Abstract
Hypoxia benefits undifferentiated pluripotent stem cell renewal, and 2-oxoglutarate (2OG) dioxygenases have been implicated in pluripotent stem cell induction and renewal. We show in human embryonic stem cells (hESC) that an ambient oxygen-induced oxidative stress response elicited by culture in a hypoxic atmosphere (0.5% O2) correlates with the expression of 2OG dioxygenases, which oxidise DNA (TET1, 2, 3) and histone H3 (KDM4C), the former reflected by elevation in genomic 5-hydroxymethylcytosine (5hmC). siRNA-mediated targeting of KDM4C and TET1–3 induces hESC differentiation. Under ambient atmospheric oxygen (21% O2), exposure to a low inhibitory concentration of sodium arsenite (NaAsO2, IC10), as a model of chemically-induced oxidative stress, suppresses antioxidant gene expression, reduces mitochondrial membrane potential and induces hESC differentiation. Co-administration of the antioxidant N-acetyl-L-cysteine promoted anti-oxidant, pluripotency and 2OG dioxygenase gene expression, elevated genomic hydroxymethylation and blocked induction of differentiation. Transient ectopic expression of KDM4C or TET1 in ambient atmospheric oxygen achieved the same. Our study substantiates a role for 2OG-dependent dioxygenases in hypoxia's promotion of undifferentiated hESC self-renewal.
Highlights
Efforts to exploit the potential of human embryonic stem cells and analogous induced pluripotent stem cells in regenerative medicine are ongoing
Conserved outcomes between cell lines are denoted throughout all figures by a solid inverted triangle (▼) with degree of statistical significance noted by asterisks as follows: *p
Using RT-qPCR to assess mRNA steady state levels hypoxia increased the relative expression of pluripotency-associated transcription factors (OCT4, NANOG; p
Summary
Efforts to exploit the potential of human embryonic stem cells (hESCs) and analogous induced pluripotent stem cells (hiPSCs) in regenerative medicine are ongoing These include 5 their use as source material for cell therapy products such as for the treatment of conditions characterised by acute or chronic cell loss (e.g. macular degeneration, repair of spinal cord trauma, Parkinson’s and Huntington’s diseases, cardiomyopathy; for up to date accounting go to www.clinicaltrials.gov) and in cell banking initiatives to provide a standardised resource for discovery An improved understanding of the mechanisms of hypoxia-mediated benefits is likely to assist in changing practices
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