A74-41 Pyruvate Anaplerosis Determines Fibroblast Collagen Output in Lung Fibrosis

Abstract Background Idiopathic pulmonary fibrosis (IPF) features bioenergetic failure with ATP depletion and a rerouting of pyruvate away from mitochondrial oxidation. In fibroblasts, pyruvate dehydrogenase kinase 1 (PDK1) inhibits pyruvate dehydrogenase (PDH), limiting oxidative entry of pyruvate into the tricarboxylic acid (TCA) cycle and biasing flux toward pyruvate carboxylase (PC)-mediated replenishment of TCA metabolites (a process termed anaplerosis). We investigated whether this switch toward pyruvate anaplerosis is a key determinant of pathologic fibroblast activation and collagen output. Methods We performed metabolomics on control and IPF lung tissue to quantify ATP and glycolytic intermediates; measured oxygen consumption (OCR) and extracellular acidification (ECAR) in control and IPF fibroblasts under substrate-defined conditions to assess pyruvate-supported respiration and glycolysis; and conducted laser-capture proteomics of the pathognomonic fibroblastic foci. We queried public lung datasets for PDK1 expression and assessed Pdk1 as well as Pc in fibroblasts isolated after bleomycin injury in mice. To modulate pyruvate fate, we inhibited PDK with dichloroacetate (DCA) and quantified PDH phosphorylation, mitochondrial energetics, and collagen protein and mRNA. Results IPF lungs showed reduced ATP with elevated glycolysis and lactate. Compared with control cells, IPF fibroblasts exhibited reduced pyruvate-supported OCR with compensatory increases in ECAR, indicating impaired mitochondrial pyruvate utilization. Fibroblastic foci were enriched for glycolytic enzymes and relatively depleted of fatty acid β-oxidation proteins. Analysis of public datasets localized increased PDK1 to fibroblastic regions; fibroblasts from bleomycin-injured mice displayed higher Pdk1 and Pc. PDK inhibition in lung fibroblasts in vitro activated PDH, improved mitochondrial energetics, and markedly reduced steady-state collagen protein. Notably, collagen mRNA changed minimally, indicating a disconnect between transcription and protein abundance consistent with metabolite-dependent, translational/post-translational control. Conclusions In IPF fibroblasts, mitochondrial routing of pyruvate—oxidation versus PC-mediated anaplerosis—serves as the proximal control point for collagen output. Redirecting pyruvate into oxidative metabolism with PDK inhibition restores bioenergetics and lowers collagen despite minimal transcript change, implicating TCA-intermediate-dependent, translational/post-translational control of collagen. Because dichloroacetate is a clinically explored PDK inhibitor, this pathway is immediately actionable and merits evaluation as an antifibrotic strategy in IPF. This abstract is funded by: NIH