VPB0898 - Utilizing a Quartz Crystal Microbalance with dissipation (QCM-D) to Study Platelet NO-Signalling Under Flow Conditions

Background: Platelet function testing has been a subject of research due to its potential role in predicting platelet responses and acute coronary syndromes risk. Human platelets produce nitric oxide (NO), an important factor that negatively regulates platelet adhesion and aggregation, and previous studies have shown that platelets of patients with acute coronary syndromes have reduced NO production. While previous studies have used Quartz Crystal Microbalance with dissipation (QCM-D) under flowing conditions to assess platelet aggregation and stiffness of the resulting aggregate, QCM-D has not been used to study platelet NO-signalling.

Aims: To assess the effects of the nitric oxide synthase substrate L-arg and inhibitor L-NAME on the platelet aggregation profile to collagen-coated sensors using QCM-D

Methods: Platelets were isolated from healthy donors and resuspended to a concentration of 2.0 x 108/ml. QCM-D sensors were coated with collagen (10ug/ml) for 1 hour. Platelets were pre-incubated for 15 minutes with either vehicle, L-Arg (100uM), or L-NAME (1mM) at 37ºC. The platelets were then flowed through the QCM at 1.410 ml/minute and the negative change in frequency (ΔF) and dissipation (D) was measured over 5 minutes.

Results: L-arg significantly reduced the -ΔF to 70.3 ± 8.4% of control (Fig.1), demonstrating a reduction in mass on the sensor consistent with enhanced NO production and inhibition of platelet adhesion/aggregation. Conversely, L-NAME caused a -ΔF of 115.5 ± 12.3% of control. Similarly, L-arg caused a significant reduction in D, consistent with a reduced stiffness of the platelet layer, an effect not observed with L-NAME (Fig.2).

Conclusion(s): L-arg reduces platelet aggregation on collagen-coated sensors as measured by QCM under flow conditions, consistent with an enhanced NO inhibitory effect. Further, the QCM assay may be a novel and highly sensitive technique to assess platelet NO-signalling and the function of platelets of patients at increased risk of acute coronary syndromes.