Introduction: Coronary artery disease (CAD) is a leading cause of morbidity and mortality worldwide. In the last decade, large-scale genetic studies discovered variants in genes encoding the soluble guanylyl cyclase (sGC) to be associated with coronary artery disease (CAD) risk. Platelets are strongly affected by impaired sGC function, while their role in atherosclerosis remains incompletely understood. Here we sought to investigate the contribution of platelet sGC to atherosclerosis and the therapeutic potential of targeting sGC in atherosclerosis.

Methods and Results: We genetically deleted sGC in platelets of atherosclerosis-prone Ldlr^-/- mice. By intravital fluorescence microscopy, such Pf4-Cre⁺Gucy1b1^flox/floxLdlr^-/- mice displayed enhanced leukocyte adhesion to atherosclerotic plaques in comparison with their litter mates (20.9 ± 1.5 vs. 13.6 ± 1.4 Cd11b⁺ cells, p<0.01). Moreover, histological and flow cytometry analyses revealed more numerous inflammatory leukocytes (Ly6C^high monocytes: 1,829 ± 262 vs. 964 ± 129* cells/aorta; neutrophils: 974 ± 170 vs. 392 ± 60* cells/aorta; and macrophages: 14,895 ± 1,912 vs. 9,463 ± 1,211* cells/aorta; *p≤0.02) and larger plaque sizes (246,998 ± 22,162 vs. 189,843 ± 15,156 μm², p=0.04) in aortic tissue of Ldlr^-/- mice lacking sGC in platelets. In vitro, supernatant from activated, sGC-deficient platelets promoted leukocyte adhesion to endothelial cells (monocytes: 33,125 ± 1,313 vs. 27,039 ± 555 rfu, p<0.01; neutrophils: 33,810 ± 1,139 vs. 27,824 ± 758 rfu, p<0.001) via increasing activation of endothelial cells. Using cytokine profiling, we identified reduced angiopoietin-1 release by Pf4-Cre⁺Gucy1b1^flox/flox (30.4 ± 6.4 vs. 60.3 ± 4.7 ng/ml, p<0.01) and human sGC risk allele carrier platelets (4.5 ± 0.7 vs. 8.3 ± 1.4 ng/ml, p=0.04) to be responsible for enhanced activation of endothelial cells and subsequent leukocyte adhesion. Pharmacological sGC stimulation increased platelet angiopoietin-1 release in vitro (133.8 ± 6.5 vs. 64.0 ± 4.3 ng/ml, p<0.01) and reduced recruitment of adoptively transferred leukocytes 21.4 ± 3.3 vs. 42.8 ± 6.9 cells, p=0.01) in Ldlr^-/- mice fed a Western diet. Pharmacological sGC stimulation further reduced atherosclerotic plaque formation (62.5 ± 16.1 vs. 123.1 ± 18.6 μm², p=0.03) and vascular inflammation (macrophages: 13,915 ± 1,550 vs. 22,156 ± 2,737 cells/aorta, p=0.02; Ly6C^high monocytes: 1,485 ± 348 vs. 2,688 ± 531 cells/aorta, p=0.07; and neutrophils: 1,201 ± 192 vs. 1,885 ± 232 cells/aorta, p=0.04).

Conclusion: Loss of sGC in platelets contributes to atherosclerotic plaque formation via reduced release of the soluble factor angiopoietin-1 and, subsequently, enhanced leukocyte recruitment. Pharmacological sGC stimulation might represent a novel therapeutic strategy to prevent and treat CAD.