Homo sapiens
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ABSTRACT:
PROVIDER: PRJNA208331 | ENA |
REPOSITORIES: ENA
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Project description:The self-renewal and differentiation capacities of human pluripotent stem cells (hPSCs) make them good sources of cells 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, to on the genetic and epigenetic stability and the phenotypic characteristics of hPSCs. In extensive experiments in hEScs involving over 100 continuous passages, , we observed that both enzymatic passaging and feeder-free culture were associated with genetic instability, higher cell proliferation, and persistence of OCT4-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 observedculture-dependent variations in global gene expression and DNA methylation. Verification of the negative effects of enzymatic passaging and feeder-free conditions was performed in hiPSC cultures. Our results highlight the need for careful assessment of effects of culture conditions on cells intended for clinical therapies. Three independent hiPSC clones reprogrammed from human fetal dermal fibroblasts: HDF51iPS1, HDF51iPS7, and HDF51iPS11.
2015-03-29 | E-GEOD-56834 | biostudies-arrayexpress
Project description:The self-renewal and differentiation capacities of human pluripotent stem cells (hPSCs) make them good sources of cells 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, to on the genetic and epigenetic stability and the phenotypic characteristics of hPSCs. In extensive experiments in hEScs involving over 100 continuous passages, , we observed that both enzymatic passaging and feeder-free culture were associated with genetic instability, higher cell proliferation, and persistence of OCT4-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 observedculture-dependent variations in global gene expression and DNA methylation. Verification of the negative effects of enzymatic passaging and feeder-free conditions was performed in hiPSC cultures. Our results highlight the need for careful assessment of effects of culture conditions on cells intended for clinical therapies.
2015-03-29 | GSE56834 | GEO
Project description:Genomic instability in human embryonic stem cells is associated to culture density and medium acidification.
| PRJNA251888 | ENA
Project description:The self-renewal and differentiation capacities of human pluripotent stem cells (hPSCs) make them good sources of cells 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 applications require extensive expansion of hPSC cultures, a process that has been associated with genetic and epigenetic alterations. We have performed a systematic study over more than 100continuous passages to identify characteristics of culture conditions (including passage method, substrate, and media type) that influence the genetic and epigenetic stability and the phenotypic characteristics of hPSCs. The predominant effects we observed were increased genetic instability with enzymatic passage, higher cell proliferation with feeder-free substrate, and variations among cultures in global gene expression and DNA methylation with time in culture. We observed recurrent duplications in two genomic regions that have been noted in earlier studies to be hotspots for duplication in hPSCs, as well as a previously unreported recurrent deletion of the tumor suppressor gene TP53 in all but one of the long-term culture conditions; the exception was the condition using mechanical passaging on feeder layers. The deletion of TP53 is associated with decreased mRNA expression of TP53, as well as alterations in the expression of several other genes in the TP53 pathway, which taken together indicate a decrease in the function of the TP53 pathway. Our results highlight the need for careful assessment of effects of culture conditions on cells intended for clinical therapies. Total RNA extracted at different passages from Human Embryonic Stem Cells in different culture conditions. Total DNA extracted at different passages from Human Embryonic Stem Cells in different culture conditions.
2015-03-29 | E-GEOD-34982 | biostudies-arrayexpress
Project description:Transcriptome of embryonic stem cells cultured in enzymatic and mechanic passaging methods [RNA-seq]
| PRJNA208433 | ENA
Project description:Induced pluripotent stem cells (iPSCs) outwardly appear to be indistinguishable from embryonic stem cells (ESCs). A study of gene expression profiles of mouse and human ESCs and iPSCs suggests that, while iPSCs are quite similar to their embryonic counterparts, a recurrent gene expression signature appears in iPSCs regardless of their origin or the method by which they were generated. Upon extended culture, hiPSCs adopt a gene expression profile more similar to hESCs; however, they still retain a gene expression signature unique from hESCs that extends to miRNA expression. Genome-wide data suggested that the iPSC signature gene expression differences are due to differential promoter binding by the reprogramming factors. High-resolution array profiling demonstrated that there is no common specific subkaryotypic alteration that is required for reprogramming and that reprogramming does not lead to genomic instability. Together, these data suggest that iPSCs should be considered a unique subtype of pluripotent cell. Detailed analysis comparing induced pluripotent stem cells revealed functional gene expression differences to hESCs that are changed with long term passaging. We used microarrays to detail the changes that are made throughout passaging to hiPSC lines.
2009-07-10 | GSE16654 | GEO
Project description:Recurrent genomic instability of chromosome 1q in neural derivatives of human embryonic stem cells
| PRJNA148481 | ENA
Project description:By analyzing five human embryonic stem (hES) cell lines over long-term culture, we detected a recurrent genomic instability in the human genome. An amplification of 2.5–4.6 Mb at 20q11.21, encompassing about 23 genes in common, appeared in four cell lines of different origins. This amplification, which has been associated with oncogenic transformation, may provide a selective advantage to hES cells in culture
2008-11-19 | GSE13565 | GEO
Project description:There are a total of four samples each for this analysis. Each sample consists of the cells grown on three 10 cm culture plates. Each plate should have 2x106 cells for a total of 6x106 cells per sample when all three plates are combined. The first sample is undifferentiated human embryonic stem cells, the second sample is human glutamatergic neurons derived from those human embryonic stem cells, the third sample is undifferentiated human induced pluripotent stem cells and the fourth sample is human glutamatergic neurons derived from those human induced pluripotent stem cells.
2017-06-19 | ST000768 | MetabolomicsWorkbench
Project description:By analyzing five human embryonic stem (hES) cell lines over long-term culture, we detected a recurrent genomic instability in the human genome. An amplification of 2.5â4.6 Mb at 20q11.21, encompassing about 23 genes in common, appeared in four cell lines of different origins. This amplification, which has been associated with oncogenic transformation, may provide a selective advantage to hES cells in culture Array-based comparative genomic hybridization, was performed on genomic DNA samples from different human embryonic stem cell lines, all cultured in our laboratory under the same conditions. VUB05-HD is an hES cell line carrying the Huntingtin mutant gene H1, H9, SA01, and VUB01 are various hES cell lines GSM341552 and GSM341553: SNP analysis GSM341589 to GSM341669: CGH analysis
2010-05-16 | E-GEOD-13565 | biostudies-arrayexpress