Atherogenesis may be associated with increased transport and accumulation of low density lipoprotein (LDL) in the arterial wall. The long-term objective of our research is to obtain a quantitiative physicochemical understanding of the LDL uptake process and of the effects of normal and pathophysiological variables which influence uptake. We have discovered microscopic sites of markedly increased permeability to horseradish peroxidase (HRP) and radiolabelled LDL in certain preferred regions of the rabbit aorta, some of which are also characterized by abnormal, enlarged endothelial cells and other mophological features different from normal tissue. The goal of our proposed research is to obtain a better fundamental understanding of the nature of these high permeability sites, including (1) the nature and causes of the abnormal cells, (2) the morphology of the underlying subendothelial tissue, (3) the mechanism(s) of enhanced transendothelial transport, (4) the binding of LDL to extracellular matrix, (5) the role of receptor- mediated and receptor-independent LDL degradation processes, (6) the temporal history of these sites, (whether they are transient or permanent), the (7) correlation of their spatial distribution with hemodynamic factors, and (8) their possilbe predisposition to development of early focal atherosclerotic lesions in hypercholesterolemic animals. These issues will be addressed in a variety of experiments with rabbits using light and electron microscopy techniques in order to investigate the cells and tissues in these sites at the ultrastructural level. En face mapping of HRP spots and 3H-thymidine incorporation will be compared to determine if some of these abnormal cells are replicating endothelial cells. In vivo uptake experiments with radiodinated and tritium-labeled LDL will be analyzed with quantitative autoradiography. LDL will be conjugated with fluorescence and colloidal gold markers for improved light and electron microscopy localization. In addition, binding of LDL to extracellular matrix and stress fiber expression in high permeability sites will be evaluated. The relationship between local flow patterns and wall shear stress and the local density and distribution of HRP spots, especially at sites with a predilection for atherosclerosis, will be studeid using cinemicrographic techniques in the rabbit aorta rendered transparent by a tissue clearing technique. Theoretical mathematical models will be developed to describe LDL uptake and accumulation in these high permeability sites.
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