The dynamic alternation of monocytes phenotypes and gene signature-related biomarkers in the airways and blood reveals myeloid subset-driven inflammation and lung fibrosis in hypercapnia following pulmonary sepsis- associated ARDS

Abstract

Abstract Background ARDS, a severe complication of sepsis, often exhibits hypercapnia, whose immunological mechanisms remain elusive. Our study aimed to investigate the pathogenesis of hypercapnia in pulmonary sepsis-associated ARDS and identify potential therapeutic targets. Methods Between December 2022 and April 2023, we conducted a single-center, prospective study with sixty-one severe pneumonia patients. Eleven patients without sepsis served as controls based on Sepsis-3 criteria. The remaining fifty patients with pulmonary sepsis met ESICM criteria for ARDS, with twenty-six developing hypercapnia. We comprehensively collected clinical data, respiratory parameters, and serial specimens of peripheral blood mononuclear cells (PBMCs) and bronchoalveolar lavage fluid (BALF) at early (within 24 hours) and late (Day 7) time points post-intubation. These specimens were analyzed using flow cytometry and a cytokine panel. Additionally, we performed single-cell RNA sequencing (scRNA-seq) on blood samples from four ARDS patients to characterize immunological and cellular profiles following pneumonia sepsis. Results Compared with non-hypercapnia ARDS patients and non-sepsis controls, septic associated-ARDS patients with hypercapnia exhibit poor prognosis and high mortality. To investigate their immunological response, we conducted scRNA-seq on PBMCs from four ARDS patients (one without hypercapnia and three with hypercapnia). Results showed a significant increase in monocytes, particularly classical monocytes (CD14+CD16−, CM), in hypercapnia patients. Flow cytometry analysis revealed a similar increase in CM throughout sepsis stages. This underscores the role of monocytes in hypercapnia ARDS. ScRNA-seq analysis also revealed elevated expression of cytokine storm-related genes, including IL-1β signaling, and genes linked to myeloid cell activation and recruitment. Elevated levels of IL-1β, IL-12p40, and IL-23 were observed in both blood and alveolar lavage fluid. Additionally, hypercapnia patients exhibited enrichment of profibrotic genes in monocytes and accumulation of monocyte-derived macrophages in airways, supported by clinical parameters, specifically the increased driving pressure and decreased static respiratory lung compliance, indicating pulmonary fibrosis. Conclusions We offer novel perspectives on monocyte-centered clusters and associated biomarkers, which play a pivotal role in driving hypercapnia-induced lung fibrosis. Our study provides fresh insights into the immunological mechanisms underlying hypercapnia in pulmonary sepsis-associated ARDS, laying the foundation for future therapeutic interventions to improve patient outcomes.