Proliferation of vascular smooth muscle cells (VSMC) is important in the pathogenesis of atherosclerosis, post-transplantation atherosclerosis, vascular graft occlusion, post-angioplasty restenosis, and vascular remodeling in hypertension. The insulin-like growth factor I receptor (IGF IR) is a membrane tyrosine-kinase receptor that is expressed at high levels on VSMC. We have shown that this receptor is an important mediator of VSMC growth responses in vitro. The major objective of this proposal is to study the interactions between this receptor and other heterologous growth factors that are important in VSMC growth. These include platelet- derived growth factor (PDGF), basic fibroblast growth factor (bFGF), angiotensin II (ang II), and alpha-thrombin. We have cloned a full-length cDNA encoding the rat IGF IR and shown that antisense transcription of an IGF IR cDNA in VSMC has marked inhibitory effects on VSMC growth responses. The IGF IR appears to be at a convergence point for the action of other growth factors. Thus one can inhibit ang II and alpha-thrombin induced DNA synthesis in VSMC by downregulating this receptor. Multiple growth factors upregulate IGF IR number on VSMC, and this upregulation is likely essential for their mitogenic effects.
The specific aims of the grant are: 1) To determine biochemical and molecular signaling events important in growth factor regulation of the IGF I receptor on VSMC. 2) To characterize crosstalk between the IGF IR and other growth factors by determining the effects of changes in IGF IR concentration on growth factor-triggered distal signaling events in VSMC. 3) Exploration of the role of the IGF IR in vascular growth responses in vivo through use of expression cassettes encoding a dominant-negative IGF IR mutant or antisense IGF IR RNA. We will use recombinant adenovirus to express kinase-deficient receptors or antisense receptor RNA transcripts in VSMC. This technology will allow us to address directly the function of the IGF receptor in vascular growth in vivo. Methodologies to be used include northern and solution hybridization/RNase protection assays, in situ hybridization, gene delivery techniques through adenoviral vectors, immunohistochemistry, western ligand blotting, radioligand binding assays, radioimmunoassays, immunoprecipitation and western blotting, kinase assays, morphometry, and studies of promoter/reporter constructs. These studies are critical to understanding the function of the IGF IR as it relates to mitogenic responses to other growth factors. It should provide the basis for the development of rational approaches to address vascular complications of hypertension, atherosclerosis, post-angioplasty restenosis, and post-transplantation atherosclerosis.
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