Platelet ADP Research Articles

METABOLIC AND HEMOSTATIC PARAMETERS IN PRE-DIABETES AND NEWLY DIAGNOSED TYPE 2 DIABETES

Metabolic changes in diabetes mellitus are associated with hemostatic disorders. However, the sequence of hemostatic events in the early stages of disease remains unclear. Aim. To assess metabolic and hemostatic parameters and their interaction in pre-diabetes and newly diagnosed type 2 diabetes mellitus (ND T2DM). Material and methods. The study enrolled volunteers of 40 to 65 years who considered themselves healthy and did not get any medication therapy. Of 170 examined individuals, 46 had impaired carbohydrate exchange (ICE) — 13 with impaired fasting glucose, 17 — impaired glucose tolerance and 16 with ND T2DM. The control group comprised healthy volunteers with normal body mass index and without signs of metabolic abnormalities. The metabolic (carbohydrate, lipid, protein exchange, hepatic transaminase), platelet and plasma hemostatic parameters (mean platelet volume, ADP- &collagen-induced platelet aggregation, coagulation profile, fibrinogen, plasminogen) were investigated. We identified the peculiarities initiating impact of changed parameters on different hemostatic components in patients with pre-diabetes and ND T2DM. Results. Concentration of insulin, C-peptide, Homa-IR, total cholesterol demonstrated increase in groups with ICE. ADP-induced platelet aggregation, fibrinogen increased in ICE, however these changes were not statistically significant. Mean platelet volume and plasminogen had the tendency to be elevated in pre-diabetes and demonstrated significant increase in ND T2DM. Conclusion. Metabolic disorders in prediabetic stage initiate changes in platelet hemostasis and fibrinolysis. The increase of MPV and higher concentrations of plasminogen are considered to be significant in ND T2DM.

Evaluation on antithrombotic effect of aspirin eugenol ester from the view of platelet aggregation, hemorheology, TXB2/6-keto-PGF1α and blood biochemistry in rat model.

BackgroundBased on the prodrug principle, aspirin and eugenol, as starting precursors, were esterified to synthesize aspirin eugenol ester (AEE). The aim of the present study was to evaluate the antithrombotic effect of AEE in an animal disease model. In order to compare the therapeutic effects of AEE and its precursors, aspirin, eugenol and a combination of aspirin and eugenol were designed at the same molar quantities as the AEE medium dose in the control group.MethodsAfter oral administration of AEE (dosed at 18, 36 and 72 mg/kg) for seven days, rats were treated with k-carrageenan to induce tail thrombosis. Following the same method, aspirin (20 mg/kg), eugenol (18 mg/kg) and 0.5 % CMC-Na (30 mg/kg) were administered as control drug. Different drug effects on platelet aggregation, hemorheology, TXB2/6-keto-PGF1α ratio and blood biochemistry were studied.ResultsAEE significantly inhibited ADP and AA-induced platelet aggregation in vivo. AEE also significantly reduced blood and plasma viscosity. Moreover, AEE down-regulated TXB2 and up-regulated 6-keto-PGF1α, normalizing the TXB2/6-keto-PGF1α ratio and blood biochemical profile. In comparison with aspirin and eugenol, AEE produced more positive therapeutic effects than its precursors under the same molar quantity.ConclusionIt may be concluded that AEE was a good candidate for new antithrombotic and antiplatelet medicine. Additionally, this study may help to understand how AEE works on antithrombosis in different ways.

SIRT1 prevents pulmonary thrombus formation induced by arachidonic acid via downregulation of PAF receptor expression in platelets

SIRT1, a class III histone deacetylase, is critically involved in cellular response to stress and modulates cardiovascular risk factors. However, its role in thrombus formation is largely unknown. Thus, this study investigated the effect of SIRT1 on pulmonary thrombus formation, and then identified its role in the modulation of platelet aggregation. In isolated human platelets, cell aggregation was increased by various platelet activators, such as platelet activating factor (PAF), arachidonic acid (AA), ADP, and thrombin. AA- and PAF-mediated platelet aggregations were suppressed by WEB2086, a PAF receptor (PAFR) antagonist. Pulmonary thrombus formation induced by PAF or AA was also attenuated by WEB2086, suggesting that PAFR plays a key role in AA-induced platelet aggregation. In platelets isolated from SIRT1-TG mice as well as in platelets treated with resveratrol or reSIRT1, PAFR expression was decreased, whereas this expressional downregulation by SIRT1 activators was inhibited in platelets treated with MG132 (a proteasome inhibitor) or NH4Cl (a lysosome inhibitor). Furthermore, platelet aggregation induced by AA was markedly attenuated by resveratrol and reSIRT1. Likewise, the increased pulmonary thrombus formation in mice treated with AA was also attenuated by SIRT1 activators. In line with these results, pulmonary thrombus formation was markedly attenuated in SIRT1-TG mice. Taken together, this study showed that SIRT1 downregulates PAFR expression on platelets via proteasomal and lysosomal pathways, and that this downregulation inhibits platelet aggregation in vitro and pulmonary thrombus formation in vivo.

FAK regulates platelet extravasation and tumor growth after antiangiogenic therapy withdrawal.

Recent studies in patients with ovarian cancer suggest that tumor growth may be accelerated following cessation of antiangiogenesis therapy; however, the underlying mechanisms are not well understood. In this study, we aimed to compare the effects of therapy withdrawal to those of continuous treatment with various antiangiogenic agents. Cessation of therapy with pazopanib, bevacizumab, and the human and murine anti-VEGF antibody B20 was associated with substantial tumor growth in mouse models of ovarian cancer. Increased tumor growth was accompanied by tumor hypoxia, increased tumor angiogenesis, and vascular leakage. Moreover, we found hypoxia-induced ADP production and platelet infiltration into tumors after withdrawal of antiangiogenic therapy, and lowering platelet counts markedly inhibited tumor rebound after withdrawal of antiangiogenic therapy. Focal adhesion kinase (FAK) in platelets regulated their migration into the tumor microenvironment, and FAK-deficient platelets completely prevented the rebound tumor growth. Additionally, combined therapy with a FAK inhibitor and the antiangiogenic agents pazopanib and bevacizumab reduced tumor growth and inhibited negative effects following withdrawal of antiangiogenic therapy. In summary, these results suggest that FAK may be a unique target in situations in which antiangiogenic agents are withdrawn, and dual targeting of FAK and VEGF could have therapeutic implications for ovarian cancer management.

Impact of the CYP2C19 Gene Polymorphism on Clopidogrel Personalized Drug Regimen and the Clinical Outcomes.

Although the relationship between CYP2C19 genotype and clopidogrel metabolism has been studied clearly, we have not seen the report that clopidogrel was administered at a dose adjusted based on genotyping. The two main polymorphism loci of CYP2C19 gene were detected by the CYP2C19 genetic testing. ADP platelet aggregation technology was used to investigate the correlation between clinical effect and the clopidogrel dose, genetic metabolic type, physiological, pathological and other factors, to provide new ideas for clopidogrel therapy for percutaneous coronary intervention postoperation patients. A total of 48 patients were enrolled. All patients were given clopidogrel routine maintenance dose treatment and underwent CYP2C19 genotyping and platelet function testing. Patients were divided into extensive metabolizers, intermediate metabolizers, and poor metabolizers based on the different CYP2C19 genotypes. The clopidogrel dosage was adjusted to double the maintenance dose for the ineffective patients. The study showed that all patients had no toxic side effects. The low responsiveness to clopidogrel in patients with diabetes is closely related to insulin resistance. Patients with hypertension and hyperlipidemia may increase the risk of clopidogrel resistance. The occurrence of clopidogrel resistance is associated with the CYP2C19 polymorphism in the < 65-year-old female patients. For percutaneous coronary intervention in postoperative patients, research data is still lacking regarding clopidogrel dosage on the basis of different CYP2C19 genotypes. Looking forward, more rigorous research programs need to be designed to bring more guidance for clinical application.

Viscoelastic measurements of platelet function, not fibrinogen function, predicts sensitivity to tissue‐type plasminogen activator in trauma patients

Systemic hyperfibrinolysis is a lethal phenotype of trauma-induced coagulopathy. Its pathogenesis is poorly understood. Recent studies have support a central role of platelets in hemostasis and in fibrinolysis regulation, implying that platelet impairment is integral to the development of postinjury systemic hyperfibrinolysis. The objective of this study was to identify if platelet function is associated with blood clot sensitivity to fibrinolysis. We hypothesize that platelet impairment of the ADP pathway correlates with fibrinolysis sensitivity in trauma patients. A prospective observational study of patients meeting the criteria for the highest level of activation at an urban trauma center was performed. Viscoelastic parameters associated with platelet function (maximum amplitude [MA]) were measured with native thrombelastography (TEG), and TEG platelet mapping of the ADP pathway (ADP-MA). The contribution of fibrinogen to clotting was measured with TEG (angle) and the TEG functional fibrinogen (FF) assay (FF-MA). Another TEG assay containing tissue-type plasminogen activator (t-PA) (75 ng mL(-1) ) was used to assess clot sensitivity to an exogenous fibrinolytic stimulus by use of the TEG lysis at 30 min (LY30) variable. Multivariate linear regression was used to identify which TEG variable correlated with t-PA-LY30 (quantification of fibrinolysis sensitivity). Fifty-eight trauma patients were included in the analysis, with a median injury severity score of 17 and a base deficit of 6 mEq L(-1) . TEG parameters that significantly predicted t-PA-LY30 were related to platelet function (ADP-MA, P = 0.001; MA, P < 0.001) but not to fibrinogen (FF-MA, P = 0.773; angle, P = 0.083). Clinical predictors of platelet ADP impairment included calcium level (P = 0.001), base deficit (P = 0.001), and injury severity (P = 0.001). Platelet impairment of the ADP pathway is associated with increased sensitivity to t-PA. ADP pathway inhibition in platelets may be an early step in the pathogenesis of systemic hyperfibrinolysis.

Assessment of platelet function in endogenous hypercortisolism.

It is commonly known that glucocorticoids exert a significant effect on haemostasis. Studies that have analysed the plasmatic coagulation system and fibrinolysis parameters in hypercortisolaemic patients are abundant. Platelet function, which plays a vital role in primary haemostasis, is much less clear in this context. We aimed at assessing platelet function in endogenous hypercortisolaemic patients. Twenty-five hypercortisolaemic patients were included in the study. Twelve of them were diagnosed with overt Cushing's syndrome (OCS) and 13 had subclinical Cushing's syndrome (SCS). Thirty healthy volunteers comprised the control group. In all subjects platelet function parameters were examined: ADP- and collagen-induced platelet aggregation (ADP-IPA and Col-IPA, respectively), IMPACT R (expressed as percentage of surface covered (SC) by platelets and average size (AS) of the adhering particles in μm2), as well as closure time (CT) after platelet activation with agonists: ADP and Col or Col and epinephrine (EPI). The statistical significance level was set at 0.05. There was no significant difference in mean values of ADP-IPA, Col-IPA, Col/Epi CT, Col/ADP CT, SC, and AS between hypercortisolaemic subjects and controls. No statistically significant differences in means of examined parameters were found between overt and subclinical Cushing's syndrome patients. Furthermore, no statistically significant relationships were found between these parameters and hormonal indicators of hypercortisolism: 24-hour urinary cortisol excretion, morning and evening serum cortisol level, and overnight-test cortisol concentration. In hypercortisolaemic patients no primary haemostasis disorders are present, as reflected by platelet adhesion and ADP- and collagen-induced aggregation measurements.

Synthesis and Antiplatelet Aggregation Activity Evaluation of some 2-Aminopyrimidine and 2-Substituted-4,6-diaminopyrimidine Derivatives.

A series of novel 2-aminopyrimidine and 2-Substituted-4,6-diaminopyrimidine derivatives have been synthesized and their antiplatelet aggregation activities were assessed against ADP and arachidonic acid-induced platelet aggregation in human plasma using light transmission aggregometry. Among the tested derivatives, compounds Ia, Ib, IB and II16 exhibited the highest antiplatelet aggregation activity (36.75, 72.4, 62.5 and 80 µM). None of the compounds showed satisfactory activity against the aggregation induced by ADP but acceptable activities were observed against the aggregation induced by arachidonic acid. 2- aminopyrimidines were more active than 4,6- diaminopyrimidines in this respect.

Managing Antiplatelet Therapy

1. George A. Fritsma, MS, MLS[⇑][1] 1. The Fritsma Factor, Your Interactive Hemostasis Resource, Fritsma & Fritsma LLC, Birmingham, AL <!-- --> 1. Address for Correspondence: George A. Fritsma, MS, MLS, The Fritsma Factor, Your Interactive Hemostasis Resource, Fritsma & Fritsma LLC, 153 Redwood Drive, Birmingham, AL 35173, George{at}fritsmafactor.com 1. List antiplatelet drugs that are used in dual antiplatelet therapy. 2. Describe the basis for aspirin and thienopyridine antiplatelet property. 3. Define aspirin and clopidogrel low response. 4. List and describe platelet function testing that is applied to detecting aspirin and clopidogrel low response. Dual Antiplatelet Therapy In 1955, President Dwight Eisenhower, recovering from an acute myocardial infarction (MI), was treated with warfarin (Coumadin®) to prevent a repeat MI (Figures 1 and 2).1 Coumadin, FDA-cleared in 1954, had become an essential part of acute coronary syndrome treatment; it was intended to reduce the risk of a secondary MI, peripheral artery disease, ischemic stroke, and venous thromboembolic disease subsequent to an MI. Coumadin continues today as the most-prescribed anticoagulant in North America, though now its most frequent indication is prophylaxis to reduce the risk of ischemic stroke in non-valvular atrial fibrillation. Because of its narrow therapeutic and safety range, Coumadin requires monthly laboratory monitoring using the prothrombin time assay, which, beginning in 1987, was enhanced through computation of the international normalized ratio (PT/INR).2 The accepted therapeutic range worldwide is an INR of 2–3. An INR below 2 signals increased risk of thrombosis, above 3, risk of hemorrhage.3 In 1992, thienopyridine, a nucleic acid derivative distributed as ticlopidine (Ticlid®), was added to post-MI Coumadin therapy (Figure 3).4 Ticlopidine is an antiplatelet drug that reversibly occupies the platelet membrane ADP receptor, P2Y12 (Figure 3). Ticlopidine competes with ADP for P2Y12 receptor sites, suppressing ADP's platelet activation property. From 1992 to 2000, Coumadin and Ticlid administered together were the most commonly used post-MI antithrombotic regimen. This regimen was typically discontinued six months to two years after the event. Regrettably, 1 in 3–5000 ticlopidine patients developed life-threatening thrombotic thrombocytopenic purpura or aplastic anemia, creating a negative public perception.5 This… ABBRVIATIONS: AA-arachidonic acid, ACS-acute coronary syndrome, ADP-adenosine diphosphate, ALR-aspirin low responder, AMI-acute myocardial infarction, ARU-aspirin reaction (or resistance) units, ASA-acetylsalicylic acid, CABG-coronary artery bypass graft, CLR-clopidogrel low responder, COX-cyclooxygenase, CT-closure time, DAPT-dual antiplatelet therapy, CLIA-clinical laboratory improvements amendment, FDA-US Food and Drug Administration, LTA-light transmittance aggregometry, MI-myocardial infarction, NSAID-non-steroidal anti-inflammatory drug, CYP-cytochrome oxidase pathway, PCI-percutaneous intervention, PGE1-prostaglandin E1, PGG2-prostaglandin G2, PGH2-prostaglandin H2, POC-point of care, PRU-P2Y12 reaction units, PT/INR-prothrombin time with international normalized ratio, TEG-thromboelastography, TEM-thromboelastometry, TXA2-thromboxane A2, WBA-whole blood aggregometry, VASP-vasodilator-stimulated phosphoprotein 1. List antiplatelet drugs that are used in dual antiplatelet therapy. 2. Describe the basis for aspirin and thienopyridine antiplatelet property. 3. Define aspirin and clopidogrel low response. 4. List and describe platelet function testing that is applied to detecting aspirin and clopidogrel low response. [1]: #corresp-1

Effects of chronic administration of valproic acid to epileptic patients on coagulation tests and primary hemostasis.

Valproic acid (VPA) is an antiepileptic drug that has been associated with impaired hemostasis and increased risk for postsurgical bleeding. However, the published reports provide controversial results. We measured parameters of primary hemostasis in VPA-treated patients with epilepsy, focusing on adenosine nucleotide-dependent platelet responses, which play a central role in primary hemostasis. We enrolled 20 cases (epileptic patients receiving treatment with VPA) and 20 controls (12 epileptic patients receiving treatment with drugs different from VPA and 8 healthy subjects). Measurements included prothrombin time (PT), activated partial thromboplastin time (APTT), platelet count, platelet function analyzer (PFA)-100 closure times, plasma von Willebrand factor levels, platelet content of ADP, ATP, and serotonin (all stored in platelet dense granules), and platelet shape change and aggregation induced by ADP and other platelet agonists, including the ATP analog α,β-methylene-ATP. The plasma concentration of VPA was in the therapeutic range in 17 patients and slightly above the upper limit in 3 patients. There were no statistically significant differences in any of the studied parameters in cases versus controls. Our thorough controlled study failed to show that chronic treatment with VPA induces significant abnormalities of coagulation and primary hemostasis. Therefore, VPA, when present in the circulation in the therapeutic range, does not impair hemostasis.

Dl-3-n-butylphthalide inhibits platelet activation via inhibition of cPLA2-mediated TXA2 synthesis and phosphodiesterase

Aberrant platelet activation plays a critical role in the pathogenesis of heart attack and stroke. dl-3-n-butylphthalide (NBP) has been approved in China to treat stroke with multiple mechanisms. The anti-stroke effects of NBP may be related to its antiplatelet effects reported in rats in addition to its antioxidative, antiapoptotic, and angiogenic effects. However, the effects and the underlying mechanisms of NBP on human platelets are not yet clear. In this study, we found that NBP concentration-dependently inhibited human platelet aggregation and ATP release induced by ADP, thrombin, U46619, arachidonic acid, or collagen. NBP also inhibited PAC-1 binding induced by ADP or thrombin and platelet spreading on immobilized fibrinogen. NBP reduced TXA2 synthesis induced by thrombin or collagen via inhibiting cPLA2 phosphorylation, concomitantly with a marked decrease in intracellular calcium mobilization. Moreover, NBP also inhibited human platelet phosphodiesterase (PDE) and elevated 3,5-cyclic adenosine monophosphate level in platelets. In conclusion, NBP significantly inhibits human platelet activation via inhibition of cPLA2-mediated TXA2 synthesis and PDE, and may be effective as an antiplatelet drug to treat other arterial thrombotic diseases.

Abstract T P111: General Anesthetic Use and Clopidogrel Resistance During Neurovascular Stenting Procedures

Purpose: To determine the effect of general anesthesia on clopidogrel resistance among patients undergoing intracranial stenting procedures Background: Clopidogrel resistance is associated with increased periprocedural thromboembolic complications in neurovascular stenting procedures. Clopidogrel is metabolized by cytochrome P450 system to an active drug that irreversibly binds to the platelet ADP receptor P2Y12 and inhibits downstream platelet aggregation. General anesthetics are commonly used in intracranial stenting procedures and are known to inhibit cytochrome P450 monooxygenases. Therefore we hypothesized that the use of common general anesthetics would be associated with increased clopidogrel resistance. Methods: We conducted a retrospective chart review of 41 patients who underwent intracranial stenting and coiling procedures under general anesthesia from 2010-2013. We analyzed the 35/41 who had platelet assays performed within twenty-four hours both before and after the procedure (VerifyNow, Accumetrics, Inc., San Diego, CA, USA). P2Y12 reaction unit (PRU) was used to measure platelet resistance, with values ≤178 indicating increased bleeding risk, 179-238 therapeutic, and ≥239 thrombogenic. Pre-procedural and post-procedural mean PRU values were compared using t tests to determine the effect of general anesthesia on clopidogrel resistance. Results: We found significantly increased post-procedure PRUs compared to pre-procedure PRUs. Mean PRU twenty-four hours before procedure was 214±34 (95% CI 180-248) compared with 260±25.24 (95% CI 235-285) twenty-four hours after the procedure (p&lt;0.05). Out of 35 patients, 27 (77%) had increased PRUs post- procedure compared to pre-procedure. Conclusion: Clopidogrel resistance is increased following endovascular stenting procedures. We propose that this occurs through competitive interaction with CYP isoforms by general anesthetics. We are not aware of any prior studies demonstrating an association between general anesthetics and clopidogrel resistance. Further prospective studies are needed to further define this observation.

Quantification of ATP and ADP levels in platelets from healthy human donors: a potential novel indicator of cardiovascular risk

Background Understanding the biology of platelets is essential for the management of thrombotic diseases like stroke and cardiovascular disease. Platelets store ATP and ADP biomolecules and secrete them in response to thrombotic stimuli. However, patient-to-patient variability in storage of these biomolecules in platelets is poorly studied [1]. This study assesses ATP and ADP concentration in platelets and determines (1) the absolute values of these biomolecules per platelet, (2) the ratio of ATP and ADP in platelets and (3) the quantities released from dense granule storage sites in response to a discrete stimulus (Thrombin receptor activating peptide (TRAP); 10μM).

Experimental evaluation of analgesic, anti-inflammatory and anti-platelet potential of Dashamoola.

Background:Dashamoola, in the form of arishta and kwath, is a commonly used classical Ayurvedic multi-ingredient formulation for management of pain, arthritis and inflammatory disorders.Objective:To study analgesic, anti-inflammatory and anti-platelet activity of Dashamoola and its combination with aspirin.Materials and Methods:Wistar albino rats (180-200 g) and Swiss albino mice (20-25 g) of either sex were divided randomly into five groups: Distilled water, aspirin (500mg/kg in rats; 722.2 mg/kg in mice), Dashamoolarishta (1.8 mL/kg in rats; 2.5 mL/kg in mice) and Dashamoolarishta with aspirin. Anti-inflammatory activity was measured by change in paw volume in carrageenan-induced inflammation, protein content in model of peritonitis and granuloma weight in cotton pellet granuloma. Analgesic effect was evaluated by counting number of writhes in writhing model. Maximum platelet aggregation and percentage inhibition of ADP and collagen-induced platelet aggregation were estimated in vitro. Statistical analysis was done using one way ANOVA (post hoc Tukey's test) and P < 0.05 was considered significant.Results:Dashamoolarishta and its combination with aspirin showed significantly (P < 0.01) less number of writhes. It showed significant (P < 0.001) anti-inflammatory activity by paw edema reduction in rats, decrease in proteins in peritoneal fluid (P < 0.001) and decrease in granuloma weight (P < 0.05) as compared to respective vehicle control groups. Dashamoola kwath alone and in combination with aspirin inhibited maximum platelet aggregation and percent inhibition of platelets as compared to vehicle (P < 0.001).Conclusion:Dashamoola formulation alone and its combination with aspirin showed comparable anti-inflammatory, analgesic and anti-platelet effects to aspirin.

Genetic polymorphisms of CYP2C19*2 and ABCB1 C3435T affect the pharmacokinetic and pharmacodynamic responses to clopidogrel in 401 patients with acute coronary syndrome

Backgrounds and objectivesClopidogrel, an inhibitor of platelet ADP P2Y12 receptors, plays an important role in the prevention of stent thrombosis. However, some patients do not attain adequate antiplatelet effects. Studies have shown that the genetic variation in CYP2C19*2 is associated with an impaired response to clopidogrel. This study was designed to investigate the genetic variants of 21 genes involving in the absorption, metabolism, and pharmacodynamics of clopidogrel. The effects of these genes on the plasma level of clopidogrel and its metabolites (active clopi-H4 and inactive CLPM) and platelet reactivity were also studied. Methods and results401 acute coronary syndrome (ACS) patients received either a 300mg loading dose following 75mg maintenance dose daily or a 75mg maintenance dose daily of clopidogrel. The inhibition of platelets was assessed using light transmittance aggregometry. Plasma concentrations of clopidogrel as well as its active (clopi-H4) and inactive (CLPM) metabolites were measured using HPLC-MS-MS method. Among 21 genes, the carriers of CYP2C19*2 were associated with lower exposure to its active (clopi-H4) and inactive (CLPM) metabolites (both P<0.05 vs. non-carriers) and thus decreased platelet inhibition (P<0.05 vs. non-carriers). Notably, the carriers of ABCB1 C3435T were associated with lower levels of plasma clopidogrel and its active (clopi-H4) and inactive (CLPM) metabolites (all P<0.05 vs. non-carriers) which also correlated with subsequently decreased platelet inhibition (P<0.05 vs. non-carriers). There were no obvious effects of other studied genes on clopidogrel. ConclusionsCYP2C19*2 is a determinant for the formation of the active metabolite of clopidogrel and its antiplatelet effects. Meanwhile, ABCB1 C3435T plays an important role in intestinal absorption of clopidogrel which further affects the exposure to the active metabolite of clopidogrel and platelet aggregation.

Association Between the P2RY12 Receptor Gene Polymorphism and Aspirin Resistance in Patients with Coronary Artery Disease.

IntroductionPlatelet activation and aggregation are key elements in the development of coronary atherosclerosis. Recent studies have shown that the two polymorphisms of platelet ADP receptor P2RY12 (haplotypes H2 and 34T) are associated with increased platelet aggregation and atherothrombotic risk. It was shown that these polymorphisms promote reduced body response to antiplatelet therapy.AimWe investigated the association of P2RY12 gene polymorphisms with aspirin resistance in patients with coronary artery disease (CAD).MethodsThis case-control study included 100 cases with CAD (mean age 57.6 ± 2.8 years) treated in the cardiology department of the city hospital Semey, Kazakhstan, 90 of whom suffered from myocardial infarction. The control group (n = 100) were healthy people without a history of CAD, matched on sex and age. Genotyping of polymorphisms H1/H2 in P2RY12 gene was performed by PCR. Statistical analysis was performed using SPSS v.19.0.ResultsThe distribution of H1/H2 genotypes P2RY12 was 42%, 34%, and 24%, respectively, in cases and 42%, 58%, and 0%, respectively, in controls. All allele frequencies were consistent with the Hardy Weinberg equilibrium (p = 0.0036 and p = 0.0001 in cases and controls, respectively). Genotype H2 was associated with risk of CAD with aspirin resistance (co-dominant model: OR = 3.75, 95% CI 0.14 – 99.88, p = 0.05 and dominant model: OR = 2.78, 95% CI 0.11 – 70.93, p = 0.05). We found significant differences in the distribution of the mutant genotype H2 between CAD patients with aspirin resistance and healthy controls (χ2 = 30.3, p < 0.05).ConclusionWe found an association of H2 haplotype in P2RY12 gene with aspirin resistance in patients with CAD. However, in order to obtain definitive conclusions about the role of genetic variants with the development of aspirin resistance in patients with CAD, there is a need for further research with a larger sample size as well as the use of selective thromboxane receptor antagonists for studying functional effects of genetic variants.

Pretreatment with P2Y12 inhibitors in non-ST-Segment-elevation acute coronary syndrome: an outdated and harmful strategy.

Coronary plaque rupture that occurs during the acute coronary syndrome (ACS) results in the deposition and activation of platelets and the formation of thrombi. The platelet mediators thromboxane A2 and ADP have been demonstrated to play a dominant role in the initiation and propagation of coronary thrombosis, representing key therapeutic targets in the management of ACS. Early inhibition of these biomarkers with oral administration of aspirin and a P2Y12 receptor inhibitor together with anticoagulation has been recommended to avoid complete thrombotic occlusion, which may occur during the initial platelet response to vascular injury when the diagnosis of non–ST-segment–elevation ACS (NSTE-ACS) is made or after percutaneous coronary intervention (PCI). An early invasive strategy of cardiac catheterization and revascularization is recommended in patients in whom the thrombotic burden is the highest. A routine invasive versus selective invasive strategy was shown to reduce the rates of death and myocardial infarction, with the most pronounced difference in these high-risk patients.1 In particular, an early catheterization followed by coronary intervention on the first day of hospitalization was shown to be safe and superior in terms of lower risk of recurrent ACS (−41%) and shorter hospital stay (−28%).2 The strategy of “sooner is better” for catheterization suggests also having the same strategy for antiplatelet strategy, which has remained unproven. Guidelines have recommended early initiation of dual antiplatelet therapy, a combination of aspirin and clopidogrel, as soon as possible when the diagnosis of high-risk NSTE-ACS is made, although the timing of loading remained unclear.3,4 This concept of treatment of NSTE-ACS at the time of presentation or admission is known as pretreatment, a treatment given when the diagnosis is suspected and always before the coronary status is known to confirm the diagnosis of coronary artery disease (CAD) and to make the …

Impaired platelet nitric oxide response in patients with new onset atrial fibrillation

BackgroundClinical factors associated with thromboembolic risk in AF patients are well characterized and include new onset AF. Biochemically, AF is associated with inflammatory activation and impairment of nitric oxide (NO) signalling, which may also predispose to thromboembolism: the bases for variability in these anomalies have not been identified. We therefore sought to identify correlates of impaired platelet NO signalling in patients hospitalized with atrial fibrillation (AF), and to evaluate the impact of acuity of AF. Methods87 patients hospitalized with AF were evaluated. Platelet aggregation, and its inhibition by the NO donor sodium nitroprusside, was evaluated using whole blood impedance aggregometry. Correlates of impaired NO response were examined and repeated in a “validation” cohort of acute cardiac illnesses. ResultsWhilst clinical risk scores were not significantly correlated with integrity of NO signalling, new onset AF was associated with impaired NO response (6±5% inhibition versus 25±4% inhibition for chronic AF, p<0.01). New onset AF was a multivariate correlate (p<0.01) of impaired NO signalling, along with platelet ADP response (p<0.001), whereas the associated tachycardia was not. Platelet ADP response was predicted by elevation of plasma thrombospondin-1 concentrations (p<0.01). Validation cohort evaluations confirmed that acute AF was associated with significant (p<0.05) impairment of platelet NO response, and that neither acute heart failure nor acute coronary syndromes were associated with similar impairment. ConclusionRecent onset of AF is associated with marked impairment of platelet NO response. These findings may contribute to thromboembolic risk in such patients.

Isolation and characterization of four medium-size disintegrins from the venoms of Central American viperid snakes of the genera Atropoides, Bothrops, Cerrophidion and Crotalus.

Four disintegrins were isolated from the venoms of the Central American viperid snakes Atropoides mexicanus (atropoimin), Bothrops asper (bothrasperin), Cerrophidion sasai (sasaimin), and Crotalus simus (simusmin). Purifications were performed by reverse-phase HPLC. The four disintegrins have biochemical characteristics, i.e. molecular mass and location of Cys, which allow their classification within the group of medium-size disintegrins. All of them present the canonical RGD sequence, which determines their interaction with integrins in cell membranes. The disintegrins inhibited ADP and collagen-induced human platelet aggregation, with similar IC50s in the nM range. In addition, disintegrins inhibited the adhesion of an endothelial cell line and a melanoma cell line to the extracellular matrix proteins type I collagen, laminin, fibronectin, and vitronectin, albeit showing variable ability to exert this activity. This study expands the inventory of this family of viperid venom proteins, and reports, for the first time, disintegrins from the venoms of species of the genera Atropoides and Cerrophidion.

Effects of acute and chronic psychological stress on platelet aggregation in mice

Although psychological stress has long been known to alter cardiovascular function, there have been few studies on the effect of psychological stress on platelets, which play a pivotal role in cardiovascular disease. In the present study, we investigated the effects of acute and chronic psychological stress on the aggregation of platelets and platelet cytosolic free calcium concentration ([Ca2+]i). Mice were subjected to both transportation stress (exposure to novel environment, psychological stress) and restraint stress (psychological stress) for 2 h (acute stress) or 3 weeks (2 h/day) (chronic stress). In addition, adrenalectomized mice were subjected to similar chronic stress (both transportation and restraint stress for 3 weeks). The aggregation of platelets from mice and [Ca2+]i was determined by light transmission assay and fura-2 fluorescence assay, respectively. Although acute stress had no effect on agonist-induced platelet aggregation, chronic stress enhanced the ability of the platelet agonists thrombin and ADP to stimulate platelet aggregation. However, chronic stress failed to enhance agonist-induced increase in [Ca2+]i. Adrenalectomy blocked chronic stress-induced enhancement of platelet aggregation. These results suggest that chronic, but not acute, psychological stress enhances agonist-stimulated platelet aggregation independently of [Ca2+]i increase, and the enhancement may be mediated by stress hormones secreted from the adrenal glands.