Elevated Circulating Endothelial Microparticles in Acute Stroke Patients: A Correlation with Brain Lesion Volume and Outcome.
Authors: Simak Jan , Gelderman Monique P. , Yu Hua , Wright Violet , Alberts-Grill Noah , Stranix John T. , Baird Alison E. 
Published: October 2019

Abstract
Elevated endothelial cell membrane microparticles (EC MP) in blood have been demonstrated in various diseases with a vascular injury component. The aim of this study was to investigate if circulating EC MP show a relationship with outcome after acute stroke and with the ischemic brain lesion volume measured by magnetic resonance diffusion-weighted imaging (DWI). We analyzed EC MP in the blood of 42 acute stroke patients (AS): 20 patients with National Institutes of Health Stroke Scale (NIHSS) scores < 5 were classified as mild stroke (MS) (median NIHSS= 2; 25th–75th%: 0–2), while the other 22 patients with NIHSS ≥5 (NIHSS=12; 6–21) were classified as moderate to severe stroke (SS). Peripheral venous blood samples were collected at a median time of 36 hours (18–52) after the onset of clinical symptoms. The patients outcome was based on the Rankin disability score at the time of hospital discharge. Blood samples of 23 age matched control volunteers (CTRL) were used for comparison. EC MP analysis used a three-color flow cytometry assay (Simak et al, British J Haematol125, 804–813, 2004). EC MP were identified by antibodies to EC antigen CD105 (endoglin) and the highly specific CD144 (VE-cadherin). Platelet, white, and red blood cell MP were identified using cell specific antibodies to CD41, CD45, and CD235a, respectively. Plasma counts of CD105+CD41−CD45- EC MP were elevated in SS (median: 840/μL; 25th–75th%: 565–1079/μL) as compared to CTRL (415/μL; 201–624/μL; p=0.014). Moreover, CD105+CD144+ EC MP were elevated in SS (261/μL; 137–433/μL) when compared to MS (154/μL; 99–182/μL; p=0.031) or CTRL group (140/μL; 79–247/μL; p=0.031). Interestingly, CD105+CD41−CD45- EC MP, but not CD105+CD144+ EC MP, exhibited a significant correlation (p=0.005; r=0.45) with DWI brain lesion volume in AS group. However, CD105+CD144+ EC MP in the admission samples highly correlated (p=0.0007; r=0.54) with the Rankin disability score in the AS group at hospital discharge, while correlation of CD105+CD41−CD45- EC MP with the Rankin score was not as significant (p=0.007; r=0.44). We further analyzed 12 MS and 12 SS follow-up samples collected at a median period of 10 days (7–14) after the first sampling. Surprisingly, in SS follow-up samples, CD105+ EC MP populations decreased, while CD144+CD105−CD41- EC MP significantly increased, as compared to the samples at admission. In conclusion, the SS patient group had elevated different phenotypes of EC MP in the plasma samples at admission when compared to MS or CTRL groups. This is likely a reflection of the severity of ischemic-reperfusion injury of the brain vasculature. Elevated endoglin-positive EC MP were associated with brain ischemic lesion volume, whereas EC MP positive for both endoglin and VE-cadherin in the admission samples showed highly significant correlation with the patients disability outcome. The increased VE-cadherin-positive EC MP in follow-up samples may reflect a continuing endothelial injury in SS patients. Analysis of different phenotypes of EC MP in peripheral blood of stroke patients may be indicative of volume, character and severity of brain vascular injury and could be of diagnostic and prognostic use.