Atherosclerotic vascular disease continues to be a major cause of morbidity and mortality in the U.S despite aggressive medical and surgical therapy. Vascular cell (endothelial and smooth muscle) responses to injury elicited by angioplasty and bypass grafting are major factors in determining the ultimate functional state of the particular vessel. The vascular cell responses to injury are known to be modulated by complex, dynamic cell-cell interactions and cellular interactions with the underlying and surrounding extracellular martix via integrins. In addition, the roles of a variety of growth factors (PDGF, TGF-b, bFGF), although still incompletely understood, are thought to be critical in directing the outcome following vessel injury. The long-term goal of this proposal is to elucidate the interactive signal transduction mechanisms which are active following the engagement of selected cell- cell adhesion molecules with their ligands, integrins with ECM components and selected growth factors with their receptors; and to understand the interrelationships among these systems in the vessel wall. Specifically, tissue culture models of large vessel endothelial cell denudation injury- repair and microvascular endothelial cell angiogenesis will be used to characterize and determine the mechanism(s) of signal transduction following cell adhesion molecule, integrin and growth factor receptor engagement during endothelial cell responses to injury. Several methodologies will be employed including tissue culture, immunolabeling at light, confocal and electron microscopic levels, Northern, Southern and in situ hybridizations, biosynthetic labeling, immunoprecipitation, immunoblotting, retroviral transduction and over-expression of genes of choice. Experiments will center around the use of antibodies, cDNA probes and peptides directed against specific kinases thought to be involved in these signalling cascades, selected growth factor receptors and their ligands, matrix molecules and matrix binding proteins and selected domains of these molecules, cRNA and cDNA probes specific for these molecules and specific chimeric TGF-b molecules in in vitro large vessel endothelial cell migrations and during in vitro angiogenesis studies with microvascular endothelial cells. A better understanding of the signalling cascades that occur during the processes of vascular cell response to injury may lead to the design, production and implementation of improved synthetic grafting materials, agents that promote optimal vascular cell behavior following therapeutic intervention, and agents that can be used to modulate the angiogenic response and a somatic cell gene therapy approach which has the potential to deliver long-term parenteral therapy.
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