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...
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2015-01-01
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| Series: | PLoS ONE |
| Online Access: | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0118307&type=printable |
| _version_ | 1826586542057979904 |
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| author | Ibon Garitaonandia Hadar Amir Francesca Sesillo Boscolo Gerald K Wambua Heather L Schultheisz Karen Sabatini Robert Morey Shannon Waltz Yu-Chieh Wang Ha Tran Trevor R Leonardo Kristopher Nazor Ileana Slavin Candace Lynch Yingchun Li Ronald Coleman Irene Gallego Romero Gulsah Altun David Reynolds Stephen Dalton Mana Parast Jeanne F Loring Louise C Laurent |
| author_facet | Ibon Garitaonandia Hadar Amir Francesca Sesillo Boscolo Gerald K Wambua Heather L Schultheisz Karen Sabatini Robert Morey Shannon Waltz Yu-Chieh Wang Ha Tran Trevor R Leonardo Kristopher Nazor Ileana Slavin Candace Lynch Yingchun Li Ronald Coleman Irene Gallego Romero Gulsah Altun David Reynolds Stephen Dalton Mana Parast Jeanne F Loring Louise C Laurent |
| author_sort | Ibon Garitaonandia |
| collection | DOAJ |
| 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. |
| first_indexed | 2024-12-20T05:14:18Z |
| format | Article |
| id | doaj.art-7a2844f2408f4d81be97633913b4efe8 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2025-03-14T16:11:11Z |
| publishDate | 2015-01-01 |
| publisher | Public Library of Science (PLoS) |
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| series | PLoS ONE |
| spelling | doaj.art-7a2844f2408f4d81be97633913b4efe82025-02-23T05:32:11ZengPublic Library of Science (PLoS)PLoS ONE1932-62032015-01-01102e011830710.1371/journal.pone.0118307Increased risk of genetic and epigenetic instability in human embryonic stem cells associated with specific culture conditions.Ibon GaritaonandiaHadar AmirFrancesca Sesillo BoscoloGerald K WambuaHeather L SchultheiszKaren SabatiniRobert MoreyShannon WaltzYu-Chieh WangHa TranTrevor R LeonardoKristopher NazorIleana SlavinCandace LynchYingchun LiRonald ColemanIrene Gallego RomeroGulsah AltunDavid ReynoldsStephen DaltonMana ParastJeanne F LoringLouise C LaurentThe 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.https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0118307&type=printable |
| spellingShingle | Ibon Garitaonandia Hadar Amir Francesca Sesillo Boscolo Gerald K Wambua Heather L Schultheisz Karen Sabatini Robert Morey Shannon Waltz Yu-Chieh Wang Ha Tran Trevor R Leonardo Kristopher Nazor Ileana Slavin Candace Lynch Yingchun Li Ronald Coleman Irene Gallego Romero Gulsah Altun David Reynolds Stephen Dalton Mana Parast Jeanne F Loring Louise C Laurent Increased risk of genetic and epigenetic instability in human embryonic stem cells associated with specific culture conditions. PLoS ONE |
| 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 |
| url | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0118307&type=printable |
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