Ginsenoside Rs3 (G-Rs3) inhibits agonist- and oxLDL-mediated platelet activation and attenuates in vivo arterial thrombosis.

Platelet hyperactivation and thrombo-inflammatory processes are critical drivers of cardiovascular diseases. Although conventional antiplatelet therapies reduce thrombotic risk, their use is limited by bleeding complications and drug resistance. Ginsenoside Rs3 (G-Rs3) exhibits anti-inflammatory potential; however, the specific mechanisms underlying the antiplatelet and thrombo-inflammatory activity of ginsenoside Rs3 (G-Rs3) remain unclear. Human platelets were treated with G-Rs3 in vitro to evaluate its effects on aggregation, intracellular signaling, granule secretion, and thrombo-inflammatory signaling markers. Network pharmacological analyses were performed to predict pathway-level modulations. The in vivo antithrombotic efficacy of G-Rs3 was assessed using a murine FeCl3-induced thrombosis model. G-Rs3 inhibited platelet aggregation in a dose-dependent manner following stimulation with collagen, thrombin, and U46619. It suppressed Ca2+ mobilization, dense granule secretion, and phosphorylation of key signaling molecules, including InsP3R, ERK1/2, MAPKs, PI3K, Akt, GSK3, and Syk. Additionally, G-Rs3 inhibited integrin αIIbβ3 activation, reduced platelet adhesion, and enhanced cyclic nucleotide-mediated negative regulation, as evidenced by elevated VASP phosphorylation and increased cAMP/cGMP levels. Notably, G-Rs3 attenuated oxLDL-enhanced upregulation of the pro-inflammatory platelet markers CD62P and CD162, highlighting its potential to suppress platelet-driven thrombo-inflammatory responses under atherogenic conditions. Further, network pharmacology analysis revealed that G-Rs3 targets multiple interconnected thrombo-inflammatory pathways, including MAPK, PI3K-Akt, Ras, calcium, and cyclic nucleotide signaling. In vivo, G-Rs3 significantly delayed thrombus formation and moderately prolonged occlusion time. These findings highlight G-Rs3 as a novel natural antithrombotic agent with anti-thrombo-inflammatory properties. Further studies are required to determine its translational potential and establish its therapeutic viability and safety profile.