Electronic Thesis and Dissertation Repository

Thesis Format

Monograph

Degree

Master of Science

Program

Pathology and Laboratory Medicine

Supervisor

Hoffman, Lisa M

Abstract

In Duchenne muscular dystrophy (DMD), angiogenesis appears to be attenuated. Local administration of angiopoietin 1 (Ang1) has been shown to reduce inflammation, ischemia, and fibrosis in DMD mice. Ang1 is a vital vascular stabilizing factor that activates the endothelial cell receptor Tie2, leading to downstream pro-survival PI3K/Akt pathway activation and eNOS phosphorylation. In this study, we aim to characterize the Ang/Tie2 signaling pathway within the skeletal muscle of mouse models of DMD. Utilizing immunoblots and RT-qPCR, we show that Ang1 is downregulated, while the antagonist angiopoietin 2 (Ang2) is upregulated, leading to a decreased Ang1/Ang2 ratio. This correlates with a reduction in phosphorylated Tie2/total Tie2 ratio. Interestingly, no significant differences in Akt or eNOS phosphorylation were observed, although DMD murine models did have elevated total Akt protein concentrations. These observations suggest that Ang1/Tie2 signaling may be dysregulated in DMD and further investigations may lead to new therapeutic interventions for DMD.

Summary for Lay Audience

Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disease that affects 1 in every 3000 boys. The most common symptom of DMD is muscle cell death and tissue degeneration. The body will naturally attempt to repair the damages by triggering an inflammatory response to remove the cellular debris of cells that undergo apoptosis, also known as programmed cell death. Muscle stem cells, cells that can become muscle cells, will then be activated and undergo cell division to repair the damages. Additionally, nearby blood vessels will undergo angiogenesis, the process of forming new blood vessels from preexisting vessels, to supply the regenerating tissue with sufficient nutrients and oxygen that is needed for tissue repair. Unfortunately, angiogenesis appears to be impaired in DMD, leading to the formation of immature vessels that are incapable of functional perfusion (i.e. supplying the necessary nutrients and oxygen needed for repair). To treat this vascular defect, our lab has shown that the administration of angiopoietin-1 (Ang1) can stimulate the formation of new functional vessels that are capable of functional perfusion. Ang1 is a vascular stabilizing factor that binds to the Tie2 receptor on the surface of endothelial cells and triggers a cascade of signaling effects that help maintain vasculature integrity and help the maturation of angiogenic vessels. Although we have shown that Ang1 can help repair vascular defects, Ang1 signaling has not been previously characterized in DMD. In this study, we aimed to evaluate Ang1 signaling, including the expression of its antagonist angiopoietin-2 (Ang2), receptor Tie2 expression and activation, and further downstream targets protein kinase B (Akt) and endothelial nitric oxide synthase (eNOS) activation. From our results, we show that Ang1 is expressed less in DMD mouse models compared to healthy mice, while more Ang2 is being produced. This difference may shift the Ang1/Ang2 concentration ratio towards Ang2 and lead to the decrease in activated Tie2/total Tie2 ratio seen in this study. These findings suggest that Ang1 signaling is reduced in DMD and warrants further investigation that may need to development of new therapeutic options for DMD patients.

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