Induction of phase 2 enzymes by serum oxidized polyamines through activation of Nrf2: effect of the polyamine metabolite acrolein

Abstract

The naturally occurring polycationic polyamines including putrescine, spermidine, and spermine play an important role in cell growth, differentiation, and gene expression. However, circulating polyamines are potential substrates for several oxidizing enzymes including copper-containing serum amine oxidase. These enzymes are capable of oxidizing serum polyamines to several toxic metabolites including aldehydes and H 2O 2. In this study, we investigated the effects of polyamines as inducers of phase 2 enzymes and other genes that promote cell survival in a cell culture system in the presence of bovine serum. Spermidine and spermine (50 μM) increased NAD(P)H quinone oxidoreductase (NQO1) activity up to 3-fold in murine keratinocyte PE cells. Transcript levels for glutathione S-transferase (GST) A1, GST M1, NQO1, γ-glutamylcysteine ligase regulatory subunit, and UDP-glucuronyltransferase 1A6 were significantly increased by spermidine and this effect was mediated through the antioxidant response element (ARE). The ARE from the mouse GST A1 promoter was activated about 9-fold by spermine and 5-fold by spermidine treatment, but could be inhibited by the amine oxidase inhibitor, aminoguanidine, suggesting that acrolein or hydrogen peroxide generated from polyamines by serum amine oxidase may be mediators for phase 2 enzyme induction. Elevations of ARE-luciferase expression and NQO1 enzyme activity by spermidine were not affected by catalase, while both were completely repressed by aldehyde dehydrogenase treatment. Direct addition of acrolein to PE cells induced multiple phase 2 genes and elevated nuclear levels of Nrf2, a transcription factor that binds to the ARE. Expression of mutant Nrf2 repressed the activation of the ARE-luciferase reporter by polyamines and acrolein. These results indicate that spermidine and spermine increase the expression of phase 2 genes in cells grown in culture through activation of the Nrf2–ARE pathway by generating the sulfhydryl reactive aldehyde, acrolein.