The anti-inflammatory and anti-oxidative effect of a classical hypnotic bromovalerylurea mediated by the activation of NRF2

Abstract

The Kelch-like ECH-associated protein 1-nuclear factor erythroid 2-related factor 2 (KEAP1-NRF2) system plays a central role in redox homeostasis and inflammation control. Oxidative stress or electrophilic compounds promote NRF2 stabilization and transcriptional activity by negatively regulating its inhibitor, KEAP1. We have previously reported that bromovalerylurea (BU), originally developed as a hypnotic, exerts anti-inflammatory effects in various inflammatory disease models. However, the molecular mechanism underlying its effect remains uncertain. Herein, we found that by real-time multicolor luciferase assay using stable luciferase red3 (SLR3) and green-emitting emerald luciferase (ELuc), BU potentiates NRF2-dependent transcription in the human hepatoblastoma cell line HepG2 cells, which lasted for more than 60 h. Further analysis revealed that BU promotes NRF2 accumulation and the transcription of its downstream cytoprotective genes in the HepG2 and the murine microglial cell line BV2. Keap1 knockdown did not further enhance NRF2 activity, suggesting that BU upregulates NRF2 by targeting KEAP1. Knockdown of Nfe2l2 in BV2 cells diminished the suppressive effects of BU on the production of pro-inflammatory mediators, like nitric oxide (NO) and its synthase NOS2, indicating the involvement of NRF2 in the anti-inflammatory effects of BU. These data collectively suggest that BU could be repurposed as a novel NRF2 activator to control inflammation and oxidative stress.

Keywords: Abbreviations: ARE, antioxidant responsive element; BU, bromovalerylurea; CCL2, C-C motif chemokine 2; DMF, dimethyl fumarate; GCLC, glutamate–cysteine ligase catalytic subunit; GCLM, glutamate–cysteine ligase modifier subunit; GSS, glutathione synthetase; GSH, glutathione; Hmox-1, heme oxygenase-1; IL-1β, interluekin-1β; IL-6, interluekin-6; JAK, Janus kinase; KEAP1, Kelch-like ECH-associated protein; NO, nitric oxide; NOS2, NO synthase 2; NRE, NF-κB responsive element; NQO-1, NAD(P)H quinone dehydrogenase; NRF2, nuclear factor erythroid 2-related factor 2/nuclear factor erythroid-derived 2-like 2; TXNRD, thioredoxin–disulfide reductase; ROS, reactive oxygen species; KEAP1–NRF2; anti-inflammation; anti-oxidant oxygen; bromovalerylurea; drug action toxins/drugs/xenobiotics.