The endothelium is well recognized as a mechanotranduction interface influencing vessel physiology and pathology. The prevalence of atherogenesis in regions associated with complex disturbed flows in vivo has long been recognized yet the factors that predispose such sites and potentiate preferential atherosclerosis there are poorly understood. We propose that hemodynamic forces determine the susceptibility to atherosclerosis by regulation of the regional phenotype of endothelium at such sites. Preliminary profiling studies in the normal adult porcine aorta using microarray have shown differences in endothelial gene expression between areas prone to develop atherosclerotic lesions (disturbed flow) and those areas that are typically spared. Project 5 will address the hypothesis that at lesion-susceptible locations, the endothelial phenotype is in an equilibrium state that is primed to develop atherosclerosis by expression of pro-inflammatory genes but is protected by the up-regulation of anti-oxidant mechanisms and other genes that prevent the initiation/early development of atherosclerosis. Exposure to one or more risk factors (additional to hemodynamics) is proposed to be necessary for imbalances that lead to the development of vascular pathology. In a collaboration with the Center for Devices and Radiological Health animal facility at the FDA Laurel, MD, we will conduct the first multi-gene expresssion studies of endothelium in vivo in defined hemodynamic locations. Regional endothelial phenotypes will be defined in normal adult porcine arteries, and the effects of the important risk factors hypercholesterolemia, gender, and hormonal manipulations upon atherosclerosis-primed regions will be determined. The critical role of hemodynamics in regional susceptibility will be tested by creating flow disturbances in the common carotid artery, a location that is normally exposed to undisturbed laminar flow and which is resistant to atherogenesis; we propose that a susceptible phenotype will result from such intervention. The effects of hemodynamic forces upon anti-oxidant, coagulation, inflammatory and other gene classifications will be measured in vitro where transcription profiles of porcine endothelial cells exposed to oscillating and unidirectional flow will be analyzed. Using small interfering RNA, specific gene 'knock-downs' will be created (gene silencing) to determine the effect upon hemodynamic responses in vitro including perturbations of the phenotype as evaluated by microarray profiling with follow-up protein and proteomic studies.
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