Proteomic profiling and pathway analyses reveal molecular signatures and immune networks in pediatric sepsis

Background: Sepsis remains a leading cause of childhood mortality worldwide. Most deaths occur within the first few days of presentation, underscoring the urgent need for early recognition and biologically informed treatment strategies. The heterogeneous etiology of sepsis involves complex, intertwined biological networks, explaining why single-biomarker approaches have proven inadequate for capturing this complexity. We used high-throughput proximity extension assay technology to comprehensively profile plasma proteins in critically ill pediatric sepsis patients, aiming to identify dysregulated biological pathways that could inform risk stratification and therapeutic development. Methods: Study participants were prospectively enrolled based on established pediatric sepsis criteria. Plasma proteins were quantified using the Olink proximity extension assay, with differential expression, machine learning, and pathway enrichment analyses performed to define molecular signatures of pediatric sepsis. Results: Analysis of plasma samples from 17 pediatric sepsis patients and 17 age- and sex-matched healthy controls revealed 626 significantly differentially expressed proteins: 399 upregulated and 227 downregulated. The most significantly elevated proteins included calcitonin-related polypeptide α (CALCA), tumor necrosis factor superfamily member 14 (TNFSF14), and asialoglycoprotein receptor 1 (ASGR1). Machine learning identified a minimal 9-protein signature accounting for 90% of discriminatory power between groups. Pathway enrichment analysis revealed profound dysregulation of immune and inflammatory networks. Interleukin-10 (IL-10) signaling emerged as the most significantly enriched pathway, alongside extracellular matrix degradation, IL-4 and IL-13 signaling, and other cytokine signaling pathways. Dysregulated pathways were associated with clinical variables, particularly gram-negative infections and respiratory infection sources. Conclusions: Pediatric sepsis is characterized by dysregulation of multiple immune and inflammatory pathways rather than isolated protein abnormalities. IL-10 and related cytokine signaling emerged as central nodes, providing insights into the balance between hyperinflammation and immunosuppression in critically ill children. Associations between pathways and clinical variables suggest that specific pathogen types and infection sources trigger distinct patterns of biological network activation, offering potential targets for patient stratification and pathway-directed therapeutics