Single-cell RNA sequencing of carotid plaques from 46 endarterectomy patients reveals four major macrophage archetypes: inflammatory macrophages, lipid-associated macrophages (LAMs), tissue-resident-like LAMs, and inflammatory LAMs.
Deconvolution of bulk transcriptome data from 656 patients shows that overall macrophage burden and, specifically, LAM and inflammatory LAM foam cell markers PLIN2 and TREM1 are associated with increased risk of secondary major adverse cardiovascular events (MACE) within 3 years after surgery.
These associations were validated in an independent cohort of 82 patients, supporting the clinical relevance of plaque macrophage subtyping in predicting secondary cardiovascular risk.
The study provides a detailed cellular atlas linking monocyte differentiation pathways to plaque macrophage heterogeneity and clinical outcomes.
Study Snapshot
Design: Single-cell RNA sequencing on blood and plaques from 46 carotid endarterectomy patients (AtheroExpress cohort); deconvolution on bulk transcriptome data from 656 patients; validation in 82 patients (Carotid Plaque Imaging Project).
Population: Patients undergoing carotid endarterectomy for symptomatic or asymptomatic carotid stenosis.
Primary outcome: Secondary major adverse cardiovascular events (MACE) during 3-year follow-up after endarterectomy.
Key markers: PLIN2 and TREM1 (LAM and inflammatory LAM foam-cell markers) significantly associated with increased MACE risk.
Funding/Registration: Not specified in abstract; part of the AtheroExpress biobank study.
Why This Study Matters
Atherosclerosis remains a leading cause of cardiovascular morbidity and mortality. Although macrophages are central to plaque progression, the relationship between specific macrophage subpopulations within carotid plaques and future cardiovascular events has not been clearly established. This study leverages single-cell resolution and large-scale transcriptomic deconvolution to identify precise cell states that may improve risk stratification after carotid endarterectomy.
How the Study Was Conducted
Researchers used single-cell RNA sequencing to profile immune cells from blood and atherosclerotic plaques obtained from 46 patients enrolled in the AtheroExpress cohort. They identified macrophage subtypes and reconstructed cellular differentiation trajectories. Using the AtheroExpress bulk RNA-seq dataset (n=656), they performed deconvolution to estimate cell-type proportions and assessed associations with symptom status at surgery and with secondary major adverse cardiovascular events (MACE) over 3 years of follow-up. Key associations were then validated in an independent cohort of 82 patients from the Carotid Plaque Imaging Project.
What the Researchers Found
Four major macrophage archetypes were identified: inflammatory macrophages, lipid-associated macrophages (LAMs), tissue-resident-like LAMs, and inflammatory LAMs. Trajectory analysis suggested that these subtypes differentiate from both classical and non-classical monocytes, with inflammatory LAMs arising via intermediate LAM stages. In the bulk deconvolution analysis, macrophages were the only cell population significantly associated with symptomatic status at surgery and with increased risk of secondary MACE during follow-up. Within the macrophage compartment, high expression of LAM and inflammatory LAM markers—particularly PLIN2 and TREM1—correlated with a greater risk of MACE. These findings were confirmed in the validation cohort.
What the Findings May Mean
The results suggest that not all plaque macrophages carry the same prognostic weight; specifically, lipid-associated and inflammatory lipid-associated macrophage markers may help identify patients at elevated risk for subsequent cardiovascular events after endarterectomy. If replicated in prospective studies, these markers could inform post-surgical surveillance intensity or therapeutic targeting of residual inflammatory risk. However, the study does not establish causation, and the clinical utility of these markers in routine practice remains to be determined.
Strengths and Limitations
Strengths include the combination of single-cell transcriptomics with large bulk-cohort deconvolution and external validation. Limitations include the relatively small single-cell discovery cohort (n=46), the inherent bias of endarterectomy patients (predominantly symptomatic), the reliance on association rather than causation, and the need for prospective validation of the predictive markers in broader populations. Residual confounding from unmeasured clinical factors cannot be excluded.
Implications for Practice and Research
For clinicians, these findings highlight that plaque macrophage composition—especially the presence of lipid-associated and inflammatory foam cells—may contribute to residual cardiovascular risk after revascularization. For researchers, the study provides a refined classification of plaque macrophages and suggests that PLIN2 and TREM1 could be developed as biomarkers or therapeutic targets. Future work should focus on standardizing assays for these markers and testing whether targeting LAM pathways reduces secondary events.
Funding, Disclosures, and Registration
The abstract does not detail funding, disclosures, or registration. The study uses the AtheroExpress biobank, a long-standing repository for atherosclerotic tissue research.
References
Prange KHM, Bel-Bordes G, Depuydt MAC, et al. Carotid plaque macrophage burden and inflammatory lipid-associated macrophage markers predict secondary major adverse cardiovascular events after endarterectomy. European Heart Journal. 2026;47(28):3821-3836. PMID: 41758068. DOI: not provided.
