Activated polymorphonuclear neutrophils disseminate anti-inflammatory microparticles by ectocytosis

Abstract

On activation, human neutrophils release microparticles, called ectosomes, directly from the cell surface membrane. Microparticles from platelets, endothelial cells, and monocytes were reported to support coagulation or to modulate vascular homeostasis by activating monocytes as well as endothelial cells. We find that neutrophil ectosomes have no proinflammatory activity on human macrophages as assessed by the release of interleukin 8 (IL-8) and tumor necrosis factor ␣ (TNF␣). On the contrary, ectosomes increase the release of transforming growth factor ␤1 (TGF␤1), suggesting that ectosomes down-modulate cellular activation in macrophages. Polymorphonuclear neutrophil (PMN) ectosomes are able to block inflammatory response of macrophages to zymosan and lipopolysaccharide (LPS). We show that an early-phase TGF␤1 secretion and the exposure of phosphatidylserine on the surface of ectosomes independently contribute to this effect. IntroductionPolymorphonuclear neutrophils (PMNs) constitute the bulk of the leukocytes found in our blood. PMNs ingest and eventually kill invading pathogens by means of potent antimicrobial agents released during the process of degranulation. Because this microbicidal weaponry largely lacks specificity, it can lead to severe tissue damage if not controlled or secluded adequately from surrounding tissue. 1 At the time of degranulation, activated PMNs release small microvesicles directly from the cell surface membrane. Many eukaryotic cells, including tumor cells, release vesicles by ectocytosis (ectosomes), either spontaneously or in response to various stimuli. [2][3][4][5][6][7][8][9][10][11][12][13][14] Data on the function(s) of ectosomes have accumulated. 6,7,9,11,[15][16][17] So far, ectosomes have been associated with procoagulant and proinflammatory effects. Ectosomes derived from endothelial cells have been described to induce procoagulant activity in monocytic cells, 18 whereas ectosomes from platelets and monocytes were shown to directly promote hemostasis and induce inflammation by activating endothelial cells. 9,11 MacKenzie et al 6 described monocyte-derived ectosomes as being important proinflammatory agents by mediating the rapid secretion of interleukin 1␤ (IL-1␤).We recently described characteristics and molecular properties of ectosomes released by human PMNs. 19,20 Importantly for the work presented herein, PMN ectosomes expose phosphatidylserine on their outer membrane leaflet, and all ectosome preparations used in our assays are devoid of any cellular debris or apoptotic cells/bodies, as shown by electron microscopy. 20 Here, we show that PMN-derived ectosomes, unexpectedly, feature immunosuppressive/anti-inflammatory functions. Challenging a linear model of induction and resolution of inflammation, our data suggest a more complex balance of proinflammatory and anti-inflammatory events initiated in a parallel manner. Material and methods Antibodies and reagentsZymosan A from Saccharomyces cerevisiae, cytochalasin D, actinomycin D, formyl-methionine-leucine-phenylalan...

Cell-Cell Communication Via Extracellular Membrane Vesicles and Its Role in the Immune Response

“…Incubation of a macrophage cell line with PMN-derived ectosomes attenuates its activation by LPS and zymosan A and reduces the levels of inflammatory cytokines (i.e., IL-8, TNF-β) produced. PS present in the outer surface of ectosomes is found to be responsible for the immunomodulatory property of ectosomes (Gasser and Schifferli, 2004), and a specific receptor protein tyrosine kinase (i.e., Mer receptor tyrosine kinase) is involved in the intracellular signaling process (Scott et al, 2001). PMN-derived ectosomes also have an inhibitory effect on immature DCs (Eken et al, 2008;Gasser and Schifferli, 2004).…”

“…PS present in the outer surface of ectosomes is found to be responsible for the immunomodulatory property of ectosomes (Gasser and Schifferli, 2004), and a specific receptor protein tyrosine kinase (i.e., Mer receptor tyrosine kinase) is involved in the intracellular signaling process (Scott et al, 2001). PMN-derived ectosomes also have an inhibitory effect on immature DCs (Eken et al, 2008;Gasser and Schifferli, 2004). Thus, when immature DCs are cultured with LPS in the presence of PMN-derived ectosomes, expression levels of the cytokines and costimulatory and adhesion ligands required for T cell activation are significantly reduced, compromising T cell activation.…”

Changing world of neutrophils

“…Most of the data on PMN-derived EVs (called in these studies ectosomes) come from the laboratory of Dr. Schifferli [26,[34][35][36]39,40,45,[93][94][95][96]. They characterized the size and surface properties of EVs released upon fMLP-stimulation and came to the conclusion that both the in vitro and the in vivo generated EVs are right-side out [39,45].…”

“…They characterized the size and surface properties of EVs released upon fMLP-stimulation and came to the conclusion that both the in vitro and the in vivo generated EVs are right-side out [39,45]. Further, they demonstrated that PMN-derived EVs increased the secretion of the anti-inflammatory cytokine TGF-β1 from monocytes, whereas the stimulated release of IL-8, IL-10 and TNFα was decreased [40]. Secretion of TGF-β1 was induced also in monocyte-derived dendritic cells, where incubation with PMN-EVs interfered with the LPS-induced differentiation process indicated by a decrease of release of inflammatory cytokines, decrease of phagocytosis and T-cell stimulation [34].…”

P2X7 receptors regulate multiple types of membrane trafficking responses and non-classical secretion pathways

“…Small membrane vesicles released into the circulation from activated platelets are described in the literature as microparticles (MP); these have major roles in blood coagulation by providing a large surface area for the assembly of clotting factors that bind to the exposed phosphatidylserine of the released MPs [30,31]. Activated human neutrophils release "ectosomes" that exhibit anti-inflammatory activity and mediate inhibition of DC maturation from cultured blood monocytes [26,27,32]. Particles secreted from the basolateral membranes of various epithelial cells during tissue development are called "argosomes" due to their involvement in the transfer of developmental morphogens between cells and the maintenance of morphogen gradients [33].…”