Apolipoprotein-deficient mice are more susceptible to mild traumatic brain injury than are wild-type mice

Study objectives: Apolipoprotein E (apoE) is involved in redistributing cholesterol and phospholipids during compensatory synaptogenesis in the injured central nervous system and is believed to participate in repair, growth, and maintenance of myelin and axonal membranes. Three known isoforms of apoE (E2, E3, and E4) exist in humans and confer various degrees of protection or increasing risk of sequelae from traumatic brain injury (TBI). We hypothesize that apolipoprotein-deficient (apoE KO) C57BL6 mice will demonstrate greater neuronal injury from mild TBI than C57BL6 wild-type (WT) mice with mouse apoE. Methods: The weight-drop apparatus we used for impacts consisted of a 21-g weight, guided onto a force transducer located on the mouse head, midline, just cephalad to the interaural line. The weight fell freely from a 35-cm height. A counterweight avoided rebound impacts. Data were digitized using SigLab 20-22a dynamic signal analyzer data acquisition hardware and software. Dental molding material was used to make a mold of the mouse body and provided a firm (24.6 N/mm), consistent support for the head and body. Animals were anesthetized using 5% isoflurane (in air at 2.5 L/min) for 60 seconds, positioned in the holder, and subjected to a single weight drop. Sham controls were anesthetized and placed in the holder but not subjected to impact. A second group of animals underwent a different protocol. Using a small scalp incision and stereotactic head holder, an electromagnetic impactor was placed on the skull 2 mm left and 3 mm anterior to the lambda suture at an angle of impact of 10 degrees from vertical. Each animal had 1 impact; velocity 3.5 m/second and depth of displacement 3.0 mm. Sham mice had anesthesia and scalp incision but did not receive an impact. There were 38 male C57BL6 mice aged 8 to 12 weeks involved in these experiments, 20 apoE KO and 18 WT. All mice had histopathology using the de Olmos cupric silver-staining technique. Brains were removed, postfixed in perfusate for 24 hours, and sectioned (50 μm) throughout their extent. Eight coronal sections at the following levels were examined: 2.0, 1.2, 0.8, −0.8, −2.0, −2.8, −3.3, and −4.2 mm relative to bregma. One group of animals was killed and perfused 24 hours after impact; a second was perfused 96 hours after impact. Results: There were 21 mice in the weight-drop group (10 WT, 11 apoE KO) and 17 in the stereotactic impact group (8 WT, 9 apoE KO). The weight-drop group had histologic damage concentrated in the superior colliculus, optic tract, lateral geniculate, cingulated, and motor cortex. Damage was greater in apoE KO mice than WT. Sham mice did not show brain damage. The electromagnetic impact caused damage in apoE KO and WT mice, concentrated anteriorly in the ipsilateral cingulate, endopyriform, and bilateral motor cortices and posteriorly in the ipsilateral visual cortex and retrosplenial cortex. ApoE KO mice had more extensive injury. Conclusion: ApoE is important in repairing injured myelin and axonal membranes. Two previous rodent studies suggest there is increased neuronal damage in animals lacking apoE. Our findings further support this contention but also elucidate the variability of results, depending on methodology. Changes in force and location of impact and skull displacement can change extent and location of neuronal injury. Histology may differ according to staining technique as well. Most of the histologic damage seen in our slides results from fiber staining and would not be detectable using hematoxylin and eosin staining. ApoE KO mice show more extensive neuronal damage after mild TBI than WT mice, but the extent and location of injury is highly dependent on the characteristics of the impact model.