Publications by authors named "Sylvie Cointe"

12 Publications

  • Page 1 of 1

Granulocyte microvesicles with a high plasmin generation capacity promote clot lysis and improve outcome in septic shock.

Blood 2022 Jan 13. Epub 2022 Jan 13.

AP-HM, France.

Microvesicles (MVs) have previously been shown to exert profibrinolytic capacity, which is increased in patients with septic shock (SS) with a favorable outcome. We therefore hypothesized that the plasmin generation capacity (PGC) could confer to MVs a protective effect supported by their capacity to lyse a thrombus, and we investigated the mechanisms involved. Using a MV-PGC kinetic assay, ELISA and flow cytometry, we found that granulocyte MVs (Gran-MVs) from SS patients display a heterogeneous PGC profile driven by the uPA (urokinase)/uPAR system. In vitro, these MVs lyse a thrombus according to their MV-PGC levels in a uPA/uPAR-dependent manner, as shown in a fluorescent clot lysis test and a lysis front retraction assay. Fibrinolytic activators conveyed by MVs contribute to approximately 30% of the plasma plasminogenolytic capacity of SS patients. In a murine model of SS, the injection of high PGC Gran-MVs significantly improved mouse survival and reduced the number of thrombi in vital organs. This was associated with a modification of the mouse coagulation and fibrinolysis properties toward a more fibrinolytic profile. Interestingly, mouse survival was not improved when soluble uPA was injected. Finally, using a multiplex array on plasma from SS patients, we found that neutrophil elastase correlates with the effect of high-PGC-capacity plasma and modulates the Gran-MV plasmin generation capacity by cleaving uPA-PAI-1 complexes. In conclusion, we show that high PGC level displayed by Gran-MVs reduce thrombus formation and improve survival conferring to Gran-MVs a protective role in a murine model of sepsis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2021013328DOI Listing
January 2022

The Interaction of Platelets with Colorectal Cancer Cells Inhibits Tumor Growth but Promotes Metastasis.

Cancer Res 2020 01 14;80(2):291-303. Epub 2019 Nov 14.

Aix Marseille Univ, INSERM, INRA, Marseille, France.

Platelets promote metastasis, however, their role in tumor growth remains controversial. Here, we investigated the effect of platelet interactions with colorectal tumor cells. Platelets extravasated into the tumor microenvironment and interacted with tumor cells in a cadherin-6-dependent manner. The interaction induced platelet spreading, release of their granule content, and the generation of three types of microparticles (iMP) that expressed platelet markers, tumor markers, or both. The presence of iMPs was confirmed in colorectal cancer tissue specimens. Platelets significantly reduced tumor growth and increased intratumoral macrophages. This was mediated by iMP recruitment of macrophages via the chemoattractants RANTES, MIF, CCL2, and CXCL12 and activation of their tumor cell killing capacity through IFNγ and IL4, which led to cell-cycle arrest of tumor cells in a p21-dependent manner. In contrast, in the bloodstream, iMPs activated endothelial cells and platelets and induced epithelial-to-mesenchymal transition of tumor cells, promoting metastasis. Altogether, these results indicate that depending on the environment, local or bloodstream, the consequences of the interactions between platelets and a tumor may promote or prevent cancer progression. SIGNIFICANCE: Tumor cell interaction with platelets produces chimeric extracellular vesicles that suppress primary tumor growth by activating tumor-eliminating macrophages, while promoting metastasis through EMT and endothelial activation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/0008-5472.CAN-19-1181DOI Listing
January 2020

Comparison of the Response to Rituximab between Myelin Oligodendrocyte Glycoprotein and Aquaporin-4 Antibody Diseases.

Ann Neurol 2020 02 27;87(2):256-266. Epub 2019 Nov 27.

Public Assistance Hospitals of Marseille, Department of Neurology, Timone Hospital, Marseille.

Objective: To compare response to rituximab (RTX) between adult patients positive for myelin oligodendrocyte glycoprotein (MOG) and aquaporin-4 (AQP4) antibodies.

Methods: We prospectively studied adult patients with MOG or AQP4 antibodies who received RTX under an individualized dosing schedule adapted to the biological effect of RTX monitored by memory B-cell measurement. Memory B cells were counted monthly and when relapse occurred. The biological effect of RTX was considered significant with <0.05% memory B cells in peripheral blood lymphocytes.

Results: In 16 patients with MOG antibodies and 29 with AQP4 antibodies, mean follow-up was 19 (range = 9-38) and 38 (13-79) months. Under RTX, 10 relapses occurred in 6 of 16 (37.5%) patients with MOG antibodies, and 13 occurred in 7 of 29 (24%) with AQP4 antibodies. The median time of relapse after the most recent infusion was 2.6 (0.6-5.8) and 7 (0.8-13) months, respectively (p < 0.001). Memory B cells had reemerged in 2 of 10 (20%) relapses in patients with MOG antibodies and 12 of 13 (92.5%) with AQP4 antibodies (p < 0.001).

Interpretation: In AQP4 antibody-associated disorder, relapse mostly occurs when the biological effect of RTX decreases, which argues for treatment efficacy. In MOG antibody-associated disorder, the efficacy of RTX is not constant, because one-third of patients showed relapse despite an effective biological effect of RTX. In this subpopulation, memory B-cell depletion was unable to prevent relapse, which was probably caused by different immunological mechanisms. These findings should be used to improve treatment strategies for MOG antibody-associated disorder. ANN NEUROL 2020;87:256-266.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/ana.25648DOI Listing
February 2020

A new assay to evaluate microvesicle plasmin generation capacity: validation in disease with fibrinolysis imbalance.

J Extracell Vesicles 2018 16;7(1):1494482. Epub 2018 Jul 16.

Aix-Marseille Université, C2VN, UMR-1263, INSERM, INRA 1260, UFR de Pharmacie, Marseille, France.

Among extracellular vesicles, leukocyte-derived microvesicles (LMVs) have emerged as complex vesicular structures. Primarily identified as procoagulant entities, they were more recently ascribed to plasmin generation capacity (MV-PGC). The objectives of this work were (1) to develop a new hybrid bio-assay combining the specific isolation of LMVs and measurement of their PGC, and compare its performance to the original method based on centrifugation, (2) to validate MV-PGC in septic shock, combining increased levels of LMVs and fibrinolytic imbalance. Using plasma sample spiked with LMVs featuring different levels of PGC, we demonstrated that CD15-beads specifically extracted LMVs. The MV dependency of the test was demonstrated using electron microscopy, high speed centrifugation, nanofiltration and detergent-mediated solubilization and the MV-PGC specificity using plasmin-specific inhibitors, or antibodies blocking elastase or uPA. Thanks to a reaction booster (ε-ACA), we showed that the assay was more sensitive and reproducible than the original method. Moreover, it exhibited a good repeatability, inter-operator and inter-experiment reproducibility. The new immunomagnetic bio-assay was further validated in patients with septic shock. As a result, we showed that MV-PGC values were significantly lower in septic shock patients who died compared to patients who survived, both at inclusion and 24 h later (1.4 [0.8-3.0] 3.1 [1.7-18]  × 10/min,  = 0.02; 1.4 [1-1.6] 5.2 [2.2-16]  × 10/min,  = 0.004). Interestingly, combining both MV-PGC and PAI-1 in a ratio significantly improved the predictive value of PAI-1. This strategy, a hybrid capture bioassay to specifically measure LMV-PGC using for the first time, opens new perspectives for measuring subcellular fibrinolytic potential in clinical settings with fibrinolytic imbalance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/20013078.2018.1494482DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6052415PMC
July 2018

Increased serum levels of fractalkine and mobilisation of CD34CD45 endothelial progenitor cells in systemic sclerosis.

Arthritis Res Ther 2017 03 20;19(1):60. Epub 2017 Mar 20.

Internal Medicine Department, Assistance Publique-Hôpitaux de Marseille (APHM), CHU Nord, 13015, Marseilles, France.

Background: The disruption of endothelial homeostasis is a major determinant in the pathogenesis of systemic sclerosis (SSc) and is reflected by soluble and cellular markers of activation, injury and repair. We aimed to provide a combined assessment of endothelial markers to delineate specific profiles associated with SSc disease and its severity.

Methods: We conducted an observational, single-centre study comprising 45 patients with SSc and 41 healthy control subjects. Flow cytometry was used to quantify circulating endothelial microparticles (EMPs) and CD34 progenitor cell subsets. Colony-forming unit-endothelial cells (CFU-ECs) were counted by culture assay. Circulating endothelial cells were enumerated using anti-CD146-based immunomagnetic separation. Blood levels of endothelin-1, vascular endothelial growth factor (VEGF) and soluble fractalkine (s-Fractalkine) were evaluated by enzyme-linked immunosorbent assay. Disease-associated markers were identified using univariate, correlation and multivariate analyses.

Results: Enhanced numbers of EMPs, CFU-ECs and non-haematopoietic CD34CD45 endothelial progenitor cells (EPCs) were observed in patients with SSc. Patients with SSc also displayed higher serum levels of VEGF, endothelin-1 and s-Fractalkine. s-Fractalkine levels positively correlated with CD34CD45 EPC numbers. EMPs, s-Fractalkine and endothelin-1 were independent factors associated with SSc. Patients with high CD34CD45 EPC numbers had lower forced vital capacity values. Elevated s-Fractalkine levels were associated with disease severity, a higher frequency of pulmonary fibrosis and altered carbon monoxide diffusion.

Conclusions: This study identifies the mobilisation of CD34CD45 EPCs and high levels of s-Fractalkine as specific features of SSc-associated vascular activation and disease severity. This signature may provide novel insights linking endothelial inflammation and defective repair processes in the pathogenesis of SSc.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13075-017-1271-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5359964PMC
March 2017

Thrombospondin-1-Derived Peptide RFYVVMWK Improves the Adhesive Phenotype of CD34 Cells From Atherosclerotic Patients With Type 2 Diabetes.

Cell Transplant 2017 02 6;26(2):327-337. Epub 2016 Oct 6.

CD34+ progenitor cells are growing in use for vascular repair. However, in diabetic individuals with cardiovascular diseases, these cells have dysfunctional engraftment capabilities, which compromise their use for autologous cell therapy. The thrombospondin-1-derived peptide RFYVVMWK has previously been reported to stimulate cell adhesiveness through CD47 and integrin activation pathways. Our aim was to test whether RFYVVMWK preconditioning could modulate CD34+ cell phenotype and enhance its proadhesive properties in diabetic patients. Peripheral blood mononuclear CD34+ cells isolated from 40 atherosclerotic patients with type 2 diabetes (T2D; n = 20) or without (non-T2D; n = 20) were preconditioned with 30 μM RFYVVMWK or truncated peptide RFYVVM. CD34+ cell adhesion was assessed on a vitronectin-collagen matrix and on TNF-α or IL-1β-stimulated HUVEC monolayers. Adhesion receptors, platelet/CD34+ cell conjugates, and cell viability were analyzed by flow cytometry and confocal microscopy. RFYVVMWK increased the adhesion of T2D CD34+ cells by eightfold to the vitronectin-collagen matrix (p < 0.001) corresponding to a threefold increase compared to unstimulated non-T2D CD34+ cells. The peptide induced the formation of platelet/CD34+ conjugates and increased the expression of TSP-1, CD29, CD51/CD61, and CD62P in both T2D and non-T2D cells. However, RFYVVMWK treatment did not affect the viability/apoptosis of CD34+ progenitor cells. In conclusion, priming CD34+ cells with RFYVVMWK may enhance their vascular engraftment during autologous proangiogenic cell therapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3727/096368916X693329DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5657766PMC
February 2017

Microparticles and Fibrinolysis.

Semin Thromb Hemost 2017 Mar 6;43(2):129-134. Epub 2016 Dec 6.

Division of Hematology/Oncology, Northwestern University Feinberg School of Medicine, Chicago, Illinois.

Microparticles (MPs) are submicronic vesicles which are formed by budding of the cellular membrane of virtually any cell type in response to cell activation or apoptosis. Both circulating MPs and MPs generated within tissues harbor molecules with a large repertoire of biological activities and transfer material to target cells. Depending on their cellular origin, the stimuli triggering their formation, or their localization, they may participate in the maintenance of organ or vascular homeostasis as well as inducing dysfunction. MPs have mostly been described as having procoagulant properties. However, the fact that some MP subsets are able to efficiently generate plasmin suggests that the role of MPs in hemostasis is more complex than initially thought. In this review, we summarize key findings showing that MPs provide a heterogeneous catalytic surface for plasmin generation, according to their cellular origin. We further address the specific features of the MP-dependent fibrinolytic system. Potential consequences of this MP-associated fibrinolytic activity in pathology are illustrated in cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1055/s-0036-1592301DOI Listing
March 2017

Detection of EpCAM-positive microparticles in pleural fluid: A new approach to mini-invasively identify patients with malignant pleural effusions.

Oncotarget 2016 Jan;7(3):3357-66

VRCM, UMR-S1076, Aix-Marseille Université, INSERM, Faculté de Pharmacie, Marseille, France.

Pleural biomarkers allowing to mini-invasively discriminate benign from malignant pleural effusions are needed. Among potential candidates, microparticles (MPs) are extracellular vesicles that vectorize antigen derived from the parent cell. We hypothesized that tumor-derived MPs could be present in the pleural liquid and help to identify patients with malignant pleural effusions. Using highly sensitive flow cytometry and cryo-electron microscopy, we showed that large amounts of MPs from hematopoïetic and vascular origin could be detectable in pleural fluids. Their level did not differ between benign (n = 14) and malignant (n = 71) pleural effusions. Analysis of selected tumoral associated antigens (podoplanin, mucin 1 and EpCAM, epithelial-cell-adhesion-molecule) evidenced for the first time the presence of tumor-derived MPs expressing EpCAM in malignant pleural fluids only (Specificity = 93%, Sensitivity = 49% and 45% for flow cytometry and ELISA, respectively). The detection of EpCAM-positive-MPs (EpCAM + MPs) by flow cytometry showed a better specificity and sensitivity than ELISA to distinguish between pleural carcinoma and the others malignant pleural effusions (MPE; Sp: 96% vs 89%; Se: 79% vs 66%). Combining EpCAM+ MPs and cytology improved the diagnosis of MPE compared to cytology alone. This study establishes the basis for using EpCAM+ MPs as a promising new biomarker that could be added to the armamentarium to mini-invasively identify patients with malignant pleural effusions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.6581DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4823111PMC
January 2016

Maintenance chemotherapy in children with ALL exerts metronomic-like thrombospondin-1 associated anti-endothelial effect.

Oncotarget 2015 Sep;6(26):23008-14

Aix-Marseille Université INSERM, Vascular Research Center of Marseille UMRS-1076, Marseille, France.

Maintenance chemotherapy is an important part of the treatment of ALL in children. It relies on the long-term oral administration of daily low-dose mercaptopurin and weekly low-dose methotrexate. Although it has been used in the clinic for decades, its mechanisms of action remain unclear. Here, we investigated different angiogenic and immune biomarkers to gain insights into the mechanisms of action of maintenance therapy in children with ALL. We thus monitored circulating endothelial cells (CEC), endothelial progenitor cells (EPC) and endothelial microparticles (EMP), pro-angiogenic factors (VEGF, VEGFR-1 and Ang-2), anti-angiogenic factor thrombospondin-1 (THBS1) and regulatory T lymphocytes (Treg) in 47 children with ALL during the maintenance phase of their treatment (at treatment initiation and after 6, 12 and 18 months). We observed a statistically significant decrease in EPC and EMP counts throughout the maintenance phase associated with a significant increase in THBS1 levels. No significant change was detected in other angiogenic markers or in Treg numbers.The results presented here indicate that maintenance therapy in children with ALL exerts its antitumor activity at least in part through anti-angiogenic effects, similar to those induced by metronomic chemotherapy. Larger studies are now warranted to validate these findings and determine their clinical implications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.3984DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4673217PMC
September 2015

Platelet and not erythrocyte microparticles are procoagulant in transfused thalassaemia major patients.

Br J Haematol 2015 Nov 24;171(4):615-24. Epub 2015 Jul 24.

Centre de Référence Maladies Rares Thalassémies, Marseille-Lyon, Service d'Hémato-Oncologie Pédiatrique, Hôpital de la Timone, APHM, Marseille, France.

The level of circulating platelet-, erythrocyte-, leucocyte- and endothelial-derived microparticles detected by high-sensitivity flow cytometry was investigated in 37 β-thalassaemia major patients receiving a regular transfusion regimen. The phospholipid procoagulant potential of the circulating microparticles and the microparticle-dependent tissue factor activity were evaluated. A high level of circulating erythrocyte- and platelet-microparticles was found. In contrast, the number of endothelial microparticles was within the normal range. Platelet microparticles were significantly higher in splenectomized than in non-splenectomized patients, independent of platelet count (P < 0·001). Multivariate analysis indicated that phospholipid-dependent procoagulant activity was influenced by both splenectomy (P = 0·001) and platelet microparticle level (P < 0·001). Erythrocyte microparticles were not related to splenectomy, appear to be devoid of proper procoagulant activity and no relationship between their production and haemolysis, dyserythropoiesis or oxidative stress markers could be established. Intra-microparticle labelling with anti-HbF antibodies showed that they originate only partially (median of 28%) from thalassaemic erythropoiesis. In conclusion, when β-thalassaemia major patients are intensively transfused, the procoagulant activity associated with thalassaemic erythrocyte microparticles is probably diluted by transfusions. In contrast, platelet microparticles, being both more elevated and more procoagulant, especially after splenectomy, may contribute to the residual thrombotic risk reported in splenectomized multi-transfused β-thalassaemia major patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/bjh.13609DOI Listing
November 2015

Ticagrelor increases endothelial progenitor cell level compared to clopidogrel in acute coronary syndromes: A prospective randomized study.

Int J Cardiol 2015 31;187:502-7. Epub 2015 Mar 31.

Vascular Research Center of Marseille, INSERM UMR-S 1076, Aix-Marseille Université, Marseille, France; Laboratoire d'Hématologie et de Biologie Vasculaire, Assistance-Publique Hôpitaux de Marseille, Centre hospitalo-universitaire de la Conception, Marseille, France.

Background: The clinical benefit of ticagrelor compared to clopidogrel in ACS patients suggested off-target property. Such pleiotropic effect could be mediated by circulating endothelial progenitor cells (EPC) which are critical for vascular healing. We aimed to investigate the impact of ticagrelor on EPC in acute coronary syndrome (ACS) patients undergoing percutaneous coronary intervention (PCI).

Methods: We prospectively randomized 106 ACS patients to ticagrelor or clopidogrel. Sub-populations of CD34+ circulating progenitor cells (PC) were analyzed by flow cytometry allowing one to determine the levels of CD34+ PC, CD34+CD45+ Hematopoietic PC, CD34+133+ immature PC and CD34+KDR+ EPC on admission and at 1 month. Changes in PC level were calculated as the difference between 1 month and baseline value.

Results: The 2 groups were similar regarding baseline characteristics including PC numbers on admission. The 2 groups had similar change in overall CD34+ PC and hematopoietic CD34+45+ PC level (p=0.2). On the contrary, when considering CD34+133+ PC and CD34+KDR+ EPC, we observed that patients treated by ticagrelor had a significantly higher increase in levels of these PC subtypes compared to those treated by clopidogrel (0.23 (-0.33; 0.79) vs 0.00 (-0.5; 0.34); p=0.04 and 0.01 (-0.04; 0.05) vs -0.01 (-0.06; 0.03); p=0.02). Changes in the level of CD34+CD133+ PC correlated with platelet activity measured by the VASP index (r=-0.30; p=0.008). By contrast the increase in the level of CD34+KDR+ EPC in the ticagrelor group was independent of platelet activity.

Conclusions: Ticagrelor increases the number of EPC in ACS patients suggesting a benefit on endothelial regeneration that may participate in the pleiotropic property of the drug.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijcard.2015.03.414DOI Listing
March 2016

Requirements for membrane attack complex formation and anaphylatoxins binding to collagen-activated platelets.

PLoS One 2011 Apr 15;6(4):e18812. Epub 2011 Apr 15.

Department of Medicine, Montreal Heart Institute and University of Montreal, Montreal, Quebec, Canada.

Background: The activation of complement during platelet activation is incompletely understood.

Objectives: We sought to explore the formation of C5b-9 and anaphylatoxins binding to collagen-activated platelets.

Methods: C5b-9, anaphylatoxins C3a, C4a and C5a, and anaphylatoxin receptors C3aR1 and C5aR were measured by flow cytometry and/or confocal microscopy. Platelet microparticles were quantified by flow cytometry, and their C5b-9 content was determined by western blot analyses. In all experiments, sodium citrate was used for blood anticoagulation.

Results: C5b-9 rapidly formed on the platelet surface following activation with collagen, TRAP, ADP or A23187, but was surprisingly restricted to a subset of platelets (1 to 15%) independently of P-selectin or phosphatidylserine exposure. Following collagen activation, C5b-9-positive platelets in thrombi were found associated with collagen fibres. C5b-9 formation was obliterated by Mg(2+)-EGTA and significantly reduced by the thrombin inhibitor hirudin (-37%, p<0.05), but was unaffected by chondroitinase, compstatin, SCH79797 (PAR-1 inhibitor), or in the PRP of a MBL-deficient donor. Compstatin and Mg(2+)-EGTA, but not hirudin, SCH79797 or chondroitinase, inhibited the formation of collagen-induced microparticles (-71% and -44%, respectively, p<0.04). These microparticles contained greater amounts of C5b-9 compared with the other agonists. Platelet activation by collagen or convulxin resulted in the strong binding of anaphylatoxins and the exposure of receptors C3aR1 and C5aR (CD88) on their surface.

Conclusions: C5b-9 formation on collagen-activated platelets is i) partially controlled by thrombin, ii) restricted to a subset of platelets, and iii) can occur without P-selectin expression or phosphatidylserine exposure. Activated platelets bind anaphylatoxins on their surface and express C3a and C5a receptors, which may contribute to the localization of inflammatory processes during thrombosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0018812PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3078139PMC
April 2011
-->