Homocysteine (Hcy) could induce apoptosis of vascular smooth muscle cells (VSMC). Asymmetric dimethylarginine (ADMA) has been thought as a novel risk factor for cardiovascular diseases. We hypothesized that ADMA mediates homocysteine-induced apoptosis of VSMC. In this experiment the level of ADMA in the medium measured by high-performance liquid chromatography (HPLC) was elevated when the apoptosis of T/G HA-VSMC was induced by Hcy which was detected by Hoechst33342 staining or flow cytometry (FCM) with Annecin V+Propidium Iodide (PI). Exogenous ADMA induced the apoptosis of VSMC. At the same time, ADMA elevated the level of intracellular reactive oxidative species (ROS) determined by fluorescent ROS detection kit. The activation of JNK and p38MAPK contributed to ADMA-induced apoptosis of VSMC. The present results suggest that endogenous ADMA is involved in apoptosis of VSMC induced by Hcy, and the effects of ADMA is related to elevation of intracellular ROS and activation of JNK/p38MAPK signaling pathways.