The mechanisms of contractile dysfunction following chronic limited platelet activation in (pro)thrombotic conditions

Abstract

Background

Patients with (pro)thrombotic conditions often exhibit a combination of platelet activation and dysfunction, including impaired contractility, which may contribute to thrombosis progression.

Aims

To study the mechanisms of secondary platelet dysfunction developed during prolonged limited activation in vitro.

Methods

Platelet-rich plasma from healthy subjects was incubated with 10 μM thrombin receptor-activating peptide (PAR1-receptor agonist) for 6 h at 37 °C to simulate continuous incomplete platelet activation. The dynamic functional and morphological state of platelets was assessed at specific time points by the rate and degree of thrombin-induced clot contraction, the mitochondrial membrane potential, intracellular ATP content, expression of phosphatidylserine, P-selectin and active integrin αIIbβ3 (before and after hyper-stimulation), platelet aggregation, as well as by scanning electron microscopy.

Results

In long-term activated platelets, a significant decrease in contractility was observed, while in control untreated plasma samples the contractility remained relatively stable throughout the 6-h observation period. Continuing platelet activation was accompanied by a decrease in the mitochondrial membrane potential and intracellular ATP level. An increase in expression of phosphatidylserine and P-selectin was steady, while the expression of active integrin αIIbβ3 was initially elevated, but dropped eventually, which disrupts platelet aggregation and mechanotransmission from contracting platelets to fibrin. Morphologically, the activated platelets initially formed aggregates followed by disaggregation along with progressive platelet inactivation and fragmentation.

Conclusions

The reduced platelet contractility in (pro)thrombotic conditions results from continuing limited activation of circulating platelets, leading to multiple functional and structural impairments, altogether underlying ‘activation-induced platelet dysfunction’, a particular and common type of acquired thrombocytopathy.