Browsing by Subject "Inflammasome"

Now showing 1 - 13 of 13
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    4-octyl itaconate reduces NLRP3 inflammasome constitutive activation with cryopyrin-associated periodic syndrome p.R262W, p.D305N and p.T350M variants
    (Springer, Birkhäuser Verlag, 2025-05-23) Molina-López, Cristina; Hurtado-Navarro, Laura; O'Neill, Luke A.J.; Pelegrin, Pablo; Bioquímica y Biología Molecular B e Inmunología
    Cryopyrin-associated periodic syndrome (CAPS) is a condition characterized by dominant genetic variants in the NLRP3 gene, leading to the formation of constitutively active inflammasomes. These inflammasomes play a crucial role in the inflammatory episodes experienced by CAPS patients, primarily driven by the production of interleukin (IL)-1. Although treatment with IL-1 blockers is effective for CAPS, some patients develop refractory responses and adverse reactions to these therapies. Consequently, there is a need for novel treatments for CAPS patients. Promising candidates are the derivatives of itaconate, which have been shown to impair NLRP3 inflammasome activation and IL-1 release in blood mononuclear cells from CAPS patients. In this study, we provide insight into the inhibitory mechanisms of the itaconate derivative 4-octyl itaconate (4-OI) on NLRP3 with different (p.R262W, p.D305N and p.T350M) gain-of-function mutations associated with CAPS. Notably, 4-OI effectively blocks the basal auto-activation of the inflammasome formed by NLRP3 p.R262W, p.D305N and p.T350M variants, resulting in reduced caspase-1 activation, gasdermin D processing, and IL-18 release. Furthermore, after lipopolysaccharide priming of macrophages, 4-OI also decreases IL-1 gene expression and release. Overall, 4-OI impairs CAPS p.D305N-associated inflammasome function at multiple levels, and therapeutic agents based on itaconate could be a promising therapeutic approach to managing inflammatory episodes in CAPS patients carrying p.R262W, p.D305N or p.T350M variants.
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    Chloride regulates dynamic NLRP3-dependent ASC oligomerization and inflammasome priming
    (National Academy of Sciences, 2018-09-19) Martín Sánchez, María Rosario Fátima; Pelegrín Vivancos, Pablo; Green, Jack Peter; Yu, Shi; Lopez-Castejon, Gloria; Lawrence, Catherine B.; Brough, David; Farmacología
    The NLRP3 inflammasome is an important regulator of inflammation and immunity. It is a multi-molecular platform formed within cells that facilitates the activation of pro-inflammatory caspases to drive secretion of cytokines such as IL-1β. Knowledge of the mechanisms regulating formation of the NLRP3-inflammasome is incomplete. Here we report Cl- channel dependent formation of dynamic ASC oligomers and inflammasome specks that remain inactive in the absence of K+ efflux. Formed after Cl- efflux exclusively ASC specks are NLRP3 dependent, reversible, and inactive, though they further prime inflammatory responses accelerating and enhancing release of IL-1β in response to a K+ efflux inducing stimulus. NEK7 is a specific K+ sensor and does not associate with NLRP3 under conditions stimulating exclusively Cl- efflux, but does after K+ efflux, activating the complex driving inflammation. Our investigation delivers new mechanistic understanding into inflammasome activation and the regulation of inflammatory responses.
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    Danger signals released during cold ischemia storage activate NLRP3 inflammasome in myeloid cells and influence early allograft function in liver transplantation
    (Elsevier, 2022-12-19) Lucas-Ruiz, Fernando; Mateo, Sandra V.; Jover-Aguilar, Marta; Alconchel, Felipe; Martínez-Alarcón, Laura; Torre-Minguela, Carlos de; Vidal-Correoso, Daniel; Villalva-López, Francisco; López-López, Víctor; Rios-Zambudio, Antonio; Pons Miñano, José Antonio; Ramírez, Pablo; Baroja-Mazo, Alberto; Pelegrín Vivancos, Pablo; Medicina
    Background Innate immunity plays a fundamental role in solid organ transplantation. Myeloid cells can sense danger signals or DAMPs released after tissue or cell damage, such as during ischemia processes. This study aimed to identify DAMPs released during cold ischemia storage of human liver and analyze their ability to activate the inflammasome in myeloid cells and the possible implications in terms of short-term outcomes of liver transplantation. Methods 79 samples of organ preservation solution (OPS) from 79 deceased donors were collected after cold static storage. We used different analytical methods to measure DAMPs in these end-ischemic OPS (eiOPS) samples. We also used eiOPS in the human macrophage THP-1 cell line and primary monocyte cultures to study inflammasome activation. Findings Different DAMPs were identified in eiOPS, several of which induced both priming and activation of the NLRP3 inflammasome in human myeloid cells. Cold ischemia time and donation after circulatory death negatively influenced the DAMP signature. Moreover, the presence of oligomeric inflammasomes and interleukin-18 in eiOPS correlated with early allograft dysfunction in liver transplant patients. Interpretation DAMPs released during cold ischemia storage prime and activate the NLRP3 inflammasome in liver macrophages after transplantation, inducing a pro-inflammatory environment that will complicate the outcome of the graft. The use of pharmacological blockers targeting DAMPs or the NLRP3 inflammasome in liver ischemia during static cold storage or through extracorporeal organ support could be a suitable strategy to increase the success of liver transplantation.
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    Decreased survival in patients with pancreatic cancer may be associated with an increase in histopathological expression of inflammasome marker NLRP3
    (Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Fraile Martínez, Oscar; García Montero, Cielo; Pekarek, Leonel; Saz, Jose V.; Álvarez Mon, Miguel Ángel; Barrena Blázquez, Silvestra; García Honduvilla, Natalio; Buján, Julia; Asúnsolo, Ángel; Coca, Santiago; Álvarez Mon, Melchor; Guijarro, Luis G.; Saez, Miguel A.; Ortega, Miguel A.
    Pancreatic cancer is a malignant neoplasm that, despite its low frequency, has a 5-year survival rate of less than 10%. The study of different histopathological markers has allowed a better understanding of the onset and development of this type of tumor as well as facilitating an approach to clinical variables based on their diagnostic, prognostic, and predictive value. In this sense, the NLRP3 protein of the inflammasome has been shown to be a component of great relevance in the initiation and progression of pancreatic cancer, although the value of this biomarker in patients has not yet been clarified. In this study, we selected 41 patients with pancreatic cancer and followed them for 60 months (5 years), evaluating their NLRP3 expression using immunohistochemical techniques. Furthermore, by performing Kaplan-Meier curves, we evaluated the survival of these patients in relation to their NLRP3 expression. Our results show that a significant percentage of our cohort had high expression of this component (90.74%) and that there is an inverse relationship between the expression of NLRP3 and patient survival. High levels of NLRP3 expression are related to lower survival and worse prognosis in these patients, possibly due to an ineffective immune system response and increased tumor-promoted inflammation. Future studies should be aimed at confirming these results in larger groups and evaluating various clinical strategies based on this knowledge.
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    Development of an Acrylate Derivative Targeting the NLRP3 Inflam- masome for the Treatment of Inflammatory Bowel Disease
    (ACS Publications, 2017) Cocco, Mattia; Pellegrini, Carolina; Martínez-Banaclocha, Helios; Giorgis, Marta; Marini, Elisabetta; Costale, Annalisa; Miglio, Gianluca; Fornai, Matteo; Antonioli, Luca; López-Castejón, Gloria; Angosto, Diego; Hafner-Bratkovic, Iva; Regazzoni, Luca; Blandizzi, Corrado; Pelegrin, Pablo; Bertinaria, Massimo; Tapia Abellán, Ana; Bioquímica y Biología Molecular B e Inmunología
    Pharmacological inhibition of NLRP3 inflammasome activation may offer a new option in the treatment of Inflamma- tory Bowel Disease (IBD). In this work, we report the design, the synthesis, and the biological screening of a series of acrylate derivatives as NLRP3 inhibitors. The in vitro determination of reactivity, cytotoxicity, NLRP3 ATPase inhibition, and antipyroptotic properties allowed the selection of 11 (INF39), a stable, non toxic compound inhibiting interleukin 1β release from macrophages. Bioluminescence resonance energy transfer experiments proved that this compound was able to directly interfere with NLRP3 acti- vation in cells. In vivo studies confirmed the ability of the selected lead to alleviate the effects of DNBS-induced colitis in rats after oral administration.
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    Gasdermin D mediates a fast transient release of ATP after NLRP3 inflammasome activation before ninjurin 1-induced lytic cell death
    (Cell Press, 2025) Schachter, Julieta; Guijarro, Adriana; Angosto-Bazarra, Diego; Pinilla, Miriam; Hurtado-Navarro, Laura; Meunier, Etienne; Perez-Oliva, Ana Belen; Schwarzbaum, Pablo J; Pelegrín Vivancos, Pablo; Pinilla Marquinez, Míriam; Bioquímica y Biología Molecular B e Inmunología
    Pyroptosis is a lytic cell death triggered by the cleavage of gasdermin (GSDM) proteins and subsequent pore formation by the N-terminal domain oligomerization in the plasma membrane. GSDMD is cleaved by caspase-1/-4/-5/-11 upon inflammasome activation and mediates interleukin (IL)-1β and IL-18 release. GSDMD pores favor ninjurin 1 (NINJ1)-induced plasma membrane rupture and cell death. Here, we demonstrate that GSDMD mediates early ATP release upon NLRP3 inflammasome activation independently of NINJ1, occurring before IL-1β release and cell death and constituting an early danger signal. The release of ATP is a transient signal terminated before the cells continue to permeabilize and die. The different N termini of GSDMA to -E are also able to release ATP and induce monocyte migration toward pyroptotic cells. This study reveals ATP release as an early and transient danger signal depending on GSDMD plasma membrane permeabilization, independently of the late stages of lytic cell death.
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    Luteoloside ameliorates sepsis-induced acute lung injury via AMPK-ULK1 pathway-mediated autophagy
    (2025) Min Huang; Hang Qi; Cheng Liu; Hongzhou Xu; Liang Cai; Bo Xu; Biología Celular e Histología
    Background. Septic patients are at high risk of acute lung injury (ALI). Luteoloside is a flavonoid isolated from natural herbs and has many beneficial effects. This study aimed to investigate the protective role of luteoloside in sepsis-induced ALI. Methods. Sepsis was induced by cecal ligation and puncture (CLP) in C57BL/6 mice. Inflammation was induced by lipopolysaccharide (LPS) in MLE-12 cells. The survival rate over 12 days, histological changes in lung and heart, pulmonary edema, vascular leakage, hypoxemia, and inflammation were examined. Apoptosis was detected by TUNEL staining in vivo and flow cytometry in vitro. The levels of autophagy-related proteins, the AMPK/ULK1 pathway, and the NLRP3 inflammasome were evaluated by western blotting. Cell viability was estimated by MTT assays. LC3 expression was evaluated by immunofluorescence staining. Results. Luteoloside attenuated lung and cardiac injury, pulmonary edema, vascular leakage, hypoxemia, and inflammation and improved the survival of septic mice. Luteoloside (20 mg/kg) had no toxic effect on the heart, liver, spleen, and kidney in normal mice. Luteoloside enhanced autophagy to inhibit apoptosis in vivo and in vitro, and autophagy induction was responsible for the protective effect of luteoloside. Luteoloside activated AMPK/ULK1 signaling to enhance autophagy. Luteoloside also inhibited the activation of the NLRP3 inflammasome in LPS-challenged MLE-12 cells. Conclusion. Overall, luteoloside activates AMPK/ ULK1 signaling to stimulate autophagy, thereby inhibiting apoptosis and alleviating sepsis-induced ALI.
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    Neutrophils mediate Salmonella Typhimurium clearance through the GBP4 inflammasome-dependent production of prostaglandins
    (Nature Research, 2016-07-01) Martín Sánchez, María Rosario Fátima; Tyrkalska, Sylwia D.; Candel Camacho, Sergio; Angosto, Diego; Gómez Abellán, Victoria; García Moreno, Diana; Zapata Pérez, Rubén; Sánchez Ferrer, Álvaro; Pelegrín Vivancos, Pablo; Mulero Méndez, Victoriano Francisco; Sepulcre Cortés, María Pilar; Farmacología
    Inflammasomes are cytosolic molecular platforms that alert the immune system about thepresence of infection. Here we report that zebrafish guanylate-binding protein 4 (Gbp4),an IFNg-inducible GTPase protein harbouring a C-terminal CARD domain, is required for theinflammasome-dependent clearance of Salmonella Typhimurium (ST) by neutrophils in vivo.Despite the presence of the CARD domain, Gbp4 requires the universal inflammasomeadaptor Asc for mediating its antibacterial function. In addition, the GTPase activity of Gbp4is indispensable for inflammasome activation and ST clearance. Mechanistically, neutrophilsare recruited to the infection site through the inflammasome-independent production of thechemokine (CXC motif) ligand 8 and leukotriene B4, and then mediate bacterial clearancethrough the Gbp4 inflammasome-dependent biosynthesis of prostaglandin D2. Our resultspoint to GBPs as key inflammasome adaptors required for prostaglandin biosynthesis andbacterial clearance by neutrophils and suggest that transient activation of the inflammasomemay be used to treat bacterial infections.
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    NLRP3 lacking the leucine-rich repeat domain can be fully activated via the canonical inflammasome pathway
    (Nature, 2018) Hafner-Bratkovic, Iva; Susjan, Petra; Lainscek, Dusko; Cerovic, Kosta; Kadunc, Lucija; Angosto-Bazarra, Diego; Pelegrin, Pablo; Jerala, Roman; Tapia Abellán, Ana; Bioquímica y Biología Molecular B e Inmunología
    NLRP3 is a cytosolic sensor triggered by different pathogen- and self-derived signals that plays a central role in a variety of pathological conditions, including sterile inflammation. The leucine- rich repeat domain is present in several innate immune receptors, where it is frequently responsible for sensing danger signals and regulation of activation. We show by reconstitution of truncated and chimeric variants into NLRP3-/- macrophages that the leucine-rich repeat domain is dispensable for activation and self-regulation of NLRP3 by several different triggers. The pyrin domain on the other hand is required to maintain NLRP3 in the inactive conformation. A fully responsive minimal NLRP3 truncation variant reconstituted peritonitis in NLRP3-/- mice. We demonstrate that in contrast to pathogen-activated NLRC4, the constitutively active NLRP3 molecule cannot engage wild-type NLRP3 molecules in a self-catalytic oligomerization. This lack of signal amplification is likely a protective mechanism to decrease sensitivity to endogenous triggers to impede autoinflammation.
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    Pathogenic NLRP3 mutants form constitutively active inflammasomes resulting in immune-metabolic limitation of IL-1β production
    (Nature Research, 2024-02-06) Molina-López, Cristina; Hurtado-Navarro, Laura; Garcia, Carlos J.; Angosto-Bazarra, Diego; Vallejo, Fernando; Marques-Soares, Joana R.; Vargas, Carmen; Bujan-Rivas, Segundo; Tomás-Barberán, Francisco A.; Arostegui, Juan I.; Pelegrín Vivancos, Pablo; Tapia Abellán, Ana; Bioquímica y Biología Molecular B e Inmunología
    Cryopyrin-associated periodic syndrome (CAPS) is an autoinflammatory condition resulting from monoallelic NLRP3 variants that facilitate IL-1b production. Although these are gain-of-function variants characterised by hypersensitivity to cell priming, patients with CAPS and animal models of the disease may present inflammatory flares without identifiable external triggers. Here we find that CAPS-associated NLRP3 variants are forming constitutively active inflammasome, which induce increased basal cleavage of gasdermin D, IL-18 release and pyroptosis, with a concurrent basal pro-inflammatory gene expression signature, including the induction of nuclear receptors 4A. The constitutively active NLRP3-inflammasome is responsive to the selective NLRP3 inflammasome inhibitor MCC950 and its activation is regulated by deubiquitination. Despite their preactivated state, the CAPS inflammasomes are responsive to activation of the NF-kB pathway. NLRP3-inflammasomes with CAPS-associated variants affect the immunometabolism of the myeloid compartment, leading to disruptions in lipids and amino acid pathways and impaired glycolysis, limiting IL-1b production. In summary, NLRP3 variants causing CAPS form a constitutively active inflammasome inducing pyroptosis and IL-18 release without cell priming, which enables the host's innate defence against pathogens while also limiting IL-1b–dependent inflammatory episodes through immunometabolism modulation.
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    Priming is dispensable for NLRP3 inflammasome activation in human monocytes in vitro
    (Frontiers Media, 2020-09-30) Martín Sánchez, María Rosario Fátima; Gritsenko, Anna; Yu, Shi; Díaz del Olmo, Inés; Nichols, Eva María; Davis, Daniel M.; Brough, David; López Castejón, Gloria; Farmacología
    Interleukin (IL)-18 and IL-1b are potent pro-inflammatory cytokines that contribute toinflammatory conditions such as rheumatoid arthritis and Alzheimer’s disease. They areproduced as inactive precursors that are activated by large macromolecular complexescalled inflammasomes upon sensing damage or pathogenic signals. NLRP3inflammasome activation is regarded to require a priming step that causes NLRP3 andIL-1b gene upregulation, and also NLRP3 post-translational licencing. A subsequentactivation step leads to the assembly of the complex and the cleavage of pro-IL-18 andpro-IL-1b by caspase-1 into their mature forms, allowing their release. Here we show thathuman monocytes, but not monocyte derived macrophages, are able to form canonicalNLRP3 inflammasomes in the absence of priming. NLRP3 activator nigericin caused theprocessing and release of constitutively expressed IL-18 in an unprimed setting. This wasmediated by the canonical NLRP3 inflammasome that was dependent on K + and Cl−efflux and led to ASC oligomerization, caspase-1 and Gasdermin-D (GSDMD) cleavage.IL-18 release was impaired by the NLRP3 inhibitor MCC950 and by the absence ofNLRP3, but also by deficiency of GSDMD, suggesting that pyroptosis is the mechanism ofrelease. This work highlights the readiness of the NLRP3 inflammasome to assemble inthe absence of priming in human monocytes and hence contribute to the very early stagesof the inflammatory response when IL-1b has not yet been produced. It is important toconsider the unprimed setting when researching the mechanisms of NLRP3 activation, asto not overshadow the pathways that occur in the absence of priming stimuli, which mightonly enhance this response.
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    Sensing low intracellular potassium by NLRP3 results in a stable open structure that promotes inflammasome activation
    (American Association for the Advancement of Science, 2021-09-15) Angosto-Bazarra, Diego; Alarcón-Vila, Cristina; Baños, Maria C; Hafner-Bratkovič, Iva; Oliva, Baldomero; Pelegrín Vivancos, Pablo; Tapia Abellán, Ana; Bioquímica y Biología Molecular B e Inmunología
    The NLRP3 inflammasome is activated in response to a wide range of stimuli and drives diverse inflammatory diseases. The decrease of intracellular K+ concentration is a minimal upstream signal to most of the different NLRP3 activation models. Here we found that cellular K+ efflux induces a stable structural change in the inactive NLRP3 promoting an open conformation as a step preceding activation. This conformational change is facilitated by the presence of the specific NLRP3 FISNA domain and a unique flexible linker sequence between the PYD and FISNA domains. This linker is also important to facilitate the ensemble of NLRP3PYD into a seed structure for ASC oligomerization. The introduction of the NLRP3 PYD-linker-FISNA sequence into NLRP6 resulted in a chimeric receptor able to be activated by K+ efflux-specific NLRP3 activators and promoted an in vivo inflammatory response to uric acid crystals. Our results establish that the N-terminal sequence between PYD and NACHT domain of NLRP3 is key for inflammasome activation.
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    The NLRP3 inflammasome is released as a particulate danger signal that amplifies the inflammatory response
    (Nature Research, 2014-08) Baroja Mazo, Alberto; Gómez, Ana I.; Amores Iniesta, Joaquín; Compan, Vincent; Barberà Cremades, María; Yagüe, Jordi; Ruiz Ortiz, Estibaliz; Antón, Jordi; Buján, Segundo; Coullin, Isabelle; Brough, David; Arostegui, Juan I.; Martínez Cáceres, Carlos Manuel; Martín Sánchez, María Rosario Fátima; Pelegrín Vivancos, Pablo; Anatomía y Anatomía Patológica Comparadas
    Assembly of the NLRP3 inflammasome activates caspase-1 and mediates the processing and release of the leaderless cytokine IL-1β and thereby serves a central role in the inflammatory response and in diverse human diseases. Here we found that upon activation of caspase-1, oligomeric NLRP3 inflammasome particles were released from macrophages. Recombinant oligomeric protein particles composed of the adaptor ASC or the p.D303N mutant form of NLRP3 associated with cryopyrin-associated periodic syndromes (CAPS) stimulated further activation of caspase-1 extracellularly, as well as intracellularly after phagocytosis by surrounding macrophages. We found oligomeric ASC particles in the serum of patients with active CAPS but not in that of patients with other inherited autoinflammatory diseases. Our findings support a model whereby the NLRP3 inflammasome, acting as an extracellular oligomeric complex, amplifies the inflammatory response.