The embryo-like structures (embryoids) constructed by aggregating embryonic stem cells (ESCs) and trophoblast stem cells (TSCs) have provided revolutionary tools for studying the intricate interaction between embryonic and extra-embryonic tissues during early embryonic development, which has been achieved in mice. However, due to the opposite dependence on some signalling pathways for in vitro culture of human ESCs (hESCs) and TSCs (hTSCs), particularly WNT and TGFβ signalling pathways, which limits the construction of human post-implantation embryoids by aggregating hESCs and hTSCs. To overcome this challenge, here, by screening 1639 chemicals, we found that an inhibitor of integrated stress response, ISRIB, can replace WNT agonists and TGFβ inhibitors to maintain the stemness and differentiation capacity of hTSCs. Thus, we developed an ISRIB-dependent in vitro culture medium for hTSCs, namely nTSM. Furthermore, we demonstrated that ISRIB could also maintain the hESC stemness. Using a 3D co-culture system (hESCs and hTSCs aggregate, ETA), we demonstrated that a 1:1 mixture of hESC culture medium (ESM) and nTSM improved the cell proliferation and organisation of both hESC- and hTSC-compartments and the lumenogenesis of hESC-compartment in ETAs. Overall, our study provided an ISRIB-dependent system for co-culturing hESCs and hTSCs, which facilitated the construction of human embryoids by aggregating hESCs and hTSCs.

中文翻译：

---

ISRIB 促进人类滋养层干细胞和胚胎干细胞的共培养

由胚胎干细胞（ESC）和滋养层干细胞（TSC）聚集构建的胚胎样结构（胚状体）为研究早期胚胎发育过程中胚胎和胚胎外组织之间复杂的相互作用提供了革命性的工具。老鼠。然而，由于人ESCs（hESCs）和TSCs（hTSCs）体外培养对某些信号通路的相反依赖性，特别是WNT和TGFβ信号通路，这限制了通过聚集hESCs和hTSCs构建人植入后胚状体。为了克服这一挑战，我们在这里通过筛选1639种化学物质，发现一种整合应激反应抑制剂ISRIB可以替代WNT激动剂和TGFβ抑制剂来维持hTSC的干性和分化能力。因此，我们开发了一种依赖ISRIB的hTSC体外培养基，即nTSM。此外，我们证明ISRIB还可以维持hESC的干性。使用 3D 共培养系统（hESC 和 hTSC 聚合体，ETA），我们证明 hESC 培养基 (ESM) 和 nTSM 的 1:1 混合物改善了 hESC 和 hTSC 区室的细胞增殖和组织以及腔生成ETA 中的 hESC 区室。总体而言，我们的研究提供了一种依赖于 ISRIB 的 hESC 和 hTSC 共培养系统，该系统通过聚集 hESC 和 hTSC 促进了人类胚状体的构建。