Extracellular vesicles in atherothrombosis: From biomarkers and precision medicine to therapeutic targets

Abstract

Atherosclerotic cardiovascular disease (ASCVD) remains the leading cause of global mortality. Extracellular vesicles (EVs) are small phospholipid vesicles that convey molecular bioactive cargoes and play essential roles in intercellular communication and, hence, a multifaceted role in health and disease. The present review offers a glimpse into the current state and up-to-date concepts on EV field. It also covers their association with several cardiovascular risk factors and ischemic conditions, being subclinical atherosclerosis of utmost relevance for prevention. Interestingly, we show that EVs hold promise as prognostic and diagnostic as well as predictive markers of ASCVD in the precision medicine era. We then report on the role of EVs in atherothrombosis, disentangling the mechanisms involved in the initiation, progression, and complication of atherosclerosis and showing their direct effect in the context of arterial thrombosis. Finally, their potential use for therapeutic intervention is highlighted.

Keywords: atherosclerosis; extracellular vesicles; inflammation; ischemic disease; microvesicles; thrombosis.

FIGURE 1

Distinct subsets of circulating extracellular vesicle EVs (cEVs) as predictive, prognostic, and diagnostic biomarkers of the entire spectrum of atherosclerotic cardiovascular disease (ASCVD). Asymptomatic high cardiovascular risk (HCVR) patients with familial hypercholesterolemia (FH) patients about to suffer an ischemic event presented increased numbers of blood pan‐leukocyte‐ (CD45⁺) and activated neutrophil‐released EVs (CD15⁺) as well as levels of EVs (TSP1⁺/CD62P⁺) bearing markers of platelet activation. In asymptomatic patients with FH, lymphocyte‐derived (CD3⁺/CD45⁺) EVs signal for lipid‐rich atherosclerotic plaques, despite receiving lipid‐lowering therapy as per guidelines, likely reflecting their lifetime chronic exposure to high LDL plasma levels. HCVR patients showed a significant high number of blood platelet‐released (TSP1⁺/CD142⁺) EVs carrying markers of platelet activation and tissue factor. Thus, EVs from platelets and inflammatory cells provide valuable predictive information to map atherosclerotic plaque burden and stratify patient risk. Glycophorin A (CD235a⁺)‐rich erythrocyte‐derived EVs are released from evolving growing thrombi into the distal perfusing blood and might be useful as a systemic biomarker of ongoing thrombosis. A prothrombotic, pro‐inflammatory and endothelial dysfunction‐related (CD66b⁺/CD62E⁺/CD142⁺) cEV signature in the systemic circulation also reflects the formation of coronary thrombotic occlusions in patients with acute myocardial infarction (MI). Those ST‐elevation MI patients complicated with cardiogenic shock revealed that granulocyte‐ (CD66b⁺) and leukocyte−/neutrophil‐derived (CD45⁺/CD15⁺) EVs could serve as biomarkers of adverse prognosis in cardiogenic shock. CD indicates cluster of differentiation; EC, endothelial cell; TSP1, thrombospondin 1.

FIGURE 2

Effect of extracellular vesicles (EVs) on blood thrombogenicity. Schematic representation showing that increased numbers of circulating EVs and, specifically, platelet‐derived extracellular vesicles (pEVs) enhance platelet adhesion thrombus growth on thromboactive substrates (atherosclerotic and injured vessel wall). Thrombin activation of platelets causes proteostatic changes on the released pEVs. For instance, dysregulation of proteins involved in cell adhesion and prothrombotic signaling underlying thrombus formation. Representative confocal microscopy photomicrograph depicts perfused whole blood with pEVs in the flat perfusion chamber over collagen type‐I surface under flow (high shear rate) conditions. Platelets were labelled by calcein (green) and pEVs by BODIPY (red). CDCP1 indicates CUB domain‐containing protein 1; FERMT3, fermitin family homolog 3 or kindlin‐3; PAR‐1 or PAR‐4, protease‐activated receptor 1 or 4; PCDHα4, protocadherin‐α4.

FIGURE 3

Molecular mechanisms of extracellular vesicle (EV)‐mediated angiogenic activity and neovascularization. (a) Tissue factor (TF)‐rich extracellular vesicles released by ischaemic endothelial cells (TF⁺‐eEVs) reach the circulation and induce the polarization of circulating blood monocytes, their recruitment into the damaged tissue, and their differentiation to endothelial cell (EC)‐like cells in a β1‐integrin and TF‐dependent fashion. Similarly, (b) TF‐positive microvascular endothelium‐derived EVs (TF⁺‐meEVs) interact with cell surface β1‐integrin to activate a Rac1‐ERK1/2‐ETS1 signaling cascade that leads to CCL2 production and angiogenesis in vivo after ischemic hindlimb femoral arteriectomy. Thus, TF⁺‐eEVs exert a paracrine cellular crosstalk (either EC‐EC or monocyte‐EC) that is able to promote postischemic neovascularization and tissue repair. TF⁺‐eEVs can also transfer microRNA‐126 and induce monocyte reprogramming of angiogenic genes, thereby stimulating new vessel formation and attenuating inflammation. AKT indicates Ak strain transforming factor; CCL2, chemokine (C‐C motif) ligand 2; EC, endothelial cell; eEV, endothelial‐derived extracellular vesicle; eNOS, endothelial nitric oxide synthase; ERK1/2, extracellular signal‐regulated kinases 1 and 2; ETS1, external transcribed spacer region 1; M APK, mitogen‐activated protein kinase; mECs, microvascular endothelial cells; miR, microRNA; Mo, monocyte; Rac1, Ras‐related C3 botulinum toxin substrate 1; SPRED1, sprouty‐related EVH1 domain‐containing protein 1; PI3KR2, phosphoinositol‐3 kinase regulatory subunit 2; TF, tissue factor; VECM, vascular endothelial cell marker; VCAM1, vascular cell adhesion molecule 1; VE‐Cad, vascular endothelial cadherin; vWF, von Willebrand factor.