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Dysregulated neutrophil responses and neutrophil extracellular trap formation and degradation in PAPA syndrome.

ABSTRACT: OBJECTIVES:Pyogenic arthritis, pyoderma gangrenosum and acne (PAPA) syndrome is characterised by flares of sterile arthritis with neutrophil infiltrate and the overproduction of interleukin (IL)-1?. The purpose of this study was to elucidate the potential role of neutrophil subsets and neutrophil extracellular traps (NET) in the pathogenesis of PAPA. METHODS:Neutrophils and low-density granulocytes (LDG) were quantified by flow cytometry. Circulating NETs were measured by ELISA and PAPA serum was tested for the ability to degrade NETs. The capacity of NETs from PAPA neutrophils to activate macrophages was assessed. Skin biopsies were analysed for NETs and neutrophil gene signatures. RESULTS:Circulating LDGs are elevated in PAPA subjects. PAPA neutrophils and LDGs display enhanced NET formation compared with control neutrophils. PAPA sera exhibit impaired NET degradation and this is corrected with exogenous DNase1. Recombinant human IL-1? induces NET formation in PAPA neutrophils but not healthy control neutrophils. NET formation in healthy control neutrophils is induced by PAPA serum and this effect is inhibited by the IL-1 receptor antagonist, anakinra. NETs from PAPA neutrophils and LDGs stimulate IL-6 release in healthy control macrophages. NETs are detected in skin biopsies of patients with PAPA syndrome in association with increased tissue IL-1?, IL-8 and IL-17. Furthermore, LDG gene signatures are detected in PAPA skin. CONCLUSIONS:PAPA syndrome is characterised by an imbalance of NET formation and degradation that may enhance the half-life of these structures in vivo, promoting inflammation. Anakinra ameliorates NET formation in PAPA and this finding supports a role for IL-1 signalling in exacerbated neutrophil responses in this disease. The study also highlights other inflammatory pathways potentially pathogenic in PAPA, including IL-17 and IL-6, and these results may help guide new therapeutic approaches in this severe and often treatment-refractory condition.

SUBMITTER: Mistry P

PROVIDER: S-EPMC6728908 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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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 Streptomyces conglobatus) 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.

Project description:Schnitzler's syndrome is a rare autoinflammatory disorder characterized by interleukin-1ß-mediated and neutrophil-dominated inflammation. Neutrophil extracellular traps (NETs) are web-like structures of decondensed chromatin, histones, and antimicrobial peptides released by neutrophils. NETs were initially described in the context of pathogen defense but are also involved in autoimmune-mediated skin diseases. Here, we assessed the role of neutrophil extracellular trap formation (NETosis) in Schnitzler's syndrome. Immunofluorescence co-staining of myeloperoxidase and subnucleosomal complex was performed on lesional skin samples from patients with Schnitzler's syndrome, other neutrophilic dermatoses (cryopyrin-associated periodic syndrome, Sweet syndrome, and pyoderma gangrenosum), urticarial vasculitis and chronic spontaneous urticaria as well as healthy control skin. Blood neutrophils from patients with Schnitzler's syndrome and controls were isolated, and NETosis was induced by phorbol 12-myristate 13-acetate (PMA). Also, NETosis of control neutrophils induced by symptomatic Schnitzler's syndrome sera, cytokines and sub-threshold PMA doses was studied. Immunofluorescence co-staining revealed widespread and substantial NET formation in lesional skin of Schnitzler's syndrome patients but absence of NETs in chronic spontaneous urticaria and control skin. Neutrophils undergoing NETosis were observed in the skin of other neutrophilic diseases too. Correspondingly, blood neutrophils from Schnitzler's syndrome patients showed significantly elevated NETosis rates compared to control neutrophils following stimulation with PMA. Increased NETosis correlated well with high levels of C-reactive protein (CRP). SchS patients with the lowest NETosis rates had persistent joint and bone pain despite IL-1 blockade. Stimulation of control neutrophils and sub-threshold PMA with sera of symptomatic Schnitzler's syndrome patients disclosed enhanced NETosis as compared to control sera. Our results suggest that the induction of NET formation by neutrophils contributes to skin and systemic inflammation and may support the resolution of local inflammation in Schnitzler's syndrome.

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Dysregulated neutrophil responses and neutrophil extracellular trap formation and degradation in PAPA syndrome.

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