The role of 12-lipoxygenase in regulating the effects of EPA on platelet activation and thrombosis

PROJECT SUMMARY Cardiovascular disease (CVD) is the leading cause of death in the US, accounting for nearly 1 in 4 deaths and over $400 billion in annual health care cost in the US. Platelet-mediated thrombosis is most common underlying pathophysiology of cardiovascular diseases and leads to life-threatening clinical cardiovascular events, such as myocardial infarction or stroke. Under normal physiological conditions, platelet activation is critical for maintaining normal blood flow following vascular injury, however in pathologic conditions hyperactive platelets can form occlusive thrombi. Fish oil supplements are one of the most common over-the-counter supplements due to previous research showing their ability to slow the progression of cardiovascular disease. Fish oil supplements are highly enriched in the omega-3 polyunsaturated fatty acid (PUFA), eicosapentaenoic acid (EPA). Supplementation with EPA alone reduces the risk for major cardiovascular events, suggesting the cardiovascular protection is regulated by EPA, however the exact mechanisms regulating these benefits remain unknown. Our group has shown that several PUFAs inhibit platelet activation, and these antiplatelet effects are realized through the production of 12-lipoxygenase (12-LOX) metabolites. Further, we have shown both EPA and its 12-LOX metabolite, 12-hydroxyeicosapentaenoic acid (HEPE) inhibit platelet activation. However, the exact signaling mechanisms regulating the antiplatelet effects of EPA and 12-HEPE, as well as the role of 12-LOX in regulating the effects of EPA on thrombosis are unknown. We will use both pharmacological and genetic techniques to determine the role of peroxisome proliferator activated receptors (PPARs) and Gαs-coupled GPRCs in regulating the antiplatelet effects of EPA and 12-HEPE, both of which have been implicated in the signaling of other 12- LOX metabolites. Finally, we will investigate the effects of supplementation with EPA and icosapent ethyl (IPE), an FDA approved esterified form of EPA, on the ability to alter plasma 12-HEPE concentrations, platelet membrane lipid composition, ex vivo platelet activation, in vivo thrombosis, and hemostasis. Additionally, 12- LOX knockout mice will be utilized to determine if 12-LOX is required for the regulation of platelet activation and thrombosis by EPA and IPE. Successful completion of this project will, for the first time, elucidate the mechanisms regulating the ability of EPA to alter platelet function. The findings will be used to inform the use of fish oil supplements and IPE as a therapeutic intervention for thrombotic diseases to improve cardiovascular outcomes. Improved use of already available and widely used supplements could reduce the overall burden of CVD.