Osteoarthritis (OA) is a debilitating, progressive disease of diarthrodial joints associated with aging. At the molecular level, OA is characterized by an imbalance between anabolic (i.e. extracellular matrix biosynthesis) and catabolic (i.e. extracellular matrix degradation) pathways in which articular cartilage is the principal site of tissue injury responses. The pathophysiology of OA also involves the synovium in that ‘nonclassical’ inflammatory synovial processes contribute to OA progression. Chondrocytes are critical to the OA process in that the progression of OA can be judged by the vitality of chondrocytes and their ability to resist apoptosis. Growth factors exemplified by insulin-like growth factor-1, its binding proteins and transforming growth factor-β contribute to anabolic pathways including compensatory biosynthesis of extracellular matrix proteins. Catabolic pathways are altered by cytokine genes such as interleukin-1 (IL-1) and tumor necrosis factor-α (TNF-α) which are upregulated in OA. In addition, IL-1 and TNF-α downregulate extracellular matrix protein biosynthesis while concomitantly upregulating matrix metalloproteinase (MMP) gene expression. When MMPs are activated, cartilage extracellular matrix degradation ensues apparently because levels of endogenous cartilage MMP inhibitors cannot regulate MMP activity. Therapeutic strategies designed to modulate the imbalance between anabolic and catabolic pathways in OA may include neutralizing cytokine activity or MMP gene expression or inhibiting signaling pathways which result in apoptosis dependent on mature caspase activity or mitogen-activated protein kinase (MAPK) activity. MAPK activity appears critical for regulating chondrocyte and synoviocyte apoptosis and MMP genes.

Objective. The proinflammatory cytokine interleukin-1␤ (IL-1␤) induces the production of high levels of nitric oxide (NO) in human chondrocytes. Green tea (Camellia sinensis) polyphenols are potent antiinflammatory agents and have been shown to inhibit NO production in tumor cell lines. In the present study, we examined the effect of epigallocatechin-3-gallate (EGCG), a green tea polyphenol, on IL-1␤-induced production of NO in primary human osteoarthritis (OA) chondrocytes.Methods. Human chondrocytes were derived from OA cartilage and were treated with EGCG (100 M) and IL-1␤ (2 ng/ml) for different periods, and inducible nitric oxide synthase (iNOS) messenger RNA and protein expression was determined by real-time quantitative reverse transcriptase-polymerase chain reaction and Western blotting, respectively. Production of NO was determined as nitrite in culture supernatant. Activation and translocation of nuclear factor B (NF-B), levels of inhibitor of nuclear factor B (I B), and NF-B DNA binding activity were determined by Western blotting and a highly sensitive and specific enzymelinked immunosorbent assay. Activity of I B kinase was determined using in vitro kinase assay.Results. Human chondrocytes cotreated with EGCG produced significantly less NO compared with chondrocytes stimulated with IL-1␤ alone (P < 0.005). The inhibition of NO production correlated with the suppression of induction and expression of NF-Bdependent gene iNOS. EGCG inhibited the activation and translocation of NF-B to the nucleus by suppressing the degradation of its inhibitory protein I B␣ in the cytoplasm.Conclusion. Our results indicate that EGCG inhibits the IL-1␤-induced production of NO in human chondrocytes by interfering with the activation of NF-B through a novel mechanism. Our data further suggest that EGCG may be a therapeutically effective inhibitor of IL-1␤-induced inflammatory effects that are dependent on NF-B activation in human OA chondrocytes.The proinflammatory cytokine interleukin-1␤ (IL-1␤), produced in an arthritic joint by activated synovial cells and infiltrating macrophages, is considered to be one of the most potent catabolic factors in joint diseases (1). IL-1␤ induces the production of several mediators of cartilage degradation, such as nitric oxide (NO) and matrix metalloproteinases (1-3), and inhibits the concentration of tissue inhibitor of metalloproteinases in arthritic joints (3). IL-1␤ also suppresses the biosynthesis of type II collagen and aggrecan (4-6) and the proliferation of chondrocytes (7-9), thus inhibiting the repair process in the cartilage. Additional evidence indicating the involvement of IL-1 in cartilage degrada-

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(131)I-mIBG therapy produces symptomatic and hormonal improvement and moderate tumour regression/stabilization in patients with metastatic neuroendocrine tumours with minimal adverse effects. It may be a valuable alternative or additional therapeutic option to the currently available conventional treatment modalities.

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Hydrostatic pressure (HP) is thought to increase within cartilage extracellular matrix as a consequence of fluid flow inhibition. The biosynthetic response of human articular chondrocytes to HP in vitro varies with the load magnitude, load frequency, as well as duration of loading. We found that continuous cyclic HP (5 MegaPascals (MPa) for 4 h; 1 Hz frequency) induced apoptosis in human chondrocytes derived from osteoarthritic cartilage in vitro as evidenced by reduced chondrocyte viability which was independent of initial cell densities ranging from 8.1 x 10(4) to 1.3 x 10(6) cells ml(-1). HP resulted in internucleosomal DNA fragmentation, activation of caspase-3, and cleavage of poly-ADP-ribose polymerase (PARP). At the molecular level, induction of apoptosis by HP was characterized by up-regulation of p53, c-myc, and bax-alpha after 4 h with concomitant down-regulation of bcl-2 after 2 h at 5 MPa as measured by RT-PCR. In contrast, beta-actin expression was unchanged. Real-time quantitative RT-PCR confirmed a HP-induced (5 MPa) 1.3-2.6 log-fold decrease in bcl-2 mRNA copy number after 2 and 4 h, respectively, and a significant increase (1.9-2.5 log-fold) in tumor necrosis factor-alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS) mRNA copy number after 2 and 4 h, respectively. The up-regulation of p53 and c-myc, and the down-regulation of bcl-2 caused by HP were confirmed at the protein level by Western blotting. These results indicated that HP is a strong inducer of apoptosis in osteoarthritic human chondrocytes in vitro.

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