Serum EV-enriched proteomics reveals host-response subtypes and molecular similarity to organ dysfunction phenotypes in sepsis.

Introduction: Sepsis is a heterogeneous syndrome characterized by dysregulated host responses and diverse patterns of organ dysfunction. This study aimed to characterize serum extracellular vesicle (EV)-enriched proteomic heterogeneity in sepsis and assess whether proteomic subtypes identified among Sepsis-only patients showed molecular similarity to established organ dysfunction phenotypes. Methods: In this single-center observational study, serum EV-enriched preparations were analyzed from 250 patients with sepsis and 50 healthy controls enrolled between June 2021 and December 2022. ARDS and AKI status was determined at blood sampling, and patients with neither condition were classified as Sepsis-only. After proteomic quality control, 231 patients and 48 controls were included. Non-negative matrix factorization was used to identify proteomic subtypes among Sepsis-only patients. Centroid projection and clinically adjusted sensitivity analyses evaluated cross-sectional molecular similarity between these subtypes and established S-AKI, S-ARDS, and S-ARDS&AKI phenotypes. An exploratory elastic-net protein panel was derived, and the downstream supervised workflow was evaluated using repeated nested cross-validation conditional on the original subtype labels. Results: A total of 4,754 protein groups were quantified across 279 samples. Three proteomic subtypes were identified among 96 Sepsis-only patients. S1 showed less pronounced inflammatory features, whereas S2 was characterized by cellular immune and metabolic-stress pathways and S3 by humoral immune, complement/coagulation, hemostatic, and acute-phase processes. These molecular patterns were not fully captured by conventional severity scores. The organ dysfunction groups showed partial and reference-dependent molecular similarity to S3 after clinical adjustment. A hypothesis-generating 15-protein panel captured features shared by S3 and established organ dysfunction relative to S2. Conclusions: Serum EV-enriched proteomics revealed distinct host-response subtypes within the Sepsis-only group and contemporaneous molecular features shared with established organ dysfunction. These findings advance the molecular characterization of sepsis heterogeneity and provide a focused framework for future longitudinal and external validation.