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Dussiot M.,French Institute of Health and Medical Research | Dussiot M.,University of Paris Pantheon Sorbonne | Dussiot M.,French National Center for Scientific Research | Dussiot M.,Laboratory of Excellence GR Ex | And 57 more authors.
Nature Medicine | Year: 2014

The pathophysiology of ineffective erythropoiesis in β-thalassemia is poorly understood. We report that RAP-011, an activin receptor IIA (ActRIIA) ligand trap, improved ineffective erythropoiesis, corrected anemia and limited iron overload in a mouse model of β-thalassemia intermedia. Expression of growth differentiation factor 11 (GDF11), an ActRIIA ligand, was increased in splenic erythroblasts from thalassemic mice and in erythroblasts and sera from subjects with β-thalassemia. Inactivation of GDF11 decreased oxidative stress and the amount of α-globin membrane precipitates, resulting in increased terminal erythroid differentiation. Abnormal GDF11 expression was dependent on reactive oxygen species, suggesting the existence of an autocrine amplification loop in β-thalassemia. GDF11 inactivation also corrected the abnormal ratio of immature/mature erythroblasts by inducing apoptosis of immature erythroblasts through the Fas-Fas ligand pathway. Taken together, these observations suggest that ActRIIA ligand traps may have therapeutic relevance in β-thalassemia by suppressing the deleterious effects of GDF11, a cytokine which blocks terminal erythroid maturation through an autocrine amplification loop involving oxidative stress and α-globin precipitation. © 2014 Nature America, Inc. All rights reserved.


Zumerle S.,French National Center for Scientific Research | Zumerle S.,University of Paris Descartes | Zumerle S.,Laboratory of Excellence GR Ex | Mathieu J.R.R.,French National Center for Scientific Research | And 17 more authors.
Blood | Year: 2014

Hepcidin is a 25-amino-acid peptide demonstrated to be the iron regulatory hormone capable of blocking iron absorption from the duodenum and iron release from macrophages. Mutations affecting hepcidin regulators or the hepcidin gene itself cause hemochromatosis, a common genetic disorder. Hepcidin is produced mainly by the liver, but many cells and tissues express low levels of the hormone. To determine the contribution of these hepcidin-producing tissues in body iron homeostasis, we have developed a new mouse model in which the hepcidin gene can be conditionally inactivated. Here we compare a liver-specific knockout (KO) mousemodel with total KO mice.We showthat the liver-specificKOmice fully recapitulate the severe iron overload phenotype observed in the total KO mice, with increased plasma iron and assive parenchymal iron accumulation. This result demonstrates that the hepatocyte constitutes the predominant reservoir for systemic hepcidin and that the other tissues are unable to compensate. © 2014 by The American Society of Hematology.


Brousse V.,Hopital University Necker Enfants Malades | Brousse V.,University of Paris Descartes | Brousse V.,Laboratory of Excellence GR Ex | Buffet P.,Laboratory of Excellence GR Ex | And 2 more authors.
British Journal of Haematology | Year: 2014

The spleen has a combined function of immune defence and quality control of senescent or altered red cells. It is the first organ injured in sickle cell anaemia (SCA) with evidence of hyposplenism present before 12 months in the majority of children. Repeated splenic vaso-occlusion leads to fibrosis and progressive atrophy of the organ (autosplenectomy), which is generally complete by 5 years in SCA. The precise sequence of pathogenic events leading to hyposplenism is unknown. Splenic injury is generally silent and progressive. It can be clinically overt with acute splenic sequestration of red cells, an unpredictable and life-threatening complication in infants. Splenomegaly, with or without hypersplenism, can also occur and can coexist with loss of function. Hyposplenism increases the susceptibility of SCA children to infection with encapsulated bacteria, which is notably reduced by penicillin prophylaxis and immunization. Whether hyposplenism indirectly increases the risk of vaso-occlusion or other circulatory complications remains to be determined. © 2014 John Wiley & Sons Ltd.


Konz T.,University of Oviedo | Montes-Bayon M.,University of Oviedo | Vaulont S.,French Institute of Health and Medical Research | Vaulont S.,French National Center for Scientific Research | And 2 more authors.
Metallomics | Year: 2014

Hepcidin is a 25-amino acid peptide hormone that is produced and secreted predominantly by hepatocytes, circulates in the bloodstream, and is excreted by the kidneys. Since the discovery of hepcidin and the elucidation of its important role in iron homeostasis, hepcidin has been suggested as a promising diagnostic marker for iron-related disorders. In this regard, a number of analytical methods have been developed in order to assess hepcidin concentration in different biological fluids, particularly serum and urine. In this critical review we have tried to address the issues still pending in accurate determination of this peptide by evaluating the available analytical methodologies. Among them, the use of ELISA strategies (in competitive or sandwich formats) and molecular mass spectrometry (MS) including MALDI and/or LC-MS has been critically compared. The use of elemental mass spectrometry (ICP-MS) has also been included as a possible complementary tool to the previous ones. In addition, this manuscript has revised the existing and potentially emerging clinical applications of hepcidin testing for diagnosis. These include the iron disorders such as iron deficiency anemia (IDA, low hepcidin), anemia of chronic disease (ACD, high hepcidin) and the combined state of ACD and IDA or hemochromatosis. Other applications such as using hepcidin in assessing the response to existing therapies in cancer have also been revised in the manuscript. © 2014 The Royal Society of Chemistry.


Rossi G.,French Institute of Health and Medical Research | Rossi G.,University Paris Diderot | Rossi G.,Laboratory of Excellence GR Ex | Barnoud J.,French Institute of Health and Medical Research | And 5 more authors.
Physica Scripta | Year: 2013

Carbon nanoparticles (CNPs) are considered to be among the most promising nanomaterials, with applications in many different areas of technology. Most CNPs can enter both artificial lipid membranes and living cells, and are biologically active. The interaction of CNPs with lipid membranes is of great interest because biological activity requires crossing or breaking lipid membranes. Moreover, lipid bilayers have been proposed to be efficient solubilizing agents for C60 and C70 fullerenes. In this comment, we review the literature on fullerene partitioning and dispersion in lipid membranes, considering both the experimental and the simulation literature, and highlighting similarities and differences. Both experiments and simulations confirm that fullerenes partition to the membrane interior, although experimental information on the location of fullerene molecules is only qualitative. On the other hand, the fullerene dispersion state is difficult to assess experimentally, and appears to depend on the details of the methodology used for the preparation of fullerene-loaded liposomes. Although some degree of aggregation is confirmed by most experiments, the extent of the aggregation is uncertain. Large aggregates observed in the presence of lipid membranes are unlikely to be found within the membrane, as they are orders of magnitude larger than the membrane thickness. Simulations carried out so far yielded contrasting results. Both atomistic and some coarse-grained simulations indicated that fullerene dimerization in lipid membranes should be significantly less favorable than that in bulk alkanes, but the physical reasons for this are still unclear. © 2013 The Royal Swedish Academy of Sciences.


De Grandis M.,French Institute of Health and Medical Research | De Grandis M.,University Paris Diderot | De Grandis M.,Sanguine | De Grandis M.,Laboratory of Excellence GR Ex | And 21 more authors.
Blood | Year: 2013

Polycythemia vera (PV) is characterized by an increased RBC mass, spontaneous erythroid colony formation, and the JAK2V617F mutation. PV is associated with a high risk of mesenteric and cerebral thrombosis. PV RBC adhesion to endothelial laminin is increased and mediated by phosphorylated erythroid Lu/BCAM. In the present work, we investigated the mechanism responsible for Lu/BCAM phosphorylation in the presence of JAK2V617F using HEL and BaF3 cell lines as well as RBCs from patients with PV. High levels of Rap1-GTP were found in HEL and BaF3 cells expressing JAK2V617F compared with BaF3 cells with wild-type JAK2. This finding was associated with increased Akt activity, Lu/BCAM phosphorylation, and cell adhesion to laminin that were inhibited by the dominant-negative Rap1S17N or by the specific Rap1 inhibitor GGTI-298. Surprisingly, knocking-down EpoR in HEL cells did not alter Akt activity or cell adhesion to laminin. Our findings reveal a novel EpoR-independent Rap1/Akt signaling pathway that is activated by JAK2V617F in circulating PV RBCs and responsible for Lu/BCAM activation. This new characteristic of JAK2V617F could play a critical role in initiating abnormal interactions among circulating and endothelial cells in patients with PV. © 2013 by The American Society of Hematology.


Olivieri L.,French Institute of Health and Medical Research | Olivieri L.,University of Reunion Island | Olivieri L.,Sanguine | Olivieri L.,Laboratory of Excellence GR Ex | And 4 more authors.
Journal of Molecular Graphics and Modelling | Year: 2014

FKBP12 is an important target in the treatment of transplant rejection and is also a promising target for cancer and neurodegenerative diseases. We determined for two ligands of nanomolar affinity the set of parameters in the CHARMM force field. The fitting procedure was based on reproducing the quantum chemistry data (distances, angles, and energies). Since the dynamical behavior of such ligands strongly depends on the dihedral angles, care was taken to derive the corresponding parameters. Moreover, since each of the central core region of these two ligands is similar to other known ligands or drugs of other proteins, part at least of these parameters could also be useful for these other ligands. © 2014 Elsevier Inc.


Moura D.S.,French Institute of Health and Medical Research | Moura D.S.,University of Paris Pantheon Sorbonne | Moura D.S.,French National Center for Scientific Research | Moura D.S.,Laboratory of Excellence GR Ex | And 10 more authors.
Immunology and Allergy Clinics of North America | Year: 2014

In approximately one-third of cases, patients with mastocytosis can display various disabling general and neuropsychological symptoms. General signs may have a major impact on quality of life. Neurologic symptoms are less frequent. In a majority of cases, the pathophysiology of these symptoms is not known but could be linked to tissular mast cell infiltration, mast cell mediator release, or both. Treatments aiming at reducing mast cell number and/or stabilizating mast cells may be useful. Preliminary results suggest that treatment with kinase inhibitors may improve symptoms of depression and cognitive impairment. © 2014 Elsevier Inc.


PubMed | Laboratory of Excellence GR Ex, French Institute of Health and Medical Research, Tunis el Manar University and CNRS Gustave Roussy Institute
Type: | Journal: DNA repair | Year: 2016

The family of Ten-Eleven Translocation (TET) proteins is implicated in the process of active DNA demethylation and thus in epigenetic regulation. TET 1, 2 and 3 proteins are oxygenases that can hydroxylate 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC) and further oxidize 5-hmC into 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC). The base excision repair (BER) pathway removes the resulting 5-fC and 5-caC bases paired with a guanine and replaces them with regular cytosine. The question arises whether active modification of 5-mC residues and their subsequent elimination could affect the genomic DNA stability. Here, we generated two inducible cell lines (Ba/F3-EPOR, and UT7) overexpressing wild-type or catalytically inactive human TET2 proteins. Wild-type TET2 induction resulted in an increased level of 5-hmC and a cell cycle defect in S phase associated with higher level of phosphorylated P53, chromosomal and centrosomal abnormalities. Furthermore, in a thymine-DNA glycosylase (Tdg) deficient context, the TET2-mediated increase of 5-hmC induces mutagenesis characterized by GC>AT transitions in CpG context suggesting a mutagenic potential of 5-hmC metabolites. Altogether, these data suggest that TET2 activity and the levels of 5-hmC and its derivatives should be tightly controlled to avoid genetic and chromosomal instabilities. Moreover, TET2-mediated active demethylation might be a very dangerous process if used to entirely demethylate the genome and might rather be used only at specific loci.


PubMed | Laboratory of Excellence GR Ex and French Institute of Health and Medical Research
Type: | Journal: Malaria journal | Year: 2015

Malaria is still one of the most prevalent infectious diseases in the world. Sequestration of infected erythrocytes (IEs) is the prime mediator of disease. Cytoadhesion of IEs is mediated by members of the highly diverse Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). A restricted sub-set of var genes encoding for PfEMP1s possessing the domain cassettes DC8 and DC13 were found to bind to the endothelial protein C receptor (EPCR). These var genes were shown to be highly expressed by parasites from patients with severe malaria clinical outcomes compared to those from patients with uncomplicated symptoms.In order to further study the molecular mechanisms underlying DC8/DC13 expressing IEs adhesion to EPCR, a method was developed to produce highly pure recombinant EPCR. The IT4 parasite strain was selected on either anti-IT4-VAR19 purified IgG, EPCR or human brain endothelial cell line and their var gene expression profiles as well as their binding phenotypes were compared. The N-terminal region of IT4-VAR19 comprising a full-length DC8 cassette as well as the single EPCR binding CIDR1.1 domain were also produced, and their immune recognition (IgG) was assessed using plasma samples from Beninese children presenting acute mild malaria, severe malaria or cerebral malaria at the time of their admission to the clinic, and from convalescent-phase plasma collected 30 days after anti-malarial treatment.The multi-domain VAR19-NTS-DBL6 binds to EPCR with a greater affinity than the CIDR1.1 domain alone and this study also demonstrates that VAR19-NTS-DBL6 binding to the EPCR-expressing endothelial cell line (HBEC5i) is more pronounced than that of the CIDR1.1 domain alone. IT4-VAR19 represents the preferentially expressed-PfEMP1 when FCR3-IEs are selected based on their capability to bind EPCR. Notably, no significant difference in the levels of antibodies towards IT4-VAR19 antigens was observed within all clinical groups between plasma samples collected during the acute malaria phase compared to samples collected 30 days after anti-malaria treatment.These data indicate that even being the preferentially selected IT4-EPCR-binding variant, the IT4-VAR19-DC8 region does not appear to be associated with the acquisition of antibodies during a single severe paediatric malaria episode in Benin.

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