Emerging evidence suggests that matrix metalloproteases (MMPs) play a pivotal role in vascular integrity and wound healing, however, inappropriate MMP activity may underlie the pathogenesis of several diseases including restenosis, atherosclerosis, and myocardial dysfunction. Selectively controlling defined aspects of MMP activity in restenosis and end-stage coronary artery disease may be a valid target for therapy. We recently made the unanticipated discovery that the matrix metalloprotease MMP-1, cleaves and activates protease-activated receptor 1 (PAR1) in a variety of cells including endothelium, vascular SMCs, platelets and carcinoma cells. Both MMP-1 and PAR1 are highly upregulated by several cell types in the blood vessel wall in the context of intimal hyperplasia and inflammatory states. However, it is completely unknown what role(s), MMP1-PAR1 signaling plays in either normal blood vessel biology or in vascular disease. Moreover, activation of PAR1 can lead to decreased barrier function, dysregulated vasomotor tone and control of SMC functions through inter-cellular communication pathways.
In aim 1, we will investigate the molecular mechanism and (patho)physiologic relevance of MMP1-PAR1 signaling in SMC proliferation, migration and invasion, survival and de-differentiation in the arterial wall. Modulation of PAR1 signaling with cell- penetrating pepducins will be used to evaluate the function(s) of MMP1-PAR1 signaling in in vivo arterial injury and restenosis models using wild-type mice and strains deficient in PAR1, PAR2 and PAR4. Furthermore, prior studies suggested that PAR1 promotes the conversion of the endothelium to a pro- thrombotic and pro-inflammatory state. We made the recent observation that PAR1 can also confer protective effects in endothelial barrier function and survival in animal models of systemic inflammation by transactivation of the structurally-related PAR2. Our preliminary data suggest that the signaling outputs of PAR1 and PAR2 are entwined and need to be examined as a potential functional unit.
Aim 2 will use a combination of biochemical, pharmacological and RNA interference techniques to determine how PAR1 and PAR2 regulate Rac versus Rho signaling. We will delineate the components and kinetics of downstream pro- inflammatory versus anti-inflammatory signaling pathways emanating from PAR1 and PAR2 in the context of barrier disruption and leukocyte transmigration. Critical downstream chemokine pathways known to mediate both autocrine and paracrine signaling during these processes will be interrupted with our newly validated chemokine receptor pepducins. Lastly, we will investigate the molecular basis of PAR1-PAR2 interactions and liganding using biochemical and genetic approaches. Relevance to public health: The studies outlined here may lead to the eventual discovery of novel therapeutics that may prevent heart attacks and stroke in patients with heart disease, and could be used in other conditions such as life-threatening sepsis.
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