WNT16 has been shown to play important roles in joint formation, bone homeostasis and knee joint osteoarthritis. However, whether WNT16 has any effect during temporomandibular joint osteoarthritis (TMJOA) is still unknown. Here, we first established a surgically induced TMJOA model by performing partial discectomy in discs of TMJ in mice. Further, we investigated the role of WNT16 during the initiation and progression of TMJOA. Our results showed that WNT16 expression is upregulated early at 4 weeks after initiation of osteoarthritis by partial discectomy in mouse TMJ cartilage, but decreased after 12 weeks post-surgery. Further cellular and molecular analyses revealed that WNT16 signals via both the canonical WNT/β-catenin and non-canonical WNT/JNK-cJUN pathways, upregulates the expression of Lubricin and SOX9, and protects against IL-1β induced inflammatory response by regulation of RUNX2/MMP13 cascade in fibrochondrocytes. In conclusion, WNT16 may play an important role in the early stage of TMJOA by regulating cartilage anabolic and catabolic factors, and may serve as novel therapeutic targets for TMJOA.