Aspirin,Platelet Aggregation,and the Circadian Variation of Acute Thrombotic Events

The onset of several acute cardiovascular diseases occurs in a circadian pattern, with a peak incidence in the hours soon after awakening. This finding, coupled with laboratory data that confirm a surge in platelet activation during the early morning hours, suggests that acute changes in platelet aggregability may be an important trigger of thrombosis. Therefore, the efficacy of antiplatelet agents, such as aspirin, in reducing risks of vascular occlusion may result, at least in part, from a blunting of these short-term changes in platelet aggregability. In this review, clinical and laboratory evidence describing these cyclical changes is discussed, as is current evidence of the effects of aspirin on platelet function and the circadian variation of acute thrombosis.     

Epidemiological characteristics of platelet aggregability.

The epidemiological characteristics of platelet aggregability were established in 958 participants in the Northwick Park Heart Study. The main analyses were based on the dose of adenosine diphosphate at which primary aggregation occurred at half its maximum velocity. Aggregability increased with age in both sexes, was greater in whites than blacks (particularly among men), and tended to decrease with the level of habitual alcohol consumption. Aggregability was, however, greater in women than men and in nonsmokers than smokers. There was no relation between aggregability on the one hand and obesity, current or past oral contraceptive use, menopausal state, or blood cholesterol and triglyceride concentrations on the other. Aggregability was somewhat, though not significantly, higher in men with a history of ischaemic heart disease and in those with electrocardiographic evidence of ischaemia than in those without. There was a strong association between the plasma fibrinogen concentration and aggregability. The widely held concept of platelet aggregability and its implications is probably an oversimplification. In the prevention of thrombosis it may be as useful to consider modifying external influences on platelet behaviour, such as plasma fibrinogen concentration or thrombin production, as it is to rely solely on platelet active agents.     

A regimen for low-dose aspirin?

The effects of different regimens of 40 mg aspirin on platelet thromboxane A2 synthesis and vascular prostacyclin synthesis were determined in patients who were undergoing elective surgery for removal of varicose veins. Aspirin 40 mg taken at intervals of 48 hours consistently reduced platelet thromboxane A2 synthesis to a level at which it failed to support platelet aggregation and the associated release reaction. This effect lasted for at least 36 hours. In contrast, aspirin 40 mg every 72 hours did not have the same consistent effect. Both dose regimens led to a reduction in vascular prostacyclin synthesis 12 hours after the last dose, but 36 or 72 hours after the last dose prostacyclin synthesis was not reduced; thus the inhibition of prostacyclin synthesis was short lived. If the balance between platelet thromboxane A2 and vascular prostacyclin synthesis is important in thrombosis 40 mg aspirin every 48 hours may have the maximum antithrombotic effect.     

Flow Cytometric Analysis of Circadian Changes in Platelet Activation Using Anti-Gmp-140 Monoclonal Antibody

The hemostatic activity of blood shows a circadian variation with a higher frequency of acute coronary events in the morning. The thrombotic tendency of blood is influenced by many factors, including platelets. Diurnal changes of in vivo platelet activation were investigated by whole blood flow cytometry in 10 young healthy male volunteers using anti-GMP-140 (anti-α-granule membrane protein 140 kD) monoclonal antibody at 3h intervals from 06:00 to 24:00. We also studied circulating platelet aggregates to investigate whether there exists a similarity between the results of these methods. Results of flow cytometric analysis indicate that there is an increase in platelet activation during the period from 06:00 to 09:00. Platelet activation then decreases gradually during the period from noon to midnight. These changes are accompanied by a similar trend in circulating platelet aggregates. This suggests that GMP-140 expression on platelets is synchronized with or followed by platelet aggregate formation in vivo, and increased platelet activation may predispose individuals to thrombosis at this time.     

Flow cytometric analysis of circadian changes in platelet activation using anti-GMP-140 monoclonal antibody.

The hemostatic activity of blood shows a circadian variation with a higher frequency of acute coronary events in the morning. The thrombotic tendency of blood is influenced by many factors, including platelets. Diurnal changes of in vivo platelet activation were investigated by whole blood flow cytometry in 10 young healthy male volunteers using anti-GMP-140 (anti-alpha-granule membrane protein 140 kD) monoclonal antibody at 3h intervals from 06:00 to 24:00. We also studied circulating platelet aggregates to investigate whether there exists a similarity between the results of these methods. Results of flow cytometric analysis indicate that there is an increase in platelet activation during the period from 06:00 to 09:00. Platelet activation then decreases gradually during the period from noon to midnight. These changes are accompanied by a similar trend in circulating platelet aggregates. This suggests that GMP-140 expression on platelets is synchronized with or followed by platelet aggregate formation in vivo, and increased platelet activation may predispose individuals to thrombosis at this time.     

The human endogenous circadian system causes greatest platelet activation during the biological morning independent of behaviors

Background

Platelets are involved in the thromboses that are central to myocardial infarctions and ischemic strokes. Such adverse cardiovascular events have day/night patterns with peaks in the morning (∼9AM), potentially related to endogenous circadian clock control of platelet activation. The objective was to test if the human endogenous circadian system influences (1) platelet function and (2) platelet response to standardized behavioral stressors. We also aimed to compare the magnitude of any effects on platelet function caused by the circadian system with that caused by varied standardized behavioral stressors, including mental arithmetic, passive postural tilt and mild cycling exercise.

Methodology/Principal Findings

We studied 12 healthy adults (6 female) who lived in individual laboratory suites in dim light for 240 h, with all behaviors scheduled on a 20-h recurring cycle to permit assessment of endogenous circadian function independent from environmental and behavioral effects including the sleep/wake cycle. Circadian phase was assessed from core body temperature. There were highly significant endogenous circadian rhythms in platelet surface activated glycoprotein (GP) IIb-IIIa, GPIb and P-selectin (6–17% peak-trough amplitudes; p≤0.01). These circadian peaks occurred at a circadian phase corresponding to 8–9AM. Platelet count, ATP release, aggregability, and plasma epinephrine also had significant circadian rhythms but with later peaks (corresponding to 3–8PM). The circadian effects on the platelet activation markers were always larger than that of any of the three behavioral stressors.

Conclusions/Significance

These data demonstrate robust effects of the endogenous circadian system on platelet activation in humans—independent of the sleep/wake cycle, other behavioral influences and the environment. The ∼9AM timing of the circadian peaks of the three platelet surface markers, including platelet surface activated GPIIb-IIIa, the final common pathway of platelet aggregation, suggests that endogenous circadian influences on platelet function could contribute to the morning peak in adverse cardiovascular events as seen in many epidemiological studies.     

Circadian and seasonal variations of physiological and biochemical determinants of acute myocardial infarction

Almost all cardiovascular events occur according to a circadian rhythm with a greater frequency in the morning on waking and when resuming activity, the mechanism and precise triggering events for myocardial infarction (MI) are not yet fully known. Multiple biologic functions show a diurnal and/or seasonal variation that may contribute to adverse cardiac outomes. Exogenous factors may also modulates these variations. The MI peak usually occurs between 07:00 and 12:00 h. This timing corresponds to the concurrent increase in platelet aggregability, blood concentration of cortisol, catecholamines, angiotensin II, myocardial oxygen demand and coagulation activity, while fibrinolytic activity is decreased. In this review paper we will point out the biological rhythms of a number of functions involved in acute myocardial infarction e.g. blood pressure, hormonal determinants, cholesterol, among others.     

Postprandial hyperlipemia inhibits platelet aggregation without affecting prostanoid metabolism

The objective of this work was to characterize changes in platelet aggregability during postprandial hypertriglyceridemia with special emphasis on arachidonic acid metabolism. Ten healthy young men consumed 100 g fat after a fasting period of 12 hr. In-vitro platelet aggregation induced by ADP and collagen was measured at 0, 3, 5, and 9 hours after the fat intake. The major arachidonic acid metabolites, 12-hydroxyeicosatetraenoic acid (12-HETE), thromboxane A2 (TXA2), prostaglandin F2 alpha (PGF2a), and prostaglandin E2 (PGE2) produced during collagen-induced platelet activation were quantified by gas chromatography/mass spectrometry. A significant decrease in platelet aggregability induced by both ADP and collagen was detected during the postprandial hyperlipemia. No significant changes could be found in the prostanoid pattern of collagen activated platelets. There was no correlation between the degree of the inhibition of platelet aggregation and the relative or absolute increase of triglyceride-levels in the plasma during the postprandial hyperlipemia.     

Successful therapy of the prostaglandin defect "Wien-Hietzing" (lack of platelet high-affinity binding sites for prostaglandin I2)

Some years ago we detected a lack of platelet high-affinity PGI2 binding sites in a 10 year-old girl who presented to the outpatient unit with clinical symptoms and signs of acute popliteal artery occlusion. We named this new defect in the prostaglandin system "Wien-Hietzing". Acute surgery was successful. On the basis of earlier findings that aspirin is able to sensitize platelets to the action of PGI2 and produce beneficial changes in platelet sensitivity, we decided to treat this girl with a daily dosage of 20 mg aspirin orally. Repeated control examinations during the total follow-up period of about 6 years revealed normalized platelet sensitivity and normalized receptor behaviour. The girl is symptom-free to date. It is concluded that this prostaglandin defect may be successfully treated with long-term, low-dose aspirin administration.     

Preventing venous thromboembolism in elderly patients with hip fractures: studies of low-dose heparin,dipyridamole, aspirin,and flurbiprofen.

Low-dose heparin, dipyridamole (alone and in combination with aspirin), and flurbiprofen were evaluated as potential prophylactic agents against deep venous thrombosis in elderly patients with hip fractures. None of the agents that modify platelet behaviour could reduct the frequency of isotopically diagnosed venous thrombosis. Low-dose heparin reduced the overall frequency of venous thrombosis and its extent as judged by the frequency of bilaterally abnormal scans, but this reduction did not achieve statistical significance.     

Plateletworks: a novel point of care platelet function screen

Platelet function is critically important in the acute-care settings of cardiopulmonary bypass surgery and percutaneous coronary intervention, which are commonly associated with the adverse vascular events of hemorrhage and thrombosis, respectively. To improve outcomes, it has been suggested that patients should be screened for platelet count and function periprocedurally, and therapeutic intervention including the possible use of thrombolytics and adequate anticoagulation or administration of antiplatelet agents, should be utilized. Antiplatelet therapy including aspirin (acetylsalicylic acid), the thienopyridines (clopidopgrel), and parenteral anti-glycoprotein (GP) IIb/IIIa agents (abciximab, tirofiban, and eptifibatide) are recognized as clinically important in patients at risk of developing thrombotic events. Recently, it has been recognized that empiric therapeutic administration of these agents may be suboptimal in clinical environments because of interpatient variability with regard to platelet count, platelet response, receptor concentration on the platelet, and other factors. Hence there is a clinical need to monitor such therapies on an individual basis. Traditional platelet tests including light transmission aggregometry (LTA) are inconvenient for acute diagnostic testing because of the complexity of the test and the requirement for specialty training. Hence, 'near-patient' test systems have recently been introduced. Plateletworks is an in vitro diagnostic, point-of-care test platform that has demonstrated utility in monitoring platelet response to all current antiplatelet agents including aspirin and clopidogrel.     

Inhibition of platelet function by high-density lipoprotein from a patient with apolipoprotein E deficiency.

Apolipoprotein E-(apoE-) rich high-density lipoprotein (HDL) of normal subjects showed marked inhibitory effects on platelet aggregation and ATP release as compared with apoE-poor HDL, suggesting that apoE has inhibitory effects on platelet function (Desai et al. J. Lipid Res. 30:831, 1989; Higashihara et al. FEBS Lett. 282:82, 1991). A patient with apoE deficiency showed evidence of decreased platelet aggregability in platelet-rich plasma, but normal aggregability in washed platelets. Both patient's plasma and HDL fraction inhibited platelet aggregation of normal subjects. Patient's HDL reconstituted with recombinant apoE showed further inhibitory effects on platelet function. These results suggest that apoE is a potent, but not unique, inhibitory factor for HDL.     

External triggers of onset of myocardial infarction-an update

Coronary heart disease is one of the leading causes of death in both industrialised and developing countries. About two thirds of all coronary deaths occur outside the hospital and before any medical care can be reached. Therefore, the prevention of coronary events appears to be of utmost importance. Rupture of an atherosclerotic plaque and subsequent coronary thrombosis is the most common underlying pathophysiological mechanism of coronary events. External stresses or 'triggers' imposing on coronary plaques may precipitate plaque rupture. External triggers include physical activity, various emotional stresses, eating, environmental factors, and sexual activity. The increased relative risk of myocardial infarction may be induced by external triggers via the activation of internal triggering mechanisms such as biomechanical and hemodynamic stresses and changes in platelet aggregability and blood viscosity. Recent prospective studies have confirmed the results of earlier retrospective studies and have, similarly, shown the importance of external triggers in increasing the risk of myocardial infarction and other coronary events. Particularly, sedentary individuals with underlying coronary heart disease appear to be at risk when exposed to external triggers. Regular physical activity, on the other hand, is a protective factor against the increased risk associated with external triggers. However, health education needs to be provided about the risks associated with strenuous exercise in untrained individuals. Apart from lifestyle factors, pharmacological protection of the population at risk during vulnerable periods plays an important role as well. Medication such as beta-blockers and aspirin may lower the increased risk of coronary events during exposure to external triggers. Also, further research is needed with regard to psychosocial stressors and their potential role as external triggers of myocardial infarction and other coronary events.     

血栓弹力图指导支架内血栓后不稳定心绞痛治疗1 例及文献复习

目的:探讨在血栓弹力图监测血小板抑制率的情况下,调整氯吡格雷及阿司匹林用量,治疗冠心病、PCI术后支架内再发血栓患者的临床意义。方法:报告中国人民解放军总医院1例支架内亚急性血栓患者的临床资料并复习相关文献,对其临床表现、诊断、在血栓弹力图指导下的治疗进行分析。结果:1例支架内亚急性血栓患者经治疗病情好转出院,出院后继续调整氯吡格雷及阿司匹林用量,达到满意血小板抑制率,患者症状消失。结论:冠状动脉介入治疗后发生支架内血栓的患者,应用血栓弹力图指导氯吡格雷及阿司匹林用量,可达到令人满意的血小板抑制率,并防止出血情况发生。     

Role of platelets in hemostasis and thrombosis.

Platelets are central to both normal hemostasis and abnormal thrombotic states along with the vessel wall, coagulation elements, and blood flow. The platelets play a pivotal role in the reaction that occurs after vessel injury, during which platelets first adhere to the vessel wall, undergo a release reaction and then aggregate, probably as a result of the materials released from platelets. These processes can be studied by a series of in vitro tests which form the basis of our knowledge of platelets in hemostasis. While the hemostatic plug is usually microscopic in size, this same plug (platelet thrombus) may contribute to the pathogenesis of several arterial diseases such as transient ischemic attacks, sudden blindness, sudden cardiac death and acute respiratory death syndrome. Careful microscopic examinations have shown that platelet aggregates may be found in the microcirculation which could affect vital structures such as the conduction system of the heart. Both anatomic and therapeutic evidence evidence suggests that platelets play a role in venous thrombosis. Recent evidence suggests increased levels of materials known to be released from platelets in patients with both arterial and venous thrombi along with increased platelet coagulant activities in patients with venous thrombosis.     

生物节律影响巨核细胞发育和血小板产生的研究进展

自然界中生物体的生命活动、生活习性都存在着一定的周期性变化。生物昼夜节律的产生是以内源性的生物钟系统为基础的。生物钟不仅易受到外界环境的影响,而且可以通过调控一系列特定的下游基因的表达,影响生物体的生理生化过程。巨核细胞是生成血小板的前体细胞,经过分化、增殖、成熟和裂解,最终生成血小板。血小板是一种没有细胞核的特殊细胞,在生理性止血和器官修复上发挥着重要作用,同时参与血栓等多种疾病的发生。近几年借助现代分子生物学和细胞生物学手段。证实了哺乳动物的巨核细胞和血小板的生成呈现明显的周期性的变化,利用生物钟基因缺失模型进一步发现了生物钟基因对巨核细胞和血小板的影响。本文概述了生物节律对巨核细胞和血小板的影响,为进一步研究巨核细胞的发育和血小板生成机制提供了参考。     

Equivalent inhibition of in vivo platelet function by low dose and high dose aspirin treatment

In vitro platelet function was inhibited in healthy volunteers by two different doses of aspirin, as confirmed by measurement of maximum serum production of thromboxane B2 (TXB2) by platelets. 75 mg aspirin did not fully inhibit serum TXB2 production after 24 hours, whereas 300 mg aspirin did. Inhibition of platelet function in vitro was maintained by both 75 mg/day aspirin or 300 mg/alternate day aspirin. In contrast, in vivo production of TXB2, measured as urinary levels of the 11-keto-TXB2 metabolite, was inhibited similarly by both doses of aspirin throughout the study. These findings suggest that 75 mg/day aspirin may be sufficient adequately to inhibit platelet aggregation in vivo.     

The Effect of Exercise on Platelet Aggregability and Lipid Peroxidation

The effects of single bicycle ergometric exercises of a moderate or high intensity on platelet ADP-dependent aggregation, content of malonic dialdehyde (MDA), and activity of catalase were studied in young men with relatively high working capacities. Platelet aggregability either increased or decreased in response to the exercise. Hyperaggregation was recorded in about two-thirds of the subjects and hypoaggregation, in one-third. Changes in the aggregation parameters correlated with their basal values at rest. Muscular activity raised the aggregability of platelets when it was initially low and lowered it when it was initially high. An increased intensity of exercise made these correlations stronger. The exercise-caused changes in the MDA content in platelets also correlated with its basal level. A low MDA content at rest increased, and an initially high MDA content decreased. The response of hypoaggregation was associated with moderate and strong correlations between the basal content of MDA and catalase activity in platelets and the exercise-induced changes in the MDA content and platelet aggregability. It was concluded that a high basal level of MDA in platelets is a factor decreasing platelet aggregability during muscular activity.     

Effect of aspirin on prostaglandin synthesis by human platelets

When platelet rich plasma is exposed to N-ethylmaleimide, a ten fold increase in measurable prostaglandin E synthesis occurs. This effect is almost completely abolished within 2 hours of ingestion of 600 mg of aspirin by human volunteers. Recovery of this platelet function is slow for the first two days, returning sharply to normal over the next six days and plateauing approximately 8 days following initial removal from aspirin. It is suggested from these studies that platelet prostaglandin E production following NEM may be a useful test of platelet function.     

Secretory dysfunction of vascular endothelium limits the effect of angiotensin converting enzyme inhibitor quinapril on aggregation of erythrocytes in experimental hypertension.

Using automatic erythrocyte aggregometer type MA-1 (Myrenne gmbh, Germany), we investigated the hypothesis that therapeutic effectiveness of quinapril--angiotensin converting enzyme inhibitor (ACEI)--in the treatment of hypertension would correlate with improvement of red blood cell (RBC) aggregability. Experiments were performed on commercially available inbred strain of spontaneously hypertensive male rats (SHR) aged 19-21 weeks. Age-matched normotensive Wistar-Kyoto (WKY) rats genetically related to SHR were used as a control. Aggregability of RBC in hypertensive rats was significantly higher than in control WKY animals. Quinapril (100 microg/kg) administered i.p. for 8 days improved RBC aggregability in normotensive rats but surprisingly not in SHR animals. Beneficial effect of quinapril on RBC aggregation observed in normotensive animals did not occur when this drug was injected in combination with aspirin (1 or 50 mg/kg) or with indomethacin (20 mg/kg) or with L-NAME (10 mg/kg). However, much the same damaging effects on RBC aggregability were observed when aspirin, indomethacin or L-NAME were each administered into normotensive animals without quinapril. In contrast with normotensive rats, aggregability of RBC in SHR was not affected either by quinapril or by indomethacin and by L-NAME, given separately or in combination. The only compound significantly worsening RBC aggregability in SHR was aspirin but this effect was not dose-dependent. Quinapril-induced improvement of RBC aggregability in normotensive rats (but not in SHR) was completely abolished by simultaneous administration of B2 receptor antagonist icatibant and successfully mimicked by 8 days of treatment with bradykinin. In vitro aggregability of RBC isolated from WKY was not affected by previous incubation (30 min at 37 degrees C) with quinapril, indomethacin or L-NAME. Only aspirin (3 mM) significantly increased RBC aggregability as compared to placebo. It is concluded that under physiological conditions quinapril efficiently inhibits RBC aggregability and this effect is modulated by secretion of endothelial mediators, mainly prostacyclin and nitric oxide. In hypertension quinapril, in spite of lowering of arterial blood pressure, is unable to display its beneficial effects on RBC aggregability possibly due to the hypertension-induced/accompanied dysfunction of vascular endothelium. Aspirin revealed unique erythrocyte damaging properties, presumably independent of inhibition of cyclooxygenase but related to a direct membrane protein acetylation.