Kayla M. Fantone
Naveen Gokanapudi
Balázs Rada- Department of Infectious Diseases, College of Veterinary Medicine, The University of Georgia, Athens, GA, United States
Cystic fibrosis (CF) lung disease manifests through abnormally thick mucus, persistent bacterial infections and a dysregulated innate immune system that involves significant neutrophilic inflammation. Neutrophils, immune cells essential to fight infections, accumulate in large numbers in CF airways and release neutrophil extracellular traps (NETs) into the airway lumen that deliver extracellular DNA, granule content and cytokines including IL-1β. Interleukin-1β, a powerful, proinflammatory cytokine, represents another, significant component of the innate immune system that is dysregulated in CF. Both defense mechanisms become problematic as NETs and IL-1β are present at elevated levels in CF airways, potentially creating a destructive cycle that exacerbates lung damage rather than protects against infections. Therefore, understanding the interplay between IL-1β and NETs is crucial for addressing CF lung disease progression. This review examines the general mechanisms of IL-1β release and NET formation, with particular focus on their role in CF lung disease, and proposes that a self-perpetuating, positive feedback loop between these two innate immune processes represents a major driving force in disease progression. This understanding suggests potential therapeutic targets for interrupting the cycle of inflammation and tissue damage in CF airways.
1 Introduction
1.1 Cystic fibrosis
Cystic fibrosis (CF) is a genetic disease affecting an estimated 160,000 people worldwide (1, 2). It is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene encoded on chromosome 7 (7q31.2) in humans. CF is an autosomal recessive disease that requires two copies of mutated CFTR genes to manifest (3). Carriers with a single copy of the mutant CFTR gene will pass the mutation to their children but demonstrate no symptoms. An effected individual carrying two mutated copies of CFTR can develop CF. One in 30 people are carriers in the USA. Around 2,000 different CFTR gene mutations have been identified that are associated with CF (3). The CFTR protein functions as a chloride channel regulating the Na+/Cl- balance across mucosal surfaces, especially in the respiratory and digestive tracts. Normal CFTR function facilitates the regular maintenance of mucus levels which is disrupted in CF causing mucus buildup (4–6). CF affects multiple organs that have a secretory function including the digestive and respiratory systems. The absence of a fully functional CFTR anion channel in CF impacts multiple physiological mechanisms indicating its complex and intricate function. Mutant CFTR can lead to -among others- abnormal mucus production in the lung and gastrointestinal system, pancreatic problems, reduced glucose metabolism and impacted circadian rhythm. One of the most impacted organs in CF is the lung and lung disease reduces life expectancy for people with CF (PwCF) (7).
1.2 CF lung disease: inflammation and infections
CF airway disease is characterized by chronic inflammation and persistent polymicrobial infections. The defect of the CFTR gene alters anion transport across airway epithelial cells which causes dehydration of the airway surface liquid and excessive mucin secretion (7, 8). The excess fluid and mucus in the lungs contribute to airway obstruction and microbial infections leading to chronic inflammation (7, 9). The most abundant inflammatory cells recruited to the CF airway are neutrophils whose main function in the body is to fight off infections (10). The CF lung hosts polymicrobial infections (11, 12). The major pathogens infecting the CF lung are bacteria such as Staphylococcus aureus, Pseudomonas aeruginosa, Haemophilus influenzae, Stenotrophomonas maltophila, Achromobacter species and Burkholderia cepacia (9). S. aureus and P. aeruginosa represent the two most prevalent respiratory pathogens in PwCF. P. aeruginosa has been observed for a long time as the dominant airway pathogen, infecting 60-75% adult PwCF, but in the past two decades S. aureus has become the most common in CF (9, 13, 14). The prevalence of non-tuberculous mycobacterial (NTM) infections has also been rising among PwCF, with Mycobacterium avium and M. abscessus being the most common ones (11, 12, 15). In addition, PwCF can also be infected with fungal pathogens, such as Aspergillus fumigatus (16–18).
Neutrophils are the primary innate cells that respond to S. aureus and P. aeruginosa challenge in humans (19–22). In CF, neutrophils are recruited to the airways in high numbers but fail to clear a select group of pathogens listed above (23–25). Neutrophils represent a major cause of CF lung damage by releasing their granular and nuclear content driving chronic inflammation (10, 23–25). Proteolytic stress carried out by neutrophil-derived proteases such as neutrophil elastase (NE) and oxidative stress mediated by oxidants produced by neutrophils represent two major mechanisms by which neutrophils directly contribute to lung damage in CF (23, 26). In addition to causing direct tissue damage, neutrophil components released into the CF airway lumen also stimulate the release of cytokines including IL-1β from epithelial and immune cells which attracts additional neutrophils, thereby fueling a feed-forward inflammatory process. Two important components of the innate immune system, and the interplay between them, will be discussed in this article that are hypothesized to be part of a self-perpetuating, proinflammatory process contributing to CF airway disease progression: neutrophil extracellular traps (NETs) and IL-1β (Figure 1).
Figure 1. A schematic illustration indicating the hypothesis of this review article that NETs and IL-1β mutually enhance each other and assemble into a positive, innate immune feedback loop that contributes to the chronic inflammation and tissue damage observed in the lungs of PwCF.
2 NETs
2.1 NETs: general mechanisms
Neutrophils are abundant innate immune cells equipped with diverse antimicrobial molecules, making them crucial to combat microbial pathogens (27). Neutrophils combat bacteria through 1) intracellular killing by phagocytosis or 2) extracellular killing by forming neutrophil extracellular traps (NETs) or releasing granules by degranulation (27–29). Neutrophils identify pathogens via surface receptors such as pattern recognition receptors, Fc receptors and complement receptors, and engulf them through phagocytosis (27). Once the pathogen is in the phagosome, granules of the neutrophil fuse with the phagosome, and proteases, antimicrobial molecules and oxidants are released into the phagolysosome to create a toxic environment for the engulfed and isolated pathogen (27). Neutrophils also kill pathogens by another mechanism called NET formation (29). During NET formation, neutrophils release their decondensed chromatin decorated with granular content including histones and granule proteins intro the extracellular environment (30). The nucleus of neutrophils loses its lobed morphology in the course of NET formation and chromatin decondenses which is potentiated by enzymes such as NE and myeloperoxidase (MPO) (30). NET formation can be induced by many agonists such as bacterial pathogens and can require the production of reactive oxygen species (ROS) or the activation of different kinases (31). Activation of kinases causes chromatin decondensation leading to histone modifications catalyzed by peptidyl deaminase 4 (PAD4) that converts arginine residues to citrulline (29, 31). In the final stages of NET formation, the nuclear membrane disintegrates, the cell membrane ruptures leading to the release of granular proteins such as NE, citrullinated histones, MPO, defensins and cathelicidins (30, 31).
In addition to the originally described route of NET formation that requires the cell to die (suicidal NETs), another form of NET release has also been documented. In certain cases, neutrophils do not become lysed but only release small amounts of NETs, much faster than suicidal NET formation, remain alive for hours and perform tasks such as migration, ROS production and phagocytosis. This alternative NET pathway has been termed as “viable” NET release and was proposed to mainly involve mitochondrial, not nuclear, DNA (32, 33).
Neutrophils release NETs in response to various stimuli. Different mechanisms of NET formation have been described which include NADPH oxidase (Nox)-dependent and Nox-independent pathways (31). Nox-dependent NET formation is induced by lipopolysaccharide (LPS) found in gram-negative bacteria that binds to TLR4 on the surface of neutrophils and induces ROS production by the NADPH oxidase (Nox2) (31, 34). During Nox-dependent NET release, ROS generated by Nox2 induces the disintegration of granule and nuclear membranes, allowing for NE and MPO to interact with the nucleus to cleave histones and cause chromatin decondesation (30, 31, 34). On the other hand, calcium ionophores can also induce NET formation in a Nox-independent manner (35). PAD4 is abundantly present in the cytosol of neutrophils, can bind to calcium and translocate into the nucleus (31, 36). The PAD4 enzyme deiminates histone arginine residues into neutral citrulline amino acids causing chromatin decondensation (24, 31). There is further evidence to suggest that calcium-activated potassium channel of small conductance (SK3 channel) and mitochondrial ROS (mROS) are also required to induce Nox-independent NET formation (37). Calcium ionophores stimulate mROS through influx via the SK3 channel to induce Nox-independent NET extrusion (37). Another important step underlying NET formation is transcriptional firing at promoter regions that help to mediate DNA decondensation (31, 38). It has been found that transcription of Erk-, Akt-, p38- and cSrc-regulated genes are the primary drivers of Nox-dependent NET formation, whereas transcription of Akt-, p38-, cSrc-, PyK2- and Jnk-regulated genes drive Nox-independent production of NETs (31, 38). Lastly, histone modifications are also important components for NET formation in neutrophils. Histone acetylation plays a relevant role in NET formation and causes neutralization of the positive charges on the histones that promotes chromatin decondensation (31, 39).
Dysregulation of NET formation and/or NET clearance have been associated with the severity of several lung diseases such as CF, acute respiratory distress syndrome, acute lung injury and airway infections including COVID-19 (40, 41). COVID-19 caused by the respiratory virus SARS-COV-2 is characterized by severe lung disease that requires hospitalization in some patients (42). In the NETCOV2 study, the number of days with severe hypoxemia in the intensive care unit patients with SARS-CoV-2-related pneumonia correlated negatively with blood NET levels measured at 1-day post-admission (42). The absence of decrease of the blood NET levels between day-1 and day-3 discriminated patients who died within days (42). This and other studies strongly suggest a pathologic role of NETs in severe SARS-CoV-2-associated pneumonia (43, 44). A pathologic role of exaggerated neutrophil recruitment and NET release in lung function decline, and a prognostic role of the blood neutrophil:lymphocyte ratio for future disease severity have been indicated in severe COVID-19 (45–47). NETs have been proposed as the delivery platform to bring the tissue-damaging intracellular cargo of neutrophils to the airway lumen including reactive oxidants leading to lung tissue damage in severe COVID-19 (48). Overall, while NETs play an important role in innate host defense by trapping extracellular pathogens, they have been proposed to contribute to the pathologies of several illnesses including CF lung disease.
2.2 NETs in CF
In CF, chronic inflammation in the airways is characterized by a massive neutrophil influx with subsequent release of NETs (34) (Figure 2). All these changes lead to persistent lung injury through the accumulation of dysfunctional neutrophils releasing their DNA and cytotoxic antimicrobial content such as MPO, lysozyme, lactoferrin, NE, defensins, gelatinase, cathelicidins and cathepsins. The CF sputum and bronchioalveolar lavage fluid contain high levels of neutrophil granular components whose concentrations correlate with the severity of lung disease in CF (10, 49–51). The massive influx of neutrophils and the release of neutrophil DNA in the bronchioles in the CF lung aggravate mucus viscosity providing a suitable environment for the colonization of infectious bacteria (10, 49–51). The excessive mucus production in CF airways not only contributes to the establishment and persistence of bacterial infections but also to the clogging of the airways resulting in tissue damage and subsequent disease pathology. S. aureus and P. aeruginosa are strong NET inducers (52–58) (Figure 2). DNA released from neutrophils via NETs has been proposed to promote bacterial colonization and biofilm formation in the CF airways (59, 60). CF neutrophils were shown to have more robust, spontaneous NET formation than non-CF control cells due to their delayed apoptotic response and longer survival (61–63). DNA in the CF sputum demonstrates characteristics of NETs and NET levels correlate with lung disease in CF (50, 64).
Figure 2. Proposed mechanisms connecting NETs, IL-1β and other components of the airways that support chronic inflammation in CF. CFTR deficiency in airway epithelial cells leads to excessive release of mucus and cytokines including IL-1β. Produced cytokines recruit innate immune cells, macrophages and neutrophils. Mucus overproduction and impaired mucociliary clearance attract bacterial pathogens that stimulate the recruited immune cells and the airway epithelium to generate cytokines and to recruit additional leukocytes, leading to a self-perpetuating inflammatory process. In addition, macrophages produce IL-1β and undergo pyroptosis while neutrophils release NETs, which overall deliver the intracellular content of these innate cells to the airway lumen. NETs activate macrophages yielding IL-1β release and pyroptosis, while microbial and host molecules, likely including IL-1β, induce further NET extrusion. The complex network of the proposed innate immune mechanisms indicated by the arrows fuel chronic inflammation and lung disease in CF in a positive feed-forward fashion. Prepared with BioRender.
PwCF develop autoantibodies that target neutrophil or NET components, such as bactericidal permeability-increasing protein, carbamylated proteins, PAD4 and DNA (65–69). Blood levels of anti-PAD4 antibodies and anti-double-stranded DNA IgA autoantibodies are elevated in CF and associate with worsened lung function (67, 68). The reported autoantibody pattern in CF is different from that described in well-established and characterized autoimmune diseases, such as rheumatoid arthritis or systemic lupus erythematosus (SLE) (65, 67). Not only IgG but also IgA autoantibodies can be strong stimulants of neutrophils (70, 71). These autoantibodies could represent additional stimuli for neutrophils to release NETs and to fuel NET-mediated inflammation in CF (65, 67).
Overall, NETs have been documented in the CF airways in abundance, correlate with lung function decline, are not capable of clearing bacterial infections, and likely provide a platform for delivering the proinflammatory and tissue-damaging neutrophil intracellular content to the airway lumen to fuel chronic inflammation (Figures 1, 2).
3 Interleukin-1β
3.1 IL-1β in CF
Progressive lung disease in CF results in excess mucus in the airways and trapping of bacteria, leading to infections, inflammation, and overall respiratory failure. Release of NETs and microbial molecules originating from bacterial infections activate a multimeric protein complex known as the inflammasome, which is present in many innate immune cell types including neutrophils and macrophages (72). Macrophages infiltrate the CF airways to eliminate neutrophil debris and help fight microbial pathogens, contributing to chronic inflammation (Figure 2).
IL-1β serves as a key inflammatory mediator in the airway, and its secretion involves inflammasome-mediated processing (72). In PwCF, IL-1β was detected in the bronchoalveolar lavage fluid even without infection, its airway levels increased in the presence of bacterial infection and correlated with the neutrophil count and NE activity in CF airways (73). These data and others suggest that IL-1β plays an important role in recruiting and/or stimulating neutrophils in CF. Indeed, IL-1β was shown to recruit neutrophils to the airways in sterile lung injury and airway infection models in mice (74). It is unclear whether PwCF have increased IL-1β production in the absence of infection due to an intrinsic increase in NF-κB activity or because of the loss of CFTR function (75). However, without a functional CFTR, ENaC channels are dysregulated leading to increased intracellular Na+ levels and increased efflux of K+ (76). Upregulation of K+ efflux in the CF airways is an extracellular stimulus for NLRP3 inflammasome activation, leading to subsequent release of IL-1β and IL-18. The combination of K+ efflux and microbial stimuli such as pathogen-associated molecular patterns (PAMP), lead to excessive NLRP3 inflammasome activation and downstream proinflammatory cytokine release (Figure 3). P. aeruginosa and S. aureus lung infections increase airway levels of IL-1β in mice (77, 78). Levels of IL-1β in the lung are also higher in CF mice infected with either P. aeruginosa or Aspergillus fumigatus, a fungal CF respiratory pathogen (79). Anakinra, a recombinant non-glycosylated homolog of the human IL-1 receptor antagonist (IL-1Ra), protects CF mice from infections and NLRP3-mediated inflammation following either P. aeruginosa or A. fumigatus infection (79).
Figure 3. Proposed molecular mechanisms driving the generation of bioactive IL-1β in innate immune cells in CF airways. Innate immune cells in the CF lung are stimulated by microbial molecules derived from respiratory pathogens via patter recognition receptors (PRRs) or by host molecules originating from neutrophils that had undergone NET formation or pyroptosed macrophages via receptors recognizing danger-associated molecular patterns (DAMP-R). DAMP-Rs and PRRs activate the transcription factor NF-κB that up-regulates the expression of the genes encoding inflammasome components, pro-IL-1β and pro-caspase-1. Ion channels and transporters localized to the plasma membrane can influence signaling pathways as indicated. Inflammasomes (NLRP3, NLRC4) cleave pro-caspase-1 into active caspase-1. Caspase-1 cleaves pro-IL-1β into bioactive IL-1β and gasdermin-D into two fragments, the secreted component, gasdermin-C (GSDMD-C) and the pore-forming unit, gasdermin-N (GSDMD-N). IL-1β leaves the cell via the gasdermin pore. When innate immune cells die by pyroptosis, necrosis or NET formation, pro-IL-1β has been proposed be released into the airway lumen. Extracellular pro-IL-1β can be cleaved by neutrophil proteases (neutrophil elastase, NE; proteinase 3, PR3) released from neutrophils via the indicated, different mechanisms or by the lasB protease derived from P. aeruginosa. The red cross indicates CFTR deficiency. Prepared with BioRender.
One study found that blocking ENaC channels in human bronchial epithelial cells with a naturally occurring 18-residue peptide restores Na+ and K+ levels, reduces NLRP3-mediated IL-1β and IL-18 release and attenuates inflammation (76). Sterile inflammation in PwCF displays IL-1β-induced expression of secreted airway mucins in primary human bronchial epithelial cells, which can be prevented by IL-1Ra treatment (80–82). Evidence suggests an important role of the IL-1 signaling pathway early in CF lung disease by contributing to neutrophilic airway inflammation and mucus hypersecretion, in the absence of any detectable infection (83). IL-1β may be the dominant cytokine in the inflammatory environment of the CF lung, especially after the onset of bacterial infections (84) (Figure 2). The IL-1R is predominantly expressed on cells that also produce IL-1β: innate immune cells including neutrophils and macrophages, and airway epithelial cells; suggesting a positive feedback loop for production and release of IL-1β in the CF airways (84). Airway epithelial cells undergoing necrosis release IL-1β through activation of the IL-1R-MyD88 signaling pathway that is known to induce neutrophilic inflammation in mice with CF-like lung disease (85). The inflammasome has been demonstrated to be important in the pathogenesis of lung fibrosis and studies suggest its dependence on the IL-1R pathway (85). PwCF may also have a predisposition to dysregulated IL-1β signaling due to genetic polymorphisms (86). IL-1β polymorphisms are present among PwCF and specific single nucleotide polymorphism (SNP) in the IL-1β gene have been associated with lung disease in CF. IL-1β levels are enhanced in PwCF harboring these SNPs compared to patients without them, and IL-1β-mediated inflammation is further increased in the presence of respiratory infection (86, 87).
On a per cell basis, macrophages represent the major source of IL-1β, while neutrophils and airway epithelial cells also release this cytokine at lower levels per cell (88–90). Considering their larger cell number in the CF airways, neutrophils and airway epithelial cells also represent significant sources of IL-1β (Figure 2). P. aeruginosa stimulates IL-1β release in vitro in macrophages (91, 92), neutrophils (93) and airway epithelial cells (90, 94). S. aureus also induces IL-1β synthesis in innate immune cells (95–97) in vitro.
CF airway epithelial cells demonstrated significantly higher levels of released IL-8 and IL-6 in response to IL-1β stimulation, in the absence of any infection or microbial stimuli in vitro (98–100). This proinflammatory phenotype of CF airway epithelial cells is likely due to a combination of different factors including 1) endoplasmic reticulum stress that develops because of the intracellular accumulation of misfolded CFTR proteins in case of certain CFTR mutations, impaired regulation of 2) ion transport and 3) intracellular signaling pathways (101, 102). Macrophages isolated from PwCF have also been reported to release larger amounts of cytokines including IL-1β when stimulated with LPS in vitro (103). These data indicate that endogenous CFTR deficiency can push airway epithelial cells and macrophages towards a “proinflammatory” phenotype that manifests -among other things- in heightened proinflammatory cytokine release.
Overall, several mechanisms are likely responsible for the increased IL-1β levels observed in the CF lung.
3.2 Inflammasomes
The inflammasome provides a protective innate immune mechanism that activates caspase-1 and leads to the proteolytic cleavage of pro-IL-1β and pro-IL-18 to induce inflammation in response to PAMPs and danger-associated molecular patterns (DAMPs) (104) (Figure 2). Toll-like receptors (TLRs) are the key inducers of the transcription of proinflammatory cytokines, IL-1β and IL-18, through NF-κB stimulation (Figure 3). Nod-like receptor (NLR) family protein members contain a nucleotide binding domain (NBD), a variable N-terminal domain [either pyrin domain (PYD) or the caspase recruitment domain (CARD)], and a C-terminal leucin-rich repeat (LRR) region (105, 106). The assembly of the inflammasome requires interaction between the CARD-CARD or PYD-PYD domains. The CARD domain assembles with the pyrin domain to recruit caspase-1 through the adaptor molecule apoptosis-associated speck-like protein containing a CARD (ASC) (107). Two inflammasomes have been under investigation in CF lung inflammation in greater detail: nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3 (NLRP3) and NLR family CARD domain-containing protein 4 (NLRC4) (79). NLRC4 has been more extensively studied in CF than NLRP3, however, NLRP3 has been investigated in general in more detail (79, 108). The NLRP3 inflammasome is comprised of a N-terminal PYD domain, a central binding domain and a C-terminal LRR domain (105, 106, 109). The CARD-domain in the inflammasome contains the NLRs which assemble with the pyrin domain to recruit caspase-1 through the adaptor molecule, ASC (105, 106, 109). NLRs are named according to their domain structure and contain a nucleotide-binding domain (NBD), a signaling domain (CARD or PYRIN) and a leucine-rich repeat (LRR) domain that mediates ligand binding (110). Two signals are required for the NRLP3 inflammasome to produce bioactive IL-1β and IL-18: priming and activation. Priming of the inflammasome is the first signal and it manifests in the transcription of inflammasome proteins, pro-IL-1β and pro-IL-18 themselves. This priming step is typically initiated by TLR or cytokine receptor activation via NF-κB-dependent transcriptional upregulation of the affected genes. Activation of the inflammasome is the secondary signal resulting in the proteolytic cleavage of its target protein, pro-caspase-1, into enzymatically active caspase-1, that will then next cleave pro-IL-1β and pro-IL-18 into their bioactive, final forms (Figure 3). The NLRP3 inflammasome can be activated by a wide variety of signals ranging from bacterial toxins, pore-forming complexes, extracellular ATP, microcrystals and stimuli leading to lysosome rupture to release calcium, potassium ions and ROS (105, 106, 109, 111) (Figure 3). Accordingly, the activation of the NLRP3 inflammasome is triggered by an abundance of receptors and particles during infections and sterile inflammation. Caspase-1 also processes cytosolic gasdermin-D to unleash its pore-forming, N-terminal domain, resulting in the release of mature cytokines, alarmins and pyroptotic cell lysis (111) (Figure 3).
While the NLRP3 inflammasome is an essential part of the innate immune system, its overactivation can result in enhanced inflammation, tissue damage and contribute to disease pathologies in inflammatory conditions (112). Therefore, understanding the precise regulation of NLRP3 inflammasome activation is critical for fighting infections and curbing inflammation (113). Although cleavage of IL-18 and IL-1β takes place in the cytoplasm, IL-1β, in particular, has more than one pathway to exit the cell such as via exocytosis in secretory lysosomes, shedding of the plasma membrane in microvesicles, direct release by exosomes or through pyroptosis (114–116). Pyroptosis is a form of cell death dependent on caspase-1 and initiated by bacteria and a member of the NLR family receptors (117) (Figures 2, 3). IL-1β is involved in a variety of cellular activities such as cell proliferation, differentiation and apoptosis (113). IL-1β may also be processed independently of caspase-1 suggesting a larger role in the inflammatory responses (113). In summary, IL-1β has been detected at elevated levels in CF airways and thought to be a significant factor in driving chronic lung inflammation (Figures 1–3).
3.3 Inflammasome-dependent IL-1β release in CF
A line of several, in vitro and in vivo, studies suggest a role of inflammasome-mediated IL-1β activation in lung diseases including CF. Mice deficient in Asc have decreased IL-1β levels and are protected against lung fibrosis (118). IL-1Ra inhibits inflammasome activation in human CF bronchial epithelial cells yielding reduced IL-1β production (79). Anakinra, an IL-1R antagonist, displayed protective effects on neutrophilic inflammation in a Scn1b-Tg mice model of CF lung disease (85). Anakinra has also been shown to reduce NLRP3-mediated inflammation in Cftr-deficient mice and in human CF bronchial epithelial cells (79). Anakinra also regulates mROS production that is required for downstream activation of the NLRP3 inflammasome (118). NLRP3 depletion leads to robust reduction of IL-1β production in mice and human bronchial epithelial cells (79). Therefore, NLRP3 is likely the most significant mediator of IL-1β generation and release in the CF lung.
Neutrophils and macrophages not only respond to IL-1β but also produce bioactive IL-1β themselves. These cell types express more than one inflammasome type and therefore multiple inflammasomes may play a role in their IL-1β secretion. Macrophages secrete proinflammatory cytokines that enhance neutrophil responses, and vice versa, neutrophils release cytokines that increase the responsiveness of macrophages (Figures 1, 2). It is thought that macrophage dysfunction contributes to the early cascade of inflammatory events leading to chronic infection and inflammation in CF (119). CF macrophages demonstrate enhanced IL-1β secretion and reduced surface expression of TLR5 leading to the diminished bacterial phagocytosis and promotion of chronic inflammation (119). The excessive inflammatory response of CF macrophages also likely affects CF neutrophils contributing to a cascade of inflammatory signals originating from both cell types in CF.
3.4 Inflammasome-independent IL-1β synthesis in CF
While IL-1β is typically formed as a result of inflammasome activation, there is evidence that IL-1β can be generated independently of any inflammasome. Neutrophil-derived proteases, for instance, have been proposed to cause proteolytic cleavage of extracellular pro-IL-1β to bioactive IL-1β, contributing to the sustained inflammatory airway environment in CF (120) (Figure 3). Specifically, proteinase 3 (PR3) that colocalizes with NE in neutrophil primary granules, enhances bioactive IL-1β secretion (121–123). Neutrophil serine proteases have been shown to cleave IL-1β in several murine models of sterile inflammation (122–124). Inhibition of NE activity in Nlrp3-deficient mice infected with S. aureus significantly reduced airway levels of IL-1β, suggesting that NE-mediated conversion of IL-1β takes place in vivo (77). Furthermore, it has been shown that IL-1β is detectable in the lavage fluid of CF children (mean range of 3.8 years of age) in the absence of any infection, which is associated with enhanced NE activity and worsened structural lung damage (87). In a corneal, not pulmonary, P. aeruginosa infection mouse model, most of IL-1β was generated by NE, not capsase-1 or inflammasomes (125). Therefore, neutrophil-derived and NE-cleaved IL-1β could be a significant contributor to the total IL-1β released into the CF airway (Figure 2).
In addition, the P. aeruginosa protease lasB has also been shown to cleave IL-1β in vitro and to contribute to airway inflammation in a P. aeruginosa lung infection model in mice (126, 127). These findings suggest that not only host (inflammasome-dependent and -independent), but microbial proteases can also contribute to the generation of bioactive IL-1β in the CF airways and to ongoing inflammation (Figure 3).
3.5 Pathogen-independent activation of the NLRP3 inflammasome
NLRP3 inflammasome activation provides a vital element of host defense against invading pathogens in the lung. NLRP3 is, however, also activated in response to signals of cellular stress, independent of microbial stimuli. CFTR dysfunction leads to impaired endosomal trafficking, cytoskeleton disassembly and inflammasome activation through NF-κB to produce bioactive IL-1β in the CF airway epithelium (128). CFTR deficiency also results in the accumulation of ceramide that activates the NLPR3 inflammasome (129). βENaC mice harboring mucus-obstructed bronchioles characteristic of CF lung disease presented enhanced NLRP3 inflammasome activation in their epithelial cells and infiltrating leukocytes and a decrease of intracellular sphingosine-1 phosphate (S1P) signaling (130). Ceramide is the central sphingolipid metabolite that precedes S1P production to induce inflammatory responses (131, 132). S1P and ceramide signaling are part of the sphingolipid rheostat regulatory system with opposing effects (132). It is not fully understood how ceramide activates the NLRP3 inflammasome, but studies suggest that intracellular fatty acid crystals may recruit ASC and trigger ROS release (129, 133, 134). Pretreatment of airway epithelial cells of the βENaC mice with NLRP3 inflammasome inhibitors reduced their exaggerated inflammatory response (76). CF monocytes display enhanced NLRP3 inflammasome activity with increased IL-18, IL-1β, caspase-1 and ASC specks (76). In addition, treatment of CF monocytes with CFTR modulators, ivacaftor and tezacaftor, shows a reduction in IL-18, IL-1β, caspase-1 and ASC specks (135). Levels of IL-18 and IL-1β are also reduced in the serum of PwCF following treatment with CFTR modulators (135). The CFTR modulators partially restore CFTR function and alter CFTR-ENaC coupling to reduce the elevated amiloride-sensitive Na+ transport (136, 137). CFTR dysfunction also influences Ca2+ influx promoting a proinflammatory response in mitochondria (138) (Figure 3). Ca2+ influx promotes NLRP3 inflammasome activation (138). In human macrophages, Ca2+ is essential for the release of IL-1β and NLRP3 inflammasome activation (139) (Figure 3). Also, CF airway neutrophils exposed to LPS demonstrate increased cytoplasmic levels of the M2 isoform of pyruvate kinase that is known to increase pro-IL-1β synthesis while affecting the mitochondria to shuffle glucose between the tricarboxylic acid cycle to glycolysis (140). This results in increased glycolysis in neutrophils mitochondria and enhanced mROS generation, which is a major activator of the NLRP3 inflammasome (140). The signaling pathways for activation of the NLRP3 inflammasome in CF are not well-understood and require further investigations.
3.6 CF pathogen-dependent activation of the NLRP3 inflammasome
While the NLPR3 inflammasome can be activated by a broad range of different, non-microbial stimuli described above, bacterial pathogens and their PAMPs are also its strong activators. Pathogens in the lung trigger NLRP3 signaling directly through PAMPs and indirectly via host-stress signals through DAMPs. Pathogens can affect host cells by inducing K+ efflux, stimulating ROS production, Ca2+ mobilization, mitochondrial destabilization or lysosome rupture, all of which act as DAMPs for the innate immune system (Figure 3). P. aeruginosa is known to trigger the NLRC4 inflammasome in response to its flagellin and type 3 secretion system, however, P. aeruginosa loses those virulence factors over time to evade immune recognition and to develop chronic infection (141, 142). P. aeruginosa isolates from PwCF failed to induce inflammasome activation, IL-1β release and pyroptotic cell death in primary macrophages isolated during both stable infection and exacerbation (141). This was attributed to diminished expression of inflammasome ligands and reduced bacterial motility (141). CF human bronchial epithelial cells infected with a P. aeruginosa reference strain displayed mitochondrial perturbation to trigger NLRP3 activation, IL-1β and IL-18 processing (138). The flagellin was shown to be the responsible inducer for the mitochondrial Ca2+ uniporter to signal enhanced NLRP3 activation (138). Therefore, one of the reasons why P. aeruginosa loses its flagellum overtime is thought to prevent NLRP3 inflammasome activation while another important reason is to avoid recognition by neutrophils and to evade neutrophil-mediated killing (52). Clinical isolates of P. aeruginosa from PwCF collected at the beginning of infection induce inflammasome signaling, cell death and expression of IL-1β in macrophages, however, chronic isolates displayed poor inflammasome activation and proinflammatory cytokine release (143). Also, genetic polymorphisms in NLRP3 have been linked to higher rates of P. aeruginosa colonization in CF macrophages resulting in worsened lung function overtime (144).
As the currently most predominant respiratory pathogen in CF, S. aureus is also known to activate the NLRP3 inflammasome. A pore forming toxin of S. aureus, the Panton-Valentine Leukocidin (PVL), induces NLRP3 inflammasome in human primary monocytes, macrophages, and neutrophils (145). This suggests that toxins released by this pathogen may trigger inflammasome signaling, however, it remains unclear whether S. aureus CF clinical isolates produce these toxins (145). It is believed that PVL is the predominant factor of S. aureus to trigger inflammasome activation in human phagocytic cells to facilitate inflammation in the lung (145, 146). IL-1β released by PVL-intoxicated macrophages causes secretion of IL-8 and monocyte chemotactic protein-1 to recruit neutrophils (146). NLRP3 inflammasome activation in CF macrophages and neutrophils infected with bacterial clinical isolates have yet to be characterized, therefore, it remains largely unknown how chronic infection in PwCF affects inflammasome-mediated inflammation.
4 Interplay between NETs and IL-1β
4.1 NETs and IL-1β in disease pathologies
NETs and IL-1β have been studied in CF separately but their potential interactions remain largely unexplored. This work proposes that NETs and IL-1β mutually enhance each other’s generation and are therefore parts of a positive feedback loop that is a significant force in the chronic inflammatory process in the CF lung.
Neutrophils play a primary role in inflammation in many autoimmune and chronic inflammatory diseases such as adult-onset Still’s disease (AOSD), SLE, rheumatoid arthritis, gout, asthma and COPD (147–150). In several autoimmune diseases it has been shown that NETs trigger enhanced activation of NLRP3 expression in macrophages (151–153). In rheumatoid arthritis, treatment with an NLRP3 inhibitor improved inflammation indicating a crucial role of IL-1β in disease pathogenesis (152). It remains, however, largely unknown how NETs trigger NLRP3 inflammasome activation in rheumatoid arthritis. In AOSD, galectin-3, a protein known for macrophage activation, was shown to promote ASC and NLRP3 association and inflammasome activation (151). Lupus macrophages have increased inflammasome activation and IL-1β release in response to NETs and the NET-associated protein LL-37 (153). NETs have been described to play a central role in activating the NLRP3 inflammasome in macrophages via LL-37 triggering K+ efflux from the cell via the purinergic receptor P2X7R (154) (Figure 3). A cooperation between NETs and IL-1β has also been proposed to be behind the pathologies in conditions such as venous thrombosis (155), acute respiratory distress syndrome/acute lung injury (156) and cancer (157, 158).
The sputum of neutrophilic asthma and COPD patients contain elevated levels of extracellular DNA and increased gene expressions of NLRP3 and IL-1β, which correlate with the severity of lung disease (159). The major difference between these autoimmune diseases and CF is that chronic microbial infections are abundant in CF. More recently, in severe cases of COVID-19, IL-1β and NETs were proposed to lead to excessive alveolar and endothelial damage, suggesting a feed-forward loop involving both mechanisms (160). It is believed that the increased levels of IL-1β during SARS-CoV-2 infection activate more neutrophils resulting in increased NET extrusion, which in turn enhances clot formation, endothelial and alveolar damage in the lung (160).
4.2 The potential role of NETs in IL-1β production in CF
NETs can trigger macrophages and monocytes to release IL-1β through NLRP3 inflammasome activation causing persistent proinflammatory signals (Figures 2, 3). NETs were shown in vitro to promote the activation of the NLRP3 inflammasome and pyroptosis in peripheral blood mononuclear, pulmonary microvascular endothelial cells and keratinocytes (161–163).
In CF, NLRP3 has been found to be the dominant inflammasome, mediating IL-1β release in neutrophils (164). Intracellular accumulation of Cl- in PwCF stimulates the secretion of IL-1β, acting as an autocrine positive feedback loop by regulating NLRP3 and caspase-1 activation (165). P2X7R promotes inflammasome activation in monocytes/macrophages (166). P2X7R is overexpressed in CF monocytes and its inhibition decreases NLRP3 expression and IL-1β release (167). The NLRP3 inflammasome results in damaging levels of inflammation that may be beneficial to the pathogens surviving in the airways of PwCF. NETs are present in large amounts in the CF lung; however, pathogens are able to evade being killed by NETs, therefore, NETs may contribute to the enhanced NLRP3 activation observed in CF. Although chronic P. aeruginosa infection has been shown to evade NLRP3 activation, type 3 secretion system-negative P. aeruginosa is detected by guanylate binding protein 2 and interferon-inducible protein which causes bacterial lysis (168). The bacterial lysis activates caspase-11 which inhibits proliferation of bacteria and activates the NLRP3 inflammasome (168). On the other hand, Type III interferon has been shown to contribute to the pathogenesis of S. aureus infection in the airway, influencing the NLRP3 inflammasome and associated proinflammatory cytokine production (77). It was also demonstrated that IL-1β production during early infection was dependent on caspase-1, whereas during chronic infection, IL-1β generation was dependent on NE (77). Type III IFN activates inflammasome signaling through the JAK/STAT pathway (77, 169). NE and caspase-1 are required for IL-1β processing in response to S. aureus lung infection and the inhibition of both, NE and NLRP3, reduce airway levels of IL-1β and clears bacterial airway infection (77).
Lethal NET formation also represents a process by which the intracellular neutrophil content is released into the extracellular environment, and NETs have also been proposed as a platform for extracellular delivery of pro-IL-1β, the inactive preform of IL-1β (170) (Figure 3). Pyroptosis, necrosis, exosome release or breakdown following apoptosis of immune cells have all been proposed as alternative mechanisms for the extracellular delivery of pro-IL-1β (170, 171). Once in the extracellular space, pro-IL-1β can be cleaved into bioactive IL-1β by neutrophil-derived (NE, PR3) or bacterial proteases (P. aeruginosa lasB) (Figure 3) (121–123, 126, 127).
In summary, the NLRP3 inflammasome is activated by the innate immune system to aid in recruitment of immune cells for bacterial clearance, however, in the CF lung this may be detrimental by feeding lung tissue damage and chronic infections.
4.3 The potential role of IL-1β in NET formation in CF
While it has been studied more exhaustedly how NETs affect inflammasome activation, interestingly, much less information is available about how IL-1β interferes with NET release. The best this topic has been studied is in the context of gout, an autoinflammatory condition characterized by monosodium urate (MSU) crystal-induced inflammasome activation and IL-1β release in macrophages, followed by neutrophil recruitment and NET extrusion (172). Neutrophils play a major role in mediating inflammation in gout, and IL-1β is a crucial cytokine being released in large quantities from macrophages activated by MSU crystals (173). While IL-1β itself does not stimulate NETs in vitro, it was shown to significantly enhance MSU crystal-induced NET release in neutrophils, suggesting that the two innate immune mechanisms could promote each other in a positive feed-forward manner (173). Anakinra inhibited the potentiation of MSU crystal-stimulated NET formation by IL-1β in vitro (173, 174). It remains an interesting question whether IL-1β also promotes NET formation induced by stimuli other than MSU crystals that are clinically relevant for CF lung disease such as P. aeruginosa, S. aureus, other CF pathogens or microbial or host molecules.
5 Clinical considerations
The dysregulated and overactive innate immune system represents a clinically attractive target in CF to improve lung function. NETs and IL-1β represent two important components of CF lung inflammation. Degrading or inhibiting the formation or activities of NETs, IL-1β, or both, are expected to benefit PwCF. Extracellular DNA is hydrolyzed by the human endonuclease, deoxyribonuclease I (DNase I). Recombinant DNase targeting NETs significantly reduces airway levels of extracellular DNA and improves lung function in PwCF (175). DNase I is the only mucolytic agent with proven efficacy in CF and remains recommended to be given, even in the era of effective CFTR modulators (175). Currently, there are no therapies approved for PwCF that specifically target NET formation (40).
Targeting IL-1β therapeutically became first feasible by the introduction of anakinra, a recombinant and non-glycosylated form of the naturally occurring IL-1 receptor antagonist (IL-1Ra) (176). While anakinra has not been approved for PwCF to date, results of animal models and preclinical studies suggest it could represent a beneficial anti-inflammatory therapy in CF with no or minimal side effects (177). Canakinumab (ACZ885) is a human anti-IL-1β monoclonal neutralizing antibody (Novartis) that was approved by the Food and Drug Administration in the U.S.A. in 2009 for the treatment of familial cold auto-inflammatory syndrome and Muckle-Wells syndrome (178) but it has not been approved for PwCF.
Accumulating data suggest that CFTR correctors have beneficial effects on chronic lung inflammation in CF but failed to resolve it entirely (167, 179, 180). Trikafta therapy significantly increases CFTR protein expression and reduces ATP/P2X7R-induced NLRP3 inflammasome activation, reducing airway inflammation in PwCF (167).
As theoretical, additional possibilities, activation of the inflammasomes, caspase-1 or NET formation could be inhibited by specific inhibitors. Whether these, other, anti-inflammatory approaches, or their combinations with each other or CFTR correctors, will be clinically feasible in CF, will be decided in the future.
6 Conclusions
Altogether, cells in the CF airway demonstrate dysregulated NLRP3 inflammasome signaling characterized by enhanced IL-1β secretion. A characteristic of CF lung disease is the infiltration of neutrophils releasing NETs that could be potentially enhanced by IL-1β. Microbial infections in the CF lung induce NET release which induces more NLRP3 activation in macrophages and keeps feeding chronic inflammation. We acknowledge that focusing on NETs and IL-1β in this manuscript is somewhat subjective and several other mechanisms of the innate immune system take place in the lungs of PwCF that could drive lung disease. The data summarized in this review article propose, however, that a self-perpetuating, positive feedback loop consisting of IL-1β and NETs represent a significant driving force behind CF lung disease progression (Figure 1). While both mechanisms are complex and relevant to fight infections in healthy individuals, data accumulated thus far mainly suggests their overall pathologic, not beneficial, role in CF. Therefore, understanding the pathways associated with inflammasome activation and NET formation could aid in developing new therapeutics for PwCF to prevent or reduce chronic lung disease.
Author contributions
KF: Conceptualization, Writing – original draft. NG: Writing – review & editing. BR: Writing – review & editing, Conceptualization, Funding acquisition.
Funding
The author(s) declare that financial support was received for the research and/or publication of this article. This work was funded by the National Institutes of Health awards (R01HL136707 R21AI183057 to B. Rada). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Conflict of interest
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.
Generative AI statement
The author(s) declare that no Generative AI was used in the creation of this manuscript.
Publisher’s note
All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.
Abbreviations
AOSD, adult-onset Still’s disease; ASC, apoptosis-associated speck-like protein containing a CARD; CARD, caspase recruitment domain; CF, cystic fibrosis; CFTR, cystic fibrosis transmembrane conductance regulator; COPD, chronic obstructive pulmonary disease; DAMP, danger-associated molecular pattern; DNase I, deoxyribonuclease I; IL-1β, interleukin-1 beta; IL-1Ra, Interleukin-1 receptor antagonist; LPS, lipopolysaccharide; LRR, leucine-rich repeat; MPO, myeloperoxidase; mROS, mitochondrial ROS; MSU, monosodium urate; NBD, nucleotide binding domain; NE, neutrophil elastase; NET; neutrophil extracellular traps; NLR, Nod-like receptor; Nox, NADPH oxidase; NLRC4, NLR family CARD domain-containing protein 4; NLRP3; nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3; NTM, non-tuberculous mycobacteria; PAD4, peptidyl deaminase 4; PAMP, pathogen-associated molecular pattern; PR3, proteinase 3; PRR, pattern recognition receptor; PVL, Panton-Valentine Leukocidin; PwCF, people with cystic fibrosis; PYD, pyrin domain; ROS, reactive oxygen species; S1P, sphingosine-1 phosphate; SK channels, small-conductance, calcium-activated potassium channels; SNP, single nucleotide polymorphism; SLE, systemic lupus erythematosus; TLR, Toll-like receptor.
References
1. Brady E, Perkins RC, Cullen K, Sawicki GS, Kaplan RS, and Doyle G. Innovations in evaluating ambulatory costs of cystic fibrosis care: A comparative study across multidisciplinary care centers in Ireland and the United States. NEJM Catal Innov Care Delivery. (2025) 6. doi: 10.1056/CAT.24.0095
2. Guo J, Garratt A, and Hill A. Worldwide rates of diagnosis and effective treatment for cystic fibrosis. J Cyst Fibros. (2022) 21:456–62. doi: 10.1016/j.jcf.2022.01.009
3. Tsui LC and Dorfman R. The cystic fibrosis gene: a molecular genetic perspective. Cold Spring Harb Perspect Med. (2013) 3:a009472. doi: 10.1101/cshperspect.a009472
4. Saint-Criq V and Gray MA. Role of CFTR in epithelial physiology. Cell Mol Life Sci. (2017) 74:93–115. doi: 10.1007/s00018-016-2391-y
5. Anwar S, Peng JL, Zahid KR, Zhou YM, Ali Q, and Qiu CR. Cystic fibrosis: understanding cystic fibrosis transmembrane regulator mutation classification and modulator therapies. Adv Respir Med. (2024) 92:263–77. doi: 10.3390/arm92040026
6. Farinha CM and Callebaut I. Molecular mechanisms of cystic fibrosis - how mutations lead to misfunction and guide therapy. Biosci Rep. (2022) 42. doi: 10.1042/BSR20212006
7. Doring G and Gulbins E. Cystic fibrosis and innate immunity: how chloride channel mutations provoke lung disease. Cell Microbiol. (2009) 11:208–16. doi: 10.1111/j.1462-5822.2008.01271.x
8. Boucher RC. Airway surface dehydration in cystic fibrosis: pathogenesis and therapy. Annu Rev Med. (2007) 58:157–70. doi: 10.1146/annurev.med.58.071905.105316
9. 2017 Annual Data Report. Cystic fibrosis foundation patient registry. Bethesda, Maryland, USA: Cystic Fibrosis Foundation (2018).
10. Khan MA, Ali ZS, Sweezey N, Grasemann H, and Palaniyar N. Progression of cystic fibrosis lung disease from childhood to adulthood: neutrophils, neutrophil extracellular trap (NET) formation, and NET degradation. Genes (Basel). (2019) 10. doi: 10.3390/genes10030183
11. Rodriguez-Sevilla G, Garcia-Coca M, Romera-Garcia D, Aguilera-Correa JJ, Mahillo-Fernandez I, Esteban J, et al. Non-Tuberculous Mycobacteria multispecies biofilms in cystic fibrosis: development of an in vitro Mycobacterium abscessus and Pseudomonas aeruginosa dual species biofilm model. Int J Med Microbiol. (2018) 308:413–23. doi: 10.1016/j.ijmm.2018.03.003
12. Baker EJ, Allcott G, and Cox JAG. Polymicrobial infection in cystic fibrosis and future perspectives for improving Mycobacterium abscessus drug discovery. NPJ Antimicrob Resist. (2024) 2:38. doi: 10.1038/s44259-024-00060-5
13. Cystic Fibrosis Foundation. Patient Registry Annual Data Report. Bethesda, Maryland, USA: Cystic Fibrosis Foundation (2015).
14. C.F. Foundation. Patient Registry Annual Data Report. Bethesda, Maryland, USA: Cystic Fibrosis Foundation (2023).
15. Nick JA, Daley CL, Lenhart-Pendergrass PM, and Davidson RM. Nontuberculous mycobacteria in cystic fibrosis. Curr Opin Pulm Med. (2021) 27:586–92. doi: 10.1097/MCP.0000000000000816
16. Hong G. Progress and challenges in fungal lung disease in cystic fibrosis. Curr Opin Pulm Med. (2022) 28:584–90. doi: 10.1097/MCP.0000000000000921
17. Poore TS, Hong G, and Zemanick ET. Fungal infection and inflammation in cystic fibrosis. Pathogens. (2021) 10. doi: 10.3390/pathogens10050618
18. Schwarz C, Eschenhagen P, and Bouchara JP. Emerging fungal threats in cystic fibrosis. Mycopathologia. (2021) 186:639–53. doi: 10.1007/s11046-021-00574-w
19. Nasser A, Moradi M, Jazireian P, Safari H, Alizadeh-Sani M, Pourmand MR, et al. Staphylococcus aureus versus neutrophil: Scrutiny of ancient combat. Microb Pathog. (2019) 131:259–69. doi: 10.1016/j.micpath.2019.04.026
20. Nickerson R, Thornton CS, Johnston B, Lee AHY, and Cheng Z. Pseudomonas aeruginosa in chronic lung disease: untangling the dysregulated host immune response. Front Immunol. (2024) 15:1405376. doi: 10.3389/fimmu.2024.1405376
21. Cohen-Cymberknoh M. Infection and Inflammation in the Cystic fibrosis (CF) airway. Pediatr Pulmonol. (2025) 60:S82–3. doi: 10.1002/ppul.27355
22. Rada B. Interactions between neutrophils and pseudomonas aeruginosa in cystic fibrosis. Pathogens. (2017) 6.doi: 10.3390/pathogens6010010
23. Mall MA, Davies JC, Donaldson SH, Jain R, Chalmers JD, and Shteinberg M. Neutrophil serine proteases in cystic fibrosis: role in disease pathogenesis and rationale as a therapeutic target. Eur Respir Rev. (2024) 33. doi: 10.1183/16000617.0001-2024
24. Wang G and Nauseef WM. Neutrophil dysfunction in the pathogenesis of cystic fibrosis. Blood. (2022) 139:2622–31. doi: 10.1182/blood.2021014699
25. Law SM and Gray RD. Neutrophil extracellular traps and the dysfunctional innate immune response of cystic fibrosis lung disease: a review. J Inflammation (Lond). (2017) 14:29. doi: 10.1186/s12950-017-0176-1
26. Meyer KC. Neutrophils, myeloperoxidase, and bronchiectasis in cystic fibrosis: green is not good. J Lab Clin Med. (2004) 144:124–6. doi: 10.1016/j.lab.2004.05.014
27. Burn GL, Foti A, Marsman G, Patel DF, and Zychlinsky A. The neutrophil. Immunity. (2021) 54:1377–91. doi: 10.1016/j.immuni.2021.06.006
28. Teng TS, Ji AL, Ji XY, and Li YZ. Neutrophils and immunity: from bactericidal action to being conquered. J Immunol Res. (2017) 2017:9671604. doi: 10.1155/2017/9671604
29. Brinkmann V, Reichard U, Goosmann C, Fauler B, Uhlemann Y, Weiss DS, et al. Neutrophil extracellular traps kill bacteria. Science. (2004) 303:1532–5. doi: 10.1126/science.1092385
30. Delgado-Rizo V, Martinez-Guzman MA, Iniguez-Gutierrez L, Garcia-Orozco A, Alvarado-Navarro A, and Fafutis-Morris M. Neutrophil extracellular traps and its implications in inflammation: an overview. Front Immunol. (2017) 8:81. doi: 10.3389/fimmu.2017.00081
31. Ravindran M, Khan MA, and Palaniyar N. Neutrophil extracellular trap formation: physiology, pathology, and pharmacology. Biomolecules 9. (2019). doi: 10.3390/biom9080365
32. Cristinziano L, Modestino L, Loffredo S, Varricchi G, Braile M, Ferrara AL, et al. Anaplastic thyroid cancer cells induce the release of mitochondrial extracellular DNA traps by viable neutrophils. J Immunol. (2020) 204:1362–72. doi: 10.4049/jimmunol.1900543
33. Yousefi S, Mihalache C, Kozlowski E, Schmid I, and Simon HU. Viable neutrophils release mitochondrial DNA to form neutrophil extracellular traps. Cell Death Differ. (2009) 16:1438–44. doi: 10.1038/cdd.2009.96
34. Khan MA, Farahvash A, Douda DN, Licht JC, Grasemann H, Sweezey N, et al. JNK activation turns on LPS- and gram-negative bacteria-induced NADPH oxidase-dependent suicidal NETosis. Sci Rep. (2017) 7:3409. doi: 10.1038/s41598-017-03257-z
35. Parker H, Dragunow M, Hampton MB, Kettle AJ, and Winterbourn CC. Requirements for NADPH oxidase and myeloperoxidase in neutrophil extracellular trap formation differ depending on the stimulus. J Leukoc Biol. (2012) 92:841–9. doi: 10.1189/jlb.1211601
36. Papayannopoulos V, Metzler KD, Hakkim A, and Zychlinsky A. Neutrophil elastase and myeloperoxidase regulate the formation of neutrophil extracellular traps. J Cell Biol. (2010) 191:677–91. doi: 10.1083/jcb.201006052
37. Douda DN, Khan MA, Grasemann H, and Palaniyar N. SK3 channel and mitochondrial ROS mediate NADPH oxidase-independent NETosis induced by calcium influx. Proc Natl Acad Sci U.S.A. (2015) 112:2817–22. doi: 10.1073/pnas.1414055112
38. Khan MA and Palaniyar N. Transcriptional firing helps to drive NETosis. Sci Rep. (2017) 7:41749. doi: 10.1038/srep41749
39. Hamam HJ, Khan MA, and Palaniyar N. Histone acetylation promotes neutrophil extracellular trap formation. Biomolecules. (2019) 9. doi: 10.3390/biom9010032
40. Keir HR and Chalmers JD. Neutrophil extracellular traps in chronic lung disease: implications for pathogenesis and therapy. Eur Respir Rev. (2022) 31. doi: 10.1183/16000617.0241-2021
41. Pan T and Lee JW. A crucial role of neutrophil extracellular traps in pulmonary infectious diseases. Chin Med J Pulm Crit Care Med. (2024) 2:34–41. doi: 10.1016/j.pccm.2023.10.004
42. Godement M, Zhu J, Cerf C, Vieillard-Baron A, Maillon A, Zuber B, et al. Neutrophil extracellular traps in SARS-coV2 related pneumonia in ICU patients: the NETCOV2 study. Front Med (Lausanne). (2021) 8:615984. doi: 10.3389/fmed.2021.615984
43. Thierry AR and Roch B. SARS-CoV2 may evade innate immune response, causing uncontrolled neutrophil extracellular traps formation and multi-organ failure. Clin Sci (Lond). (2020) 134:1295–300. doi: 10.1042/CS20200531
44. Li S, Wang H, and Shao Q. The central role of neutrophil extracellular traps (NETs) and by-products in COVID-19 related pulmonary thrombosis. Immun Inflammation Dis. (2023) 11:e949. doi: 10.1002/iid3.949
45. Kyala NJ, Mboya I, Shao E, Sakita F, Kilonzo KG, Shirima L, et al. Neutrophil-to-lymphocyte ratio as a prognostic indicator in COVID-19: Evidence from a northern Tanzanian cohort. PloS One. (2025) 20:e0300231. doi: 10.1371/journal.pone.0300231
46. Nathan C. Neutrophils and COVID-19: nots, NETs, and knots. J Exp Med. (2020) 217. doi: 10.1084/jem.20201439
47. Zuo Y, Yalavarthi S, Shi H, Gockman K, Zuo M, Madison JA, et al. Neutrophil extracellular traps (NETs) as markers of disease severity in COVID-19. medRxiv. (2020). doi: 10.1101/2020.04.09.20059626
48. Laforge M, Elbim C, Frere C, Hemadi M, Massaad C, Nuss P, et al. Tissue damage from neutrophil-induced oxidative stress in COVID-19. Nat Rev Immunol. (2020) 20:515–6. doi: 10.1038/s41577-020-0407-1
49. Rada B. Neutrophil extracellular trap release driven by bacterial motility: Relevance to cystic fibrosis lung disease. Commun Integr Biol. (2017) 10:e1296610. doi: 10.1080/19420889.2017.1296610
50. Marcos V, Zhou-Suckow Z, Onder Yildirim A, Bohla A, Hector A, Vitkov L, et al. Free DNA in cystic fibrosis airway fluids correlates with airflow obstruction. Mediators Inflammation. (2015) 2015:408935. doi: 10.1155/2015/408935
51. Davies JC, Ebdon AM, and Orchard C. Recent advances in the management of cystic fibrosis. Arch Dis Child. (2014) 99:1033–6. doi: 10.1136/archdischild-2013-304400
52. Floyd M, Winn M, Cullen C, Sil P, Chassaing B, Yoo DG, et al. Swimming motility mediates the formation of neutrophil extracellular traps induced by flagellated pseudomonas aeruginosa. PloS Pathog. (2016) 12:e1005987. doi: 10.1371/journal.ppat.1005987
53. Martinez-Aleman S, Bustamante AE, Jimenez-Valdes RJ, Gonzalez GM, and Sanchez-Gonzalez A. Pseudomonas aeruginosa isolates from cystic fibrosis patients induce neutrophil extracellular traps with different morphologies that could correlate with their disease severity. Int J Med Microbiol. (2020) 310:151451. doi: 10.1016/j.ijmm.2020.151451
54. Young RL, Malcolm KC, Kret JE, Caceres SM, Poch KR, Nichols DP, et al. Neutrophil extracellular trap (NET)-mediated killing of Pseudomonas aeruginosa: evidence of acquired resistance within the CF airway, independent of CFTR. PloS One. (2011) 6:e23637. doi: 10.1371/journal.pone.0023637
55. Berends ET, Horswill AR, Haste NM, Monestier M, Nizet V, and von Kockritz-Blickwede M. Nuclease expression by Staphylococcus aureus facilitates escape from neutrophil extracellular traps. J Innate Immun. (2010) 2:576–86. doi: 10.1159/000319909
56. Speziale P and Pietrocola G. Staphylococcus aureus induces neutrophil extracellular traps (NETs) and neutralizes their bactericidal potential. Comput Struct Biotechnol J. (2021) 19:3451–7. doi: 10.1016/j.csbj.2021.06.012
57. Yoo DG, Winn M, Pang L, Moskowitz SM, Malech HL, Leto TL, et al. Release of cystic fibrosis airway inflammatory markers from Pseudomonas aeruginosa-stimulated human neutrophils involves NADPH oxidase-dependent extracellular DNA trap formation. J Immunol. (2014) 192:4728–38. doi: 10.4049/jimmunol.1301589
58. Yoo DG, Floyd M, Winn M, Moskowitz SM, and Rada B. NET formation induced by Pseudomonas aeruginosa cystic fibrosis isolates measured as release of myeloperoxidase-DNA and neutrophil elastase-DNA complexes. Immunol Lett. (2014) 160:186–94. doi: 10.1016/j.imlet.2014.03.003
59. Herzog S, Dach F, de Buhr N, Niemann S, Schlagowski J, Chaves-Moreno D, et al. High nuclease activity of long persisting staphylococcus aureus isolates within the airways of cystic fibrosis patients protects against NET-mediated killing. Front Immunol. (2019) 10:2552. doi: 10.3389/fimmu.2019.02552
60. Alhede M, Alhede M, Qvortrup K, Kragh KN, Jensen PO, Stewart PS, et al. The origin of extracellular DNA in bacterial biofilm infections in vivo. Pathog Dis. (2020) 78. doi: 10.1093/femspd/ftaa018
61. Moriceau S, Kantari C, Mocek J, Davezac N, Gabillet J, Guerrera IC, et al. Coronin-1 is associated with neutrophil survival and is cleaved during apoptosis: potential implication in neutrophils from cystic fibrosis patients. J Immunol. (2009) 182:7254–63. doi: 10.4049/jimmunol.0803312
62. Moriceau S, Lenoir G, and Witko-Sarsat V. In cystic fibrosis homozygotes and heterozygotes, neutrophil apoptosis is delayed and modulated by diamide or roscovitine: evidence for an innate neutrophil disturbance. J Innate Immun. (2010) 2:260–6. doi: 10.1159/000295791
63. Gray RD, Hardisty G, Regan KH, Smith M, Robb CT, Duffin R, et al. Delayed neutrophil apoptosis enhances NET formation in cystic fibrosis. Thorax. (2018) 73:134–44. doi: 10.1136/thoraxjnl-2017-210134
64. Law SM, Hardisty G, Gillan JL, Robinson NJ, Davidson DJ, Whyte MKB, et al. Neutrophil extracellular traps are associated with airways inflammation and increased severity of lung disease in cystic fibrosis. ERJ Open Res. (2024) 10. doi: 10.1183/23120541.00312-2024
65. Skopelja S, Hamilton BJ, Jones JD, Yang ML, Mamula M, Ashare A, et al. The role for neutrophil extracellular traps in cystic fibrosis autoimmunity. JCI Insight. (2016) 1:e88912. doi: 10.1172/jci.insight.88912
66. Zhao MH, Jayne DR, Ardiles LG, Culley F, Hodson ME, and Lockwood CM. Autoantibodies against bactericidal/permeability-increasing protein in patients with cystic fibrosis. QJM. (1996) 89:259–65. doi: 10.1093/qjmed/89.4.259
67. Yadav R, Yoo DG, Kahlenberg JM, Bridges SL Jr., Oni O, Huang H, et al. Systemic levels of anti-PAD4 autoantibodies correlate with airway obstruction in cystic fibrosis. J Cyst Fibros. (2019) 18:636–45. doi: 10.1016/j.jcf.2018.12.010
68. Linnemann RW, Yadav R, Zhang C, Sarr D, Rada B, and Stecenko AA. Serum anti-PAD4 autoantibodies are present in cystic fibrosis children and increase with age and lung disease severity. Autoimmunity. (2022) 55:109–17. doi: 10.1080/08916934.2021.2021193
69. Yadav R, Linnemann RW, Kahlenberg JM, Bridges LS Jr., Stecenko AA, and Rada B. IgA autoantibodies directed against self DNA are elevated in cystic fibrosis and associated with more severe lung dysfunction. Autoimmunity. (2020) 53:476–84. doi: 10.1080/08916934.2020.1839890
70. van Delft MAM, Aleyd E, van der Mast R, de Jong N, Boon L, Simons PJ, et al. Antagonizing FcalphaR1 (CD89) as treatment in IgA-mediated chronic inflammation and autoimmunity. Front Immunol. (2023) 14:1118539. doi: 10.3389/fimmu.2023.1118539
71. Bos A, Aleyd E, van der Steen LPE, Winter PJ, Heemskerk N, Pouw SM, et al. Anti-fcalphaRI monoclonal antibodies resolve igA autoantibody-mediated disease. Front Immunol. (2022) 13:732977. doi: 10.3389/fimmu.2022.732977
72. Martinon F, Burns K, and Tschopp J. The inflammasome: a molecular platform triggering activation of inflammatory caspases and processing of proIL-beta. Mol Cell. (2002) 10:417–26. doi: 10.1016/S1097-2765(02)00599-3
73. Montgomery ST, Dittrich AS, Garratt LW, Turkovic L, Frey DL, Stick SM, et al. Interleukin-1 is associated with inflammation and structural lung disease in young children with cystic fibrosis. J Cyst Fibros. (2018) 17:715–22. doi: 10.1016/j.jcf.2018.05.006
74. Tian X, Sun H, Casbon AJ, Lim E, Francis KP, Hellman J, et al. NLRP3 Inflammasome Mediates Dormant Neutrophil Recruitment following Sterile Lung Injury and Protects against Subsequent Bacterial Pneumonia in Mice. Front Immunol. (2017) 8:1337. doi: 10.3389/fimmu.2017.01337
75. Tang A, Sharma A, Jen R, Hirschfeld AF, Chilvers MA, Lavoie PM, et al. Inflammasome-mediated IL-1beta production in humans with cystic fibrosis. PloS One. (2012) 7:e37689. doi: 10.1371/journal.pone.0037689
76. Scambler T, Jarosz-Griffiths HH, Lara-Reyna S, Pathak S, Wong C, Holbrook J, et al. ENaC-mediated sodium influx exacerbates NLRP3-dependent inflammation in cystic fibrosis. Elife. (2019) 8. doi: 10.7554/eLife.49248
77. Pires S and Parker D. IL-1beta activation in response to Staphylococcus aureus lung infection requires inflammasome-dependent and independent mechanisms. Eur J Immunol. (2018) 48:1707–16. doi: 10.1002/eji.201847556
78. Palomo J, Marchiol T, Piotet J, Fauconnier L, Robinet M, Reverchon F, et al. Role of IL-1beta in experimental cystic fibrosis upon P. aeruginosa infection. PloS One. (2014) 9:e114884. doi: 10.1371/journal.pone.0114884
79. Iannitti RG, Napolioni V, Oikonomou V, De Luca A, Galosi C, Pariano M, et al. IL-1 receptor antagonist ameliorates inflammasome-dependent inflammation in murine and human cystic fibrosis. Nat Commun. (2016) 7:10791. doi: 10.1038/ncomms10791
80. Chen G, Sun L, Kato T, Okuda K, Martino MB, Abzhanova A, et al. IL-1beta dominates the promucin secretory cytokine profile in cystic fibrosis. J Clin Invest. (2019) 129:4433–50. doi: 10.1172/JCI125669
81. Chen Y, Garvin LM, Nickola TJ, Watson AM, Colberg-Poley AM, and Rose MC. IL-1beta induction of MUC5AC gene expression is mediated by CREB and NF-kappaB and repressed by dexamethasone. Am J Physiol Lung Cell Mol Physiol. (2014) 306:L797–807. doi: 10.1152/ajplung.00347.2013
82. Fujisawa T, Chang MM, Velichko S, Thai P, Hung LY, Huang F, et al. NF-kappaB mediates IL-1beta- and IL-17A-induced MUC5B expression in airway epithelial cells. Am J Respir Cell Mol Biol. (2011) 45:246–52. doi: 10.1165/rcmb.2009-0313OC
83. Balazs A and Mall MA. Mucus obstruction and inflammation in early cystic fibrosis lung disease: Emerging role of the IL-1 signaling pathway. Pediatr Pulmonol. (2019) 54:S5–S12. doi: 10.1002/ppul.24462
84. Dinarello CA, Simon A, and van der Meer JW. Treating inflammation by blocking interleukin-1 in a broad spectrum of diseases. Nat Rev Drug Discov. (2012) 11:633–52. doi: 10.1038/nrd3800
85. Fritzsching B, Zhou-Suckow Z, Trojanek JB, Schubert SC, Schatterny J, Hirtz S, et al. Hypoxic epithelial necrosis triggers neutrophilic inflammation via IL-1 receptor signaling in cystic fibrosis lung disease. Am J Respir Crit Care Med. (2015) 191:902–13. doi: 10.1164/rccm.201409-1610OC
86. Levy H, Murphy A, Zou F, Gerard C, Klanderman B, Schuemann B, et al. IL1B polymorphisms modulate cystic fibrosis lung disease. Pediatr Pulmonol. (2009) 44:580–93. doi: 10.1002/ppul.21026
87. de Vries L, Griffiths A, Armstrong D, and Robinson PJ. Cytokine gene polymorphisms and severity of CF lung disease. J Cyst Fibros. (2014) 13:699–705. doi: 10.1016/j.jcf.2014.04.007
88. Chen KW, Bezbradica JS, Gross CJ, Wall AA, Sweet MJ, Stow JL, et al. The murine neutrophil NLRP3 inflammasome is activated by soluble but not particulate or crystalline agonists. Eur J Immunol. (2016) 46:1004–10. doi: 10.1002/eji.201545943
89. Karmakar M, Katsnelson MA, Dubyak GR, and Pearlman E. Neutrophil P2X7 receptors mediate NLRP3 inflammasome-dependent IL-1beta secretion in response to ATP. Nat Commun. (2016) 7:10555. doi: 10.1038/ncomms10555
90. Yang Y, Bin W, Aksoy MO, and Kelsen SG. Regulation of interleukin-1beta and interleukin-1beta inhibitor release by human airway epithelial cells. Eur Respir J. (2004) 24:360–6. doi: 10.1183/09031936.04.00089703
91. Britton N, Villabona-Rueda A, Whiteside SA, Mathew J, Kelley M, Agbor-Enoh S, et al. Pseudomonas-dominant microbiome elicits sustained IL-1beta upregulation in alveolar macrophages from lung transplant recipients. J Heart Lung Transplant. (2023) 42:1166–74. doi: 10.1016/j.healun.2023.04.005
92. Miao EA, Ernst RK, Dors M, Mao DP, and Aderem A. Pseudomonas aeruginosa activates caspase 1 through Ipaf. Proc Natl Acad Sci U.S.A. (2008) 105:2562–7. doi: 10.1073/pnas.0712183105
93. Minns MS, Liboro K, Lima TS, Abbondante S, Miller BA, Marshall ME, et al. NLRP3 selectively drives IL-1beta secretion by Pseudomonas aeruginosa infected neutrophils and regulates corneal disease severity. Nat Commun. (2023) 14:5832. doi: 10.1038/s41467-023-41391-7
94. Mills PR, Davies RJ, and Devalia JL. Airway epithelial cells, cytokines, and pollutants. Am J Respir Crit Care Med. (1999) 160:S38–43. doi: 10.1164/ajrccm.160.supplement_1.11
95. Shimada T, Park BG, Wolf AJ, Brikos C, Goodridge HS, Becker CA, et al. Staphylococcus aureus evades lysozyme-based peptidoglycan digestion that links phagocytosis, inflammasome activation, and IL-1beta secretion. Cell Host Microbe. (2010) 7:38–49. doi: 10.1016/j.chom.2009.12.008
96. Munoz-Planillo R, Franchi L, Miller LS, and Nunez G. A critical role for hemolysins and bacterial lipoproteins in Staphylococcus aureus-induced activation of the Nlrp3 inflammasome. J Immunol. (2009) 183:3942–8. doi: 10.4049/jimmunol.0900729
97. Kremserova S and Nauseef WM. Frontline Science: Staphylococcus aureus promotes receptor-interacting protein kinase 3- and protease-dependent production of IL-1beta in human neutrophils. J Leukoc Biol. (2019) 105:437–47. doi: 10.1002/JLB.4HI0918-346R
98. Stecenko AA, King G, Torii K, Breyer RM, Dworski R, Blackwell TS, et al. Dysregulated cytokine production in human cystic fibrosis bronchial epithelial cells. Inflammation. (2001) 25:145–55. doi: 10.1023/A:1011080229374
99. Ruef C, Jefferson DM, Schlegel-Haueter SE, and Suter S. Regulation of cytokine secretion by cystic fibrosis airway epithelial cells. Eur Respir J. (1993) 6:1429–36. doi: 10.1183/09031936.93.06101429
100. Muselet-Charlier C, Roque T, Boncoeur E, Chadelat K, Clement A, Jacquot J, et al. Enhanced IL-1beta-induced IL-8 production in cystic fibrosis lung epithelial cells is dependent of both mitogen-activated protein kinases and NF-kappaB signaling. Biochem Biophys Res Commun. (2007) 357:402–7. doi: 10.1016/j.bbrc.2007.03.141
101. Courtney JM, Ennis M, and Elborn JS. Cytokines and inflammatory mediators in cystic fibrosis. J Cyst Fibros. (2004) 3:223–31. doi: 10.1016/j.jcf.2004.06.006
102. Bradley KL, Stokes CA, Marciniak SJ, Parker LC, and Condliffe AM. Role of unfolded proteins in lung disease. Thorax. (2021) 76:92–9. doi: 10.1136/thoraxjnl-2019-213738
103. Meyer M, Huaux F, Gavilanes X, van den Brule S, Lebecque P, Re S, et al. Azithromycin reduces exaggerated cytokine production by M1 alveolar macrophages in cystic fibrosis. Am J Respir Cell Mol Biol. (2009) 41:590–602. doi: 10.1165/rcmb.2008-0155OC
104. Franchi L, Eigenbrod T, Munoz-Planillo R, and Nunez G. The inflammasome: a caspase-1-activation platform that regulates immune responses and disease pathogenesis. Nat Immunol. (2009) 10:241–7. doi: 10.1038/ni.1703
105. Franchi L, Warner N, Viani K, and Nunez G. Function of Nod-like receptors in microbial recognition and host defense. Immunol Rev. (2009) 227:106–28. doi: 10.1111/j.1600-065X.2008.00734.x
106. Davis BK, Wen H, and Ting JP. The inflammasome NLRs in immunity, inflammation, and associated diseases. Annu Rev Immunol. (2011) 29:707–35. doi: 10.1146/annurev-immunol-031210-101405
107. Srinivasula SM, Poyet JL, Razmara M, Datta P, Zhang Z, and Alnemri ES. The PYRIN-CARD protein ASC is an activating adaptor for caspase-1. J Biol Chem. (2002) 277:21119–22. doi: 10.1074/jbc.C200179200
108. Yang Y, Wang H, Kouadir M, Song H, and Shi F. Recent advances in the mechanisms of NLRP3 inflammasome activation and its inhibitors. Cell Death Dis. (2019) 10:128. doi: 10.1038/s41419-019-1413-8
109. Swanson KV, Deng M, and Ting JP. The NLRP3 inflammasome: molecular activation and regulation to therapeutics. Nat Rev Immunol. (2019) 19:477–89. doi: 10.1038/s41577-019-0165-0
110. Ting JP, Lovering RC, Alnemri ES, Bertin J, Boss JM, Davis BK, et al. The NLR gene family: a standard nomenclature. Immunity. (2008) 28:285–7. doi: 10.1016/j.immuni.2008.02.005
111. Zhang Y, Chen X, Gueydan C, and Han J. Plasma membrane changes during programmed cell deaths. Cell Res. (2018) 28:9–21. doi: 10.1038/cr.2017.133
112. Shaw PJ, McDermott MF, and Kanneganti TD. Inflammasomes and autoimmunity. Trends Mol Med. (2011) 17:57–64. doi: 10.1016/j.molmed.2010.11.001
113. Dinarello CA. Overview of the IL-1 family in innate inflammation and acquired immunity. Immunol Rev. (2018) 281:8–27. doi: 10.1111/imr.12621
114. Andrei C, Dazzi C, Lotti L, Torrisi MR, Chimini G, and Rubartelli A. The secretory route of the leaderless protein interleukin 1beta involves exocytosis of endolysosome-related vesicles. Mol Biol Cell. (1999) 10:1463–75. doi: 10.1091/mbc.10.5.1463
115. Qu Y, Franchi L, Nunez G, and Dubyak GR. Nonclassical IL-1 beta secretion stimulated by P2X7 receptors is dependent on inflammasome activation and correlated with exosome release in murine macrophages. J Immunol. (2007) 179:1913–25. doi: 10.4049/jimmunol.179.3.1913
116. Bergsbaken T, Fink SL, and Cookson BT. Pyroptosis: host cell death and inflammation. Nat Rev Microbiol. (2009) 7:99–109. doi: 10.1038/nrmicro2070
117. Suzuki T, Franchi L, Toma C, Ashida H, Ogawa M, Yoshikawa Y, et al. Differential regulation of caspase-1 activation, pyroptosis, and autophagy via Ipaf and ASC in Shigella-infected macrophages. PloS Pathog. (2007) 3:e111. doi: 10.1371/journal.ppat.0030111
118. Gasse P, Mary C, Guenon I, Noulin N, Charron S, Schnyder-Candrian S, et al. IL-1R1/MyD88 signaling and the inflammasome are essential in pulmonary inflammation and fibrosis in mice. J Clin Invest. (2007) 117:3786–99. doi: 10.1172/JCI32285
119. Simonin-Le Jeune K, Le Jeune A, Jouneau S, Belleguic C, Roux PF, Jaguin M, et al. Impaired functions of macrophage from cystic fibrosis patients: CD11b, TLR-5 decrease and sCD14, inflammatory cytokines increase. PloS One. (2013) 8:e75667. doi: 10.1371/journal.pone.0075667
120. Coeshott C, Ohnemus C, Pilyavskaya A, Ross S, Wieczorek M, Kroona H, et al. Converting enzyme-independent release of tumor necrosis factor alpha and IL-1beta from a stimulated human monocytic cell line in the presence of activated neutrophils or purified proteinase 3. Proc Natl Acad Sci U.S.A. (1999) 96:6261–6. doi: 10.1073/pnas.96.11.6261
121. Cassel SL, Janczy JR, Bing X, Wilson SP, Olivier AK, Otero JE, et al. Inflammasome-independent IL-1beta mediates autoinflammatory disease in Pstpip2-deficient mice. Proc Natl Acad Sci U.S.A. (2014) 111:1072–7. doi: 10.1073/pnas.1318685111
122. Guma M, Ronacher L, Liu-Bryan R, Takai S, Karin M, and Corr M. Caspase 1-independent activation of interleukin-1beta in neutrophil-predominant inflammation. Arthritis Rheum. (2009) 60:3642–50. doi: 10.1002/art.24959
123. Schreiber A, Pham CT, Hu Y, Schneider W, Luft FC, and Kettritz R. Neutrophil serine proteases promote IL-1beta generation and injury in necrotizing crescentic glomerulonephritis. J Am Soc Nephrol. (2012) 23:470–82. doi: 10.1681/ASN.2010080892
124. Clancy DM, Sullivan GP, Moran HBT, Henry CM, Reeves EP, McElvaney NG, et al. Extracellular neutrophil proteases are efficient regulators of IL-1, IL-33, and IL-36 cytokine activity but poor effectors of microbial killing. Cell Rep. (2018) 22:2937–50. doi: 10.1016/j.celrep.2018.02.062
125. Karmakar M, Sun Y, Hise AG, Rietsch A, and Pearlman E. Cutting edge: IL-1beta processing during Pseudomonas aeruginosa infection is mediated by neutrophil serine proteases and is independent of NLRC4 and caspase-1. J Immunol. (2012) 189:4231–5. doi: 10.4049/jimmunol.1201447
126. Everett MJ, Davies DT, Leiris S, Sprynski N, Llanos A, Castandet JM, et al. Chemical optimization of selective pseudomonas aeruginosa lasB elastase inhibitors and their impact on lasB-mediated activation of IL-1beta in cellular and animal infection models. ACS Infect Dis. (2023) 9:270–82. doi: 10.1021/acsinfecdis.2c00418
127. Sun J, LaRock DL, Skowronski EA, Kimmey JM, Olson J, Jiang Z, et al. The Pseudomonas aeruginosa protease LasB directly activates IL-1beta. EBioMedicine. (2020) 60:102984. doi: 10.1016/j.ebiom.2020.102984
128. Villella VR, Venerando A, Cozza G, Esposito S, Ferrari E, Monzani R, et al. A pathogenic role for cystic fibrosis transmembrane conductance regulator in celiac disease. EMBO J. (2019) 38. doi: 10.15252/embj.2018100101
129. Grassme H, Carpinteiro A, Edwards MJ, Gulbins E, and Becker KA. Regulation of the inflammasome by ceramide in cystic fibrosis lungs. Cell Physiol Biochem. (2014) 34:45–55. doi: 10.1159/000362983
130. Tran HB, Macowan MG, Abdo A, Donnelley M, Parsons D, and Hodge S. Enhanced inflammasome activation and reduced sphingosine-1 phosphate S1P signalling in a respiratory mucoobstructive disease model. J Inflammation (Lond). (2020) 17:16. doi: 10.1186/s12950-020-00248-2
131. Hait NC and Maiti A. The role of sphingosine-1-phosphate and ceramide-1-phosphate in inflammation and cancer. Mediators Inflammation. (2017) 2017:4806541. doi: 10.1155/2017/4806541
132. Maceyka M, Harikumar KB, Milstien S, and Spiegel S. Sphingosine-1-phosphate signaling and its role in disease. Trends Cell Biol. (2012) 22:50–60. doi: 10.1016/j.tcb.2011.09.003
133. Camell CD, Nguyen KY, Jurczak MJ, Christian BE, Shulman GI, Shadel GS, et al. Macrophage-specific de novo synthesis of ceramide is dispensable for inflammasome-driven inflammation and insulin resistance in obesity. J Biol Chem. (2015) 290:29402–13. doi: 10.1074/jbc.M115.680199
134. Karasawa T, Kawashima A, Usui-Kawanishi F, Watanabe S, Kimura H, Kamata R, et al. Saturated fatty acids undergo intracellular crystallization and activate the NLRP3 inflammasome in macrophages. Arterioscler Thromb Vasc Biol. (2018) 38:744–56. doi: 10.1161/ATVBAHA.117.310581
135. Jarosz-Griffiths HH, Scambler T, Wong CH, Lara-Reyna S, Holbrook J, Martinon F, et al. Different CFTR modulator combinations downregulate inflammation differently in cystic fibrosis. Elife 9. (2020). doi: 10.7554/eLife.54556
136. Pranke IM, Hatton A, Simonin J, Jais JP, Le Pimpec-Barthes F, Carsin A, et al. Correction of CFTR function in nasal epithelial cells from cystic fibrosis patients predicts improvement of respiratory function by CFTR modulators. Sci Rep. (2017) 7:7375. doi: 10.1038/s41598-017-07504-1
137. Cholon DM, Quinney NL, Fulcher ML, Esther CR Jr., Das J, Dokholyan NV, et al. Potentiator ivacaftor abrogates pharmacological correction of DeltaF508 CFTR in cystic fibrosis. Sci Transl Med. (2014) 6:246ra96. doi: 10.1126/scitranslmed.3008680
138. Rimessi A, Bezzerri V, Patergnani S, Marchi S, Cabrini G, and Pinton P. Mitochondrial Ca2+-dependent NLRP3 activation exacerbates the Pseudomonas aeruginosa-driven inflammatory response in cystic fibrosis. Nat Commun. (2015) 6:6201. doi: 10.1038/ncomms7201
139. Rada B, Park JJ, Sil P, Geiszt M, and Leto TL. NLRP3 inflammasome activation and interleukin-1beta release in macrophages require calcium but are independent of calcium-activated NADPH oxidases. Inflammation Res. (2014) 63:821–30. doi: 10.1007/s00011-014-0756-y
140. Cantin AM. Cystic fibrosis lung disease and immunometabolism. Targeting the NLRP3 inflammasome. Am J Respir Crit Care Med. (2019) 200:1335–7. doi: 10.1164/rccm.201908-1558ED
141. Huus KE, Joseph J, Zhang L, Wong A, Aaron SD, Mah TF, et al. Clinical Isolates of Pseudomonas aeruginosa from Chronically Infected Cystic Fibrosis Patients Fail To Activate the Inflammasome during Both Stable Infection and Pulmonary Exacerbation. J Immunol. (2016) 196:3097–108. doi: 10.4049/jimmunol.1501642
142. Kay C, Wang R, Kirkby M, and Man SM. Molecular mechanisms activating the NAIP-NLRC4 inflammasome: Implications in infectious disease, autoinflammation, and cancer. Immunol Rev. (2020) 297:67–82. doi: 10.1111/imr.12906
143. Phuong MS, Hernandez RE, Wolter DJ, Hoffman LR, and Sad S. Impairment in inflammasome signaling by the chronic Pseudomonas aeruginosa isolates from cystic fibrosis patients results in an increase in inflammatory response. Cell Death Dis. (2021) 12:241. doi: 10.1038/s41419-021-03526-w
144. Graustein AD, Berrington WR, Buckingham KJ, Nguyen FK, Joudeh LL, Rosenfeld M, et al. Inflammasome genetic variants, macrophage function, and clinical outcomes in cystic fibrosis. Am J Respir Cell Mol Biol. (2021) 65:157–66. doi: 10.1165/rcmb.2020-0257OC
145. Holzinger D, Gieldon L, Mysore V, Nippe N, Taxman DJ, Duncan JA, et al. Staphylococcus aureus Panton-Valentine leukocidin induces an inflammatory response in human phagocytes via the NLRP3 inflammasome. J Leukoc Biol. (2012) 92:1069–81. doi: 10.1189/jlb.0112014
146. Perret M, Badiou C, Lina G, Burbaud S, Benito Y, Bes M, et al. Cross-talk between Staphylococcus aureus leukocidins-intoxicated macrophages and lung epithelial cells triggers chemokine secretion in an inflammasome-dependent manner. Cell Microbiol. (2012) 14:1019–36. doi: 10.1111/j.1462-5822.2012.01772.x
147. Kim JW, Ahn MH, Jung JY, Suh CH, and Kim HA. An update on the pathogenic role of neutrophils in systemic juvenile idiopathic arthritis and adult-onset still’s disease. Int J Mol Sci. (2021) 22. doi: 10.3390/ijms222313038
148. O’Neil LJ, Kaplan MJ, and Carmona-Rivera C. The role of neutrophils and neutrophil extracellular traps in vascular damage in systemic lupus erythematosus. J Clin Med. (2019) 8. doi: 10.3390/jcm8091325
149. Frade-Sosa B and Sanmarti R. Neutrophils, neutrophil extracellular traps, and rheumatoid arthritis: An updated review for clinicians. Reumatol Clin (Engl Ed). (2023) 19:515–26. doi: 10.1016/j.reuma.2023.08.001
150. Meijer M, Rijkers GT, and van Overveld FJ. Neutrophils and emerging targets for treatment in chronic obstructive pulmonary disease. Expert Rev Clin Immunol. (2013) 9:1055–68. doi: 10.1586/1744666X.2013.851347
151. Hu Q, Shi H, Zeng T, Liu H, Su Y, Cheng X, et al. Increased neutrophil extracellular traps activate NLRP3 and inflammatory macrophages in adult-onset Still’s disease. Arthritis Res Ther. (2019) 21:9. doi: 10.1186/s13075-018-1800-z
152. Guo C, Fu R, Wang S, Huang Y, Li X, Zhou M, et al. NLRP3 inflammasome activation contributes to the pathogenesis of rheumatoid arthritis. Clin Exp Immunol. (2018) 194:231–43. doi: 10.1111/cei.13167
153. Kahlenberg JM, Carmona-Rivera C, Smith CK, and Kaplan MJ. Neutrophil extracellular trap-associated protein activation of the NLRP3 inflammasome is enhanced in lupus macrophages. J Immunol. (2013) 190:1217–26. doi: 10.4049/jimmunol.1202388
154. Bonaventura A, Vecchie A, Abbate A, and Montecucco F. Neutrophil extracellular traps and cardiovascular diseases: an update. Cells. (2020) 9. doi: 10.3390/cells9010231
155. Campos J, Ponomaryov T, De Prendergast A, Whitworth K, Smith CW, Khan AO, et al. Neutrophil extracellular traps and inflammasomes cooperatively promote venous thrombosis in mice. Blood Adv. (2021) 5:2319–24. doi: 10.1182/bloodadvances.2020003377
156. Li J, Fang Z, Xu S, Rao H, Liu J, Lei K, et al. The link between neutrophils, NETs, and NLRP3 inflammasomes: The dual effect of CD177 and its therapeutic potential in acute respiratory distress syndrome/acute lung injury. Biomol BioMed. (2024) 24:798–812. doi: 10.17305/bb.2023.10101
157. Wang Y, Liu F, Chen L, Fang C, Li S, Yuan S, et al. Neutrophil extracellular traps (NETs) promote non-small cell lung cancer metastasis by suppressing lncRNA MIR503HG to activate the NF-kappaB/NLRP3 inflammasome pathway. Front Immunol. (2022) 13:867516. doi: 10.3389/fimmu.2022.867516
158. Ye X, Fang C, Hong W, Qian X, Yu B, Zhou B, et al. LncRNA MIR503HG regulates NETs-mediated NLRP3 inflammasome activation and NSCLC metastasis by enhancing the ubiquitination of C/EBPbeta. Clin Transl Med. (2025) 15:e70342. doi: 10.1002/ctm2.70342
159. Wright TK, Gibson PG, Simpson JL, McDonald VM, Wood LG, and Baines KJ. Neutrophil extracellular traps are associated with inflammation in chronic airway disease. Respirology. (2016) 21:467–75. doi: 10.1111/resp.12730
160. Yaqinuddin A and Kashir J. Novel therapeutic targets for SARS-CoV-2-induced acute lung injury: Targeting a potential IL-1beta/neutrophil extracellular traps feedback loop. Med Hypotheses. (2020) 143:109906. doi: 10.1016/j.mehy.2020.109906
161. Jin J, Zhao Y, Fang Y, Pan Y, Wang P, Fan Z, et al. Neutrophil extracellular traps promote the activation of the NLRP3 inflammasome and PBMCs pyroptosis via the ROS-dependent signaling pathway in Kawasaki disease. Int Immunopharmacol. (2025) 145:113783. doi: 10.1016/j.intimp.2024.113783
162. Zhao P, Zhu J, Bai L, Ma W, Li F, Zhang C, et al. Neutrophil extracellular traps induce pyroptosis of pulmonary microvascular endothelial cells by activating the NLRP3 inflammasome. Clin Exp Immunol. (2024) 217:89–98. doi: 10.1093/cei/uxae028
163. Cao T, Yuan X, Fang H, Chen J, Xue K, Li Z, et al. Neutrophil extracellular traps promote keratinocyte inflammation via AIM2 inflammasome and AIM2-XIAP in psoriasis. Exp Dermatol. (2023) 32:368–78. doi: 10.1111/exd.14711
164. McElvaney OJ, Zaslona Z, Becker-Flegler K, Palsson-McDermott EM, Boland F, Gunaratnam C, et al. Specific inhibition of the NLRP3 inflammasome as an antiinflammatory strategy in cystic fibrosis. Am J Respir Crit Care Med. (2019) 200:1381–91. doi: 10.1164/rccm.201905-1013OC
165. Clauzure M, Valdivieso AG, Dugour AV, Mori C, Massip-Copiz MM, Aguilar MA, et al. NLR family pyrin domain containing 3 (NLRP3) and caspase 1 (CASP1) modulation by intracellular Cl(-) concentration. Immunology. (2021) 163:493–511. doi: 10.1111/imm.13336
166. Hu S, Yu F, Ye C, Huang X, Lei X, Dai Y, et al. The presence of P2RX7 single nuclear polymorphism is associated with a gain of function in P2X7 receptor and inflammasome activation in SLE complicated with pericarditis. Clin Exp Rheumatol. (2020) 38:442–9.
167. Gabillard-Lefort C, Casey M, Glasgow AMA, Boland F, Kerr O, Marron E, et al. Trikafta Rescues CFTR and lowers monocyte P2X7R-induced inflammasome activation in cystic fibrosis. Am J Respir Crit Care Med. (2022) 205:783–94. doi: 10.1164/rccm.202106-1426OC
168. Balakrishnan A, Karki R, Berwin B, Yamamoto M, and Kanneganti TD. Guanylate binding proteins facilitate caspase-11-dependent pyroptosis in response to type 3 secretion system-negative Pseudomonas aeruginosa. Cell Death Discov. (2018) 4:3. doi: 10.1038/s41420-018-0068-z
169. Cohen TS and Parker D. Microbial pathogenesis and type III interferons. Cytokine Growth Factor Rev. (2016) 29:45–51. doi: 10.1016/j.cytogfr.2016.02.005
170. Lopez-Castejon G and Brough D. Understanding the mechanism of IL-1beta secretion. Cytokine Growth Factor Rev. (2011) 22:189–95. doi: 10.1016/j.cytogfr.2011.10.001
171. Forrest OA, Dobosh B, Ingersoll SA, Rao S, Rojas A, Laval J, et al. Neutrophil-derived extracellular vesicles promote feed-forward inflammasome signaling in cystic fibrosis airways. J Leukoc Biol. (2022) 112:707–16. doi: 10.1002/JLB.3AB0321-149R
172. Ahn EY and So MW. The pathogenesis of gout. J Rheum Dis. (2025) 32:8–16. doi: 10.4078/jrd.2024.0054
173. Sil P, Wicklum H, Surell C, and Rada B. Macrophage-derived IL-1beta enhances monosodium urate crystal-triggered NET formation. Inflammation Res. (2017) 66:227–37. doi: 10.1007/s00011-016-1008-0
174. Mitroulis I, Kambas K, Chrysanthopoulou A, Skendros P, Apostolidou E, Kourtzelis I, et al. Neutrophil extracellular trap formation is associated with IL-1beta and autophagy-related signaling in gout. PloS One. (2011) 6:e29318. doi: 10.1371/journal.pone.0029318
175. Roesch EA, Rahmaoui A, Lazarus RA, and Konstan MW. The continuing need for dornase alfa for extracellular airway DNA hydrolysis in the era of CFTR modulators. Expert Rev Respir Med. (2024) 18:677–91. doi: 10.1080/17476348.2024.2394694
176. Cavalli G and Dinarello CA. Anakinra therapy for non-cancer inflammatory diseases. Front Pharmacol. (2018) 9:1157. doi: 10.3389/fphar.2018.01157
177. Cohen SB. The use of anakinra, an interleukin-1 receptor antagonist, in the treatment of rheumatoid arthritis. Rheum Dis Clin North Am. (2004) 30:365–80. doi: 10.1016/j.rdc.2004.01.005
179. Garcia MS, Madrid-Carbajal CJ, Pelaez A, Moreno RMG, Alonso EF, Garcia BP, et al. The role of triple CFTR modulator therapy in reducing systemic inflammation in cystic fibrosis. Lung. (2025) 203:55. doi: 10.1007/s00408-025-00806-6
Keywords: cystic fibrosis - CF, neutrophil extracellular traps (NET), neutrophil, lung disease, inflammasome, interleukin-1 (IL-1β)
Citation: Fantone KM, Gokanapudi N and Rada B (2025) Neutrophil extracellular traps and interleukin-1β in cystic fibrosis lung disease. Front. Immunol. 16:1595994. doi: 10.3389/fimmu.2025.1595994
Received: 19 March 2025; Accepted: 26 June 2025;
Published: 28 July 2025.
Edited by:
Mahadevappa Hemshekhar, University of Manitoba, CanadaReviewed by:
Giulio Cabrini, University of Ferrara, ItalyKartiga Natarajan, Houston Methodist Research Institute, United States
Copyright © 2025 Fantone, Gokanapudi and Rada. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Balázs Rada, cmFkYWJAdWdhLmVkdQ==