An in vitro model of hemogenic endothelium commitment and hematopoietic production.

Abstract

Adult-type hematopoietic stem and progenitor cells are formed during ontogeny from a specialized subset of endothelium, named as hemogenic endothelium, via an endothelial-to-hematopoietic transition (EHT) that occurs in the embryonic aorta and the associated arteries. Despite efforts in generating models, little is known about the mechanisms driving endothelial cells to the hemogenic fate and about the subsequent molecular control of the EHT. Here we have designed a stromal line-free controlled culture system utilizing the embryonic pre-somitic mesoderm to obtain large numbers of endothelial cells that subsequently commit into hemogenic endothelium before undergoing EHT. Follow up of the culture for up to twelve days using key molecular markers reveals stepwise commitment into the blood-forming system that is reminiscent of the cellular and molecular changes occurring during hematopoietic development at the level of the aorta. Long-term, single cell imaging, allows tracking the EHT of newly formed blood cells from the layer of hemogenic endothelial cells. By modifying the culture conditions, it is also possible to modulate the endothelial cell commitment or the EHT or to produce smooth muscle cells at the expense of endothelial cells thereby demonstrating the versatility of the cell culture system. This method will improve our understanding of the intimate cellular changes associated with hemogenic endothelium commitment and EHT and, by unfolding these earliest steps of the hematopoietic program, will pave the way for future ex vivo production of blood cells.