Improving stroke risk prediction and individualised treatment in carotid atherosclerosis

Background: Unstable carotid atherosclerosis causes stroke, but methods to identify patients and lesions at risk are lacking. Currently, this risk estimation is based on measurements of stenosis and neurological symptoms, which determines the therapy of either medical treatment with or without carotid endarterectomy. The efficacy of this therapy is low and higher accuracy of diagnosis and therapy is warranted. Imaging of carotid plaque morphology using software for visualisation of plaque components may improve assessment of plaque phenotype and stroke risk. These studies aimed firstly to investigate if, and if yes, how, the carotid plaque morphology with image analysis of CTA associated with on-going biology in the corresponding specimen. Secondly, if risk stratification in clinical risk scores can be linked to the aforementioned associations. Finally, if the on-going biological processes can be specifically predicted out of the CTA imaging analysis. Methods: Plaque features were analysed in pre-operative CTA with dedicated software. In study I and II, the plaques were stratified according to quantified high and low of each feature, profiled with microarrays, followed by bioinformatic analyses. Immunohistochemistry was performed to evaluate the findings in plaques. In study III, patient phenotype, according to clinical stroke risk scores of CAR and ABCD2 stratified the cohorts of high vs low scores which were subsequently profiled with microarrays, followed by bioinformatic analyses and correlation analyses of plaque morphology in CTA. In study IV, the microarray transcriptomes were individually coupled to morphological data from the CTA analysis, developing models with machine intelligence to predict the gene expression from a CTA image. The models were then tested in unseen patients. Results: In study I, stabilising markers and processes related to SMCs and ECM organisation were associated with highly calcified plaques, while inflammatory and lipid related processes were repressed. PRG4, a novel marker for atherosclerosis, was identified as the most up-regulated gene in highly calcified plaques. Study II showed that carotid lesions with large lipid rich necrotic core, intraplaque haemorrhage or plaque burden were characterized by molecular signatures coupled with inflammation and extracellular matrix degradation, typically linked with instability. Symptomatology associated with large lipid rich necrotic core and plaque burden. Cross-validated prediction model for symptoms, showed that plaque morphology by CTA alone was superior to stenosis degree. Study III revealed that a high clinical risk score in CAR and ABCD2, reflect a plaque phenotype linked to immune response and coagulation, where the novel ABCB5, was one of the most up-regulated genes. The high risk scores correlated with the plaque components matrix and calcification but no positive association with stenosis degree. Study IV resulted in 414 robustly predicted transcripts from the CTA image analysis, of which pathway analysis showed biological processes associated with typical pathophysiology of atherosclerosis and plaque instability. The model testing demonstrated a good correlation between predicted and observed transcript expression levels and pathway analysis revealed a unique dominant mechanism for each individual. Conclusions: Biological processes in carotid plaques associated to vulnerability, can be linked to plaque morphology analysed with CTA image analysis. Patient phenotype classified with clinical risk scores associates to plaque phenotype and morphology in CTA. The biological processes in the atherosclerotic plaque can be predicted with plaque morphology CTA analysis in this small pilot study, providing a possibility to precision medicine after validation in larger scale studie

Characterization of Carotid Smooth Muscle Cells during Phenotypic Transition

Vascular smooth muscle cells (VSMCs) are central players in carotid atherosclerosis plaque development. Although the precise mechanisms involved in plaque destabilization are not completely understood, it is known that VSMC proliferation and migration participate in plaque stabilization. In this study, we analyzed expression patterns of genes involved in carotid atherosclerosis development (e.g., transcription factors of regulation of SMC genes) of VSMCs located inside or outside the plaque lesion that may give clues about changes in phenotypic plasticity during atherosclerosis. VSMCs were isolated from 39 carotid plaques extracted from symptomatic and asymptomatic patients by endarterectomy. Specific biomarker expression, related with VSMC phenotype, was analyzed by qPCR, western immunoblot, and confocal microscopy. MYH11, CNN1, SRF, MKL2, and CALD1 were significantly underexpressed in VSMCs from plaques compared with VSMCs from a macroscopically intact (MIT) region, while SPP1, KLF4, MAPLC3B, CD68, and LGALS3 were found significantly upregulated in plaque VSMCs versus MIT VSMCs. The gene expression pattern of arterial VSMCs from a healthy donor treated with 7-ketocholesterol showed high similarity with the expression pattern of carotid plaque VSMCs. Our results indicate that VSMCs isolated from plaque show a typical SMC dedifferentiated phenotype with macrophage-like features compared with VSMCs isolated from a MIT region of the carotid artery. Additionally, MYH11, KLF5, and SPP1 expression patterns were found to be associated with symptomatology of human carotid atherosclerosis.This work was supported by Spanish Institute for Health Carlos III, RETICS program (Grant Number RD16/0019/0007) and by Basque Government, Education Department, Consolidated Groups program (Grant Number IT512-10)

Smooth muscle cell characterization and transcriptomic analysis in human carotid atherosclerotic plaques

145 p.Las enfermedades cardiovasculares (CV) son una de las causas más importantes de discapacidad y muerte prematura en todo el mundo. Una de las causas subyacentes de las CV es la aterosclerosis. La aterosclerosis de carótida se caracteriza por el desarrollo de una placa de ateroma, la cual puede producir oclusión del vaso sangúineo, causando una esquemia cerebral o ictus. La formación de estas placas está fuertemente influenciada por las células de músculo liso (CML) que se encuentran en la pared arterial y que participan tanto en la formación como ruptura de la placa (y posterior isquemia) gracias a la plasticidad fenotípica que retienen. Sin embargo, a día de hoy, no se conoce el rol exacto de las CML en la aterosclerosis, ni tampoco el mecanismo de rutpura de la placa. Por ello, con el objetivo de buscar un biomarcador que identifique una placa con altas probabilidades de romperese, y así, poder evitar el ictus., se ha realizado la caracterización y secuenciación del tansciptoma completo de las CML de placas de ateroma carotideas provenientes de pacientes sintomáticos (han sufrido un ictus) y asintomáticos (sin ningún síntoma). Se ha conseguido definir varios roles para las CML: por un lado, se ha asociado la senescencia celular a un tipo de placa inestable (propensa a romperse), mientras que la calcificación es más frecuente en placas estables (sin ruptura). Además, se han detectado varias moléculas (BMP2, ID1 e ID4) de alto interés pronóstico.Achucarro Basque Center for Neuroscienc

Multimodality carotid plaque tissue characterization and classification in the artificial intelligence paradigm: a narrative review for stroke application

Cardiovascular disease (CVD) is one of the leading causes of morbidity and mortality in the United States of America and globally. Carotid arterial plaque, a cause and also a marker of such CVD, can be detected by various non-invasive imaging modalities such as magnetic resonance imaging (MRI), computer tomography (CT), and ultrasound (US). Characterization and classification of carotid plaque-type in these imaging modalities, especially into symptomatic and asymptomatic plaque, helps in the planning of carotid endarterectomy or stenting. It can be challenging to characterize plaque components due to (I) partial volume effect in magnetic resonance imaging (MRI) or (II) varying Hausdorff values in plaque regions in CT, and (III) attenuation of echoes reflected by the plaque during US causing acoustic shadowing. Artificial intelligence (AI) methods have become an indispensable part of healthcare and their applications to the non-invasive imaging technologies such as MRI, CT, and the US. In this narrative review, three main types of AI models (machine learning, deep learning, and transfer learning) are analyzed when applied to MRI, CT, and the US. A link between carotid plaque characteristics and the risk of coronary artery disease is presented. With regard to characterization, we review tools and techniques that use AI models to distinguish carotid plaque types based on signal processing and feature strengths. We conclude that AI-based solutions offer an accurate and robust path for tissue characterization and classification for carotid artery plaque imaging in all three imaging modalities. Due to cost, user-friendliness, and clinical effectiveness, AI in the US has dominated the most

Smooth muscle cell characterization and transcriptomic analysis in human carotid atherosclerotic plaques

145 p.Las enfermedades cardiovasculares (CV) son una de las causas más importantes de discapacidad y muerte prematura en todo el mundo. Una de las causas subyacentes de las CV es la aterosclerosis. La aterosclerosis de carótida se caracteriza por el desarrollo de una placa de ateroma, la cual puede producir oclusión del vaso sangúineo, causando una esquemia cerebral o ictus. La formación de estas placas está fuertemente influenciada por las células de músculo liso (CML) que se encuentran en la pared arterial y que participan tanto en la formación como ruptura de la placa (y posterior isquemia) gracias a la plasticidad fenotípica que retienen. Sin embargo, a día de hoy, no se conoce el rol exacto de las CML en la aterosclerosis, ni tampoco el mecanismo de rutpura de la placa. Por ello, con el objetivo de buscar un biomarcador que identifique una placa con altas probabilidades de romperese, y así, poder evitar el ictus., se ha realizado la caracterización y secuenciación del tansciptoma completo de las CML de placas de ateroma carotideas provenientes de pacientes sintomáticos (han sufrido un ictus) y asintomáticos (sin ningún síntoma). Se ha conseguido definir varios roles para las CML: por un lado, se ha asociado la senescencia celular a un tipo de placa inestable (propensa a romperse), mientras que la calcificación es más frecuente en placas estables (sin ruptura). Además, se han detectado varias moléculas (BMP2, ID1 e ID4) de alto interés pronóstico.Achucarro Basque Center for Neuroscienc

Correlation of computed tomography with carotid plaque transcriptomes associates calcification with lesion-stabilization

Background and aims: Unstable carotid atherosclerosis causes stroke, but methods to identify patients and lesions at risk are lacking. We recently found enrichment of genes associated with calcification in carotid plaques from asymptomatic patients. Here, we hypothesized that calcification represents a stabilising feature of plaques and investigated how macro-calcification, as estimated by computed tomography (CT), correlates with gene expression profiles in lesions. Methods: Plaque calcification was measured in pre-operative CT angiographies. Plaques were sorted into high- and lowcalcified, profiled with microarrays, followed by bioinformatic analyses. Immunohistochemistry and qPCR were performed to evaluate the findings in plaques and arteries with medial calcification from chronic kidney disease patients. Results: Smooth muscle cell (SMC) markers were upregulated in high-calcified plaques and calcified plaques from symptomatic patients, whereas macrophage markers were downregulated. The most enriched processes in high-calcified plaques were related to SMCs and extracellular matrix (ECM) organization, while inflammation, lipid transport and chemokine signaling were repressed. These findings were confirmed in arteries with high medial calcification. Proteoglycan 4 (PRG4) was identified as the most upregulated gene in association with plaque calcification and found in the ECM, SMA+ and CD68+/TRAP + cells. Conclusions: Macro-calcification in carotid lesions correlated with a transcriptional profile typical for stable plaques, with altered SMC phenotype and ECM composition and repressed inflammation. PRG4, previously not described in atherosclerosis, was enriched in the calcified ECM and localized to activated macrophages and smooth muscle-like cells. This study strengthens the notion that assessment of calcification may aid evaluation of plaque phenotype and stroke risk.The European Union’s Horizon 2020/Marie Sklodowska-Curie grant agreement No 722609 (INTRICARE);Swedish Heart and Lung FoundationSwedish Research Council (K2009-65X-2233-01-3, K2013- 65X-06816-30-4, 349-2007-8703)Uppdrag Besegra Stroke (P581/ 2011-123)Stockholm County Council (ALF2011-0260, ALF-2011- 0279)Swedish Society for Medical ResearchTore Nilsson’s FoundationMagnus Bergvall’s FoundationKarolinska Institutet FoundationEuropean Commission (722609)Publishe

Carotid Artery Wall Imaging: Perspective and Guidelines from the ASNR Vessel Wall Imaging Study Group and Expert Consensus Recommendations of the American Society of Neuroradiology

SUMMARY: Identification of carotid artery atherosclerosis is conventionally based on measurements of luminal stenosis and surface irregularities using in vivo imaging techniques including sonography, CT and MR angiography, and digital subtraction angiography. However, histopathologic studies demonstrate considerable differences between plaques with identical degrees of stenosis and indicate that certain plaque features are associated with increased risk for ischemic events. The ability to look beyond the lumen using highly developed vessel wall imaging methods to identify plaque vulnerable to disruption has prompted an active debate as to whether a paradigm shift is needed to move away from relying on measurements of luminal stenosis for gauging the risk of ischemic injury. Further evaluation in randomized clinical trials will help to better define the exact role of plaque imaging in clinical decision-making. However, current carotid vessel wall imaging techniques can be informative. The goal of this article is to present the perspective of the ASNR Vessel Wall Imaging Study Group as it relates to the current status of arterial wall imaging in carotid artery disease

The impact of carotid plaque presence and morphology on mortality outcome in cardiological patients

BACKGROUND: Carotid plaque severity and morphology can affect cardiovascular prognosis. We evaluate both the importance of echographically assessed carotid artery plaque geometry and morphology as predictors of death in hospitalised cardiological patients. METHODS: 541 hospitalised patients admitted in a cardiological division (age = 66 ± 11 years, 411 men), have been studied through ultrasound Duplex carotid scan and successively followed-up for a median of 34 months. Echo evaluation assessed plaque severity and morphology (presence of heterogeneity and profile). RESULTS: 361 patients showed carotid stenosis (67% with <50% stenosis, 18% with 50–69% stenosis, 9% with >70% stenosis, 4% with near occlusion and 2% with total occlusion). During the follow-up period, there were 83 all-cause deaths (15% of the total population). Using Cox's proportional hazard model, age (RR 1.06, 95% CI 1.03–1.09, p = 0.000), ejection fraction > 50% (RR = 0.62, 95% CI 0.4–0.96, p = 0.03), treatment with statins (RR = 0.52, 95% CI 0.29–0.95, p = 0.34) and the presence of a heterogeneous plaque (RR 1.6; 95% CI, 1.2 to 2.14, p = 0.002) were independent predictors of death. Kaplan – Meier survival estimates have shown the best outcome in patients without plaque, intermediate in patients with homogeneous plaques and the worst outcome in patients with heterogeneous plaques (90% vs 79% vs 73%, p = 0.0001). CONCLUSION: In hospitalised cardiological patients, carotid plaque presence and morphology assessed by ultrasound are independent predictors of death

Carotid Plaque Imaging with SPECT/CT and PET/CT

A major contributor to the occurrence of ischemic stroke is the existence of carotid atherosclerosis. A vulnerable carotid atherosclerotic plaque may rupture or erode, thus causing a thrombotic event. Currently, clinical decision-making with regard to carotid endarterectomy or stenting is still primarily based on the extent of luminal stenosis, estimated with CT angiography and/or (duplex) ultrasonography. However, there is growing evidence that the anatomic impact of stenosis alone has limited value in predicting the exact consequences of plaque vulnerability. Various molecular processes have, independently of degree of stenosis, shown to be importantly associated with the plaque's capability to cause thrombotic events. These molecular processes can be visualized with nuclear medicine techniques allowing the identification of vulnerable patients by non-invasive in vivo SPECT(/CT) and PET(/CT) imaging. This chapter provides an overview of SPECT(/CT) and PET(/CT) imaging with specific radiotracers that have been evaluated for the detection of plaques together with a future perspective in this field of imaging.</p

International Union of Angiology (IUA) consensus paper on imaging strategies in atherosclerotic carotid artery imaging: From basic strategies to advanced approaches

Cardiovascular disease (CVD) is the leading cause of mortality and disability in developed countries. According to WHO, an estimated 17.9 million people died from CVDs in 2019, representing 32% of all global deaths. Of these deaths, 85% were due to major adverse cardiac and cerebral events. Early detection and care for individuals at high risk could save lives, alleviate suffering, and diminish economic burden associated with these diseases. Carotid artery disease is not only a well-established risk factor for ischemic stroke, contributing to 10%–20% of strokes or transient ischemic attacks (TIAs), but it is also a surrogate marker of generalized atherosclerosis and a predictor of cardiovascular events. In addition to diligent history, physical examination, and laboratory detection of metabolic abnormalities leading to vascular changes, imaging of carotid arteries adds very important information in assessing stroke and overall cardiovascular risk. Spanning from carotid intima-media thickness (IMT) measurements in arteriopathy to plaque burden, morphology and biology in more advanced disease, imaging of carotid arteries could help not only in stroke prevention but also in ameliorating cardiovascular events in other territories (e.g. in the coronary arteries). While ultrasound is the most widely available and affordable imaging methods, computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET), their combination and other more sophisticated methods have introduced novel concepts in detection of carotid plaque characteristics and risk assessment of stroke and other cardiovascular events. However, in addition to robust progress in usage of these methods, all of them have limitations which should be taken into account. The main purpose of this consensus document is to discuss pros but also cons in clinical, epidemiological and research use of all these techniques