Mapping Inflammation Dynamics Across Brain, CSF, and Serum after TBI: A Multi-Compartment and Machine Learning Analysis of Injury and Celastrol Effects

Traumatic brain injury (TBI) involves a transition from acute to chronic neuroinflammation that contributes to lasting neurological deficits; however, the mechanisms by which brain-derived inflammation orchestrates brain-periphery interactions and how to modulate them remain incompletely understood. This multi-compartment study assessed cytokine concentrations across paired serum, cerebrospinal fluid, and ipsilateral hippocampal lysate samples 48 h after controlled cortical impact. Celastrol, an understudied anti-inflammatory compound in TBI, was used as a model therapeutic to evaluate the utility of multi-compartment sampling in pre-clinical therapy screening. Multi-compartment factor analysis (MCFA) was applied as a novel machine learning workflow to identify covariance patterns across fluid and tissue TBI biomarkers. Injury-induced responses were greatest in the hippocampus (IL-6, KC/GRO, and tumor necrosis factor-alpha TNF-α increases), with serum decreases in IL-6 and TNF-α. Celastrol restored serum TNF-α to control levels. Sensitivity analyses after outlier removal additionally revealed hippocampal IFN-γ suppression, CSF IL-1β and TNF-α elevation, and celastrol-associated increases in hippocampal IL-10 and IFN-γ. MCFA uncovered coordinated CSF shifts in IL-6, IL-1β, and KC/GRO not individually significant by standard methods and identified a serum inflammatory pattern tracking hippocampal inflammation. Cross-compartment correlations were limited, supporting local regulation. Multivariate dispersion analysis revealed that untreated injury doubled inter-animal inflammatory heterogeneity across the factor space, while celastrol restored dispersion to sham-like levels, indicating treatment re-consolidated the multi-compartment inflammatory response. These findings demonstrate a compartment-specific inflammatory response 48 h post-TBI and suggest celastrol exerts cytokine- and compartment-specific immunomodulatory effects. MCFA provided insights beyond traditional analyses and a standardizable framework for characterizing serum signatures of active brain injury with translational potential for broader TBI biomarker datasets.