Troglitazone Attenuates TGF-[beta]1-Induced EMT in Alveolar Epithelial Cells via a PPAR[gamma]-Independent Mechanism

Peroxisome proliferator activated receptor [gamma] (PPAR[gamma]) agonists are effective antifibrotic agents in a number of tissues. Effects of these agents on epithelial-mesenchymal transition (EMT) of primary alveolar epithelial cells (AEC) and potential mechanisms underlying effects on EMT have not been well delineated. We examined effects of troglitazone, a synthetic PPAR[gamma] agonist, on transforming growth factor (TGF)-[beta]1-induced EMT in primary rat AEC and an alveolar epithelial type II (AT2) cell line (RLE-6TN). TGF-[beta]1 (2.5 ng/mL) induced EMT in both cell types, as evidenced by acquisition of spindle-like morphology, increased expression of the mesenchymal marker [alpha]-smooth muscle actin ([alpha]-SMA) and downregulation of the tight junctional protein zonula occludens-1 (ZO-1). Concurrent treatment with troglitazone (or rosiglitazone), ameliorated effects of TGF-[beta]1. Furthermore, following stimulation with TGF-[beta]1 for 6 days, troglitazone reversed EMT-related morphological changes and restored both epithelial and mesenchymal markers to control levels. Treatment with GW9662 (an irreversible PPAR[gamma] antagonist), or overexpression of a PPAR[gamma] dominant negative construct, failed to inhibit these effects of troglitazone in AEC. Troglitazone not only attenuated TGF-[beta]1-induced phosphorylation of Akt and glycogen synthase kinase (GSK)-3[beta], but also inhibited nuclear translocation of [beta]-catenin, phosphorylation of Smad2 and Smad3 and upregulation of the EMT-associated transcription factor SNAI1. These results demonstrate inhibitory actions of troglitazone on TGF-[beta]1-induced EMT in AEC via a PPAR[gamma]-independent mechanism likely through inhibition of [beta]-catenin-dependent signaling downstream of TGF-[beta]1, supporting a role for interactions between TGF-[beta] and Wnt/[beta]-catenin signaling pathways in EMT.