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Embryonic stem cells: modelling effects ofearly embryo environment

Embryonic stem cells: modelling effects ofearly embryo environment
Embryonic stem cells: modelling effects ofearly embryo environment
The Developmental Origins of Health and Disease (DOHaD) hypothesis proposes that embryonic environment can induce permanent changes in metabolism during development, increasing the risk of disease in adults. Adverse environments during critical stages of gestation are sufficient to induce adaptations in offspring and disease susceptibility in later life. Rodent models show that maternal diet exclusively during preimplantation development induces cardiovascular and metabolic disease in adult offspring. Changes must therefore occur within the distinct cell populations of the early embryo and be maintained throughout development. Determining adaptive mechanisms has been challenging due to the small size of the early embryo, and genetic variability in outbred strains previously used. We generated mouse embryonic stem (ES) cells from inbred C57BL/6 mice as a model to overcome these problems. These were used to characterise mechanisms associated with the embryo’s adaptive responses to maternal diet. ES cell lines were derived from blastocysts of C57BL/6 mice assigned to either an isocaloric low protein diet (LPD), or a control diet exclusively through preimplantation development. ES cell lines were characterised for karyotype, sex, gene expression, and functional characteristics including proliferation, death, and metabolism at standardised passages. LPD had no impact on blastocyst formation in vivo or blastocyst cell lineage allocation. Experimental conditions did affect blastocyst outgrowth development in vitro. LPDoutgrowths cultured with less feeder fibroblasts showed slower development than controls. Although LPD blastocyst outgrowth was comparable to controls under high feeder growth conditions, there was a significant reduction in the capacity for ES cell derivation. There was a prominent sex bias towards male ES cell lines. These ES cells retained similar levels of gene expression related to pluripotency, housekeeping and developmental functions irrespective of diet. LPD did not affect growth or metabolism. These cells however showed increased basal apoptosis, and reduced levels of phosphorylated Extracellular signal-regulated kinase (ERK). The reduced ES cell isolation efficiency may indicate a reduced number of pluripotent cells present within the early embryo or increased sensitivity of these cells in response to maternal LPD. Increased apoptosis in ES cells derived from LPD-blastocysts reveal that these cells are indeed more sensitive. Reduced activated ERK may suggest that dysregulated ERK-mediated survival signalling causes enhanced apoptosis. Such adaptations in the early embryo may impact on lineage allocation as differentiation occurs. These ES cell lines may provide a model to investigate such mechanistic adaptations in post-implantation tissues providing further insight into foetal responses to poor nutrition and the induction of adult onset disease.
Cox, Andrew
43b91b2e-2d5e-4484-bc3c-cb8eaa1d367c
Cox, Andrew
43b91b2e-2d5e-4484-bc3c-cb8eaa1d367c
Smyth, Neil
0eba2a40-3b43-4d40-bb64-621bd7e9d505
Fleming, Thomas
2abf761a-e5a1-4fa7-a2c8-12e32d5d4c03

Cox, Andrew (2013) Embryonic stem cells: modelling effects ofearly embryo environment. University of Southampton, Bological Sciences, Doctoral Thesis, 236pp.

Record type: Thesis (Doctoral)

Abstract

The Developmental Origins of Health and Disease (DOHaD) hypothesis proposes that embryonic environment can induce permanent changes in metabolism during development, increasing the risk of disease in adults. Adverse environments during critical stages of gestation are sufficient to induce adaptations in offspring and disease susceptibility in later life. Rodent models show that maternal diet exclusively during preimplantation development induces cardiovascular and metabolic disease in adult offspring. Changes must therefore occur within the distinct cell populations of the early embryo and be maintained throughout development. Determining adaptive mechanisms has been challenging due to the small size of the early embryo, and genetic variability in outbred strains previously used. We generated mouse embryonic stem (ES) cells from inbred C57BL/6 mice as a model to overcome these problems. These were used to characterise mechanisms associated with the embryo’s adaptive responses to maternal diet. ES cell lines were derived from blastocysts of C57BL/6 mice assigned to either an isocaloric low protein diet (LPD), or a control diet exclusively through preimplantation development. ES cell lines were characterised for karyotype, sex, gene expression, and functional characteristics including proliferation, death, and metabolism at standardised passages. LPD had no impact on blastocyst formation in vivo or blastocyst cell lineage allocation. Experimental conditions did affect blastocyst outgrowth development in vitro. LPDoutgrowths cultured with less feeder fibroblasts showed slower development than controls. Although LPD blastocyst outgrowth was comparable to controls under high feeder growth conditions, there was a significant reduction in the capacity for ES cell derivation. There was a prominent sex bias towards male ES cell lines. These ES cells retained similar levels of gene expression related to pluripotency, housekeeping and developmental functions irrespective of diet. LPD did not affect growth or metabolism. These cells however showed increased basal apoptosis, and reduced levels of phosphorylated Extracellular signal-regulated kinase (ERK). The reduced ES cell isolation efficiency may indicate a reduced number of pluripotent cells present within the early embryo or increased sensitivity of these cells in response to maternal LPD. Increased apoptosis in ES cells derived from LPD-blastocysts reveal that these cells are indeed more sensitive. Reduced activated ERK may suggest that dysregulated ERK-mediated survival signalling causes enhanced apoptosis. Such adaptations in the early embryo may impact on lineage allocation as differentiation occurs. These ES cell lines may provide a model to investigate such mechanistic adaptations in post-implantation tissues providing further insight into foetal responses to poor nutrition and the induction of adult onset disease.

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Published date: 31 May 2013
Organisations: University of Southampton, Centre for Biological Sciences

Identifiers

Local EPrints ID: 354427
URI: https://eprints.soton.ac.uk/id/eprint/354427
PURE UUID: 6867a4e2-9a36-4678-940d-d51b2831bcf9

Catalogue record

Date deposited: 22 Oct 2013 10:13
Last modified: 02 Feb 2018 05:02

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Contributors

Author: Andrew Cox
Thesis advisor: Neil Smyth
Thesis advisor: Thomas Fleming

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