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Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells
Rationale: In recent decades, the great potential of human epicardium-derived cells (EPDCs) as an endogenous cell source for cardiac regeneration has been recognized. The limited availability and low proliferation capacity of primary human EPDCs and phenotypic differences between EPDCs obtained from...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8391799/ https://www.ncbi.nlm.nih.gov/pubmed/34440833 http://dx.doi.org/10.3390/cells10082064 |
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author | Ge, Yang Smits, Anke M. Liu, Jia Zhang, Juan van Brakel, Thomas J. Goumans, Marie José T. H. Jongbloed, Monique R. M. de Vries, Antoine A. F. |
author_facet | Ge, Yang Smits, Anke M. Liu, Jia Zhang, Juan van Brakel, Thomas J. Goumans, Marie José T. H. Jongbloed, Monique R. M. de Vries, Antoine A. F. |
author_sort | Ge, Yang |
collection | PubMed |
description | Rationale: In recent decades, the great potential of human epicardium-derived cells (EPDCs) as an endogenous cell source for cardiac regeneration has been recognized. The limited availability and low proliferation capacity of primary human EPDCs and phenotypic differences between EPDCs obtained from different individuals hampers their reproducible use for experimental studies. Aim: To generate and characterize inducible proliferative adult human EPDCs for use in fundamental and applied research. Methods and results: Inducible proliferation of human EPDCs was achieved by doxycycline-controlled expression of simian virus 40 large T antigen (LT) with a repressor-based lentiviral Tet-On system. In the presence of doxycycline, these inducible EPDCs (iEPDCs) displayed high and long-term proliferation capacity. After doxycycline removal, LT expression ceased and the iEPDCs regained their cuboidal epithelial morphology. Similar to primary EPDCs, iEPDCs underwent an epithelial-to-mesenchymal transition (EMT) after stimulation with transforming growth factor β3. This was confirmed by reverse transcription-quantitative polymerase chain reaction analysis of epithelial and mesenchymal marker gene expression and (immuno) cytochemical staining. Collagen gel-based cell invasion assays demonstrated that mesenchymal iEPDCs, like primary EPDCs, possess increased invasion and migration capacities as compared to their epithelial counterparts. Mesenchymal iEPDCs co-cultured with sympathetic ganglia stimulated neurite outgrowth similarly to primary EPDCs. Conclusion: Using an inducible LT expression system, inducible proliferative adult human EPDCs were generated displaying high proliferative capacity in the presence of doxycycline. These iEPDCs maintain essential epicardial characteristics with respect to morphology, EMT ability, and paracrine signaling following doxycycline removal. This renders iEPDCs a highly useful new in vitro model for studying human epicardial properties. |
format | Online Article Text |
id | pubmed-8391799 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-83917992021-08-28 Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells Ge, Yang Smits, Anke M. Liu, Jia Zhang, Juan van Brakel, Thomas J. Goumans, Marie José T. H. Jongbloed, Monique R. M. de Vries, Antoine A. F. Cells Article Rationale: In recent decades, the great potential of human epicardium-derived cells (EPDCs) as an endogenous cell source for cardiac regeneration has been recognized. The limited availability and low proliferation capacity of primary human EPDCs and phenotypic differences between EPDCs obtained from different individuals hampers their reproducible use for experimental studies. Aim: To generate and characterize inducible proliferative adult human EPDCs for use in fundamental and applied research. Methods and results: Inducible proliferation of human EPDCs was achieved by doxycycline-controlled expression of simian virus 40 large T antigen (LT) with a repressor-based lentiviral Tet-On system. In the presence of doxycycline, these inducible EPDCs (iEPDCs) displayed high and long-term proliferation capacity. After doxycycline removal, LT expression ceased and the iEPDCs regained their cuboidal epithelial morphology. Similar to primary EPDCs, iEPDCs underwent an epithelial-to-mesenchymal transition (EMT) after stimulation with transforming growth factor β3. This was confirmed by reverse transcription-quantitative polymerase chain reaction analysis of epithelial and mesenchymal marker gene expression and (immuno) cytochemical staining. Collagen gel-based cell invasion assays demonstrated that mesenchymal iEPDCs, like primary EPDCs, possess increased invasion and migration capacities as compared to their epithelial counterparts. Mesenchymal iEPDCs co-cultured with sympathetic ganglia stimulated neurite outgrowth similarly to primary EPDCs. Conclusion: Using an inducible LT expression system, inducible proliferative adult human EPDCs were generated displaying high proliferative capacity in the presence of doxycycline. These iEPDCs maintain essential epicardial characteristics with respect to morphology, EMT ability, and paracrine signaling following doxycycline removal. This renders iEPDCs a highly useful new in vitro model for studying human epicardial properties. MDPI 2021-08-12 /pmc/articles/PMC8391799/ /pubmed/34440833 http://dx.doi.org/10.3390/cells10082064 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/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 (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Ge, Yang Smits, Anke M. Liu, Jia Zhang, Juan van Brakel, Thomas J. Goumans, Marie José T. H. Jongbloed, Monique R. M. de Vries, Antoine A. F. Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells |
title | Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells |
title_full | Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells |
title_fullStr | Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells |
title_full_unstemmed | Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells |
title_short | Generation, Characterization, and Application of Inducible Proliferative Adult Human Epicardium-Derived Cells |
title_sort | generation, characterization, and application of inducible proliferative adult human epicardium-derived cells |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8391799/ https://www.ncbi.nlm.nih.gov/pubmed/34440833 http://dx.doi.org/10.3390/cells10082064 |
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