Abstract TP075: Circulating extracellular vesicles from the plasma of patients with silent brain infarction induce a senescence-associated secretory phenotype in human brain endothelial cells

Silent brain infarcts (SBIs), or silent strokes, are small areas of cortical or subcortical gliosis found incidentally on brain imaging without clinical deficits on examination. Although individuals with silent brain infarcts (SBI) are at increased risk for stroke and cognitive decline, the underlying mechanisms are unclear, and there are no robust SBI biomarkers. Recent studies have established a role for extracellular vesicles (EVs) in the pathophysiology of acute ischemic stroke (AIS) and identified specific EV cargo as stroke biomarkers. Emerging evidence also suggests a link between cellular senescence and stroke that is regulated through EVs. To better understand a mechanistic role for circulating EVs in SBI and AIS, the objective of this study was to compare circulating EV profiles and their effects on cellular senescence. We hypothesized that circulating plasma EVs from AIS patients (AIS-EVs) would induce a more inflammatory cellular senescence phenotype compared to EVs from SBI patients (SBI-EVs). In this IRB-approved study, SBI and AIS patients were recruited from the neurology inpatient service at WVU Medicine Ruby Memorial Hospital based on eligibility criteria. EVs were purified from plasma followed by Nanoparticle Tracking Analysis (NTA) and measurement of zeta potential (ZP). Preliminary analysis of 15 subjects [SBI (n=7) and AIS (n=8)] revealed no significant differences between the groups in terms of age, sex, or common stroke risk factors. NTA found that SBI subjects had a higher EV concentration and EV size distribution compared to AIS subjects, while no differences in ZP were observed. Next, we tested the functional ability of SBI-EVs and AIS-EVs to induce an inflammatory senescence phenotype in human brain endothelial cells (hBECs). Surprisingly, treatment of hBECs with SBI-EVs decreased viability, increased cytotoxicity, and reduced wound healing compared to AIS-EVs (all p<0.001). Next, we found that SBI-EVs significantly increased expression of senescence genes (CDKN1A/p21, IL-1α, TNFRSF1A, PRODH, and DAO) in hBECs compared to AIS-EVs (all p<0.01). Lastly, we found that shortened telomere length, a marker of senescence, was also increased in hBECs treated with SBI-EVs versus AIS-EVs (p<0.0001). In contrast to our hypothesis, these results demonstrate a novel mechanism wherein plasma EVs from SBI patients generate a more severe inflammatory cellular senescence phenotype compared to their AIS counterparts.