Aim: Inflammatory cell recruitment and intimal neovascularization contribute to atherosclerotic plaque destabilization. The anti-inflammatory red wine polyphenol, resveratrol, has been implicated in cardiovascular protection. In this study, we investigated the effects of resveratrol on endothelial and monocytic cell migration.
Methods: Human umbilical vein endothelial cell (EC) migration was assessed in a modified barrier assay. Chemotaxis of THP-1 monocytic cells towards monocyte chemoattractant protein (MCP)-1 was determined using a Boyden chamber. Erk phosphorylation downstream of MCP-1 receptor and activation of myosin phosphatase targeting subunit 1 (pMYPT1) downstream of Rho kinase were determined by Western blotting.
Results: In resveratrol-treated cells, progressive shape elongation was observed, evident after 6h of treatment. Treatment with resveratrol (1-20 µmol/L) dose-dependently inhibited EC migration. This effect of resveratrol was independent of nuclear factor (NF)-kappaB and sirtuin 1, but was abrogated in the presence of Rho kinase inhibitors. Moreover, resveratrol induced pMYPT1 activation, indicating a novel mechanism of resveratrol activity in EC. In monocytic cells, treatment with resveratrol significantly inhibited chemotaxis towards MCP-1 already at 1 µmol/L. At a resveratrol concentration of 10 µmol/L, chemotaxis was reduced nearly to the negative control (unstimulated with MCP-1) levels. This effect was independent of NF-kappaB and RhoA signaling. In resveratroltreated monocytic cells, MCP-1-induced Erk phosphorylation downstream of CCR2 receptor was dose-dependently inhibited, as observed by Western blot analysis.
Conclusions: Resveratrol dose-dependently inhibited endothelial cell migration and MCP-1-induced monocytic cell chemotaxis. This activity may contribute to the cardioprotective effects of resveratrol by inhibition of intimal neovascularization and monocyte recruitment into the artery wall.

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Aim: The use of currently marketed drug-eluting stents (DES) presents safety concerns, including an increased risk for late thrombosis in the range of 0.6% per year in patients, including acute coronary syndrome, which is thought to result from delayed endothelial healing effects. A new DES system targeting vascular smooth muscle cells without adverse effects on endothelial cells is therefore needed. Platelet-derived growth factor (PDGF) plays a central role in the pathogenesis of restenosis; therefore, we hypothesized that imatinib mesylate (PDGF receptor tyrosine kinase inhibitor) encapsulated bioabsorbable polymeric nanoparticle (NP)-eluting stent attenuates in-stent neointima formation.
Methods: Effects of imatinib-incorporated NP-eluting stent on neointima formation and endothelial healing were examined in a pig coronary artery stent model. Effects of imatinib-NP were also examined in cultured cells.
Results: In a cultured cell study, imatinib-NP attenuated the proliferation of vascular smooth muscle cells associated with inhibition of the target molecule (phosphorylation of PDGF receptor-β), but showed no effect on endothelial proliferation. In a pig coronary artery stent model, imatinib-NP-eluting stent markedly attenuated in-stent neointima formation and stenosis by approximately 50% as assessed by angiographic, histopathological, and intravascular ultrasound imaging analyses. Imatinib-NP-eluting stent also attenuated MAP kinase activity, but did not affect inflammation and re-endothelialization.
Conclusion: These data suggest that suppression of neointima formation by a imatinib-NP-eluting stent holds promise as a molecular-targeting NP delivery system for preventing in-stent restenosis.

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Aim: We investigated the relationship between the lipid levels achieved in a high-risk group of patients undergoing primary prevention with pravastatin and the incidence of cardiovascular disease, and moreover the influence of lifestyle compliance on lipid control.
Methods: In the APPROACH-J study, 6,229 patients who were treated with pravastatin were enrolled, and will be followed for two years. The subjects are men aged 20 years or older and women aged 55 years or older (or postmenopausal women) receiving primary prevention, who are categorized as high-risk patients with 3 or more major risk factors other than the LDL-C level according to the Japan Atherosclerosis Society guideline. Achieved LDL-C levels will be used to categorize the subjects into four quartiles for this analysis, and the maximum contrast method based on the Cox proportional hazards model will be used to investigate the relations between achieved LDL-C and the incidence of vascular event. Multiple linear regression analysis will be used to investigate the relations between adherence scores and achieved lipid level.
Results: In 5,871 patients (58.5% women) who were eligible for analysis, the mean age was 66.4years. The mean LDL-C at baseline was 135.9 mg/dL. The risk factors other than LDL-C were aging in 95.4%, diabetes in 76.9%, hypertension in 72.2%, a family history of coronary artery disease (CAD) in 20.9%, smoking in 20.0%, and low high-density lipoprotein cholesterol in 11.1%.
Conclusions: The results of this study are expected to confirm the validity of the target LDL-C level for high-risk patients receiving primary prevention and the importance of improving adherence for primary prevention.

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Aim: Postprandial hyperlipidemia (PH) is thought to be caused by the impaired postprandial metabolism of triglycerides (TG)-rich lipoproteins in both endogenous and exogenous pathways; however, there is no consensus. It is difficult to estimate the presence of PH without performing a time-consuming oral fat loading (OFL) test, so postprandial lipoprotein metabolism was analyzed by measuring the postprandial levels of apolipoprotein (apo) B-48 and apo B-100, and the correlation between postprandial TG increase and fasting apoB-48 levels was assessed to establish a good marker of PH without performing an OFL test.
Methods: Ten male normolipidemic subjects were loaded with a high-fat (HF, 1045 kcal) or standard (ST, 566 kcal) meal, and the lipids, apolipoproteins and lipoprotein profiles were analyzed after each meal.
Results: TG, apo B-48, remnant-like particles (RLP)-cholesterol and RLP-TG levels were increased and their levels were significantly higher after intake of the HF meal than the ST meal; however, there was no postprandial increase in apo B-100 and LDL-C levels. Postprandial increases in TG levels of CM, VLDL, LDL and HDL were significantly higher after intake of the HF meal than the ST meal. Fasting apo B-48 levels were strongly correlated with the incremental area under the curve of TG after intake of the HF meal, but not the ST meal.
Conclusion: Postprandial TG increase was mainly due to increased CM and CM-R, but not VLDL. Measurement of fasting serum apo B-48 may be a simple and useful method for assessment of the existence of PH.

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Aim: Although serum albumin and C-reactive protein (CRP) levels and pulse wave velocity (PWV) are known to be associated with the clinical outcome of hemodialysis (HD) patients, it is unknown which of these parameters are more predictive of the long-term mortality of such patients.
Methods: We measured biochemical parameters, including serum albumin and CRP, and the PWV of 202 patients on maintenance HD therapy and followed their course for 4 years, and 186 of the patients were enrolled in the current study analyses. We divided the 186 patients into three tertiles according to their serum albumin and CRP levels and PWV values, and conducted multivariate analyses to examine the impact of the tertiles on 4-year mortality.
Results: Twenty-three (12.4%) patients died during the follow-up period, and the serum albumin of the group that died was significantly lower than in the group that survived, but the CRP levels and PWV were significantly higher in the group that died. The results of Kaplan-Meier analyses revealed a significantly higher risk of all-cause mortality in HD patients with higher CRP based on the results of Cox proportional hazards analyses; however, the serum albumin and PWV values were not associated with all-cause mortality.
Conclusion: The results of this study suggest that serum CRP levels are a better mortality predictor than serum albumin or PWV values of chronic HD patients.

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Aim: Ezetimibe, an inhibitor of cholesterol intestinal absorption, is a lipid lowering agent. However, anti-atherogenic effects of ezetimibe have not been fully elucidated. Therefore, the objective in this study was to clarify the vascular protective effects of ezetimibe in patients with hypercholesterolemia.
Methods: Ezetimibe was administered to 20 patients with hypercholesterolemia (group E), and 20 age- and sex-matched patients with hypercholesterolemia were followed as controls (group C). Difference in metabolic profiles and cardiovascular surrogate markers before ezetimibe treatment and after 12 weeks of ezetimibe treatment were statistically evaluated.
Results: Ezetimibe treatment significantly reduced serum levels of low-density lipoprotein cholesterol (LDL-C) and malondialdehyde-modified low-density lipoprotein (MDA-LDL). In addition, the values of body mass index, body weight, waist circumference, plasma HbA1c and urinary albumin were significantly decreased in group E compared to those in group C. On the other hand, high-density lipoprotein cholesterol (HDL-C) and adiponectin levels were significantly increased in group E compared to those in group C. The values of brachial-ankle pulse wave velocity (ba-PWV), mean arterial blood pressure (m-ABP), and % of flow-mediated dilation (FMD) were significantly improved in group E. Furthermore, ultrasonic studies demonstrated amelioration of the vascular stiffness of common carotid arteries in group E but not in group C. These vascular protective effects of ezetimibe were statistically correlated with the decreased values of MDA-LDL and MDA-LDL-to-LDL-C ratio but not with those of LDL-C.
Conclusion: Ezetimibe has a lipid lowering-independent vascular protective effect in patients with hypercholesterolemia through decreasing oxidative stress.

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Aim: It has been considered that interleukin (IL)-18, a T helper 1(Th1) type cytokine, has a promoting effect on atherosclerosis development. A previous mouse study demonstrated that short-term exogenous IL-18 promoted atherosclerosis through a Th1 type immune response; however, the serum IL-18 may have increased greatly beyond its physiological range, and the effect of increased serum IL-18 on atherosclerosis development has not been investigated under different conditions of dietary fat content. The purpose of this study was to reveal the effect of increased serum IL-18 within its physiological fluctuations on atherosclerosis development under different conditions of dietary fat content.
Methods: Spontaneously hyperlipidemic (SHL) mice were systemically supplied with IL-18 for 10 weeks by means of an in vivo gene transfer system with a high-fat diet containing 0.15% cholesterol or a normal diet.
Results: Serum IL-18 steadily elevated within its physiological fluctuations. An atherosclerotic lesion area in the aortic root significantly increased with a high-fat diet. Systemic cytokine balance shifted to a Th1-dominant state, and IL-12 mRNA in the arterial wall significantly increased with a high-fat diet; however, these findings were not observed with a normal diet.
Conclusions: It was suggested that the proatherogenic effect of IL-18 is physiologically exerted exclusively with a high-fat diet through Th1 type immune responses, but not with a normal diet.

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Aim: To examine whether diabetic duration and insulin use are independently associated with the prognosis of critical limb ischemia (CLI) after endovascular therapy.
Methods: We recruited 312 Japanese patients who underwent endovascular therapy for CLI. The outcome measures were major amputation and mortality. Cox proportional hazards regression analyses were performed.
Results: The prevalence of diabetes mellitus (DM) was 69%, and 47% of the DM population were treated with insulin. DM patients with insulin use had hemoglobin A1c (HbA1c) of 7.5±1.3% and diabetic duration of 21±11 years, whereas those without insulin use had HbA1c of 6.6±1.4% and diabetic duration of 19±11 years. Follow-up period was 93±72 weeks; 55 patients underwent major amputations and 102 died. Diabetic duration and insulin use had significant associations with major amputation in each univariate model, with an unadjusted hazard ratio (HR) and 95% confidence interval (CI) of 1.019 [1.000, 1.039] in one-year increments and 2.321 [1.368, 3.938], respectively. In the multivariate model, however, diabetic duration and insulin use were not significantly associated with limb prognosis, as HbA1c level was, with an adjusted HR [95% CI] of 1.332 [1.114, 1.593] in 1% increment. Mortality had no statistical association with any of these DM-related variables.
Conclusion: Diabetic duration and insulin use were not independent risk factors for the prognosis of CLI after endovascular therapy.

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Aim: In heterozygous familial hypercholesterolaemic (FH) patients, study of the arterial wall and its function are of particular interest. Arterial stiffness has been shown to be associated with increased cardiovascular risk (CVR). In this study, we examined arterial stiffness in FH patients and its association with biochemical and vascular parameters.
Methods: In this cross-sectional study, we included 125 FH patients (20-60 years old) and 59 gender- and age-matched healthy controls (CG). Clinical, anthropometry and biochemical data were obtained. Arterial stiffness determined based on the augmentation index (AIx) was assessed with peripheral artery tonometry. Carotid intima-media thickness (cIMT) and ankle-brachial index (ABI) were also assessed.
Results: FH patients displayed a significant increase in AIx with respect to CG subjects (9.6±17.2 vs. 2.6±10.3%, P= 0.011). FH patients also had a thicker cIMT (0.758±0.280 vs. 0.635±0.160 mm, P< 0.001), while their ABIs were not different from CG subjects. AIx values were positively correlated with LDLc, non-HDLc, apolipoprotein B100, triglyceride and sE-selectin levels. Moreover, apolipoprotein B100-rich particles, along with systolic blood pressure and glucose levels, were the main determinants of AIx. In addition, we found that AIx (β= 0.224, P= 0.014) was an independent determinant of cIMT.
Conclusions: FH patients have increased arterial stiffness, despite advancements in typical clinical management. AIx is clearly associated with apolipoprotein B100 concentrations, and it is a determinant of cIMT. AIx can be utilised as a vascular risk marker in FH patients.

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Aim: Atherosclerosis is influenced by multiple environmental factors that involve a complex interaction between blood components and the arterial wall and is characterized by inflammatory reactions. Melittin has been used in treatment of various chronic inflammatory diseases. We investigated the effects of melittin regulated atherosclerotic changes in an animal model of atherosclerosis.
Methods: Atherosclerotic mice were induced by intraperitoneal (i.p) injection of lipopolysaccharide (LPS, 2 mg/kg) three times a week and an atherogenic diet for 12 weeks.
Results: Melittin (0.1 mg/kg) treatment was administered with i.p injection. Melittin treatment showed that total cholesterol and triglyceride levels decreased in atherosclerotic mice however, high-density lipoprotein cholesterol (HDL-C) levels were higher in atherosclerotic mice treated with melittin than in atherosclerotic mice. H&E staining showed that heart and descending aorta were significantly recovered by melittin, compared to atherosclerotic mice. In addition, melittin decreased the expression levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, vascular cell adhesion molecule (VCAM)-1, intercellular adhesion molecule (ICAM)-1, fibronectin and transforming growth factor (TGF)-β1 in atherosclerotic mice. In vitro, melittin decreased LPS-induced THP-1 cells-derived macrophages TNF-α and IL-1β expression levels and nuclear factor (NF)-κB signal pathway.
Conclusions: These results demonstrate that melittin has an anti-atherogenic effect by suppression of pro-inflammatory cytokines and adhesion molecules.

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According to many prospective cohort studies and meta-analyses of those studies, physical inactivity and/or low levels of physical fitness are associated with an elevated risk for the development of metabolic syndrome, type 2 diabetes, hypertension, coronary artery disease (CAD), and stroke, and with an increased risk of cardiovascular disease (CVD) mortality and all-cause mortality. Most of these analyses, however, were conducted on non-Japanese populations in the West. This report summarizes prospective observational and clinical studies in Japan. The annual national nutrition survey has shown a gradual decline in the number of walking steps in both genders and in all age groups over the last 10 years. While exercise habits have been gradually increasing in the elderly, only one-fifth of young and middle-aged people undertake leisure-time physical activity. Prospective cohort studies have shown that increased physical fitness and greater physical activity in either daily life or leisure time are of benefit in preventing all-cause mortality and CVD mortality. The daily number of walking steps is positively associated with HDL cholesterol levels and negatively associated with triglyceride levels. According to a random-effects model meta-analysis of 4 randomized controlled trials comparing supervised aerobic exercise training with non-exercise control in subjects without CAD, exercise resulted in a significant increase in HDL-cholesterol (10.01 mg/dL, 95% CI 5.38 to 14.65, p< 0.0001). While this confirms the importance of physical activity in preventing CVD mortality and all-cause mortality, the levels of physical activity are on a declining trend in Japan, particularly among the young.

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