Didymin alleviates neuropathic pain by targeting RKIP-mediated NF-κB/NLRP3 crosstalk to inhibit pyroptosis and neuroinflammation

ETHNOPHARMACOLOGICAL RELEVANCE: Mentha haplocalyx Briq. (Mentha) is recorded in the Pharmacopoeia of the People's Republic of China. It has the effect of relieving pain. Its main active component, Didymin, exhibits a variety of pharmacological activities, yet its mechanism of action in neuropathic pain remains not fully clarified. AIM OF THE STUDY: This study focuses on elucidating the analgesic efficacy of Didymin and delineating its underlying molecular pathways. METHODS: Network pharmacology and molecular docking were used to identify potential targets of Didymin in neuropathic pain. The analgesic effects were assessed in rats with chronic constriction injury (CCI) following Didymin treatment. Transcriptomic profiling (RNA-seq), in vitro microglial experiments, and pharmacological inhibition using Locostatin (an RKIP inhibitor) were performed to elucidate mechanistic pathways. RESULTS: Didymin significantly alleviated mechanical allodynia in CCI rats and upregulated mitophagy-related proteins (LC3, TOM20). It decreased NLRP3, ASC, GSDMD, and phosphorylated NF-κB levels, while enhancing RKIP expression. Molecular docking and dynamics confirmed stable binding between Didymin and RKIP. The protective effects were abolished by Locostatin, confirming RKIP dependence. CONCLUSION: Didymin ameliorates neuropathic pain by promoting RKIP expression and suppressing NF-κB/NLRP3-mediated pyroptosis and neuroinflammation. These findings bridge traditional Chinese medicine with molecular neuroscience, supporting Didymin as a promising candidate for pain management.