Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Aug;122(1):e52.
doi: 10.1002/cpim.52. Epub 2018 Jul 20.

Pyroptosis Induction and Detection

Andreas B den Hartigh  1 , Susan L Fink  1
Affiliations

Pyroptosis Induction and Detection

Andreas B den Hartigh et al. Curr Protoc Immunol. 2018 Aug.

Abstract

Pyroptosis is a form of programmed pro-inflammatory cell death that plays a protective role in the host response to infection, but can also promote pathogenic inflammation. Pyroptosis is mediated by the cysteine protease, caspase-1. Caspase-1 cleaves gasdermin D, releasing the N-terminal pore-forming domain, which inserts into the plasma membrane and drives osmotic lysis. Caspase-1 also proteolytically activates the inflammatory cytokines interleukin 1β (IL-1β) and IL-18. This unit describes methods for stimulating pyroptosis and assessing subsequent loss of plasma membrane integrity. We also describe an ELISA to quantify released IL-1β. These methods can be applied to many different types of experiments. © 2018 by John Wiley & Sons, Inc.

Keywords: caspase-1; cell death; inflammasome; pyroptosis.

PubMed Disclaimer

Figures

Figure 1
Figure 1
LDH release following pyroptosis induction. BMMs were treated with nigericin or infected with Salmonella as described in basic protocol 1 and alternative protocol 3. Pyroptosis was induced in the presence of 5 mM glycine as indicated. LDH release was measured as described in basic protocol 1. Data are means ± SD of three replicates.
Figure 2
Figure 2
EtBr uptake following pyroptosis induction. BMMs were infected with Salmonella as described in alternative protocol 3, or treated with PBS as a negative control. After 30 minutes, cells were stained with ethidium bromide (membrane impermeant, red) and Syto62 (membrane permeant, blue) for 5 min before imaging by confocal microscopy as described in basic protocol 2. White arrows in Salmonella field point to cells not undergoing pyroptotic cell death.
Figure 3
Figure 3
Quantification of secreted IL-1β following pyroptosis induction. BMMs were treated with nigericin, transfected with poly(dA:dT) or infected with Salmonella as described in basic protocol 1 and alternative protocols 2 and 3. IL-1β in the supernatant was measured by ELISA as described in basic protocol 3. Data are means ± SD of three replicates. ND (none detected)
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Bergsbaken T, Fink SL, Cookson BT. Pyroptosis: host cell death and inflammation. Nat Rev Microbiol. 2009;7(2):99–109. doi: 10.1038/nrmicro2070. - DOI - PMC - PubMed
    1. Bergsbaken T, Fink SL, den Hartigh AB, Loomis WP, Cookson BT. Coordinated host responses during pyroptosis: caspase-1-dependent lysosome exocytosis and inflammatory cytokine maturation. J Immunol. 2011;187(5):2748–2754. doi: 10.4049/jimmunol.1100477. - DOI - PMC - PubMed
    1. Boyden ED, Dietrich WF. Nalp1b controls mouse macrophage susceptibility to anthrax lethal toxin. Nat Genet. 2006;38(2):240–244. doi: 10.1038/ng1724. - DOI - PubMed
    1. Broz P, Newton K, Lamkanfi M, Mariathasan S, Dixit VM, Monack DM. Redundant roles for inflammasome receptors NLRP3 and NLRC4 in host defense against Salmonella. J Exp Med. 2010;207(8):1745–1755. doi: 10.1084/jem.20100257. - DOI - PMC - PubMed
    1. Cookson BT, Brennan MA. Pro-inflammatory programmed cell death. Trends Microbiol. 2001;9(3):113–114. - PubMed

LinkOut - more resources