Aim: The aim of the present study was to investigate how small dense low-density lipoprotein cholesterol (sdLDL-C) compared with LDL-C affect the long-term prognosis in patients with stable coronary artery disease (CAD). Methods: sdLDL-C measured by heparin magnesium precipitation and LDL particle size measured by non-denatured gradient-gel electrophoresis were compared in 190 consecutive CAD patients who underwent coronary arteriography between 2003 and 2004 who did or did not develop cardiovascular events during a seven-year follow-up period. Cardiovascular events were death caused by cardiovascular diseases(CVDs), onset of acute coronary syndrome, need for coronary and peripheral arterial revascularization, hospitalization for heart failure, surgical procedure for any CVDs, and/or hospitalization for stroke. Results: First-time cardiovascular events were observed in 72 patients. Those who experienced cardiovascular events were older and had higher prevalence rates of hypertension and diabetes; significantly higher Gensini coronary atherosclerotic scores; significantly higher levels of sdLDL-C, sdLDL-C/LDL-C, and LDL-C/high-density lipoprotein cholesterol (HDL-C) ratios; and greater glycated hemoglobin(Hb)A1c and brain natriuretic peptide (BNP) levels. They also had significantly smaller LDL particle sizes, HDL-C, apolipoprotein A-1, and estimated glomerular filtration rate (GFR) compared with patients without cardiovascular events. Conversely, LDL-C, non-HDL-C, apolipoprotein B, remnantlike particle cholesterol, and high-sensitivity C-reactive protein (hs-CRP) levels were similar between the two groups. A Kaplan-Meyer event-free survival curve demonstrated that patients with sdLDL-C≥35 mg/dL (median level) had significantly poorer prognosis compared with those with lower sdLDL-C levels, while patients with LDL-C ≥100 mg/dL had a non-significantly lower survival rate. Conclusion: These results confirm that sdLDL-C is a very promising biomarker to predict future cardiovascular events in the secondary prevention of stable CAD.
Aim: Inflammation and oxidative stress are now recognized to be two important contributing factors to the development of atherosclerosis(AS). NADPH oxidase-4 (Nox4)-derived reactive oxygen species(ROS), NF-κB and MAPK play crucial roles in these processes. Luteolin, a flavone rich in many plants, can interrupt the molecular expression and inhibit the progression of inflammation and oxidative stress. The present study was designed to test whether luteolin inhibits TNF-α-induced inflammation and oxidative stress in human umbilical vein endothelial cells(HUVECs) and identify some of the mechanisms underlying these effects. Methods: HUVECs were treated with luteolin in the presence/absence of TNF-α. The mechanism of luteolin against TNF-α-induced cell injury was evaluated using Western blotting, real-time RT-PCR and flow cytometry analyses. Results: Luteolin suppressed the TNF-α-activated ROS generation, as well as the Nox4, p22phox, and ICAM-1 and VCAM-1 expression. Luteolin also enhanced the Bcl-2 and reduced caspase-3, -9 expression in the TNF-α-treated HUVECs. Finally, luteolin inhibited the TNF-α-induced transcriptional activity of NF-κB and p38 in addition to ERK1/2 phosphorylation. The inhibitors and siRNA of Nox4 and NF-κB not only reduced ROS generation, p38, ERK1/2 phosphorylation and the ICAM-1 and VCAM-1 expression, but also enhanced Bcl-2 expression. The inhibitor of p38 had the same effect on the expression of ICAM-1, VCAM-1 and Bcl-2, while the inhibitor of ERK1/2 increased the Bcl-2 expression rather than reducing the ICAM-1 and VCAM-1 expression. Conclusions: Luteolin attenuates TNF-α-induced oxidative stress and inflammation via its effects on the Nox4/ROS-NF-κB and MAPK pathways. These results suggest that luteolin may provide a beneficial effect in treating vascular diseases associated with oxidative stress and inflammation.
Aim: The Framingham risk score (FRS) is one of the standard tools used to predict the incidence of coronary heart disease (CHD). No previous study has investigated its efficacy for a Japanese population cohort. The purpose of this study was to develop new coronary prediction algorithms for the Japanese population in the manner of the FRS, and to compare them with the original FRS. Methods: Our coronary prediction algorithms for Japanese were based on a large population-based cohort study (Suita study). The study population comprised 5,521 healthy Japanese. They were followed-up for 11.8 years on average, and 213 cases of CHD were observed. Multiple Cox proportional hazard model by stepwise selection was used to construct the prediction model. Results: Our coronary prediction algorithms for Japanese patients were based on a large populationbased cohort study (the Suita study). A multiple Cox proportional hazard model by stepwise selection was used to construct the prediction model. The C-statistics showed that the new model had better accuracy than the original and recalibrated Framingham scores. The net reclassification improvement (NRI) by the Suita score with the inclusion of CKD was 41.2% (P＜0.001) compared with the original FRS. The recalibration of the FRS slightly improved the efficiency of the prediction, but it was still worse than the Suita score with the CKD model. The calibration analysis suggested that the original FRS and the recalibrated FRS overestimated the risk of CHD in the Japanese population. The Suita score with CKD more accurately predicted the risk of CHD. Conclusion: The FRS and recalibrated FRS overestimated the 10-year risk of CHD for the Japanese population. A predictive score including CKD as a coronary risk factor for the Japanese population was more accurate for predicting CHD than the original Framingham risk scores in terms of the C-statics and NRI.
Aims: To compare the efficacy of Framingham Risk Score (FRS), UK Prospective Diabetes Study (UKPDS) risk engine, a risk score based on the Japanese Atherosclerosis Longitudinal Study-Existing Cohorts Combine (JALS-ECC), the maximum intima-media thickness (max-IMT) determined on coronary computed tomography angiography (CCTA) and their combination in asymptomatic patients with type 2 diabetes. Methods: A total of 116 Japanese patients with type 2 diabetes underwent CCTA. The risk of coronary heart disease was calculated according to the FRS, UKPDS and JALS-ECC. We evaluated the reclassification of coronary artery stenosis (CAS) based on the risk score categories after adding each IMT related variable. Results: Sixty-eight patients had CAS. The areas under the curves (AUCs) in the receiver operating characteristic curve analyses of FRS, UKPDS and JALS-ECC were 0.763 (95% confidence interval [CI]: 0.674-0.853), 0.785 (95% CI: 0.703-0.868) and 0.767 (95% CI: 0.681-0.853), respectively. The AUCs for FRS, UKPDS and JALS-ECC combined with the max-IMT were 0.788 (95% CI: 0.705-0.872), 0.800 (95% CI: 0.720-0.879) and 0.786 (95% CI: 0.703-0.869), respectively. Combining the max-IMT with the risk scores improved the identification of subjects with stenotic lesions, in particular, those in the first, second and third tertiles of the FRS, first and second tertiles of the UKPDS and first and second tertiles of the JALS-ECC (P=0.054, P=0.056, P=0.015, P=0.082, P=0.060, P=0.007, and P=0.080, respectively). The net reclassification improvement increased following the addition of a max-IMT of ≥ 1.9 mm (32.4% in FRS, 19.9% in UKPDS and 51.7% in JALS-ECC). Conclusions: These data suggest that combining a risk score with the max-IMT improves the prediction of CAS in comparison with the risk score alone.
Aim: The study aim is to investigate synergistic effects of cilostazol and probucol combination therapy on coronary plaque volume and composition. Methods: A total of 119 patients undergoing coronary stenting were treated with probucol and cilostazol combination therapy (group Ⅰ) or with cilostazol monotherapy (group Ⅱ) in a double-blind, randomized multicenter trial, and evaluated by virtual histology intravascular ultrasound (VH-IVUS) at baseline and 9-month follow-up for changes in coronary plaque volume and composition at an index intermediate lesion with luminal narrowing ≥30% and ＜70% and for neointimal hyperplasia at the stented segment. In all patients simvastatin 20 mg was started with enrollment. Results: Qualifying VH-IVUS data from 91 patients were included in the final analysis. There were no significant differences between group Ⅰ and Ⅱ with respect to the primary endpoint, nominal change in normalized total atheroma volume (TAV) of the index intermediate coronary lesion (Δ−12.6±17.7 vs. −14.2±20.2 mm3, p=0.691), or plaque composition. Plaque regression was observed in more than 70% of patients in both groups. Diabetes was the only significant independent determinant of changes in TAV (β=0.22, p=0.037). There were greater decreases in total cholesterol (Δ−51.8±33.0 vs. −25.4±39.1 mg/dL, p＜0.001) and LDL (Δ−33.5±30.5 vs. −20.3±30.8 mg/dL, p=0.044) levels in group Ⅰ than in group Ⅱ. However, HDL cholesterol (Δ−11.2±8.5 vs. 2.7±7.7 mg/dL, p＜0.001) and apoA1 (Δ−18.2±21.4 vs. 10.0±16.5 mg/dL, p＜0.001) levels were also significantly decreased in group Ⅰ compared with group Ⅱ. Conclusions: There were no significant differences in changes in plaque volume or composition between the cilostazol and probucol combination therapy and cilostazol monotherapy group despite different impacts of the treatments on lipid biomarkers.
Aim: To clarify the contribution of hemodynamic factors to the onset of plaque erosion in smooth muscle cell (SMC)-rich atherosclerotic plaque. Methods: We developed a rabbit model of SMC-rich atherosclerotic plaque with various degree of stenosis induced by incomplete ligation and generated three-dimensional models of five rabbit femoral arteries based on 130-162 serial histological cross-sections at 100-μm intervals per artery. We performed a computational blood flow simulation using the Reynolds-averaged Navier-Stokes model and calculated the wall shear stress (WSS), turbulence kinetic energy (TKE), blood pressure (BP) and blood pressure gradients (BPG) in eight sections (the inlet, the stenotic portion and areas 1, 2 and 5mm from the stenotic portion) in each rabbit. We also investigated whether the magnitude of WSS or TKE was related to the presence or absence of erosive injury by evaluating six points (the locally highest, median and lowest of WSS or TKE) in each section. Results: The magnitudes of WSS, TKE and BPG, but not BP, correlated significantly with the extent of histologically-defined plaque erosion (WSS, r=0.55, p＜0.001; TKE, r=0.53, p＜0.001; BPG, r=0.61, p＜0.0001, n=40). The values for WSS and TKE were significantly larger at sites with, compared to without, erosive injury (n=107 and n=119 points, respectively; both p＜0.0001). Conclusions: These results suggest that increased values of WSS, TKE and BPG considerably contribute to the onset of plaque erosion.
Aim: Mutations in lipoprotein-associated phospholipase A2 (Lp-PLA2) are related to atherosclerosis. However, the molecular effects of Lp-PLA2 on atherosclerosis have not been fully investigated. Therefore, this study attempted to elucidate this issue. Methods: Monocytes were isolated from randomly selected healthy male volunteers according to each Lp-PLA2 genotype (wild-type Lp-PLA2 [Lp-PLA2 (V/V)], the heterozygous V279F mutation [LpPLA2 (V/F)] and the homozygous V279F mutation [Lp-PLA2 (F/F)]) and differentiated into macrophages. The level of apoptosis in the macrophages following incubation without serum was measured using the annexin V/propidium iodide double staining method, and the underlying mechanisms were further examined using a culture cell line. Results: The average plasma Lp-PLA2 concentration [Lp-PLA2 (V/V): 129.4 ng/mL, Lp-PLA2 (V/F): 70.7 ng/mL, Lp-PLA2 (F/F): 0.4 ng/mL] and activity [Lp-PLA2 (V/V): 164.3 nmol/min/mL, LpPLA2 (V/F): 100.9 nmol/min/mL, Lp-PLA2 (F/F): 11.6 nmol/min/mL] were significantly different between each genotype, although the basic clinical characteristics were similar. The percentage of apoptotic cells was significantly higher among the Lp-PLA2 (F/F) macrophages compared with that observed in the Lp-PLA2 (V/V) macrophages. This induction of apoptosis was independent of the actions of acetylated low-density lipoproteins. In addition, the transfection of the expression plasmid of V279F mutant Lp-PLA2 into Cos-7 cells or monocyte/macrophage-like U937 cells promoted apoptosis. The knockdown of Lp-PLA2 also increased the number of apoptotic cells. Among the cells expressing mutant Lp-PLA2, the caspase-7 activity was increased, while the activated Akt level was decreased. Conclusions: The V279F mutation of Lp-PLA2 positively regulates the induction of apoptosis in macrophages and Cos-7 cells. An increase in the caspase-7 activity and a reduction in the activated Akt level are likely to be involved in this phenomenon.
Aim: The mechanism through which SAA contributes to the pathogenesis of atherosclerosis remains unclear. The objective of this study was therefore to determine whether SAA plays a role in the onset of atherosclerosis via inflammatory and apoptotic pathways. Methods: An array of experiments, including cell cultures, MTT assays, quantitative real-time PCR and Western blotting, were carried out in order to assess the effects of human recombinant SAA on NF-κB activation and the role of NF-κB in the onset of SAA-induced inflammation and apoptosis. Results: We found that SAA (80μg/mL) induced 3.5- to 37.8-fold increases in the expression of targets known to play important roles in the initiation and progression of atherosclerosis (i.e., ICAM-1, MCP-1, MMP-9 and TF) via NF-κB regulation within one hour after exposure. RAW264.7 cells treated with increasing doses of SAA showed regulation of apoptotic targets and a dose-dependent reduction in cell viability, with 69% cell viability observed following exposure to 80μg/mL of SAA for 24 hours. Conclusions: Our results suggest that SAA contributes to the pathogenesis of atherosclerosis via both inflammatory and apoptotic mechanisms.
Aim: We hypothesized that, in stable angina patients, atorvastatin therapy lowers the cathepsin S (CTSS) concentrations, as assessed non-invasively according to a plasma analysis. In addition, the low-density lipoprotein (LDL) and high-density lipoprotein (HDL) size and subclasses in the plasma were analysed to establish the association between CTSS and lipoprotein metabolism and determine whether this association is atorvastatin-sensitive.
Methods: A total of 43 patients with stable angina received atorvastatin therapy (20 mg/day, 10 weeks). The plasma CTSS mRNA levels, CTSS protein concentrations and CTSS activity, as well as LDL and HDL size and subclasses, were analysed before and after treatment. Results: Atorvastatin treatment did not change the plasma CTSS mRNA levels, although it lowered the plasma CTSS concentrations and activity. An increased plasma CTSS concentration and activity were found to be associated with a more atherogenic LDL subclass profile (a decreased dominant LDL size and increased percentage of small, dense LDL particles). The atorvastatin-induced CTSS-loweringeffect was concomitant with an improvement in the LDL subclass profile, and the changes were found to be interrelated. Concomitant, interrelated changes in the CTSS levels and LDL subclass profiles were found in the LDL phenotype B patients only (a dominant LDL diameter of ≤ 25.5 nm at the start of the study). In this subgroup, lowering of the plasma CTSS mRNA level also correlated with lowering of the proportion of small, dense LDL particles. Conclusions: Atorvastatin-induced CTSS-lowering and LDL subclass profile improvements in the plasma of LDL phenotype B patients with stable angina are concomitant and interrelated.
Aim: Stroke is a leading cause of death and disability worldwide. Most ischemic strokes (IS) are caused by atherosclerosis. Recently, the pivotal role of ADAM17 in atherosclerosis has been thoroughly addressed. However, the association between ADAM17 and IS has not yet been thoroughly explored. The present study therefore aimed to investigate the association between disintegrin and metalloproteinase 17 (ADAM17) gene polymorphisms and the risk of ischemic stroke (IS). Methods: The associations between five ADAM17 promoter polymorphisms and the risk of IS were assessed in 342 patients with IS and 296 age-matched healthy individuals in a case-control study. Results: The allele and genotype frequencies of the ADAM17 polymorphisms (rs11684747, rs11689958, rs12692386, rs55790676 and rs1524668) did not differ significantly between the IS patients and healthy control group subjects. In addition, no significant associations were detected between the ADAM17 haplotypes and IS. The mean intima-media thickness in the IS patients was not associated with the ADAM17 polymorphisms. When the IS patients were stratified according to their OCSP classification, the genotype frequencies of the ADAM17-rs1524668 polymorphism exhibited a significant association with the PACI subtype of IS. Moreover, the ADAM17-rs12692386 A＞G polymorphism was found to be associated with a higher ADAM17 mRNA expression. Conclusions: The SNPs in the ADAM17 promoter region do not appear to be major contributors to the pathogenesis of IS. However, the rs12692386 G ADAM17 allele is correlated with a higher expression of ADAM17 mRNA, which may play a role in increasing inflammation in IS patients. Furthermore, the ADAM17-rs1524668 polymorphism is linked to a higher risk of PACI-type stroke, confirming the role of ADAM17 in the pathophysiology of PACI, with potentially important therapeutic implications.