EV Flow Series | Aleksandra Gasecka - Concentration of EVs in patients with myocardial infarction.

This video is part of the ISEV-ISAC-ISTH EV Flow Cytometry Working Group Seminar Series. Visit our website at http://www.evflowcytometry.org/ for more information. Subscribe to our seminar series and get access to our Slack channel to continue the discussion by completing this form: https://bit.ly/3eKmlNP Date: 3rd June 2020 Topic: Research Talk Title: Calibrated flow cytometry to measure the concentration of extracellular vesicles in patients with myocardial infarction. Moderators: André Görgens [1], Paul Harrison [2] Karolinska Institute, Sweden [1], University of Birmingham, UK Speaker: Aleksandra Gąsecka, Medical University of Warsaw, Poland & Amsterdam University Medical Centers, Netherlands Abstract: Acute myocardial infarction (AMI) is a major cause of human death and disability, but early biomarkers for AMI are lacking. AMI is due to atherothrombosis, i.e. formation of platelet aggregates (thrombi) on ruptured atherosclerotic plaques. Because platelets release extracellular vesicles (EVs) during thrombus formation, we hypothesized that EVs are a biomarker of atherothrombosis and an early biomarker of AMI. Venous blood was collected 24 hours (acute phase), 72 hours (hospital discharge) and 6 months (late phase) after AMI from fasting patients (n=60, age 64.5±10.8 years, 68% male), and once from fasting healthy volunteers (n=30, mean age 57.7±6.6 years, 62% male). Flow cytometry (Apogee A60-Micro) was used to determine concentrations of plasma EVs labelled with markers for endothelial cells (CD146), leukocytes (CD45), phosphatidylserine (lactadherin), platelets (CD31, CD61, CD62p), and fibrinogen. Analysis of the 1,224 flow cytometry data files was fully automated with in-house developed software (MATLAB R2018a), enabling automatic flow rate stabilization, application of Rosetta Calibration (Exometry) and Flow-SR for diameter and refractive index determination, size distribution fitting, MESF calibration, fluorescent gate determination, and statistics reporting. To differentiate between EVs and small platelets, only particles < 1,000 nm were included. Populations for which an unpaired t-test or one-way ANOVA with Bonferroni correction resulted in p < 0.05 were considered significant. EV concentrations from leukocytes and endothelial cells were lower in patients in the acute AMI phase, compared to healthy volunteers (p < 0.05 for both), and increased to the level observed in healthy volunteers in the late phase (p < 0.05 for both). Concentrations of PS-exposing EVs and platelet EVs (CD31, CD61) were decreased in the acute AMI phase, compared to the late phase (p < 0.05 for all). We identified decreased concentrations of EVs from leukocytes and endothelial cells as new candidate biomarkers to differentiate patients with atherothrombosis from healthy volunteers. Further, we identified decreased concentrations of PS-exposing EVs and EVs from platelets as new candidate biomarkers to differentiate patients in the acute and late AMI phase. Because the flow rate, fluorescence detectors and scatter detectors of our flow cytometer were calibrated, our data can be compared to future clinical studies, which essential to confirm the clinical utility of EVs as biomarkers of atherothrombosis.