Role of TLR1, TLR2 and TLR6 in the modulation of intestinal inflammation and Candida albicans elimination

Toll-like receptors (TLRs) are the major pattern recognition receptors that mediate sensing of a wide range of microorganisms. TLR2 forms heterodimers with either TLR1 or TLR6, broadening its ligand diversity against pathogens. TLR1, TLR2 and TLR6 have been implicated in the recognition of Candida albicans, an opportunistic fungal pathogen that colonizes the gastrointestinal tract. In this study, we explored whether the deficiency in TLR1, TLR2 or TLR6 impacts C. albicans colonization and inflammation-associated colonic injury in the dextran sulfate sodium (DSS)-induced colitis in mice. DSS treatment and C. albicans challenge induced greater weight loss, worse clinical signs of inflammation, higher histopathologic scores, and increased mortality rates in TLR1(-/-) and TLR2(-/-) mice when compared to TLR6(-/-) and wild-type mice. The number of C. albicans colonies in the stomach, colon and feces was decreased in TLR6(-/-) mice as compared to TLR2(-/-), TLR1(-/-) and wild-type mice. Interestingly, the population of E. coli in colonic luminal contents, intestinal permeability to FITC-dextran and cytokine expression were significantly increased in TLR1(-/-) and TLR2(-/-) mice, while they were decreased in TLR6(-/-) mice. In contrast to TLR6, both TLR1 and TLR2 deficiencies increased intestinal inflammation, and the overgrowth of C. albicans and E. coli populations in the colitis model, suggesting the involvement of TLR1 and TLR2 in epithelial homeostasis, and a role of TLR6 in increasing intestinal inflammation in response to pathogen-sensing

The first sawfly from the Oligocene of Céreste (Southern France) (Hymenoptera: Tenthredinidae)

Luberotenthredo cerestensis gen. et sp. nov. is the first record of the sawfly family Tenthredinidae from the Oligocene of Céreste (Southern France). This taxon is described and illustrated based on a well-preserved specimen. This genus resembles the extant genus Perineura (subfamily Tenthredininae, tribe Perineurini) with which it shares forewing venation similarities and numerous morphological characters. This new taxon is the first fossil representative of the tribe Perineurini and can be used as a calibration point for future investigation of the diversification of the family Tenthredinidae

Quantification des "F-cells" par cytométrie en flux, comparaison avec la mesure de l'hémoglobine F pr chromatographie en milieu liquide (étude de 75 patients drépanocytaires)

Les F-cells sont définies comme des globules rouges matures contenant une proportion variable d'hémoglobine fœtale, la concentration d'HbF dans ces cellules restant toujours minoritaire par rapport à celle des autres hémoglobines. Chez le patient drépanocytaire, l'hémoglobine F en ne s'incorporant pas dans le polymère d'hémoglobine S en interrompt la polymérisation et diminue ainsi le phénomène de falciformation caractéristique de la maladie. Le traitement par Hydroxyurée a pour but d'augmenter le pourcentage d'HbF fœtale et c'est pour cette raison que la détermination de l'HbF et des F-cells est un atout essentiel au suivi de ces patients. Nous avons donc étudié une méthode par cytométrie en flux pour la quantification des F-cells et comparé ces résultats à ceux de l'HbF obtenus par chromatographie en milieu liquide chez 75 patients drépanocytaires répartis en trois groupes (hétérozygotes, homozygotes non traités et homozygotes traités par Hydroxyurée). Cette étude a montré une excellente corrélation entre les deux techniques (r = 0,96). Les taux de F-cells étaient significativement plus élevés chez les patients traités. Il a été retrouvé une grande variation interindividuelle des pourcentages de F-cells chez les patients homozygotes. La quantification des F-cells associée à celle de l'HbF a permis d'évaluer le contenu en HbF par F-cells. Une étude de ces paramètres associée à la quantification des F-réticulocytes permettrait une approche plus complète pour le suivi des sujets drépanocytaires traités par HydroxyuréePARIS12-CRETEIL BU Médecine (940282101) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

La numération des érythroblastes circulants, étude comparative de 133 échantillons par trois méthodes (automate compte-globules, cytométrie en flux, microscopie optique)

PARIS-BIUP (751062107) / SudocSudocFranceF

h-Goliath, paralog of GRAIL, is a new E3 ligase protein, expressed in human leukocytes.

In Drosophila, the RING finger protein d-Goliath was originally identified as a transcription factor involved in the embryo mesoderm formation [Bouchard, M.L., Cote, S., 1993. The Drosophila melanogaster developmental gene g1 encodes a variant zinc-finger-motif protein. Gene 125, 205-209]. In mouse, the m-Goliath mRNA level was shown to be increased in growth factor withdrawal-induced apoptosis of myeloid cells [Baker, S.J., Reddy, E.P., 2000. Cloning of murine G1RP, a novel gene related to Drosophila melanogaster g1. Gene 248, 33-40]. Due to its putative function of transcription factor in apoptosis, we cloned the human cDNA for h-Goliath and characterized the expression of the protein in blood and bone marrow cells. The human protein of 419 aa (44 kDa) contains a protease-associated domain, a transmembrane domain and a RING-H2 motif. This structure classifies h-Goliath as a new member of a human family of ubiquitin ligases with GRAIL (gene related to anergy in lymphocytes) as founder. This E3 ligase controls the development of T cell clonal anergy by ubiquitination [Anandasabapathy, N., Ford, G.S., Bloom, D., Holness, C., Paragas, V., Seroogy, C., Skrenta, H., Hollenhorst, M., Fathman, C.G., Soares, L., 2003. GRAIL: an E3 ubiquitin ligase that inhibits cytokine gene transcription is expressed in anergic CD4+ T cells. Immunity 18, 535-547]. In vitro ubiquitination studies support the E3 ubiquitin ligase activity of h-Goliath. In human, the protein is expressed under 3 isoforms, a major one at 28 kDa and two others at 46 and 55 kDa. These proteins come from a common precursor (44 kDa) as we observed using in vitro transcription-translation. Using immunohistochemistry on blood or bone marrow smears, of healthy or leukemia samples, we found that the protein expression was restricted to the cytoplasm of progenitors and fully differentiated leukocyte populations. We did not observe any modification of h-Goliath expression or localization in leukemia. In these cells, this new E3 ubiquitin ligase protein does not seem associated with a differentiation state of the cell or with apoptosis

Erythrocyte density in sickle cell syndromes is associated with specific clinical manifestations and hemolysis.

International audienceDense, dehydrated red blood cells (DRBCs) are a characteristic feature of sickle-cell disease (SCD). DRBCs play a role in the pathophysiology of SCD acute and chronic organ damage because of heightened tendency to undergo polymerization and sickling because of their higher hemoglobin S concentration. Relations between red cell density (assessed with phthalate density-distribution profile method) and several hematologic, biochemical, genetic parameters, and clinical manifestations were studied in a large cohort of homozygous patients. The percentage of DRBCs was significantly higher in patients who experienced skin ulcers, priapism, or renal dysfunction. Presence of α-thalassemia deletions was associated with fewer DRBCs. A multivariable analysis model showed DRBCs to be positively associated with hemolytic parameters such as lactate dehydrogenase and bilirubin and negatively with fetal hemoglobin. The percentage of DRBCs decreased by 34% at 6 months of hydroxycarbamide (xydroxyurea) therapy. Thus, DRBCs are associated with specific clinical manifestations and biologic markers and may be a useful addition to the biologic and clinical evaluation of patients with SCD, because they can easily be measured in a hematocrit tube

A mouse model provides evidence that genetic background modulates anemia and liver injury in erythropoietic protoporphyria

International audienceErythropoietic protoporphyria is an inherited disorder of heme biosynthesis caused by partial ferrochelatase deficiency, resulting in protoporphyrin (PP) overproduction by erythrocytes. In humans, it is responsible for painful skin photosensitivity and, occasionally, liver failure due to accumulation of PP in the liver. The ferrochelatase deficiency mouse mutation is the best animal model available for human erythropoietic protoporphyria. The original description, based on mice with a BALB/cByJCrl genetic background, reported a disease resembling the severe form of the human disease, with anemia, jaundice, and liver failure. Using congenic strains, we investigated the effect of genetic background on the severity of the phenotype. Compared with BALB/cByJCrl, C57BL/6JCrl mice developed moderate but increasing anemia and intense liver accumulation of PP with severe hepatocyte damage and loss. Bile excretory function was not affected, and bilirubin remained low. Despite the highest PP concentration in erythrocytes, anemia was mild and there were few PP deposits in the liver in SJL/JOrlCrl homozygotes. Discriminant analysis using six hematologic and biochemical parameters showed that homozygotes of the three genetic backgrounds could be clustered in three well-separated groups. These three congenic strains provide strong evidence for independent genetic control of bone marrow contribution of PP overproduction to development of liver disease and biliary PP excretion. They provide a tool to investigate the physiological mechanisms involved in these phenotypic differences and to identify modifying genes

Four- and five-color flow cytometry analysis of leukocyte differentiation pathways in normal bone marrow: a reference document based on a systematic approach by the GTLLF and GEIL.

International audienceBACKGROUND: The development of multiparameter flow cytometry (FCM) and increasingly sophisticated analysis software has considerably improved the exploration of hematological disorders. These tools have been widely applied in leukaemias, lymphomas, and myelodysplasias, yet with very heterogeneous approaches. Consequently, there is no extensive reference document reporting on the characteristics of normal human bone marrow (BM) in multiparameter FCM. Here, we report a reference analysis procedure using relevant antibody combinations in normal human BM. METHODS: A first panel of 23 antibodies, constructed after literature review, was tested in four-color combinations (including CD45 in each) on 30 samples of BM. After evaluation of the data, a second set of 22 antibodies was further applied to another 35 BM samples. All list-modes from the 65 bone marrow samples were reviewed collectively. A systematised protocol for data analysis was established including biparametric representations and color codes for the three major lineages and undifferentiated cells. RESULTS: This strategy has allowed to obtain a reference atlas of relevant patterns of differentiation antigens expression in normal human BM that is available within the European LeukemiaNet. This manuscript describes how this atlas was constructed. CONCLUSIONS: Both the strategy and atlas could prove very useful as a reference of normality, for the determination of leukemia-associated immunophenotypic patterns, analysis of myelodysplasia and, ultimately, investigation of minimal residual disease in the BM

Characterization of New Oligosaccharides Obtained by An Enzymatic Cleavage of the Exopolysaccharide Produced by the Deep-Sea Bacterium Alteromonas infernus Using its Cell Extract

Bacteria from deep-sea hydrothermal vents constitute an attractive source of bioactive molecules. In particular, exopolysaccharides (EPS) produced by these bacteria become a renewable source of both biocompatible and biodegradable molecules. The low molecular weight (LMW) derivatives of the GY785 EPS produced by the deep-sea hydrothermal vent strain Alteromonas infernus have previously displayed some biological properties, similar to those of glycosaminoglycans (GAG), explored in cancer and tissue engineering. These GAG-mimetic derivatives are obtained through a free radical depolymerization process, which could, however, affect their structural integrity. In a previous study, we have shown that A. infernus produces depolymerizing enzymes active on its own EPS. In the present study, an enzymatic reaction was optimized to generate LMW derivatives of the GY785 EPS, which could advantageously replace the present bioactive derivatives obtained by a chemical process. Analysis by mass spectrometry of the oligosaccharide fractions released after enzymatic treatment revealed that mainly a lyase activity was responsible for the polysaccharide depolymerization. The repeating unit of the GY785 EPS produced by enzyme cleavage was then fully characterized