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The objective of this research is to understand the molecular events occurring in the endothelial cells of the arterial wall that either control or are induced by changes in permeability and fluid shear stress. The endothelium consists of a single, continuous layer of cells on the innermost surface of blood vessels. Among its roles is to regulate the entry of blood components, such as cholesterol-rich low-density lipoprotein (LDL), inflammatory cells, and clotting agents, into the vessel wall, where they can promote the development of dangerous atherosclerotic lesions. Certain daily activities, such as exercise, smoking, and stress, result in sudden changes in local arterial blood flow. In response to such changes in the fluid dynamic environment, the cytoskeletal components of endothelial cells undergo reorganization. It is hypothesized that this period of adaptation presents an opportunity for macromolecules like LDL to penetrate the endothelial cell layer and enter the underlying tissue. It is also hypothesized that shear stress and permeability changes in vivo affect the distribution, expression and/or activation of a variety of proteins. Some of these events may lead to the development of atherosclerotic lesions.
To elucidate these molecular events, experiments using porcine arteries are performed in which the relationships between local fluid shear stress, uptake of Evans blue dye (EBD), cellular morphology, and the distribution, expression, and activation of a variety of proteins are studied. EBD binds to albumin in the blood, and its uptake reflects the local permeability of the wall to macromolecules such as LDL. The proteins in which we are interested are involved in cellular junctions, cytoskeletal structure, cellular adhesion, signaling, transport, and the regulation of transcription. During the experiments, albumin-bound EBD is injected into the bloodstream of a pig. The iliac and coronary arteries are dissected post-mortem and used in a variety of molecular studies at the genomic and protein levels. The cellular response is related to the local fluid dynamic environment, as determined from computational fluid dynamics. Here we illustrate immunofluorescence studies, which reveal the cellular distributions of proteins of interest. Below are sample immunofluorescence images for vimentin, a cytoskeletal protein (left, 40x), and PECAM, a protein found a junctions between cells (right, 10x), within the endothelial monolayer of an iliac artery.
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