Formative pluripotent stem cells show features of epiblast cells poised for gastrulation

The pluripotency of mammalian early and late epiblast could be recapitulated by naïve embryonic stem cells (ESCs) and primed epiblast stem cells (EpiSCs), respectively. However, these two states of pluripotency may not be sufficient to reflect the full complexity and developmental potency of the epiblast during mammalian early development. Here we report the establishment of self-renewing formative pluripotent stem cells (fPSCs) which manifest features of epiblast cells poised for gastrulation. fPSCs can be established from different mouse ESCs, pre-/early-gastrula epiblasts and induced PSCs. Similar to pre-/early-gastrula epiblasts, fPSCs show the transcriptomic features of formative pluripotency, which are distinct from naïve ESCs and primed EpiSCs. fPSCs show the unique epigenetic states of E6.5 epiblast, including the super-bivalency of a large set of developmental genes. Just like epiblast cells immediately before gastrulation, fPSCs can efficiently differentiate into three germ layers and primordial germ cells (PGCs) in vitro. Thus, fPSCs highlight the feasibility of using PSCs to explore the development of mammalian epiblast. Overall design: We established formative pluripotent stem cells (fPSCs) with long-term self-renewal capacities from mESCs, and mouse pre-gastrula epiblast cells, through mimicking the in vivo environment of postimplantation developing embryos combined with suppression of the Wnt/ß-catenin pathway. Then we performed transcriptome and epigenome analyses to investigate whether fPSCs represent a stable intermediate state between nai¨ve ESCs and primed EpiSCs.

哺乳动物早期及晚期上胚层的多能性，可分别由初始态胚胎干细胞（naïve embryonic stem cells, ESCs）与始发态上胚层干细胞（primed epiblast stem cells, EpiSCs）模拟。然而，这两种多能性状态或许不足以反映哺乳动物早期发育过程中上胚层的全部复杂性与发育潜能。本研究报道了可自我更新的形态发生态多能干细胞（formative pluripotent stem cells, fPSCs）的建立，该细胞系具备待原肠胚形成阶段上胚层细胞的特征。fPSCs可由多种小鼠ESCs、胚前/早期原肠胚上胚层以及诱导多能干细胞（induced PSCs）诱导建立。与胚前/早期原肠胚上胚层类似，fPSCs具有形态发生态多能性的转录组特征，这一特征与初始态ESCs及始发态EpiSCs显著不同。fPSCs还展现出E6.5时期上胚层特有的表观遗传状态，包括大量发育基因的超级二价结构。正如原肠胚形成前夕的上胚层细胞，fPSCs可在体外高效分化为三胚层与原始生殖细胞（primordial germ cells, PGCs）。因此，fPSCs为利用多能干细胞探索哺乳动物上胚层发育提供了可行途径。整体实验设计：本研究通过模拟植入后发育胚胎的体内环境，并联合抑制Wnt/β-连环蛋白（Wnt/β-catenin）通路，分别从小鼠ESCs以及小鼠胚前原肠胚上胚层中，成功建立具备长期自我更新能力的形态发生态多能干细胞（fPSCs）。随后通过转录组与表观基因组分析，探究fPSCs是否代表初始态ESCs与始发态EpiSCs之间的稳定中间状态。