Factor VIII as Positive Regulator of Activated Platelets

Abstract

Deficiency of factor VIII (FVIII) as an important cofactor in the tenase coagulation factor complex causes severe bleeding, whereas high FVIII levels seem to be associated with venous and arterial thromboembolism. Resting platelets do not bind FVIII, but activated platelets bind inactivated FVIII if vWF is not present. In this study the influence of platelet bound FVIII on platelet function itself was investigated as a direct effect of FVIII on platelets has not been shown so far. Methods: The influence of platelet bound FVIII on platelet function was investigated by flow cytometric analysis of P-selectin expression (CD62P) and PAC-1 binding before and after submaximal stimulation with TRAP-6 (5 μM final concentration) and by confocal microscopy. Blood was incubated with human recombinant FVIII concentrate (Helixate, ZLB Behring, Marburg, Germany) for 5 minutes at 37 °C. Results: There were no significant changes in P-selectin expression or PAC-1 binding respectively in FVIII-pretreated platelets. Stimulation of platelets with TRAP-6 increased the expression of Pselectin (445 %) and PAC-1 binding (934 %) as expected. These effects were significantly further increased when platelets were stimulated with TRAP-6 and FVIII (Pselectin 499 %, PAC-1 1626 %; Student's t-test, p < 0.05. Values were expressed in %, related to unstimulated, Helixate buffer treated platelets). The assessment of platelet spreading by the means of confocal microscopy showed an increased spreading on fibrinogen matrix after stimulation with TRAP-6 and FVIII. This effect was higher, compared to platelets stimulated with TRAP-6 alone. Conclusions: TRAP-co-stimulated platelets seem to be positively regulation by additional FVIII. Thus a FVIIIinduced increase in platelet activation might contribute to venous and even more arterial thrombus formation in patients with high FVIII levels.