Creation of organoid systems to model human endodermal organ development in vitro
Organoid systems have revolutionised the study of adult stem cells and organ homeostasis. However, there have been limited attempts to use the technology to study human organ development. The work of this thesis established two novel hepatic organoid systems; one based upon the archetypal liver stem cell, the hepatoblast, whilst the other was based upon a hitherto undescribed biliary progenitor cell. The work continued with exploration of the development of the human intestine, characterising how intestinal organoids develop in vitro, and going on to assess human intestinal maturation in vivo at single cell resolution. First, the developing human liver was characterised during the first trimester and the cellular composition at this crucial developmental timepoint was investigated at single cell resolution. The hepatoblast organoids (HBO) were then demonstrated to retain their hepatoblast profile in culture and demonstrated the functional capacity of this fetal cell-type. Exploiting this new model, the molecular processes of differentiation were then explored as the HBO were demonstrated to have bipotential capacity to generate hepatocytes and cholangiocytes. Furthermore, these organoids were then transplanted into mice and were able to differentiate into complex functional tissues. Next, a newly discovered stem cell was characterised and termed the ‘biliary progenitor cell’. This new cell type was able to generate three-dimensional organoids, fetal biliary organoids (FBO), that could be transplanted into mice and provide insight into biliary development. Next induced pluripotent stem cell (iPSC)-derived and primary fetal intestinal derived organoids were characterised against purified primary intestinal epithelial tissue, demonstrating a dynamic transcriptome of fetal organoids in vitro and a lack of tissue specificity of iPSC-derived organoids. The fetal and paediatric intestine were assessed at single cell level to assess the cell types and developmental pathways involved in the process of development, and this information was then utilised to try to model development in vitro using small molecule stimulation of fetal intestinal organoids. The work described in this thesis develops our understanding of human endodermal organ development. The newly described organoid models present an exciting opportunity to model human liver development, generate cells for drug-screening, and offer the potential for cell-based therapies, whilst the exploration of the human intestine during development may aide our understanding of how such pathways may be abrogated in developmental defects, or reverted to during disease states.