Aim To investigate the efffect of breviscapine on the proliferation, cell ultrastructure, cell cycle and nuclear factor-κB (NF-κB) activity of rabbit vascular smooth muscle cell (VSMC) in order to find out the mechanism of its action. Methods By techniques of cell culture in vitro, VSMC stimulated by fetal bovine serum were treated by breviscapine in different concentration for a certain period, then the inhibitory ratio of cell proliferation was measured by MTT, cell cycle distribution and NF-κB activity of the cell were measured by Flow-Cytometry respectively, and the cell ultrastructure was observed by transmission electron microscopy (TEM). Results Breviscapine could obviously inhibit the proliferation of VSMC stimulated by fetal bovine serum (IC50 value was 17.9 mg/L), and change the cell ultrastructure, which included that electron density increased, a part of cytomembrane lysised, endoplasmic reticulum and Golgi apparatus showed vacuolar degeneration, some mitochondrion showed coagulation and mitochondrial crista disappeared or showed coagulation. Breviscapine could also decrease S phase cells, increase G 0/G 1 phase cells and down-regulate NF-κB activity of VSMC by concentration-dependent fashion, and the results were significant compared with the control (p<0.01). Conclusion Breviscapine could obviously inhibit the proliferation of VSMC and may prevent atherosclerosis, which mechanism may be realized partly by regulating NF-κB activity of VSMC.