Skip to main content

Main menu

  • Home
  • Our journals
    • Clinical Medicine
    • Future Healthcare Journal
  • Subject collections
  • About the RCP
  • Contact us

Clinical Medicine Journal

  • ClinMed Home
  • Content
    • Current
    • Ahead of print
    • Archive
  • Author guidance
    • Instructions for authors
    • Submit online
  • About ClinMed
    • Scope
    • Editorial board
    • Policies
    • Information for reviewers
    • Advertising

User menu

  • Log in

Search

  • Advanced search
RCP Journals
Home
  • Log in
  • Home
  • Our journals
    • Clinical Medicine
    • Future Healthcare Journal
  • Subject collections
  • About the RCP
  • Contact us
Advanced

Clinical Medicine Journal

clinmedicine Logo
  • ClinMed Home
  • Content
    • Current
    • Ahead of print
    • Archive
  • Author guidance
    • Instructions for authors
    • Submit online
  • About ClinMed
    • Scope
    • Editorial board
    • Policies
    • Information for reviewers
    • Advertising

A novel assay of neutrophil extracellular trap (NET) formation identifies anti-IL-8 therapies to reduce disseminated intravascular coagulation and mortality in the intensive care unit

Simon Abrams, Ben Morton, Yasir Alhamdi, Mohmad Alsabani, Zhenxing Cheng, Steven Lane, Ingeborg Welters, Guozheng Wang and Cheng-Hock Toh
Download PDF
DOI: https://doi.org/10.7861/clinmed.20-2-s114
Clin Med March 2020
Simon Abrams
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Ben Morton
BLiverpool School of Tropical Medicine, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yasir Alhamdi
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Mohmad Alsabani
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Zhenxing Cheng
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Steven Lane
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Ingeborg Welters
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Guozheng Wang
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Cheng-Hock Toh
AUniversity of Liverpool, Liverpool, UK
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • Article
  • Info & Metrics
Loading

Introduction

Neutrophils are the first line of defence against bacterial infection, and formation of neutrophil extracellular traps (NETs) is an important protective mechanism. NETs can also be harmful by inducing intravascular coagulation and multi-organ failure (MOF) in animal models.1–6 Although increasingly considered as important therapeutic targets,7–9 there is currently no robust and specific measure of NET formation to inform clinical care and enable precision medicine in patients on the intensive care unit (ICU). The aim of this study is to establish a novel assay for measuring NETs and assess its clinical significance.

Materials and methods

A prospective cohort of 341 consecutive adult ICU patients was recruited at the Royal Liverpool University Hospital, following written informed consent. The NET-forming capacity of ICU admission blood samples was semi-quantified by directly incubating patient plasma with isolated healthy neutrophils ex vivo. Associations of NET-forming capacity with sequential organ failure assessment (SOFA) scores, disseminated intravascular coagulation (DIC) and 28-day mortality were analysed and compared with available NET assays. Cytokine analysis and inhibitor studies were performed to determine the driving factors of NET formation in patients. To determine the pathological relevance of NETs, complementary in vivo studies were performed in mouse models of sepsis (caecal ligation and puncture (CLP) or intraperitoneal injection of Escherichia coli), without or with anti-NET therapy.

Results and discussion

We observed that NETs were directly induced by heterologous healthy neutrophils incubated with plasma taken from ICU patients on ICU admission, but not from healthy donors (unless incubated with 100 nM PMA). Using this novel assay we could stratify patients into four groups: those with absent (22.0%), mild (49.9%), moderate (14.4%) and strong (13.8%) NET formation. Strong NET formation was predominantly found in sepsis (p<0.0001) and was associated with higher SOFA scores. Adjusted by APACHE II, multivariate regression showed that measuring the degree of NET formation on ICU admission could independently predict DIC and mortality, whereas other NET assays, eg cell-free DNA, myeloperoxidase and myeloperoxidase–DNA complexes, could not. Interleukin (IL)-8 levels were found to be strongly associated with NET formation, and inhibiting IL-8 significantly attenuated NETosis.

Using mouse models of sepsis, we could monitor NET formation using plasma, which was associated with NET-positive staining (cit-H3) in the lung tissue. This was associated with increased fibrin deposition within the lung tissue, along with lung injury scores and circulating markers of liver (blood urea nitrogen; CLP: p=0.005, E coli: p<0.001), kidney (alanine aminotransferase; CLP: p=0.01, E coli: p=0.002) and cardiac injury (cardiac troponin I; CLP: p<0.001, E coli: p<0.001). By targeting IL-8 (using a clinically relevant compound, reparixin) in septic mice, we were able to significantly inhibit NET formation, fibrin deposition and organ injury, and improve survival times (p=0.004).

Conclusion

Our new NET assay directly measures the NET-forming capacity in patient plasma. This could guide clinical management and enable identification of NET-inducing factors in individual patients for targeted treatment and personalised ICU medicine. We identify IL-8 as a major driving factor in sepsis, with anti-IL-8 therapy in septic mice significantly reducing NET-induced organ damage and mortality.

Conflicts of interest

None declared.

  • © Royal College of Physicians 2020. All rights reserved.

References

  1. ↵
    1. Fuchs TA
    , Brill A, Duerschmied D, et al. Extracellular DNA traps promote thrombosis. Proc Natl Acad Sci U S A 2010;107:15880–5.
    OpenUrlAbstract/FREE Full Text
  2. ↵
    1. Gould TJ
    , Lysov Z, Liaw PC. Extracellular DNA and histones: double-edged swords in immunothrombosis. J Thromb Haemost 2015;13 Suppl 1:S82–91.
    OpenUrlCrossRefPubMed
  3. ↵
    1. Gould TJ
    , Vu TT, Swystun LL, et al. Neutrophil extracellular traps promote thrombin generation through platelet-dependent and platelet-independent mechanisms. Arterioscler Thromb Vasc Biol 2014;34:1977–84.
    OpenUrlAbstract/FREE Full Text
  4. ↵
    1. Delabranche X
    , Stiel L, Severac F, et al. Evidence of netosis in septic shock-induced disseminated intravascular coagulation. Shock 2017;47:313–7.
    OpenUrl
  5. ↵
    1. McDonald B
    , Davis R, Jenne CN. Neutrophil extracellular traps (NETs) promote disseminated intravascular coagulation in sepsis. J Immunol 2016;196 (1 Suppl):60.8.
    OpenUrl
  6. ↵
    1. Fuchs TA
    , Brill A, Wagner DD. Neutrophil extracellular trap (NET) impact on deep vein thrombosis. Arterioscler Thromb Vasc Biol 2012;32:1777–83.
    OpenUrlAbstract/FREE Full Text
  7. ↵
    1. Caudrillier A
    , Kessenbrock K, Gilliss BM, et al. Platelets induce neutrophil extracellular traps in transfusion-related acute lung injury. J Clin Invest 2012;122:2661–71.
    OpenUrlCrossRefPubMed
  8. ↵
    1. Brill A
    , Fuchs TA, Savchenko AS, et al. Neutrophil extracellular traps promote deep vein thrombosis in mice. J Thromb Haemost 2012;10:136–44.
    OpenUrlCrossRefPubMed
  9. ↵
    1. Thomas GM
    , Carbo C, Curtis BR, et al. Extracellular DNA traps are associated with the pathogenesis of TRALI in humans and mice. Blood 2012;119:6335–43.
    OpenUrlAbstract/FREE Full Text
Back to top
Previous articleNext article

Article Tools

Download PDF
Article Alerts
Sign In to Email Alerts with your Email Address
Citation Tools
A novel assay of neutrophil extracellular trap (NET) formation identifies anti-IL-8 therapies to reduce disseminated intravascular coagulation and mortality in the intensive care unit
Simon Abrams, Ben Morton, Yasir Alhamdi, Mohmad Alsabani, Zhenxing Cheng, Steven Lane, Ingeborg Welters, Guozheng Wang, Cheng-Hock Toh
Clinical Medicine Mar 2020, 20 (Suppl 2) s114-s115; DOI: 10.7861/clinmed.20-2-s114

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Share
A novel assay of neutrophil extracellular trap (NET) formation identifies anti-IL-8 therapies to reduce disseminated intravascular coagulation and mortality in the intensive care unit
Simon Abrams, Ben Morton, Yasir Alhamdi, Mohmad Alsabani, Zhenxing Cheng, Steven Lane, Ingeborg Welters, Guozheng Wang, Cheng-Hock Toh
Clinical Medicine Mar 2020, 20 (Suppl 2) s114-s115; DOI: 10.7861/clinmed.20-2-s114
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Jump to section

  • Article
    • Introduction
    • Materials and methods
    • Results and discussion
    • Conclusion
    • Conflicts of interest
    • References
  • Info & Metrics

Related Articles

  • No related articles found.
  • PubMed
  • Google Scholar

Cited By...

  • No citing articles found.
  • Google Scholar

More in this TOC Section

  • Complex care liaison preoperative assessment clinic: a 1-year review
  • The acceptability and utility of different diagnostic tests and sample types for the surveillance of trachoma in the Bijagos Islands, Guinea Bissau
  • Machine learning methods in predicting chemotherapy-induced neutropenia in oncology patients using clinical data
Show more Research

Similar Articles

Navigate this Journal

  • Journal Home
  • Current Issue
  • Ahead of Print
  • Archive

Related Links

  • ClinMed - Home
  • FHJ - Home
clinmedicine Footer Logo
  • Home
  • Journals
  • Contact us
  • Advertise
HighWire Press, Inc.

Follow Us:

  • Follow HighWire Origins on Twitter
  • Visit HighWire Origins on Facebook

Copyright © 2021 by the Royal College of Physicians