Introduction: We previously identified that the activity of Factor Inhibiting HIF (FIH), a major regulator of HIF, is reduced in human lung fibroblasts from patients with IPF and that this promotes increased tissue stiffness. Here we investigated the transcriptomic and metabolic consequences of the loss of activity of FIH within lung fibroblasts.

Methods: We performed siRNA-mediated knockdown and CRISPR/Cas9-mediated knockout of FIH within human lung fibroblasts. The global transcriptome was determined by bulk RNA-seq and Seahorse real-time cell metabolic analysis was used to investigate effects upon glycolysis and mitochondrial function.

Results: Global transcriptomic analysis following FIH depletion in human lung fibroblasts identified 2,987 differential expressed genes (DEGs). Hallmark pathway enrichment analysis revealed that the glycolysis pathway was enriched in both up-regulated (p=1.9E-10) and down-regulated DEGs (p=7.7E-10), meanwhile, oxidative phosphorylation pathway is enriched in down-regulated DEGs (p=5.6E-11). Seahorse analysis suggested that loss of FIH reduced basal respiration oxygen consumption rate (OCR) and maximal respiration OCR in a HIF-dependent manner. In contrast the effects of FIH on basal extracellular acidification rate (ECAR) and glycolysis capacity ECAR was HIF-independent.

Conclusion: Our results identify that FIH activity has an important role in the transcriptional regulation of lung fibroblasts. Furthermore, we identify evidence that FIH may function as a metabolic switch in lung fibroblasts via both HIF-dependent and HIF-independent mechanisms. Further investigation into the consequences of these findings within human lung fibrosis is ongoing.