Atherosclerotic Segments Research Articles

Macrophage-specific deletion of Notch-1 induced M2 anti-inflammatory effect in atherosclerosis via activation of the PI3K-oxidative stress axis.

Notch-1 signaling is significantly associated with the occurrence and development of atherosclerosis (AS). However, the molecular mechanisms underlying the specific deletion of Notch-1 in AS-associated macrophages are not fully understood. This study aimed to investigate the effects of Notch-1 in AS. Tissue samples were obtained from atherosclerotic segments of human carotid arteries. Immunofluorescence staining showed that Notch-1 was significantly colocalized with macrophages (CD68+), and Notch-1 staining was increased in human vulnerable plaques. Notch-1MAC-KO/ApoE-/- mice were generated in which Notch-1 was selectively inactivated in macrophages, and WT for littermate control mice (ApoE-/-/Notch-1WT). A control group was then established. All mice fed with a high-fat and Oil Red O, Movat, a-SMA, CD68, and Sirius red staining were used to evaluate the morphology. Specific deletion of Notch-1 in macrophages repressed the pathophysiology of AS. Immunofluorescent staining and Western blotting revealed that Notch-1MAC-KO repressed M1 and M2 responses in AS. Here, GSEA revealed that Notch-1 activation and PI3K signaling were statistically significantly correlated with each other, and Notch-1 was involved in the regulation of the PI3K signaling pathway. In the in vitro experiments, the secretion of Arg-1 and exosomes was classified by peritoneal macrophages of Notch-1MAC-KO/ApoE-/- and Notch-1WT/ApoE-/- mice. Immunohistochemistry staining and Western blotting were used to measure the expression levels of Notch1, PI3K, p-PI3K, AKT, p-AKT, Arg-1, IL-6, CD36, SREBP-1, CD206, iNOS, cleaved-caspase-3/-9, Bax, CD9, Alix and TSG101 in the peritoneal macrophages and exosomes, respectively. The specific deletion of Notch-1 in macrophage represses the formation and development of AS via the PI3K/AKT signaling pathway.

Phosphorylation of USP20 on Ser334 by IRAK1 promotes IL-1β-evoked signaling in vascular smooth muscle cells and vascular inflammation

Reversible lysine-63 (K63) polyubiquitination regulates proinflammatory signaling in vascular smooth muscle cells (SMCs) and plays an integral role in atherosclerosis. Ubiquitin-specific peptidase 20 (USP20) reduces NFκB activation triggered by proinflammatory stimuli, and USP20 activity attenuates atherosclerosis in mice. The association of USP20 with its substrates triggers deubiquitinase activity; this association is regulated by phosphorylation of USP20 on Ser334 (mouse) or Ser333 (human). USP20 Ser333 phosphorylation was greater in SMCs of atherosclerotic segments of human arteries as compared with nonatherosclerotic segments. To determine whether USP20 Ser334 phosphorylation regulates proinflammatory signaling, we created USP20-S334A mice using CRISPR/Cas9-mediated gene editing. USP20-S334A mice developed ∼50% less neointimal hyperplasia than congenic WT mice after carotid endothelial denudation. WT carotid SMCs showed substantial phosphorylation of USP20 Ser334, and WT carotids demonstrated greater NFκB activation, VCAM-1 expression, and SMC proliferation than USP20-S334A carotids. Concordantly, USP20-S334A primary SMCs invitro proliferated and migrated less than WT SMCs in response to IL-1β. An active site ubiquitin probe bound to USP20-S334A and USP20-WT equivalently, but USP20-S334A associated more avidly with TRAF6 than USP20-WT. IL-1β induced less K63-linked polyubiquitination of TRAF6 and less downstream NFκB activity in USP20-S334A than in WT SMCs. Using invitro phosphorylation with purified IRAK1 and siRNA-mediated gene silencing of IRAK1 in SMCs, we identified IRAK1 as a novel kinase for IL-1β-induced USP20 Ser334 phosphorylation. Our findings reveal novel mechanisms regulating IL-1β-induced proinflammatory signaling: by phosphorylating USP20 Ser334, IRAK1 diminishes the association of USP20 with TRAF6 and thus augments NFκB activation, SMC inflammation, and neointimal hyperplasia.

Radial Wall Strain Assessment From AI-Assisted Angiography: Feasibility and Agreement With OCT as Reference Standard.

High-strain spots in coronary arteries are associated with plaque vulnerability and predict future events. Artificial intelligence currently enables the calculation of radial wall strain (RWS) from coronary angiography (RWSAngio). This study aimed to determine the agreement between novel RWSAngio and RWS derived from optical coherence tomography (OCT) followed by finite element analysis, as the established reference standard (RWSOCT). All lesions from a previous OCT study were enrolled. OCT was automatically coregistered with angiography. RWSAngio was computed as the relative luminal deformation throughout the cardiac cycle, whereas RWSOCT was analyzed using finite element analysis on OCT cross-sections at 1-mm intervals. The luminal deformation in the direction of minimal lumen diameter was used to derive RWSOCT, using the same definition as RWSAngio. The maximal RWSOCT and RWSAngio at healthy segments adjacent to the interrogated lesion were also analyzed. Finite element analysis was performed in 578 OCT cross-sections from 45 lesions stemming from 36 patients. RWSAngio showed good correlation and agreement with RWSOCT (r = 0.91; P < .001; Lin coefficient = 0.85). RWSAngio in atherosclerotic segments was significantly higher than that in healthy segments (12.6% [11.0, 16.0] vs 4.5% [2.9, 5.5], P < .001). The intraclass correlation coefficients for intra- and interobserver variability in repeated RWSAngio analysis were 0.92 (95% CI, 0.87-0.95) and 0.88 (95% CI, 0.81-0.92), respectively. The mean analysis time of RWSOCT and RWSAngio for each lesion was 95.0 ± 41.1 and 0.9 ± 0.1 minutes, respectively. Radial wall strain from coronary angiography can be rapidly and easily computed solely from angiography, showing excellent agreement with strain derived from coregistered OCT. This novel and simple method might provide a cost-effective biomechanical assessment in large populations.

Differential Proteomics of Cardiovascular Risk and Coronary Artery Disease in Humans.

BackgroundProteomics of atypical phenotypes may help unravel cardiovascular disease mechanisms.AimWe aimed to prospectively screen the proteome of four types of individuals: with or without coronary artery disease (CAD), each with or without multiple risk factors. Associations with individual risk factors and circulating biomarkers were also tested to provide a functional context to the protein hits.Materials and MethodsThe CAPIRE study (ClinicalTrials.gov Identifier: NCT02157662) is a cross-sectional study aimed at identifying possible new mechanisms promoting or protecting against atherothrombosis. Quantification (by aptamer technology), ranking (using partial least squares), and correlations (by multivariate regression) of ~5000 plasma proteins were performed in consecutive individuals aged 45–75 years, without previous cardiovascular disease, undergoing computed tomography angiography for suspected CAD, showing either >5/16 atherosclerotic segments (CAD+) or completely clean arteries (CAD−) and either ≤ 1 risk factor (RF+) or ≥3 risk factors (RF−) (based on history, blood pressure, glycemia, lipids, and smoking).ResultsOf 544 individuals, 39% were atypical (93 CAD+/RF−; 120 CAD−/RF+) and 61% typical (102 CAD+/RF+; 229 CAD−/RF−). In the comparison with CAD+/RF− adjusted for sex and age, CAD−/RF+ was associated with increased atrial myosin regulatory light chain 2 (MYO) and C-C motif chemokine-22 (C-C-22), and reduced protein shisa-3 homolog (PS-3) and platelet-activating factor acetylhydrolase (PAF-AH). Extending the analysis to the entire cohort, an additional 8 proteins were independently associated with CAD or RF; by logistic regression, the 12-protein panel alone discriminated the four groups with AUCROC's of 0.72–0.81 (overall p = 1.0e−38). Among them, insulin-like growth factor binding protein-3 is positively associated with RF, lower BMI, and HDL-cholesterol, renin with CAD higher glycated hemoglobin HbA1c, and smoking.ConclusionsIn a CCTA-based cohort, four proteins, involved in opposing vascular processes (healing vs. adverse remodeling), are specifically associated with low CAD burden in high CV-risk individuals (high MYO and C-C-22) and high CAD burden in low-risk subjects (high PS-3 and PAF-AH), in interaction with BMI, smoking, diabetes, HDL-cholesterol, and HbA1c. These findings could contribute to a deeper understanding of the atherosclerotic process beyond traditional risk profile assessment and potentially constitute new treatment targets.

Assessment of neovascularization in atherosclerotic carotid sinus plaques using quantitative contrast-enhanced ultrasound perfusion imaging

Aim. To assess the prospects of using quantitative contrast-enhanced ultrasound perfusion imaging of atherosclerotic carotid sinus plaques. Material and methods. The study included 5 men and 1 woman (59-76 years old, median 72) with symptomatic coronary sinus atherosclerosis. The inclusion criterion was history of ischemic stroke due to internal carotid artery lesion (NASCET &gt;60%). We performed contrast-enhanced ultrasound perfusion imaging of the carotid arteries, endarterectomy, studying pathomorphology of the removed plaque with the calculation of the neovascular density and the total number of neovessels with a diameter &lt;40 μm. Neovascularization was assessed by quantitative contrast-enhanced ultrasound 20 seconds after the 1 ml infusion of Sonovia (Bracco, Italy) and subsequent application of the flash. The analysis of dynamics of ultrasonic signal intensity in the atherosclerotic plaque was carried out by creating the curves of the ultrasonic signal intensity (dB)/time (s) over 3 segments of the cross section of the internal carotid artery long axis. The automatic calculation of the intensity dynamics took into account the parameter values in the studied areas within 20 s after the flash. The calculated coefficients (A, B, β) of the exponential equation for 3 atherosclerotic segments were recorded. Results. Perfusion and neovascularization were assessed in 27 segments of atherosclerotic plaques. The correlation relationships between the ultrasonic parameters of plaque perfusion and the severity of neovascularization were assessed according to the histological data. Significant correlations of the exponential curve coefficient β and histological parameters characterizing the prevalence of “young” vessels (&lt;40 microns) in the atherosclerotic plaque were revealed. Spearman’s R for the density of neovessels was 0,54; for the number of neovessels with a diameter &lt;40 μm. —0,66 (p&lt;0,01). Conclusion. Diagnosis of atherosclerotic plaque neovascularization becomes possible to quantify, assessing not only the presence of neovascular vessels, but also the perfusion intensity. The novel approach replaces the qualitative and semi-quantitative method for calculating the number of carotid plaques neovessels in vivo.

Abstract 230: Matrix Regulation of Lymphangiogenesis in Atherosclerosis

Background: Atherosclerosis is a consequence of an imbalance of cellular uptake of lipoproteins versus removal of cholesterol from the arterial wall. The lymphatic vasculature represents the primary route of cholesterol removal from atherosclerotic vessels. Previous studies have linked matrix protein thrombospondin-1 (TSP1), a well-known inhibitor of angiogenesis, with human atherosclerotic disease; however the mechanisms by which TSP1 contributes to atherosclerosis and its effect on arterial lymphangiogenesis remain unknown. Methods: The effect of TSP1 on human lymphatic endothelial cell (LEC) proliferation, migration and tube formation was investigated in vitro . TSP1 protein expression was analyzed in human and murine (ApoE -/- , 12 weeks Western diet) atherosclerotic aortic tissue and lesion free segments using Western blotting. The role of TSP1 and its cognate receptor CD47 in the pathogenesis of atherosclerosis was investigated using knockout animals. Results: TSP1 expression was significantly increased in both human and ApoE -/- atherosclerotic arteries compared to plaque free segments in human and wild type aortic tissue. TSP1 at physiologically relevant concentrations (11-22 nM, 24 hrs) inhibited LEC proliferation, migration, and tube formation. RT-PCR data demonstrated that CD47 is the predominant TSP1 receptor expressed in LEC (&gt; 50-fold over CD36 mRNA levels). Finally, genome-wide deficiency in TSP1 and CD47 receptor attenuated atherosclerotic lesion formation in the aorta and brachiocephalic artery by ~50% compared to wild type controls. Importantly, there was no difference in plasma cholesterol levels, body weight, fat and lean mass, fasting plasma glucose, and blood pressure between the experimental groups. Conclusions: The present study suggests that matrix protein TSP1 via CD47 receptor contributes to the pathogenesis of atherosclerosis. Inhibition of arterial lymphangiogenesis by TSP1-CD47 signaling may contribute to arterial lipid accumulation and atherosclerosis.

First Observation of a “Golden Tube” After Complete Resorption of a Bioresorbable Vascular Scaffold in a Transplanted Patient With Cardiac Allograft Vasculopathy

The promise of complete histological reconstruction of the arterial wall after implantation of a bioresorbable vascular scaffold to treat atherosclerotic segments has challenged the way in which endovascular interventions are conceived. However, the observation of this phenomenon is not yet part of

The relationship between location-specific epicardial adipose tissue volume and coronary atherosclerotic plaque burden in type 2 diabetic patients.

Epicardial adipose tissue (EAT) is a metabolically active visceral fat depot that plays an important role in the pathogenesis of coronary atherosclerosis. Due to its asymmetrical distribution, the relation between location-specific EAT measurements and coronary atherosclerosis remains unclear. Our study investigated the relationship between location-specific EAT volume and coronary atherosclerotic plaque burden that was detected by coronary computed tomography angiography (CCTA) in type 2 diabetic patients without coronary artery disease (CAD) history. A total of 157 consecutive diabetic patients who had undergone CCTA were included retrospectively. After evaluation of the CCTA images, the study population was divided into two groups according to the presence of coronary atherosclerosis. In both groups, total and left atrioventricular groove EAT volumes were measured. Total and left atrioventricular groove EAT volumes were significantly associated with coronary atherosclerosis, but only left atrioventricular groove EAT volumes were an independent predictor for CAD. Also, total and left atrioventricular groove EAT volumes were positively correlated with C-reactive protein values (p = 0.0001/p = 0.0001) and the number of coronary atherosclerotic segments (p = 0.0001/p = 0.0001). Left atrioventricular groove EAT volume is an independent predictor of CAD in type 2 diabetic patients without CAD history. Left atrioventricular groove EAT volume may be used to identify type 2 diabetic patients who may require early CAD intervention because of the potential risk of coronary atherosclerosis.

Expansive arterial remodeling of the carotid arteries and its effect on atherosclerotic plaque composition and vulnerability: an in-vivo black-blood 3T CMR study in symptomatic stroke patients.

BackgroundBased on intravascular ultrasound of the coronary arteries expansive arterial remodeling is supposed to be a feature of the vulnerable atheroslerotic plaque. However, till now little is known regarding the clinical impact of expansive remodeling of carotid lesions. Therefore, we sought to evaluate the correlation of expansive arterial remodeling of the carotid arteries with atherosclerotic plaque composition and vulnerability using in-vivo Cardiovascular Magnetic Resonance (CMR).MethodsOne hundred eleven symptomatic patients (74 male/71.8 ± 10.3y) with acute unilateral ischemic stroke and carotid plaques of at least 2 mm thickness were included. All patients received a dedicated multi-sequence black-blood carotid CMR (3Tesla) of the proximal internal carotid arteries (ICA). Measurements of lumen, wall, outer wall, hemorrhage, calcification and necrotic core were determined. Each vessel-segment was classified according to American Heart Association (AHA) criteria for vulnerable plaque. A modified remodeling index (mRI) was established by dividing the average outer vessel area of the ICA segments by the lumen area measured on TOF images in a not affected reference segment at the distal ipsilateral ICA. Correlations of mRI and clinical symptoms as well as plaque morphology/vessel dimensions were evaluated.ResultsSeventy-eight percent (157/202) of all internal carotid arteries showed atherosclerotic disease with AHA Lesion-Type (LT) III or higher. The mRI of the ICA was significantly different in normal artery segments (AHA LT I; mRI 1.9) compared to atherosclerotic segments (AHA LT III-VII; mRI 2.5; p < 0.0001). Between AHA LT III-VII there was no significant difference of mRI. Significant correlations (p < 0.05) of the mRI with lumen-area (LA), wall-area (WA), vessel-area (VA) and wall-thickness (WT), necrotic-core area (NC), and ulcer-area were observed. With respect to clinical presentation (symptomatic/asymptomatic side) and luminal narrowing (stenotic/non-stenotic) no relevant correlations or significant differences regarding the mRI were found.ConclusionExpansive arterial remodeling exists in the ICA. However, no significant association between expansive arterial remodeling, stroke symptoms, complicated AHA VI plaque, and luminal stenosis could be established. Hence, results of our study suggest that expansive arterial remodeling is not a very practical marker for plaque vulnerability in the carotid arteries.

HIV and coronary arterial remodeling from the Multicenter AIDS Cohort Study (MACS).

Positive remodeling (PR), a coronary artery characteristic associated with risk for myocardial infarction (MI), may be more prevalent in HIV-infected (HIV+) people. We evaluated the prevalence of PR using coronary CT angiography (CCTA) in HIV+ and HIV-uninfected (HIV-) men. Men enrolled in the Multicenter AIDS Cohort Study underwent CCTA if they were 40-70 years, had normal kidney function and no history of coronary revascularization. Multivariable logistic regression models were used to estimate the odds ratio (OR) of PR by HIV serostatus, adjusting for demographics and coronary artery disease (CAD) risk factors. Analysis of PR among atherosclerotic segments further adjusted for plaque type and stenosis. The prevalence of PR was 8.4% versus 12.1% (p=0.10) for HIV- and HIV+men, respectively. After demographic adjustment, HIV+men had twice the odds of PR [OR 2.01(95% CI 1.20-3.38)], which persisted after CAD risk factor adjustment [1.76(1.00-3.10)]. Higher systolic blood pressure, total cholesterol, diabetes medication use, older age, segment number with plaque present, mixed and non-calcified plaque, and stenosis>50%, were associated with increased odds of PR, while higher HDL cholesterol, higher nadir CD4 count, and black race were associated with lower PR odds. Among atherosclerotic segments, the association between HIV infection and PR persisted, but was not statistically significantly. HIV+men have more positively remodeled arterial segments, which may be due to more coronary segments with atherosclerosis or HIV-related immunosuppression. Further studies are needed to evaluate whether PR contributes to higher rates of MI in HIV+individuals.

The 24-Month Prognosis of Patients With Positive or Intermediate Results in the Intracoronary Ergonovine Provocation Test.

This study was an observational, multicenter registry to determine clinical characteristics and 24-month prognosis of patients who underwent intracoronary ergonovine provocation tests. The clinical characteristics and prognosis of patients who underwent the ergonovine provocation for vasospastic angina were not fully elucidated. A total of 2,129 patients in the VA-KOREA (Vasospastic Angina in Korea) registry were classified into positive (n = 454), intermediate (n = 982), and negative (n = 693) groups by intracoronary ergonovine provocation tests. The 24-month incidences of cardiac death, new-onset arrhythmia, and acute coronary syndrome were determined (mean 26.7 ± 8.8 months). The number of smokers, frequency of angina before angiography, high-sensitivity C-reactive protein, and triglyceride were higher in the positive group than in other groups. The clinical characteristics of the intermediate and the negative groups were very similar. In the positive group, the incidences of diffuse, focal, and mixed spasm were 65.9%, 23.6%, and 10.6%. Coronary spasm was more frequently provoked on atherosclerotic segments. The 24-month incidences of cardiac death, arrhythmia, and acute coronary syndrome were low (0.9%, 1.6%, and 1.9%, respectively) in the positive group, and there was no cardiac death in the intermediate group (p = 0.02). In the positive group, frequent angina, current smoking, and multivessel spasm were independent predictors for adverse events. The 24-month prognosis of the positive group in the intracoronary ergonovine provocation test was relatively worse than that of the intermediate group. More intensive clinical attention should be paid to vasospastic angina patients with high-risk factors including frequent angina before angiography, current smoking, and multivessel spasm.

Arterial clamping leads to stenosis at clamp sites after femoropopliteal bypass surgery

BackgroundTo date, the incidence and clinical relevance of arterial stenosis at clamp sites after femoropopliteal bypass surgery is unknown. MethodsNinety-four patients underwent a femoropopliteal bypass in which the arterial inflow and outflow clamp sites were controlled by the Fogarty-Soft-Inlay clamp and marked with an hemoclip. The number of pre-existing atherosclerotic segments, clamp force, and clamp time were recorded and the occurrence of a stenosis at the clamp site was determined. ResultsAfter a mean follow-up of 83 months, a significant stenosis was confirmed at 23 of the 178 clamp sites (12.9%; 95% confidence interval 8.4 to 18.8). The mean number of pre-existing atherosclerotic segments (P = .28) and the mean clamp force (P = .55) was similar between the groups with and without a stenosis. There was a significant difference regarding clamp time between the group with and without a stenosis (38 minutes and 26 minutes, P = .001). ConclusionArterial clamping, even with the Fogarty-Soft-Inlay clamp, can lead to clamp stenosis and seems to be related to the duration of clamping, but not to pre-existent atherosclerotic burden.

Different Plaque Composition and Progression in Patients with Stable and Unstable Coronary Syndromes Evaluated by Cardiac CT

Objective. To compare the quantity, subtype, and progression of atherosclerosis by cardiac computed tomography (CT) and intravascular ultrasound (IVUS) in patients with stable (SAP) and unstable angina pectoris or non-ST-elevation myocardial infarction (UAP/n-STEMI). Methods. Forty patients with SAP and 20 with UAP/n-STEMI underwent cardiac CT and angiography with IVUS at baseline and after one year. Atherosclerotic segments were divided into calcified, mixed, or noncalcified subtypes, and significant stenoses were registered. Results. Thirty-two SAP and 15 UAP/n-STEMI patients completed the CT follow-up. At baseline, the number of atherosclerotic segments was higher in UAP/n-STEMI than in SAP (P = 0.039). UAP/n-STEMI patients had more segments with noncalcified plaques (P = 0.0005) whereas SAP patients had more segments with calcified plaques (P = 0.013). The number of segments with significant stenosis did not differ between the groups, but noncalcified plaques more frequently caused significant stenoses in UAP/n-STEMI than in SAP patients (P = 0.0002). After one year the number of segments with atherosclerosis increased in SAP patients (P = 0.0001). The number of atherosclerotic segments remained unchanged in UAP/n-STEMI patients. However, composition was altered as the number of segments with noncalcified plaques decreased (P = 0.018). IVUS data confirmed the CT findings. Conclusion. Quantity, subtype, and progression of atherosclerosis differ between SAP and UAP/n-STEMI patients.

Towards non-invasive imaging of vulnerable atherosclerotic plaques by targeting co-stimulatory molecules

BackgroundMyocardial infarction and stroke are the life-threatening consequences after plaque rupture in coronary or carotid arteries. Positron emission tomography employing [18F]fluorodeoxyglucose can visualize plaque inflammation; however, the question remains whether this is specific for plaque vulnerability. The pathophysiology of vulnerable plaques suggests several molecular processes. Here, we propose the co-stimulatory molecules CD80 and CD86 as potential new targets for non-invasive imaging. Methods and resultsHuman atherosclerotic segments were obtained from carotid endarterectomy and classified into stable and vulnerable plaques. We identified CD80 and CD86 with significantly higher mRNA levels in vulnerable than stable plaques. CD80+ and CD86+ cells were found in spatial proximity to CD83+ dendritic cells and CD68+ macrophages of atherosclerotic plaques. As a proof of target-expression we labeled a low molecular weight ligand, which has a high affinity for human CD80, with carbon-11 to perform in vitro autoradiography with human plaque slices. We observed 3-fold higher binding to vulnerable than stable plaques, demonstrating a first approach towards discriminating between the two plaque types. Positron emission tomography studies showed accumulation in CD80+ Raji xenografts, low radioactivity in myocardium and rapid clearance from the blood pool in mice. ConclusionIn human carotid arteries, the co-stimulatory molecules CD80 and CD86 show significantly higher expression levels in vulnerable compared to stable plaques. With the novel CD80-specific radiotracer we are able to discriminate between stable and vulnerable atherosclerotic plaques in vitro. This is an important step towards non-invasive imaging of the life-threatening vulnerable lesions in humans.

High Susceptibility of Atherosclerotic Coronary Arteries to the Onset of Vasospasm and Angina Pectoris-like Symptoms due to Coronary Spasm in WHHLMI Rabbits

Objectives: We examined the relationship between atherosclerosis and provocation of coronary spasm as well as influence of coronary spasm on the onset of acute ischemic myocardial disease. Methods and Results: Coronary spasm was provoked in anesthetized normal Japanese white (JW) rabbits and WHHLMI rabbits, an animal model for coronary atherosclerosis and myocardial infarction, by injecting ergonovine during the infusion of norepinephrine through a marginal ear vein. A decrease in contrast flow in the left circumflex artery was observed on coronary angiograms. Ischemic changes were observed on the electrocardiogram of 29% (2/7) of JW and 79% (27/34) of WHHLMI rabbits. The frequency of coronary spasm was significantly high in rabbits with severe coronary plaques showing diffuse lesions. In addition, the degree of contraction of coronary strips with atherosclerotic plaques was higher than that of normal coronary strips excised from JW rabbits stimulated by a combination of ergonovine and norepinephrine. Left ventricle motility in these vasospasm-positive rabbits, which was evaluated with echocardiograms was decreased by 29% following the ergonovine injection (P<0.001), and every serum ischemic marker examined markedly increased 4 hours after the provocation of vasospasm. Conclusions: The results of the present study demonstrated that atherosclerotic coronary arteries were positively related to the provocation of vasospasm, and vasospasm in severe atherosclerotic coronary segments evoked angina pectoris and/or non-fatal myocardial infarction.

Brief Report: Citrullination Within the Atherosclerotic Plaque: A Potential Target for the Anti–Citrullinated Protein Antibody Response in Rheumatoid Arthritis

To investigate whether citrullinated proteins within the atherosclerotic plaque can be targeted by anti-citrullinated protein antibodies (ACPAs), forming stimulatory immune complexes that propagate the progression of atherosclerosis. Protein lysates prepared from atherosclerotic segments of human aorta were assessed for the presence of citrulline-modified proteins, and specifically citrullinated fibrinogen (Cit-fibrinogen), by immunoprecipitation and/or immunoblotting followed by mass spectrometry. Immunohistochemical analysis of coronary artery plaque was performed to determine the presence of citrullinated proteins and peptidylarginine deiminase type 4 (PAD-4). Serum levels of anti-cyclic citrullinated peptide (anti-CCP), anti-citrullinated vimentin (anti-Cit-vimentin), and anti-Cit-fibrinogen antibodies were measured in 134 women with seropositive rheumatoid arthritis; these subjects had previously been characterized for the presence of subclinical atherosclerosis, by electron beam computed tomography scanning. Western blot analysis of atherosclerotic plaque lysates demonstrated several citrullinated proteins, and the presence of Cit-fibrinogen was confirmed by immunoprecipitation and mass spectrometry. Immunohistochemical analysis showed colocalization of citrullinated proteins and PAD-4 within the coronary artery plaque. In age-adjusted regression models, antibodies targeting Cit-fibrinogen and Cit-vimentin, but not CCP-2, were associated with an increased aortic plaque burden. Citrullinated proteins are prevalent within atherosclerotic plaques, and certain ACPAs are associated with the atherosclerotic burden. These observations suggest that targeting of citrullinated epitopes, specifically Cit-fibrinogen, within atherosclerotic plaques could provide a mechanism for the accelerated atherosclerosis observed in patients with RA.

Abstract 213: Notch Activation in Aortic Adventitial Fibroblasts upregulates TGF-ß and may have Implications in Aortic Aneurysms

Objectives Aortic adventitial fibroblasts (AoAF) are a rich source of cytokines and extracellular matrix (ECM). They critically participate in arterial wall remodeling and are implicated in the pathophysiology of aortic aneurysms (AA) and atherosclerosis. Little is known about signaling pathway(s) that regulate AoAF in cardiovascular diseases. Here, we examined and compared the status of Notch signaling in AoAF in human samples of normal, AA and atherosclerosis samples. We also explored the role of Notch signaling on AoAF cytokines and ECM production. Methods Immunohistochemistry was conducted to examine the Notch activity in AoAF of normal human aorta, segments with atherosclerotic plaque and segments of AA wall (N=4/type of aorta). Normal human AoAF were tested in vitro to study the effects of enforced Notch activation using lentiviral vector encoding the Notch1 intracellular domain, NIC/Lenti. The effect of Notch activation on cell proliferation was tested by MTT assay, and differentiation into myofibroblasts was tested by smooth muscle actin (α-SMA). The effect of Notch signaling on AoAF release of cytokines and ECM was studied using Pathway-Focused Human Fibrosis PCR Array (QIAGEN). Array findings were validated by Western blot. Results Compared to normal human aorta, significantly increased Notch activity was observed in AoAF from AA samples whereas decreased Notch activity was found in AoAF of atherosclerotic aorta (p&lt;0.01). Notch activation inhibited cell proliferation (~30%, p&lt;0.01) while induced differentiation of AoAF to myofibroblasts in vitro. Notch activation also resulted in a fibrotic response in AoAF as determined by increased expression of genes of collagen I/III, connective tissue growth factor (CTGF) and TGF-β1/2/3. Elevated expression of TGF-β 1/2/3(2.4-5.2-fold, p&lt;0.001) in NIC/Lenti-transduced AoAF was confirmed by Western blot. Conclusions Notch signaling is activated in aortic adventitial fibroblasts at the site of aortic aneurysms and results in increased production of TGF-β by these cells.Our novel findings implicate a potential involvement of Notch-TGF-β signaling in the pathophysiology of aortic aneurysms.

Detection and quantification of coronary atherosclerotic plaque by 64-slice multidetector CT: A systematic head-to-head comparison with intravascular ultrasound

ObjectiveWe evaluated the ability of 64-slice multidetector computed tomography (MDCT)-derived plaque parameters to detect and quantify coronary atherosclerosis, using intravascular ultrasound (IVUS) as the reference standard. MethodsIn 32 patients, IVUS and 64-MDCT was performed. The MDCT and IVUS datasets of 44 coronary arteries were co-registered using a newly developed fusion technique and quantitative parameters were derived from both imaging modalities. The threshold of >0.5mm of maximum wall thickness was used to establish plaque presence on MDCT and IVUS. ResultsWe analyzed 1364 coregistered 1-mm coronary cross-sections and 255 segments of 5-mm length. Compared with IVUS, 64-MDCT enabled correct detection in 957 of 1109 cross-sections containing plaque (sensitivity 86%). In 180 of 255 cross-sections atherosclerosis was correctly excluded (specificity 71%). On the segmental level, MDCT detected 213 of 220 segments with any atherosclerotic plaque (sensitivity 96%), whereas the presence of any plaque was correctly ruled out in 28 of 32 segments (specificity 88%). Interobserver agreement for the detection of atherosclerotic cross-sections was moderate (Cohen's kappa coefficient K=0.51), but excellent for the atherosclerotic segments (K=1.0). Pearson's correlation coefficient for vessel plaque volumes measured by MDCT and IVUS was r=0.91 (p<0.001). Bland–Altman analysis showed a slight non-significant underestimation of any plaque volume by MDCT (p=0.5), with a trend to underestimate noncalcified and overestimate mixed/calcified plaque volumes (p=0.22 and p=0.87 respectively). ConclusionMDCT is able to detect and quantify atherosclerotic plaque. Further improvement in CT resolution is necessary for more reliable assessment of very small and distal coronary plaques.

Atherosclerotic reference segments influence assessment of coronary remodeling by intravascular ultrasound in patients with acute coronary syndrome

Objective To investigate the influence of atherosclerotic reference segments on coronary remodeling by intravascular ultrasound(IVUS) in patients with acute coronary syndrome(ACS). Methods One hundred and three consecutively enrolled patients [male 68 cases,female 35 cases,average age (60 ±11) years] with ACS were undergone by coronary artery angiography and IVUS with time selecting operation. The remodeling index(RI) was defined as lesion external elastic membrane cross section aere (EEM CSA) divided by the mean reference EEM CSA at the culprit vessel. Positive remodeling was RI≥ (67 cases), negative remodeling was RI <1(36 cases). The parameters of plaque were analysed between two remodeling groups, as well as compared between distal and proximal references. Results Only one patients do not have plaque at reference sites. In the analysis of 103 lesions and 102 reference sites, there were not significant different in plaque burden,plaque CSA,lumen CSA and lesion EEM CSA between two groups. Distal plaque burden[(39. 76 ±12. 54)% vs (32.38 ± 13.97)%, P <0.001] and plaque CSA [(6.14 ± 3.20)mm2 vs (4. 75 ± 3. 07) mm2, P = 0. 001] were larger than those at proximal reference. EEM CSA at distal reference was smaller than proximal one[(14. 99 ± 4. 12)mm2 vs (28. 37 ± 4. 48)mm2 , P <0. 001]. Plaque burden, plaque CSA and lumen CSA persented positive corelation between distal and proximal reference sites. Conclusions The reference segments lay a severe diffuse atherosclerosis in the group of ACS, which may influence assessment of coronary remodeling. Coronary remodeling as a process, a accurate evaluation could be result from the combination of static and serial approach by IVUS. Key words: Endosonography; Coronary disease; Ventricular remodeling; Acute coronary syndrome

The dermatan sulfate-dependent anticoagulant pathway is mostly preserved in aneurysm and in severe atherosclerotic lesions while the heparan sulfate pathway is disrupted

BackgroundThe pathogenesis of abdominal aortic aneurysm is associated with changes of several components of arterial wall. Vascular glycosaminoglycans contribute to the non-thrombogenic activity of blood vessels. We investigated whether modifications of glycosaminoglycans in human abdominal aortic aneurysm affect their anticoagulant properties. MethodsGlycosaminoglycans were extracted from abdominal aortic aneurysms (n=11) derived from reconstitution surgeries, human abdominal aortas (n=9) from normal organ transplant donors and from preserved (n=10) and atherosclerotic (n=17) segments obtained from autopsy of an old patient. Glycosaminoglycan composition, concentration and anticoagulant activity were determined. ResultsGlycosaminoglycans extracted from aneurysms have a more potent anticoagulant activity than those from normal arteries of young adults, mostly due to a relative enrichment of dermatan sulfate, which potentiates heparin cofactor II inhibition of thrombin. Arterial segments of aged patient with severe atherosclerosis showed a glycosaminoglycan composition similar to aneurysms samples. Glycosaminoglycans extracted from these regions showed also a more potent heparin cofactor II-dependent anticoagulant activity than lesion-free areas due to the relative enrichment of dermatan sulfate. ConclusionThe anticoagulant activity from abdominal aortic aneurysms is preserved. No modifications particular to the aneurysms were dissociated from those observed in atherosclerosis.