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    Determining the suitability of Human Umbilical Vein Endothelial Cells (HUVEC) and Endothelial Colony Forming Cells (ECFC) against Human Coronary Artery Endothelial Cells (HCAEC) as cell models for studying cardiovascular disease

    Filidoro, J F (2019) Determining the suitability of Human Umbilical Vein Endothelial Cells (HUVEC) and Endothelial Colony Forming Cells (ECFC) against Human Coronary Artery Endothelial Cells (HCAEC) as cell models for studying cardiovascular disease. Masters by Research thesis (MSc), Manchester Metropolitan University for the degree of Master of Science (by Research).

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    Abstract

    Cardiovascular diseases (CVD) are the most common cause of mortality globally and can be defined as disorders of the heart and blood vessels. CVD mortality rate and complications are on the rise, especially in developing countries. Age, obesity, smoking, diet, hypertension, genetics and diabetes are a few of the risk factors associated with CVD. Atherosclerosis is a significant contributor to CVD and is characterised by an accumulation of atherosclerotic plaque in the arteries, over time causing narrowing of the vessel lumen, limiting flow and causing tissue ischemia as well as acute occlusion due to atherothrombosis. Atherosclerosis initiates via dysfunction or damage of the endothelial cells (EC) that line the vessel wall. Endothelial dysfunction is amplified at branch points and curved sections in the vessel wall, exposed to disturbed blood flow patterns and limited in regions of arteries exposed to laminar flow, leading to focal development of disease. The protection in sections that experience laminar flow is predominantly driven through shear-regulated activation of KLF2, KLF4 and Nrf2 and suppression of NFκB activation in endothelial cells. In turn regulating the expression of over 1000 genes to modify endothelial behaviour and limit oxidative stress, inflammation and permeability. Huge amounts of research have been carried out to understand the processes, using a number of cell models to perform the research, including human coronary artery endothelial cells (HCAEC), which might be considered the gold standard, as well as human umbilical vein endothelial cells (HUVEC), because of their lower cost. This project assessed the relative gene expression of HCAECs and HUVECs cultured under identical conditions: static 4 | Page culture, ‘plaque promoting’ oscillatory shear stress (±5 dynes/cm2 , 1Hz), or ‘plaque-limiting’ physiological laminar shear stress (15 dynes/cm2 ). We concurrently evaluated the response of endothelial colony forming cells (ECFC), under the same conditions, evaluating whether these blood isolated cells can also be used as a model to investigate pathological processes involved in atherosclerosis from different patient groups. Our data concluded that KLF2 is significantly upregulated by laminar flow in HCAECs, whilst KLF4 is significantly upregulated by flow in all 3 cell types. The KLF2 and KLF4 responsive genes demonstrated a range of responses, with only eNOS showing significant upregulation in HCAECs by laminar flow. Additionally, Nrf2 regulated genes showed the largest upregulation in HCAECs under laminar flow whereas little difference was seen in within the NFκB regulated genes. The results obtained from the project provide evidence that HCAEC remain the most suitable cell model, with HUVECs demonstrating potential with additional work however ECFCs are rendered as unsuitable, due to their inability to provide significant upregulation of atherosclerosis-relevant genes.

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