Subphenotype-specific serum proteomic signatures in patients with ARDS

Abstract Background Acute respiratory distress syndrome (ARDS) is associated with a substantial burden of morbidity and mortality. Previous studies have identified reproducible hyperinflammatory and hypoinflammatory ARDS subphenotypes with distinct clinical outcomes and treatment responses. Although recent transcriptomic studies have provided insights into biological differences between these subphenotypes, the role of the circulating proteome remains incompletely understood. Methods In this prospective study, patients with ARDS were classified into hyperinflammatory or hypoinflammatory subphenotypes using a previously validated 3-variable classifier based on bicarbonate, interleukin-6, and soluble tumor necrosis factor receptor 1. Serum proteomic profiling was performed to identify differentially abundant proteins between subphenotypes. Biological pathways were examined using gene set enrichment analysis. To provide cellular expression context for the subphenotype-associated serum proteins, serum proteomic data were mapped onto publicly available single-cell RNA-seq datasets from peripheral blood mononuclear cells, neutrophils, and tracheal aspirates. Results Among 31 patients with ARDS, 13 (42%) were classified as having the hyperinflammatory subphenotype and 18 (58%) as having the hypoinflammatory subphenotype. Both 28-day and in-hospital mortality were higher in the hyperinflammatory subphenotype than in the hypoinflammatory subphenotype (46% vs. 11%, P=0.04; 62% vs. 11%, P=0.01, respectively). In total, 54 serum proteins were differentially abundant between subphenotypes. These proteins represented biologically relevant categories, including endothelial injury, immunoregulation, coagulation/fibrinolysis, complement regulation, tissue remodeling, and metabolism-related proteins. Gene set enrichment analysis identified pathways differentially enriched between subphenotypes, including cell adhesion, cell–cell signaling, and cytokine responses (all FDR < 0.10). In external single-cell RNA-seq reference datasets, genes encoding the subphenotype-associated proteins were expressed predominantly in CD14 and CD16 monocytes in PBMCs, showed distinct expression patterns across immature and CD163+IL1R2^hi^ circulating neutrophil states, and were expressed mainly in macrophage and monocyte populations, with additional epithelial expression, in tracheal aspirates. Conclusions Serum proteomic profiling identified distinct protein patterns associated with established ARDS inflammatory subphenotypes, providing insights into the biological heterogeneity of ARDS.