Human chondrocytes, reactive oxygen species production and Nox
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KeywordsReactive Oxygen Species Reactive Oxygen Species Production NADPH Oxidase Respiratory Burst Ionomycin
Joint destruction is a key point in many inflammatory and degenerative diseases. This is mainly mediated by inflammation cells and resident cells such as chondrocytes. Matrix metalloproteinases and reactive oxygen species are widely involved.
This work shows that three different human immortalized chondrocyte (HIC) cell lines (a gift from M Goldring) and native chondrocytes can generate superoxide anions (O2-) constitutively and after activation with PMA or ionomycin. The O2- produced by chondrocyte NADPH oxidase is about 0.1% of the respiratory burst measured in phagocytic cells such as neutrophils. All the components of NADPH oxidase were detected by Western blot analysis: the cytosolic factors p47phox, p67phox and p40phox and the two subunits of the cytochrome b558, gp91phox (Nox2) and p22phox. The presence of cytochrome b558 was also demonstrated by reduced minus oxidized differential spectra in 1% Triton X-100-soluble extract from the three HIC cell lines. The cytochrome b558 content in chondrocytes was about 10 pmol/mg of proteins: this amount is lower than in neutrophils (200 pmol/mg of proteins). The presence of Nox4 and Nox2 mRNA in the three HIC cell lines was demonstrated by RT-PCR and Northern blot analysis.
The three HIC cell lines and native chondrocytes contain phagocyte NADPH oxidase components, and Nox2 (gp91phox) analogs as Nox4 and Nox5. The low level of superoxide anions O2- produced by chondrocytes in comparison with neutrophils may be involved in signal transduction and may be implicated in the structural and functional alterations of cartilage matrix observed in arthritis and osteoarthritis.