Embyronic stem cell differentiation

Lead Research Organisation: University of Edinburgh
Department Name: UNLISTED

Abstract

Appropriate control of cellular differentiation is critical for coordinating growth and patterning during embryonic development and is essential for maintaining the health and reproductive fitness of adult organisms. Differences in the onset and extent of cellular differentiation underlie much of what is manifest as phenotypic variation in natural populations. However, when the processes are severely deregulated or disrupted they can lead to disease and compromise viability. Cellular differentiation is therefore an area of fundamental importance in biological research and improved understanding of the underlying processes has many beneficial applications in biotechnology, agriculture and biomedicine. All differentiated cells in a mammalian embryo originate from a cluster of unspecialised cells within the inner cell mass of the blastocyst. When these founder cells are explanted in culture they can give rise to pluriptotent embryonic stem (ES) cell lines that will proliferate indefinitely and retain the capacity to differentiate into all embryonic cell types. At present, ES cell lines are routinely derived from blastocysts of mice and a limited number of primates, including human, and represent a unique resource for investigating the basic molecular mechanisms that control differentiation. The ability of ES cells to differentiate into germ cells and reprogramme efficiently upon nuclear transfer also makes them an important tools for genetic modification. Efficient proliferation of undifferentiated ES cells in vitro requires continual suppression of differentiation by extrinsic factors. Disruption of prodifferentiative signaling pathways by mutation or pharmacological inhibition enhances ES cell self-renewal and reduces dependence on extrinsic factors. It seems therefore that tight regulation of prodifferentiative signals is critical for maintaining ES cell pluripotency and self-renewal, as well as driving acquisition of the differentiated phenotype.
 
Description 1. Demonstrated that induced pluripotent stem cell lines derived from pig fibroblasts acquired a biological response to the embryonic stem cell self-renewal factor Leukaemia inhibitory factor
2. Isolated sheep induced pluripotent stem cells that exhibit embryonic stem cell-like characteristics.
3. We assisted, through collaboration with Dr Tilo Kunath, in the generation of the first human induced pluripotent stem cell lines from patient derived fibroblasts carrying a triplication of the alpha-synuclein gene. These pluripotent cell lines were differentiated into human neurons where they can be used to investigate the biochemical effects of ASN over-expression in normal human cells.
4. Demonstrated efficient derivation of rat embryonic stem cells from different genetic strains, and efficient gene targeting at the hprt locus in these cells using homologous recombination.
5. Derived feeder-free cultures of rat embryonic stem cells, that allowed chromatin IP to be performed in relatively pure cultures of rat pluripotent stem cells.
Exploitation Route 1. Pig and sheep induced pluripotent stem cell lines have been requested by other laboratories (Campbell, Nottingham: Smith , Cambridge) .
2. The rat hprt gene targeting vector has been used by a S Korean colleague to target the gene in rat hypoblast stem cells ( Han and Binas, BBRC 2012).
3. We have provided rat embryonic stem cell lines to a number of European laboratories to aid in their research ( Pederson, Aarhus; Geijsen, Utrecht: Gribnau Erasmus Rotterdam.
4. The gene targeting work in rat ESC has lead to a number of collaborations employing rat ESC targeting technology to generate new genetic models in the rat. ( CSFR1- Hume/UoE: CFTR-McLachlan/UoE: VAPB- Skehel/UoE: ICE-HIP - Boland,Luo/Aarhus)
Sectors Education,Pharmaceuticals and Medical Biotechnology

 
Description European Large-scale functional genomics in the rat for translational research
Amount € 357,000 (EUR)
Funding ID 241504 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 04/2010 
End 03/2015
 
Title HPRT knock-out rat 
Description Transgenic rat that carries a mutation in the hprt gene. Similar mutations in humans are responsible for Lesch Nyhan Disease, an X-chromosome linked severe neurological disorder. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Provided To Others? No  
Impact The rat is the preferred rodent model for brain and behaviour studies. This transgenic rat will be useful in assessing behavioural effects of hprt deficiency in a laboratory animal - and may provide useful insights into the molecular basis of Lesch Nyhan Disease. 
 
Description Gene targeting in rat embryonic stem cells 
Organisation University of Edinburgh
Country United Kingdom 
Sector Academic/University 
PI Contribution We provided embryonic stem cell lines and provided expertise in their maintenance and manipulation in culture
Collaborator Contribution Provision of rat embryos and advice on selection of target genes and rat strains
Impact S. Meek, M. Buehr, L. Sutherland, A. Thomson, J. J. Mullins, A. J. Smith, and T. Burdon, 'Efficient Gene Targeting by Homologous Recombination in Rat Embryonic Stem Cells', PLoS One, 5 (2010), e14225. 2010-2013 Gene targeting in rat ES cells by homologous recombination. WP2.2 A Smith , T Burdon, J Mullins. European large-scale functional genomics in the rat for translational research (EURATRANS) consortium £340,000
Start Year 2008
 
Description Identification of Oct4 target genes in human embryonic stem cells 
Organisation Max Planck Society
Department Max Planck Institute for Molecular Cell Biology and Genetics
Country Germany 
Sector Academic/University 
PI Contribution Gene knock-down in human embryonic stem cells
Collaborator Contribution Microarray analysis and bioinfomatics
Impact Y. Babaie, R. Herwig, B. Greber, T. C. Brink, W. Wruck, D. Groth, H. Lehrach, T. Burdon, and J. Adjaye, 'Analysis of Oct4-Dependent Transcriptional Networks Regulating Self-Renewal and Pluripotency in Human Embryonic Stem Cells', Stem Cells, 25 (2007), 500-10.