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Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment

Large tumor-derived Extracellular Vesicles (tdEVs) detected in blood of metastatic prostate, breast, colorectal, and non-small cell lung cancer patients after enrichment for Epithelial Cell Adhesion Molecule (EpCAM) expression and labeling with 4′,6-diamidino-2-phenylindole (DAPI), phycoerythrin-con...

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Autores principales: Nanou, Afroditi, Zeune, Leonie L., Terstappen, Leon W.M.M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6721753/
https://www.ncbi.nlm.nih.gov/pubmed/31434250
http://dx.doi.org/10.3390/cells8080937
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author Nanou, Afroditi
Zeune, Leonie L.
Terstappen, Leon W.M.M.
author_facet Nanou, Afroditi
Zeune, Leonie L.
Terstappen, Leon W.M.M.
author_sort Nanou, Afroditi
collection PubMed
description Large tumor-derived Extracellular Vesicles (tdEVs) detected in blood of metastatic prostate, breast, colorectal, and non-small cell lung cancer patients after enrichment for Epithelial Cell Adhesion Molecule (EpCAM) expression and labeling with 4′,6-diamidino-2-phenylindole (DAPI), phycoerythrin-conjugated antibodies against Cytokeratins (CK-PE), and allophycocyanin-conjugated antibody against the cluster of differentiation 45 (CD45-APC), are negatively associated with the overall survival of patients. Here, we investigated whether, similarly to tdEVs, leukocyte-derived EVs (ldEVs) could also be detected in EpCAM-enriched blood. Presence of ldEVs and leukocytes in image data sets of EpCAM-enriched samples of 25 healthy individuals and 75 metastatic cancer patients was evaluated using the ACCEPT software. Large ldEVs could indeed be detected, but in contrast to the 20-fold higher frequency of tdEVs as compared to Circulating Tumor Cells (CTCs), ldEVs were present in a 5-fold lower frequency as compared to leukocytes. To evaluate whether these ldEVs pre-exist in the blood or are formed during the CellSearch procedure, the blood of healthy individuals without EpCAM enrichment was labelled with the nuclear dye Hoechst and fluorescently tagged monoclonal antibodies recognizing the leukocyte-specific CD45, platelet-specific CD61, and red blood cell-specific CD235a. Fluorescence microscopy imaging using a similar setup as the CellSearch was performed and demonstrated the presence of a similar population of ldEVs present at a 3-fold lower frequency as compared to leukocytes.
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spelling pubmed-67217532019-09-10 Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment Nanou, Afroditi Zeune, Leonie L. Terstappen, Leon W.M.M. Cells Article Large tumor-derived Extracellular Vesicles (tdEVs) detected in blood of metastatic prostate, breast, colorectal, and non-small cell lung cancer patients after enrichment for Epithelial Cell Adhesion Molecule (EpCAM) expression and labeling with 4′,6-diamidino-2-phenylindole (DAPI), phycoerythrin-conjugated antibodies against Cytokeratins (CK-PE), and allophycocyanin-conjugated antibody against the cluster of differentiation 45 (CD45-APC), are negatively associated with the overall survival of patients. Here, we investigated whether, similarly to tdEVs, leukocyte-derived EVs (ldEVs) could also be detected in EpCAM-enriched blood. Presence of ldEVs and leukocytes in image data sets of EpCAM-enriched samples of 25 healthy individuals and 75 metastatic cancer patients was evaluated using the ACCEPT software. Large ldEVs could indeed be detected, but in contrast to the 20-fold higher frequency of tdEVs as compared to Circulating Tumor Cells (CTCs), ldEVs were present in a 5-fold lower frequency as compared to leukocytes. To evaluate whether these ldEVs pre-exist in the blood or are formed during the CellSearch procedure, the blood of healthy individuals without EpCAM enrichment was labelled with the nuclear dye Hoechst and fluorescently tagged monoclonal antibodies recognizing the leukocyte-specific CD45, platelet-specific CD61, and red blood cell-specific CD235a. Fluorescence microscopy imaging using a similar setup as the CellSearch was performed and demonstrated the presence of a similar population of ldEVs present at a 3-fold lower frequency as compared to leukocytes. MDPI 2019-08-20 /pmc/articles/PMC6721753/ /pubmed/31434250 http://dx.doi.org/10.3390/cells8080937 Text en © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Nanou, Afroditi
Zeune, Leonie L.
Terstappen, Leon W.M.M.
Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment
title Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment
title_full Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment
title_fullStr Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment
title_full_unstemmed Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment
title_short Leukocyte-Derived Extracellular Vesicles in Blood with and without EpCAM Enrichment
title_sort leukocyte-derived extracellular vesicles in blood with and without epcam enrichment
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6721753/
https://www.ncbi.nlm.nih.gov/pubmed/31434250
http://dx.doi.org/10.3390/cells8080937
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