Cardiovascular disease is a major cause of morbidity and mortality. A major complication in mamy forms of therapy for cardiovascular disorders is restenosis resulting from intimal hyperplasia. Intimal hyperplasia occurs when the intravascular healing process to vascular injury goes out of control. Division and migration of medial smooth muscle cells across the internal elastic lamina are important features of intimal hyperplasia. Although the molecular mechanisms regulating smooth muscle proliferation and migration are incompletely understood, the conversion of smooth muscle cells from a contractile, quiescent state to a proliferative, migratory, synthetic state appears to be important. This conversion is a complex process that appears to result from the activation of pro-proliferative and/or the inactivation of anti-proliferative signalling pathways. Studies on smooth muscle and other cells have shown that small G proteins and the activation of the MAP kinase family of enzymes play central roles in regulating proliferation and migration. Small G protein/MAP kinase signalling involves changes in the actin cytoskeleton and myosin II has been implicated in these changes. The actin-myosin II interaction in smooth muscle is regulated by myosin light chain kinase. Myosin light chain kinase is also thought to regulate the timing of mitosis. The investigators have recently found that myosin light chain kinase is phosphorylated and differentially regulated by site specific phosphorylation by two different members of the MAP kinase family of proteins. Myosin light chain kinase activity by site-specific phosphorylation by extracellular receptor kinase or p21 activated kinase is one of the key events regulating smooth muscle cells proliferation.
The specific aims are to a) characterize the proliferation of smooth muscle cells and the formation of a neo-intima when blood vessels are cultured, b) establish the temporal sequences of MAP kinase activation/inactivation, changes in the phosphorylation and activity of myosin light chain kinase and smooth muscle proliferation and c) test the hypothesis by expressing the catalytic domain of myosin light chain kinase or constitutively-active p21 activated kinase on smooth muscle proliferation. These experiments will be performed on cultured blood vessels, which develop a neo-intima when cultured under defined conditions, and on isolated smooth muscle cells. Thus, these experiments will test a novel hypothesis, they will be performed on a potentially important model of intimal proliferation and they will increase our understanding of the molecular mechanisms that regulate smooth muscle proliferation. This, in turn could result in improved therapy for intimal hyperplasia.
