Role of Staphylococcus aureus surface proteins in experimental endocarditis

Résumé Le staphylocoque doré est un pathogène responsable d'une grande variété de maladies chez l'être humain. Il est extrêmement bien équipé de facteurs de virulence, dont les adhésines. Jusqu'à présent, 21 protéines liant des composants de tissus de l'hôte ("microbial surface components reacting with adherence matrix molecules, MSCRAMM") ont été identifiées, par exemple le "clumping factor" A (CIfA) ou la "fibronectin-binding protein" A (FnBPA). Néanmoins, pour la plupart de ces protéines, leur rôle dans la pathogénie des infections à staphylocoque doré reste à être élucidé. Le but de cette thèse est de contribuer à ce processus. Premièrement, les "MSCRAMM" CIfA, CIfB, FnBPA, FnBPB, Cna, SpA, Pls, SdrC, SdrD, SdrE, SasD, SasE, SasF, SasG, Sasl, SasJ et SasK ont été exprimés dans une bactérie substitut, Lactococcus lactis, et testés pour leurs propriétés adhésives et leur pathogénicité dans un modèle d'endocardite expérimentale (voir chapitre 1). Cette technique a préalablement été utilisée avec succès et a l'avantage d'éviter le contexte complexe des redondances et systèmes de régulations propres au staphylocoque doré. Les résultats montrent que, de tous les facteurs de virulence testés, seuls CIfA et FnBPA sont d'importance primordiale dans le développement d'endocardite expérimentale. En ce qui concerne l'internalisation dans les cellules endothéliales, seulement FnPBA et FnBPB en sont capables. En outre, l'adhérence à chacun des ligands testés (fibrinogène, fibronectine, kératine, élastine, collagène, et les caillots de fibrine et plaquettes) est très spécifique et est médiée par une ou plusieures adhésines provenant du staphylocoque doré. Par conséquence, ces protéines pourraient représenter des cibles potentielles pour de futures thérapies anti-adhésives contre le staphylocoque doré. Deuxièmement, l'expression des facteurs de virulence décrits dans le chapitre 1 par les souches recombinantes de lactocoques a été vérifiée par une nouvelle méthode utilisant la spectrométrie de masse (voir chapitre 2). L'expression de toutes ces protéines par les souches recombinantes a pu être confirmée. Cette méthode pourrait être de grande valeur dans la vérification de la présence de protéines quelconques dans toutes sortes d'applications. Troisièmement, deux facteurs de virulence du staphylocoque, CIfA et une forme tronquée de FnBPA, ont été exprimés de façon simultanée dans une souche recombinante de lactocoque (voir chapitre 3}. Contrairement à une souche exprimant la FnBPA entière, une souche exprimant la forme tronquée de FnPBA, qui ne contient plus le domaine capable de lier le fibrinogène, perd complètement sa capacité d'infecter dans le modèle d'endocardite expérimentale. Par contre, il est montré que, en cas de complémentation de la forme tronquée de FnPBA avec le domaine de liaison au fibrinogène de CIfA dans la souche double recombinante, le phénotype intégral de FnBPA est récupéré. En conséquence, les facteurs de virulence sont capables de coopérer dans le but de la pathogénie des infections à staphylocoque doré. Summary Staphylococcus aureus is a human pathogen causing a wide variety of disease. It is extremely well equipped with both secreted and surface-attached virulence factors, which can act as adhesins to host tissues. In total, twenty-one microbial surface components reacting with adherence matrix molecules (MSCRAMMs) have been identified, so far. These include well-characterized adhesins such as clumping factor A (CIfA) or fibronectin-binding protein A (FnBPA). However, for most of them their potential role in the pathogenesis of staphylococcal infections remains to be elucidated. This has been attempted in this thesis work. Firstly, the staphylococcal MSCRAMMs CIfA, CIfB, FnBPA, FnBPB, Cna, SpA, Pls, SdrC, SdrD, SdrE, SasD, SasE, SasF, SasG, Sasl, SasJ, and SasK have been expressed in a surrogate bacterium, Lactococcus lactis, and tested for their in vitro adherence properties and their pathogenicity in the rat model of experimental endocarditis (see chapter 1). This model has successfully been used previously, and has the advantage of bypassing the complex S. aureus background of redundancies and differential regulation. Here, it is shown that of the seventeen tested potential virulence factors, only CIfA and FnBPA are critical for the pathogenesis of experimental endocarditis in rats, while internalization into bovine endothelial cells is mediated exclusively by FnBPA and FnBPB. In addition, the adherence to specific host ligands (fibrinogen, fibronectin, keratin, elastin, collagen, and fibrin-platelet clots) is highly specific and mediated by one or few staphylococcal adhesins, respectively. Thus, these surface proteins may represent potential targets for an anti-adhesive strategy against S. aureus infections. Secondly, the expression of the staphylococcal proteins by L. lactis recombinants described in chapter 1 was tested by a novel method using mass spectrometry (see chapter 2). The expression of all the staphylococcal proteins by the respective recombinant lactococcal strain could be confirmed. This method may prove to be of great value in the confirmation of the presence of any given protein in various experimental settings. Thirdly, two staphylococcal virulence factors, CIfA and a truncated form of FnBPA, were expressed simultaneously in one recombinant lactococcal strain (see chapter 3). In contrast to a recombinant strain expressing full-length FnPBA, a recombinant strain expressing a truncated FnPBA, lacking the domain capable of binding fibrinogen, completely lost infectivity in experimental endocarditis. However, it is shown that the complementation of the truncated form of FnBPA with the fibrinogenbinding domain of CIfA in a double recombinant strain results in the recovery of the complete phenotype of full-length FnBPA. Thus, virulence factors can cooperate in the pathogenesis of staphylococcal infections.

Double-blind comparison of indapamide with a placebo in hypertensive patients treated by practicing physicians.

The antihypertensive effect of indapamide (2.5 mg/day) was compared to that obtained with a placebo in a controlled trial carried out by 11 physicians in their private practice. Thirty-one patients with uncomplicated essential hypertension were included. After a run-in period of 3 weeks without any treatment, either indapamide (n = 16) or a placebo (n = 15) were administered for 8 weeks in double-blind fashion. Blood pressure decreased in both groups. In patients treated with indapamide, systolic pressure was significantly lower than in those given the placebo at 3 out of the 4 follow-up visits; diastolic pressure, however, was significantly lower only at the end of the trial. Both the active drug and the placebo were well tolerated. No significant change in body weight, plasma potassium and uric acid occurred during the study in either group of patients. It appears therefore that indapamide, at a dose which apparently has no major diuretic effect, may be useful for practitioners in managing patients with mild to moderate hypertension.

Issue review : Iowa State Patrol and the Office of Motor Vehicle Enforcement potential integration.

This issue review analyzes the duties and responsibilities of troopers in the Iowa State Patrol, or ISP, and Motor Vehicle Enforcement, MVE, officers in the Department of Transportation, or DOT, as well as the differences such as funding, pay and pension. In addition, this issue review discusses the proposal for a potential integration of the offices under one system.

Identification of Wax Esters in Latent Print Residues by Gas Chromatography-Mass Spectromertry and Their Potential Use as Aging Parameters

Recent studies show that the composition of fingerprint residue varies significantly from the same donor as well as between donors. This variability is a major drawback in latent print dating issues. This study aimed, therefore, at the definition of a parameter that is less variable from print to print, using a ratio of peak area of a target compound degrading over time divided by the summed area of peaks of more stable compounds also found in latent print residues.Gas chromatography-mass spectrometry (GC/MS) analysis of the initial lipid composition of latent prints identifies four main classes of compounds that can be used in the definition of an aging parameter: fatty acids, sterols, sterol precursors, and wax esters (WEs). Although the entities composing the first three groups are quite well known, those composing WEs are poorly reported. Therefore, the first step of the present work was to identify WE compounds present in latent print residues deposited by different donors. Of 29 WEs recorded in the chromatograms, seven were observed in the majority of samples.The identified WE compounds were subsequently used in the definition of ratios in combination with squalene and cholesterol to reduce the variability of the initial composition between latent print residues from different persons and more particularly from the same person. Finally, the influence of a latent print enhancement process on the initial composition was studied by analyzing traces after treatment with magnetic powder, 1,2-indanedione, and cyanoacrylate.

Double incretin receptor knock-out (DIRKO) mice present with alterations of trabecular and cortical micromorphology and bone strength.

A role for gut hormone in bone physiology has been suspected. We evidenced alterations of microstructural morphology (trabecular and cortical) and bone strength (both at the whole-bone - and tissue-level) in double incretin receptor knock-out (DIRKO) mice as compared to wild-type littermates. These results support a role for gut hormones in bone physiology. INTRODUCTION: The two incretins, glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1), have been shown to control bone remodeling and strength. However, lessons from single incretin receptor knock-out mice highlighted a compensatory mechanism induced by elevated sensitivity to the other gut hormone. As such, it is unclear whether the bone alterations observed in GIP or GLP-1 receptor deficient animals resulted from the lack of a functional gut hormone receptor, or by higher sensitivity for the other gut hormone. The aims of the present study were to investigate the bone microstructural morphology, as well as bone tissue properties, in double incretin receptor knock-out (DIRKO) mice. METHODS: Twenty-six-week-old DIRKO mice were age- and sex-matched with wild-type (WT) littermates. Bone microstructural morphology was assessed at the femur by microCT and quantitative X-ray imaging, while tissue properties were investigated by quantitative backscattered electron imaging and Fourier-transformed infrared microscopy. Bone mechanical response was assessed at the whole-bone- and tissue-level by 3-point bending and nanoindentation, respectively. RESULTS: As compared to WT animals, DIRKO mice presented significant augmentations in trabecular bone mass and trabecular number whereas bone outer diameter, cortical thickness, and cortical area were reduced. At the whole-bone-level, yield stress, ultimate stress, and post-yield work to fracture were significantly reduced in DIRKO animals. At the tissue-level, only collagen maturity was reduced by 9 % in DIRKO mice leading to reductions in maximum load, hardness, and dissipated energy. CONCLUSIONS: This study demonstrated the critical role of gut hormones in controlling bone microstructural morphology and tissue properties.

TLR3 and Rig-like receptor on myeloid dendritic cells and Rig-like receptor on human NK cells are both mandatory for production of IFN-gamma in response to double-stranded RNA.

Cross-talk between NK cells and dendritic cells (DCs) is critical for the potent therapeutic response to dsRNA, but the receptors involved remained controversial. We show in this paper that two dsRNAs, polyadenylic-polyuridylic acid and polyinosinic-polycytidylic acid [poly(I:C)], similarly engaged human TLR3, whereas only poly(I:C) triggered human RIG-I and MDA5. Both dsRNA enhanced NK cell activation within PBMCs but only poly(I:C) induced IFN-gamma. Although myeloid DCs (mDCs) were required for NK cell activation, induction of cytolytic potential and IFN-gamma production did not require contact with mDCs but was dependent on type I IFN and IL-12, respectively. Poly(I:C) but not polyadenylic-polyuridylic acid synergized with mDC-derived IL-12 for IFN-gamma production by acting directly on NK cells. Finally, the requirement of both TLR3 and Rig-like receptor (RLR) on mDCs and RLRs but not TLR3 on NK cells for IFN-gamma production was demonstrated using TLR3- and Cardif-deficient mice and human RIG-I-specific activator. Thus, we report the requirement of cotriggering TLR3 and RLR on mDCs and RLRs on NK cells for a pathogen product to induce potent innate cell activation.

Neurophysiological effects of vasopressin and oxytocin in the central amygdala of the rat : a potential mechanism for the opposite modulation of anxiety and fear behavior

Abstract The amygdala is a group of nuclei in the temporal lobe of the brain that plays a crucial role in anxiety and fear behavior. Sensory information converges in the basolateral and lateral nuclei of the amygdala, which have been the first regions in the brain where the acquisition of new (fear) memories has been associated with long term changes in synaptic transmission. These nuclei, in turn, project to the central nucleus of the amygdala. The central amygdala, through its extensive projections to numerous nuclei in the midbrain and brainstem, plays a pivotal role in the orchestration of the rapid autonomic and endocrine fear responses. In the central amygdala a large number of neuropeptides and receptors is expressed, among which high levels of vasopressin and oxytocin receptors. Local injections of these peptides into the amygdala modulate several aspects of the autonomic fear reaction. Interestingly, their effects are opposing: vasopressin tends to enhance the fear reactions, whereas oxytocin has anxiolytic effects. In order to investigate the neurophysiological mechanisms that could underlie this opposing modulation of the fear behavior, we studied the effects of vasopressin and oxytocin on the neuronal activity in an acute brain slice preparation of the rat central amygdala. We first assessed the effects of vasopressin and oxytocin on the spontaneous activity of central amygdala neurons. Extracellular single unit recordings revealed two major populations of neurons: a majority of neurons was excited by vasopressin and inhibited by oxytocin, whereas other neurons were only excited by oxytocin receptor activation. The inhibitory effect of oxytocin could be reduced by the block of GABAergic transmission, whereas the excitatory effects of vasopressin and oxytocin were not affected. In a second step we identified the cellular mechanisms for the excitatory effects of both peptides as well as the morphological and biochemical mechanisms underlying the opposing effects, by using sharp electrode recordings together with intracellular labelings. We revealed that oxytocin-excited neurons are localized in the lateral part (CeL) whereas vasopressin excited cells are found in the medial part of the central amygdala (CeM). The tracing of the neuronal morphology showed that the axon collaterals of the oxytocin-excited neurons project from the CeL, far into the CeM. Combined immunohistochemical stainings indicated that these projections are GABAergic. In the third set of experiments we investigated the synaptic interactions between the two identified cell populations. Whole-cell patch-clamp recordings in the CeM revealed that the inhibitory effect of oxytocin was caused by the massive increase of inhibitory GABAergic currents, which was induced by the activation of CeL neurons. Finally, the effects of vasopressin and oxytocin on evoked activity were investigated. We found on the one hand, that the probability of evoking action potentials in the CeM by stimulating the basolateral amygdala afferents was enhanced under vasopressin, whereas it decreased under oxytocin. On the other hand, the impact of cortical afferents stimulation on the CeL neurons was enhanced by oxytocin application. Taken together, these findings have allowed us to develop a model, in which the opposing behavioral effects of vasopressin and oxytocin are caused by a selective activation of two distinct populations of neurons in the GABAergic network of the central amygdala. Our model could help to develop new anxiolytic treatments, which modulate simultaneously both receptor systems. By acting on a GABAergic network, such treatments can further be tuned by combinations with classical benzodiazepines. Résumé: L'amygdale est un groupe de noyaux cérébraux localisés dans le lobe temporal. Elle joue un rôle essentiel dans les comportements liés à la peur et l'anxiété. L'information issue des aires sensorielles converge vers les noyaux amygdaliens latéraux et basolatéraux, qui sont les projections vers différents noyaux du tronc cérébral et de l'hypothalamus, joue un rôle clef premières régions dans lesquelles il a été démontré que l'acquisition d'une nouvelle mémoire (de peur) était associée à des changements à long terme de la transmission synaptique. Ces noyaux envoient leurs projections sur l'amygdale centrale, qui à travers ses propres dans l'orchestration des réponses autonomes et endocrines de peur. Le contrôle de l'activité neuronale dans l'amygdale centrale module fortement la réaction de peur. Ainsi, un grand nombre de neuropeptides sont spécifiquement exprimés dans l'amygdale centrale et un bon nombre d'entre eux interfère dans la réaction de peur et d'anxiété. Chez les rats, une forte concentration de récepteurs à l'ocytocine et à la vasopressine est exprimée dans le noyau central, et l'injection de ces peptides dans l'amygdale influence différents aspects de la réaction viscérale associée à la peur. Il est intéressant de constater que ces peptides exercent des effets opposés. Ainsi, la vasopressine augmente la réaction de peur alors que l'ocytocine a un effet anxiolytique. Afin d'investiguer les mécanismes neurophysiologiques responsables de ces effets opposés, nous avons étudié l'effet de la vasopressine et de l'ocytocine sur l'activité neuronale de préparations de tranches de cerveau de rats contenant entre autres de l'amygdale centrale. Tout d'abord, notre intérêt s'est porté sur les effets de ces deux neuropeptides sur l'activité spontanée dans l'amygdale centrale. Des enregistrements extracellulaires ont révélé différentes populations de neurones ; une majorité était excitée par la vasopressine et inhibée par l'ocytocine ; d'autres étaient seulement excités par l'activation du récepteur à l'ocytocine. L'effet inhibiteur de l'ocytocine a pu être réduit par l'inhibition de la transmission GABAergique, alors que ses effets excitateurs n'étaient pas affectés. Dans un deuxième temps, nous avons identifié les mécanismes cellulaires responsables de l'effet excitateur de ces deux peptides et analysé les caractéristiques morphologiques et biochimiques des neurones affectés. Des enregistrements intracellulaires ont permis de localiser les neurones excités par l'ocytocine dans la partie latérale de l'amygdale centrale (CeL), et ceux excités par la vasopressine dans sa partie médiale (CeM). Le traçage morphologique des neurones a révélé que les collatérales axonales des cellules excitées par l'ocytocine projetaient du CeL loin dans le CeM. De plus, des colorations immuno-histochimiques ont révélé que ces projections étaient GABAergiques. Dans un troisième temps, nous avons étudié les interactions synaptiques entre ces deux populations de cellules. Les enregistrements en whole-cell patch-clamp dans le CeM ont démontré que les effets inhibiteurs de l'ocytocine résultaient de l'augmentation massive des courants GABAergique résultant de l'activation des neurones dans le CeL. Finalement, les effets de l'ocytocine et de la vasopressine sur l'activité évoquée ont été étudiés. Nous avons pu montrer que la probabilité d'évoquer un potentiel d'action dans le CeM, par stimulation de l'amygdale basolatérale, était augmentée sous l'effet de la vasopressine et diminuée sous l'action de l'ocytocine. Par contre, l'impact de la stimulation des afférences corticales sur les neurones du CeL était augmenté par l'application de l'ocytocine. L'ensemble de ces résultats nous a permis de développer un modèle dans lequel les effets comportementaux opposés de la vasopressine et de l'ocytocine sont causés par une activation sélective des deux différentes populations de neurones dans un réseau GABAergique. Un tel modèle pourrait mener au développement de nouveaux traitements anxiolytiques en modulant l'activité des deux récepteurs simultanément. En agissant sur un réseau GABAergique, les effets d'un tel traitement pourraient être rendus encore plus sélectifs en association avec des benzodiazépines classiques.

Traitement de la dépression résistante par l'association citalopram-lithium. Méthodologie d'une étude multicentrique en double aveugle et résultats préliminaires [Treatment of resistant depression with the citalopram-lithium combination. Methodology of a double-blind multicenter study and preliminary results].

Citalopram, a new bicyclic antidepressant, is the most selective serotonin reuptake inhibitor. In a number of double-blind controlled studies, citalopram was compared to placebo and to known tricyclic antidepressants. These studies have shown their efficacy and good safety. The inefficacy of a psychotropic treatment in at least 20% of depressives has led a number of authors to propose original drug combinations and associations, like antidepressant/lithium (Li), antidepressant/sleep deprivation (agrypnia), antidepressant/ECT, or antidepressant/LT3. The aim of this investigation is to evaluate the clinical effectiveness and safety of a combined citalopram/lithium treatment in therapy-resistant patients, taking account of serotonergic functions, as tested by the fenfluramine/prolactin test, and of drug pharmacokinetics and pharmacogenetics of metabolism. DESIGN OF THE STUDY: A washout period of 3 days before initiating the treatment is included. After an open treatment phase of 28 days (D) with citalopram (20 mg D1-D3; 40 mg D4-D14; 40 or 60 mg D15-D28; concomitant medication allowed: chloral, chlorazepate), the nonresponding patients [less than 50% improvement in the total score on the 21 item-Hamilton Depression Rating Scale (HDRS)] are selected and treated with or without Li (randomized in double-blind conditions: citalopram/Li or citalopram/placebo) during the treatment (D29-D35). Thereafter, all patients included in the double-blind phase subsequently receive an open treatment with citalopram/Li for 7 days (D36-D42). The hypothesis of a relationship between serotoninergic functions in patients using the fenfluramine/prolactin test (D1) and the clinical response to citalopram (and Li) is assessed. Moreover, it is evaluated whether the pharmacogenetic status of the patients, as determined by the mephenytoin/dextromethorphan test (D0-D28), is related to the metabolism of fenfluramine and citalopram, and also to the clinical response. CLINICAL ASSESSMENT: Patients with a diagnosis of major depressive disorders according to DSM III are submitted to a clinical assessment of D1, D7, D14, D28, D35, D42: HDRS, CGI (clinical global impression), VAS (visual analog scales for self-rating of depression), HDRS (Hamilton depression rating scale, 21 items), UKU (side effects scale), and to clinical laboratory examens, as well as ECG, control of weight, pulse, blood pressure at D1, D28, D35. Fenfluramine/prolactin test: A butterfly needle is inserted in a forearm vein at 7 h 45 and is kept patent with liquemine. Samples for plasma prolactin, and d- and l-fenfluramine determinations are drawn at 8 h 15 (base line). Patients are given 60 mg fenfluramine (as a racemate) at 8 h 30. Kinetic points are determined at 9 h 30, 10 h 30, 11 h 30, 12 h 30, 13 h 30. Plasma levels of d- and l-fenfluramine are determined by gas chromatography and prolactin by IRNA. Mephenytoin/dextromethorphan test: Patients empty their bladders before the test; they are then given 25 mg dextropethorphan and 100 mg mephenytoin (as a racemate) at 8 h 00. They collect all urines during the following 8 hours. The metabolic ratio is determined by gas chromatography (metabolic ratio dextromethorphan/dextrorphan greater than 0.3 = PM (poor metabolizer); mephenytoin/4-OH-mephenytoin greater than 5.6, or mephenytoin S/R greater than 0.8 = PM). Citalopram plasma levels: Plasma levels of citalopram, desmethylcitalopram and didesmethylcitalopram are determined by gas chromatography--mass spectrometry. RESULTS OF THE PILOT STUDY. The investigation has been preceded by a pilot study including 14 patients, using the abovementioned protocol, except that all nonresponders were medicated with citalopram/Li on D28 to D42. The mean total score (n = 14) on the 21 item Hamilton scale was significantly reduced after the treatment, ie from 26.93 +/- 5.80 on D1 to 8.57 +/- 6.90 on D35 (p less than 0.001). A similar patCitalopram, a new bicyclic antidepressant, is the most selective serotonin reu

Shikonin-loaded antibody-armed nanoparticles for targeted therapy of ovarian cancer.

Conventional chemotherapy of ovarian cancer often fails because of initiation of drug resistance and/or side effects and trace of untouched remaining cancerous cells. This highlights an urgent need for advanced targeted therapies for effective remediation of the disease using a cytotoxic agent with immunomodulatory effects, such as shikonin (SHK). Based on preliminary experiments, we found SHK to be profoundly toxic in ovarian epithelial cancer cells (OVCAR-5 and ID8 cells) as well as in normal ovarian IOSE-398 cells, endothelial MS1 cells, and lymphocytes. To limit its cytotoxic impact solely to tumor cells within the tumor microenvironment (TME), we aimed to engineer SHK as polymeric nanoparticles (NPs) with targeting moiety toward tumor microvasculature. To this end, using single/double emulsion solvent evaporation/diffusion technique with sonication, we formulated biodegradable NPs of poly(lactic-co-glycolic acid) (PLGA) loaded with SHK. The surface of NPs was further decorated with solubilizing agent polyethylene glycol (PEG) and tumor endothelial marker 1 (TEM1)/endosialin-targeting antibody (Ab) through carbodiimide/N-hydroxysuccinimide chemistry. Having characterized the physicochemical and morphological properties of NPs, we studied their drug-release profiles using various kinetic models. The biological impact of NPs was also evaluated in tumor-associated endothelial MS1 cells, primary lymphocytes, and epithelial ovarian cancer OVCAR-5 cells. Based on particle size analysis and electron microscopy, the engineered NPs showed a smooth spherical shape with size range of 120 to 250 nm and zeta potential value of -30 to -40 mV. Drug entrapment efficiency was ~80%-90%, which was reduced to ~50%-60% upon surface decoration with PEG and Ab. The liberation of SHK from NPs showed a sustained-release profile that was best fitted with Wagner log-probability model. Fluorescence microscopy and flow cytometry analysis showed active interaction of Ab-armed NPs with TEM1-positive MS1 cells, but not with TEM1-negative MS1 cells. While exposure of the PEGylated NPs for 2 hours was not toxic to lymphocytes, long-term exposure of the Ab-armed and PEGylated NPs was significantly toxic to TEM1-positive MS1 cells and OVCAR-5 cells. Based on these findings, we propose SHK-loaded Ab-armed PEGylated PLGA NPs as a novel nanomedicine for targeted therapy of solid tumors.

Potential Impacts of Climate Change on Ecosystem Services in Europe: the Case of Pest Control by Vertebrates. BioScience.

Global environmental changes threaten ecosystems and cause significant alterations to the supply of ecosystem services that are vital for human well-being. We provide an assessment of the potential impacts of climate change on European diversity of vertebrates and their associated pest control services. We modeled the distributions of the species that provide this service using ensembles of forecasts from bioclimatic envelope models and then used their results to generate maps of potential species richness among vertebrate providers of pest control services. We assessed how potential richness of pest control providers would change according to different climate and greenhouse emissions scenarios. We found that potential richness of pest control providers was likely to face substantial reductions, especially in southern European countries that had economies highly dependent on agricultural yields. In much of central and northern Europe, where countries had their economies less dependent on agriculture, climate change was likely to benefit pest control providers

Application of RNAi to silence tartrate-resistant acid phosphatase: unexpected effects on the monocyte-macrophage lineage

Tartraatti-resistentin happaman fosfataasin hiljentäminen RNAi menetelmällä: odottamaton vaikutus monosyytti-makrofagi linjan soluissa RNA interferenssi (RNAi) eli RNA:n hiljentyminen löydettiin ensimmäisenä kasveissa, ja 2000-luvulla RNAi menetelmä on otettu käyttöön myös nisäkässoluissa. RNAi on mekanismi, jossa lyhyet kaksi juosteiset RNA molekyylit eli siRNA:t sitoutuvat proteiinikompleksiin ja sitoutuvat komplementaarisesti proteiinia koodaavaan lähetti RNA:han katalysoiden lähetti RNA:n hajoamisen. Tällöin RNA:n koodaamaa proteiinia ei solussa tuoteta. Tässä työssä on RNA interferenssi menetelmän avuksi kehitetty uusi siRNA molekyylien suunnittelualgoritmi siRNA_profile, joka etsii lähetti RNA:sta geenin hiljentämiseen sopivia kohdealueita. Optimaalisesti suunnitellulla siRNA molekyylillä voi olla mahdollista saavuttaa pitkäaikainen geenin hiljeneminen ja spesifinen kohdeproteiinin määrän aleneminen solussa. Erilaiset kemialliset modifikaatiot, mm. 2´-Fluoro-modifikaatio, siRNA molekyylin riboosirenkaassa lisäsivät siRNA molekyylin stabiilisuutta veren plasmassa sekä siRNA molekyylin tehokkuutta. Nämä ovat tärkeitä siRNA molekyylien ominaisuuksia kun RNAi menetelmää sovelletaan lääketieteellisiin tarkoituksiin. Tartraatti-resistentti hapan fosfataasi (TRACP) on entsyymi, joka esiintyy luunsyöjäsoluissa eli osteoklasteissa, antigeenejä esittelevissä dendiriittisissä soluissa sekä eri kudosten makrofageissa, jotka ovat syöjäsoluja. TRACP entsyymin biologista tehtävää ei ole saatu selville, mutta oletetaan että TRACP entsyymin kyvyllä tuottaa reaktiivisia happiradikaaleja on tehtävä sekä luuta hajoittavissa osteoklasteissa sekä antigeenia esittelevissä dendriittisissä soluissa. Makrofageilla, jotka yliekpressoivat TRACP entsyymiä, on myös solunsisäinen reaktiivisten happiradikaalien tuotanto sekä bakteerin tappokyky lisääntynyt. TRACP-geenin hiljentämiseen tarkoitetut spesifiset DNA ja siRNA molekyylit aiheuttivat monosyytti-makrofagilinjan soluviljelymallissa TRACP entsyymin tuoton lisääntymistä odotusten vastaisesti. DNA ja RNA molekyylien vaikutusta TRACP entsyymin tuoton lisääntymiseen tutkittiin myös Tolllike reseptori 9 (TLR9) poistogeenisestä hiirestä eristetyissä monosyyttimakrofaagisoluissa. TRACP entsyymin tuoton lisääntyminen todettiin sekvenssistä ja TLR9:stä riippumattomaksi vasteeksi solun ulkopuolisia DNA ja RNA molekyylejä vastaan. Havainto TRACP entsyymin tuoton lisääntymisestä viittaa siihen, että TRACP entsyymillä on tehtävä solun immuunipuolustusjärjestelmässä.

Hybrid model of a double-fed induction machine and back-to-back converter

This report details the port interconnection of two subsystems: a power electronics subsystem (a back-to-back AC/AC converter (B2B), coupled to a phase of the power grid), and an electromechanical subsystem (a doubly-fed induction machine (DFIM), coupled mechanically to a flywheel and electrically to the power grid and to a local varying load). Both subsystems have been essentially described in previous reports (deliverables D 0.5 and D 4.3.1), although some previously unpublished details are presented here. The B2B is a variable structure system (VSS), due to the presence of control-actuated switches: however from a modelling and simulation, as well as a control-design, point of view, it is sensible to consider modulated transformers (MTF in the bond-graph language) instead of the pairs of complementary switches. The port-Hamiltonian models of both subsystems are presents and coupled through a power-preserving interconnection, and the Hamiltonian description of the whole system is obtained; detailed bond-graphs of all the subsystems and the complete system are provided.

A methodology to estimate the potential to move inpatient to one day surgery.

BACKGROUND: The proportion of surgery performed as a day case varies greatly between countries. Low rates suggest a large growth potential in many countries. Measuring the potential development of one day surgery should be grounded on a comprehensive list of eligible procedures, based on a priori criteria, independent of local practices. We propose an algorithmic method, using only routinely available hospital data to identify surgical hospitalizations that could have been performed as one day treatment. METHODS: Moving inpatient surgery to one day surgery was considered feasible if at least one surgical intervention was eligible for one day surgery and if none of the following criteria were present: intervention or affection requiring an inpatient stay, patient transferred or died, and length of stay greater than four days. The eligibility of a procedure to be treated as a day case was mainly established on three a priori criteria: surgical access (endoscopic or not), the invasiveness of the procedure and the size of the operated organ. Few overrides of these criteria occurred when procedures were associated with risk of immediate complications, slow physiological recovery or pain treatment requiring hospital infrastructure. The algorithm was applied to a random sample of one million inpatient US stays and more than 600 thousand Swiss inpatient stays, in the year 2002. RESULTS: The validity of our method was demonstrated by the few discrepancies between the a priori criteria based list of eligible procedures, and a state list used for reimbursement purposes, the low proportion of hospitalizations eligible for one day care found in the US sample (4.9 versus 19.4% in the Swiss sample), and the distribution of the elective procedures found eligible in Swiss hospitals, well supported by the literature. There were large variations of the proportion of candidates for one day surgery among elective surgical hospitalizations between Swiss hospitals (3 to 45.3%). CONCLUSION: The proposed approach allows the monitoring of the proportion of inpatient stay candidates for one day surgery. It could be used for infrastructure planning, resources negotiation and the surveillance of appropriate resource utilization.

Characterization of rue extract and its potential for controlling rice blast

Abstract: The objective of this work was to purify and standardize the rue (Ruta graveolens) extract and evaluate its effect on Magnaporthe oryzae as an alternative to the integrated management of rice blast. The drug was characterized, the liquid extract was obtained, and the methodology for quantifying the standard markers psoralen and bergapten was validated. Rue extract and the markers, solely or in combination, were assayed in vitro, as well as in greenhouse conditions, for their ability to suppress leaf blast, by the evaluation of mycelial growth, conidial germination, and appressorium formation. Rue extract inhibited M. oryzae mycelial growth (100%), conidial germination (LD50=0.237 mg), and the appressorium formation (LD50=0.121 mg); besides, the extract reduced leaf blast severity by 80.84%. Fluorescence microscopy showed that rue extract did not damage M. oryzae cell wall and plasma membrane, indicating another mode of action. Rue extract has a great potential for controlling rice leaf blast.

Potential impact of environmental bacteriophages in spreading antibiotic resistance genes

The idea that bacteriophage transduction plays a role in the horizontal transfer of antibiotic resistance genes is gaining momentum. Such transduction might be vital in horizontal transfer from environmental to human bodyassociated biomes and here we review many lines of evidence supporting this notion. It is well accepted that bacteriophages are the most abundant entities in most environments, where they have been shown to be quite persistent. This fact, together with the ability of many phages to infect bacteria belonging to different taxa, makes them suitable vehicles for gene transfer. Metagenomic studies confirm that substantial percentages of the bacteriophage particles present in most environments contain bacterial genes, including mobile genetic elements and antibiotic resistance genes. When specific genes of resistance to antibiotics are detected by real-time PCR in the bacteriophage populations of different environments, only tenfold lower numbers of these genes are observed, compared with those found in the corresponding bacterial populations. In addition, the antibiotic resistance genes from these bacteriophages are functional and generate resistance to the bacteria when these genes are transfected. Finally, reports about the transduction of antibiotic resistance genes are on the increase.