Coenzyme Q10 (CoQ10) exists in a wide variety of foods and has promising cardiovascular benefits. However, its effects on platelets and integrin αIIbβ3 signaling during atherosclerosis have not been previously explored. Here, apolipoprotein E-deficient (ApoE−/−) mice were fed a standard diet, high-fat diet (HFD) or CoQ10-supplemented HFD for 12 weeks. We found that CoQ10 supplementation in ApoE−/− mice significantly alleviated formation of HFD-induced atherosclerotic lesions, and attenuated platelet hyper-aggregation and granule secretion, including CD62P, CD63 and CD40 ligand (CD40L) expression and platelet factor-4, β-thromboglobulin and activation normal T cell expressed and secreted (CCL5) release. CoQ10 supplementation decreased soluble fibrinogen and JON/A binding to αIIbβ3 on activated platelets, indicating that αIIbβ3-mediated inside-out signaling was attenuated. Additionally, CoQ10 down-regulated platelet αIIbβ3 outside-in signaling including decreasing phosphorylation of the β3 intracellular tail, cellular and sarcoma tyrosine-protein kinase (c-Src), and myosin light chain (MLC), and consistently attenuating platelet spreading and clot retraction. Importantly, platelet–monocyte aggregation that was primarily mediated by αIIbβ3 and can be blocked using an αIIbβ3-specific antagonist tirofiban was also markedly diminished by CoQ10. Thus, CoQ10 supplementation attenuates platelet hyper-reactivity via down-regulating both αIIbβ3 inside-out and outside-in signaling, which may play important preventive roles in atherothrombosis.