Abstract
Nitric oxide (NO) is a free radical that plays an important role in modulating platelet adhesion and aggregation. Platelets are a source of vascular NO, but since erythrocytes avidly scavenge NO, the functional significance of platelet-derived NO is not clear. Our purpose was to determine if NO from platelets affects platelet thrombus formation in the presence of anticoagulated whole blood in an in vitro parallel plate flow system. We studied platelet adhesion and aggregation on a collagen type III surface in the presence of physiologically relevant fluid mechanical shear stress. We found that certain receptor mediated agonists (insulin and isoproterenol) caused a concentration dependent reduction in thrombus formation at a shear rate of 1000 s-1. This effect was mediated by NO since it was abolished in the presence of the NO inhibitor L-nitro-arginine-methyl-ester (L-NAME). As expected, at venous levels of shear rate (100 s-1) neither of the agonists had any effect on thrombus formation since platelet adhesion does not depend on activation at these low levels of shear. Interestingly, at a shear rate of 2000 s-1 the addition of L-NAME caused an increase in platelet coverage suggesting that shear, by itself, induces NO production by platelets. This is the first demonstration of shear stress causing platelets to produce an inhibitor of platelet activation. These results demonstrate that the development of a platelet thrombus is regulated in a complex way and that platelets produce functionally significant amounts of NO even in the presence of whole blood.
Highlights
Platelet activation and aggregation play an important role in the development of cardiovascular disease, which is the leading cause of death in the United States
We examined the effect of several external factors on platelet thrombus formation, including insulin, the β-adrenoceptor agonist isoproterenol, and shear stress
Effect of insulin and isoproterenol on guanylate cyclase activity We further investigated the mechanism of platelet derived nitric oxide inhibition of thrombus formation by using the guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo [4, 3-a]quinozalin-1-one (ODQ), since the primary target of NO in platelets is guanylate cyclase [18,19]
Summary
Platelet activation and aggregation play an important role in the development of cardiovascular disease, which is the leading cause of death in the United States. A general paradigm has emerged that endothelial cells synthesize and release substances that inhibit platelet activation (e.g. nitric oxide and prostacyclin) except near a site of vascular injury, while platelets synthesize and release substances that promote further platelet activation. In contrast to this paradigm, several recent studies have suggested that, in the presence of certain agonists (IGF-1, adenosine diphosphate, insulin, and isoproterenol), platelets will produce nitric oxide (NO), a potent inhibitor of platelet activation [1,2,3,4,5]. There is a need to determine if the NO derived from platelets is (page number not for citation purposes)
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