PD-L1+ neutrophils trigger pulmonary endothelial pyroptosis via an oxidative phosphorylation-dependent mechanism in sepsis

Abstract Background Sepsis-induced acute respiratory distress syndrome (ARDS) is characterized by microvascular dysfunction, uncontrolled inflammation, and pulmonary edema, leading to high morbidity and mortality. Despite their clinical importance, targeted therapies are lacking. Neutrophils play a critical role in sepsis-induced lung injury, but the specific contributions of PD-L1+ neutrophils remain poorly understood. The aim of this study was to elucidate the role of PD-L1+ neutrophils in endothelial injury and the underlying mechanisms during sepsis. Methods We employed single-cell RNA sequencing (scRNA-seq) to analyze neutrophil heterogeneity in septic lungs and identified a distinct PD-L1 + neutrophil subpopulation. Using a murine model of sepsis induced by cecal ligation and puncture (CLP), we isolated PD-L1+ neutrophils and assessed their effects on pulmonary vascular endothelial cells (ECs) through coculture experiments. In vivo, PD-L1 was systemically neutralized using a neutralizing antibody to assess the impact of PD-L1 on lung injury and inflammatory responses. Results scRNA-seq revealed a unique PD-L1+ neutrophil subpopulation that infiltrated the lungs during sepsis. These neutrophils exacerbated endothelial pyroptosis, leading to vascular barrier dysfunction and increased inflammatory cytokine release. Mechanistically, PD-L1+ neutrophils exhibited a metabolic shift from oxidative phosphorylation (OXPHOS) to glycolysis, which amplified their proinflammatory effects. The systemic neutralization of PD-L1 significantly reduced pulmonary endothelial dysfunction, inflammatory responses, and lung injury in septic mice, as evidenced by decreased vascular permeability, reduced inflammatory cytokine levels, and improved histopathological injury. Conclusion This study demonstrated that PD-L1+ neutrophils play pivotal roles in driving endothelial pyroptosis and vascular injury during sepsis. The metabolic reprogramming of these neutrophils, characterized by a shift from OXPHOS to glycolysis, underlies their proinflammatory effects. Targeting PD-L1 or modulating metabolic pathways may offer novel therapeutic strategies to improve the clinical prognosis of sepsis-induced ARDS patients.