Dataset Information

Acetylated Histones in Apoptotic Microparticles Drive the Formation of Neutrophil Extracellular Traps in Active Lupus Nephritis.

ABSTRACT:

Objective

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the presence of autoantibodies against nuclear components. Lupus nephritis (LN) is the major cause of morbidity and mortality in patients with SLE. Central to the pathogenesis of SLE is the accumulation of cellular waste, especially apoptotic microparticles (MPs), which stimulates diverse immune reactions including the formation of neutrophil extracellular traps (NETs). In this study, we investigated the content of MPs from SLE patients with and without (active) LN, their capacity to stimulate NET release, and assessed the molecular mechanisms underlying MP-induced NETosis.

Methods

MPs from SLE patients with biopsy-proven active LN, remissive LN, without LN, and healthy controls were characterized by flow cytometry. Isolated neutrophils were exposed to MPs derived from either patient plasma or apoptotic human umbilical vein endothelial cells, and NET release was quantified by immunofluorescence imaging, spectrofluorometry or an in-house developed NET ELISA.

Results

MPs from SLE patients with active LN contain higher levels of acetylated chromatin compared to MPs from those with remissive LN, without LN, or healthy controls. MPs enriched in hyperacetylated chromatin are more potent in inducing NETosis when compared to MPs containing moderate acetylated chromatin. The release of NETs in response to MPs occurs rapidly in a concentration-dependent manner and proceeds independent from the formation of reactive oxygen species (ROS).

Conclusion

Our data suggest that MPs containing acetylated chromatin drive ROS-independent NET release in SLE patients with active LN, which may lead to the glomerular deposition of NETs and subsequent NET-driven LN.

SUBMITTER: Rother N

PROVIDER: S-EPMC5604071 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Publications

Acetylated Histones in Apoptotic Microparticles Drive the Formation of Neutrophil Extracellular Traps in Active Lupus Nephritis.

Rother Nils N Pieterse Elmar E Lubbers Jelle J Hilbrands Luuk L van der Vlag Johan J

Frontiers in immunology 20170914

<h4>Objective</h4>Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the presence of autoantibodies against nuclear components. Lupus nephritis (LN) is the major cause of morbidity and mortality in patients with SLE. Central to the pathogenesis of SLE is the accumulation of cellular waste, especially apoptotic microparticles (MPs), which stimulates diverse immune reactions including the formation of neutrophil extracellular traps (NETs). In this study, we investigated t ...[more]

PMID: 28959262

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Project description:BackgroundThe release of neutrophil extracellular traps (NETs), a mesh of DNA, histones and neutrophil proteases from neutrophils, was first demonstrated as a host defence against pathogens. Recently it became clear that NETs are also released in pathological conditions. NETs released in the blood can activate thrombosis and initiate a cascade of platelet responses. However, it is not well understood if these responses are mediated through direct or indirect interactions. We investigated whether cell-free NETs can induce aggregation of washed human platelets in vitro and the contribution of NET-derived extracellular DNA and histones to platelet activation response.MethodsIsolated human neutrophils were stimulated with PMA to produce robust and consistent NETs. Cell-free NETs were isolated and characterised by examining DNA-histone complexes and quantification of neutrophil elastase with ELISA. NETs were incubated with washed human platelets to assess several platelet activation responses. Using pharmacological inhibitors, we explored the role of different NET components, as well as main platelet receptors, and downstream signalling pathways involved in NET-induced platelet aggregation.ResultsCell-free NETs directly induced dose-dependent platelet aggregation, dense granule secretion and procoagulant phosphatidyl serine exposure on platelets. Surprisingly, we found that inhibition of NET-derived DNA and histones did not affect NET-induced platelet aggregation or activation. We further identified the molecular pathways involved in NET-activated platelets. The most potent single modulator of NET-induced platelet responses included NET-bound cathepsin G, platelet Syk kinase, and P2Y12 and ?IIb?3 receptors.ConclusionsIn vitro-generated NETs can directly induce marked aggregation of washed human platelets. Pre-treatment of NETs with DNase or heparin did not reduce NET-induced activation or aggregation of human washed platelets. We further identified the molecular pathways activated in platelets in response to NETs. Taken together, we conclude that targeting certain platelet activation pathways, rather than the NET scaffold, has a more profound reduction on NET-induced platelet aggregation.

Project description:Neutrophil infiltration and neutrophil extracellular traps (NET) in solid cancers are associated with poorer prognosis, but the mechanisms are incompletely understood. We hypothesized that NETs enhance mitochondrial function in tumor cells, providing extra energy for accelerated growth. Metastatic colorectal cancer tissue showed increased intratumoral NETs and supranormal preoperative serum MPO-DNA, a NET marker. Higher MPO-DNA correlated with shorter survival. In mice, subcutaneous tumor implants and hepatic metastases grew slowly in PAD4-KO mice, genetically incapable of NETosis. In parallel experiments, human cancer cell lines grew slower in nu/nu mice treated with DNAse, which disassembles NETs. PAD4-KO tumors manifested decreased proliferation, increased apoptosis, and increased evidence of oxidative stress. PAD4-KO tumors had decreased mitochondrial density, mitochondrial DNA, a lesser degree of ATP production, along with significantly decreased mitochondrial biogenesis proteins PGC1?, TFAM, and NRF-1. In vitro, cancer cells treated with NETs upregulated mitochondrial biogenesis-associated genes, increased mitochondrial density, increased ATP production, enhanced the percentage of cancer cells with reduced mitochondrial membrane potential, and increased the oxygen consumption rate. Furthermore, NETs increased cancer cells' expression of fission and fusion-associated proteins, DRP-1 and MFN-2, and mitophagy-linked proteins, PINK1 and Parkin. All of which were decreased in PAD4-KO tumors. Mechanistically, neutrophil elastase released from NETs activated TLR4 on cancer cells, leading to PGC1? upregulation, increased mitochondrial biogenesis, and accelerated growth. Taken together, NETs can directly alter the metabolic programming of cancer cells to increase tumor growth. NETs represent a promising therapeutic target to halt cancer progression. SIGNIFICANCE: Neutrophils through the release of NETs facilitate the growth of stressed cancer cells by altering their bioenergetics, the inhibition of which induces cell death.

Project description:Systemic lupus erythematosus (SLE) is associated with increased risk of adverse pregnancy outcomes, including pre-eclampsia, particularly in association with antiphospholipid antibody syndrome (APS). While significant placental abnormalities are expected in pre-eclampsia, less is known about how lupus activity and APS in pregnancy affect the placenta. We describe placental pathology from a population of lupus pregnancies, several of which were complicated by APS-related thromboses, in which pre-eclampsia and other complications developed. We performed standard histopathological placental review and quantified neutrophils and neutrophil extracellular traps (NETs) in the intervillous space, given the recognised association of NETs with lupus, APS and pre-eclampsia.Pre-eclampsia, SLE and control placentas were scored for histological features, and neutrophils were quantified on H&E and immunohistochemical staining for the granular protein myeloperoxidase. NETs were identified by extracellular myeloperoxidase staining in the setting of decondensed nuclei. Non-parametric analysis was used to evaluate differences in netting and intact neutrophils between groups, with Kruskal-Wallis testing for associations between histological findings and neutrophils.Placentas were evaluated from 35 pregnancies: 10 controls, 11 pre-eclampsia, 4 SLE+pre-eclampsia and 10 SLE, including one complicated by catastrophic APS and one complicated by hepatic and splenic vein thromboses during pregnancy. Intrauterine growth restriction and oligohydramnios were observed in lupus cases but not controls. Significantly more NETs were found infiltrating placental intervillous spaces in pre-eclampsia, SLE+pre-eclampsia and all 10 SLE non-pre-eclampsia cases. The ratio of NETs to total neutrophils was significantly increased in all case groups compared with controls. When present, NETs were associated with maternal vasculitis, laminar decidual necrosis, maternal-fetal interface haemorrhage and non-occlusive fetal thrombotic vasculopathy.In this pilot study of placental tissue from lupus pregnancies, outcomes were more complicated, particularly if associated with APS. Placental tissue revealed marked inflammatory and vascular changes that were essentially indistinguishable from placental tissue of pre-eclampsia pregnancies.

Dataset Information

Acetylated Histones in Apoptotic Microparticles Drive the Formation of Neutrophil Extracellular Traps in Active Lupus Nephritis.

Objective

Methods

Results

Conclusion

Publications

Acetylated Histones in Apoptotic Microparticles Drive the Formation of Neutrophil Extracellular Traps in Active Lupus Nephritis.

Similar Datasets

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