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Comparison of gene expression in human embyronic stem cells grown in MEF-CM or defined media conditions

ABSTRACT: A fully defined media for human embryonic stem cells (hESCs) was designed after examining the phosphorylation status of 42 receptor tyrosine kinases (RTKs) on hESCs exposed to mouse embryonic fibroblast conditioned medium (CM). Activation of the insulin and insulin-like growth factor-1 receptors (IR, IGF1R) as well as EGFR family members including ERBB2 and ERBB3 led to the design of a simple defined media which supported robust long-term growth of multiple hESC lines and allowed massive expansion of undifferentiated cells. Illumina bead arrays were used to demonstrate the maintenance of gene expression profiles characteristic of pluripotent cells in cultures grown in CM or defined media. Keywords: Comparison of cells in different culture conditions

ORGANISM(S): Homo sapiens

PROVIDER: GSE6645 | GEO | 2007/06/24

SECONDARY ACCESSION(S): PRJNA98823

REPOSITORIES: GEO

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Project description:The realization of human embryonic stem cells (hESC) as a model for human developmental hematopoiesis and potential cell replacement strategies relies on an improved understanding of the extrinsic and intrinsic factors regulating hematopoietic-specific hESC differentiation. Mesenchymal stem cells (hMSCs) are multipotent cells of mesodermal origin that form part of hematopoietic stem cell niches and have an important role in the regulation of hematopoiesis through production of secreted factors and/or cell-to-cell interactions. We have previously shown that hESCs may be successfully maintained feeder-free using hMSC-conditioned media (MSC-CM). Here, we hypothesized that hESCs maintained in MSC-CM may be more prone to differentiation towards hematopoietic lineage than hESCs grown in standard human foreskin fibroblast (HFF)-conditioned media (HFF-CM). We report that specification into hemogenic progenitors and subsequent hematopoietic differentiation and clonogenic progenitor capacity is robustly enhanced in hESC lines maintained in MSC-CM. Interestingly, co-culture of hESCs on hMSCs fully abrogates hematopoietic specification of hESCs suggesting that the improved hematopoietic differentiation is mediated by MSC-secreted factors rather than by MSC-hESC physical interactions. To investigate the molecular mechanism involved in this process, we analyzed global (LINE-1) methylation and genome-wide promoter DNA methylation. Human ESCs grown in MSC-CM showed a decrease of 20% in global DNA methylation and a promoter DNA methylation signature consisting in 45 genes commonly hypomethylated and 102 genes frequently hypermethylated. Our data indicate that maintenance of hESCs in MSC-CM robustly augments hematopoietic specification and that the process seems mediated by MSC-secreted factors conferring a DNA methylation signature to undifferentiated hESCs which may influence further predisposition towards hematopoietic specification. Total DNA isolated by standard procedures from human embryonic stem cells (hESC) cultured in different conditioned media

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Comparison of gene expression in human embyronic stem cells grown in MEF-CM or defined media conditions

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