactivity and completely abolished mtv-8-specific V(beta)5, -11, and -12 interactions. These new functional superantigens with mixed V(beta) recognition patterns allowed us to precisely delineate sites relevant for molecular interactions between the SIM or mtv-8 superantigen and the T-cell receptor V(beta) domain within the 30 C-terminal residues of the viral superantigen.

Assessment of statistical iterative reconstructions to optimize CT examination protocols for children and young adults

La tomodensitométrie (CT) est une technique d'imagerie dont l'intérêt n'a cessé de croître depuis son apparition dans le début des années 70. Dans le domaine médical, son utilisation est incontournable à tel point que ce système d'imagerie pourrait être amené à devenir victime de son succès si son impact au niveau de l'exposition de la population ne fait pas l'objet d'une attention particulière. Bien évidemment, l'augmentation du nombre d'examens CT a permis d'améliorer la prise en charge des patients ou a rendu certaines procédures moins invasives. Toutefois, pour assurer que le compromis risque - bénéfice soit toujours en faveur du patient, il est nécessaire d'éviter de délivrer des doses non utiles au diagnostic.¦Si cette action est importante chez l'adulte elle doit être une priorité lorsque les examens se font chez l'enfant, en particulier lorsque l'on suit des pathologies qui nécessitent plusieurs examens CT au cours de la vie du patient. En effet, les enfants et jeunes adultes sont plus radiosensibles. De plus, leur espérance de vie étant supérieure à celle de l'adulte, ils présentent un risque accru de développer un cancer radio-induit dont la phase de latence peut être supérieure à vingt ans. Partant du principe que chaque examen radiologique est justifié, il devient dès lors nécessaire d'optimiser les protocoles d'acquisitions pour s'assurer que le patient ne soit pas irradié inutilement. L'avancée technologique au niveau du CT est très rapide et depuis 2009, de nouvelles techniques de reconstructions d'images, dites itératives, ont été introduites afin de réduire la dose et améliorer la qualité d'image.¦Le présent travail a pour objectif de déterminer le potentiel des reconstructions itératives statistiques pour réduire au minimum les doses délivrées lors d'examens CT chez l'enfant et le jeune adulte tout en conservant une qualité d'image permettant le diagnostic, ceci afin de proposer des protocoles optimisés.¦L'optimisation d'un protocole d'examen CT nécessite de pouvoir évaluer la dose délivrée et la qualité d'image utile au diagnostic. Alors que la dose est estimée au moyen d'indices CT (CTDIV0| et DLP), ce travail a la particularité d'utiliser deux approches radicalement différentes pour évaluer la qualité d'image. La première approche dite « physique », se base sur le calcul de métriques physiques (SD, MTF, NPS, etc.) mesurées dans des conditions bien définies, le plus souvent sur fantômes. Bien que cette démarche soit limitée car elle n'intègre pas la perception des radiologues, elle permet de caractériser de manière rapide et simple certaines propriétés d'une image. La seconde approche, dite « clinique », est basée sur l'évaluation de structures anatomiques (critères diagnostiques) présentes sur les images de patients. Des radiologues, impliqués dans l'étape d'évaluation, doivent qualifier la qualité des structures d'un point de vue diagnostique en utilisant une échelle de notation simple. Cette approche, lourde à mettre en place, a l'avantage d'être proche du travail du radiologue et peut être considérée comme méthode de référence.¦Parmi les principaux résultats de ce travail, il a été montré que les algorithmes itératifs statistiques étudiés en clinique (ASIR?, VEO?) ont un important potentiel pour réduire la dose au CT (jusqu'à-90%). Cependant, par leur fonctionnement, ils modifient l'apparence de l'image en entraînant un changement de texture qui pourrait affecter la qualité du diagnostic. En comparant les résultats fournis par les approches « clinique » et « physique », il a été montré que ce changement de texture se traduit par une modification du spectre fréquentiel du bruit dont l'analyse permet d'anticiper ou d'éviter une perte diagnostique. Ce travail montre également que l'intégration de ces nouvelles techniques de reconstruction en clinique ne peut se faire de manière simple sur la base de protocoles utilisant des reconstructions classiques. Les conclusions de ce travail ainsi que les outils développés pourront également guider de futures études dans le domaine de la qualité d'image, comme par exemple, l'analyse de textures ou la modélisation d'observateurs pour le CT.¦-¦Computed tomography (CT) is an imaging technique in which interest has been growing since it first began to be used in the early 1970s. In the clinical environment, this imaging system has emerged as the gold standard modality because of its high sensitivity in producing accurate diagnostic images. However, even if a direct benefit to patient healthcare is attributed to CT, the dramatic increase of the number of CT examinations performed has raised concerns about the potential negative effects of ionizing radiation on the population. To insure a benefit - risk that works in favor of a patient, it is important to balance image quality and dose in order to avoid unnecessary patient exposure.¦If this balance is important for adults, it should be an absolute priority for children undergoing CT examinations, especially for patients suffering from diseases requiring several follow-up examinations over the patient's lifetime. Indeed, children and young adults are more sensitive to ionizing radiation and have an extended life span in comparison to adults. For this population, the risk of developing cancer, whose latency period exceeds 20 years, is significantly higher than for adults. Assuming that each patient examination is justified, it then becomes a priority to optimize CT acquisition protocols in order to minimize the delivered dose to the patient. Over the past few years, CT advances have been developing at a rapid pace. Since 2009, new iterative image reconstruction techniques, called statistical iterative reconstructions, have been introduced in order to decrease patient exposure and improve image quality.¦The goal of the present work was to determine the potential of statistical iterative reconstructions to reduce dose as much as possible without compromising image quality and maintain diagnosis of children and young adult examinations.¦The optimization step requires the evaluation of the delivered dose and image quality useful to perform diagnosis. While the dose is estimated using CT indices (CTDIV0| and DLP), the particularity of this research was to use two radically different approaches to evaluate image quality. The first approach, called the "physical approach", computed physical metrics (SD, MTF, NPS, etc.) measured on phantoms in well-known conditions. Although this technique has some limitations because it does not take radiologist perspective into account, it enables the physical characterization of image properties in a simple and timely way. The second approach, called the "clinical approach", was based on the evaluation of anatomical structures (diagnostic criteria) present on patient images. Radiologists, involved in the assessment step, were asked to score image quality of structures for diagnostic purposes using a simple rating scale. This approach is relatively complicated to implement and also time-consuming. Nevertheless, it has the advantage of being very close to the practice of radiologists and is considered as a reference method.¦Primarily, this work revealed that the statistical iterative reconstructions studied in clinic (ASIR? and VECO have a strong potential to reduce CT dose (up to -90%). However, by their mechanisms, they lead to a modification of the image appearance with a change in image texture which may then effect the quality of the diagnosis. By comparing the results of the "clinical" and "physical" approach, it was showed that a change in texture is related to a modification of the noise spectrum bandwidth. The NPS analysis makes possible to anticipate or avoid a decrease in image quality. This project demonstrated that integrating these new statistical iterative reconstruction techniques can be complex and cannot be made on the basis of protocols using conventional reconstructions. The conclusions of this work and the image quality tools developed will be able to guide future studies in the field of image quality as texture analysis or model observers dedicated to CT.

Mécanismes moléculaires de la fonction anti-apoptotique des HDLs sur la cellule béta pancréatique : 4E-BP1 comme potentielle cible thérapeutique. [Molecular mechanisms of anti-apoptotic function of HDLs on pancreatic beta cells : 4E-BP1 as a potential therapeutic target]

Introduction : Les particules de HDL (High Density Lipoprotein) ont des fonctions diverses notamment en raison de leur structure très hétérogène. Tout d'abord, les HDLs assurent le transport du cholestérol de la périphérie vers le foie mais sont également dotées de nombreuses vertus protectrices. Un grand nombre d'études démontre les mécanismes de protection des HDL sur les cellules endothéliales. Sachant que les patients diabétiques ont ses niveaux bas de HDL, le but de cette étude est d'investiguer les mécanismes moléculaires de protection sur la cellule beta pancréatique. Résultats : Une étude « microarray » nous a permis d'obtenir une liste de gènes régulés par le stress, comme la privation de sérum, en présence ou en absence de HDL. Parmi ces gènes, nous nous sommes particulièrement intéressés à un répresseur de la synthèse protéique « cap » -dépendante, 4EBP1. Dans notre étude transcriptomique, les niveaux d'ARNm de 4E-BP1 augmentaient de 30þ% dans des conditions sans sérum alors que les HDLs bloquaient cette élévation. Au niveau protéique, les niveaux totaux de 4EBP1 étaient augmentés dans les conditions de stress et cette élévation était contrée par les HDLs. D'autres expériences de transfection ou d'infection de 4E-BP1 ont montrés que cette protéine était capable d'induire l'apoptose dans les cellules beta, imitant ainsi l'effet de la privation de sérum. Afin de déterminer le rôle direct de 4E-BP1 dans la mort cellulaire, ses niveaux ont été réduits par interférence ARN. Le niveau de mort cellulaire induit par l'absence de sérum était moins élevé dans des cellules à taux réduits de 4EBP1 par RNAi que dans des cellules contrôle. Conclusion : Ces données montrent que les HDL protègent les cellules beta suite à différents stress et que 4E-BP1 est une des protéines pro-apoptotiques inhibées par les HDL. 4E-BP1 est capable d'induire la mort cellulaire dans les cellules bêta et cette réponse peut-être réduite en diminuant l'expression de cette protéine. Nos données suggèrent que 4E-BP1 est une cible potentielle pour le traitement du diabète.

Endocarditis infecciosa por estreptococos beta-hemolíticos. Características y curso clínico de una cohorte multinacional.

Se trata de un estudio descriptivo de una cohorte prospectiva y multinacional de pacientes adultos diagnosticados de endocarditis infecciosa, de la cual se seleccionaron los pacientes diagnosticados de endocarditis infecciosa por estreptococos beta-hemolíticos y por estreptococos orales del grupo viridans. Los objetivos del estudio fueron describir las características clínicas, ecocardiográficas y el pronóstico de los pacientes con endocarditis infecciosa causada por estreptococos beta-hemolíticos y comparar dichas características con la endocarditis causada por estreptococos del grupo viridans. Las principales conclusiones fueron que la endocarditis infecciosa por estreptococos beta-hemolíticos es una entidad poco frecuente, que presenta un curso agresivo con una alta frecuencia de complicaciones y una necesidad quirúrgica alta.

Ab initio absorption spectrum of NiO combining molecular dynamics with the embedded cluster approach in a discrete reaction field

We developed a procedure that combines three complementary computational methodologies to improve the theoretical description of the electronic structure of nickel oxide. The starting point is a Car-Parrinello molecular dynamics simulation to incorporate vibrorotational degrees of freedom into the material model. By means ofcomplete active space self-consistent field second-order perturbation theory (CASPT2) calculations on embedded clusters extracted from the resulting trajectory, we describe localized spectroscopic phenomena on NiO with an efficient treatment of electron correlation. The inclusion of thermal motion into the theoretical description allowsus to study electronic transitions that, otherwise, would be dipole forbidden in the ideal structure and results in a natural reproduction of the band broadening. Moreover, we improved the embedded cluster model by incorporating self-consistently at the complete active space self-consistent field (CASSCF) level a discrete (or direct) reaction field (DRF) in the cluster surroundings. The DRF approach offers an efficient treatment ofelectric response effects of the crystalline embedding to the electronic transitions localized in the cluster. We offer accurate theoretical estimates of the absorption spectrum and the density of states around the Fermi level of NiO, and a comprehensive explanation of the source of the broadening and the relaxation of the charge transferstates due to the adaptation of the environment

Genetic structure of a linear population of Beta vulgaris ssp maritima (sea beet) revealed by isozyme and RFLP analysis

Knowledge of the genetic structure of plant populations is necessary for the understanding of the dynamics of major ecological processes. It also has applications in conservation biology and risk assessment for genetically modified crops. This paper reports the genetic structure of a linear population of sea beet, Beta vulgaris ssp. maritima (the wild relative of sugar beet), on Furzey Island, Poole Harbour. The relative spatial positions of the plants were accurately mapped and the plants were scored for variation at isozyme and RFLP loci. Structure was analysed by repeated subdivision of the population to find the average size of a randomly mating group. Estimates of F-ST between randomly mating units were then made, and gave patterns consistent with the structure of the population being determined largely by founder effects. The implications of these results for the monitoring of transgene spread in wild sea beet populations are discussed.