Date of Award

8-2018

Degree Type

Thesis

Degree Name

Biomedical Sciences (M.H.S.)

Department

Biomedical Sciences

First Advisor

David Kurjiaka

Second Advisor

Christopher Pearl

Third Advisor

Frank Sylvester

Academic Year

2017/2018

Abstract

Cardiovascular disease is the number one killer of adults in the US (~800,000 lives every year:1 in 3 deaths). Evidence suggests omega-3 fatty acids like eicosapentaenoic acid (EPA), found in fish oils, reduce the risk of developing cardiovascular diseases. Many of these diseases are caused by atherosclerotic plaque development in vessels due to endothelial cell damage initiating inflammatory responses. Recent work argues omega-3 fatty acids stimulate anti-inflammatory responses by binding to Free Fatty Acid (FFA) receptors. Inflammatory responses in endothelial cells alter the expression of many proteins that affect the function of those cells. In particular, gap junction protein (connexin: Cx) expression is affected by inflammatory signals with Cx43 increasing by NF-kB signaling. This study will evaluate the impact of EPA on endothelial cell expression of Cx43. We hypothesize EPA treatment will decrease endothelial cell expression of Cx43 via the FFA-4 receptor. Mouse endothelial cells (bEnd.3) were treated with 30 μM EPA for up to 48 hours, with protein isolated from a control (0.1% ethanol) and EPA treated cells at specific times. The expression of Cx43 was evaluated via Western Blot. The Cx43 signal in the EPA treated samples was normalized to the Cx43 signal in control samples isolated at the same time (a ratio of 1 indicates no change). EPA decreased (p<0.05) endothelial cell expression of Cx43 after 48 hours. However, addition of AH-7614 (FFA-4 receptor antagonist) to EPA treated cells did not alter the EPA induced decrease (p<0.05) in Cx43 expression at 48 hours. More importantly, blocking the FFA-4 receptor for 48 hours decreased (p<0.05) Cx43 expression, suggesting something in the cell culture media caused basal FFA-4 receptor stimulation. Treatment with the FFA-4 agonist (TUG- 891) for 48 hours caused a non-significant increase in Cx43 expression. Thus, the reduction in expression of Cx43 at 48 hours did not involve the FFA-4 receptor. In fact, the response of endothelial cells to FFA-4 receptor activation is consistent with increased inflammation (i.e. increased Cx43 expression). EPA has an anti-inflammatory effect on mouse endothelial cells at 48 hours that is not mediated by the FFA-4 receptor.

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