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Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics

Pulmonary artery, capillary, and vein endothelial cells possess distinctive structures and functions, which represent a form of vascular segment specific macroheterogeneity. However, within each of these segmental populations, individual cell functional variability represents a poorly characterized...

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Autores principales: Lee, Ji Young, McMurtry, Sarah A., Stevens, Troy
Formato: Online Artículo Texto
Lenguaje:English
Publicado: SAGE Publications 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5703126/
https://www.ncbi.nlm.nih.gov/pubmed/28841087
http://dx.doi.org/10.1177/2045893217731295
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author Lee, Ji Young
McMurtry, Sarah A.
Stevens, Troy
author_facet Lee, Ji Young
McMurtry, Sarah A.
Stevens, Troy
author_sort Lee, Ji Young
collection PubMed
description Pulmonary artery, capillary, and vein endothelial cells possess distinctive structures and functions, which represent a form of vascular segment specific macroheterogeneity. However, within each of these segmental populations, individual cell functional variability represents a poorly characterized microheterogeneity. Here, we hypothesized that single cell clonogenic assays would reveal microheterogeneity among the parent cell population and enable isolation of highly representative cells with committed parental characteristics. To test this hypothesis, pulmonary microvascular endothelial cells (PMVECs) and pulmonary arterial endothelial cells (PAECs) were isolated from different Sprague Dawley rats. Serum stimulated proliferation of endothelial populations and single cell clonogenic potential were evaluated. In vitro Matrigel assays were utilized to analyze angiogenic potential and the Seahorse assay was used to evaluate bioenergetic profiles. PMVEC populations grew faster and had a higher proliferative potential than PAEC populations. Fewer PMVECs were needed to form networks on Matrigel when compared with PAECs. PMVECs primarily utilized aerobic glycolysis, while PAECs relied more heavily on oxidative phosphorylation, to support bioenergetic demands. Repeated single cell cloning and expansion of PAEC colonies generated homogeneous first-generation clones that were highly reflective of the parental population in terms of growth, angiogenic potential, and bioenergetic profiles. Repeated single cell cloning of the first-generation clones generated second-generation clones with increased proliferative potential while maintaining other parental characteristics. Second-generation clones were highly homogeneous populations. Thus, single cell cloning reveals microheterogeneity among the parent cell population and enables isolation of highly representative cells with parental characteristics.
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spelling pubmed-57031262017-12-04 Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics Lee, Ji Young McMurtry, Sarah A. Stevens, Troy Pulm Circ Research Articles Pulmonary artery, capillary, and vein endothelial cells possess distinctive structures and functions, which represent a form of vascular segment specific macroheterogeneity. However, within each of these segmental populations, individual cell functional variability represents a poorly characterized microheterogeneity. Here, we hypothesized that single cell clonogenic assays would reveal microheterogeneity among the parent cell population and enable isolation of highly representative cells with committed parental characteristics. To test this hypothesis, pulmonary microvascular endothelial cells (PMVECs) and pulmonary arterial endothelial cells (PAECs) were isolated from different Sprague Dawley rats. Serum stimulated proliferation of endothelial populations and single cell clonogenic potential were evaluated. In vitro Matrigel assays were utilized to analyze angiogenic potential and the Seahorse assay was used to evaluate bioenergetic profiles. PMVEC populations grew faster and had a higher proliferative potential than PAEC populations. Fewer PMVECs were needed to form networks on Matrigel when compared with PAECs. PMVECs primarily utilized aerobic glycolysis, while PAECs relied more heavily on oxidative phosphorylation, to support bioenergetic demands. Repeated single cell cloning and expansion of PAEC colonies generated homogeneous first-generation clones that were highly reflective of the parental population in terms of growth, angiogenic potential, and bioenergetic profiles. Repeated single cell cloning of the first-generation clones generated second-generation clones with increased proliferative potential while maintaining other parental characteristics. Second-generation clones were highly homogeneous populations. Thus, single cell cloning reveals microheterogeneity among the parent cell population and enables isolation of highly representative cells with parental characteristics. SAGE Publications 2017-09-12 /pmc/articles/PMC5703126/ /pubmed/28841087 http://dx.doi.org/10.1177/2045893217731295 Text en © The Author(s) 2017 http://creativecommons.org/licenses/by-nc/4.0/ This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 License (http://www.creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).
spellingShingle Research Articles
Lee, Ji Young
McMurtry, Sarah A.
Stevens, Troy
Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
title Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
title_full Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
title_fullStr Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
title_full_unstemmed Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
title_short Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
title_sort single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5703126/
https://www.ncbi.nlm.nih.gov/pubmed/28841087
http://dx.doi.org/10.1177/2045893217731295
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