Important Regulatory Roles of Erythrocytes in Platelet Adhesion to the von Willebrand Factor on the Wall under Blood Flow Conditions

Abstract

The roles of erythrocytes on platelet adhesion to von Willebrand factor (VWF) on the vessel wall through their membrane glycoprotein (GP)Ibα under blood flow condition is still to be elucidated. Blood specimens containing fluorescently labeled platelet and native, biochemically fixed, or artificial erythrocytes, at various hematocrits were perfused on a surface of VWF immobilized on the wall at a shear rate of 1,500 s-1. Rates of platelet adhesions were measured in each condition. Computer simulation of platelet adhesion to the VWF on the wall at the same shear rate was conducted by solving governing equations with a finite-difference method on K-computer. The rates of platelet adhesion were calculated at various hematocrits conditions in the computational domain of 100 µm (x-axis) x 400 µm (y-axis) x 100 µm (z-axis). Biological experiments demonstrated the positive correlation between the rates of platelet adhesion and hematocrit values in native, fixed, and artificial erythrocytes. (r=0.992, 0.934, and 0.825 respectively, p<0.05 for all). The computer simulation results supported the hematocrit dependent increase in platelet adhesion rates on VWF (94.3/sec at 10%, 185.2/sec at 20%, and 327.9/sec at 30%, respectively). These results suggest the important contributing role of erythrocytes on platelet adhesion to the VWF. The augmented z-axis fluctuation of flowing platelet caused by the physical presence of erythrocytes is speculated as the cause for hematocrit dependent increase in platelet adhesion.

Effects of caplacizumab, a specific inhibitor of the A1 domain of von Willebrand factor binding with platelet membrane glycoprotein (GP) Ibα, on the length of platelet pseudopods supporting platelet adhesions on immobilized von Willebrand factor under blood flow condition

“…It is important to note that the distributions of GPIbα on platelet surface was heterogeneous [18]. Thus, the region(s) of platelet with higher densities of GPIbα tend to bind more likely to bind with VWF under blood flow conditions [38].…”

“…(Fig. 2) The longer pseudopods formed on platelet adhering to VWF were also [38] (1). Then, the platelet is captured on the vessel wall through GPIbα binding with VWF (2).…”

“…All 4 participants did not take any medications known to influence platelet functions including aspirin and P2Y 12 inhibitors for at least a month before the blood drawing. A blood specimen of 4.5 mL was obtained and immediately transferred into plastic tubes containing 0.5 mL of HEPES Tyrode buffer containing argatroban at 100 μM according to the previous publications [22,23].…”

“…(gift from Dr. Nakada in Centcore Co., USA) The FITC-conjugated abciximab rather than commonly used mepacrine was used for fluorinated platelets to block platelet adhesion mediated by GPIIb/IIIa [24]. The details of experimental methods were published elsewhere [16,22].…”

“…Figure 1 Measurements of the Length of the Pseudopods Supporting Platelet AdhesionsPanel A illustrated the behavior of platelets adhering to VWF under blood flow conditions. Single platelet expressing glycoprotein (GP) Ibα is flowing with the direction of flow toward the vessel wall induced by positional fluctuations of erythrocytes[38] (1). Then, the platelet is captured on the vessel wall through GPIbα binding with VWF (2).…”