Effect of TGF-β/Smad signaling pathway on lung myofibroblast differentiation

Aim:Myofibroblasts play important roles in the pathogenesis of lung fibrosis.Transforming growth factor(TGF)-β1has been widely recognized as a keyfibrogenic cytokine.The major signaling pathway of TGF--β1is through cytoplas-mic Smad proteins.Our study investigated the role of individual TGF--β1/Smadsignal proteins in mediating α-smooth muscle actin(α-SMA)gene expression,which is a well-known key marker of myofibroblast differentiation.Methods:Wetransiently cotransfected α-SMA promoter-luciferase fusion plasmid(p895-Luc)and Smad expression plasmids and measured Luc activity in TGF--β1-treated hu-man fetal lung fibroblasts.We induced Smad3 knockout mice lung fibrosis bybleomycin,α-SMA protein expression was assessed by Western blotting.Col-lagen protein was analyzed by measuring hydroxyprolin.Myofibroblast morphol-ogy was assessed by immunohistochemistry.Results:We found that theoverexpression of Smad3,not Smad2 markedly increased TGF--β1-induced α-SMApromoter activity and α-SMA protein expression in vitro,whereas the over-expression of dominant negative mutant Smad3 and Smad7 repressed TGF--β1-induced α-SMA gene expression.Compared to wild-type mice,Smad3 knockoutmice showed attenuated lung fibrosis after bleomycin treatment,manifested bylower collagen production and myofibroblast differentiation.Conclusion:Ourstudy suggested TGF--β1/Smad3 is a major pathway which regulated themyofibroblast differentiation.This result indicates a potential significance forfuture attempts of attenuating the progression of human lung fibrosis by theinhibition of the Smad3 cascade.