Neutrophils are the principal trouper of the innate immune system. Activated neutrophils undergo a noble cell death termed NETosis and release a mesh-like structure called neutrophil extracellular traps (NETs) as a part of their defensive strategy against microbial pathogen attack. This web-like architecture includes a DNA backbone embedded with antimicrobial proteins like myeloperoxidase (MPO), neutrophil elastase (NE), histones and deploys in the entrapment and clearance of encountered pathogens. Thus NETs play an inevitable beneficial role in the host’s protection. However, recent accumulated evidence shows that dysregulated and enhanced NET formation has various pathological aspects including the promotion of sepsis, pulmonary, cardiovascular, hepatic, nephrological, thrombotic, autoimmune, pregnancy, and cancer diseases, and the list is increasing gradually. In this review, we summarize the NET-mediated pathophysiology of different diseases and focus on some updated potential therapeutic approaches against NETs.

Background: The purpose of this study is to investigate the time course of syndecan-1 (Syn-1) plasma levels, the correlation between Syn-1 and organ damage development, and the associations of Syn-1 level with cumulative fluid balance and ventilator-free days (VFD) in patients with septic shock. Methods: We collected blood samples from 38 patients with septic shock upon their admission to ICU and for the first 7 days of their stay. Syn-1 plasma level, acute respiratory distress syndrome (ARDS), other organ damage, VFD, and cumulative fluid balance were assessed daily. Results: Over the course of 7 days, Syn-1 plasma levels increased significantly more in patients with ARDS than in those without ARDS. Patients with high levels of Syn-1 in the 72 h after ICU admission had significantly higher cumulative fluid balance, lower PaO₂/FiO₂, and fewer VFD than patients with low levels of Syn-1. Syn-1 levels did not correlate with sequential organ failure assessment score or with APACHE II score. Conclusions: In our cohort of patients with septic shock, higher circulating level of Syn-1 of cardinal glycocalyx component is associated with more ARDS, cumulative positive fluid balance, and fewer VFD. Measurement of Syn-1 levels in patients with septic shock might be useful for predicting patients at high risk of ARDS.

Components of neutrophil extracellular traps (NETs) are released into the circulation by neutrophils and contribute to microcirculatory disturbance in sepsis. Removing NET components (DNA, histones, and proteases) from the circulation could be a new strategy for counteracting NET-dependent tissue damage. We evaluated the effect of hemoperfusion with a polymyxin B (PMX) cartridge, which was originally developed for treating gram-negative infection, on circulating NET components in patients with septic shock, as well as the effect on phorbol myristate acetate (PMA)-stimulated neutrophils obtained from healthy volunteers. Ex vivo closed loop hemoperfusion was performed through PMX filters in a laboratory circuit. Whole blood from healthy volunteers (incubated with or without PMA) or from septic shock patients was perfused through the circuit. For in vivo experiment blood samples were collected before and immediately after hemoperfusion with PMX to measure the plasma levels of cell-free NETs. The level of cell-free NETs was assessed by measuring myeloperoxidase-associated DNA (MPO-DNA), neutrophil elastase-associated DNA (NE-DNA), and cell-free DNA (cf-DNA). Plasma levels of MPO-DNA, NE-DNA, and cf-DNA were significantly increased after 2 h of PMA stimulation. When the circuit was perfused with blood from septic shock patients or PMA-stimulated neutrophils from healthy volunteers, circulating levels of MPO-DNA, NE-DNA, and cf-DNA were significantly reduced after 1 and 2 h of perfusion with a PMX filter compared with perfusion without a PMX filter. In 10 patients with sepsis, direct hemoperfusion through filters with immobilized PMX significantly reduced plasma levels of MPO-DNA and NE-DNA. These ex vivo and in vivo findings demonstrated that hemoperfusion with PMX removes circulating NET components. Selective removal of circulating NET components from the blood could be effective for prevention/treatment of NET-related inappropriate inflammation and thrombogenesis in patients with sepsis.

Certain stimuli, such as microorganisms, cause neutrophils to release neutrophil extracellular traps (NETs), which are basically web-like structures composed of DNA with granule proteins, such as myeloperoxidase (MPO) and neutrophil elastase (NE), and cytoplasmic and cytoskeletal proteins. Although interest in NETs has increased recently, no sensitive, reliable assay method is available for measuring NETs in clinical settings. This article describes a modified sandwich enzyme-linked immunosorbent assay to quantitatively measure two components of circulating NETs, MPO-DNA and NE-DNA complexes, which are specific components of NETs and are released into the extracellular space as breakdown products of NETs. The assay uses specific monoclonal antibodies for MPO or NE as the capture antibodies and a DNA-specific detection antibody. MPO or NE binds to one site of the capture antibody during the initial incubation of samples containing MPO-DNA or NE-DNA complexes. This assay shows good linearity and high inter-assay and intra-assay precision. We used it in 16 patients with COVID-19 with accompanying acute respiratory distress syndrome and found that the plasma concentrations of MPO-DNA and NE-DNA were significantly higher than in the plasma obtained from healthy controls. This detection assay is a reliable, highly sensitive, and useful method for investigating the characteristics of NETs in human plasma and culture supernatants.

Neutrophils are the principal trouper of innate immune system. Activated neutrophils undergo a noble cell death termed NETosis and release a mesh-like structure called neutrophil extracellular traps (NETs) as a part of their defensive strategy against microbial pathogen attack. This web-like architecture includes a DNA backbone embedded with antimicrobial proteins like myeloperoxidase (MPO), neutrophil elastase (NE), histones etc. and deploys in the entrapment and clearance of encountered pathogens. Thus NETs play an inevitable beneficial role in the host's protection. However, recent accumulated evidence shows that dysregulated and enhanced NET formation has various pathological aspects including promotion of sepsis, pulmonary, cardiovascular, hepatic, nephrological, thrombotic, autoimmune, pregnancy, cancer diseases etc. and the list is increasing gradually. In this review, we summarize NETs mediated pathophysiology of different diseases, focus on some updated potential therapeutic approaches against NETs and share our future perspectives.

Arsenic, a major environmental toxicant and pollutant, is a global public health concern. Among its many adverse effects, arsenic is immunotoxic, but its effects on human neutrophil functions are not yet well-defined. In this study, we aimed to evaluate the in vitro effects of acute low-dose NaAsO₂ exposure on human polymorphonuclear neutrophils (PMNs) for 12 h on the following innate defense mechanisms: formation of neutrophil extracellular traps (NETs), production of reactive oxygen species (ROS), and phagocytosis. Phorbol myristate acetate (PMA) was added to induce NETs formation, which was quantified by measuring cell-free extracellular DNA (cf-DNA), myeloperoxidase-conjugated (MPO)-DNA and neutrophil elastase-conjugated (NE)-DNA, and confirmed by immunofluorescence labeling and imaging. Extracellular bactericidal activity by NETs was evaluated by co-culturing Escherichia coli and PMNs in the presence of a phagocytic inhibitor. Levels of NETs in the culture medium after PMA stimulation was significantly lower in PMNs pre-exposed to arsenic than those not exposed to arsenic. Immunofluorescence staining and extracellular bactericidal activity by NETs revealed similar results. Phagocytosis and ROS production by PMNs were also significantly reduced by arsenic pre-exposure. Together, our findings provide new insights in arsenic immunotoxicity and suggest how it increases susceptibility to infectious diseases in humans.