Inhibition and Role of let-7d in Idiopathic Pulmonary Fibrosis

• Published in: American journal of respiratory and critical care medicine 2010-07, Vol.182 (2), p.220-229
• Main Authors: PANDIT, Kusum V, CORCORAN, David, RICHARDS, Thomas, SELMAN, Moises, WATKINS, Simon C, PARDO, Annie, BEN-YEHUDAH, Ahmi, BOUROS, Demosthenes, EICKELBERG, Oliver, RAY, Prabir, BENOS, Panayiotis V, KAMINSKI, Naftali, YOUSEF, Hanadie, YARLAGADDA, Manohar, TZOUVELEKIS, Argyris, GIBSON, Kevin F, KONISHI, Kazuhisa, YOUSEM, Samuel A, SINGH, Mandal, HANDLEY, Daniel
• Format: Article
• Language: English
• Subjects: Actins - metabolism; Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy; Animals; Biological and medical sciences; Cadherins - metabolism; Cells, Cultured; Down-Regulation; Epithelial Cells - metabolism; F. Interstitial Lung Disease; HMGA2 Protein - metabolism; Humans; Idiopathic Pulmonary Fibrosis - metabolism; Idiopathic Pulmonary Fibrosis - pathology; In Situ Hybridization; Intensive care medicine; Lung - metabolism; Medical sciences; Mice; Mice, Inbred C57BL; MicroRNAs - metabolism; Pneumology; Polymerase Chain Reaction; Pulmonary Alveoli - metabolism; Pulmonary hypertension. Acute cor pulmonale. Pulmonary embolism. Pulmonary vascular diseases; S100 Calcium-Binding Protein A4; S100 Proteins - metabolism; Smad3 Protein - metabolism; Transforming Growth Factor beta - physiology; Vimentin - metabolism
• ISSN: 1073-449X; 1535-4970
• DOI: 10.1164/rccm.200911-1698oc

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and usually lethal fibrotic lung disease characterized by profound changes in epithelial cell phenotype and fibroblast proliferation. To determine changes in expression and role of microRNAs in IPF. RNA from 10 control and 10 IPF tissues was hybridized on Agilent microRNA microarrays and results were confirmed by quantitative real-time polymerase chain reaction and in situ hybridization. SMAD3 binding to the let-7d promoter was confirmed by chromatin immunoprecipitation, electrophoretic mobility shift assay, luciferase assays, and reduced expression of let-7d in response to transforming growth factor-beta. HMGA2, a let-7d target, was localized by immunohistochemistry. In mice, let-7d was inhibited by intratracheal administration of a let-7d antagomir and its effects were determined by immunohistochemistry, immunofluorescence, quantitative real-time polymerase chain reaction, and morphometry. Eighteen microRNAs including let-7d were significantly decreased in IPF. Transforming growth factor-beta down-regulated let-7d expression, and SMAD3 binding to the let-7d promoter was demonstrated. Inhibition of let-7d caused increases in mesenchymal markers N-cadherin-2, vimentin, and alpha-smooth muscle actin (ACTA2) as well as HMGA2 in multiple epithelial cell lines. let-7d was significantly reduced in IPF lungs and the number of epithelial cells expressing let-7d correlated with pulmonary functions. HMGA2 was increased in alveolar epithelial cells of IPF lungs. let-7d inhibition in vivo caused alveolar septal thickening and increases in collagen, ACTA2, and S100A4 expression in SFTPC (pulmonary-associated surfactant protein C) expressing alveolar epithelial cells. Our results indicate a role for microRNAs in IPF. The down-regulation of let-7d in IPF and the profibrotic effects of this down-regulation in vitro and in vivo suggest a key regulatory role for this microRNA in preventing lung fibrosis. Clinical trial registered with www.clinicaltrials.gov (NCT 00258544).