Inhibition Of Adenosine Diphosphate-induced Platelet Aggregation Research Articles

Effect of Routine Clopidogrel Use on Bleeding Complications After Endobronchial Ultrasound-guided Fine Needle Aspiration.

Endobronchial ultrasonography has proven to be highly sensitive and specific in the diagnoses of patients with mediastinal and hilar adenopathy. Many of these patients are on a combination of clopidogrel (a compound that inhibits adenosine diphosphate-induced platelet aggregation) and aspirin due to neurological and/or cardiac-related comorbidities, and stopping anticoagulation may place these patients at high risk for potential complications. Our group has previously showed that thoracentesis with an 8-french catheter is safe in patients receiving clopidogrel and aspirin with low risk of complications. In this manuscript, we report the outcomes of the largest prospective multicenter series of patients undergoing endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) while receiving clopidogrel. Patients presenting to our institutions with mediastinal/hilar adenopathy, requiring EBUS-TBNA, and actively taking clopidogrel and aspirin were included in the study. If the medication could be held for 5 to 7 days before the procedure, the patient was excluded. EBUS-TBNA was performed by an interventional pulmonology faculty on a total of 42 patients. All patients received total intravenous anesthesia, and a total of 92 nodes were sampled. First, 3 passes were performed with a 22-G needle. If no complications were encountered, we followed with additional 3 passes with a 21 G. Rapid onsite evaluation was performed in all patients. Bleeding at the puncture site was considered significant if it required cold saline, topical sympathomimetic, or balloon tamponade for hemostasis. Bleeding was considered nonsignificant if no interventions were required to achieve hemostasis. We were able to perform all procedures successfully using both the 21 and 22-G needles. One patient required 30 mL cold saline installation to accomplish hemostasis with the 21 and 22-G needles. Our yield was comparable with the current literature. No statistically significant complications occurred during the procedure. All patients were contacted within 24 hours, and none reported bloody sputum. We suggest that EBUS-TBNA, using 22 and 21-G needles, is safe with high yields in patients with mediastinal/hilar adenopathy, actively taking clopidogrel and aspirin, and are at high risk for thrombotic complications if the medication is discontinued.

Study on antiplatelet effect of a new thiophenopyridine platelets P2Y12 receptor antagonist DV-127

P2Y12 receptor antagonists are a class of drugs that act on platelet P2Y12 receptors and inhibit adenosine diphosphate-induced platelet aggregation. As a thienopyridine antiplatelet agent which is approved by the US Food and Drug Administration for the treatment of cardiovascular diseases, currently, clopidogrel was once considered to be the most safe and effective antiplatelet drug in the P2Y12 receptor antagonists, however, it has become increasingly clear that clopidogrel does not satisfactorily inhibit the platelets of approximately one-third of patients. This is in part due to clopidogrel is a prodrug and reliance on multiple cytochrome P450 enzymes for conversion into its active metabolite. Prasugrel and ticagrelor reduces the risk of adverse cardiovascular events compared to clopidogrel in acute coronary syndromes patients, however, the cardiovascular benefit of both drugs is counter-balanced by increased rates of spontaneous bleeding. Unlike clopidogrel, which is a prodrug, cangrelor is an active drug not requiring metabolic conversion, despite fewer bleeding events during cardiac surgery, cangrelor carries the risk of potential autoimmune reactions manifesting as breathlessness.DV-127 was synthesized by using three generations of thienopyridine P2Y12 receptor antagonists as research models, using high resolution mass spectrometry, selective deuteration, and 2,7-position replacement groups technologies in order to maximize cardiovascular benefit while minimizing adverse effects on hemostasis. Our results show that although the dose of DV-127 is greatly reduced, it can achieve similar anticoagulant and antiplatelet effects as clopidogrel, and DV-127 can more strongly inhibit the release of α-granules even though the inhibitory effect of dense granules is similar to clopidogrel.

Identification of active ingredients mediating anti-platelet aggregation effects of BuyangHuanwu decoction using a platelet binding assay, solid phase extraction, and HPLC-MS/MS

BuyangHuanwu decoction (BHD) is widely used as a traditional herbal medicine because of its antithrombotic effect, which is attributed to the inhibition of platelet aggregation; however, its active compounds remain unknown. In this study, we developed a method involving platelet binding, solid-phase extraction, and HPLC-MS/MS for screening BHD compounds with potential anti-platelet aggregation properties. Five compounds showing platelet binding affinity were identified as 6-hydroxykaempferol-di-O-glucoside, paeoniflorin, calycosin-7-O-β-d-glucoside, galloylpaeoniflorin, and formononetin-7-O-β-d-glucoside. The results of anti-platelet aggregation experiments in vitro confirmed that these compounds inhibited adenosine diphosphate-induced platelet aggregation. Our results suggest that a platelet binding assay combined with solid-phase extraction and HPLC-MS/MS is an effective method for screening anti-platelet aggregation agents in traditional Chinese medicines.

Aqueous Extract of Tournefortia sarmentosa Stem Inhibits ADP-induced Platelet Aggregation

Tournefortia sarmentosa is a Chinese herbal medicine used as an antioxidant, detoxicant, and anti-inflammatory agent. The present study evaluated the effect of T. sarmentosa on adenosine diphosphate-induced platelet aggregation. Our results showed that aqueous stem extract of T. sarmentosa inhibited adenosine diphosphate-induced platelet aggregation. In addition, we found components in T. sarmentosa, including caffeic acid, rosmarinic acid, salvianolic acid, play important roles in mediating adenosine diphosphate-induced platelet aggregation suppression. The stem extract of T. sarmentosa inhibited the adenosine diphosphate receptor P2Y12-mediated cyclic AMP production. Caffeic acid inhibited P2Y1-induced calcium influx. Furthermore, treatment of platelets with T. sarmentosa, or the components in stem extract of T. sarmentosa, suppressed adenosine diphosphate-induced release of thromboxane A2 and arachidonic acid and surface PAC-1 expression. These data demonstrate the aqueous stem extract of T. sarmentosa significantly suppressed platelet aggregation through P2Y1 and P2Y12 receptor signal pathways. The antiaggregation properties found in the stem extract of T. sarmentosa might help to prevent cardiovascular disease.

Purification and characterization of a platelet aggregation inhibitor and anticoagulant Cc 5_NTase, CD 73-like, from Cerastes cerastes venom.

The present study is the first attempt to report the characterization of a nucleotidase from Cerastes cerastes venom. A 70kDa 5'-nucleotidase (Cc-5'NTase) was purified to homogeneity. The amino acid sequence of Cc-5'NTase displayed high homology with many nucleotidases. Its activity was optimal at pH 7 with a specific hydrolytic activity toward mono-, di-, and triphosphate adenylated nucleotides. Cc-5'NTase preferentially hydrolyzed ADP and obeyed Michaelis-Menten kinetics. Among the metals and inhibitors tested, Ni2+ and Mg2+ completely potentiated enzyme activity, whereas EGTA, PMSF, iodoacetamide, vanillic acid, vanillyl mandelic acid, and 1,10-phenanthroline partially abolished its activity. Cc-5'NTase was not lethal for mice at 5mg/kg and exhibited in vivo anticoagulant effect. It also dose-dependently inhibited adenosine diphosphate-induced platelet aggregation by converting adenosine diphosphate to adenosine and prohibited arachidonic acid-induced aggregation but was not effective on fibrinogen-induced aggregation. Cc-5'NTase could be a good tool as pharmacological molecule in thrombosis diagnostic and/or therapy.

The potential of anthocyanin-rich Queen Garnet plum juice supplementation in alleviating thrombotic risk under induced oxidative stress conditions

Increased oxidant production in humans induces a number of thrombotic consequences; including platelet hyperactivity/aggregability, which could be countered through specifically developed functional foods. We sought to determine the antithrombotic properties of anthocyanin-rich Queen Garnet plum juice (QGPJ) supplementation with and without exercise-induced oxidative stress. Thirteen healthy participants were investigated in a randomised, double-blind, placebo-controlled, cross-over trial. Participants consumed 200 mL/day of QGPJ and placebo juice for 28-days, with treatments separated by a two-week wash-out period. Blood samples were collected at baseline and after 1 h of exercise (70% peak-O2 uptake) both before and after oral supplementation and evaluated for platelet function and haemostatic activity. QGPJ supplementation inhibited adenosine diphosphate-induced platelet aggregation both without and under exercise-induced oxidative stress by 10.7% (P < 0.01) and 12.7% (P < 0.001) respectively; arachidonic acid-induced aggregation under oxidative stress by 28.8% (P < 0.05); reduced platelet activation-dependant P-selectin expression by 32.9% (P < 0.01) and 38.7% (P < 0.001) both without and under oxidative stress respectively; and exhibited favourable effects on coagulation parameters both with and without oxidative stress. The anti-thrombotic activity exhibited by anthocyanin-rich QGPJ suggests a potential for cardiovascular disease risk reduction and may be considered as complementary anti-platelet nutritional therapy in pro-thrombotic population.

Effect of Routine Clopidogrel Use on Bleeding Complications After Ultrasound-guided Thoracentesis

Thoracentesis is one of the most commonly performed medical procedures with an excellent safety profile. Clopidogrel (a compound that inhibits adenosine diphosphate-induced platelet aggregation) is often prescribed for primary or secondary prevention of cardiovascular disease and has been associated with bleeding complications in patients undergoing surgical procedures. The purpose of this study was to assess the safety of ultrasound (US)-guided thoracentesis in patients receiving clopidogrel therapy. Data were collected on 30 consecutive patients taking clopidogrel without other known underlying coagulation problems. These patients underwent 45 US-guided thoracenteses over 26 months. Clopidogrel was not discontinued before the thoracentesis in patients presenting with symptomatic pleural effusion. Thoracenteses were performed in these patients and the incidence of bleeding and other complications among patients was reported. Between June 2009 and August 2011, there were 30 consecutive patients on clopidogrel at the time of thoracenteses. These patients presented with respiratory distress because of pleural effusion and underwent a total of 45 thoracenteses. There was no significant bleeding or other complications in this patient population. No patient required transfusion after the procedure. Patients who are receiving clopidogrel and present with symptomatic pleural effusion can safely undergo US-guided thoracentesis without interrupting clopidogrel before the procedure. Larger studies are required to confirm these results.

Detection of Thromboembolism with 99mTc-labeled F(ab)2 Fragment of Anti-glycoprotein IIIa Chimeric Monoclonal Antibody in Beagle Canines

IntroductionRapid and timely diagnosis of pulmonary embolism (PE) and deep venous thrombosis (DVT) is important to improve patient outcome. The goal of this study was using 99mTc-chSZ21-F(ab)2, F(ab)2 fragment of anti-glycoprotein IIIa chimeric monoclonal antibody, to image experimental thromboembolism (DVT and PE) in dogs. Materials and MethodsFlow cytometry assay and adenosine diphosphate (ADP) stimulated platelet aggregation was performed to determine the specificity and affinity of chSZ21-F(ab)2 to the GPIIb/IIIa receptor on human or canine platelets. Both PE and DVT were induced in 12 beagle canines by catheter under X-ray direction. After 99mTc-chSZ21-F(ab)2 injection,animals were imaged for up to 3hours then heparinized and sacrificed. ResultsSpecific binding of chSZ21-F (ab)2 to GPIIb/IIIa on human or canine platelets was verified by flow cytometry assay. chSZ21-F (ab)2 inhibited ADP induced platelet aggregation with a dose-dependent manner, the concentration required to inhibit 50% (IC50) of platelet aggregation was 11.6±7.9nM and 24.9±18.8nM for human and canine, respectively. In vivo, focal uptake was observed in planar images as early as 30min (DVT) and 60min (PE), and became clearer within 3hours after injection. Lesion-to-background ratio averaged 12.8 (PE-to-lung), 7.2 (DVT-to-blood), and 117.0(DVT-to-muscle), respectively. ConclusionsThese results suggested that 99mTc-chSZ21-F(ab)2 with high DVT and PE uptake is a promising agent for imaging vascular thrombosis.

Pharmacokinetics and Tolerability of Single and Multiple Doses of Ticagrelor in Healthy Chinese Subjects

Ticagrelor (Brilinta™) is an antithrombotic agent that reversibly binds to P2Y(12) receptors and inhibits adenosine diphosphate-induced platelet aggregation. Ticagrelor has undergone evaluation in the phase III PLATO trial, which enrolled 18 624 patients with acute coronary syndromes (ACS) from 43 countries, and 6% of patients were Asian. Subsequently, ticagrelor has now been approved in more than 40 countries for the prevention of atherothrombotic events in adult patients with ACS. Gene polymorphisms in drug-metabolizing enzymes may vary with ethnicity, and can alter drug exposure, potentially impacting drug efficacy and/or tolerability. The objectives of this study were to assess the pharmacokinetic parameters of ticagrelor and its active metabolite AR-C124910XX, and the safety and tolerability of ticagrelor in healthy Chinese subjects, following single and multiple oral doses of ticagrelor. This trial was an open-label, sequential, two-cohort, single-centre study investigating 90 mg and 180 mg doses of ticagrelor in healthy Chinese subjects. On day 1, 12 subjects received a single oral dose of ticagrelor 90 mg. Following a 2-day washout period, ticagrelor was administered twice daily (90 mg twice daily) on days 4-9 and as a single dose on day 10. After completion of this phase, additional subjects (n = 14) were recruited into the ticagrelor 180 mg group, and received ticagrelor 180 mg under the same dosing schedule. On days 1 and 10 of both dosing schedules, blood samples for pharmacokinetic analyses were collected for 72 hours. Following single and multiple doses at both dose levels, ticagrelor was rapidly absorbed (median time [t(max)] to reach maximum plasma concentration [C(max)] 2 hours) with a mean elimination half-life (t(1/2;)) of 10.9-14.9 hours. AR-C124910XX was rapidly formed (median t(max) 2.0-3.0 hours; mean t(1/2;) 9.1-11.9 hours). Steady-state concentrations of ticagrelor and AR-C124910XX were rapidly established with both dosing regimens. Both parent and metabolite exhibited linear and predictable pharmacokinetics with single and multiple dosing as mean minimum plasma concentration (C(min)), C(max) and area under the plasma concentration-time curve from time zero to infinity (AUC(∞)) were approximately 2-fold higher with ticagrelor 180 mg (e.g. multiple-dosing, geometric mean [% coefficient of variation]: C(max) 1973 [27] and AUC(∞) 18 035 [46]) versus 90 mg (e.g. multiple dosing: C(max) 915 [32] and AUC(∞) 7168 [35]) dosing regimens. Overall, ticagrelor was generally well tolerated in healthy Chinese subjects. Two subjects discontinued in the ticagrelor 90 mg group due to elevated serum levels of ALT and AST. Mild ticagrelor-associated adverse events were seen: bleeding events (90 mg: epistaxis n = 1; 180 mg: gingival bleeding n = 1) and dyspnoea (180 mg: n = 3). In healthy Chinese subjects, ticagrelor and AR-C124910XX pharmacokinetics were linear and predictable. Although ticagrelor and AR-C124910XX exposure at steady state were found to be slightly higher in Chinese subjects, these results were broadly similar to previous data in Caucasian subjects. Overall, ticagrelor was well tolerated in healthy Chinese subjects. ClinicalTrials.gov identifier: NCT00721448.

Boosting platelet inhibition in poor responder to aspirin and clopidogrel undergoing percutaneous coronary intervention: role of tirofiban

Nowadays, aspirin (acetylsalicylic acid) and clopidogrel form the cornerstone in prevention of cardiovascular events and their clinical effectiveness has been well established. The thienopyridine clopidogrel is a prodrug that, after hepatic metabolization, strongly inhibits adenosine diphosphate-induced platelet aggregation. Aspirin is a non-steroidal anti-inflammatory drug that exerts its anti-platelet action through the irreversible acetylation of platelet cyclooxygenase (COX)-1, blocking thromboxane A2 production. However, despite dual-antiplatelet therapy, some patients still develop recurrent cardiovascular ischemic events. Many studies have clearly showed that a marked variability exists in the responsiveness to aspirin and clopidogrel, being the poor responder patients at higher risk of short (peri-procedural) and long-term ischemic complications. In particular, these patients showed a major recurrence of myocardial infarction and, after stent implantation, of stent thrombosis. The mechanisms of aspirin and clopidogrel poor response are numerous and not fully elucidated, and are likely multifactorial (eg, genetic polymorphisms, elevated baseline platelet reactivity, drug interaction). How to improve the short- and long-term outcome of these patients is currently unknown. Recently published and ongoing clinical trials are evaluating different strategies for the acute and chronic treatments (eg, reload of clopidogrel, double clopidogrel maintenance dose, switching to prasugrel). In this paper, we reviewed all available evidence on aspirin and clopidogrel resistance and focused our attention on tirofiban, a glycoprotein IIb/IIIa inhibitor that may be used to obtain a better platelet inhibition in poor responder patients during the acute phase and in particular during percutaneous coronary intervention.

Selective activation of the prostaglandin E2 receptor subtype EP2 or EP4 leads to inhibition of platelet aggregation

The effect of selective activation of platelet prostaglandin (PG) E2 receptor subtype EP2 or EP4 on platelet aggregation remains to be determined. In platelets prepared from wild-type mice (WT platelets), high concentrations of PGE2 inhibited platelet aggregation induced by U-46619, a thromboxane receptor agonist. However, there was no significant change in the inhibitory effect of PGE2 on platelets lacking EP2 (EP2-/- platelets) and EP4 (EP4-/- platelets) compared with the inhibitory effect on WT platelets. On the other hand, AE1-259 and AE1-329, agonists for EP2 and EP4, respectively, potently inhibited U-46619 -induced aggregation with respective IC50 values of 590 ± 14 and 100 ± 4.9 nM in WT platelets, while the inhibition was significantly blunted in EP2-/- and EP4-/- platelets. In human platelets, AE1-259 and AE1-329 inhibited U-46619-induced aggregation with respective IC50 values of 640 ± 16 and 2.3 ± 0.3 nM. Notably, the inhibitory potency of AE1-329 in human platelets was much higher than that in murine platelets, while such a difference was not observed in the inhibitory potency of AE1-259. AE1-329 also inhibited adenosine diphosphate-induced platelet aggregation, and the inhibition was almost completely blocked by AE3-208, an EP4 antagonist. In addition, AE1-329 increased intracellular cAMP concentrations in a concentration- and EP4-dependent manner in human platelets. These results indicate that selective activation of EP2 or EP4 can inhibit platelet aggregation and that EP4 agonists are particularly promising as novel anti-platelet agents.

Advances in antiplatelet therapy: overview of new P2Y12 receptor antagonists in development

Platelets play an important role in thrombus formation. A number of new antiplatelet agents currently in development are anticipated to improve clinical outcomes and safety benefits in patients with acute coronary syndrome (ACS). This manuscript reviews the pharmacology and clinical development of three of these agents: prasugrel, cangrelor, and AZD6140. Prasugrel, a third-generation, oral thienopyridine, has been shown to be superior to clopidogrel, the current gold standard, in preventing ischaemic events in patients with ACS undergoing percutaneous coronary intervention (PCI), although the bleeding rate was higher. Cangrelor, a chemical analogue of adenosine triphosphate, is a potent direct platelet P2Y12 antagonist. In development as an intravenous agent, cangrelor is currently being evaluated in two phase III studies in patients requiring PCI. AZD6140 is the first of a novel new class of antiplatelet agents that inhibits adenosine diphosphate-induced platelet aggregation at the level of the P2Y12 receptor. AZD6140 was shown to have a good safety profile in phase II studies and is currently being studied in a phase III trial in patients with ACS.

Variability of Response to Clopidogrel

Clopidogrel has been shown to inhibit adenosine diphosphate-induced platelet aggregation and has been demonstrated to be effective in reducing the risk of arterial thrombotic events in several large clinical studies. However, the clinical benefit could be attenuated by the variability of response to the antiplatelet effects of clopidogrel in as many as 30% of patients. Multiple mechanisms likely contribute to clopidogrel variability of response, including inappropriate dosing or underdosing of clopidogrel, drug-drug interactions, and genetic polymorphisms. The best laboratory procedure to screen for possible clopidogrel variability of response remains to be determined.

Characterization of novel dihydrothienopyridinium and thienopyridinium metabolites of ticlopidine in vitro: role of peroxidases, cytochromes p450, and monoamine oxidases.

Ticlopidine is an agent that inhibits adenosine diphosphate-induced platelet aggregation. Metabolic studies with ticlopidine have indicated that the principal routes of metabolism are N-dealkylation, N-oxidation, and oxidation of the thiophene ring. However, ticlopidine shares some structural features that are similar to those of cyclic tertiary amines such as 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and tetrahydroisoquinolines, which are converted to neurotoxic pyridinium metabolites, via the iminium (dihydropyridinium) species. The current in vitro studies examined the potential of ticlopidine to undergo a similar conversion by cytochrome P450 (P450), peroxidases, and monoamine oxidase (MAO). The results from these studies have suggested that ticlopidine undergoes an overall 4-electron oxidation to the novel thienopyridinium metabolite (M6) via the intermediate 2-electron oxidation product, the thienodihydropyridinium metabolite (M5) by P450, horseradish peroxidase, and myeloperoxidase and, to a lesser extent, by MAO. The structures of these metabolites were characterized by liquid chromatography (LC)-tandem mass spectrometry and LC-NMR. Qualitative studies with baculovirus-expressed P450s revealed the involvement of P450 3A4 in this conversion. Interestingly, M5 was the primary metabolite in the peroxidase-mediated reactions and was quite stable to air oxidation or disproportionation. It was less electrophilic and did not form cyanide, glutathione, or N-acetylcysteine adducts. On the other hand, M6 was the major metabolite in P450-catalyzed oxidation of ticlopidine. The results from this study have revealed that in addition to metabolism of the thiophene ring of ticlopidine, the tetrahydropyridine moiety of the compound is susceptible to a 2-electron and a 4-electron oxidation like other cyclic tertiary amines.

Safety profile and tolerability of intravenous AR-C69931MX, a new antiplatelet drug, in unstable angina pectoris and non—Q-wave myocardial infarction

Background: Thrombin generation and platelet aggregation in the disrupted atherosclerotic plaque are the major reasons for thrombus formation associated with acute coronary events. AR-C69931MX is a new agent that inhibits adenosine diphosphate—induced platelet aggregation by antagonism of the P2T purinoceptor.Objective: This study assessed the safety profile, tolerability, and plasma concentrations at steady state of intravenous AR-C69931MX in patients with unstable angina pectoris or non—Q-wave myocardial infarction (MI).Methods: This was a Phase II, multicenter, double-blind, randomized, placebo-controlled trial. Patients with unstable angina or non—Q-wave MI were randomized to a 72-hour infusion of AR-C69931MX or placebo as adjunctive therapy to aspirin and low-molecular-weight heparin. Other treatment was at the discretion of the local investigator. Outcomes were assessed at 30 days.Results: Ninety-four patients were randomized and 91 received treatment (45 AR-C69931MX, 46 placebo). Plasma concentrations of AR-C69931MX were within the expected range, there were no signs of accumulation, and interindividual variability in clearance was low. Four patients receiving AR-C69931MX discontinued treatment due to minor bleeding events, and 5 patients receiving placebo discontinued treatment due to other adverse events or deterioration in their condition. No serious bleeding events were seen during treatment. The incidence of ≥1 episode of minor bleeding was slightly higher in patients receiving AR-C69931MX compared with those receiving placebo (38% vs 26%, respectively). The drug was well tolerated hemodynamically, and there were no significant changes in other laboratory values between groups.Conclusions: As adjunctive therapy to aspirin and low-molecular-weight heparin in patients with unstable angina or non—Q-wave MI, intravenous AR-C69931MX was well tolerated, with no difference in the incidence of serious adverse events compared with placebo.

The Effects of Aprotinin on Platelets In Vitro Using Whole Blood Flow Cytometry

We sought to evaluate the effects of aprotinin on the number and function of the platelet glycoprotein (GP) IIb-IIIa receptor and on the expression of P-selectin in vitro in order to gain insight into the potential mechanisms involved in the platelet-protective action of aprotinin during cardiopulmonary bypass. Aprotinin at 50 to 200 kallikrein inhibiting units/mL decreased the expression of activated GP IIb-IIIa complex in response to adenosine diphosphate or thrombin receptor activator peptide 6 in a dose-dependent manner in both citrated and heparinized whole blood experiments. Aprotinin inhibited adenosine diphosphate-induced platelet aggregation, but it exhibited no effect on the expression of GP IIIa and P-selectin. These results indicate that aprotinin interferes with the platelet fibrinogen receptor function during pharmacological activation. Reduced aggregability and platelet adhesion to fibrinogen adsorbed to synthetic surfaces in the presence of aprotinin may prevent platelet consumption during clinical cardiopulmonary bypass. This in vitro study demonstrates that aprotinin decreases the agonist-induced expression of activated GP IIb-IIIa receptors that play a major role in platelet aggregation and adhesion to biomaterial surfaces. This in vitro study demonstrates that aprotinin decreases the agonist-induced expression of activated glycoprotein IIb-IIIa receptors that play a major role in platelet aggregation and adhesion to biomaterial surfaces.

Sevoflurane Does Not Inhibit Human Platelet Aggregation Induced by Thrombin

Sevoflurane reportedly inhibits adenosine diphosphate-induced platelet aggregation by suppressing thromboxane A2 formation. The increase in intracellular calcium concentration that fosters platelet aggregation, however, is also induced by other cell signaling pathways, such as activation of the production of inositol 1,4,5-triphosphate by thrombin. The current study aimed to clarify the net influence of sevoflurane on thrombin-induced platelet aggregation. Washed platelets were stimulated by thrombin after incubation with 0.5, 1.0, or 1.5 mM sevoflurane, halothane, or isoflurane. Aggregation curves were measured by an aggregometer. Intracellular calcium concentration was measured fluorometrically using fura-2. Calcium mobilization via plasma membrane calcium channels and the dense tubular system was assessed differentially. Intracellular inositol 1,4,5-triphosphate was measured by radioimmunoassay. Halothane significantly suppressed aggregation ratios at 5 min compared with those in controls (89 +/- 7%) to 71 +/- 10% (1.0 mM) and 60 +/- 11% (1.5 mM) and the increase in intracellular calcium concentration (controls, 821 +/- 95 nM vs. 440 +/- 124 nM [1.0 mM] or 410 +/- 74 nM [1.5 mM]). Halothane also significantly inhibited release of calcium from the dense tubular system (controls, 220 +/- 48 nM vs. 142 +/- 31 nM [1.0 mM]). Neither sevoflurane nor isoflurane produced a net change in aggregation ratios, intracellular calcium concentration, or calcium mobilization. Halothane (1 mM) significantly suppressed inositol 1,4,5-triphosphate concentrations, whereas neither 1 mM isoflurane nor 1 mM sevoflurane had any effect. Although sevoflurane has been reported to inhibit human platelet aggregation induced by weak agonists such as adenosine diphosphate, it does not inhibit human platelet aggregation induced by strong agonists such as thrombin.

Effects of Clopidogrel on Platelet Activation and Coagulation of Non-Anticoagulated Rat Blood under High Shear Stress

Clopidogrel is a new thienopyridine derivative similar to ticlopidine, which inhibits adenosine diphosphate-induced platelet aggregation. The in vitro effects of clopidogrel on shear-induced platelet activation and coagulation were assessed after oral administration to rats, by subjecting non-anticoagulated blood to haemostatometry. Clopidogrel significantly inhibited shear-induced platelet activation and coagulation 2 h after administration at doses of 7.5 and 15 mg/kg. Both ticlopidine (200 mg/kg) and aspirin (200 mg/kg) inhibited shear-induced platelet activation, but not coagulation. The peak inhibition of plaetelet activation by clopidogrel occurred 2 h after oral administration, but significant inhibition persisted even after 24 h. These results suggest that clopidogrel could be a more potent antithrombotic agent than ticlopidine or aspirin, and also that ADP plays an important role in shear-induced platelet activation.

Temporary and partial inhibition of platelets by SM-20302 prevents coronary artery thrombosis in a chronic canine model

We proposed that temporary and partial platelet inhibition by a GPIIb/IIIa receptor antagonist, SM-20302, would provide sustained antithrombotic efficacy in a chronic model of coronary artery thrombosis. Instrumented, conscious dogs received vehicle (Group I, n=7), low dose SM-20302 (30 μg/kg bolus+1 μg/kg/min infusion for 6 h) (Group II, n=7), or high dose SM-20302 (100 μg/kg bolus+1 μg/kg/min infusion for 6 h) (Group III, n=7). Thrombosis was initiated by electrolytic injury to the circumflex coronary artery. Coronary blood flow was monitored for 6 h on day 1 and days 2–6. Platelet aggregation was performed in platelet-rich plasma prepared from citrated or heparinized blood. At 6 h, both doses of SM-20302 inhibited adenosine diphosphate-induced platelet aggregation completely (>90%) in citrated platelet-rich plasma, but incompletely (57–59%) in heparinized platelet-rich plasma. Platelet reactivity returned to baseline values at 24 h. Control animals developed thrombotic occlusion on Day 1. Both doses of SM-20302 maintained vessel patency during the infusion period (Day 1) and the subsequent 5 days. Myocardial infarct size and mortality in the drug treated groups were reduced compared to the vehicle group. Thus, temporary inhibition of platelet reactivity by SM-20302 is associated with sustained prevention of primary thrombus formation, and reduction in infarct size and mortality.

The in vitro pharmacological profile of SK&F 106760, a novel GPIIB/IIIA antagonist

The properties of SK&F 106760 [N α-acetyl-cyclo(S,S)-cysteinyl-N α-methylarginylglycyl-aspartyl-penicillamine-amide] as a GPIIb/IIIa antagonist have been studied in vitro and compared with those of the parent molecule, Ac-RGDS-NH 2. Ac-RGDS-NH 2 inhibited biotinylated fibrinogen binding to purified human GPIIb/IIIa immobilized on plastic microtitre plates with a Ki of 530 ± 73 nM. In canine platelet rich plasma Ac-RGDS-NH 2 produced a concentration related inhibition of adenosine diphosphate-induced platelet aggregation following preincubation for 3 min with an IC50 of 91 ± 1 μM. However, incubation in platelet rich plasma for 3 hr abolished the activity of Ac-RGDS-NH 2. SK&F 106760 inhibited biotinylated fibrinogen binding to purified human GPIIb/IIIa immobilized on plastic microtitre plates with a K i of 477 ± 57 pM. SK&F 106760 inhibited adenosine diphosphate-induced platelet aggregation in human platelet rich plasma with an IC50 of 230 ± 60 nM but did not inhibit the von Willebrand Factor receptor (GPIb/IX)-mediated platelet agglutination produced by ristocetin. In canine platelet rich plasma SK&F 106760 inhibited aggregation produced by adenosine diphosphate, collagen and epinephrine/U-46619 with IC 50 values of 355 ± 35, 260 ± 20, and 490 ± 90 nM, respectively and in gel filtered platelets inhibited thrombin-mediated aggregation with an IC50 of 188 ± 10 nM. Preincubation of SK&F 106760 in platelet rich plasma for three hours had no significant effect on its ability to inhibit adenosine diphosphate-induced platelet aggregation. SK&F 106760 produced insurmountable inhibition of adenosine diphosphate-induced platelet aggregation in the presence of constant fibrinogen concentrations, but produced competitive inhibition of the concentration-response curve to fibrinogen in adenosine diphosphate-activated platelets with a Kb of 8.0 ± 1.0 nM. Thus, SK&F 106760 is a potent, stable competitive GPIIb/IIIa antagonist with no detectable activity at the von Willebrand Factor receptor (GPIb/IX).