Fuyuan Huoxue decoction ameliorates CFA-induced pain by modulating chemokine and IL-17 signaling pathways

Ethnopharmacology relevance: Fuyuan Huoxue Decoction (FYHX), originally documented in Yixue Faming, is traditional Chinese medicine used to promote blood circulation, remove blood stasis, and soothe liver meridians. It is mainly prescribed for traumatic injuries and blood stasis-related syndromes. In modern clinical practice, FYHX has been widely applied in the treatment of soft tissue injuries, neuropathic pain, and chondritis. However, its bioactive constituents and the multi-target mechanisms underlying its analgesic effects remain largely unclear.

Aim of the study: This study aimed to evaluate the analgesic effects of FYHX, identify its active small-molecule constituents, and elucidate the molecular mechanisms involved using a CFA-induced pain model.

Materials and methods: CFA was injected into the hind paw of rats. Pain behaviors were examined using von-Frey filaments and the Hargreaves' test. High-performance liquid chromatography-mass spectrometry (HPLC-MS) was used to identify the circulating bioactive components and metabolites of FYHX. Network pharmacology, quantitative transcriptomics and molecular docking were integrated to explore potential mechanisms in the dorsal root ganglia (DRG) and spinal cord. Key molecular targets were validated using Quantitative Real-Time PCR (qPCR) and Western blotting.

Results: FYHX significantly alleviated CFA-induced mechanical and thermal pain behaviors. A total of 42 molecules and metabolites were identified as the circulating components of FYHX. Integrated analysis revealed that FYHX primarily modulates the chemokine signaling pathway in the DRG and the IL-17 signaling pathway in the spinal cord. Validation experiments confirmed that downregulation of critical molecules in these pathways was associated with the analgesic of FYHX. Furthermore, four compounds-Scutellarein, 3,4'-Dibydroxyflavone, 5,6,7-Trimethoxyflavone and Armillarisin A-were shown to significantly reduce pain behaviors in the CFA model.

Conclusion: FYHX effectively alleviates CFA-induced chronic pain, potentially through the suppression of chemokine and IL-17 signaling pathways in the DRG and spinal cord, respectively. The identified bioactive compounds may contribute to its analgesic effects. Overall, FYHX and its active ingredients might be further developed to be new analgesic treatment for chronic pain.

Chemokine; Chronic pain; Dorsal root ganglion; Fuyuan Huoxue decoction; IL-17; Spinal cord.