Hideo Ikarugi

Because of the high mortality from myocardial infarction and stroke, there is a great demand for finding novel methods of diagnosis, prevention and treatment of these diseases. Most of the current tests measure important determinants of thrombosis such as platelet function, coagulation and fibrinolysis in isolation; therefore, a global test measuring the actual thrombotic status would be more useful in clinical conditions. We obtained considerable experience by using the global thrombosis test, which determines the actual thrombotic status by taking into account the measured platelet reactivity, coagulation and fibrinolytic activities. In animal experiments, we found significant correlation between the ex vivo global thrombosis test measurements and the in vivo thrombotic status. The published evidence for the benefit of an antithrombotic diet with regular physical exercise is also described.

Prevention of thrombotic disorders has priority over treatment. There are only two pathologically relevant tests which are suitable for measuring the overall thrombotic status both in experimental conditions and in humans. The Global Thrombosis Test (GTT) and the Global Parallel-Plate Thrombosis Test can detect the pathologically relevant global thrombotic status. These tests have been successfully used for monitoring the effect of antithrombotic drugs and for developing novel antithrombotic agents. By using GTT, varieties of fruits, vegetables, and regular physical exercise have been tested for the effect on global thrombotic status. This review discusses the published evidence for the benefit of diet of selected fruit and vegetable varieties and doing regular physical exercise on improving thrombotic status. Future clinical trials monitored by GTT or Global Parallel-Plate Thrombosis Test could decide on the effectiveness of an experimentally proven antithrombotic diet with regular physical exercise in the prevention of thrombotic diseases.

A study was conducted to investigate the relationship between sprinting ability and agility using a longitudinal design. The subjects were 70 junior athletes (39 boys and 31 girls) aged 9 to 12 years participating in a Talent Identification and Development program. Physical constitution (height and body weight), 20-m sprint test time, rebound jump index, and ability to change direction (agility test and T-run test) were measured periodically for three years. The main results were as follows:<br>  1.&nbsp;&nbsp;Physical constitution, sprinting ability, rebound jump index, and ability to change direction increased significantly along with development in both boys and girls.<br>  2.&nbsp;&nbsp;Performances in the 20-m sprint, agility test, and T-run test in boys were significantly better than in girls, but no significant sex difference was observed in the rebound jump index.<br>  3.&nbsp;&nbsp;Single correlation analysis demonstrated significant correlations between all of the tests.<br> 4.&nbsp;&nbsp;Partial correlation analysis (using age in months as the control variable) revealed significant relationships between the agility test and T-run test in boys and girls, but correlations between 20-m sprint ability and the agility test or T-run test were not significant.<br>  5.&nbsp;&nbsp;There was little difference in the rebound jump index between boys and girls, and it was suggested that ability to change running direction was less affected by simple sprinting or strength-shortening cycle abilities during the developmental period.<br>

<p>While exercise is widely believed to prevent atherothrombotic diseases, it occasionally causes sudden death. This exercise paradox may be due to the inadequate testing of the thrombotic and thrombolytic status. A recently developed shear-induced thrombosis/endogenous fibrinolysis test performed with non-anticoagulated blood samples allows the assessment of the thrombotic state of an individual both at rest and after exercise. This sensitive and physiologically relevant test may help to solve the aforementioned exercise paradox.</p>

Introduction: Prevention of arterial thrombotic diseases has high priority in developed countries. Taurine (2-aminomethylsulfonic acid), which is rich in sea foods, showed antithrombotic effect in animal models of thrombosis. The present study aimed to investigate such effect in healthy human volunteers. Methods and Results: In 101 healthy Japanese people the overall thrombotic status was accessed from non-anticoagulated blood sample by the Global Thrombosis Test (GTT). There was no significant correlation between taurine concentration in urine samples and GTT-Occlusion Times (OT; mainly reactivity of platelets). In contrast, a significant inverse correlation was demonstrated between urine taurine concentrations and GTT-Lysis Times (LT; showing spontaneous thrombolytic activity). Conclusions: Our findings suggest that taurine enhances endogenous thrombolytic activity which could be a mechanism of the earlier observed cardioprotective and antithrombotic effect. (C) 2012 Elsevier Ltd. All rights reserved.

Objective: To determine and compare thrombotic and endogenous thrombolytic status in Japanese and Western populations. Background: Incidence of coronary heart disease (CHD) and AMI in Japan remains lower than in Western countries. Primary genetic effects are unlikely, given the increased CHD in Japanese migrants. For men, cholesterol and blood pressure have been similar in Japan and the U.S. Dietary factors are implicated, but how these effect CHD is unclear. We postulated that differences in thrombotic and/or thrombolytic status may contribute. Methods: We measured thrombotic and thrombolytic status in 100 healthy Japanese (J) from Japan and 100 healthy Westerners (W) from the U.K. using the Global Thrombosis Test (GTT). The GTT employs non-anticoagulated blood to create platelet-rich thrombi under high shear (occlusion time OT: seconds), and then measures the restart of blood flow, due to spontaneous thrombolysis (lysis time LT; seconds). Results: OT was longer in (J) compared to (W) (545 vs. 364, p&lt;0.0001). LT was longer in (J) than in (W) (1753 vs. 1052, p&lt;0.0001). Distribution of LT in (J) did not conform to a normal population, with markedly impaired thrombolytic status (LT&gt;3000 s) in 18%, compared to none of the Westerners (p&lt;0.0001). Conclusions: There are marked differences in thrombotic and thrombolytic status, with (J) having less prothrombotic (longer OT) but less favourable endogenous thrombolytic profile (longer LT). This may be important in the aetiology of thrombotic events. Since platelets and thrombolysis were both inhibited in (J) and yet incidence of AMI is lower, OT would seem more important than LT as a determinant of overall thrombotic risk in this population. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Introduction: We have shown earlier that diacylglycerol (DAG) but not triacylglycerol (TAG) inhibited thrombus formation. The aim of the present study was to investigate the mechanism of this antithrombotic effect of DAG. Materials and Methods: Four different diets, the (1) Western-style high-fat diet (HFD) containing 20% lipid and 0.05% cholesterol (w/w), (2) TAG-rich and (3) DAG-rich HFDs containing 20% lipid and 0.05% cholesterol, but all lipid replaced by TAG or DAG oil with very similar fatty acid composition and the (4) Japanese-style low-fat diet (LFD) containing 7% oil but no cholesterol were given to apolipoprotein E and low-density lipoprotein (LDL) receptor double-deficient mice. Atherogenicity was assessed by morphology, mapping the whole aorta and measuring the total area of lipid-stained lesions. Endothelial function was measured by the flow-mediated vasodilation test. Platelet reactivity was assessed from native blood sample by a shear-induced platelet function test (hemostatometry). Serum lipoprotein profile was measured by HPLC. Results: Both the Western-style and the TAG-rich HFDs have accelerated atherosclerosis. In contrast, DAG-rich HFD inhibited the atherosclerotic process to an extent comparable with the Japanese-style LFD. There was no significant difference in platelet and coagulant activity between the studied diet groups. DAG-rich but not the TAG-rich HFD significantly suppressed serum LDL cholesterol level. Conclusions: The present findings suggest that the mechanism of antithrombotic and anti-atherogenic effect of DAG may involve the protection of the vascular endothelium from injury and lowered serum LDL cholesterol. Copyright © 2006 S. Karger AG.

Introduction: Earlier we have demonstrated a prothrombotic state in spontaneously atherogenic rodents kept on Western-style high fat diet. The aim of the present study was to investigate the cellular mechanism of such prothrombotic state. Materials and methods: Two kinds of diets, Western-style high fat diet containing 20% fat (w/w) and 0.05% cholesterol (w/w) and low fat diet containing 7% fat without cholesterol based on AIN93G, were added to diet-sensitive apolipoprotein E and low-density lipoprotein receptor double deficient male mice for 12 or 18 weeks from 6 weeks of age. Atherosclerosis was assessed by morphometry of the aortic wall or lipid-stained lesions. Endothelial function was measured by flow-mediated vasodilation (FMV) of the femoral artery. Platelet reactivity was measured ex vivo by a shear-induced platelet aggregation test. Results and conclusions: 12 weeks feeding of mice with high fat diet significantly impaired FMV, as compared with mice fed with low fat diet (P &lt 0.05). In contrast, there was no significant difference in the lipid-stained areas and in the reactivity of platelets between the two groups. 18 weeks feeding with high fat diet significantly impaired FMV (P &lt 0.05) and enhanced both lipid-stained areas (P &lt 0.05) and platelet reactivity (P &lt 0.01). These findings show that in high fat diet-induced prothrombotic state, endothelial dysfunction precedes both the morphologically detectable lesions and the enhancement of platelet reactivity. © 2005 Elsevier Ltd. All rights reserved.

The aim of the present study was to investigate the effect of a NO donor (GSNO) and a plasma kallikrein inhibitor (PKSI-527) alone and in combination on global haemostatic status. A new in vitro test was employed which allows the measurement of both platelet function and spontaneous thrombolysis. Sixteen healthy young and 18 elderly volunteers were enrolled in this study. When GSNO (1 mM) or PKSI-527 (20 μM) was added to native human blood, platelet reactivity was significantly inhibited in both age groups. The combination of GSNO and PKSI-527 had additive inhibitory effect on platelets. Addition of either GSNO or PKSI-527 to blood samples did not significantly affect spontaneous thrombolysis, while added together, spontaneous thrombolysis was significantly enhanced. The thrombolysis enhancing effect was more prominent in elderly subjects. Our present findings suggest that the combination of NO donor and plasma kallikrein inhibitor may have clinical antithrombotic potential. © 2005 Elsevier Ltd. All rights reserved.

Background: Although incidence of arterial thrombotic events increases with advancing age, the mechanism of such increased risk is poorly understood. There is also a gender difference coronary heart disease is more common in men than women. The aim of the present study was to investigate the possible contribution from platelet reactivity and spontaneous thrombolysis, determinants of arterial thrombosis, to the increased risk of thromboembolism in aging, but otherwise healthy subjects. Method: One hundred and forty-five normal subjects (61 men, 82 women) of age between 19 and 77 years were grouped into groups of young (&lt 30 years) middle aged (31-50 years), and older (&gt 51 years), according to their gender and body mass index (BMI). A new in vitro test [Gorog Thrombosis Test (GTT)] was used to measure in sequence both platelet reactivity [occlusion time (OT)] and spontaneous thrombolysis [lysis time (LT)] from one non-anticoagulated blood sample. Result: OT in all women was 315±9.2 s (mean±S.E.M.) , in all men OT was 300±10.6 s. The mean LT in all women was 2557±201.5 s, and in all men LT was 2493±198.6 s. Advanced age did not enhance platelet reactivity (OT), but increased BMI did (P=0.039). Spontaneous thrombolysis (LT) was impaired in older men but not in women (difference between young vs. middle age: P=0.019 young vs. older: P=0.0002). Conclusion: Our findings suggest that in men, spontaneous thrombolytic activity is reduced with age, and this may explain the increased frequency and severity of thromboembolic events. Interestingly, in women, spontaneous thrombolytic activity did not change after menopause. © 2004 Elsevier Ltd. All rights reserved.

Platelets play a crucial role in the pathogenesis of acute cardiac events, such as angina, myocardial infarction and sudden death. It is believed that regular low-intensity exercise can reduce, while high-intensity exercise may provoke acute cardiac events. The aim of the present study was to investigate the effect of acute exercise both at low and high intensities on the ventilatory threshold (VT), platelet reactivity and coagulation before and after exercise. Platelet reactivity and coagulation were measured under flow condition, using native blood, by hemostatometry. Seven healthy young men (age: 20-29 years) performed bicycle ergometer exercise for 30 min at intensities of 90% (Ex-VT90% or approximately 55% VO 2max) and 130% (Ex-VT130% or 80% VO 2max) of individual VT. Blood cell counts, hematocrit, blood lactic acid and plasma catecholamine levels were slightly but significantly increased after Ex-VT90% and markedly after Ex-VT130% after 30 min exercise. Subsequent to the exercise, the elevated blood cell counts decreased to the resting levels both at Ex-VT90% and at Ex-VT130%. Platelet reactivity to shear stress and dynamic coagulation were significantly enhanced immediately and 30 min after Ex-130%VT. In contrast, no significant changes occurred in those of Ex-90%VT. The present study suggests that high-intensity exercise-induced platelet hyperreactivity and hypercoagulable state may pose an increased risk for acute, sometimes fatal cardiac event. On the other hand, our findings support the view that low-intensity exercise does not present a risk of thrombosis. Copyright © 2003 S. Karger AG, Basel.

This is the first laboratory evaluation of a new instrument, designed to test both platelet function and thrombolytic activity from a native blood sample, in vitro. The inventor assumed that the reduction and arrest of blood flow was due to activation, aggregation and stabilized thrombus formation by shear-activated platelets, and that re-establishment of flow was due to thrombolysis. Morphologic and functional studies presented here confirm these mechanisms. In vitro tests provided incontestable evidence for the principal role of platelets in the obstruction of flow (occlusion time) and for thrombolysis as the principal mechanism underlying the restoration of blood flow (lysis time). In addition to aggregation, it is the explosive generation of thrombin by shear-activated platelets that results in the formation of an occlusive haemostatic thrombus. Anticoagulation of blood completely prevented occlusion. Platelet-rich thrombus formation (occlusion time) was dose-dependently inhibited by monoclonal antibody against platelet glycoprotein (GP) Ib (6B4 and 12E4), aurin tricarboxylic acid, monoclonal antibody against platelet GPIIb/IIIa (MA-16N7C2 and abciximab), a synthetic GPIIb/IIIa antagonist (TAK-029), thrombin inhibitor (argatroban), and anti-von Willebrand factor, but not by anti-fibrinogen. Plasminogen activator streptokinase (Varidase) and tissue-type plasminogen activator (Monteplase) dose-dependently enhanced thrombolysis (lysis time) without affecting platelet function (occlusion time). The test is specific for thrombolysis. The plasmin inhibitor tranexamic acid prevented plasminogen activator-induced thrombolysis, while inhibition of clot retraction by cytochalasin B did not affect the lysis time. This rapid and sensitive global test of platelet function and thrombolytic activity could be of great value both in research and in clinical practice. © 2003 Lippincott Williams &amp Wilkins.

The effects of exercise and catecholamines on platelet reactivity or coagulation and fibrinolysis appear to be inconsistent. This may be partly due to the methods employed in previous studies. In the present study, we investigated the effects of acute aerobic exercise and catecholamines on the thrombotic status by a novel in vitro method, shear-induced hemostatic plug formation (hemostatometry), using nonanticoagulated (native) blood. Aerobic exercise (60% maximal O₂consumption) was performed by healthy male volunteers for 20 min, and the effect on platelet reactivity and coagulation was assessed by performing hemostatometry before and immediately after exercise. Exercise significantly increased shear-induced platelet reactivity, coagulation, and catecholamine levels. The effect of catecholamines on platelet reactivity and coagulation was assessed in vitro by adding catecholamines to blood collected in the resting state. The main findings of the present study are that elevation of circulating norepinephrine at levels that are attained during exercise causes platelet hyperreactivity and a platelet-mediated enhanced coagulation. This may be a mechanism of an association of aerobic exercise with thrombotic risk.