Targeted single-marker extracellular vesicle profiles in acute ischemic stroke and small vessel disease

Introduction: Extracellular vesicles (EVs) are increasingly recognized as circulating biomarkers reflecting endothelial dysfunction, thrombo-inflammatory activity, and microvascular injury. Their association with cerebral small vessel disease (SVD) in acute ischemic stroke (AIS) remains insufficiently characterized. We investigated the relationship between circulating EV subpopulations, vascular risk factors, and magnetic resonance imaging (MRI)-defined SVD in AIS. Materials and methods: Medium-sized EVs were quantified and phenotyped by flow cytometry in platelet-free plasma from AIS patients and healthy controls. EVs were classified according to cellular origin, and binary logistic regression was used to assess their associations with AIS and SVD, adjusting for demographic and vascular risk factors. Results: AIS patients demonstrated significantly elevated concentrations of circulating CD146+ EVs (endothelial-origin), CD41+ EVs (platelet-origin), CD62P+ EVs (activated platelet-origin), CD235a+ EVs (erythrocyte-origin), and CD66b+ EVs (granulocyte-origin) compared with controls. CD62P+ and CD235a+ EVs were most strongly associated with AIS with adjusted odds ratios (AORs) of 1.10 and 1.09, respectively. Notably, CD235a+ EVs (erythrocyte-derived) showed an inverse association with SVD burden (AOR = 0.77), although this association was attenuated in penalized sensitivity analysis. SVD was frequently accompanied by hypertension, white matter hyperintensities, silent infarcts, basis pontis ischemia, and cerebral microbleeds, with hypertension on admission emerging as the strongest independent predictor of SVD (AOR = 9.33). Conclusions: Circulating EVs, particularly those derived from activated platelets, erythrocytes, and granulocytes, demonstrated distinct associations with AIS and SVD phenotypes, suggesting differing underlying thrombo-inflammatory and microvascular mechanisms. Although the observed odds ratios do not currently support their use as standalone diagnostic biomarkers for AIS or SVD burden, these findings provide a foundation for future longitudinal and mechanistic studies investigating EV-based biomarkers and their temporal dynamics in cerebrovascular disease.