Simultaneous binding of quercetin and catechin to FOXO3 enhances IKKα transcription inhibition and suppression of oxidative stress-induced acute alcoholic liver injury in rats

Introduction: Oxidative stress is one of the major contributors to acute alcoholic liver injury (AALI), which is a common alcoholic liver disease. Quercetin and catechin are flavonoid antioxidants present in plant foods and possess chemopreventive and chemotherapeutic activities. Quercetin and catechin are often included in the same meal and ingested together. While they show cooperative actions against oxidative damage, the underlying mechanisms behind their counteracting effects against oxidative stress-induced AALI remain poorly understood.

Objectives: The aim of this study was to understand the mechanism underlying the enhanced antioxidant effect of quercetin-catechin combination to alleviate AALI in rats.

Methods: The ethanol (EtOH)-treated rats and H₂O₂-treated liver cells were used to demonstrate the enhanced antioxidant effect of quercetin and catechin. Then we used RNA-sequencing to compare quercetin alone, catechin alone and quercetin-catechin combination and then identified the critical role of IKKα combining with gene silencing and overexpression techniques. Its transcription factor, FOXO3 was found through yeast one-hybrid assay, luciferase reporter assay, EMSA and ChIP assay. Finally, the interaction between quercetin, catechin and FOXO3 was verified through molecular docking, UV-Vis absorption spectroscopy, fluorescence spectroscopy, and CD spectroscopy.

Results: The study demonstrated the enhanced antioxidant effect of a quercetin-catechin combination in EtOH-treated rats and in H₂O₂-treated liver cells. Quercetin and catechin cooperatively inhibited IKKα/p53 pathway and activated Nrf2 signaling pathway. IKKα was a critical negative regulator in their joint action. FOXO3 bound to IKKα promoter to regulate IKKα transcription. Quercetin and catechin influenced FOXO3-IKKα binding through attaching directly to FOXO3 at different sites and altering FOXO3's secondary structures.

Conclusion: Our study revealed the mechanism of quercetin and catechin against oxidative stress-induced AALI through jointly interacting with transcription factor. This research opens new vistas for examining the joint effect of therapeutics towards functional proteins and confirms the chemopreventive effects of multiple flavonoids via co-regulation.