Temporal metabolic alterations in cerebrospinal fluid-derived small extracellular vesicles following severe traumatic brain injury

Severe traumatic brain injury (sTBI) initiates complex secondary injury mechanisms that evolve over time and are difficult to monitor using current clinical tools. Small extracellular vesicles (sEVs) in cerebrospinal fluid (CSF) represent a promising source for tracking molecular alterations linked to injury progression. We performed untargeted metabolomics profiling of phosphatidylserine-positive sEV-enriched fractions from the CSF of sTBI patients at Days 1, 3, and 5 post-injury, alongside healthy controls. LC-MS revealed distinct temporal alterations, with 53, 77, and 15 differentially abundant metabolites at Days 1, 3, and 5, respectively, and a persistent core of 12 metabolites altered across all time points. Sustained reductions in cholic acid and trans-2-dodecenoylcarnitine suggest prolonged bile acid dysregulation and impaired mitochondrial fatty acid oxidation. Early alterations in glycochenodeoxycholic acid, phosphate, and N-arachidonoyl dopamine are consistent with disturbed bile acid homeostasis, energy metabolism, and endocannabinoid signaling, while Day 3 reductions in N-acetyl-L-aspartic acid and sex steroids suggest neuronal injury and steroid dysregulation. Exploratory pathway analysis nominally implicated bile acid biosynthesis, androgen and estrogen metabolism, and aspartate metabolism. These findings demonstrate the utility of CSF-sEV metabolomics for capturing dynamic biochemical changes post-sTBI. This work provides a foundation for evaluating sEV-based biomarkers for monitoring injury progression in TBI patients.