ABSTRACT: Antagonism of ROS signaling can inhibit cell apoptosis and autophagy, thus favoring the maintenance and expansion of hematopoietic stem cells. Alpha lipoic acid (ALA), a small antioxidant molecule, affects cell apoptosis by lowering the ROS level. In this study, we show that ALA promoted production of human pluripotent stem cells (hPSCs) derived hemogenic endothelial cells and hematopoietic stem/progenitor cells in vitro. Transcriptome analysis of hPSCs derived hemogenic endothelial cells showed that ALA promoted endothelial-to-hematopoietic transition by up-regulating RUNX1, GFI1, GFI1B, MEIS2, and HIF1A and down-regulating SOX17, TGFB1, TGFB2, TGFB3, TGFBR1, and TGFBR2. ALA also up-regulated sensor genes of ROS signals, including HIF1A, FOXO1, FOXO3, ATM, PETEN, SIRT1, and SIRT3, during