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International audience; Background: Bleeding during oral anticoagulant therapy is currently codified by expert guidelines. Monitoring of coagulation during bleeding events is challenging. Our study sought to assess thrombin generation assay (TGA) in direct oral anticoagulant-treated patients without bleeding (WB), bleeding without reversal therapy (BR-), and bleeding with reversal therapy (BR+).Methods: We conducted a prospective, monocentric study from June 2015 to June 2018. For all bleeding groups, TGA was evaluated using platelet-poor plasma collected upon arrival at emergency (T0), and 30 min (T1), 6 h (T2) and 24 h (T3) after reversal therapy (if indicated) following activation by tissue factor 5 pM and phospholipids.Results: Overall, 292 patients participated, including 91 BR+, 94 BR-, and 107 WB patients. At T0, vitamin K antagonist reversed (VKA-BR+) patients experienced a significant decrease in TGA parameters (ETP and peak) compared with VKA without bleeding (VKA-WB). Compared with healthy controls, VKA-BR+ patients reversed by four-factor prothrombin complex concentrate (4F-PCC) displayed comparable TGA 's ETP and peak at T1, T2, and T3, whereas direct anti-Xa BR+ patients reversed by 4F-PCC or activated prothrombin complex concentrate (aPCC) reached thrombin generation parameters that exceeded normal range at T2 and T3.Conclusions: In VKA-treated patients reversed by 4F-PCC, TGA parameters were normalized, whereas in rivar-oxaban or apixaban-treated patients reversed by 4F-PCC or aPCC, TGA parameters exceeded normal range. Further studies are needed to compare the efficacy and safety of a different dose of reversal therapy and the impact on coagulation parameters.

Additional file 1: Table 1. Stability of samples collected in citrate-containing tubes for UFH monitoring. Table 2. Stability of samples collected in CTAD tubes for UFH monitoring. Table 3. Stability of blood samples collected in citrate-containing tubes for LMWH monitoring.

BACKGROUND: Storage of frozen plasma samples for hemostasis testing is a key step to obtain reliable results. Variables that can affect the quality of plasma during storage include the cryotube type and volume and the tube filling level that conditions the residual air volume. To date, there are only few data on which to base recommendations. OBJECTIVES: The aim of this study was to investigate the influence of the tube filling volume (20%, 40%, and 80%) of 2-mL microtubes on frozen plasma for a large panel of hemostasis assays. METHODS: For this study, 85 subjects were included, and blood samples were collected from them by venipuncture. After double centrifugation, each sample was aliquoted in 3 2-mL microtubes with different volumes (0.4, 0.8, and 1.6 mL) and stored at −80 °C. At the end of the frozen storage period (3 months ± 1 week), all aliquots from the sample were tested in the same analytical series for a large panel of hemostasis analyses. RESULTS: Compared with completely filled microtubes (1.6/2 mL), storing frozen plasma in smaller volumes (0.4/2 mL) significantly decreased prothrombin time and activated partial thromboplastin time. Conversely, factor II, V, VII, and X levels were increased. Antithrombin, Russell’s viper venom time, and anti-Xa activity in patients treated with heparin were also increased. CONCLUSION: To store plasma at −80 °C for hemostasis analysis, samples should be frozen in small-volume microtubes (

National audience

Flavonoids are the most widespread polyphenols in the plant kingdom. They are also associated to many of the beneficial effects of plant-based food and diet, especially in cardiovascular health. Among them, epicatechin, a flavonoid from the flavan-3-ols family is known for its antioxidant or anti-inflammatory properties and capacity to protect from a range of dysfunctions related to chronic diseases. Hemostasis, which contributes to the pathophysiology of cardiometabolic diseases, could also be modulated by flavonoids. The first part of this thesis was to study the impact of epicatechin on the three main stages of hemostasis which are primary hemostasis, plasma coagulation and fibrinolysis, using global tools to identify the hemostatic phenotype induced by epicatechin. We have shown that epicatechin at high concentration inhibits platelet function and modulates the coagulant and fibrinolytic profile. In the second part, we tried to determine the effect of epicatechin on microparticles and their procoagulant functions. While the overall microparticle concentration is not impacted by epicatechin, the latter appears to attenuate their procoagulant potential when detected by the formation of thrombin. Finally in the third part, we studied the final process of fibrin clot formation. Using microscopy, spectrophotometric and functional techniques, we were able to determine that the clot formed in the presence of epicatechin is less dense and more permeable. These in vitro data have yet to be confirmed in vivo to objectify the impact of epicatechin and its metabolites at concentrations reachable through dietary intakes., Les flavonoïdes sont les polyphénols les plus largement répandus dans le règne végétal. Ils sont également à l’origine de nombreux effets bénéfiques liés à l’alimentation notamment dans la santé cardiovasculaire. Parmi eux, l’épicatéchine, un flavonoïde de la famille des flavan-3-ols est connu pour ses propriétés antioxydantes ou anti-inflammatoires. L’hémostase, qui participe à la physiopathologie des maladies cardiométaboliques pourrait également être modulée par les flavonoïdes. La première partie de ces travaux de thèse a consisté à étudier l’impact de l’épicatéchine sur les trois principales étapes de l’hémostase que sont l’hémostase primaire, la coagulation plasmatique et la fibrinolyse, grâce à des outils globaux, afin de tenter de dégager le phénotype hémostatique induit par l’épicatéchine. Nous avons pu montrer qu’à forte concentration l’épicatéchine inhibe la fonction plaquettaire et module le profil coagulant et fibrinolytique. Dans la deuxième partie, nous avons tenté de déterminer l’effet de l’épicatéchine sur les microparticules et leurs fonctions procoagulantes. Alors que la concentration globale en microparticules n’est pas impactée par l’épicatéchine, cette dernière semble atténuer leur potentiel procoagulant lorsqu’il est détecté par la formation de thrombine. Enfin dans la troisième partie, nous avons étudié le processus final de formation du caillot de fibrine. Par des techniques de microscopie, spectrophotométrique et fonctionnelle nous avons pu déterminer que le caillot formé en présence d’épicatéchine est moins dense et plus perméable. Ces données in vitro restent encore à être confirmées in vivo afin d’objectiver l’impact de l’épicatéchine et de ses métabolites à des concentrations compatibles avec l’alimentation.

International audience; Le STA R Max® (Stago) est un coagulomètre à détection viscosimétrique. Il est doté d’un logiciel innovant (le système STA Coag Expert®) comportant un menu « accréditation » et permet une traçabilité totale de 5 ans. Le but de cette étude était d’évaluer ses performances avec les 3 méthodes de détection embarquées en vue d’une vérification de méthode selon la norme ISO 15189. Ce travail s’est déroulé au sein du service d’hématologie biologique du CHU de Clermont-Ferrand qui possède 4 STA R Max® répartis sur 2 sites et dédiés à l’activité de routine d’hémostase. Les paramètres étudiés étaient les suivants : le taux de prothrombine (TP), les temps de céphaline avec activateurs (TCA et TCK), le fibrinogène, l’activité anti-Xa et les D-dimères ; chacun testé dans les zones normales et pathologiques. Pour les analyses TP, TCA, TCK et fibrinogène, les coefficients de variation (CV) des répétabilités étaient inférieurs à 4,0 %. Les CV de répétabilité de l’activité anti-Xa et des D-dimères étaient respectivement au maximum de 4,0 % et 7,9 %. Les CV de la fidélité intermédiaire étaient inférieurs à 5,0 % pour TP, TCA, TCK et fibrinogène, 14,9 % pour l’activité anti-Xa et 8,6 % pour les D-dimères. Le biais (évaluant l’inexactitude) était inférieur à 8,7 % pour le TP, TCA et TCK, à 5,0 % pour le fibrinogène et inférieur à 15,5 % pour l’activité anti-Xa. L’étude de la concordance entre les 4 automates (corrélation intra-classe) retrouvait des coefficients de corrélation intra-classe de 0,99 pour tous les paramètres sauf le TCA (0,98). La concordance entre les automates était excellente. L’ensemble des résultats obtenus sont satisfaisants au regard des recommandations du Groupe français d’études sur l’hémostase et la thrombose (GFHT) et le fournisseur. Le STA R Max® est un analyseur performant pour gérer l’activité d’hémostase de routine d’un laboratoire et présente l’avantage de faciliter la démarche d’accréditation.

Cirrhosis significantly changes all hemostasis steps. Recent studies suggest that cirrhosis is associated with a coagulopathy leading to a hypercoagulable state. The underlying mechanisms are not fully understood, but protein C deficiency is probably a major determinant of this phenotype. The aim of this study was to compare the results of thrombin generation assays performed with addition of thrombomodulin or activated protein C to assess the effect of by-passing the protein C activation step in cirrhotic patients and healthy controls.Fifty-eight patients with cirrhosis and 26 healthy controls were prospectively included in this study. Thrombin generation was determined in platelet-poor plasma using 5 pM of tissue factor and 4 nM of phospholipids, without and with external addition of 1 nM thrombomodulin or 4 nM activated protein C. All results were normalized with the values of a pool of normal plasma samples to limit inter-plate variability.When thrombin generation assays were performed in the presence of thrombomodulin, endogenous thrombin potential (ETP) and ETP with/ETP without TM ratio were significantly higher in cirrhotic patients than in healthy controls (P 0.0001). Moreover, these values progressively increased with cirrhosis severity. When thrombin generation assays were performed with activated protein C, all thrombin generation parameters were comparable between healthy controls and cirrhotic patients, despite an acquired protein S deficiency.In the presence of activated protein C, no hypercoagulability was observed, adding to the current evidence that acquired protein C deficiency plays a key role in the coagulation imbalance.