scholarly journals Candida albicans triggers NADPH oxidase-independent neutrophil extracellular traps through dectin-2

2019 ◽  
Vol 15 (11) ◽  
pp. e1008096 ◽  
Author(s):  
Sheng-Yang Wu ◽  
Chia-Lin Weng ◽  
Min-Jhen Jheng ◽  
Hung-Wei Kan ◽  
Sung-Tsang Hsieh ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Nadph Oxidase ◽  
Neutrophil Extracellular Traps ◽  
Extracellular Traps

scholarly journals Inactivation of α1-proteinase inhibitor by Candida albicans aspartic proteases favors the epithelial and endothelial cell colonization in the presence of neutrophil extracellular traps.

2015 ◽  
Vol 63 (1) ◽  
Author(s):  
Mariusz Gogol ◽  
Dominika Ostrowska ◽  
Kinga Klaga ◽  
Oliwia Bochenska ◽  
Natalia Wolak ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Proteinase Inhibitor ◽  
Proteolytic Enzymes ◽  
Fungal Infections ◽  
Critical Role ◽  
Neutrophil Extracellular Traps ◽  
Host Cells ◽  
Inhibition Mechanism ◽  
Aspartic Proteases ◽  
Extracellular Traps

Candida albicans, a causative agent of opportunistic fungal infections in immunocompromised patients, uses ten secreted aspartic proteases (SAPs) to deregulate the homeostasis of the host organism on many levels. One of these deregulation mechanisms involves a SAP-dependent disturbance of the control over proteolytic enzymes of the host by a system of dedicated proteinase inhibitors, with one important example being the neutrophil elastase and alpha1-proteinase inhibitor (A1PI). In this study, we found that soluble SAPs 1-4 and the cell membrane-anchored SAP9 efficiently cleaved A1PI, with the major cleavage points located at the C-terminal part of A1PI in a close vicinity to the reactive-site loop that plays a critical role in the inhibition mechanism. Elastase is released by neutrophils to the environment during fungal infection through two major processes, a degranulation or formation of neutrophil extracellular traps (NET). Both, free and NET-embedded elastase forms, were found to be controlled by A1PI. A local acidosis, resulting from the neutrophil activity at the infection sites, favors A1PI degradation by SAPs. The deregulation of NET-connected elastase affected a NET-dependent damage of epithelial and endothelial cells, resulting in the increased susceptibility of these host cells to candidal colonization. Moreover, the SAP-catalyzed cleavage of A1PI was found to decrease its binding affinity to a proinflammatory cytokine, interleukin-8. The findings presented here suggest a novel strategy used by C. albicans for the colonization of host tissues and overcoming the host defense.

β-Conglycinin induces the formation of neutrophil extracellular traps dependent on NADPH oxidase-derived ROS, PAD4, ERK1/2 and p38 signaling pathways in mice

2021 ◽  
Author(s):  
Xiao Liu ◽  
Yunhe Fu ◽  
Jingjing Wang ◽  
Di Wu ◽  
Shuangqiu Li ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Signaling Pathways ◽  
Neutrophil Extracellular Traps ◽  
Schematic Representation ◽  
Extracellular Traps

Schematic representation of β-conglycinin induced NETosis.

scholarly journals Extracellular Nucleic Acids Present in the Candida albicans Biofilm Trigger the Release of Neutrophil Extracellular Traps

2021 ◽  
Vol 11 ◽  
Author(s):  
Magdalena Smolarz ◽  
Marcin Zawrotniak ◽  
Dorota Satala ◽  
Maria Rapala-Kozik
Keyword(s):  
Candida Albicans ◽  
Nucleic Acids ◽  
Fungal Infection ◽  
Fungal Infections ◽  
Neutrophil Extracellular Traps ◽  
Human Neutrophils ◽  
High Concentration ◽  
First Line ◽  
Extracellular Traps ◽  
Biofilm Structure

Neutrophils, the first line of the host’s defense, use a variety of antimicrobial mechanisms to fight invading pathogens. One of the most crucial is the production of neutrophil extracellular traps (NETs) in the process called NETosis. The unique structure of NETs effectively inhibits the spread of pathogens and ensures their exposure to a high concentration of NET-embedded antimicrobial compounds. NETosis strategy is often used by the host to defend against fungal infection caused by Candida albicans. In immunocompromised patients, this microorganism is responsible for developing systemic fungal infections (candidiasis). This is correlated with the use of a vast array of virulence factors, leading to the acquisition of specific resistance to host defense factors and available drug therapies. One of the most important features favoring the development of drug resistance is a C. albicans ability to form biofilms that protect fungal cells mainly through the production of an extracellular matrix (ECM). Among the main ECM-building macromolecules extracellular nucleic acids have been identified and their role is probably associated with the stbilization of the biofilm structure. The complex interactions of immune cells with the thick ECM layer, comprising the first line of contact between these cells and the biofilm structure, are still poorly understood. Therefore, the current studies aimed to assess the release of extracellular nucleic acids by C. albicans strains at different stages of biofilm formation, and to determine the role of these molecules in triggering the NETosis. We showed for the first time that fungal nucleic acids, purified directly from mature C. albicans biofilm structure or obtained from the whole fungal cells, have the potential to induce NET release in vitro. In this study, we considered the involvement of TLR8 and TLR9 in NETosis activation. We showed that DNA and RNA molecules initiated the production of reactive oxygen species (ROS) by activation of the NADPH oxidase complex, essential for ROS-dependent NETosis. Furthermore, analysis of the cell migration showed that the nucleic acids located in the extracellular space surrounding the biofilm may be also effective chemotactic factors, driving the dynamic migration of human neutrophils to the site of ongoing fungal infection.

The Role of Rac and Pak in Neutrophil Histone Hypercitrullination and Neutrophil Extracellular Traps Formation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 462-462 ◽  
Author(s):  
Mathilde Gavillet ◽  
Kimberly Martinod ◽  
Denisa D. Wagner ◽  
David A. Williams
Keyword(s):  
Flow Cytometry ◽  
Nadph Oxidase ◽  
Autoimmune Diseases ◽  
Actin Polymerization ◽  
Genetic Model ◽  
Neutrophil Extracellular Traps ◽  
Extracellular Traps ◽  
Downstream Effector ◽  
Group A

Abstract Under specific activating conditions, polymorphonuclear neutrophils (PMNs) release neutrophil extracellular traps (NETs) composed of decondensed chromatin lined with microbicidal protein such as neutrophil elastase and myeloperoxidase. NETs contribute to innate immunity but can also foster autoimmune diseases and thrombus formation. NET formation (NETosis) requires reactive oxygen species (ROS) production by NADPH oxidase and histone hypercitrullination by peptidylarginine deiminase 4 (PAD4), allowing for chromatin decondensation. Rac GTPases are expressed in three isoforms: Rac1 is ubiquitously expressed and plays a role in PMN migration and oxidase function; Rac2 is hematopoietic-specific and the major isoform in PMNs and Rac3 is mostly neuronal. Rac1 and Rac2 regulate the cytoskeleton in PMNs, controlling actin polymerization, cell shape, adhesion and migration and are essential components of the NADPH oxidase complex. The present study aimed to explore the role of the Rac pathway on NETosis in PMNs, including the upstream guanosine exchange factor (GEF) activator, Vav, and a downstream effector of Rac, p21 activated kinase, Pak. We developed a flow cytometry-based quantification of H3 hypercitrullination (H3Cit). In response to phorbol myristate ester (PMA) stimulation, H3Cit is increased to 136% of basal in WT cells, compared with 103% in Rac2-/- (P<0.01) (Table). H3Cit levels observed by flow were confirmed in a NET formation assay. Rac2-/- PMNs formed significantly fewer NETs both spontaneously and after PMA stimulation (WT unstimulated 2.79%, Rac2-/- unstimulated 0.72%, WT+PMA 10.84%, Rac2-/-+PMA 1.39%, P< 0.05 for all pair comparisons). Furthermore, Rac2-/- mice demonstrated a trend towards reduced frequency of provoked thrombosis in an in vivo vena cava stenosis model (WT 78% and Rac2-/- 56% of mice with thrombus). Deletion of floxed Rac1 sequences in a Rac2-/- background in vivoallows generation and purification of PMNs lacking both Rac isoforms. Rac1Δ/Δ,Rac2-/- PMNs, which are defective in actin polymerization, had reduced basal H3Cit and a nearly complete lack of PMA-induced increase in H3Cit (136% vs 69%, WT vs Rac1Δ/Δ,Rac2-/-, P<0.01) (Table). Null knockouts of the GEFs Vav1 (hematopoietic-specific), Vav2, Vav3 or both Vav1 and 3 did not impair H3Cit response to PMA (Table). We next studied downstream effectors of Rac. Group A Paks include Pak1, 2 and 3 isoforms. Pretreatment of wild-type PMNs with either PF3758309 or IPA-3, two group A Pak inhibitors with distinct mechanisms of action, led to reduced H3Cit after PMA stimulation (induction reduced from 36% to 11% for both PF3758309 and IPA-3 treated (Table). To validate this in a genetic model, we studied Pak2Δ/Δ PMNs, since we have recently demonstrated the dependence of hematopoietic stem cell migration on Pak2. Pak2Δ/Δ demonstrated a reduced basal level of H3Cit and a significantly reduced PMA-induced increase in H3Cit (136% vs 94%, WT vs Pak2Δ/Δ, P<0.05, Table). In summary, we describe a flow-based assay that quantitates the early processes of NET formation and validated that this assay reliably predicts agonist-induced NET formation in a genetic model. The results establish that both Rac1 and Rac2, and the downstream effector Pak2, regulate histone H3 hypercitrullination and NET formation in PMNs, while suggesting that Vav does not activate the Rac pathway in PMA-induced NET formation. These data further delineate the role of the Rac pathway in NETosis, linking cytoskeleton and oxidase functions. Furthermore, these data indicate Pak could represent a therapeutic target for a wide array of pathological processes related to NETosis such as thrombosis and numerous autoimmune diseases. Table Intensity of H3Cit staining as determined by flow cytometry-based assay. Basal H3Cit level PMA-induced H3Cit PMA-induced change WT PMN 100±2% 136±5% 36% Rac2-/- 82±9% ns 103±15.3% ** 21% Rac1Δ/Δ, Rac2-/- 64±10% *** 69±10% ** 5% Vav1-/- 86±9% ns 145±15% ns 59% Vav2-/- 91±4% ns 134±18% ns 43% Vav3-/- 153±30% * 171±28% ns 18% Vav1,3-/- 125±20% ns 144±24% ns 19% WT+ PF 5nM 76±17% ns 87±8% * 11% WT+ IPA 5µM 100±4% ns 111±10% ns 11% Pak2Δ/Δ 75±7% * 94±11% * 19% Results are expressed as mean±SEM % of the untreated WT control of each experiment. Results are from ≥3 independent experiments. * P<0.05, **P<0.01. P<0.001, ns non-significant, by two-tailed t-test. Disclosures No relevant conflicts of interest to declare.

Killing by neutrophil extracellular traps: fact or folklore?

Blood ◽  
2012 ◽  
Vol 119 (5) ◽  
pp. 1214-1216 ◽  
Author(s):  
Renzo Menegazzi ◽  
Eva Decleva ◽  
Pietro Dri
Keyword(s):  
Staphylococcus Aureus ◽  
Candida Albicans ◽  
Neutrophil Extracellular Traps ◽  
Dna Structures ◽  
Extracellular Traps ◽  
Cell Medium

AbstractNeutrophil extracellular traps (NETs) are DNA structures released by dying neutrophils and claimed to constitute a new microbicidal mechanism. Killing by NET-forming cells is ascribed to these structures because it is prevented by preincubation with DNase, which has been shown to dismantle NETs, before addition of the target microorganisms. Curiously, the possibility that the microorganisms ensnared in NETs are alive has not been considered. Using Staphylococcus aureus and Candida albicans blastospores, we demonstrate that the microorganisms captured by NETs and thought to be killed are alive because they are released and recovered in cell medium by incubation with DNase. It is concluded that NETs entrap but do not kill microbes.

scholarly journals Novel cell death program leads to neutrophil extracellular traps

2007 ◽  
Vol 176 (2) ◽  
pp. 231-241 ◽  
Author(s):  
Tobias A. Fuchs ◽  
Ulrike Abed ◽  
Christian Goosmann ◽  
Robert Hurwitz ◽  
Ilka Schulze ◽  
...  
Keyword(s):  
Cell Death ◽  
Nadph Oxidase ◽  
Neutrophil Extracellular Traps ◽  
Death Process ◽  
Extracellular Traps ◽  
Cell Death Pathway ◽  
Antimicrobial Function ◽  
Cell Death Program ◽  
Cell Death Process ◽  
Extracellular Structures

Neutrophil extracellular traps (NETs) are extracellular structures composed of chromatin and granule proteins that bind and kill microorganisms. We show that upon stimulation, the nuclei of neutrophils lose their shape, and the eu- and heterochromatin homogenize. Later, the nuclear envelope and the granule membranes disintegrate, allowing the mixing of NET components. Finally, the NETs are released as the cell membrane breaks. This cell death process is distinct from apoptosis and necrosis and depends on the generation of reactive oxygen species (ROS) by NADPH oxidase. Patients with chronic granulomatous disease carry mutations in NADPH oxidase and cannot activate this cell-death pathway or make NETs. This novel ROS-dependent death allows neutrophils to fulfill their antimicrobial function, even beyond their lifespan.

scholarly journals Restoration of NET formation by gene therapy in CGD controls aspergillosis

Blood ◽  
2009 ◽  
Vol 114 (13) ◽  
pp. 2619-2622 ◽  
Author(s):  
Matteo Bianchi ◽  
Abdul Hakkim ◽  
Volker Brinkmann ◽  
Ulrich Siler ◽  
Reinhard A. Seger ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Antimicrobial Activity ◽  
Nadph Oxidase ◽  
Invasive Pulmonary Aspergillosis ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Neutrophil Extracellular Traps ◽  
Pulmonary Aspergillosis ◽  
Extracellular Traps ◽  
Healthy Persons

AbstractChronic granulomatous disease (CGD) patients have impaired nicotinamide adenine dinucleotide phosphate (NADPH) oxidase function, resulting in poor antimicrobial activity of neutrophils, including the inability to generate neutrophil extracellular traps (NETs). Invasive aspergillosis is the leading cause of death in patients with CGD; it is unclear how neutrophils control Aspergillus species in healthy persons. The aim of this study was to determine whether gene therapy restores NET formation in CGD by complementation of NADPH oxidase function, and whether NETs have antimicrobial activity against Aspergillus nidulans. Here we show that reconstitution of NET formation by gene therapy in a patient with CGD restores neutrophil elimination of A nidulans conidia and hyphae and is associated with rapid cure of preexisting therapy refractory invasive pulmonary aspergillosis, underlining the role of functional NADPH oxidase in NET formation and antifungal activity.

scholarly journals Neutrophil extracellular traps capture and kill Candida albicans yeast and hyphal forms

2006 ◽  
Vol 8 (4) ◽  
pp. 668-676 ◽  
Author(s):  
Constantin F. Urban ◽  
Ulrike Reichard ◽  
Volker Brinkmann ◽  
Arturo Zychlinsky
Keyword(s):  
Candida Albicans ◽  
Neutrophil Extracellular Traps ◽  
Extracellular Traps
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