Evaluation of neutrophil extracellular trap deregulated formation in pyoderma gangrenosum.
Ontology highlight
ABSTRACT:
SUBMITTER: Croia C
PROVIDER: S-EPMC8453546 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:BackgroundTriptolide (PG490), as a triterpene dicyclic oxide has been reported to increase the generation of reactive oxygen species (ROS) and nitric oxide (NO) and induce apoptosis of RAW 264.7 cells in a dose-dependent manner. The activity of death NETs plays an important role in anti-bacterial processes in the human body. This study aimed to investigate the effect of triptolide (PG490) on neutrophil extracellular traps (NETs) formation.MethodsAfter isolating peripheral blood neutrophils from healthy volunteers, cells were incubated with PG490 to observe and detect the level of NETs and detect the level of reactive oxygen species (ROS). The cells were cultured, stained and analyzed by fluorescence microscopy.ResultsCompared with the 12-myristate-13-acetate (PMA) group, the average fluorescence intensity of SYTOX Green in the PG490 + PMA group, as detected by a multifunctional microplate reader, was significantly decreased. Intracellular ROS were labeled by fluorescence, with fluorescence intensity then measured by multifunctional microplate reader and flow cytometry. The results showed that compared with the control group, the fluorescence intensity of the PMA group was significantly increased, while there was no significant difference between PMA group and PG490 + PMA group.ConclusionsThe production of NETs is inhibited by PG490 in vitro, which is not associated with the level of cellular ROS. This suggests that PG490in Tripterygium wilfordii Hook F can suppress related diseases.
| S-EPMC8506553 | biostudies-literature
Project description:Neutrophils undergo a unique form of cell death to generate neutrophil extracellular traps (NETs). It is well established that citrullination of histones (e.g., CitH3) facilitates chromatin decondensation during NET formation (NETosis), particularly during calcium-induced NETosis that is independent of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activation. However, the importance of other forms of histone modifications in NETosis has not been established. We considered that acetylation of histones would also facilitate NETosis. To test this hypothesis, we induced NOX-dependent NETosis in human neutrophils with phorbol myristate acetate or lipopolysaccharide (from Escherichia coli 0128), and NOX-independent NETosis with calcium ionophores A23187 or ionomycin (from Streptomycesconglobatus) in the presence or absence of two pan histone deacetylase inhibitors (HDACis), belinostat and panobinostat (within their half maximal inhibitory concentration (IC50) range). The presence of these inhibitors increased histone acetylation (e.g., AcH4) in neutrophils. Histone acetylation was sufficient to cause a significant increase (~20%) in NETosis in resting neutrophils above baseline values. When acetylation was promoted during NOX-dependent or -independent NETosis, the degree of NETosis additively increased (~15⁻30%). Reactive oxygen species (ROS) production is essential for baseline NETosis (mediated either by NOX or mitochondria); however, HDACis did not promote ROS production. The chromatin decondensation step requires promoter melting and transcriptional firing in both types of NETosis; consistent with this point, suppression of transcription prevented the NETosis induced by the acetylation of histones. Collectively, this study establishes that histone acetylation (e.g., AcH4) promotes NETosis at baseline, and when induced by both NOX-dependent or -independent pathway agonists, in human neutrophils. Therefore, we propose that acetylation of histone is a key component of NETosis.
| S-EPMC6359456 | biostudies-literature
Project description:Neutrophil extracellular traps (NETs) are formed after neutrophils expelled their chromatin content in order to primarily capture and eliminate pathogens. However, given their characteristics due in part to DNA and different granular proteins, NETs may induce a procoagulant response linking inflammation and thrombosis. Unraveling NET formation molecular mechanisms as well as the intracellular elements that regulate them is relevant not only for basic knowledge but also to design diagnostic and therapeutic tools that may prevent their deleterious effects observed in several inflammatory pathologies (e.g., cardiovascular and autoimmune diseases, cancer). Among the potential elements involved in NET formation, several studies have investigated the role of microRNAs (miRNAs) as important regulators of this process. miRNAs are small non-coding RNAs that have been involved in the control of almost all physiological processes in animals and plants and that are associated with the development of several pathologies. In this review, we give an overview of the actual knowledge on NETs and their implication in pathology with a special focus in cardiovascular diseases. We also give a brief overview on miRNA biology to later focus on the different miRNAs implicated in NET formation and the perspectives opened by the presented data.
| S-EPMC7924615 | biostudies-literature
Project description:Histones, the major protein components of chromatin, are released into the extracellular space during sepsis, trauma, and ischemia-reperfusion injury, and subsequently mediate organ failure. Extracellular histones can promote endothelial damage and platelet aggregation, which can be suppressed by administration of recombinant thrombomodulin (rTM). The present study aimed to clarify whether histones can activate neutrophils to induce NET formation and whether rTM can prevent histone-induced NET formation. NET formation was analyzed in vitro by stimulating human neutrophils with histones in the absence or presence of rTM. NET formation was further analyzed in vivo by intravenous infusion of histones into rats with or without rTM. Histones induced NET release in a dose-dependent manner in vitro and NET release was induced as early as 1 h after stimulation. Histone-induced NET release was independent of NADPH oxidase. rTM suppressed histone-induced NET release in vitro as well as in vivo. The suppression might be mediated by rTM binding to histones, as suggested by analysis using a quartz crystal microbalance system. The present findings suggest that histones can activate neutrophils to form NETs and that rTM can inhibit histone-induced NET formation.
| S-EPMC6828967 | biostudies-literature
Project description:Neutrophils are the first immune cells to kill invading microbes at sites of infection using a variety of processes, including the release of proteases, phagocytosis and the production of neutrophil extracellular traps (NETs). NET formation, or NETosis, is a specific and highly efficient process, which is induced by a variety of stimuli leading to expulsion of DNA, proteases and antimicrobial peptides to the extracellular space. However, uncontrolled NETosis may lead to adverse effects and exert tissue damage in pathological conditions. Here, we show that the ATP channel pannexin1 (Panx1) is functionally expressed by bone marrow-derived neutrophils (BMDNs) of wild-type (WT) mice and that ATP contributes to NETosis induced in vitro by the calcium ionophore A23187 or phorbol 12-myristate 13-acetate (PMA). Interestingly, neutrophils isolated from Panx1-/- mice showed reduced and/or delayed induction of NETosis. Brilliant blue FCF dye (BB-FCF), a Panx1 channel inhibitor, decreased NETosis in wild-type neutrophils to the extent observed in Panx1-/- neutrophils. Thus, we demonstrate that ATP and Panx1 channels contribute to NETosis and may represent a therapeutic target.
| S-EPMC6851112 | biostudies-literature
Project description:Because neutrophil extracellular trap (NET) formation is involved in the pathology of a wide variety of diseases, NET-regulating compounds are expected to be useful for the therapies of these diseases. In this study, we identified sulfasalazine (SSZ) as a potent enhancer of NET formation both in vitro and in vivo. Although SSZ did not increase the amount of ROS generated, it accelerated the generation of ether-linked oxidized phospholipids, such as PE (18;1e/15-HETE) and PC (16;0e/13-HODE). Trolox, but not 2-ME, effectively suppressed lipid oxidation and NET formation that were induced by SSZ. SSZ is known as a potent inducer of ferroptosis in cancer cells by inhibiting xCT, a component of the cystine transporter. However, we found that SSZ accelerated NET formation in an xCT-independent manner. Structure-activity relationship studies revealed that the sulfapyridine moiety of SSZ plays a central role in enhancing NET formation. Furthermore, we found that two additional sulfonamide and sulfone derivatives possess NET-inducing activity by accelerating lipid oxidation. These results indicate that the hyperoxidation of ether-linked phospholipids is a key mechanism for accelerating NET formation.
| S-EPMC5700140 | biostudies-literature
Project description:IntroductionNeutrophil extracellular trap (NET) formation was recently identified as a novel mechanism to kill pathogens. However, excessive NET formation in sepsis can injure host tissues. We have recently shown that parenteral vitamin C (VitC) is protective in sepsis. Whether VitC alters NETosis is unknown.MethodsWe used Gulo-/- mice as they lack the ability to synthesize VitC. Sepsis was induced by intraperitoneal infusion of a fecal stem solution (abdominal peritonitis, FIP). Some VitC deficient Gulo-/- mice received an infusion of ascorbic acid (AscA, 200 mg/kg) 30 min after induction of FIP. NETosis was assessed histologically and by quantification for circulating free DNA (cf-DNA) in serum. Autophagy, histone citrullination, endoplasmic reticulum (ER) stress, NF?B activation and apoptosis were investigated in peritoneal PMNs.ResultsSepsis produced significant NETs in the lungs of VitC deficient Gulo-/- mice and increased circulating cf-DNA. This was attenuated in the VitC sufficient Gulo-/- mice and in VitC deficient Gulo-/- mice infused with AscA. Polymorphonuclear neutrophils (PMNs) from VitC deficient Gulo-/- mice demonstrated increased activation of ER stress, autophagy, histone citrullination, and NF?B activation, while apoptosis was inhibited. VitC also significantly attenuated PMA induced NETosis in PMNs from healthy human volunteers.ConclusionsOur in vitro and in vivo findings identify VitC as a novel regulator of NET formation in sepsis. This study complements the notion that VitC is protective in sepsis settings.
| S-EPMC3775246 | biostudies-literature
Project description:The endosymbiotic bacteria, Wolbachia, induce neutrophilic responses to the human helminth pathogen Onchocerca volvulus. The formation of Neutrophil Extracellular Traps (NETs), has been implicated in anti-microbial defence, but has not been identified in human helminth infection. Here, we demonstrate NETs formation in human onchocerciasis. Extracellular NETs and neutrophils were visualised around O. volvulus in nodules excised from untreated patients but not in nodules from patients treated with the anti-Wolbachia drug, doxycycline. Whole Wolbachia or microspheres coated with a synthetic Wolbachia lipopeptide (WoLP) of the major nematode Wolbachia TLR2/6 ligand, peptidoglycan associated lipoprotein, induced NETosis in human neutrophils in vitro. TLR6 dependency of Wolbachia and WoLP NETosis was demonstrated using purified neutrophils from TLR6 deficient mice. Thus, we demonstrate for the first time that NETosis occurs during natural human helminth infection and demonstrate a mechanism of NETosis induction via Wolbachia endobacteria and direct ligation of Wolbachia lipoprotein by neutrophil TLR2/6.
| S-EPMC5067710 | biostudies-literature
Project description:Neutrophils form neutrophil extracellular traps (NETs), which have been implicated in microcirculatory plugging. NET formation (NETosis) involves the fusion of granule and nuclear contents, which are then released in the extracellular space. Myeloperoxidase (MPO) plays a major role in NETosis leading to the dissociation of DNA from histones. During neutrophil activation, MPO is released and activated to convert hydrogen peroxide and chloride to hypochlorous acid (HOCl). HOCl targets plasmalogens leading to the production of the chlorinated lipids, 2-chlorofatty aldehyde and 2-chlorofatty acid (2-ClFA). Here, we tested the hypothesis that 2-ClFAs are important lipid mediators of NETosis. Human neutrophils treated with physiological levels of 2-ClFAs formed NETs, characterized by MPO association with DNA and neutrophil elastase (NE) redistribution to the perinuclear area. 2-ClFA-induced NETs reduced Escerichia coli colony forming units. 2-ClFA-induced NETosis is calcium- and protein arginine deiminase 4-dependent. Interestingly, unlike PMA, 2-ClFA initiates the NETosis process without neutrophil activation and degranulation. Furthermore, 2-ClFA elicits NETosis in bone-marrow derived neutrophils from MPO-deficient mice. Taken together, these findings suggest 2-ClFA as an MPO product that triggers the NETosis pathway following neutrophil activation.
| S-EPMC6071778 | biostudies-literature
Project description:BackgroundNeutrophil extracellular traps (NETs) constitute antimicrobial function of neutrophils but have also been linked to perpetuation of inflammation. Despite this evident physiological relevance, mechanistic understanding of NET formation is poor. In this study, we examined the mechanism by which Mincle, a C-type lectin receptor, regulates NET formation.MethodsNET formation, reactive oxygen species, autophagy activation and intracellular signaling pathways were analyzed in Mincle-sufficient and -deficient neutrophils stimulated in vitro with various stimuli and in vivo during Klebsiella infection.ResultsWe found that Mincle mediates NET formation in response to several activation stimuli in vitro and in vivo during pneumoseptic infection with Klebsiella pneumoniae, indicating its regulatory role in NET formation. Mechanistically, we show that attenuated NET formation in Mincle-/- neutrophils correlates with an impaired autophagy activation in vitro and in vivo, whereas reactive oxygen species (ROS) formation in these neutrophils remained intact. The requirement of autophagy in Mincle-mediated NET formation was further supported by exogenous treatment with autophagy inducer tamoxifen, which rescued the NET formation defect in Mincle-/- neutrophils.ConclusionsOur findings identify a previously unrecognized role of Mincle as a regulator of autophagy, which mediates NET formation without affecting ROS generation. Our study addresses a major challenge in the field by positing this pathway to be targeted for modulation of NETs while preserving ROS production, an important innate immune defense.
| S-EPMC5426377 | biostudies-literature