Human Coronary Atherosclerotic Plaques Research Articles

3D MRI-based multicomponent thin layer structure only plaque models for atherosclerotic plaques

MRI-based fluid-structure interactions (FSI) models for atherosclerotic plaques have been developed to perform mechanical analysis to investigate the association of plaque wall stress (PWS) with cardiovascular disease. However, the time consuming 3D FSI model construction process is a great hinder for its clinical implementations.In this study, a 3D thin-layer structure only (TLS) plaque model was proposed as an approximation with much less computational cost to 3D FSI models for better clinical implementation potential. 192 TLS models were constructed based on 192 ex vivo MRI Images of 12 human coronary atherosclerotic plaques. Plaque stresses were extracted from all lumen nodal points. The maximum value of Plaque wall stress (MPWS) and average value of plaque wall stress (APWS) of each slice were used to compare with those from corresponding FSI models. The relative errors for MPWS and APWS were 9.76% and 9.89%, respectively. Both MPWS and APWS values obtained from TLS models showed very good correlation with those from 3D FSI models. Correlation results from TLS models were in consistent with FSI models.Our results indicated that the proposed 3D TLS plaque models may be used as a good approximation to 3D FSI models with much less computational cost. With further validation, 3D TLS models may be possibly used to replace FSI models to save time and perform mechanical analysis for atherosclerotic plaques for clinical implementation.

AHA classification of coronary and carotid atherosclerotic plaques by grating-based phase-contrast computed tomography.

To evaluate the potential of grating-based phase-contrast computed-tomography (gb-PCCT) to classify human carotid and coronary atherosclerotic plaques according to modified American Heart Association (AHA) criteria. Experiments were carried out at a laboratory-based set-up consisting of X-ray tube (40kVp), grating-interferometer and detector. Eighteen human carotid and coronary artery specimens were examined. Histopathology served as the standard of reference. Vessel cross-sections were classified as AHA lesion type I/II, III, IV/V, VI, VII or VIII plaques by two independent reviewers blinded to histopathology. Conservative measurements of diagnostic accuracies for the detection and differentiation of plaque types were evaluated. A total of 127 corresponding gb-PCCT/histopathology sections were analyzed. Based on histopathology, lesion type I/II was present in 12 (9.5%), III in 18 (14.2%), IV/V in 38 (29.9%), VI in 16 (12.6%), VII in 34 (26.8%) and VIII in 9 (7.0%) cross-sections. Sensitivity, specificity and positive and negative predictive value were ≥0.88 for most analyzed plaque types with a good level of agreement (Cohen's kappa = 0.90). Overall, results were better in carotid (kappa = 0.97) than in coronary arteries (kappa = 0.85). Inter-observer agreement was high with kappa = 0.85, p < 0.0001. These results indicate that gb-PCCT can reliably classify atherosclerotic plaques according to modified AHA criteria with excellent agreement to histopathology. • Different atherosclerotic plaque types display distinct morphological features in phase-contrast CT. • Phase-contrast CT can detect and differentiate AHA plaque types. • Calcifications caused streak artefacts and reduced sensitivity in type VI lesions. • Overall agreement was higher in carotid than in coronary arteries.

Fibronectin Splicing Variants Containing Extra Domain A Promote Atherosclerosis in Mice Through Toll-Like Receptor 4

Cellular fibronectin containing extra domain A (EDA(+)-FN) is abundant in the arteries of patients with atherosclerosis. Several in vitro studies suggest that EDA(+)-FN interacts with Toll-like receptor 4 (TLR4). We tested the hypothesis that EDA(+)-FN exacerbates atherosclerosis through TLR4 in a clinically relevant model of atherosclerosis, the apolipoprotein E-deficient (Apoe(-/-)) mouse. The extent of atherosclerosis was evaluated in whole aortae and cross sections of the aortic sinus in male and female EDA(-/-)Apoe(-/-) mice (which lack EDA(+)-FN), EDA(fl/fl)Apoe(-/-) mice (which constitutively express EDA(+)-FN), and control Apoe(-/-) mice fed a high-fat Western diet for 14 weeks. Irrespective of sex, EDA(fl/fl)Apoe(-/-) mice exhibited a 2-fold increase in atherosclerotic lesions (aorta and aortic sinus) and macrophage content within plaques, whereas EDA(-/-)Apoe(-/-) mice exhibited reduced atherosclerotic lesions (P<0.05 versus Apoe(-/-), n=10-12 mice/group), although cholesterol and triglyceride levels and circulating leukocytes were similar. Genetic ablation of TLR4 partially reversed atherosclerosis exacerbation in EDA(fl/fl)Apoe(-/-) mice (P<0.05) but had no effect on atherosclerotic lesions in EDA(-/-)Apoe(-/-) mice. Purified cellular FN, which contains EDA, potentiated dose-dependent NFκB-mediated inflammation (increased phospho-NFκB p65/NFκB p65, tumor necrosis factor-α, and interleukin-1β) in bone marrow-derived macrophages from EDA(-/-)Apoe(-/-) mice but not from EDA(-/-)TLR4(-/-)Apoe(-/-) mice. Finally, using immunohistochemistry, we provide evidence for the first time that EDA(+)-FN colocalizes with macrophage TLR4 in murine aortic lesions and human coronary artery atherosclerotic plaques. Our findings reveal that TLR4 signaling contributes to EDA(+)-FN-mediated exacerbation of atherosclerosis. We suggest that EDA(+)-FN could be a therapeutic target in atherosclerosis.

MicroRNA-136 Promotes Vascular Muscle Cell Proliferation Through the ERK1/2 Pathway by Targeting PPP2R2A in Atherosclerosis

Aberrant proliferation of vascular smooth muscle cells [VSMCs] is implicated in the pathogenesis of vascular pathologies such as atherosclerosis and restenosis. Accumulating evidences have revealed that microRNAs are involved in cell proliferation in various pathological conditions. In the present study, we showed that miR-136 was up regulated in human coronary atherosclerotic plaques when compared with normal coronary artery tissues. Moreover, miR-136 levels were up regulated in proliferative vascular smooth muscle cells induced by platelet-derived growth factor [PDGF] or serum. In cultured VSMCs, over expression of miR-136 stimulated cell proliferation. PPP2R2A was proved to be the direct target gene of miR-136 and knockdown of PPP2R2A had a proliferative effect on VSMCs. miR-136-induced PPP2R2A down-regulation was accompanied by increased expression of ERK1/2 phosphorylation. Inhibition of ERK1/2 abolished the effect of miR-136 and knockdown of PPP2R2A on VSMCs proliferation. In summary, aberrant miR-136 up regulation in atherosclerosis contributes to abnormal VSMC proliferation through suppressing the ERK1/2 pathway by targeting PPP2R2A. Our study also suggested that specific modulation of miR-136 in human VSMCs may provide a potential approach for the treatment of atherosclerosis.

Extended output phasor representation of multi-spectral fluorescence lifetime imaging microscopy.

In this paper, we investigate novel low-dimensional and model-free representations for multi-spectral fluorescence lifetime imaging microscopy (m-FLIM) data. We depart from the classical definition of the phasor in the complex plane to propose the extended output phasor (EOP) and extended phasor (EP) for multi-spectral information. The frequency domain properties of the EOP and EP are analytically studied based on a multiexponential model for the impulse response of the imaged tissue. For practical implementations, the EOP is more appealing since there is no need to perform deconvolution of the instrument response from the measured m-FLIM data, as in the case of EP. Our synthetic and experimental evaluations with m-FLIM datasets of human coronary atherosclerotic plaques show that low frequency indexes have to be employed for a distinctive representation of the EOP and EP, and to reduce noise distortion. The tissue classification of the m-FLIM datasets by EOP and EP also improves with low frequency indexes, and does not present significant differences by using either phasor.

Prevalence of M4 macrophages within human coronary atherosclerotic plaques is associated with features of plaque instability

BackgroundThe platelet chemokine CXCL4 induces monocyte differentiation resulting in a macrophage phenotype called “M4”, which co-expresses CD68, MMP7, and S100A8. We hypothesized that M4 macrophages are associated with plaque destabilization. MethodsAtherosclerotic arteries were obtained from explanted hearts of patients with severe coronary artery disease (CAD, n=32) and of patients with dilated cardiomyopathy and no or mild CAD (controls, n=19). Coronary arteries were stained with H&E, and immuno-fluorescence was performed against CD68, MMP7, and S100A8. ResultsBoth CD68+ macrophages representing the entire macrophage population and MMP7+S100A8+CD68+ M4 macrophages could be reproducibly identified within all arterial layers. The average proportion of the M4 macrophage phenotype amongst all CD68+ macrophages was 31.7±16.2%. The highest number of M4 macrophages was found in the adventitia, followed by the intima. CD68+ and M4 macrophage numbers were significantly higher in patients with severe CAD. The presence of M4 macrophages within the intima and the media was significantly associated with plaque instability as determined by Stary class. Multivariate analysis showed a highly significant contribution of cardiovascular risk factors (P=0.008) to plaque instability, while only trends were observed for age (P=0.060) and intimal prevalence of M4 macrophages (P=0.098). ConclusionsWe demonstrate for the first time that M4 macrophages can be reproducibly found in coronary artery plaques. The prevalence of M4 macrophages is associated with indexes of plaque instability, most likely representing a surrogate marker of inflammatory activity. These findings suggest a pathogenetic role of M4 macrophages in vulnerable atherosclerotic plaques.

MicroRNA-24 regulates macrophage behavior and retards atherosclerosis.

Our recent studies have highlighted membrane type-1 matrix metalloproteinase (MMP)-14 as a selective marker for an invasive subset of macrophages potentially related to atherosclerotic plaque progression. Moreover, colony stimulating factors (CSF) may exert divergent effects on macrophage MMP expression, possibly through microRNAs. We, therefore, aim to identify and test the pathophysiological role of microRNAs, which modulate macrophage MMP-14 expression in atherosclerotic plaque progression. Compared with macrophage CSF-differentiated macrophages, granulocyte/macrophage CSF-matured macrophages exhibited reduced MMP-14 mRNA levels but increased protein expression and activity, which resulted in heightened macrophage invasion. MicroRNA-24, identified to target MMP-14, was accordingly increased in macrophage CSF compared with granulocyte/macrophage CSF macrophages. Silencing microRNA-24 in macrophage CSF macrophages significantly increased MMP-14 expression and enhanced their invasive capacity, mimicking granulocyte/macrophage CSF macrophages, and suggesting that granulocyte/macrophage CSF modulates MMP-14 protein expression and subsequent macrophage invasion in a microRNA-24-dependent manner. In human coronary atherosclerotic plaques, increased MMP-14 protein expression in foam cell macrophages was associated with lesions exhibiting histological characteristics associated with an unstable phenotype. Furthermore, microRNA-24 expression in these atherosclerotic plaques was inversely related to MMP-14 protein expression. Moreover, stable plaques contained higher microRNA-24 levels than unstable plaques, and microRNA-24 colocalized with foam cell macrophages that exhibited low MMP-14 protein expression. Finally, in atherosclerotic mice (apolipoprotein E-deficient), microRNA-24 inhibition increased plaque size and macrophage MMP-14 expression. Taken together, our data demonstrates that downregulation of microRNA-24 promotes an invasive macrophage subset and plays a novel regulatory role in MMP-14 proteolytic activity and, therefore, plaque stability, highlighting its therapeutic potential.

Data Mining of Atherosclerotic Plaque Transcriptomes Predicts STAT1-Dependent Inflammatory Signal Integration in Vascular Disease

Atherosclerotic plaque development involves multiple extra- and intra-cellular signals engaging cells from the immune system and from the vasculature. Pro-inflammatory pathways activated by interferon gamma (IFNγ) and toll-like receptor 4 (TLR4) ligands are profoundly involved in plaque formation and have been shown to involve cross-talk in all atheroma-interacting cell types leading to increased activation of signal transducer and activator of transcription-1 (STAT1) and elevated expression of pro-inflammatory mediators. Here we demonstrate that in Gene Expression Omnibus repository (GEO) deposited microarray datasets, obtained from human coronary and carotid atherosclerotic plaques, a significant increase in expression of pro-inflammatory and immunomodulatory genes can be detected. Moreover, increased expression of multiple chemokines, adhesion molecules and matrix-remodeling molecules was commonly detected in both plaque types and correlated with the presence of putative STAT1 binding sites in their promoters, suggesting strong involvement of STAT1 in plaque development. We also provide evidence to suggest that STAT1-nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) or STAT1-interferon-regulated factor (IRF) regulatory modules are over-represented in the promoters of these inflammatory genes, which points to a possible contribution of IFNγ and TLR4 cross-talk in the process of atherogenesis. Finally, a subset of these genes encodes for secreted proteins that could serve as a basis of a non-invasive diagnostic assay. The results of our in silico analysis in vitro provide potential evidence that STAT1-dependent IFNγ-TLR4 cross-talk plays a crucial role in coronary and carotid artery plaque development and identifies a STAT1-dependent gene signature that could represent a novel diagnostic tool to monitor and diagnose plaque progression in human atherosclerosis.

Photon counting spectral CT component analysis of coronary artery atherosclerotic plaque samples.

To evaluate the capabilities of photon counting spectral CT to differentiate components of coronary atherosclerotic plaque based on differences in spectral attenuation and iodine-based contrast agent concentration. 10 calcified and 13 lipid-rich non-calcified histologically demonstrated atheromatous plaques from post-mortem human coronary arteries were scanned with a photon counting spectral CT scanner. Individual photons were counted and classified in one of six energy bins from 25 to 70 keV. Based on a maximum likelihood approach, maps of photoelectric absorption (PA), Compton scattering (CS) and iodine concentration (IC) were reconstructed. Intensity measurements were performed on each map in the vessel wall, the surrounding perivascular fat and the lipid-rich and the calcified plaques. PA and CS values are expressed relative to pure water values. A comparison between these different elements was performed using Kruskal-Wallis tests with pairwise post hoc Mann-Whitney U-tests and Sidak p-value adjustments. RESULTS for vessel wall, surrounding perivascular fat and lipid-rich and calcified plaques were, respectively, 1.19 ± 0.09, 0.73 ± 0.05, 1.08 ± 0.14 and 17.79 ± 6.70 for PA; 0.96 ± 0.02, 0.83 ± 0.02, 0.91 ± 0.03 and 2.53 ± 0.63 for CS; and 83.3 ± 10.1, 37.6 ± 8.1, 55.2 ± 14.0 and 4.9 ± 20.0 mmol l(-1) for IC, with a significant difference between all tissues for PA, CS and IC (p < 0.012). This study demonstrates the capability of energy-sensitive photon counting spectral CT to differentiate between calcifications and iodine-infused regions of human coronary artery atherosclerotic plaque samples by analysing differences in spectral attenuation and iodine-based contrast agent concentration. Photon counting spectral CT is a promising technique to identify plaque components by analysing differences in iodine-based contrast agent concentration, photoelectric attenuation and Compton scattering.

Estimation of the number of fluorescent end-members for quantitative analysis of multispectral FLIM data.

Multispectral fluorescence lifetime imaging (m-FLIM) can potentially allow identifying the endogenous fluorophores present in biological tissue. Quantitative description of such data requires estimating the number of components in the sample, their characteristic fluorescent decays, and their relative contributions or abundances. Unfortunately, this inverse problem usually requires prior knowledge about the data, which is seldom available in biomedical applications. This work presents a new methodology to estimate the number of potential endogenous fluorophores present in biological tissue samples from time-domain m-FLIM data. Furthermore, a completely blind linear unmixing algorithm is proposed. The method was validated using both synthetic and experimental m-FLIM data. The experimental m-FLIM data include in-vivo measurements from healthy and cancerous hamster cheek-pouch epithelial tissue, and ex-vivo measurements from human coronary atherosclerotic plaques. The analysis of m-FLIM data from in-vivo hamster oral mucosa identified healthy from precancerous lesions, based on the relative concentration of their characteristic fluorophores. The algorithm also provided a better description of atherosclerotic plaques in term of their endogenous fluorophores. These results demonstrate the potential of this methodology to provide quantitative description of tissue biochemical composition.

Abstract 59: Fibronectin Extradomain A Modulates Atherosclerotic Lesion Progression in Hypercholesterolemic Mice Through Toll-Like Receptor 4

Background and Hypothesis: Fibronectin (FN) containing the alternatively-spliced extra domain A (EDA + -FN) is absent in the arteries or circulation of healthy humans and mice, but specifically expressed in the endothelium of atherosclerotic arteries and elevated in circulation during pathological settings including stroke and atherosclerosis. We tested the hypothesis that EDA + -FN exacerbates atherosclerosis. Model and Methods: We generated EDA hi/hi /ApoE -/- mice (over express EDA + -FN in plasma) and EDA -/- /ApoE -/- mice (EDA-deleted allele of FN). ApoE -/- mice, which express EDA + -FN in plasma (~3-5 μg/mL) similar to levels found in human pathological settings including stroke and atherosclerosis, were used as control. Male mice (n=14/group) were fed a high-fat Western diet beginning at 6 weeks until they were sacrificed at 5 and 8 months of age. Extent of atherosclerosis was evaluated in whole aortae (Oil Red O stain) and in the cross section area of the aortic sinus (Verhoeff-Van Gieson stain). Markers of inflammation were quantified by immunohistochemistry. Results: We report that atherosclerotic plaque formation in the aortae and aortic sinus of EDA hi/hi /ApoE -/- mice were increased by 2-fold, concomitant with increased plaque inflammatory content (neutrophils, macrophages, and NF-κB p65 expression; P&lt;0.01 vs. ApoE -/- ). Conversely, EDA -/- /ApoE -/- mice had smaller plaques (P&lt;0.01 vs. ApoE -/- mice), concordant with decreased inflammatory content within plaque. Because EDA + -FN activates toll-like receptor 4 (TLR4) in vitro , we investigated the role of TLR4 in the EDA + -FN-mediated accelerated atherosclerosis. Genetic ablation of TLR4 in EDA hi/hi /ApoE -/- mice partially reversed exacerbated atherosclerosis (P&lt;0.05 vs. EDA hi/hi /ApoE -/- mice), whereas TLR4 deficiency in EDA -/- /ApoE -/- had no effect. Total cholesterol and triglycerides levels were similar among groups. Finally, we report co-localization of EDA + -FN with TLR4 on macrophages in human coronary atherosclerotic plaque samples. Conclusions: Endogenous EDA + -FN exacerbates atherosclerosis via TLR4. Targeting EDA + -FN/TLR4 pathway may have the potential to reduce atherosclerosis and improve outcomes in patients at high risk for coronary artery disease.

Blind End-Member and Abundance Extraction for Multispectral Fluorescence Lifetime Imaging Microscopy Data

This paper proposes a new blind end-member and abundance extraction (BEAE) method for multispectral fluorescence lifetime imaging microscopy (m-FLIM) data. The chemometrical analysis relies on an iterative estimation of the fluorescence decay end-members and their abundances. The proposed method is based on a linear mixture model with positivity and sum-to-one restrictions on the abundances and end-members to compensate for signature variability. The synthesis procedure depends on a quadratic optimization problem, which is solved by an alternating least-squares structure over convex sets. The BEAE strategy only assumes that the number of components in the analyzed sample is known a spriori. The proposed method is first validated by using synthetic m-FLIM datasets at 15, 20, and 25dB signal-to-noise ratios. The samples simulate the mixed response of tissue containing multiple fluorescent intensity decays. Furthermore, the results were also validated with six m-FLIM datasets from fresh postmortem human coronary atherosclerotic plaques. A quantitative evaluation of the BEAE was made against two popular techniques: minimum volume constrained nonnegative matrix factorization (MVC-NMF) and multivariate curve resolution-alternating least-squares (MCR-ALS). Our proposed method (BEAE) was able to provide more accurate estimations of the end-members: 0.32% minimum relative error and 13.82% worst-case scenario, despite different initial conditions in the iterative optimization procedure and noise effect. Meanwhile, MVC-NMF and MCR-ALS presented more variability in estimating the end-members: 0.35% and 0.34% for minimum errors and 15.31% and 13.25% in the worst-case scenarios, respectively. This tendency was also maintained for the abundances, where BEAE obtained 0.05 as the minimum absolute error and 0.12 in the worst-case scenario; MCR-ALS and MVC-NMF achieved 0.04 and 0.06 for the minimum absolute errors, and 0.15 and 0.17 under the worst-case conditions, respectively. In addition, the average computation time was evaluated for the synthetic datasets, where MVC-NMF achieved the fastest time, followed by BEAE and finally MCR-ALS. Consequently, BEAE improved MVC-NMF in convergence to a local optimal solution and robustness against signal variability, and it is roughly 3.6 time faster than MCR-ALS.

Spectroscopic intravascular photoacoustic imaging of lipids in atherosclerosis

The natural history of atherosclerosis is marked by changes in the lipid biochemistry in the diseased arterial wall. As lesions become more vulnerable, different cholesterol species accumulate in the plaque. Understanding unstable atherosclerosis as a pharmacological and interventional therapeutic target requires chemically specific imaging of disease foci. In this study, we aim to image atherosclerotic plaque lipids and other vessel wall constituents with spectroscopic intravascular photoacoustics (sIVPA). sIVPA imaging can identify lipids in human coronary atherosclerotic plaque by relying on contrast in the near-infrared absorption spectra of the arterial wall components. Using reference spectra acquired on pure compounds, we analyzed sIVPA data from human coronary plaques ex vivo, to image plaque composition in terms of cholesterol and cholesterol ester content. In addition, we visualized the deeper lying connective tissue layers of the adventitia, as well as the fatty acid containing adipose cells in the peri-adventitial tissue. We performed simultaneous coregistered IVUS imaging to obtain complementary morphological information. Results were corroborated by histopathology. sIVPA imaging can distinguish the most prevalent lipid components of human atherosclerotic plaques and also visualize the connective tissue layers of the adventitia and the fatty acid containing adipose cells in the peri-adventitial tissue.

Expression of stanniocalcin-1 in culprit coronary plaques of patients with acute myocardial infarction or stable angina

BackgroundStanniocalcin-1 (STC1) is involved in fundamental biological processes such as angiogenesis, inflammation and wound healing, but little is known about its expression in human coronary atherosclerotic plaques or its relationship...

Classification of Coronary Atherosclerotic Plaques Ex Vivo With T1, T2, and Ultrashort Echo Time CMR

This study sought to determine whether the classification of human coronary atherosclerotic plaques with T1, T2, and ultrashort echo time (UTE) cardiac magnetic resonance (CMR) would correlate well with atherosclerotic plaque classification by histology. CMR has been extensively used to classify carotid plaque, but its ability to characterize coronary plaque remains unknown. In addition, the detection of plaque calcification by CMR remains challenging. Here, we used T1, T2, and UTE CMR to evaluate atherosclerotic plaques in fixed post-mortem human coronary arteries. We hypothesized that the combination of T1, T2, and UTE CMR would allow both calcified and lipid-rich coronary plaques to be accurately detected. Twenty-eight plaques from human donor hearts with proven coronary artery disease were imaged at 9.4-T with a T1-weighted 3-dimensional fast low-angle shot (FLASH) sequence (250-μm resolution), a T2-weighted rapid acquisition with refocused echoes (RARE) sequence (in-plane resolution 0.156 mm), and an UTE sequence (300-μm resolution). Plaques showing selective hypointensity on T2-weighted CMR were classified as lipid-rich. Areas of hypointensity on the T1-weighted images, but not the UTE images, were classified as calcified. Hyperintensity on the T1-weighted and UTE images was classified as hemorrhage. Following CMR, histological characterization of the plaques was performed with a pentachrome stain and established American Heart Association criteria. CMR showed high sensitivity and specificity for the detection of calcification (100% and 90%, respectively) and lipid-rich necrotic cores (90% and 75%, respectively). Only 2 lipid-rich foci were missed by CMR, both of which were extremely small. Overall, CMR-based classification of plaque was in complete agreement with the histological classification in 22 of 28 cases (weighted κ = 0.6945, p < 0.0001). The utilization of UTE CMR allows plaque calcification in the coronary arteries to be robustly detected. High-resolution CMR with T1, T2, and UTE contrast enables accurate classification of human coronary atherosclerotic plaque.

Salusins: Potential Use as a Biomarker for Atherosclerotic Cardiovascular Diseases

Human salusin-α and salusin-β are related peptides produced from prosalusin. Bolus injection of salusin-β into rats induces more profound hypotension and bradycardia than salusin-α. Central administration of salusin-β increases blood pressure via release of norepinephrine and arginine-vasopressin. Circulating levels of salusin-α and salusin-β are lower in patients with essential hypertension. Salusin-β exerts more potent mitogenic effects on human vascular smooth muscle cells (VSMCs) and fibroblasts than salusin-α. Salusin-β accelerates inflammatory responses in human endothelial cells and monocyte-endothelial adhesion. Human macrophage foam cell formation is stimulated by salusin-β but suppressed by salusin-α. Chronic salusin-β infusion into apolipoprotein E-deficient mice enhances atherosclerotic lesions; salusin-α infusion reduces lesions. Salusin-β is expressed in proliferative neointimal lesions of porcine coronary arteries after stenting. Salusin-α and salusin-β immunoreactivity have been detected in human coronary atherosclerotic plaques, with dominance of salusin-β in macrophage foam cells, VSMCs, and fibroblasts. Circulating salusin-β levels increase and salusin-α levels decrease in patients with coronary artery disease. These findings suggest that salusin-β and salusin-α may contribute to proatherogenesis and antiatherogenesis, respectively. Increased salusin-β and/or decreased salusin-α levels in circulating blood and vascular tissue are closely linked with atherosclerosis. Salusin-α and salusin-β could be candidate biomarkers and therapeutic targets for atherosclerotic cardiovascular diseases.

Optimizing the Imaging Protocol forEx VivoCoronary Artery Wall Using High-Resolution MRI: An Experimental Study on Porcine and Human

ObjectiveTo optimize the MR imaging protocol for coronary arterial wall depiction in vitro and characterize the coronary atherosclerotic plaques.Materials and MethodsMRI examination was prospectively performed in ten porcine hearts in order to optimize the MR imaging protocol. Various surface coils were used for coronary arterial wall imaging with the same parameters. Then, the image parameters were further optimized for high-resolution coronary wall imaging. The signal-noise ratio (SNR) and contrast-noise ratio (CNR) of images were measured. Finally, 8 human cadaver hearts with coronary atherosclerotic plaques were prospectively performed with MRI examination using optimized protocol in order to characterize the coronary atherosclerotic plaques.ResultsThe SNR and CNR of MR image with temporomandibular coil were the highest of various surface coils. High-resolution and high SNR and CNR for ex vivo coronary artery wall depiction can be achieved using temporomandibular coil with 512 × 512 in matrix. Compared with histopathology, the sensitivity and specificity of MRI for identifying advanced plaques were: type IV-V (lipid, necrosis, fibrosis), 94% and 95%; type VI (hemorrhage), 100% and 98%; type VII (calcification), 91% and 100%; and type VIII (fibrosis without lipid core), 100% and 98%, respectively.ConclusionTemporomandibular coil appears to be dramatically superior to eight-channel head coil and knee coil for ex vivo coronary artery wall imaging, providing higher spatial resolution and improved the SNR. Ex vivo high-resolution MRI has capability to distinguish human coronary atherosclerotic plaque compositions and accurately classify advanced plaques.

Subcellular localization of tissue factor and human coronary artery smooth muscle cell migration

Tissue factor (TF) is the most relevant physiological trigger of thrombosis. Additionally TF is a transmembrane receptor with cell signaling functions. The aim of this study was to investigate TF subcellular localization, function and signaling in human coronary artery smooth muscle cell migration. Coronary arteries and primary cultures of vascular smooth muscle cells (HVSMC) were obtained from human explanted hearts. Wound repair and Boyden chamber assays were used to measure migration in vitro. TF-pro-coagulant activity (TF-PCA) was measured in extracted cellular membranes. Analysis of TF distribution was performed by confocal microscopy. A nucleofector device was used for TF and protease activated receptor 2 (PAR2) silencing. mRNA levels were analyzed by RT-PCR. In migrating HVSMC TF translocates to the leading edge of the cells showing an intense patch-like staining in the lamellipodia. In the migrating front TF colocalizes with filamin (FLN) in the polarized lipid rafts. TF-PCA was increased in migrating cells. Silencing of the TF gene inhibits RSK-induced FLN-Ser-2152 phosphorylation, down-regulates CDC42, RhoA, and Rac1 protein expression and significantly inhibits cell migration. Silencing PAR2 also inhibits cell migration; however, silencing both TF and PAR2 induces a significantly higher effect on migration. Smooth muscle cells expressing TF have been identified in non-lipid-rich human coronary artery atherosclerotic plaques. TF translocates to the cell front in association with cytoskeleton proteins and regulates HVSMC migration by mechanisms dependent and independent of factor (F)VIIa/PAR2. These results extend the functional role of TF to smooth muscle cell trafficking in vessel wall remodeling.

Classification of Human Coronary Atherosclerotic Plaques Using Ex Vivo High-Resolution Multicontrast-Weighted MRI Compared With Histopathology

The objective of our study was to evaluate the feasibility of ex vivo high-resolution multicontrast-weighted MRI to accurately classify human coronary atherosclerotic plaques according to the American Heart Association classification. Thirteen human cadaver heart specimens were imaged using high-resolution multicontrast-weighted MR technique (T1-weighted, proton density-weighted, and T2-weighted). All MR images were matched with histopathologic sections according to the landmark of the bifurcation of the left main coronary artery. The sensitivity and specificity of MRI for the classification of plaques were determined, and Cohen's kappa analysis was applied to evaluate the agreement between MRI and histopathology in the classification of atherosclerotic plaques. One hundred eleven MR cross-sectional images obtained perpendicular to the long axis of the proximal left anterior descending artery were successfully matched with the histopathologic sections. For the classification of plaques, the sensitivity and specificity of MRI were as follows: type I-II (near normal), 60% and 100%; type III (focal lipid pool), 80% and 100%; type IV-V (lipid, necrosis, fibrosis), 96.2% and 88.2%; type VI (hemorrhage), 100% and 99.0%; type VII (calcification), 93% and 100%; and type VIII (fibrosis without lipid core), 100% and 99.1%, respectively. Isointensity, which indicates lipid composition on histopathology, was detected on MRI in 48.8% of calcified plaques. Agreement between MRI and histopathology for plaque classification was 0.86 (p < 0.001). Ex vivo high-resolution multicontrast-weighted MRI can accurately classify advanced atherosclerotic plaques in human coronary arteries.

The fibrillogenic L178H variant of apolipoprotein A-I forms helical fibrils

A number of amyloidogenic variants of apoA-I have been discovered but most have not been analyzed. Previously, we showed that the G26R mutation of apoA-I leads to increased β-strand structure, increased N-terminal protease susceptibility, and increased fibril formation after several days of incubation. In vivo, this and other variants mutated in the N-terminal domain (residues 26 to ∼90) lead to renal and hepatic accumulation. In contrast, several mutations identified within residues 170 to 178 lead to cardiac, laryngeal, and cutaneous protein deposition. Here, we describe the structural changes in the fibrillogenic variant L178H. Like G26R, the initial structure of the protein exhibits altered tertiary conformation relative to wild-type protein along with decreased stability and an altered lipid binding profile. However, in contrast to G26R, L178H undergoes an increase in helical structure upon incubation at 37°C with a half time (t1/2) of about 12 days. Upon prolonged incubation, the L178H mutant forms fibrils of a diameter of 10 nm that ranges in length from 30 to 120 nm. These results show that apoA-I, known for its dynamic properties, has the ability to form multiple fibrillar conformations, which may play a role in the tissue-specific deposition of the individual variants.