Preclinical Models for Translational Investigations of Left Ventricular Assist Device‐Associated von Willebrand Factor Degradation

Abstract

Evidence suggests a major role for von Willebrand factor (vWF) in left ventricular assist device (LVAD)-associated bleeding. However, the mechanisms of vWF degradation during LVAD support are not well understood. We developed: (i) a simple and inexpensive vortexer model; and (ii) a translational LVAD mock circulatory loop to perform preclinical investigations of LVAD-associated vWF degradation. Whole blood was obtained from LVAD patients (n = 8) and normal humans (n = 15). Experimental groups included: (i) blood from continuous-flow LVAD patients (baseline vs. post-LVAD, n = 8); (ii) blood from normal humans (baseline vs. 4 h in vitro laboratory vortexer, ∼ 2400 rpm, shear stress ∼175 dyne/cm(2) , n = 8); and (iii) blood from normal humans (baseline vs. 12 h HeartMate II mock circulatory loop, 10 000 rpm, n = 7). vWF multimers and degradation fragments were characterized with electrophoresis and immunoblotting. Blood from LVAD patients, blood exposed to in vitro supraphysiologic shear stress, and blood circulated through an LVAD mock circulatory loop demonstrated a similar profile of decreased large vWF multimers and increased vWF degradation fragments. A laboratory vortexer and an LVAD mock circulatory loop reproduced the pathologic degradation of vWF that occurs during LVAD support. Both models are appropriate for preclinical studies of LVAD-associated vWF degradation.

Clinical and In Vitro Evidence That Subclinical Hemolysis Contributes to LVAD Thrombosis

“…To reproduce blood trauma from a continuous-flow LVAD, we previously developed a tightly controlled ex vivo LVAD model that generates continuous-flow LVAD-like supraphysiologic shear stress. The model produces the same profile of vWF degradation [5,12,13] and platelet activation [14] as patients with a continuousflow LVAD. Briefly, plasma was exposed to a continuous cyclone spin in a laboratory vortexer (Vortex Genie-2, Scientific Industries, Bohemia, NY) at a shear stress of approximately 175 dyne/cm 2 .…”

“…High-molecular-weight plasma vWF multimers and fragments were evaluated with standard agarose and polyacrylamide electrophoresis, respectively, as previously described in detail [5,[12][13][14][15]. Proteins were transferred to polyvinylidine difluoride membranes, blocked, and incubated with rabbit antihuman vWF primary antibody (1:500, Dako, Carpinteria, CA) and goat antirabbit immunoglobulin G horseradish peroxidase (HRP)-conjugated secondary antibody (1:3,000, Cell Signaling, Danvers, MA).…”

Artificial Organs 2015: A Year in Review

“…David J. Restle et al of the Hospital of the University of Pennsylvania, Philadelphia, PA, USA reported on the development of preclinical models for investigation of left ventricular assist device (LVAD) on von Willebrand factor (vWF) degradation and associated bleeding. They used a vortexer model and a translational LVAD mock circulatory loop to perform preclinical investigations.…”

Acquired Von Willebrand Syndrome and Blood Pump Design

“…While studying AVWS, Goda et al found that patients with LVADs had a high incidence of bleeding complications from the immediate postoperative phase throughout the entire observation and that the von Willebrand factor (vWF) ristocetin cofactor activity (Rco)/antigen ratio of patients with the HMII and CircuLite (HeartWare Inc., Framingham, MA, USA) devices was consistently lower compared with heart transplantation patients. Restle et al , using a HMII on a mock loop, concluded that blood from LVAD patients exposed to in vitro supraphysiologic shear stress, and blood circulated through an LVAD mock circulatory loop demonstrated a similar profile of decreased large vWF multimers and increased vWF degradation fragments. Morrison et al similarly reported that during CentriMag (Levitronix LLC, Waltham, MA, USA) support, AVWS was common in early post implantation, and associated with frequent bleeding events requiring a high volume of blood products.…”