36 Endothelial microparticles: investigating their role on endothelial cells in vitro

Endothelial microparticles (EMPs) are complex structures with pleiotropic properties and are emerging as an index of endothelial damage. Previous studies have shown elevated circulating EMP levels in inflammatory disorders, which raises the question about their physiological role. This study aims to investigate the autocrine effects of EMPs on TNFα-mediated EMP release, as well as on endothelial cell activation and migration. Two groups of EMPs were generated in vitro from TNF-stimulated human umbilical vein endothelial cells (HUVECs; sEMPs) or untreated HUVECs (uEMPs) and collected by ultracentrifugation. HUVECs were incubated with either 106 sEMPs or uEMPs/ml in the presence or absence of TNFα (10 ng/ml) for 24 h. EMP levels were enumerated by flow cytometry; RT-PCR and functional scratch assays were also carried out. HUVECs treated with either sEMP or uEMP showed increased ICAM-1 mRNA abundance (p < 0.05) and enhanced cell migration, compared to untreated cells (p < 0.005). Both sEMPs and uEMPs reduced TNFα-mediated migration in a scratch assay (p < 0.005). In addition, uEMP treatment reduced TNFα-induced EMP release (p < 0.05) whereas sEMPs did not. sEMPs and uEMPs have been subjected to proteomic and microRNA screening to identify pathways and interrogate these findings further. We conclude that EMPs may exhibit different properties depending upon the environment in which they are produced and may have dual properties in the endothelial layer, depending on the micro-environment, by accelerating cell migration and endothelial cell activation. Further mechanistic studies are required to understand cell-EMP interaction and to determine how they contribute to pathophysiological mechanisms in inflammation-associated disorders.