ObjectiveTo investigate the role of osteopontin (OPN) on the expressions of matrix metalloproteinase 13 (MMP-13) mRNA and protein in human knee osteoarthritic chondrocytes and to find the optimal time and concentration for OPN treatment. MethodsChondrocytes were isolated from articular cartilage tissues of patients with primary osteoarthritis (OA) and cultured using one step digestive treatment with collagenase typeⅡ. The chondrocytes were then identified using immunohistochemistry of collagen typeⅡ. The first generation of chondrocytes were stimulated with OPN at a concentration of 1μg/mL for 0, 24, 48, and 72 hours, and with OPN at the concentrations of 0, 0.5, 1, 2, and 4μg/mL for 48 hours. The levels of MMP-13 mRNA and protein expressions were measured with real-time fluorescent quantitative PCR and Western blot. ResultsThe immunohistochemical staining showed that first generation of chondrocytes expressed collagen typeⅡ. Both MMP-13 mRNA and protein expression levels in OA chondrocytes increased significantly in the presence of OPN (1μg/ mL) and peaked at 48 hours after incubation, showing significant difference between different time points (P < 0.05). The MMP-13 mRNA expression level in OA chondrocytes at the OPN concentration of 1μg/mL was significantly higher than those at the other concentrations (P < 0.05), and the MMP-13 protein expression level at the OPN concentration of 1μg/mL was significantly higher than that at 0μg/mL (P < 0.05). MMP-13 protein expression level at the OPN concentrations of 0.5, 2, and 4μg/mL were significantly higher than that at 0μg/mL (P < 0.05). ConclusionOPN induces up-regulation of MMP-13 mRNA and protein expressions in human knee osteoarthritic chondrocytes in time-and dose-dependent manners. The optimal time and concentration for OPN treatment are 48 hours and 1μg/mL, respectively.