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Oxysterols have been implicated in the pathogenesis of cardiovascular diseases. Serum levels of oxysterols could be positively correlated with cholesterol absorption and synthesis. However, physiological regulation of various serum oxysterols is largely unknown. The aim of this study was to investigate the relationship between clinical factors and cholesterol metabolism markers, and identify oxysterols associated with cholesterol absorption and synthesis in patients with coronary artery disease. Subjects (n = 207) who underwent coronary stenting between 2011 and 2013 were studied cross-sectionally. We measured lipid profiles including serum oxysterols. As for the serum biomarkers of cholesterol synthesis and absorption, oxysterol levels were positively correlated with campesterol and lathosterol. Covariance structure analysis revealed that dyslipidemia and statin usage had a positive correlation with “cholesterol absorption”. Statin usage also had a positive correlation with “cholesterol synthesis”. Several oxysterols associated with cholesterol absorption and/or synthesis. In conclusion, we elucidated the potential clinical factors that may affect cholesterol metabolism, and the associations between various oxysterols with cholesterol absorption and/or synthesis in patients with coronary artery disease.

Aims Previous studies suggested that implantation with a 1st-generation DES was associated with coronary endothelial dysfunction, which was associated with Rho-kinase activation. Second-generation drug-eluting stents (DESs) may preserve coronary endothelial function in stented coronary arteries; however, because of methodological limitations, further study is needed to clarify the association between 2nd-generation DESs and coronary endothelial dysfunction. Methods We retrospectively analysed the CuVIC trial database, where we identified 112 patients who underwent coronary stenting in the left coronary arteries with either a bare metal stent (BMS, n=53) or 2nd-generation DES (n=59). We compared vasomotions of target vessels with stents and non-target vessels without stents. Furthermore, we measured the Rho-kinase activation detected in mononucleocytes from aortic and coronary sinus blood. Results ACh-induced vasoconstrictive responses of target vessels were not enhanced with a 2nd-generation DES (45±21% vs. 44±20%, P=0.56, paired t-test), but significantly enhanced in the coronary arteries with a BMS (50±18% vs. 42±20%, P=0.002). Rho-kinase activation did not differ between patients with a BMS and 2nd-generation DES. In the target vessels with a BMS, large late lumen loss and acute coronary syndrome (ACS) at the index percutaneous coronary intervention (PCI) were associated with ACh-induced enhanced coronary vasoconstrictive responses. Conclusions Evaluation of ACh-induced vasomotion of target vessels comparing with non-target vessels revealed that 2nd-generation DESs were not associated with coronary endothelial dysfunction in target vessels, nor activation of Rho-kinase in the coronary sinus blood 6-8 months after stenting.

Aim Abdominal aortic aneurysm (AAA) is a lethal and multifactorial disease. To prevent a rupture and dissection of enlarged AAA, prophylactic surgery and stenting are currently available. There are, however, no medical therapies preventing these complications of AAA. Statin is one of the candidates, but its efficacy on AAA formation/progression remains controversial. We have previously demonstrated that nanoparticles (NPs) incorporating pitavastatin (Pitava-NPs)-clinical trials using these nanoparticles have been already conducted-suppressed progression of atherosclerosis in apolipoprotein E-deficient (Apoe－/－) mice. Therefore, we have tested a hypothesis that monocytes/macrophages-targeting delivery of pitavastatin prevents the progression of AAA. Methods Angiotensin II was intraperitoneally injected by osmotic mini-pumps to induce AAA formation in Apoe－/－ mice. NPs consisting of poly(lactic-co-glycolic acid) were used for in vivo delivery of pitavastatin to monocytes/macrophages. Results Intravenously administered Pitava-NPs (containing 0.012 mg/kg/week pitavastatin) inhibited AAA formation accompanied with reduction of macrophage accumulation and monocyte chemoattractant protein-1 (MCP-1) expression. Ex vivo molecular imaging revealed that Pitava-NPs not only reduced macrophage accumulation but also attenuated matrix metalloproteinase activity in the abdominal aorta, which was underpinned by attenuated elastin degradation. Conclusion These results suggest that Pitava-NPs inhibit AAA formation associated with reduced macrophage accumulation and MCP-1 expression. This clinically feasible nanomedicine could be an innovative therapeutic strategy that prevents devastating complications of AAA.

Background: Activated macrophages contribute to the pathogenesis of vascular disease. Vein graft failure is a major clinical problem with limited therapeutic options. PCSK9 (proprotein convertase subtilisin/kexin 9) increases low-density lipoprotein (LDL)-cholesterol levels via LDL receptor (LDLR) degradation. The role of PCSK9 in macrophage activation and vein graft failure is largely unknown, especially through LDLR-independent mechanisms. This study aimed to explore a novel mechanism of macrophage activation and vein graft disease induced by circulating PCSK9 in an LDLR-independent fashion. Methods: We used Ldlr -/- mice to examine the LDLR-independent roles of circulating PCSK9 in experimental vein grafts. Adeno-associated virus (AAV) vector encoding a gain-of-function mutant of PCSK9 (rAAV8/D377Y-mPCSK9) induced hepatic PCSK9 overproduction. To explore novel inflammatory targets of PCSK9, we used systems biology in Ldlr -/- mouse macrophages. Results: In Ldlr -/- mice, AAV-PCSK9 increased circulating PCSK9, but did not change serum cholesterol and triglyceride levels. AAV-PCSK9 promoted vein graft lesion development when compared with control AAV. In vivo molecular imaging revealed that AAV-PCSK9 increased macrophage accumulation and matrix metalloproteinase activity associated with decreased fibrillar collagen, a molecular determinant of atherosclerotic plaque stability. AAV-PCSK9 induced mRNA expression of the pro-inflammatory mediators IL-1β (interleukin-1 beta), TNFα (tumor necrosis factor alpha), and MCP-1 (monocyte chemoattractant protein-1) in peritoneal macrophages underpinned by an in vitro analysis of Ldlr -/- mouse macrophages stimulated with endotoxin-free recombinant PCSK9. A combination of unbiased global transcriptomics and new network-based hyperedge entanglement prediction analysis identified the NF-κB (nuclear factor-kappa B) signaling molecules, lectin-like oxidized LOX-1 (LDL receptor-1), and SDC4 (syndecan-4) as potential PCSK9 targets mediating pro-inflammatory responses in macrophages. Conclusions: Circulating PCSK9 induces macrophage activation and vein graft lesion development via LDLR-independent mechanisms. PCSK9 may be a potential target for pharmacologic treatment for this unmet medical need.

Background: Myocardial ischemia-reperfusion (IR) injury limits the beneficial effects of early reperfusion therapy for acute myocardial infarction. Emerging evidence suggests that 7-ketocholesterol (7-KC), one of the most common dietary oxysterols, has pro-inflammatory properties and correlates with cardiovascular diseases. However, the underlying mechanisms remain scant. Here we investigated the effects of 7-KC on myocardial IR injury in mice. Methods and Results: Wild type mice were fed either a control high-fat high-cholesterol diet (HFHCD) or HFHCD containing 7-KC (7KC-HFHCD) for three weeks. In a murine model of 30-min myocardial ischemia and subsequent reperfusion, dietary 7KC-HFHCD increased plasma 7-KC level (145 ± 89.7 ng/mL vs 399 ± 175 ng/mL, Phigh inflammatory monocytes to total monocytes increased in 7KC-HFHCD group as assessed by flow cytometric analysis (49 ± 15% vs 61 ± 9.3%, Pin vitro that 7-KC induced ER stress, mitochondrial reactive oxygen species, and nuclear factor-kappa B activation associated with increased mRNA levels of pro-inflammatory cytokines such as MCP-1 and TNF-α in murine peritoneal macrophages. Administration of N-acetyl-L-cysteine (5mM), an antioxidant, decreased 7-KC-induced pro-inflammatory cytokines, and this decrease was not observed in Tlr4 -/- murine macrophages, or in macrophages transfected with liver X receptor siRNA. Conclusions: Dietary 7-KC exacerbated myocardial IR injury through macrophage-mediated inflammation in mice. Oxidative stress is involved in the 7-KC-induced pro-inflammatory response in macrophages. Dietary oxysterols are a promising therapeutic target for IR injury.

Previous studies suggested that implantation with a 1st-generation DES was associated with coronary endothelial dysfunction, which was associated with Rho-kinase activation. Second-generation drug-eluting stents (DESs) may preserve coronary endothelial function in stented coronary arteries; however, because of methodological limitations, further study is needed to clarify the association between 2We retrospectively analysed the CuVIC trial database, where we identified 112 patients who underwent coronary stenting in the left coronary arteries with either a bare metal stent (BMS, n=53) or 2ACh-induced vasoconstrictive responses of target vessels were not enhanced with a 2Evaluation of ACh-induced vasomotion of target vessels comparing with non-target vessels revealed that 2

Introduction: Macrophage-mediated inflammation promotes various vascular diseases, but how macrophage functions are regulated has not been fully understood. Notch signaling is a fundamental pathway regulating cell differentiation in developmental stages, but its role in adult vascular diseases are largely unknown. Aim: The aim of this study is to explore the role of Notch ligands in macrophage activation and subsequent arteriosclerosis. Methods: 1) To screen the Notch ligands, blocking antibodies of each Notch ligands (Jagged-1, Jagged-2, Dll1, and Dll4) were prepared. After pre-treatment with these antibodies, mouse peritoneal macrophages were stimulated with 10 ng/mL LPS + 10 ng/mL IFN-γ or 10 ng/mL IL-4 for 24 hours. mRNA expressions were quantified by real-time PCR. 2) C57BL/6J mice were subjected to femoral arterial wire injury and blocking antibodies (250 μg/injection) were intraperitoneally injected twice a week for 4 weeks, 3) Apolipoprotein E-deficient mice ( Apoe -/- ) at 8 weeks of age were fed a high-fat diet and blocking antibodies (250 μg/injection) were intraperitoneally injected twice a week for 12 weeks. Results: Blockade of Jagged 2, Dll1 and Dll4 decreased expression of iNOS and IL-1β in macrophages treated with LPS and IFN-γ. In contrast, blockade of Dll1 increased arginase-1 and Ym1 in IL-4 treated macrophages. Immobilized recombinant Dll1 increased prototypical Notch-target gene Hes-1 and induced subsequent IL-1β expression. Therefore, we have tested Dll1 blocking antibodies in in vivo models. After femoral arterial injury, administration of Dll1 blocking antibodies attenuated intimal hyperplasia, perivascular fibrosis, and negative remodeling of injured arteries. In Apoe -/- mice, a high-fat diet feeding increased Dll1 expression in atherosclerotic plaques, especially in Mac-3 positive macrophages. Blockade of Dll1 decreased atherosclerotic plaque area and lipid content determined by oil red O staining. In addition, blockade of Dll1 decreased accumulation of Mac-3 positive macrophages to the plaque. Conclusions: These results indicate that Dll1 promotes macrophage activation and arteriosclerosis. Dll1 may be a potential therapeutic target for vascular diseases promoted by macrophage-mediated inflammation.

Objective: Mitochondria consistently change their morphology in a process regulated by proteins, including Drp1 (dynamin-related protein 1), a protein promoting mitochondrial fission. Drp1 is involved in the mechanisms underlying various cardiovascular diseases, such as myocardial ischemia/reperfusion injury, heart failure, and pulmonary arterial hypertension. However, its role in macrophages, which promote various vascular diseases, is poorly understood. We therefore tested our hypothesis that macrophage Drp1 promotes vascular remodeling after injury. Method and Results: To explore the selective role of macrophage Drp1, we created macrophage-selective Drp1-deficient mice and performed femoral arterial wire injury. In these mice, intimal thickening and negative remodeling were attenuated at 4 weeks after injury when compared with control mice. Deletion of macrophage Drp1 also attenuated the macrophage accumulation and cell proliferation in the injured arteries. Gain- and loss-of-function experiments using cultured macrophages indicated that Drp1 induces the expression of molecules associated with inflammatory macrophages. Morphologically, mitochondrial fission was induced in inflammatory macrophages, whereas mitochondrial fusion was induced in less inflammatory/reparative macrophages. Pharmacological inhibition or knockdown of Drp1 decreased the mitochondrial reactive oxygen species and chemotactic activity in cultured macrophages. Co-culture experiments of macrophages with vascular smooth muscle cells indicated that deletion of macrophage Drp1 suppresses growth and migration of vascular smooth muscle cells induced by macrophage-derived soluble factors. Conclusions: Macrophage Drp1 accelerates intimal thickening after vascular injury by promoting macrophage-mediated inflammation. Macrophage Drp1 may be a potential therapeutic target of vascular diseases.

Objective: Vascular calcification is a cardiovascular risk factor and accelerated in diabetes mellitus. Previous work has established a role for calcification-prone extracellular vesicles in promoting vascular calcification. However, the mechanisms by which diabetes mellitus provokes cardiovascular events remain incompletely understood. Our goal was to identify that increased S100A9 promotes the release of calcification-prone extracellular vesicles from human macrophages in diabetes mellitus. Approach and Results: Human primary macrophages exposed to high glucose (25 mmol/L) increased S100A9 secretion and the expression of receptor for advanced glycation end products (RAGE) protein. Recombinant S100A9 induced the expression of proinflammatory and osteogenic factors, as well as the number of extracellular vesicles with high calcific potential (alkaline phosphatase activity, P −/− S100a9 −/− mice and mice treated with S100a9 siRNA encapsulated in macrophage-targeted lipid nanoparticles showed decreased inflammation and microcalcification in atherosclerotic plaques, as gauged by molecular imaging and comprehensive histological analysis. In human carotid plaques, comparative proteomics in patients with diabetes mellitus and histological analysis showed that the S100A9-RAGE axis associates with osteogenic activity and the formation of microcalcification. Conclusions: Under hyperglycemic conditions, macrophages release calcific extracellular vesicles through mechanisms involving the S100A9-RAGE axis, thus contributing to the formation of microcalcification within atherosclerotic plaques.

A 31-year-old man was admitted to our hospital with atrial tachycardia and cardiogenic shock. He had been diagnosed with hereditary spherocytosis (HS) during childhood, but he never received any red blood cell transfusions. Right ventricular endomyocardial biopsy revealed multiple myocardial hemosiderin deposits, and he was diagnosed with cardiac hemochromatosis. In addition to the iron deposition in the heart, the loss of myocyte and severe interstitial fibrosis were present. His cardiac function did not improve even after the cardioversion for atrial tachycardia, and he suffered from recurrent heart failure. Despite intensive medical treatment for heart failure and arrhythmias in combination with iron chelation therapy, he eventually died of progressive and refractory heart failure. Hemochromatosis is a systemic disorder characterized by the excessive deposition of iron in multiple organs. The occurrence of hemochromatosis in HS is extremely rare, and previous reports have shown that the coexistence of heterozygosity for the HFE gene mutation in HS patients causes excess iron storage. The prognosis is poor due to progressive congestive heart failure and refractory arrhythmias. Here we report a rare case of fatal cardiac hemochromatosis associated with HS. The possibility of cardiac hemochromatosis needs to be considered in cases of heart failure or arrhythmia in patients with HS.

Objectives— We sought to investigate whether treatment with ezetimibe in combination with statins improves coronary endothelial function in target vessels in coronary artery disease patients after coronary stenting. Approach and Results— We conducted a multicenter, prospective, randomized, open-label, blinded-end point trial among 11 cardiovascular treatment centers. From 2011 to 2013, 260 coronary artery disease patients who underwent coronary stenting were randomly allocated to 2 arms (statin monotherapy, S versus ezetimibe [10 mg/d]+statin combinational therapy, E+S). We defined target vessel dysfunction as the primary composite outcome, which comprised target vessel failure during treatment and at the 6- to 8-month follow-up coronary angiography and coronary endothelial dysfunction determined via intracoronary acetylcholine testing performed in cases without target vessel failure at the follow-up coronary angiography. Coadministration of ezetimibe with statins further lowered low-density lipoprotein cholesterol levels (83±23 mg/dL in S versus 67±23 mg/dL in E+S; P P =0.0256), and the incidence of target vessel dysfunction at follow-up was significantly decreased in the E+S arm (69/112 (62%) in S versus 47/109 (43%) in E+S; P =0.0059). A post hoc analysis of post-treatment low-density lipoprotein cholesterol–matched subgroups revealed that the incidence of both target vessel dysfunction and coronary endothelial dysfunction significantly decreased in the E+S arm, with significant reductions in oxysterol levels. Conclusions— The CuVIC trial (Effect of Cholesterol Absorption Inhibitor Usage on Target Vessel Dysfunction after Coronary Stenting) has shown that ezetimibe with statins, compared with statin monotherapy, improves functional prognoses, ameliorating endothelial dysfunction in stented coronary arteries, and was associated with larger decreases in oxysterol levels.

Although susceptibility to cardiovascular disease (CVD) is different for every patient, why some patients with type 2 diabetes mellitus (T2DM) develop CVD while others are protected has not yet been clarified. Using T2DM-patient-derived human induced pluripotent stem cells (hiPSCs), we found that in patients protected from CVD, there was significantly elevated expression of an esterase, arylacetamide deacetylase (AADAC), in vascular smooth muscle cells (VSMCs). We overexpressed this esterase in human primary VSMCs and VSMCs differentiated from hiPSCs and observed that the number of lipid droplets was significantly diminished. Further metabolomic analyses revealed a marked reduction in storage lipids and an increase in membrane phospholipids, suggesting changes in the Kennedy pathway of lipid bioassembly. Cell migration and proliferation were also significantly decreased in AADAC-overexpressing VSMCs. Moreover, apolipoprotein E (Apoe)-knockout mice overexpressing VSMC-specific Aadac showed amelioration of atherosclerotic lesions. Our findings suggest that higher AADAC expression in VSMCs protects T2DM patients from CVD.

Background: Chronic kidney disease (CKD) increases cardiovascular risk. Underlying mechanisms, however, remain obscure. The uremic toxin indoxyl sulfate is an independent cardiovascular risk factor in CKD. We explored the potential impact of indoxyl sulfate on proinflammatory activation of macrophages and its underlying mechanisms. Methods: We examined in vitro the effects of clinically relevant concentrations of indoxyl sulfate on proinflammatory responses of macrophages and the roles of organic anion transporters and organic anion transporting polypeptides (OATPs). A systems approach, involving unbiased global proteomics, bioinformatics, and network analysis, then explored potential key pathways. To address the role of Delta-like 4 (Dll4) in indoxyl sulfate–induced macrophage activation and atherogenesis in CKD in vivo, we used 5/6 nephrectomy and Dll4 antibody in low-density lipoprotein receptor–deficient (Ldlr-/-) mice. To further determine the relative contribution of OATP2B1 or Dll4 to proinflammatory activation of macrophages and atherogenesis in vivo, we used siRNA delivered by macrophage-targeted lipid nanoparticles in mice. Results: We found that indoxyl sulfate–induced proinflammatory macrophage activation is mediated by its uptake through transporters, including OATP2B1, encoded by the SLCO2B1 gene. The global proteomics identified potential mechanisms, including Notch signaling and the ubiquitin-proteasome pathway, that mediate indoxyl sulfate-triggered proinflammatory macrophage activation. We chose the Notch pathway as an example of key candidates for validation of our target discovery platform and for further mechanistic studies. As predicted computationally, indoxyl sulfate triggered Notch signaling, which was preceded by the rapid induction of Dll4 protein. Dll4 induction may result from inhibition of the ubiquitin-proteasome pathway, via the deubiquitinating enzyme USP5. In mice, macrophage-targeted OATP2B1/Slco2b1 silencing and Dll4 antibody inhibited proinflammatory activation of peritoneal macrophages induced by indoxyl sulfate. In low-density lipoprotein receptor–deficient mice, Dll4 antibody abolished atherosclerotic lesion development accelerated in Ldlr-/- mice. Moreover, coadministration of indoxyl sulfate and OATP2B1/Slco2b1 or Dll4 siRNA encapsulated in macrophage-targeted lipid nanoparticles in Ldlr-/- mice suppressed lesion development. Conclusions: These results suggest that novel crosstalk between OATP2B1 and Dll4-Notch signaling in macrophages mediates indoxyl sulfate–induced vascular inflammation in CKD.

A 73-year-old woman presented with diarrhea, leg edema, and exertional dyspnea. Six years before admission, she had been hospitalized for heart failure with moderate-to-severe tricuspid regurgitation because of incomplete leaflet coaptation (Figure 1A; Movie I in the Data Supplement). She was treated with furosemide and has remained on this therapy since then. In the same period of time, an ovarian tumor was detected after total hysterectomy and unilateral salpingo-oophorectomy for uterine myoma. Although the surgeons tried to resect the tumor, it was inoperable and could not be diagnosed definitively at that time because of postoperative adhesion. Figure 1. The time-course of changes in the echocardiograms of the right-sided heart valves. A and C , Tricuspid and pulmonary valves recorded 6 years earlier (arrows). B and D , Tricuspid and pulmonary valves recorded on admission (arrows). Left , The B-mode image; and right , the color Doppler image. Ao indicates aorta; PA, pulmonary artery; RA, right atrium; and RV, right ventricle. On admission, physical examination revealed jugular venous distention, hepatomegaly, and ascites. Compared with the echocardiogram at the previous hospitalization (Figure 1A), the tricuspid valve leaflets were further retracted with severe regurgitation (Figure 1B; Movie II in the Data Supplement). The pulmonary valve was also retracted with moderate regurgitation (Figure 1C and 1D; Movies III and IV in the Data Supplement). It was noteworthy that the aortic and mitral valves became thickened and moderately regurgitant (Figure 2A through 2D; Movies …

Myocardial ischemia-reperfusion (IR) injury limits the therapeutic effect of early reperfusion therapy for acute myocardial infarction (AMI), in which the recruitment of inflammatory monocytes plays a causative role. Here we develop bioabsorbable poly-lactic/glycolic acid (PLGA) nanoparticles incorporating irbesartan, an angiotensin II type 1 receptor blocker with a peroxisome proliferator-activated receptor (PPAR)γ agonistic effect (irbesartan-NP). In a mouse model of IR injury, intravenous PLGA nanoparticles distribute to the IR myocardium and monocytes in the blood and in the IR heart. Single intravenous treatment at the time of reperfusion with irbesartan-NP (3.0 mg kg−1 irbesartan), but not with control nanoparticles or irbesartan solution (3.0 mg kg−1), inhibits the recruitment of inflammatory monocytes to the IR heart and reduces the infarct size via PPARγ-dependent anti-inflammatory mechanisms and ameliorates left ventricular remodeling 21 days after IR. Irbesartan-NP is a novel approach to treat myocardial IR injury in patients with AMI.

An 85-year-old woman with chronic kidney disease was referred to us for the examination of a tumour in the left ventricle that was not observed 2 years earlier. She was afebrile associated with negative blood cultures. Transthoracic and transoesophageal echocardiography revealed mitral annular calcification (MAC) and a highly mobile, hyperechoic, and spindle-shaped tumour …

Background— Preventing atherosclerotic plaque destabilization and rupture is the most reasonable therapeutic strategy for acute myocardial infarction. Therefore, we tested the hypotheses that (1) inflammatory monocytes play a causative role in plaque destabilization and rupture and (2) the nanoparticle-mediated delivery of pitavastatin into circulating inflammatory monocytes inhibits plaque destabilization and rupture. Methods and Results— We used a model of plaque destabilization and rupture in the brachiocephalic arteries of apolipoprotein E–deficient (ApoE −/− ) mice fed a high-fat diet and infused with angiotensin II. The adoptive transfer of CCR2 +/+ Ly-6C high inflammatory macrophages, but not CCR2 −/− leukocytes, accelerated plaque destabilization associated with increased serum monocyte chemoattractant protein-1 (MCP-1), monocyte-colony stimulating factor, and matrix metalloproteinase-9. We prepared poly(lactic-co-glycolic) acid nanoparticles that were incorporated by Ly-6G − CD11b + monocytes and delivered into atherosclerotic plaques after intravenous administration. Intravenous treatment with pitavastatin-incorporated nanoparticles, but not with control nanoparticles or pitavastatin alone, inhibited plaque destabilization and rupture associated with decreased monocyte infiltration and gelatinase activity in the plaque. Pitavastatin-incorporated nanoparticles inhibited MCP-1–induced monocyte chemotaxis and the secretion of MCP-1 and matrix metalloproteinase-9 from cultured macrophages. Furthermore, the nanoparticle-mediated anti–MCP-1 gene therapy reduced the incidence of plaque destabilization and rupture. Conclusions— The recruitment of inflammatory monocytes is critical in the pathogenesis of plaque destabilization and rupture, and nanoparticle-mediated pitavastatin delivery is a promising therapeutic strategy to inhibit plaque destabilization and rupture by regulating MCP-1/CCR2–dependent monocyte recruitment in this model.

Aim No prior studies have investigated the effects of dietary cholesterol oxidation products (oxysterols) on atherosclerotic plaque destabilization and rupture. We used an atherosclerotic mouse model with histological features similar to those seen in ruptured human plaques to test the hypothesis that (1) dietary oxysterols accelerate plaque destabilization and rupture and (2) a NPC1L1 inhibitor, ezetimibe, has therapeutic effects on these processes. Methods and results Advanced atherosclerotic plaques were examined in innominate arteries of ApoE(-/-) mice that were fed either a regular high-fat diet (HFD) or HFD containing oxysterols (oxysterol-HFD; 6.8% of added cholesterol was oxidized) and infused with angiotensin II. Compared with HFD, oxysterol-HFD did not affect plasma lipid levels but did accelerate plaque destabilization and rupture, which was associated with increased monocyte infiltration/activation, monocyte chemoattractant protein-1 (MCP-1) expression, and matrix metalloproteinase (MMP) activity. Dietary oxysterol-induced plaque destabilization and rupture were blunted in ApoE(-/-) CCR2(-/-) mice. Oral treatment with ezetimibe, significantly decreased plasma lipid levels and prevented the acceleration of plaque destabilization and rupture induced by dietary oxysterol. These data indicate a primary role for monocyte-mediated inflammation via the MCP-1-CCR2 pathway and the resultant increase in MMP activity in plaque destabilization and rupture induced by dietary oxysterols in ApoE(-/-) mice. These data also provide a mechanism by which dietary oxysterols are connected with the pathogenesis of plaque destabilization and rupture. Conclusions These data suggest that inhibition of the absorption of oxysterols by ezetimibe may be useful for the treatment of high-risk patients with high oxysterol intake.