Platelet Interactions with Von Willebrand Factor: Comparing Platelet Function in Acute and Stable Coronary Syndromes
Authors: Dunne Eimear , Ralph Adam , Foley David , Ricco Antonio J , Kenny Dermot 
Published: October 2019

Abstract
In unstable coronary syndromes, von Willebrand Factor (VWF) adheres to exposed subendothelial matrix. Initial platelet interaction with immobilised VWF is rapidly reversible, shear dependent, and includes characteristic start-stop translocation of platelets. Platelets tether to bound VWF via the glycoprotein (GP)Ib receptor and initiate a complex signalling cascade, ultimately activating the integrin αIIbβ3 receptor that crosslinks fibrinogen and causes platelet arrest. We have developed a microfluidic assay utilizing video microscopy to accurately measure dynamic platelet behaviour in microliters of blood perfused across VWF at arterial shear rates (1500 s-1). Tracking multiple individual platelets from frame to frame with unique motion-analysis software, the assay measures the total number of platelets 1) interacting with VWF, 2) translocating across VWF (GPIb dependent), and 3) stably adhered to the surface (αIIbβ3 dependent), as well as 4) the development rate of thrombi, and 5) the percent surface coverage at 17 sec (the end of the assay).
Variations in platelet function associated with acute coronary syndromes (ACS) and stable coronary artery disease (CAD) are poorly understood. We hypothesise that platelet function differs between normal donors, patients with stable CAD taking aspirin (ASA), and patients with ACS on dual anti-platelet therapy (DAPT). We characterised dynamic platelet function in 66 healthy donors, 67 patients with stable CAD on long-term ASA alone and 85 patients recruited within 3 days of a cardiac event on DAPT (ACS group). All patients had an appropriate response to either ASA alone or DAPT as defined by consensus guidelines using light transmission aggregometry (LTA) in response to 500 mg/mL arachidonic acid and 20 mM ADP. Compared to healthy controls and despite an adequate LTA response to DAPT, patients with ACS had significantly more platelets interacting with VWF (534±281 vs 424±198 (mean±SD), p ² 0.01), stably adhered platelets (237±118 vs 189±69, p ² 0.01), translocating platelets (365±186 vs 303±141, p ² 0.04) and final surface coverage (12.8±3.7% vs 11.2±2.7%, p ² 0.002). The differences between CAD patients on ASA alone and those with ACS were even more striking: long-term-ASA CAD patients had fewer stably adhered (181±83 vs 237±118, p ² 0.002) and translocating platelets (271±141 vs 365±186, p ² 0.0006). Both the rate of thrombus growth (9.8±2.7 vs 8.6±2.7, p ² 0.005) and the percent surface coverage (12.8±3.7% vs 10.8±3.4%, p ² 0.0007) were significantly greater in ACS patients.
Conclusion: Despite nominal LTA-measured response to DAPT, the profile of dynamic platelet behaviour measured by novel platelet function parameters is remarkably different in patients with ACS from those with stable CAD (Figure 1). Platelets from patients with ACS interact more with VWF compared to healthy controls and patients with stable coronary artery disease. Our dynamic platelet function assay describes for the first time novel platelet interactions with VWF suggesting a new way to guide antiplatelet therapy.
Figure 1 The profile of platelet interactions with VWF is markedly different in patients with ACS. Dynamic platelet interaction with VWF in patients with ACS and stable CAD was normalised against healthy controls (bars represent normalised 95% confidence intervals). Platelets from patients with ACS have a higher rate of interaction with the surface compared to healthy controls and stable patients on aspirin alone. Figure 1. The profile of platelet interactions with VWF is markedly different in patients with ACS. Dynamic platelet interaction with VWF in patients with ACS and stable CAD was normalised against healthy controls (bars represent normalised 95% confidence intervals). Platelets from patients with ACS have a higher rate of interaction with the surface compared to healthy controls and stable patients on aspirin alone.

Disclosures
Dunne: Science Foundation Ireland: Research Funding. Ralph:Science Foundation Ireland: Research Funding. Ricco:Science Foundation Ireland: Research Funding. Kenny:Science Foundation Ireland: Research Funding; Enterprise Ireland: Research Funding.