Females live longer than males. The higher levels of oestrogens in females protect them against ageing, by up-regulating the expression of antioxidant, longevity-related genes (Borras et al. 2003). Oestradiol and genistein share chemical properties which confer antioxidant features to these compounds. The low concentration of them make it unlikely that they exhibit significant antioxidant capacity in the organism. The aim of this study is to elucidate the mechanism through which oestrogens and phytoestrogens up-regulate the expression of antioxidant enzymes. We have used a mammary gland tumour cell line(MCF-7). Phosphorylation of mitogen activated protein kinase (MAPK) was determined by western blotting,translocation of p50 subunit of nuclear factor κ B(NFκB) to the nucleus in nuclear extracts by ELISA,expression of antioxidant enzymes by quantitative real-time RT-PCR,and peroxide levels was measured fluorimetrically (Barja et al. 1999). Our results show that physiological concentrations of oestrogens (0.02 nM) and concentrations of genistein equivalent to those present in blood of oriental people (0.5 μM), activate the MAPK pathway. Oestrogens (n=6) and phytoestrogens (n=4) increase ERK1 and ERK2 phosphorylation when compared with controls (43.1±0.3 vs 41.2±1.7 arbitrary units, p<0.01 and 39.0±1.0 vs 37.6±0.3 arbitrary units, p<0.05, unpaired Student's t test, respectively). All the results are means±S.E.M. These, in turn, increase p50 subunit of NFκB in nuclear extracts from cells treated with oestrogens or phytoestrogens. The Mn-SOD and the GPx promoter region contain putative NFκB-binding motifs. Thus, activation of NFκB by oestrogens subsequently activates the expression of Mn-SOD(3.5-fold, n=3, p<0.001, ANOVA-Tukey test) and GPx (2.2-fold, n=3, p<0.001, ANOVA-Tukey test), but genistein is only capable of activating Mn-SOD expression (2.9-fold, n=3, p<0.01, ANOVA-Tukey test). This could be due to the fact that genistein binds preferently to oestrogen receptor β. This antioxidant protection is reflected in the lower peroxide levels we find in cells treated with oestrogens or phytoestrogens when compared with controls. The activation of signalling pathways and up-regulation of antioxidant enzyme expression is prevented when cells are co-treated with either oestrogens or phytoestrogens and the MAPK phosphorylation inhibitor UO126 (1 μM), thus reflecting the role of this signalling pathway in the antioxidant action of oestrogens and phytoestrogens. We conclude that oestrogens and phytoestrogens up-regulate expression of antioxidant enzymes via the MAPK activation, which in turn activate the NFkB signalling pathway.