The role of low oxygen in the self-renewal and neural differentiation of human pluripotent stem cells

Steeg, Rachel 2015. The role of low oxygen in the self-renewal and neural differentiation of human pluripotent stem cells. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

Human embryonic stem cells are derived from the pre-implantation blastocyst, residing in an extremely low oxygen environment. Application of this in vivo oxygen concentration to an in vitro setting has previously shown to be essential for driving the self-renewal and directed differentiation of human pluripotent stem cells. However, studies on hESCs at low oxygen have been frequently contra-indicatory. Here, comparative analysis of a hESC and iPSC line at a low oxygen concentration resulted in divergent effects across the two cell lines. Augmented TGFβ signalling was observed in conjunction with up-regulated transcription of pluripotency markers NANOG and POU5F1 in hESCs but not iPSCs. IPSCs also shifted to a state of increased proliferation whereas hESCs did not. It was also observed that exposure of hESCs and iPSCs to light throughout low oxygen culture induced large amounts of apoptosis, highlighting the requirement for careful selection of cell culture equipment for environmental oxygen control. Both the embryonic and adult brain retain tissue specific oxygen concentrations far below that at atmospheric oxygen concentrations. Previous studies showed that embryonic silencing of factors responsive to low oxygen caused a range of embryonic abnormalities, including deformation of the neural plate and tube. Here, differentiation of hESCs to an early neuroectodermal identity at low O2 did not definitively augment production of NPCs but did additively suppress BMP signalling above that at atmospheric O2. A concordant rise in apoptosis was also observed in a manner both independent of, and augmented by, exogenous BMP inhibition. Subsequently, neurogenesis at low oxygen produced terminal neurons with accelerated and augmented synaptic and induced excitability. This was characterised by a rise in the rate of membrane depolarisation, increased action potential overshoot and accelerated expression of pre-synaptic marker Synaptophysin. These results highlight a novel, critical role for low oxygen in augmenting the excitability of hESC derived neurons.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Biosciences
Subjects:	Q Science > QH Natural history > QH301 Biology Q Science > QH Natural history > QH426 Genetics
Funders:	KESS
Date of First Compliant Deposit:	30 March 2016
Last Modified:	21 Feb 2022 16:23
URI:	https://orca.cardiff.ac.uk/id/eprint/69784

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