Hemangiopoietin (HAPO) is a growth factor that significantly stimulates proliferation and survival of the primitive cells of hematopoietic and endothelial lineages. To determine the mechanism of action of HAPO, the anti-apoptotic activity and signal transduction pathway of HAPO were investigated using a factor-dependent leukemia cell line, the MO7e cells. Recombinant human HAPO (rhHAPO) was produced in Escherichia coli and purified by a series of column chromatography with a purity of more than 95%. rhHAPO significantly supported the survival of MO7e cells after deprivation of granulocyte-macrophage colony stimulating factor and activated phosphatidylinositol 3-kinase (P13K). When the MO7e cells were treated with two specific inhibitors to P13K (LY294002 or wortmannin), a significant loss of cell viability with evidence of apoptosis was observed. Moreover, the protein kinase B (Akt), one of the downstream effectors of P13K-dependent survival signaling, was activated in HAPO-stimulated MO7e cells. Phosphorylation of Akt at serine 473 and its downstream molecular Bad at serine 136 was induced by HAPO, but was blocked by two P13K inhibitors, LY294002 and wortmannin. In addition, HAPO inhibited caspase-3 activities and poly(ADP-ribose) polymerase degradation. Such an effect of HAPO was also significantly blocked by either LY294002 or wortmannin. These results indicate that HAPO protects MO7e cells from apoptotic death through a P13K-Akt pathway. (C) 2004 Elsevier Inc. All rights reserved.