Genistein targets Wnt5a to alleviate temporomandibular joint osteoarthritis by suppressing JNK/JUN-dependent MMP9 activation.

Abstract

Temporomandibular joint osteoarthritis (TMJ-OA) is a progressive degenerative joint disease characterized by cartilage destruction, extracellular matrix (ECM) degradation, and chronic inflammation, yet effective disease-modifying therapeutic targets and pharmacological interventions remain limited. Using a monosodium iodoacetate (MIA)-induced rat TMJ-OA model and interleukin-1β (IL-1β)-stimulated chondrocytes, we integrated transcriptomic analysis, chromatin accessibility profiling, and molecular screening to elucidate disease mechanisms and identify candidate compounds. Integrated RNA sequencing (RNA-seq) analysis revealed Wnt family member 5a (Wnt5a) as a critical upstream regulator significantly associated with TMJ-OA progression, while assay for transposase-accessible chromatin using sequencing (ATAC-seq) uncovered an inflammatory chromatin landscape characterized by enhanced activator protein-1 (AP-1) activity and JUN-dependent transcriptional regulation ofmatrix metalloproteinase 9 (Mmp9). Mechanistically, Wnt5a activated c-Jun N-terminal kinase (JNK) signaling to promote JUN nuclear accumulation and enhance AP-1-mediatedMmp9transcription, leading to ECM degradation and chondrocyte dysfunction. Structure-based virtual screening and biophysical assays, including surface plasmon resonance (SPR), cellular thermal shift assay (CETSA), and drug affinity responsive target stability (DARTS), identified genistein (GEN) as a small-molecule compound capable of directly binding Wnt5a, supported by cellular target-engagement evidence. Functionally, genistein suppressed the Wnt5a/JNK/JUN/MMP9 signaling axis, partially restored type II collagen (COL2A1) expression, alleviated oxidative stress, and preserved cartilage integrity in TMJ-OA rats. Collectively, these findings establish Wnt5a as a therapeutically actionable driver of TMJ-OA progression and identify genistein as a Wnt5a-targeting compound with cartilage-protective activity, providing a mechanism-based rationale for developing disease-modifying interventions for degenerative joint diseases.