Controlled activin A signaling enables liver organoid generation from human pluripotent stem cells

Morphogen gradients critically regulate the coordinated emergence of endodermal and mesodermal lineages during liver bud formation, yet current human hepatic organoid models inadequately replicate these spatial cues. Here, we examine how modulation of activin A signaling within a 3D vertical-wheel bioreactor culture system shapes early morphogenetic organization in human pluripotent stem cell aggregates. We observe that defined activin A conditions promote the spatial organization of human pluripotent stem cells during hepatic differentiation. High activin levels induce uniform definitive endoderm and enhanced hepatic competence, whereas low levels support the emergence of epithelial tube-like structures encased by mesenchymal cells, recapitulating early liver bud-like architecture. By leveraging differential activin A concentrations, this approach modulates the balance between morphogenesis and hepatic maturation in vertical-wheel-derived liver organoids, providing a scalable platform to model human liver organogenesis and engineer ventral foregut-derived tissues.