Establishment of Mouse Primed Stem Cells by Combination of Activin and LIF Signaling.

Mengyi Wei,Li Zheng,Baojiang Wu,Siqin Bao,Chen Chen,Xihe Li,Chaoyue Zhao,Yanglin Chen

doi:10.3389/fcell.2021.713503

Abstract

In mice, embryonic stem cells (ESCs) and epiblast stem cells (EpiSCs) are established from pre- and post-implantation embryos and represent the naive and primed state, respectively. Herein we used mouse leukemia inhibitory factor (LIF), which supports ESCs self-renewal and Activin A (Act A), which is the main factor in maintaining EpiSCs in post-implantation epiblast cultures, to derive a primed stem cell line named ALSCs. Like EpiSCs, ALSCs express key pluripotent genes Oct4, Sox2, and Nanog; one X chromosome was inactivated; and the cells failed to contribute to chimera formation in vivo. Notably, compared to EpiSCs, ALSCs efficiently reversed to ESCs (rESCs) on activation of Wnt signaling. Moreover, we also discovered that culturing EpiSCs in AL medium for several passages favored Wnt signaling-driven naive pluripotency. Our results show that ALSCs is a primed state stem cell and represents a simple model to study the control of pluripotency fate and conversion from the primed to the naive state.

Highlights

Embryonic stem cells (ESCs) are known for their potential of self-renewal and differentiating into different embryonic tissues (Evans and Kaufman, 1981)
We did not observe any distinctive morphological change when AFSCs were placed into AL medium for more than 10 passages (Supplementary Figure 1B), and afALSCs maintained the characteristics of pluripotency by AP staining and immunofluorescence of NANOG and OCT4 (Supplementary Figures 1B,C)
EpiALSC colonies developed to become GOF/GFP+ when cultured in CL or 2iL for 4 days (Figure 4B), the transition rate was lower than for ALSCs (Table 4). These results suggested that the successive conversion from epiblast stem cells (EpiSCs) to reversed to ESCs (rESCs) was accomplished in two stages, with the first step involving the replacement of bFGF in AF medium to leukemia inhibitory factor (LIF) and the second step involving the use of CHIR instead of Activin A (Act A)

Summary

Introduction

Embryonic stem cells (ESCs) are known for their potential of self-renewal and differentiating into different embryonic tissues (Evans and Kaufman, 1981). Yu S. et al (2020) proposed that BMP4 plays an essential role in primed-to-naive transition (PNT) by opening up chromatin loci to activate critical regulators of PNT These reports demonstrate that the state of pluripotent stem cells can be reversed to some extent by factors in their culture conditions. Two recent studies have suggested that there is a “formative” state in ESCs which is between the “naive” and “primed” state and presents formative features of human stem cells and horse stem cells (Kinoshita et al, 2020; Yu L. et al, 2020) These two studies describe different culture systems. All intermediate stem cells, including the formative stem cells (FS cells), XPSCs and rosette-like stem cells (RSCs), encode a higher pluripotent gene expression than EpiSCs and contribute to chimera formation; different culture conditions and the unique properties of stem cells still require further exploration (Kinoshita et al, 2020; Neagu et al, 2020; Yu L. et al, 2020)

Methods

Results

Conclusion