Emese Toth-Zsamboki

The Platelet ATP and ADP Receptors

Adenine nucleotides, ADP and ATP, are coreleased from dense granules during platelet activation, as well as from endothelial cells and damaged red blood cells following vascular injury. Through autocrine and paracrine mechanisms, these extracellular signaling molecules interact with the platelet P2 receptors to amplify ongoing platelet activation. Two receptors for ADP, the G(q)-protein-coupled P2Y1 and G(i)-protein-coupled P2Y12 and one receptor for ATP, the P2X1 ion channel, have been identified on platelets. Due to distinct pharmacological properties and differential regulation, the P2Y and P2X receptors essentially operate on different scales of time and distance and trigger selective intracellular signaling cascades. Recent advances in the understanding of the P2Y receptor physiology have reinforced the concept of these receptors as useful targets for antithrombotic therapy. The function of P2X1 in platelet activation only recently started to be unraveled. This review focuses on recent findings on the physiology of these platelet ADP and ATP receptors, their distinct downstream intracellular signaling pathways as well as on the available agonists, antagonists and inhibitors that allow their pharmacological discrimination.

Analysis of platelet α2-adrenergic receptor activity in stable coronary artery disease patients on dual antiplatelet therapy

Combined antiplatelet therapy reduces recurrent atherothrombotic events in stable coronary disease patients; however, high residual platelet reactivity measured ex vivo still raises concerns as a condition related to treatment failure. Alpha-2 adrenoceptor enhances platelet reactivity and might contribute to this phenomenon. For the present study, 121 stable angina patients on standard dual antiplatelet therapy (75 mg clopidogrel and 100 mg acetylsalicylic acid) were recruited. Born aggregometry was performed with adenosine diphosphate (ADP),collagen and epinephrine. To verify platelet adrenergic activity, potentiation by low-dose epinephrine and inhibition by selective alpha-2 receptor blocker atipamezole were determined. To assess the P2Y12-specific residual activity, cangrelor was used. Plasma norepinephrine, soluble CD40-ligand, high-sensitivity-C-reactive protein (hsCRP) – and in 24 subjects platelet P-selectin positivity were measured. Epinephrine – at very low concentration (10–9g/ml) – significantly potentiates (1.25 μM ADP: 26.5% vs. 43%; 5 μM ADP: 53% vs. 64.5%; collagen: 17% vs 42%, p

ADP receptors in platelet activation and aggregation

ADP plays a central role in platelet aggregation. Activation of platelets via ADP proceeds via three membrane receptor proteins, two of which are coupled to a G protein and the third one constituting an ion channel mediating rapid Ca2+-influx. Via Ca2+-mobilization, the Gq-coupled P2Y1 receptor acts in concert with the Gi-coupled P2TAC receptor, which functions by lowering intracellular cAMP levels. The importance of Ca2+-influx via the P2X1 ion channel remains to be elucidated.ADP plays a central role in platelet aggregation. Activation of platelets via ADP proceeds via three membrane receptor proteins, two of which are coupled to a G protein and the third one constituting an ion channel mediating rapid Ca2+-influx. Via Ca2+-mobilization, the Gq-coupled P2Y 1 receptor acts in concert with the Gicoupled P2T C receptor, which functions by lowering intracellular cAMP levels. The importance of Ca2+-influx via the P2X 1 A ion channel remains to be elucidated.

High on clopidogrel treatment platelet reactivity is frequent in acute and rare in elective stenting and can be functionally overcome by switch of therapy

UNLABELLEDnThe benefit of adjusted antiplatelet therapy in patients with myocardial infarction after primary percutaneous coronary intervention is not well elucidated. We aimed to identify patients with high on treatment platelet reactivity and to gradually adjust antiplatelet therapy.nnnMATERIALS AND METHODSnWe enrolled 133 acute myocardial infarction and 67 stable angina patients undergoing intracoronary stenting into our study. Maximal aggregation was determined with light transmission aggregometry. Aggregation >50% induced by 5 μM ADP was indexed with high on-clopidogrel treatment platelet reactivity. In these cases 75 mg clopidogrel was doubled and control test was performed. Patients effectively inhibited with 150 mg clopidogrel were defined as clopidogrel pseudo non-responders. Patients with high platelet reactivity even on 150 mg clopidogrel were considered as clopidogrel real non-responders and were switched to ticlopidine.nnnRESULTSnAggregations (5ADP; p=0.046) and the ratio of real non-responders (p=0.013) were significantly higher in the myocardial infarction group. Most real non-responders were effectively treated with switch of therapy. The ratio of pseudo non-responders also tended to be higher in myocardial infarction. Platelet reactivity remained constant during follow-up; however, a new appearance of high platelet reactivity was observed at 6 and at 12 months.nnnCONCLUSIONSnPatients with acute myocardial infarction undergoing percutaneous coronary intervention may benefit from prospective platelet function testing, because of higher platelet reactivity and much higher ratio of clopidogrel real non-response. Switch of therapy may effectively overcome clopidogrel non-response. A new appearance of high platelet reactivity with unknown clinical significance is observed in both groups among the patients on clopidogrel.

Cardiac rehabilitation programme as a non-pharmacological platelet inhibitory tool in acute coronary syndrome survivors

Background Acute coronary syndrome is associated with platelet hyperactivity, which in its persistent form, promotes recurrent thrombotic events. Complex cardiac rehabilitation after acute coronary syndrome improves clinical outcome; however, its effect on platelet hyperactivity is unknown. Design and methods We enrolled 84 acute coronary syndrome patients on dual antiplatelet therapy, who underwent a new complex cardiac rehabilitation programme (NovaCord physiotherapy, lifestyle counselling, strict diet, stress management and regular coaching) and 51 control acute coronary syndrome patients with traditional cardiac rehabilitation. Platelet functionality was determined at enrolment and at three months follow-up by aggregometry, serum platelet-derived growth factor levels, total- and platelet-derived microvesicle counts (PMV; CD41a+/CD61+, CD62P+). Results Platelet aggregation parameters and platelet-derived growth factor levels were significantly decreased in the complex cardiac rehabilitation group at three months (1u2009µg/ml collagen, median (interquartile range): 22 (10–45) vs 14 (7.5–25.5)%, pu2009=u20090.0015; 2u2009µg/ml collagen: 36 (22–60) vs 26.5 (16–37)%, pu2009=u20090.0019; 1.25u2009µM adenosine-diphosphate: 4.5 (1–10) vs 1 (0–3)%, pu2009=u20090.0006; 5u2009µM adenosine-diphosphate: 27 (16–38) vs 22 (12–31)%, pu2009=u20090.0078; epinephrine: 33 (15–57) vs 27 (12–43)%, pu2009=u20090.01; platelet-derived growth factor: 434.6 (256.0–622.7) vs 224.8 (148.5–374.1) pg/ml, pu2009=u20090.0001). In contrast, these changes were absent or did not reach statistical significance in the traditional cardiac rehabilitation group. Platelet–derived microvesicle counts were significantly decreased in both groups, while total microvesicle count was significantly reduced only in the complex cardiac rehabilitation group (median (interquartile range): 3945.5 (2138–5661) vs 1739 (780–2303) count/µl; pu2009=u20090.0001). Conclusions Platelet hyperactivity three months after acute coronary syndrome significantly decreased in patients undergoing complex cardiac rehabilitation. Besides dual antiplatelet therapy, effective management and comprehensive control of cardiovascular risk factors might represent a new, non-pharmacological approach to influence platelet functionality.

Predictors of high on-clopidogrel platelet reactivity in patients with acute coronary syndrome.

Abstract High on-clopidogrel platelet reactivity (HPR) is a predictor of ischemic events after percutaneous coronary intervention. We conducted a prospective cohort study to identify variables related to HPR in acute coronary syndrome patients who are at high thrombotic risk. We enrolled 463 patients undergoing urgent coronary angiography. Platelet reactivity was measured 12–36 hours after 600u2009mg clopidogrel loading with multiple electrode aggregometry (Multiplate® analyzer, Roche, Basel, Switzerland, 6.4u2009µM ADP). HPR was defined by the consensus cut-off area under the curve >46u2009U. The rate of HPR was 16.0%. We analyzed simple clinical and laboratory parameters with backward multivariate logistic regression and identified the following predictors of HPR: platelet count (per G/L, OR: 1.0073, 95% CI: 1.0035–1.0112, pu2009=u20090.0002), CRP level (per mg/L, OR: 1.0077, 95% CI: 1.0016–1.01372, pu2009=u20090.01), and active smoking (OR: 0.51, 95% CI: 0.29–0.89, pu2009=u20090.02). We developed and internally validated a risk prediction model demonstrating moderate discriminative capacity (area-under-the-receiver operating characteristic curveu2009=u20090.67). In conclusion, we found a relatively low rate of high on-clopidogrel platelet reactivity (16.0%) even in an acute patient cohort. HPR measured by Multiplate was associated with high platelet count and CRP level on admission and was inversely related to active smoking. The model with rapidly available simple parameters might help to identify individuals at risk for HPR in the acute setting.

Non-invasive, Complex Examination of Micro- and Macrovascular System of Patients with Type 1 Diabetes Mellitus with or Without Vascular Complications

Abstract Objective: We examined the vascular system, from the microvasculature to the aorta, in diabetes mellitus, using non-invasive methods. Methods: We enrolled patients with type 1 diabetes: 17 patients without complications (DMW) and 19 patients with clinically manifest complications (DMC). Control group was represented by 34 healthy volunteers (C). We examined microvascular function with laser-Doppler flowmetry, using post-occlusive reactive hyperemia test and local heating. Arterial stiffness was studied by arteriograph, determining augmentation index and pulse wave velocity. We measured serum levels of sE-selectin and sICAM-1, markers of endothelial dysfunction. Results: Microvascular reactivity was significantly reduced in DMC-, and tendentiously in DMW groups. sE-selectin level was significantly higher in DMC group than in controls. Arterial stiffness was the highest in the DMC group and the lowest in the DMW group. Heart rate was significantly higher in both diabetic groups compared to controls. Time to maximum flow during PORH test tended to be the shortest in DMW group. Conclusions: Our results confirm impairment of the microvascular system in diabetic patients, even in early, uncomplicated stage of the disease, and might demonstrate diffuse hyperkinesis in the vascular system, resulting from the insulin effect or refering to the “vasodilation phase” of diabetes mellitus.