Hydrogen peroxide-mediated transcriptional induction of macrophage colony-stimulating factor by TGF-beta1

TGF-beta1 and macrophages are important regulators of tissue fibrosis and remodeling. Here we show that TGF-beta1 induces the expression of macrophage-CSF (M-CSF) in vascular endothelial cells via a signaling pathway(s) involving hydrogen peroxide (H2O2). In a time-dependent manner, TGF-beta1 produced a 10- and a 6-fold increase in M-CSF mRNA and protein levels after 12 h, respectively. This increase in M-CSF expression was attenuated by a nitric oxide donor, S-nitrosoglutathione (GSNO), and by a nonspecific oxidase inhibitor, diphenylene iodonium. Furthermore, the TGF-beta1-induced M-CSF mRNA expression was inhibited by catalase, but not by superoxide dismutase, suggesting that H2O2 rather than superoxide anion (O2.-) is the primary mediator of the effects of TGF-beta1. Transient transfection studies using deletional M-CSF promoter constructs demonstrated that TGF-beta1 produced a 13-fold induction in M-CSF promoter activity that was repressed by >85% with GSNO and catalase, in part through inhibitory effects on kappaB cis-acting elements. Electrophoretic mobility shift assays revealed that the activation of nuclear factor-kappaB by TGF-beta1 was also inhibited by GSNO and catalase, but not by superoxide dismutase. In a concentration-dependent manner, treatment with exogenous H2O2 produced 14- and 4.6-fold increases in M-CSF promoter activity and mRNA expression, respectively. These results indicate that the generation of H2O2 and activation of NF-kappaB by TGF-beta1 are required for the induction of M-CSF gene transcription.