Single cell analysis provides insight into haematopoietic differentiation of human embryonic stem cells

Abstract

One of the challenges in regenerative medicine remains generating functional haematopoietic stem cells (HSCs) of clinical quantity. Utilizing data from single cell RNA sequencing (scRNA-seq) of human embryonic stem cells (hESC) derived cells, I explored the differences that exist between in vitro and in vivo haematopoiesis, with the aim that understanding these differences could help improve future chances of generating HSCs in vitro. In the first part of this project, functional analysis into in vitro differentiation using an established haematopoietic differentiation protocol, confirmed that the protocol is myeloid and NK lineage biased compared to umbilical cord blood, with the population lineage dynamics changing over time. Using scRNA-seq, I investigated dynamic gene expression of cell populations emerging during hESC differentiation. I show that the haematogenic endothelium capable of generating the haematopoietic progenitors emerges as early as day 6 of differentiation. I also show that priming of the arterial endothelial cells to the haematogenic endothelium, and subsequent emergence of haematopoietic progenitors is associated with an increase in ribosomal and mitochondrial activity. Finally, a comparative analysis of the hESCs generated dataset with publicly available dataset of embryonic haematopoietic tissues revealed that hESC derived haematopoiesis, associated with increased mitochondrial gene activity, is likely mimicking events of the yolk sac haematopoiesis, rather than the haematopoiesis occurring in the AGM region. Based on the findings presented here, I hypothesize that increased mitochondrial activity, likely due to the normoxic culture conditions, may be contributing to the challenge in generating HSCs from hESCs in the dish.