Worsened axonal injury and gliosis following diffuse brain injury in male and female mice with neurofilament light gene deletion

Neurofilaments have been theorized to make neurons more vulnerable to damage with injury or disease. To investigate how neurofilaments influence traumatic brain injury (TBI) pathophysiology, we examined neuropathology and glial response in neurofilament light chain knockout (NFL-KO) mice. Given the critical role of neurofilaments in maintaining axonal structure, we hypothesized that NFL-KO mice would exhibit altered neuropathology and gliosis after diffuse TBI. Male and female NFL-KO and wild-type mice were subjected to midline fluid percussion injury. Brains were collected at 3 h, 1 day, or 3 days post-injury for immunohistochemistry and compared with naïve controls. The number of APP-positive (APP+) profiles (indicator of axon pathology) was increased across time post-injury throughout the brain. Microglial morphological changes were evident by 3 h post-injury where cells appeared hypertrophic (increased cell volume) with hyper-ramified processes (increased branch points). This was followed by an increase in the density of microglia and colocalization with a surrogate marker of phagocytosis (CD68) at 3 days post-injury compared to naïve. Furthermore, astrocyte (GFAP) immunoreactivity was increased across time post-injury that was accompanied by morphological changes including retracted processes (reduced branch length). Notably, NFL-KO mice exhibited more severe axonal pathology and microgliosis, but a blunted astrocyte response compared to wild-type mice. Biological sex differences were also evident, with males showing more neuropathology and gliosis, particularly at acute time points post-injury. These findings highlight the influence of axonal cytoskeletal structure and biological sex on the acute glial response to TBI, suggesting new avenues for sex-informed and cytoskeleton-targeted interventions.