Retinoic acid regulates the development of human definitive hematopoiesis in a non-cell-autonomous manner

The developing embryo harbors multiple hematopoietic programs, categorized as either intra-embryonic or extra-embryonic yolk-sac, that differ in their spatio-temporal origins and developmental potential. In the vertebrate embryo, the hematopoietic stem cell derives from the definitive intra-embryonic hematopoietic program and is dependent on stage-specific retinoic acid (RA) signaling. We have recently modelled aspects of this developmental process in vitro using human pluripotent stem cells (hPSCs) and identified a KDR+CXCR4+ mesodermal population that generates definitive hematopoietic progeny in a uniquely RA-dependent manner. A subpopulation of this mesoderm expresses ALDH1A2, an enzyme involved in RA synthesis. Here, we sought to characterize the role of ALDH1A2 in the development of the human RA-dependent hematopoietic lineage and to map its mesodermal origin. Using two different engineered reporter hPSC lines, we show that specification of this lineage requires a functional ALDH1A2 enzyme at the mesoderm stage. Through functional analyses of different mesoderm subpopulations, we demonstrate that this RA-dependent lineage derives from ALDH1A2neg mesoderm by non-cell-autonomous RA signaling. Collectively, these studies provide new insight into the differentiation trajectory of hPSCs towards the definitive hematopoietic lineage.