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Effect on Multipotency and Phenotypic Transition of Unrestricted Somatic Stem Cells from Human Umbilical Cord Blood after Treatment with Epigenetic Agents.

ABSTRACT: The epigenetic mechanism of DNA methylation is of central importance for cellular differentiation processes. Unrestricted somatic stem cells (USSCs) from human umbilical cord blood, which have a broad differentiation spectrum, reside in an uncommitted epigenetic state with partial methylation of the regulatory region of the gene coding for the pluripotency master regulator OCT4. Thus we hypothesized that further opening of this "poised" epigenetic state could broaden the differentiation potential of USSCs. Here we document that USSCs drastically change their phenotype after treatment by a new elaborated cultivation protocol which utilizes the DNA hypomethylating compound 5'-aza-2-deoxycytidine (5-Aza-CdR) and the histone deacetylase inhibitor trichostatin A (TSA). This treatment leads to a new stable, spheroid-forming cell type which we have named SpheUSSC. These cells can be stably propagated over at least 150 cell divisions, express OCT4, retain the potential to undergo osteogenic differentiation, and have additionally acquired the ability to uniformly differentiate into adipocytes, unlike the source USSC population. Here we describe our treatment protocol and provide evidence that it induces a dedifferentiation step and concomitantly the acquisition of an extended differentiation capability of the new SpheUSSC type.

SUBMITTER: Ghanjati F

PROVIDER: S-EPMC4691642 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Effect on Multipotency and Phenotypic Transition of Unrestricted Somatic Stem Cells from Human Umbilical Cord Blood after Treatment with Epigenetic Agents.

Ghanjati Foued F Santourlidis Simeon S

Stem cells international 20151214

The epigenetic mechanism of DNA methylation is of central importance for cellular differentiation processes. Unrestricted somatic stem cells (USSCs) from human umbilical cord blood, which have a broad differentiation spectrum, reside in an uncommitted epigenetic state with partial methylation of the regulatory region of the gene coding for the pluripotency master regulator OCT4. Thus we hypothesized that further opening of this "poised" epigenetic state could broaden the differentiation potentia ...[more]

PMID: 26788071

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Project description:Induced pluripotent stem (iPS) cells have been generated from mouse and human somatic cells by ectopic expression of the transcription factors OCT4, SOX2, KLF4, c-MYC as well as NANOG and LIN28. Here we report generation of induced pluripotent stem cells from human umbilical cord blood derived unrestricted somatic stem cells (USSC) using retroviral expression of the transcription factors OCT4, SOX2, KLF4 and C-MYC and evaluation of their molecular signature and differentiation potential in comparison to human embryonic stem cells. The reprogrammed cells (HUiPS) were analysed morphologically, by qRT-PCR, global miRNA and epigenetic profiling and gene expression microarrays, as well as in their in vitro and in vivo differentiation potential by embryoid body formation and teratoma assay. The cord blood iPS cells are highly similar to human embryonic stem cells morphologically, at their molecular signature as well as in their in vitro and in vivo differentiation potential. Human cord blood derived unrestricted somatic stem cells offer an attractive source of cells for the generation of induced pluripotent stem cells. Our findings open novel perspectives to generate HLA matched pluripotent stem cell banks based on existing cord blood banks. Besides its obvious relevance of such a second generation cord blood iPS bank for pharmacological and toxicological testing, its application for autologous or allogenic regenerative cell transplantation appears feasible. For transcriptome profiling, 400 ng of total DNA-free RNA was used as input for labelled cRNA synthesis (Illumina TotalPrep RNA Amplification Kit - Ambion) following the manufacturer's instructions (IVT: 10h). Quality-checked cRNA samples were hybridized as biological or technical duplicates for 18 h onto HumanRef-8 v3 expression BeadChips (Illumina), washed, stained, and scanned following guidelines and using materials / instrumentation supplied / suggested by the manufacturer. Six sample types were analyzed, each one of them in duplicate. USSC: human umbilical cord blood unrestricted somatic stem cells (duplicates) HUiPS: human iPS cells from human umbilical cord blood USSC, hand-picked cols (duplicates) H9 hESC: H9 human ESCs grown on low-density CF1 MEFs (duplicates) H1 hESC: H1 human ESCs grown on low-density CF1 MEFs (duplicates) 1F hNiPS: One factor (Oct4) human iPS cells from hNSCs, hand-picked cols (duplicates) 2F hNiPS: Two factors (Oct4, Klf4) human iPS cells from hNSCs, hand-picked cols (duplicates)

Dataset Information

Effect on Multipotency and Phenotypic Transition of Unrestricted Somatic Stem Cells from Human Umbilical Cord Blood after Treatment with Epigenetic Agents.

Publications

Effect on Multipotency and Phenotypic Transition of Unrestricted Somatic Stem Cells from Human Umbilical Cord Blood after Treatment with Epigenetic Agents.

Similar Datasets

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