GPV is a marker of in vivo platelet activation - Study in a rat thrombosismodel

Thrombin plays a central role in the genesis of thrombotic events and is th e most potent known platelet agonist. This enzyme activates platelets by cl eaving G-protein coupled protease activated receptors (PARs) and by binding to glycoprotein (GP) Ib. Thrombin also cleaves platelet GPV to liberate a soluble 69 kDa fragment (GPVIf1), leaving a 20 kDa fragment (GPVf2) attache d to the membrane. The aim of this study was to assess the value of GPV as an in vivo marker of the activation of platelets by thrombin. Newly develop ed monoclonal and polyclonal antibodies recognizing rat GPVf1 and GPVf2 res pectively were used to detect soluble GPV by ELISA and the new NH2-terminus exposed by thrombin using flow cytometry. These assays were employed in a rat thrombosis model designed to trigger thrombin formation in vivo. When t hromboplastin (4.8 ml/kg/h) was infused for 30 min, thrombin generation was reflected by a rapid increase in thrombin-antithrombin (TAT) complexes in plasma and by the appearance of GPVf2 at the surface of circulating platele ts. Simultaneously, GPVf1 disappeared from the surface of platelets and acc umulated as a soluble fragment in plasma, where it was detected by GPV ELIS A. These effects were inhibited by pretreatment of the rats with hirudin. L evels of plasma PF4 also increased in this model, but unlike GPV levels whi ch returned slowly (> 2 hours) to baseline, PF4 had a very short half-life. In conclusion, GPV is cleaved by thrombin in vivo, circulates and is a reli able in vivo marker of the activation of platelets by thrombin. Monitoring of GPV levels in rats should be useful to evaluate the effects of antithrom bolic and antiplatelet drugs, while further studies will be required to con firm the potential interest of GPV as a marker of thrombotic states in huma ns.