Toll-Interacting Protein Deficiency Leads to Increased Susceptibility to Acute and Chronic Colitis.

BACKGROUND: Sensing of bacterial products via Toll-like receptors is critical to maintain gut immune homeostasis. The Toll-Interacting Protein (Tollip) inhibits downstream signaling through the IL-1 receptor, TLR-2 and TLR-4. Here,we aimed to address the role of Tollip in acute and chronic inflammatory responses in the gut. MATERIAL AND METHODS: WT or Tollip-deficient mice were exposed to dextran sulfate sodium (DSS) 1.5% in the drinking water during 7 days. To generate bone-marrow chimeras, WT or Tollip deficient mice were 900-rads irradiated, transplanted with WT or Tollip deficient bone-marrow cells and challenged with DSS 2-3 months after transplantation. IL-10 deficient mice were bred with Tollip deficient mice and colitis was compared at various time points. RESULTS: Upon DSS exposure, Tollip-deficient mice had increased body weight loss and increased pro-inflammatory cytokine expression compared to WT controls. Challenge of bone-marrow chimeras showed that colitis susceptibility was also increased when Tollip deficiency was restricted to non-hematopoietic cells. DSS-exposure lead to a disorganized distribution of zona-occludens-1, a tight junction marker and increased number of apoptotic, cleaved caspase 3 positive, epithelial cells in Tollip-deficient compared to WT mice. Chronic colitis was also affected by Tollip deficiency as Tollip/IL-10 deficient mice had more severe histological stigmata of colitis and higher IL-17 expression than IL-10 deficient controls. CONCLUSION: Tollip in non-hematopoietic cells is critical for adequate response to a chemical-induced stress in the gut and to hamper chronic bacteria-driven colitis. Modulation of epithelial cell integrity via Tollip likely contributes to the observed defects.

Glycogen storage capacity and de novo lipogenesis during massive carbohydrate overfeeding in man.

The metabolic balance method was performed on three men to investigate the fate of large excesses of carbohydrate. Glycogen stores, which were first depleted by diet (3 d, 8.35 +/- 0.27 MJ [1994 +/- 65 kcal] decreasing to 5.70 +/- 1.03 MJ [1361 +/- 247 kcal], 15% protein, 75% fat, 10% carbohydrate) and exercise, were repleted during 7 d carbohydrate overfeeding (11% protein, 3% fat, and 86% carbohydrate) providing 15.25 +/- 1.10 MJ (3642 +/- 263 kcal) on the first day, increasing progressively to 20.64 +/- 1.30 MJ (4930 +/- 311 kcal) on the last day of overfeeding. Glycogen depletion was again accomplished with 2 d of carbohydrate restriction (2.52 MJ/d [602 kcal/d], 85% protein, and 15% fat). Glycogen storage capacity in man is approximately 15 g/kg body weight and can accommodate a gain of approximately 500 g before net lipid synthesis contributes to increasing body fat mass. When the glycogen stores are saturated, massive intakes of carbohydrate are disposed of by high carbohydrate-oxidation rates and substantial de novo lipid synthesis (150 g lipid/d using approximately 475 g CHO/d) without postabsorptive hyperglycemia.

In vitro activity of gallium maltolate against Staphylococci in logarithmic, stationary, and biofilm growth phases: comparison of conventional and calorimetric susceptibility testing methods.

Ga(3+) is a semimetal element that competes for the iron-binding sites of transporters and enzymes. We investigated the activity of gallium maltolate (GaM), an organic gallium salt with high solubility, against laboratory and clinical strains of methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), methicillin-susceptible Staphylococcus epidermidis (MSSE), and methicillin-resistant S. epidermidis (MRSE) in logarithmic or stationary phase and in biofilms. The MICs of GaM were higher for S. aureus (375 to 2000 microg/ml) than S. epidermidis (94 to 200 microg/ml). Minimal biofilm inhibitory concentrations were 3,000 to >or=6,000 microg/ml (S. aureus) and 94 to 3,000 microg/ml (S. epidermidis). In time-kill studies, GaM exhibited a slow and dose-dependent killing, with maximal action at 24 h against S. aureus of 1.9 log(10) CFU/ml (MSSA) and 3.3 log(10) CFU/ml (MRSA) at 3x MIC and 2.9 log(10) CFU/ml (MSSE) and 4.0 log(10) CFU/ml (MRSE) against S. epidermidis at 10x MIC. In calorimetric studies, growth-related heat production was inhibited by GaM at subinhibitory concentrations; and the minimal heat inhibition concentrations were 188 to 4,500 microg/ml (MSSA), 94 to 1,500 microg/ml (MRSA), and 94 to 375 microg/ml (MSSE and MRSE), which correlated well with the MICs. Thus, calorimetry was a fast, accurate, and simple method useful for investigation of antimicrobial activity at subinhibitory concentrations. In conclusion, GaM exhibited activity against staphylococci in different growth phases, including in stationary phase and biofilms, but high concentrations were required. These data support the potential topical use of GaM, including its use for the treatment of wound infections, MRSA decolonization, and coating of implants.

Hypoxia-inducible miR-210 regulates the susceptibility of tumor cells to lysis by cytotoxic T cells.

Hypoxia in the tumor microenvironment plays a central role in the evolution of immune escape mechanisms by tumor cells. In this study, we report the definition of miR-210 as a miRNA regulated by hypoxia in lung cancer and melanoma, documenting its involvement in blunting the susceptibility of tumor cells to lysis by antigen-specific cytotoxic T lymphocytes (CTL). miR-210 was induced in hypoxic zones of human tumor tissues. Its attenuation in hypoxic cells significantly restored susceptibility to autologous CTL-mediated lysis, independent of tumor cell recognition and CTL reactivity. A comprehensive approach using transcriptome analysis, argonaute protein immunoprecipitation, and luciferase reporter assay revealed that the genes PTPN1, HOXA1, and TP53I11 were miR-210 target genes regulated in hypoxic cells. In support of their primary importance in mediating the immunosuppressive effects of miR-210, coordinate silencing of PTPN1, HOXA1, and TP53I11 dramatically decreased tumor cell susceptibility to CTL-mediated lysis. Our findings show how miR-210 induction links hypoxia to immune escape from CTL-mediated lysis, by providing a mechanistic understanding of how this miRNA mediates immunosuppression in oxygen-deprived regions of tumors where cancer stem-like cells and metastatic cellular behaviors are known to evolve.

Host genome influences on susceptibility to HIV-1

In vitro and in vivo analyses identified a significant component of heritability in cellular or host susceptibility to HIV-1. The bases for susceptibility can be traced to genetic differences (inter-species) resulting from evolutionary adaptation to exogenous (and endogenous) retroviral infections, and to intra-species and inter-individual (human) differences associated with genetic variation. We have completed large scale evolutionary analysis of genes involved in HIV life cycle and pathogenesis, as well as participating and conducting genome-wide association studies, linkage analysis, and transcriptome analysis. These studies allowed a better understanding of the influence of common human variants in HIV-1 susceptibility and define a number of experimental challenges in the filed: understanding of the role of rare and private mutations in susceptibility, and the development of better tools for the integration of data from large-scale studies.

Une méta-analyse GWA de la glycémie après 2h d'HGPO révèle que GIPR est associé avec la sécrétion de l'insuline en réponse au glucose et que l'ADCY5 est un nouveau gène de suseptibilité au diabète de type 2. [A meta-analysis of GWA blood glucose after 2 h of OGTT revealed that GIPR is associated with the secretion of insulin based on glucose and ADCY5 is a new susceptibility gene for type 2 diabetes.]

Introduction: Les études GVvA (Genome-wide association ,-studies) ont identifié et confirmé plus de 20 gènes de susceptibilité au DT2 et ont contribué à mieux comprendre sa physiopathologie. L'hyperglycémie à jeun (GJ), et 2 heures après une HGPO (G2h) sont les deux mesures cliniques du diagnostic du DT2. Nous avons identifié récemment la G6P du pancréas (G6PC2) comme déterminant de la variabilité physiologique de la GJ puis Ie récepteur à la mélatonine (MTNRIB) qui de plus lie la régulation du rythme circadien au DT2. Dans ce travail nous avons étudié la génétique de la G2h à l'aide de l'approche GWA. Résultats: Nous avons réalisé une méta-analyse GWA dans le cadre de MAGIC (Meta-Analysis of Glucose and Insulin related traits Consortium) qui a inclus 9 études GWA (N=15'234). La réplication de 29 loci (N=6958-30 121, P < 10-5 ) a confirmé 5 nouveaux loci; 2 étant connus comme associés avec Ie DT2 (TCF7L2, P = 1,6 X 10-10 ) et la GJ (GCKR, p = 5,6 X 10-10 ); alors que GIPR (p= 5,2 X 10-12), VSP13C (p= 3,9 X 10-8) et ADCY5 (p = 1,11 X 10-15 ) sont inédits. GIPR code Ie récepteur au GIP (gastric inhibitory polypeptide) qui est sécrété par les ceIlules intestinales pour stimuler la sécrétion de l'insuline en réponse au glucose (l'effet incrétine). Les porteurs du variant GIPR qui augmente la G2h ont également un indice insulinogénique plus bas, (p= 1,0 X 10-17) mais ils ne présentent aucune modification de leur glycémie suite à une hyperglycémie provoquée par voie veineuse (p= 0,21). Ces résultats soutiennent un effet incrétine du locus GIPR qui expliquerait ~9,6 % de la variance total de ce trait. La biologie de ADCY5 et VPS13C et son lien avec l'homéostasie du glucose restent à élucider. GIPR n'est pas associé avec le risque de DT2 indiquant qu'il influence la variabilité physiologique de la G2h alors que le locus ADCY5 est associé avec le DT2 (OR = 1,11, P = 1,5 X 10-15). Conclusion: Notre étude démontre que l'étude de la G2h est une approche efficace d'une part pour la compréhension de la base génétique de la physiologie de ce trait clinique important et d'autre part pour identifier de nouveaux gènes de susceptibilité au DT2.

Methicillin-resistant Staphylococcus aureus with intermediate susceptibility to vancomycin: are strict additional contact measures sufficient?

Background: A hospitalised patient infected with MRSA was found to harbour a VISA strain after 6 weeks of treatment with vancomycin. Additional contact measures were reinforced according to CDCs recommendations. We decide to evaluate if these applied control measures were effective. Objective: To evaluate the efficacy of strict additional contact measures to contain the dissemination of VISA from an infected patient. Methods: All patients from the unit were screened weekly for MRSA during a 6-week period, whereas health care workers (HCW) were screened only once. Screening specimen included nose, throat, groin, and clinical specimens for patients, and only nose and throat for HCW. Broth enrichment and chromogenic agar (MRSA-select) were used for MRSA detection. All MRSA isolates were tested on Van screen plates, and growing colonies were tested for MIC of vancomycin. MIC was performed using Etest. Population analysis was done for VISA confirmation. One strain per person was typed by Double Locus Sequence Typing (based on clfB and spa sequencing). Results: 66 patients hospitalized in the same service during the 6 weeks and 55 HCW were screened for MRSA and VISA. MRSA was found in 16/66 (24%) patients and 1/55 (2%) HCW. 16/17 MRSA from patients belonged to the same genotype that the VISA strain. The remaining patient had a MRSA identical to the HCW isolate. Among the 16 MRSA isolates sharing the same genotype than the VISA strain, two showed Etests vancomycin MIC of only 4 mg/L. MIC results were confirmed by the population analysis. They were not considered as VISA, but as MRSA with increased vancomycin MICs. Both isolates were obtained from two roommates. Conclusion: Strict additional contact measures were found to be effective to contain VISA dissemination. However, the identification of two isolates with increased vancomycin MIC (4 mg/L) in two roommates raised the question of the need to routinely test this susceptibility and of adequate control measures for patients harbouring such isolates.

Kernel-based Mapping of Meteorological Fields in Complex Orography

Due to the advances in sensor networks and remote sensing technologies, the acquisition and storage rates of meteorological and climatological data increases every day and ask for novel and efficient processing algorithms. A fundamental problem of data analysis and modeling is the spatial prediction of meteorological variables in complex orography, which serves among others to extended climatological analyses, for the assimilation of data into numerical weather prediction models, for preparing inputs to hydrological models and for real time monitoring and short-term forecasting of weather.In this thesis, a new framework for spatial estimation is proposed by taking advantage of a class of algorithms emerging from the statistical learning theory. Nonparametric kernel-based methods for nonlinear data classification, regression and target detection, known as support vector machines (SVM), are adapted for mapping of meteorological variables in complex orography.With the advent of high resolution digital elevation models, the field of spatial prediction met new horizons. In fact, by exploiting image processing tools along with physical heuristics, an incredible number of terrain features which account for the topographic conditions at multiple spatial scales can be extracted. Such features are highly relevant for the mapping of meteorological variables because they control a considerable part of the spatial variability of meteorological fields in the complex Alpine orography. For instance, patterns of orographic rainfall, wind speed and cold air pools are known to be correlated with particular terrain forms, e.g. convex/concave surfaces and upwind sides of mountain slopes.Kernel-based methods are employed to learn the nonlinear statistical dependence which links the multidimensional space of geographical and topographic explanatory variables to the variable of interest, that is the wind speed as measured at the weather stations or the occurrence of orographic rainfall patterns as extracted from sequences of radar images. Compared to low dimensional models integrating only the geographical coordinates, the proposed framework opens a way to regionalize meteorological variables which are multidimensional in nature and rarely show spatial auto-correlation in the original space making the use of classical geostatistics tangled.The challenges which are explored during the thesis are manifolds. First, the complexity of models is optimized to impose appropriate smoothness properties and reduce the impact of noisy measurements. Secondly, a multiple kernel extension of SVM is considered to select the multiscale features which explain most of the spatial variability of wind speed. Then, SVM target detection methods are implemented to describe the orographic conditions which cause persistent and stationary rainfall patterns. Finally, the optimal splitting of the data is studied to estimate realistic performances and confidence intervals characterizing the uncertainty of predictions.The resulting maps of average wind speeds find applications within renewable resources assessment and opens a route to decrease the temporal scale of analysis to meet hydrological requirements. Furthermore, the maps depicting the susceptibility to orographic rainfall enhancement can be used to improve current radar-based quantitative precipitation estimation and forecasting systems and to generate stochastic ensembles of precipitation fields conditioned upon the orography.

Isothermal microcalorimetry for antifungal susceptibility testing of Mucorales, Fusarium spp., and Scedosporium spp.

We evaluated isothermal microcalorimetry for real-time susceptibility testing of non-Aspergillus molds. MIC and minimal effective concentration (MEC) values of Mucorales (n = 4), Fusarium spp. (n = 4), and Scedosporium spp. (n = 4) were determined by microbroth dilution according to the Clinical Laboratory Standard Institute M38-A2 guidelines. Heat production of molds was measured at 37 °C in Sabouraud dextrose broth inoculated with 2.5 × 10(4) spores/mL in the presence of amphotericin B, voriconazole, posaconazole, caspofungin, and anidulafungin. As determined by microcalorimetry, amphotericin B was the most active agent against Mucorales (MHIC 0.06-0.125 μg/mL) and Fusarium spp. (MHIC 1-4 μg/mL), whereas voriconazole was the most active agent against Scedosporium spp. (MHIC 0.25 to 8 μg/mL). The percentage of agreement (within one 2-fold dilution) between the MHIC and MIC (or MEC) was 67%, 92%, 75%, and 83% for amphotericin B, voriconazole, posaconazole, and caspofungin, respectively. Microcalorimetry provides additional information on timing of antifungal activity, enabling further investigation of drug-mold and drug-drug interaction, and optimization of antifungal treatment.

Susceptibility of clinical isolates of Leishmania aethiopica to miltefosine, paromomycin, amphotericin B and sodium stibogluconate using amastigote-macrophage in vitro model.

Cutaneous Leishmaniasis (CL) caused by Leishmania aethiopica is a public health and social problem with a sequel of severe and mutilating skin lesions. It is manifested in three forms: localized CL (LCL), mucosal CL (MCL) and diffuse CL (DCL). Unresponsiveness to sodium stibogluconate (Sb(V)) is common in Ethiopian CL patients. Using the amastigote-macrophage in vitro model the susceptibility of 24 clinical isolates of L. aethiopica derived from untreated patients was investigated. Eight strains of LCL, 9 of MCL, and 7 of DCL patients together with a reference strain (MHOM/ET/82/117/82) were tested against four antileishmanial drugs: amphotericin B, miltefosine, Sb(V) and paromomycin. In the same order of drugs, IC(50) (μg/ml±SD) values for the 24 strains tested were 0.16±0.18, 5.88±4.79, 10.23±8.12, and 13.63±18.74. The susceptibility threshold of isolates originating from the 3 categories of patients to all 4 drugs was not different (p>0.05). Maximal efficacy was superior for miltefosine across all the strains. Further susceptibility test could validate miltefosine as a potential alternative drug in cases of sodium stibogluconate treatment failure in CL patients.

Storage and uptake of D-serine into astrocytic synaptic-like vesicles specify gliotransmission.

Glial cells are increasingly recognized as active players that profoundly influence neuronal synaptic transmission by specialized signaling pathways. In particular, astrocytes have been shown recently to release small molecules, such as the amino acids l-glutamate and d-serine as "gliotransmitters," which directly control the efficacy of adjacent synapses. However, it is still controversial whether gliotransmitters are released from a cytosolic pool or by Ca(2+)-dependent exocytosis from secretory vesicles, i.e., by a mechanism similar to the release of synaptic vesicles in synapses. Here we report that rat cortical astrocytes contain storage vesicles that display morphological and biochemical features similar to neuronal synaptic vesicles. These vesicles share some, but not all, membrane proteins with synaptic vesicles, including the SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) synaptobrevin 2, and contain both l-glutamate and d-serine. Furthermore, they show uptake of l-glutamate and d-serine that is driven by a proton electrochemical gradient. d-Serine uptake is associated with vesicle acidification and is dependent on chloride. Whereas l-serine is not transported, serine racemase, the synthesizing enzyme for d-serine, is anchored to the membrane of the vesicles, allowing local generation of d-serine. Finally, we reveal a previously unexpected mutual vesicular synergy between d-serine and l-glutamate filling in glia vesicles. We conclude that astrocytes contain vesicles capable of storing and releasing d-serine, l-glutamate, and most likely other neuromodulators in an activity-dependent manner.

NR2A at CA1 synapses is obligatory for the susceptibility of hippocampal plasticity to sleep loss.

A loss in the necessary amount of sleep alters expression of genes and proteins implicated in brain plasticity, but key proteins that render neuronal circuits sensitive to sleep disturbance are unknown. We show that mild (4-6 h) sleep deprivation (SD) selectively augmented the number of NR2A subunits of NMDA receptors on postsynaptic densities of adult mouse CA1 synapses. The greater synaptic NR2A content facilitated induction of CA3-CA1 long-term depression in the theta frequency stimulation range and augmented the synaptic modification threshold. NR2A-knock-out mice maintained behavioral response to SD, including compensatory increase in post-deprivation resting time, but hippocampal synaptic plasticity was insensitive to sleep loss. After SD, the balance between synaptically activated and slowly recruited NMDA receptor pools during temporal summation was disrupted. Together, these results indicate that NR2A is obligatory for the consequences of sleep loss on hippocampal synaptic plasticity. These findings could advance pharmacological strategies aiming to sustain hippocampal function during sleep restriction.

Loss of mating flight and shift in the pattern of carbohydrate storage in sexuals of ants (Hymenoptera; Formicidae)

In ants, energy for flying is derived from carbohydrates (glycogen and free sugars). The amount of these substrates was compared in sexuals participating or not participating in mating flights. Results show that in participating females (Lasius niger, L. flavus, Myrmica scabrinodis, Formica rufa, F. polyctena, F. lugubris), the amount of carbohydrates, especially glycogen, was higher than in non-participating females (Cataglyphis cursor, Iridomyrmex humilis). Similarly, male C. cursor and I. humilis which fly, exhibit a much higher carbohydrate content than do the non-flying females of these species. Furthermore, the quantity of carbohydrates stored was generally higher in males than in females for each species. These results are discussed with regard to the loss of the nuptial flight by some species of ants.

Susceptibility and adaptation to human TRIM5α alleles at positive selected sites in HIV-1 capsid.

Numerous in vitro studies attribute to human TRIM5α some modest anti-HIV-1 activity and human population studies suggest some differential effect of TRIM5α polymorphisms on disease progression. If the activity of TRIM5α were relevant in vivo, it could result in positive selection on the viral capsid. To address this issue, we identified 10 positively selected sites in HIV-1 capsid from multiple viral strains and generated 17 clade B viruses carrying a minor (i.e. low frequency) residue or an alanine at those positions. All recombinant viruses were susceptible to the modest effect of common human TRIM5α and allelic variants R136Q, and H419Y; H43Y and G249D TRIM5α were generally inactive. Increased sensitivity to TRIM5α was observed for some capsid variants, suggesting that minor residues are selected against in human populations. On the other hand, the modest potency of human TRIM5α does not translate in escape mutations in the viral capsid.