Vascular smooth muscle cells (VSMCs) proliferate in response to arterial injury. Recent findings suggest that, in addition to platelet-derived growth factors, growth factors from inflammatory cells and endothelial cells at the site of injury may contribute to VSMC proliferation. We hypothesized that a common mechanism by which endothelial cells and inflammatory cells stimulate VSMC growth could be the active oxygen species (i.e., O2, H2O2, and ·OH) generated during arterial injury. Using xanthine/xanthine oxidase to generate active oxygen species, we studied the effects of these agents on VSMC growth. Xanthine/xanthine oxidase (100 μM xanthine and 5 microunits/ml xanthine oxidase) stimulated DNA synthesis in growth-arrested VSMCs by 180% over untreated cells. Administration of the scavenging enzymes superoxide dismutase and catalase demonstrated that H2O2was primarily responsible for xanthine/xanthine oxidase-induced VSMC DNA synthesis. H2O2directly increased VSMC DNA synthesis and cell number (maximal at 200 μM) but decreased DNA synthesis of endothelial cells and fibroblasts. This effect was protein kinase C independent: sphingosine, a potent protein kinase C inhibitor, failed to block H2O2-induced VSMC DNA synthesis. H2O2(200 μM) stimulated c-mycand c-fosmRNA levels by fourfold and 20-fold, respectively, as compared with quiescent levels. In contrast to DNA synthesis, H2O2induction of c-mycand c-fosmRNA was primarily protein kinase C dependent. These findings show that H2O2specifically increases VSMC DNA synthesis and suggest a role for this oxidant in intimal proliferation, especially after arterial injury. (Circulation Research1992;70:593–599)