Antithrombotic properties of pravastatin reducing intra-thrombus fibrin deposition under high shear blood flow conditions

Summary

In addition to lowering cholesterol, the 3-hydroxyl-3-methylglutaryl coenzyme A reductase inhibitors (statins) have a range of pleiotropic effects that help reduce the risk of adverse cardiovascular events. We sought to understand the molecular mechanisms by which statins could exert anti-platelet actions under physiologic whole blood flow conditions. Using an in vitroperfusion chamber system, we examined the anti-platelet effects of pravastatin under whole blood flow conditions with high or low shear rates. We determined that pravastatin significantly suppressed platelet activation-dependent procoagulant activity, decreasing P-selectin membrane expression, tissue factor accumulation, and thrombin binding within platelet thrombi generated on a von Willebrand factor-surface under high shear rate conditions. These effects resulted in reductions of intra-thrombus fibrin deposition. These antithrombotic properties of pravastatin, which were comparable to those of atorvastatin, could be abrogated by mevalonate. Our experimental approach revealed a novel mechanism mediating the anti-platelet action of statins. Shear rate-dependent antithrombotic activity may explain the favourable effect of pravastatin on the reduction in cardiovascular events that typically occur in vivounder whole blood flow conditions with high shear rates.

Keywords

Statins - high fluid shear - thrombus - von Willebrand factor - fibrinogen/fibrin

• References

• 1 Maron DJ, Fazio S, Linton MF. Current perspectives on statins. Circulation 2000; 101: 207-213.
• 2 Baigent C, Keech A, Kearney PM. et al. Efficacy and safety of cholesterol-lowering treatment: prospective meta-analysis of data from 90,056 participants in 14 randomised trials of statins. Lancet 2005; 366: 1267-1278.
• 3 Wolfrum S, Jensen KS, Liao JK. Endothelium-dependent effects of statins. Arterioscler Thromb Vasc Biol 2003; 23: 729-736.
• 4 Diamant M, Tushuizen ME, Abid-Hussein MN. et al. Simvastatin-induced endothelial cell detachment and microparticle release are prenylation dependent. Thromb Haemost 2008; 100: 489-497.
• 5 Kleemann R, Verschuren L, de Rooij B-J. et al. Evidence for anti-inflammatory activity of statins and PPAR? activators in human C-reactive protein transgenic mice in vivo and in cultured human hepatocytes in vitro. Blood 2004; 103: 4188-4194.
• 6 Kruithof EK. Regulation of plasminogen activator inhibitor type 1 gene expression by inflammatory mediators and statins. Thromb Haemost 2008; 100: 969-975.
• 7 Waters D, Higginson L, Gladstone P. et al. Effects of monotherapy with an HMGCoA reductase inhibitor on the progression of coronary atherosclerosis as assessed by serial quantitative arteriography: the Canadian Coronary Atherosclerosis Intervention Trial. Circulation 1994; 89: 959-968.
• 8 Sanguigni V, Pignatelli P, Lenti L. et al. Short-term treatment with atorvastatin reduces platelet CD40 ligand and thrombin generation in hypercholesterolemic patients. Circulation 2005; 111: 412-419.
• 9 Redecha P, van Rooilen N, Torry D. et al. Pravastatin prevents miscarriages in mice: role of tissue factor in placental and fatal injury. Blood 2009; 113: 4101-4109.
• 10 Aberg M, Johnell M, Wickström M. et al. Simvastatin reduces the production of prothrombotic prostasomes in human prostate cancer cells. Thromb Haemost 2008; 100: 655-662.
• 11 Ramacciotti E, Hawley AE, Farris DM. et al. Leukocyte- and platelet-derived microparticles correlate with thrombus weight and tissue factor activity in an experimental mouse model of venous thrombosis. Thromb Haemost 2009; 101: 748-754.
• 12 Sugimoto M, Mohri H, McClintock RA. et al. Identification of discontinuous von Willebrand factor sequences involved in complex formation with botrocetin. A model for the regulation of von Willebrand factor binding to platelet glycoprotein Ib. J Biol Chem 1991; 266: 18172-18178.
• 13 Kansas G. Selectins and their ligands: Current concepts and controversies. Blood 1996; 88: 3259-3287.
• 14 Crawley J, Lupu F, Westmuckett AD. et al. Expression, localization, and activity of tissue factor pathway inhibitor in normal and atherosclerotic human vessels. Arterioscler Thromb Vasc Biol 2000; 20: 1362-1373.
• 15 Abcam Inc. product data sheet: Protocol for mouse monoclonal [109–04] to thrombin (ab17199). Available at http://www.abcam.com/Thrombin-antibody-5G9-ab17199.html
• 16 Kudryk B, Rohoza A, Ahadi M. et al. Specificity of a monoclonal antibody for the NH2-terminal region of fibrin. Molec Immun 1984; 21: 89-94.
• 17 Mizuno T, Sugimoto M, Matsui H. et al. Visual evaluation of blood coagulation during mural thrombogenesis under high shear flow. Thromb Res 2008; 121: 855-864.
• 18 Matsui H, Sugimoto M, Mizuno T. et al. Distinct and concerted functions of von Willebrand factor and fibrinogen in mural thrombus growth under high shear flow. Blood 2002; 100: 3604-3610.
• 19 Sugimoto M, Matsui H, Mizuno T. et al. Mural thrombus generation in type 2A and 2B von Willebrand disease under flow conditions. Blood 2003; 101: 915-920.
• 20 Shida Y, Nishio K, Sugimoto M. et al. Functional imaging of shear-dependent activity of ADAMTS13 in regulating mural thrombus growth under whole blood flow conditions. Blood 2008; 111: 1295-1298.
• 21 Giessen PLA, Rauch U, Bohmann B. et al. Blood-borne tissue factor: another view of thrombosis. Proc Natl Acad Sci USA 1999; 96: 2311-2315.
• 22 Balasubramanian V, Vele O, Nemerson Y. Local shear conditions and platelet aggregates regulate the incorporation and activity of circulating tissue factor in exvivo thrombi. Thromb Haemost 2002; 88: 822-826.
• 23 Falati S, Liu Q, Gross P. et al. Accumulation of tissue factor into developing thrombi in vivo is dependent upon microparticle P-selectin glycoprotein ligand 1 and platelet P-selectin. J Exp Med 2003; 197: 1585-1598.
• 24 Undas A, Brummel KE, Musial J. et al. Simvastatin depresses blood clotting by inhibiting activation of prothrombin, factor V, and factor XIII and by enhancing factor Va inactivation. Circulation 2001; 103: 2248-2253.
• 25 Dajani EZ, Shahwan TG, Dajani NE. Statins, platelet aggregation and coronary heart disease. J Assoc Acad Minor Phys 2002; 13: 27-31.
• 26 Cortellaro M, Cofrancesco E, Arbustini E. et al. Atorvastatin and thrombogenicity of the carotid atherosclerotic plaque: the ATROCAP study. Thromb Haemost 2002; 88: 41-47.
• 27 Santos MT, Fuset MP, Ruano M. et al. Effect of atorvastatin on platelet thromboxane A(2) synthesis in aspirin-treated patients with acute myocardial infarction. Am J Cardiol 2009; 104: 1618-1623.
• 28 Savage B, Almus-Jacobs F, Ruggeri ZM. Specific synergy of multiple substrate-receptor interactions in platelet thrombus formation under flow. Cell 1998; 94: 657-666.
• 29 Tsuji S, Sugimoto M, Miyata S. et al. Real-time analysis of mural thrombus formation in various platelet aggregation disorders: distinct shear-dependent roles of platelet receptors and adhesive proteins under flow. Blood 1999; 94: 968-975.
• 30 Savage B, Saldivar E, Ruggeri ZM. Initiation of platelet adhesion by attachment onto fibrinogen or translocation on von Willebrand factor. Cell 1996; 84: 289-297.
• 31 Kuwahara M, Sugimoto M, Tsuji S. et al. Cytosolic calcium changes in a process of platelet adhesion and cohesion on a von Willebrand factor-coated surface under flow conditions. Blood 1999; 94: 1149-1155.
• 32 Kuwahara M, Sugimoto M, Tsuji S. et al. Platelet shape changes and adhesion under high shear flow. Arterioscler Thromb Vasc Biol 2002; 22: 329-334.
• 33 Fuster V, Badimon L, Badimon JJ. et al. The pathogenesis of coronary artery disease and the acute coronary syndromes (part I). N Engl J Med 1992; 326: 242-250.
• 34 Pan HY, DeVault AR, Wang-Iverson D. et al. Comparative pharmacokinetics and pharmacodynamics of pravastatin and lovastatin. J Clin Pharmacol 1990; 30: 1128-1135.
• 35 Ohkawara H, Ishibashi T, Sakamoto T. et al. Thrombin-induced rapid geranylgeranylation of RhoA as an essential process for RhoA activation in endothelial cells. J Biol Chem 2005; 280: 10182-10188.