Li Gao1,2, Dingyuan Jiang3, Jing Geng3, Run Dong4, Huaping Dai1,3. 1. Department of Pulmonary and Critical Care Medicine, Beijing Chao-Yang Hospital, Capital Medical University, Beijing, China. 2. Department of Pulmonary and Critical Care Medicine, Inner Mongolia Autonomous Region People's Hospital, Hohhot, China. 3. Department of Pulmonary and Critical Care Medicine, Center of Respiratory Medicine, China-Japan Friendship Hospital, National Clinical Research Center for Respiratory Diseases, Beijing, China. 4. Department of Respiratory Medicine, Zhengzhou Central Hospital, Zhengzhou, China.
Abstract
NEW FINDINGS: • What is the central question of this study? The aim was to explore the effects and underlying mechanisms of H2 on bleomycin-induced pulmonary fibrosis. • What are the main findings and its importance? Our results indicate that, in bleomycin-induced pulmonary fibrosis, H2 inhalation attenuated oxidative stress and reversed the pulmonary epithelial-to-mesenchymal transition process by reducing reactive oxygen species production and inhibiting the expression of transforming growth factor-β1, α-smooth muscle actin and collagen I to improve fibrotic injury and exert anti-fibrogenic effects. Thus, H2 inhalation has promising therapeutic potential as a useful adjuvant treatment for patients with idiopathic pulmonary fibrosis, which deserves further study and evaluation. ABSTRACT: Hydrogen (H2 ) can protect against tissue damage. The effect of H2 inhalation therapy on the pathogenesis of pulmonary fibrosis remains unknown. This study was designed to explore the effects and underlying mechanisms of H2 inhalation on bleomycin (BLM)-induced pulmonary fibrosis. A rat model of pulmonary fibrosis was established with BLM. Rats were randomly divided into control and H2 inhalation groups. Haematoxylin and Eosin staining and Mason's Trichrome staining were performed to evaluate pulmonary fibrosis injury, inflammatory cell infiltration, structural disorder and collagen deposition. qRT-PCR and western blot assays were used to determine the expression of TNF-α, TGF-β1, α-SMA, E-cadherin, N-cadherin, vimentin, VEGF and collagen type I at both mRNA and protein levels. The contents of reactive oxygen species, TGF-β1, TNF-α, malondialdehyde and hydroxyproline were determined with biochemical test kits or ELISA kits. Bleomycin-stimulated rats exhibited typical symptoms of pulmonary fibrosis, which featured an increase in collagen deposition, alveolitis, fibrosis and parenchymal structural disorder in the lung. However, BLM-induced oxidative stress was attenuated by H2 inhalation therapy, which reduced the contents of reactive oxygen species, malondialdehyde and hydroxyproline, enhanced the activity of glutathione peroxidase and decreased the expression of TGF-β1 and TNF-α. In addition, H2 inhalation also inhibited BLM-induced epithelial-to-mesenchymal transition by inhibiting TGF-β1, increasing the expression level of the epithelial cell marker E-cadherin, and decreasing the expression level of the mesenchymal cell marker vimentin in a time-dependent manner. In addition, H2 inhalation downregulated α-SMA expression and suppressed collagen I generation, exerting anti-fibrogenic effects. Hydrogen inhalation therapy attenuates BLM-induced pulmonary fibrosis by inhibiting TGF-β1, relevant oxidative stress and epithelial-to-mesenchymal transition.
NEW FINDINGS: • What is the central question of this study? The aim was to explore the effects and underlying mechanisms of H2 on bleomycin-induced pulmonary fibrosis. • What are the main findings and its importance? Our results indicate that, in bleomycin-induced pulmonary fibrosis, H2 inhalation attenuated oxidative stress and reversed the pulmonary epithelial-to-mesenchymal transition process by reducing reactive oxygen species production and inhibiting the expression of transforming growth factor-β1, α-smooth muscle actin and collagen I to improve fibrotic injury and exert anti-fibrogenic effects. Thus, H2 inhalation has promising therapeutic potential as a useful adjuvant treatment for patients with idiopathic pulmonary fibrosis, which deserves further study and evaluation. ABSTRACT: Hydrogen (H2 ) can protect against tissue damage. The effect of H2 inhalation therapy on the pathogenesis of pulmonary fibrosis remains unknown. This study was designed to explore the effects and underlying mechanisms of H2 inhalation on bleomycin (BLM)-induced pulmonary fibrosis. A rat model of pulmonary fibrosis was established with BLM. Rats were randomly divided into control and H2 inhalation groups. Haematoxylin and Eosin staining and Mason's Trichrome staining were performed to evaluate pulmonary fibrosis injury, inflammatory cell infiltration, structural disorder and collagen deposition. qRT-PCR and western blot assays were used to determine the expression of TNF-α, TGF-β1, α-SMA, E-cadherin, N-cadherin, vimentin, VEGF and collagen type I at both mRNA and protein levels. The contents of reactive oxygen species, TGF-β1, TNF-α, malondialdehyde and hydroxyproline were determined with biochemical test kits or ELISA kits. Bleomycin-stimulated rats exhibited typical symptoms of pulmonary fibrosis, which featured an increase in collagen deposition, alveolitis, fibrosis and parenchymal structural disorder in the lung. However, BLM-induced oxidative stress was attenuated by H2 inhalation therapy, which reduced the contents of reactive oxygen species, malondialdehyde and hydroxyproline, enhanced the activity of glutathione peroxidase and decreased the expression of TGF-β1 and TNF-α. In addition, H2 inhalation also inhibited BLM-induced epithelial-to-mesenchymal transition by inhibiting TGF-β1, increasing the expression level of the epithelial cell marker E-cadherin, and decreasing the expression level of the mesenchymal cell marker vimentin in a time-dependent manner. In addition, H2 inhalation downregulated α-SMA expression and suppressed collagen I generation, exerting anti-fibrogenic effects. Hydrogen inhalation therapy attenuates BLM-induced pulmonary fibrosis by inhibiting TGF-β1, relevant oxidative stress and epithelial-to-mesenchymal transition.