scRNA-seq of hPSC-derived human intestinal organoids with endogenous vascular endothelial cells

The development of human pluripotent stem cell (hPSC)- derived small intestinal organoids (HIOs) has led to an improved understanding of human intestinal development and physiology. HIOs generated using directed differentiation lack some cellular populations found in the native organ, including vasculature. We performed single cell RNA sequencing (scRNA-seq) on approximately 13,000 cells at various timepoints (0, 3, 7, and 14 days) across HIO in vitro development and observed a transient population of endothelial-like cells (ECs) present within HIOs early during differentiation. Our data demonstrate that EC-like cells fail to be robustly maintained in long term culture. Here, we have developed a new directed differentiation approach to enhance co-differentiation and maintenance of ECs within HIOs, leading to the development of vascularized HIOs (vHIOs). scRNAseq was used to compare vHIOs to control HIOs after 59d months in culture.