PB0802 - The VWF Propeptide Is A Novel Component Of Venous Thrombi In A Mouse Model Of DVT

Background: The essential roles of the von Willebrand Factor propeptide (VWFpp) in mediating VWF multimerization and trafficking to Weibel-Palade are well documented. However, whether plasma VWFpp exerts haemostatic activity or contributes to thrombus formation has not been determined.

Aims: To investigate whether VWFpp contributes to venous thrombus formation in vivo.

Methods: The inferior vena cava (IVC) stenosis model of deep vein thrombosis (DVT) was used. Anesthetized male normal or VWF-/- mice on a C57Bl/6 background underwent a midline laparotomy to expose the IVC. The vessel was ligated around a blunted 30-gauge needle which was subsequently removed to create a 90% stenosis. Mice were recovered for 24 hours at which point the IVC was removed. Thrombi were dissected from the IVC wall and were formalin-fixed, paraffin-embedded. Longitudinal 5 µm sections were prepared and stained. Images were captured using an Aperio ScanScope CS slide scanner and image analysis was performed in ImageJ. Additional experiments involving VWFpp overexpression were carried out via hydrodynamic injection of murine VWFpp cDNA in normal mice one week prior to IVC stenosis.

Results: Thrombus development was observed in 70% of normal C57Bl/6 mice with a mean thrombus weight of 14.8±1.4mg versus an incidence of 30% in VWF-/- mice with a mean thrombus weight of 6.9±0.3mg. Immunofluorescent analysis carried out using a polyclonal anti-VWFpp antibody revealed distinct VWFpp staining in normal thrombi compared with VWF-/- thrombi. Fluorescent VWFpp staining partially co-localized with VWF staining. Dense VWFpp staining was observed at the thrombus tail, the region of active thrombus growth and resolution. Comparative analysis with H&E-stained sections also showed concentration of VWFpp in white thrombus regions. Subsequent overexpression of VWFpp had no effect on the incidence of thrombus formation, thrombus size or amount of VWFpp incorporated into the thrombus.

Conclusion(s): VWFpp constitutes a novel component of venous thrombus architecture in a mouse model of DVT.