Abstract

Early neurological deterioration due to ischemic progression (ENDi) remains a poorly understood complication in acute ischemic stroke (AIS). We aim to uncover exosomal protein signatures for ENDi risk prediction and potential targets for intervention. From 110 prospectively enrolled AIS patients undergoing thrombolysis, we selected 3 ENDi and 5 age- and sex-matched early neurological improvement (ENI) patients. Plasma exosomes isolated before and 24 h post-thrombolysis were subjected to 4D-DIA proteomics, followed by Gene Set Enrichment Analysis (GSEA) and cellular origin analyses. Baseline differentially expressed proteins (DEPs) were further analyzed via functional enrichment and protein-protein interaction networks to identify candidate biomarkers and validated by ELISA in an independent cohort. We identified 1,238 exosomal proteins, among which 69 DEPs at baseline distinguished the ENDi from ENI patients. Cellular origin analysis revealed a post-thrombolytic reduction in monocyte-derived exosomal proteins in the ENDi group, while the ENI group showed upregulation of naive B cell-derived exosomal proteins. Regarding biological processes, ENDi after thrombolysis was characterized by activation of the coagulation and neuroinflammation pathways, along with suppression of the complement pathway and humoral immunity. From thrombo-inflammatory-related DEPs, we identified C-C motif chemokine 5 (CCL5) as a promising biomarker, and independent validation confirmed that elevated baseline plasma exosomal CCL5 was correlated with ENDi. In conclusion, our study highlights the role of exosomal proteins in immunothrombosis and establishes CCL5 as a potential predictor for ENDi. Importantly, we propose a novel mechanism involving immunoglobulin- and complement-related humoral immune suppression in the pathogenesis of ENDi, offering new directions for therapeutic intervention.