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Thesis University of Iceland > Heilbrigðisvísindasvið > B.S. verkefni - Heilbrigðisvísindasvið >

Please use this identifier to cite or link to this item: http://hdl.handle.net/1946/24639

Title: 
  • Changes in protein expression and activation in rat arteriovenous fistula in vitro
  • is Breytingar á prótíntjániningu og -virkjun í rottuæðafistli in vitro
Advisor: 
Submitted: 
  • May 2016
Abstract: 
  • Introduction: Over 400,000 people were being treated with hemodialysis because of end stage renal disease in 2013. Of these 62.5% had arteriovenous fistula as their vascular access. Arteriovenous fistula are the preferred vascular access for hemodialysis as they are associated with lower infection rates and lower all-cause mortality than other methods of establishing vascular access. The drawback to using arteriovenous fistula is that they need to mature before being used. This maturation involves vessel wall thickening and vessel dilation. Studies today show that up to 60% of arteriovenous fistula do not mature enough to be used. Recent studies have shown that several proteins, including Eph-B4, caveolin-1, Akt-1 and eNOS can affect fistula maturation. The aim of this study was to assess the changes in protein expression and activation in a rat arteriovenous fistula using a novel bioreactor flow chamber.
    Methods and Materials: A bioreactor flow chamber that can subject an arteriovenous fistula to arterial levels of flow and shear stress was created. A rat fistula made from the jugular vein and carotid artery was placed in the bioreactor and connected to needles. These needles lead to tubing that runs through a peristaltic pump. A media of endothelial basal media and xanthan gum was prepared so that the viscosity of the media is similar to that of blood and used as intra- and extraluminal fluid. Each sample was ran in the bioreactor for an hour. As a control the other jugular vein was also harvested and kept in static conditions while the sample was in the bioreactor. One sample was analyzed with an immunofluorescence assay to assess the state of the endothelium. A western blot assay was then performed where total amounts of Eph-B4, caveolin-1, Akt-1 and eNOS were assessed. Phosphorylated versions of caveolin-1, Akt-1 and eNOS were also assessed. GAPDH was used as loading control.
    Results: The immunofluorescence assay showed that the endothelium was not destroyed in the bioreactor. Only two proteins assessed showed bands in the western blot, total caveolin-1 and total Akt-1. They showed no trends towards changes. A band was also observed for GAPDH which showed that the loading for the samples was uneven.
    Discussion: These results from the immunofluorescence assay show that arteriovenous fistula can be put under arterial conditions in the bioreactor without their endothelium being destroyed. The results from the western blot are unreliable but the fact that there do not seem to be changes in the total amount of caveolin-1 and Akt-1 also suggest that the endothelium is preserved. No phosphorylated proteins showed bands so no conclusions about protein activation can be drawn. We conclude that the bioreactor flow chamber model is viable for further, exciting research into fistula maturation.

Accepted: 
  • May 13, 2016
URI: 
  • http://hdl.handle.net/1946/24639


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