[Clinical and biological features of patients with essential thrombocythaemia according to their mutational status JAK2 or CALR: Single-center study of 40 patients and review of the literature]. / Caractéristiques clinico-biologiques des patients avec thrombocytémie essentielle en fonction de leur statut mutationnel JAK2 ou CALR : étude monocentrique de 40 patients et revue de la littérature.

BACKGROUND: Somatic mutations in the calreticulin gene (CALR) were recently described in essential thrombocythemia (ET) and primary myelofibrosis with non-mutated JAK2 or MPL. The aim of this single-center study was to compare the clinical and biological features of ET patients according to their mutational status. METHODS: We included 40 patients with ET followed in hematology consultation. The JAK2 V617F mutation was assessed by quantitative PCR. For the detection of CALR mutations, we performed a PCR amplification of CALR exon 9 followed by direct sequencing. RESULTS: Among 40 study patients, 23 (57.5%) harbored V617F JAK2, 12 of the 17 patients without JAK2 mutation harbored CALR, no patient expressed MPL mutation and 5 were negative for all three mutations. Five types of mutations were identified with predominance of 52bp deletion and 5bp insertion (7/12 and 2/12 respectively). The incidence of thrombotic events at diagnosis was significantly higher in JAK2 mutated patients (P<0.05). Biologically, patients with CALR mutation had significantly higher platelet count (P<0.01) and significantly lower hemoglobin level (P<0.05) than those with V617F JAK2 mutation. CONCLUSION: JAK2 and CALR mutation screening in ET has a diagnostic value. Each mutation displays a distinct phenotype with uncertain impact on long-term outcome.

Compositional variations in In(0.5)Ga(0.5)N nanorods grown by molecular beam epitaxy.

The composition of InxGa1 - xN nanorods grown by molecular beam epitaxy with nominal x = 0.5 has been mapped by electron microscopy using Z-contrast imaging and x-ray microanalysis. This shows a coherent and highly strained core-shell structure with a near-atomically sharp boundary between a Ga-rich shell (x âˆ¼ 0.3) and an In-rich core (x âˆ¼ 0.7), which itself has In- and Ga-rich platelets alternating along the growth axis. It is proposed that the shell and core regions are lateral and vertical growth sectors, with the core structure determined by spinodal decomposition.

End-to-end learning with interpretation on electrohysterography data to predict preterm birth.

Prediction of preterm birth is a difficult task for clinicians. By examining an electrohysterogram, electrical activity of the uterus that can lead to preterm birth can be detected. Since signals associated with uterine activity are difficult to interpret for clinicians without a background in signal processing, machine learning may be a viable solution. We are the first to employ Deep Learning models, a long-short term memory and temporal convolutional network model, on electrohysterography data using the Term-Preterm Electrohysterogram database. We show that end-to-end learning achieves an AUC score of 0.58, which is comparable to machine learning models that use handcrafted features. Moreover, we evaluate the effect of adding clinical data to the model and conclude that adding the available clinical data to electrohysterography data does not result in a gain in performance. Also, we propose an interpretability framework for time series classification that is well-suited to use in case of limited data, as opposed to existing methods that require large amounts of data. Clinicians with extensive work experience as gynaecologist used our framework to provide insights on how to link our results to clinical practice and stress that in order to decrease the number of false positives, a dataset with patients at high risk of preterm birth should be collected. All code is made publicly available.

Cytomegalovirus-specific regulatory and effector T cells share TCR clonality--possible relation to repetitive CMV infections.

Cytomegalovirus (CMV) infections have a major impact on morbidity and mortality of transplant patients. Among the complex antiviral T-cell response, CMV-IE-1 antigen-specific CD8+ cells are crucial for preventing CMV disease but do not protect from recurring/lasting CMV reactivation. Recently, we confirmed that adoptive transfer of autologous IE-1/pp65-specific T-cell lines was able to combat severe CMV disease; however, the control of CMV infection was only temporary. We hypothesized that CMV-induced regulatory T cells (iTreg) might be related to recurring/lasting CMV infection. In fact, kidney transplant patients with recurring CMV infections expressed enhanced suppression on CMV response. Analysis of in vitro expanded CD4+ epitope-specific cells revealed that CMV-specific CD4+CD25(high) Treg cells functionally suppress CD25(low) effector T cells (Teff) upon epitope-specific reactivation. Their phenotype is similar to iTreg - CD39(high) /Helios-/IL-2(low) /IFNγ(high) /IL-10±/TGFß-LAP±/FOXP3+ and methylated foxp3 locus. Remarkably, in vitro expanded CD4+CD25(high) iTreg share the same dominant TCR-Vß-CDR3 clones with functionally distinct CD4+CD25(low) Teff. Moreover, the same clones were present in freshly isolated CD4+CD25(high) and CD4+CD25(low) T cells suggesting their in vivo generation. These findings directly demonstrate that Teff and iTreg can differentiate from one "mother" clone with specificity to the same viral epitope and indicate that peripheral iTreg generation is related to frequent antigen appearance.

Arterial and venous thrombosis is associated with different angiogenic cytokine patterns in patients with antiphospholipid syndrome.

Antiphospholipid syndrome (APS) is defined as a combination of antiphospholipid antibodies, arterial and/or venous thrombosis, and, in women, recurrent fetal loss. The mechanisms underlying this prothrombotic tendency are unclear. Here we determined plasma levels of the angiogenic growth factors vascular endothelial growth factor (VEGF), placental growth factor (PlGF) and stromal cell-derived factor-1 (SDF-1) in 34 patients with APS (median age 40 years) compared with 180 healthy controls and with 80 age-matched deep-venous thrombosis patients in whom the diagnosis of APS had been excluded. All of the patients met updated APS criteria and two-thirds of them were triply positive for antiphospholipid antibodies (lupus anticoagulant, anti-beta2-glycoprotein I and anti-cardiolipin antibodies). Angiogenic cytokines were quantified at least 6 months after an acute thrombotic event. VEGF levels were similar in the patients and controls, but were significantly higher in the patients with arterial thrombosis than in the patients with venous thrombosis. Plasma levels of SDF-1 and PlGF were significantly elevated in the patients, regardless of the arterial/venous nature of the thrombosis. Together, these results suggest that APS is associated with an angiogenic process, but that the angiogenic signal differs between patients with arterial and venous thrombosis. Lupus (2010) 19, 837-843.

SEM characterization of silicon nanostructures: can we meet the challenge?

The current semiconductor technology road map for device scaling champions a 4.5 nm gate length in production by 2022. The scanning electron microscope (SEM) as applied to critical dimensions (CD) metrology and associated characterization modes such as electron beam-induced current and cathodoluminescence (CL) has proved to be a workhorse for the semiconductor industry during the microelectronics era. We review some of the challenges facing these techniques in light of the silicon nanotechnology road map. We present some new results using voltage contrast imaging and CL spectroscopy of top-down fabricated silicon nanopillar/nanowires (<100 nm diameter), which highlight the visualization challenge. However, both techniques offer the promise of providing process characterization on the 10-20 nm scale with existing technology. Visualization at the 1 nm scale with these techniques may have to wait for aberration-corrected SEM to become more widely available. Basic secondary electron imaging and CD applications may be separately addressed by the He-ion microscope.

Control of human B cell tumor growth in severe combined immunodeficiency mice by monoclonal anti-B cell antibodies.

Severe combined immunodeficiency (scid) mice develop EBV (+)B cell tumors after infusion of EBV(+)B cells or of B cells and EBV. In this study, scid mice were infused with B cell lines derived from three patients who developed a B lymphocyte proliferative disorder after bone marrow or organ transplantation. Intraperitoneal injection of 5 x 10(6) B cells induced tumor growth in all mice, leading to death within 60 d. Human B cells were identified in spleen and bone marrow by means of immunofluorescence or EBV genome amplification, and human IgM was detected in serum. Infusion of murine monoclonal antibodies specific for human B cell membrane antigens CD21, CD24, and CD23 was effective in 80% of animals, against two of the three cell lines preventing tumor development or inducing remission according to the time of treatment. The effect was antibody dose dependent and was optimal with four intravenous infusions of at least 0.1 mg 4 d apart. Human IgM in serum and human B cells in spleen and bone marrow became undetectable when peritoneal tumors regressed completely. Infusions of IgG1 isotype-matched anti-CD4 antibody or anti-CD3 antibody had no effect. Tumors developed or recurred in 50% of these animals injected with one of the B cell line 3 mo after treatment was stopped. The same anti-CD21 and anti-CD24 antibodies had been used to treat the three patients, and shown similar degrees of effectiveness as in the scid mouse model. These results indicate that scid mice may be suitable for assessing therapeutic approaches to human B cell proliferation.

Molecular characterization of antithrombin III (ATIII) variants using polymerase chain reaction. Identification of the ATIII Charleville as an Ala 384 Pro mutation.

The genes of seven structural mutants of antithrombin III (ATIII), presenting either defective serine protease reactivity or abnormal heparin binding, were analyzed. The polymerase chain reaction (PCR) was used to amplify the corresponding gene exon and the mutation was identified by either dot blot analysis using a battery of allele-specific oligonucleotide probes or sequencing. Variants Paris and Paris 2 were identified as Arg 47 Cys mutations, and Clichy, Clichy 2, and Franconville were found to be Pro 41 Leu mutations. All five are heparin binding-site variants. ATIII Avranches is an Arg 393 His mutation and ATIII Charleville is an Ala 384 Pro mutation. These two mutations impair the reactive site of the molecule. ATIII Charleville is a new mutation of the reactive center, as predicted by previous biochemical data. The position of this new mutation, together with the other previously described mutations of the reactive center, sheds light on the molecular function of this site in inhibiting thrombin. Finally, genomic amplification by PCR is a powerful technique for the fast identification of antithrombin III mutations and their homozygous/heterozygous status, and should be useful for predicting thrombotic risk.

[State of the art: Direct oral anticoagulants and transfusion]. / État de l'art : nouveaux anticoagulants et transfusion.

Direct oral anticoagulants (DOAC) are indicated for stroke prevention in atrial fibrillation and for the prevention and treatment of venous thromboembolism. As any anticoagulant, they are associated with a bleeding risk. Management of DOAC-induced bleeding is challenging. Idarucizumab, antidote for dabigatran, is currently available and is part of the therapeutic strategy, whereas antidotes for anti-Xa agents are under development. Activated or non-activated prothrombin concentrates are proposed, although their efficacy to reverse DOAC is uncertain. We propose an update on DOAC-associated bleeding management, integrating the availability of idarucizumab and the critical place of DOAC concentration measurements.

Roles of "Wanting" and "Liking" in Motivating Behavior: Gambling, Food, and Drug Addictions.

The motivation to seek out and consume rewards has evolutionarily been driven by the urge to fulfill physiological needs. However in a modern society dominated more by plenty than scarcity, we tend to think of motivation as fueled by the search for pleasure. Here, we argue that two separate but interconnected subcortical and unconscious processes direct motivation: "wanting" and "liking." These two psychological and neuronal processes and their related brain structures typically work together, but can become dissociated, particularly in cases of addiction. In drug addiction, for example, repeated consumption of addictive drugs sensitizes the mesolimbic dopamine system, the primary component of the "wanting" system, resulting in excessive "wanting" for drugs and their cues. This sensitizing process is long-lasting and occurs independently of the "liking" system, which typically remains unchanged or may develop a blunted pleasure response to the drug. The result is excessive drug-taking despite minimal pleasure and intense cue-triggered craving that may promote relapse long after detoxification. Here, we describe the roles of "liking" and "wanting" in general motivation and review recent evidence for a dissociation of "liking" and "wanting" in drug addiction, known as the incentive sensitization theory (Robinson and Berridge 1993). We also make the case that sensitization of the "wanting" system and the resulting dissociation of "liking" and "wanting" occurs in both gambling disorder and food addiction.

Mechanism of factor IXa inhibition by antithrombin in the presence of unfractionated and low molecular weight heparins and fucoidan.

Heparin exerts its anticoagulant activity by catalysing the inhibition of coagulation proteases by antithrombin (AT). Its main target is thrombin but it also catalyses the inhibition of the other serine-proteases of the coagulation cascade, such as factor IXa (fIXa). The aim of this study was to compare the catalysis of inhibition of blood fIXa by antithrombin in the presence of several sulfated polysaccharides with anticoagulant activity, i.e. heparin, three widely used in therapeutics low molecular weight heparins (LMWH) and fucoidan. Plots of the second-order rate constants of the fIXa-antithrombin reaction vs. the concentration of added heparin and LMWH are bell-shaped and fit the kinetic model established for thrombin-antithrombin reaction by Jordan R., Beeler D., Rosenberg R. (1979) J. Biol. Chem., 254, 2902-2913. In the ascending branch, the catalyst (C) binds quickly to the inhibitor (I) to form a catalyst-inhibitor (CI) complex which is more reactive towards the enzyme (E) than the free inhibitor, leading to the formation of an inactive enzyme-inhibitor complex (EI) and the release of free catalyst, in a rate-limiting second step. After a maximum corresponding to an optimal catalyst concentration, the decrease in the reaction rate was in keeping with the formation of a catalyst-enzyme (CE) complex, whose inactivation by the CI complex was slower than that of the free enzyme. Maximum second-order rate constants for the inhibition of fIXa by AT were 105, 6.8, 12.24 and 22 microM-1 min-1 with heparin, Enoxaparin, Fraxiparin and Fragmin, respectively, leading to 3500-, 225-, 405- and 728-fold increases in the inhibition rate in the absence of polysaccharide, respectively. Fucoidan yielded 23-fold increase in the fIXa-antithrombin interaction rate. The kinetic profiles obtained with this polysaccharide exhibited ascending branch which correlated well with the kinetic model based on the formation of binary complexes (CI or CE). Fucoidan was covalently conjugated with a fluorescent probe (DTAF) and used in conjunction with fluorescence anisotropy to follow its binding to antithrombin, heparin cofactor II (HCII), thrombin and fIXa. The binding of fucoidan to these proteins occurred with low affinities when compared to heparin and LMWH. Fucoidan had higher affinity for the inhibitor HCII compared to antithrombin and enzymes. These data suggest that binding of heparins and fucoidan to the inhibitor (CI) is required for the polysaccharide-dependent enhancement in the rate of neutralization of the enzyme by the inhibitor.

Characterization, chemical modifications and in vitro anticoagulant properties of an exopolysaccharide produced by Alteromonas infernus.

A new low-molecular-weight 'heparin-like' component was obtained from an exopolysaccharide produced by a mesophilic strain found in deep-sea hydrothermal vents. Data concerning the structure of the native high-molecular-weight exopolysaccharide (10(6) g/mol, 10% sulfate content) are reported for the first time. Two depolymerization processes were used to obtain low-molecular-weight (24-35x10(3) g/mol) oversulfated fractions (sulfate content 20 or 40%). Nuclear magnetic resonance studies indicated that after sulfation (40%), the low-molecular-weight fraction obtained by free radical depolymerization was less sulfated in the 6-O-position than the fraction depolymerized by acid hydrolysis. The free radical depolymerized product also had sulfated residues in the 4-O-position and disulfated ones in the 2,3-O-positions. Moreover, the compounds generated by the free radical process were more homogeneous with respect to molecular mass. Also for the first time, the anticoagulant activity of the low-molecular-weight exopolysaccharide fractions is reported. When the fractions obtained after sulfation and depolymerization were compared with heparins, anticoagulant activity was detected in oversulfated fractions, but not in native exopolysaccharide. The free radical depolymerized fraction inhibited thrombin generation in both contact-activated and thromboplastin-activated plasma, showing a prolonged lag phase only in the contact-activated assay. Affinity co-electrophoresis studies suggested that a single population of polysaccharide chains binds to antithrombin and that only a subpopulation strongly interacts with heparin cofactor II.

Modulation of human endothelial cell proliferation and migration by fucoidan and heparin.

Fucoidan is a sulfated polysaccharide extracted from brown seaweeds. It has anticoagulant and antithrombotic properties and inhibits, as well as heparin, vascular smooth muscle cell growth. In this study, we investigated, in the presence of serum and human recombinant growth factors, the effects of fucoidan and heparin on the growth and migration of human umbilical vein endothelial cells (HUVEC) in culture. We found that fucoidan stimulated fetal bovine serum-induced HUVEC proliferation, whereas heparin inhibited it. In the presence of fibroblast growth factor-1 (FGF-1), both fucoidan and heparin potentiated HUVEC growth. In contrast, fucoidan and heparin inhibited HUVEC proliferation induced by FGF-2, but did not influence the mitogenic activity of vascular endothelial growth factor (VEGF). In the in vitro migration assay from a denuded area of confluent cells, the two sulfated polysaccharides markedly enhanced the migration of endothelial cells in the presence of FGF-1. Finally, a weak inhibitory effect on cell migration was found only with the two polysaccharides at high concentrations (> or = 100 micro/ml) in presence of serum or combined with FGF-2. All together, the results indicated that heparin and fucoidan can be used as tools to further investigate the cellular mechanisms regulating the proliferation and migration of human vascular cells. Moreover, the data already suggest a potential role of fucoidan as a new therapeutic agent of vegetal origin in the vascular endothelium wound repair.

The frequency of selective IgA deficiency in myasthenia gravis.

The frequency of selective IgA deficiency is unknown in myasthenia gravis. We screened a series of 333 consecutive adult myasthenic patients for selective IgA deficiency and found only one patient with a selective IgA deficiency, which is not significantly different from the normal population. Organ-specific autoimmune diseases may differ from systemic autoimmune diseases concerning their frequency of association with selective IgA deficiency.

Prospective study of the occurrence of monoclonal gammapathies following bone marrow transplantation in young children.

We have prospectively studied the occurrence of monoclonal serum immunoglobulins in 38 recipients of BMT. Patients were young children with primary immunodeficiencies (n = 31), other inherited diseases (n = 4), leukemia (n = 2), or aplastic anemia (n = 1). Twenty-nine received an HLA-nonidentical marrow and nine an HLA-identical marrow. Serum monoclonal immunoglobulins were detected by the immunofixation method. Monoclonal immunoglobulins were found in 26 patients. Monoclonal components were more frequently detected in patients with primary severe T cell deficiencies (21/25) rather than in the other patients (6/13). In 7 of 29 recipients of HLA-nonidentical transplants, versus 0 out of 9 recipients of HLA-identical transplants, serum monoclonal immunoglobulins were found associated with a B lymphocyte proliferation syndrome due to an Epstein-Barr virus infection. In this group, monoclonal immunoglobulins were detected early, prior to the onset of the clinical syndrome. The simultaneous occurrence of several monoclonal immunoglobulins was more frequent in these patients, while monoclonal immunoglobulin concentrations increased faster, especially those of IgM isotype. These characteristics may allow in patients at risk (recipients with primary T cell immunodeficiencies and receiving HLA-nonidentical transplantation) an earlier diagnosis of B lymphocyte proliferative syndrome that may eventually lead to early and more efficient therapy.