Studies on the activation mechanisms of MMP-2 and ADAM17: identification of regulatory proteins, oxidant production pathways, epigenetics and proteolytic targets

Abstract

The role of Matrix Metalloproteinases (MMPs) in the degradation and extracellular matrix remodeling is but one aspect of the role played by this family of proteases that has 27 members. Once activated, the MMPs cleave cleave substrates such as matrix proteins, proinflammatory mediators, and growth factors, which will activate several signalling passways that depend on MAP kinases, and are important in processes of vascular tonus regulation, cardiac hypertrophy, and response to lesion. It is now known that activation of MMPs by agonists is a fast process that involves PKC, production of oxidants and other MMPs (like membrane-type MMPs), but it is still necessary to know in more detail about the activation/functions of each MMP, specially because of the great therapeutic potential of these metaloenzymes. Furthermore, MMP-7 has been described to increase the concentrations of oxidants, a very important finding, since it changes the paradigm nowadays used to think about the regulation of cardiovascular effects evoked by MMPs and redox processes. In this thematic project we aim at studying MMP-2 and ADAM17 in the cardiovascular system, with particular interest in the mechanism of action, intracellular oxidant production, epigenetic regulation, and the identification of novel substrates. We have 4 main questions: 1- to study a few of the mechanisms by which MMP-2 is associated with an increase in oxidants, and how this increase is associated to altered function of smooth muscle cells in culture, in an ex-vivo model (in rabbit aortas) and in experimental atherosclerosis in mice; 2- to elucidate the activation of ADAM17 by thioredoxin-1 (Trx-1), both in the direct interaction between the proteins as well as in the redox regulation of Trx-1; 3- to study which epigenetic alterations (genomic DNA methylation) are associated with the increase in the expression of MMP-2 in the heart/kidney of mice; 4- to analyse the secretome of cardiomyocytes exposed to ADAM17 or MMP-2 to be able to depict the proteolytic cascades that are specific for each of these two metaloproteinases. The 4 subprojects are related to each other and are also complementary, both conceptually and methodologically. We propose 4 complexity levels regarding experimental approaches. The results obtained here will contribute for the advancement of the understanding of the interfaces between redox processes and metaloproteinases, and will also enable the identification of pathways activated specifically by MMP-2 and ADAM17 in the cardiovascular system, with potential implication for the design of future interventions. (AU)