Hypoblast from human pluripotent stem cells regulates epiblast development [scRNA-seq]

Recently, several studies using cultures of human embryos together with single-cell RNA-seq (scRNA-seq) analyses have revealed differences between humans and mice, necessitating the study of human embryos. Despite the importance of human embryology, ethical and legal restrictions have limited post-implantation stage studies. Thus, recent efforts have focused on developing in vitro self-organising models using human stem cells. Here, we established a genetic approach to generate authentic hypoblast cells (nHyC) - known to give rise to one of the two extraembryonic tissues essential for embryonic development - from naïve human pluripotent stem cells (hPSCs). Our nHyCs spontaneously assemble with naïve hPSCs to form a three-dimensional bilaminar structure (bilaminoids) with a pro-amniotic-like cavity. In the presence of additional naïve hPSC-derived analogues of the second extraembryonic tissue, the trophectoderm, the efficiency of bilaminoid formation increases from 20% to 40%, and the epiblast within the bilaminoids continues to grow due to IL6 secreted by the trophectoderm. Furthermore, we show that bilaminoids robustly recapitulate the patterning of the anterior-posterior axis and the formation of cells reflecting the pre-gastrula stage, whose emergence can be shaped by genetically manipulating the DKK1/OTX2 hypoblast-like domain. We therefore model and reveal mechanisms by which the two extraembryonic tissues efficiently guide the stage-specific growth and progression of the epiblast as it establishes the post-implantation landmarks of human embryogenesis. Overall design: Analyzing transcriptional profiles in naïve PSCs, primed PSCs, PDGFRA positive cells induced by GATA6 o/e from naive and primed PSCs; induced by 7F, 4F, and 2F from naïve PSCs and bilaminoids at day 6, day9.