Aim: To determine whether high plasma levels or activities of different hemostatic proteins contribute to the development of early atherosclerotic vessel wall changes. Elevated levels of various hemostatic proteins and markers of inflammation have been linked to an increased risk of ischemic cardiovascular events; however, the mechanisms by which these molecules might contribute to this increased risk is not clear.
Methods: The intima-media thickness of the common carotid arteries (CCA-IMT) of 125 healthy young volunteers without known cardiovascular risk factors was measured by high-resolution ultrasound. Plasma concentrations of fibrinogen, thrombomodulin, protein Z and CRP were quantified, and the plasma activities of protein C, plasminogen and factor VIII were measured. Other established risk factors, such as body mass index (BMI) and plasma levels of cholesterol, triglycerides and homocysteine, were also determined. Furthermore, the carotid arteries were examined for the presence of plaques and stenoses.
Results: Univariate analysis showed a significant negative correlation between CCA-IMT and HDL cholesterol, and positive correlations with age, BMI, LDL cholesterol, triglycerides, homocysteine, fibrinogen and thrombomodulin, but not with total cholesterol, lipoprotein(a) and hsCRP. CCA-IMT was also statistically independent of the activities of protein C, factor VIII and plasminogen. Multivariate analysis revealed a significant correlation of CCA-IMT with age, BMI and fibrinogen.
Conclusion: Our data suggest that fibrinogen levels correlate with early atherosclerotic changes of the carotid artery in a population with very low cardiovascular risk.

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Aim: The abnormal proliferation of vascular smooth muscle cells (VSMCs) in arterial walls is an important pathogenic factor of vascular disorders such as atherosclerosis and restenosis after angioplasty. During atherogenesis or in response to vessel injury, VSMC proliferation is induced by a number of peptide growth factors released from platelets and VSMCs. Cilostazol is a phosphodiesterase (PDE) 3 inhibitor that increases intracellular cAMP levels and decreases intracellular Ca²⁺ levels, inhibiting platelet aggregation and inducing vasodilatation. Cilostazol is also known to have an inhibitory effect on the proliferation of VSMCs, but the anti-proliferative mechanism of cilostazol in VSMCs has not yet been established. In the present study, we investigated whether the anti-proliferative mechanism of cilostazol is associated with the suppression of extracellular signal-regulated kinases (ERK) and phosphatidylinositol 3 kinase (PI3K) signaling pathways.
Methods: To confirm the anti-proliferative effects of cilostazol on VSMCs, VSMCs were induced to proliferate by serum-induced mitogenesis and then were treated with cilostazol for 24 h. And, to investigate whether the anti-proliferative mechanism of cilostazol in VSMCs involves the suppression of the ERK and PI3K pathways, expression of the phosphorylated forms of ERK1/2, Raf, Akt, and glycogen synthase kinase (GSK)-3 were evaluated by western blot.
Results: Cilostazol inhibited VSMC proliferation in a dose-dependent manner. Phosphorylated ERK1/2 and Raf were significantly reduced in a dose-dependent manner, whereas phosphorylated Akt and GSK-3 were not changed.
Conclusion: These results suggest that suppression of the ERK pathway but not the PI3K pathway is an important mechanism in the anti-proliferative effect of cilostazol on VSMCs.

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Aim: Present study aimed to elucidate the suppression of serum lipids by gamma- and delta-tocotrienol (γδT3).
Methods: The lipid-lowering effects of γδT3 were investigated using HepG2 liver cell line, hypercholesterolemic mice and borderline-high cholesterol patients.
Results: In-vitro results demonstrated two modes of action. First, γδT3 suppressed the upstream regulators of lipid homeostasis genes (DGAT2, APOB100, SREBP1/2 and HMGCR) leading to the suppression of triglycerides, cholesterol and VLDL biosyntheses. Second, γδT3 enhanced LDL efflux through induction of LDL receptor (LDLr) expression. Treatment of LDLr-deficient mice with 1 mg/day (50 mg/kg/day) γδT3 for one-month showed 28%, 19% reduction in cholesterol and triglyceride levels respectively, whereas HDL level was unaltered. The lipid-lowering effects were not affected by alpha-tocopherol (αTP). In a placebo-controlled human trial using 120 mg/day γδT3, only serum triglycerides were lowered by 28% followed by concomitant reduction in the triglyceriderich VLDL and chylomicrons. In contrast, total cholesterol, LDL and HDL remained unchanged in treated and placebo groups. The discrepancies between in-vitro, in-vivo and human studies may be attributed to the differential rates of post-absorptive γδT3 degradation and LDL metabolism.
Conclusion: Reduction in triglycerides synthesis and transport may be the primary benefit caused by ingesting γδT3 in human.

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Aim: Recent studies have suggested that hyperinsulinemia is associated with high cardiovascular risk. The purpose of this study was to assess the relationship between the serum insulin level and coronary artery calcification (CAC).
Methods: We performed a cross-sectional study of 582 consecutive and nondiabetic participants with clinical suspicion of coronary heart disease, and assessed the CAC score determined by multislice computed tomography. A standard 75-g oral glucose tolerance test was performed and venous blood was collected at 0, 30, 60 and 120 min for the measurement of plasma glucose and serum insulin. Statistical analyses were conducted for 4 subgroups according to fasting insulin and insulin area under the concentration time curve (InsAUC).
Results: Mean log (CAC+1) and InsAUC were 1.6 and 109.1 μIU/mL, respectively. Unadjusted analysis demonstrated that the fasting insulin quartiles (p=0.0256) and InsAUC quartiles (p<0.0001) were significantly associated with log (CAC+1), and the lowest fasting insulin quartiles (p<0.0001) and the lowest InsAUC quartile (p=0.0006) had lower glucose AUC. Analysis of covariance demon-strated that the lowest InsAUC quartile had the highest log (CAC+1), and the highest InsAUC quar-tile had a higher log (CAC+1) than the second and third InsAUC quartiles, adjusted for several coro-nary risk factors (p<0.0001).
Conclusion: The lowest InsAUC quartile was related to CAC, although the lowest InsAUC quartile maintained glucose homeostasis, in this study population. Not only hyperinsulinemia but also a low insulin level are independently associated with CAC.

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Aim: Helicobacter Pylori infection was associated with the risk of cardiovascular disease; however, the relation between Helicobacter Pylori infection and the lipid profile has not been fully established.
Methods: We measured anti-Helicobacter Pylori antibody concentration and lipid profiles in 6,289 Japanese subjects aged 21-64 years (5,077 male and 1,212 female).
Results: The prevalence of Helicobacter Pylori-seropositive subjects was 46.8% and 39.6% in men and women, respectively. Adjusted mean values of low-density lipoprotein (LDL) cholesterol and high-density lipoprotein (HDL) cholesterol in men were significantly higher and lower in Helicobacter Pylori-seropositive than-negative subjects, respectively, (LDL-cholesterol: 129.0±0.8 vs. 125.3±0.7 mg/dL, p< 0.001, HDL-cholesterol: 54.6±0.3 vs. 56.6±0.3 mg/dL, p< 0.01), whereas these associations were not significant in female subjects. Moreover, the odds ratio of Helicobacter Pylori infection for high LDL-cholesteremia and low HDL-cholesteremia in male subjects was 1.23 (95% CI (confidence interval): 1.08-1.40, p< 0.05) and 1.29 (95% CI: 1.03-1.59, p< 0.05), respectively. Female subjects did not have such associations.
Conclusions: The present study demonstrates that Helicobacter Pylori infection is significantly associ-ated with high LDL-cholesteremia and low HDL-cholesteremia in Japanese male subjects.

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Aim: The effects of statins on the distribution of oxidized LDL in plasma LDL subfractions have not been well defined. Effects of 12-month treatment with low-dose simvastatin on the distribution of cholesterol and oxidized LDL in 3 ultracentrifugally separated plasma LDL subfractions were compared in patients with hypercholesterolemia.
Methods: Simvastain was administered to 30 hypercholesterolemic subjects for 12 months at an initial dose of 5 mg/day, which was increased to 20 mg/day via 10mg/day to decrease plasma LDL-cholesterol (C) lower than 130 mg/dL. Simvastatin dose was fixed after 3 months of treatment. The amounts of cholesterol and oxidized LDL in 3 ultracentrifugally separated plasma LDL subfractions were compared between 0 and 12 months of treatment.
Results: The distribution of ox-LDL skewed to denser LDL fractions, compared with cholesterol in plasma LDL subfractions. Plasma cholesterol in low-density LDL, medium-density LDL and high-density LDL decreased significantly by 31%, 30%, and 25%, respectively (p<0.0001) after 12 months of simvastatin treatment. Plasma oxidized LDL was decreased from 70 U/L to 56 U/L in medium-density LDL (p=0.042). Oxidized LDL in low-density LDL and high-density LDL did not change significantly after 12 months of treatment.
Conclusion: Treatment with low-dose simvastatin decreased plasma cholesterol in 3 LDL subfractions and oxidized LDL in medium-density LDL. The decrease of oxidized LDL seemed to be not due to the decrease of cholesterol in plasma LDL subfractions because the decreasing patterns of cholesterol and ox-LDL were different in 3 LDL subfractions.

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Aim: Peripheral arterial disease (PAD) is a common cause of cardiovascular morbidity and an inde-pendent predictor of cardiovascular mortality. However, little is known about the genetic basis of PAD. To elucidate this, we performed a two-staged genome-wide association study in Japanese indi-viduals.
Methods: We initially tested 222,285 single-nucleotide polymorphisms (SNPs). After the first screen-ing in a panel of 195 PAD cases and 1,358 controls, 2,696 SNPs (1.2%) were further genotyped in the second screening using another panel of 699 PAD cases and 1540 controls. In both screenings, controls were subjects affected with some diseases other than PAD.
Results: When analyzed in the combined panel, the strongest signal of PAD association was observed at rs1902341 in the intron of OSBPL10 (p=4.7E-7 for trend test; OR=1.31, 95% CI 1.18-1.46). Also, PAD was modestly associated at several other loci such as rs2554503 in CSMD1 (p=5.7E-5; OR=1.32, 95% CI 1.15-1.51) or rs235243 in VSP13D (p=0.04; OR=1.18, 95% CI 1.01-1.37).
Conclusion: Our genome-wide exploration identified suggestive evidence of PAD association at the OSBPL10 locus. Because the association has not reached a genome-wide significant level, further replication study is warranted for verification in the Japanese population.

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Aim: A reduced risk of type 2 diabetes has been reported following treatment with pravastatin. Adiponectin is an adipocyte-derived protein that has an antidiabetic property. The objective of this study was to evaluate the effect of pravastatin on serum adiponectin concentration and other influencing factors.
Methods: This study was a multicenter observational study: Dyslipidemia Open-labeled observa-tional study by Lipid-lowering therapy with Pravastatin of the effect on High-molecular weight adi-ponectin in Nippon Yokohama (DOLPHIN). The protocol was registered in the UMIN Clinical Trial Registry as UMIN000000791. All patients received pravastatin 10 mg/day for 6 months and the change in concentration of total and high molecular weight adiponectin was assessed before and after follow-up. The difference in the change in total adiponectin concentration by patient characteristics was analyzed by an unpaired t-test. Influences of continuous variable factors on the change in total adiponectin concentration were estimated by simple linear regression analyses. Finally, in order to estimate the influences of factors that potentially affect the change in total adiponectin concentration induced by pravastatin, multiple linear regression analysis was conducted.
Results: After 6 months, total adiponectin concentration was increased significantly by 23.2% from 11.7±6.4 to 13.7±8.6 μg/mL (p=0.002). The use of thiazolidinedione as a concomitant medication was the only significant influencing factor (β=0.580, p<0.001).
Conclusion: Pravastatin increased the serum adiponectin concentration in Japanese dyslipidemic patients without previous coronary artery disease. Interestingly, this effect was seen synergistically in combination with thiazolidinedione.

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Aim: High cholesterol absorption in the small intestine has been proposed to be a risk factor of atherosclerosis. In this study, we evaluated the effect of ezetimibe monotherapy on arterial stiffness in type 2 diabetic patients.
Methods: Forty type 2 diabetes mellitus patients with high serum low-density lipoprotein cholesterol (LDL-C) were enrolled and treated with ezetimibe 10 mg/day for 6 months. HbA1c, serum lipids, remnant-like particle-cholesterol (RLP-C), serum lipoprotein lipase mass (LPL mass) and the cardioankle vascular index (CAVI) were measured before and after ezetimibe treatment.
Results: After 6 months of ezetimibe treatment, significant decreases in LDL-C, RLP-C and CAVI were observed. In the group that achieved the LDL-C goal of <120 mg/dL after 6 months of ezeti-mibe treatment, the pretreatment CAVI was markedly high, and CAVI decreased significantly after ezetimibe treatment.
Conclusions: In type 2 diabetic patients, ezetimibe monotherapy may have the potential to ameliorate arterial stiffness in addition to lowering LDL-C and RLP-C.

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Aim and Methods: We assessed the relationship of the body mass index (BMI) of 187 college female students aged 18 years with the reported BMI of their middle-aged biological parents measured on 2 occasions: when the parents were 18-20 years old and at the time of the study. The relationships of fat mass measured using whole body dual energy X-ray absorptiometry (DXA) and serum leptin levels were also determined between 148 daughters and middle-aged parents (148 mothers and 59 fathers).
Results: The BMI of daughters was associated with their mothers' BMI (r=0.30, p<0.0001) but not with their fathers' BMI measured when they were 18 years old. Daughters' BMI showed a stronger association with the current BMI of their mothers BMI (r=0.36, p<0.0001) than that of their fathers' BMI (r=0.19, p=0.01). In addition, the serum leptin levels of daughters were correlated with their mothers' leptin values (r=0.22, p=0.04). Further, not only total body fat mass (r=0.19, p<0.05) but also fat mass in the trunk (r=0.18, p<0.05) and legs (r=0.17, p<0.05) was associated between daughters and their mothers.
Conclusion: The significant correlation between daughters' and mothers' BMI measured when their mothers were 18 years old did not result from shared environmental factors, including the intrauterine environment. The results in the present study therefore suggest that adiposity in 18-year-old daughters may be influenced by the maternal effect. The associations of serum leptin and DXA-derived fat mass between daughters and their mothers may support our hypothesis.

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Aims: This study investigated the association between dietary glycemic index (GI)/glycemic load (GL) and serum lipids in middle-aged Japanese men and women.
Methods: The study participants were employees of a metal products factory in Japan: 2,257 men and 1,598 women aged 35 years or older. Dietary GI and GL were assessed using a self-administered diet history questionnaire. Serum lipid levels, adjusted for age, body mass index, alcohol consumption, smoking, physical activity, menopause status, and dietary intake of total energy, saturated fatty acids, monounsaturated fatty acids, polyunsaturated fatty acids, cholesterol and fiber, were compared among GI/GL quintiles for each gender.
Results: No significant associations were observed between GI and adjusted serum lipids in men or women. In contrast, GL was inversely associated with HDL-cholesterol in men and women (p for trend=0.001 for men and < 0.001 for women), and positively associated with non-HDL-cholesterol (p for trend=0.010), LDL-cholesterol (p for trend=0.035) and triglycerides (p for trend=0.011) in women; however, alcohol drinking affected these associations; there was no association between GL and serum lipids in male nondrinkers and between GL and LDL-cholesterol in female nondrinkers.
Conclusion: GL was inversely associated with HDL-cholesterol and positively associated with non-HDL-cholesterol in Japanese women. These associations in men were not observed in nondrinkers. A high-GL diet for women may have an atherogenic effect through these serum lipid abnormalities.

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Aim: We have shown that aggressive lipid lowering by pitavastatin and atorvastatin results in marked regression of atherosclerotic coronary lesions after acute coronary syndrome (ACS). The purpose of this study was to address the association of lipid levels after statin therapy with regression of atherosclerotic coronary lesions and major cardiovascular events in patients after ACS.
Methods: JAPAN-ACS is a prospective, randomized open-label study performed at 33 centers in Japan. Patients with ACS undergoing intravascular ultrasound (IVUS)-guided percutaneous coronary intervention (PCI) were randomly assigned to receive either 4 mg/day pitavastatin or 20 mg/day atorvastatin within 72 hours after PCI. IVUS image was obtained in 251 patients, including 73 diabetic patients. Lipid profiles at the end of the study were divided into quartiles and the association with the percent change in non-culprit coronary plaque volume (PV) was assessed in total and diabetic patients. We also studied whether baseline and follow-up levels of HDL-cholesterol are associated with restenosis after PCI.
Results: Decreasing LDL-cholesterol, non-HDL-cholesterol, LDL-C/HDL-C ratio, apolipoprotein B quartiles were associated with a progressively smaller plaque burden in total and diabetic patients. In diabetic patients, further reduction of these parameters was associated with a significantly greater reduction in PV. We also found that patients with lower HDL-cholesterol had a significantly higher incidence of target lesion revascularization.
Conclusions: Early intensive statin therapy in patients after ACS results in remarkable regression of coronary PV. Diabetic patients can have a benefit with more intensive therapy to achieve a lower target level in Japanese.

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