Evidence Linking Chondrocyte Lipid Peroxidation to Cartilage Matrix Protein Degradation

Abstract

Reactive oxygen species (ROS) are implicated in both cartilage aging and the pathogenesis of osteoarthritis. We developed an in vitro model to study the role of chondrocyte-derived ROS in cartilage matrix protein degradation. Matrix proteins in cultured primary articular chondrocytes were labeled with [(3)H]proline, and the washed cell matrix was returned to a serum-free balanced salt solution. Exposure to hydrogen peroxide resulted in oxidative damage to the cell matrix as established by monitoring the release of labeled material into the medium. Calcium ionophore treatment of chondrocytes, in a dose-dependent manner, significantly enhanced the release of labeled matrix, suggesting a chondrocyte-dependent mechanism of matrix degradation. Antioxidant enzymes such as catalase or superoxide dismutase did not influence matrix release by the calcium ionophore-activated chondrocytes. However, vitamin E, at physiological concentrations, significantly diminished the release of labeled matrix by activated chondrocytes. The fact that vitamin E is a chain-breaking antioxidant indicates that the mechanism of matrix degradation and release is mediated by the lipid peroxidation process. Lipid peroxidation was measured in chondrocytes loaded with cis-parinaric acid. Both resting and activated cells showed constitutive and enhanced levels of lipid peroxidation activity, which were significantly reduced in the presence of vitamin E. In an immunoblot analysis, malondialdehyde and hydroxynonenal adducts were observed in chondrocyte-matrix extracts, and the amount of adducts increased with calcium ionophore treatment. Furthermore, vitamin E diminished aldehyde-protein adduct formation in activated extracts, which suggests that vitamin E has an antioxidant role in preventing protein oxidation. This study provides in vitro evidence linking chondrocyte lipid peroxidation to cartilage matrix protein (collagen) oxidation and degradation and suggests that vitamin E has a preventive role. These observations indicate that chondrocyte lipid peroxidation may have a role in the pathogenesis of cartilage aging and osteoarthritis.