Paper BI-MoM4
Surface Activation of the VWF A1 Domain: The Relationship between Platelet Activity and Absorbed A1 Structure

Monday, December 12, 2016, 9:00 am, Room Milo

When a material is placed in a biological environment, the surface of the material acts as the interface between that material and the biological environment. Upon contacting blood, plasma proteins attach to these surfaces and mediate platelet adhesion and activation and thrombosis. A key protein in this process is the clotting protein von Willebrand Factor (VWF) which binds to platelet receptor glycoprotein 1bα (GPIbα) when VWF is activated by chemicals, high shear stress, or immobilization onto surfaces. Activation of VWF by surface immobilization is an important problem in the failure of cardiovascular implants, but is poorly understood. Here we investigate whether some or all surfaces can activate VWF at least in part by affecting the orientation or conformation of the immobilized GPIbα-binding A1 domain of VWF. Platelets translocate rapidly on A1 adsorbed onto PS surfaces, and demonstrate shear-enhanced adhesion in that they detach at low rather than high shear stress. In contrast, platelets translocate more slowly on A1 adsorbed onto TCPS surfaces and are nearly stationary on A1 adsorbed onto glass surfaces, and demonstrate shear-inhibited adhesion in that they detach at high but not low shear stress. Both X-ray photoelectron spectroscopy and conformation independent antibodies reported comparable A1 amounts on all surfaces. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) suggested differences in orientation on the three surfaces, but none that could explain the biological data. Instead, ToF-SIMS determined Cys exposure and conformation-sensitive antibody binding suggest that A1 retains its native conformation when adsorbed onto PS surfaces, while TCPS surfaces and especially glass surfaces stabilized an alternative activated conformation of A1 that likely resembles the activated form of A1 that is also stabilized by disease-causing mutations. Regardless of the specific structure of the activated forms of A1, these studies demonstrate that it is not enough to determine the amount of various proteins that bind to different biomaterials placed in contact with the blood; instead, it is necessary to understand how different surfaces control the conformation of the many blood proteins that are capable of undergoing activating conformational changes. In particular, these studies demonstrate that the A1 domain of VWF has fundamentally different biological activity when adsorbed onto different surfaces. This is important when interpreting or designing in vitro experiments with surface-adsorbed A1 domain, and is also of likely relevance for blood-contacting biomaterials.