{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T22:09:18Z","timestamp":1791929358692,"version":"4.3.4"},"reference-count":48,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2020,1,22]],"date-time":"2020-01-22T00:00:00Z","timestamp":1579651200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["JCM"],"abstract":"<jats:p>Neutrophil extracellular traps (NETs) are supposed to play a central role in atherothrombosis. We measured circulating citrullinated histone H3 (H3Cit) and cell-free DNA (cfDNA), which serve as surrogate markers of NET formation, in 79 patients with peripheral artery disease (PAD) following infrainguinal angioplasty with stent implantation. Analysis of cfDNA and H3Cit was performed using Quant-iT\u2122 PicoGreen\u00ae dsDNA Assay Kit or an ELISA, respectively. Within two years of follow-up, the primary endpoint defined as nonfatal myocardial infarction, stroke or transient ischemic attack, cardiovascular death, and &gt;80% target vessel restenosis occurred in 34 patients (43%). Both H3Cit (HR per 1-SD: 2.72; 95% CI: 1.2\u20136.3; p = 0.019) and cfDNA (HR per 1-SD: 2.15; 95% CI: 1.1\u20134.2; p = 0.028) were associated with the primary endpoint in a univariate Cox regression analysis. Multivariate linear regression analyses showed associations between cfDNA and platelet surface expression of P-selectin (p = 0.006) and activated glycoprotein IIb\/IIIa (p &lt; 0.001) in response to arachidonic acid (AA) after adjustment for age, sex, clinical risk factors, and inflammatory markers. H3Cit was also associated with P-selectin expression in response to thrombin-receptor activating peptide (p = 0.048) and AA (p = 0.032). Circulating H3Cit and cfDNA predict ischemic outcomes after peripheral angioplasty with stent implantation, and are associated with on-treatment platelet activation in stable PAD.<\/jats:p>","DOI":"10.3390\/jcm9020304","type":"journal-article","created":{"date-parts":[[2020,1,24]],"date-time":"2020-01-24T11:01:00Z","timestamp":1579863660000},"page":"304","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":33,"title":["Surrogate Markers of Neutrophil Extracellular Trap Formation are Associated with Ischemic Outcomes and Platelet Activation after Peripheral Angioplasty and Stenting"],"prefix":"10.3390","volume":"9","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0601-0667","authenticated-orcid":false,"given":"Svitlana","family":"Demyanets","sequence":"first","affiliation":[{"name":"Department of Laboratory Medicine, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stefan","family":"Stojkovic","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lisa-Marie","family":"Mauracher","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christoph W.","family":"Kopp","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Johann","family":"Wojta","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"},{"name":"Core Facilities, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"},{"name":"Ludwig Boltzmann Institute for Cardiovascular Research, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Johannes","family":"Thaler","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Simon","family":"Panzer","sequence":"additional","affiliation":[{"name":"Department of Blood Group Serology and Transfusion Medicine, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9554-7292","authenticated-orcid":false,"given":"Thomas","family":"Gremmel","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Medical University of Vienna, Waehringer-Guertel 18-20, 1090 Vienna, Austria"},{"name":"Department of Internal Medicine, Cardiology and Nephrology, Landesklinikum Wiener Neustadt, Corvinusring 3-5, 2700 Wiener Neustadt, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,1,22]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"e1020","DOI":"10.1016\/S2214-109X(19)30255-4","article-title":"Global, regional, and national prevalence and risk factors for peripheral artery disease in 2015: An updated systematic review and analysis","volume":"7","author":"Song","year":"2019","journal-title":"Lancet Glob. Health"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"187","DOI":"10.2147\/VHRM.S209241","article-title":"The burden of critical limb ischemia: A review of recent literature","volume":"15","author":"Duff","year":"2019","journal-title":"Vasc. Health Risk Manag."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"332","DOI":"10.1038\/s41569-018-0005-0","article-title":"Interventions for lower extremity peripheral artery disease","volume":"15","author":"Hiramoto","year":"2018","journal-title":"Nat. Rev. Cardiol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"507","DOI":"10.1016\/j.atherosclerosis.2015.05.007","article-title":"The role of vascular biomarkers for primary and secondary prevention. A position paper from the European Society of Cardiology Working Group on peripheral circulation: Endorsed by the Association for Research into Arterial Structure and Physiology (ARTERY) Society","volume":"241","author":"Vlachopoulos","year":"2015","journal-title":"Atherosclerosis"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1532","DOI":"10.1126\/science.1092385","article-title":"Neutrophil extracellular traps kill bacteria","volume":"303","author":"Brinkmann","year":"2004","journal-title":"Science"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1394","DOI":"10.1093\/eurheartj\/ehv044","article-title":"TLR2 and neutrophils potentiate endothelial stress, apoptosis and detachment: Implications for superficial erosion","volume":"36","author":"Quillard","year":"2015","journal-title":"Eur. Heart J."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1405","DOI":"10.1093\/eurheartj\/ehv007","article-title":"Expression of functional tissue factor by neutrophil extracellular traps in culprit artery of acute myocardial infarction","volume":"36","author":"Stakos","year":"2015","journal-title":"Eur. Heart J."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1182","DOI":"10.1161\/CIRCRESAHA.116.304944","article-title":"Coronary neutrophil extracellular trap burden and deoxyribonuclease activity in ST-elevation acute coronary syndrome are predictors of ST-segment resolution and infarct size","volume":"116","author":"Mangold","year":"2015","journal-title":"Circ. Res."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1078","DOI":"10.1055\/s-0038-1641749","article-title":"Neutrophil Extracellular Traps Participate in All Different Types of Thrombotic and Haemorrhagic Complications of Coronary Atherosclerosis","volume":"118","author":"Pertiwi","year":"2018","journal-title":"Thromb. Haemost."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Novotny, J., Chandraratne, S., Weinberger, T., Philippi, V., Stark, K., Ehrlich, A., Pircher, J., Konrad, I., Oberdieck, P., and Titova, A. (2018). Histological comparison of arterial thrombi in mice and men and the influence of Cl-amidine on thrombus formation. PLoS ONE, 13.","DOI":"10.1371\/journal.pone.0190728"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1016\/j.thromres.2019.01.006","article-title":"Neutrophil extracellular traps in thrombi retrieved during interventional treatment of ischemic arterial diseases","volume":"175","author":"Farkas","year":"2019","journal-title":"Thromb. Res."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1977","DOI":"10.1161\/ATVBAHA.114.304114","article-title":"Neutrophil extracellular traps promote thrombin generation through platelet-dependent and platelet-independent mechanisms","volume":"34","author":"Gould","year":"2014","journal-title":"Arter. Thromb. Vasc. Biol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"659","DOI":"10.1160\/TH14-05-0455","article-title":"Neutrophil extracellular traps (NETs) and the role of platelets in infection","volume":"112","author":"Andrews","year":"2014","journal-title":"Thromb. Haemost."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"271","DOI":"10.3389\/fimmu.2016.00271","article-title":"Platelets: New Bricks in the Building of Neutrophil Extracellular Traps","volume":"7","author":"Carestia","year":"2016","journal-title":"Front. Immunol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"2689","DOI":"10.4049\/jimmunol.1201719","article-title":"Neutrophil extracellular traps: Double-edged swords of innate immunity","volume":"189","author":"Kaplan","year":"2012","journal-title":"J. Immunol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"887","DOI":"10.1038\/nm.2184","article-title":"Reciprocal coupling of coagulation and innate immunity via neutrophil serine proteases","volume":"16","author":"Massberg","year":"2010","journal-title":"Nat. Med."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"819","DOI":"10.1084\/jem.20112322","article-title":"Monocytes, neutrophils, and platelets cooperate to initiate and propagate venous thrombosis in mice in vivo","volume":"209","author":"Stark","year":"2012","journal-title":"J. Expr. Med."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"15880","DOI":"10.1073\/pnas.1005743107","article-title":"Extracellular DNA traps promote thrombosis","volume":"107","author":"Fuchs","year":"2010","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1364","DOI":"10.1093\/eurheartj\/ehv105","article-title":"Neutrophil extracellular traps: A new source of tissue factor in atherothrombosis","volume":"36","author":"Badimon","year":"2015","journal-title":"Eur. Heart J."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"542","DOI":"10.1055\/s-0039-1678664","article-title":"Therapeutic Targeting of Neutrophil Extracellular Traps in Atherogenic Inflammation","volume":"119","author":"Maegdefessel","year":"2019","journal-title":"Thromb. Haemost."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1266","DOI":"10.1093\/cvr\/cvz084","article-title":"Novel findings in neutrophil biology and their impact on cardiovascular disease","volume":"115","author":"Bonaventura","year":"2019","journal-title":"Cardiovasc. Res."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"706","DOI":"10.1007\/s12026-017-8905-3","article-title":"Validation of an enzyme-linked immunosorbent assay for the quantification of citrullinated histone H3 as a marker for neutrophil extracellular traps in human plasma","volume":"65","author":"Thalin","year":"2017","journal-title":"Immunol. Res."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1919","DOI":"10.1160\/TH17-02-0130","article-title":"Neutrophil extracellular traps are increased in patients with acute ischemic stroke: Prognostic significance","volume":"117","author":"Valles","year":"2017","journal-title":"Thromb. Haemost."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"508","DOI":"10.1111\/jth.13951","article-title":"Citrullinated histone H3, a biomarker of neutrophil extracellular trap formation, predicts the risk of venous thromboembolism in cancer patients","volume":"16","author":"Mauracher","year":"2018","journal-title":"J. Thromb. Haemost."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Thalin, C., Lundstrom, S., Seignez, C., Daleskog, M., Lundstrom, A., Henriksson, P., Helleday, T., Phillipson, M., Wallen, H., and Demers, M. (2018). Citrullinated histone H3 as a novel prognostic blood marker in patients with advanced cancer. PLoS ONE, 13.","DOI":"10.1371\/journal.pone.0191231"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"311","DOI":"10.1111\/bjh.15906","article-title":"Citrullinated histone H3, a biomarker for neutrophil extracellular trap formation, predicts the risk of mortality in patients with cancer","volume":"186","author":"Grilz","year":"2019","journal-title":"Br. J. Haematol."},{"key":"ref_27","first-page":"474","article-title":"In vivo and protease-activated receptor-1-mediated platelet activation but not response to antiplatelet therapy predict two-year outcomes after peripheral angioplasty with stent implantation","volume":"111","author":"Gremmel","year":"2014","journal-title":"Thromb. Haemost."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"738","DOI":"10.1177\/1526602816655521","article-title":"Serum Cholinesterase Levels Are Associated With 2-Year Ischemic Outcomes After Angioplasty and Stenting for Peripheral Artery Disease","volume":"23","author":"Gremmel","year":"2016","journal-title":"J. Endovasc. Ther."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"333","DOI":"10.1160\/TH08-09-0577","article-title":"Comparison of methods to evaluate clopidogrel-mediated platelet inhibition after percutaneous intervention with stent implantation","volume":"101","author":"Gremmel","year":"2009","journal-title":"Thromb. Haemost."},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Gremmel, T., Koppensteiner, R., and Panzer, S. (2015). Comparison of Aggregometry with Flow Cytometry for the Assessment of Agonists -Induced Platelet Reactivity in Patients on Dual Antiplatelet Therapy. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0129666"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"119","DOI":"10.1016\/j.atherosclerosis.2013.10.027","article-title":"Response to antiplatelet therapy and platelet reactivity to thrombin receptor activating peptide-6 in cardiovascular interventions: Differences between peripheral and coronary angioplasty","volume":"232","author":"Gremmel","year":"2014","journal-title":"Atherosclerosis"},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Budczies, J., Klauschen, F., Sinn, B.V., Gyorffy, B., Schmitt, W.D., Darb-Esfahani, S., and Denkert, C. (2012). Cutoff Finder: A comprehensive and straightforward Web application enabling rapid biomarker cutoff optimization. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0051862"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"327","DOI":"10.23736\/S0392-9590.18.03996-2","article-title":"Epidemiology of peripheral artery disease in Europe: VAS Educational Paper","volume":"37","author":"Olinic","year":"2018","journal-title":"Int. Angiol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"290","DOI":"10.1160\/TH12-06-0425","article-title":"Neutrophils, neutrophil extracellular traps and interleukin-17 associate with the organisation of thrombi in acute myocardial infarction","volume":"109","author":"Li","year":"2013","journal-title":"Thromb. Haemost."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"2032","DOI":"10.1161\/ATVBAHA.113.301627","article-title":"Elevated levels of circulating DNA and chromatin are independently associated with severe coronary atherosclerosis and a prothrombotic state","volume":"33","author":"Borissoff","year":"2013","journal-title":"Arterioscler. Thromb. Vasc. Biol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1538","DOI":"10.1093\/eurheartj\/ehv419","article-title":"Histopathological evaluation of thrombus in patients presenting with stent thrombosis. A multicenter European study: A report of the prevention of late stent thrombosis by an interdisciplinary global European effort consortium","volume":"37","author":"Riegger","year":"2016","journal-title":"Eur. Heart J."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1055\/s-0035-1564835","article-title":"Platelet Physiology","volume":"42","author":"Gremmel","year":"2016","journal-title":"Semin. Thromb. Hemost."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"610","DOI":"10.1016\/j.jvs.2005.01.013","article-title":"Association of neutrophils and future cardiovascular events in patients with peripheral artery disease","volume":"41","author":"Haumer","year":"2005","journal-title":"J. Vasc. Surg."},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Mauracher, L.M., Buchtele, N., Schorgenhofer, C., Weiser, C., Herkner, H., Merrelaar, A., Spiel, A.O., Hell, L., Ay, C., and Pabinger, I. (2019). Increased Citrullinated Histone H3 Levels in the Early Post-Resuscitative Period Are Associated with Poor Neurologic Function in Cardiac Arrest Survivors-A Prospective Observational Study. J. Clin. Med., 8.","DOI":"10.3390\/jcm8101568"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1021","DOI":"10.1182\/blood-2016-06-722298","article-title":"In vitro activation of coagulation by human neutrophil DNA and histone proteins but not neutrophil extracellular traps","volume":"129","author":"Noubouossie","year":"2017","journal-title":"Blood"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1795","DOI":"10.1111\/j.1538-7836.2011.04422.x","article-title":"Extracellular histones increase plasma thrombin generation by impairing thrombomodulin-dependent protein C activation","volume":"9","author":"Ammollo","year":"2011","journal-title":"J. Thromb. Haemost."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"e12871","DOI":"10.1111\/eci.12871","article-title":"Decision shaping neutrophil-platelet interplay in inflammation: From physiology to intervention","volume":"48","author":"Soehnlein","year":"2018","journal-title":"Eur. J. Clin. Investig."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"806","DOI":"10.1160\/TH14-08-0690","article-title":"Impact of variables of the P-selectin\u2014P-selectin glycoprotein ligand-1 axis on leukocyte-platelet interactions in cardiovascular disease","volume":"113","author":"Gremmel","year":"2015","journal-title":"Thromb. Haemost."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"2573","DOI":"10.1182\/blood-2013-07-516484","article-title":"Synchronized integrin engagement and chemokine activation is crucial in neutrophil extracellular trap-mediated sterile inflammation","volume":"123","author":"Rossaint","year":"2014","journal-title":"Blood"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"1234","DOI":"10.1126\/science.1256478","article-title":"Neutrophils scan for activated platelets to initiate inflammation","volume":"346","author":"Sreeramkumar","year":"2014","journal-title":"Science"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"242","DOI":"10.1182\/blood-2015-01-624023","article-title":"P-selectin promotes neutrophil extracellular trap formation in mice","volume":"126","author":"Etulain","year":"2015","journal-title":"Blood"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1952","DOI":"10.1182\/blood-2011-03-343061","article-title":"Extracellular histones promote thrombin generation through platelet-dependent mechanisms: Involvement of platelet TLR2 and TLR4","volume":"118","author":"Semeraro","year":"2011","journal-title":"Blood"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"430","DOI":"10.1124\/jpet.112.202879","article-title":"Regulation of neutrophil extracellular trap formation by anti-inflammatory drugs","volume":"345","author":"Lapponi","year":"2013","journal-title":"J. Pharmacol. Expr. Ther."}],"container-title":["Journal of Clinical Medicine"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2077-0383\/9\/2\/304\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,13]],"date-time":"2025-10-13T13:44:24Z","timestamp":1760363064000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2077-0383\/9\/2\/304"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,1,22]]},"references-count":48,"journal-issue":{"issue":"2","published-online":{"date-parts":[[2020,2]]}},"alternative-id":["jcm9020304"],"URL":"https:\/\/doi.org\/10.3390\/jcm9020304","relation":{},"ISSN":["2077-0383"],"issn-type":[{"value":"2077-0383","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,1,22]]}}}