Rat post-implantation epiblast-derived pluripotent stem cells produce functional germ cells

Kenyu Iwatsuki; Mami Oikawa; Hisato Kobayashi; Christopher A Penfold; Makoto Sanbo; Takuya Yamamoto; Shinichi Hochi; Kazuki Kurimoto; Masumi Hirabayashi; Toshihiro Kobayashi

doi:10.1016/j.crmeth.2023.100542

Rat post-implantation epiblast-derived pluripotent stem cells produce functional germ cells

Cell Rep Methods. 2023 Jul 27;3(8):100542. doi: 10.1016/j.crmeth.2023.100542. eCollection 2023 Aug 28.

Authors

Kenyu Iwatsuki^{1

2}, Mami Oikawa^{1

3}, Hisato Kobayashi⁴, Christopher A Penfold^{5

6

7}, Makoto Sanbo⁸, Takuya Yamamoto^{9

10

11}, Shinichi Hochi^{2

12}, Kazuki Kurimoto⁴, Masumi Hirabayashi^{8

13}, Toshihiro Kobayashi^{1

8}

Affiliations

¹ Division of Mammalian Embryology, Center for Stem Cell Biology and Regenerative Medicine, The Institute of Medical Science, The University of Tokyo, Tokyo 108-8639, Japan.
² Graduate School of Medicine, Science and Technology, Shinshu University, Nagano 386-8567, Japan.
³ Laboratory of Regenerative Medicine, Tokyo University of Pharmacy and Life Sciences, Tokyo 192-0392, Japan.
⁴ Department of Embryology, Nara Medical University, Nara 634-0813, Japan.
⁵ Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Site, Cambridge CB2 3EG, UK.
⁶ Centre for Trophoblast Research, University of Cambridge, Downing Site, Cambridge CB2 3EG, UK.
⁷ Wellcome Trust - Cancer Research UK Gurdon Institute, Henry Wellcome Building of Cancer and Developmental Biology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.
⁸ Center for Genetic Analysis of Behavior, National Institute for Physiological Sciences, Aichi 444-8787, Japan.
⁹ Center for iPS Cell Research and Application, Kyoto University, Kyoto 606-8507, Japan.
¹⁰ Institute for the Advanced Study of Human Biology (ASHBi), Kyoto University, Kyoto 606-8501, Japan.
¹¹ Medical-risk Avoidance Based on iPS Cells Team, RIKEN Center for Advanced Intelligence Project, Kyoto 606-8501, Japan.
¹² Faculty of Textile Science and Technology, Shinshu University, Nagano 386-8567, Japan.
¹³ The Graduate University of Advanced Studies, Aichi 444-8787, Japan.

Abstract

In mammals, pluripotent cells transit through a continuum of distinct molecular and functional states en route to initiating lineage specification. Capturing pluripotent stem cells (PSCs) mirroring in vivo pluripotent states provides accessible in vitro models to study the pluripotency program and mechanisms underlying lineage restriction. Here, we develop optimal culture conditions to derive and propagate post-implantation epiblast-derived PSCs (EpiSCs) in rats, a valuable model for biomedical research. We show that rat EpiSCs (rEpiSCs) can be reset toward the naive pluripotent state with exogenous Klf4, albeit not with the other five candidate genes (Nanog, Klf2, Esrrb, Tfcp2l1, and Tbx3) effective in mice. Finally, we demonstrate that rat EpiSCs retain competency to produce authentic primordial germ cell-like cells that undergo functional gametogenesis leading to the birth of viable offspring. Our findings in the rat model uncover principles underpinning pluripotency and germline competency across species.

Keywords: Klf4; epiblast stem cell; germline competency; pluripotency; primordial germ cell-like cell; rat; reprogramming.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Biomedical Research*
Embryo Implantation
Germ Cells
Germ Layers
Kruppel-Like Transcription Factors
Mammals
Mice
Pluripotent Stem Cells*
Rats

Substances

Klf2 protein, rat
Kruppel-Like Transcription Factors