Sex-dependent blood-brain barrier alterations following repeated mild blast traumatic brain injury at varying inter-injury intervals

BACKGROUND: Traumatic brain injury (TBI) is a worldwide epidemic and a major cause of disability, morbidity, and mortality. TBI is a major risk factor for the development of late-life dementia, especially Alzheimer's Disease, and other neurological conditions, such as epilepsy. The most prevalent form of TBI is mild TBI (mTBI), which is characterized by cognitive and psychological deficits as well as metabolic and vascular mechanisms of neuropathobiology. mTBI can be induced by either impact or blast insults and multiple mTBIs can result in worsened outcomes. There is a need to understand pathological impairments in the blood-brain barrier (BBB) following mTBI. METHODS: To model repeated mild blast traumatic brain injury (rmbTBI), male and female rats (N = 6/group) were exposed to repeated low-level, 11 psi static peak overpressure blast waves using the McMillan blast device. rmbTBI was produced with either 1 h or 24 h inter-injury interval. Sham animals undergo all procedures except for the blast. Animals performed open field and elevated plus maze (EPM) behavior tests before euthanasia at 7d post-rmbTBI. Hemibrains were taken separately for immunohistochemistry and western blot analysis. Brain capillaries were isolated from fresh brain tissue and taken for immunofluorescent (IF) staining. RESULTS: To examine BBB-specific deficits, pericyte (PDGFRβ), tight junction (TJ) protein (zonula occludens-1 (ZO1), occludin and Claudin-5), astrocytic end-feet (AQP4), and BBB integrity (SMI-71) markers were analyzed at 7d post-rmbTBI. Deficits in cortical AQP-4 and SMI-71 levels were observed in male rmbTBI-24 h group compared to sham while female rmbTBI groups displayed no deficits in these markers compared to sham. There were deficits in TJ markers in both male rmbTBI groups that were not apparent in female-derived capillaries. Western blot analysis demonstrates that PDGFRβ is significantly decreased in male rmbTBI-24 h animals but not male rmbTBI-1 h or female rmbTBI animals. Male rmbTBI-1 h group displayed lower levels of GFAP and higher levels of IBA-1 in the cortex as compared to sham; female rmbTBI groups displayed similar levels of cortical GFAP and IBA-1 as sham. Male rmbTBI and female 1 h-interval rmbTBI groups displayed significantly higher closed arm entrances during EPM as compared to respective sham groups. CONCLUSION: Our findings demonstrate that rmbTBI produces robust on-going deficits in BBB and glial outcomes that correspond with behavioral abnormalities in male animals. The extent of these outcomes is dependent upon inter-injury interval. Female rmbTBI animals display anxiety-related behavior that is not driven by BBB-related impairments.