Publications by authors named "Pierre Fontana"

134 Publications

A universal anti-Xa assay for rivaroxaban, apixaban, and edoxaban measurements: method validation, diagnostic accuracy and external validation.

Br J Haematol 2021 May 6. Epub 2021 May 6.

Department of Clinical Chemistry, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland.

A universal anti-Xa assay for the determination of rivaroxaban, apixaban and edoxaban drug concentrations would simplify laboratory procedures and facilitate widespread implementation. Following two pilot studies analysing spiked samples and material from 698 patients, we conducted a prospective multicentre cross-sectional study, including 867 patients treated with rivaroxaban, apixaban or edoxaban in clinical practice to comprehensively evaluate a simple, readily available anti-Xa assay that would accurately measure drug concentrations and correctly predict relevant levels in clinical practice. Anti-Xa activity was measured by an assay calibrated with low-molecular-weight heparin (LMWH) in addition to ultra-high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). As an external validation, LMWH-calibrated anti-Xa activity was also determined in nine external laboratories. The LMWH-calibrated anti-Xa activity correlated strongly with rivaroxaban, apixaban or edoxaban drug levels [r  = 0·98, 95% confidence interval (CI) 0·98-0·98]. The sensitivity for the clinically relevant cut-off levels of 30, 50 and 100 µg/l was 96·2% (95% CI 94·4-97·4), 96·4% (95% CI 94·4-97·7) and 96·7% (95% CI 94·3-98·1) respectively. Concordant results were obtained in the external validation study. In conclusion, a universal, LMWH-calibrated anti-Xa assay accurately measured rivaroxaban, apixaban and edoxaban concentrations and correctly predicted relevant drug concentrations in clinical practice.
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http://dx.doi.org/10.1111/bjh.17470DOI Listing
May 2021

miR-204-5p and platelet function regulation: insight into a mechanism mediated by CDC42 and GPIIbIIIa.

Thromb Haemost 2021 May 3. Epub 2021 May 3.

Geneva Platelet Group Geneva, Faculty of Medicine, University of Geneva, Geneva, Switzerland.

Background: Several platelet-derived miRNAs are associated with platelet reactivity (PR) and clinical outcome in cardiovascular patients. We previously showed an association between miR-204-5p and PR in stable cardiovascular patients, but data on functional mechanisms are lacking.

Aims: To validate miR-204-5p as a regulator of PR in platelet-like structures (PLS) derived from human megakaryocytes and to address mechanistic issues.

Methods: Human hematopoietic stem cells were differentiated into megakaryocytes, enabling the transfection of miR-204-5p and the recovery of subsequent PLS. The morphology of transfected megakaryocytes and PLS was characterized using flow cytometry and microscopy. The functional impact of miR-204-5p was assessed using a flow assay, the quantification of the activated form of the GPIIbIIIa receptor and a fibrinogen-binding assay. qPCR and western blot were used to evaluate the impact of miR-204-5p on a validated target, CDC42. The impact of CDC42 modulation was investigated using a silencing strategy.

Results: miR-204-5p transfection induced cytoskeletal changes in megakaryocytes associated with the retracted protrusion of proPLS, but it had no impact on the number of PLS released. Functional assays showed that the PLS produced by megakaryocytes transfected with miR-204-5p were more reactive than controls. This phenotype is mediated by the regulation of GPIIbIIIa expression, a key contributor in platelet-fibrinogen interaction. Similar results were obtained after CDC42 silencing, suggesting that miR-204-5p regulates PR, at least in part, via CDC42 downregulation.

Conclusions: We functionally validated miR-204-5p as a regulator of the PR that occurs through CDC42 downregulation and regulation of fibrinogen receptor expression.
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http://dx.doi.org/10.1055/a-1497-9649DOI Listing
May 2021

An Ex Vivo and In Silico Study Providing Insights into the Interplay of Circulating miRNAs Level, Platelet Reactivity and Thrombin Generation: Looking beyond Traditional Pharmacogenetics.

J Pers Med 2021 Apr 21;11(5). Epub 2021 Apr 21.

Geneva Platelet Group, Faculty of Medicine, University of Geneva, 1205 Geneva, Switzerland.

Platelet reactivity (PR), a key pharmacodynamic (PD) component of the action of antiplatelet drugs in cardiovascular disease (CVD) patients, is highly variable. PR is associated with occurrence or recurrence of thrombotic and bleeding events, but this association is modulated by several factors. Conventional pharmacogenetics explains a minor part of this PR variability, and among determinants of PR, circulating microRNAs (miRNAs) have been the focus of attention during these last years as biomarkers to predict PR and clinical outcomes in CVD. This being said, the impact of miRNAs on platelet function and the mechanisms behind it are largely unknown. The level of a set of candidate miRNAs including miR-126-3p, miR-150-5p, miR-204-5p and miR-223-3p was quantified in plasma samples of stable CVD patients and correlated with PR as assessed by light-transmission aggregometry and in vivo thrombin generation markers. Finally, miRNA target networks were built based on genes involved in platelet function. We show that all candidate miRNAs were associated with platelet aggregation, while only miR-126-3p and miR-223-3p were positively correlated with in vivo thrombin generation markers. In silico analysis identified putative miRNA targets involved in platelet function regulation. Circulating miRNAs were associated with different aspects of platelet reactivity, including platelet aggregation and platelet-supported thrombin generation. This paves the way to a personalized antithrombotic treatment according to miRNA profile in CVD patients.
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http://dx.doi.org/10.3390/jpm11050323DOI Listing
April 2021

The ISTH bleeding assessment tool as predictor of bleeding events in inherited platelet disorders: Communication from the ISTH SSC Subcommittee on Platelet Physiology.

J Thromb Haemost 2021 May;19(5):1364-1371

Department of Internal Medicine, IRCCS Policlinico S. Matteo Foundation, University of Pavia, Pavia, Italy.

Background: The ISTH Bleeding Assessment Tool (ISTH-BAT) has been validated for clinical screening of suspected von Willebrand disease (VWD) and for bleeding prediction. Recently it has been validated for subjects with inherited platelet disorders (IPD) (BAT-VAL study).

Objectives: To determine whether the ISTH-BAT bleeding score (BS) predicts subsequent bleeding events requiring treatment in IPD patients.

Methods: Patients with IPD, type 1 VWD (VWD-1) and age- and sex-matched healthy controls enrolled in the BAT-VAL study were prospectively followed-up for 2 years and bleeding episodes requiring treatment were recorded.

Results: Of the 1098 subjects initially enrolled, 955 were followed-up and 124 suffered hemorrhages during follow-up, 60% of whom had inherited platelet function disorders (IPFD). Total number of events was significantly higher in IPFD (n = 235) than VWD-1 (n = 52) or inherited thrombocytopenia (IT; n = 20). Events requiring transfusions were 66% in IPFD, 5.7% in VWD-1, and 3% in IT. Baseline BS was significantly higher in IPFD patients with a bleeding event at follow-up than in those without (p < .01) and the percentage of subjects suffering a bleeding event increased proportionally to baseline BS quartile. A significant association between the BS and the chance of suffering severe bleeding was found in the overall, IPFD, and VWD-1 populations. Similar results were obtained for the pediatric population.

Conclusions: Inherited platelet function disorder patients with high BS at enrollment are more likely to suffer from bleeding events requiring treatment at follow-up. Moreover, the higher the baseline BS quartile the greater the incidence of subsequent events, suggesting that independently from diagnosis a high BS is associated with a greater risk of subsequent hemorrhage.
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http://dx.doi.org/10.1111/jth.15263DOI Listing
May 2021

COVID-19 patients often show high-titer non-platelet-activating anti-PF4/heparin IgG antibodies.

J Thromb Haemost 2021 05 7;19(5):1294-1298. Epub 2021 Apr 7.

Institut für Immunologie und Transfusionsmedizin, Universitätsmedizin Greifswald, Greifswald, Germany.

Background: Heparin-induced thrombocytopenia (HIT) is a severe adverse reaction to heparin caused by heparin-dependent, platelet-activating anti-platelet factor 4 (PF4)/heparin antibodies. Heparin is a cornerstone of treatment in critically ill COVID-19 patients. HIT antibodies can be detected by antigen tests and functional tests. Often strong reactivity in the antigen test is used as a surrogate marker for the presence of clinically relevant, platelet-activating antibodies. We observed an unexpectedly high percentage of COVID-19 patients, clinically suspected to have HIT, with high titer anti-PF4/heparin antibodies, but a negative functional test.

Objective: We investigated whether in COVID-19 patients a serum-derived factor inhibits the heparin-induced platelet activation test (HIPA).

Methods And Results: Twelve COVID-19 patients with suspected HIT were tested. Three samples tested negative in all assays; nine samples tested positive by antigen tests, among which only three tested also positive by HIPA. When we spiked COVID-19 serum or control serum with the human HIT antibody like monoclonal antibody 5B9, reactivity of 5B9 remained the same. Also, the purified IgG fractions of COVID-19 sera testing strongly positive in the PF4/heparin antigen test but negative in the functional test did not show increased reactivity in the functional test in comparison to the original serum. Both results make a functionally inhibitory factor in the serum/plasma of COVID-19 patients highly unlikely.

Conclusion: COVID-19 patients often present with strong reactivity in PF4/heparin antigen tests without the presence of platelet-activating antibodies. Diagnosis of HIT requires confirmation of heparin-dependent, platelets activating antibodies to avoid overdiagnosis and overtreatment with non-heparin anticoagulants.
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http://dx.doi.org/10.1111/jth.15262DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8013750PMC
May 2021

[Towards new indications for direct oral anticoagulants].

Rev Med Suisse 2021 Jan;17(720-1):20-23

Service d'angiologie et d'hémostase, HUG, 1211 Genève 14.

Direct oral anticoagulants (DOAC) represent commonly prescribed drugs in everyday clinical practice for indications such as atrial fibrillation, prevention of venous thromboembolic disease (VTE) after major orthopaedic surgery, treatment of deep vein thrombosis and pulmonary embolism, and long term prevention of VTE recurrence. More recently, the efficacy of DOAC has been demonstrated in new clinical situations, such as heparin-induced thrombocytopenia, cancer-associated VTE and secondary prevention of cardiovascular events in patients with atherothrombotic arterial disease. This article's aims is to present the recent data on which these new indications are based.
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January 2021

Management of bleeding events and invasive procedures in patients with haemophilia A without inhibitors treated with emicizumab.

Swiss Med Wkly 2020 Dec 18;150:w20422. Epub 2020 Dec 18.

Department of Haematology and Central Haematology Laboratory, Inselspital, Bern University Hospital, University of Bern, Switzerland.

Introduction: Emicizumab (Hemlibra®, Hoffmann-La Roche, Switzerland) is now available for haemophilia A patients with or without factor VIII inhibitors. Management of bleeding events and replacement therapy for invasive procedures have to be adapted.

Objective: To provide a practical guidance for the management of breakthrough bleeding events and elective or urgent surgery in adult and paediatric patients with haemophilia A without inhibitors treated with emicizumab.

Methods: Based on the available literature and the experiences collected from adult and paediatric patients treated in Switzerland, the Working Party on Haemostasis of the Swiss Society of Haematology and the Swiss Haemophilia Network worked together to reach a consensus on the management of bleeding events and invasive procedures.

Results And Conclusion: Minor bleeding events and invasive procedures associated with low bleeding risk can be treated without factor replacement therapy in most cases, whereas major bleeding events and high-risk surgery require additional factor VIII replacement at usual doses, at least for the first days. Emicizumab treatment should be continued throughout the procedure and during the postoperative period. Elective major surgery should be planned according to emicizumab dosing for patients with a once-a-month posology. Of note, so far only few data are available on the management of major bleeds and surgery in patients with haemophilia A treated with emicizumab and this practical guidance will have to be regularly updated with growing experience.  .
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http://dx.doi.org/10.4414/smw.2020.20422DOI Listing
December 2020

[Contraception and venous thromboembolism].

Rev Med Suisse 2020 Dec;16(718):2383-2386

Service de gynécologie, Groupe hospitalier Paris Saint-Joseph, INSERM, Obstetrical, perinatal and pediatric epidemiology research team, Center for epidemiology and statistics, Rue Raymond-Losserand 185, 75014 Paris.

Combined oral contraceptives remain in 2020 the most used contraceptive method in Switzerland and Europe, and are found in about half of venous thromboembolism (VTE) occurring in women aged up to 50 years. In this narrative review, we describe the determinants of the VTE risk, related to the types of oral contraceptives and to genetic or acquired risk factors of users, while summarizing several current recommendations of prescription for contraceptives. The complex management of contraception at the time of VTE should be discussed with patients, in order to minimize the risks of undesired pregnancy, abnormal uterine bleeding and recurrent VTE.
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December 2020

Implémentation des stratégies diagnostiques de l’embolie pulmonaire : encore et toujours un défi.

Rev Med Suisse 2020 12;16(718):2359-2360

Service d'angiologie et d'hémostase, HUG, Genève.

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December 2020

Estimating the risk thresholds used by guidelines to recommend postpartum thromboprophylaxis.

J Thromb Haemost 2021 02 17;19(2):452-459. Epub 2020 Dec 17.

Faculty of Medicine, University of Geneva, Geneva, Switzerland.

Background: Guidelines for postpartum thromboprophylaxis are mostly expert based. Acceptable postpartum venous thromboembolism (VTE) risk thresholds to justify the use of thromboprophylaxis are ill defined.

Objective: To compare the proportion of postpartum women with recommended thromboprophylaxis according to the Royal College of Obstetricians and Gynecologists (RCOG), American College of Obstetricians and Gynecologists (ACOG), and American College of Chest Physicians (ACCP) guidelines, and to estimate their threshold of risk of VTE.

Methods: We collected maternal, obstetrical, and fetal characteristics among all women who delivered at the Geneva University Hospitals in January 2019 through medical chart review. We determined for each woman the recommended strategy of thromboprophylaxis according to each guideline. We indirectly estimated individual absolute VTE risks using a validated risk prediction model (Sultan).

Results: Among 344 women (mean age 32.2 years), with 23.3% of cesarean deliveries (CD), the RCOG guideline categorized 40.1% of all deliveries, 25.4% of vaginal deliveries (VD), and 88.8% of CD as requiring thromboprophylaxis. The ACOG and ACCP guidelines, which focus on CD, categorized fewer women as high risk (35.0% and 40.0% of CD, respectively). The mean estimated risk of VTE was 0.07% in all participants, 0.12% in those with recommended thromboprophylaxis according to the RCOG, and 0.20% among women after CD with recommended thromboprophylaxis by the ACOG and ACCP guidelines.

Conclusions: Our data highlight important variations in the proportion of thromboprophylaxis recommendation between guidelines. Risk thresholds to recommend the use of heparin appear very low, and may correlate with a high, and perhaps undesirable, number needed to treat to prevent VTE events.
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http://dx.doi.org/10.1111/jth.15166DOI Listing
February 2021

Methods to Investigate miRNA Function: Focus on Platelet Reactivity.

Thromb Haemost 2021 Apr 29;121(4):409-421. Epub 2020 Oct 29.

Geneva Platelet Group, Faculty of Medicine, University of Geneva, Geneva, Switzerland.

MicroRNAs (miRNAs) are small noncoding RNAs modulating protein production. They are key players in regulation of cell function and are considered as biomarkers in several diseases. The identification of the proteins they regulate, and their impact on cell physiology, may delineate their role as diagnostic or prognostic markers and identify new therapeutic strategies. During the last 3 decades, development of a large panel of techniques has given rise to multiple models dedicated to the study of miRNAs. Since plasma samples are easily accessible, circulating miRNAs can be studied in clinical trials. To quantify miRNAs in numerous plasma samples, the choice of extraction and purification techniques, as well as normalization procedures, are important for comparisons of miRNA levels in populations and over time. Recent advances in bioinformatics provide tools to identify putative miRNAs targets that can then be validated with dedicated assays. In vitro and in vivo approaches aim to functionally validate candidate miRNAs from correlations and to understand their impact on cellular processes. This review describes the advantages and pitfalls of the available techniques for translational research to study miRNAs with a focus on their role in regulating platelet reactivity.
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http://dx.doi.org/10.1055/s-0040-1718730DOI Listing
April 2021

Mixing Drugs and Genetics: A Complex Hemorrhagic Cocktail.

Am J Med 2021 03 25;134(3):e211-e212. Epub 2020 Aug 25.

Division of General Internal Medicine, Geneva University Hospitals, Switzerland; Geneva Platelet Group, Faculty of Medicine, University of Geneva, Switzerland.

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http://dx.doi.org/10.1016/j.amjmed.2020.07.032DOI Listing
March 2021

Studies on hemostasis in COVID-19 deserve careful reporting of the laboratory methods, their significance, and their limitations.

J Thromb Haemost 2020 11;18(11):3121-3124

Université catholique de Louvain, CHU UCL Namur, Hematology Laboratory, Namur Thrombosis and Hemostasis Center (NTHC), NARILIS, Yvoir, Belgium.

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http://dx.doi.org/10.1111/jth.15061DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7436855PMC
November 2020

Venous thromboembolism in COVID-19: systematic review of reported risks and current guidelines.

Swiss Med Wkly 2020 06 21;150:w20301. Epub 2020 Jun 21.

Division of Angiology and Haemostasis, Geneva University Hospitals and Faculty of Medicine, Geneva, Switzerland.

Aims Of The Study: Many centres have noticed a high number of venous thromboembolism (VTE) events among critically ill inpatients with COVID-19 pneumonia. The aims of this study were (1) to summarise the reported risk of VTE associated with COVID-19 infections and (2) to summarise guidance documents on thromboprophylaxis in COVID-19 patients, in a systematic review.

Methods: We systematically searched for peer-reviewed evidence on the risk of VTE in patients with COVID-19, in PubMed, Embase and Twitter, and for guidelines or guidance documents for thromboprophylaxis, from international or national societies relevant to the field of thrombosis and haemostasis, up to April 30 2020.

Results: We found 11 studies (1 clinical trial, 7 retrospective cohorts and 3 prospective cohorts), which included a range of 16 to 388 in patients with COVID-19 (total of 1369 inpatients). The diagnoses of COVID-19 and VTE were of high quality, but the follow-up was often unclear. Most studies reported universal in-hospital thromboprophylaxis. Among all inpatients and among intensive care unit (ICU) inpatients with COVID-19, reported risks of VTE were 4.4–8.2% (three studies) and 0–35.3% (six studies), respectively. Two studies at least partially screened for VTE in ICU inpatients with COVID-19, and found risks of 24.7–53.8%. We found 12 guidelines for thromboprophylaxis of COVID-19 patients. The majority suggested universal pharmacological thromboprophylaxis in all COVID-19 inpatients, but there was heterogeneity in the suggested intensity of thromboprophylaxis: seven advised considering intensified doses of heparin according to the clinical or biological severity of the disease, especially in the ICU setting.

Conclusions: Venous thromboembolism very commonly complicates the clinical course of inpatients with COVID-19, despite thromboprophylaxis. The risk appears highest among critically ill inpatients. We found no estimates of risks among outpatients. Many questions remain unresolved, as delineated by the heterogeneity of national and international guidelines. This situation calls for fast randomised clinical trials, comparing different schemes of thromboprophylaxis in COVID-19 inpatients.
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http://dx.doi.org/10.4414/smw.2020.20301DOI Listing
June 2020

MicroRNA-126 is a regulator of platelet-supported thrombin generation.

Platelets 2020 Aug 12;31(6):746-755. Epub 2020 Jun 12.

Geneva Platelet Group, Faculty of Medicine, University of Geneva , Geneva, Switzerland.

Circulating microRNA (miRNA) expression profiles correlate with platelet reactivity. MiR-126 is a promising candidates in this regard. We generated a transgenic zebrafish line with thrombocyte-specific overexpression of miR-126. Laser injury of the posterior cardinal vein of 5 day-old larvae was performed with or without antithrombotic pre-treatment. Platelet-like structures (PLS) derived from human megakaryocytes transfected with miR-126 were also evaluated for procoagulant activity. Finally, we studied the correlation between miR-126 level and thrombin generation markers in a cohort of stable cardiovascular patients. Control zebrafish developed small thrombocyte-rich thrombi at the site of vessel injury, without vessel occlusion. The miR-126 transgenic line developed an occluding thrombus in 75% (95% CI: 51-91%) of larvae. Pre-treatment with the direct thrombin inhibitor argatroban, but not aspirin, prevented vessel occlusion in the transgenic line (0% occlusion, 95%CI: 0-18%). Upon activation, human PLS showed an increased procoagulant profile after miR-126 transfection compared to control. Finally, the plasma levels of miR-126, but not a control platelet-derived miRNA, correlated with markers of thrombin generation in a cohort of 185 cardiovascular patients. Our results from three complementary approaches support a key role for miR-126 in platelet-supported thrombin generation and open new avenues in the tailoring of antithrombotic treatment.
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http://dx.doi.org/10.1080/09537104.2020.1775804DOI Listing
August 2020

Genomewide Association Study of Platelet Reactivity and Cardiovascular Response in Patients Treated With Clopidogrel: A Study by the International Clopidogrel Pharmacogenomics Consortium.

Clin Pharmacol Ther 2020 Nov 9;108(5):1067-1077. Epub 2020 Jul 9.

Department of Medicine and Program for Personalized and Genomic Medicine, University of Maryland, Baltimore, Maryland, USA.

Antiplatelet response to clopidogrel shows wide variation, and poor response is correlated with adverse clinical outcomes. CYP2C19 loss-of-function alleles play an important role in this response, but account for only a small proportion of variability in response to clopidogrel. An aim of the International Clopidogrel Pharmacogenomics Consortium (ICPC) is to identify other genetic determinants of clopidogrel pharmacodynamics and clinical response. A genomewide association study (GWAS) was performed using DNA from 2,750 European ancestry individuals, using adenosine diphosphate-induced platelet reactivity and major cardiovascular and cerebrovascular events as outcome parameters. GWAS for platelet reactivity revealed a strong signal for CYP2C19*2 (P value = 1.67e-33). After correction for CYP2C19*2 no other single-nucleotide polymorphism reached genomewide significance. GWAS for a combined clinical end point of cardiovascular death, myocardial infarction, or stroke (5.0% event rate), or a combined end point of cardiovascular death or myocardial infarction (4.7% event rate) showed no significant results, although in coronary artery disease, percutaneous coronary intervention, and acute coronary syndrome subgroups, mutations in SCOS5P1, CDC42BPA, and CTRAC1 showed genomewide significance (lowest P values: 1.07e-09, 4.53e-08, and 2.60e-10, respectively). CYP2C19*2 is the strongest genetic determinant of on-clopidogrel platelet reactivity. We identified three novel associations in clinical outcome subgroups, suggestive for each of these outcomes.
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http://dx.doi.org/10.1002/cpt.1911DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7689744PMC
November 2020

Platelet Transforming Growth Factor-β1 Induces Liver Sinusoidal Endothelial Cells to Secrete Interleukin-6.

Cells 2020 05 25;9(5). Epub 2020 May 25.

Faculty of Science and Medicine, Section of Medicine, University of Fribourg, Route Albert-Gockel 1, 1700 Fribourg, Switzerland.

The roles and interactions of platelets and liver sinusoidal endothelial cells in liver regeneration are unclear, and the trigger that initiates hepatocyte proliferation is unknown. We aimed to identify the key factors released by activated platelets that induce liver sinusoidal endothelial cells to produce interleukin-6 (IL-6), a cytokine implicated in the early phase of liver regeneration. We characterized the releasate of activated platelets inducing the in vitro production of IL-6 by mouse liver sinusoidal endothelial cells and observed that the stimulating factor was a thermolabile protein. Following gel filtration, a single fraction of activated platelet releasate induced a maximal IL-6 secretion by liver sinusoidal endothelial cells (90.2 ± 13.9 versus control with buffer, 9.0 ± 0.8 pg/mL, < 0.05). Mass spectroscopy analysis of this fraction, followed by in silico processing, resulted in a reduced list of 18 candidates. Several proteins from the list were tested, and only recombinant transforming growth factor β1 (TGF-β1) resulted in an increased IL-6 production up to 242.7 ± 30.5 pg/mL, which was comparable to non-fractionated platelet releasate effect. Using neutralizing anti-TGF-β1 antibody or a TGF-β1 receptor inhibitor, IL-6 production by liver sinusoidal endothelial cells was dramatically reduced. These results support a role of platelet TGF-β1 β1 in the priming phase of liver regeneration.
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http://dx.doi.org/10.3390/cells9051311DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7290849PMC
May 2020

Platelet Interactions with Liver Sinusoidal Endothelial Cells and Hepatic Stellate Cells Lead to Hepatocyte Proliferation.

Cells 2020 05 18;9(5). Epub 2020 May 18.

Faculty of Science and Medicine, University of Fribourg, 1700 Fribourg, Switzerland.

(1) Background: Platelets were postulated to constitute the trigger of liver regeneration. The aim of this study was to dissect the cellular interactions between the various liver cells involved in liver regeneration and to clarify the role of platelets. (2) Methods: Primary mouse liver sinusoidal endothelial cells (LSECs) were co-incubated with increasing numbers of resting platelets, activated platelets, or platelet releasates. Alterations in the secretion of growth factors were measured. The active fractions of platelet releasates were characterized and their effects on hepatocyte proliferation assessed. Finally, conditioned media of LSECs exposed to platelets were added to primary hepatic stellate cells (HSCs). Secretion of hepatocyte growth factor (HGF) and hepatocyte proliferation were measured. After partial hepatectomy in mice, platelet and liver sinusoidal endothelial cell (LSEC) interactions were analyzed in vivo by confocal microscopy, and interleukin-6 (IL-6) and HGF levels were determined. (3) Results: Co-incubation of increasing numbers of platelets with LSECs resulted in enhanced IL-6 secretion by LSECs. The effect was mediated by the platelet releasate, notably a thermolabile soluble factor with a molecular weight over 100 kDa. The conditioned medium of LSECs exposed to platelets did not increase proliferation of primary hepatocytes when compared to LSECs alone but stimulated hepatocyte growth factor (HGF) secretion by HSCs, which led to hepatocyte proliferation. Following partial hepatectomy, in vivo adhesion of platelets to LSECs was significantly increased when compared to sham-operated mice. Clopidogrel inhibited HGF secretion after partial hepatectomy. (4) Conclusion: Our findings indicate that platelets interact with LSECs after partial hepatectomy and activate them to release a large molecule of protein nature, which constitutes the initial trigger for liver regeneration.
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http://dx.doi.org/10.3390/cells9051243DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7290338PMC
May 2020

[Venous thrombotic risk related to SARS-CoV-2 : prevalence, recommendations and perspectives].

Rev Med Suisse 2020 May;16(692):951-954

Service d'angiologie et d'hémostase, HUG et Faculté de médecine, 1205 Genève.

Since the SARS-CoV-2 pandemic outbreak, growing evidence suggests that patients suffering from COVID-19 are at increased risk of thrombotic events. The sepsis-related activation of the coagulation combined with a high prevalence of common thrombotic risk factors could contribute to this prothrombotic state. Coagulation biomarkers could help in the identification of patients at risk of complications and mortality. The incidence of venous thromboembolic events appears to be increased, especially in severe COVID-19 patients. Based on that knowledge, several societies have provided recommendation on the prevention of venous thromboembolism. In this narrative review, we summarize available epidemiologic data on venous thromboembolism and recommendations on thromboprophylaxis in COVID-19.
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May 2020

Recommendations on the use of anticoagulants for the treatment of patients with heparin-induced thrombocytopenia in Switzerland.

Swiss Med Wkly 2020 04 24;150:w20210. Epub 2020 Apr 24.

Division of Hematology and Central Hematology Laboratory, Cantonal Hospital of Lucerne and University of Bern, Switzerland.

Heparin-induced thrombocytopenia (HIT) is an immune-mediated adverse drug effect that occurs in 0.1–5% of heparin treated patients. Management of acute HIT currently involves (1) cessation of heparin exposure, and (2) inhibition of coagulation with an anticoagulant other than heparin. Several anticoagulants can be considered for the treatment of HIT. Anticoagulant monitoring, management of drug-induced adverse events including bleeding, and therapeutic dosing schedules in selected clinical settings represent challenges to the clinician treating HIT patients. Moreover, the fact that not all registered anticoagulants are approved for HIT in Switzerland further complicates the management of HIT. The present recommendations on the anticoagulant treatment of HIT in Switzerland have been elaborated by a panel of Swiss experts belonging to the Working Party Hemostasis (WPH) of the Swiss Society of Hematology (SGH-SSH). They are intended to support clinicians in their decision making when treating HIT patients.
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http://dx.doi.org/10.4414/smw.2020.20210DOI Listing
April 2020

A Unique Case of Acquired Hemophilia A Presenting with Transient Ischemic Attack.

Acta Haematol 2021 10;144(1):88-90. Epub 2020 Mar 10.

Division of Angiology and Haemostasis, Geneva University Hospitals and Faculty of Medicine, Geneva, Switzerland,

Acquired hemophilia A (AHA) is a rare but serious condition, usually associated with significant spontaneous or traumatic bleeding and a high mortality rate. In this report, we describe the case of an elderly patient presenting a transient ischemic attack concurrently with AHA. A thrombotic event in AHA is occasionally associated with the use of bypassing agents for treatment, but a spontaneous thrombotic event has not ever been described.
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http://dx.doi.org/10.1159/000506273DOI Listing
March 2021

Contribution of exome sequencing to the identification of genes involved in the response to clopidogrel in cardiovascular patients.

J Thromb Haemost 2020 06 20;18(6):1425-1434. Epub 2020 Mar 20.

Geneva Platelet Group, Faculty of Medicine, University of Geneva, Geneva, Switzerland.

Background: On-clopidogrel platelet reactivity (PR) is associated with the risk of thrombotic or bleeding event in selected populations of high-risk patients. PR is a highly heritable phenotype and a few variants of cytochrome genes, essentially CYP2C19, are associated with PR but only explain 5% to 12% of the variability.

Objective: The aim of this study is to delineate genetic determinants of on-clopidogrel PR using high-throughput sequencing.

Methods: We performed a whole exome sequencing of 96 low- and matched high-PR patients in a discovery cohort. Exomes from genes with variants significantly associated with PR were sequenced in 96 low- and matched high-PR patients from an independent replication cohort.

Results: We identified 585 variants in 417 genes with an adjusted P value < .05. In the replication cohort, all top variants including CYP2C8, CYP2C18, and CYP2C19 from the discovery population were found again. An original network analysis identified several candidate genes of potential interest such as a regulator of PI3K, a key actor in the downstream signaling pathway of the P2Y receptor.

Conclusion: This study emphasizes the role of CYP-related genes as major regulators of clopidogrel response, including the poorly investigated CYP2C8 and CYP2C18.
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http://dx.doi.org/10.1111/jth.14776DOI Listing
June 2020

Platelet Function Test Use for Patients with Coronary Artery Disease in the Early 2020s.

J Clin Med 2020 Jan 10;9(1). Epub 2020 Jan 10.

Geneva Platelet Group, Faculty of Medicine, University of Geneva, 1206 Geneva, Switzerland.

In the field of antithrombotics, precision medicine is of particular interest, as it may lower the incidence of potentially life-threatening side effects. Indeed, antiplatelet drugs such as P2Y inhibitors are one of the most common causes of emergency admissions for drug-related adverse events. The last ten years have seen a continuous debate on whether platelet function tests (PFTs) should be used to tailor antiplatelet drugs to cardiovascular patients. Large-scale randomized studies investigating the escalation of antiplatelet therapies according to the results of PFTs were mostly negative. Potent P2Y inhibitors are recommended as a first-line treatment in acute coronary syndrome patients, bringing the bleeding risk at the forefront. De-escalation from prasugrel or ticagrelor to clopidogrel is now considered, with or without the use of a PFT. This review covers recent advances in escalation and de-escalation strategies based on PFTs in various clinical settings. It also describes the main features of the most popular platelet function tests as well as the potential added value of genetic testing. Finally, we detail practical suggestions on how PFTs could be used in clinical practice.
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http://dx.doi.org/10.3390/jcm9010194DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7019825PMC
January 2020

An unusual thrombus location in a Heartmate 3 device with fatal outcome.

Perfusion 2020 07 9;35(5):442-446. Epub 2019 Dec 9.

Geneva Hemodynamic Research Group, Geneva, Switzerland.

Even if the HeartMate 3 left ventricular assist device is associated with excellent outcomes, complications, such as pump thrombosis continue to affect patients on hemodynamic support. We report the history of a 68-year-old man who underwent implantation of an HeartMate 3 as a bridge to transplantation. Nineteen months later, he developed signs of heart failure leading to cardiogenic shock. Neither clinical examination nor parameters from the device allowed a clear-cut diagnosis. Only surgical exploration revealed the presence of clots between the polyethylene terephthalate (Dacron®) and polytetrafluoroethylene tubes. This constitutes a weakness of this device for which we propose to the manufacturer for minimal modifications to overcome the problem.
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http://dx.doi.org/10.1177/0267659119890218DOI Listing
July 2020

Maladies vasculaires ­périphériques et prévention secondaire : un appel à agir.

Rev Med Suisse 2019 Dec;15(674):2227

Service d'angiologie et hémostase, Département de médecine, HUG, Genève.

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December 2019

Validation of the ISTH/SSC bleeding assessment tool for inherited platelet disorders: A communication from the Platelet Physiology SSC.

J Thromb Haemost 2020 03 16;18(3):732-739. Epub 2019 Dec 16.

Department of Internal Medicine, IRCCS Policlinico S. Matteo Foundation, University of Pavia, Pavia, Italy.

Background: Careful assessment of bleeding history is the first step in the evaluation of patients with mild/moderate bleeding disorders, and the use of a bleeding assessment tool (BAT) is strongly encouraged. Although a few studies have assessed the utility of the ISTH-BAT in patients with inherited platelet function disorders (IPFD) none of them was sufficiently large to draw conclusions and/or included appropriate control groups.

Objectives: The aim of the present study was to test the utility of the ISTH-BAT in a large cohort of patients with a well-defined diagnosis of inherited platelets disorder in comparison with two parallel cohorts, one of patients with type-1 von Willebrand disease (VWD-1) and one of healthy controls (HC).

Patients/methods: We enrolled 1098 subjects, 482 of whom had inherited platelet disorders (196 IPFD and 286 inherited platelet number disorders [IT]) from 17 countries.

Results: IPFD patients had significantly higher bleeding score (BS; median 9) than VWD-1 patients (median 5), a higher number of hemorrhagic symptoms (4 versus 3), and higher percentage of patients with clinically relevant symptoms (score > 2). The ISTH-BAT showed excellent discrimination power between IPFD and HC (0.9 < area under the curve [AUC] < 1), moderate (0.7 < AUC < 0.9) between IPFD and VWD-1 and between IPFD and inherited thrombocytopenia (IT), while it was inaccurate (AUC ≤ 0.7) in discriminating IT from HC.

Conclusions: The ISTH-BAT allows to efficiently discriminate IPFD from HC, while it has lower accuracy in distinguishing IPFD from VWD-1. Therefore, the ISTH-BAT appears useful for identifying subjects requiring laboratory evaluation for a suspected IPFD once VWD is preliminarily excluded.
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http://dx.doi.org/10.1111/jth.14683DOI Listing
March 2020

Selective inhibition of Panx1 channels decreases hemostasis and thrombosis in vivo.

Thromb Res 2019 Nov 18;183:56-62. Epub 2019 Oct 18.

Dept of Pathology and Immunology, University of Geneva, Geneva, Switzerland; Dept of Medical Specializations - Cardiology, University of Geneva, Geneva, Switzerland. Electronic address:

Background: Hemostasis is a tightly regulated physiological process to rapidly induce hemostatic plugs at sites of vascular injury. Inappropriate activation of this process may lead to thrombosis, i.e. pathological blood clot formation in uninjured vessels or on atherosclerotic lesions. ATP release through Pannexin1 (Panx1) membrane channels contributes to collagen-induced platelet aggregation in vitro.

Objective: To investigate the effects of genetic and pharmacological inhibition of Panx1 on hemostasis and thrombosis in vivo.

Results: Bleeding time after tail clipping was increased by 2.5-fold in Panx1 mice compared to wild-type controls, suggesting that Panx1 deficiency impairs primary hemostasis. Wire myography on mesenteric arteries revealed diminished vasoconstriction in response to phenylephrine or U446619 in Panx1 mice. Mice with platelet-specific deletion of Panx1 (Panx1) displayed 2-fold longer tail bleeding times than Panx1 controls. Moreover, venous thromboembolism (VTE) after injection of collagen/epinephrine in the jugular vein was reduced in Panx1 and Panx1 mice. Panx1 mice also showed reduced FeCl-induced thrombosis in mesenteric arteries. BrilliantBlue-FCF, a Panx1 channel inhibitor, decreased collagen-induced platelet aggregation in vitro, increased tail bleeding time and reduced VTE in wild-type mice. Furthermore, we developed a specific Panx1 blocking antibody targeting a Panx1 extracellular loop, which reduced ATP release from platelets in vitro. Treating wild-type mice with this antibody increased tail bleeding time and decreased VTE compared to control antibody.

Conclusions: Panx1 channel deletion or inhibition diminishes clot formation during hemostasis and thrombosis in vivo. Blocking Panx1 channels may be an attractive strategy for modulating platelet aggregation in thrombotic disease.
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http://dx.doi.org/10.1016/j.thromres.2019.09.028DOI Listing
November 2019

Thrombin generation and fibrin clot structure after vitamin D supplementation.

Endocr Connect 2019 Nov;8(11):1447-1454

Division of Angiology and Hemostasis, Geneva University Hospitals and Faculty of Medicine, University of Geneva, Geneva, Switzerland.

Objective: Vitamin D deficiency is associated with increased risks of arterial and venous cardiovascular events. Hypothetically, supplementation with vitamin D may lead to a less prothrombotic phenotype, as measured by global coagulation assays and fibrin clot structure.

Methods: In this prospective cohort study, we enrolled adult outpatients attending the Primary Care Division of the Geneva University Hospitals with a severe vitamin D deficiency (25-hydroxyvitamin-D3 (25-OHD) <25 nmol/L), excluding obese patients or with a recent acute medical event. We evaluated changes in coagulation times, thrombin generation assay, clot formation and clot lysis time, 25-OHD and parathormone before and 1-3 months after cholecalciferol oral supplementation with one-time 300,000 IU then 800 IU daily. Paired t-tests with a two-sided alpha of 0.05 compared absolute mean differences.

Results: The 48 participants had a mean age of 43.8 ± 13.8 years. After supplementation, 25-OHD levels increased from 17.9 ± 4.6 nmol/L to 62.5 ± 20.7 nmol/L 6.4 ± 3.0 weeks after inclusion. Endogenous thrombin potential and thrombin generation peak values both decreased significantly (-95.4 nM × min (95%CI -127.9 to -62.8), P < 0.001; -15.1 nM (-23.3 to -6.8), P < 0.001). The maximum absorbance by turbidimetry decreased significantly (P = 0.001) after supplementation. There was no change in clot lysis time, coagulation times or plasminogen activator inhibitor-1 and homocysteine levels.

Conclusions: In severe vitamin D deficiency, a high-dose cholecalciferol supplementation was associated with a reduction in thrombin generation and an average decreased number of fibrin protofibrils per fibers and fibrin fiber size measured by turbidimetry. This suggests that severe vitamin D deficiency may be associated with a potentially reversible prothrombotic profile.
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http://dx.doi.org/10.1530/EC-19-0429DOI Listing
November 2019

Platelet Function Testing or Genotyping: Which Should We Trust?

JACC Cardiovasc Interv 2019 09;12(18):1867-1868

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http://dx.doi.org/10.1016/j.jcin.2019.07.034DOI Listing
September 2019