It is now about 20 years since the first case of a gain-of-function mutation involving the as-yet-unknown actor in cholesterol homeostasis, proprotein convertase subtilisin/kexin type 9 (PCSK9), was described. It was soon clear that this protein would have been of huge scientific and clinical value as a therapeutic strategy for dyslipidemia and atherosclerosis-associated cardiovascular disease (CVD) management. Indeed, PCSK9 is a serine protease belonging to the proprotein convertase family, mainly produced by the liver, and essential for metabolism of LDL particles by inhibiting LDL receptor (LDLR) recirculation to the cell surface with the consequent upregulation of LDLR-dependent LDL-C levels. Beyond its effects on LDL metabolism, several studies revealed the existence of additional roles of PCSK9 in different stages of atherosclerosis, also for its ability to target other members of the LDLR family. PCSK9 from plasma and vascular cells can contribute to the development of atherosclerotic plaque and thrombosis by promoting platelet activation, leukocyte recruitment and clot formation, also through mechanisms not related to systemic lipid changes. These results further supported the value for the potential cardiovascular benefits of therapies based on PCSK9 inhibition. Actually, the passive immunization with anti-PCSK9 antibodies, evolocumab and alirocumab, is shown to be effective in dramatically reducing the LDL-C levels and attenuating CVD. While monoclonal antibodies sequester circulating PCSK9, inclisiran, a small interfering RNA, is a new drug that inhibits PCSK9 synthesis with the important advantage, compared with PCSK9 mAbs, to preserve its pharmacodynamic effects when administrated every 6 months. Here, we will focus on the major understandings related to PCSK9, from its discovery to its role in lipoprotein metabolism, involvement in atherothrombosis and a brief excursus on approved current therapies used to inhibit its action.

Platelets are key players in the thrombotic processes. The alterations of platelet function due to the occurrence of metabolic disorders contribute to an increased trend to thrombus formation and arterial occlusion, thus playing a major role in the increased risk of atherothrombotic events in patients with cardiometabolic risk factors. Several lines of evidence strongly correlate metabolic disorders such as obesity, a classical condition of insulin resistance, dyslipidemia, and impaired glucose homeostasis with cardiovascular diseases. The presence of these clinical features together with hypertension and disturbed microhemorrheology are responsible for the prothrombotic tendency due, at least partially, to platelet hyperaggregability and hyperactivation. A number of clinical platelet markers are elevated in obese and type 2 diabetes (T2DM) patients, including the mean platelet volume, circulating levels of platelet microparticles, oxidation products, platelet-derived soluble P-selectin and CD40L, thus contributing to an intersection between obesity, inflammation, and thrombosis. In subjects with insulin resistance and T2DM some defects depend on a reduced sensitivity to mediators—such as nitric oxide and prostacyclin—playing a physiological role in the control of platelet aggregability. Furthermore, other alterations occur only in relation to hyperglycemia. In this review, the main cardiometabolic risk factors, all components of metabolic syndrome involved in the prothrombotic tendency, will be taken into account considering some of the mechanisms involved in the alterations of platelet function resulting in platelet hyperactivation.

Summary: Glucagon-like peptide 1 (GLP-1) is object of intensive investigation for not only its metabolic effects but also the protective vascular actions. Since platelets exert a primary role in the pathogenesis of atherosclerosis, inflammation and vascular complications, we investigated whether GLP-1 directly influences platelet reactivity. For this purpose, in platelets from 72 healthy volunteers we evaluated GLP-1 receptor (GLP-1R) expression and the effects of a 15-minute incubation with the native form GLP-1(7–36), the N-terminally truncated form GLP-1(9–36) and the GLP-1 analogue Liraglutide (100 nmol/l) on: i) aggregation induced by collagen or arachidonic acid (AA); ii) platelet function under shear stress; iii) cGMP and cAMP synthesis and cGMP-dependent protein kinase (PKG)-induced Vasodilator-Stimulated-Phosphoprotein (VASP) phosphorylation; iv) activation of the signalling molecules Phosphatidylinositol 3-Kinase (PI3-K)/Akt and Mitogen Activated Protein Kinase (MAPK)/ERK-1/2; and v) oxidative stress. Experiments were repeated in the presence of the nitric oxide donor Na–nitroprusside. We found that platelets constitutively express GLP-1R and that, independently of GLP-1R, GLP-1(7–36), GLP-1(9–36) and Liraglutide exert platelet inhibitory effects as shown by: a) increased NO-antiaggregating effects, b) increased the activation of the cGMP/PKG/VASP pathway, c) reduced the activation of PI3-K/Akt and MAPK/ERK-2 pathways, d) reduced the AA-induced oxidative stress. When the experiments were repeated in the presence of the antagonist of GLP-1R Exendin(9–39), the platelet inhibitory effects were maintained, thus indicating a mechanism independent of GLP-1R. In conclusion, GLP-1(7–36), its degradation product GLP-1(9–36) and Liraglutide exert similar inhibitory effects on platelet activation, suggesting a potential protective effect on the cardiovascular system.

Background. Beside the lipid-lowering effect, statins slow the progression of atherosclerosis by exerting anti-inflammatory and platelet inhibiting effects. We investigated whether platelet inhibition by simvastatin correlates with the statin effects on lipid lowering, inflammation, oxidative stress, and endothelial and platelet activation. Methods. In hypercholesterolemic patients allocated to diet (n=20) or a 2-month treatment with diet plus 40 mg simvastatin (n=25), we evaluated platelet aggregating responses to ADP, collagen, and arachidonic acid (AA), the effect of aspirin on AA-induced aggregation, pro- and anti-inflammatory and atherogenic mediators (IL-1β, -5, -6, -7, -8, -9, -10, -12, and -13, IFN-γ, IP-10, Eotaxin, and sRAGE), markers of endothelium (sE-selectin, VEGF, and MCP-1) and platelet activation (sP-selectin, sCD-40L, RANTES, and PDGF-bb), and oxidative stress (8-OH-2’-deoxyguanosine). Results. After treatment, beside the improvement of lipid profile, we observed the following: a reduction of platelet aggregation to ADP (p=0.0001), collagen (p=0.0001), AA (p=0.003); an increased antiaggregating effect of aspirin in the presence of AA (p=0.0001); a reduction of circulating levels of IL-6 (p=0.0034), IL-13 (p<0.0001), IFN-γ (p<0.0001), VEGF (p<0.0001), sE-selectin (p<0.0001), sCD-40L (p<0.0001), sP-selectin (p=0.003), and 8-OH-2’-deoxyguanosine (p<0.0001); an increase of IL-10 and sRAGEs (p=0.0001 for both). LDL-cholesterol levels (i) positively correlated with IL-6, IFN-γ, E-selectin, sCD-40L, 8-OH-2’-deoxyguanosine, platelet aggregation to ADP, collagen, AA, and aspirin IC-50 and (ii) negatively correlated with IL-10 and sRAGE. In multiple regression analyses, LDL-cholesterol was the strongest predictor for most parameters of platelet reactivity. Conclusion. In primary hypercholesterolemia, simvastatin treatment reduced platelet activation and subclinical inflammation and improved endothelial dysfunction. LDL-cholesterol levels were the major correlate of platelet reactivity; however, other effects of statins may contribute to reducing the progression of atherosclerosis.

Nitric oxide (NO) and carbon monoxide (CO) represent a pair of biologically active gases with an increasingly well-defined range of effects on circulating platelets. These gases interact with platelets and cells in the vessels and heart and exert fundamentally similar biological effects, albeit through different mechanisms and with some peculiarity. Within the cardiovascular system, for example, the gases are predominantly vasodilators and exert antiaggregatory effects, and are protective against damage in myocardial ischemia-reperfusion injury. Indeed, NO is an important vasodilator acting on vascular smooth muscle and is able to inhibit platelet activation. NO reacts with superoxide anion (O₂(⁻•)) to form peroxynitrite (ONOO(⁻)), a nitrosating agent capable of inducing oxidative/nitrative signaling and stress both at cardiovascular, platelet, and plasma levels. CO reduces platelet reactivity, therefore it is an anticoagulant, but it also has some cardioprotective and procoagulant properties. This review article summarizes current knowledge on the platelets and roles of gas mediators (NO, and CO) in cardioprotection. In particular, we aim to examine the link and interactions between platelets, NO, and CO and cardioprotective pathways.