GENE EXPRESSION IN VASCULAR REMODELING

The biology of the vascular smooth muscle cell (VSMC) is of central importance in understanding the behavior of the vascular wall in some of the most prevalent and costly diseases in developed societies, including hypertension, peripheral vascular disease, and coronary artery disease. Pathologic VSMC activity is still more evident in the syndromes of accelerated arteriosclerosis seen after coronary bypass surgery, cardiac transplantation, and coronary angioplasty. Balloon injury of the rat carotid artery (RCBI) is an animal model of vascular response to injury bearing greatest resemblance to the clinical problem of restenosis following angioplasty. The kinetics of cellular response to denudation and stretch injury to the carotid artery have been well delineated. Yet despite intensive investigation, the factors controlling such fundamental VSMC functions as migration, proliferation, and synthesis and secretion of proteins comprising extracellular matrix (ECM) remain incompletely understood. The scientific approach described in this proposal is to apply a state-of-the-art molecular biologic technique, differential mRNA display, to an in vivo model system, RCBI, in which these VSMC behaviors have been well described. To date, 3 genes with increased expression after RCBI have been confirmed by Northern analysis; preliminary 3' sequence data do not demonstrate high homology with any entries in Genbank. Full length cDNAs will be cloned, sequenced, and examined for sequence similarities to known genes, which may suggest gene function. Cellular localization within the vessel wall and tissue expression patterns will be defined. Fusion proteins will be expressed and evaluated in vitro with migration and proliferation assays and in Northern analysis of ECM gene expression. Blocking antibodies raised against fusion proteins will be used in similar studies. Analysis will then return to the in vivo process by assessing effects of both fusion protein and blocking antibody on vascular remodeling in the RCBI model and through the study of transgenic mice. Finally, correlation with clinical restenosis and atherosclerosis will be evaluated by immunohistochemistry, in situ hybridization, and quantitative polymerase chain reaction studies using atherectomy tissue and other specimens of human vascular pathology. Following the hypothesis that these changes in cellular activity result from changes in transcription level of particular mRNA species within the vascular wall, this approach should provide a unique avenue of insight into the molecular mechanisms determining in vivo smooth muscle cell behavior during vascular remodeling.