Inflammatory responses in the blood vessel play a pivotal role in the pathogenesis of atherosclerosis. Eupatilin, a flavone derived from Artemisia princepsPampanini, has various pharmacological activities, including antioxidant, anti-tumor, and anti-inflammatory capacities. However, there has been no research examining the function of eupatilin on vascular inflammation. Therefore, the aim of this study was to investigate the effects of eupatilin on tumor necrosis factor (TNF)-α-induced human umbilical vein endothelial cells (HUVECs) activation and the underlying molecular mechanisms. Our findings showed that eupatilin reduced U937 cells adhesion to TNF-α-stimulated HUVECs and attenuated TNF-α-induced the expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in HUVECs, as well as the production of intracellular reactive oxygen species (ROS). Moreover, eupatilininhibits TNF-α-induced phosphorylation of NF-kB p65 and MAPKs in HUVECs. Taken together, the results of the present study suggest that eupatilin inhibited inflammatory reaction through suppressing the ROS/MAPK-NF-ĸB pathway in HUVECs. Thus, eupatilin is proposed as an effective new anti-inflammatory agent to suppress vascular inflammation, and further prevent atherosclerosis.