Diverse Activators of the NLRP3 Inflammasome Promote IL-1β Secretion by Triggering Necrosis

Cullen, Sean P.; Kearney, Conor J.; Clancy, Danielle M.; Martin, Séamus J.

doi:10.1016/j.celrep.2015.05.003

Cited by 199 publications

(188 citation statements)

References 36 publications

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“…As such, we could not conclusively determine if IL-1β secretion was an active or passive process during necroptosis. This question of whether IL-1β can be actively secreted from cells after caspase-1 activation remains controversial (20,23,(35)(36)(37). These experiments nonetheless suggested that NLRP3 activation could be cell-intrinsic, because glycine treatment had no effect on mature IL-1β secretion following MLKL activation.…”

Section: Discussionmentioning

confidence: 99%

Active MLKL triggers the NLRP3 inflammasome in a cell-intrinsic manner

Conos

Chen

Nardo

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

366

298

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Necroptosis is a physiological cell suicide mechanism initiated by receptor-interacting protein kinase-3 (RIPK3) phosphorylation of mixed-lineage kinase domain-like protein (MLKL), which results in disruption of the plasma membrane. Necroptotic cell lysis, and resultant release of proinflammatory mediators, is thought to cause inflammation in necroptotic disease models. However, we previously showed that MLKL signaling can also promote inflammation by activating the nucleotide-binding oligomerization domain (NOD)-like receptor protein 3 (NLRP3) inflammasome to recruit the adaptor protein apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC) and trigger caspase-1 processing of the proinflammatory cytokine IL-1β. Here, we provide evidence that MLKL-induced activation of NLRP3 requires (i) the death effector four-helical bundle of MLKL, (ii) oligomerization and association of MLKL with cellular membranes, and (iii) a reduction in intracellular potassium concentration. Although genetic or pharmacological targeting of NLRP3 or caspase-1 prevented MLKLinduced IL-1β secretion, they did not prevent necroptotic cell death. Gasdermin D (GSDMD), the pore-forming caspase-1 substrate required for efficient NLRP3-triggered pyroptosis and IL-1β release, was not essential for MLKL-dependent death or IL-1β secretion. Imaging of MLKL-dependent ASC speck formation demonstrated that necroptotic stimuli activate NLRP3 cell-intrinsically, indicating that MLKL-induced NLRP3 inflammasome formation and IL-1β cleavage occur before cell lysis. Furthermore, we show that necroptotic activation of NLRP3, but not necroptotic cell death alone, is necessary for the activation of NF-κB in healthy bystander cells. Collectively, these results demonstrate the potential importance of NLRP3 inflammasome activity as a driving force for inflammation in MLKLdependent diseases.aspase-dependent apoptotic cell death is required for mammalian development and the prevention of autoimmune and neoplastic diseases. Programmed cell death can also act to eliminate pathogen-infected cells, with recent studies highlighting how targeted apoptosis-inducing anticancer compounds can treat viral and intracellular bacterial infections (1, 2). On the other hand, the recently characterized caspase-independent necroptotic cell death pathway is dispensable for organism development but, like apoptosis, can be triggered to kill cells harboring pathogenic microbes (3). A number of studies have also reported how pathological activation of necroptotic signaling may contribute to diverse disease states, such as ischemia-reperfusion injury, atherosclerosis, and liver disease, presumably through cell death and the release of proinflammatory mediators (4).The execution of necroptosis is dependent on receptor interacting serine-threonine protein kinase 3 (RIPK3) phosphorylation of mixed-lineage kinase domain-like protein (MLKL), and MLKL's association with, and disruption of, plasma membrane integrity (5).In the absence of caspase activit...

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Section: Discussionmentioning

confidence: 99%

Active MLKL triggers the NLRP3 inflammasome in a cell-intrinsic manner

Conos

Chen

Nardo

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

366

298

View full text Add to dashboard Cite

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“…TNF-α can activate pro-apoptotic pathways 27 and production and release of mature Il-1β may occur when the permeability of the cell membrane is increased, potentially triggering cell necrosis 28 . To determine if bactericidal antibiotic treatment is also associated with cell death, we measured excretion of LDH by SNECs - LDH is released by dying mammalian cells and has been used as a marker of cytotoxicity.…”

Section: Resultsmentioning

confidence: 99%

“…In addition to their roles in inflammation, both TNF-α and IL-1β are also associated with activation of pro-apoptotic or necrosis death pathways 27,28 . In this study, we also demonstrate that treatment with levofloxacin and amoxicillin for 48 and 96-hours is cytotoxic (as measured by LDH secretion) to human SNECs compared to untreated SNECs (Figure 5).…”

Section: Discussionmentioning

confidence: 99%

Bactericidal antibiotics promote reactive oxygen species formation and inflammation in human sinonasal epithelial cells

Kohanski

Tharakan

Lane

et al. 2015

Int Forum Allergy Rhinol

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Background Bactericidal antibiotics have been shown to stimulate reactive oxygen species (ROS) formation in mammalian cells through mitochondrial dysfunction. This results in oxidative tissue damage that may have negative consequences for long-term antibiotic use. Antibiotics are widely and heavily used in the treatment of acute and chronic sinusitis, however the relationship between antibiotics and ROS formation in sinonasal epithelial cells (SNECs) has not yet been demonstrated. Methods Human SNECs were collected from patients during endoscopic sinus surgery and grown in culture at the air-liquid interface. Differentiated SNECs were stimulated with the bactericidal antibiotics amoxicillin and levofloxacin and the bacteriostatic antibiotic clarithromycin for 24-hours. Reactive oxygen species were quantified via fluorescence. Cell death was quantified by LDH secretion. Expression of inflammatory markers such as TNF-α and Nrf2 mediated antioxidant genes were measured by RT-PCR. Results Cultured SNECs treated with the bactericidal antibiotics amoxicillin and levofloxacin resulted in a significant increase in production of ROS (p<0.05) and secretion of LDH (p<0.05). The increase in ROS formation correlated with an increase in expression of Nrf-2 mediated antioxidant genes as well as the expression and production of pro-inflammatory cytokine TNF- α, and IL-1β (p<0.05). SNECs treated with clarithromycin did not demonstrate statistically significant increases in ROS or pro-inflammatory cytokine production. Discussion In this study, we demonstrate that treatment of cultured human SNECs with bactericidal antibiotics leads to formation of ROS with an associated increase in inflammatory and antioxidant gene expression and cell death. This suggests that long term or inappropriate antibiotic use in the treatment of sinusitis, may result in oxidative tissue damage to the sinonasal epithelium. Future studies will explore the clinical implications of such damage to the sinonasal epithelium.

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“…[50][51] It was shown that only when cell membrane disruption and necrotic cell death was initiated by wellknown activators of the NLRP3 inflammasome, IL-1β could be released from the cytosol. 54 Corroborating this finding and underpinning the importance of plasma membrane rupture, cleavage of gasdermin D by both caspase-1 and caspase-4/-5/-11 appeared crucial for the execution of pyroptosis and the release of mature IL-1β. [55][56][57] An alternative (or integrated) pathway of execution of pyroptosis could be the caspase-11-dependent cleavage of pannexin-1, which leads to ATP release and subsequent purinergic P2X7-receptor-mediated plasma membrane permeabilization.…”

Section: Pyroptosis: Inflammatory Cell Deathmentioning

confidence: 92%