Abstract 541: RAGE Regulates Cellular Migration and Adhesion Through Proteolysis of Its Ectodomain
Introduction Enhanced expression of the Receptor for Advanced Glycation End-products (RAGE) not only plays deleterious roles in various vascular diseases but also affects many vascular properties including endothelial permeability, leukocyte adhesion and transmigration. Decreased serum level of soluble RAGE (sRAGE), which inhibits RAGE functions, has been associated with various inflammatory diseases including atherosclerosis. Notably, several studies have shown that the release of sRAGE ectodomain (ECD) occurs by proteolysis from the cell surface. However, little is known about how this process is regulated and the functional effects of RAGE ECD shedding.
Hypothesis We hypothesize that shedding of RAGE ECD is a fundamental process regulating cell signaling pathways leading to altered cell function.
Methods & Results Toward this end, we first demonstrated that RAGE ECD shedding is a conserved mechanism in humans and mice. Next, we showed that common activators of ECD shedding, such as the protein kinase C activator PMA; the metalloproteinase-activator APMA; and the calcium ionophore ionomycin, can induce RAGE ECD release. Furthermore, various inflammatory mediators including CRP and TNF-alpha can induce RAGE ECD shedding. Inhibition studies revealed RAGE ECD shedding could be blocked by metalloproteinase, PKC, and various MAP kinase inhibitors. Finally, utilizing a proteolytically resistant splice variant of RAGE lacking the ECD shedding site (RAGEv4), we investigated the cellular function of RAGE shedding. Compared to cell expressing RAGE or empty vector, the expression of RAGEv4 resulted in impeded cell migration. Moreover, RAGEv4 reduced cellular adhesion as compared to vector control and RAGE expressing cells. Key downstream signaling pathways affected by RAGE ECD shedding are being currently investigated.
Conclusions RAGE ECD shedding is a conserved and tightly regulated mechanism involving distinct signaling pathways. Shedding of RAGE regulates key cellular properties including adhesion and migration. Our data suggest that proteolysis of RAGE is a major regulatory process and may emerge as a novel therapeutic target for cardiovascular diseases.
- © 2013 by American Heart Association, Inc.