High Density Lipoprotein and Apolipoprotein A-I Bind Von Willebrand Factor and Prevent Its Self-Association Into Thick Fibers
Authors: Chung Dominic W. , Zheng Ying , Chen Junmei , Ling Minhua , Fu Xiaoyun , López José A. 
Published: October 2019

Von Willebrand factor (VWF) is a large plasma protein secreted constitutively or upon activation by endothelial cells. On activated endothelium, a portion of the VWF molecules remain attached to the surface, where they self-associate to produce long hyperadhesive strands and fibers capable of capturing platelets and other blood cells if not removed by the plasma metalloprotease ADAMTS13. Failure to clear these fibers underlies the pathophysiology of many microangiopathies, most prominently thrombotic thrombocytopenic purpura, in which accumulation of VWF-platelet aggregates in the microvasculature leads to ischemia and organ failure. VWF self-association also plays a major role in the adsorption of purified VWF to surfaces. We observed that adsorption of VWF to surfaces began with contact and direct binding of VWF to the surface via hydrophobic interactions. After saturating the binding sites on the surface, VWF continued to bind to and associate with the immobilized VWF molecules via hydrophilic VWF-VWF interactions, depositing multiple layers of VWF molecules on the surface until essentially all VWF molecules were depleted from the fluid phase. Using the method of Magnani and coworkers, we eluted VWF molecules that were bound to the surface via VWF-VWF interactions with the ionic detergent SDS, and then eluted VWF molecules that were bound to the surface via VWF-surface interactions with the zwitterion detergent CHAPS. Quantification of the eluted VWF showed that self-association of VWF onto the surface accounted for >80% of the adsorptive loss. Using the disappearance of purified VWF from the fluid phase as a measurement of self-association, we fractionated human plasma by heat and identified that apolipoprotein A-I (ApoAI), the major apolipoprotein component in high density lipoprotein (HDL) particles, which is stable to heat at 100° C, stabilized and prevented VWF surface adsorption by interfering with VWF self-association. Commercial preparations of ApoAI and HDL, prepared without exposure to heat, similarly prevented adsorption of purified VWF to surfaces. Half-maximal stabilization occurred at a molar ratio of eight ApoAI molecules to each VWF subunit. We assessed the role of HDL in VWF self-association that leads to the assembly of ultra-large VWF (ULVWF) strings on the surface of phorbol myristyl acetate-stimulated endothelial cells in flow chambers. We observed that HDL reduced the number and length of platelet-decorated ULVWF strings on the endothelial surface, consistent with the ability of HDL to interfere with VWF self-association and ULVWF assembly. We also studied the role of ApoAI in the recruitment of fluid-phase VWF to hyperadhesive ULVWF fibers in a synthetic microvessel system. We observed that ApoAI directly bound to hyperadhesive transluminal ULVWF fibers under flow, and this binding completely blocked the recruitment of fluid phase VWF molecules to the immobilized ULVWF fibers. These results showed that ApoAI or HDL interacted with hyperadhesive forms of VWF and modified the adhesive properties of the ULVWF strings and fibers. Consistent with its antithrombotic properties, the level of ApoAI in patients with hyperadhesive forms of VWF, such as thrombotic thrombocytopenic purpura and sepsis, was significantly reduced. These results suggest that regulation of VWF self-association may be another mechanism by which HDL protects against cardiovascular disease and extends its protective effects from large arteries to the microvasculature.

Disclosures:

No relevant conflicts of interest to declare.