OC 13.1 - Scavenger Receptor Stabilin-2 is a Major Regulator of VWF Propeptide Clearance

Sunday, June 25, 2023

10:45 – 11:00 ET

Room: 710B

Session Available As: On-Demand

Presenting Author(s)

Orla Rawley, PhD (she/her/hers)

Post-Doctoral Fellow
Queen's University, Pathology and Molecular Medicine
Kingston, Ontario, Canada

Co-Author(s)

CB

Christine Brown

Research Assistant
Queen's University
Kingston, Ontario, Canada
EB

Erin Burnett

Veterinary Technician
Queen's University, Pathology and Molecular Medicine
Kingston, Ontario, Canada
CD

Courtney Dwyer

Veterinary Technician
Queen's University
Kingston, Ontario, Canada
KN

Kate Nesbitt

Veterinary Technician
Queen's University
Kingston, Ontario, Canada
MM

Miranda Mathieson

Veterinary Technician
Queen's University
Kingston, Ontario, Canada
LS

Laura Swystun

Post-doctoral fellow
Queen's Universty
Kingston, Ontario, Canada
EH

Edward Harris

Professor
University of Nebraska-Lincoln
Kingston, Ontario, Canada
David Lillicrap, MD

Professor
Department of Pathology and Molecular Medicine, Queen's University
Kingston, Ontario, Canada

Background: The VWF propeptide (VWFpp) to VWF antigen (VWF:Ag) ratio is widely used as a marker for increased VWF clearance and to determine VWD subtype. Despite this, the mechanisms regulating clearance of VWFpp itself remain unknown.

Aims: To investigate the role of stabilin-2 in mediating VWFpp clearance.

Methods: VWFpp was expressed in HEK293T cells and was subsequently infused (200U mVWFpp/kg) into VWF-/- or Stab2-/-/VWF-/- mice. Mice were then retro-orbitally sampled and VWFpp was quantified by ELISA. Solid-phase binding assays were carried out utilizing the secreted ectodomain of the short stabilin-2 isoform. Age and sex-matched wild-type C57Bl6 mice and STAB2-/- mice were retro-orbitally sampled and endogenous VWF and VWFpp were quantified by ELISA.

Results: Clearance studies involving VWF-/- and Stab2-/-/VWF-/- mice revealed that stabilin-2 deficiency significantly prolonged VWFpp survival (7.8 mins versus 18.2 mins, p < 0.0001). Subsequent immunohistochemical analysis of VWF-/- mouse liver sections harvested following VWFpp infusion, demonstrated co-localization of both VWFpp and stabilin-2 with the liver sinusoidal endothelial cell markers LYVE1 and FcΥRIIB. To further investigate this interaction, solid-phase binding assays were carried out. Binding of VWFpp to stabilin-2 was found to be dose-dependent and saturable. Furthermore, the interaction was inhibited by the presence of propeptide-deleted full-length VWF (31.5 ± 8.1%, p < 0.0001) and was enhanced by removal of the VWFpp N-linked glycans (180.0 ± 0.6%, p = 0.0079). Notably, although STAB2-/- mice were found to have normal VWF plasma levels (STAB2-/- versus wild-type C57BL/6 mice, 0.9 ± 0.1 versus 1.0 ± 0.9, p > 0.05), significantly elevated endogenous VWFpp levels were observed (3.1 ± 0.2 versus 1.0 ± 0.1, p < 0.0001). Consequently, STAB2-/- mice had elevated VWFpp/VWF:Ag ratios (2.7 ± 0.4 versus 0.7 ± 0.1, p < 0.0001) that result from altered VWFpp clearance rather than altered VWF clearance.

Conclusion(s): Stabilin-2 is a major regulator of VWFpp clearance.

Learning Objectives:

Upon completion, participant will be able to describe the factors regulating VWF propeptide clearance.
Upon completion, participant will be able to identify stabilin-2 as a key modifier of VWF propeptide clearance.
Upon completion, participant will be to recognise the complexities of the overlapping VWF and VWFpp clearance pathways.