Effects of FGF2 and oxygen in the BMP4-driven differentiation of trophoblast from human embryonic stem cells

Human embryonic stem cells (hESC) differentiate into trophoblast when treated with BMP4. Here we studied the effects of either low (4% O₂, L) or atmospheric O₂ (20% O₂, A) in the presence and absence of FGF2 on H1 hESC cultured in the presence of BMP4. Differentiation progressed from the periphery toward the center of colonies. It occurred most quickly in the absence of FGF2 and under A and was slowest in the presence of FGF2 and under L. Chorionic gonadotropin (CG) production required A, while FGF2 suppressed progesterone synthesis under both A and L. FGF2 was then omitted while we examined the trophoblast markers SSEA-1 and cytokeratin-7 and-8, whose expression also progressed inward from the periphery of colonies and occurred more rapidly under A than under L. By day 5, most cells outside central islands of Oct4-positive cells were positive for these antigens under both conditions and many also expressed HLA-G, a marker of extravillous cytotrophoblast. Under A, but not L, CG and CGβ became prominent in GATA2-positive, peripherally located, multinucleated cells. In conclusion, BMP4 induced conversion of hESC exclusively toward trophoblast; FGF2 slowed differentiation, while O₂ accelerated this process and promoted syncytiotrophoblast formation.