Specific Increase in Joint Neutrophil Extracellular Traps and Its Relation to Interleukin 6 in Autoimmune Arthritis

Neutrophils and their extracellular traps have been shown to play an important role in the pathogenesis of rheumatoid arthritis (RA), but the detailed mechanisms in joints are still unclear, and their regulation remains to be solved. Here, we explored neutrophil extracellular trap (NET)osis in experimental models of arthritis and further investigated the effects of interleukin-6 (IL-6) inhibition in neutrophils and NETosis. In skins of peptide GPI-induced arthritis (pGIA), citrullinated protein was detected as well as citrullinated histone expression in immunized skin but this was not specific to pGIA. Citrullinated histone expression in pGIA joints was specific to pGIA and was merged with neutrophil elastase, suggesting NETosis. Neutrophils in joints tend to upregulate IL-6 receptors when compared with bone marrow neutrophils. Administration of mouse anti-IL-6 receptor antibodies in pGIA suppressed arthritis in association with a decrease in neutrophil infiltration and NETosis in joints. In the plasma of RA patients, citrullinated protein was significantly reduced after tocilizumab treatment. Our results suggest that IL-6 enhances neutrophil chemotaxis and NETosis in inflammatory joints and could be the source of citrullinated proteins.

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Anti-Citrullinated Protein Antibodies in Patients with Rheumatoid Arthritis: Biological Effects and Mechanisms of Immunopathogenesis

International Journal of Molecular Sciences ◽

10.3390/ijms21114015 ◽

2020 ◽

Vol 21 (11) ◽

pp. 4015 ◽

Cited By ~ 1

Author(s):

Chao-Yi Wu ◽

Huang-Yu Yang ◽

Jenn-Haung Lai

Keyword(s):

Rheumatoid Arthritis ◽

Biological Effects ◽

Cross Reactivity ◽

Neutrophil Extracellular Trap ◽

Gamma Receptor ◽

Increased Risk ◽

Citrullinated Proteins ◽

Loss Of Tolerance ◽

System Activation ◽

Citrullinated Protein

Individuals with high anti-citrullinated protein antibody (ACPA) titers have an increased risk of developing rheumatoid arthritis (RA). Although our knowledge of the generation and production of ACPAs has continuously advanced during the past decade, our understanding on the pathogenic mechanisms of how ACPAs interact with immune cells to trigger articular inflammation is relatively limited. Citrullination disorders drive the generation and maintenance of ACPAs, with profound clinical significance in patients with RA. The loss of tolerance to citrullinated proteins, however, is essential for ACPAs to exert their pathogenicity. N-linked glycosylation, cross-reactivity and the structural interactions of ACPAs with their citrullinated antigens further direct their biological functions. Although questions remain in the pathogenicity of ACPAs acting as agonists for a receptor-mediated response, immune complex (IC) formation, complement system activation, crystallizable fragment gamma receptor (FcγR) activation, cross-reactivity to joint cartilage and neutrophil extracellular trap (NET)-related mechanisms have all been suggested recently. This paper presents a critical review of the characteristics and possible biological effects and mechanisms of the immunopathogenesis of ACPAs in patients with RA.

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Role of Neutrophils on the Ocular Surface

International Journal of Molecular Sciences ◽

10.3390/ijms221910386 ◽

2021 ◽

Vol 22 (19) ◽

pp. 10386

Author(s):

Yongseok Mun ◽

Jin Sun Hwang ◽

Young Joo Shin

Keyword(s):

Ocular Surface ◽

Innate Immune System ◽

Neutrophil Infiltration ◽

Neutrophil Extracellular Traps ◽

Therapeutic Agents ◽

Innate Immune ◽

Neutrophil Extracellular Trap ◽

Extracellular Traps ◽

Ocular Surface Diseases

The ocular surface is a gateway that contacts the outside and receives stimulation from the outside. The corneal innate immune system is composed of many types of cells, including epithelial cells, fibroblasts, natural killer cells, macrophages, neutrophils, dendritic cells, mast cells, basophils, eosinophils, mucin, and lysozyme. Neutrophil infiltration and degranulation occur on the ocular surface. Degranulation, neutrophil extracellular traps formation, called NETosis, and autophagy in neutrophils are involved in the pathogenesis of ocular surface diseases. It is necessary to understand the role of neutrophils on the ocular surface. Furthermore, there is a need for research on therapeutic agents targeting neutrophils and neutrophil extracellular trap formation for ocular surface diseases.

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Mast Cell Tryptase Potentiates Neutrophil Extracellular Trap Formation

Journal of Innate Immunity ◽

10.1159/000520972 ◽

2021 ◽

pp. 1-14

Author(s):

Gunnar Pejler ◽

Sultan Alanazi ◽

Mirjana Grujic ◽

Jeremy Adler ◽

Anna-Karin Olsson ◽

...

Keyword(s):

Neutrophil Extracellular Trap ◽

Human Neutrophils ◽

Functional Communication ◽

Extracellular Milieu ◽

Extracellular Traps ◽

Inflammatory Conditions ◽

Histone 3 ◽

Activated Neutrophils ◽

Core Histones

Previous research has indicated an intimate functional communication between mast cells (MCs) and neutrophils during inflammatory conditions, but the nature of such communication is not fully understood. Activated neutrophils are known to release DNA-containing extracellular traps (neutrophil extracellular traps [NETs]) and, based on the known ability of tryptase to interact with negatively charged polymers, we here hypothesized that tryptase might interact with NET-contained DNA and thereby regulate NET formation. In support of this, we showed that tryptase markedly enhances NET formation in phorbol myristate acetate-activated human neutrophils. Moreover, tryptase was found to bind vividly to the NETs, to cause proteolysis of core histones and to cause a reduction in the levels of citrullinated histone-3. Secretome analysis revealed that tryptase caused increased release of numerous neutrophil granule compounds, including gelatinase, lactoferrin, and myeloperoxidase. We also show that DNA can induce the tetrameric, active organization of tryptase, suggesting that NET-contained DNA can maintain tryptase activity in the extracellular milieu. In line with such a scenario, DNA-stabilized tryptase was shown to efficiently degrade numerous pro-inflammatory compounds. Finally, we showed that tryptase is associated with NET formation in vivo in a melanoma setting and that NET formation in vivo is attenuated in mice lacking tryptase expression. Altogether, these findings reveal that NET formation can be regulated by MC tryptase, thus introducing a novel mechanism of communication between MCs and neutrophils.

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Preclinical characterization of the selective JAK1/2 inhibitor INCB018424: therapeutic implications for the treatment of myeloproliferative neoplasms

Blood ◽

10.1182/blood-2009-04-214957 ◽

2010 ◽

Vol 115 (15) ◽

pp. 3109-3117 ◽

Cited By ~ 449

Author(s):

Alfonso Quintás-Cardama ◽

Kris Vaddi ◽

Phillip Liu ◽

Taghi Manshouri ◽

Jun Li ◽

...

Keyword(s):

Polycythemia Vera ◽

Interleukin 6 ◽

Myeloproliferative Neoplasms ◽

Primary Cultures ◽

Myeloproliferative Neoplasm ◽

Primary Myelofibrosis ◽

Experimental Models ◽

Jak2v617f Mutation ◽

Erythroid Progenitor ◽

Therapeutic Implications

AbstractConstitutive JAK2 activation in hematopoietic cells by the JAK2V617F mutation recapitulates myeloproliferative neoplasm (MPN) phenotypes in mice, establishing JAK2 inhibition as a potential therapeutic strategy. Although most polycythemia vera patients carry the JAK2V617F mutation, half of those with essential thrombocythemia or primary myelofibrosis do not, suggesting alternative mechanisms for constitutive JAK-STAT signaling in MPNs. Most patients with primary myelofibrosis have elevated levels of JAK-dependent proinflammatory cytokines (eg, interleukin-6) consistent with our observation of JAK1 hyperactivation. Accordingly, we evaluated the effectiveness of selective JAK1/2 inhibition in experimental models relevant to MPNs and report on the effects of INCB018424, the first potent, selective, oral JAK1/JAK2 inhibitor to enter the clinic. INCB018424 inhibited interleukin-6 signaling (50% inhibitory concentration [IC50] = 281nM), and proliferation of JAK2V617F+ Ba/F3 cells (IC50 = 127nM). In primary cultures, INCB018424 preferentially suppressed erythroid progenitor colony formation from JAK2V617F+ polycythemia vera patients (IC50 = 67nM) versus healthy donors (IC50 > 400nM). In a mouse model of JAK2V617F+ MPN, oral INCB018424 markedly reduced splenomegaly and circulating levels of inflammatory cytokines, and preferentially eliminated neoplastic cells, resulting in significantly prolonged survival without myelosuppressive or immunosuppressive effects. Preliminary clinical results support these preclinical data and establish INCB018424 as a promising oral agent for the treatment of MPNs.

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Differential Use of Human Neutrophil FcγReceptors for Inducing Neutrophil Extracellular Trap Formation

Journal of Immunology Research ◽

10.1155/2016/2908034 ◽

2016 ◽

Vol 2016 ◽

pp. 1-17 ◽

Cited By ~ 38

Author(s):

Omar Rafael Alemán ◽

Nancy Mora ◽

Ricarda Cortes-Vieyra ◽

Eileen Uribe-Querol ◽

Carlos Rosales

Keyword(s):

Monoclonal Antibodies ◽

Human Neutrophil ◽

Neutrophil Extracellular Traps ◽

Neutrophil Extracellular Trap ◽

Cross Linking ◽

Erk Activation ◽

Extracellular Traps ◽

Extracellular Trap ◽

Antigen Antibody ◽

Insight Into

Neutrophils (PMN) are the most abundant leukocytes in the blood. PMN migrate from the circulation to sites of infection, where they are responsible for antimicrobial functions. PMN use phagocytosis, degranulation, and formation of neutrophil extracellular traps (NETs) to kill microbes. NETs are fibers composed of chromatin and neutrophil-granule proteins. Several pathogens, including bacteria, fungi, and parasites, and also some pharmacological stimuli such as phorbol 12-myristate 13-acetate (PMA) are efficient inducers of NETs. Antigen-antibody complexes are also capable of inducing NET formation. However the particular Fcγreceptor involved in triggering this function is a matter of controversy. In order to provide some insight into what Fcγreceptor is responsible for NET formation, each of the two human Fcγreceptors was stimulated individually by specific monoclonal antibodies and NET formation was evaluated. FcγRIIa cross-linking did not promote NET formation. Cross-linking other receptors such as integrins also did not promote NET formation. In contrast FcγRIIIb cross-linking induced NET formation similarly to PMA stimulation. NET formation was dependent on NADPH-oxidase, PKC, and ERK activation. These data show that cross-linking FcγRIIIb is responsible for NET formation by the human neutrophil.

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Degranulation of gastrointestinal mast cells contributes to hepatic ischemia–reperfusion injury in mice

Clinical Science ◽

10.1042/cs20180662 ◽

2018 ◽

Vol 132 (20) ◽

pp. 2241-2259 ◽

Cited By ~ 3

Author(s):

Zhigang He ◽

Yue Li ◽

Sunqiang Ma ◽

Muqing Yang ◽

Yuanyuan Ma ◽

...

Keyword(s):

Mast Cells ◽

Liver Damage ◽

Ischemia Reperfusion Injury ◽

Ischemia Reperfusion ◽

Neutrophil Infiltration ◽

Neutrophil Extracellular Trap ◽

Liver Sinusoidal Endothelial Cell ◽

Hepatic Ischemia ◽

Endothelial Cell Death ◽

Hepatic Ischemia Reperfusion

The pathological changes following liver damage, including those caused by ischemia and reperfusion (I/R), are closely related to gastrointestinal dysregulation. Mast cells (MCs) are tissue-resident immune cells abundant in the gastrointestinal system that play diverse roles. In view of the characteristic localization of MCs around the microvasculature, we hypothesized that a stimulus-specific set of mediators released through degranulation of gastrointestinal MCs, which are enriched in hepatic sinusoids via the hepatic system, subsequently participate in associated pathological development within the liver. To elucidate the biological role of gastrointestinal MC granules in liver damage, we employed an experimental liver I/R model that allows conditional ablation of MCs. Marked degranulation was detected during I/R, which showed a significant positive correlation with liver damage. Our experiments further disclosed that MC degranulation primarily enhanced the cycle of inflammatory damage in I/R liver consisting of liver sinusoidal endothelial cell death, neutrophil infiltration, and formation of a neutrophil extracellular trap, with a concomitant increase in adhesion molecules, inflammatory cytokines, chemokines, and oxidative stress. Based on the collective results, we propose that suppression of activity or number of MCs may present an effective strategy for protection against hepatic I/R injury.

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Anticitrullinated protein antibodies facilitate migration of synovial tissue-derived fibroblasts

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2018-214967 ◽

2019 ◽

Vol 78 (12) ◽

pp. 1621-1631 ◽

Cited By ~ 9

Author(s):

Meng Sun ◽

Bence Rethi ◽

Akilan Krishnamurthy ◽

Vijay Joshua ◽

Alexandra Circiumaru ◽

...

Keyword(s):

Joint Inflammation ◽

Arginine Deiminase ◽

Kinase Activation ◽

Cellular Effects ◽

Joint Pathology ◽

Protein Arginine Deiminase ◽

Citrullinated Proteins ◽

Phosphoinositide 3 Kinase ◽

Citrullinated Protein ◽

Fibroblast Like Synoviocytes

ObjectivesRheumatoid arthritis (RA)-specific anti-citrullinated protein/peptide antibodies (ACPAs) might contribute to bone loss and arthralgia before the onset of joint inflammation. We aimed to dissect additional mechanisms by which ACPAs might contribute to development of joint pathology.MethodsFibroblast-like synoviocytes (FLS) were isolated from the synovial membrane of patients with RA. The FLS cultures were stimulated with polyclonal ACPAs (anti-CCP-2 antibodies) purified from the peripheral blood of patients with RA or with monoclonal ACPAs derived from single synovial fluid B cells. We analysed how ACPAs modulate FLS by measuring cell adhesion and mobility as well as cytokine production. Expression of protein arginine deiminase (PAD) enzymes and protein citrullination were analysed by immunofluorescence, and signal transduction was studied using immunoblotting.ResultsChallenge of FLS by starvation-induced stress or by exposure to the chemokine interleukin-8 was essential to sensitise the cells to ACPAs. These challenges led to an increased PAD expression and protein citrullination and an ACPA-mediated induction of FLS migration through a mechanism involving phosphoinositide 3-kinase activation. Inhibition of the PAD enzymes or competition with soluble citrullinated proteins or peptides completely abolished the ACPA-induced FLS migration. Different monoclonal ACPAs triggered distinct cellular effects in either fibroblasts or osteoclasts, suggesting unique roles for individual ACPA clones in disease pathogenesis.ConclusionWe propose that transient synovial insults in the presence of a certain pre-existing ACPA repertoire might result in an ACPA-mediated increase of FLS migration.

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Transplantable myeloproliferative disease induced in mice by an interleukin 6 retrovirus.

Journal of Experimental Medicine ◽

10.1084/jem.176.4.1149 ◽

1992 ◽

Vol 176 (4) ◽

pp. 1149-1163 ◽

Cited By ~ 125

Author(s):

R G Hawley ◽

A Z Fong ◽

B F Burns ◽

T S Hawley

Keyword(s):

Bone Marrow ◽

Interleukin 6 ◽

Inflammatory Responses ◽

Bone Marrow Cells ◽

Constitutive Expression ◽

Fold Increase ◽

Neutrophil Infiltration ◽

Microcytic Anemia ◽

Myeloproliferative Disease

Lethally irradiated mice transplanted with bone marrow cells infected with a novel recombinant retrovirus (murine stem cell virus-interleukin 6 [MSCV-IL-6]) bearing a mouse IL-6 gene developed a fatal myeloproliferative disease within 4 wk of engraftment. The hematologic manifestations of the syndrome included elevated peripheral leukocyte counts (up to 430 x 10(3) cells/mm3) with a predominance of neutrophilic granulocytes, microcytic anemia, and thrombocytosis or thrombocytopenia. The mice showed extensive neutrophil infiltration of the lungs, liver, and occasionally lymph nodes, plus splenomegaly resulting from enhanced splenic myelopoiesis (30-60-fold increase in progenitor numbers). Despite the chronic stimulation of neutrophil excess by IL-6, bone marrow from affected mice was capable of repopulating the hematopoietic tissues (bone marrow and spleen) of lethally irradiated hosts during repeated serial transplantation. In the longest documented case, the progeny of a single MSCV-IL-6-marked cell transferred the myeloproliferative disease to two secondary, four tertiary, and two quaternary recipients (the clone endured for a total of 72 wk). These results, demonstrating considerable proliferative longevity of the IL-6-producing cells, support an in vivo role of IL-6 in the maintenance of hematopoietic precursors. Dysregulated IL-6 production also had significant systemic effects. The mice displayed increased mesangial cell proliferation in the kidney, frequent liver abnormalities, and marked alterations in plasma protein levels. Unlike previous studies where constitutive expression of exogenous IL-6 genes resulted in lymphoproliferative disorders characterized by massive plasmacytosis, minimal plasma cell expansion occurred in the MSCV-IL-6 mice during the observation period. Potential explanations for the differences in disease phenotypes observed in the present and previous studies are different cell types expressing the exogenous IL-6 genes, higher sustained circulating levels of IL-6 achieved using the MSCV-IL-6 retroviral delivery system, and/or the premature death (3-15 wk after transplantation) of the MSCV-IL-6 mice before the onset of plasmacytosis. This animal model should prove useful for further investigation of the function of IL-6 in normal and abnormal hematopoiesis and in inflammatory responses.

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