Illicit drug N-ethylpentylone potentiates platelet activation through 5-hydroxytryptamine2A receptor-mediated mitogen-activated protein kinase signaling.

BACKGROUND: Several illicit drugs have been reported to be associated with ischemic stroke and myocardial infarction. However, their underlying mechanisms remain poorly investigated.

OBJECTIVES: In this study, we focused on N-ethylpentylone (NEP), a synthetic cathinone with potent hallucinogenic properties, and explored its role in platelet activation.

METHODS: Platelet function assays and flow cytometry were used to evaluate the effects of NEP on platelet aggregation, spreading, clot retraction, and integrin αIIbβ3 activation. The role of 5-hydroxytryptamine2A receptor (5-HT2AR) in NEP-mediated platelet responses was investigated using the selective 5-HT2AR antagonist M100907 in both ex vivo and in vivo experiments. Phosphoproteomics identified NEP-regulated phosphorylation sites, and Western blotting validated mitogen-activated protein kinase pathway activation by targeting key phosphorylation nodes.

RESULTS: NEP significantly potentiated platelet aggregation, spreading, clot retraction, and integrin αIIbβ3 activation in a concentration-dependent manner. The NEP-potentiated platelet responses were suppressed by M100907 in both ex vivo and in vivo models, confirming the critical regulatory role of 5-HT2AR. Phosphoproteomics revealed NEP-triggered upregulation of phosphorylation, which was enriched in mitogen-activated protein kinase pathways. Western blotting confirmed selective extracellular signal-regulated kinase and p38 phosphorylation, with no effect on C-Jun NH2-terminal kinase.

CONCLUSION: NEP directly enhances agonist-induced platelet activation by targeting 5-HT2AR on platelet membranes. The findings of this study provide valuable insights for elucidating the hematotoxic mechanisms of NEP and other illicit drugs.