Effects of TNF-α and TGF-β1 on IL-1β-stimulated MMP-12 activity, mRNA expression and c-Jun activation. ASMC were treated with 10 ng/ml of TNF-α or TGF-β1 alone, or in combination with 10 ng/ml IL-1β. (A) MMP-12 activity in condition media was detected by zymography after 48 hours, and the relevant band intensities were quantified by densitometric analysis and normalized to 5 × 105 cells. (B) MMP-12 mRNA expression was determined by real-time RT-PCR after 24 hours and expressed as a ratio to GAPDH mRNA. (C) c-Jun activation and DNA-binding analysed by the TransAM AP-1 family kit. Data are the mean ± SEM from three ASMC donors. *P < 0.05, **P < 0.01 compared with control; +P < 0.05, ++P < 0.01, compared with IL-1β alone.