Fig. 1.

Downregulation of FAO and upregulation of glycolysis in lung fibrosis. A Representative image showing α-SMA (green) and nuclei (blue) in rat fibroblasts from healthy and fibrotic lung (cells were isolated from SD rats after 28 days of PBS or bleomycin treatment). B FAO and glycolysis gene transcription alterations in rat fibroblasts from normal and fibrotic lung. mRNA levels of FAO and glycolysis genes are relative to healthy control (n = 3). C Western blot analysis of FAO and glycolysis protein in rat fibroblast from the groups indicated. D Representative images of Oil Red O staining of lung samples from healthy control and fibrotic lung. E Triglyceride abundance in the rat lung tissues indicated (n = 12). F Gating strategy for detecting LipidTOX ⁺ cells by flow cytometry and quantification of LipidTOX ⁺ cell abundance in rat fibroblast from healthy and fibrotic lung (n = 3). G Staining of lipid droplets in fibroblasts using LipidTOX (green). Nuclei were counterstained with DAPI (blue). H PKM2 staining of lung sections from the groups indicated. I Relative lactate production of lung sections from the groups indicated (n = 6). J Glucose consumption in rat fibroblast from healthy and fibrotic lung (n = 6). K Fatty acid oxidation (n = 4) in rat fibroblast from the healthy and fibrotic lung. Y-axis represents the oxygen consumption rate (OCR), a measure of metabolite oxidation over time. Palmitate (170μM), etomoxir (40μM) and oligomycin (2 μM). Etomoxir inhibits FAO by irreversibly binding to CPT-1A, which catalyzed the transport of fatty acids, such as palmitate, into the mitochondria for β-oxidation. L Glycolysis stress test (n = 4) in rat fibroblast from the healthy and fibrotic lung. Y-axis plots the extracellular acidification rate (ECR) which is a measure of glycolysis via the production of lactic acid. Glucose (10 mM), oligomycin (1 μM), and 2-DG (2-Deoxy-Glucose, 100 mM). ECAR, prior to glucose injection, is referred to as non-glycolytic acidification which is caused by other processes rather than glycolysis, and glucose-induced response is reported as the rate of glycolysis under basal conditions. M qRT-PCR of FAO and glycolysis genes are relative to control (n = 3). In vitro profiling of primary lung fibroblast with hypoxia (2 %), TGF-β1(5 ng/ml), and PGDF-BB (5 ng/ml). Bars represent 50 μm (A), 125 μm (D), 25 μm (G), 200 μm (H). The results are presented as the means ± S.D. (*p < 0.05, **P < 0.01; student's t-test). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)