Abstract 4345057: CCL2 Aggravates Thrombosis by Upregulating Platelet P2Y12 Receptor via CCR2: Mechanisms and Therapeutic Implications

Introduction: The P2Y12 receptor mediates platelet aggregation, but current therapies (e.g., clopidogrel) face interindividual variability and resistance. While CCL2 may regulate platelets via paracrine signaling, its direct effect on P2Y12 remains unclear. This study explores the CCL2/CCR2 axis in platelet activation to advance anti-thrombotic strategies. Hypothesis: CCL2-CCR2 axis upregulates P2Y12, aggravating platelet activation and ischemic injury. Methods: CCL2 levels were measured by ELISA. Platelet aggregation was assessed by Light transmission aggregometry (LTA), while P2Y12 expression and activation markers were analyzed via flow cytometry. Cardiac function was evaluated by echocardiography in a rat MI model induced by left anterior descending ligation. Results: 1. Elevated CCL2 predicts adverse outcomes in STEMI via P2Y12 upregulation: ELISA showed higher CCL2 in STEMI patients versus controls (Fig1A). Kaplan-Meier survival analysis demonstrated marked differences in 5-year major adverse cardiovascular event rates among three groups stratified by baseline plasma CCL2 levels (Fig1B). Flow cytometry showed a strong positive correlation between platelet surface P2Y12 receptor expression and plasma CCL2 concentration (Fig1C).The treatment of SD rats with CCL2 antibody alone or in combination with clopidogrel significantly improved LVEF and FS after myocardial infarction (Figs1D–E). 2. CCL2 dose-dependently upregulates platelet P2Y12 via CCR2: CCL2 dose-dependently increased platelet P2Y12 expression (Fig2A), exceeding ADP/thrombin effects (Fig2B), CCR2 inhibitor prevented this upregulation (Fig2C). While total P2Y12 remained unchanged(Fig2D).The expression of P2Y12 on the membrane was decreased by ciliobrevin(dynein inhibitor) and nocodazole (microtubule disruptor)(Fig2E), suggesting that CCL2 may affect the membrane transport of P2Y12. 3. CCL2 enhances platelet activation: LTA showed that CCL2 dose-dependently increased platelet aggregation (Figs3A–B) and ATP release (Fig3C). Flow cytometry demonstrated upregulated expression of P-selectin and Glycoprotein IIb/IIIa (Figs3D-E). Clot retraction assays revealed accelerated dynamics and enhanced retraction extent with CCL2 treatment (Figs3F–G). Conclusions: CCL2 aggravates myocardial injury through CCR2-mediated P2Y12 upregulation and platelet activation. Targeting the CCL2/CCR2 axis may improve antiplatelet efficacy, particularly in clopidogrel-resistant cases, suggesting potential for personalized therapeutic approaches.