Platelet aggregation in patients with various forms of left ventricular hypertrophy and its changes at long-term follow-up and treatment

AIM: To study platelet aggregation (PA) in patients with left ventricular hypertrophy (LVH) in essential hypertension (EH) and hypertrophic cardiomyopathy (HTCM), and to assess aspirin treatment effects. MATERIAL AND METHODS: A general clinical examination, echocardiography, 24-h monitoring of ECG and arterial pressure, bicycle exercise, test for platelet aggregation, routine blood biochemical tests were performed in 30 males with EH and LVH, 30 males with HTCM and 10 healthy controls. RESULTS: It was found that the patients had high platelet sensitivity to aggregation inductors as well as high spontaneous aggregation. LVH severity correlated positively with activation of spontaneous and induced platelet aggregation. This dependence is unrelated to arterial pressure or the disease duration. Patients with episodes of silent myocardial ischemia had a more pronounced rise in spontaneous and induced platelet aggregation than those without silent ischemia. A long-term administration of aspirin significantly lowered spontaneous and induced platelet aggregation, depression of ST segment. The above changes occurred both in HTCM and in hypertensive patients with LVH. CONCLUSION: Platelet aggregation correlates with severity of LVH and episodes of silent myocardial ischemia both in patients with hypertension, LVH and HTCM.     

Von Willebrand''s disease with spontaneous platelet aggregation induced by an abnormal plasma von Willebrand factor.

We have investigated and characterized the abnormalities in four unrelated patients with von Willebrand's disease (vWd) who have (a) enhanced ristocetin-induced platelet aggregation (RIPA) at low ristocetin concentrations, (b) absence of the largest plasma von Willebrand factor (vWf) multimers, and (c) thrombocytopenia. The platelet-rich plasma of these patients aggregates spontaneously without the addition of any agonists. When isolated normal platelets are resuspended in patient plasma spontaneous aggregation occurs; however, the patients' plasmas did not induce platelet aggregation of normal washed formalinized platelets. When the patients' platelets are suspended in normal plasma, spontaneous aggregation is not observed. The spontaneous platelet aggregation (SPA) is associated with dense granule secretion as measured by ATP release and alpha granule release as measured by beta-thromboglobulin and platelet factor 4 release. The SPA is totally inhibited by 5 mM EDTA, prostaglandin I2, and dibutryl cyclic AMP, while it is only partially inhibited by 1 mM EDTA, acetylsalicylic acid, or apyrase. A monoclonal antibody directed against glycoprotein Ib (GPIb) and/or a monoclonal antibody against the glycoprotein IIb/IIIa (GPIIb/IIIa) complex totally inhibits the SPA. The vWf was isolated from the plasma of one of these patients. The purified vWf induced platelet aggregation of normal platelets resuspended in either normal or severe vWd plasma, but the vWf did not induce platelet aggregation of normal platelets resuspended in afibrinognemic plasma. Sialic acid and galactose quantification of the patient's vWf revealed approximately a 50% reduction compared with normal vWf. These studies indicate that a form of vWd exists, which is characterized by SPA that is induced by the abnormal plasma vWf. The SPA is dependent on the presence of plasma fibrinogen, and the availability of the GPIb and the GPIIb/IIIa complex. In this variant form of vWd the abnormal vWf causes enhanced RIPA, SPA, and thrombocytopenia.     

The pre-procedural platelet state predicts clinical recurrence after coronary angioplasty

Percutaneous transluminal coronary angioplasty is an established therapy for coronary artery disease, but restenosis still occurs at a rate of 25%-40%. The aim of this study was to investigate the acute effect of percutaneous transluminal coronary angioplasty on platelet function and the relationship between platelet function and clinical recurrence. Spontaneous platelet aggregation was assessed before and after successful coronary angioplasty in 155 patients (120 men, 35 women). Patients were followed for a mean time of 20 months; follow-up angiography was performed only in patients with clinical recurrence. In 122 of 155 patients (79%) a significant increase in spontaneous platelet aggregation was observed immediately after coronary angioplasty. Baseline spontaneous platelet aggregation in platelet-rich plasma was significantly lower in patients with clinical recurrence than in those without (P<0.05). Kaplan-Meier event-free survival estimate showed a significant difference in clinical recurrence between patients with spontaneous platelet aggregation in platelet-rich plasma below and above the first quintile (P<0.05) with a relative risk of 2.5. In conclusion. these results indicate that percutaneous transluminal coronary angioplasty enhances spontaneous platelet aggregation and that the platelet state before coronary angioplasty affects the risk of clinical recurrence after the procedure.     

Effects of the new glycopeptide antibiotic teicoplanin on platelet function and blood coagulation.

Teicoplanin, a new glycopeptide antibiotic, is structurally related to ristocetin, an antibiotic known to induce human platelet agglutination and, thus, thrombocytopenia and thromboembolic side effects. The aim of this study was to evaluate the effects of teicoplanin on platelet function in vitro and ex vivo and on blood coagulation ex vivo. In the in vitro studies, spontaneous platelet aggregation; platelet aggregation induced by ADP, collagen, and ristocetin; and the release of beta-thromboglobulin from platelets were assessed. Platelets from healthy subjects were incubated with teicoplanin at final concentrations of 100, 1,500, 5,000, and 10,000 micrograms/ml. The maximal achievable concentration with therapeutic doses is 100 micrograms/ml. When compared with saline, teicoplanin at concentrations of 100 and 1,500 micrograms/ml had no effect on platelet function, but at concentrations of 5,000 and 10,000 micrograms/ml, it induced greater spontaneous platelet aggregation (P less than 0.01) and inhibited platelet aggregation induced by ADP, collagen, and ristocetin (P less than 0.01). Teicoplanin at concentrations of 100, 1,500, and 5,000 micrograms/ml did not induce the release of beta-thromboglobulin, in contrast to teicoplanin at a concentration of 10,000 micrograms/ml and ristocetin at a concentration of 1.5 mg/ml (P less than 0.01). In the ex vivo studies, platelet count, bleeding time, plasma beta-thromboglobulin, platelet aggregation induced by ADP, ristocetin, and epinephrine, activated partial thromboplastin time, prothrombin time, thrombin clotting time, and serum fibrinogen degradation products were evaluated at days 0, 3, and 6 and at 72 h after the end of therapy. All subjects completed the study without evidence of side effects. When compared with the pretreatment values, none of the values from these assays showed a significant change at any time during and after treatment. We concluded that platelet function and blood coagulation are not affected by therapeutic concentrations of teicoplanin and that in vitro platelet function is affected only by concentrations of teicoplanin far in excess of those that are clinically achievable.     

Adenoviral vectors do not induce, inhibit, or potentiate human platelet aggregation.

Adenoviruses are commonly used as vectors in human clinical gene therapy trials. High doses of intravenous adenovirus vectors have been associated with development of thrombocytopenia of undetermined origin. Viral internalization requires the presence cell surface integrins, alpha(v)beta(3) or alpha(v)beta(5), that can blind ligands with a arginine-glycine-aspartic acid (RGD) sequence. This sequence is found in the adenovirus penton base. Platelets express the alpha(v)beta(3) integrin and other integrins that bind the RGD sequence of ligands such as fibrinogen, laminin, vitronectin, and von Willebrand factor (vWF). Platelet aggregation is mediated, in part, by the binding of the RGD sequence of fibrinogen to a platelet surface integrin, glycoprotein IIb/IIIa (GP IIb/IIIa). We investigated whether adenovirus particles could interfere with or potentiate agonist-induced platelet aggregation. Incubation of platelet-rich plasma with adenovirus under stirred conditions did not promote spontaneous aggregation. The addition of physiological platelet agonists, ADP, collagen, or epinephrine, induced platelet aggregation. However, the presence of adenovirus in a wide range of concentrations did not inhibit or potentiate agonist-induced aggregation. These results suggest that the adenovirus-associated thrombocytopenia observed in vivo is independent of a direct effect of the virus on platelet aggregation.     

具有异常结构的巨大血小板的自发性聚集

目的：研究1例具有异常结构巨大血小板的自发性聚集患者的病理和临床特征。方法：光学显微镜与电子显微镜观察血小板形态与结构。比浊法测定血小板聚集。流式细胞术检测血小板膜糖蛋白（GP）。PCR扩增与DNA序列分析确定基因异常。结果：患者的血小板体积巨大，胞膜粗厚，胞浆内颗粒增多而形态异常。血小板自发性聚集，阿司匹林或噻氯 氯匹啶无抑制作用。血小板膜GPⅠb、GPⅡb、GPⅢa和P选择素的表达正常。GPⅠbα、GPⅠbβ与GPⅨ基因分析正常。结论：本例患者的血小板结构和功能异常不同于文献报道的各种遗传性巨大血小板疾病，可能代表了一种新的先天性的血小板病。     

A new method to evaluate spontaneous platelet aggregation in type 2 diabetes by Cellfacts

BACKGROUND: The alterations in the functional activities of platelets in diabetes produce an increase of spontaneous platelet aggregation (SPA) and release of platelet-derived microparticles. Platelet-derived microparticles are shed from platelets during activation by high shear stress, collagen and certain agonists. Although the physiologic role of microparticles has been difficult to assess, the characterization of their biological activity is of interest in view of a possible role in hemostasis and coagulation and their reported involvement in thrombotic disease. METHODS: We propose a new, simple method to evaluate spontaneous platelet aggregation and release of platelet-derived microparticles by the Cellfacts analyser (Microbial System Limited (MSL), Coventry, England) that uses electrical sensing flow impedance determination to detect the size particles and the cells in a conductive fluid. Platelet-rich plasma (PRP) from type 2 diabetes was employed for this study. The importance of platelet-activating factor (PAF) on spontaneous platelet aggregation was evaluated and the effect of vitamin E and WEB 2086-BS, an antagonist of platelet-activating factor, was measured. RESULTS AND CONCLUSIONS: Data presented show that Cellfacts could be an easy and fast instrument to check the state of platelets in patients with alterations in the functionality of platelets, and to follow the effect of pharmacological therapy on spontaneous platelet aggregation and the release of platelet-derived microparticles.     

Rapid dissociation of platelet-rich fibrin clots in vitro by a combination of plasminogen activators and antiplatelet agents.

Thrombin promotes the formation of arterial thrombi by converting fibrinogen to fibrin and by causing platelets to aggregate. We have examined the combined effects of plasminogen activators and inhibitors of platelet aggregation on the lysis of platelet-rich fibrin clots formed by alpha-thrombin in citrated platelet-rich plasma. The extent of platelet aggregation and clot formation were measured by recording light transmission in an aggregometer. Immediately after the formation of platelet-rich fibrin clots, addition of 2,000 U/ml streptokinase or 50 micrograms/ml recombinant tissue-type plasminogen activator alone resulted in the degradation of polymerized fibrin and the release of trapped platelet aggregates without causing significant platelet deaggregation. Preincubation of the platelet-rich plasma with 20 microM indomethacin for 1 min before thrombin stimulation or simultaneous addition of prostaglandin E1 (10 microM) with the plasminogen activators after thrombin stimulation resulted in spontaneous platelet deaggregation. Because platelet aggregation is, in part, mediated by the binding of Arg-Gly-Asp-containing adhesive proteins to activated platelets, the effect of Arg-Gly-Asp peptides on platelet deaggregation was examined. By itself, Gly-Arg-Gly-Asp-Ser-Pro specifically caused dose- and time-dependent deaggregation of platelet aggregates formed by ADP or by thrombin in the presence of 1 mM Gly-Pro-Arg-Pro, but had no effect on the dissociation of thrombin-induced platelet-rich fibrin clots. In combination with streptokinase or recombinant tissue-type plasminogen activator, Gly-Arg-Gly-Asp-Ser-Pro enhanced the rate of lysis of platelet-rich fibrin clots. The control Gly-Arg-Gly-Glu-Ser-Pro peptide was completely ineffective.(ABSTRACT TRUNCATED AT 250 WORDS)     

Platelet aggregation in patients with hypertrophic cardiomyopathy

AIM: To examine correlation between platelet aggregation and the degree of left ventricular hypertrophy (LVH) in patients with hypertrophic cardiomyopathy (HC). MATERIAL AND METHODS: Routine clinical examination, ECG, echo-CG, measurement of platelet aggregation, routine biochemical blood tests were made in 45 HC males and 15 healthy controls. RESULTS: HC patients were characterized with high induced and spontaneous platelet aggregation which increased with left ventricular myocardial mass. In LVH, platelets were especially sensitive to the aggregation inductor U-46619, thromboxane A2 analogue. CONCLUSION: It is demonstrated that there is a positive correlation between platelet aggregation and the degree of LVH in HC patients.     

Formation of prostacyclin-sensitive platelet aggregates in human whole blood in vitro. Part I. The occurrence of the phenomenon in healthy male volunteers

When unstirred citrated blood of young males was left to stand at 21 degrees C, the number of free platelets decreased as measured with a whole blood platelet counter. This decrease indicated spontaneous platelet aggregation (SPA) since it was accompanied by the time-dependent increase in platelet micro-aggregates and plasma beta-thromboglobulin while lactate dehydrogenase level did not change significantly. Addition of PGI2 to whole blood increased free platelet count and decreased the number of platelet micro-aggregates. The de-aggregatory effect of PGI2 (measured as a percentage increase in free platelet number) was correlated with the initial amount of platelet micro-aggregates. Blood storage enhanced SPA and de-aggregatory effect of prostacyclin. Immediately after blood collection de-aggregatory effect of prostacyclin was absent. Our results favour an assumption that prostacyclin-sensitive platelet aggregates in blood of young volunteers are formed in vitro rather than in vivo.     

Exposure of fibrinogen receptors in human platelets by surface proteolysis with elastase.

Human platelets that were preincubated with porcine elastase aggregated spontaneously upon the addition of fibrinogen. Maximal aggregation to fibrinogen was observed with platelets pretreated with an elastase concentration of 111 micrograms/ml, and half-maximal aggregation occurred after treatment with 11 micrograms/ml elastase. Binding of radiolabeled fibrinogen to elastase-treated platelets was specific, saturable, and showed a single class of 48,400 +/- 9,697 fibrinogen-binding sites per platelet with a dissociation constant of 6.30 +/- 1.48 X 10(-7) M. ATP, apyrase, and the stimulators of platelet adenylate cyclase forskolin, prostaglandin E1, prostacyclin, and N6, 2''-O-dibutyryl cyclic AMP did not inhibit the fibrinogen-induced aggregation of elastase-treated platelets. EDTA completely blocked the initiation of aggregation and reversed the fibrinogen-induced aggregation of elastase-treated platelets. Monoclonal and polyclonal antibodies directed against glycoproteins (GP) IIb and IIIa completely blocked the fibrinogen-induced aggregation of elastase-treated platelets. Immunoprecipitates with these antibodies obtained from detergent extracts of surface-radiolabeled, intact, and elastase-treated platelets contained the glycoproteins IIb and IIIa. We conclude that surface proteolysis by low concentrations of elastase can expose fibrinogen-binding sites associated with GPIIb and GPIIIa on the platelet surface, resulting in spontaneous aggregation upon the addition of fibrinogen. These findings may be relevant to hemostatic changes observed in patients with increased levels of circulating elastase.     

Red cell aggregation and the echogenicity of whole blood.

To study the relationship between red cell aggregation and whole blood echogenicity, red cell aggregation was quantitated by a photometric method, whole blood echogenicity was quantitated by videodensitometry and sedimentation rate was quantitated by a modified Westergren method. Changes in red cell aggregation were produced by alterations in the hematocrit. The results showed that red cell aggregation increased in a linear fashion with increases in hematocrit. The sedimentation rate decreased in a linear manner with increases in hematocrit. Whole blood echogenicity showed a biphasic response, with an initial increase in echogenicity, peaking at hematocrits varying from 14-24% and decreasing thereafter. Over the physiologic range of hematocrits, an increase in the formation of red cell aggregates is associated with a decrease in the echogenicity of whole blood. Thus, red cell aggregates were not visible using our ultrasound equipment at physiologic hematocrits, and the echo contrast in blood under our experimental conditions at these hematocrits must represent either plasma spaces, platelet aggregates or possibly white cell aggregates. The association between spontaneous contrast and a propensity for thromboembolism imply that platelet aggregates are the most likely origin of in vivo echo contrast in flowing blood.     

Intravascular platelet aggregation and spontaneous contrast.

The purpose of this study was to determine why spontaneous contrast developed after general anesthesia in dogs. Twenty-seven dogs underwent echocardiography before and after pentobarbital or chloralose-urethane general anesthesia. The results showed that none of the 12 dogs receiving pentobarbital and 10 of 15 dogs receiving chloralose-urethane anesthesia developed contrast, in association with large platelet and platelet-neutrophil aggregates (P < 0.01); this effect could also be reproduced in vitro. The administration of adenosine diphosphate or antiplatelet antibody to nine dogs confirmed that intravascular platelet aggregation can cause ultrasonic contrast. The implications of these findings for patients with spontaneous contrast are discussed.     

Coagulation-physiological studies on the prevention of thromboembolism with dextran 60. I. The effect on platelet function

Platelet aggregation, spreading capacity, adhesiveness (methods according to BREDDIN) have been investigated in 3 groups of 20 patients each after gynecological surgery. Group A (control group) got only 4-hydroxycumarin (Sintrom), group B--in addition to that--intraoperative 500 ml Dextran 60 (Macrodex 6%), group C 500 ml Dextran 60 as well intraoperative as on the 1., 2., 4., 7. and 10. day after surgery. The present data show that depending on the duration of Dextran application, platelet spreading capacity and adhesiveness are inhibited differently as to duration and extent. There was, above all, a reduction and with that a normalisation of the postoperatively increased spontaneous platelet aggregation. It is possible to influence the phase of early postoperative embolism in a favorable way by applicating Dextran already intraoperatively. After that one can use further infusions of Dextran, a prophylaxis by Cumarins or other anticoagulants. Giving dextran only once during surgery one can prove this normalisation of the increased platelet aggregation up to 24 hours postoperatively. By further infusions of Dextran 60 on the 1., 2., 4., 7. and 10. postoperative day it is possible to keep the extent of aggregation on a significantly reduced level till the complete mobilisation of the patient and consequently attain a protective effect on a possible development of deep venous thrombosis of the legs.     

State of the microcirculatory component of hemostasis and microhemodynamics in patients with diabetic angiopathies

A total of 150 patients presenting with diabetes mellitus were examined. Patients with diabetic angiopathy demonstrated substantial impairment of platelet aggregation (secondary adhesion, spontaneous aggregation), of the functional activity of heparinocytes, lipid metabolism and microhemodynamics.     

糖尿病患者白细胞自发性活化的变化

为探讨糖尿病及其血管病变时白细胞自发性活化的变化，对30例糖尿病患者及23例正常人进行了白细胞自发活化率(SAR)、血浆胆固醇、纤维蛋白原浓度及血小板聚集功能的测定。结果显示糖尿病患者白细胞SAR(%)显著高于正常对照组(P＜0．01)；胆固醇、纤维蛋白原及血小板聚集率也较正常对照组增高。提示：白细胞自发性活化与糖尿病及其血管病变的发生发展有关；胆固醇、纤维蛋白原及血小板聚集与白细胞自发性活化有关。     

Adenosine-mediated inhibition of platelet aggregation by acadesine. A novel antithrombotic mechanism in vitro and in vivo.

Inhibition of platelet aggregation by acadesine was evaluated both in vitro and ex vivo in human whole blood using impedance aggregometry, as well as in vivo in a canine model of platelet-dependent cyclic coronary flow reductions. In vitro, incubation of acadesine in whole blood inhibited ADP-induced platelet aggregation by 50% at 240 +/- 60 microM. Inhibition of platelet aggregation was time dependent and was prevented by the adenosine kinase inhibitor, 5'-deoxy 5-iodotubercidin, which blocked conversion of acadesine to its 5'-monophosphate, ZMP, and by adenosine deaminase. Acadesine elevated platelet cAMP in whole blood, which was also prevented by adenosine deaminase. In contrast, acadesine had no effect on ADP-induced platelet aggregation or platelet cAMP levels in platelet-rich plasma, but inhibition of aggregation was restored when isolated erythrocytes were incubated with acadesine before reconstitution with platelet-rich plasma. Acadesine (100 mg/kg i.v.) administered to human subjects also inhibited platelet aggregation ex vivo in whole blood. In the canine Folts model of platelet thrombosis, acadesine (0.5 mg/kg per min, i.v.) abolished coronary flow reductions, and this activity was prevented by pretreatment with the adenosine receptor antagonist, 8-sulphophenyltheophylline. These results demonstrate that acadesine exhibits antiplatelet activity in vitro, ex vivo, and in vivo through an adenosine-dependent mechanism. Moreover, the in vitro studies indicate that inhibition of platelet aggregation requires the presence of erythrocytes and metabolism of acadesine to acadesine monophosphate (ZMP).     

Effect of filtration, storage and platelet suspension media on platelet indices

We describe a new approach for assessing filtration-induced changes in cellular indices of platelet concentrates at the beginning and the end of storage, using pairs of identical packs. The results revealed that post-filtered products did not store as well as their counterparts. Filtration did not induce any significant changes on aggregation as determined by spontaneous aggregation nor was there a disparity between leucocyte peroxidase/basophil count. We recommend filtration on day 2 which causes minimal loss of platelets and less change in the mean platelet volume.     

Effect of enalapril maleate on vascular endothelial function and platelet-endothelial interactions in patients with essential hypertension

AIM: Evaluation of endothelial function and platelet-endothelial interactions in patients with essential hypertension and dynamics of these changes in the course of treatment with enalapril maleate. MATERIALS AND METHODS: The study included 37 patients with essential hypertension and 22 normotensive volunteers. 17 of hypertensive patients received enalapril maleate (enap, KRKA) 5-20 mg/day during the period of 1.5 months. The complex of investigations included: measurement of total plasma cholesteroi, 12-lead ECG, echocardiography, high-resolution ultrasound investigation of brachio-cephalic arteries, evaluation of flow-mediated dilation, measurement of von Willebrand's factor, spontaneous and induced platelet aggregation. RESULTS: Patients with essential hypertension exhibited higher levels of von Willebrand's factor in plasma and degree of spontaneous and induced platelet aggregation as well as lower responses of vessel wall to hemodynamic stimuli compared to normotensive healthy individuals. There was a strong correlation between endothelial function markers and CAD risk factors, elevation of platelet activity. Treatment with enalapril maleate led to a statistically significant decrease of von Willebrand's factor in plasma and ex vivo platelet aggregation whereas flow-mediated dilatation increased. Values of endothelial function markers and platelet activity approached to those of normotensive subjects and these changes were accompanied by a decrease of ECG signs of left ventricular hypertrophy. CONCLUSION: Patients with essential hypertension were found to have compromised endothelial function. However, the degree of endothelial dysfunction depends not on hemodynamic parameters, but on the cumulative effect of CAD risk factors. Treatment with enalapril maleate may lead to normalisation of endothelial function and decrease of platelet activity.     

Vitamin E. An inhibitor of the platelet release reaction.

Formation of lipid peroxides rises sharply when platelets undergo the release reaction. In this study the in vitro effect of vitamin E on platelet aggregation was investigated. alpha-Tocopherol, an anitoxidant of known inhibitory action on lipid peroxidation, was added to platelet suspensions in concentrations up to 1.5 mM. A dose-dependent reduction in platelet aggregation was observed, with complete inhibition of the secondary wave of aggregation at greater than or equal to 0.9 mM alpha-tocopherol. The inhibitory effect of alpha-tocopherol on the platelet release reaction was further documented by the decrease in aggregation-induced release of [14C]5-hydroxytryptamine from prelabeled platelets and by the reduction of N-acetylglucosaminidase activity released into the medium. The sharp rise in lipid peroxides normally associated with platelet aggregation was markedly reduced by alpha-tocopherol and also by acetylsalicylic acid, a known inhibitor of the platelet release reaction. In vivo studies examined the effect of oral vitamin E administration (1,200-2,400 IU daily) on plasma and platelet levels of alpha-tocopherol. Up to 1,800 IU daily, increasing dosages of vitamin E resulted in increasing concentrations of alpha-tocopherol in plasma and platelets, but intake of vitamin E in excess of this dosage failed to show any further increase in plasma or platelet levels.