ABSTRACT

lide has been used for treating various autoimmune diseases. However, it remains unclear whether triptolide exerts effects on extracellular matrix (ECM) synthesis, which plays an important role in renal fibrosis. Methods. NRK-49F cells stimulated with TGF-β1 were incubated with triptolide in various concentrations. ECM proteins, including collagen type III and fibronectin, were detected using the reverse transcription real-time PCR and ELISAmethods.MAPK and Smad2/3 phosphorylationwere measured withwestern blot. P38 and ERK1/2 pathwayswere inhibited with the specific inhibitors, SB203580 and PD98059. The Smad2 signal was blocked with the siRNA method. Results. Triptolide inhibited ECM synthesis in TGF-β1-stimulated NRK-49F cells in a concentration-dependent manner. Triptolide enhanced TGF-β1-induced activation of the p38, ERK 1/2 signals, whereas it inhibited Smad2 activation.There was no crosstalk between the p38, ERK 1/2 and Smad2 pathways in NRK-49F cells. Inhibition of either the p38 or ERK 1/2 signals reduced ECM synthesis. Triptolide downregulated synthesis of fibronectin and collagen type 3180 B. Zhu et al.? The Author 2010. Published by Oxford University Press on behalf of ERA-EDTA.III in TGF-β1-stimulated cells treatedwith SB203580 and/or PD98059.SB203580 and/or PD98059 significantly repressed synthesis of fibronectin and collagen type III in TGF-β1-stimulated cells treated with triptolide. Smad2 inhibition by siRNA significantly reduced ECMsynthesis. However,ECM synthesis in NRK-49F cells transfected with Smad2 siRNA and treated by triptolide was increased compared with Smad2 siRNA-transfected cells.Conclusion. The effect of triptolide to suppress ECM synthesis by inhibiting Smad2 activation may surpass its stimulating effect on ECM synthesis by activation of p38 and ERK 1/2, leading to a total inhibition of ECM synthesis in TGF-β1-stimulated NRK-49F cells.