Characterization of mechanisms through which coagulation proteases differentially regulate hyperglycemia and hyperlipidemia induced accelerated atherosclerosis.

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The role of coagulation proteases, in particular protease dependent signalling, for the establishment of atherosclerosis is unknown. Thrombin and activated protein C (PC) are two key regulators of the coagulation system with opposing effects in regard to coagulation activation. They provoke differential intracellular signalling despite activation of the same receptor, protease activated receptor 1 (PAR-1). Activity of thrombin and activated PC is determined by the thrombin binding endothelial protein thrombomodulin (TM), which inhibits thrombin and activates PC. Thus, TM has a key role in determining activity of thrombin, activated PC and secondary effects mediated through these proteases. Expression of receptors for coagulation proteases (PAR´s, TM, EPCR (endothelial protein C receptor)) has been established in cell-types relevant for cardiovascular disease. However, the mechanisms through which TM, thrombin and activated PC evoke differential intracellular signalling and the relation to chronic cardiovascular disease are not understood. We have been able to show that TM modulates atherosclerosis and cardiac hypertrophy in vivo. In vitro results suggest that TM might modulate cellular function (e.g. NF-kB activation, expression of adhesion molecules) through two mechanisms: (A) inhibition of thrombin dependent cell activation and (B) promotion of activated PC dependent cytoprotection. The aim of the proposal is to identify the mechanism through which TM modulates differential cell activation and cardiovascular disease. Using in vivo and in vitro experiments we intend to: 1) Identify the mechanism through which thrombin and activated PC differentially regulate NF-kB activation and expression of adhesion molecules; 2) Determining whether TM has direct signalling properties; 3) Define the role of TM-dependent regulation of thrombin and activated PC for atherosclerosis in vivo; 4) Determine whether the TM-PC system regulates cardiac hypertrophy indirectly (via regulation of arterial blood pressure) or directly through receptor dependent mechanisms. The current proposal will provide an in depth analysis of the mechanisms through which thrombin, activated PC, and TM regulate cardiovascular disease. Animal models of accelerated atherosclerosis (high fat diet or hyperglycemia) and with genetically altered activity of the TM-PC system will be used. These in vivo experiments will be paired with complementary in vitro studies. The long term aim of the proposed studies is an in depth understanding of the mechanisms through which the coagulation system regulates cardiovascular disease. This work will lay ground for novel targeted therapeutic intervention based on the mechanisms regulated by coagulation proteases, but circumventing associated problems such as increased risk of hemorrhage.