Nitric oxide attenuates epithelial-mesenchymal transition in alveolar epithelial cells

Patients with interstitial lung diseases, such as idiopathic pulmonary fibrosis ( IPF) and bronchopulmonary dysplasia ( BPD), suffer from lung fibrosis secondary to myofibroblast- mediated excessive ECM deposition and destruction of lung architecture. Transforming growth factor ( TGF)-beta 1 induces epithelial- mesenchymal transition ( EMT) of alveolar epithelial cells ( AEC) to myofibroblasts both in vitro and in vivo. Inhaled nitric oxide ( NO) attenuates ECM accumulation, enhances lung growth, and decreases alveolar myofibroblast number in experimental models. We therefore hypothesized that NO attenuates TGF-beta 1-induced EMT in cultured AEC. Studies of the capacity for endogenous NO production in AEC revealed that endothelial nitric oxide synthase ( eNOS) and inducible nitric oxide synthase ( iNOS) are expressed and active in AEC. Total NOS activity was 1.3 pmol . mg protein(-1) . min(-1) with 67% derived from eNOS. TGF-beta 1 ( 50 pM) suppressed eNOS expression by more than 60% and activity by 83% but did not affect iNOS expression or activity. Inhibition of endogenous NOS with L- NAME led to spontaneous EMT, manifested by increased alpha-smooth muscle actin ( alpha-SMA) expression and a fibroblastlike morphology. Provision of exogenous NO to TGF-beta 1-treated AEC decreased stress fiber- associated alpha-SMA expression and decreased collagen I expression by 80%. NO-treated AEC also retained an epithelial morphology and expressed increased lamellar protein, E-cadherin, and pro- surfactant protein B compared with those treated with TGF-beta alone. These findings indicate that NO serves a critical role in preserving an epithelial phenotype and in attenuating EMT in AEC. NO- mediated regulation of AEC fate may have important implications in the pathophysiology and treatment of diseases such as IPF and BPD.