Role of Notch Signaling in Human Pluripotent Stem Cell-derived Hematopoiesis

Hematopoietic development of pluripotent stem cells (PSCs) begins with the emergence of hemogenic endothelial (HE) precursors, followed by endothelial-to-hematopoietic transition (EHT). To test the effect of Notch signaling on HE specification, PSCs were differentiated on OP9-control or OP9-DL1 cells expressing the Notch ligand Delta-like 1 (Dll1), and then induced to become T-cells. There was high variability in T-cell generation from many experimental repeats. As a result, we tested different serum lots and also administered anti-TGF-β treatments during co-culture. However, the inconsistency of T-cell generation using this protocol remained unaffected. Next, we engineered OP9 cells to ectopically express human JAG1 (OP9-JAG1), and evaluated their ability to promote EHT by culturing HE cells on OP9-JAG1 or OP9-DL4 cells. Of note, cells undergoing EHT in the presence of JAG1 showed better hematopoietic conversion than those on OP9-DL4. Collectively, these data suggest a preferential role for JAG1 during human EHT in vitro.M.Sc