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Thrombosis is the main cause of morbidity and mortality in patients with JAK2 V617F myeloproliferative neoplasms. Recent studies have reported the presence of JAK2 V617F in endothelial cells of some patients with myeloproliferative neoplasms. We investigated the role of endothelial cells that express JAK2 V617F in thrombus formation using an in vitro model of human endothelial cells overexpressing JAK2 V617F and an in vivo model of mice with endothelial-specific JAK2 V617F expression. Interestingly, these mice displayed a higher propensity for thrombus. When deciphering the mechanisms by which JAK2 V617F -expressing endothelial cells promote thrombosis, we observed that they have a pro-adhesive phenotype associated with increased endothelial P-selectin exposure, secondary to degranulation of Weibel-Palade bodies. We demonstrated that P-selectin blockade was sufficient to reduce the increased propensity of thrombosis. Moreover, treatment with hydroxyurea also reduced thrombosis and decreased the pathological interaction between leukocytes and JAK2 V617F -expressing endothelial cells through direct reduction of endothelial P-selectin expression. Taken together, our data provide evidence that JAK2 V617F -expressing endothelial cells promote thrombosis through induction of endothelial P-selectin expression, which can be reversed by hydroxyurea. Our findings increase our understanding of thrombosis in patients with myeloproliferative neoplasms, at least those with JAK2 V617F -positive endothelial cells, and highlight a new role for hydroxyurea. This novel finding provides the proof of concept that an acquired genetic mutation can affect the pro-thrombotic nature of endothelial cells, suggesting that other mutations in endothelial cells could be causal in thrombotic disorders of unknown cause, which account for 50% of recurrent venous thromboses., (Copyright© 2019 Ferrata Storti Foundation.)

Glycoprotein VI, a major platelet activation receptor for collagen and fibrin, is considered a particularly promising, safe antithrombotic target. In this study, we show that human glycoprotein VI signals upon platelet adhesion to fibrinogen. Full spreading of human platelets on fibrinogen was abolished in platelets from glycoprotein VI- deficient patients suggesting that fibrinogen activates platelets through glycoprotein VI. While mouse platelets failed to spread on fibrinogen, human-glycoprotein VI-transgenic mouse platelets showed full spreading and increased Ca 2+ signaling through the tyrosine kinase Syk. Direct binding of fibrinogen to human glycoprotein VI was shown by surface plasmon resonance and by increased adhesion to fibrinogen of human glycoprotein VI-transfected RBL-2H3 cells relative to mock-transfected cells. Blockade of human glycoprotein VI with the Fab of the monoclonal antibody 9O12 impaired platelet aggregation on preformed platelet aggregates in flowing blood independent of collagen and fibrin exposure. These results demonstrate that human glycoprotein VI binds to immobilized fibrinogen and show that this contributes to platelet spreading and platelet aggregation under flow., (Copyright © 2018 Ferrata Storti Foundation.)