Self-organized blastoids from extended pluripotent stem (EPS) cells possess enormous potential for investigating postimplantation embryo development and related diseases. However, the limited ability of postimplantation development of EPS-blastoids hinders its further application. In this study, single-cell transcriptomic analysis indicated that the "trophectoderm (TE)-like structure" of EPS-blastoids was primarily composed of primitive endoderm (PrE)-related cells instead of TE-related cells. We further identified PrE-like cells in EPS cell culture that contribute to the blastoid formation with TE-like structure. Inhibition of PrE cell differentiation by inhibiting MEK signaling or knockout of Gata6 in EPS cells markedly suppressed EPS-blastoid formation. Furthermore, we demonstrated that blastocyst-like structures reconstituted by combining the EPS-derived bilineage embryo-like structure (BLES) with either tetraploid embryos or tetraploid TE cells could implant normally and develop into live fetuses. In summary, our study reveals that TE improvement is critical for constructing a functional embryo using stem cells in vitro.

中文翻译：

---

来自 EPS 细胞的双系胚胎样结构可以产生具有四倍体滋养外胚层的活小鼠。

来自扩展多能干 (EPS) 细胞的自组织母细胞具有研究植入后胚胎发育和相关疾病的巨大潜力。然而，EPS-母细胞的植入后发育能力有限阻碍了其进一步应用。在这项研究中，单细胞转录组学分析表明，EPS 母细胞的“滋养外胚层 (TE) 样结构”主要由原始内胚层 (PrE) 相关细胞而非 TE 相关细胞组成。我们进一步鉴定了 EPS 细胞培养物中的 PrE 样细胞，这些细胞有助于具有 TE 样结构的母细胞形成。通过抑制 MEK 信号传导或敲除 EPS 细胞中的 Gata6 来抑制 PrE 细胞分化，显着抑制了 EPS 母细胞的形成。此外，我们证明了通过将 EPS 衍生的双系胚胎样结构 (BLES) 与四倍体胚胎或四倍体 TE 细胞相结合而重建的囊胚样结构可以正常植入并发育成活胎儿。总之，我们的研究表明，TE 改善对于在体外使用干细胞构建功能性胚胎至关重要。