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Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions
The self-renewal and differentiation capacities of human pluripotent stem cells (hPSCs) make them a promising source of material for cell transplantation therapy, drug development, and studies of cellular differentiation and development. However, the large numbers of cells necessary for many of thes...
Autores principales: | , , , , , , , , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4340884/ https://www.ncbi.nlm.nih.gov/pubmed/25714340 http://dx.doi.org/10.1371/journal.pone.0118307 |
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author | Garitaonandia, Ibon Amir, Hadar Boscolo, Francesca Sesillo Wambua, Gerald K. Schultheisz, Heather L. Sabatini, Karen Morey, Robert Waltz, Shannon Wang, Yu-Chieh Tran, Ha Leonardo, Trevor R. Nazor, Kristopher Slavin, Ileana Lynch, Candace Li, Yingchun Coleman, Ronald Gallego Romero, Irene Altun, Gulsah Reynolds, David Dalton, Stephen Parast, Mana Loring, Jeanne F. Laurent, Louise C. |
author_facet | Garitaonandia, Ibon Amir, Hadar Boscolo, Francesca Sesillo Wambua, Gerald K. Schultheisz, Heather L. Sabatini, Karen Morey, Robert Waltz, Shannon Wang, Yu-Chieh Tran, Ha Leonardo, Trevor R. Nazor, Kristopher Slavin, Ileana Lynch, Candace Li, Yingchun Coleman, Ronald Gallego Romero, Irene Altun, Gulsah Reynolds, David Dalton, Stephen Parast, Mana Loring, Jeanne F. Laurent, Louise C. |
author_sort | Garitaonandia, Ibon |
collection | PubMed |
description | The self-renewal and differentiation capacities of human pluripotent stem cells (hPSCs) make them a promising source of material for cell transplantation therapy, drug development, and studies of cellular differentiation and development. However, the large numbers of cells necessary for many of these applications require extensive expansion of hPSC cultures, a process that has been associated with genetic and epigenetic alterations. We have performed a combinatorial study on both hESCs and hiPSCs to compare the effects of enzymatic vs. mechanical passaging, and feeder-free vs. mouse embryonic fibroblast feeder substrate, on the genetic and epigenetic stability and the phenotypic characteristics of hPSCs. In extensive experiments involving over 100 continuous passages, we observed that both enzymatic passaging and feeder-free culture were associated with genetic instability, higher rates of cell proliferation, and persistence of OCT4/POU5F1-positive cells in teratomas, with enzymatic passaging having the stronger effect. In all combinations of culture conditions except for mechanical passaging on feeder layers, we noted recurrent deletions in the genomic region containing the tumor suppressor gene TP53, which was associated with decreased mRNA expression of TP53, as well as alterations in the expression of several downstream genes consistent with a decrease in the activity of the TP53 pathway. Among the hESC cultures, we also observed culture-associated variations in global gene expression and DNA methylation. The effects of enzymatic passaging and feeder-free conditions were also observed in hiPSC cultures. Our results highlight the need for careful assessment of the effects of culture conditions on cells intended for clinical therapies. |
format | Online Article Text |
id | pubmed-4340884 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-43408842015-03-04 Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions Garitaonandia, Ibon Amir, Hadar Boscolo, Francesca Sesillo Wambua, Gerald K. Schultheisz, Heather L. Sabatini, Karen Morey, Robert Waltz, Shannon Wang, Yu-Chieh Tran, Ha Leonardo, Trevor R. Nazor, Kristopher Slavin, Ileana Lynch, Candace Li, Yingchun Coleman, Ronald Gallego Romero, Irene Altun, Gulsah Reynolds, David Dalton, Stephen Parast, Mana Loring, Jeanne F. Laurent, Louise C. PLoS One Research Article The self-renewal and differentiation capacities of human pluripotent stem cells (hPSCs) make them a promising source of material for cell transplantation therapy, drug development, and studies of cellular differentiation and development. However, the large numbers of cells necessary for many of these applications require extensive expansion of hPSC cultures, a process that has been associated with genetic and epigenetic alterations. We have performed a combinatorial study on both hESCs and hiPSCs to compare the effects of enzymatic vs. mechanical passaging, and feeder-free vs. mouse embryonic fibroblast feeder substrate, on the genetic and epigenetic stability and the phenotypic characteristics of hPSCs. In extensive experiments involving over 100 continuous passages, we observed that both enzymatic passaging and feeder-free culture were associated with genetic instability, higher rates of cell proliferation, and persistence of OCT4/POU5F1-positive cells in teratomas, with enzymatic passaging having the stronger effect. In all combinations of culture conditions except for mechanical passaging on feeder layers, we noted recurrent deletions in the genomic region containing the tumor suppressor gene TP53, which was associated with decreased mRNA expression of TP53, as well as alterations in the expression of several downstream genes consistent with a decrease in the activity of the TP53 pathway. Among the hESC cultures, we also observed culture-associated variations in global gene expression and DNA methylation. The effects of enzymatic passaging and feeder-free conditions were also observed in hiPSC cultures. Our results highlight the need for careful assessment of the effects of culture conditions on cells intended for clinical therapies. Public Library of Science 2015-02-25 /pmc/articles/PMC4340884/ /pubmed/25714340 http://dx.doi.org/10.1371/journal.pone.0118307 Text en © 2015 Garitaonandia et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Garitaonandia, Ibon Amir, Hadar Boscolo, Francesca Sesillo Wambua, Gerald K. Schultheisz, Heather L. Sabatini, Karen Morey, Robert Waltz, Shannon Wang, Yu-Chieh Tran, Ha Leonardo, Trevor R. Nazor, Kristopher Slavin, Ileana Lynch, Candace Li, Yingchun Coleman, Ronald Gallego Romero, Irene Altun, Gulsah Reynolds, David Dalton, Stephen Parast, Mana Loring, Jeanne F. Laurent, Louise C. Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions |
title | Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions |
title_full | Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions |
title_fullStr | Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions |
title_full_unstemmed | Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions |
title_short | Increased Risk of Genetic and Epigenetic Instability in Human Embryonic Stem Cells Associated with Specific Culture Conditions |
title_sort | increased risk of genetic and epigenetic instability in human embryonic stem cells associated with specific culture conditions |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4340884/ https://www.ncbi.nlm.nih.gov/pubmed/25714340 http://dx.doi.org/10.1371/journal.pone.0118307 |
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