The intracellular mechanisms which are required to induce vascular smooth muscle cell migration and proliferation in response to injury are poorly understood. Although several receptor systems are capable of initiating smooth muscle cell migration and proliferation following the binding of ligand (bFGF, PDGF and most recently, NGF) the specific intracellular pathways which are activated by these receptor tyrosine kinases in vascular smooth muscle cells have yet to be characterized. Recent studies from our laboratory have demonstrated the expression of a novel growth factor/receptor system, the neurotrophins and their receptors, trk receptor tyrosine kinases and p75, in medial smooth muscle cells of both rat and human aorta and in cultured vascular human and rat smooth muscle cells. Moreover, their expression is exquisitely regulated during the development of vascular restenosis, as demonstrated by their expression in human atherosclerotic lesions and in the neointima which forms following balloon injury to the rat aorta. Finally, neurotrophins are potent mediators of vascular smooth muscle cell migration, in a response comparable to PDGF. The coexpression of the neurotrophins and trk receptors and their effect on smooth muscle cell migration suggests that these growth factors play an important role in regulating the response of smooth muscle cells to vascular injury. Trk is a receptor tyrosine kinase which has been shown to activate a number of intracellular signaling pathways in neuronal cells, such as the ras/MAP kinase pathway, phospholipase C (PLC-gamma) and phosphotidyl inositol 3-OH kinase (P13-kinase). The mechanisms of p75 intracellular signaling have not yet been defined, although an activation of sphingomyelinase and subsequent release of intracellular ceramide has recently been demonstrated by several laboratories. The overall aim of this grant proposal is to define the signaling pathways for smooth muscle cell migration in response to the neurotrophins. These pathways will be compared with those activated by the PDGF and bFGF receptors, to define those pathways which are required for the induction of vascular smooth muscle cell migration. Specifically, we intend to: I. Establish a vascular smooth muscle cell system to express trk receptors, as well as other receptor tyrosine kinases implicated in initiating vascular smooth muscle cell migration using gene transfer techniques. II. Identify the post-receptor pathways which mediate smooth muscle cell migration using cells which express trk, PDGF-beta and FGF-1 receptor tyrosine kinases. Mutant receptors, incapable of activating defined signaling enzymes, will be used to confirm that these specific pathways mediate cell migration. III. Examine the role of p75 receptor signal transduction in neurotrophin- mediated smooth muscle cell migration. These studies involve critical interactions with Dr. Gross, and Drs. Hajjar, Pomerantz, and Lander.
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