The emergence of human gastrulation upon in vitro attachment.

While studied extensively in model systems, human gastrulation remains obscure. The scarcity of fetal biological material as well as ethical considerations limit our understanding of this process. In vitro attachment of natural blastocysts shed light on aspects of the second week of human development in the absence of the morphological manifestation of gastrulation. Stem cell-derived blastocyst models, blastoids, provide the opportunity to reconstitute pre- to post-implantation development in vitro. Here we show that upon in vitro attachment, human blastoids self-organize a BRA+ population and undergo gastrulation. Single-cell RNA sequencing of these models replicates the transcriptomic signature of the human gastrula. Analysis of developmental timing reveals that in both blastoid models and natural human embryos, the onset of gastrulation as defined by molecular markers, can be traced to timescales equivalent to 12 days post fertilization. In all, natural human embryos and blastoid models self-organize primitive streak and mesoderm derivatives upon in vitro attachment.

In vitro attachment and symmetry breaking of a human embryo model assembled from primed embryonic stem cells.

Our knowledge of the molecular mechanisms surrounding human embryo implantation and gastrulation is lacking, largely due to technical and ethical limitations of experimenting with human embryos. Alternatives to human embryos have been reported, in which 3D clusters of embryonic stem cells are differentiated in a stepwise manner to model aspects of human embryogenesis. Yet it remains challenging to model the events past attachment. We propose a strategy of modeling the post-attachment human embryo by assembling a pre-formed polarized epithelial epiblast and extraembryonic cells, allowing them to self-organize into a structure that mimics the dish-attached human embryo. The model attaches in vitro and, in the absence of exogenous morphogens, breaks anteroposterior symmetry, giving rise to early gastrulation cell types. Our assembloid approach enables in a modular way to upgrade or exchange extraembryonic tissues to access more advanced stages of post-attachment development while complying with ethical policies.