HOXA9 promotes specification rather than survival or proliferation of HEPs. (A) Apoptosis and cell cycle analysis on empty vector (EV)- or HOXA9-hESC–derived HEPs. Similar numbers of dead cells (Annexin V⁺/7AAD⁺), apoptosis-undergoing cells (Annexin V⁺), and cycling cells (S, G2, and M) were found in EV-HEPs versus HOXA9-HEPs. (B) HEPs were purified at day 10 of differentiation and cocultured with OP9 for 4 days. HOXA9-HEPs differentiated faster toward CD34⁺CD45⁺, and CD45⁺ hematopoietic cells. Data represent mean ± standard error of the mean (SEM) for 4 independent experiments. (C) Gene expression kinetics of the mesendodermal transcription factors Brachyury and MixL1 during EV- and HOXA9-EB hematopoietic differentiation. Data represent mean ± SEM for 2 independent experiments. SSC, side scatter.