SCUBE1 (signal peptide-CUB-EGF domain-containing protein 1), expressed in endothelial cells (ECs) and platelets, exists in soluble or membrane forms. We previously showed that soluble SCUBE1 is a biomarker for platelet activation and also an active participant of thrombosis. However, whether the adhesive module of its EGF-like repeats is essential and the specific contribution of SCUBE1 synthesized in ECs or platelets to thrombosis in vivo remain unclear.

We generated new mutant (∆2) mice lacking the entire EGF-like repeats to evaluate the moduleʼs functional importance during thrombogenesis in vivo. ∆2 platelet-rich plasma showed markedly impaired platelet aggregation induced by agonists including ADP, collagen, the thrombin agonist PAR-4 peptide and the thromboxane A2 analog U46619. Consistently, genetic ablation of the EGF-like repeats diminished arterial thrombosis and protected ∆2 mice against lethal thromboembolism. On flow chamber assay, whole blood isolated from ∆2 or wild-type (WT) mice pretreated with blocking antibodies against the EGF-like repeats showed a significant decrease in platelet deposition and thrombus formation on collagen-coated surfaces under arterial shear rates. Moreover, we created animals expressing SCUBE1 only in ECs (S1-EC) or platelets (S1-PLT) by reciprocal bone-marrow transplantation between WT and ∆2 mice. The time of carotid arterial thrombosis induced by ferric chloride was normal in S1-PLT chimeric mice but much prolonged in S1-EC animals.

We demonstrate that platelet-derived SCUBE1 plays a critical role in arterial thrombosis via its adhesive EGF-like repeats in vivo and suggest targeting these adhesive motifs of SCUBE1 for potential anti-thrombotic strategy.