Pharmacological Research 57 (2008) 274–282 TGF-1 targets the GSK-3/-catenin pathway via ERK activation in the transition of human lung fibroblasts into myofibroblasts Filippo Caraci a , Elisa Gili b , Marco Calafiore a , Marco Failla b , Cristina La Rosa b , Nunzio Crimi b , Maria Angela Sortino c , Ferdinando Nicoletti d,e , Agata Copani a,f , Carlo Vancheri b,∗ a Department of Pharmaceutical Sciences, University of Catania, Viale Andrea Doria 6, 95125 Catania, Italy b Department of Internal and Specialistic Medicine, Section of Respiratory Medicine, University of Catania, Via Passo Gravina 187, 95125 Catania, Italy c Department of Experimental and Clinical Pharmacology, University of Catania, Viale Andrea Doria 6, 95125 Catania, Italy d Department of Human Physiology and Pharmacology, University of Rome La Sapienza, Piazzale Aldo Moro 5, 00185 Rome, Italy e I.N.M. Neuromed, Localit` a Camerelle, 86077 Pozzilli, Italy f I.B.B., CNR-Catania, Italy Accepted 4 February 2008 Abstract Transforming growth factor-1 (TGF-1) is known to induce the transition of human lung fibroblasts to myofibroblasts, a primary event in the pathogenesis of idiopathic pulmonary fibrosis. The molecular pathways involved in myofibroblast transformation are only partially identified. We found that a 24-h treatment with TGF-1 (10 ng/ml) induced -smooth actin (SMA) expression and collagen production in human lung fibroblasts. These effects were abrogated by PD98059, a specific inhibitor of the mitogen-activated protein kinase (MAPK) pathway. TGF-1 treatment activated the MAPK pathway, as shown by an increased phosphorylation of extracellular-regulated kinases (ERK)1/2 after 30min of exposure. TGF-1 also increased the expression of the Ser-9-phosphorylated inactive form of glycogen synthase kinase-3(GSK-3), an effect that was largely attenuated by PD98059. A nuclear translocation of -catenin in human lung fibroblasts was observed 2 h after TGF-1 addition both by confocal microscopy and nuclear protein analysis. At this time, TGF-1 also increased the total levels of -catenin, an effect that was prevented by PD98059. Similarly to TGF-1, the GSK-3inhibitor lithium chloride (10 mM), increased the total levels of -catenin and promoted -SMA expression and collagen production. This study demonstrates that TGF-1 induces -SMA expression and collagen production in human lung fibroblasts via ERK1/2 activation, GSK-3inhibition and nuclear -catenin translocation. The evidence that the silencing of -catenin by siRNAs was able to prevent the induction of -SMA expression in TGF-1-treated fibroblasts further supports the hypothesis of a contribution of the GSK-3/-catenin pathway in the pathogenesis of idiopathic pulmonary fibrosis. © 2008 Elsevier Ltd. All rights reserved. Keywords: TGF-1; ERK1/2; GSK-3; -Catenin; Fibroblast; -SMA 1. Introduction Idiopathic pulmonary fibrosis (IPF) is a chronic interstitial lung disease of unknown etiology characterized by increased fibroblastic proliferation and extracellular matrix remodeling resulting into a loss of lung function and, eventually, respira- ∗ Corresponding author. Tel.: +39 095 7594503. E-mail address: vancheri@unict.it (C. Vancheri). tory failure [1]. Recent insights suggest that foci of dysregulated fibroblasts driven by profibrotic cytokines, such as transform- ing growth factor-beta1 (TGF-1), are at the roots of IPF. In particular, TGF-1 is known to induce -smooth-muscle-actin (-SMA) expression and is considered by far the most potent inducer of myofibroblast transformation and activation [2,3]. Myofibroblasts are mainly found within areas of active pul- monary fibrosis termed fibroblastic foci [4], the extension of which correlates with disease progression and survival [5,6]. As such, the understanding of the molecular mechanisms respon- 1043-6618/$ – see front matter © 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.phrs.2008.02.001