Antigenic characterization of endothelial cell-derived microparticles and their detection ex vivo

Abstract

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)-lα 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 (w=ll) and healthy individuals (n = 10). Results: Platelet-endothelial cell adhesion molecule-1 (PECAM-1), α ν and β 3 were constitutively exposed on HUVEC, but (almost) absent on EMP (<15% positive for α ν and β 3), 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, β 3, and glycoprotein (GP)Ib (CD42b), but not α ν or E-selectin. GPIb and P-selectin (CD62P) were absent on EMP. Plasma samples contained 0.5% MP staining for E-selectin and/or α ν. Plasma from one SLE patient contained E-selectin exposing MP (21%), but little α ν-positive MP. Conclusions: EC release EMP in vitro. The antigenic phenotype of EMP released from resting and IL-lα-stimulated EC differs among each other as well as from resting and stimulated EC, respectively. E-selection on IL-lα-stimulated EC is a valid marker for EMP detection ex vivo to establish endothelial cell activation. © 2003 International Society on Thrombosis and Haemostasis.