Platelets compensate for poor thrombin generation in type 3 von Willebrand disease

Abstract

In type 3 von Willebrand disease (VWD3), the most severe form with absent von Willebrand factor (VWF), the bleeding phenotype is variable. Platelet contribution to the hemostatic defect in VWD3 calls upon further studies. We investigated the contribution of platelets to in vitro thrombin generation (TG) and platelet procoagulant activity in VWD3. TG was assessed by calibrated automated thrombogram (CAT) in platelet-poor (PPP) and –rich plasma (PRP) from 9 patients before and in 6 patients also 30 min after receiving their regular VWF therapy. Responsiveness of PPP to FVIII and protein S was also investigated. TG data were compared with routine laboratory variables, rotational thromboelastometry (ROTEM) and platelet expression of P-selectin and phosphatidylserine in flow cytometry. Compared with healthy controls, TG was markedly decreased in VWD3 PPP (peak thrombin was 16% of normal median), but not in PRP (77% of normal median) (p = 0.002). Six out of nine patients (67%) were high responders in their platelet P-selectin, and 5/9 (56%) in phosphatidylserine expression. Replacement therapy improved TG in PPP, while in PRP TG only modestly increased or was unaffected. In PPP, FVIII levels associated with TG and in vitro FVIII-supplemented TG inclined up to threefold. Conversely, a FVIII inhibitory antibody reduced plasma TG in all, but especially in patients with remnant FVIII levels. Inhibition of protein S improved plasma TG, particularly at low FVIII levels. ROTEM failed to detect VWD3.

In VWD3, TG is reduced in PPP and regulated by FVIII and protein S, but TG is close to normal in PRP. VWD3 platelets seem to compensate for the FVIII-associated reduction in TG by their exposure of P-selectin and phosphatidylserine.

Keywords:

• FVIII
• platelet procoagulant activity
• protein S
• thrombin generation
• von Willebrand disease type 3
• von Willebrand factor

Acknowledgements

We would like to acknowledge our VWD3 patients in their participation to this study. We acknowledge the research group of Prof. Per Morten Sandset (Oslo University Hospital, Research Institute for Internal Medicine) for TFPI measurements.

Statement of contribution

TS and RL designed the research study. TS and VN performed the research and analyzed the data, AJ performed part of the TG measurements in PRP and ROTEM, and HJB performed TG in plasmas spiked with FVIII, VK3 and CLB-PS13. All authors contributed to the interpretation of the results. The first draft was written by VN, RL, and TS, and all authors contributed to and edited all subsequent drafts. TS and VN report equal contribution.

Supplementary material

Supplemental data for this article can be accessed here.

Additional information

Funding

This work was supported by the State funding for university level health research, Finland, Helsinki University Hospital (EVO 2015223); CSL Behring [The CSL Behring Nordic von Willebrand Scholarship].