Spontaneous induction of neural crest cells in human pluripotent stem cell-derived aggregates revealed by single-cell RNA sequencing

Neural crest cells (NCCs) are a transient population which contributes to the majority of craniofacial tissue. The development of craniofacial tissue involves highly complex intermingled processes of cell fate specification and migration in densely packed mesenchyme. The establishment of appropriate in vitro models would be helpful for understanding these complicated processes. Thus, we developed a new method to induce NCCs in aggregates of human pluripotent stem cells (hPSCs). To gain insight into the process of aggregate development we performed single cell RNA sequencing (scRNA-seq) on day 2, 3, 4, and 5. scRNA-seq analysis revealed that hPSCs in the aggregates acquired neural plate border NPB-like identity before NCC induction. scRNA-seq also revealed that NCCs showed changes in the gene expression associated with epithelial mesenchymal transition as in vivo developmental process. Single-cell RNA sequencing was performed on human pluripotent stem cell-derived aggregates at differentiation day 2, 3, 4, and 5.