Skip to main content
SpringerLink
Log in

Detection of SLE Antigens in Neutrophil Extracellular Traps (NETs)

  • Protocol
  • First Online:
Systemic Lupus Erythematosus

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1134))

Abstract

Neutrophils are sentinel cells of the innate immune system with a primary role of clearing extracellular pathogens. The release of weblike structures decorated with granular proteins called neutrophil extracellular traps (NETs) has recently been implicated in the pathogenesis of inflammatory and autoimmune diseases. Indeed, NETs may represent an important source of autoantigens and immunostimulatory proteins in systemic lupus erythematosus (SLE). In this chapter, we describe protocols to isolate human peripheral neutrophils, to generate and isolate NETs, and to detect SLE antigens in NETs using immunofluorescence and immunoblot.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Borregaard N (2010) Neutrophils, from marrow to microbes. Immunity 33:657–670

    Article  CAS  PubMed  Google Scholar 

  2. Amulic B, Cazalet C, Hayes GL, Metzler KD, Zychlinsky A (2012) Neutrophil function: from mechanisms to disease. Annu Rev Immunol 30:459–489

    Article  CAS  PubMed  Google Scholar 

  3. Brinkmann V, Reichard U, Goosmann C, Fauler B, Uhlemann Y, Weiss DS, Weinrauch Y, Zychlinsky A (2004) Neutrophil extracellular traps kill bacteria. Science 303:1532–1535

    Article  CAS  PubMed  Google Scholar 

  4. Villanueva E, Yalavarthi S, Berthier CC, Hodgin JB, Khandpur R, Lin AM, Rubin CJ, Zhao W, Olsen SH, Klinker M, Shealy D, Denny MF, Plumas J, Chaperot L, Kretzler M, Bruce AT, Kaplan MJ (2011) Netting neutrophils induce endothelial damage, infiltrate tissues, and expose immunostimulatory molecules in systemic lupus erythematosus. J Immunol 187:538–552

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  5. Wang Y, Li M, Stadler S, Correll S, Li P, Wang D, Hayama R, Leonelli L, Han H, Grigoryev SA, Allis CD, Coonrod SA (2009) Histone hypercitrullination mediates chromatin decondensation and neutrophil extracellular trap formation. J Cell Biol 184:205–213

    Article  CAS  PubMed  Google Scholar 

  6. Knight JS, Zhao W, Luo W, Subramanian V, O’Dell AA, Yalavarthi S, Hodgin JB, Eitzman DT, Thompson PR, Kaplan MJ (2013) Peptidylarginine deiminase inhibition is immunomodulatory and vasculoprotective in murine lupus. J Clin Invest 123(7):2981–2983

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Papayannopoulos V, Metzler KD, Hakkim A, Zychlinsky A (2010) Neutrophil elastase and myeloperoxidase regulate the formation of neutrophil extracellular traps. J Cell Biol 191:677–691

    Article  CAS  PubMed  Google Scholar 

  8. Lande R, Ganguly D, Facchinetti V, Frasca L, Conrad C, Gregorio J, Meller S, Chamilos G, Sebasigari R, Riccieri V, Bassett R, Amuro H, Fukuhara S, Ito T, Liu YJ, Gilliet M (2011) Neutrophils activate plasmacytoid dendritic cells by releasing self-DNA-peptide complexes in systemic lupus erythematosus. Sci Transl Med 3:73ra19

    Article  PubMed Central  PubMed  Google Scholar 

  9. Kahlenberg JM, Carmona-Rivera C, Smith CK, Kaplan MJ (2012) Neutrophil extracellular trap-associated protein activation of the NLRP3 inflammasome is enhanced in lupus macrophages. J Immunol 190(3):1217–1226

    Article  PubMed Central  PubMed  Google Scholar 

  10. Knight JS, Carmona-Rivera C, Kaplan MJ (2012) Proteins derived from neutrophil extracellular traps may serve as self-antigens and mediate organ damage in autoimmune diseases. Front Immunol 3:380

    Article  PubMed Central  PubMed  Google Scholar 

  11. Tamiya H, Tani K, Miyata J, Sato K, Urata T, Lkhagvaa B, Otsuka S, Shigekiyo S, Sone S (2006) Defensins- and cathepsin G-ANCA in systemic lupus erythematosus. Rheumatol Int 27:147–152

    Article  CAS  PubMed  Google Scholar 

  12. Mosca M, Chimenti D, Pratesi F, Baldini C, Anzilotti C, Bombardieri S, Migliorini P (2006) Prevalence and clinico-serological correlations of anti-alpha-enolase, anti-C1q, and anti-dsDNA antibodies in patients with systemic lupus erythematosus. J Rheumatol 33:695–697

    CAS  PubMed  Google Scholar 

  13. Mansour RB, Lassoued S, Gargouri B, El Gaid A, Attia H, Fakhfakh F (2008) Increased levels of autoantibodies against catalase and superoxide dismutase associated with oxidative stress in patients with rheumatoid arthritis and systemic lupus erythematosus. Scand J Rheumatol 37:103–108

    Article  PubMed  Google Scholar 

  14. Zhao MH, Liu N, Zhang YK, Wang HY (1998) Antineutrophil cytoplasmic autoantibodies (ANCA) and their target antigens in Chinese patients with lupus nephritis. Nephrol Dial Transplant 13:2821–2824

    Article  CAS  PubMed  Google Scholar 

  15. Manolova I, Dancheva M, Halacheva K (2001) Antineutrophil cytoplasmic antibodies in patients with systemic lupus erythematosus: prevalence, antigen specificity, and clinical associations. Rheumatol Int 20:197–204

    Article  CAS  PubMed  Google Scholar 

  16. Nassberger L, Jonsson H, Sjoholm AG, Sturfelt G, Heubner A (1989) Circulating anti-elastase in systemic lupus erythematosus. Lancet 1:509

    Article  CAS  PubMed  Google Scholar 

  17. Monestier M, Decker P, Briand JP, Gabriel JL, Muller S (2000) Molecular and structural properties of three autoimmune IgG monoclonal antibodies to histone H2B. J Biol Chem 275:13558–13563

    Article  CAS  PubMed  Google Scholar 

  18. Robinson WH, DiGennaro C, Hueber W, Haab BB, Kamachi M, Dean EJ, Fournel S, Fong D, Genovese MC, de Vegvar HE, Skriner K, Hirschberg DL, Morris RI, Muller S, Pruijn GJ, van Venrooij WJ, Smolen JS, Brown PO, Steinman L, Utz PJ (2002) Autoantigen microarrays for multiplex characterization of autoantibody responses. Nat Med 8:295–301

    Article  CAS  PubMed  Google Scholar 

  19. van Bavel CC, Dieker JW, Kroeze Y, Tamboer WP, Voll R, Muller S, Berden JH, van der Vlag J (2011) Apoptosis-induced histone H3 methylation is targeted by autoantibodies in systemic lupus erythematosus. Ann Rheum Dis 70:201–207

    Article  PubMed  Google Scholar 

  20. Lee SS, Lawton JW, Chan CE, Li CS, Kwan TH, Chau KF (1992) Antilactoferrin antibody in systemic lupus erythematosus. Br J Rheumatol 31:669–673

    Article  CAS  PubMed  Google Scholar 

  21. Caccavo D, Rigon A, Picardi A, Galluzzo S, Vadacca M, Ferri GM, Amoroso A, Afeltra A (2005) Anti-lactoferrin antibodies in systemic lupus erythematosus: isotypes and clinical correlates. Clin Rheumatol 24:381–387

    Article  PubMed  Google Scholar 

  22. Nassberger L, Sjoholm AG, Jonsson H, Sturfelt G, Akesson A (1990) Autoantibodies against neutrophil cytoplasm components in systemic lupus erythematosus and in hydralazine-induced lupus. Clin Exp Immunol 81:380–383

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  23. Cambridge G, Wallace H, Bernstein RM, Leaker B (1994) Autoantibodies to myeloperoxidase in idiopathic and drug-induced systemic lupus erythematosus and vasculitis. Br J Rheumatol 33:109–114

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Systemic Autoimmunity Branch, Intramural Research Program, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD, USA

    Carmelo Carmona-Rivera & Mariana J. Kaplan M.D.

Authors
  1. Carmelo Carmona-Rivera
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mariana J. Kaplan M.D.
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. University of Exeter Medical School, Devon, United Kingdom

    Paul Eggleton

  2. Section of Immunology and Infection, Division of Applied Medicine, Institute of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom

    Frank J. Ward

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media, New York

About this protocol

Cite this protocol

Carmona-Rivera, C., Kaplan, M.J. (2014). Detection of SLE Antigens in Neutrophil Extracellular Traps (NETs). In: Eggleton, P., Ward, F. (eds) Systemic Lupus Erythematosus. Methods in Molecular Biology, vol 1134. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0326-9_11

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-0326-9_11

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-0325-2

  • Online ISBN: 978-1-4939-0326-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation