L008 Défaut de différenciation veino-lymphatique embyonnaire par modulation de l’ARN interférence

L’ARN interférence, mécanisme de régulation de l’expression des gènes, est médiée par les siARNs et les microARNs, ARN non-codants de 20 à 22 nucléotides affectant la régulation post-transcriptionnelle d’ARNm cibles avec lesquels ils s’apparient.La RNase DICER est une enzyme centrale de la biosynthèse des siARNs et microARNs. Les souris dont le gène dicer est invalidé ont un phénotype complexe, et meurent très tôt pendant le développement, notamment à cause d’un défaut d’angiogenèse.Afin d’étudier l’ARN interférence au cours de l’angiogenèse embryonnaire, des souris dont le gène dicer est floxé (mutant conditionnel) sont croisées avec des souris exprimant la recombinase Cre, de manière constitutive, sous le contrôle du promoteur du gène tie2, dirigeant ainsi son expression dans les cellules endothéliales (CE) et les cellules hématopiétiques.Nos résultats montrent que l’invalidation de dicer sous le contrôle du promoteur du gène tie2 entraine une mortalité embryonnaire suite à un œdème et des hémorragies au treizième jour du développement (E13,5). L’analyse histologique montre des vaisseaux lymphatiques remplis de sang, suggérant une mauvaise séparation du réseau sanguin et lymphatique. Cette hypothèse est étudiée par marquage des vaisseaux lymphatiques (LYVE-1) et des vaisseaux sanguins (PECAM) sur embryon entier et peaux isolées à différents stades précédant la mort.Ces embryons présentent également un problème de développement du foie, probablement dû à l’activité du promoteur tie2 dans les lignées hématopoiétiques. La mise en culture de ces foies fœtaux à E13,5 révèle une atteinte des précurseurs hématopoétiques.L’étude de ces précurseurs à des stades plus précoces (E8,5) est en cours au laboratoire.Nos résultats démontrent donc un rôle important de l’ARN interférence dans le contrôle épigénétique de l’angiogenèse et de la lymphangiogenèse embryonnaire mais également dans le développement de l’hématopoièse, suggérant son implication dans la différenciation veino-lymphangiogenèse, dont les mécanismes moléculaires seront discutés

An activating mutation in the CSF3R gene induces a hereditary chronic neutrophilia

We identify an autosomal mutation in the CSF3R gene in a family with a chronic neutrophilia. This T617N mutation energetically favors dimerization of the granulocyte colony-stimulating factor (G-CSF) receptor transmembrane domain, and thus, strongly promotes constitutive activation of the receptor and hypersensitivity to G-CSF for proliferation and differentiation, which ultimately leads to chronic neutrophilia. Mutant hematopoietic stem cells yield a myeloproliferative-like disorder in xenotransplantation and syngenic mouse bone marrow engraftment assays. The survey of 12 affected individuals during three generations indicates that only one patient had a myelodysplastic syndrome. Our data thus indicate that mutations in the CSF3R gene can be responsible for hereditary neutrophilia mimicking a myeloproliferative disorder

Gene profiling of the erythro- and megakaryoblastic leukaemias induced by the Graffi murine retrovirus

<p>Abstract</p> <p>Background</p> <p>Acute erythro- and megakaryoblastic leukaemias are associated with very poor prognoses and the mechanism of blastic transformation is insufficiently elucidated. The murine Graffi leukaemia retrovirus induces erythro- and megakaryoblastic leukaemias when inoculated into NFS mice and represents a good model to study these leukaemias.</p> <p>Methods</p> <p>To expand our understanding of genes specific to these leukaemias, we compared gene expression profiles, measured by microarray and RT-PCR, of all leukaemia types induced by this virus.</p> <p>Results</p> <p>The transcriptome level changes, present between the different leukaemias, led to the identification of specific cancerous signatures. We reported numerous genes that may be potential oncogenes, may have a function related to erythropoiesis or megakaryopoiesis or have a poorly elucidated physiological role. The expression pattern of these genes has been further tested by RT-PCR in different samples, in a Friend erythroleukaemic model and in human leukaemic cell lines.</p> <p>We also screened the megakaryoblastic leukaemias for viral integrations and identified genes targeted by these integrations and potentially implicated in the onset of the disease.</p> <p>Conclusions</p> <p>Taken as a whole, the data obtained from this global gene profiling experiment have provided a detailed characterization of Graffi virus induced erythro- and megakaryoblastic leukaemias with many genes reported specific to the transcriptome of these leukaemias for the first time.</p

Microhardness variation and related microstructure in Al–Cu alloys prepared by HF induction melting and RF sputtering

The materials under consideration are binary aluminium-copper alloys (10 at% to 90.3 at%Cu) produced by HF melting and RF magnetron sputtering. The resulting micro structures have been observed by standard metallographic techniques, X-ray powder diffraction, scanning electron microscopy and transmission electron microscopy. Vickers microhardness of bulk Al–Cu alloys reaches a maximum of 1800 MPa at 70.16 at%Cu. An unexpected metastable $\theta '$ phase has been observed within aluminium grain in Al-37 at%Cu. The mechanical properties of a family of homogeneous Al$_{1-x}$Cux (0 < x < 0.92) thin films made by radiofrequency (13.56 MHz) cathodic magnetron sputtering from composite Al–Cu targets have been investigated. The as-deposited microstructures for all film compositions consisted of a mixture of the two expected face-centred-cubic (fcc) Al solid solution and tetragonal θ (Al2Cu) phases. The microhardness regularly increases and the grain size decreases both with copper concentration. This phenomenon of significant mechanical strengthening of aluminium by means of copper is essentially due to a combination between solid solution effects and grain size refinement. This paper reports some structural features of different Al–Cu alloys prepared by HF melting and RF magnetron on glass substrate sputtering

Thrombopoietin dose-response curves reflect the number of doublings undergone by human megakaryocyte progenitors

audience: researcherTo assess the variation of thrombopoietin (TPO) responsiveness associated with MK progenitor amplification, TPO dose-response curves were obtained for MK clones derived from normal human CD34+CD41+ cells which had been single-cell plated in serum-deprived medium with TPO concentrations of 0-1000 pg/mL per well. On day 5, the number of MK cells per well was determined in situ and expressed as number of doublings (NbD). Dose-response curves of the means of the frequency of clones of each size vs logTPO concentration showed highly significant differences in the TPO concentration needed for half-maximum generation of clones of different sizes (TPO50). Based on 3 independent experiments, TPO50, expressed in pg/mL, was 1.89 ±SD 0.87 for 1 cell MK clone; 7.75 ±1.40 for 2-3 cell MK clone; 38.5 ±8.73 for 4-7 cell MK clone and 93.6 ±27.7 for 8-15 cell MK clone (p<0.001). Compared to the CD42- fraction of CD34+CD41+ progenitors, the corresponding CD42+ fraction had a lower TPO50 (p<0.001), underwent lesser NbD (p<0.001) and expressed a 10-fold greater mean mpl receptor density. In order to interpret the conclusion that the most responsive progenitors were those which proliferated less, we tested a prediction of the generation-age model, namely that the percentage of progenitors undergoing a given number of residual doublings in vitro (NbDr) depends on the number of previous doublings they had undergone in vivo (NbDp). It was demonstrated that logTPO50 had a positive correlation with NbDr and negative correlation with NbDp (p<0.001). The reciprocal relationship between NbDp and NbDr was evidence for a generation-age model in which TPO responsiveness increases by a factor of approximately 4 per doubling. This model suggests that progenitors which have completed their proliferative program have maximum factor responsiveness and are preferentially induced to terminal differentiation