Besnoitia besnoiti tachyzoites induce monocyte extracellular trap formation

Abstract

Extracellular trap (ET) formation has been demonstrated as an important novel effector mechanism of polymorphonuclear neutrophils (PMN), eosinophils, mast cells and macrophages acting extracellularly against pathogens. In the present study, we show that tachyzoites of the emerging apicomplexan parasite Besnoitia besnoiti, that have recently been reported as potent inducers of PMN-derived ETosis, also trigger the release of ETs in an additional cell type, namely in monocytes. Fluorescence illustrations as well as scanning electron microscopy analyses (SEM) showed monocyte-promoted ET formation to be rapidly induced upon exposure to viable tachyzoites of B. besnoiti. Classical characteristics of ETs were confirmed by the co-localization of extracellular DNA with histones (H3) or myeloperoxidase (MPO) in parasite-entrapping structures. Monocyte-derived ETs were efficiently abolished by DNase I treatment and significantly reduced by treatments with inhibitors of MPO and NADPH oxidase, thus strengthening the key roles of reactive oxygen species (ROS) and MPO in monocyte ET formation. For comparative reasons, we additionally tested sporozoite stages of the closely related parasite Eimeria bovis for their capacity to induce monocyte-derived ETs and showed that these stages indeed induce ETs. To our best knowledge, we here report for the first time on monocyte ETs against the apicomplexan parasites B. besnoiti and E. bovis. Our results indicate that monocyte-triggered ETs may represent an important effector mechanism of the host early innate immune response against B. besnoiti and add a new cell type to the list of cells capable to release ETs.

Macrophage Extracellular Traps: A Scoping Review

“…Yousefi et al [9] described DNA release from eosinophils within 5 min of stimulation with complement component C5a or lipopolysaccharide (LPS), and the maximum effect, measured as fluorescence of a cell-impermeable DNA-staining dye, occurred within 30 min. METosis has been described as a rapid process that can occur in <30 min [22,23].…”

“…Pretreatment with the actin cytoskeletal inhibitor, cytochalasin D, often used to block phagocytosis, has demonstrated the ability to decrease MET formation in some studies [25,26]. Cytochalasin D treatment did not have a significant effect on METosis of bovine monocytes exposed to B. besnoiti or murine J774A.1 macrophage-like cells in response to E. coli or C. albicans [23,28]. Aulik et al [25] reported that cytochalasin D reduced the bovine peripheral-blood and alveolar macrophage METosis response to M. haemolytica and its leukotoxin, but, in this same study, cytochalasin D had no significant effect on MET release from THP-1 or RAW 264.7 macrophagelike cells in response to E. coli hemolysin.…”

“…Elastase, often considered a neutrophil-specific marker, has been identified in the METs of human peripheral-blood monocytes and in THP-1 macrophage-like cells [22,26]. Similarly, MPO has been identified in the METs of diverse macrophage populations including human glomerular macrophages, human peripheral-blood monocytes, THP-1 macrophage-like cells, murine J774A.1 macrophage-like cells, bovine monocytes, and caprine monocytes [22,23,[26][27][28][29] (Table 1).…”

“…Reports have demonstrated that inhibitors of the NADPH-oxidase system including diphenylene iodonium reduce MET formation, although this has not been consistent across all studies [23,25,28,29,40]. King et al [47] used a fluorescent readout of ROS production (dihydrorhodamine 123) to indicate that human alveolar macrophages producing METs had a 2-fold increase in ROS fluorescence compared to cells not forming METs.…”

“…In contrast, Je et al [26] noted AAPV-treated THP-1 macrophage-like cells infected with M. massili ense were unaffected with regard to MET release despite elastase being identified within the MET structures. Muñoz-Caro et al [23] inhibited MET release by bovine monocytes infected with Besnoitia besnotii trachyzoites using the MPO inhibitor, 4-aminobenzoic acid hydrazide (ABAH). These studies all suggest an active role for elastase and MPO in METosis, but it remains to be determined if the mechanisms by which these enzymes contribute actually mirror those seen in neutrophils.…”