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Cytokine-mediated induction and overexpression of matrix metalloproteinases (MMPs) is recognized as an important factor in the pathogenesis of arthritis. Interleukin (IL)-1␤ is a proinflammatory cytokine that is known to superinduce the expression and production of MMP-13 in many cell types. Phenyl N-tert-butylnitrone (PBN), a spin trap agent, inhibited the IL-1␤-induced expression of MMP-13 in human osteoarthritis (OA) chondrocytes. Downregulation of MMP-13 expression correlated with the inhibition of mitogen-activated protein kinase (MAPK) subgroups c-Jun NH 2 -terminal kinase (JNK) and p38-MAPK activation, accumulation of phospho-c-jun, and the DNA binding activity of activating protein-1 (AP-1). Results of in vitro kinase assays showed that exogenously added PBN completely blocked the c-Jun phosphorylating activity of JNK. Interestingly, using in vitro kinase assay, we also found that chondrocyte p38-MAPK phosphorylate c-Jun and that PBN was not very effective in inhibiting c-Jun phosphorylating activity of p38-MAPK. In addition, PBN did not block the ATF-2 phosphorylating activity of p38-MAPK and Elk-1 phosphorylating activity of extracellular regulated kinase p44/p42 in vitro, indicating that PBN may act selectively to inhibit the phosphorylation of c-Jun in OA chondrocytes. Together, our results for the first time demonstrate that PBN suppresses the IL-1␤-stimulated expression of MMP-13 in OA chondrocytes and that this was achieved by inhibiting the activation of JNK and AP-1. These results suggest that use of PBN or compounds derived from it may be of potential benefit in inhibiting signaling events associated with cartilage degradation in arthritis.

The major allergens in fish‐food are parvalbumins (PVs). Muscle PVs of four fish species of genus Channa: Channa gachua, Channa punctatus, Channa striatus and Channa marulius were purified, characterized and used to differentiate (heated) fish specimens. Two to three proteins have been detected as PV‐isoforms by immunological probing through Western blotting. They were purified to homogeneity on diethyl amino ethyl‐cellulose. Modified systems of nondenaturing and denaturing polyacrylamide gel electrophoresis were used to resolve low molecular weight proteins, distinctly. Isoelectric points (pIs) were determined on native isoelectric focusing gels, which lie in the pI range of 4.55–5.10. The electrophoretic profiles remained species‐specific even after heating the PVs for 3 h at 90C and would be of immense help in discerning cooked fish specimens. Further, the addition of 5 mM β‐mercaptoethanol in the PVs has showed protection against heat denaturation of the protein and resulted in more compatible protein patterns. This study also demonstrates the relevance of the thermostable capacity of PVs in the differentiation of heat‐processed fish through electrophoretic analysis of heated PVs. Also, they represent a promising and simple biochemical tool in discerning closely related species.

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