Aim To investigate the effects of gambogic acid(GA)on rat vascular smooth muscle cell(VSMC) proliferation and migration stimulated by epithelial growth factor(EGF) and its underlying molecular mechanism. Methods The inhibitory effect of GA at various concentration on the proliferation induced by EGF was measured by using CCK8 assay and 3H-thymidine incorporation.The effects of GA on the cell cycle progression stimulated by EGF were analyzed by flow cytometry.The effect of GA EGF receptor(EGFR) and pi-Tyrosine was measured by Western blotting.The capacity of GA binding with EGF was also measured. Results GA inhibited EGF-induced 3H-thymidine incorporation into DNA and proliferation on VSMC.The cell cycle progression was blocked significantly in the GA pretreated cells.GA significantly inhibited expression of pi-EGFR,pi-Tyrosine stimulated by EGF compared with the control group without GA.Dot binding assay shown that a negative binding signal was detected on the GA and BSA spot incubated with EGF. Conclusions These observations show that inhibition effects on VSMC proliferation of GA are mediated by the inhibition of EGFR tyrosine phosphrylation.The inhibitory effects of GA are due to directly tyrosine phosphorylation inhibition,rather than binding with EGF.