Transforming growth factor-β1 induces heparan sulfate 6-O-endosulfatase 1 expression in vitro and in vivo

Transforming growth factor (TGF)-beta 1 plays an important role in the development of pulmonary fibrosis. In this study we examined the relationship between TGF-beta 1 stimulation and the expression of heparan sulfate (HS) 6-O-endosulfatase 1 (Sulf1) in cultured normal human lung fibroblasts (NHLFs) and in murine lungs in vivo. By removing 6-O-sulfates from specific HS intrachain sites on the cell surface, Sulf1 has been shown to modulate the activities of many HS binding growth factors and morphogens including fibroblast growth factor (FGF)-2. Real time reverse transcription-PCR analysis revealed that TGF-beta 1 increased Sulf1 expression in NHLFs in a dose- and time-dependent manner which was accompanied by a decrease in 6-O-sulfated disaccharides as revealed by high performance liquid chromatography analysis. Decreased ERK activation after FGF-2 stimulation was observed in TGF-beta 1-treated NHLFs compared with control cells without changes in HS-dependent FGF-2 binding or FGF-2 center dot FR1c complex formation. To study the function of Sulf1, negative control or Sulf1-specific small interference RNA (siRNA)-transfected NHLFs were stimulated with TGF-beta 1. Enhanced Smad2/3 phosphorylation and elevated total Smad2 protein level were observed in Sulf1 siRNA-transfected cells and were accompanied by enhanced expression of alpha-smooth muscle actin and fibronectin. In addition, Sulf1 siRNA transfection enhanced the anti-proliferative effect of TGF-beta 1. Finally Sulf1 expression was up-regulated in the lungs of mice treated with adenovirus encoding active TGF-beta 1. Taken together, our data indicate that Sulf1 is a TGF-beta 1-responsive gene both in vitro and in vivo and may function as a negative regulator of TGF-beta 1-induced fibrogenesis.