Gout is characterized by an acute inflammatory reaction and the accumulation of neutrophils in response to monosodium urate (MSU) crystals. Inflammation resolves spontaneously within a few days, although MSU crystals can still be detected in the synovial fluid and affected tissues. Here we report that neutrophils recruited to sites of inflammation undergo oxidative burst and form neutrophil extracellular traps (NETs). Under high neutrophil densities, these NETs aggregate and degrade cytokines and chemokines via serine proteases. Tophi, the pathognomonic structures of chronic gout, share characteristics with aggregated NETs, and MSU crystals can induce NETosis and aggregation of NETs. In individuals with impaired NETosis, MSU crystals induce uncontrolled production of inflammatory mediators from neutrophils and persistent inflammation. Furthermore, in models of neutrophilic inflammation, NETosis-deficient mice develop exacerbated and chronic disease that can be reduced by adoptive transfer of aggregated NETs. These findings suggest that aggregated NETs promote the resolution of neutrophilic inflammation by degrading cytokines and chemokines and disrupting neutrophil recruitment and activation.

Background Coronavirus induced disease 2019 (COVID-19) can be complicated by severe organ damage leading to dysfunction of the lungs and other organs. The processes that trigger organ damage in COVID-19 are incompletely understood. Methods Samples were donated from hospitalized patients. Sera, plasma, and autopsy-derived tissue sections were examined employing flow cytometry, enzyme-linked immunosorbent assays, and immunohistochemistry. Patient findings Here, we show that severe COVID-19 is characterized by a highly pronounced formation of neutrophil extracellular traps (NETs) inside the micro-vessels. Intravascular aggregation of NETs leads to rapid occlusion of the affected vessels, disturbed microcirculation, and organ damage. In severe COVID-19, neutrophil granulocytes are strongly activated and adopt a so-called low-density phenotype, prone to spontaneously form NETs. In accordance, markers indicating NET turnover are consistently increased in COVID-19 and linked to disease severity. Histopathology of the lungs and other organs from COVID-19 patients showed congestions of numerous micro-vessels by aggregated NETs associated with endothelial damage. Interpretation These data suggest that organ dysfunction in severe COVID-19 is associated with excessive NET formation and vascular damage. Funding Deutsche Forschungsgemeinschaft (DFG), EU, Volkswagen-Stiftung

Since the discovery and definition of neutrophil extracellular traps (NETs) 14 years ago, numerous characteristics and physiological functions of NETs have been uncovered. Nowadays, the field continues to expand and novel mechanisms that orchestrate formation of NETs, their previously unknown properties, and novel implications in disease continue to emerge. The abundance of available data has also led to some confusion in the NET research community due to contradictory results and divergent scientific concepts, such as pro-and anti-inflammatory roles in pathologic conditions, demarcation from other forms of cell death, or the origin of the DNA that forms the NET scaffold. Here, we present prevailing concepts and state of the science in NET-related research and elaborate on open questions and areas of dispute.

SARS-CoV-2 infection poses a major threat to the lungs and multiple other organs, occasionally causing death. Until effective vaccines are developed to curb the pandemic, it is paramount to define the mechanisms and develop protective therapies to prevent organ dysfunction in patients with COVID-19. Individuals that develop severe manifestations have signs of dysregulated innate and adaptive immune responses. Emerging evidence implicates neutrophils and the disbalance between neutrophil extracellular trap (NET) formation and degradation plays a central role in the pathophysiology of inflammation, coagulopathy, organ damage, and immunothrombosis that characterize severe cases of COVID-19. Here, we discuss the evidence supporting a role for NETs in COVID-19 manifestations and present putative mechanisms, by which NETs promote tissue injury and immunothrombosis. We present therapeutic strategies, which have been successful in the treatment of immunο-inflammatory disorders and which target dysregulated NET formation or degradation, as potential approaches that may benefit patients with severe COVID-19.

Neutrophil extracellular trap (NET) formation contributes to gout, autoimmune vasculitis, thrombosis, and atherosclerosis. The outside-in signaling pathway triggering NET formation is unknown. Here, we show that the receptor-interacting protein kinase (RIPK)-1-stabilizers necrostatin-1 or necrostatin-1s and the mixed lineage kinase domain-like (MLKL)-inhibitor necrosulfonamide prevent monosodium urate (MSU) crystal-or PMAinduced NET formation in human and mouse neutrophils. These compounds do not affect PMA-or urate crystal-induced production of ROS. Moreover, neutrophils of chronic granulomatous disease patients are shown to lack PMA-induced MLKL phosphorylation. Genetic deficiency of RIPK3 in mice prevents MSU crystal-induced NET formation in vitro and in vivo. Thus, neutrophil death and NET formation may involve the signaling pathway defining necroptosis downstream of ROS production. These data imply that RIPK1, RIPK3, and MLKL could represent molecular targets in gout or other crystallopathies. Additional supporting information may be found in the online version of this article at the publisher's web-site Keywords IntroductionNeutrophil extracellular trap (NET) formation was first described to enhance bacterial killing via the release of histones, ROS, and proteases [1,2], but also contributes to autoinflammatory Correspondence: Prof. Hans-Joachim Anders e-mail: hjanders@med.uni-muenchen.de and autoimmune disorders [3][4][5][6][7]. For example, monosodium urate (MSU) crystals trigger NET formation, which first drives massive inflammation and subsequently fosters the resolution of inflammation, explaining both gouty arthritis and tophusrelated immune energy [8]. MSU crystals, cytokines, and bacterial * These authors contributed equally to this work. Results and discussionPMA triggered death and NET formation of human neutrophils release around 50% of total DNA after 2 h (Supporting Information Fig. 1A). During these processes neutrophils release IL-1β (but not TNF-α) in time-dependent manner into the supernatant. However, pretreatment with IL-1R antagonist anakinra, TNF blocker etanercept or anti-TLR4 as well as lack of Fas (Fas lpr mice) had no effect on DNA release upon PMA or MSU stimulation as compared to their respective controls, which excludes their role in this process (Supporting Information Fig. 1B-F).In addition, neither zVAD-FMK nor ferrostatin-1 (Fer-1) had an effect (Fig. 1A), excluding caspase-mediated extrinsic apoptosis or pyroptosis, ferroptosis [12], respectively. In contrast, firstgeneration necrostatin (Nec)-1, Nec-1s, and the MLKL inhibitor necrosulfonamide (NSA) decreased overall cell death and NET formation as assessed by nuclear Sytox uptake, the release of DNA using PICO green dye, and chromatin release to induce NET structures. Nec-1 inhibits necroptosis via modulating RIPK1 and preventing RIPK3 and MLKL phosphorylation and necrosome formation [13][14][15][16]. Indeed, MSU-, LPS-, and PMA-induced cell death and NET formation of human neutrophils-induced RIPK3 expression and phospho...

ABSTRACTTermites and cockroaches are closely related, with molecular phylogenetic analyses even placing termites within the radiation of cockroaches. The intestinal tract of wood-feeding termites harbors a remarkably diverse microbial community that is essential for the digestion of lignocellulose. However, surprisingly little is known about the gut microbiota of their closest relatives, the omnivorous cockroaches. Here, we present a combined characterization of physiological parameters, metabolic activities, and bacterial microbiota in the gut ofShelfordella lateralis, a representative of the cockroach family Blattidae, the sister group of termites. We compared the bacterial communities within each gut compartment using terminal-restriction fragment length polymorphism (T-RFLP) analysis and made a 16S rRNA gene clone library of the microbiota in the colon—the dilated part of the hindgut with the highest density and diversity of bacteria. The colonic community was dominated by members of theBacteroidetes,Firmicutes(mainlyClostridia), and someDeltaproteobacteria. SpirochaetesandFibrobacteres, which are abundant members of termite gut communities, were conspicuously absent. Nevertheless, detailed phylogenetic analysis revealed that many of the clones from the cockroach colon clustered with sequences previously obtained from the termite gut, which indicated that the composition of the bacterial community reflects at least in part the phylogeny of the host.

Highlights d Gallstones contain extracellular DNA and neutrophil-derived granular enzymes d Neutrophils form extended aggregates of gallstone building blocks in vitro d Uptake of crystals by neutrophils causes lysosomal leakage and NET formation d NOX2 and PADI4 activities are required for the generation and growth of gallstones Authors

The critical size for strong interaction of hydrophobic particles with phospholipid bilayers has been predicted to be 10 nm. Because of the wide spreading of nonpolar nanoparticles (NPs) in the environment, we aimed to reveal the ability of living organisms to entrap NPs via formation of neutrophil extracellular traps (NETs). Upon interaction with various cell types and tissues, 10-to 40-nmsized NPs induce fast (<20 min) damage of plasma membranes and instability of the lysosomal compartment, leading to the immediate formation of NETs. In contrast, particles sized 100-1,000 nm behaved rather inertly. Resulting NET formation (NETosis) was accompanied by an inflammatory reaction intrinsically endowed with its own resolution, demonstrated in lungs and air pouches of mice. Persistence of small NPs in joints caused unremitting arthritis and bone remodeling. Small NPs coinjected with antigen exerted adjuvant-like activity. This report demonstrates a cellular mechanism that explains how small NPs activate the NETosis pathway and drive their entrapping and resolution of the initial inflammatory response.nanoparticles | size | neutrophils | NETosis | inflammation