Traumatic brain injury (TBI) can cause a number of deleterious effects to the neurovascular system, including reduced cerebral blood flow (CBF), vascular regression, and ischemia, resulting in cognitive decline. Research into therapeutic targets to restore neurovascular function following injury has identified endothelial EphA4 receptor tyrosine kinase as a major regulator of vascular regrowth. The research outlined herein utilizes an endothelial-specific EphA4 knockout mouse model (KO-EphA4flf/Tie2-Cre) to determine the extent to which this receptor may influence vascular regrowth following TBI. Analysis of the colocalization and proximity of endothelial and mural cell markers (i.e. PECAM-1 and PDGFRβ, respectively) in immunohistochemically-stained brain sections demonstrates that EphA4 silencing does not seem to affect the physical association between, nor total amounts of, endothelial cells and pericytes, between genotypes by 4 days post-injury (dpi). Nevertheless, these measures demonstrate that these cell types may preferentially proliferate and/or expand into peri-lesion tissue in both KO-EphA4flf/Tie2-Cre) and WT-EphA4fl/fl mice. These data further suggest that both genotypes experience homogeneity of PECAM-1 and PDGFRβ expression between regions of the injury cavity. Gene expression analysis using mRNA samples from both genotypes reveals that KO-EphA4flf/Tie2-Cre CCI-injured mice experience increased expression of Vegfa, Flt1, and Fn (Fibronectin) compared to sham-injured condition knockouts. These results demonstrate changes in expression of angiogenic factors in the absence of early differences in patterns of vessel formation, which may underlie improved vascular regrowth, as well as outline a potential mechanism wherein the interplay between these factors and EphA4 silencing may lead to improved cognitive outcomes following TBI. / Master of Science / Every day in the United States, an average of 155 people die due to the consequences of traumatic brain injury (TBI), with many survivors suffering life-long debilitating effects, including deficits in behavior, mobility, and cognitive ability. Because of this, there is a need for researchers to identify therapeutic strategies to stimulate recovery and improve patient outcomes. Recent advancements in the field of vascular biology have identified the regrowth of the blood vessels in the brain following TBI-induced damage as an important step in the recovery process, since the resulting increases in blood flow to damaged tissue will provide oxygen and nutrients necessary to fuel recovery. The work presented in this Masters thesis follows in this vein by examining a protein receptor known as EphA4, which is found on cells within blood vessels and has been implicated in reducing the rate of vessel growth under injury conditions. By blocking the activity of EphA4, we hoped to find increased vascular regrowth following brain injury in mice. During the experiments outlined herein, it was found that there were no statistically significant differences in vessel-associated cell densities between mice with or without EphA4 activity 4 days after injury, but there were differences in the levels of proteins and/or signals associated with vessel growth. Based on these results, we conclude that removing EphA4 activity increases expression of these pro-vessel growth proteins in mouse brains following injury at these early time points, potentially leading to increased vessel growth and improved recovery over subsequent weeks following injury.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/99318 |
| Date | 09 July 2020 |
| Creators | McGuire, David Robert |
| Contributors | Translational Biology, Medicine, and Health, Chappell, John C., Robel, Stefanie, Theus, Michelle H., Fox, Michael A. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Thesis |
| Format | ETD, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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