Jiawei Yanghe decoction (JWYHD) is a Traditional Chinese Medicine (TCM) formula for the treatment of osteoarthritis (OA), however the underlying mechanisms of action of JWYHD in OA are not fully explored. This study investigates how JWYHD protects cartilage from degradation via Wnt/β-catenin signaling pathway. The chondroprotective and anti-inflammatory effect of JWYHD on chondrocytes in vitro and on MIA-induced OA rat model in vivo were investigated. In vitro, JWYHD increased the chondrocyte viability against interleukin (IL)-1β-induced chondrocytes apoptosis and preserved glycosaminoglycans in the extracellular matrix. JWYHD promoted chondrocyte viability against apoptosis, decreased MMP-3, MMP-13, Caspase-3, Caspase-9 via Wnt/β-catenin signaling pathway in both IL-1β-induced and Licl-induced chondrocytes. The qRT-PCR and western blot results showed that mRNA and protein expressions of Wnt signaling pathway related genes β-catenin and CyclinD1, apoptosis related genes Casapase-3 and Caspase-9, collagen degradation related genes Metalloproteinase (MMP)-3 and MMP-13 were up-regulated, and Col2a1 was down-regulated on IL-1β-induced chondrocytes. Treatment with JWYHD reversed these effects in a dose-dependent manner. Licl was used as Wnt/β-catenin signaling pathway activator in chondrocytes to determine the molecular mechanisms. Activation of Wnt signaling pathway by Licl up-regulated β-catenin, CyclinD1, Axin2, Caspase-3, Caspase-9, MMP-3, MMP-13 and IL-1β. These effects were blocked by JWYHD treatment. Furthermore, 75 Sprawl-Dawley rats were used to verify the results obtained in vitro. A total of 75 rats were randomly divided into the control group (no MIA-induced OA, received intragastric administration of an equivalent amount of saline), the OA group (MIA-induced OA, received intragastric administration of an equivalent amount of saline), and the JWYHD treatment group (MIA-induced OA, received intragastric administration of an equivalent amount of various concentrations of JWYHD at 1.4/2.7/5.5 g/kg). After 8 weeks of administration, all rats were sacrificed. JWYHD decreased the MIA-induced up-regulation of β-catenin, CyclinD1, Caspase-3, Caspase-9, MMP-3 and MMP-13 protein expressions in cartilage. It was also demonstrated that JWYHD decreased serum and synovium pro-inflammatory cytokines, IL-1β, IL-6 and TNF-α in MIA-induced OA rats and ameliorated the cartilage degradation. Histopathological staining, macroscopic observation and micro-CT scan with 3-dimension remodeling showed a cartilage protective effect of JWYHD. In conclusion, JWYHD possess multiple capabilities including preventing chondrocyte apoptosis, preserving integrity of extracellular matrix and anti-inflammatory effect in the treatment of OA both in vitro and in vivo.