Research on the Mechanism of Isoliquiritigenin Improving Pain Sensitivity in KOA Based on the Nrf2/HO-1 Signaling Pathway

OBJECTIVE To investigate the mechanism by which isoliquiritigenin (ISL) improves pain hypersensitivity in knee osteoarthritis (KOA) by regulating the Nrf2/HO-1 signaling pathway.

METHODS Fifty C57BL/6J mice were randomly divided into five groups (n=10): control group, KOA group, low-dose ISL group (10 mg·kg^-1), high-dose ISL group (40 mg·kg^-1), and celecoxib group (43.3 mg·kg^-1). A KOA model was established using the destabilization of the medial meniscus (DMM) surgery, and pain hypersensitivity was assessed. In vivo experiments included ELISA to measure serum levels of TNF-α, IL-1β, and IL-6; immunofluorescence to observe TRPA1, TRPV1, and TRPM8 ion channel fluorescence intensity in dorsal root ganglia (DRG); and Western blot to quantify Nrf2, HO-1, PGP9.5, NGF, and Substance P protein expression in DRG. For in vitro experiments, ML385 was used to inhibit the Nrf2/HO-1 pathway, and the effects of ISL on neurons were evaluated after pathway inhibition.

RESULTS Compared with the control group, the KOA group exhibited significantly reduced cold and mechanical stimulation thresholds (P<0.01), increased TRPA1, TRPV1, and TRPM8 fluorescence intensity (P<0.01), elevated serum TNF-α, IL-1β, and IL-6 levels (P<0.01), decreased Nrf2 and HO-1 expression, and increased PGP9.5, NGF, and Substance P expression (P<0.01). Compared with the KOA group, both ISL dose groups and the celecoxib group showed significantly improved cold and mechanical stimulation thresholds (P<0.01), reduced TRP channel fluorescence intensity (P<0.01), decreased inflammatory cytokine levels (P<0.05,P<0.01), upregulated Nrf2 and HO-1 expression, and downregulated PGP9.5, NGF, and Substance P expression (P<0.05, P<0.01). In vitro experiments revealed that LPS treatment reduced Nrf2/HO-1 levels (P<0.01), increased PGP9.5, NGF, and Substance P expression (P<0.01), and elevated reactive oxygen species (ROS) (P<0.01). ISL reversed these effects (P<0.01), while ML385 co-treatment abolished ISL’s benefits, reactivating ROS and pain-related proteins (P<0.01).

CONCLUSION Isoliquiritigenin alleviates KOA pain hypersensitivity by activating the Nrf2/HO-1 signaling pathway.