The Role of Platelet-Derived Extracellular Vesicles in Immune-Mediated Thrombosis

Abstract

Platelet-derived extracellular vesicles (PEVs) play important roles in hemostasis and thrombosis. There are three major types of PEVs described based on their size and characteristics, but newer types may continue to emerge owing to the ongoing improvement in the methodologies and terms used to define various types of EVs. As the literature on EVs is growing, there are continuing attempts to standardize protocols for EV isolation and reach consensus in the field. This review provides information on mechanisms of PEV production, characteristics, cellular interaction, and their pathological role, especially in autoimmune and infectious diseases. We also highlight the mechanisms through which PEVs can activate parent cells in a feedback loop.

Keywords: immune mediated; platelet-derived extracellular vesicles; thrombosis.

Figure 1

Subtypes of Extracellular Vesicles. There are three main subtypes of extracellular vesicles (EVs) that have been characterized to date, apoptotic bodies, ectosomes (microparticles/microvesicles), and exosomes. Apoptotic bodies are large (greater than 1000 nm in diameter) and heterogenous in size and shape due to them originating from plasma membrane blebs. Ectosomes, are also heterogenous in size and shape and can range from 150–1000 nm in diameter. Exosomes are more homogenous in size and shape, being 30–150 nm in diameter and mostly round, as they are produced via a regulated process., Apoptotic bodies and ectosomes originate from the plasma membrane and can express specific markers ARF6, VCAMP3, and membrane glycoproteins. Exosomes can be identified by the markers ALIX, CD9, CD63, CD81, and TSG101. While these three EV types have been characterized, emerging technologies are revealing new EV populations that have yet to be characterized. These include EV populations that are smaller, are non-membranous, and/or have different contents and mechanisms of production.

Figure 2

Effect of pH on Exosome Biogenesis. When the endosome, or the α-granule in platelets, undergoes invagination of its membrane, a multivesicular body (MVB) is formed. An acidic pH in the MVB destines the created exosomes for degradation via the lysosome, while a more basic pH destines the exosomes for secretion. H⁺ ATPase controls the acidification of the MVB. Cholesterol in the MVB activates H⁺ ATPase creating an acidic environment. On the other hand, TNFα activates neutral sphingomyelinase 2 which in turn inactivates H⁺ ATPase leading to a more basic pH within the MVB.

Figure 3

EV Uptake by Cells. EVs can be taken up and cleared from circulation via three methods. (1) The EV membrane can fuse with the plasma membrane of the cell, releasing the EV contents into the cytoplasm. (2) Receptor-mediated endocytosis can occur when a receptor on the cell membrane, such as lactadherin, attaches to a molecule on the EV, such as negatively charged phosphatidylserine. (3) EVs can be endocytosed or phagocytosed. When EVs are coated with molecules like immunoglobulins, phagocytosis of the EVs is increased. When coated with IgG, EVs are endocytosed to a greater degree regardless of their size, however, when they are coated with IgM, phagocytosis is preferential for smaller EVs.

Figure 4

Diverse production of platelet-derived extracellular vesicles with different agonists. Platelets stimulated with different agonists produce extracellular vesicles (EVs) that are characteristically different. ADP and collagen stimulation produces spherical and smooth EVs and a higher number of EVS are produced with collagen. Thrombin stimulation produces an even larger number of EVs, but they are smaller, both spherical and elongated in shape, and have rough surfaces with thin offshoots. Calcium ionophore stimulation produces EVs that are larger and more heterogenous in size and have a rougher membrane surface. Included here is also LPS, a non-host derived stimuli, that leads to PEV production.

Figure 5

Mechanisms by which Extracellular Vesicles (EVs) can Contribute to Thrombosis. There are three known pathways through which EVs can influence thrombosis: (A). Phosphatidylserine on the surface of EVs provides a negatively charged surface on which the prothrombinase complex (comprised of factors Va, Xa, and II) can form, (B). Tissue factor (TF) on EVs interacts with factor VIIa to initiate the coagulation cascade, (C). Molecules such as clotting factors, MHC-1, complement proteins, CXCL4, CXCL7, CD40 ligand (CD40L), and immunoglobulins (IgG and IgM) on the vesicle membrane can interact with other cell types to produce a pro-inflammatory and pro-thrombotic environment.

Figure 6

Platelet Extracellular Vesicles (PEVs) Stimulate Megakaryocyte Maturation and Platelet Production. (A). Megakaryopoiesis is normally induced by thrombopoietin (TPO), where megakaryocytes become polyploid and platelets are produced from pseudopodia extending from the mature megakaryocyte. (B). PEVs can stimulate megakaryopoiesis independent of TPO. When PEVs stimulate megakaryocyte maturation and platelet production, the resulting megakaryocytes are larger and more polyploid and as a result, there is increased platelet production.