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Objective : The interaction between the renin-angiotensin system and toll-like receptors (TLRs) in the pathogenesis of advanced atherosclerotic plaques is not well understood. We studied the effects of the renin inhibitor aliskiren on the progression of advanced atherosclerotic plaque in apolipoprotein E-deficient (ApoE −/− ) mice with a special focus on plaque neovessel formation. Methods and results : Four-wk-old ApoE −/− mice were fed a high-fat diet for 8 wks, and the mice were randomly assigned to one of three groups and administered a vehicle, hydralazine, or aliskiren for an additional 12 wks. Aliskiren reduced the atherosclerotic plaque area and plaque neovessel density. It increased the plaque collagen and elastin contents, and reduced plasma angiotensin II levels and plaque macrophage infiltration and cathepsin S (CatS) protein. Aliskiren also decreased the levels of AT1R, gp91phox, TLR2, monocyte chemotactic protein-1, and CatS mRNAs in the aortic roots. Hydralazine had no beneficial vascular effects, although its administration resulted in the same degree of blood pressure reduction as aliskiren. CatS deficiency mimicked the aliskiren-mediated vasculoprotective effect in the ApoE −/− mice, but aliskiren showed no further benefits in ApoE −/− CatS −/− mice. In vitro , TLR2 silencing reduced CatS expression induced by angiotensin II. Moreover, aliskiren or the inhibition of CatS impaired the endothelial cell angiogenic action in vitro or/and ex vivo . Conclusion : Renin inhibition appears to inhibit advanced plaque neovessel formation in ApoE −/− mice and to decrease the vascular inflammatory action and extracellular matrix degradation, partly by reducing AT1R/TLR2-mediated CatS activation and activity, thus regressing advanced atherosclerosis. [ABSTRACT FROM AUTHOR]

Abstract: Background: Cathepsin K (CatK) is one of the most potent mammalian collagenases involved in atherosclerosis-based vascular disease. We investigated whether circulating CatK is associated with the prevalence of coronary artery disease (CAD). Methods: Two-hundred fifty-two consecutive subjects were enrolled from among patients who underwent coronary angiography and intravascular ultrasound analyses. One-hundred thirty-two age-matched subjects served as controls. Plasma CatK, intact procollagen type I N-terminal propeptide (I-PINP), and linked carboxy-terminal telopeptide of collagen type I (ICTP) were measured. Results: Patients with CAD had higher CatK levels (44.0 ± 31.2 versus 15.5 ± 8.3 ng/mL, P < 0.001) and ICTP/I-PINP ratios (0.2 ± 0.1 versus 0.04 ± 0.03, P < 0.001) than the controls. Patients with acute coronary syndrome had higher CatK levels than those with stable angina pectoris. Overall, linear regression analysis showed that the CatK levels correlated positively with ICTP/I-PINP ratios (r = 0.41, P < 0.001). Multiple logistic regression analysis showed that CatK levels were independent predictors of CAD (odds ratio, 1.15; 95% CI, 1.07 to 1.23; P < 0.01). Furthermore, CatK levels were also correlated positively with percent plaque volumes and inversely with percent fibrous volumes by intravascular ultrasound. Conclusions: These data indicated that high levels of CatK are closely linked with the presence of CAD and that CatK serves as a novel biomarker for CAD. [Copyright &y& Elsevier]

Abstract: Although it has been demonstrated that statins stabilize atherosclerotic lesions in animal models of advanced atherosclerosis, there is little evidence to suggest that statins have a preventive effect on plaque rupture itself. In the present study, we examined the effect of fluvastatin on plaque disruption using a simple and quick method of plaque disruption in carotid artery lesions in apolipoprotein E-deficient mice. Male apolipoprotein E-deficient mice received normal chow and underwent ligation of the left common carotid artery just proximal to its bifurcation. Four weeks later, a polyethylene cuff was placed around the artery immediately proximal to the ligation site. Fluvastatin (10mg/kg per day) was given by oral gavage every day starting at 3 days before cuff placement. The administration of fluvastatin suppressed atherosclerotic plaque disruption accompanied by luminal thrombi by 31.5% compared with controls at 4 days after the cuff was placed at the ligated carotid artery. Fluvastatin administration decreased matrix metalloproteinase-9 expression, gelatinolytic activity, endothelial adhesion molecules expression and neutrophil infiltration, and increased type I collagen content in the cuffed region. In summary, fluvastatin was found to prevent plaque disruption through pleiotropic effect on acute inflammation in an animal model using apolipoprotein E-deficient mice. [Copyright &y& Elsevier]

Epidemiological studies suggest that green tea consumption is associated with a reduced risk of cardiovascular disease. Antioxidative properties of green tea flavonoids, catechins, have been believed to be involved in the antiatherogenic effect of green tea, since catechins inhibit low density lipoprotein oxidation. The migration of vascular smooth muscle cells (SMCs) from the tunica media to the subendothelial region is a key event in the development and progression of atherosclerosis and post-angioplasty vascular remodeling. Matrix metalloproteinases (MMPs) play a key role in these processes of SMC migration. In the present study, we investigated the effect of catechins on the gelatinolytic activity of MMP-2 that was derived from cultured bovine aortic SMCs. We also investigated the effect of catechins on the SMC invasion through the reconstituted basement membrane barrier. A major constituent of green tea catechins, (−)-epigallocatechin gallate (EGCG), inhibited the gelatinolytic activity of MMP-2 and concanavalin A (ConA)-induced pro-MMP-2 activation without the influence of membrane-type MMP expression in SMCs. EGCG also inhibited the SMC invasion through the basement membrane barrier in a concentration-dependent manner without any influence of SMC migration across the basement membrane protein thin-coated filter. The antagonistic effects of other catechins, namely (−)-epigallocatechin (EGC) and (−)-epicatechin gallate (ECG), on gelatinolytic activity of MMP-2, ConA-induced pro-MMP-2 activation, or PDGF-BB-directed SMC invasion were much less pronounced than those of EGCG. Also, (+)-catechin and (−)-epicatechin failed to show any effect. These findings may suggest that the anti-invasive and anti-metalloproteinase activities involve at least part of the anti-atherogenic action of catechin in accordance with the antioxidant properties of catechin. [Copyright &y& Elsevier]

Background and aims Exposure to psychosocial stress is a risk factor for cardiovascular disorders. Because the glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) agonist prevents cardiovascular injury, we investigated the beneficial effects and mechanism of the GLP-1 analogue exenatide on stress-related vascular senescence and atherosclerosis in apolipoprotein E-deficient ( ApoE −/− ) mice fed a high-fat (HF) diet. Methods ApoE −/− mice fed the HF diet were assigned to non-stressed and immobilized-stress groups for 12 weeks. Mice fed the HF diet were divided into 2 groups and administered vehicle or exenatide for 12 weeks under stress conditions. Results Chronic stress enhanced vascular endothelial senescence and atherosclerotic plaque growth. The stress increased the levels of plasma depeptidyl peptidase-4 activity and decreased the levels of plasma GLP-1 and both plasma and adipose adiponectin (APN). As compared with the mice subjected to stress alone, the exenatide-treated mice had decreased plaque microvessel density, macrophage accumulation, broken elastin, and enhanced plaque collagen volume, and lowered levels of peroxisome proliferator-activated receptor-α, gp91phox osteopontin, C-X-C chemokine receptor-4, toll-like receptor-2 (TLR2), TLR4, and cathepsins K, L, and S mRNAs and/or proteins. Exenatide reduced aortic matrix metalloproteinase-9 ( MMP-9 ) and MMP-2 gene expression and activities. Exenatide also stimulated APN expression of preadipocytes and inhibited ox-low density lipoprotein-induced foam cell formation of monocytes in stressed mice. Conclusions These results indicate that the exenatide-mediated beneficial vascular actions are likely attributable, at least in part, to the enhancement of APN production and the attenuation of plaque oxidative stress, inflammation, and proteolysis in ApoE −/− mice under chronic stress. [ABSTRACT FROM AUTHOR]

Abstract: Objective: Matrix metalloproteinases (MMPs) have been implicated in the process of vascular calcification. However, the exact roles of individual MMPs in vascular calcification are poorly understood. To study the putative role of MMP-2 in atherogenic calcification in vivo and in vitro, we investigate whether or not MMP-2 deficiency affects aortic atherosclerotic calcification in apolipoprotein E-deficient (Apoe−/−) mice and cultured smooth muscle cell (SMC) calcification. Methods and results: The area of calcified lesions in aortic intima was significantly larger in MMP-2+/+ Apoe−/− mice at 45 and 60 weeks of age than in MMP-2−/− Apoe−/− mice. In these aortic calcified atherosclerotic lesions, the expression of type II collagen, osteoprotegerin, bone morphogenetic protein (BMP)-2 and osteocalcin which are chondrocyte maker and bone-related proteins, was observed. MMP-2 deficiency reduced BMP-2 and osteocalcin expression in aortae in Apoe−/− mice at 30 weeks and 45–60-weeks-old, respectively. The expressions of MMP-2 and that of α-SMC actin were observed in chondrocyte-like cells of calcified lesions. Beta-glycerophosphate administration induced calcium deposition in MMP-2+/+ aorta-derived cultured SMCs, and this calcium deposition was significantly suppressed in MMP-2−/− aorta-derived cultured SMCs. Conclusions: These results suggest that MMP-2 may contribute to the mechanisms of calcification associated with atherosclerosis. [Copyright &y& Elsevier]

Abstract: Aims: Imaging mass spectrometry (IMS) enables the visualization of individual molecules present on tissue sections. We attempted to identify and visualize specific markers for aortic atherosclerotic lesions. Methods and results: Atherosclerotic lesions were obtained from aortic roots of apolipoprotein E (ApoE)-deficient mice at 60 weeks of age and from femoral arteries of humans with peripheral artery occlusive disease. IMS was performed with a matrix-assisted laser desorption/ionization mass spectrometry time-of-flight (TOF)/TOF-type instrument. The molecular ions at m/z 671.6 and 673.6 were found to be specific molecules in the mouse and human lipid-rich regions. These molecules were assigned as cholesterol linoleate (CE 18:2) and cholesterol oleate (CE 18:1). In the case of the human samples, triacylglycerol was also localized in the lipid-rich regions. The distributions of the molecular ions at m/z 804.5 and 832.5 were the same as the distribution of both the mouse and the human SMCs. These molecules were assigned as phosphatidylcholine (PC) (diacyl 16:0/20:4) and PC (diacyl 18:0/20:4). The molecular ion at m/z 566.9 was localized in the mouse calcified regions, and the molecular ions at m/z 539.0 were localized in the human calcified regions. Conclusions: The IMS-based histopathologic examination (IbHE) revealed the characteristic peaks of lipid-rich regions, SMCs, and calcified regions in the atherosclerotic lesions. In addition, IbHE revealed the characteristic distribution of lipids in human atherosclerotic lesions. These data indicate that an IMS-based pathologic approach is of considerable value as a new histopathologic examination. [Copyright &y& Elsevier]

Abstract: Although it has been suggested that the renin–angiotensin (RA) system and cathepsins contribute to the development and vulnerability of atherosclerotic plaque, the interaction of the RA system and cathepsins is unclear. Thus, we investigated the effects of an angiotensin II type 1 receptor (AT1) antagonist, olmesartan, on the levels of cathepsins in brachiocephalic atherosclerotic plaque and plaque stabilization in apolipoprotein E (apoE)-deficient mice receiving a high-fat diet. Under a high fat diet, treatment with olmesartan (3mg/kg per day) maintained collagen and elastin at high levels and attenuated the plaque development and cathepsin S (Cat S) level in the atherosclerotic plaque of apoE-deficient mice. The administration of olmesartan suppressed the accumulation of macrophages in plaque. Immunoreactivities of Cat S and AT1 were observed in macrophages. The amount of Cat S mRNA and the macrophage-mediated collagenolytic and elastolytic activities in cultured macrophages were increased by exposure to angiotensin II (Ang II), and these effects were diminished by olmesartan and the NADPH-oxidase inhibitor apocynin. These results suggested that Cat S derived from macrophages is involved in the mechanisms of atherosclerotic plaque vulnerability, and AT1 blocker maintained the plaque stabilization alongside the suppression of Cat S and macrophage activities. [Copyright &y& Elsevier]