Nrf2-Keap1 Antioxidant Defense and Cell Survival Signaling Are Upregulated by 17β-Estradiol in Homocysteine-Treated Dopaminergic SH-SY5Y Cells

<b><i>Background and Aims:</i></b> A recent neuroimaging study discovered the neurotoxicity effects of homocysteine (Hcy), which is only seen in elderly women. Estrogens exert a variety of actions on brain function that influence cognitive function, mood, and neuroprotection. The Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant defense pathway has been well-known to afford neuroprotection. Here, we first demonstrate the roles of Nrf2-Keap1 in 17β-estradiol (E₂) cytoprotection and Hcy toxicity and the protective mechanisms of E₂ on Hcy cytotoxicity in human dopaminergic SH-SY5Y cells. <b><i>Methods:</i></b> Cell viability was determined by trypan blue method. Protein expression was determined by Western blot analysis. Superoxide dismutase (SOD) activity was determined by ELISA. Reactive oxygen species (ROS) production was determined by flow cytometry. <b><i>Results:</i></b> In Hcy-treated SH-SY5Y cells, E₂ increased cell viability, attenuated ROS production, activated Akt signaling and inhibited glycogen synthase kinase-3β (GSK-3β), a kinase known to participate in neurodegeneration. Moreover, E₂ treatment led to Nrf2 dissociation from Keap1, the main negative regulator of Nrf2 activity in the cytoplasm, and increased the protein level of Nrf2 in the nucleus, with a significant increase in HO-1 expression and SOD activity in Hcy-treated cells. E₂-induced Nrf2 activation was attenuated by the PI3K inhibitor LY294002 and the estrogen receptor antagonist ICI 182,780. Further, E₂ decreased Hcy-induced apoptotic death by upregulating the antiapoptotic protein Bcl-2, decreasing cytochrome <i>c</i> release from mitochondria, and attenuating apoptotic cascade activation (Bax, caspase-9, and caspase-3). <b><i>Conclusion:</i></b> E₂ activates cell survival signaling and Nrf2-Keap1 antioxidant defense pathway and attenuates Hcy cytotoxicity.