Abstract Number: PB0905
Meeting: ISTH 2021 Congress
Background: The bulk of von Willebrand factor (VWF) in plasma is synthesized in endothelial cells. During biosynthesis, VWF dimerizes in the ER and multimerizes in the Golgi via various intra- and interchain cysteines in the CK and D3 domain, respectively. Mutations in the D3 domain can lead to von Willebrand’s disease (VWD) type 2A, which is characterized by a striking loss of high molecular weight (HMW) VWF multimers in plasma. Whether this arises due to reduced half-life in circulation or is caused by biosynthetic defects is incompletely understood.
Aims: To study VWF trafficking and secretion in endothelial colony forming cells (ECFCs) from VWD 2A patients with mutations in the D3 domain.
Methods: ECFCs were isolated from venous blood of healthy control donors and three patients from the Willebrand in the Netherlands (WiN) cohort with heterozygous c.3569 G>A (p.C1190Y) and c.3568 T>C (p.C1190R) mutations in VWF. Secretion of VWF by ECFCs was determined by ELISA and multimer assay while morphology of intracellular compartments was assessed using confocal microscopy. The study was approved by the medical ethical committee of the Erasmus MC and all patients gave informed consent.
Results: Unstimulated (basal and constitutive) release of VWF was ~50% lower in patient ECFCs compared to healthy ECFCs. In addition, patient ECFCs secreted less VWF after Ca2+– and cAMP-dependent stimulation, although equal amounts of VWF were present intracellularly. A large pool of intracellular VWF colocalized with protein disulfide isomerase-positive spherical structures (Figure 1), suggestive of retention in the ER. In addition, patient ECFCs no longer secreted HMW VWF multimers, resembling the loss of HMW multimers in their plasma.
Conclusions: These findings suggest that mutations in the D3 domain of VWF (p.C1190R and p.C1190Y) cause VWD as a result of VWF ER retention.
To cite this abstract in AMA style:Van Moort I, Bürgisser PE, Swinkels MW, Yildiz S, Atiq F, Leebeek FW, Bierings R, for the SYMPHONY consortium . Mutations at p.C1190 in VWF D3 Retain VWF in the ER in VWD 2A Patient-derived ECFCs [abstract]. Res Pract Thromb Haemost. 2021; 5 (Suppl 1). https://abstracts.isth.org/abstract/mutations-at-p-c1190-in-vwf-d3-retain-vwf-in-the-er-in-vwd-2a-patient-derived-ecfcs/. Accessed September 24, 2021.
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