Figure 4

Kinetics of IL-1β and IL-1F8 mRNA production by HSFs and hCAs in response to IL-1 and/or TNF-α. (a) Analysis of IL-1F8 mRNA levels in hSFs treated or not treated for 8 hours with IL-1β (1 ng/ml) and/or TNF-α (10 ng/ml), as detailed under Materials and method. A representative agarose gel electrophoresis of PCR products is shown. (b) Real-time PCR analysis of IL-1F8 mRNA levels in hSFs stimulated (black columns) or not stimulated (white columns) for 8 hours with IL-1β (1 ng/ml) alone, IL-1β (1 ng/ml) plus TNF-α (10 ng/ml), or IL-1α (1 ng/ml) alone, as indicated. The amount of 28S rRNA was monitored as an internal control. The expression of IL-1F8 mRNA was corrected for 28S rRNA levels and the IL-1F8/28S ratios were normalized to the maximal value observed in each experiment, which was set to 100%. The results shown represent the mean ± standard error of data obtained with samples from three independent cultures. *P < 0.05 versus, as determined by analysis of variance. (c) Fold increase (after correction for 28S RNA levels) in IL-1β (dashed line) and IL-1F8 (solid line) mRNA levels after treatment of hSFs with 1 ng/ml IL-1β for the indicated times, as revealed by real-time PCR analysis. Basal IL-1F8/28S and IL-1β/28S levels were respectively 6.4 and 20 (arbitrary units). (d) Real-time PCR analysis of IL-1F8 and IL-1β mRNA levels in hACs stimulated (black columns) or not stimulated (white columns) with IL-1β (1 ng/ml) and TNF-α (10 ng/ml) for 8 hours. The amount of 28S rRNA was monitored as an internal control. The expression of IL-1F8 and IL-1β mRNA was corrected for 28S rRNA levels and the IL/28S ratios were normalized to the maximal value observed in each experiment, which was set to 100%. The results shown represent the mean ± standard error of data obtained with samples from six independent cultures. *P < 0.05 versus, as determined by analysis of variance. hAC, human articular chondrocyte; hSF, human synovial fibroblast; IL, interleukin; RT-PCR, reverse transcriptase polymerase chain reaction; TNF, tumour necrosis factor.