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Papers by Cédric Garcia

Haematologica

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare acquired hemopathy (about 1.3 individuals per... more Paroxysmal nocturnal hemoglobinuria (PNH) is a rare acquired hemopathy (about 1.3 individuals per million incidence) characterized by hemolytic anemia and venous thrombosis. 1 The molecular defect involved is glycosylphosphatidylinositol (GPI) anchor loss related to X-linked PIGA gene mutations. Other genes responsible for the GPI anchor biosynthesis pathway could also be involved, 2-5 and, research on their involvement in the pathophysiogenesis of PNH and its borderline forms is being uncovered. Here, we report a case of essential thrombocythemia (ET) caused by a somatic mutation in MPL (c.1544G>T:p.W515L) shortly preceded by copy-neutral loss of heterozygosity (CN-LOH) in cis on chromosome 1, leading to homozygosity of both the acquired MPL missense mutation and an inherited heterozygous stop-gain mutation in PIGV (c.1405C>T:p.R469X) causing PNH. Clinical presentation In July 2017, a 73-year-old Caucasian patient was referred to the cardiology intensive care untit due to pulmonary embolism associated with cardiogenic shock. Thrombolysis was performed, followed by anticoagulant therapy. Active neoplasm and the antiphospholipid syndrome were excluded. The persistence of thrombocytosis after 3 months (platelets >600 10 9 /L) without any sign of infection or iron deficiency, justified a myelogram revealing ET with a clone size of 74% (Figure 1). Concomitantly, glomerulopathy was identified (urinary protein to creatinine ratio [uPCr] 5.3 g/g; serum albumin 33 g/L; 39 urinary red cells/mL). Renal biopsy showed interstitial fibrosis with focal segmental glomerulosclerosis (FSGS). Causes of secondary FSGS were ruled out. Increased level of the nephrotic syndrome-associated soluble urokinase-type plasminogen activated receptor (suPAR) was identified (731 pg/mL). Blood tests showed hemolysis (lactate dehydrogenase 1,270 IU/L, undetectable haptoglobin) without schistocytes or positive Coombs test (Figure 1). PNH was suspected then confirmed by flow cytometry (80.4% CD14-CD55-FLAERmonocytes and 88.8% CD24-CD16-CD55-FLAERgranulocytes) with an 88% clone size at Q4 2017 (Figure 1). Thus, this patient developed a massive pulmonary embolism, with two underlying clonal myeloid disorders (ET and PNH) and a nephrotic syndrome. After 24 months of treatment, combining hydroxycarbamide (6,500 mg/week) and the C5 complement inhibitor eculizumab (900 mg/2 weeks), the clones size dramatically decreased, cell counts normalized, hemolysis stopped, and uPCr decreased below 2 g/g

Blood Advances

Mild thrombocytopenia, changes in platelet gene expression, enhanced platelet functionality, and ... more Mild thrombocytopenia, changes in platelet gene expression, enhanced platelet functionality, and presence of platelet-rich thrombi in the lung have been associated with thromboinflammatory complications of patients with COVID-19. However, whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) gets internalized by platelets and directly alters their behavior and function in infected patients remains elusive. Here, we investigated platelet parameters and the presence of viral material in platelets from a prospective cohort of 29 patients with severe COVID-19 admitted to an intensive care unit. A combination of specific assays, tandem mass spectrometry, and flow cytometry indicated high levels of protein and lipid platelet activation markers in the plasma from patients with severe COVID-19 associated with an increase of proinflammatory cytokines and leukocyte-platelets interactions. Platelets were partly desensitized, as shown by a significant reduction of αIIbβ3 activati...

Journal of NeuroInterventional Surgery

BackgroundIntracranial occlusion recanalization fails in 20% of endovascular thrombectomy procedu... more BackgroundIntracranial occlusion recanalization fails in 20% of endovascular thrombectomy procedures, and thrombus composition is likely to be an important factor. In this study, we demonstrate that the combination of electrical impedance spectroscopy (EIS) and machine learning constitutes a novel and highly accurate method for the identification of different human thrombus types.Methods134 samples, subdivided into four categories, were analyzed by EIS: 29 ‘White’, 26 ‘Mixed’, 12 ‘Red’ thrombi, and 67 liquid ‘Blood’ samples. Thrombi were generated in vitro using citrated human blood from five healthy volunteers. Histological analysis was performed to validate the thrombus categorization based on red blood cell content. A machine learning prediction model was trained on impedance data to differentiate blood samples from any type of thrombus and in between the four sample categories.ResultsHistological analysis confirmed the similarity between the composition of in vitro generated thr...

Cells

Platelets are mainly known for their key role in hemostasis and thrombosis. However, studies over... more Platelets are mainly known for their key role in hemostasis and thrombosis. However, studies over the last two decades have shown their strong implication in mechanisms associated with inflammation, thrombosis, and the immune system in various neoplastic, inflammatory, autoimmune, and infectious diseases. During sepsis, platelets amplify the recruitment and activation of innate immune cells at the site of infection and contribute to the elimination of pathogens. In certain conditions, these mechanisms can lead to thromboinflammation resulting in severe organ dysfunction. Here, we discuss the interactions of platelets with leukocytes, neutrophil extracellular traps (NETs), and endothelial cells during sepsis. The intrinsic properties of platelets that generate an inflammatory signal through the NOD-like receptor family, pyrin domain-containing 3 (NLRP3) inflammasome are discussed. As an example of immunothrombosis, the implication of platelets in vaccine-induced immune thrombotic thr...

Journal of Thrombosis and Haemostasis, 2020

Essentials-Compared to first generation Btk inhibitors, the novel Btk inhibitor tirabrutinib has ... more Essentials-Compared to first generation Btk inhibitors, the novel Btk inhibitor tirabrutinib has only a modest inhibitory effect on platelet activation in vitro and ex vivo. Accepted Article This article is protected by copyright. All rights reserved-The novel Syk inhibitor entospletinib efficiently inhibits washed platelet aggregation in response to collagen, but weakly affects platelet activation in platelet-richplasma and in whole blood.-Combination of tirabrutinib and entospletinib induces a significant decrease in platelet response to collagen in vitro and ex vivo compared to each agent alone and may expose patients to bleeding risk.

Advances in Biological Regulation, 2020

The novel Corona virus infection (Covid-19) first identified in China in December 2019 has rapidl... more The novel Corona virus infection (Covid-19) first identified in China in December 2019 has rapidly progressed in pandemic leading to significant mortality and unprecedented challenge for healthcare systems. Although the clinical spectrum of Covid-19 is variable, acute respiratory failure and systemic coagulopathy are common in severe Covid-19 patients. Lung is an important target of the SARS-CoV-2 virus causing eventually acute respiratory distress syndrome associated to a thromboinflammatory state. The cytokinic storm, thromboinflammation and pulmonary tropism are the bedrock of tissue lesions responsible for acute respiratory failure and for prolonged infection that may lead to multiple organ failure and death. The thrombogenicity of this infectious disease is illustrated by the high frequency of thromboembolic events observed even in Covid-19 patients treated with anticoagulation. Increased D-Dimers, a biomarker reflecting activation of hemostasis and fibrinolysis, and low platelet count (thrombocytopenia) are associated with higher mortality in Covid-19 patients. In this review, we will summarize our current knowledge on the thromboembolic manifestations, the disturbed hemostatic parameters, and the thromboinflammatory conditions associated to Covid-19 and we will discuss the modalities of anticoagulant treatment or other potential antithrombotic options.

International Journal of Stroke, 2021

Background The composition of the thrombus influences its retrievability by mechanical thrombecto... more Background The composition of the thrombus influences its retrievability by mechanical thrombectomy. Purpose Our study aimed to report on thrombi resistant to aspiration, regarding susceptibility vessel sign and histologic composition. Methods This observational study was based on a prospective database of acute anterior circulation ischemic strokes treated by mechanical thrombectomy. Endovascular first-line strategy was aspiration and in case of failure, combined therapy-rescue was performed. The positivity of susceptibility vessel sign (SVS+) or its negativity (SVS−) was assessed on T2* sequences. The thrombus composition was analyzed with hematoxylin eosin staining. Results Histological analysis was performed on 102 clots. Thrombi with SVS− were significantly richer in fibrin/platelets, p = 0.04. Out of 210 mechanical thrombectomy, aspiration first pass strategy was performed in 131/210 (62%) patients. Combined therapy-rescue was needed in 37% of aspiration first pass strategy ca...

TH Open, 2019

Transcatheter aortic valve implantation (TAVI) is an established treatment option for symptomatic... more Transcatheter aortic valve implantation (TAVI) is an established treatment option for symptomatic patients with severe aortic valve stenosis (AS). During and early after the procedure, both ischemic events (predominantly stroke) and bleedings remain prevalent. The optimal antithrombotic regimen is still debated. Single- versus dual-antiplatelet therapy is associated with a lower rate of severe bleeding, without difference in thrombotic complications. Although platelets have been empirically targeted, little is known on their contribution to these events primarily related to embolization of thrombotic material and tissue-derived debris from the wounded aortic valve and large vessels. The objective of this study was to assess local platelet activation in blood sampled in the ascending aorta immediately before and within minutes postimplantation. A series of 18 patients with AS on monotherapy with aspirin successfully underwent TAVI with the self-expandable Medtronic CoreValve by trans...

Fundamental & Clinical Pharmacology, 2020

A 71‐year‐old man is admitted for nose bleeds recurring for several days. His medical background ... more A 71‐year‐old man is admitted for nose bleeds recurring for several days. His medical background shows in particular major depression for which he has been receiving sertraline for several years. The workup shows anemia, and no anomalies on head and neck CT angiography. However, further explorations suggest an acquired thrombopathy that could have contributed to the bleeding. During sertraline exposure, platelet functional exploration and platelet secretion were abnormal. Sertraline is often used as first‐line treatment of depression. Pharmacological data and spontaneous notifications suggest increased potential risk with sertraline. It appears necessary to pay attention to bleeding with sertraline use.

International Journal of Molecular Sciences, 2019

Host defense against infection is based on two crucial mechanisms: the inflammatory response and ... more Host defense against infection is based on two crucial mechanisms: the inflammatory response and the activation of coagulation. Platelets are involved in both hemostasis and immune response. These mechanisms work together in a complex and synchronous manner making the contribution of platelets of major importance in sepsis. This is a summary of the pathophysiology of sepsis-induced thrombocytopenia, microvascular consequences, platelet-endothelial cells and platelet–pathogens interactions. The critical role of platelets during sepsis and the therapeutic implications are also reviewed.

Scientific Reports, 2018

Sepsis is associated with thrombocytopenia and microvascular thrombosis. Studies have described p... more Sepsis is associated with thrombocytopenia and microvascular thrombosis. Studies have described platelets implication in this pathology but their kinetics of activation and behavior remain poorly known. We show in a mouse model of peritonitis, the appearance of platelet-rich thrombi in organ microvessels and organ damage. Complementary methods are necessary to characterize platelet activation during sepsis as circulating soluble markers and platelet-monocyte aggregates revealed early platelet activation, while surface activation markers were detected at later stage. A microfluidic based ex-vivo thrombosis assay demonstrated that platelets from septic mice have a prothrombotic behavior at shear rate encountered in microvessels. Interestingly, we found that even though phosphoinositide-3-kinase β−deficient platelet mice formed less thrombi in liver microcirculation, peritoneal sepsis activates a platelet alternative pathway to compensate the otherwise mandatory role of this lipid-kina...

Journal of Thrombosis and Haemostasis, 2019

First description of an IgM monoclonal antibody causing a IIb b 3 integrin activation and acquire... more First description of an IgM monoclonal antibody causing a IIb b 3 integrin activation and acquired Glanzmann thrombasthenia associated with macrothrombocytopenia.

Journal of Thrombosis and Haemostasis, 2018

Essentials • HDL subclasses were studied in acute coronary syndrome (ACS). • HDL2 from ACS patien... more Essentials • HDL subclasses were studied in acute coronary syndrome (ACS). • HDL2 from ACS patients have better antiplatelet potency than HDL from non ACS subjects. • ACS remodels the antiplatelet properties of HDL subclasses. • Oxidized polyunsaturated fatty acids content of HDL is modified by ACS. Summary. Background: Although HDLs have antithrombotic effects by reducing platelet activation, the relationship between HDL levels and the risk of acute coronary syndrome (ACS) is unclear, as HDL particles are heterogeneous in composition and biological properties. Objective: To characterize the effects of HDL2 and HDL3 subclasses from ACS patients and non-coronary artery disease (CAD) subjects on platelet activation. Methods: We measured platelet aggregation and ex vivo thrombus formation, analyzed signaling pathways by flow cytometry, and performed a targeted lipidomics analysis on HDL subclasses. Results: Analysis of human platelet aggregation in suspension, adhesion on von Willebrand factor and thrombus formation on collagen under arterial shear demonstrated that HDL2 from ACS patients had higher antiplatelet potency than HDL3 from ACS patients and HDL from non-CAD subjects. HDL binding to scavenger receptor class B type I was essential for this effect. A lipidomics analysis revealed that HDL2 from ACS patients had more oxidized polyunsaturated fatty acids (PUFAs). An inverse correlation between the concentrations of 9-hydroxyoctadecadienoic acid (9-HODE), 13-hydroxyoctadecadienoic acid (13-HODE), the eicosapentaenoic acid metabolite 18-hydroxyeicosapentaenoic acid (18-HEPE) and hydroxyeicosatetraenoic acid isomers in HDL2 and platelet aggregation was observed. This relationship was further demonstrated by the direct inhibitory effects of 18-HEPE, 9-HODE, 13-HODE, 17hydroxydocosahexaenoic acid and 14-hydroxydocosahexaenoic acid on collagen-related peptide-induced platelet aggregation, indicating that oxidized PUFAs contribute to the antithrombotic effect of ACS HDL2. Conclusions: Our data shed new light on the antiplatelet effects of HDL subclasses, and suggest physiological adaptation through the modulation of HDL properties in ACS patients that may limit their platelet-dependent thrombotic risk.

Haematologica, 2019

Two groups in the healthy population and in chronic lymphocytic leukemia patients were characteri... more Two groups in the healthy population and in chronic lymphocytic leukemia patients were characterized by low or high sensitivity to ibrutinib in terms of platelet aggregation response to collagen in platelet-rich-plasma in vitro.-The better profile of acalabrutinib being mainly observed in the ibrutinib low sensitive group, replacement therapy in patients may not systematically reduce the bleeding risk, particularly in patients taking anti-platelet drugs.

Stroke, 2019

Background and Purpose— Our goal was to evaluate whether the presence of a low signal intensity k... more Background and Purpose— Our goal was to evaluate whether the presence of a low signal intensity known as susceptibility vessel sign (SVS) on T2*-gradient echo imaging sequence was predictive of arterial recanalization and the early clinical improvement after mechanical thrombectomy. Methods— This observational study was based on a prospective database of acute ischemic strokes treated by mechanical thrombectomy. Inclusion criteria were patients with acute anterior ischemic stroke, diagnosed by magnetic resonance imaging, including a T2*-gradient echo imaging sequence, and treated by mechanical thrombectomy. Two independent readers assessed the presence of an SVS. Successful recanalization was defined as a Thrombolysis in Cerebral Infarction score of 2b-3 after mechanical thrombectomy. Early clinical improvement was estimated by the difference between the baseline National Institutes of Health Stroke Scale and the National Institutes of Health Stroke Scale on day 1 after treatment Re...

Molecular and cellular endocrinology, Jan 5, 2018

Estetrol (E4) is a natural estrogen synthesized exclusively during pregnancy by the human fetal l... more Estetrol (E4) is a natural estrogen synthesized exclusively during pregnancy by the human fetal liver, and the physiological role of this hormone is unknown. Interestingly, E4 was recently evaluated in preclinical and phase II-III clinical studies in combination with a progestin, with the advantage to not increase the circulating level of coagulation factors, at variance to oral estradiol or ethinylestradiol. Here, we evaluated the effect of E4 on hemostasis and thrombosis in mouse. Following chronic E4 treatment, mice exhibited a prolonged tail-bleeding time and were protected from arterial and also venous thrombosis in vivo. In addition, E4 treatment decreased ex vivo thrombus growth on collagen under arterial flow conditions. We recently showed that E4 activates uterine epithelial proliferation through nuclear estrogen receptor (ER) α. To analyze the impact of nuclear ERα actions on hemostasis and thrombosis, we generated hematopoietic chimera with bone marrow cells deficient for...

Cellular and Molecular Life Sciences, 2018

P2Y 12 receptor (P2Y 12-R) is one of the major targets for drug inhibiting platelet aggregation i... more P2Y 12 receptor (P2Y 12-R) is one of the major targets for drug inhibiting platelet aggregation in the treatment/prevention of arterial thrombosis. However, the clinical use of P2Y 12-R antagonists faces some limitations, such as a delayed onset of action (clopidogrel) or adverse effect profile (ticagrelor, cangrelor), justifying the development of a new generation of P2Y 12-R antagonists with a better clinical benefit-risk balance. Although the recent concept of biased agonism offers the possibility to alleviate undesirable adverse effects while preserving therapeutic outcomes, it has never been explored at P2Y 12-R. For the first time, using highly sensitive BRET2-based probes, we accurately delineated biased ligand efficacy at P2Y 12-R in living HEK293T cells on G protein activation and downstream effectors. We demonstrated that P2Y 12-R displayed constitutive Gi/o-dependent signaling that is impaired by the R122C mutation, previously associated with a bleeding disorder. More importantly, we reported the biased inverse agonist efficacy of cangrelor and ticagrelor that could underlie their clinical efficacy. Our study points out that constitutive P2Y 12-R signaling is a normal feature of the receptor that might be essential for platelets to respond faster to a vessel injury. From a therapeutic standpoint, our data suggest that the beneficial advantages of antiplatelet drugs might be more related to inverse agonism at P2Y 12-R than to antagonism of ADP-mediated signaling. In the future, deciphering P2Y 12-R constitutive activity should allow the discovery of more selective biased P2Y 12-R blockers demonstrating therapeutic advantages over classical antiplatelet drugs by improving therapeutic outcomes and concomitantly relieving undesirable adverse effects.