Journal
OSTEOARTHRITIS AND CARTILAGE
Volume 9, Issue 4, Pages 294-299Publisher
W B SAUNDERS CO LTD
DOI: 10.1053/joca.2000.0388
Keywords
stress-activated protein kinase; nitric oxide; osteoarthritic cartilage
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Objective: We have demonstrated in bovine chondrocytes that nitric oxide (NO) mediates IL1 dependent apoptosis under conditions of oxidant stress. This process is accompanied by activation of c-Jun NH2-terminal kinase (JNK; also called stress-activated protein kinase). In these studies we examined activation of JNK in explant cultures of human osteoarthritic cartilage obtained at joint replacement surgery and we characterized the role of peroxynitrite to act as an upstream trigger. Design: A novel technique to isolate chondrocyte proteins (<10% of total cartilage protein) from cartilage specimens was developed. It was used to analyse JNK activation by a western blot technique. To examine the hypothesis that chondrocyte JNK activation is a result of increased peroxynitrite, in vitro experiments were performed in which cultured chondrocytes were incubated with this oxidant. Results: Activated JNK was detected in the cytoplasm of osteoarthritis (OA) affected chondrocytes but not in that of controls. In vitro, chondrocytes produce NO and superoxide anion. IL-1 (48 h), which induces nitric oxide synthase, resulted in an activation of JNK; this effect was reversed by N-monomethylarginine (NMA). TNF treated chondrocytes at 48 h produce superoxide anion (EPR method). Exposure of cells to peroxynitrite led to an accumulation of intracellular oxidants, in association with JNK activation and cell death by apoptosis. Conclusion: We suggest that JNK activation is among the IL-1 elicited responses that injure articular chondrocytes and this activation of JNK is dependent on intracellular oxidant formation (including NO peroxynitrite). In addition, the extraction technique here described is a novel method that permits the quantitation and study of proteins such as JNK involved in the signaling pathways of chondrocytes within osteoarthritic cartilage. (C) 2001 OsteoArthritis Research Society International.
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