Cellular Models and Assays to Study NLRP3 Inflammasome Biology

Zito, Giovanni; Buscetta, Marco; Cimino, Maura; Dino, Paola; Bucchieri, Fabio; Cipollina, Chiara

doi:10.3390/ijms21124294

Cited by 47 publications

(27 citation statements)

References 134 publications

(220 reference statements)

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“…Therefore, THP-1 cells have been widely used as a suitable in vitro model of human macrophages in mechanistic studies of external stimuli. Macrophages are used as model cell lines to research inflammasome activation for therapeutic and/or mechanistic studies [25]. The upper airways, including the nasopharynx and paranasal sinuses, have been shown to be a silent reservoir for P. aeruginosa [26].…”

Section: Discussionmentioning

confidence: 99%

Benzyl Isothiocyanate Attenuates Inflammasome Activation in Pseudomonas aeruginosa LPS-Stimulated THP-1 Cells and Exerts Regulation through the MAPKs/NF-κB Pathway

Park

Lee

et al. 2022

IJMS

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Inflammasomes are a group of intracellular multiprotein platforms that play important roles in immune systems. Benzyl isothiocyanate (BITC) is a constituent of cruciferous plants and has been confirmed to exhibit various biological activities. The modulatory effects of BITC on inflammasome-mediated interleukin (IL)-1β expression and its regulatory mechanisms in Pseudomonas aeruginosa (P. aeruginosa) LPS/ATP-stimulated THP-1 cells was investigated. Monocytic THP-1 cells were treated with phorbol myristate acetate (PMA) to induce differentiation into macrophages. Enzyme-linked immunosorbent assays (ELISA) were performed to measure the levels of IL-1β produced in P. aeruginosa LPS/ATP-exposed THP-1 cells. Western blotting was performed to examine the BITC modulatory mechanisms in inflammasome-mediated signaling pathways. BITC inhibited IL-1β production in P. aeruginosa LPS/ATP-induced THP-1 cells. BITC also inhibited activation of leucine-rich repeat protein-3 (NLRP3) and caspase-1 in P. aeruginosa LPS/ATP-induced THP-1 cells. Furthermore, we show that mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) activation in P. aeruginosa LPS was attenuated by BITC. These BITC-mediated modulatory effects on IL-1β production may have therapeutic potential for inflammasome-mediated disorders such as a nasal polyp.

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Section: Discussionmentioning

confidence: 99%

Benzyl Isothiocyanate Attenuates Inflammasome Activation in Pseudomonas aeruginosa LPS-Stimulated THP-1 Cells and Exerts Regulation through the MAPKs/NF-κB Pathway

Park

Lee

et al. 2022

IJMS

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“…Different techniques could be used to study NLRP3 inhibitors [ 47 ]; the use of differentiated THP-1 cells treated with NLRP3 activating stimuli is suitable for the screening of large series of compounds [ 48 ]. All the synthesised compounds ( Figure 2 ) were evaluated for their NLRP3 inhibitory activity by measuring their ability to prevent NLRP3-dependent pyroptosis in differentiated THP-1 cells.…”

Section: Resultsmentioning

confidence: 99%

Chemical Modulation of the 1-(Piperidin-4-yl)-1,3-dihydro-2H-benzo[d]imidazole-2-one Scaffold as a Novel NLRP3 Inhibitor

et al. 2021

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In the search for new chemical scaffolds able to afford NLRP3 inflammasome inhibitors, we used a pharmacophore-hybridization strategy by combining the structure of the acrylic acid derivative INF39 with the 1-(piperidin-4-yl)1,3-dihydro-2H-benzo[d]imidazole-2-one substructure present in HS203873, a recently identified NLRP3 binder. A series of differently modulated benzo[d]imidazole-2-one derivatives were designed and synthesised. The obtained compounds were screened in vitro to test their ability to inhibit NLRP3-dependent pyroptosis and IL-1β release in PMA-differentiated THP-1 cells stimulated with LPS/ATP. The selected compounds were evaluated for their ability to reduce the ATPase activity of human recombinant NLRP3 using a newly developed assay. From this screening, compounds 9, 13 and 18, able to concentration-dependently inhibit IL-1β release in LPS/ATP-stimulated human macrophages, emerged as the most promising NLRP3 inhibitors of the series. Computational simulations were applied for building the first complete model of the NLRP3 inactive state and for identifying possible binding sites available to the tested compounds. The analyses led us to suggest a mechanism of protein–ligand binding that might explain the activity of the compounds.

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“…We used acute glucose shift conditions as changes between 5 mM and 25 mM glucose to emulate the commonly observed glycemic variability owing to postprandial hyperglycemia in patients with type 2 diabetes, rather than rare severe hypoglycemia. Owing to the difficulties in collecting human primary monocytes, the THP-1 monocyte-like cell line, derived from acute monocytic leukemia, has been extensively used in the study of NLRP3 inflammasome biology, despite its tumoral origin [28]. Emerging evidence has indicated that the activation of the NLRP3 inflammasome might lead to the maturation and secretion of IL-1β, which has been implicated in the pathological development of type 2 diabetes and cardiovascular diseases [29].…”

Section: Discussionmentioning

confidence: 99%

Acute Glucose Shift Induces the Activation of the NLRP3 Inflammasome in THP-1 Cells

Lee

Kang

Kim

et al. 2021

IJMS

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We aimed to investigate the effect of acute glucose shift on the activation of the NLRP3 inflammasome, IL-1β secretion, and underlying signaling pathways in THP-1 cells. THP-1 cells were divided into four groups and exposed to the following glucose concentrations for 24 h: constant normal glucose (NG, 5.5 mM), constant high glucose (HG, 25 mM), normal to high glucose shift (NG-to-HG, 5.5 to 25 mM), and high to normal glucose shift (HG-to-NG, 25 to 5.5 mM). Cell viability, oxidative stress, and the levels of NLRP3 inflammasome components were assessed. Both directions of the acute glucose shift increased the activation of the NLRP3 inflammasome, generation of reactive oxygen species (ROS), and expression of phosphorylated p38 MAPK, JNK, and NF-κB compared with either constant NG or HG. Treatment with N-acetylcysteine, a pharmacological antioxidant, inhibited the acute glucose shift-induced generation of ROS, activation of NLRP3 inflammasome, and upregulation of MAPK-NF-κB. Further analysis using inhibitors of p38 MAPK, JNK, and NF-κB indicated that acute glucose shifts promoted IL-1β secretion by activating the signaling pathway in a ROS-MAPK-NF-κB-NLRP3 inflammasome in THP-1 cells. These findings suggested that acute changes in glucose concentration might cause monocyte inflammation, which is associated with diabetic complications.

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Cellular Models and Assays to Study NLRP3 Inflammasome Biology

Cited by 47 publications

References 134 publications

Benzyl Isothiocyanate Attenuates Inflammasome Activation in Pseudomonas aeruginosa LPS-Stimulated THP-1 Cells and Exerts Regulation through the MAPKs/NF-κB Pathway

Benzyl Isothiocyanate Attenuates Inflammasome Activation in Pseudomonas aeruginosa LPS-Stimulated THP-1 Cells and Exerts Regulation through the MAPKs/NF-κB Pathway

Chemical Modulation of the 1-(Piperidin-4-yl)-1,3-dihydro-2H-benzo[d]imidazole-2-one Scaffold as a Novel NLRP3 Inhibitor

Acute Glucose Shift Induces the Activation of the NLRP3 Inflammasome in THP-1 Cells

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