Abstract: AIM To investigate the non-genomic effect on inhibition of the proliferation of vascular smooth cells (VSMCs) via activation of iNOS and mitogen-activated protein kinase (p42/44MAPK) by 17β-estradiol. METHODSiNOS activation was determined by radioimmunoassay. Expressions of iNOS, p42/44 MAPK and p42/44 MAPK phosphorylation were measured by Western blot and expression of NO content was detected by colorimetric method. RESULTS In cultured VSMCs, activation of iNOS began to increase after pretreatment with estrogen for 5 min, and the maximum effect took place at 15 min and rapidly reverted thereafter. The effect of E2 on iNOS activation was not affected by actinomycin D but was obviously inhibited by tamoxifen. At the same time, NO production increased significantly after treatment with 17β-estradiol within 15 min. Compared with fetal calf serum, E2 pretreatment of VSMCs for 15 min decreased p42/44MAPK activity and this effect was blocked by L-NAME. CONCLUSION 17β-estradiol rapidly upregulates iNOS activation and NO production and downregulates p42/44 MAPK activation to inhibit VSMC proliferation through a non-genomic effect.