BACKGROUND It is believed that midkine (MK), a heparin-binding growth factor, plays an important role in carcinogenesis. However, the biologic mechanism of MK in hepatocellular carcinoma has not been clarified to date. The objective of the current study was to investigate the antiapoptotic role of MK in a human hepatoma cell line. METHODS The human hepatoma cell line HepG2 was used to study the antiapoptotic effect of MK. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/actinomycin D (ActD)-induced apoptosis was detected using a 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulphophenyl)-2H-tetrazolium monosodium salt (WST-8) assay, a caspase-3 activity assay, a caspase-8 activity assay, and flow cytometric analysis. RESULTS TRAIL had a potent, dose-dependent inductive effect on cell death in HepG2 cells, for which viable cell counts decreased to 6.3% of the control count at a TRAIL concentration of 100 ng/mL in the presence of 500 ng/mL ActD. Flow cytometry was used to demonstrate that apoptosis induced by TRAIL/ActD was in fact the cause of cell death. According to the WST-8 assay, MK pretreatment resulted in the suppression of TRAIL/ActD-mediated apoptosis in HepG2 cells, although cell viability did not increase when HepG2 cells were treated with MK alone. Caspase-3 activity was down-regulated when MK was added, but caspase-8 activity was high in both the absence and presence of MK. CONCLUSIONS The results of the current study indicate that MK acts as an antiapoptotic factor in HepG2 cells through the down-regulation of caspase-3 activity.