Aim: Up-regulation of soluble CD40 ligand (sCD40L) and of monocyte chemoattractant protein-1 (MCP-1) has been found in diabetes and in patients with acute cerebral ischemia. We asked whether (i) the two molecules are similarly upregulated among non-lacunar and lacunar diabetic strokes and (ii) sCD40L and/or MCP-1 predict the risk of cardiovascular events in this setting.
Methods: Ninety patients with type 2 diabetes mellitus presenting with an acute ischemic stroke (compared with 45 control subjects) were evaluated on admission and up to 36 months (median 24 months) after the event.
Results: Diabetic patients with acute stroke had higher plasma CD40L and MCP-1 than controls (p<0.0001), with no significant differences among lacunar and non-lacunar strokes. On multiple regression analysis, only higher sCD40L quartiles and older age were associated with higher MCP-1 quartiles. Forty-eight percent of patients experienced vascular events. Cox regression analysis showed that only the presence of higher sCD40L values independently predicted the recurrence of vascular events.
Conclusion: Up-regulation of inflammatory molecules, such as CD40L and MCP-1, is involved in the advanced stage of atherosclerotic cerebro-vascular disease and is associated with increased risk of recurrence of cardiovascular events.

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Aim: To examine whether fasting blood lipid profiles are associated with the risk of myocardial infarc-tion among Japanese men.
Methods: We conducted a nested case-control study in the Morbidity of Myocardial Infarction Mul-ticenter Study in Japan (3M Study). For each case of myocardial infarction entered in the 3M Study between 1997 and 2000, we randomly selected two controls, matched for age (±3 years), from among the participants in risk factor surveys with no history of myocardial infarction. A total of 723 male employees (241 cases and 482 controls) aged 35 to 65 years were enrolled in the present study.
Results: The subjects had significantly higher mean fasting LDL-cholesterol and triglyceride, and lower mean HDL-cholesterol than controls. The multivariable conditional odds ratio (95% confi-dence interval) for myocardial infarction after adjustment for known cardiovascular risk factors was 3.87 (1.27-11.7, p for trend <0.001) for total cholesterol [-6.71 vs <4.65 mmol/L], 3.28 (1.12-9.60, p for trend=0.001) for LDL-cholesterol [-4.64 vs <2.59 mmol/L], 0.17 (0.07-0.43, p for trend=0.001) for HDL-cholesterol [-1.55 vs <1.03 mmol/L] and 3.03 (1.37-6.70, p for trend= 0.01) for triglycerides [-2.26 vs <1.13 mmol/L].
Conclusion: High total and LDL-cholesterol, low HDL-cholesterol and high triglycerides levels were independently associated with an increased risk of myocardial infarction among middle-aged Japa-nese male workers.

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Aim: Atherosclerosis is a cardiovascular disease; however, there is little information on signal transduction for vascular function in the early stage of atherosclerosis. In this work, we investigated the role of Rho/Rho-kinase and nitrogen oxide (NO)/cyclic GMP (cGMP) signaling pathways in the aorta prior to atherosclerosis.
Methods: Tension, the expression of RhoA protein, Rho-kinase activity and the cGMP level were measured using endothelium-intact or -denuded aorta prepared from apolipoprotein E-deficient (apoE-KO) and C57BL/6 wild-type (WT) mice at 2 months of age.
Results: Phenylephrine (PE) induced less maximal contraction in the endothelium-denuded aorta from apoE-KO than from WT mice. A Rho-kinase inhibitor (Y-27632) reduced more effectively the contraction of apoE-KO than WT mice, but their RhoA proteins and Rho-kinase activities were not so different. Acetylcholine caused larger relaxation of the PE-stimulated, endothelium-intact aorta in apoE-KO due to endothelial NO release than WT mice. The basal cGMP level in the endotheliumintact aorta of apoE-KO mice was higher than that of WT.
Conclusions: Smooth muscle contraction via α₁-adrenergic receptor shows higher dependency on Rho-kinase activity, suggesting down-regulation of the mechanism different from Rho/Rho kinase signaling in the aorta prior to atherosclerosis. Endothelium-dependent relaxation is also intensified through the NO/cGMP pathway.

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Aim: To determine the effects of exercise training on total and high molecular weight (HMW) adiponectin and adiponectin oligomeric distribution.
Methods: A randomized parallel-design study (n=53; 40 women and 13 men; age range, 32?65 years) was conducted at a fitness club between April 2006 and July 2007. Participants were randomly assigned to the exercise (n=26) or control (n=27) group and received exercise training for 70 min 2 times per week for 12 weeks and exercise advice at the baseline, respectively. The primary outcomes were changes in total adiponectin, HMW adiponectin and percentage of HMW adiponectin.
Results: Muscle strength improved in the exercise group but remained stable in the control group. Body weight and BMI were not modified by exercise training. After 12 weeks, there were no differences between groups in total adiponectin levels, HMW adiponectin levels, or percentage of HMW adiponectin.
Conclusion: In the absence of weight loss, twice-weekly exercise training does not change HMW adi-ponectin levels or the adiponectin oligomer composition.

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Aim: Remnant lipoproteins are well known to play a pivotal role in atherosclerosis. In patients with postprandial dyslipidemia, metabolic pathways for exogenous lipoproteins are generally disturbed, resulting in accumulation of chylomicron remnants. Although it has been difficult to make a specific antibody against apolipoprotein B48 (apoB48) , a constituent of exogenous lipoproteins, we succeeded in creating a specific monoclonal antibody against apoB48. In this study, we isolated apoB48-containing lipoproteins from lipoproteins with a density less than 1.019 g/mL using this anti-apoB48 monoclonal antibody (4C8) .
Methods: Apolipoproteins and lipids were analyzed to confirm whether apoB48-containing lipoproteins are isolated from other lipoproteins. The characteristics of apoB48-containing lipoproteins in the plasma of patients were compared with type 2 diabetes mellitus and non-diabetic patients. Furthermore, the uptake of apoB48-containing lipoproteins by THP-1 cells (a human acute monocytic leukemia cell line) , HepG2 cells (a human hepatoma cell line) , and human umbilical vein endothelial cells (HUVEC) was investigated. Also, the expression of apoB48 receptors in these cells was tested with RT-PCR.
Results: Apolipoprotein analysis of 4C8-bound lipoproteins indicated the isolation of apoB48-containing lipoproteins, because the content of apoB100 was quite low (less than 5%) . Compared with lipoproteins that were not bound to the antibody, apoB48-containing lipoproteins had a higher content of triglycerides. There was no significant difference in the composition of apoB48-containing lipo-proteins between patients with and without type 2 diabetes. Uptake of fluorescence-labeled apoB48-containing lipoproteins by THP-1-derived macrophages and HepG2 cells, but not by HUVEC, was observed. The specificity of this uptake was confirmed because the fluorescent signal was competed out by an excess amount of the same unlabeled lipoproteins. RT-PCR revealed the expression of apoB48 receptors in THP-1 and HepG2 cells but not in HUVEC. These results suggest that specific uptake of apoB48-containing lipoproteins may occur via apoB48 receptors.
Conclusion: ApoB48-containing lipoproteins have a higher triglyceride content and are taken up into THP-1-derived macrophages and HepG2 cells by a specific pathway.

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Aim: Atherosclerosis is a systemic disease with focal cardiovascular events. Although the accelerated development of peripheral arterial disease in diabetic patients is well known, the pathogenic mechanism of site-specific susceptibility to glycemia is uncertain.
Objective: To investigate the association of fasting and post-challenge glucose levels with intimamedia thickness (IMT) at different arterial sites.
Methods: Forty consecutive middle-aged volunteers aged 37 to 53 years were recruited to define the association of IMT with cardiovascular risk factors at 12 carotid and 6 lower-limb arterial sites. A linear mixed model was used to regress the primary outcome measures, which were repeated measures of IMT at multiple arterial sites (18 sites per participant), on fixed-effect predictors of various conventional cardiovascular risk factors, while accounting for the interdependence of repeated measures taken from the same participant with unstructured covariance.
Results: Carotid IMTs were associated independently with waist circumference and systolic blood pressure, whereas lower-limb IMTs were associated with waist circumference, low-density lipoprotein cholesterol (LDL-C), glycated hemoglobin A1C (HbA1C), and fasting and 2-hour post-challenge plasma glucose levels; these associations were stronger in overall arteries. Independent associations of ALT and smoking with IMT appeared only in overall arteries.
Conclusion: In a middle-aged, nonclinical sample, lower-limb but not carotid IMTs are associated independently with HbA1C, and fasting and 2-hour post-challenge plasma glucose levels.

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Aim: This study was designed to clarify the current measurement performance of 7 reagent manufacturers for high-density lipoprotein cholesterol (HDLC), low-density lipoprotein cholesterol (LDLC) and triglycerides (TG) specified for the metabolic syndrome (MetS)-focused health checkups program in Japan.
Methods: Twenty HDLC, 21 LDLC and 9 TG analytical reagent/instrument/calibrator systems (system), and combinations of reagent lots, instrument models and calibrator lots, underwent Centers for Disease Control and Prevention (CDC)/Cholesterol Reference Method Laboratory Network (CRMLN) lipid standardization. Eighty and 100% systems were requested to achieve an accuracy of within ±1% and ±2% of the reference value, so that a clinical laboratory can meet the CDC criteria.
Results: The CDC performance criteria of HDLC, LDLC and TG require an accuracy of within ±5%, ±4% and ±5%, respectively. For HDLC, all 20 systems met the criteria. Fourteen (70.0%) and 18 (90.0%) systems were within ±1% and ±2%, respectively. For LDLC, 14 (66.7%) of 21 systems met the criteria, but 7 (33.3%) failed. Five (23.8%) and 17 (81.0%) systems were within ±1% and ±2%, respectively. For TG, 8 of 9 systems met the criteria. Two (22.2%) and 4 (44.4%) systems were within ±1% and ±2%, respectively. The minimum and maximum differences of a specified sample among manufacturers were 1.6 and 11.0 mg/dL for HDLC, 7.8 and 33.0 mg/dL for LDLC, and 2.8 and 27.4 mg/dL for TG, respectively.
Conclusion: Homogeneous HDLC methods are acceptable for MetS, but further accuracy improvement of homogeneous LDLC and TG methods will be needed because of their poor performance.

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Aim: Loss or gain in obesity indexes, such as body mass index (BMI) and waist circumference (WC), may affect serum lipid parameters. We therefore analyzed the impact of changes in WC and BMI over a one-year period on serum levels of LDL cholesterol (LDL-C), HDL cholesterol (HDL-C), and triglycerides (TG).
Methods: We analyzed the data of 3,111 individuals who were not on lipid- lowering medication and who underwent general health screening two years running.
Results: The correlation between percent changes of WC (%dWC) and BMI (%dBMI) were both statistically significantly correlated with percent changes in LDL-C (%dLDL), HDL-C (%dHDL), and TG (%dTG) except that between %dWC and %dHDL in women. In multiple regression analysis, %dBMI, but not %dWC, was found to be an independent predictor of %dLDL, %dHDL, and %dTG. When %dBMI was excluded from the variables, %dWC was identified as an independent factor predicting %dLDL and %dTG; however, in individuals with %dBMI of ≥0, %dWC was not found to be a predictor of percent changes in any lipid parameters tested in this model.
Conclusion: Percent changes in BMI were found to be an independent predictor of adverse changes in lipid parameters in both genders. Although percent changes in WC (%dWC) also tended to confer adverse changes in lipid parameters, this relationship did not remain statistically significant after controlling for %dBMI. It is suggested that changes in obesity parameters are an important goal to avoid adverse lipid changes, although there might be some gender differences.

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Aim: Increased clusterin mRNA and protein levels have been detected in various tissues undergoing stress, and we previously reported that clusterin is markedly induced in media and neointima following vascular injury. The present study therefore investigated the impact of clusterin on neointimal hyperplasia following vascular injury.
Methods and Results: As compared with wild-type mice, clusterin knockout mice (clusterin-KO) demonstrated a significant decrease of the intima/media ratio 4 weeks after cuff placement. Immunohistochemical analysis of injured femoral arteries in clusterin-KO demonstrated the accumulation of p53 in nuclei of neointimal vascular smooth muscle cells (VSMCs). Moreover, VSMCs from either clusterin-KO or rat VSMCs treated with clusterin-short-interfering (si) RNA subjected to static stretch exhibited significantly increased p53 and p21, and increased G1 cell cycle arrest as indicated by flow cytometry compared with VSMCs from wild-type mice.
Conclusion: Reduced clusterin expression reduced the proliferation of VSMCs and induced G1 arrest via p53 and p21. Clusterin therefore represents a promising molecular target to limit restenosis after coronary intervention.

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Aim: We investigated the relationship between coronary risk factors and the intima-media thickness (IMT) and wall elasticity of carotid arteries. A new real-time ultrasonic measurement system that allows measurement of the elasticity of the carotid arterial intima-media complex was applied to 350 subjects, who were also checked for the presence/absence of hypertension, dyslipidemia, diabetes mellitus and regular smoking. Simultaneous measurement of the elastic modulus and IMT was conducted at four sites in the bilateral carotid arteries.
Methods and Results: In the group with maximum IMT (max IMT) <1.1 mm, the IMT, as well as the mean elastic modulus in the circumferential direction (Eθ), showed positive correlations with age and coronary risk factors. Multiple regression analysis showed that age, systolic blood pressure and pulse rate remained independent determinants of Eθ. The number of criteria fulfilling the diagnosis of metabolic syndrome showed a good positive correlation with the value of Eθ in the group with max IMT values <1.1 mm.
Conclusion: Measurement of carotid arterial wall elasticity together with IMT is useful to detect dis-tortions in intramural elasticity distribution occurring in the early stages of atherosclerosis.

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Aim: Lower levels of high-density lipoprotein cholesterol (HDL-C) have been recognized as a risk factor for coronary artery disease (CAD), but the relationship between HDL-C values and the occurrence of CAD has not been fully established in the Japanese general population.
Methods: A cohort study of 5,371 Japanese men with 12 years of follow-up was conducted to identify risk factors for the occurrence of CAD.
Results: One hundred and twelve subjects had CAD (acute myocardial infarction in 67 patients and angina in 45 patients) during the follow-up period. Adjustment for variables including age, body mass index, smoking habit, alcohol intake, systolic blood pressure, low-density lipoprotein cholesterol, triglyceride, fasting plasma glucose, hypertension, diabetes mellitus and hyperlipidemia, adjusted hazard ratio (HR) of lower levels of HDL-C for the occurrence of CAD was 1.21 (95% confidence interval (CI): 1.11-1.33, p<0.001). Serum HDL-C concentration less than 51 mg/dL was a significant risk for CAD.
Conclusions: Low HDL-C was identified as a significant and independent risk for CAD in Japanese men using long-term follow-up data.

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Aims: In Japan, it is believed that guidelines for lifestyle-related disease are used in routine clinical practice, however, there are few reports on the actual rate of healthcare conducted in accordance with these guidelines by general practitioners and on their usefulness in preventing cardiovascular events. Therefore, the Heart Care Network (HCN) groups were organized mainly by general practitioners treating lifestyle diseases in 62 areas of Japan.
Methods: The HCN has collected data on lifestyle diseases in high-risk patients in routine practices and investigated management conditions, guideline target achievement rates and medication. Additionally, the incidence of cardiovascular events was assessed.
Results: We analyzed 14,064 cases. The lipid profile, blood pressure, glycemic control were significantly improved over the 3 years. The incidence of cardiovascular events were significantly reduced by the achievement of target LDL cholesterol, systolic blood pressure and hemoglobin A1c and even after adjustment for age, gender, history of myocardial infarction, the reduction of these lifestylerelated parameters remains significant.
Conclusion: These results revealed the current trends in the healthcare activities of general practitioners, the management conditions for lifestyle diseases in CHD high-risk patients and their effects on reducing cardiovascular events.

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Aim: We investigated thrombus formation at the site of functional injury to endothelial cells by FeCl₃.
Methods: After preparation of cremasteric arteries of mice, controlled endothelial injury was induced by application of FeCl₃. Endothelial cells were rendered fluorescent by addition of FITC (fluorescein isothiocyanate)-labeled isolectin B4. Circulating platelets and leukocytes were made fluorescent by rhodamine 6G. Three-dimensional (3D) growth of thrombi was visualized in real time. Effects of aspirin and clopidogrel pre-treatments on the growth of thrombi were investigated in vivo as well as in an ex vivo flow chamber system.
Results: Endothelial cells were tightly bound to each other to protect local thrombus formation. Platelets started to adhere to endothelial cells when FeCl₃ was applied. Three-dimensional growth of thrombi, which takes 10.6±7.5 minutes for complete occlusion in control, can be visualized with our imaging system. Aspirin pre-treatment at the dose tested did not influence either endothelial injury or platelet thrombus growth, while clopidogrel pretreatment significantly inhibited 3D growth and prolonged occlusion time up to 64.6±25.3 minutes (100 mg/kg). A similar inhibiting effect of clopidogrel was reproduced in ex vivo flow chamber experiments.
Conclusions: We have developed an in vivo system to detect thrombus formation after functional damage to the endothelium.

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Aim: to investigate the association of C677T polymorphism in the methylene tetrahydrofolate reductase (MTHFR) gene, homocysteine plasma values (Hcy), and plasma HDL cholesterol in heterozy-gous familial hypercholesterolemia (hFH).
Methods: One hundred and twenty-five hFH subjects were studied. Plasma lipid, lipoprotein, vitamin B12, folic acid and Hcy values were determined. C677T polymorphism in the MTHFR gene was detected by SSCP-PCR. Genetic diagnosis of FH was determined by a three-step protocol using SSCP-PCR, Southern blot, long PCR and automatic sequencing.
Results: We found significant differences in plasma HDL-C (CC 1.39±0.34, CT 1.33±0.39 and TT 1.14±0.26 mmol/L, p=0.028) between the C677T MTHFR genotypes, that were also found when gender age, and BMI were included as covariables. In addition, Hcy values were significantly different between C/T MTHFR genotypes (CC 11.75±2.9, CT 12.69±2.88, TT 15.34±2.1 μmol/L). The distribution of gender, smoking habit and LDLR gene mutations was similar among the three groups.
A significant correlation was found between Hcy plasma values and plasma HDL-C (-0.370, p= 0.003), but no correlations were found with age, BMI or other lipid and apo B plasma values.Conclusion: In hFH subjects, the genotype TT and higher plasma Hcy levels were associated with lower HDL-C plasma values in FH subjects. More studies are needed to confirm our results and also to elucidate the exact mechanism of interaction between plasma homocysteine and lipid metabolism.

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Aim: To evaluate carotid intima-media thickness (IMT) in patients with long-term early-onset type 1 diabetes, and investigate the associations between IMT, diabetic retinopathy and clinical characteristics.
Methods: We evaluated anthropometric measurements, biochemical parameters and carotid IMT. Ultrasonography was performed on 73 patients diagnosed with type 1 diabetes before 30 years of age, and who have been living with diabetes for 20 years or more.
Results: The mean max-IMT (maximal thickness of whole carotid artery) and IMT-Cmax (maximal thickness of common carotid artery) values were 0.94 mm and 0.67 mm; 21 patients had proliferative diabetic retinopathy and 21 patients had plaque (IMT ≥1.1 mm). Age, age at diagnosis of diabetes and adolescent HbA_1C level (HbA_1C at 18 years old or the earliest measurement available) were higher in patients with plaque than in those without. Max-IMT was greater in patients with proliferative retinopathy than in those without. Similarly, there were significant differences in current HbA_1C level, and the prevalence of hypertension and dyslipidemia. In multivariate analysis, age and dyslipidemia were independently associated with max-IMT and IMT-Cmax.
Conclusions: Age and dyslipidemia were associated with IMT. In contrast, glycemic control was closely associated with diabetic retinopathy, but weakly associated with IMT.

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Aim: Scavenger receptors type I and II (SRBI/II) have dual roles in both atherogenic and antiatherogenic functions through interactions with lipoproteins and their expression in macrophages; how-ever, the distribution and density of SRBI/II-positive macrophages and smooth muscle cells (SMCs) as well as their association with lipid metabolism-related proteins in atherosclerotic intima of the human aorta remain unclear.
Methods: Autopsied aortic tissues were double-immunostained with SRBI/BII and smooth muscle actin or macrophage-specific antibodies. The density of SRBI/BII-positive SMCs and macrophages in intimal lesion was measured. They were also immunostained with antibodies against four apolipoproteins, four phospholipase A2s, and CETP.
Results: SRBI/II was expressed in both macrophages and SMCs distributed in various intimal lesions. The density of SRBI/II-positive SMCs in intimal lesions significantly decreased with the advance of atherosclerosis, whereas the density of SRBI/II-positive macrophages significantly increased with atherosclerotic development. In addition, functional proteins, such as apolipoproteins, secretory phospholipase A2s, and CETP, were distributed in the intimal stroma around SRBI/II-positive cells in all lesion types.
Conclusion: The results indicated that SMCs are involved in lipid metabolism via SRBI/II expression mainly in the early stages of atherosclerosis evolution, and that SRBI/II-positive macrophages are mainly involved in advanced stages.

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Aim: Chronic kidney disease (CKD) is associated with an increased risk of cardiovascular disease, although it has yet to be established whether CKD is an independent risk factor for arterial stiffness in community residents. The purpose of this study was to determine the correlation between the cardio-ankle vascular index (CAVI) and estimated glomerular filtration rate (eGFR) in the general population.
Methods: We studied 881 consecutively enrolled subjects undergoing health checkups. CAVI was calculated automatically from the pulse volume record, blood pressure and the vascular length from the heart to the ankle. CKD was evaluated by the eGFR.
Results: The distribution of eGFR was as follows: 241 with eGFR (mL/min/1.73m²) ≥90; 572 with eGFR 60-89; 65 with eGFR 30-59; 3 with eGFR 15-29; 0 with eGFR <15. Linear regression analysis showed that CAVI was negatively correlated significantly with eGFR, while multiple regression analysis using CAVI as an objective variable, adjusted for conventional atherosclerotic risk factors and eGFR as explanatory variables, demonstrated that CAVI was an independent determinant of eGFR. We also showed that stepwise increments of CAVI occurred with progressive deterioration of CKD.
Conclusion: CAVI was independently correlated with eGFR indicating that CKD is associated with arterial stiffness in the general population.

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Aim: The activation of RhoA and Rac1 is crucial for the pathogenesis of atherosclerosis. This study investigated the changes of unprocessed and mature forms of RhoA and Rac1 in the progression of atherosclerosis.
Methods: Unprocessed and geranylgeranylated forms of RhoA and Rac1 in aortic atherosclerotic lesions were separated by the Triton X-114 partition method using Watanabe heritable hyperlipidemic (WHHLMI) rabbits prone to myocardial infarction. The activation of RhoA and Rac1 was determined by membrane translocation and pull-down assays.
Results: The levels of unprocessed RhoA and Rac1 of the aortas were higher at 7 months than 3 months, accompanied by increased levels of total RhoA and Rac1. Membrane-bound RhoA and Rac1 levels of the aortas at 7 months were significantly increased compared with those at 3 months, consistent with the results of GTP-loading. Unprocessed and activated forms of RhoA and Rac1 had gradually decreas at 15 and 24 months compared to 7 months.
Conclusions: We show evidence of marked increases in unprocessed RhoA and Rac1 with enhanced activities in the progression of atherosclerosis in WHHLMI rabbits. This is important for better understanding of the pathogenesis of hyperlipidemia-dependent atherosclerosis.

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Aim: In 2003, the American Diabetes Association recommended that the threshold for diagnosing impaired fasting glucose (IFG) should be lowered from 6.1 mmol/L (110 mg/dL) to 5.6 mmol/L (100 mg/dL). To discuss the diagnostic threshold for IFG, the association between fasting plasma glucose (FPG) and the risk of future diabetes must be known; however, data regarding this relation in the Japanese population are scarce. The aim of this study was to determine the relation between FPG and the risk of future diabetes in the Japanese general population.
Methods: A retrospective cohort study was conducted using data from annual health check-ups performed in Omiya city. A total of 11,369 subjects between the ages of 40-79 years who were not dia-betic at baseline were followed for seven years. Diabetes was defined as FPG ≥126 mg/dL or self-report.
Results: The incidence of diabetes increased as the baseline FPG level increased and a similar pattern was observed irrespective of sex or age. The hazard ratios compared with subjects with FPG <85 mg/ dL adjusted for possible confounding factors were 3.83 (95% confidence interval (95% CI); 2.41-6.08) for subjects with 100 to 104 mg/dL FPG and 7.97 (95% CI; 4.98-12.4) for subjects with 105 to 109 mg/dL FPG.
Conclusions: Subjects with 100-109 mg/dL FPG have an appreciable risk of diabetes that cannot be considered as “normal” and should be notified of their potential risk of developing diabetes.

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Aim: Obstructive sleep apnea (OSA) is a risk factor for cardiovascular diseases. Platelets play key roles in the development of atherothrombosis. Several studies assessing platelet activation in patients with OSA have been published; however, there have been only a few studies with a small number of patients with OSA investigating platelet aggregability, which evaluates platelet aggregation more directly than the platelet activation status. We aimed to investigate the effects of OSA and nasal continuous positive airway pressure (nCPAP) therapy, a well-established treatment for OSA, on platelet aggregability.
Methods and Results: We examined 124 consecutive patients with snoring in whom the 3% oxygen desaturation index (3%ODI), a severity marker of OSA, and ADP- and collagen-induced platelet aggregability measured with the optical aggregometer were analyzed. ADP-induced platelet aggre-gability was increased more in patients with moderate-to-severe OSA (3%ODI>15) than in patients with non-to-mild OSA (p=0.029). In multiple linear models, 3%ODI significantly contributed to increased platelet aggregability induced by both ADP and collagen among 59 subjects with one or more risk factors for vascular diseases, such as smoking, hypertension, diabetes mellitus or hyperlipidemia. In 23 patients treated by nCPAP, collagen-induced platelet aggregability was ameliorated on Day 90, compared to at the baseline.
Conclusion: The severity of OSA significantly contributed to platelet aggregability, which was improved by nCPAP treatment partially at three months.

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Aim: Statins are effective in lowering cholesterol levels, but cause fatal rhabdomyolysis in susceptible individuals. Because it has been hypothesized that muscle damage could result from alterations in Ca²⁺ homeostasis in muscle cells, we tested whether measuring statin-induced changes in intracellular calcium ([Ca²⁺]_i) is useful for predicting susceptibility to statin-muscle damage, using human CD19+ primary B lymphocytes.
Methods: Statin-induced alterations in [Ca²⁺]_i were studied using the human THP-1 cell line and CD19+ primary B lymphocytes. Changes in [Ca²⁺]_i were measured directly in fluo-3- loaded cells using either single or dual-color flow cytometry.
Results: The Ca²⁺ release study suggested that statin-induced changes in [Ca²⁺]_i were due to Ca²⁺ release from ryanodine-sensitive Ca²⁺ stores and mitochondrial compartments. Further, statin users who experienced elevated creatine kinase (n=8) exhibited significantly greater statin-induced Ca²⁺ release in B cells than healthy volunteers (n=45) and statin users without elevated creatine kinase (n=16), while no difference was seen between the latter two groups.
Conclusion: Statin-induced Ca²⁺ release from ryanodine-sensitive stores and mitochondria may contribute to myotoxicity. The laboratory test for Ca²⁺ release using CD19+ primary B lymphocytes may be useful to predict susceptibility to statin-induced muscle toxicity prior to statin use.

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Aim: Platelet-derived microparticles (PDMPs) play roles in normal hemostatic responses to vascular injury because they possess prothrombinase activity. Although the most widely used method for studying PDMP is flow cytometry, we previously developed an enzyme-linked immunosorbent assay (ELISA) method as an easier and more reproducible PDMP assay. The purpose of this study was to use various clinical settings to verify whether this ELISA method can produce equivalent results to flow cytometry for PDMP.
Methods: We performed a large-scale clinical study for various thrombotic and subatherothrombotic diseases using an ELISA kit. The study group included 692 patients with cerebral infarction, heart failure, acute coronary syndrome or diabetes mellitus.
Results: When baseline PDMP values in the various diseases were compared with those in healthy controls, significant differences were noted in all cases. There were significantly elevated levels of PDMPs in diabetic patients with complications but no thrombosis. When baseline PDMP values in cerebral infarction were compared within the subclassifications, atheroma and other types of infarction exhibited significantly elevated PDMP levels compared with lacunar infarction. Cerebral infarction exhibited a significant change in PDMPs after therapy compared with the baseline (before therapy), but not in acute coronary syndrome and heart failure. The ELISA method exhibited results almost identical to flow cytometry for PDMP in various atherothrombotic diseases.
Conclusion: Although further examinations to evaluate the therapeutic usefulness of these diseases are necessary, ELISA kits possibly represent a new tool for PDMP related to atherothrombosis.

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Aim: Bitter melon (Momordica charantia L.) is a common vegetable grown in Okinawa that has also been used recently in medicine for the treatment of diseases such as diabetes, hypertension, and dyslipidemia. Among Bitter melon extracts compounds, we focused on an extract known as momordin in the present study, to examine its effect on peroxisome-proliferator activated-receptor (PPAR) δ (also called PPARδ in rodents) expression and promoter activity of the human PPARδ gene.
Methods: A human PPARδ promoter-reporter plasmid was made as a template from a BAC CLONE (RPCI-11C) containing a -3076 bp (BglI site) +74 bp (EcoRI site) sequence. Luciferase assay of PPARδ promoter activity was performed using HepG2 cells.
Results: 10 and 25 nM Momordin significantly increased the expression of PPARδ mRNA 1.5-fold (relative to the control). Moreover, 10 and 25 nM Momordin significantly increased PPARδ promoter activity in a dose-dependent manner, reaching more than 1.5-fold relative to the control.
Conclusion: Our present data obtained through successful cloning of the PPARδ promoter demonstrate that PPARδ production and activation are upregulated through PPARδ promoter activity following momordin treatment.

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Aims: Perilipin and adipophilin, PAT family proteins, play important roles in lipid metabolism. Although nonalcoholic fatty liver disease (NAFLD) is initiated by hepatocyte lipidation, little is known about the relationship between these proteins and hepatocellular injury. We investigated the expressions of perilipin and adipophilin and their relation to inflammation, fibrosis, hepatocellular ballooning, and oxidized phosphatidylcholine (oxPC) localization in human NAFLD.
Methods and Results: Liver biopsies of nonalcoholic steatohepatitis (NASH, n=39) or simple steatosis (n=9) were studied by immunohistochemical techniques using anti-perilipin, anti-adipophilin and anti-oxPC antibodies. The severity of liver damage was histologically assessed by the Brunt system and NAFLD activity score (NAS). Enlarged hepatocytes usually containing Mallory-Denk bodies were defined as ballooned. Perilipin and adipophilin were detected on the rim of lipid droplets in both NASH and simple steatosis. Perilipin was more evident in larger lipid droplets while adipophilin expression was frequent in lipid droplets of ballooned hepatocytes. The frequency of adipophilin-positive ballooned hepatocytes was correlated to inflammation (Rs=0.72, p<0.0001), fibrosis (Rs=0.46, p=0.005), NAS (Rs=0.47, p=0.004) and oxPC-positive ballooned hepatocytes (Rs=0.35, p=0.033).
Conclusions: Expression patterns of perilipin and adipophilin in NASH livers varied with the size of lipid droplets. In partiew or, adipophilin expression in ballooned hepatocytes was closely associated with oxidative damage.

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Aim: The aim of this study was to evaluate the usefulness of glutamate pyruvate transaminase (GPT) levels in the diagnosis of metabolic syndrome (MS) in obese Japanese children.
Methods: We examined 193 obese boys (mean age: 12.1 yrs; mean percent overweight [POW]: 53.9%) and 37 obese girls (mean age: 11.4 yrs; mean POW: 57.2%). Anthropometric measurements, blood pressure and levels of liver transaminases, serum lipids and lipoproteins, fasting blood glucose (FBG), serum insulin and adiponectin were measured. The subjects were divided into either an MS or a non-MS group according to the MS definition criteria for Japanese children.
Results: The level of GPT was significantly higher in the MS group in both genders. Correlation analysis revealed positive correlations between GPT and waist circumference, blood pressure, maximum preperitoneal fat thickness, serum insulin and homeostasis model assessment-insulin resistance (HOMA-R), but no correlation between GPT and FBG. ANOVA showed a significant difference in GPT levels between MS and non-MS subgroups, whereas there was no difference in FBG between the two groups. Receiver operating characteristic curves demonstrated that GPT was clearly superior to FBG as a diagnostic marker of MS.
Conclusion: We conclude that an elevation in GPT in obese children most likely reflects insulin resistance and that GPT is superior to FBG as a marker of MS.

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Aim: Recently, a novel device for measuring the cardio-ankle vascular index (CAVI) as an arterial stiffness parameter has been developed. In this study, we evaluated the effect of angiotensin II receptor blocker (ARB) and calcium channel (Ca) blocker on CAVI in type 2 diabetic patients with hypertension.
Methods: Seventy type 2 diabetes mellitus patients with hypertension were enrolled and randomly divided into two groups. One group was administered olmesartan medoxomil 20 mg/day for 12 months (ARB group), and the other group was administered amlodipine besilate 5 mg/day for 12 months (Ca blocker group).
Results: In the ARB group, a significant decrease in CAVI was observed after 12 months; however, no significant change in CAVI was observed in the Ca blocker group although changes in blood pressure were almost the same. By simple regression analyses, CAVI changes correlated positively with 8-OHdG changes.
Conclusions: Olmesartan, an ARB, improved arterial stiffness more than amlodipine, and this effect might be due to not only the blood pressure-lowering effect but also to reducing the potential of oxidative stress recognized in olmesartan.

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