Mitochondrial proteomics reveals reductive metabolism dependent on glutamine in fibroblasts of idiopathic pulmonary fibrosis under hypoxia.

Pulmonary fibrosis, particularly idiopathic pulmonary fibrosis (IPF), is a chronic disease characterized not only by a transcriptionally active signature associated with hypoxia but also by feedback loops that may underlie disease progression. IPF fibroblasts are known to contribute to disease by the pronounced differentiation of fibroblasts into myofibroblasts, which results in the accumulation of excessive extracellular matrix, creating a hypoxic microenvironment that supports the characteristic phenotype observed in the fibroblasts of these patients. Although several changes have been linked to fibroblast metabolism, the hypoxic conditions in mitochondria generally go unreported. This study aimed to characterize the differences in the mitochondrial proteomic profile between healthy lung fibroblasts and those affected by IPF under hypoxic conditions. We isolated mitochondria and validated the extraction of mitochondrial proteins using electron microscopy and western blotting. Subsequently, we performed label free proteomic analysis to obtain a proteomic profile and validated. Our results revealed that in controls, there is a metabolism of fatty acids and acetyl-CoA regulation, with a slight increase in mitophagy to utilize different substrates as energy sources in an appropriate response to low oxygen conditions. On the other hand, the mitochondria of fibroblasts from patients with IPF show a particular adaptation in glutamine metabolism, which may participate in its commitment to myofibroblast differentiation and the alteration in collagen production. These results allow us to visualize the importance of the proteins that we found deregulated or altered in the mitochondrial context. This helps us to have a basis for future research on their function and possible participation in specific biological processes, especially in the fibrotic process.