Abstract 5428: Effect of amentoflavone on Nrf2 signaling in HaCaT cells

Abstract

Abstract Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key mediator of cell response to oxidative stress and xenobiotics. Multiple lines of evidence suggest that Nrf2 activation protects against many diseases, aging, inflammation, and cancer. To date, Nrf2 activators from natural products have been widely studied. In this study we investigated the potential effect of Amentoflavone (AFN), a biflavonoid in many natural plants, on Nrf2 signaling in human keratinocytes cells (HaCaT). As results, AFN significantly increased ARE-luciferase activity and nuclear accumulation of Nrf2. Furthermore, a Nrf2 target protein, NQO-1, was significantly increased by AFN in dose- and time-dependent manner. To verify the Nrf2 signaling mechanism by AFN, firstly we measured reactive oxygen species (ROS) using DCFDA because mild intracellular ROS could stimulate Nrf2 activation. Surprisingly AFN triggered the production of ROS in 1 h and AFN-induced Nrf2 was inhibited by N-acetyl cysteine (NAC). Thus, we studied the ROS-related signaling on Nrf2 by measuring the activation of Akt and mitogen-activated protein (MAP) kinase family such as extracellular signal-regulated kinase (ERK) and p38. As results, pharmacology inhibitor of PI3K/Akt (LY294002) or p38 (SB 203580), but not ERK (U0126), abrogated AFN-activated Nrf2. Furthermore, silencing p38 using p38siRNA or inhibition using its pharmacology inhibitor decreased phosphorylation of Akt and attenuated Nrf2 activation. We also confirmed that AFN stabilized Nrf2 by inhibiting the ubiquitination. Taken together, AFN activates Nrf2 through ROS-mediated phosphorylation of p38 and followed by phosphorylation of Akt. Citation Format: Lilik Duwi Wahyudi, Jiwon Jeong, Heejung Yang, Jung-Hwan Kim. Effect of amentoflavone on Nrf2 signaling in HaCaT cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5428.