| Summary: | abstract: Changes to the microenvironment of the endothelium can produce significant changes in the response of endothelial cells to stimuli. Human Aortic Endothelial Cells (HAECs) are tested in vitro for their fluid shear stress response when their substrates, and the solute concentrations of the fluids to which they are exposed, are modulated, and for their nitric oxide expression when they are exposed to hyperglycemic conditions. ImageJ is used to quantify either the degree of cellular alignment and elongation with the direction of flow, or the relative NO expression using the fluorochrome DAF-2. First, the results of Brower, et.al. are replicated: HAECs under normal glucose (4mM) conditions align and elongate with flow (p<<0.05), while high glucose (30.5mM) conditions negate this effect (p<<0.05) and is likely the result of Advanced Glycation End-products (AGEs). Then, in this study it is found that substitution of fibronectin for gelatin substrates does not impair flow (p<<0.05), indicating that fibronectin likely does not participate in the initiation of vascular lesions. High palmitic acid also does not prevent HAEC shear response (p<<0.05), which is consistent with Brower's predictions that AGEs are responsible for impaired elongation and alignment. NO production is significantly increased (p<<0.025) in HAECs cultured 24 hours under high glucose (30.5mM) conditions compared with normal glucose (4mM) conditions, indicating the presence of inducible nitric oxide as part of an inflammatory response. Aminoguanidine (5mM) added to high glucose concentrations reduces, but does not eliminate NO production (p<<0.05), likely due to insufficient concentration. Modulation of the endothelial microenvironment leads to pronounced changes in HAEC behavior with regards to NO production under hyperglycemic conditions. Diabetic model rat aortas are explanted and imaged for the purpose of detecting aortic endothelial cell alignment and elongation; improvements in this method are discussed. A microvessel chamber used with explanted human tissue is re-fit to reduce required volumes of solutions and allow more effective experimentation. === Dissertation/Thesis === M.S. Bioengineering 2013
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