Download PDF
Thromb Haemost 2017; 117(10): 1919-1929
DOI: 10.1160/TH17-02-0130
Stroke, Systemic or Venous Thromboembolism
Schattauer GmbH

Neutrophil extracellular traps are increased in patients with acute ischemic stroke: prognostic significance

Juana Vallés
1   Hemostasis, Thrombosis, Atherosclerosis, and Vascular Biology Unit, Health Research Institute La Fe, Valencia, Spain
,
Aida Lago
2   Department of Neurology, University and Polytechnic Hospital La Fe, Valencia, Spain
,
María Teresa Santos
1   Hemostasis, Thrombosis, Atherosclerosis, and Vascular Biology Unit, Health Research Institute La Fe, Valencia, Spain
,
Ana María Latorre
1   Hemostasis, Thrombosis, Atherosclerosis, and Vascular Biology Unit, Health Research Institute La Fe, Valencia, Spain
,
José I. Tembl
2   Department of Neurology, University and Polytechnic Hospital La Fe, Valencia, Spain
,
Juan B. Salom
3   Cerebrovascular Research Joint Unit, Health Research Institute La Fe, Valencia, Spain
4   Department of Physiology, University of Valencia, Valencia, Spain
,
Candela Nieves
2   Department of Neurology, University and Polytechnic Hospital La Fe, Valencia, Spain
,
Antonio Moscardó
1   Hemostasis, Thrombosis, Atherosclerosis, and Vascular Biology Unit, Health Research Institute La Fe, Valencia, Spain
› Author Affiliations Financial support: This work was supported in part by grants from the Spanish Fondo de Investigaciones Sanitarias Carlos III [FIS13/00016] Fondos FEDER, una forma de hacer Europa, Fundación Española de Trombosis y Hemostasia, Sociedad Valenciana de Cardiología, Consellería Valenciana d’Educació (ACIF/2016/465), GV and RETICS networks INVICTUS (RD12/0014/0004) and INVICTUS+ (RD16/0019/0008) ‘Instituto de Salud Carlos III’.
Further Information

Publication History

Received: 23 February 2017

Accepted after major revision: 16 June 2017

Publication Date:
28 November 2017 (online)

    Permissions and Reprints

Summary

Neutrophil extracellular traps (NETs) are networks of DNA, histones, and proteolytic enzymes produced by activated neutrophils through different mechanisms. NET formation is promoted by activated platelets and can in turn activate platelets, thus favoring thrombotic processes. NETs have been detected in venous and arterial thrombosis, but data in stroke are scarce. The aim of this study was to evaluate NETs in the plasma of patients with acute ischemic stroke and their potential association with baseline clinical characteristics, stroke severity, and one-year clinical outcomes. The study included 243 patients with acute ischemic stroke. Clinical and demographic data and scores of stroke severity (NIHSS and mRs) at onset and discharge were recorded. Markers of NETs (cell-free DNA, nucleosomes, and citrullinated histone 3 (citH3)), were determined in plasma. Patients were followed-up for 12 months after the ischemic event. NETs were significantly elevated in the plasma of patients with acute ischemic stroke when compared to healthy subjects. NETs were increased in patients who were over 65 years of age and in those with a history of atrial fibrillation (AF), cardioembolic stroke, high glucose levels, and severe stroke scores at admission and discharge. In multivariate analysis, elevated levels of citH3, the most specific marker of NETs, at onset were independently associated with AF and all-cause mortality at oneyear follow-up. NETs play a role in the pathophysiology of stroke and are associated with severity and mortality. In conclusion, citH3 may constitute a useful prognostic marker and therapeutic target in patients with acute stroke.

Keywords

Stroke - extracellular traps - atrial fibrillation - glucose - immunity
 

  • References

  • 1 Krishnamurthi RV, Feigin VL, Forouzanfar MH. et al. Global and regional burden of first-ever ischaemic and haemorrhagic stroke during 1990-2010: findings from the Global Burden of Disease Study 2010. Lancet Glob Health 2013; 01: 259-281.
    CrossrefPubMedGoogle Scholar
  • 2 Chamorro A, Dirnagl U, Urra X. et al. Neuroprotection in acute stroke: targeting excitotoxicity, oxidative and nitrosative stress, and inflammation. Lancet Neurol 2016; 15: 869-881.
    CrossrefPubMedGoogle Scholar
  • 3 Kehrel BE, Fender AC. Resolving Thromboinflammation in the Brain After Ischemic Stroke?. Circulation 2016; 133: 2128-2131.
    CrossrefPubMedGoogle Scholar
  • 4 Moscardo A, Latorre A, Santos MT. et al. Platelet function in malignant hematological disorders. Curr Opin Oncol 2015; 27: 522-531.
    CrossrefPubMedGoogle Scholar
  • 5 Valles J, Santos MT, Aznar J. et al. Platelet-erythrocyte interactions enhance alpha(IIb)beta(3) integrin receptor activation and P-selectin expression during platelet recruitment: down-regulation by aspirin ex vivo. Blood 2002; 99: 3978-3984.
    CrossrefPubMedGoogle Scholar
  • 6 Santos MT, Valles J, Marcus AJ. et al. Enhancement of platelet reactivity and modulation of eicosanoid production by intact erythrocytes. A new approach to platelet activation and recruitment. J Clin Invest 1991; 87: 571-580.
    CrossrefPubMedGoogle Scholar
  • 7 Valles J, Lago A, Moscardo A. et al. TXA2 synthesis and COX1-independent platelet reactivity in aspirin-treated patients soon after acute cerebral stroke or transient ischaemic attack. Thromb Res 2013; 132: 211-216.
    CrossrefPubMedGoogle Scholar
  • 8 Lago A, Parkhutik V, Tembl JI. et al. Assessment of platelet function in acute ischemic stroke patients previously treated with aspirin. J Stroke Cerebrovasc Dis 2014; 23: 2794-2799.
    CrossrefPubMedGoogle Scholar
  • 9 Brinkmann V, Reichard U, Goosmann C. et al. Neutrophil extracellular traps kill bacteria. Science 2004; 303: 1532-1535.
    CrossrefPubMedGoogle Scholar
  • 10 Fuchs TA, Brill A, Wagner DD. Neutrophil extracellular trap (NET) impact on deep vein thrombosis. Arterioscler Thromb Vasc Biol 2012; 32: 1777-1783.
    CrossrefPubMedGoogle Scholar
  • 11 Li P, Li M, Lindberg MR. et al. PAD4 is essential for antibacterial innate immunity mediated by neutrophil extracellular traps. J Exp Med 2010; 207: 1853-1862.
    CrossrefPubMedGoogle Scholar
  • 12 Martinod K, Demers M, Fuchs TA. et al. Neutrophil histone modification by peptidylarginine deiminase 4 is critical for deep vein thrombosis in mice. Proc Natl Acad Sci USA 2013; 110: 8674-8679.
    CrossrefPubMedGoogle Scholar
  • 13 Etulain J, Martinod K, Wong SL. et al. P-selectin promotes neutrophil extracellular trap formation in mice. Blood 2015; 126: 242-246.
    CrossrefPubMedGoogle Scholar
  • 14 Adams Jr HP, Del Zoppo G, Alberts MJ. et al. Guidelines for the early management of adults with ischemic stroke: a guideline from the American Heart Association/American Stroke Association Stroke Council, Clinical Cardiology Council, Cardiovascular Radiology and Intervention Council, and the Atherosclerotic Peripheral Vascular Disease and Quality of Care Outcomes in Research Interdisciplinary Working Groups: The American Academy of Neurology affirms the value of this guideline as an educational tool for neurologists. Circulation 2007; 115: 478-534.
    CrossrefPubMedGoogle Scholar
  • 15 Banks JL, Marotta CA. Outcomes validity and reliability of the modified Rankin scale: implications for stroke clinical trials: a literature review and synthesis. Stroke 2007; 38: 1091-1096.
    CrossrefPubMedGoogle Scholar
  • 16 Borissoff JI, Joosen IA, Versteylen MO. et al. Elevated levels of circulating DNA and chromatin are independently associated with severe coronary atherosclerosis and a prothrombotic state. Arterioscler Thromb Vasc Biol 2013; 33: 2032-2040.
    CrossrefPubMedGoogle Scholar
  • 17 Wong SL, Demers M, Martinod K. et al. Diabetes primes neutrophils to undergo NETosis, which impairs wound healing. Nat Med 2015; 21: 815-819.
    CrossrefPubMedGoogle Scholar
  • 18 Adams Jr HP, Davis PH, Leira EC. et al. Baseline NIH Stroke Scale score strongly predicts outcome after stroke: A report of the Trial of Org 10172 in Acute Stroke Treatment (TOAST). Neurology 1999; 53: 126-131.
    CrossrefPubMedGoogle Scholar
  • 19 Fuchs TA, Brill A, Duerschmied D. et al. Extracellular DNA traps promote thrombosis. Proc Natl Acad Sci USA 2010; 107: 15880-15885.
    CrossrefPubMedGoogle Scholar
  • 20 Mangold A, Alias S, Scherz T. et al. Coronary neutrophil extracellular trap burden and deoxyribonuclease activity in ST-elevation acute coronary syndrome are predictors of ST-segment resolution and infarct size. Circ Res 2015; 116: 1182-1192.
    CrossrefPubMedGoogle Scholar
  • 21 Savchenko AS, Martinod K, Seidman MA. et al. Neutrophil extracellular traps form predominantly during the organizing stage of human venous thromboembolism development. J Thromb Haemost 2014; 12: 860-870.
    CrossrefPubMedGoogle Scholar
  • 22 van Montfoort ML, Stephan F, Lauw MN. et al. Circulating nucleosomes and neutrophil activation as risk factors for deep vein thrombosis. Arterioscler Thromb Vasc Biol 2013; 33: 147-151.
    CrossrefPubMedGoogle Scholar
  • 23 Diaz JA, Fuchs TA, Jackson TO. et al. Plasma DNA is Elevated in Patients with Deep Vein Thrombosis. J Vasc Surg Venous Lymphat Disord 2013; 01: 10-1016.
    PubMedGoogle Scholar
  • 24 Geiger S, Holdenrieder S, Stieber P. et al. Nucleosomes in serum of patients with early cerebral stroke. Cerebrovasc Dis 2006; 21: 32-37.
    CrossrefPubMedGoogle Scholar
  • 25 Lam NY, Rainer TH, Wong LK. et al. Plasma DNA as a prognostic marker for stroke patients with negative neuroimaging within the first 24 h of symptom onset. Resuscitation 2006; 68: 71-78.
    CrossrefPubMedGoogle Scholar
  • 26 Tsai NW, Lin TK, Chen SD. et al. The value of serial plasma nuclear and mitochondrial DNA levels in patients with acute ischemic stroke. Clin Chim Acta 2011; 412: 476-479.
    CrossrefPubMedGoogle Scholar
  • 27 Rainer TH, Wong LK, Lam W. et al. Prognostic use of circulating plasma nucleic acid concentrations in patients with acute stroke. Clin Chem 2003; 49: 562-569.
    CrossrefPubMedGoogle Scholar
  • 28 Perez-de-Puig I, Miro-Mur F, Ferrer-Ferrer M. et al. Neutrophil recruitment to the brain in mouse and human ischemic stroke. Acta Neuropathol 2015; 129: 239-257.
    CrossrefPubMedGoogle Scholar
  • 29 Thalin C, Demers M, Blomgren B. et al. NETosis promotes cancer-associated arterial microthrombosis presenting as ischemic stroke with troponin elevation. Thromb Res 2016; 139: 56-64.
    CrossrefPubMedGoogle Scholar
  • 30 De Meyer SF, Denorme F, Langhauser F. et al. Thromboinflammation in Stroke Brain Damage. Stroke 2016; 47: 1165-1172.
    CrossrefPubMedGoogle Scholar
  • 31 Gunzer M. Traps and hyper inflammation – new ways that neutrophils promote or hinder survival. Br J Haematol 2014; 164: 189-199.
    CrossrefPubMedGoogle Scholar
  • 32 Caudrillier A, Kessenbrock K, Gilliss BM. et al. Platelets induce neutrophil extracellular traps in transfusion-related acute lung injury. J Clin Invest 2012; 122: 2661-2671.
    CrossrefPubMedGoogle Scholar
  • 33 Gupta S, Agrawal A. Inflammation & autoimmunity in human ageing: dendritic cells take a center stage. Indian J Med Res 2013; 138: 711-716.
    PubMedGoogle Scholar
  • 34 Lindsberg PJ, Roine RO. Hyperglycemia in acute stroke. Stroke 2004; 35: 363-364.
    CrossrefPubMedGoogle Scholar
  • 35 Quagliaro L, Piconi L, Assaloni R. et al. Intermittent high glucose enhances apoptosis related to oxidative stress in human umbilical vein endothelial cells: the role of protein kinase C and NAD(P)H-oxidase activation. Diabetes 2003; 52: 2795-2804.
    CrossrefPubMedGoogle Scholar
  • 36 Rodriguez-Espinosa O, Rojas-Espinosa O, Moreno-Altamirano MM. et al. Metabolic requirements for neutrophil extracellular traps formation. Immunology 2015; 145: 213-224.
    CrossrefPubMedGoogle Scholar
  • 37 Menegazzo L, Ciciliot S, Poncina N. et al. NETosis is induced by high glucose and associated with type 2 diabetes. Acta Diabetol 2015; 52: 497-503.
    CrossrefPubMedGoogle Scholar
  • 38 Hosseinzadeh A, Thompson PR, Segal BH, Urban CF. Nicotine induces neutrophil extracellular traps. J Leukoc Biol 2016; 100: 1105-1112.
    CrossrefPubMedGoogle Scholar
  • 39 Lee J, Luria A, Rhodes C. et al. Nicotine drives neutrophil extracellular traps formation and accelerates collagen-induced arthritis. Rheumatology 2017; 56: 644-653.
    PubMedGoogle Scholar
  • 40 Mehta H, Nazzal K, Sadikot RT. Cigarette smoking and innate immunity. Inflamm Res 2008; 57: 497-503.
    CrossrefPubMedGoogle Scholar
  • 41 Ali SF, Smith EE, Bhatt DL. et al. Paradoxical association of smoking with in-hospital mortality among patients admitted with acute ischemic stroke. J Am Heart Assoc 2013; 02: 000171.
    CrossrefPubMedGoogle Scholar
  • 42 Kufner A, Nolte CH, Galinovic I. et al. Smoking-thrombolysis paradox: recanalization and reperfusion rates after intravenous tissue plasminogen activator in smokers with ischemic stroke. Stroke 2013; 44: 407-413.
    CrossrefPubMedGoogle Scholar
  • 43 Fekete K, Szatmari S, Szocs I. et al. Prestroke alcohol consumption and smoking are not associated with stroke severity, disability at discharge, and case fatality. J Stroke Cerebrovasc Dis 2014; 23: 31-37.
    CrossrefPubMedGoogle Scholar
  • 44 Engelmann MD, Svendsen JH. Inflammation in the genesis and perpetuation of atrial fibrillation. Eur Heart J 2005; 26: 2083-2092.
    CrossrefPubMedGoogle Scholar
  • 45 Harada M, Van Wagoner DR, Nattel S. Role of inflammation in atrial fibrillation pathophysiology and management. Circ J 2015; 79: 495-502.
    CrossrefPubMedGoogle Scholar
  • 46 Conway DS, Buggins P, Hughes E. et al. Prognostic significance of raised plasma levels of interleukin-6 and C-reactive protein in atrial fibrillation. Am Heart J 2004; 148: 462-466.
    CrossrefPubMedGoogle Scholar
  • 47 Aviles RJ, Martin DO, Apperson-Hansen C. et al. Inflammation as a risk factor for atrial fibrillation. Circulation 2003; 108: 3006-3010.
    CrossrefPubMedGoogle Scholar
  • 48 Boos CJ, Anderson RA, Lip GY. Is atrial fibrillation an inflammatory disorder?. Eur Heart J 2006; 27: 136-149.
    CrossrefPubMedGoogle Scholar
  • 49 Marin F, Corral J, Roldan V. et al. Factor XIII Val34Leu polymorphism modulates the prothrombotic and inflammatory state associated with atrial fibrillation. J Mol Cell Cardiol 2004; 37: 699-704.
    CrossrefPubMedGoogle Scholar
  • 50 Shantsila E, Lip GY. Stroke in atrial fibrillation and improving the identification of ’high-risk’ patients: the crossroads of immunity and thrombosis. J Thromb Haemost 2015; 13: 1968-1970.
    CrossrefPubMedGoogle Scholar
  • 51 Licata G, Tuttolomondo A, Di Raimondo D. et al. Immuno-inflammatory activation in acute cardio-embolic strokes in comparison with other subtypes of ischaemic stroke. Thromb Haemost 2009; 101: 929-937.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 52 Piccardi B, Giralt D, Bustamante A. et al. Blood markers of inflammation and endothelial dysfunction in cardioembolic stroke: systematic review and meta-analysis. Biomarkers 2017; 22: 200-209.
    CrossrefPubMedGoogle Scholar
  • 53 Vogel B, Shinagawa H, Hofmann U. et al. Acute DNase1 treatment improves left ventricular remodeling after myocardial infarction by disruption of free chromatin. Basic Res Cardiol 2015; 110 (15) 015-0472.
    CrossrefPubMedGoogle Scholar
  • 54 Lewis HD, Liddle J, Coote JE. et al. Inhibition of PAD4 activity is sufficient to disrupt mouse and human NET formation. Nat Chem Biol 2015; 11: 189-191.
    CrossrefPubMedGoogle Scholar