Skull Fractures Induce Neuroinflammation and Worsen Outcomes after Closed Head Injury in Mice

Abstract

The weight-drop model is used widely to replicate closed-head injuries in mice; however, the histopathological and functional outcomes may vary significantly between laboratories. Because skull fractures are reported to occur in this model, we aimed to evaluate whether these breaks may influence the variability of the weight-drop (WD) model. Male Swiss Webster mice underwent WD injury with either a 2 or 5 mm cone tip, and behavior was assessed at 2 h and 24 h thereafter using the neurological severity score. The expression of interleukin (IL)-6, IL-1b, tumor necrosis factor-a, matrix metalloproteinase-9, and tissue inhibitor of metalloproteinase-1 genes was measured at 12 h and 1, 3, and 14 days after injury. Before the injury, micro-computed tomography (micro-CT) was performed to quantify skull thickness at the impact site. With a conventional tip diameter of 2 mm, 33% of mice showed fractures of the parietal bone; the 5 mm tip produced only 10% fractures. Compared with mice without fractures, mice with fractures had a severity-dependent worse functional outcome and a more pronounced upregulation of inflammatory genes in the brain. Older mice were associated with thicker parietal bones and were less prone to skull fractures. In addition, mice that underwent traumatic brain injury (TBI) with skull fracture had macroscopic brain damage because of skull depression. Skull fractures explain a considerable proportion of the variability observed in the WD model in mice-i.e., mice with skull fractures have a much stronger inflammatory response than do mice without fractures. Using older mice with thicker skull bones and an impact cone with a larger diameter reduces the rate of skull fractures and the variability in this very useful closed-head TBI model.

Mast Cell Activation, Neuroinflammation, and Tight Junction Protein Derangement in Acute Traumatic Brain Injury

“…The weight drop model is widely used as an animal model of TBI. However, the histopathological changes, immune and inflammatory responses, and behavioral changes vary, as there are many variations in the experimental procedures and animals used [1,27]. The mechanism and the nature of the neuroimmune response, including mast cell response, are more complex to understand after neurotrauma/TBI.…”

“…Then, the skull was exposed on the top with an incision. Following this, a metal weight (35 g) was released to free fall from 80 cm above the head through a vertical tube (guided path) to induce TBI without craniotomy, as previously reported by others for closed-head injury weight drop model [25][26][27][28]. Then, the skin was sutured for the recovery.…”

Time-dependent cytokine and chemokine changes in mouse cerebral cortex following a mild traumatic brain injury

“…Also, it may be worth considering the effects of skull fractures in weight drop models as described by Zvejniece and co-workers. They describe work in an open head, close skull weight drop model (drop height of 8 cm with a weight of 90 g) examining the effects of different weight cone diameters (2 or 5 mm) on inducing TBIs associated with frequent skull fractures in mice ( Zvejniece et al, 2020 ). Their observations describe worsened functional outcomes and more pronounced inflammatory responses for animals that received the weight drops which resulted in the occurrence of skull fractures.…”

Acute Traumatic Brain Injury-Induced Neuroinflammatory Response and Neurovascular Disorders in the Brain

“…Closed head acute TBI was induced in 8-week-old male mice (CD-1 mice; the University of Missouri, Mutant Mouse Resource and Research Center and the University of Missouri Animal Modeling Core, Columbia, MO) by standard weight-drop procedure, as reported previously with slight modifications (Milman et al 2005 ; Shishido et al 2019 ; Zohar et al 2003 ; Zvejniece et al 2019 ). Briefly, the mouse was first anesthetized with isoflurane, and then the head was positioned on a spongy cushioned surface.…”

“…Immediately, the mouse skull was exposed on the top with a small incision. Then a metal weight of 35 g was allowed to fall from 80 cm above the head through a vertical tube and induced cortical contusion without craniotomy (Milman et al 2005 ; Shishido et al 2019 ; Zohar et al 2003 ; Zvejniece et al 2019 ). Only the skin incision was made and then sutured in sham control mice instead of the weight-drop procedure.…”