Ligand-independent transforming growth factor-ß type I receptor signalling mediates type I collagen-induced epithelial-mesenchymal transition

Evidence suggests epithelialmesenchymal transition (EMT) as one potential source of fibroblasts in idiopathic pulmonary fibrosis. To assess the contribution of alveolar epithelial cell (AEC) EMT to fibroblast accumulation in vivo following lung injury and the influence of extracellular matrix on AEC phenotype in vitro, Nkx2.1-Cre;mT/mG mice were generated in which AECs permanently express green fluorescent protein (GFP). On days 1721 following intratracheal bleomycin administration, similar to 4% of GFP-positive epithelial-derived cells expressed vimentin or a-smooth muscle actin (a-SMA). Primary AECs from Nkx2.1-Cre;mT/mG mice cultured on laminin-5 or fibronectin maintained an epithelial phenotype. In contrast, on type I collagen, cells of epithelial origin displayed nuclear localization of Smad3, acquired spindle-shaped morphology, expressed a-SMA and phospho-Smad3, consistent with activation of the transforming growth factor-beta (TGF beta) signalling pathway and EMT. a-SMA induction and Smad3 nuclear localization were blocked by the TGF beta type I receptor (T beta RI, otherwise known as Alk5) inhibitor SB431542, while AEC derived from Nkx2.1-Cre;Alk5(flox/KO) mice did not undergo EMT on collagen, consistent with a requirement for signalling via Alk5 in collagen-induced EMT. Inability of a pan-specific TGF beta neutralizing antibody to inhibit effects of collagen together with absence of active TGF beta in culture supernatants is consistent with TGF beta ligand-independent activation of Smad signalling. These results support the notion that AECs can acquire a mesenchymal phenotype following injury in vivo and implicate type I collagen as a key regulator of EMT in AECs through signalling via Alk5, likely in a TGF beta ligand-independent manner. Copyright (C) 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.