Antioxidants and inhibitors of flavoprotein-dependent oxidases abrogate TGF-beta induction of biglycan: Evidence for a role of reactive oxygen species

Transforming growth factor-beta (TGF-β) in excess evokes inflammation, fibrosis, and desmoplasia in part by stimulating extracellular matrix synthesis including expression of the small leucine-rich proteoglycan biglycan (Bgn). Since TGF-β exerts some of its effects by stimulating the generation of reactive oxygen species (ROS) in cells, we investigated the possibility that ROS signaling is involved in Bgn induction by TGF-β. In pancreatic carcinoma PANC-1 cells, antioxidants (N-acetyl-L-cysteine, ascorbic acid), ROS scavengers (Tiron) and inhibitors of flavoprotein-dependent oxidases such as the ROS producing enzyme NADPH oxidase (apocynin and diphyleneiodonium) all inhibited TGF-β1-induced Bgn expression. In agreement with a role of the transcription factor NFκB in ROS cellular actions, the NFκB inhibitor pyrrolidinecarbodithiocarbamate also reduced the TGF-β effect on Bgn, while S-M-sulfate, an inhibitor of the nitric oxide-generating enzyme iNOS was ineffective in this respect. TGF-β time-dependently and moderately induced ROS production that was suppressed by diphenyleneiodonium. RT-PCR revealed that PANC-1 cells expressed various subunits of NADPH oxidase, namely p22phox, p47phox and p67phox as well as Nox isoforms 2, 4, 5 and 6, but lacked mRNA for Nox1 and 3. Blocking TGF-β-induced Bgn expression by modulating cellular redox status could have beneficial effects on fibrotic disorders caused by TGF-β hyperactivity.