This application for a FIRST Award examines the role of activation of protein kinase C in immediate early gene expression following balloon catheter-induced injury of the rat thoracic aorta (BAL). Enhanced smooth muscle cell (SMC) proliferation and migration account for the intimal thickening which results from BAL of the rat thoracic aorta. The imposition of stretch upon SMC is inevitable in BAL, which produces a greater SMC proliferative response as compared to endothelial denudation alone. Protein kinase C (PKC) inhibitors and calcium channel blockers are known to modulate SMC proliferation in vitro, and the latter compounds reduce proliferation following BAL in vivo. The fundamental goal of this application is to delineate early calcium dependent signal transduction mechanisms responsible for stretch-induced activation of quiescent SMC in vivo, which in association with humoral factors, results in activation of the SMC cell cycle. The first two Aims center upon BAL stretch-induced regulation of PKC in an in situ perfused aortic preparation.
Aim 1 covers the time course and calcium dependence for stretch-induced activation and membrane translocation of both the calcium sensitive and insensitive PKC isoforms.
Aim 2 characterizes the time course and requirements for biphasic generation of the PKC cofactor diacylglycerol and inositol 3,4,5-trisphosphate. These findings will be extended using SMC in vitro in Aim 3.
This Aim will examine translocation and down regulation of specific PKCs during the G1 phase of the cell cycle in vitro. The SMC in the vessel wall possesses a """"""""contractile"""""""" phenotype whose growth is inhibited by heparinoid molecules associated with the extracellular matrix. Compared to proliferating """"""""synthetic"""""""" SMC in vitro, the perfusion model is more relevant in that stretch-related signal transduction mechanisms in situ are more likely to mimic the earliest activation response of quiescent SMC in vivo. It is recognized the PKC and cell cycle studies in the in vitro system reflects growth control in cells that are persistently proliferative and in which the modeling of stretch may be limited. However, these studies serve as a basis of subsequent investigation of the role of PKC on the regulation of the cell cycle in differentiated vessel wall SMC.
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