Myeloid HIF-1α Couples Glycolytic Energy Supply with NCF2-Dependent Oxidative Killing to Protect Against Klebsiella pneumoniae Pneumonia

BACKGROUND: Hypervirulent Klebsiella pneumoniae (hvKp) induces severe pneumonia and sepsis. HIF-1α coordinates metabolic and immune responses in myeloid cells, but its role in hvKp-mediated pulmonary defense remains undefined. METHODS: Monocyte HIF-1α expression was assessed in patients with Klebsiella pneumoniae (K. pneumoniae) pneumonia. Myeloid-specific Hif-1α knockout mice and BMDMs were used to examine survival, bacterial burden, and macrophage function. RNA-seq, Seahorse flux analysis, and confocal microscopy were employed to investigate the regulatory effects of HIF-1α on phagocytosis and ROS production. The Hif-1α-NCF2-ROS signaling pathway was substantiated through the application of small interfering RNA (siRNA), JASPAR prediction tools, dual-luciferase reporter assays, chromatin immunoprecipitation followed by quantitative PCR (ChIP-qPCR), metabolic inhibitors, and dimethyloxalylglycine (DMOG), a prolyl hydroxylase inhibitor that stabilizes HIF-1α. RESULTS: Clinical data showed a negative correlation between monocyte HIF-1α levels and serum CRP, procalcitonin, ICU stay duration, and SOFA scores. In vivo, myeloid Hif-1α knockout mice demonstrated heightened susceptibility to hvKp, with markedly reduced survival and widespread bacterial dissemination. Mechanistically, Hif-1α-deficient macrophages displayed impaired phagocytosis, phagolysosomal maturation, and glycolytic reprogramming in response to infection. RNA-seq identified NCF2, encoding p67-phox, as a critical HIF-1α-dependent component of the NADPH oxidase complex. JASPAR prediction, dual-luciferase reporter assays, and ChIP-qPCR further demonstrated that NCF2 is directly transcriptionally regulated by HIF-1α. HIF-1α deficiency impaired both glycolytic ATP production and NCF2-mediated ROS generation, thereby compromising macrophage antibacterial activity. Inhibition of glycolysis or silencing NCF2 abolished HIF-1α-dependent defense, whereas pharmacological stabilization of HIF-1α using DMOG significantly enhanced host resistance. CONCLUSION: HIF-1α serves as a pivotal regulator of host defense in experimental hvKp pneumonia and is clinically associated with disease severity in K. pneumoniae pneumonia, linking glycolytic metabolism to the NCF2-ROS bactericidal pathway. These findings highlight the potential of targeting immunometabolic pathways to improve host defense against severe K. pneumoniae infections.