The Impact of Various Reactive Oxygen Species on the Formation of Neutrophil Extracellular Traps

The formation of neutrophil extracellular traps (NETs) depends on the generation of reactive oxygen species (ROS). Previous studies revealed that both NADPH oxidase and myeloperoxidase (MPO) are required for NET release. However, the contribution of various ROS as well as the role of mitochondria-derived ROS has not been addressed so far. In the present study we aimed to investigate in a systematic and comprehensive manner the contribution of various ROS and ROS-generating pathways to the PMA-induced NET release. By using specific inhibitors, the role of both NADPH oxidase- and mitochondria-derived ROS as well as the contribution of superoxide dismutase (SOD) and MPO on the NET release was assessed. We could demonstrate that NADPH oxidase function is crucial for the formation of NETs. In addition, we could clearly show the involvement of MPO-derived ROS in NET release. Our results, however, did not provide evidence for the role of SOD- or mitochondria-derived ROS in NET formation.

[1]  B Chance,et al.  The mitochondrial generation of hydrogen peroxide. General properties and effect of hyperbaric oxygen. , 1973, The Biochemical journal.

[2]  V. Wahn,et al.  Myeloperoxidase is required for neutrophil extracellular trap formation: implications for innate immunity. , 2011, Blood.

[3]  A. Kettle,et al.  Mechanism of inhibition of myeloperoxidase by anti-inflammatory drugs. , 1991, Biochemical pharmacology.

[4]  S. Nakaji,et al.  Analysis and assessment of the capacity of neutrophils to produce reactive oxygen species in a 96-well microplate format using lucigenin- and luminol-dependent chemiluminescence. , 1997, Journal of immunological methods.

[5]  T. Hartung,et al.  Highly purified lipoteichoic acid activates neutrophil granulocytes and delays their spontaneous apoptosis via CD14 and TLR2 , 2004, Journal of leukocyte biology.

[6]  M. Trush,et al.  Diphenyleneiodonium, an NAD(P)H oxidase inhibitor, also potently inhibits mitochondrial reactive oxygen species production. , 1998, Biochemical and biophysical research communications.

[7]  Abdul Hakkim,et al.  Impairment of neutrophil extracellular trap degradation is associated with lupus nephritis , 2010, Proceedings of the National Academy of Sciences.

[8]  R. El Bekay,et al.  Stimulators of AMP‐activated protein kinase inhibit the respiratory burst in human neutrophils , 2004, FEBS letters.

[9]  Carl Nathan,et al.  Neutrophils and immunity: challenges and opportunities , 2006, Nature Reviews Immunology.

[10]  Volker Brinkmann,et al.  Beneficial suicide: why neutrophils die to make NETs , 2007, Nature Reviews Microbiology.

[11]  C. Dahlgren,et al.  Intra- and extracellular events in luminol-dependent chemiluminescence of polymorphonuclear leukocytes , 1984, Infection and immunity.

[12]  R. D. del Maestro,et al.  Generation of hydrogen peroxide by brain mitochondria: the effect of reoxygenation following postdecapitative ischemia. , 1989, Archives of biochemistry and biophysics.

[13]  H. Gyllenhammar,et al.  Lucigenin chemiluminescence in the assessment of neutrophil superoxide production. , 1987, Journal of immunological methods.

[14]  Z. Werb,et al.  Netting neutrophils in autoimmune small-vessel vasculitis , 2009, Nature Medicine.

[15]  B Chance,et al.  The cellular production of hydrogen peroxide. , 1972, The Biochemical journal.

[16]  Abdul Hakkim,et al.  Neutrophil elastase and myeloperoxidase regulate the formation of neutrophil extracellular traps , 2010, The Journal of cell biology.

[17]  J. Chappell,et al.  Dihydrorhodamine 123: a fluorescent probe for superoxide generation? , 1993, European journal of biochemistry.

[18]  S. Walrand,et al.  Is the neutrophil reactive oxygen species production measured by luminol and lucigenin chemiluminescence intra or extracellular? Comparison with DCFH-DA flow cytometry and cytochrome c reduction. , 2002, Clinica chimica acta; international journal of clinical chemistry.

[19]  M. Surette,et al.  A Novel Mechanism of Rapid Nuclear Neutrophil Extracellular Trap Formation in Response to Staphylococcus aureus , 2010, The Journal of Immunology.

[20]  E. Abraham,et al.  Mitochondrial respiratory complex I regulates neutrophil activation and severity of lung injury. , 2008, American journal of respiratory and critical care medicine.

[21]  J. Arnhold,et al.  Human myeloperoxidase in innate and acquired immunity. , 2010, Archives of biochemistry and biophysics.

[22]  P. Ganey,et al.  Impairment of human neutrophil oxidative burst by polychlorinated biphenyls: inhibition of superoxide dismutase activity , 1998, Journal of leukocyte biology.

[23]  A. Zychlinsky,et al.  Neutrophil Extracellular Traps Kill Bacteria , 2004, Science.

[24]  S. Adachi,et al.  Singlet oxygen is essential for neutrophil extracellular trap formation. , 2011, Biochemical and biophysical research communications.

[25]  S. Grinstein,et al.  Unconventional Roles of the NADPH Oxidase: Signaling, Ion Homeostasis, and Cell Death , 2007, Science's STKE.

[26]  D. Tew,et al.  Studies on the inhibitory mechanism of iodonium compounds with special reference to neutrophil NADPH oxidase. , 1993, The Biochemical journal.

[27]  A. Verhoeven,et al.  Human neutrophils lose their surface Fc gamma RIII and acquire Annexin V binding sites during apoptosis in vitro. , 1995, Blood.

[28]  F. Fonnum,et al.  Evaluation of the probes 2',7'-dichlorofluorescin diacetate, luminol, and lucigenin as indicators of reactive species formation. , 2003, Biochemical pharmacology.

[29]  A. Kettle,et al.  Reactions of superoxide with myeloperoxidase and its products. , 2004, Japanese journal of infectious diseases.

[30]  Helmut Laufs,et al.  Inhibition of the Spontaneous Apoptosis of Neutrophil Granulocytes by the Intracellular Parasite Leishmania major1 , 2002, The Journal of Immunology.

[31]  Paul Stevens,et al.  The role of myeloperoxidase and superoxide anion in the luminol- and lucigenin-dependent chemiluminescence of human neutrophils , 1984 .

[32]  M. M. Lieberman,et al.  Comparative study of neutrophil activation by chemiluminescence and flow cytometry , 1996, Clinical and diagnostic laboratory immunology.

[33]  R. Moots,et al.  The Mitochondrial Network of Human Neutrophils: Role in Chemotaxis, Phagocytosis, Respiratory Burst Activation, and Commitment to Apoptosis 1 , 2003, The Journal of Immunology.

[34]  J. Paul Robinson,et al.  Mitochondrial Complex I Inhibitor Rotenone Induces Apoptosis through Enhancing Mitochondrial Reactive Oxygen Species Production* , 2003, The Journal of Biological Chemistry.

[35]  E. Fernandes,et al.  Inhibition of human neutrophil oxidative burst by pyrazolone derivatives. , 2006, Free radical biology & medicine.