Follistatin-like1Is A Critical Regulator Of Lung Fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and often fatal lung disease of unknown etiology, characterized by (myo)fibroblasts accumulation and extracellular matrix (ECM) deposition. Fstl1(follistatin-like1), an inducible extracellular glycoprotein by transforming growth factor β1(TGF-β1), has been observed increased expression in lung tissues from IPF patients compared with controls by microarray analysis. In this study, we identified Fstll as a gene up-regulated in patients with IPF and in the bleomycin model of pulmonary fibrosis. Here, we hypothesized that Fstll has a potential role for the pathogenesis of IPF.To confirm this hypothesis, bleomycin was intra-tracheally administrated into Fstl1+/-mice and WT littermate controls to develop fibrosis. Compared with WT controls, Fstll+/-mice had a normal acute response to bleomycin injury, but they exhibited markedly less extent of fibrosis, as indicated by survival curve, histological analysis, hydroxyproline content, and immunoblot analysis.Exaggerated deposition and remodeling of ECM proteins leads ultimately to fibrosis and failure of the alveolar units. Hyploinsufficiency of Fstl1significantly reduced bleomycin-induced ECM components like collagen and fibronectin in lung tissues and primary pulmonary fibroblasts. Additionally, TGF-β1, implicated as the most potent inducer of ECM production, induced Fstll expression in mouse lung fibroblasts. We found reduced expression of ECM proteins in TGF-β1-treated fibroblasts from Fstl1+/-or Fstll lungs compared with WT controls, which suggested that TGF-β1-induced ECM production was dependent on the expression of Fstl1.Myofibroblast, the key effector cell of IPF, are the primary cell type responsible for the synthesis and deposition of ECM. In our murine model of bleomycin-induced lung fibrosis, we observed markedly decreased expression and immunostaining of a-SMA in lung tissues of Fstl1+/-mice compared with WT controls, suggesting that endogenous Fstll regulated myofibroblasts accumulation during fibrogenesis. TGF-β1has been implicated in fibroblast activation through a-SMA induction. Loss of Fstll prevented TGF-β1-induced a-SMA expression in primary mouse lung fibroblasts, which further suggested that Fstll is involved in local fibroblasts activation following bleomycin injury.Myofibroblasts involved in the fibrotic process is thought to originate from lung epithelial cells via epithelial-mesenchymal transition (EMT), a process frequently mediated by TGF-β.Examination by double staining of a-SMA and Pro-SPC revealed that haploinsufficiency of Fstll significantly reduced EMT progress in vivo. Further in vitro experiments in A549cells showed that over-expression of Fstl1increased TGF-β1-induced EMT via Smad-dependent and MAPK-dependent pathway in A549cells, as chemically selectively blocked Smad2/3, ERK or JNK phosphorylation led to diminish the alternation of EMT marker proteins. Finally, we first proved that Fstl1can directly bind to TGF-β1or TGF-β receptor Ⅱ (TβRⅡ) but not TGF-β receptor Ⅰ (TβRI) in vitro.Collectively, haploinsufficiency of Fstll markedly ameliorated bleomycin-induced pulmonary fibrosis partially by reduced ECM deposition and EMT progress but not influencing acute inflammation. And Fstll modulated TGF-β1-induced EMT through the interacting with TGF-β1-receptor complex and regulating the downstream Smad and MAPK. signaling. This study indicates a pivotal role for Fstll in the development of bleomycin-induced pulmonary fibrosis and suggests a novel therapeutic target for patients with pulmonary fibrosis.