Abstract
BackgroundEpithelial cell death is a major contributor to fibrogenesis in the lung. In this study, we sought to determine the function of mitochondria and their clearance (mitophagy) in alveolar epithelial cell death and fibrosis.MethodsWe studied markers of mitochondrial injury and the mitophagy marker, PTEN-induced putative kinase 1 (PINK1), in IPF lung tissues by Western blotting, transmission electron microscopy (TEM), and immunofluorescence. In vitro experiments were carried out in lung epithelial cells stimulated with transforming growth factor-β1 (TGF-β1). Changes in cell function were measured by Western blotting, flow cytometry and immunofluorescence. In vivo experiments were performed using the murine bleomycin model of lung fibrosis.ResultsEvaluation of IPF lung tissue demonstrated increased PINK1 expression by Western blotting and immunofluorescence and increased numbers of damaged mitochondria by TEM. In lung epithelial cells, TGF-β1 induced mitochondrial depolarization, mitochondrial ROS, and PINK1 expression; all were abrogated by mitochondrial ROS scavenging. Finally, Pink1 -/- mice were more susceptible than control mice to bleomycin induced lung fibrosis.ConclusionTGF-β1 induces lung epithelial cell mitochondrial ROS and depolarization and stabilizes the key mitophagy initiating protein, PINK1. PINK1 ameliorates epithelial cell death and may be necessary to limit fibrogenesis.
Highlights
Mitochondria play a vital metabolic role but can cause cellular damage via the toxic byproducts of oxidative phosphorylation[1,2]
transforming growth factor-β1 (TGF-β1) induced mitochondrial depolarization, mitochondrial ROS, and PINK1 expression; all were abrogated by mitochondrial ROS scavenging
PINK1 accumulation on the outer membrane of mitochondria is the key signal for triggering mitophagy[8]
Summary
Mitochondria play a vital metabolic role but can cause cellular damage via the toxic byproducts of oxidative phosphorylation[1,2]. Damaged mitochondria trigger apoptosis through release of cytochrome c into the cytosol[3] and activation of cell death pathways. Cellular survival depends upon efficient clearance of dysfunctional mitochondria. A selective form of autophagy, dubbed mitophagy, is a major avenue for the removal of these mitochondria. The pathways for generating mitophagy overlap considerably with those required for general autophagy; in mammals, mitochondriaspecific mechanisms have been recently identified and are primarily mediated by PTENInduced Putative Kinase 1 (PINK1)[4] and the E3 ubiquitin ligase, Parkin[5]. Epithelial cell death is a major contributor to fibrogenesis in the lung. We sought to determine the function of mitochondria and their clearance (mitophagy) in alveolar epithelial cell death and fibrosis
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