Differences In Platelet Reactivity Research Articles

Inflammation and platelet reactivity during adjunctive colchicine versus aspirin in patients with acute coronary syndrome treated with potent P2Y12 inhibitor

BackgroundIn patients undergoing percutaneous coronary intervention (PCI), the use of anti-inflammatory therapy with colchicine is associated with a reduction of recurrent ischemic events. The mechanisms of such findings are not fully elucidated.ObjectivesTo investigate the effects of colchicine versus aspirin on inflammation and platelet reactivity in patients with acute coronary syndrome (ACS) undergoing PCI.MethodsThis observational study compared laboratory measurements in ACS patients receiving single antiplatelet therapy with ticagrelor or prasugrel plus colchicine (MACT) (n = 185) versus conventional dual-antiplatelet therapy (DAPT) with aspirin plus ticagrelor or prasugrel (n = 497). The primary outcome was the frequency of high residual inflammation, defined as high-sensitivity C-reactive protein (hs-CRP) ≥2 mg/L at 1 month post-PCI. Multiple sensitivity analyses were performed for the primary outcome, including multivariable adjustment, propensity-score matching, and inverse-probability weighted methods.ResultsOne month after PCI, patients treated with MACT had significantly lower levels of hs-CRP compared to those treated with DAPT (0.6 [0.4–1.2] vs. 0.9 [0.6–2.3] mg/L, p < 0.001). The frequency of high residual inflammation was also lower in the MACT group (10.8% vs. 27.2%, p < 0.001) (odds ratio [95% confidence interval] = 0.33 [0.20–0.54], p < 0.001). This effect was consistent across sensitivity analyses. There was no difference in platelet reactivity between MACT and DAPT (49.6 ± 49.0 vs. 51.5 ± 66.4 P2Y12 reaction unit [PRU] measured by VerifyNow, p = 0.776).ConclusionIn ACS patients undergoing PCI, MACT was associated with a lower rate of high residual inflammation without increasing platelet reactivity compared to conventional DAPT.Clinical trial registrationNCT04949516 for MACT pilot trial and NCT04650529 for Gyeongsang National University Hospital registry.

Quantitative Label-Free Mass Spectrometry Identifies Molecular Content and Signaling Differences between Neonatal and Adult Platelets

Observational and randomized studies have associated platelet transfusions with increased morbidity and mortality in preterm neonates. Clinical practice changes have been hindered by a limited understanding of why adult donor-derived platelets might be inappropriate or harmful to infants. Neonatal and adult platelets differ in reactivity and function, but we lack an understanding of underlying molecular differences. As proteins and kinase-driven phosphorylation signaling dictate platelet biology, we aimed to characterize altered protein and phosphoprotein content between neonatal and adult platelets to help explain differences in platelet reactivity and function. We isolated resting platelets from full term cord blood (n=9) and adult peripheral blood (n=7) and used mass spectrometry-based platforms to ascertain protein and phosphoprotein content. We identified 4745 high-confidence proteins in adult and neonatal platelets, including 331 differentially abundant proteins (≥1.5-fold change, p<0.05). Neonatal platelets had increased metabolic and ribosomal proteins, consistent with transcriptional data. Adult platelets were enriched for inflammatory proteins, including complement components, consistent with pro-inflammatory function. We also identified 2115 phospho-proteins (17852 unique phosphopeptides) across all samples. These impacted actin cytoskeletal biology, cell adhesion, and GTPase signaling that regulate platelet functions. Kinase enrichment analysis identified FYN, SRC, ABL1, and AKT1 as kinases most responsible for platelet phosphorylation activities. Granule trafficking and degranulation proteins were among 1183 differentially abundant phosphoproteins in neonatal vs adult samples (p<0.05). For example, 31 different Reticulon 1 (RTN1) phosphopeptides were enriched in adult platelets (p<0.05). RTN1 regulates membrane trafficking and participates in SNARE-mediated exocytosis. Increased RTN1-directed membrane trafficking may promote increased activation and degranulation in adult vs neonatal platelets. Indeed, SNARE complex protein deficiency was previously suggested to mediate hyporeactive degranulation in neonatal platelets. We detected 2 phosphopeptides in adult samples that were absent from any neonatal sample. We reasoned that these could represent developmental stage-specific differences that exist only in adult platelets. One of the adult-specific phosphopeptides was in Rap1GAP2 (phospho-S588). Rap1GAP2 is a highly phosphorylated GTPase-activating protein that inhibits Rap1, a small guanine-nucleotide-binding protein that facilitates integrin activation in platelets. Rap1GAP2 binds synaptotagmin-like protein 1 (Slp1) to regulate dense granule secretion. These findings may link altered developmentally regulated Rap1GAP2 activities with platelet degranulation. A Platelet Cell Adhesion Molecule (PECAM) phosphopeptide (phospho-S726) was also found exclusively in adult platelets. PECAM facilitates inflammation and leukocyte transendothelial migration. PECAM phosphorylation in adult platelets may reflect an enhanced ability to participate in these processes. These findings reveal key differences in protein content and signaling that underlie differential neonatal vs adult platelet reactivity and function, including pro-inflammatory activities and enhanced activation and/or degranulation propensity in adult platelets. RTN1-mediated SNARE complex function and Rap1 signaling activities represent novel and potentially modifiable targets to manipulate platelet activation and/or degranulation. Our results also help provide a biological rationale for the higher morbidity and mortality observed in preterm infants transfused with adult platelets, including the inadvertent introduction of serum complement and other pro-inflammatory molecules within transfusions of donated adult platelets.

Abstract 12411: Fatty Acid Saturation Status in Murine High Fat Diets Modulates Platelet Reactivity

Introduction: We have recently shown that membrane fatty acids are key modulators of megakaryocyte differentiation and platelet formation. In obesity, patients have an altered plasma lipid composition, which coincides with enhanced platelet responsiveness to activation. However, the mechanism underlying this platelet phenotype remains unknown. Hypothesis: We hypothesized that the obese plasma lipidome alters the composition of the megakaryocyte membrane, which ultimately leads to the generation of hyper-reactive platelets. Aim: To investigate the impact of high fat diets with different fatty acid compositions on megakaryocyte development and platelet reactivity. Results: Mice were fed chow or high fat diets enriched in either saturated fatty acids (50% SFAs) or polyunsaturated fatty acids (50% PUFAs, omega-6-enriched). Neither high fat diet resulted in changes in megakaryocyte numbers, but both caused a significant increase in the size of bone marrow megakaryocytes, suggesting diet-derived lipid uptake into megakaryocytes. When examining platelet reactivity, we found that mice fed the SFA-enriched high fat diet exhibited increased expression of the active form of integrin αIIbβ3 (JON/A) in resting platelets and that both JON/A and P-selectin expression were significantly decreased after platelets were stimulated with ADP and the thromboxane A 2 analogue U46619. In contrast, platelets from mice fed the PUFA omega-6-enriched high fat diet had no differences in platelet reactivity compared to mice fed normal chow. Conclusions: Our data reveal that dietary fatty acid saturation status affects platelet reactivity, consistent with previous human studies showing variability in platelet reactivity in obese individuals. Further explorations will elucidate if plasma and/or platelet lipid content may be an indicator of risk for thrombosis or cardiovascular disease.

Genetic Association Study and Machine Learning to Investigate Differences in Platelet Reactivity in Patients with Acute Ischemic Stroke Treated with Aspirin.

Aspirin resistance (AR) is a pressing problem in current ischemic stroke care. Although the role of genetic variations is widely considered, the data still remain controversial. Our aim was to investigate the contribution of genetic features to laboratory AR measured through platelet aggregation with arachidonic acid (AA) and adenosine diphosphate (ADP) in ischemic stroke patients. A total of 461 patients were enrolled. Platelet aggregation was measured via light transmission aggregometry. Eighteen single-nucleotide polymorphisms (SNPs) in ITGB3, GPIBA, TBXA2R, ITGA2, PLA2G7, HMOX1, PTGS1, PTGS2, ADRA2A, ABCB1 and PEAR1 genes and the intergenic 9p21.3 region were determined using low-density biochips. We found an association of rs1330344 in the PTGS1 gene with AR and AA-induced platelet aggregation. Rs4311994 in ADRA2A gene also affected AA-induced aggregation, and rs4523 in the TBXA2R gene and rs12041331 in the PEAR1 gene influenced ADP-induced aggregation. Furthermore, the effect of rs1062535 in the ITGA2 gene on NIHSS dynamics during 10 days of treatment was found. The best machine learning (ML) model for AR based on clinical and genetic factors was characterized by AUC = 0.665 and F1-score = 0.628. In conclusion, the association study showed that PTGS1, ADRA2A, TBXA2R and PEAR1 polymorphisms may affect laboratory AR. However, the ML model demonstrated the predominant influence of clinical features.

Manipulation of Fatty Acid Metabolism Impairs Megakaryocyte Differentiation and Platelet Production

Background: Obesity is associated with an altered plasma lipid composition that impacts platelet activation, indicating that lipids directly affect platelet function and reactivity. However, very little is known about which essential lipids are important to enable platelet production from their precursors, megakaryocytes (MKs), and whether obesity influences lipid composition and consequently affects thrombopoiesis.Aim: We aimed to determine which lipids and associated pathways play a significant role in MK differentiation and platelet production in health and upon obesity. Our long-term goal is to determine if we can manipulate MK maturation and platelet production through lipid incorporation or inhibition.Methods: In order to determine the lipid profile of MKs through maturation, we performed an extensive lipidomics screen on primary MK progenitors, mature MKs, and platelets. Based on our findings, we then targeted de novo fatty acid (FA) synthesis in MKs to determine whether manipulation of this pathway attenuates MK differentiation and platelet formation. In parallel, we investigated megakaryo- and thrombopoiesis in an experimental setting of diet-induced obesity (DIO) in mice to examinate how altered lipid compositions within the plasma affect MK maturation.Results: Lipidomic data revealed increased polyunsaturated FA and plasmalogen content in the late stages of MKs maturation, suggesting that lipid composition is undergoing changes that may enable platelet production. To validate and extend these results, we inhibited multiple enzymes in de novo lipogenesis and FA synthesis. Using two different sources of primary murine hematopoietic stem cells (HSCs), we found that MK differentiation and maturation from HSCs was significantly decreased up to 60% after inhibition of acetyl-coA carboxylase (ACC) and 80% after fatty acid synthetase (FASN) inhibition, both enzymes from the de novo lipogenesis pathway. Moreover, we also found that MK differentiation and maturation were highly decreased (up to 70%) after the inhibition of acyl-coA synthetase (ACS). To further examine the role of fatty acids incorporation on platelet production, we next treated mature MKs with an ACS inhibitor to investigate the direct effect of FA incorporation on proplatelet formation; we observed a significant, multifoldreduction in proplatelet area. While dihydroceramides were altered in the lipidomic data, impairment of the de novo ceramide synthesis pathway did not affect either MK maturation or platelet production. Moreover, we observed an increased abundance of HSCs, myeloid cells, and MK progenitors in the bone marrow of DIO mice compared to control mice on a chow diet. This further supports an important role for lipids in megakaryopoiesis and suggests that differences in platelet reactivity during obesity may be caused by altered MK maturation and/or platelet production.Conclusions: Our results elucidate an important and previously unrecognized role for fatty acid synthesis in megakaryopoiesis and platelet production. The findings imply that an altered lipid content, as observed in patients with obesity, may not only impact platelet reactivity but also MK maturation and function. Consequently, platelets with an altered lipid content might originate from pathogenic MKs in obesity, which makes lipid-regulating proteins in MKs novel and viable therapeutic targets. DisclosuresMachlus: KEROS Therapeutics: Consultancy, Honoraria; STRM.BIO: Consultancy, Honoraria.

Heterogeneity in Bleeding Tendency and Arthropathy Development in Individuals with Hemophilia.

People with hemophilia (PWH) have an increased tendency to bleed, often into their joints, causing debilitating joint disease if left untreated. To reduce the incidence of bleeding events, PWH receive prophylactic replacement therapy with recombinant factor VIII (FVIII) or FIX. Bleeding events in PWH are typically proportional to their plasma FVIII or IX levels; however, in many PWH, bleeding tendency and the likelihood of developing arthropathy often varies independently of endogenous factor levels. Consequently, many PWH suffer repeated bleeding events before correct dosing of replacement factor can be established. Diagnostic approaches to define an individual's bleeding tendency remain limited. Multiple modulators of bleeding phenotype in PWH have been proposed, including the type of disease-causing variant, age of onset of bleeding episodes, plasma modifiers of blood coagulation or clot fibrinolysis pathway activity, interindividual differences in platelet reactivity, and endothelial anticoagulant activity. In this review, we summarize current knowledge of established factors modulating bleeding tendency and discuss emerging concepts of additional biological elements that may contribute to variable bleeding tendency in PWH. Finally, we consider how variance in responses to new gene therapies may also necessitate consideration of patient-specific tailoring of treatment. Cumulatively, these studies highlight the need to reconsider the current "one size fits all" approach to treatment regimens for PWH and consider therapies guided by the bleeding phenotype of each individual PWH at the onset of therapy. Further characterization of the biological bases of bleeding heterogeneity in PWH, combined with the development of novel diagnostic assays to identify those factors that modulate bleeding risk in PWH, will be required to meet these aspirations.

Cangrelor in addition to standard therapy reduces cardiac damage and inflammatory markers in patients with ST-segment elevation myocardial infarction.

Although P2Y12 receptor blockers have become a standard, adjunctive therapy in patients with ST-segment elevation myocardial infarction (STEMI) undergoing percutaneous coronary intervention (PCI), the optimal regimen has not been established. We performed a prospective, open-label, randomized study to investigate the effect of cangrelor administration on platelet function and inflammation in patients with primary PCI (PPCI). Twenty-two patients were randomized to receive either cangrelor and ticagrelor or ticagrelor alone (standard group) before PPCI. Platelet reactivity was evaluated at baseline (before PCI), 10min and the end of the procedure. At baseline, there was no significant difference in platelet reactivity between both groups, whereas platelets were significantly inhibited at 10min after initiating cangrelor vs. standard (adenosine-diphosphate-induced aggregation 102.2 ± 24.88 vs. 333.4 ± 63.3, P < 0.05 and thrombin-receptor-activating-peptide-induced aggregation 285.8 ± 86.1 vs. 624.8 ± 106.0, P < 0.05). Lower platelet aggregation in the cangrelor group persisted but the difference was reduced by the end of the procedure. Circulating inflammatory cells, pro-inflammatory cytokines, total elastase, and surrogates of neutrophil extracellular traps (total elastase-myeloperoxidase complexes) were significantly lower in the cangrelor compared to the standard therapy group at 6h after randomization. There was a trend towards reduction in cardiac damage in the cangrelor group as reflected by the changes in late gadolinium enhancement between 48h and 3months after STEMI. Early administration of cangrelor in STEMI patients was associated with more effective platelet inhibition during PPCI and significantly dampened the deleterious inflammatory response compared to standard therapy (NCT03043274).

Influence of Sex on Platelet Reactivity in Response to Aspirin.

BackgroundThere are sex differences in the efficacy and safety of aspirin for the prevention of myocardial infarction and stroke. Whether this is explained by underlying differences in platelet reactivity and aspirin response remains poorly understood.Methods and ResultsHealthy volunteers (n=378 208 women) and patients with coronary artery disease or coronary artery disease risk factors (n=217 112 women) took aspirin for 4 weeks. Light transmittance aggregometry using platelet‐rich plasma was used to measure platelet reactivity in response to epinephrine, collagen, and ADP at baseline, 3 hours after the first aspirin dose, and after 4 weeks of daily aspirin therapy. A subset of patients underwent pharmacokinetic and pharmacodynamic assessment with levels of salicylate and cyclooxygenase‐1–derived prostaglandin metabolites and light transmittance aggregometry in response to arachidonic acid and after ex vivo exposure to aspirin. At baseline, women had increased platelet aggregation in response to ADP and collagen. Innate platelet response to aspirin, assessed with ex vivo aspirin exposure of baseline platelets, did not differ by sex. Three hours after the first oral aspirin dose, platelet aggregation was inhibited in women to a greater degree in response to epinephrine and to a lesser degree with collagen. After 4 weeks of daily therapy, despite higher salicylate concentrations and greater cyclooxygenase‐1 inhibition, women exhibited an attenuation of platelet inhibition in response to epinephrine and ADP.ConclusionsWe observed agonist‐dependent sex differences in platelet responses to aspirin. Despite higher cyclooxygenase‐1 inhibition, daily aspirin exposure resulted in a paradoxical attenuation of platelet inhibition in response to epinephrine and ADP over time in women but not in men.

Sex differences in flow cytometry–based platelet reactivity in stable outpatients suspected of myocardial ischemia

BackgroundAntiplatelet therapy is the mainstay of secondary prevention of cardiovascular events. Studies suggest that women do not obtain equal therapeutic benefit from antiplatelet therapy compared with men. The link between sex differences in platelet biology and response to antiplatelet therapies is unclear. We therefore investigated the role of sex differences in platelet reactivity in a cohort of outpatients with chest pain, in response to treatment with antiplatelet agents. MethodsPlatelet reactivity was measured in 382 randomly selected patients participating in the Myocardial Ischemia Detection by Circulating Biomarkers (MYOMARKER) study, an observational cohort study of outpatients suspected of myocardial ischemia. In all patients, blood was collected during diagnostic workup, and platelet reactivity was assessed with a flow cytometry–based platelet activation test that quantifies both platelet degranulation (P‐selectin expression) and platelet aggregation (fibrinogen binding to integrin αIIbβ3) in whole blood. ResultsPlatelet reactivity was higher in women compared with men when activated with protease activating receptor 1–activating peptide SFLLRN (PAR1‐AP) and adenosine 5′‐phosphate (ADP), independent of age, basal activation status, estimated glomerular filtration rate < 60, platelet count, statin use, the use of P2Y12 inhibitors, or the use of aspirin. P2Y12 inhibitor use strongly reduced fibrinogen binding after stimulation with PAR1‐AP, but only slightly reduced platelet P‐selectin expression. Calculation of the relative inhibition in P2Y12 users indicated 62% inhibition of the response toward ADP. Stratified analysis showed that women (n = 14) using P2Y12 inhibitors showed less inhibition of fibrinogen binding after PAR1‐AP stimulation than men (n = 38) using P2Y12 inhibitors. ConclusionsThese findings call for further study of differential effects of P2Y12 inhibitors in women with suspected myocardial ischemia.

Sex differences in platelet reactivity in patients with myocardial infarction treated with triple antiplatelet therapy - results from assessing platelet activity in coronary heart disease (APACHE)

)Several earlier studies have reported increased risk of bleeding in women with myocardial infarction, (MI) compared to men. The reasons for the observed difference are incompletely understood, but one suggested explanation has been excess dosing of antithrombotic drugs in women. The aim of this prospective observational study was to assess sex differences in platelet activity in patients treated with three different platelet inhibitors. We recruited 125 patients (37 women and 88 men) with MI, scheduled for coronary angiography. All patients received clopidogrel and aspirin. A subgroup of patients received glycoprotein (GP) IIb/IIIa-inhibitor. Platelet aggregation in whole blood was assessed at several time points, using impedance aggregometry. Soluble P-selectin was measured 3 days after admission. There were no significant differences between women and men in baseline features or comorbidities except higher frequency of diabetes, lower hemoglobin value, and lower estimated glomerular filtration rate, in women on admission. We observed significantly more in-hospital bleeding events in women compared to men (18.9% vs. 6.8%, p = .04). There were no differences in platelet aggregation using three different agonists, reflecting treatment effect of GPIIb/IIIa-inhibitors, clopidogrel, and aspirin, 6–8 hours, 3 days, 7–9 days, or 6 months after loading dose. Moreover, there was no significant difference in soluble P-selectin. The main finding of this study was a consistent lack of difference between the sexes in platelet aggregation, using three different agonists at several time-points. Our results do not support excess dosing of anti-platelet drugs as a major explanation for increased bleeding risk in women.

Establishment of a megakaryoblastic cell line for conventional assessment of platelet calcium signaling

Platelet function tests utilizing agonists or patient serum are generally performed to assess platelet activation ex vivo. However, inter-individual differences in platelet reactivity and donor requirements make it difficult to standardize these tests. Here, we established a megakaryoblastic cell line for the conventional assessment of platelet activation. We first compared intracellular signaling pathways using CD32 crosslinking in several megakaryoblastic cell lines, including CMK, UT-7/TPO, and MEG-01 cells. We confirmed that CD32 was abundantly expressed on the cell surface, and that intracellular calcium mobilization and tyrosine phosphorylation occurred after CD32 crosslinking. We next employed GCaMP6s, a highly sensitive calcium indicator, to facilitate the detection of calcium mobilization by transducing CMK and MEG-01 cells with a plasmid harboring GCaMP6s under the control of the human elongation factor-1α promoter. Cells that stably expressed GCaMP6s emitted enhanced green fluorescent protein fluorescence in response to intracellular calcium mobilization following agonist stimulation in the absence of pretreatment. In summary, we have established megakaryoblastic cell lines that mimic platelets by mobilizing intracellular calcium in response to several agonists. These cell lines can potentially be utilized in high-throughput screening assays for the discovery of new antiplatelet drugs or diagnosis of disorders caused by platelet-activating substances.

Gender Differences in Platelet Reactivity in Diabetic Patients Receiving Dual Antiplatelet Therapy

BackgroundIncreased comorbidities and a perceived high-bleeding risk often prevent the use of dual antiplatelet therapy (DAPT) in female patients. However, more aggressive antiplatelet treatment would certainly offer additional outcome benefits in coronary artery disease, especially among diabetic patients. The aim of the present study was to evaluate the gender differences in high-residual on treatment platelet reactivity (HRPR) among diabetic patients treated with DAPT. MethodsOur population is represented by a consecutive cohort of diabetic patients treated with DAPT (ASA + clopidogrel, ticagrelor or dose-adjusted prasugrel) for an acute coronary syndrome or elective PCI, undergoing platelet reactivity assessment at 30–90 days post-discharge. Aggregation was assessed by multiple-electrode aggregometry and in diabetic patients naïve to antiplatelet therapy, by light transmission aggregometry, surface expression of P-selectin and plasma concentration of Thromboxane B2. ResultsWe included 472 patients, 113 (23.9%) women. Female gender was associated with more advanced age, and increased comorbidities. Mean platelet reactivity did not differ according to gender. The rate of HRPR was similar in women as compared to men (for ASA: adjusted OR[95%CI] = 0.59[0.27–1.33], p = 0.21, for ADP-antagonists: adjusted OR[95%CI] = 1.24[0.25–1.80], p = 0.27), however, the benefits of the new ADP-antagonists on platelet reactivity were lower in women than in men (p interaction = 0.01).No impact of gender on platelet reactivity was confirmed among 50 diabetic patients naïve to antiplatelet therapy. ConclusionsAmong diabetic patients receiving dual antiplatelet therapy gender does not affect platelet reactivity or high-on treatment platelet reactivity. However, the enhanced platelet inhibition provided by the new-ADP antagonists of new-ADP antagonists could be mitigated in women.

P4744Patients with atrial fibrillation exhibit a systemic prothrombotic state attributable to impaired endogenous fibrinolysis

Abstract Background The association of atrial fibrillation (AF) with thromboembolic stroke due to stasis in the left atrium and left atrial appendage is well described. Whether AF is associated with a systemic prothrombotic state, detectable in peripheral blood, unclear. Previous studies have been inconsistent, with some very small previous studies (&lt;30 patients each) variably indicating that patients with AF may have raised platelet reactivity and levels of antithrombin III, d-dimer, PAI-1 and t-PA-PAI complexes. These cumbersome laboratory tests of coagulation and fibrinolysis are not readily available in the clinical setting. Purpose It was our aim to compare, in peripheral venous blood, thrombotic and endogenous fibrinolytic profile of healthy volunteers and patients with newly diagnosed nonvalvular atrial fibrillation (NVAF), using a point-of-care technique. Methods In a prospective observational study, venous blood samples were taken from 98 healthy volunteers and 100 patients with newly diagnosed NVAF in the out-patient setting. Patients with newly diagnosed NVAF had venous blood tested before any treatment was initiated with aspirin or oral anticoagulation. Thrombotic status was assessed using the Global Thrombosis Test (GTT), a point-of-care test using native non-coagulated blood, assessed within 15 sec of blood withdrawal. The time to form an occlusive venous thrombus in native (non-citrated) blood, a measure of platelet reactivity (occlusion time, OT) and the time taken to spontaneous endogenous fibrinolysis to restore flow (lysis time, LT) were assessed. Results Basic blood tests (full blood count, renal and liver function, inflammatory markers) were normal in all subjects. The groups were matched for sex and race. Mean age of the healthy cohort was 34±8 years and patients 65±10 years. Endogenous fibrinolysis was markedly impaired in patients with NVAF compared to healthy individuals as shown by markedly prolonged LT (median 2015s [interquartile range IQR 1555–2507] vs. 1124s [IQR 919–1554], p&lt;0.ehz745.11201). There was no difference in platelet reactivity between patients and normal volunteers (369s [IQR 308–445]vs 368s [IQR 309–441], p=0.704). Sensitivity analysis was performed on a subgroup matched for age, sex and race. LT remained significantly longer in patients with NVAF compared to controls (1569s [IQR 1499–2244] vs. 1219s [IQR 943–1560], p=0.03), with no difference in platelet reactivity (p=NS). Conclusion In the largest study to date and using a clinically-friendly automated point-of-care technique, we show that patients with NVAF exhibit a systemic prothrombotic state, attributable to significantly impaired endogenous fibrinolysis compared with healthy volunteers. Further studies are needed to see if this could become a screening test for the prothrombotic state in patients with NVAF. Acknowledgement/Funding None

Assessment of the relationship between platelet reactivity, vascular risk factors and gender in cerebral ischaemia patients.

Excessive activation and platelet aggregation play important roles in the aetiopathogenesis of cerebral ischaemia. The aim of this study was to assess the relationship between platelet reactivity, gender and vascular risk factors in cerebral ischaemia patients. The research is useful because we found high risk groups of inefficient aspirin treatment in cerebral ischaemia patients. The study involved 101 patients, including 69 patients with ischaemic stroke and 32 patients with transient ischaemic attack. The assessment of platelet reactivity was made within 24 hours of the disease onset using two aggregometric methods: impedance and optical. Resistance to acetylsalicylic acid among people with cerebral ischaemia was estimated at 30.69% using impedance aggregometry and 9.2% using optical aggregometry. There were no differences in platelet reactivity or ASA resistance between the groups of patients with stroke and TIA in either method. In the whole group of patients (p = 0.04), and in the group of patients with stroke (p = 0.0143), higher reactivity of platelets was observed by impedance aggregometry in men than in women. In the whole group of patients (p = 0.0229), and in the subgroup with stroke (p = 0.0123), it was shown that aspirin resistance is significantly more common in the subgroup of men than in women. In patients suffering from nicotine addiction, significantly higher platelet reactivity was found in the whole group of patients (p = 0.004), as well as in the subgroup of patients with stroke (p = 0.0135). There are no differences between platelet reactivity and the incidence of aspirin resistance in patients with stroke and TIA. Male gender and smoking are associated with greater reactivity of platelets and more frequent occurrence of acetylsalicylic acid resistance in patients with cerebral ischaemia. Dual antiplatelet therapy or clopidogrel treatment should be considered in smoking males with cerebral ischaemia due to the high risk of aspirin inefficiency.

Role of Gender in Dual Antiplatelet Therapy After Acute Coronary Syndrome.

The effect of gender on use of dual antiplatelet therapy (DAPT) in treatment of acute coronary syndrome (ACS) is not well established. The purpose of this review is to understand gender-based differences in response to DAPT, so that treatment of ACS can be optimized in women to prevent ischemic events while minimizing bleeding risk. There are innate gender differences in platelet reactivity and response. However, it is unknown if this translates into differences in clinical outcomes. In all major studies evaluating the effect of DAPT in ACS, women are underrepresented. Hence, the results from the existing trials cannot be generalizable to women. There is a significant knowledge gap regarding how to balance the bleeding and ischemic risk profile among women with ACS. Currently, there is no recommendation to consider gender as covariate in choosing the type of antiplatelet drug or duration. The existing clinical evidence is limited by under representation of women in DAPT trials. The current literature does not strongly support considering gender in decision making regarding type or duration of DAPT after ACS. Future dedicated trial designs with adequate representation from women and gender specific analysis from large registry data are warranted to enhance our understanding of the interaction of gender with DAPT after ACS.

The impact of rivaroxaban on primary hemostasis in patients with venous thrombosis

Factor Xa inhibitors are safe and effective alternatives to warfarin, but several studies indicate that rivaroxaban may cause a different risk profile for bleeding. For instance, while the risk of major bleeding in general may be lower with rivaroxaban than for warfarin, the risk of gastrointestinal bleeding or abnormal uterine bleeding may be higher. The underlying mechanisms for these differences are not known, and the effect of rivaroxaban on primary hemostasis is poorly understood. The aim of this study was to investigate the effect of rivaroxaban on platelet function, P-selectin and von Willebrand factor (VWF) antigen and activity. Patients with venous thrombosis assigned to 3 months of treatment due to temporary risk factors were included. Blood was collected both during (on-treatment) and 4–6 weeks after end of treatment (without treatment). The platelet reactivity was assessed by light transmission aggregometry. P-selectin was measured by an enzyme-linked immunosorbent assay and vWF antigen and activity by latex immunoagglutination assays. Platelet reactivity during on-treatment (trough- and peak concentration) was similar to values without treatment. There was a trend toward a reduction of P-selectin during rivaroxaban treatment (peak concentration) compared to value without treatment (p = 0.06). There were no differences in vWF antigen and activity between the different time-points. We found no difference in platelet reactivity or vWF antigen/activity during rivaroxaban treatment compared with values without treatment. Apart from possibly causing a reduction of P-selectin, rivaroxaban seems not to influence primary hemostasis.

Individual variations in platelet reactivity towards ADP, epinephrine, collagen and nitric oxide, and the association to arterial function in young, healthy adults

IntroductionPlatelet aggregation and secretion can be induced by a large number of endogenous activators, such as collagen, adenosine diphosphate (ADP) and epinephrine. Conversely, the blood vessel endothelium constitutively release platelet inhibitors including nitric oxide (NO) and prostacyclin. NO and prostacyclin are also well-known vasodilators and contribute to alterations in local blood flow and systemic blood pressure. Materials and methodsIn this study we investigated individual variations in platelet reactivity and arterial functions including blood pressure and flow-mediated vasodilation (FMD) in 43 young, healthy individuals participating in the Lifestyle, Biomarkers and Atherosclerosis (LBA) study. Platelet aggregation and dense granule secretion were measured simultaneously by light transmission and luminescence. FMD was measured with ultrasound. ResultsThe platelet function assay showed inter-individual differences in platelet reactivity. Specifically, a sub-group of individuals had platelets with an increased response to low concentrations of ADP and epinephrine, but not collagen. When the NO-donor S-nitroso-N-acetyl-DL-penicillamine (SNAP) was combined with high doses of these platelet activators, the results indicated for sub-groups of NO-sensitive and NO-insensitive platelets. The individuals with NO-sensitive platelets in response to SNAP in combination with collagen had a higher capacity of FMD of the arteria brachialis. ConclusionsPlatelet reactivity towards ADP, epinephrine and NO differs between young, healthy individuals. Some individuals have a more effective response towards NO, both in the aspect of platelet inhibition ex vivo, as well as vasodilation in vivo.

Возможные генетические предикторы развития сердечно-сосудистых осложнений после коронарного шунтирования

to investigate associations of single nucleotide polymorphisms (SNPs) rs2046934, rs1126643, rs5918, rs6065, rs4244285; rs4986893 with cardiovascular complications (CVC) in patients after CABG. We enrolled in this study 130 patients with stable functional class II-IV angina. After CABG 69 patients received ASA (100 mg of enteric form), 61 patients received dual antiplatelet therapy (ASA 100 mg + clopidogrel 75 mg). Mean follow up period was 10.9±5.2 months. The primary composite endpoint included all-cause mortality, myocardial infarction (MI), and ischemic stroke. The control group comprised 185 healthy volunteers. Platelet function was assessed using light transmission aggregometry with ADP (5μM) and arachidonic acid (1 mM). The following single nucleotide polymorphisms (SNPs) were identified by real-time PCR: rs2046934 (H1/H2) on P2RY12 [encoding platelet ADP receptor] (n=100); rs1126643 (C807T, Phe224Phe) on ITGA2 [encoding collagen receptor] (n=87); rs5918 (176T>C, Leu33Pro) on ITGB3 [encoding fibrinogen receptor) (n=91); rs6065 (Thr145Met) on GP1BA [encoding platelet receptor for Von Willebrand factor] (n=114); rs4244285 (*2) (n=84), and rs4986893 (*3) (n=83) on СYP2C19 [encoding cytochrome P450 activity]. The prevalence of rs5918, rs6065, rs4244285, rs4986893, rs2046934 did not differ significantly between patients and healthy volunteers. The mutant allele (T) of ITGA2 was detected more often in healthy volunteers: 67.2 % vs 51.7 % (р=0.021). Before and after CABG there was no significant difference in platelet aggregation between carries and non-carries of the mutant ITGA2 allele. Number of CVCs registered during follow-up was 12: 3 strokes, 6 MIs, 3 deaths. Patients with composite mutant alleles of ITGB3+СYP2C19*2 or СYP2C19*2+ITGA2, and with the mutant allele (*2) of СYP2C19 met end points more often than patients with other gene combinations (wild type homozygotes, presence of one mutant allele of ITGB3 or ITGA2, the composite of mutant alleles of ITGB3+ITGA2 or ITGB3+ITGA2+ СYP2C19*2) (hazard ratio 4.0, 95 % confidence interval 2,19-7.29, р=0.008). Carriers of ITGB3 mutant allele showed higher AA-induced platelet reactivity on the 1st-3rd day after CABG. Amplitude of platelet aggregation in carriers of mutant allele was 27.5 % vs 12.7 % in wild type (p=0.016). No differences in platelet reactivity among carriers of other mentioned mutant alleles and wild types were observed. Carriage of the combination of mutant alleles ITGB3+СYP2C19*2 or СYP2C19*2+ITGA2 or СYP2C19*2 is possible predictor of CVC in patients after CABG.

T2238C Atrial Natriuretic Peptide Gene Variant and the Response to Antiplatelet Therapy in Stable Ischemic Heart Disease Patients.

The T2238C variant of the ANP gene is associated with higher risk of major cardiovascular events. The purpose of this study is to investigate if this polymorphism influences the response to antiplatelet agents and it is responsible of increased platelet reactivity, thus contributing to the adverse outcome. In patients undergoing elective percutaneous coronary intervention (PCI), loaded with antiplatelets, blood samples were withdrawn for genotyping, platelet reactivity assessment and for troponin T measurement to investigate the association between the polymorphism with residual platelet reactivity and with the incidence of PPMI. No significant differences in platelet reactivity nor in PPMI incidence were observed between groups. Nevertheless, higher ARU, PRU, and % PI were detected in diabetic patients, with PRU significantly higher in carriers versus non-carriers. We observed increased residual platelet reactivity exclusively in diabetic carriers of the T2238C variant undergoing elective PCI, suggesting the need of a more effective platelet inhibition in this category of patients.

The clinical efficacy and safety evaluation of ticagrelor for acute coronary syndrome in general ACS patients and diabetic patients: A systematic review and meta-analysis.

ObjectiveIn this study, a systematic evaluation was conducted to estimate the efficacy and safety of ticagrelor for treating acute coronary syndrome (ACS) in general ACS patients and a diabetes mellitus (DM) group.MethodsA search of PubMed, Cochrane Central Register of Controlled Trials, Web of Science, CNKI databases was conducted to analyze relevant randomized controlled trails (RCTs) of ticagrelor treating ACS during 2007 to 2015. Article screening, quality accessing and data extracting was independently undertaken by two reviewers. A meta-analysis was performed to clarify the efficacy and safety of ticagrelor in general ACS patients, and a meta-regression analysis was taken to demonstrate the efficacy and safety of ticagrelor in DM patients compared with general ACS patients.ResultTwenty-two studies with 35004 participants were included. The meta-analysis result implicated that ticagrelor could: 1) reduce the incidence of the composite endpoint [OR = 0.83, 95%CI (0.77, 0.90), P<0.00001] and the incidence of myocardial infarction [OR = 0.81, 95%CI (0.74, 0.89), P = 0.0001]; 2) not statistically reduce the incidence of cardiovascular death, the incidence of stroke and the incidence of bleeding events; 3) increase the incidence of dyspnea [OR = 1.90, 95%CI (1.73, 2.08), P<0.00001] compared with clopidogrel. Meanwhile, compared with prasugrel, ticagrelor could 1) reduce the platelet reactivity of patients at maintenance dose [MD = -44.59, 95%CI (-59.16, -30.02), P<0.00001]; 2) not statistically reduce the incidence of cardiovascular death, the platelet reactivity of patients 6 hours or 8 hours after administration, or the incidence of bleeding events; 3) induce the incidence of dyspnea [OR = 13.99, 95%CI (2.58, 75.92), P = 0.002]. Furthermore, the result of meta-regression analysis implicated that there was a positive correlation between DM patients and the platelet reactivity of patients 6 hours and 8 hours after administration, but there was no obvious correlation between DM patients and general ACS patients in other endpoints.ConclusionTicagrelor could reduce the incidence of composite endpoint of cardiovascular death, myocardial infarction and stroke as well as platelet reactivity in DM patients with ACS, while not increasing the risk of bleeding. Because there are differences in platelet reactivity between DM patients and general ACS patients, we suggest that caution is needed when using ticagrelor in clinical applications.