Transcriptional analysis of pre and post excision human induced pluripotent stem cells
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE48830 | GEO | 2013/07/13
SECONDARY ACCESSION(S): PRJNA211868
REPOSITORIES: GEO
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Project description:Transcriptional analysis was performed on pre and post excision human induced pluripotent stem cells, the donor human dermal fibroblasts (HDFs) they were derived from and control human embryonic stem cells We isolated total RNA from pre and post excision human induced pluripotent stem cells, the donor human dermal fibroblasts (HDFs) they were derived from and control human embryonic stem cells and analyzed via Affymetrix microarray analysis.
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Project description:Induced cellular reprogramming to the pluripotent state offers a novel stem cell source for autologous transplantation. While recent studies have explored the role of factors required for induced pluripotent stem cell (iPSC) induction, the cellular and molecular basis of reprogramming from human fibroblasts remains elusive. Here, we have identified a subset of human dermal-derived fibroblasts that shares hallmark molecular and epigenetic features with pluripotent cells. Functional studies demonstrate that these cells contribute to the majority of human iPSCs generated from dermal fibroblasts and are dependent on heterogeneous fibroblast microenvironment for reprogramming competency. Molecular characterization indicated these predisposed fibroblasts were unique to other dermal derived stem cells and possessed features of proliferative selfrenewal. Our study reveals human fibroblasts are not equivalently capable of cellular reprogramming, and suggests that reprogramming factors overcome commitment steps that allow predetermined dermal fibroblasts to establish stable pluripotent state.
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Project description:Induced pluripotent stem cells (iPSCs) are usually clonally derived. The selection of fully reprogrammed cells generally involves picking of individual colonies with morphology similar to embryonic stem cells (ESCs). However, successfully reprogrammed cells are highly proliferative and escape from cellular senescence - it is therefore conceivable that they outgrow non-pluripotent and partially reprogrammed cells during culture expansion without the need of clonal selection. In this study, we have reprogrammed human dermal fibroblasts (HDFs) with episomal plasmid vectors. Colony frequency and size was higher when using murine embryonic fibroblasts (MEFs) as stromal support instead of HDFs or human mesenchymal stromal cells (MSCs). We have then compared iPSCs which were either clonally derived by manual selection of a single colony, or derived from bulk-cultures of all initial colonies. After few passages their morphology, expression of pluripotency markers, and gene expression profiles did not reveal any significant differences. Furthermore, clonally-derived and bulk-cultured iPSCs had indistinguishable in vitro differentiation potential towards the three germ layers. Therefore, manual selection of individual colonies does not appear to be necessary for the generation of iPSCs M-bM-^@M-^S this is of relevance for standardization and automation of cell culture procedures Gene expression profiles of induced fibroblasts (M-bM-^@M-^\iFM-bM-^@M-^]) from three different donors which were either clonally derived (M-bM-^@M-^\CM-bM-^@M-^]) or bulk-cultured (M-bM-^@M-^\BM-bM-^@M-^]) were compared
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Project description:Induced pluripotent stem cells (iPSCs) are usually clonally derived. The selection of fully reprogrammed cells generally involves picking of individual colonies with morphology similar to embryonic stem cells (ESCs). However, successfully reprogrammed cells are highly proliferative and escape from cellular senescence - it is therefore conceivable that they outgrow non-pluripotent and partially reprogrammed cells during culture expansion without the need of clonal selection. In this study, we have reprogrammed human dermal fibroblasts (HDFs) with episomal plasmid vectors. Colony frequency and size was higher when using murine embryonic fibroblasts (MEFs) as stromal support instead of HDFs or human mesenchymal stromal cells (MSCs). We have then compared iPSCs which were either clonally derived by manual selection of a single colony, or derived from bulk-cultures of all initial colonies. After few passages their morphology, expression of pluripotency markers, and gene expression profiles did not reveal any significant differences. Furthermore, clonally-derived and bulk-cultured iPSCs had indistinguishable in vitro differentiation potential towards the three germ layers. Therefore, manual selection of individual colonies does not appear to be necessary for the generation of iPSCs – this is of relevance for standardization and automation of cell culture procedures
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