Role of thrombin to non-physiological shear stress induced platelet activation and function alternation

Non-physiological shear stress (NPSS) and thrombin have two distinct mechanisms for activating platelets. NPSS in mechanically assisted circulation (MAC) devices can cause platelet dysfunction, e.g., by shedding its key receptors. In addition, patients with heart failure have increased levels of thrombin generation, which may further affect the NPSS-induced platelet dysfunction, resulting in device-associated complications. This study aimed to assess the combined effect of NPSS and thrombin in platelet activation, expression of adhesion receptors on the platelet surface, and alterations of platelet aggregation.

Fresh human blood from healthy donors was divided into two groups; one group was treated by adding 0.01 U/mL thrombin, and another group not treated with thrombin served as a control comparison. They were then pumped through a novel blood shearing device which produces similar shear stress conditions to those in the MAC devices. Three levels of NPSS (i.e., 75, 125, and 175 Pa) with a 1.0 s exposure time were selected for the shearing conditions. Expression of platelet activation markers (PAC-1, activated GPIIb/IIIa and CD62P, platelet surface P-selectin) were investigated along with the shedding of platelet receptors (GPIb, GPIIb/IIIa, and GPVI), generation of platelet microparticles, and Phosphatidylserine (PS)-positive platelets detected by flow cytometry. Platelet aggregation (induced by collagen/ristocetin) was measured by Lumi-aggregometry.

Platelet receptors were shed after exposure to NPSS showing a positive correlation with the level of shear stress. The generation of platelet microparticles and PS-positive platelets also increased with greater NPSS. Elevated NPSS decreased the platelet aggregation capacity. Platelet activation level increased with greater NPSS. Being treated by thrombin can further exacerbate these characteristics under same level of NPSS, except that platelet activation level drastically dropped after the exposure to 175 Pa NPSS in the thrombin-treated blood.

After being treated by thrombin, platelets became more susceptible to NPSS, resulting in more receptor shedding, platelet microparticles, and PS-positive platelets, thus limiting platelet aggregation capacity after exposure to NPSS. Platelet activation, in terms of PAC-1 and P-selectin, is an interim status competing between the expression and shedding of these makers/receptors. When platelets have reached a saturation level of activation, exposure to excessive NPSS can potentially impair activation.