Abstract
The purinoceptor P2X7 is highly expressed in cells of the innate immune system, including monocytes and macrophages. Its activation is a potent signal to activate the NLRP3 inflammasome and induce the release of proinflammatory cytokines of the IL-1 family. In this chapter, we present a method to monitor NLRP3 inflammasome activation in human monocytes upon P2X7 receptor stimulation by detecting intracellular oligomers of ASC by flow cytometry. This method could be used to evaluate the degree of inflammasome activation in blood samples from patients suffering different chronic inflammatory diseases.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Baroja-Mazo A, Barberà -Cremades M, Pelegrin P (2013) The participation of plasma membrane hemichannels to purinergic signaling. Biochim Biophys Acta 1828:79–93
Surprenant A, North RA (2009) Signaling at purinergic P2X receptors. Annu Rev Physiol 71:333–359
Amores-Iniesta J, Barberà -Cremades M, MartÃnez CM et al (2017) Extracellular ATP activates the NLRP3 inflammasome and is an early danger signal of skin allograft rejection. Cell Rep 21(12):3414–3426
Barberà -Cremades M, Baroja-Mazo A, Gomez AI et al (2012) P2X7 receptor-stimulation causes fever via PGE2 and IL-1β release. FASEB J 26:2951–2962
Pelegrin P (2011) Many ways to dilate the P2X7 receptor pore. Br J Pharmacol 163:908–911
Young MT, Pelegrin P, Pelegrin P et al (2006) Identification of Thr283 as a key determinant of P2X7 receptor function. Br J Pharmacol 149:261–268
de Torre-Minguela C, Mesa del Castillo P, PelegrÃn P (2017) The NLRP3 and pyrin inflammasomes: implications in the pathophysiology of autoinflammatory diseases. Front Immunol 8:43
Schmidt FI, Lu A, Chen JW et al (2016) A single domain antibody fragment that recognizes the adaptor ASC defines the role of ASC domains in inflammasome assembly. J Exp Med 213:771–790
Boucher D, Monteleone M, Coll RC et al (2018) Caspase-1 self-cleavage is an intrinsic mechanism to terminate inflammasome activity. J Exp Med 215(3):827–840
Shi J, Gao W, Shao F (2017) Pyroptosis: gasdermin-mediated programmed necrotic cell death. Trends Biochem Sci 42:245–254
Compan V, Baroja-Mazo A, Bragg L et al (2012) A genetically encoded IL-1β bioluminescence resonance energy transfer sensor to monitor inflammasome activity. J Immunol 189:2131–2137
de Torre-Minguela C, Barberà -Cremades M, Gómez AI et al (2016) Macrophage activation and polarization modify P2X7 receptor secretome influencing the inflammatory process. Sci Rep 6:22586
Sester DP, Sester DP, Thygesen SJ et al (2014) A novel flow cytometric method to assess inflammasome formation. J Immunol 194:455–462
Acknowledgments
This work was supported by the European Research Council (ERC-2013-CoG 614578), EU COST Program (BM1406), Fundación Séneca (20859/PI/18) and Ministerio de Economia, Industria y Competitividad–Fondo Europeo de Desarrollo Regional (project no. SAF2017-88276-R).
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2020 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
MartÃnez-Banaclocha, H., PelegrÃn, P. (2020). Detection of Inflammasome Activation by P2X7 Purinoceptor Activation by Determining ASC Oligomerization. In: PelegrÃn, P. (eds) Purinergic Signaling. Methods in Molecular Biology, vol 2041. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9717-6_25
Download citation
DOI: https://doi.org/10.1007/978-1-4939-9717-6_25
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-9716-9
Online ISBN: 978-1-4939-9717-6
eBook Packages: Springer Protocols