Flow cytometric analysis of intraplatelet VASP phosphorylation for the detection of clopidogrel resistance in patients with ischemic cardiovascular diseases.

Interindividual variability of the inhibitory effect of clopidogrel on platelet functions leading to clopidogrel resistance has been described in some patients with ischemic cardiovascular disease. A reliable laboratory test is therefore needed to identify patients insufficiently protected by this antiplatelet treatment. The phosphorylation of vasodilator-stimulated phosphoprotein (VASP), an intraplatelet actin regulatory protein, is dependent on the level of activation of the platelet P2Y12 receptor, which is targeted by clopidogrel. The aim of this study was to use a flow cytometric VASP phosphorylation assay to evaluate the efficacy of clopidogrel therapy. The platelet reactivity index (PRI), expressed as a percentage, is the difference in VASP fluorescence intensity between resting (+PGE1) and activated (+ADP) platelets. In vitro, the PRI was strongly correlated with the inhibition of platelet aggregation induced by specific blockade of the P2Y12 receptor by the competitive antagonist AR-C69931MX (R = 0.72, P < 0.0001). Ex vivo, the PRI was 78.3 +/- 4.6% in 47 healthy donors, 79.0 +/- 4.1% in 34 patients not receiving clopidogrel and 61.1 +/- 17.0% in 33 patients treated with clopidogrel (P < 0.0001). In the clopidogrel group, the PRI values were widely dispersed (from 6.6 to 85.8%) and more than 30% of these patients had a PRI equivalent of values in patients not receiving clopidogrel. The flow cytometric analysis of VASP phosphorylation seems to be a suitable test to evaluate the efficacy of clopidogrel treatment. This assay demonstrated a wide interindividual variability of the inhibitory response of platelets to clopidogrel and showed that one-third of the patients treated appeared to be 'unprotected' by this therapy.

[Progress in transfusion for storage of platelets]. / Actualités en transfusion sur la conservation des plaquettes.

Improvement of platelet concentrates efficacy and safety depends on platelet storage conditions and their improvement: storage in the cold, additive solutions and pathogen inactivation.

[The inter-individual variability of the response to clopidogrel]. / Variabilité interindividuelle de la réponse au clopidogrel.

Clopidogrel is an anti-platelet aggregation drug with proven efficacy in the prevention of atherothrombotic complications. However, according to studies, between 5 and 20% of patients have further thrombotic episodes despite treatment with clopidogrel. The principal ex vivo evaluation methods of clopidogrel's efficacy are platelet aggregometry, and using flow cytometry to measure platelet activation and the quantatitive analysis of VASP phosphorylation. VASP analysis is the most selective method for clopidogrel's effect. These tests show great inter-individual variability in the response to treatment. This variability is such that about a third of treated patients show an "insufficient" response to clopidogrel. The exact causes of this variability have not yet been clearly identified. They could correspond with observer error, inter-individual variability in intestinal absorption or hepatic metabolism, or polymorphism of clopidogrel's target, the P2Y12 receptor. Accordingly it may be necessary to adapt the dose of clopidogrel or to use an alternative treatment. Failure to respond to clopidogrel would be a risk factor for atherothrombotic complications, but this association has yet to be demonstrated.

[Prospective study of the new oral anticoagulants in private practice: cautious optimism]. / Enquête prospective sur les nouveaux anticoagulants oraux en médecine libérale : un enthousiasme prudent.

BACKGROUND: The new direct oral anticoagulants (DOA) such as dabigatran, rivaroxaban or apixaban are an evolution in the management of patients requiring curative anticoagulation. However, behind the simplicity of prescribing and monitoring, several questions remain about their daily use. The aim of this prospective study was to measure the feelings of general practitioners (GP), angiologists (AP) and cardiologists (CP), potential prescribers of this new anticoagulant family. METHOD: Between December 2012 and May 2013, a questionnaire including five open questions and 11 questions using a positioning on an analogic visual scale (AVS 0 to 10) was subjected to GP, AP and CP in Alsace. RESULTS: Responses from 224 physicians (150 GP, 35 AP and 39 CP) were collected. Thus, 83% of GP, 83% of AP and 100% of CP were prescribers of DOA. However, among these prescribing doctors, the feeling was not the same and the trend of prescription was lower in GP (2.0 [1.1-3.2] AVS units) than in AP (3.1 [2.0-5.6]) and in CP (5.0 [1.2-8.7]) (P<0.0001 in multivariate analysis). The female doctors tended to prescribe DOA in younger patients than male doctors (respectively 66.1 [52.5-76.7] vs. 75.0 [65.7-81.0] years; P=0.004). The DOA were more considered as progress by AP and CP (respectively 7.8 [5.3-9.0] and 7.9 [7.0-8.7] AVSu) than by GP (6.1 [4.8-8.2] AVSu; P=0.02 in multivariate analysis). The answer about the eventual replacement of vitamin K antagonists by the DOA was very mixed whatever the practitioner group (5.1 [3.0-7.8] AVSu; P=0.139). The ease to use and the lack of biological monitoring were the main arguments leading to the prescription but the attitude of practitioners was very balanced by the lack of experience on the bleeding risk and the lack of available antidote. CONCLUSIONS: If the DOA are considered as an improvement for the physicians, the enthusiasm remains cautious whatever the type of practiced medicine. The results of clinical trials and the clinical experience should better appreciate the ongoing change in the field of anticoagulation.

Sensitivity and long-term prognostic value of cardiac troponin-I increase shortly after percutaneous transluminal coronary angioplasty.

BACKGROUND: After successful coronary interventions, minor elevations of creatine kinase MB (CK-MB) identified a population with a worse long-term prognosis than that in patients without enzyme elevations. In that setting, cardiac troponin-I (cTn-I), a highly specific marker for myocardial injury, was considered for a small study; the results did not support the view that significant myocardial damage occurred during successful percutaneous transluminal coronary angioplasty (PTCA). HYPOTHESIS: The present study was designed to assess the rate of elevated values of cTn-I after successful PTCA and to determine its prognostic value. METHODS: CTn-I and CK-MB were measured in 44 patients before and daily for 3 days after PTCA. Two groups of patients were considered according to the presence or absence of elevated levels of cTn-I. The rate of free-event survival was estimated for the two groups using the Kaplan-Meier method and was compared with the log rank test. RESULTS: Globally, 36% of patients had an increase in cTn-I (normal values 0.35 ng/ml) and 9% had an increase in CK-MB, p = 0.002. The mean time to maximal enzyme level was 1.8 days for cTn-I and 2.2 days for CK-MB. Over a follow-up of 1375 +/- 416 days, 18% of patients experienced adverse events, and cTn-I did not identify a population of worse long-term prognosis. CONCLUSION: These results suggest that cTn-I is more sensitive than CK-MB in identifying minor myocardial damage after PTCA, but these elevated concentrations of cTn-I in the short-term aftermath of angioplasty do not seem to be a marker of worse long-term prognosis.

In vitro evaluation of Haemonetics MCS+ apheresis platelet concentrates treated with photochemical pathogen inactivation following plasma volume reduction using the INTERCEPT Preparation Set.

Pathogen inactivation using the INTERCEPT Blood System requires platelet resuspension in InterSol and reduced plasma. Platelets in plasma collected on the Haemonetics MCS+ were processed on the INTERCEPT Preparation Set for plasma volume reduction and addition of InterSol. The use of the Preparation Set resulted in a mean platelet loss of 5.6 +/- 3.4%. Subsequent photochemical treatment (PCT) with amotosalen and ultraviolet A light, and 7 days of storage, resulted in acceptable changes for platelet swirling, lactate, lactate dehydrogenase (LDH), platelet factor-4 (PF4), p-selectin, glycoprotein V (GpV), pO2, pCO2, tumour necrosis factor-alpha (TNF-alpha) and interleukin-8 (IL-8). All platelet units processed with the Preparation Set and PCT met European requirements for leucoreduction and pH values.

A study of platelet activation in atrial fibrillation and the effects of antithrombotic therapy.

BACKGROUND: Platelet function may be abnormal in patients with atrial fibrillation (AF) and could be related to abnormal thrombogenesis. The aim of this cross-sectional study was to investigate new aspects of platelet biology in AF, predominantly focusing on platelet activation and the effects of concomitant antiplatelet and anticoagulant therapy. PATIENTS AND METHODS: The study group of 238 patients were (i). 93 patients on no antithrombotic therapy, (ii). 60 patients taking 75-325 mg aspirin/day, and (iii). 85 patients on dose-adjusted warfarin (International Normalised Ratio [INR] range 2.0-3.0). Results were compared with those from 50 age- and sex-matched normal subjects. Platelet markers (plasma beta-thromboglobulin, soluble glycoprotein V [both ELISA]), coagulation markers (fibrin D-dimer [ELISA] and fibrinogen [Clauss]), and platelet aggregation in response to standard platelet agonists were studied. RESULTS: beta-thromboglobulin (P=0.01), soluble glycoprotein V (P<0.001) and fibrin D-dimer (P=0.002) were higher in untreated AF patients compared to healthy controls. AF patients on warfarin had lower fibrin D-dimer (P<0.001) when compared to AF patients on no therapy. Plasma fibrinogen and platelet aggregation was no different between patients with AF and healthy controls. Aspirin use was associated with reduced platelet aggregation response to epinephrine (P=0.01), whilst patients established on warfarin had significantly lower plasma fibrin D-dimer levels. CONCLUSION: AF patients exhibit changes in plasma markers of platelet function but no significant abnormalities of platelet aggregation. However, treatment with warfarin or aspirin failed to demonstrate any significant benefit on platelet activation, although warfarin use was associated with reduced thrombogenesis (fibrin D-dimer). We suggest that platelet activation may not play an important role in the pathogenesis of thromboembolism in AF.