Circulating S100A8/A9 drives faster platelet COX-1 recovery via MRP4, impairing the duration of aspirin response

Low-dose aspirin is a cornerstone of cardiovascular prevention in high-risk patients, inhibiting platelet cyclooxygenase (COX-1) and reducing thromboxane A 2 (TXA 2 ) production for 24 hours. However, the duration of this effect varies among individuals, and the role of inflammation and platelet microRNAs in this variability remains unclear. This study identifies a thromboinflammatory mechanism linked to shorter duration of TXB 2 suppression during aspirin therapy. This mechanism involves (1) elevated circulating S100A8/A9, indicating systemic inflammation; (2) reduced platelet microRNA-21-5p (miR-21-5p) and increased circulating miR-21-5p (c-miR-21-5p), suggesting altered microRNA regulation; (3) S100A8/A9-driven upregulation of platelet multidrug resistance protein 4 (MRP4); and (4) enhanced COX-1 expression, resulting in increased TX production. Patients with accelerated COX-1 recovery had increased circulating S100A8/A9 and platelet MRP4, directly related with serum TXB 2 and platelet COX-1 mRNA/protein levels. In vitro, recombinant S100A8/A9 (rS100A8/A9) upregulated MRP4 and COX-1 in platelets and DAMI, a human megakaryoblastic cell line, and MRP4 inhibition prevented this effect. Additionally, these patients exhibited lower platelet miR-21-5p levels and higher c-miR-21-5p, inversely and directly related, respectively, to S100A8/A9, MRP4, and serum TXB 2 slope. In DAMI cells, rS100A8/A9 reduced platelet miR-21-5p and increased c-miR-215p, effects prevented by MRP4 inhibition. During immune inflammation, reflected by high circulating S100A8/A9, once-daily aspirin may incompletely inhibit COX-1 due to miR-21-5p driving platelet activation, and c-miR-21-5p release. Circulating S100A8/A9 and miR-21-5p can serve as biomarkers for patients with shorter duration of aspirin effect over 24 hours and potential targets to optimize antiplatelet therapy for high-risk patients.