Culture condition reverses epiblast stem cells into ES-cell like cells – Until when does the pluripotency lose during development?

It is shocking! Just by using LIF and feeder cells, the authors generated ES-cell likes cells from post-implantation epiblast tissue.

Nature. 2009 Oct 29;461(7268):1292-5.
Epigenetic reversion of post-implantation epiblast to pluripotent embryonic stem cells.
Bao S, Tang F, Li X, Hayashi K, Gillich A, Lao K, Surani MA.

Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QN, UK.

The pluripotent state, which is first established in the primitive ectoderm cells of blastocysts, is lost progressively and irreversibly during subsequent development. For example, development of post-implantation epiblast cells from primitive ectoderm involves significant transcriptional and epigenetic changes, including DNA methylation and X chromosome inactivation, which create a robust epigenetic barrier and prevent their reversion to a primitive-ectoderm-like state. Epiblast cells are refractory to leukaemia inhibitory factor (LIF)-STAT3 signalling, but they respond to activin/basic fibroblast growth factor to form self-renewing epiblast stem cells (EpiSCs), which exhibit essential properties of epiblast cells and that differ from embryonic stem (ES) cells derived from primitive ectoderm. Here we show reprogramming of advanced epiblast cells from embryonic day 5.5-7.5 mouse embryos with uniform expression of N-cadherin and inactive X chromosome to ES-cell-like cells (rESCs) in response to LIF-STAT3 signalling. Cultured epiblast cells overcome the epigenetic barrier progressively as they proceed with the erasure of key properties of epiblast cells, resulting in DNA demethylation, X reactivation and expression of E-cadherin. The accompanying changes in the transcriptome result in a loss of phenotypic and epigenetic memory of epiblast cells. Using this approach, we report reversion of established EpiSCs to rESCs. Moreover, unlike epiblast and EpiSCs, rESCs contribute to somatic tissues and germ cells in chimaeras. Further studies may reveal how signalling-induced epigenetic reprogramming may promote reacquisition of pluripotency.

PMID: 19816418

What people thought before this paper? Check the F1000 review by Anton.

Anton Wutz
Wellcome Trust Centre for Stem Cell Research, United Kingdom
Genomics & Genetics

New Finding
Tech Advance
This study is the first to show the derivation of embryonic stem cells (ESCs) from mouse post-implantation embryo epiblast.

Previously, mouse ESCs have been derived from blastocyst stage embryos. However, attempts to obtain such cells after implantation have been, thus far, unsuccessful. Instead, a distinct epiblast SC (EpiSC) has been derived from this developmental stage {1,2}. ESCs and EpiSCs are distinguished by their growth factor requirements, gene expression patterns, X inactivation status and their ability to contribute to chimaeric embryos and germline tissue when injected into blastocysts {1-3}.

Here, a procedure is described that involves the disaggregation of dissected epiblast tissue into single cells and prolonged culture under conditions favourable for maintaining ESCs, including with LIF (leukemia inhibitory factor) and other “feeders”. The resulting rESCs are shown to have properties of mouse ESCs, including reactivation of the inactive X chromosome and contribution to chimaeric embryos and germline tissue after injection into blastocysts. The study also shows that established EpiSC lines can be reprogrammed by culturing under ESC conditions. This is remarkable as EpiSCs depend critically on bFGF (basic fibroblast growth factor) and Activin for their growth and survival, and previously it was thought that a conversion to an ES-like cell would require genetic reprogramming {3}. It now appears that the two pluripotent states of ESCs and EpiSCs can be interconverted by solely adjusting culture parameters, and without exogenous factors. Importantly, this study carefully rules out the possibility that rESCs resemble pluripotent germ cells (EGCs), which have been established previously from primordial GCs isolated from mouse post-implantation embryos {4}. In contrast to EGCs, the newly derived rESCs show a somatic pattern of imprinted loci. Furthermore, the generation of rESCs from EpiSCs is consistent with the possibility of a conversion of the somatic epiblast to an earlier ESC-like fate.

References: {1} Brons et al. Nature 2007, 448:191-5 [PMID: 17597762]. {2} Tesar et al. Nature 2007, 448:196-9 [PMID: 17597760]. {3} Guo et al. Development 2009, 136:1063-9 [PMID: 19224983]. {4} Labosky et al. Development 1994, 120:3197-204 [PMID: 7720562].

“This observation is significant for human ES cells, which resemble mouse EpiSCs 3,4,26. Further studies may provide critical insights into signal-induced epigenetic reprogramming, including DNA demethylation and X-reactivation, which also represent a major barrier during reprogramming of somatic cells to iPS cells 9,20,27,28.”