Reactive oxygen species (ROS) have emerged as important signaling molecules in the regulation of various cellular processes. In our study, we investigated the effect of a wide range of ROS on Chinese hamster lung fibroblast (V79) cell proliferation. Treatment with H2O2 (100 microM), superoxide anion (generated by 1 mM xanthine and 1 mU/ml xanthine oxidase), menadione, and phenazine methosulfate increased the cell proliferation by approximately 50%. Moreover, a similar result was observed after partial inhibition of superoxide dismutase (SOD) and glutathione peroxidase. This upregulation of cell proliferation was suppressed by pretreatment with hydroxyl radical scavengers and iron chelating agents. In addition to ROS, treatment with exogenous catalase and SOD mimic (MnTMPyP) suppressed the normal cell proliferation. Short-term exposure of the cells to 100 microM H2O2 was sufficient to induce proliferation, which indicated that activation of the signaling pathway is important as an early event. Accordingly, we assessed the ability of H2O2 to activate mitogen-activated protein kinases (MAPK). Jun-N-terminal kinase (JNK) and p38 MAPK were both rapidly and transiently activated by 100 microM H2O2, with maximal activation 30 min after treatment. However, the activity of extracellular signal-regulated kinase (ERK) was not changed. Pretreatment with SB203580 and SB202190, specific inhibitors of p38 MAPK, reduced the cell proliferation induced by H2O2. The activation of both JNK and p38 MAPK was also suppressed by pretreatment with hydroxyl radical scavenger and iron chelating agents. Our results suggest that the trace metal-driven Fenton reaction is a central mechanism that underlies cell proliferation and MAPK activation.