Publications by authors named "Dominique Lasne"

53 Publications

Functional Flow Cytometric Assay for Reliable and Convenient Heparin-Induced Thrombocytopenia Diagnosis in Daily Practice.

Biomedicines 2021 Mar 25;9(4). Epub 2021 Mar 25.

Inserm U1059 Sainbiose, Université de Lyon, 42055 Saint-Etienne, France.

Reliable laboratory diagnosis of heparin-induced thrombocytopenia (HIT) remains a major clinical concern. Immunoassays are highly sensitive, while confirmatory functional tests (based on heparin-dependent platelet activation) lack standardization. We evaluated the diagnostic performance of a functional flow cytometric assay (FCA) based on the detection of heparin-dependent platelet activation with an anti-p-selectin. A total of 288 patients were included (131 HIT-positive and 157 HIT-negative) with a HIT diagnosis established by expert opinion adjudication (EOA) considering clinical data and local laboratory results. The FCA was centrally performed in a single laboratory on platelet-rich plasma, using a very simple four-color fluorometer. The results were standardized according to the Heparin Platelet Activation (HEPLA) index. The serotonin release assay (SRA) was performed in the four French reference laboratories. Based on the final HIT diagnosis established by EOA, the sensitivity and specificity of the FCA were 88 and 95%, respectively, values very similar to those of the SRA (88 and 97%, respectively). This study showed that the FCA, based on easily implementable technology, may be routinely used as a reliable confirmatory test for HIT diagnosis.
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http://dx.doi.org/10.3390/biomedicines9040332DOI Listing
March 2021

Role of oculocerebrorenal syndrome of Lowe (OCRL) protein in megakaryocyte maturation, platelet production and functions: a study in patients with Lowe syndrome.

Br J Haematol 2021 Mar 2;192(5):909-921. Epub 2021 Feb 2.

Université de Paris, Innovations Thérapeutiques en Hémostase, Paris, INSERM U1140, France.

Lowe syndrome (LS) is an oculocerebrorenal syndrome of Lowe (OCRL1) genetic disorder resulting in a defect of the OCRL protein, a phosphatidylinositol-4,5-bisphosphate 5-phosphatase containing various domains including a Rho GTPase-activating protein (RhoGAP) homology domain catalytically inactive. We previously reported surgery-associated bleeding in patients with LS, suggestive of platelet dysfunction, accompanied with a mild thrombocytopenia in several patients. To decipher the role of OCRL in platelet functions and in megakaryocyte (MK) maturation, we conducted a case-control study on 15 patients with LS (NCT01314560). While all had a drastically reduced expression of OCRL, this deficiency did not affect platelet aggregability, but resulted in delayed thrombus formation on collagen under flow conditions, defective platelet spreading on fibrinogen and impaired clot retraction. We evidenced alterations of the myosin light chain phosphorylation (P-MLC), with defective Rac1 activity and, inversely, elevated active RhoA. Altered cytoskeleton dynamics was also observed in cultured patient MKs showing deficient proplatelet extension with increased P-MLC that was confirmed using control MKs transfected with OCRL-specific small interfering(si)RNA (siOCRL). Patients with LS also had an increased proportion of circulating barbell-shaped proplatelets. Our present study establishes that a deficiency of the OCRL protein results in a defective actomyosin cytoskeleton reorganisation in both MKs and platelets, altering both thrombopoiesis and some platelet responses to activation necessary to ensure haemostasis.
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http://dx.doi.org/10.1111/bjh.17346DOI Listing
March 2021

Dabigatran Level Before Reversal Can Predict Hemostatic Effectiveness of Idarucizumab in a Real-World Setting.

Front Med (Lausanne) 2020 16;7:599626. Epub 2020 Dec 16.

Université de Paris, Innovative Therapies in Haemostasis, INSERM, Paris, France.

Idarucizumab has been included in guidelines for the management of bleeding or surgical procedure in dabigatran-treated patients without need for biological monitoring. The aim of the study was to assess the prognostic value of dabigatran plasma level before reversal to test the hemostatic efficacy of idarucizumab. The secondary objectives were (i) to analyze plasma dabigatran level according to the risk of rebound and (ii) to evaluate the incidence of post-reversal non-favorable clinical outcomes (including thromboembolism, bleeding, antithrombotic, and death) and antithrombotic resumption. This was an observational multicentric cohort study, which included all French patients who required idarucizumab for dabigatran reversal. Between May 2016 and April 2019, 87 patients from 21 French centers were enrolled. Patients received idarucizumab for overt bleeding ( = 61), urgent procedures ( = 24), or overdose without bleeding ( = 2). Among patients with major bleeding ( = 57), treatment with idarucizumab was considered effective in 44 (77.2%) of them. Patients who did not achieve effective hemostasis after reversal had a significantly higher mean level of plasma dabigatran at baseline (524.5 ± 386 vs. 252.8 ng/mL ± 235, = 0.033). Furthermore, patients who did not achieve effective hemostasis after reversal had less favorable outcomes during follow-up (46.2 vs. 81.8%, = 0.027). ROC curve identified a cutoff of 264 ng/mL for dabigatran level at admission to be predictive of ineffective hemostasis. No plasma dabigatran rebound was observed after reversal in patients with dabigatran plasma level < 264 ng/mL at baseline. This retrospective study shows that dabigatran level before reversal could predict hemostatic effectiveness and dabigatran plasma rebound after idarucizumab injection.
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http://dx.doi.org/10.3389/fmed.2020.599626DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7772865PMC
December 2020

[GFHT proposals for management of discordance between the International normalized ratio measured in the laboratory and by self-testing].

Ann Biol Clin (Paris) 2020 Dec;78(6):655-664

Service d'hématologie-hémostase, Centre régional de traitement de l'hémophilie, Hôpital Trousseau, CHRU de Tours, EA 7501, Université François Rabelais, France.

The lack of quality control for patient point-of-care (POC) INR devices is an issue that has led the French health authorities to make recommendations: a laboratory INR (lab INR) has to be performed at the same time as the POC INR every 6 months. However, the differences observed between the two INRs, POC and lab INRs, are not necessarily due to a failure of the POC INR device. We present here a review of the different causes of discrepancies between INR results, which are the basis of the proposals of the Groupe français d'études sur l'hémostase et la thrombose (GFHT) on the management of lab and POC INR discrepancies. Pre-analytical conditions may account for discrepancies (sampling, transport and storage conditions), as well as analytical factors (mainly the nature of the thromboplastin used) and the clinical context (inflammatory or autoimmune diseases, polycythaemia...). The interpretation of INR discrepancies is not always easy and these proposals aim at standardizing the procedure to be followed in order to make the most appropriate decision for the patient.
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http://dx.doi.org/10.1684/abc.2020.1613DOI Listing
December 2020

Measuring beta-galactose exposure on platelets: Standardization and healthy reference values.

Res Pract Thromb Haemost 2020 Jul 2;4(5):813-822. Epub 2020 Jun 2.

Department of Biological Hematology Hôpital Necker AP-HP Paris France.

Background: Correct diagnosis of the cause of thrombocytopenia is crucial for the appropriate management of patients. Hyposialylation/desialylation (characterized by abnormally high β-galactose exposure) accelerates platelet clearance and can lead to thrombocytopenia. However, the reference range for β-galactose exposure in healthy individuals has not been defined previously.

Objective: The objective of the present study was to develop a standardized assay of platelet β-galactose exposure for implementation in a clinical laboratory.

Methods: β-Galactose exposure was measured in platelet-rich plasma by using flow cytometry and agglutinin (RCA). A population of 120 healthy adults was recruited to study variability.

Results: We determined an optimal RCA concentration of 12.5 μg/mL. The measure was stable for up to 4 hours (mean fluorescence intensity [MFI]-RCA: 1233 ± 329 at 0 hour and 1480 ± 410 at 4 hours). The platelet count did not induce a variation of RCA and the measure of RCA was stable when tested up to 24 hours after blood collection (MFI-RCA: 1252 ± 434 at day 0 and 1140 ± 297 24 hours after blood sampling). To take into account the platelet size, results should be expressed as RCA/forward scatter ratio. We used the assay to study variability in 120 healthy adults, and we found that the ratio is independent of sex and blood group.

Conclusion: We defined a normal range in a healthy population and several preanalytical and analytical variables were evaluated, together with positive and negative controls. This assay may assist in the diagnosis of thrombocytopenic diseases linked to changes in β-galactose exposure.
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http://dx.doi.org/10.1002/rth2.12369DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7586713PMC
July 2020

Emicizumab treatment: Impact on coagulation tests and biological monitoring of haemostasis according to clinical situations (BIMHO group proposals).

Eur J Haematol 2020 Dec 16;105(6):675-681. Epub 2020 Sep 16.

Laboratoire d'hématologie générale, Hôpital Necker, AP-HP, Paris, France.

Emicizumab, a bispecific humanised monoclonal antibody restoring to some extent the function of activated FVIII deficient in haemophilia A, represents a major therapeutic advance in the management of haemophilia A patients. No dosage adjustment is required, which leads to a major change for patients used to regular biological monitoring which is particularly burdensome in the case of substitution therapy. In some circumstances, such as before an invasive procedure or in case of bleeding, biological monitoring will be necessary and emicizumab's interference with haemostasis tests, particularly those based on an activated partial thromboplastin times (aPTT), must be known to best interpret the tests and to select the most appropriate methods to guide therapy. The normalisation of aPTT in patients treated with emicizumab is not sufficient to consider haemostasis as normalised. In the event of administration of FVIII to a patient receiving emicizumab, the determination of FVIII should use a chromogenic method using non-human reagents. Coagulation global tests have been proposed to evaluate the biological response when using bypassing agents in patients treated with emicizumab, but the usefulness must be confirmed. The French group BIMHO presents proposals for biological monitoring of a patient treated with emicizumab according to clinical situations.
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http://dx.doi.org/10.1111/ejh.13490DOI Listing
December 2020

Diagnosis and management of heparin-induced thrombocytopenia.

Anaesth Crit Care Pain Med 2020 04 13;39(2):291-310. Epub 2020 Apr 13.

Service d'anesthésie-réanimation, hôpital européen Georges-Pompidou, INSERM UMRS-1140, université Paris Descartes, 25, rue Leblanc, 75015 Paris, France. Electronic address:

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http://dx.doi.org/10.1016/j.accpm.2020.03.012DOI Listing
April 2020

Factor VIII and IX assays for post-infusion monitoring in hemophilia patients: Guidelines from the French BIMHO group (GFHT).

Eur J Haematol 2020 Aug 27;105(2):103-115. Epub 2020 May 27.

Service d'Hématologie Hémostase, Hospices Civils de Lyon, Bron, France.

Replacement therapy with plasma-derived or recombinant FVIII and FIX (pdFVIII/pdFIX or rFVIII/rFIX) concentrates is the standard of treatment in patients with haemophilia A and B, respectively. Measurement of factor VIII (FVIII:C) or factor IX (FIX:C) levels can be done by one-stage clotting assay (OSA) or chromogenic substrate assay (CSA). The French study group on the Biology of Hemorrhagic Diseases (a collaborative group of the GFHT and MHEMO network) presents a literature review and proposals for the monitoring of FVIII:C and FIX:C levels in treated haemophilia A and B patients, respectively. The use of CSA is recommended for the monitoring of patients treated with pdFVIII or rFVIII including extended half-life (EHL) rFVIII. Except for rFVIII-Fc, great caution is required when measuring FVIII:C levels by OSA in patients substituted by EHL-rFVIII. The OSA is recommended for the monitoring of patients treated with pdFIX or rFIX. Large discordances in the FIX:C levels measured for extended half-life rFIX (EHL-rFIX), depending on the method and reagents used, must lead to great attention when OSA is used for measuring FIX:C levels in patients substituted by EHL-rFIX. Data of most of recent studies, obtained with spiked plasmas, deserve to be confirmed in plasma samples of treated patients.
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http://dx.doi.org/10.1111/ejh.13423DOI Listing
August 2020

Inflammation in deep vein thrombosis: a therapeutic target?

Hematology 2019 Dec;24(1):742-750

INSERM UMR-S1176, Université Paris Saclay, Le Kremlin-Bicêtre, France.

Deep vein thrombosis is a common disease associated with a variety of complications including post-thrombotic syndrome as a late complication. It is now clear that in addition to classical deep vein thrombosis triggers such as blood flow disturbance, hypercoagulability, and vessel wall changes, inflammation has a key role in the pathophysiology of deep vein thrombosis, and there is a close relationship between inflammation and coagulation. As attested by changes in several plasma biomarkers, inflammation may have a significant role in the development of post-thrombotic syndrome. Here, we review the link between inflammation and deep vein thrombosis and thus the potential value of anti-inflammatory and/or anticoagulant drugs in the treatment of deep vein thrombosis and the prevention of post-thrombotic syndrome.
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http://dx.doi.org/10.1080/16078454.2019.1687144DOI Listing
December 2019

Management of bleeding and invasive procedures in haemophilia A patients with inhibitor treated with emicizumab (Hemlibra ): Proposals from the French network on inherited bleeding disorders (MHEMO), the French Reference Centre on Haemophilia, in collaboration with the French Working Group on Perioperative Haemostasis (GIHP).

Haemophilia 2019 Sep 11;25(5):731-737. Epub 2019 Jul 11.

Unité d'Hémostase Clinique, Louis Pradel Hospital, University Claude Bernard, Lyon, France.

Introduction: Emicizumab (Hemlibra ) recently became available and requires an adaptation for managing bleeding, suspected bleeding and emergency or scheduled invasive procedures in haemophilia A patients with inhibitor. This implicates a multidisciplinary approach and redaction of recommendations for care that must be regularly adapted to the available data.

Aim: The following text aims to provide a guide for the management of people with haemophilia A with inhibitor treated with emicizumab in case of bleeding or invasives procedures.

Methods: The French network on inherited bleeding disorders (MHEMO), the French Reference Centre on Haemophilia (CRH), in collaboration with the French Working Group on Perioperative Haemostasis (GIHP) have been working together to make proposals for the management of these situations.

Results: Haemostatic treatment and other medications should be given stepwise, according to the severity and location of the bleeding or the risk of bleeding of the procedure as well as the haemostatic response obtained at each step in order to ensure an optimal benefit/risk ratio.

Conclusion: The lack of data means that it is only possible to issue proposals rather than recommendations.
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http://dx.doi.org/10.1111/hae.13817DOI Listing
September 2019

Elevated thrombin generation in patients with congenital disorder of glycosylation and combined coagulation factor deficiencies.

J Thromb Haemost 2019 11 28;17(11):1798-1807. Epub 2019 Jul 28.

Laboratoire d'Hématologie, AP-HP, Hôpital Necker-Enfants malades, Paris, France.

Background: Congenital disorders of glycosylation are rare inherited diseases affecting many different proteins. The lack of glycosylation notably affects the hemostatic system and leads to deficiencies of both procoagulant and anticoagulant factors.

Objective: To assess the hemostatic balance in patients with multiple coagulation disorders by using a thrombin generation assay.

Method: We performed conventional coagulation assays and a thrombin generation assay on samples from patients with congenital disorder of glycosylation. The thrombin generation assay was performed before and after activation of the protein C system by the addition of soluble thrombomodulin.

Results: A total of 35 patients were included: 71% and 57% had low antithrombin and factor XI levels, respectively. Protein C and protein S levels were abnormally low in 29% and 26% of the patients, respectively, whereas only 11% displayed low factor IX levels. Under baseline conditions, the thrombin generation assay revealed a significantly higher endogenous thrombin potential and thrombin peak in patients, relative to controls. After spiking with thrombomodulin, we observed impaired involvement of the protein C system. Hence, 54% of patients displayed a hypercoagulant phenotype in vitro. All the patients with a history of stroke-like episodes or thrombosis displayed this hypercoagulant phenotype.

Conclusion: A thrombin generation assay revealed a hypercoagulant in vitro phenotype under baseline condition; this was accentuated by impaired involvement of the protein C system. This procoagulant phenotype may thus reflect the risk of severe vascular complications. Further research will have to determine whether the thrombin generation assay is predictive of vascular events.
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http://dx.doi.org/10.1111/jth.14559DOI Listing
November 2019

Relevance of platelet desialylation and thrombocytopenia in type 2B von Willebrand disease: preclinical and clinical evidence.

Haematologica 2019 12 28;104(12):2493-2500. Epub 2019 Feb 28.

HITh, UMR_S 1176, INSERM Université Paris-Sud, Université Paris-Saclay, F-94270 Le Kremlin-Bicêtre

Patients with type 2B von Willebrand disease (vWD) (caused by gain-of-function mutations in the gene coding for von Willebrand factor) display bleeding to a variable extent and, in some cases, thrombocytopenia. There are several underlying causes of thrombocytopenia in type 2B vWD. It was recently suggested that desialylation-mediated platelet clearance leads to thrombocytopenia in this disease. However, this hypothesis has not been tested The relationship between platelet desialylation and the platelet count was probed in 36 patients with type 2B von Willebrand disease (p.R1306Q, p.R1341Q, and p.V1316M mutations) and in a mouse model carrying the severe p.V1316M mutation (the 2B mouse). We observed abnormally high elevated levels of platelet desialylation in both patients with the p.V1316M mutation and the 2B mice. , we demonstrated that 2B p.V1316M/von Willebrand factor induced more desialylation of normal platelets than wild-type von Willebrand factor did. Furthermore, we found that N-glycans were desialylated and we identified αIIb and β3 as desialylation targets. Treatment of 2B mice with sialidase inhibitors (which correct platelet desialylation) was not associated with the recovery of a normal platelet count. Lastly, we demonstrated that a critical platelet desialylation threshold (not achieved in either 2B patients or 2B mice) was required to induce thrombocytopenia In conclusion, in type 2B vWD, platelet desialylation has a minor role and is not sufficient to mediate thrombocytopenia.
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http://dx.doi.org/10.3324/haematol.2018.206250DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6959185PMC
December 2019

[Factor VIII assays in treated hemophilia A patients].

Ann Biol Clin (Paris) 2019 02;77(1):53-65

CHU Lille, Institut d'hématologie-transfusion, Inserm U1011, Lille, France.

Replacement therapy with plasma-derived or recombinant FVIII (pdFVIII or rFVIII) concentrates is the standard of treatment in patients with hemophilia A. The reference method used for measuring factor VIII (FVIII:C) levels in patients treated by FVIII concentrates is the chromogenic substrate assay (CSA). However, the one-stage clotting assay (OSA) is predominantly used in current clinical practice, but this method depends on the activated partial thromboplastin time (APTT) reagent and the coagulation analyzer used, and wide variations in the measurements of FVIII recovery have been reported with some factor concentrates. The French study group on the biology of hemorrhagic diseases (a collaborative group of the GFHT and MHEMO network) presents a review of the literature and proposals for the monitoring of FVIII:C levels in treated hemophilia A patients. The use of CSA calibrated with a plasma reference tested against the current FVIII WHO (World Health Organization) International Standard is recommended for the monitoring of patients treated with pdFVIII or rFVIII including extended half-life (EHL) rFVIII. OSA are adequate for the monitoring of patients treated with pdFVIII or with most of rFVIII concentrates. However, preliminary comparison with CSA is mandatory before measuring FVIII:C by OSA in patients treated by Refacto AF. For rFVIII-EHL, OSA are only acceptable for Elocta®. Great caution is therefore required when measuring FVIII:C levels by OSA in patients substituted by other EHL-rFVIII. Indeed, most of recent studies reported data obtained with spiked plasmas, which deserve to be confirmed on plasma samples collected in treated patients.
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http://dx.doi.org/10.1684/abc.2019.1413DOI Listing
February 2019

[Factor IX assays in treated hemophilia B patients].

Ann Biol Clin (Paris) 2019 02;77(1):41-52

Service d'hématologie hémostase, Hospices civils de Lyon, Bron, France.

Replacement therapy with plasma-derived or recombinant FIX (pdFIX or rFIX) concentrates is the standard of treatment in patients with hemophilia B. The method predominantly used for measuring factor IX (FIX:C) levels is the one-stage clotting assay (OSA) but this method depends on the activated partial thromboplastin time (APTT) reagent and the coagulation analyzer used, and wide variations in the measurements of FIX recovery have been reported with some factor concentrates. The French study group on the biology of hemorrhagic diseases (a collaborative group of the GFHT and MHEMO network), presents a review of the literature and proposals for the monitoring of FIX:C levels in treated hemophilia B patients. The use of OSA calibrated with a plasma reference tested against the current FIX WHO International Standard is recommended for the monitoring of patients treated with pdFIX or rFIX. Chromogenic substrate assays (CSA) are adequate for the monitoring of patients treated with Rixubis, but data available for Benefix are currently too limited. For extended half-life rFIX (EHL-rFIX), large discordances in the FIX:C levels measured were evidenced, depending on the method and reagents used. Great attention is therefore required for measuring FIX:C levels by OSA in patients substituted by EHL-rFIX. Commercial kits for CSA are not equivalent, and although potentially useful, they are not validated for all EHL-rFIX. Most of recent studies reported data obtained with spiked plasmas, which deserve to be confirmed on plasma samples collected in treated patients.
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http://dx.doi.org/10.1684/abc.2019.1414DOI Listing
February 2019

Management of antiplatelet therapy for non-elective invasive procedures or bleeding complications: Proposals from the French Working Group on Perioperative Haemostasis (GIHP) and the French Study Group on Thrombosis and Haemostasis (GFHT), in collaboration with the French Society for Anaesthesia and Intensive Care (SFAR).

Arch Cardiovasc Dis 2019 Mar 6;112(3):199-216. Epub 2019 Jan 6.

Service d'angiologie et d'hémostase, département de spécialités de médecine, hôpitaux universitaires de Genève, 1205 Genève, Switzerland; Geneva Platelet Group, faculté de médecine, université de Genève, 1205 Genève, Switzerland.

The French Working Group on Perioperative Haemostasis (GIHP) and the French Study Group on Haemostasis and Thrombosis (GFHT), in collaboration with the French Society for Anaesthesia and Intensive Care (SFAR), drafted up-to-date proposals on the management of antiplatelet therapy for non-elective invasive procedures or bleeding complications. The proposals were discussed and validated by a vote; all proposals could be assigned with a high strength. Management of oral antiplatelet agents in emergency settings requires knowledge of their pharmacokinetic and pharmacodynamic parameters, evaluation of the degree of alteration of haemostatic competence and the associated bleeding risk. Platelet function testing may be considered. When antiplatelet agent-induced bleeding risk may worsen the prognosis, measures should be taken to neutralize antiplatelet therapy, by considering not only the efficacy of available means (which can be limited for prasugrel and even more for ticagrelor), but also the risks that these means expose the patient to. The measures include platelet transfusion at the appropriate dose and haemostatic agents (tranexamic acid; recombinant activated factor VII for ticagrelor). When possible, postponing non-elective invasive procedures at least for a few hours until the elimination of the active compound (which could compromise the effect of transfused platelets) or, if possible, for a few days (reduction of the effect of antiplatelet agents) should be considered.
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http://dx.doi.org/10.1016/j.acvd.2018.10.004DOI Listing
March 2019

TUBB1 mutations cause thyroid dysgenesis associated with abnormal platelet physiology.

EMBO Mol Med 2018 12;10(12)

INSERM U1016, Faculté de Médecine, Cochin Institute, Université Paris Descartes, Sorbonne Paris Cité, Paris, France

The genetic causes of congenital hypothyroidism due to thyroid dysgenesis (TD) remain largely unknown. We identified three novel gene mutations that co-segregated with TD in three distinct families leading to 1.1% of mutations in TD study cohort. (Tubulin, Beta 1 Class VI) encodes for a member of the β-tubulin protein family. gene is expressed in the developing and adult thyroid in humans and mice. All three mutations lead to non-functional α/β-tubulin dimers that cannot be incorporated into microtubules. In mice, knock-out disrupted microtubule integrity by preventing β1-tubulin incorporation and impaired thyroid migration and thyroid hormone secretion. In addition, mutations caused the formation of macroplatelets and hyperaggregation of human platelets after stimulation by low doses of agonists. Our data highlight unexpected roles for β1-tubulin in thyroid development and in platelet physiology. Finally, these findings expand the spectrum of the rare paediatric diseases related to mutations in tubulin-coding genes and provide new insights into the genetic background and mechanisms involved in congenital hypothyroidism and thyroid dysgenesis.
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http://dx.doi.org/10.15252/emmm.201809569DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6284387PMC
December 2018

Impact of Heparin- or Nonheparin-Coated Circuits on Platelet Function in Pediatric Cardiac Surgery.

Ann Thorac Surg 2019 04 3;107(4):1241-1247. Epub 2018 Nov 3.

Hematology Laboratory, Assistance Publique-Hôpitaux de Paris, Hôpital Necker-Enfants Malades, Paris, France; Hémostase Inflammation Thrombose, Unité Mixte de Recherche -S1176, Institut National de la Santé et de la Recherche Médicale, University Paris-Sud, Université Paris-Saclay, Le Kremlin-Bicêtre, France.

Background: Extracorporeal circuit coating has been shown to improve coagulation derangements during pediatric cardiopulmonary bypass (CPB). This study compared platelet function and hemostasis activation in pediatric cardiac surgery conducted with nonheparin coating (Balance; Medtronic, Minneapolis, MN) versus heparin-based coating (Carmeda; Medtronic) circuits.

Methods: A prospective, randomized, double-center trial was conducted in children older than 1 month undergoing congenital heart disease treatment. Blood samples were collected at baseline (T0), 15 minutes after the start of CPB (T1), and 15 minutes (T2) and 1 hour after the conclusion of CPB (T3). The primary end point of the study was to detect potential differences in β-thromboglobulin levels between the two groups at T2. Other coagulation and platelet function indicators were analyzed as secondary end points.

Results: The concentration of β-thromboglobulin increased significantly at T2 in both groups. However, there was no significant difference between the groups across all time points. There was no difference in the secondary end points between the groups.

Conclusions: The two circuits showed similar biological effects on platelet function and coagulation. This observation may be useful in optimizing the conduct of CPB and in rationalizing its cost for the treatment of congenital heart disease.
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http://dx.doi.org/10.1016/j.athoracsur.2018.09.032DOI Listing
April 2019

Pharmacokinetics of Enoxaparin After Renal Transplantation in Pediatric Patients.

J Clin Pharmacol 2018 12 26;58(12):1597-1603. Epub 2018 Sep 26.

Néphrologie pédiatrique, Centre de référence des Maladies Rénales Héréditaires de l'Enfant et de l'Adulte (MARHEA), Institut Imagine, Université Paris Descartes, Hôpital Necker-Enfants Malades, Assistance Publique - Hôpitaux de Paris (AP-HP), Paris, France.

Enoxaparin is commonly used in the prevention of renal allograft vascular thrombosis but off-label in children, and no consensus exists regarding the optimal dosing and dose adjustment. In this retrospective study, 444 anti-Xa levels were obtained from 30 pediatric renal transplant recipients in order to investigate enoxaparin population pharmacokinetics. The main results were (1) 25% of children achieved the target anti-Xa activity 36 hours after initiation of treatment, (2) anti-Xa time courses were best described by a 1-compartment open model with first-order absorption, (3) body weight but not renal function was the sole covariate influencing clearance and volume of distribution, and (4) large between-subject and between-occasion variabilities in anti-Xa activity were observed. However, creatinine-based estimated glomerular filtration rate in the first post-renal transplantation hours may not reliably reflect the actual renal function of the children. Based on the final population model, a Bayesian-based program was developed in order to estimate the individual pharmacokinetic parameters on a single anti-Xa measurement, allowing determination of the next enoxaparin dose that will quickly achieve an appropriate anti-Xa activity (targeting 0.3-0.5 IU/mL) and anticoagulation. Finally, these results should help standardize practices that remain to date largely heterogeneous in pediatric intensive care units.
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http://dx.doi.org/10.1002/jcph.1289DOI Listing
December 2018

Platelet-mapping assay for monitoring antiplatelet therapy during mechanical circulatory support in children: A retrospective observational study.

Res Pract Thromb Haemost 2017 Jul 20;1(1):120-127. Epub 2017 Jun 20.

Laboratoire d'Hématologie AP-HP Hôpital Universitaire Necker-Enfants Malades Paris France.

Introduction: The complex hemostatic changes associated with Berlin Heart (BH) implantation in children require a challenging antithrombotic treatment. The aim of this retrospective analysis was to evaluate the thromboelastography (TEG)-platelet mapping (PM) assay to monitor antiplatelet therapy in children implanted with a BH.

Methods: TEG-PM was performed in 4 BH-implanted patients receiving dipyridamole and aspirin, and 9 healthy volunteers. Patients' antiplatelet therapy was adjusted to TEG-PM results. Light transmission aggregometry (LTA) was also available for 2 of these patients.

Results: Between 2009 and 2014, 4 BH-implanted patients received a dual antiplatelet therapy monitored by TEG-PM. In 2 patients, 18 of 34 tracings were atypical, because the maximum amplitude due to fibrin never stabilized, which made difficult antiplatelet therapy adjustment as recommended by BH's guidelines. To overcome this difficulty, TEG-PM and LTA were next performed in parallel. However, both methods led to different decisions to adjust antiplatelet therapy in 57% of the cases. In order to better understand this atypical tracing, TEG-PM was also performed in 9 volunteers and surprisingly 3 of them had the same atypical tracing. This atypical tracing was corrected by adding apyrase, suggesting that adenosine diphosphate (ADP) participates to spontaneous platelet activation in heparinized samples. In addition, we evidenced a high variability in the responses of TEG-PM with ADP in volunteers.

Conclusions: Antiplatelet therapy monitoring in BH-implanted children remains challenging, as TEG-PM is sensitive to several preanalytical and analytical conditions.
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http://dx.doi.org/10.1002/rth2.12010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6058205PMC
July 2017

CD34+ Hematopoietic Stem Cell Count Is Predictive of Vascular Event Occurrence in Children with Sickle Cell Disease.

Stem Cell Rev Rep 2018 Oct;14(5):694-701

Inserm UMR-S1140, Faculté de Pharmacie, Paris, France.

Background/objectives: Sickle cell disease (SCD) complications mostly result from vascular dysfunction, concerning systemic microvasculature and cerebral large vessels. The aim of this cohort study was to identify potential circulating biomarkers predictive for further vascular event occurrence in pediatric SCD.

Methods: We consecutively enrolled 108 children with SCD at steady state, aged 3-18 years old (median 9.8 years). Hematology, coagulation, hemolysis, endothelial, platelet and vascular activation parameters were recorded at inclusion. Neurovascular and systemic vascular events were prospectively recorded during a mean follow-up period of 27 months.

Results: Patients at steady state displayed significantly higher hemolysis and platelet activation markers, higher leukocyte, CD34+ hematopoietic stem cell and microvesicle counts, and a pro-coagulant profile compared to controls matched for age and ethnicity. Circulating endothelial cell or nucleosome level did not differ. During the follow-up period, 36 patients had at least one neurovascular (n = 12) or systemic vascular event (n = 25). In a multivariate model, high CD34+ cell count was the best predictor for the occurrence of a vascular event (OR 1.2 for 1000 cell/mL increase, 95% CI [1.049-1.4], p = 0.013, sensitivity 53%, specificity 84% for a threshold of 8675 cells/mL).

Conclusion: CD34+ cell count at steady state is a promising biomarker of further vascular event in children with SCD.
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http://dx.doi.org/10.1007/s12015-018-9835-8DOI Listing
October 2018

Guidelines for certification of International Normalized Ratio (INR) for vitamin K antagonists monitoring according to the EN ISO 22870 standards.

Ann Biol Clin (Paris) 2018 Jun;76(3):271-299

Laboratoire central d'hématologie, Hôpital Necker, AP-HP, Paris, France.

Point of care testing (POCT) must comply with regulatory requirements according to standard EN ISO 22870, which identify biologists as responsible for POCT. INR for vitamin K antagonists (VKAs) monitoring is a test frequently performed in haemostasis laboratories. Bedside INR is useful in emergency room, in particular in case of VKAs overdosage but also for specific populations of patients like paediatrics or geriatrics. INR POCT devices are widely used at home by the patients for self-testing, but their use in the hospital by the clinical staff for bedside measurement is growing, with devices which now comply with standard for POCT accreditation for hospital use. The majority of point of care devices for INR monitoring has shown a good precision and accuracy with results similar to those obtained in laboratory. With the aim to help the multidisciplinary groups for POCT supervision, the medical departments and the biologists to be in accordance with the standard, we present the guidelines of the GFHT (Groupe français d'étude sur l'hémostase et la thrombose, subcommittee "CEC et biologie délocalisée") for the certification of POCT INR. These guidelines are based on the SFBC guidelines for the certification of POCT and on the analysis of the literature to ascertain the justification of clinical need and assess the analytical performance of main analysers used in France, as well as on a survey conducted with biologists.
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http://dx.doi.org/10.1684/abc.2018.1332DOI Listing
June 2018

Position of the French Working Group on Perioperative Haemostasis (GIHP) on viscoelastic tests: What role for which indication in bleeding situations?

Anaesth Crit Care Pain Med 2019 10 3;38(5):539-548. Epub 2018 Feb 3.

Laboratoire d'hématologie, hôpital Necker, 75015 Paris, France.

Purpose: Viscoelastic tests (VETs), thromboelastography (TEG) and thromboelastometry (ROTEM) are global tests of coagulation performed on whole blood. They evaluate the mechanical strength of a clot as it builds and develops after coagulation itself. The time required to obtain haemostasis results remains a major problem for clinicians dealing with bleeding, although some teams have developed a rapid laboratory response strategy. Indeed, the value of rapid point-of-care diagnostic devices such as VETs has increased over the years. However, VETs are not standardised and there are few recommendations from the learned societies regarding their use. In 2014, the recommendations of the International Society of Thrombosis and Haemostasis (ISTH) only concerned haemophilia. The French Working Group on Perioperative haemostasis (GIHP) therefore proposes to summarise knowledge on the clinical use of these techniques in the setting of emergency and perioperative medicine.

Methods: A review of the literature.

Principal Findings: The role of the VETs seems established in the management of severe trauma and in cardiac surgery, both adult and paediatric. In other situations, their role remains to be defined: hepatic transplantation, postpartum haemorrhage, and non-cardiac surgery. They must be part of the global management of haemostasis based on algorithms defined in each centre and for each population of patients. Their position at the bedside or in the laboratory is a matter of discussion between clinicians and biologists.

Conclusion: VETs must be included in algorithms. In consultation with the biology laboratory, these devices should be situated according to the way each centre functions.
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http://dx.doi.org/10.1016/j.accpm.2017.12.014DOI Listing
October 2019

Type I interferon-mediated autoinflammation due to DNase II deficiency.

Nat Commun 2017 12 19;8(1):2176. Epub 2017 Dec 19.

INSERM UMR1163, Laboratory of Neurogenetics and Neuroinflammation, Paris, 75015, France.

Microbial nucleic acid recognition serves as the major stimulus to an antiviral response, implying a requirement to limit the misrepresentation of self nucleic acids as non-self and the induction of autoinflammation. By systematic screening using a panel of interferon-stimulated genes we identify two siblings and a singleton variably demonstrating severe neonatal anemia, membranoproliferative glomerulonephritis, liver fibrosis, deforming arthropathy and increased anti-DNA antibodies. In both families we identify biallelic mutations in DNASE2, associated with a loss of DNase II endonuclease activity. We record increased interferon alpha protein levels using digital ELISA, enhanced interferon signaling by RNA-Seq analysis and constitutive upregulation of phosphorylated STAT1 and STAT3 in patient lymphocytes and monocytes. A hematological disease transcriptomic signature and increased numbers of erythroblasts are recorded in patient peripheral blood, suggesting that interferon might have a particular effect on hematopoiesis. These data define a type I interferonopathy due to DNase II deficiency in humans.
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http://dx.doi.org/10.1038/s41467-017-01932-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5736616PMC
December 2017

Quality of life in children participating in a non-selective INR self-monitoring VKA-education programme.

Arch Cardiovasc Dis 2018 Mar 1;111(3):180-188. Epub 2017 Nov 1.

Pediatric Cardiology Department, AP-HP, Necker-Enfants malades, M3C National Reference Center, Paris Descartes University, Sorbonne Paris Cité, Paris, France.

Background: The quality of life (QoL) of children receiving vitamin K antagonist (VKA) treatment has been scarcely studied.

Aim: To assess QoL of children, and its evolution, throughout our non-selective international normalized ratio (INR) self-monitoring education programme.

Methods: Children and parents completed QoL questionnaires (Qualin, PedsQL) during education sessions. Scores were compared with those from controls.

Results: A total of 111 children (mean±standard deviation age 8.7±5.4 years) were included over a 3-year period. Indications for VKA treatment were congenital heart diseases (valve replacement [42.3%], total cavopulmonary connection [29.7%]), myocardiopathy (11.7%), coronary aneurysm (7.2%), venous/intracardiac thrombosis (4.5%), pulmonary artery hypertension (1.8%), arrhythmia (0.9%) and extra-cardiac disease (1.8%). Eighty children, 105 mothers and 74 fathers completed the QoL questionnaires. QoL was good among children aged 1-4 years and moderately impaired in those aged between 5 and 18 years. There was no significant relationship between self-reported QoL and patient's sex, type of VKA, number of group sessions attended, disease duration or time of diagnosis (prenatal or postnatal). QoL scores were significantly lower among children with congenital heart diseases compared with other diseases. There were few differences in QoL between children under transient VKA treatment and those treated for life. Parental proxy QoL scoring correlated well with but was significantly lower than child self-assessments. QoL reported by mothers increased throughout the education programme, independently of any improvement of the health condition.

Conclusions: This QoL study provides original data from a large cohort of children and their parents participating in a formalized INR self-monitoring education programme for VKA treatment.
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http://dx.doi.org/10.1016/j.acvd.2017.05.013DOI Listing
March 2018

Preanalytical influence of pneumatic tube delivery system on results of routine biochemistry and haematology analysis.

Ann Biol Clin (Paris) 2017 Dec;75(6):703-712

Service de biochimie générale.

Pneumatic tube delivery system (PTS) enables to reduce considerably turnaround times. The aim of the study was to assess the influence of the PTS on the quality of routine biochemical and hematological tests in our laboratory. Blood samples from 6 hospitalized patients and 8 healthy volunteers were analyzed. Blood samples were delivered to the laboratory by a PTS and by a human courier. We performed the following analysis: ionized calcium, sodium, potassium, lactate deshydrogenase (LDH), aspartate aminotransferase (ASAT), arterial blood gas, complete blood count and coagulation test as prothrombin time, activated partial thromboplastin time, factors V and VIII. Results were compared between the both method of transport according to the recommendation of the Société française de biologie clinique and the French committee for accreditation (SH-GTA01, norme NF ISO 5275-6). The hemolysis index of plasma was similar between the groups and no morphological differences were found on blood cells. For three samples, when delivered by PTS, LDH levels (two samples) and neutrophil polynuclear count (one sample) were above the recommended guidelines compared to those delivered by courier. Conversely, LDH levels and FVIII were below in two samples delivered by PTS. LDH levels, PNN count or factor VIII can be affected by PTS without the clinical interpretation being modified. We concluded that the PTS can be used to transport blood samples for routine biochemical and hematological analysis in our hospital.
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http://dx.doi.org/10.1684/abc.2017.1287DOI Listing
December 2017

Bleeding risk assessment in hemophilia A carriers from Dakar, Senegal.

Blood Coagul Fibrinolysis 2017 Dec;28(8):642-645

Department of Hematology, Cheikh Anta Diop University, Dakar, Senegal.

: Hemophilia A carriers have an abnormal X chromosome with a molecular abnormality of FVIII gene. These carriers, long considered to be free of bleeding risk, could have the same symptoms as mild hemophiliacs. This study aim to assess bleeding risk of hemophilia A carriers monitored at the Clinical Hematology Department of Dakar. This is a prospective study of a period of 6 months including 22 hemophilia A carriers aged between 8 and 48 years. Hemophilia carriers were recruited using the genealogical tree of hemophiliacs followed in the service. Their diagnosis was carried out by long range PCR and Sanger sequencing method searching the molecular abnormality responsible for hemophilia in their family. Bleeding risk was determined using a questionnaire consisting of different bleeding symptoms quoted from -1 to 4 according to the severity. Total of different values allow to determine the bleeding score which was pathological if it was greater than or equal to 1. Medium age was 22.5 years (8-48) (SD = 9.28). Four hemophilia A carriers (18.1%) presented bleeding symptoms and had a bleeding score at least 1 (P = 0.02). Menorrhagia was predominant (13.6%) followed by epistaxis (9%), gingivorrhagia (9%), and prolonged bleeding after tooth extraction (9%). Factor VIII level was lower in hemophilia carriers who presented bleeding (42 ± 8.61 UI/l) versus hemophilia carriers without bleeding (100 ± 50.95 UI/l) (P = 0.001). There was no significant correlation between bleeding occurrence and age (P = 0.81), activated patial thromboplastin time value (P = 0.97) and FVIII/Von Willebrand Factor ratio (P = 0.12). One in five hemophilia carriers presented bleeding and the questionnaire was effective to identify hemophilia carriers who had a risk of bleeding.
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http://dx.doi.org/10.1097/MBC.0000000000000653DOI Listing
December 2017

Inactivated antithrombins as fondaparinux antidotes: a promising alternative to haemostatic agents as assessed in vitro in a thrombin-generation assay.

Thromb Haemost 2016 08 14;116(3):452-60. Epub 2016 Jul 14.

E. P. Bianchini, UMR-S1176, 80 rue du Général Leclerc, 94276 Le Kremlin-Bicêtre Cedex, France, Tel.: +33 1 49595646, Fax: +33 1 46719472, E-mail:

In the absence of specific antidote to fondaparinux, two modified forms of antithrombin (AT), one recombinant inactive (ri-AT) and the other chemically inactivated (chi-AT), were designed to antagonise AT-mediated anticoagulants, e. g. heparins or fondaparinux. These inactive ATs were previously proven to effectively neutralise anticoagulant activity associated with heparin derivatives in vitro and in vivo, as assessed by direct measurement of anti-FXa activity. This study was undertaken to evaluate in vitro the effectivity of inactive ATs to reverse anticoagulation by heparin derivatives and to compare them with non-specific fondaparinux reversal agents, like recombinant-activated factor VII (rFVIIa) or activated prothrombin-complex concentrate (aPCC), in a thrombin-generation assay (TGA). Addition of fondaparinux (3 µg/ml) to normal plasma inhibited thrombin generation by prolonging lag time (LT) as much as 244 % and lowering endogenous thrombin potential (ETP) to 17 % of their control (normal plasma) values. Fondaparinux-anticoagulant activity was reversed by ri-AT and chi-AT, as reflected by the corrections of LT up to 117 % and 114 % of its control value, and ETP recovery to 78 % and 63 %, respectively. Unlike ri-AT that had no effect on thrombin generation in normal plasma, chi-AT retained anticoagulant activity that minimises its reversal capacity. However, both ATs were more effective than rFVIIa or aPCC at neutralising fondaparinux and, unlike non-specific antidotes, inactive ATs specifically reversed AT-mediated anticoagulant activities, as suggested by their absence of procoagulant activity in anticoagulant-free plasma.
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http://dx.doi.org/10.1160/TH15-12-0927DOI Listing
August 2016