Esculetin inhibits cartilage resorption induced by interleukin 1 alpha in combination with oncostatin M

Objective-To determine if a new inhibitor, esculetin (EST), can block resor ption of cartilage. Methods-Interleukin 1 alpha (IL1 alpha, 0.04-5 ng/ml) and oncostatin M (OSM , 0.4-50 ng/ml) were used to stimulate the release of proteoglycan and coll agen from bovine nasal cartilage and human articular cartilage in explant c ulture. Proteoglycan and collagen loss were assessed by dimethyl-methylene blue and hydroxyproline assays, respectively. Collagenase levels were measu red by assay of bioactivity and by enzyme linked immunosorbent assay (ELISA ). The effects of EST on the expression of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinase-l (TIMP-1) in the transformed hu man chondrocyte cell line T/C28a4 were assessed by northern blot analysis. TIMP-1 protein levels were assayed by ELISA. The effect of EST on the MMP-1 promoter was assessed using a promoter-luciferase construct in transient t ransfection studies. Results-EST inhibited proteoglycan and collagen resorption in a dose depend ent manner with significant decreases seen at 66 muM and 100 muM EST, respe ctively. Collagenolytic activity was significantly decreased in bovine nasa l cartilage cultures. In human articular cartilage, EST also inhibited IL1 alpha + OSM stimulated resorption and decreased MMP-1 levels. TIMP-1 levels were not altered compared with controls. In T/C28a4 chondrocytes the IL1 a lpha + OSM induced expression of MMP-1, MMP-3, and MMP-13 mRNA was reduced to control levels by 250 muM EST TIMP-1 mRNA levels were unaffected by EST treatment. All cytokine stimulation of an MMP-1 luciferase-promoter constru ct was lost in the presence of the inhibitor. Conclusion-EST inhibits degradation of bovine nasal cartilage and human art icular cartilage stimulated to resorb with IL1 alpha + OSM.