Cytoskeletal perturbation leads to platelet dysfunction and thrombocytopenia in variant forms of Glanzmann thrombasthenia

Several patients with variant dominant forms of Glanzmann Thrombasthenia, associated with macrothrombocytopenia and caused by gain-of-function mutations of ITGB3 or ITGA2B leading to reduced surface expression and constitutive activation of integrin αIIbβ3, have been reported. The mechanisms leading to a bleeding phenotype of these patients have never been addressed. Aim of the present study was to unravel the mechanism by which ITGB3 mutations causing activation of αIIbβ3 lead to platelet dysfunction and macrothrombocytopenia. Here we show, using platelets from two patients carrying the β3 del647-686 mutation and CHO cells expressing different αIIbβ3-activating mutations, that reduced surface expression of αIIbβ3 is due to receptor internalization. Moreover, we demonstrate that permanent triggering of αIIbβ3-mediated outside-in signaling causes an impairment of cytoskeletal reorganization arresting actin turnover at the stage of polymerization. The induction of actin polymerization by jasplakinolide, a natural toxin that promotes actin nucleation and prevents stress fibers depolymerization, in control platelets produced an impairment of platelet function similar to that of patients with dominant Glanzmann Thrombasthenia variants. del647-686β3-transduced murine megakaryocytes, generated proplatelets with a reduced number of large tips and asymmetric barbell-proplatelets, suggesting that impaired cytoskeletal rearrangement is the cause of macrothrombocytopenia. These data show that impaired cytoskeletal remodeling caused by a constitutively activated αIIbβ3 is the main effector of platelet dysfunction and macrothrombocytopenia, and thus of bleeding, in dominant Glanzmann Thrombasthenia variants.