Neonatal Hyperoxia Induces Metabolic Reprogramming in Senescent Alveolar Macrophages, Leading to Persistent Lung Injury.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is a chronic lung disease in premature infants. Neonatal hyperoxia induces a BPD-like phenotype and lung cell senescence in rodents. In our 3-day hyperoxia model, senescent cells were predominantly lung macrophages, with their abundance peaking at postnatal day 7 (pnd7). However, the molecular and functional characteristics of these senescent macrophages remain undefined. METHODS: We reanalyzed a scRNA-seq dataset (GSE207866) generated from senescent lung cells isolated at pnd7 (SD7) following neonatal hyperoxia. Hierarchical clustering combined with manual annotation was used to compare transcriptional profiles with age-matched air-exposed controls (AirD7) and hyperoxia-exposed mice without senescent-cell enrichment (O2D7). Key molecular findings were validated by immunofluorescence. In vivo, neonatal mice received daily injections of the pyruvate dehydrogenase kinase inhibitor, dichloroacetate (DCA) from pnd4 to pnd6, and a senolytic cocktail consisting of quercetin and dasatinib from pnd4 to pnd14, following 3 days of hyperoxia exposure. RESULTS: Macrophages accounted for 65.90% of senescent cells in the SD7 group. Seven macrophage clusters were identified, enriched in M1-like and alveolar macrophage phenotypes. Two major clusters (clusters 0 and 1), together representing nearly half of all senescent macrophages, exhibited strong expression of genes associated with innate immunity, inflammation, and DNA damage responses. These clusters also showed a shift toward glycolysis, the pentose phosphate pathway, and glutamine metabolism, with reduced reliance on β-oxidation. Administration of DCA activated pyruvate dehydrogenase and attenuated hyperoxia-induced macrophage senescence and lung injury. Pathway enrichment analyses revealed enhanced metal-handling pathways, immune and stress signaling (including p38 mitogen-activated kinase, ataxia-telangiectasia mutated, and mechanistic target of rapamycin), apoptosis, and RNA regulatory processes. Conversely, genes involved in reactive oxygen species detoxification, DNA repair, phagocytosis, cytoskeletal organization, and cell adhesion were downregulated. Notably, reducing senescent cells by a senolytic cocktail during the alveolar stage mitigated hyperoxia-induced persistent lung injury. CONCLUSION: Neonatal hyperoxia drives the emergence of a heterogeneous population of senescent macrophages characterized by metabolic reprogramming and dysregulated signaling pathways, which contribute to the development and persistence of lung injury.