A novel flow cytometric approach to distinguish circulating endothelial cells from endothelial microparticles: Relevance for the evaluation of endothelial dysfunction

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• 作者：Paola Lanuti^a ; ^c ; ^{paolalanuti@yahoo.it} ; Francesca Santilli^a ; ^c ; ^{f.santilli@unich.it} ; Marco Marchisio^a ; ^c ; ^{m.marchisio@unich.it} ; Laura Pierdomenico^a ; ^c ; ^{l.pierdomenico@unich.it} ; Ester Vitacolonna^a ; ^c ; ^{e.vitacolonna@unich.it} ; Eugenio Santavenere^a ; ^{e.santavenere@unich.it} ; Antonio Iacone^d ; ^{citomorfologia@unich.it} ; Giovanni Davì ; ^a ; ^c ; ^{gdavi@unich.it} ; Mario Romano^b ; ^c ; ^{mromano@unich.it} ; Sebastiano Miscia^a ; ^c ; ^{s.miscia@unich.it}
• 关键词：Vascular endothelium ; Endothelial microparticles ; Diabetes ; Flow cytometry ; Absolute count
• 刊名：Journal of Immunological Methods
• 出版年：2012
• 期刊代码：57_00221759
• 类别：imm
• 出版时间：29 June, 2012
• 卷：380
• 期：1-2
• 页码：16-22
• 文件大小：681 K

摘要

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 渭l of peripheral blood, the large majority of events expressing an endothelial phenotype (CD45鈭?CD146+/CD34+) are due to non-nucleated particles (DRAQ5鈭? 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鈭?CD146+/CD34+/CD117+) and from mature endothelial cells (CD45鈭?CD146+/CD34+/CD117鈭?. 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.