Plasma-Derived Extracellular Vesicles as Systemic Mediators of Radiation Response and Radiation Mitigation by Activated Protein C in a Rat Model

Radiological emergencies necessitate biomarkers that not only estimate absorbed ionizing radiation (IR) dose but also guide timely interventions to prevent or delay multi-organ injury. Conventional LC–MS-based metabolomics of bulk plasma is constrained by matrix effects that mask low-abundance species. Extracellular vesicles (EVs) constitute a metabolically enriched, underexplored compartment that can provide complementary insight into systemic metabolic and redox responses to IR. Female WAG/RijCmcr rats were exposed to 13.0 Gy leg-out partial-body X-rays and treated with one of three activated protein C (APC) variants-rat wild-type (WT), rat 3K3A-APC, or human WT APC-administered 24- and 48-hours post-irradiation. Longitudinal plasma collections (days 1, 14, 30, and 90) were subjected to metabolomic and lipidomic profiling of whole plasma and matched EV-enriched fractions to define signatures of acute radiation syndrome (ARS) and delayed effects of acute radiation exposure (DEARE), and their modulation by APC. ARS was marked by early dyslipidemia and widespread metabolic disruption, evolving into DEARE with persistent alterations in energy metabolism, protein homeostasis, and nucleotide biosynthesis, consistent with sustained oxidative and inflammatory stress. EV profiles showed matrix-specific, time-dependent trajectories distinct from plasma, with prominent lipid dysregulation and enrichment of fatty acid β-oxidation, sphingolipid, and cholesterol pathway metabolites at day 90. Rat 3K3A-APC promoted early EV metabolic normalization, whereas rat WT APC more effectively mitigated late DEARE-associated changes. Elevated sphingomyelins in plasma EVs at day 90 suggested a compensatory or anti-inflammatory lipid response. These data establish plasma-derived EVs as a sensitive matrix for radiation biomarker discovery and for elucidating APC-mediated modulation of IR-induced metabolic and redox disturbances.