title: A novel flow cytometric approach to distinguish circulating endothelial cells from endothelial microparticles: relevance for the evaluation of endothelial dysfunction.
abstract: Circulating endothelial cells (CEC) and endothelial microparticles (EMP) are emerging as markers of endothelial repair and activation/apoptosis. Although significant changes in the number of CEC and EMP in pathological conditions have been reported, their reliable identification and quantification still remain a technical challenge. Here, we present a novel methodology for the identification and quantitation of CEC and EMP based on multicolor flow cytometry. Using a lyse/no wash protocol, we observed that in 50 μl of peripheral blood, the large majority of events expressing an endothelial phenotype (CD45-/CD146+/CD34+) are due to non-nucleated particles (DRAQ5-) carrying mitochondrial activity (MitoTracker+) and, therefore, classified as EMP. We enumerated circulating EMP by single platform absolute count in a lyse/no wash four-color flow-cytometric procedure, which allowed the distinction, within the whole endothelial compartment, of EMP derived from endothelial progenitors (CD45-/CD146+/CD34+/CD117+) and from mature endothelial cells (CD45-/CD146+/CD34+/CD117-). A significant increase in both subsets was observed in patients with diabetes mellitus. Thus, this simple and highly reproducible method may be useful for monitoring endothelial dysfunction in clinical settings.
authors.affiliation: Department of Medicine and Aging Science (D.M.S.I.), University G. d'Annunzio of Chieti-Pescara, Via dei Vestini 31, 66013, Chieti Scalo (CH), Italy. paolalanuti@yahoo.it
authors.forename: Paola
authors.initials: P
authors.lastname: Lanuti
authors.forename: Francesca
authors.initials: F
authors.lastname: Santilli
authors.forename: Marco
authors.initials: M
authors.lastname: Marchisio
authors.forename: Laura
authors.initials: L
authors.lastname: Pierdomenico
authors.forename: Ester
authors.initials: E
authors.lastname: Vitacolonna
authors.forename: Eugenio
authors.initials: E
authors.lastname: Santavenere
authors.forename: Antonio
authors.initials: A
authors.lastname: Iacone
authors.forename: Giovanni
authors.initials: G
authors.lastname: Davì
authors.forename: Mario
authors.initials: M
authors.lastname: Romano
authors.forename: Sebastiano
authors.initials: S
authors.lastname: Miscia
mesh_terms: D000368:Aged; D018952:Antigens, CD34 / Q000032:analysis; D051929:CD146 Antigen / Q000032:analysis; D055252:Cell-Derived Microparticles / Q000737:chemistry* / Q000276:immunology; D003924:Diabetes Mellitus, Type 2 / Q000097:blood / Q000276:immunology / Q000503:physiopathology; D003925:Diabetic Angiopathies / Q000097:blood / Q000276:immunology / Q000503:physiopathology; D042783:Endothelial Cells / Q000166:cytology* / Q000276:immunology; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D017493:Leukocyte Common Antigens / Q000032:analysis; D008297:Male; D008875:Middle Aged
chemical_list: D018952:Antigens, CD34; D051929:CD146 Antigen; D017493:Leukocyte Common Antigens