Abstract
Neutrophil extracellular traps (NETs) represent extracellular structures able to bind and kill microorganisms. It is believed that they are generated by neutrophils undergoing cell death, allowing these dying or dead cells to kill microbes. We show that, following priming with granulocyte/macrophage colony-stimulating factor (GM-CSF) and subsequent short-term toll-like receptor 4 (TLR4) or complement factor 5a (C5a) receptor stimulation, viable neutrophils are able to generate NETs. Strikingly, NETs formed by living cells contain mitochondrial, but no nuclear, DNA. Pharmacological or genetic approaches to block reactive oxygen species (ROS) production suggested that NET formation is ROS dependent. Moreover, neutrophil populations stimulated with GM-CSF and C5a showed increased survival compared with resting neutrophils, which did not generate NETs. In conclusion, mitochondrial DNA release by neutrophils and NET formation do not require neutrophil death and do also not limit the lifespan of these cells.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
Abbreviations
- Abs:
-
antibodies
- C5a:
-
complement factor 5a
- CGD:
-
chronic granulomatous disease
- DPI:
-
diphenyleneiodonium
- GM-CSF:
-
granulocyte/macrophage colony-stimulating factor
- LPS:
-
lipopolysaccharide
- MPO:
-
myeloperoxidase
- NET:
-
neutrophil extracellular trap
- PARP:
-
poly(ADP-ribose) polymerase
- PCR:
-
polymerase chain reaction
- PI:
-
propidium iodide
- PS:
-
phosphatidylserine
- ROS:
-
reactive oxygen species
- TLR:
-
toll-like receptor
References
Nathan C . Neutrophils and immunity: challenges and opportunities. Nat Rev Immunol 2006; 6: 173–182.
Brinkmann V, Reichard U, Goosmann C, Fauler B, Uhlemann Y, Weiss DS et al. Neutrophil extracellular traps kill bacteria. Science 2004; 303: 1532–1535.
Brinkmann V, Zychlinsky A . Beneficial suicide: why neutrophils die to make NETs. Nat Rev Microbiol 2007; 5: 577–582.
Martinelli S, Urosevic M, Daryadel A, Oberholzer PA, Baumann C, Fey MF et al. Induction of genes mediating interferon-dependent extracellular trap formation during neutrophil differentiation. J Biol Chem 2004; 279: 44123–44132.
Emert D, Zychlinsky A, Urban C . Fungal and bacterial killing by neutrophils. Methods Mol Biol 2009; 470: 293–312.
Clark SR, Ma AC, Tavener SA, McDonald B, Goodarzi Z, Kelly MM et al. Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood. Nat Med 2007; 13: 463–469.
Gupta AK, Hasler P, Holzgreve W, Gebhardt S, Hahn S . Induction of neutrophil extracellular DNA lattices by placental microparticles and IL-8 and their presence in preeclampsia. Hum Immunol 2005; 66: 1146–1154.
Yousefi S, Gold JA, Andina N, Lee JJ, Kelly AM, Kozlowski E et al. Catapult-like release of mitochondrial DNA by eosinophils contributes to antibacterial defense. Nat Med 2008; 14: 949–953.
von Köckritz-Blickwede M, Goldmann O, Thulin P, Heinemann K, Norrby-Teglund A, Rohde M et al. Phagocytosis-independent antimicrobial activity of mast cells by means of extracellular trap formation. Blood 2008; 111: 3070–3080.
Fuchs TA, Abed U, Goosmann C, Hurwitz R, Schulze I, Wahn V et al. Novel cell death program leads to neutrophil extracellular traps. J Cell Biol 2007; 176: 231–241.
Robinson KM, Janes MS, Pehar M, Monette JS, Ross MF, Hagen TM et al. Selective fluorescent imaging of superoxide in vivo using ethidium-based probes. Proc Natl Acad Sci USA 2006; 103: 15038–15043.
Simon HU . Neutrophil apoptosis pathways and their modification in inflammation. Immunol Rev 2003; 193: 101–110.
von Gunten S, Yousefi S, Seitz M, Jakob SM, Schaffner T, Seger R et al. Siglec-9 transduces apoptotic and nonapoptotic death signals into neutrophils depending on the proinflammatory cytokine environment. Blood 2005; 106: 1423–1431.
von Gunten S, Schaub A, Vogel M, Stadler BM, Miescher S, Simon HU . Immunologic and functional evidence for anti-Siglec-9 antibodies in intravenous immunoglobulin preparations. Blood 2006; 108: 4255–4259.
Conus S, Perozzo R, Reinheckel T, Peters C, Scapozza L, Yousefi S et al. Caspase-8 is activated by cathepsin D initiating neutrophil apoptosis during the resolution of inflammation. J Exp Med 2008; 205: 685–698.
Kostylina G, Simon D, Fey MF, Yousefi S, Simon HU . Neutrophil apoptosis mediated by nicotinic acid receptors (GPR109A). Cell Death Differ 2008; 15: 134–142.
Yousefi S, Perozzo R, Schmid I, Ziemiecki A, Schaffner T, Scapozza L et al. Calpain-mediated cleavage of Atg5 switches autophagy to apoptosis. Nat Cell Biol 2006; 8: 1124–1132.
Acknowledgements
We greatly appreciate the help obtained from Dr. Janine Reichenbach (University Children's Hospital, Zurich, Switzerland) in blood sampling of CGD patients. This work was supported by the Swiss National Science Foundation (grant No. 310000-112078 and 310000-107526).
Author information
Authors and Affiliations
Corresponding author
Additional information
Edited by L Scorrano
Supplementary Information accompanies the paper on Cell Death and Differentiation website (http://www.nature.com/cdd)
Supplementary information
Rights and permissions
About this article
Cite this article
Yousefi, S., Mihalache, C., Kozlowski, E. et al. Viable neutrophils release mitochondrial DNA to form neutrophil extracellular traps. Cell Death Differ 16, 1438–1444 (2009). https://doi.org/10.1038/cdd.2009.96
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/cdd.2009.96
Keywords
This article is cited by
-
Top Five Stories of the Cellular Landscape and Therapies of Atherosclerosis: Current Knowledge and Future Perspectives
Current Medical Science (2024)
-
NETosis and kidney disease: what do we know?
International Urology and Nephrology (2023)
-
Melanoma-derived soluble mediators modulate neutrophil biological properties and the release of neutrophil extracellular traps
Cancer Immunology, Immunotherapy (2023)
-
Neutrophil extracellular traps have auto-catabolic activity and produce mononucleosome-associated circulating DNA
Genome Medicine (2022)
-
Complement C5a induces the generation of neutrophil extracellular traps by inhibiting mitochondrial STAT3 to promote the development of arterial thrombosis
Thrombosis Journal (2022)
