Keap1 as Target of Genistein on Nrf2 Signaling Pathway Antagonizing Aβ induced Oxidative Damage of Cerebrovascular Endothelial Cells.

BACKGROUND β-amyloid peptides (Aβ) induced oxidative damage contributes to the pathogenesis of neurodegenerative diseases and cerebrovascular system is more vulnerable to oxidative stress. Our earlier study showed a clue that Genistein (Gen) might activate Nf-E2 related factor 2 (Nrf2) pathway to protect cerebrovascular cells from oxidative damage induced by Aβ, but the specific mechanisms and regulation targets are unclear. OBJECTIVE In this study, the anti-oxidative effects and the possible targets of Gen on regulating Nrf2 pathway in bEnd.3 cells were investigated. Cells were divided into control, Aβ25-35, Gen and Gen+Aβ25-35 groups. METHODS Cell viability, levels of malondialdehyde (MDA), Superoxide Dismutase (SOD) activity and nitrotyrosine were evaluated. Moreover, mRNA and/or protein expressions of Nrf2 and kelch-like ECH-associated protein 1 (Keap1) were measured. Then we transfected Keap1 over-expressed plasmid into bEnd.3 cells and measured the protein expressions of Nrf2 pathway related factors. Data showed that Gen could inhibit the over-production of MDA and nitrotyrosine and activate SOD activity. Besides we got the phenomenon that Gen could up-regulate the mRNA and protein expressions of Nrf2 and down-regulate Keap1 protein expression, the Keap1 over-expressed plasmid study indicated that the up-regulation of Nrf2 protein expression induced by pretreatment of Gen could be blocked by the transfection of Keap1 over-expressed plasmid, and the same results also occurred in Nrf2 downstream factors. CONCLUSION Gen could alleviate the cerebrovascular cells oxidative damage induced by Aβ25-35 through regulating Nrf2 pathway, and Keap1 might be one of the targets of Gen on activating Nrf2 pathway.