Large Necrotic Core Research Articles

Plaque Imaging Using Black-blood MRI

The risk of future ischemic events, such as angiographically determined stenosis and ulceration, in patients with atherosclerosis has long been mainly assessed using luminal morphology. However, recent remarkable advances in vessel wall imaging combined with a deeper understanding of vascular biology have shown that vessel wall characteristics also have a considerable influence on the onset of ischemic events. Among the modalities for vessel wall imaging, such as ultrasound, CT, and MRI, MRI is useful for plaque characterization because of its high accuracy and less invasiveness. Several features of vulnerable plaques, such as intraplaque hemorrhage, large lipid-rich necrotic cores, and ruptured fibrous caps, can be observed using contemporary high-resolution MRI. The assessment of plaque characteristics is essential for the management of atherosclerosis. In this article, the current status of carotid plaque characterization using black-blood MRI will be briefly highlighted, and potential clinical implications of MRI plaque imaging will be demonstrated.

Positive Remodeling – a Major Feature of Vulnerability in Patients with Non-Obstructive Coronary Artery Disease

Abstract The most common cause of acute coronary syndrome is thrombosis of an atheromatous plaque. Positive remodeling is the compensatory dilatation of the plaque-containing section of the vessel wall. Plaques are most commonly characterized as vulnerable when possessing some of the following features: fibrous cap thickness <65 µm, large necrotic lipid core, high degrees of inflammatory infiltrates, positive remodeling, intraplaque hemorrhage, or neoangio-genesis. The presence of these plaque features is associated with high cardiovascular risk. In the initial stage of vasculopathy, due to positive remodeling, lumen reduction is not typical; it only develops in the advanced phase of the disease, due to which, based on a lumenogram, the vascular system may appear intact. Therefore, coronary angiography can easily miss the diagnosis or underestimate its extent, since it does not inform us of the composition of the arterial wall, because the contrast agent is just filling the vessel lumen. Coronary CT angiography may fill this diagnostic gap, since changes of the vessel wall can directly be visualized. To increase diagnostic accuracy, invasive coronary angiography can be completed by intravascular ultrasound and optical coherence tomography.

The tandem stenosis mouse model: Towards understanding, imaging, and preventing atherosclerotic plaque instability and rupture.

The rupture of unstable atherosclerotic plaques is the major cause of cardiovascular mortality and morbidity. Despite significant limitations in our understanding and ability to identify unstable plaque pathology and prevent plaque rupture, most atherosclerosis research utilises preclinical animal models exhibiting stable atherosclerosis. Here, we introduce the tandem stenosis (TS) mouse model that reflects plaque instability and rupture, as seen in patients. The TS model involves dual ligation of the right carotid artery, leading to locally predefined unstable atherosclerosis in hypercholesterolaemic mice. It exhibits key characteristics of human unstable plaques, including plaque rupture, luminal thrombosis, intraplaque haemorrhage, large necrotic cores, thin or ruptured fibrous caps and extensive immune cell accumulation. Altogether, the TS model represents an ideal preclinical tool for improving our understanding of human plaque instability and rupture, for the development of imaging technologies to identify unstable plaques, and for the development and testing of plaque-stabilising treatments for the prevention of atherosclerotic plaque rupture. LINKED ARTICLES: This article is part of a themed issue on Preclinical Models for Cardiovascular disease research (BJP 75th Anniversary). To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.5/issuetoc.

Deletion or Inhibition of NOD1 Favors Plaque Stability and Attenuates Atherothrombosis in Advanced Atherogenesis †.

Atherothrombosis, the main cause of acute coronary syndromes (ACS), is characterized by the rupture of the atherosclerotic plaque followed by the formation of thrombi. Fatal plaque rupture sites show large necrotic cores combined with high levels of inflammation and thin layers of collagen. Plaque necrosis due to the death of macrophages and smooth muscle cells (SMCs) remains critical in the process. To determine the contribution of the innate immunity receptor NOD1 to the stability of atherosclerotic plaque, Apoe−/− and Apoe−/− Nod1−/− atherosclerosis prone mice were placed on a high-fat diet for 16 weeks to assess post-mortem advanced atherosclerosis in the aortic sinus. The proliferation and apoptosis activity were analyzed, as well as the foam cell formation capacity in these lesions and in primary cultures of macrophages and vascular SMCs obtained from both groups of mice. Our results reinforce the preeminent role for NOD1 in human atherosclerosis. Advanced plaque analysis in the Apoe−/− atherosclerosis model suggests that NOD1 deficiency may decrease the risk of atherothrombosis by decreasing leukocyte infiltration and reducing macrophage apoptosis. Furthermore, Nod1−/− SMCs exhibit higher proliferation rates and decreased apoptotic activity, contributing to thicker fibrous caps with reduced content of pro-thrombotic collagen. These findings demonstrate a direct link between NOD1 and plaque vulnerability through effects on both macrophages and SMCs, suggesting promising insights for early detection of biomarkers for treating patients before ACS occurs.

Contemporary rationale for non-invasive imaging of adverse coronary plaque features to identify the vulnerable patient:a Position Paper from the European Society of Cardiology Working Group on Atherosclerosis and Vascular Biology and the European Association of Cardiovascular Imaging.

Atherosclerotic plaques prone to rupture may cause acute myocardial infarction (MI) but can also heal without causing an event. Certain common histopathological features, including inflammation, a thin fibrous cap, positive remodelling, a large necrotic core, microcalcification, and plaque haemorrhage are commonly found in plaques causing an acute event. Recent advances in imaging techniques have made it possible to detect not only luminal stenosis and overall coronary atherosclerosis burden but also to identify such adverse plaque characteristics. However, the predictive value of identifying individual adverse atherosclerotic plaques for future events has remained poor. In this Position Paper, the relationship between vulnerable plaque imaging and MI is addressed, mainly for non-invasive assessments but also for invasive imaging of adverse plaques in patients undergoing invasive coronary angiography. Dynamic changes in atherosclerotic plaque development and composition may indicate that an adverse plaque phenotype should be considered at the patient level rather than for individual plaques. Imaging of adverse plaque burden throughout the coronary vascular tree, in combination with biomarkers and biomechanical parameters, therefore holds promise for identifying subjects at increased risk of MI and for guiding medical and invasive treatment.

Antibody-Mediated Inhibition of CTLA4 Aggravates Atherosclerotic Plaque Inflammation and Progression in Hyperlipidemic Mice.

T cell-driven inflammation plays a critical role in the initiation and progression of atherosclerosis. The co-inhibitory protein Cytotoxic T-Lymphocyte Associated protein (CTLA) 4 is an important negative regulator of T cell activation. Here, we studied the effects of the antibody-mediated inhibition of CTLA4 on experimental atherosclerosis by treating 6–8-week-old Ldlr−/− mice, fed a 0.15% cholesterol diet for six weeks, biweekly with 200 μg of CTLA4 antibodies or isotype control for six weeks. 18F-fluorodeoxyglucose Positron Emission Tomography—Computed Tomography showed no effect of the CTLA4 inhibition of activity in the aorta, spleen, and bone marrow, indicating that monocyte/macrophage-driven inflammation was unaffected. Correspondingly, flow cytometry demonstrated that the antibody-mediated inhibition of CTLA4 did not affect the monocyte populations in the spleen. αCTLA4 treatment induced an activated T cell profile, characterized by a decrease in naïve CD44−CD62L+CD4+ T cells and an increase in CD44+CD62L− CD4+ and CD8+ T cells in the blood and lymphoid organs. Furthermore, αCTLA4 treatment induced endothelial activation, characterized by increased ICAM1 expression in the aortic endothelium. In the aortic arch, which mainly contained early atherosclerotic lesions at this time point, αCTLA4 treatment induced a 2.0-fold increase in the plaque area. These plaques had a more advanced morphological phenotype and an increased T cell/macrophage ratio, whereas the smooth muscle cell and collagen content decreased. In the aortic root, a site that contained more advanced plaques, αCTLA4 treatment increased the plaque T cell content. The short-term antibody-mediated inhibition of CTLA4 thus accelerated the progression of atherosclerosis by inducing a predominantly T cell-driven inflammation, and resulted in the formation of plaques with larger necrotic cores and less collagen. This indicates that existing therapies that are based on αCTLA4 antibodies may promote CVD development in patients.

PET/MRI of atherosclerosis

Myocardial infarction and stroke are the most prevalent global causes of death. Each year 15 million people worldwide die due to myocardial infarction or stroke. Rupture of a vulnerable atherosclerotic plaque is the main underlying cause of stroke and myocardial infarction. Key features of a vulnerable plaque are inflammation, a large lipid-rich necrotic core (LRNC) with a thin or ruptured overlying fibrous cap, and intraplaque hemorrhage (IPH). Noninvasive imaging of these features could have a role in risk stratification of myocardial infarction and stroke and can potentially be utilized for treatment guidance and monitoring. The recent development of hybrid PET/MRI combining the superior soft tissue contrast of MRI with the opportunity to visualize specific plaque features using various radioactive tracers, paves the way for comprehensive plaque imaging. In this review, the use of hybrid PET/MRI for atherosclerotic plaque imaging in carotid and coronary arteries is discussed. The pros and cons of different hybrid PET/MRI systems are reviewed. The challenges in the development of PET/MRI and potential solutions are described. An overview of PET and MRI acquisition techniques for imaging of atherosclerosis including motion correction is provided, followed by a summary of vessel wall imaging PET/MRI studies in patients with carotid and coronary artery disease. Finally, the future of imaging of atherosclerosis with PET/MRI is discussed.

Pathologic Disparities Between Peripheral Artery Disease and Coronary Artery Disease.

Atherosclerosis is a systemic disease that involves multiple vascular beds. The pathological characteristics and clinical presentation, however, vary among the different vascular territories. Acute coronary syndrome is a relatively common manifestation of coronary atherosclerotic disease, wherein the thrombosis occurs secondary to disruption (65%-75%) and erosion (25%-35%) of the fibrous caps of atheromatous plaques. The plaques associated with plaque rupture have large necrotic cores and thin and inflamed fibrous caps. However, the pathological manifestations of peripheral artery disease result from thrombosis regardless of the extent of atherosclerosis. Approximately 75% of peripheral arteries with significant stenosis demonstrate presence of thrombi, of which two-thirds have thrombi associated with insignificant atherosclerosis. The presence of obliterative thrombi in peripheral arteries of patients with critical limb ischemia in the absence of coronary artery-like lesions suggests a locally thrombogenic or remotely embolic basis of disease. Extensive calcification of the medial vascular layer is commonly observed. In this review, we have described and compared the pathological basis of coronary and peripheral artery disease in patients with acute coronary syndrome and critical limb ischemia. It is expected that pathogenetic characterization would allow for definition of strategic targets for superior management of peripheral artery disease.

Vulnerable atherosclerotic plaques of coronary arteries in patients with stable coronary artery disease: 12-months follow-up

A key objective of modern cardiology is the assessment of acute coronary syndrome (ACS) risk in patients with coronary artery disease (CAD) to develop preventive measures and choose optimal treatment strategies. Detect vulnerable plaques of non-target coronary arteries in patients with stable CAD during routine percutaneous coronary intervention using virtual-histology intravascular ultrasound (VH-IVUS) and view their morphology over time. The prospective observational cohort study included 58 patients with stable CAD. After stenting ofatarget vessel, VH-IVUS was carried out in proximal and middle segments (6-8 cm) of a non-target coronary artery with no significant stenosis according to coronary angiography. Twelve months later, all patients underwent coronary angiography with re-IVUS of previously detected lesions. Death, myocardial infarction, rehospitalization, and unplanned myocardial revascularization due to vulnerable plaques were the endpoints of the study. IVUS with virtual histology revealed 58 lesions of non-target coronary arteries in 56 (96.5 %) patients. Two patients had no lesions in non-target coronary arteries. A large necrotic core with thin cap (thin-cap fibroatheroma) was detected in 12 (20.7 %) plaques, six of which had additional ACS risk criteria (stenosis area >70 % and / or lumen area <4 mm2). Within the 12‑month follow-up period, three patients (one with a vulnerable plaque in IVUS) were hospitalized with a clinical picture of ACS. One cardiac death was registered in a patient with the IVUS vulnerable plaque. 7 of 12 vulnerable plaques stabilized in 12 months. 1) The data presented indicate a high rate (20.7 %) of vulnerable plaques of non-target coronary arteries in patients with stable CAD who underwent stenting; 2) Two (16.6 %) patients with vulnerable plaques reached endpoints (death and rehospitalization) within the 12‑month follow-up period; 3) An analysis ofatherosclerotic plaques in non-target coronary arteries over time showed that vulnerable plaques stabilized and did not cause ACS in more than half of cases (7 of 12); 4) Plaques that were not vulnerable according to IVUS were not likely to destabilize within the 12‑month follow-up period.

Ca2+ Flux: Searching for a Role in Efferocytosis of Apoptotic Cells in Atherosclerosis.

In atherosclerosis, macrophages in the arterial wall ingest plasma lipoprotein-derived lipids and become lipid-filled foam cells with a limited lifespan. Thus, efficient removal of apoptotic foam cells by efferocytic macrophages is vital to preventing the dying foam cells from forming a large necrotic lipid core, which, otherwise, would render the atherosclerotic plaque vulnerable to rupture and would cause clinical complications. Ca2+ plays a role in macrophage migration, survival, and foam cell generation. Importantly, in efferocytic macrophages, Ca2+ induces actin polymerization, thereby promoting the formation of a phagocytic cup necessary for efferocytosis. Moreover, in the efferocytic macrophages, Ca2+ enhances the secretion of anti-inflammatory cytokines. Various Ca2+ antagonists have been seminal for the demonstration of the role of Ca2+ in the multiple steps of efferocytosis by macrophages. Moreover, in vitro and in vivo experiments and clinical investigations have revealed the capability of Ca2+ antagonists in attenuating the development of atherosclerotic plaques by interfering with the deposition of lipids in macrophages and by reducing plaque calcification. However, the regulation of cellular Ca2+ fluxes in the processes of efferocytic clearance of apoptotic foam cells and in the extracellular calcification in atherosclerosis remains unknown. Here, we attempted to unravel the molecular links between Ca2+ and efferocytosis in atherosclerosis and to evaluate cellular Ca2+ fluxes as potential treatment targets in atherosclerotic cardiovascular diseases.

107Diagnostic accuracy of optical coherence tomography for the identification of in-stent fibroatheroma following stent implantation: an ex-vivo histological validation study

Abstract Background Previous histopathological studies have demonstrated that new atherosclerotic formation within the neointima, called neoatherosclerosis, is one of the most important mechanisms leading to both very late in-stent restenosis and stent thrombosis after stent implantation. Therefore, to distinguish lipid-containing atherosclerotic neointima from other tissues using intracoronary imaging modalities is clinically important to prevent late stent failures. Purpose This study evaluated the diagnostic performance of optical coherence tomography (OCT) for the detection of “in-stent fibroatheroma” following stent implantation by comparing cross-sections of the model with the corresponding histological images. Methods Fifty stented coronary arteries from the 31 autopsy hearts were imaged by OCT. Coronary arterial histopathological specimens, all of which included more than 30% of %neointimal hyperplasia, were compared with the corresponding OCT cross-sections. Histological in-stent fibroatheroma was defined as neointima containing large necrotic core and inflammatory cells. OCT-derived in-stent fibroatheroma comprised a low-intensity tissue containing a poorly delineated region with invisible stent strut behind low signal intensity. Results A total of 122 OCT cross-sections were compared with histological images. OCT examination revealed that 24 images (20%) contained low-intensity tissue inside the neointima. Of those, 5 images, in which stent strut behind low signal intensity was invisible, were diagnosed as OCT-derived in-stent fibroatheroma (4%) (Figure A). By histological analysis, only 4 images were classified as in-stent fibroatheroma (3%) (Figure B). With histology as the gold standard, the sensitivity, specificity, positive predictive value, negative predictive value, and overall diagnostic accuracy for OCT-derived in-stent fibroatheroma were 100%, 99%, 80%, 100%, and 99%, respectively. The only histological finding underlying the false-positive-diagnosis of OCT-derived in-stent fibroatheroma was foam cells accumulation without necrotic core on the neointimal surface (Figure C and D). Most tissue that showed low-intensity tissue with visible stent strut by OCT contained proteoglycan matrix and organized thrombus in the absence of an underlying necrotic core. Coregistration of OCT with histology Conclusion This study showed the potential capability of OCT based on the visualization of stent struts behind low-intensity regions for discriminating in-stent fibroatheroma from other neointimal tissues following stent implantation.

P1941The nuclear receptor corepressor 1 blocks CD36-mediated foam cell formation in atherogenesis

Abstract Background Nuclear receptors and their cofactors regulate the expression of various target genes in different tissue and organs to orchestrate downstream (patho)physiological processes. Although the function of several nuclear receptors in atherosclerosis has been studied, very little is known about the role of nuclear receptor cofactors in atherosclerosis. Given its important role to suppress inflammatory processes, we speculated that macrophage nuclear receptor corepressor 1 (NCOR1) plays a protective function in atherosclerosis development. Purpose To evaluate the contribution of macrophage NCOR1 in atherosclerosis we used myeloid cell-specific Ncor1 knockout mice on an atherosclerosis-prone background. Methods and results 8-week-old male and female mice were exposed to a high high-cholesterol diet for 12 weeks. Our findings demonstrate that the lack of macrophage Ncor1 leads to a severe atherosclerotic phenotype in both sexes. These mice show a higher content of plaques along the thoraco-abdominal aortae as well as at the aortic sinus, which were characterized by larger necrotic cores and thinner fibrous caps, a typical signature of unstable plaques. Moreover, we found that the pro-atherogenic effects of the Ncor1 deletion are mediated via derepression of peroxisome proliferator-activated receptor gamma (PPARγ) target genes in mouse and human macrophages, especially the enhanced expression of the CD36 scavenger receptor and the subsequent rise in oxLDL uptake. Interestingly, while the expression of NCOR1 is reduced, the PPARγ signature is increased in human atherosclerotic plaques, and this signature is further pronounced in ruptured compared to stable carotid plaques. Conclusion Our findings suggest that macrophage NCOR1 blocks the pro-atherogenic functions of PPARγ in atherosclerosis and prevents the disease development. Acknowledgement/Funding The Swiss National Science Foundation, the Novartis Foundation, Olga-Mayenfisch Foundation, the OPO foundation

Invited Commentary on "Imaging-based Selection for Endovascular Treatment in Stroke".

Invited Commentary on "Imaging-based Selection for Endovascular Treatment in Stroke".

5222A distinct LDL profile to predict the risk of cardiovascular disease in familial hypercholesterolemia subjects: initial pre-clinical results

Abstract Aim Cardiovascular disease development is highly variable between patients with familial hypercholesterolemia (FH). Since current risk prediction methods fail to estimate the risk of individual patients, all patients are treated with high-intensity lipid-lowering medication like statins and PCSK9-inhibitors. To prevent overtreatment (with its associated costs and side effects) of patients with a low risk, reliable biomarkers are urgently needed. While studying atherosclerosis development in an FH porcine model, we discovered a specific LDL profile that was directly associated with the severity of atherosclerosis development. Methods 10 adult pigs with an LDLR mutation received a high-fat diet for 12 months. Atherosclerosis development in the three main coronary arteries was monitored with intravascular ultrasound (IVUS) and optical coherence tomography at three time points. After sacrifice, tissue was harvested for detailed histological analysis of the coronary plaque composition. Size-exclusion chromatography (SEC, n=10) and LC-MSMS (n=4) were used to assess the plasma lipoprotein profile, and the sphingolipid content of LDL, relative to cholesterol. Results Imaging and histology revealed a marked difference in pigs that developed large, lumen intruding plaques (IVUS-based plaque burden 13–77%) within 9 months (n=5) and pigs (n=5) that only developed early lesions (IVUS-based plaque burden 8–34%), even after 12 months of follow-up. The plaques seen in the fast responding pigs possessed distinct characteristics of advanced disease (i.e. heavy macrophage infiltration, large lipid-rich necrotic cores, neovascularisation, micro- and macrocalcifications and intraplaque haemorrhage, see figure). In these fast responders, fibrous cap atheroma occupied 34% of the total artery. SEC revealed two distinct LDL subclasses: regular and “larger” LDL particles. Fast responding pigs with advanced atherosclerosis displayed a significantly higher ratio in cholesterol concentration of regular/'larger' LDL than slow responding pigs (1.7 (1.3–1.9) vs. 0.8 (0.6–1.2); (p=0.004), see figure). Compared to regular LDL, “larger” LDL contained relatively more sphingolipids in the fast responding than in the slow responding animals (regular LDL/'larger' LDL: S1P 0.5 (0.5–0.5) vs. 1.0 (0.8–1.2); Cer16:0 0.70 (0.67–0.73) vs. 1.04 (0.95–1.13); Cer18:0 0.60 (0.58–0.61) vs. 1.15 (1.13–1.16); Cer20:0 0.73 (0.73–0.74) vs. 0.94 (0.94–0.94)). “Larger” LDL particles and comparable sphingolipid ratios were also observed in FH patients. Cardiovascular data from our FH patient cohort, coupled to the LDL subclass distribution, will provide more insight into the potential of this novel biomarker. Conclusion A distinct difference in LDL subclasses, including a new “larger” LDL particle, was found in fast versus slow responding FH animals. This finding can potentially be used to identify FH patients at the highest risk of CVD to avoid overtreatement of low risk patients. Acknowledgement/Funding ERC- starter grant (grant agreement 310457)

3284Quantification of macrophage presence and identification of thin-cap fibroatheroma by optical coherence tomography image: histopathological validation study

Abstract Background Intracoronary optical coherence tomography (OCT) is thought to be capable of identifying a vulnerable, rupture-prone plaque based on the presence of a thin-cap fibroatheroma (TCFA). Moreover, recent studies have reported that OCT may be able to identify macrophage infiltration of the fibrous cap, a key characteristic of vulnerable plaque. Purpose This study evaluated the accuracy of OCT image for characterizing TCFA and identifying macrophage infiltration in comparison with histopathology. Methods A total of 924 focal plaques in 206 coronary arteries from 78 autopsy hearts were examined to compare OCT and histological images. By histology, 16 plaques (1.7%) were classified as TCFAsthat contained a large necrotic core covered by a thin (<65μm) fibrous-cap. Correlating OCT-histological sections were identified and OCT-derived tissue property indexes named normalized standard deviation (NSD) and signal attenuation ratio were applied on the fibrous-cap to identify inflamed fibrous-cap defined as a macrophage percentage >10% by histology. Results With histology as standard, the sensitivity, specificity, and negative-predictive-value of TCFAs were extremely high (more than 90%). However, the positive-predictive-value of TCFAs was only 32%, which indicated a high proportion of false-positives. Most false-positive diagnoses of OCT for TCFAs contained large amounts of foam cell accumulations on luminal surface without necrotic core. Twelve of 16 fibrous-caps were considered as inflamed and the remaining 4 were non-inflamed on histology. However, no significant difference in NSD and signal attenuation ratio were identified between them. There was moderate correlation of the fibrous-cap thickness between OCT and histology (r2 = 0.41 and p<0.01). Conclusions OCT is a promising intracoronary imaging modality for differentiating tissue characteristics (fibrous, calcified, or lipid-rich plaque) and identifying TCFA. However, it is still challenging to precisely identify inflammation, fibrous-cap thickness, and necrotic core in the native coronary artery. Therefore, careful interpretation is required to assess coronary vulnerable plaque by OCT. Acknowledgement/Funding None

Variation in Coronary Atherosclerosis Severity Related to a Distinct LDL (Low-Density Lipoprotein) Profile: Findings From a Familial Hypercholesterolemia Pig Model.

In an adult porcine model of familial hypercholesterolemia (FH), coronary plaque development was characterized. To elucidate the underlying mechanisms of the observed inter-individual variation in disease severity, detailed lipoprotein profiles were determined. Approach and Results: FH pigs (3 years old, homozygous LDLR R84C mutation) received an atherogenic diet for 12 months. Coronary atherosclerosis development was monitored using serial invasive imaging and histology. A pronounced difference was observed between mildly diseased pigs which exclusively developed early lesions (maximal plaque burden, 25% [23%-34%]; n=5) and advanced-diseased pigs (n=5) which developed human-like, lumen intruding plaques (maximal plaque burden, 69% [57%-77%]) with large necrotic cores, intraplaque hemorrhage, and calcifications. Advanced-diseased pigs and mildly diseased pigs displayed no differences in conventional risk factors. Additional plasma lipoprotein profiling by size-exclusion chromatography revealed 2 different LDL (low-density lipoprotein) subtypes: regular and larger LDL. Cholesterol, sphingosine-1-phosphate, ceramide, and sphingomyelin levels were determined in these LDL-subfractions using standard laboratory techniques and high-pressure liquid chromatography mass-spectrometry analyses, respectively. At 3 months of diet, regular LDL of advanced-diseased pigs contained relatively more cholesterol (LDL-C; regular/larger LDL-C ratio 1.7 [1.3-1.9] versus 0.8 [0.6-0.9]; P=0.008) than mildly diseased pigs, while larger LDL contained more sphingosine-1-phosphate, ceramides, and sphingomyelins. Larger and regular LDL was also found in plasma of 3 patients with homozygous FH with varying LDL-C ratios. In our adult FH pig model, inter-individual differences in atherosclerotic disease severity were directly related to the distribution of cholesterol and sphingolipids over a distinct LDL profile with regular and larger LDL shortly after the diet start. A similar LDL profile was detected in patients with homozygous FH.

Coronary Artery Microcalcification: Imaging and Clinical Implications.

Strategies to prevent acute coronary and cerebrovascular events are based on accurate identification of patients at increased cardiovascular (CV) risk who may benefit from intensive preventive measures. The majority of acute CV events are precipitated by the rupture of the thin cap overlying the necrotic core of an atherosclerotic plaque. Hence, identification of vulnerable coronary lesions is essential for CV prevention. Atherosclerosis is a highly dynamic process involving cell migration, apoptosis, inflammation, osteogenesis, and intimal calcification, progressing from early lesions to advanced plaques. Coronary artery calcification (CAC) is a marker of coronary atherosclerosis, correlates with clinically significant coronary artery disease (CAD), predicts future CV events and improves the risk prediction of conventional risk factors. The relative importance of coronary calcification, whether it has a protective effect as a stabilizing force of high-risk atherosclerotic plaque has been debated until recently. The extent of calcium in coronary arteries has different clinical implications. Extensive plaque calcification is often a feature of advanced and stable atherosclerosis, which only rarely results in rupture. These macroscopic vascular calcifications can be detected by computed tomography (CT). The resulting CAC scoring, although a good marker of overall coronary plaque burden, is not useful to identify vulnerable lesions prone to rupture. Unlike macrocalcifications, spotty microcalcifications assessed by intravascular ultrasound or optical coherence tomography strongly correlate with plaque instability. However, they are below the resolution of CT due to limited spatial resolution. Microcalcifications develop in the earliest stages of coronary intimal calcification and directly contribute to plaque rupture producing local mechanical stress on the plaque surface. They result from a healing response to intense local macrophage inflammatory activity. Most of them show a progressive calcification transforming the early stage high-risk microcalcification into the stable end-stage macroscopic calcification. In recent years, new developments in noninvasive cardiovascular imaging technology have shifted the study of vulnerable plaques from morphology to the assessment of disease activity of the atherosclerotic lesions. Increased disease activity, detected by positron emission tomography (PET) and magnetic resonance (MR), has been shown to be associated with more microcalcification, larger necrotic core and greater rates of events. In this context, the paradox of increased coronary artery calcification observed in statin trials, despite reduced CV events, can be explained by the reduction of coronary inflammation induced by statin which results in more stable macrocalcification.

Macrophage NCOR1 protects from atherosclerosis by repressing a pro-atherogenic PPARγ signature.

Nuclear receptors and their cofactors regulate key pathophysiological processes in atherosclerosis development. The transcriptional activity of these nuclear receptors is controlled by the nuclear receptor corepressors (NCOR), scaffolding proteins that form the basis of large corepressor complexes. Studies with primary macrophages demonstrated that the deletion of Ncor1 increases the expression of atherosclerotic molecules. However, the role of nuclear receptor corepressors in atherogenesis is unknown. We generated myeloid cell-specific Ncor1 knockout mice and crossbred them with low-density lipoprotein receptor (Ldlr) knockouts to study the role of macrophage NCOR1 in atherosclerosis. We demonstrate that myeloid cell-specific deletion of nuclear receptor corepressor 1 (NCOR1) aggravates atherosclerosis development in mice. Macrophage Ncor1-deficiency leads to increased foam cell formation, enhanced expression of pro-inflammatory cytokines, and atherosclerotic lesions characterized by larger necrotic cores and thinner fibrous caps. The immunometabolic effects of NCOR1 are mediated via suppression of peroxisome proliferator-activated receptor gamma (PPARγ) target genes in mouse and human macrophages, which lead to an enhanced expression of the CD36 scavenger receptor and subsequent increase in oxidized low-density lipoprotein uptake in the absence of NCOR1. Interestingly, in human atherosclerotic plaques, the expression of NCOR1 is reduced whereas the PPARγ signature is increased, and this signature is more pronounced in ruptured compared with non-ruptured carotid plaques. Our findings show that macrophage NCOR1 blocks the pro-atherogenic functions of PPARγ in atherosclerosis and suggest that stabilizing the NCOR1-PPARγ binding could be a promising strategy to block the pro-atherogenic functions of plaque macrophages and lesion progression in atherosclerotic patients.

Angiotensin II deteriorates advanced atherosclerosis by promoting MerTK cleavage and impairing efferocytosis through the AT1R/ROS/p38 MAPK/ADAM17 pathway.

Vulnerable plaques in advanced atherosclerosis have defective efferocytosis. The role of ANG II in the progression of atherosclerosis is not fully understood. Herein, we investigated the effects and the underlying mechanisms of ANG II on macrophage efferocytosis in advanced atherosclerosis. ANG II decreased the surface expression of Mer tyrosine kinase (MerTK) in macrophages through a disintegrin and metalloproteinase17 (ADAM17)-mediated shedding of the soluble form of MerTK (sMer) in the medium, which led to efferocytosis suppression. ANG II-activated ADAM17 required reactive oxygen species (ROS) and p38 MAPK phosphorylation. Selective angiotensin II type 1 receptor (AT1R) blocker losartan suppressed ROS production, and ROS scavenger N-acetyl-l-cysteine (NAC) prevented p38 MAPK phosphorylation. In addition, mutant MERTKΔ483-488 was resistant to ANG II-induced MerTK shedding and efferocytosis suppression. The advanced atherosclerosis model that is characterized by larger necrotic cores, and less collagen content was established by feeding apolipoprotein E knockout (ApoE-/-) mice with a high-fat diet for 16 wk. NAC and losartan oral administration prevented atherosclerotic lesion progression. Meanwhile, the inefficient efferocytosis represented by decreased macrophage-associated apoptotic cells and decreased MerTK+CD68+double-positive macrophages in advanced atherosclerosis were prevented by losartan and NAC. Additionally, the serum levels of sMer were increased and positively correlated with the upregulated levels of ANG II in acute coronary syndrome (ACS) patients. In conclusion, ANG II promotes MerTK shedding via AT1R/ROS/p38 MAPK/ADAM17 pathway in macrophages, which led to defective efferocytosis and atherosclerosis progression. Defining the molecular mechanisms of defective efferocytosis may provide a promising prognosis and therapy for ACS patients.

Atherosclerotic plaque mechanical characterization coupled with vector Doppler imaging in atherosclerotic carotid arteries in-vivo.

Methods used in clinical practice to diagnose and monitor atherosclerosis present limitations. Imaging the mechanical properties of the arterial wall has demonstrated the potential evaluate plaque vulnerability and assess the risk for stroke. Adaptive Pulse Wave Imaging (PWI) is a non-invasive ultrasound imaging technique, which automatically detects points of spatial mechanical inhomogeneity along the imaged artery and provides piecewise stiffness characterization. The aims of the present study are to: 1) demonstrate the initial feasibility of adaptive PWI to image the mechanical properties of an atherosclerotic plaque 2) demonstrate the feasibility to combine adaptive PWI with vector Doppler in a single imaging modality in order to simultaneously obtain information plaque mechanical properties and plaque hemodynamics. The common carotid arteries of 1 healthy subject and 2 carotid artery disease patients were scanned in vivo. One of the patients underwent carotid endarterectomy and a plaque sample was retrieved. In this patient, a higher compliance value of the stenotic segment was estimated by Adaptive PWI as compared with the adjacent arterial wall, and the healthy carotid artery. This was corroborated by histological staining of the plaque sample, which revealed the presence of a large necrotic core and a thrombus, characteristics associated with reduced stiffness. Moreover, the same sequence demonstrated the feasibility to obtain both stiffness maps and vector flow information, showing promise in atherosclerosis diagnosis and patient care.