The aim of this study was to investigate the contribution of mitochondrial dysfunction to chemoresistance and migration of hepatoma cells. We found that inhibition of mitochondrial respiration and mitochondrial DNA (mtDNA) depletion resulted in induction of amphiregulin (AR) expression in HepG2 cells. Upon oligomycin treatment of HepG2 cells, the cytosolic Ca(2+) was significantly raised after 30 min, and the intracellular level of reactive oxygen species (ROS) was elevated 2.2-fold after 4 h. Moreover, the condition medium of oligomycin-treated HepG2 cells was found to stimulate the migration of SK-Hep-1 cells. On the other hand, oligomycin-induced cisplatin-resistance and cell migration of HepG2 cells were attenuated by AR-specific RNA interference (#L-017435, Dharmacon) and a neutralizing antibody (MAB262, R&D Systems), respectively. Together, these findings suggest that mitochondrial dysfunction induced Ca(2+) mobilization, and ROS overproduction, which modulated the chemo-resistance and migration of hepatoma cells through the induction and activation of AR.