Anti-Fas antibody- and actinomycin D (FA/AD) has been shown to have anti-tumor activity in some tumor cells. However, many of the molecular mechanism of FA/AD-induced apoptosis of human hepatoma Bel-7402 cells have not been fully clarified. In the present study, therefore, the effect of FA/AD in presence or absence of p38MAPK inhibitor SB203580 on the proliferation, apoptosis, p38MAPK, caspase-3, location of p38MAPK and caspase-3, and interaction between p38MAPK and caspase-3 in Bel-7402 cell was investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT), annexin V-FITC/propidium iodide (PI) double staining, electron microscopy, immunoblot, immunofluorescence and immunoprecipitation/immunoblot assay, respectively. We found that FA/AD significantly resulted in the inhibition of proliferation, induction of apoptosis, activation and up-regulation of p38MAPK, activation and up-regulation of caspase-3, translocation of p38MAPK and caspase-3 from cytosol to nucleus, and formation of p38MAPK/caspase-3 complex in Bel-7402 cells. In contrast, SB203580, a p38MAPK-specific inhibitor, apparently blocked induction of apoptosis, activation and up-regulation of p38MAPK, activation and up-regulation of caspase-3, and translocation of p38MAPK and caspase-3 from cytosol to nucleus in FA/AD-treated Bel-7402 cells. Taken together, we conclude that p38MAPK regulates caspase-3 by binding to caspase-3 in nucleus of Bel-7402 cells during FA/AD-induced apoptosis.