Crum, A. B. | bioRxiv Science

bioRxiv2026

Transglutaminase 2 Deletion Enhances Astrocyte-to-Neuron Metabolic Support and Attenuates Subacute Pathology Following Repetitive Mild Traumatic Brain Injury

Mild traumatic brain injury (mTBI) is the most common form of central nervous system (CNS) injury and is often characterized by persistent neuroinflammation, metabolic dysregulation, and oxidative stress. Repetitive injuries compound these pathologies and lead to multifocal axonal injuries and long-term functional deficits. Despite the prevalence of mTBIs, the cellular mechanisms that facilitate or prevent recovery following injury remain poorly defined. Here, we extend our previous work on the role of the protein transglutaminase 2 (TG2) in CNS injury and we hypothesize that transcriptional regulation by TG2 restricts metabolic versatility in astrocytes following TBI, thereby impairing neuronal energetic support and worsening pathological outcomes. We utilized an established weight-drop model of repetitive mTBI followed by multi-parametric analysis of TBI pathology in complete TG2 knockout (TG2-/-) and wild type mice. At 28 days post-injury, TG2-/- mice showed marked attenuation of TBI pathology, compared to wild type mice, in vulnerable white matter and default mode network (DMN) regions, as assessed by diffusion magnetic resonance imaging (MRI), resting-state functional MRI, and immunohistochemistry. Integrated epigenomic, proteomic, and metabolomic profiling of cortical astrocytes isolated 28 days after injury revealed a pronounced metabolic restriction in wild type astrocytes which was remarkably attenuated in the TG2-/- mice. This rescue was associated with a de-repression of gene networks involved in glutamate recycling, lipid metabolism, and metabolic homeostasis. Together, these studies provide novel mechanistic insights into the metabolic dysregulation that characterizes persistent TBI pathology, and establish a foundation for evaluating TG2 as a therapeutic target for TBI.

neuroscience↗

bioRxiv2025

Development of a modified weight-drop apparatus for closed-skull, repetitive mild traumatic brain injuries in a mouse model

Repetitive mild traumatic brain injury (rmTBI) is a major contributor to long-term neurological dysfunction, yet many preclinical models lack precise control and quantification of biomechanical forces across impacts. We developed a reproducible, closed-skull mouse model of rmTBI using a custom-built weight-drop apparatus featuring a solenoid-based rebound arrest system, integrated high-speed videography, and accelerometry to track head kinematics during impact. Adult male and female mice received either a single impact or nine daily impacts. Linear and angular acceleration data were analyzed alongside behavioral and histological outcomes. Our apparatus delivered consistent impact and velocity forces with minimal inter-subject variability. Additionally, the animals experienced consistent linear and angular acceleration as measured using high-speed video capture. These impacts did not cause skull fracture or acute vascular hemorrhage, but impacted animals had increased return of righting reflex (RoRR) time, consistent with mild, concussion-like symptoms. Behavioral testing revealed reduced performance of rmTBI-affected mice in an olfaction-mediated foraging task (buried food task), particularly at later timepoints, consistent with progressive olfactory impairment. Immunohistochemical analysis of Iba1 and CD68 in the brain demonstrated sex-dependent microglial activation, with males showing higher expression levels in both single- and nine-impact models. Among the brain regions investigated, microglial activation was most pronounced in the corpus callosum, neocortex, and olfactory tubercle. These findings underscore the importance of including sex as a biological variable in rmTBI research and support the utility of this model for probing injury thresholds, regional vulnerability, and potential therapeutic interventions in repetitive head trauma. Significance StatementMild traumatic brain injuries (mTBIs) contribute long-term sensory, motor, and cognitive dysfunction. We developed a novel approach for delivering repetitive mTBIs (rmTBIs) to mice via a custom weight-drop apparatus. The device allows precise control over impact forces and enables quantification of linear and angular acceleration during each impact. We describe the apparatus, the forces delivered, and the kinematics experienced by lightly anesthetized mice. We measured behavioral and neuroinflammatory sequelae in the brains of rmTBI-exposed mice compared to controls. rmTBI-exposed animals showed impairment in the olfaction-mediated buried food task and evidence of microglial reactivity in multiple brain regions days-to-weeks following injury. The results demonstrate the utility of this approach for studying rmTBI-associated pathophysiology, and for testing therapies or interventions for rmTBI.

neuroscience↗