Applied Computing

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Acknowledgement

I would take this opportunity to thank my research supervisor, family and friends for their support and guidance without which this research would not have been possible.

DECLARATION

I, [type your full first names and surname here], declare that the contents of this dissertation/thesis represent my own unaided work, and that the dissertation/thesis has not previously been submitted for academic examination towards any qualification. Furthermore, it represents my own opinions and not necessarily those of the University.

Signed __________________ Date _________________

1. Introduction

Qualitative modification of low-density lipoproteins (LDLs) plays a key role in the development of atherosclerotic disease. It is currently accepted that oxidative modification of LDL occurs locally in the artery wall, where oxidized LDL (old) induces endothelial dysfunction, leukocyte recruitment, monocot differentiation, smooth muscle cell proliferation, and foam cell formation . However, the presence of significant amounts of oxidative modified LDL in plasma remains a matter of controversy. An electro negatively charged subtraction of LDL (LDL(-)), isolated from human plasma, accounts for 1-10% of the total LDL in normolipemic (NL) subjects . Although discrepancies as to its origin and composition have arisen , different authors concur on the enhanced cytotoxicity of LDL(-) in endothelial cells . We recently reported that LDL(-) isolated from NL subjects induces the release of interleukin 8 (IL-8) and monocyte chemotactic protein 1 (MCP-1) by endothelial cells, which suggests a role in leukocyte recruitment . Thus, endothelium could be induced to produce chemotactic molecules, not only by oxLDL trapped in the arterial intima, but also by circulating LDL(-). Familial hypercholesterolemic (FH) subjects present a threefold higher proportion of LDL(-) than NL . Bearing in mind the elevated LDL concentration in these subjects, the absolute amount of LDL(-) in FH can be up to eightfold higher than in NL subjects. Thus, a high number of circulating lipoproteins with proinflammatory capacity could contribute to the increased atherosclerotic risk and endothelial dysfunction observed in FH . Physicochemical and biological characteristics of LDL(-) from FH subjects have not been previously studied. The aim of this work was to analyze the characteristics of LDL(-) from FH and compare them with those of NL, particularly with regard to lipid and apoprotein composition, oxidative status, and induction of chemokine production by endothelial cells.

1. Methods

1.1. Subject selection and experimental design

FH subjects were selected on the basis of their plasma lipid levels (total cholesterol>8.5 mmol/l, LDL cholesterol>5.5 mmol/l, and triglyceride<2 mmol/l) and family history of autosomal-dominant hypercholesterolemia. Control NL subjects had total cholesterol<5.5 mmol/l and triglyceride<1 mmol/l. All subjects were normoglycemic, non-smokers, and aged from 32 to 59 years. Fasting plasma samples were obtained in EDTA-containing Vacutainer tubes and stored at -80 °C until analysis. FH and NL samples were stored for a similar period that never was longer than 45 days. Freezing of plasma for this period results in the absence of oxidative modification measured as a-tocopherol consumption (a-tocopherol in LDL from fresh plasma is 8.3±1.4 mol/mol apoB; a-tocopherol in LDL from frozen plasma LDL is 8.1±1.7 mol/mol apoB; measured from eight LDL isolated from different ...