MiR-21 role in aspirin-dependent PPARα and multidrug resistance protein 4 upregulation

Abstract

Background: A mechanism involved in high on-aspirin treatment residual platelet reactivity is platelet multidrug resistance protein 4 (MRP4) overexpression. Aspirin enhances platelet MRP4 expression with a PPARα-dependent mechanism and reduces miR-21 expression that, in turn, downregulates PPARα expression. Objective: The aim of our study was to verify the relationship between miR-21 and MRP4-PPARα levels induced by aspirin treatment. Methods: We evaluated the changes in MRP4-PPARα, mRNA, MRP4 protein, and miR-21 expression induced by aspirin in: (i) in vitro–treated megakaryoblastic cell line (DAMI), (ii) primary megakaryocytes cultures and derived platelets, (iii) healthy volunteers’ platelets treated with aspirin, and (iv) aspirinated patients (aspirin-treated patients) and in a control population (control). Results: We observed an aspirin-induced reverse relationship between the expression of miR-21 and PPARα-MRP4. In DAMI cells the miR-21 mimic transfection reduces PPARα and MRP4 expression, even if cells were treated with aspirin after transfection. MiR-21 inhibitor transfection induces PPARα and MRP4 expression that are not enhanced by aspirin treatment. In human megakaryocytes, aspirin treatment lead to a miR-21 downregulation and a MRP4 upregulation and this trend is confirmed in derived platelets. In aspirin-treated volunteers, an inverse relationship between miR-21 and MRP4 platelet expression was found after aspirin treatment. A similar negative relationship was found in aspirin-treated patients vs the control population. Conclusion: The results reported in this study provide information that aspirin induces the modulation of platelet miR-21 expression levels and this modulation can be responsible for MRP4 enhancement in circulating platelets.