Deficient platelet-derived nitric oxide and enhanced hemostasis in mice lacking the NOSIII gene

Endothelial nitric oxide synthase (eNOS) has been identified inhuman platelets. Although platelet-derived nitric oxide (NO) has been shown to inhibit platelet recruitment in vitro, its role in the regulation of the hemostatic ;response in vivo has not been characterized. To define the role of platelet-derived NO in vivo, we studied mice that lacked a functional eNOS gene (NOSIII). Surface P-selectin expression in platelets from eNOS-deficient mice was not significantly altered; however, bleeding times were markedly decreased in eNOS-deficient versus wild-type mice (77.2+/-3 versus 133.4+/-3 seconds, P<0.00005). To determine the contribution of endothelium- versus platelet-derived NO to the bleeding time, isolated platelets from either eNOS-deficient or wild-type mice were transfused into a thrombocytopenic eNOS-deficient mouse and the bleeding time was measured. The bleeding times in mice transfused with eNOS-deficient platelets were significantly decreased compared with mice transfused with wild-type platelets (Delta bleeding time, -24.6 +/- 9.1 and -3.4+/-5.3 seconds, respectively; P<0.04). Platelet recruitment was studied by measuring serotonin release from a second recruitable population of platelets that were added to stimulated platelets at the peak of NO production. There was 40.3+/-3.7% and 52.0+/-2.1% serotonin release for platelets added to wild-type or eNOS-deficient platelets, respectively (P<0.05). In summary, mice that lacked eNOS had markedly decreased bleeding times even after endothelial NO production was controlled. These data suggest that the lack of platelet-derived NO alters in vivo hemostatic response by increasing platelet recruitment. Thus, these data support a role for platelet-derived NO production in the regulation of hemostasis.