Bury Loredana, Piselli Elisa, Manni Giorgia, Momi Stefania, Ciarrocca Taranta Giulia, Guglielmini Giuseppe, Fanelli Carmine, De Fano Michelantonio, Rowley Jesse W, Weyrich Andrew S, Gresele Paolo
Division of Internal and Cardiovascular Medicine (L.B., E.P., G.M., S.M., G.C.T., G.G., P.G.), University of Perugia, Italy.
Division of Endocrino-Metabolic Sciences, Department of Medicine and Surgery (C.F., M.D.F.), University of Perugia, Italy.
Circ Res. 2025 Aug 29;137(6):882-898. doi: 10.1161/CIRCRESAHA.124.325357. Epub 2025 Aug 5.
Despite being anucleate, platelets contain mRNAs and synthesize new proteins. Platelets also contain microRNAs and Dicer (ribonuclease III enzyme Dicer-1), an enzyme required for microRNA maturation. The expression of Dicer and some microRNAs is reduced in platelets from patients with type 2 diabetes (T2DM). However, the role of Dicer in the regulation of platelet function and in T2DM-associated platelet hyperreactivity is unclear. We aimed to assess whether Dicer levels are regulated in platelets upon activation, if they modulate mRNA translation by triggering pre-microRNA maturation, and whether these mechanisms are deranged in T2DM platelets.
Dicer expression in ultrapurified platelets was assessed by Western blotting, flow cytometry, and liquid chromatography-tandem mass spectrometry. P2Y (purinergic receptor P2Y) expression was assessed by Western blotting and microRNA-223 and P2RY12 transcript by real-time polymerase chain reaction. In vivo experiments were performed in Dicer-deficient and wild-type mice with alloxane-induced diabetes.
Thrombin-activated platelets from healthy individuals rapidly neosynthesize Dicer, leading to increased maturation of microRNA-223 and concomitant consumption of pre-microRNA-223. An increase in microRNA-223 was associated with a reduction of P2RY12 mRNA, one of its main targets, and platelet P2Y expression and function. All these mechanisms were significantly deranged in platelets from patients with T2DM. Similar alterations were also observed in platelets from Dicer-deficient and diabetic mice. ADP-induced platelet pulmonary thromboembolism was decreased in mice previously infused with low-dose thrombin.
The Dicer-triggered decrease in P2Y after thrombin stimulation may represent a self-regulatory mechanism of platelet activation to prevent undesired thrombus formation. The derangement of this self-regulatory mechanism in T2DM may contribute to the platelet hyperreactivity and enhanced thrombotic complications of patients with T2DM. Our results show that the complex regulatory role of microRNA neoformation during platelet activation, when deranged, may contribute to cardiovascular disease.
尽管血小板无细胞核，但含有信使核糖核酸（mRNA）并能合成新的蛋白质。血小板还含有微小核糖核酸（microRNA）和Dicer（核糖核酸酶III型酶Dicer-1），这是一种微小核糖核酸成熟所需的酶。2型糖尿病（T2DM）患者血小板中Dicer和一些微小核糖核酸的表达降低。然而，Dicer在调节血小板功能以及T2DM相关的血小板高反应性中的作用尚不清楚。我们旨在评估血小板激活后Dicer水平是否受到调节，它是否通过触发前体微小核糖核酸成熟来调节mRNA翻译，以及这些机制在T2DM血小板中是否紊乱。
通过蛋白质免疫印迹法、流式细胞术和液相色谱-串联质谱法评估超纯化血小板中Dicer的表达。通过蛋白质免疫印迹法评估嘌呤能受体P2Y（purinergic receptor P2Y）的表达，通过实时聚合酶链反应评估微小核糖核酸-223和P2RY12转录本。在链脲佐菌素诱导糖尿病的Dicer缺陷型和野生型小鼠中进行体内实验。
健康个体经凝血酶激活的血小板迅速重新合成Dicer，导致微小核糖核酸-223成熟增加，同时前体微小核糖核酸-223消耗增加。微小核糖核酸-223增加与其主要靶标之一P2RY12 mRNA减少以及血小板P2Y表达和功能降低有关。所有这些机制在T2DM患者的血小板中均明显紊乱。在Dicer缺陷型和糖尿病小鼠的血小板中也观察到类似变化。预先注入低剂量凝血酶的小鼠中，二磷酸腺苷（ADP）诱导血小板肺血栓栓塞减少。
凝血酶刺激后Dicer触发的P2Y降低可能代表血小板激活的一种自我调节机制，以防止不必要的血栓形成。T2DM中这种自我调节机制紊乱可能导致T2DM患者血小板高反应性及血栓形成并发症增加。我们的结果表明，血小板激活过程中微小核糖核酸新生的复杂调节作用一旦紊乱，可能会导致心血管疾病。
