Human pluripotent stem cells (hPSCs) are an alternative model for recapitulating key milestones of human embryogenesis during tissue development, which is an important and highly regulated process, yet poorly understood1. Currently, creating an in vitro model that provides the appropriate microenvironmental cues in a spatiotemporal manner to enable hPSCs to differentiate into complex 3D tissue structures remains a challenge2,3.
We have developed a novel three-dimensional in vitro system that has the ability to maintain and form complex tissue structures with recognisable features of various human embryonic tissues from the three germ layers by combining size controlled human embryoid bodies (hEBs) with subsequent maintenance and differentiation on a porous scaffold.
This approach extends culture time by flattening EB shape, resulting in reduced diffusion distances and increased cell viability, permitting longer term cultures, increased cell differentiation and the formation of high order structures. Current work focuses on directing the differentiation of hEB structures towards selected germ layers to study specific developmental pathways. Future work will involve the introduction of perfusion, another key microenvironmental parameter, in order to increase physiological relevance and promote even more complex tissue differentiation. Finally, human induced pluripotent stem cell populations will be used to fully validate and explore the capabilities of the model.
We hypothesize that through the development and application of new in vitro technologies, it is feasible to direct the differentiation and extend the growth/maturation of tissues derived from hPSCs. We anticipate that this novel approach will provide an opportunity to study early stages of tissue development and deliver a controllable and reproducible animal-free alternative to assess the pluripotency of hPSCs.
1.Zhu, Z., & Huangfu, D. Development, 140(4), 705-717 (2013).
2.Srivastava, P., & Kilian, K. A, Frontiers in Bioengineering and Biotechnology, 7, 357 (2019).
3.Levenberg, S., et al, Proceedings of the National Academy of Sciences, 100(22), 12741-12746 (2003).