Hericenone C exhibits anti-nociceptive effects through RORα-mediated suppression of TLR4 transcription

Introduction: Hericenone C exhibits antinociceptive effects in inflammatory pain; however, its molecular target and underlying mechanism remain unclear. Methods and results: We assessed the effect of hericenone C on formalin-induced nociceptive behavior in mice and explored its molecular target using in vitro experiments. Based on competitive affinity proteomics, we identified direct interactions between RORα and hericenone C; functional assays confirmed the role of hericenone C as a RORα antagonist that suppresses RORE-mediated transcriptional activity. Integrated bioinformatics and experimental validation indicated hericenone C-mediated suppression of TLR4 expression via inhibited RORα binding to the TLR4 promoter, which attenuates NF-κB signaling. This mechanism was further validated through pharmacological and genetic approaches, revealing that hericenone C and RORα antagonist SR3335 synergistically modulate TLR4 expression in RORα-modified macrophages. In the formalin-induced nociceptive pain model mice, formalin activated NF-κB through TLR4-dependent P65 phosphorylation, while macrophage depletion selectively suppressed phase 2 nociception. Critically, adoptive transfer of RORα-overexpressing or SR1078-pretreated monocyte-enriched PBMCs exacerbated pain, which was effectively reversed by hericenone C. Notably, hericenone C pretreatment reduced CD11c+ cell infiltration and decreased TLR4 expression in inflamed paw tissues. Conclusion: Overall, these findings establish hericenone C as a novel RORα antagonist that alleviates inflammatory pain through inhibition of the RORα-TLR4-NF-κB axis in CD11c+ cells, offering a promising therapeutic strategy for pain management.