Proteomic and functional profiling of platelet-derived extracellular vesicles released under physiological or tumor-associated conditions

During hemostasis, thrombosis, and inflammation, activated blood platelets release extracellular vesicles (PEVs) that represent biological mediators of physiological and pathological processes. We have recently demonstrated that the activation of platelets by breast cancer cells is accompanied by a massive release of PEVs, evidence that matches with the observation that breast cancer patients display increased levels of circulating PEVs. A core concept in PEVs biology is that their nature, composition and biological function are strongly influenced by the conditions that induced their release. In this study we have performed a comparative characterization of PEVs released by platelets upon activation with thrombin, a potent thrombotic stimulus, and upon exposure to the breast cancer cell line MDA-MB-231. By nanoparticle tracking analysis and tandem mass spectrometry we have characterized the two populations of PEVs, showing that the thrombotic and tumoral stimuli produced vesicles that largely differ in protein composition. The bioinformatic analysis of the proteomic data led to the identification of signaling pathways that can be differently affected by the two PEVs population in target cells. Specifically, we have demonstrated that both thrombin- and cancer-cell-induced PEVs reduce the migration and potentiate Ca²⁺-induced apoptosis of Jurkat cells, but only thrombin-derived PEVs also potentiate cell necrosis. Our results demonstrate that stimulation of platelets by thrombotic or tumoral stimuli induces the release of PEVs with different protein composition that, in turn, may elicit selective biological responses in target cells.