EphA4 Influences Blood Brain Barrier Disruption and Endothelial Cell Response following Traumatic Brain Injury in a Mouse Model

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Virginia Tech

An astonishing number of deaths and related disabilities are attributed to traumatic brain injury (TBI) in the United States per year. Due to the unforeseeable nature of TBI and its association with the sequelae of other neurological comorbidities, research is centered around the secondary responses of brain mechanisms proceeding the initial mechanical injury. Blood brain barrier disruption is a well described driver of this secondary injury response and predictive marker of prognosis following TBI. Although BBB disruption plays a role in subsequent edema, inflammation, and the overall TBI outcome, the molecular mechanisms responsible for its regulation remain to be investigated. A large family of receptor tyrosine kinases, known as Eph receptors, that are important for axon growth and guidance embryonically and early-postnatally have been implicated in brain insults. Previous findings have shown that Eph expression is upregulated at the mRNA and protein level immediately following TBI. Moreover, ablation of Eph receptors on endothelial cells (ECs) revealed improved blood flow to the lesioned cortex in knockout (KO) mice compared to wild type (WT). Based on these results, we hypothesize that Eph receptors negatively regulate BBB permeability leading to neural dysfunction and motor deficits following TBI. To investigate this hypothesis, we characterized the temporal profile of the BBB, evaluated the EC-specific effects of Eph receptors, and used RNA sequencing to assess the cell-specific contributions following TBI in WT compared to KO mice. Our results show that EC-specific loss of Eph expression ameliorated BBB permeability at 6hr, 1-, 4-, and 7-days post injury (dpi) correlating with improved motor function at 7- and 14-dpi. Furthermore, mechanistic studies revealed increased mRNA expression of Tie2, Ang1, and the tight junction proteins Zona Occludens and Occludin in KO mice compared to WT. As well as, connection with neuronal processes. Based off of these findings, we utilized a soluble Tie2 inhibitor to elucidate the influence of Eph receptors on the Tie2/Ang pathway, and their role in mediating the effects seen. Tie2 inhibition of the KO mice revealed similar BBB disruption and lesion volume as WT 1dpi, attenuating the previous protection KO mice demonstrated. Future studies are necessary to understand other pathways that may be implicated in Eph receptor influence on endothelial cells such as inflammatory mediators and neurovascular crosstalk. This data provides evidence that Eph receptors negatively mediate EC response through downstream signaling of the Tie2/Ang pathway and may be a means of therapeutic target in the future.

Traumatic Brain Injury, Blood Brain Barrier, Eph Receptors, Endothelial Cells, Tie2/Ang