A standardized flow cytometry network study for the assessment of circulating endothelial cell physiological ranges

Paola Lanuti, Pasquale Simeone, Gianluca Rotta, Camillo Almici, Giuseppe Avvisati, Rosa Azzaro, Giuseppina Bologna, Alfredo Budillon, Melania Di Cerbo, Elena Di Gennaro, Maria Luisa Di Martino, Annamaria Diodato, Paolo Doretto, Eva Ercolino, Alessandra Falda, Chiara Gregorj, Alessandra Leone, Francesca Losa, Natalia Malara, Mirella MariniPasquale Mastroroberto, Vincenzo Mollace, Michele Morelli, Emma Muggianu, Giuseppe Musolino, Arabella Neva, Laura Pierdomenico, Silvia Pinna, Giovanna Piovani, Maria Serena Roca, Domenico Russo, Lorenza Scotti, Maria Cristina Tirindelli, Roberta Venturella, Carlo Vitagliano, Fulvio Zullo, Marco Marchisio, Sebastiano Miscia

Research output: Contribution to journalArticle

Abstract

Circulating endothelial cells (CEC) represent a restricted peripheral blood (PB) cell subpopulation with high potential diagnostic value in many endothelium-involving diseases. However, whereas the interest in CEC studies has grown, the standardization level of their detection has not. Here, we undertook the task to align CEC phenotypes and counts, by standardizing a novel flow cytometry approach, within a network of six laboratories. CEC were identified as alive/nucleated/CD45negative/CD34bright/CD146positive events and enumerated in 269 healthy PB samples. Standardization was demonstrated by the achievement of low inter-laboratory Coefficients of Variation (CVL), calculated on the basis of Median Fluorescence Intensity values of the most stable antigens that allowed CEC identification and count (CVL of CD34bright on CEC ∼ 30%; CVL of CD45 on Lymphocytes ∼ 20%). By aggregating data acquired from all sites, CEC numbers in the healthy population were captured (medianfemale = 9.31 CEC/mL; medianmale = 11.55 CEC/mL). CEC count biological variability and method specificity were finally assessed. Results, obtained on a large population of donors, demonstrate that the established procedure might be adopted as standardized method for CEC analysis in clinical and in research settings, providing a CEC physiological baseline range, useful as starting point for their clinical monitoring in endothelial dysfunctions.

Original languageEnglish
Article number5823
JournalScientific Reports
Volume8
Issue number1
DOIs
Publication statusPublished - Dec 1 2018

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Flow Cytometry
Endothelial Cells
Cell Count
Population
Endothelium
Blood Cells
Fluorescence
Lymphocytes
Phenotype
Antigens

ASJC Scopus subject areas

  • General

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A standardized flow cytometry network study for the assessment of circulating endothelial cell physiological ranges. / Lanuti, Paola; Simeone, Pasquale; Rotta, Gianluca; Almici, Camillo; Avvisati, Giuseppe; Azzaro, Rosa; Bologna, Giuseppina; Budillon, Alfredo; Di Cerbo, Melania; Di Gennaro, Elena; Di Martino, Maria Luisa; Diodato, Annamaria; Doretto, Paolo; Ercolino, Eva; Falda, Alessandra; Gregorj, Chiara; Leone, Alessandra; Losa, Francesca; Malara, Natalia; Marini, Mirella; Mastroroberto, Pasquale; Mollace, Vincenzo; Morelli, Michele; Muggianu, Emma; Musolino, Giuseppe; Neva, Arabella; Pierdomenico, Laura; Pinna, Silvia; Piovani, Giovanna; Roca, Maria Serena; Russo, Domenico; Scotti, Lorenza; Tirindelli, Maria Cristina; Venturella, Roberta; Vitagliano, Carlo; Zullo, Fulvio; Marchisio, Marco; Miscia, Sebastiano.

In: Scientific Reports, Vol. 8, No. 1, 5823, 01.12.2018.

Research output: Contribution to journalArticle

Lanuti, P, Simeone, P, Rotta, G, Almici, C, Avvisati, G, Azzaro, R, Bologna, G, Budillon, A, Di Cerbo, M, Di Gennaro, E, Di Martino, ML, Diodato, A, Doretto, P, Ercolino, E, Falda, A, Gregorj, C, Leone, A, Losa, F, Malara, N, Marini, M, Mastroroberto, P, Mollace, V, Morelli, M, Muggianu, E, Musolino, G, Neva, A, Pierdomenico, L, Pinna, S, Piovani, G, Roca, MS, Russo, D, Scotti, L, Tirindelli, MC, Venturella, R, Vitagliano, C, Zullo, F, Marchisio, M & Miscia, S 2018, 'A standardized flow cytometry network study for the assessment of circulating endothelial cell physiological ranges', Scientific Reports, vol. 8, no. 1, 5823. https://doi.org/10.1038/s41598-018-24234-0
Lanuti, Paola ; Simeone, Pasquale ; Rotta, Gianluca ; Almici, Camillo ; Avvisati, Giuseppe ; Azzaro, Rosa ; Bologna, Giuseppina ; Budillon, Alfredo ; Di Cerbo, Melania ; Di Gennaro, Elena ; Di Martino, Maria Luisa ; Diodato, Annamaria ; Doretto, Paolo ; Ercolino, Eva ; Falda, Alessandra ; Gregorj, Chiara ; Leone, Alessandra ; Losa, Francesca ; Malara, Natalia ; Marini, Mirella ; Mastroroberto, Pasquale ; Mollace, Vincenzo ; Morelli, Michele ; Muggianu, Emma ; Musolino, Giuseppe ; Neva, Arabella ; Pierdomenico, Laura ; Pinna, Silvia ; Piovani, Giovanna ; Roca, Maria Serena ; Russo, Domenico ; Scotti, Lorenza ; Tirindelli, Maria Cristina ; Venturella, Roberta ; Vitagliano, Carlo ; Zullo, Fulvio ; Marchisio, Marco ; Miscia, Sebastiano. / A standardized flow cytometry network study for the assessment of circulating endothelial cell physiological ranges. In: Scientific Reports. 2018 ; Vol. 8, No. 1.
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abstract = "Circulating endothelial cells (CEC) represent a restricted peripheral blood (PB) cell subpopulation with high potential diagnostic value in many endothelium-involving diseases. However, whereas the interest in CEC studies has grown, the standardization level of their detection has not. Here, we undertook the task to align CEC phenotypes and counts, by standardizing a novel flow cytometry approach, within a network of six laboratories. CEC were identified as alive/nucleated/CD45negative/CD34bright/CD146positive events and enumerated in 269 healthy PB samples. Standardization was demonstrated by the achievement of low inter-laboratory Coefficients of Variation (CVL), calculated on the basis of Median Fluorescence Intensity values of the most stable antigens that allowed CEC identification and count (CVL of CD34bright on CEC ∼ 30{\%}; CVL of CD45 on Lymphocytes ∼ 20{\%}). By aggregating data acquired from all sites, CEC numbers in the healthy population were captured (medianfemale = 9.31 CEC/mL; medianmale = 11.55 CEC/mL). CEC count biological variability and method specificity were finally assessed. Results, obtained on a large population of donors, demonstrate that the established procedure might be adopted as standardized method for CEC analysis in clinical and in research settings, providing a CEC physiological baseline range, useful as starting point for their clinical monitoring in endothelial dysfunctions.",
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AU - Lanuti, Paola

AU - Simeone, Pasquale

AU - Rotta, Gianluca

AU - Almici, Camillo

AU - Avvisati, Giuseppe

AU - Azzaro, Rosa

AU - Bologna, Giuseppina

AU - Budillon, Alfredo

AU - Di Cerbo, Melania

AU - Di Gennaro, Elena

AU - Di Martino, Maria Luisa

AU - Diodato, Annamaria

AU - Doretto, Paolo

AU - Ercolino, Eva

AU - Falda, Alessandra

AU - Gregorj, Chiara

AU - Leone, Alessandra

AU - Losa, Francesca

AU - Malara, Natalia

AU - Marini, Mirella

AU - Mastroroberto, Pasquale

AU - Mollace, Vincenzo

AU - Morelli, Michele

AU - Muggianu, Emma

AU - Musolino, Giuseppe

AU - Neva, Arabella

AU - Pierdomenico, Laura

AU - Pinna, Silvia

AU - Piovani, Giovanna

AU - Roca, Maria Serena

AU - Russo, Domenico

AU - Scotti, Lorenza

AU - Tirindelli, Maria Cristina

AU - Venturella, Roberta

AU - Vitagliano, Carlo

AU - Zullo, Fulvio

AU - Marchisio, Marco

AU - Miscia, Sebastiano

PY - 2018/12/1

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