Unknown,Transcriptomics,Genomics,Proteomics

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Transcriptional Profiling of Hypoxic Neural Stem Cells to Identify Novel Regulators of Neural Stem Cell Biology

ABSTRACT: Neural stem cells reside in a hypoxic microenvironment within the brain. However, the crucial transcription factors that regulate neural stem cell biology under physiologic hypoxia are poorly understood. Here, we have performed microarray analysis of hypoxic versus normoxic neural stem cells with the aim of identifying pathways and transcription factors that are activated under oxygen concentrations mimicking normal brain tissue microenvironment. To identify the molecular mechanisms mediating the effect of low oxygen levels on NSC biology, we profiled the global effect of sustained, physiologic hypoxia on NSC gene expression using an unbiased approach by genome-wide microarray analysis. NSC were isolated from E13.5 mouse embryos (OF-1 strain) as previously described (Johe et al., 1996). After isolation, cells were immediately cultured at 37°C, 5% CO2 and either 5% or atmospheric (≈21%) oxygen and maintained in these conditions for 2-3 passages (minimum of 10 days). Then, total RNA was extracted, labeled and hybridized in a SurePrint G3 Mouse GE 8x60k array (Agilent Technologies). Four independent experiments were performed using different mouse donors for each experiment.

ORGANISM(S): Mus musculus

SUBMITTER: Nuria de la Iglesia

PROVIDER: E-GEOD-68572 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Reprogramming of somatic cells is a valuable tool to understand the mechanisms of regaining pluripotency and further opens up the possibility to generate patient-specific pluripotent stem cells. Reprogramming of mouse and human somatic cells into pluripotent stem cells, designated as induced pluripotent stem (iPS) cells, has been possible with the expression of the transcription factor quartet Oct4, Sox2, c-Myc, and Klf4. Considering that ectopic expression of c-Myc causes tumourigenicity in offspring and retroviruses themselves can cause insertional mutagenesis, the generation of iPS cells with a minimal number of factors may hasten the clinical application of this approach. Here, we show that adult mouse neural stem cells (NSCs) express higher endogenous levels of Sox2 and c-Myc than ES cells and that exogenous Oct4 together with either Klf4 or c-Myc are sufficient to generate iPS cells from NSCs. These two-factor (2F) iPS cells are similar to embryonic stem cells at the molecular level, contribute to development of the germline, and form chimeras. We propose that, in inducing pluripotency, the number of reprogramming factors can be reduced when using somatic cells that endogenously express appropriate levels of complementing factors. Experiment Overall Design: 8 hybridizations in total. Experiment Overall Design: NSC derived iPS cells by 2 factors (Oct4 and Klf4) in triplicate: Experiment Overall Design: - iPS cell_2F_1 Experiment Overall Design: - iPS cell_2F_2 Experiment Overall Design: - iPS cell_2F_3 Experiment Overall Design: Embryonic Stem cells (ESC) in triplicate: Experiment Overall Design: - ESC_1 Experiment Overall Design: - ESC_2 Experiment Overall Design: - ESC_3 Experiment Overall Design: NSC cultures in duplicates: Experiment Overall Design: - NSC_2 Experiment Overall Design: - NSC_3 Experiment Overall Design: NSC derived iPS cells by 4 factors (Oct4, Sox2, c-Myc and Klf4) in triplicate: Experiment Overall Design: - iPS cell_4F_1 Experiment Overall Design: - iPS cell_4F_2 Experiment Overall Design: - iPS cell_4F_3

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Transcriptional Profiling of Hypoxic Neural Stem Cells to Identify Novel Regulators of Neural Stem Cell Biology

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