Abstract Number: Esoc2026a633 Proteomic Profiling of Circulating Nervous and Immune Systems-Derived Extracellular Vesicles as Biomarkers of Intracerebral Haemorrhage Pathophysiology

Abstract Background and aims Extracellular vesicles (EVs) are emerging as biomarkers reflecting dynamic biological responses after intracerebral haemorrhage (ICH). Building on our previous work showing that EVs are involved in tissue injury and repair, we aimed to characterize the temporal proteomic signatures of nervous system (nsEVs) and immune system–derived EVs (isEVs) throughout ICH evolution, to refine the specific contribution of both systems to the pathophysiology of the disease. Methods EVs from rats subjected to striatal ICH (n=10) were isolated from blood samples collected at baseline, 24h, 48h, 72h, 7d, and 28d. nsEVs (neurons/astrocytes/microglia) and isEVs (T-cells/monocytes/macrophages) were separated by immunoprecipitation using cell-specific antibodies. Proteins were identified by DIA-SWATH-mass-spectrometry. Temporal protein dynamics were assessed, differential abundance defined as fold-change2 with adjusted p0.05, and functional enrichment was analyzed. Results We identified 1,183 proteins in nsEVs and isEVs, of which 48 showed significant changes across timepoints. nsEVs exhibited earliest and most pronounced changes at 24h, involving metabolism, synaptic and ionic regulation, inflammation, vascular responses, and cell death. At 28d, nsEVs presented ribosomal protein modulation and declining stress mediators, suggesting homeostatic recovery. In isEVs changes peaked at 7d, involving metabolic reprogramming, antioxidant defense, extracellular matrix remodeling, immune modulation, and synaptic plasticity. Despite different temporal trajectories, both EV subtypes shared core biological processes. Conclusions The nsEVs and isEVs proteomes show distinct temporal profile along the course of ICH, reflecting a time-shifted neural–immune responses. These findings provide functional insight into ICH pathophysiology and a framework to guide the investigation of new biomarkers and therapeutic candidates. Conflict of interest Gallego Ruiz, Rebeca: nothing to disclose. Pozo Novoa, Javier: nothing to disclose. Laso García, Fernando: nothing to disclose. Calzado González, Ángela: nothing to disclose. Rodríguez Pardo, Jorge: nothing to disclose. Casado Fernández, Laura: nothing to disclose. Bravo, Susana: nothing to disclose. Díez Tejedor, Exuperio: nothing to disclose. Alonso de Leciñana, María: nothing to disclose. Gutiérrez Fernández, María: nothing to disclose.