Leucocytes pp 199-222 | Cite as

Measuring the Inflammasome

Part of the Methods in Molecular Biology book series (MIMB, volume 844)


Inflammasomes are multiprotein complexes whose activity has been implicated in physiological and pathological inflammation. The hallmarks of inflammasome activation are the secretion of the mature forms of Caspase-1 and IL-1β from cells of the innate immune system. This protocol covers the methods required to study inflammasome activation using mouse bone marrow-derived dendritic cells (BMDCs) as a model system. The protocol includes the generation and handling of BMDCs, the stimulation of BMDCs with established Nlrp3 inflammasome activators, and the measurement of activation by both ELISA and western blot. These methods can be useful for the study of potential inflammasome activators, and of the signaling pathways involved in inflammasome activation. General considerations are provided that may help in the design and optimization of modified methods for the study of other types of inflammasomes and in other cell types.

Key words

Inflammasome Nlrp3 Caspase-1 Interleukin-1 Bone marrow-derived dendritic cells 



I would like to thank Michael Bscheider, Manuel Ritter, and James Vince for helpful suggestions, and Christina Thomas for proofreading and correcting the manuscript. My work is funded by an EMBO long-term fellowship. I declare no competing financial interests.


  1. 1.
    Schroder, K., Tschopp, J. (2010) The inflammasomes, Cell 140, 821–832.PubMedCrossRefGoogle Scholar
  2. 2.
    Martinon, F., Petrilli, V., Mayor, A. et al. (2006) Gout-associated uric acid crystals activate the NALP3 inflammasome, Nature 440, 237–241.PubMedCrossRefGoogle Scholar
  3. 3.
    Dostert, C., Petrilli, V., Van Bruggen, R. et al. (2008) Innate immune activation through Nalp3 inflammasome sensing of asbestos and silica, Science 320, 674–677.PubMedCrossRefGoogle Scholar
  4. 4.
    Yazdi, A. S., Guarda, G., Riteau, N. et al. (2010) Nanoparticles activate the NLR pyrin domain containing 3 (Nlrp3) inflammasome and cause pulmonary inflammation through release of IL-1alpha and IL-1beta, Proc Natl Acad Sci USA 107, 19449–19454.PubMedCrossRefGoogle Scholar
  5. 5.
    Henderson, C., Goldbach-Mansky, R. (2010) Monogenic autoinflammatory diseases: new insights into clinical aspects and pathogenesis, Curr Opin Rheumatol 22, 567–578.PubMedGoogle Scholar
  6. 6.
    Okamoto, M., Liu, W., Luo, Y. et al. (2010) Constitutively active inflammasome in human melanoma cells mediating autoinflammation via caspase-1 processing and secretion of interleukin-1beta, J Biol Chem 285, 6477–6488.PubMedCrossRefGoogle Scholar
  7. 7.
    Ghiringhelli, F., Apetoh, L., Tesniere, A. et al. (2009) Activation of the NLRP3 inflammasome in dendritic cells induces IL-1beta-dependent adaptive immunity against tumors, Nat Med 15, 1170–1178.PubMedCrossRefGoogle Scholar
  8. 8.
    Zhou, R., Yazdi, A. S., Menu, P. et al. (2010) A role for mitochondria in NLRP3 inflammasome activation, Nature doi: 10.1038/nature09663
  9. 9.
    Masters, S. L., Dunne, A., Subramanian, S. L. et al. (2010) Activation of the NLRP3 inflammasome by islet amyloid polypeptide provides a mechanism for enhanced IL-1beta in type 2 diabetes, Nat Immunol 11, 897–904.PubMedCrossRefGoogle Scholar
  10. 10.
    Duewell, P., Kono, H., Rayner, K. J. et al. (2010) NLRP3 inflammasomes are required for atherogenesis and activated by cholesterol crystals, Nature 464, 1357–1361.PubMedCrossRefGoogle Scholar
  11. 11.
    Bauer, C., Duewell, P., Mayer, C. et al. (2010) Colitis induced in mice with dextran sulfate sodium (DSS) is mediated by the NLRP3 inflammasome, Gut 59, 1192–1199.PubMedCrossRefGoogle Scholar
  12. 12.
    Gross, O., Poeck, H., Bscheider, M. et al. (2009) Syk kinase signalling couples to the Nlrp3 inflammasome for anti-fungal host defence, Nature 459, 433–436.PubMedCrossRefGoogle Scholar
  13. 13.
    Hornung, V., Bauernfeind, F., Halle, A. et al. (2008) Silica crystals and aluminum salts activate the NALP3 inflammasome through phagosomal destabilization, Nat Immunol 9, 847–856.PubMedCrossRefGoogle Scholar
  14. 14.
    Dinarello, C. A. (2009) Immunological and inflammatory functions of the interleukin-1 family, Annu Rev Immunol 27, 519–550.PubMedCrossRefGoogle Scholar
  15. 15.
    Burckstummer, T., Baumann, C., Bluml, S. et al. (2009) An orthogonal proteomic-genomic screen identifies AIM2 as a cytoplasmic DNA sensor for the inflammasome, Nat Immunol 10, 266–272.PubMedCrossRefGoogle Scholar
  16. 16.
    Fernandes-Alnemri, T., Yu, J. W., Datta, P. et al. (2009) AIM2 activates the inflammasome and cell death in response to cytoplasmic DNA, Nature 458, 509–513.PubMedCrossRefGoogle Scholar
  17. 17.
    Hornung, V., Ablasser, A., Charrel-Dennis, M. et al. (2009) AIM2 recognizes cytosolic dsDNA and forms a caspase-1-activating inflammasome with ASC, Nature 458, 514–518.PubMedCrossRefGoogle Scholar
  18. 18.
    Roberts, T. L., Idris, A., Dunn, J. A. et al. (2009) HIN-200 proteins regulate caspase activation in response to foreign cytoplasmic DNA, Science 323, 1057–1060.PubMedCrossRefGoogle Scholar
  19. 19.
    Poeck, H., Bscheider, M., Gross, O. et al. (2010) Recognition of RNA virus by RIG-I results in activation of CARD9 and inflammasome signaling for interleukin 1 beta production, Nat Immunol 11, 63–69.PubMedCrossRefGoogle Scholar
  20. 20.
    Boyden, E. D., and Dietrich, W. F. (2006) Nalp1b controls mouse macrophage susceptibility to anthrax lethal toxin, Nat Genet 38, 240–244.PubMedCrossRefGoogle Scholar
  21. 21.
    Mariathasan, S., Newton, K., Monack, D. M. et al. (2004) Differential activation of the inflammasome by caspase-1 adaptors ASC and Ipaf, Nature 430, 213–218.PubMedCrossRefGoogle Scholar
  22. 22.
    Martinon, F., Burns, K., and Tschopp, J. (2002) The inflammasome: a molecular platform triggering activation of inflammatory caspases and processing of proIL-beta, Mol Cell 10, 417–426.PubMedCrossRefGoogle Scholar
  23. 23.
    Mariathasan, S., Weiss, D. S., Newton, K. et al. (2006) Cryopyrin activates the inflammasome in response to toxins and ATP, Nature 440, 228–232.PubMedCrossRefGoogle Scholar
  24. 24.
    Bauernfeind, F. G., Horvath, G., Stutz, A. et al. (2009) Cutting edge: NF-kappaB activating pattern recognition and cytokine receptors license NLRP3 inflammasome activation by regulating NLRP3 expression, J Immunol 183, 787–791.PubMedCrossRefGoogle Scholar
  25. 25.
    Greten, F. R., Arkan, M. C., Bollrath, J. et al. (2007) NF-kappaB is a negative regulator of IL-1beta secretion as revealed by genetic and pharmacological inhibition of IKKbeta, Cell 130, 918–931.PubMedCrossRefGoogle Scholar
  26. 26.
    Bauernfeind, F., Ablasser, A., Bartok, E. et al. (2011) Inflammasomes: current understanding and open questions, Cell Mol Life Sci 68, 765–83.PubMedCrossRefGoogle Scholar
  27. 27.
    Bergsbaken, T., Fink, S. L., Cookson, B. T. (2009) Pyroptosis: host cell death and inflammation, Nat Rev Microbiol 7, 99–109.PubMedCrossRefGoogle Scholar
  28. 28.
    Keller, M., Ruegg, A., Werner, S. et al. (2008) Active caspase-1 is a regulator of unconventional protein secretion, Cell 132, 818–831.PubMedCrossRefGoogle Scholar
  29. 29.
    Lutz, M. B. (2004) IL-3 in dendritic cell development and function: a comparison with GM-CSF and IL-4, Immunobiology 209, 79–87.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Department of BiochemistryUniversity of LausanneEpalingesSwitzerland

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