Atherosclerosis Imaging Research Articles

Advancements in Precision Medicine for Imaging and Pharmacotherapy in Atherosclerosis: Recent Discoveries and Emerging Technologies

Advancements in Precision Medicine for Imaging and Pharmacotherapy in Atherosclerosis: Recent Discoveries and Emerging Technologies

Diagnostic Performance and Clinical Impact of Photon-Counting Detector Computed Tomography in Coronary Artery Disease.

Photon-counting detector-computed tomography (PCD-CT) has emerged as a promising technology, offering improved spatial resolution. This study aimed to evaluate the clinical impact and diagnostic performance of PCD-CT vs conventional energy-integrating detector computed tomography (EID-CT) for obstructive coronary artery disease (CAD). From 2022 to 2023, we retrospectively identified 7,833 consecutive patients who underwent clinically indicated coronary computed tomography angiography (CCTA) at a single center, with either PCD-CT (n=3,876; NAEOTOM Alpha [Siemens Healthineers]) or EID-CT (n=3,957; Revolution Apex 256 [GE HealthCare] or Aquilion ONE ViSION 320 [Canon Medical Systems]) scanners. Subsequent invasive coronary angiography (ICA) and percutaneous or surgical revascularization were performed as part of routine clinical care. Among those referred for ICA after coronary CTA, the presence of obstructive CAD in each vessel was determined by coronary CTA (severe stenosis on visual assessment per the Coronary Artery Disease Reporting and Data System) and ICA (≥50% diameter stenosis on quantitative coronary angiography) in a blinded fashion. The diagnostic performance of EID-CT and PCD-CT was compared by using quantitative coronary angiography as the reference standard. Patients who underwent PCD-CT were less frequently referred to subsequent ICA than those undergoing EID-CT (9.9% vs 13.1%; P< 0.001). Among those who underwent ICA, revascularization was more frequently performed in the PCD-CT group than in the EID-CT group (43.4% vs 35.5%; P=0.02). In the vessel-level analysis (n=1,686), specificity (98.0% vs 93.0%; P< 0.001), positive predictive value (83.3% vs 63.0%; P=0.002), and diagnostic accuracy (97.2% vs 92.8%; P< 0.001) were improved by PCD-CT. Sensitivity (90.9% vs 90.7%; P=0.95) and negative predictive value (98.9% vs 98.7%; P=0.83) for obstructive CAD were similar between the PCD-CT and EID-CT groups, respectively. PCD-CT exhibited excellent diagnostic performance for detecting obstructive CAD. Compared with patients undergoing conventional EID-CT, fewer patients were referred to ICA after PCD-CT, but those referred were more likely to undergo revascularization.

TCT-513 Clinical Impact and Diagnostic Performance of Photon-Counting Detector Computed Tomography in Coronary Artery Disease

TCT-513 Clinical Impact and Diagnostic Performance of Photon-Counting Detector Computed Tomography in Coronary Artery Disease

Novel Metal-Free Nanozyme for Targeted Imaging and Inhibition of Atherosclerosis via Macrophage Autophagy Activation to Prevent Vulnerable Plaque Formation and Rupture.

Atherosclerosis is a primary cause of cardiovascular and cerebrovascular diseases, with the unpredictable rupture of vulnerable atherosclerotic plaques enriched with lipid-laden macrophages being able to lead to heart attacks and strokes. Activating macrophage autophagy presents itself as a promising strategy for preventing vulnerable plaque formation and reducing the risk of rupture. In this study, we have developed a novel metal-free nanozyme (HCN@DS) that integrates the functions of multimodal imaging-guided therapy for atherosclerosis. HCN@DS has demonstrated high macrophage-targeting abilities due to its affinity toward scavenger receptor A (SR-A), along with excellent photoacoustic and photothermal imaging capabilities for guiding the precise treatment. It combines mild photothermal effects with moderate reactive oxygen species (ROS) generation to treat atherosclerosis. This controlled approach activates autophagy in atherosclerotic macrophages, inhibiting foam cell formation by reducing the uptake of oxidized low-density lipoproteins (oxLDL) and promoting efferocytosis and cholesterol efflux in macrophages. Additionally, it prevents plaque rupture by inhibiting apoptosis and inflammation within the plaque. Therefore, this metal-free nanozyme holds great potential for reducing the risk of atherosclerosis due to its high biosafety, excellent targeting ability, dual-modality imaging capability, and appropriate modulation of autophagy.

Recent Advancement of Indocyanine Green Based Nanotheranostics for Imaging and Therapy of Coronary Atherosclerosis.

Atherosclerosis is a vascular intima condition in which any part of the circulatory system is affected, including the aorta and coronary arteries. Indocyanine green (ICG), a theranostic compound approved by the FDA, has shown promise in the treatment of coronary atherosclerosis after incorporation into nanoplatforms. By integration of ICG with targeting agents such as peptides or antibodies, it is feasible to increase its concentration in damaged arteries, hence increasing atherosclerosis detection. Nanotheranostics offers cutting-edge techniques for the clinical diagnosis and therapy of atherosclerotic plaques. Combining the optical properties of ICG with those of nanocarriers enables the improved imaging of atherosclerotic plaques and targeted therapeutic interventions. Several ICG-based nanotheranostics platforms have been developed such as polymeric nanoparticles, iron oxide nanoparticles, biomimetic systems, liposomes, peptide-based systems, etc. Theranostics for atherosclerosis diagnosis use magnetic resonance imaging (MRI), computed tomography (CT), near-infrared fluorescence (NIRF) imaging, photoacoustic/ultrasound imaging, positron emission tomography (PET), and single photon emission computed tomography (SPECT) imaging techniques. In addition to imaging, there is growing interest in employing ICG to treat atherosclerosis. In this review, we provide a conceptual explanation of ICG-based nanotheranostics for the imaging and therapy of coronary atherosclerosis. Moreover, advancements in imaging modalities such as MRI, CT, PET, SPECT, and ultrasound/photoacoustic have been discussed. Furthermore, we highlight the applications of ICG for coronary atherosclerosis.

The association of cardiovascular disease risk with coronary artery calcification and thoracic aortic dilation: a study in idiopathic inflammatory myopathies and systemic lupus erythematosus.

We aim to explore the prevalence of coronary artery calcification (CAC) and ascending/descending thoracic aorta (AA/DA) dilation in idiopathic inflammatory myopathies (IIM) and systemic lupus erythematosus (SLE) patients, and to assess associations between cardiovascular disease (CVD) risk factors and these imaging signatures. This study recruited 151 IIM patients, 140 SLE patients, and 195 controls. The CAC and AA/DA diameters were quantified using non-gated chest CT images. The independent samples t-test or Mann-Whitney test was chosen for comparisons of continuous variables between patients and healthy controls. For categorical data, comparisons were made using the chi-square test or Fisher's exact test. Multivariate regression or Spearman's correlation analysis was employed to probe the associations between CVD risk factors and Framingham risk score (FRS) with imaging signatures. The IIM and SLE patients showed significantly higher prevalence of CAC and AA/DA dilatation (P < 0.01). Age was a risk factor for both CAC and AA/DA dilatation in all cohorts (P < 0.01). In IIM patients, the AA/DA dilatation was associated with BMI (P = 0.05). In SLE patients, CAC was associated with the elevated CRP level (P = 0.05). Without CAC, both IIM and SLE patients showed significant correlations between AA/DA diameters and FRS (P < 0.01, P < 0.01). Only in SLE patients, the interleukin-6 (IL-6) level correlated with AA/DA diameters. The IIM and SLE patients more commonly exhibit CAC and AA/DA dilation. These subclinical atherosclerosis signs are associated with traditional CVD risk factors. For AID patients without CAC, AA/DA diameters could serve as a potential biomarker for early CVD risk. Key Points • The study characterized the manifestation of subclinical atherosclerosis imaging biomarkers (CAC, AA/DA dilation) in IIM and SLE patients. • AA/DA diameters could serve as an early imaging biomarker in clinical management for IIM and SLE patients with early-onset and no CAC present.

Atherosclerosis quantification and cardiovascular risk: the ISCHEMIA trial.

The aim of this study was to determine the prognostic value of coronary computed tomography angiography (CCTA)-derived atherosclerotic plaque analysis in ISCHEMIA. Atherosclerosis imaging quantitative computed tomography (AI-QCT) was performed on all available baseline CCTAs to quantify plaque volume, composition, and distribution. Multivariable Cox regression was used to examine the association between baseline risk factors (age, sex, smoking, diabetes, hypertension, ejection fraction, prior coronary disease, estimated glomerular filtration rate, and statin use), number of diseased vessels, atherosclerotic plaque characteristics determined by AI-QCT, and a composite primary outcome of cardiovascular death or myocardial infarction over a median follow-up of 3.3 (interquartile range 2.2-4.4) years. The predictive value of plaque quantification over risk factors was compared in an area under the curve (AUC) analysis. Analysable CCTA data were available from 3711 participants (mean age 64 years, 21% female, 79% multivessel coronary artery disease). Amongst the AI-QCT variables, total plaque volume was most strongly associated with the primary outcome (adjusted hazard ratio 1.56, 95% confidence interval 1.25-1.97 per interquartile range increase [559 mm3]; P = .001). The addition of AI-QCT plaque quantification and characterization to baseline risk factors improved the model's predictive value for the primary outcome at 6 months (AUC 0.688 vs. 0.637; P = .006), at 2 years (AUC 0.660 vs. 0.617; P = .003), and at 4 years of follow-up (AUC 0.654 vs. 0.608; P = .002). The findings were similar for the other reported outcomes. In ISCHEMIA, total plaque volume was associated with cardiovascular death or myocardial infarction. In this highly diseased, high-risk population, enhanced assessment of atherosclerotic burden using AI-QCT-derived measures of plaque volume and composition modestly improved event prediction.

Near-Infrared II Fluorescence Imaging and Image-Guided siRNA Therapy of Atherosclerosis.

Targeting Ca2+/calmodulin-dependent protein kinase γ (CaMKIIγ) in macrophages using RNAi nanotechnology represents an innovative and promising strategy in the diagnosis and treatment of atherosclerosis. Nevertheless, it remains elusive because of the current challenges associated with the systemic delivery of siRNA nanoparticle (NP) to atheromatous plaques and the complexity of atherosclerotic plaques. Here, we demonstrate the potential of a thienothiadiazole-based near-infrared-II (NIR-II) organic aggregation-induced emission (AIE) platform encapsulated with the Camk2g siRNA to effectively target CaMKIIγ in macrophages for dynamic imaging and image-guided gene therapy of atherosclerosis. The nanoparticles effectively decreased CaMKIIγ expression and increased the expression of the efferocytosis receptor MerTK in plaque macrophages, leading to a reduction in the necrotic core area of the lesion in an aortic plaque model. Our theranostic approach highlights the substantial promise of near-infrared II (NIR-II) AIEgens for imaging and image-guided therapy of atherosclerosis.

Near-infrared multispectral photoacoustic analysis of lipids and intraplaque hemorrhage in human carotid artery atherosclerosis

Spectral photoacoustic imaging in combination with unmixing techniques may be applied to retrieve information about high-risk features present in atherosclerotic plaques, possibly providing prognostic insights into future stroke events. We present the photoacoustic spectral contrast found in 12 systematically scanned advanced atherosclerotic plaques in the near-infrared wavelength range (850–1250 nm). The main absorbers are lipid, water, and hemoglobin, with the highest photoacoustic intensities at the lipid’s second overtone at 1190 and 1210 nm. Linear unmixing resulted in visualizing regions with high lipid and hemoglobin absorption, corresponding to the histological presence of lipid and intraplaque hemorrhage. A non-negative matrix factorization approach reveals differences in lipid spectral contrast, providing potential insights into the vulnerability of atherosclerotic plaque. These results provide a reference for future, more complex, in vivo photoacoustic imaging of carotid artery atherosclerosis, potentially contributing to assessing the risk of future events and treatment decision.

Cardiovascular Imaging for Coronary Artery Disease in Patients with Diabetes Mellitus.

In patients with diabetes mellitus, accelerated progression of atherosclerosis can lead to worse clinical outcomes. Determining the best diagnostic strategy to identify patients with increased cardiovascular risk is challenging. Current guidelines recommend using both functional imaging and CT angiography to detect myocardial ischemia and coronary artery disease based on pre-test probability. Functional imaging is suggested for patients with a higher clinical likelihood due to its higher rule-in diagnostic capacity. On the other hand, CT angiography is preferred for patients with lower pre-test probability because of its excellent negative predictive value. The optimal management strategy for asymptomatic diabetic patients remains unclear. In asymptomatic diabetic patients, previous randomized trials have not shown benefits from diagnostic testing over standard care. However, these trials were methodologically inconsistent and lacked clear stratification of cardiovascular risk. In terms of invasive evaluation, a combined invasive functional and anatomic imaging approach for angiographically intermediate coronary stenosis appears to be the best, most effective decision pathway for managing diabetic patients.

Reactive oxygen species-responsive nano-platform with dual-targeting and fluorescent lipid-specific imaging capabilities for the management of atherosclerotic plaques

Atherosclerosis (AS), a pathological cause of cardiovascular disease, results from endothelial injury, local progressive inflammation, and excessive lipid accumulation. AS plaques rich in foam cells are prone to rupture and form thrombus, which can cause life-threatening complications. Therefore, the assessment of atherosclerotic plaque vulnerability and early intervention are crucial in reducing the mortality rates associated with cardiovascular disease. In this work, A fluorescent probe FC-TPA was synthesized, which switches the fluorescence state between protonated and non-protonated, reducing background fluorescence and enhancing imaging signal-to-noise ratio. On this basis, FC-TPA is loaded into cyclodextrin (CD) modified with phosphatidylserine targeting peptide (PTP) and coated with hyaluronic acid (HA) to construct the intelligent responsive diagnostic nanoplatform (HA@PCFT). HA@PCFT effectively targets atherosclerotic plaques, utilizing dual targeting mechanisms. HA binds strongly to CD44, while PTP binds to phosphatidylserine, enabling nanoparticle aggregation at the lesion site. ROS acts as a smart release switch for probes. Both in vitro and in vivo evaluations confirm impressive lipid-specific fluorescence imaging capabilities of HA@PCFT nanoparticles (NPs). The detection of lipid load in atherosclerotic plaque by fluorescence imaging will aid in assessing the vulnerability of atherosclerotic plaque. Statement of significanceCurrently, numerous fluorescent probes have been developed for lipid imaging. However, some challenges including inadequate water solubility, nonspecific distribution patterns, and fluorescence background interference, have greatly limited their further applications in vivo. To overcome these limitations, a fluorescent molecule has been designed and synthesized, thoroughly investigating its photophysical properties through both theoretical and experimental approaches. Interestingly, this fluorescent molecule exhibits the reversible fluorescence switching capabilities, mediated by hydrogen bonds, which effectively mitigate background fluorescence interference. Additionally, the fluorescent molecules has been successfully loaded into nanocarriers functionalized with the active targeting abilities, which has significantly improved the solubility of the fluorescent molecules and reduced their nonspecific distribution in vivo for an efficient target imaging in atherosclerosis. This study provides a valuable reference for evaluating the performance of such fluorescent dyes, and offers a promising perspective on the design of the target delivery systems for atherosclerosis.

Repeated Injection of Very Small Superparamagnetic Iron Oxide Particles (VSOPs) in Murine Atherosclerosis: A Safety Study.

Citrate-coated electrostatically stabilized very small superparamagnetic iron oxide particles (VSOPs) have been successfully tested as magnetic resonance angiography (MRA) contrast agents and are promising tools for molecular imaging of atherosclerosis. Their repeated use in the background of pre-existing hyperlipidemia and atherosclerosis has not yet been studied. This study aimed to investigate the effect of multiple intravenous injections of VSOPs in atherosclerotic mice. Taurine-formulated VSOPs (VSOP-T) were repeatedly intravenously injected at 100 µmol Fe/kg in apolipoprotein E-deficient (ApoE KO) mice with diet-induced atherosclerosis. Angiographic imaging was carried out by in vivo MRI. Magnetic particle spectrometry was used to detect tissue VSOP content, and tissue iron content was quantified photometrically. Pathological changes in organs, atherosclerotic plaque development, and expression of hepatic iron-related proteins were evaluated. VSOP-T enabled the angiographic imaging of heart and blood vessels with a blood half-life of one hour. Repeated intravenous injection led to VSOP deposition and iron accumulation in the liver and spleen without affecting liver and spleen pathology, expression of hepatic iron metabolism proteins, serum lipids, or atherosclerotic lesion formation. Repeated injections of VSOP-T doses sufficient for MRA analyses had no significant effects on plaque burden, steatohepatitis, and iron homeostasis in atherosclerotic mice. These findings underscore the safety of VSOP-T and support its further development as a contrast agent and molecular imaging tool.

Ros-responsive nano-platform for lipid-specific fluorescence imaging of atherosclerosis

Fluorescent probes that specifically targeting Lipid droplets (LDs) have shown potential in biological imaging. Albeit, their in vivo applications are limited due to the hydrophobicity, low signal-to-noise ratio (SNR) and LDs-specificity. Thus, we designed a novel probe namely MeOND, and a reactive oxygen species (ROS)-responsive nano-platform to improve in vivo LDs-specific imaging. MeOND exhibits a remarkable twisted intramolecular charge transfer (TICT) effect with a strongly enhanced near-infrared emission in low-polarity lipid environment. Also, MeOND demonstrates satisfactory biocompatibility and superior intracellular LDs imaging capabilities. MeOND encapsulated nano-platform (MeOND@PMM) presented favorable water solubility and biocompatibility. MeOND@PMM remains stable in physiological conditions but quickly degrades in the environment of elevated ROS level. The released MeOND could then light up the intracellular LDs in atherosclerotic plaques. The design of the probe and nano-platform is expected to provide a better tool for the scientific research of LDs and LDs-related diseases.

Uncovering atherosclerotic cardiovascular disease by PET imaging.

Assessing atherosclerosis severity is essential for precise patient stratification. Specifically, there is a need to identify patients with residual inflammation because these patients remain at high risk of cardiovascular events despite optimal management of cardiovascular risk factors. Molecular imaging techniques, such as PET, can have an essential role in this context. PET imaging can indicate tissue-based disease status, detect early molecular changes and provide whole-body information. Advances in molecular biology and bioinformatics continue to help to decipher the complex pathogenesis of atherosclerosis and inform the development of imaging tracers. Concomitant advances in tracer synthesis methods and PET imaging technology provide future possibilities for atherosclerosis imaging. In this Review, we summarize the latest developments in PET imaging techniques and technologies for assessment of atherosclerotic cardiovascular disease and discuss the relationship between imaging readouts and transcriptomics-based plaque phenotyping.

Tailoring Zinc Ferrite Nanoparticle Surface Coating for Macrophage-Affinity Magnetic Resonance Imaging of Atherosclerosis.

Atherosclerosis is a chronic inflammatory disease characterized by the formation of atherosclerotic plaques, while macrophages as key players in plaque progression and destabilization are promising targets for atherosclerotic plaque imaging. Contrast-enhanced magnetic resonance imaging (CE-MRI) has emerged as a powerful noninvasive imaging technique for the evaluation of atherosclerotic plaques within arterial walls. However, the visualization of macrophages within atherosclerotic plaques presents considerable challenges due to the intricate pathophysiology of the disease and the dynamic behavior of these cells. Biocompatible ferrite nanoparticles with diverse surface ligands possess the potential to exhibit distinct relaxivity and cellular affinity, enabling improved imaging capabilities for macrophages in atherosclerosis. In this work, we report macrophage-affinity nanoparticles for magnetic resonance imaging (MRI) of atherosclerosis via tailoring nanoparticle surface coating. The ultrasmall zinc ferrite nanoparticles (Zn0.4Fe2.6O4) as T1 contrast agents were synthesized and modified with dopamine, 3,4-dihydroxyhydrocinnamic acid, and phosphorylated polyethylene glycol to adjust their surface charges to be positively, negatively, and neutrally charged, respectively. In vitro MRI evaluation shows that the T1 relaxivity for different surface charged Zn0.4Fe2.6O4 nanoparticles was three higher than that of the clinically used Gd-DTPA. Furthermore, in vivo atherosclerotic plaque MR imaging indicates that positively charged Zn0.4Fe2.6O4 showed superior MRI efficacy on carotid atherosclerosis than the other two, which is ascribed to high affinity to macrophages of positively charged nanoparticles. This work provides improved diagnostic capability and a better understanding of the molecular imaging of atherosclerosis.

Development of a CCR2 targeted 18F-labeled radiotracer for atherosclerosis imaging with PET

Atherosclerosis is a chronic inflammatory disease and the leading cause of morbidity and mortality worldwide. CC motif chemokine ligand 2 and its corresponding cognate receptor 2 (CCL2/CCR2) signaling has been implicated in regulating monocyte recruitment and macrophage polarization during inflammatory responses that plays a pivotal role in atherosclerosis initiation and progression. In this study, we report the design and synthesis of a novel 18F radiolabeled small molecule radiotracer for CCR2-targeted positron emission tomography (PET) imaging in atherosclerosis. The binding affinity of this radiotracer to CCR2 was evaluated via in vitro binding assay using CCR2+ membrane and cells. Ex vivo biodistribution was carried out in wild type mice to assess radiotracer pharmacokinetics. CCR2 targeted PET imaging of plaques was performed in two murine atherosclerotic models. The sensitive detection of atherosclerotic lesions highlighted the potential of this radiotracer for CCR2 targeted PET and warranted further optimization.

Prognostic impact of carotid plaque imaging using total plaque area added to SCORE2 in middle-aged subjects: the ARteris Cardiovascular Outcome (ARCO) cohort study.

Many cardiovascular events occur in seemingly healthy individuals.We set out to assess the predictive value of atherosclerosis imaging in combination with cardiovascular risk calculators in subjects aged 40-65 years. We compared PROCAM (PROspective CArdiovascular Münster study), SCORE (Systematic COronary Risk Evaluation) and SCORE2 with carotid ultrasound (total plaque area, TPA) in subjects without cardiovascular disease. In this prospective cohort study, follow-up was obtained by phone or mail from patients; or from clinical records, if needed. In 2842 subjects (mean age 50±8 years; 38% women), cardiovascular events occurred in 154 (5.4%) of them over an mean follow-up period of 5.9 (range 1-12) years, specifically: 41 cases of AMI (myocardial infarction), 16 strokes, 21 CABG (coronary artery bypass grafting), 41 PTCA (percutaneous transluminal coronary angioplasty) and 35 CAD (coronary artery disease). Mean PROCAM risk was 5±6%, mean SCORE risk was 1.3±1.6% and mean SCORE2 risk was 5±3%. Both for the primary outcome (major adverse cardiovascular events, MACEs, i.e. AMI + strokes) and the secondary outcome (atherosclerotic cardiovascular disease, ASCVD, i.e. MACEs + CABG + CAD + PTCA), hazards increased significantly for TPA tertiles and SCORE2 post-test risk between 6.7 to 12.8 after adjustment for risk factors (age, smoke, sex, systolic blood pressure, lipids, medication) and after adjustment for results from PROCAM, SCORE and SCORE2. Model performance was statistically improved regarding model fit in all models using TPA. Net reclassification improvement for SCORE2 with TPA post-test risk increased significantly by 24% for MACEs (p = 0.01) and 39% for ASCVD (p <0.0001). Integration of TPA post-test risk into SCORE2 adds prognostic information, supporting the use of carotid ultrasound when assessing ASCVD risk in subjects aged 40-65 years.

Assessment of coronary flow capacity by positron emission tomography in coronary artery disease

Assessment of coronary flow capacity by positron emission tomography in coronary artery disease

Assessment of coronary flow capacity by positron emission tomography in coronary artery disease

Assessment of coronary flow capacity by positron emission tomography in coronary artery disease

Single-cell profiling of CD11c+ B cells in atherosclerosis.

Circulating CD11c+ B cells, a novel subset of activated B cells, have been linked to autoimmunity and shown to expand with age. Atherosclerosis is an age-associated disease that involves innate and adaptive immune responses to modified self-antigens. Yet, the expression of CD11c on specific B-cell subtypes and its link to atherosclerosis are poorly understood. In this study, we characterized the frequency of CD11c+ B cells in tissues in mice with aging. We observed an age-associated increase in CD11c+ B cells in the spleen and bone marrow of ApoE-/- mice, and this was associated with an increase in aortic plaque. In addition, we also utilized single-cell multi-omics profiling of 60 human subjects undergoing advanced imaging for coronary artery disease (CAD) to subtype CD11c+ B cells and determine their frequency in subjects with high and low severity of CAD. Using unsupervised clustering, we identified four distinct clusters of CD11c+ B cells, which include CD27 and IgD double negative 2 (DN2), age-associated (ABC), CD11c+ unswitched memory (USWM), and activated Naïve (aNav) B cells. We observed an increase in the frequency of both ABC B cells and DN2 B cells in patients with high CAD severity. Pathway analysis further demonstrated augmentation of autophagy, IFNg signaling, and TLR signaling in DN2 cells in high-severity CAD patients. On the other hand, an increase in the negative regulator of BCR signaling through CD72 was found in ABC cells in low-severity CAD patients. Through investigating scRNAseq of atheroma, these DN2 cells were also found to infiltrate human coronary atheroma.