| 16679036 |
Lopez-Armada MJ, Carames B, Martin MA, Cillero-Pastor B, Lires-Dean M, Fuentes-Boquete I, Arenas J, Blanco FJ: Mitochondrial activity is modulated by TNFalpha and IL-1beta in normal human chondrocyte cells. Osteoarthritis Cartilage. 2006 Oct;14(10):1011-22. Epub 2006 May 5.
OBJECTIVE: Pro-inflammatory cytokines play an important role in osteoarthritis (OA). In osteoarthritic cartilage, chondrocytes exhibit an alteration in mitochondrial activity. This study analyzes the effect of tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta) on the mitochondrial activity of normal human chondrocytes. MATERIALS AND METHODS: Mitochondrial function was evaluated by analyzing the activities of respiratory chain enzyme complexes and citrate synthase, as well as by mitochondrial membrane potential (Deltapsim) and adenosine triphosphate (ATP) synthesis. Bcl-2 family mRNA expression and protein synthesis were analyzed by RNase protection assay (RPA) and Western-blot, respectively. Cell viability was analyzed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) and apoptosis by 4', 6-diamidino-2-phenylindole dihydrochloride (DAPI) stain. Glycosaminoglycans were quantified in supernatant by a dimethyl-methylene blue binding assay. RESULTS: Compared to basal cells, stimulation with TNFalpha (10 ng/ml) and IL-1beta (5 ng/ml) for 48 h significantly decreased the activity of complex I (TNFalpha=35% and IL-1beta=35%) and the production of ATP (TNFalpha=18% and IL-1beta=19%). Both TNFalpha and IL-1beta caused a definitive time-dependent decrease in the red/green fluorescence ratio in chondrocytes, indicating depolarization of the mitochondria. Both cytokines induced mRNA expression and protein synthesis of the Bcl-2 family. Rotenone, an inhibitor of complex I, caused a significant reduction of the red/green ratio, but it did not reduce the viability of the chondrocytes. Rotenone also increased Bcl-2 mRNA expression and protein synthesis. Finally, rotenone as well as TNFalpha and IL-1beta, reduced the content of proteoglycans in the extracellular matrix of normal cartilage. CONCLUSION: These results show that both TNFalpha and IL-1beta regulate mitochondrial function in human articular chondrocytes. Furthermore, the inhibition of complex I by both cytokines could play a key role in cartilage degradation induced by TNFalpha and IL-1beta. These data could be important for understanding of the OA pathogenesis. |
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