PB1179 - Differential effects of physiological agonists on the proteome of platelet-derived extracellular vesicles

Background: Arterial thrombosis manifesting as heart attack and stroke is the leading cause of death worldwide. Platelets are central mediators of thrombosis that can be activated through multiple activation pathways. Platelet extracellular vesicles (pEVs), also known as platelet-derived microparticles, are granular mixtures of membrane structures produced by platelets in response to various activating stimuli. However, currently little is known about how physiological platelet agonists influence the pEV proteome.

Aims: We aimed to investigate how different physiological platelet agonists can impact the pEV proteome compared to their parent resting and activated platelets.

Methods: Human washed platelets were stimulated with physiological platelet agonists which reflect the microenvironments that platelets experience during thrombus formation: Adenosine diphosphate, collagen, thrombin as well as a combination of thrombin/collagen to induce platelet activation and pEV release. NanoLC-MS-based proteomic analysis was performed to ascertain the proteome landscape of platelets and pEVs and the proteomic changes between pEVs produced from different agonists and their parent platelets was examined.

Results: Proteomic profiling revealed that pEVs had an agonist dependent altered proteome in comparison to their parental activated platelets. Furthermore, we found that various protein classes including those related to coagulation and complement (prothrombin, antithrombin and plasminogen) and platelet activation (fibrinogen) are attributed to composition of platelet EVs following agonist stimulation. This occurs in the absence of de novo protein synthesis, indicating that protein packaging into pEVs is a specific and active process.

Conclusion(s): This study provides new information on how the biogenesis and proteome landscape of pEVs is impacted and conserved by physiological platelet agonist stimuli and provides insight into how pEV biogenesis might be harnessed to produce therapeutic pEVs with distinct biological functions.