CD276 regulates the immune escape of esophageal squamous cell carcinoma through CXCL1-CXCR2 induced NETs.

BACKGROUND: CD276 (B7-H3), a pivotal immune checkpoint, facilitates tumorigenicity, invasiveness, and metastasis by escaping immune surveillance in a variety of tumors; however, the underlying mechanisms facilitating immune escape in esophageal squamous cell carcinoma (ESCC) remain enigmatic. METHODS: We investigated the expression of CD276 in ESCC tissues from patients by using immunohistochemistry (IHC) assays. In vivo, we established a 4-nitroquinoline 1-oxide (4NQO)-induced CD276 knockout (CD276wKO) and K14cre; CD276 conditional knockout (CD276cKO) mouse model of ESCC to study the functional role of CD276 in ESCC. Furthermore, we used the 4NQO-induced mouse model to evaluate the effects of anti-CXCL1 antibodies, anti-Ly6G antibodies, anti-NK1.1 antibodies, and GSK484 inhibitors on tumor growth. Moreover, IHC, flow cytometry, and immunofluorescence techniques were employed to measure immune cell proportions in ESCC. In addition, we conducted single-cell RNA sequencing analysis to examine the alterations in tumor microenvironment following CD276 depletion. RESULTS: In this study, we elucidate that CD276 is markedly upregulated in ESCC, correlating with poor prognosis. In vivo, our results indicate that depletion of CD276 inhibits tumorigenesis and progression of ESCC. Furthermore, conditional knockout of CD276 in epithelial cells engenders a significant downregulation of CXCL1, consequently reducing the formation of neutrophil extracellular trap networks (NETs) via the CXCL1-CXCR2 signaling axis, while simultaneously augmenting natural killer (NK) cells. In addition, overexpression of CD276 promotes tumorigenesis via increasing NETs' formation and reducing NK cells in vivo. CONCLUSIONS: This study successfully elucidates the functional role of CD276 in ESCC. Our comprehensive analysis uncovers the significant role of CD276 in modulating immune surveillance mechanisms in ESCC, thereby suggesting that targeting CD276 might serve as a potential therapeutic approach for ESCC treatment.

The role of platelets in central hubs of inflammation: A literature review.

Platelets are increasingly recognized for their multifaceted roles in inflammation beyond their traditional involvement in haemostasis. This review consolidates knowledge on platelets as critical players in inflammatory responses. This study did an extensive search of electronic databases and identified studies on platelets in inflammation, focusing on molecular mechanisms, cell interactions, and clinical implications, emphasizing recent publications. Platelets contribute to inflammation via surface receptors, release of mediators, and participation in neutrophil extracellular trap formation. They are implicated in diseases like atherosclerosis, rheumatoid arthritis, and sepsis, highlighting their interaction with immune cells as pivotal in the onset and resolution of inflammation. Platelets are central to regulating inflammation, offering new therapeutic targets for inflammatory diseases. Future research should explore specific molecular pathways of platelets in inflammation for therapeutic intervention.

The Bifunctional Effects of Lactoferrin (LFcinB11) in Inhibiting Neural Cell Adhesive Molecule (NCAM) Polysialylation and the Release of Neutrophil Extracellular Traps (NETs).

The expression of polysialic acid (polySia) on the neuronal cell adhesion molecule (NCAM) is called NCAM-polysialylation, which is strongly related to the migration and invasion of tumor cells and aggressive clinical status. Thus, it is important to select a proper drug to block tumor cell migration during clinical treatment. In this study, we proposed that lactoferrin (LFcinB11) may be a better candidate for inhibiting NCAM polysialylation when compared with CMP and low-molecular-weight heparin (LMWH), which were determined based on our NMR studies. Furthermore, neutrophil extracellular traps (NETs) represent the most dramatic stage in the cell death process, and the release of NETs is related to the pathogenesis of autoimmune and inflammatory disorders, with proposed involvement in glomerulonephritis, chronic lung disease, sepsis, and vascular disorders. In this study, the molecular mechanisms involved in the inhibition of NET release using LFcinB11 as an inhibitor were also determined. Based on these results, LFcinB11 is proposed as being a bifunctional inhibitor for inhibiting both NCAM polysialylation and the release of NETs.

A2AR-mediated CXCL5 upregulation on macrophages promotes NSCLC progression via NETosis.

Tumor-associated macrophages (TAMs) are abundant in tumors and interact with tumor cells, leading to the formation of an immunosuppressive microenvironment and tumor progression. Although many studies have explored the mechanisms underlying TAM polarization and its immunosuppressive functions, understanding of its progression remains limited. TAMs promote tumor progression by secreting cytokines, which subsequently recruit immunosuppressive cells to suppress the antitumor immunity. In this study, we established an in vitro model of macrophage and non-small cell lung cancer (NSCLC) cell co-culture to explore the mechanisms of cell-cell crosstalk. We observed that in NSCLC, the C-X-C motif chemokine ligand 5 (CXCL5) was upregulated in macrophages because of the stimulation of A2AR by adenosine. Adenosine was catalyzed by CD39 and CD73 in macrophages and tumor cells, respectively. Nuclear factor kappa B (NFκB) mediated the A2AR stimulation of CXCL5 upregulation in macrophages. Additionally, CXCL5 stimulated NETosis in neutrophils. Neutrophil extracellular traps (NETs)-treated CD8+ T cells exhibited upregulation of exhaustion-related and cytosolic DNA sensing pathways and downregulation of effector-related genes. However, A2AR inhibition significantly downregulated CXCL5 expression and reduced neutrophil infiltration, consequently alleviating CD8+ T cell dysfunction. Our findings suggest a complex interaction between tumor and immune cells and its potential as therapeutic target.

Neutrophil extracellular traps promote proliferation of pulmonary smooth muscle cells mediated by CCDC25 in pulmonary arterial hypertension.

BACKGROUND: Previous studies have indicated that neutrophil extracellular traps (NETs) play a pivotal role in pathogenesis of pulmonary arterial hypertension (PAH). However, the specific mechanism underlying the impact of NETs on pulmonary artery smooth muscle cells (PASMCs) has not been determined. The objective of this study was to elucidate underlying mechanisms through which NETs contribute to progression of PAH. METHODS: Bioinformatics analysis was employed in this study to screen for potential molecules and mechanisms associated with occurrence and development of PAH. These findings were subsequently validated in human samples, coiled-coil domain containing 25 (CCDC25) knockdown PASMCs, as well as monocrotaline-induced PAH rat model. RESULTS: NETs promoted proliferation of PASMCs, thereby facilitating pathogenesis of PAH. This phenomenon was mediated by the activation of transmembrane receptor CCDC25 on PASMCs, which subsequently activated ILK/ß-parvin/RAC1 pathway. Consequently, cytoskeletal remodeling and phenotypic transformation occur in PASMCs. Furthermore, the level of NETs could serve as an indicator of PAH severity and as potential therapeutic target for alleviating PAH. CONCLUSION: This study elucidated the involvement of NETs in pathogenesis of PAH through their influence on the function of PASMCs, thereby highlighting their potential as promising targets for the evaluation and treatment of PAH.

Effect of F-isoprostane class on cow peripheral blood neutrophil microbicidal function in vitro.

Isoprostanes (isoP) are formed during conditions of oxidative stress (OS) through the oxidation of cell membrane fatty acids. Different classes of isoP are formed depending on the fatty acid being oxidized but the biological activity of these molecules in innate immune cells is poorly understood. Thus, the objective of this study was to compare in vitro the effects of F2- and F3-isoP on neutrophil microbicidal functions. We isolated neutrophils from 6 dairy cows and incubated them for 8 h at various concentrations of F2- and F3-isoP. Then, microbicidal function was assessed in terms of phagocytosis, respiratory burst, myeloperoxidase activity, and extracellular trap formation. In vitro supplementation with F3-isoP enhanced microbicidal capabilities whereas supplementation with F2-isoP decreased or did not impact these microbe killing functions. Hence, favoring the production of F3- over F2-isoprostanes may be a strategy to augment neutrophils' functional capacity during OS conditions. This should be tested in vivo.

Myocardial reperfusion injury exacerbation due to ALDH2 deficiency is mediated by neutrophil extracellular traps and prevented by leukotriene C4 inhibition.

BACKGROUND AND AIMS: The Glu504Lys polymorphism in the aldehyde dehydrogenase 2 (ALDH2) gene is closely associated with myocardial ischaemia/reperfusion injury (I/RI). The effects of ALDH2 on neutrophil extracellular trap (NET) formation (i.e. NETosis) during I/RI remain unknown. This study aimed to investigate the role of ALDH2 in NETosis in the pathogenesis of myocardial I/RI. METHODS: The mouse model of myocardial I/RI was constructed on wild-type, ALDH2 knockout, peptidylarginine deiminase 4 (Pad4) knockout, and ALDH2/PAD4 double knockout mice. Overall, 308 ST-elevation myocardial infarction patients after primary percutaneous coronary intervention were enrolled in the study. RESULTS: Enhanced NETosis was observed in human neutrophils carrying the ALDH2 genetic mutation and ischaemic myocardium of ALDH2 knockout mice compared with controls. PAD4 knockout or treatment with NETosis-targeting drugs (GSK484, DNase1) substantially attenuated the extent of myocardial damage, particularly in ALDH2 knockout. Mechanistically, ALDH2 deficiency increased damage-associated molecular pattern release and susceptibility to NET-induced damage during myocardial I/RI. ALDH2 deficiency induced NOX2-dependent NETosis via upregulating the endoplasmic reticulum stress/microsomal glutathione S-transferase 2/leukotriene C4 (LTC4) pathway. The Food and Drug Administration-approved LTC4 receptor antagonist pranlukast ameliorated I/RI by inhibiting NETosis in both wild-type and ALDH2 knockout mice. Serum myeloperoxidase-DNA complex and LTC4 levels exhibited the predictive effect on adverse left ventricular remodelling at 6 months after primary percutaneous coronary intervention in ST-elevation myocardial infarction patients. CONCLUSIONS: ALDH2 deficiency exacerbates myocardial I/RI by promoting NETosis via the endoplasmic reticulum stress/microsomal glutathione S-transferase 2/LTC4/NOX2 pathway. This study hints at the role of NETosis in the pathogenesis of myocardial I/RI, and pranlukast might be a potential therapeutic option for attenuating I/RI, particularly in individuals with the ALDH2 mutation.

Resveratrol prevents the release of neutrophil extracellular traps (NETs) by controlling hydrogen peroxide levels and nuclear elastase migration.

Neutrophil extracellular traps (NETs) are defense mechanisms that trap and kill microorganisms and degrade cytokines. However, excessive production, dysregulation of suppression mechanisms, or inefficient removal of NETs can contribute to increased inflammatory response and the development of pathological conditions. Therefore, research has focused on identifying drugs that inhibit or delay the NET release process. Since reactive oxygen species (ROS) play a significant role in NET release, we aimed to investigate whether resveratrol (RSV), with a wide range of biological and pharmacological properties, could modulate NET release in response to different stimuli. Thus, human neutrophils were pretreated with RSV and subsequently stimulated with PMA, LPS, IL-8, or Leishmania. Our findings revealed that RSV reduced the release of NETs in response to all tested stimuli. RSV decreased hydrogen peroxide levels in PMA- and LPS-stimulated neutrophils, inhibited myeloperoxidase activity, and altered the localization of neutrophil elastase. RSV inhibition of NET generation was not mediated through A2A or A2B adenosine receptors or PKA. Based on the observed effectiveness of RSV in inhibiting NET release, our study suggests that this flavonoid holds potential as a candidate for treating NETs involving pathologies.

Icariside II alleviates lipopolysaccharide-induced acute lung injury by inhibiting lung epithelial inflammatory and immune responses mediated by neutrophil extracellular traps.

AIMS: Acute lung injury (ALI) is a life-threatening lung disease characterized by inflammatory cell infiltration and lung epithelial injury. Icariside II (ICS II), one of the main active ingredients of Herba Epimedii, exhibits anti-inflammatory and immunomodulatory effects. However, the effect and mechanism of ICS II in ALI remain unclear. The purpose of the current study was to investigate the pharmacological effect and underlying mechanism of ICS II in ALI. MAIN METHODS: Models of neutrophil-like cells, human peripheral blood neutrophils, and lipopolysaccharide (LPS)-induced ALI mouse model were utilized. RT-qPCR and Western blotting determined the gene and protein expression levels. Protein distribution and quantification were analyzed by immunofluorescence. KEY FINDINGS: ICS II significantly reduced lung histopathological damage, edema, and inflammatory cell infiltration, and it reduced pro-inflammatory cytokines in ALI. There is an excessive activation of neutrophils leading to a significant production of NETs in ALI mice, a process mitigated by the administration of ICS II. In vivo and in vitro studies found that ICS II could decrease NET formation by targeting neutrophil C-X-C chemokine receptor type 4 (CXCR4). Further data showed that ICS II reduces the overproduction of dsDNA, a NETs-related component, thereby suppressing cGAS/STING/NF-κB signalling pathway activation and inflammatory mediators release in lung epithelial cells. SIGNIFICANCE: This study suggested that ICS II may alleviate LPS-induced ALI by modulating the inflammatory response, indicating its potential as a therapeutic agent for ALI treatment.

[Expression of neutrophil extracellular traps and phagocytic functions among patients with hepatic alveolar echinococcosis].

OBJECTIVE: To investigate the expression of neutrophil extracellular traps (NETs) and phagocytic function in the peripheral blood of patients with hepatic alveolar echinococcosis (HAE), and to examine their correlations with clinical inflamma tory indicators and liver functions. METHODS: A total of 50 patients with HAE admitted to Department of Hepatobiliary and Pancreatic Surgery, The Affiliated Hospital of Qinghai University from August 2022 to June 2023 were enrolled, while 50 age- and gender-matched healthy individuals from the Centre for Healthy Examinations of the hospital during the same period served as controls. The levels of NETs markers neutrophil myeloperoxidase (MPO) and neutrophil elastase (NE) were measured using enzyme-linked immunosorbent assay (ELISA). Peripheral blood neutrophils were isolated using density gradient centrifugation, stimulated in vitro using phorbol 12-myristate 13 acetate (PMA), and the levels of MPO and citrullination histone H3 (CitH3) released by neutrophils were quantified using flow cytometry. The phagocytic functions of neutrophils were examined using flow cytometry. In addition, the correlations of MPO and NE levels with clinical inflammatory indicators and liver biochemical indicators were examined using Spearman correlation analysis among HAE patients. RESULTS: The peripheral blood plasma MPOï¼»(417.15 ± 76.08) ng/mL vs. (255.70 ± 80.84) ng/mL; t = 10.28, P < 0.05ï¼½, NEï¼»(23.16 ± 6.75) ng/mL vs. (11.92 ± 3.17) ng/mL; t = 10.65, P < 0.05ï¼½and CitH3 levelsï¼»(33.93 ± 18.93) ng/mL vs. (19.52 ± 13.89) ng/mL; t = 4.34, P < 0.05ï¼½were all significantly higher among HAE patients than among healthy controls, and a lower phagocytosis rate of neutrophils was detected among HAE patients than among healthy controlsï¼»(70.85 ± 7.32)% vs. (94.04 ± 3.90)%; t = 20.18, P < 0.05ï¼½, and the ability to produce NETs by neutrophils was higher among HAE patients than among healthy controls following in vitro PMA stimulation. Pearson correlation analysis showed that the phagocytosis rate of neutrophils correlated negatively with platelet-to-lymphocyte ratio (PLR), neutrophil-to-lymphocyte ratio (NLR), interleukin-6 (IL-6) level and C-reactive protein (CRP) level (rs = -0.515 to -0.392, all P values < 0.05), and the MPO and NE levels positively correlated with inflammatory markers NLR, PLR, CRP and IL-6 (rs = 0.333 to 0.445, all P values < 0.05) and clinical liver biochemical indicators aspartic transaminase, alanine aminotransferase, direct bilirubin and total bilirubin among HAE patients (rs = 0.290 to 0.628, all P values < 0.001). CONCLUSIONS: Excessive formation of NETs is found among HAE patients, which affects the phagocytic ability of neutrophils and results in elevated levels of inflammatory indicators. NETs markers may be promising novel biomarkers for early diagnosis, monitoring, and severity assessment of liver disease.

Composition and Function of Neutrophil Extracellular Traps.

Neutrophil extracellular traps (NETs) are intricate fibrous structures released by neutrophils in response to specific stimuli. These structures are composed of depolymerized chromatin adorned with histones, granule proteins, and cytosolic proteins. NETs are formed via two distinct pathways known as suicidal NETosis, which involves NADPH oxidase (NOX), and vital NETosis, which is independent of NOX. Certain proteins found within NETs exhibit strong cytotoxic effects against both pathogens and nearby host cells. While NETs play a defensive role against pathogens, they can also contribute to tissue damage and worsen inflammation. Despite extensive research on the pathophysiological role of NETs, less attention has been paid to their components, which form a unique structure containing various proteins that have significant implications in a wide range of diseases. This review aims to elucidate the components of NETs and provide an overview of their impact on host defense against invasive pathogens, autoimmune diseases, and cancer.

Modelling the Impact of NETosis During the Initial Stage of Systemic Lupus Erythematosus.

The development of autoimmune diseases often takes years before clinical symptoms become detectable. We propose a mathematical model for the immune response during the initial stage of Systemic Lupus Erythematosus which models the process of aberrant apoptosis and activation of macrophages and neutrophils. NETosis is a type of cell death characterised by the release of neutrophil extracellular traps, or NETs, containing material from the neutrophil's nucleus, in response to a pathogenic stimulus. This process is hypothesised to contribute to the development of autoimmunogenicity in SLE. The aim of this work is to study how NETosis contributes to the establishment of persistent autoantigen production by analysing the steady states and the asymptotic dynamics of the model by numerical experiment.

Membrane damage by MBP-1 is mediated by pore formation and amplified by mtDNA.

Eosinophils play a crucial role in host defense while also contributing to immunopathology through the release of inflammatory mediators. Characterized by distinctive cytoplasmic granules, eosinophils securely store and rapidly release various proteins exhibiting high toxicity upon extracellular release. Among these, major basic protein 1 (MBP-1) emerges as an important mediator in eosinophil function against pathogens and in eosinophil-associated diseases. While MBP-1 targets both microorganisms and host cells, its precise mechanism remains elusive. We demonstrate that formation of small pores by MBP-1 in lipid bilayers induces membrane permeabilization and disrupts potassium balance. Additionally, we reveal that mitochondrial DNA (mtDNA) present in eosinophil extracellular traps (EETs) amplifies MBP-1 toxic effects, underscoring the pivotal role of mtDNA in EETs. Furthermore, we present evidence indicating that absence of CpG methylation in mtDNA contributes to the regulation of MBP-1-mediated toxicity. Taken together, our data suggest that the mtDNA scaffold within extracellular traps promotes MBP-1 toxicity.

Lidocaine effects on neutrophil extracellular trapping and angiogenesis biomarkers in postoperative breast cancer patients with different anesthesia methods: a prospective, randomized trial.

BACKGROUND: Anesthesia techniques and drug selection may influence tumor recurrence and metastasis. Neutrophil extracellular trapping (NETosis), an immunological process, has been linked to an increased susceptibility to metastasis in individuals with tumors. Furthermore, recurrence may be associated with vascular endothelial growth factor A (VEGF-A), a mediator of angiogenesis. This study investigates the impact of lidocaine (combined with sevoflurane or propofol anesthesia ) during breast cancer surgery inhibits the expression of biomarkers associated with metastasis and recurrence (specifically H3Cit, NE, MPO, MMP-9 and VEGF-A). METHODS: We randomly assigned 120 women undergoing primary or invasive breast tumor resection to receive one of four anesthetics: sevoflurane (S), sevoflurane plus i.v. lidocaine (SL), propofol (P), and propofol plus i.v. lidocaine (PL). Blood samples were collected before induction and 3 h after the operation. Biomarkers associated with NETosis (citrullinated histone H3 [H3Cit], myeloperoxidase [MPO], and neutrophil elastase [NE]) and angiogenesis were quantified using enzyme-linked immunosorbent assays. RESULTS: Patient and breast tumor characteristics, along with perioperative management, did not differ between study groups. In intra-group comparisons, S and P groups demonstrated a statistically significant increase in post-operative MPO (S group: 10.39[6.89-17.22] vs. 14.31[8.55-20.87] ng ml-1, P = 0.032; P group: 9.45[6.73-17.37] vs. 14.34[9.87-19.75] ng ml-1, P = 0.035)and NE(S group: 182.70[85.66-285.85] vs. 226.20[91.85-391.65] ng ml-1, P = 0.045; P group: 154.22[97.31-325.30] vs. 308.66[132.36-483.57] ng ml-1, P = 0.037) concentrations compared to pre-operative measurements, whereas SL and PL groups did not display a similar increase. H3Cit, MMP-9, and VEGF-A concentrations were not significantly influenced by the anesthesia techniques and drugs. CONCLUSIONS: Regardless of the specific technique employed for general anesthesia, there was no increase in the postoperative serum concentrations of MPO and NE after perioperative lidocaine infusion compared to preoperative serum concentrations. This supports the hypothesis that intravenous lidocaine during cancer surgery aimed at achieving a cure may potentially decrease the likelihood of recurrence. Further interpretation and discussion of clinical implications are warranted, emphasizing the significance of these findings in the context of cancer surgery and recurrence prevention. CLINICAL TRIAL REGISTRATION: ChiCTR2300068563.

Predicting the immunity landscape and prognosis with an NCLs signature in liver hepatocellular carcinoma.

BACKGROUND: Activated neutrophils release depolymerized chromatin and protein particles into the extracellular space, forming reticular Neutrophil Extracellular Traps (NETs). This process is accompanied by programmed inflammatory cell death of neutrophils, known as NETosis. Previous reports have demonstrated that NETosis plays a significant role in immune resistance and microenvironmental regulation in cancer. This study sought to characterize the function and molecular mechanism of NETosis-correlated long non-coding RNAs (NCLs) in the prognostic treatment of liver hepatocellular carcinoma (LIHC). METHODS: We obtained the transcriptomic and clinical data from The Cancer Genome Atlas (TCGA) and evaluated the expression of NCLs in LIHC. A prognostic signature of NCLs was constructed using Cox and Last Absolute Shrinkage and Selection Operator (Lasso) regression, while the accuracy of model was validated by the ROC curves and nomogram, etc. In addition, we analyzed the associations between NCLs and oncogenic mutation, immune infiltration and evasion. Finally, LIHC patients were classified into four subgroups based on consensus cluster analysis, and drug sensitivity was predicted. RESULTS: After screening, we established a risk model combining 5 hub-NCLs and demonstrated its reliability. Independence checks suggest that the model may serve as an independent predictor of LIHC prognosis. Enrichment analysis revealed a concentration of immune-related pathways in the high-risk group. Immune infiltration indicates that immunotherapy could be more effective in the low-risk group. Upon consistent cluster analysis, cluster subgroup 4 presented a better prognosis. Sensitivity tests showed the distinctions in therapeutic effectiveness among various drugs in different subgroups. CONCLUSION: Overall, we have developed a prognostic signature that can discriminate different LIHC subgroups through the 5 selected NCLs, with the objective of providing LIHC patients a more precise, personalized treatment regimen.

A promising frontier: targeting NETs for stroke treatment breakthroughs.

Stroke is a prevalent global acute cerebrovascular condition, with ischaemic stroke being the most frequently occurring type. After a stroke, neutrophils accumulate in the brain and subsequently generate and release neutrophil extracellular traps (NETs). The accumulation of NETs exacerbates the impairment of the blood‒brain barrier (BBB), hampers neovascularization, induces notable neurological deficits, worsens the prognosis of stroke patients, and can facilitate the occurrence of t-PA-induced cerebral haemorrhage subsequent to ischaemic stroke. Alternative approaches to pharmacological thrombolysis or endovascular thrombectomy are being explored, and targeting NETs is a promising treatment that warrants further investigation.

Pro-inflammatory interactions of streptolysin O toxin with human neutrophils in vitro.

The recent global resurgence of severe infections caused by the Group A streptococcus (GAS) pathogen, Streptococcus pyogenes, has focused attention on this microbial pathogen, which produces an array of virulence factors, such as the pore-forming toxin, streptolysin O (SOT). Importantly, the interactions of SOT with human neutrophils (PMN), are not well understood. The current study was designed to investigate the effects of pretreatment of isolated human PMN with purified SOT on several pro-inflammatory activities, including generation of reactive oxygen species (ROS), degranulation (elastase release), influx of extracellular calcium (Ca2+) and release of extracellular DNA (NETosis), using chemiluminescence, spectrophotometric and fluorimetric procedures, respectively. Exposure of PMN to SOT alone caused modest production of ROS and elastase release, while pretreatment with the toxin caused significant augmentation of chemoattractant (fMLP)-activated ROS generation and release of elastase by activated PMN. These effects of treatment of PMN with SOT were associated with both a marked and sustained elevation of cytosolic Ca2+concentrations and significant increases in the concentrations of extracellular DNA, indicative of NETosis. The current study has identified a potential role for SOT in augmenting the Ca2+-dependent pro-inflammatory interactions of PMN, which, if operative in a clinical setting, may contribute to hyper-activation of PMN and GAS-mediated tissue injury.

[Neutrophil extracellular traps extrusion from neutrophils stably adhered to ICAM-1 by lipoteichoic acid stimulation].

The effect of neutrophil extracellular traps (NETs) on promoting intravascular microthrombi formation and exacerbating the severity of sepsis in patients has gained extensive attention. However, in sepsis, the mechanisms and key signaling molecules mediating NET formation during direct interactions of endothelial cells and neutrophils still need further explored. Herein, we utilized lipoteichoic acid (LTA), a component shared by Gram-positive bacteria, to induce NET extrusion from neutrophils firmly adhered to the glass slides coated with intercellular adhesion molecule-1(ICAM-1). We also used Sytox green to label NET-DNA and Flou-4 AM as the intracellular Ca 2+ signaling indicator to observe the NET formation and fluctuation of Ca 2+ signaling. Our results illustrated that LTA was able to induce NET release from neutrophils firmly attached to ICAM-1-coated glass slides, and the process was time-dependent. In addition, our study indicated that LTA-induced NET release by neutrophils stably adhered to ICAM-1 depended on Ca 2+ signaling but not intracellular reactive oxygen species (ROS). This study reveals NET formation mediated by direct interactions between endothelial ICAM-1 and neutrophils under LTA stimulation and key signaling molecules involved, providing the theoretical basis for medicine development and clinical treatment for related diseases.

Neutrophils display distinct post-translational modifications in response to varied pathological stimuli.

Neutrophils play a vital role in the innate immunity by perform effector functions through phagocytosis, degranulation, and forming extracellular traps. However, over-functioning of neutrophils has been associated with sterile inflammation such as Type 2 Diabetes, atherosclerosis, cancer and autoimmune disorders. Neutrophils exhibiting phenotypical and functional heterogeneity in both homeostatic and pathological conditions suggests distinct signaling pathways are activated in disease-specific stimuli and alter neutrophil functions. Hence, we examined mass spectrometry based post-translational modifications (PTM) of neutrophil proteins in response to pathologically significant stimuli, including high glucose, homocysteine and bacterial lipopolysaccharides representing diabetes-indicator, an activator of thrombosis and pathogen-associated molecule, respectively. Our data revealed that these aforesaid stimulators differentially deamidate, citrullinate, acetylate and methylate neutrophil proteins and align to distinct biological functions associated with degranulation, platelet activation, innate immune responses and metabolic alterations. The PTM patterns in response to high glucose showed an association with neutrophils extracellular traps (NETs) formation, homocysteine induced proteins PTM associated with signaling of systemic lupus erythematosus and lipopolysaccharides induced PTMs were involved in pathways related to cardiomyopathies. Our study provides novel insights into neutrophil PTM patterns and functions in response to varied pathological stimuli, which may serve as a resource to design therapeutic strategies for the management of neutrophil-centred diseases.