Mechanistic Investigation of Aconitine Combined with Paeoniflorin Against Knee Osteoarthritis via the Ihh-Gli Signaling Pathway
This study interrogates the therapeutic efficacy and molecular mechanism of aconitine combined with paeoniflorin in a rat model of knee osteoarthritis (KOA), focusing on the Indian hedgehog (Ihh)-glioma-associated oncogene homolog (Gli) signaling axis. Anterior cruciate ligament transection (ACLT) was performed on male rats, which were then allocated to sham, model, celecoxib (24 mg/kg), and three aconitine-paeoniflorin dose groups (5+50, 10+100, 20+200 μg/kg; n=10 per group). Behavioral tests, hematoxylin-eosin staining, micro-computed tomography, ELISA for matrix metalloproteinase 13 (MMP13) and type II collagen (Col II), immunofluorescence, and qRT-PCR for Ihh, Gli, patched 1 (Ptch1), and MMP13 were conducted. Molecular docking assessed binding affinities. Safety was evaluated via serum aspartate aminotransferase, creatinine, blood urea nitrogen, urinary protein, and histopathology of heart, liver, and kidney. Results demonstrated that the combination significantly elevated mechanical and thermal pain thresholds (P<0.05, 0.01, 0.001), restored cartilage matrix integrity, improved bone microarchitecture, decreased serum MMP13, and increased Col II (P<0.05, 0.01, 0.001). Ihh, Gli, Ptch1, and MMP13 protein and gene expressions were markedly downregulated (P<0.05, 0.01, 0.001). Docking confirmed binding energies ≤−5 kcal/mol for aconitine and paeoniflorin with Ihh, Gli, ADAMTS5, and MMP13. No significant hepatic, renal, or cardiac toxicity was observed. The combination inhibits aberrant Ihh-Gli pathway activation, suppresses cartilage matrix degradation, and offers a safer, multi-target alternative to celecoxib for KOA management.