FIGURE 1.

IL-1 + OSM-stimulated MMP13 mRNA expression is preceded by nascent hnRNA formation and involves early induction of cFOS in human chondrocytes. Chondrocytes were stimulated with IL-1 (0.05 ng/ml) in combination with OSM (10 ng/ml) for the indicated durations. Total RNA was isolated, reverse transcribed, and subjected to real-time RT-PCR for MMP13 mRNA (A), cFOS mRNA (B), or MMP13 nascent hnRNA (G) as described under “Experimental Procedures.” Data are expressed relative to 18S rRNA and presented as fold-increase compared with basal expression. PCR data (n = 4) are representative of at least three separate chondrocyte populations. Cells were subjected to subcellular fractionation and protein from the cytosolic (a), membrane-bound (b), soluble nuclear (c), chromatin-bound, (d) and cytoskeletal (e) fractions resolved using SDS-PAGE and immunoblotted with antibodies to cFOS (C) or phosphorylated cJUN (E). Combined densitometric scans of five separate blots for each protein in the soluble nuclear fraction, relative to t = 0 h, are shown in D and F, respectively, and were obtained from separate chondrocyte populations. All data are presented as mean (± S.D.), where ***, p < 0.001; **, p < 0.01; *, p < 0.05 IL-1 + OSM-treated compared with control; ANOVA.