[Mechanism of MIF in regulating HIF-1α-mediated aerobic glycolysis in pulmonary fibrosis].

Objective To investigate the mechanism by which macrophage migration inhibitory factor (MIF) regulates hypoxia-inducible factor 1α (HIF-1α)-mediated aerobic glycolysis in the pathological process of pulmonary fibrosis. MethodsC57BL/6 mice were divided into four groups: control group, bleomycin (BLM) model group, BLM combined with negative control lentivirus (BLM combined with sh-NC) group, and BLM combined with MIF-shRNA lentivirus (BLM combined with sh-MIF) group. After 28 days of intratracheal administration, lung tissues, bronchoalveolar lavage fluid (BALF), and serum samples were collected. Pathological injury and the degree of fibrosis were assessed by hematoxylin and eosin (HE) staining and Masson's trichrome staining. Glycolytic capacity was evaluated using a lactate dehydrogenase (LDH) activity assay kit, and the end-product content of glycolysis was measured using a lactic acid (LA) assay kit. The protein expression of HIF-1α, 6-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase 3 (PFKFB3), and pyruvate kinase M2 (PKM2) was analyzed by Western blot. Primary mouse lung fibroblasts were divided into the Control group, recombinant MIF protein (rMIF) group, rMIF combined with PHD activator Octyl-α-ketoglutarate (rMIF combined with PHD-Act) group, MIF inhibitor (ISO-1) group, and PHD inhibitor MK-8617 (PHD-IN) groups; as well as the Control group, PHD-IN group, rMIF group, and rMIF combined with PHD-IN groups. The expression of HIF-1α and the protein levels of PHD were detected by RT-qPCR and Western blot, respectively. Cellular metabolic function was assessed by measuring the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) via a fluorescence-based assay kit. Results In animal experiments, compared with the Control group, the BLM group exhibited severe pulmonary fibrosis, along with significantly elevated LDH activity, lactate content, and protein expression of PFKFB3 and PKM2. Interference with MIF expression significantly reversed these changes and alleviated fibrosis. Cellular experiments showed that rMIF significantly upregulated HIF-1α and downregulated the expression of PHD1 and PHD2, an effect that could be reversed by the PHD activator Octyl-α-ketoglutarate. Inhibition of MIF reduced HIF-1α expression, whereas inhibition of PHD directly induced high expression of HIF-1α. Moreover, both rMIF and PHD-IN treatment significantly increased cellular OCR and ECAR. Conclusion MIF stabilizes HIF-1α by inhibiting PHD, thereby driving aerobic glycolysis metabolic reprogramming in lung fibroblasts and promoting the progression of pulmonary fibrosis.