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Updated: Aug 30, 2026
Techniques for the Analysis of Extracellular Vesicles Using Flow Cytometry
Published on: March 17, 2015
Antigenic characterization of endothelial cell-derived microparticles and their detection ex vivo
M N Abid Hussein1, E W Meesters, N Osmanovic
1Department of Clinical Chemistry, Academic Medical Center, University of Amsterdam, The Netherlands. m.n.abidhussein@amc.uva.nl
Background:
Endothelial activation and dysfunction are associated with several diseases. However, hardly any specific markers are available. Microparticles (MP) from endothelial cells (EC; EMP) were reported in patient groups and healthy individuals. The antibodies used to detect EMP, however, were mainly directed against antigens without EC specificity.
Objectives:
We evaluated the antigens on EC and EMP to establish proper markers for EMP detection.
Methods:
EMP were isolated from supernatants of resting and interleukin (IL)-1alpha activated human umbilical vein EC (HUVEC; n=3; 0-72 h), stained with annexin V and monoclonal antibodies, and analyzed by flow cytometry. Human platelet-MP (PMP), the main MP population in plasma, were prepared in vitro. EMP and PMP were studied in plasma from systemic lupus erythematosus (SLE) patients (n=11) and healthy individuals (n=10).
Results:
Platelet-endothelial cell adhesion molecule-1 (PECAM-1), alphanu and beta3 were constitutively exposed on HUVEC, but (almost) absent on EMP (<15% positive for alphanu and beta3), or only exposed on a subpopulation (PECAM-1; 30-60%). Activated HUVEC (>80%) and (subpopulations of) EMP exposed E-selectin and tissue factor. PMP strongly exposed PECAM-1, beta3, and glycoprotein (GP)Ib (CD42b), but not alphanu or E-selectin. GPIb and P-selectin (CD62P) were absent on EMP. Plasma samples contained 0.5% MP staining for E-selectin and/or alphanu. Plasma from one SLE patient contained E-selectin exposing MP (21%), but little alphanu-positive MP.
Conclusions:
EC release EMP in vitro. The antigenic phenotype of EMP released from resting and IL-1alpha-stimulated EC differs among each other as well as from resting and stimulated EC, respectively. E-selectin exposed on IL-1alpha-stimulated EC is a valid marker for EMP detection ex vivo to establish endothelial cell activation.
Insights
Endothelial microparticles (EMP) are linked to diseases, but specific markers are lacking. E-selectin on EMP from activated endothelial cells (EC) serves as a reliable marker for detecting endothelial cell activation.
Area of Science:
- Cardiovascular Biology
- Cell Biology
- Immunology
Background:
- Endothelial activation and dysfunction are implicated in various diseases.
- Specific biomarkers for endothelial cell activation are limited.
- Existing markers for endothelial microparticles (EMP) often lack endothelial cell specificity.
Purpose of the Study:
- To identify and evaluate specific antigens on endothelial cells (EC) and EMP.
- To establish reliable markers for the detection of EMP.
- To differentiate EMP from other microparticle types.
Main Methods:
- Human umbilical vein EC (HUVEC) were cultured and stimulated with IL-1alpha.
- EMP were isolated, stained with annexin V and monoclonal antibodies, and analyzed by flow cytometry.
- EMP and platelet-derived microparticles (PMP) were characterized, and their presence in plasma from SLE patients and healthy controls was assessed.
Main Results:
- Constitutive antigens like PECAM-1, alphanu, and beta3 were found on HUVEC but largely absent or reduced on EMP.
- Activated HUVEC and subpopulations of EMP expressed E-selectin and tissue factor.
- EMP lacked GPIb and P-selectin, distinguishing them from PMP.
Conclusions:
- Endothelial cells release EMP in vitro, with distinct antigenic profiles depending on activation state.
- E-selectin expression on EMP derived from IL-1alpha-stimulated EC is a valid ex vivo marker for endothelial cell activation.
- This finding aids in developing specific diagnostic tools for diseases involving endothelial dysfunction.
