Traps N' Clots: NET-Mediated Thrombosis and Related Diseases

Insights into Neutrophils and NETs: A Historical Perspective

Vessel wall injury and subsequent blood extravasation activates a series of local biological processes to prevent excess blood loss via the formation of hemostatic plug strictly restricted at the site of vascular injury with minimal or no extension in the vessel lumen.[1] In the vast majority of cases, a catastrophic systemic activation of these processes is contained by specific mechanisms. As opposed to hemostasis, thrombosis is characterized by the deregulated clot formation, various degrees of vessel occlusion, tissue ischemia, and necrosis.[1]

A large body of accumulating experimental and clinical data over the past 20 years has clearly indicated the reciprocal relationship and dynamic interplay between inflammation and thrombosis.[2] [3] [4] [5] Today, targeting inflammation to prevent thrombotic events represents a realistic and promising therapeutic approach.[6] Among immune cell subsets that are implicated in multiple molecular pathways during inflammatory response, neutrophils have a crucial role, recruited first to the site of injury following instructive signals from the tissue environment.[7]

The very first observation linking neutrophils with thromboinflammation was reported almost 70 years ago describing granulocytes as a main component of clotted blood in patients suffering from active lupus erythematosus.[8] During the following years, although several studies had described the accumulation of neutrophils at the site of thrombus formation, these cells remained neglected and less studied in many thrombotic diseases. The traditional aspect of neutrophils as dispensable, passive bystanders was dramatically revised after the milestone discovery that they represent a primary source of blood-borne tissue factor (TF), the main in vivo initiator of the extrinsic coagulation cascade, resulting in thrombin generation and ensuing thrombus formation.[9] [10] Later on, several studies from our laboratory and others provided evidence for the critical role of neutrophils in thrombosis and inflammation-mediated thrombotic complications.[3] [11] [12] Intravital microscopy studies in mouse models of venous and arterial thrombosis demonstrated neutrophil recruitment and activation at the site of endothelial damage in the early phase of thrombosis.[13] [14] Of note, neutrophils are not only implicated in thrombotic processes, but also seem to be indispensable for thrombosis. Neutropenia induced in vivo by anti-Ly6G or GR-1 antibody abrogated venous[13] and arterial[14] thrombosis, respectively. When purified neutrophils from wild-type mice were injected into transgenic mice that express no mouse TF and only minimal (< 1%) amounts of human TF (low TF mice), the defective fibrin generation in these animals was restored, indicating that TF expressing neutrophils represent the main source of TF during thrombus formation.[14] Similarly, when normal TF expressing mice were transplanted with low-human TF bone marrow cells they did not develop deep vein thrombosis (DVT).[13] However, the mechanisms underlying the activation and delivery of active TF by neutrophils remained unknown.

During the last years, advances in molecular biology provided the most exciting update of neutrophil physiology, in particular their capacity to release neutrophil extracellular traps (NETs). NETs are extracellular web-like structures of chromatin fibers lined with various highly active proteases and proteins of nuclear, granular, and cytosolic origin.[15] The release of NETs from activated neutrophils was initially described in 2004 as a novel defense mechanism able to “trap and kill” a wide range of pathogens.[16] However, increasing evidence during the past few years highlighted their fundamental role in the pathogenesis of numerous noninfectious inflammatory disorders, such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), autoimmune vasculitis, gout, ulcerative colitis, interleukin (IL)-1β-mediated autoinflammatory syndromes, and thrombosis.[15] [17] [18] [19]

Activated platelets are able to induce robust NET release within vasculature providing a scaffold for fibrin deposition and stabilization of thrombus.[20] [21] [22] [23] Notably, these NETs are decorated with functionally active TF, which explains its extracellular delivery at the site of tissue injury.[13] [19] [24] Besides TF, NETs were found to deliver several proteins and clot factors involved in thrombosis such as von Willebrand factor (vWF), XII, fibrinogen, and fibronectin.[2] [13] [25] The thrombogenic potential of NETs was further supported by experimental studies indicating that extracellular histones induce endothelial activation, platelet activation/aggregation, and thrombin generation[26] [27] ([Fig. 1]). Phylogenetically, the capacity of NETs to activate coagulation serves to trap and eliminate pathogens, resembling primitive defense systems operating several millions years ago, and it is conserved today in insects.[28] In these organisms, coagulation and immunity use common mechanisms to prevent fluid loss and pathogen invasion. These systems are tightly interrelated, since NET-associated antimicrobial proteases are able to trigger several coagulation pathways,[2] while activation of the coagulation system supports several immune responses such as bacterial compartmentalization, immobilization, and elimination especially in microvasculature (immunothrombosis). Probably, much of the NET-mediated antimicrobial effect is due to entrapment, rather than direct killing[29] [30] ([Fig. 2]).

Fig. 1 Mechanisms of neutrophil extracellular trap (NET) thrombogenicity. Left: Platelets-PolyP-neutrophils-NETs interactivation leads to NET generation. (–) denotes NET autoregulation. Right: NETs can deliver thrombogenic signals through many different mechanisms presented here. APC, activated protein C; FX, FXII, FV, coagulation factors; PolyP: polyphosphate; TF, tissue factor; TFPI, TF pathway inhibitor; tPA, tissue plasminogen activator.

Fig. 2 Skewing of immunothrombosis (tissue protection) to pathological thrombosis (tissue damage). In primitive organisms (bottom, A) immunity and coagulation use a common, hemocyte-based system to prevent fluid loss and pathogen invasion. In mammals including humans (B) hemostasis and immunity are operated by distinct systems (platelets and coagulation, left; and white blood cells such as neutrophils, right, respectively). Interaction between platelets/coagulation and neutrophils leads to neutrophil extracellular trap (NET) formation (C). In cases of a pathogen invasion, NETs are involved in pathogen entrapment and elimination in microcirculation (green path, host defense, D). On the contrary, inappropriate NET formation in cases of sterile inflammation (autoimmune or inflammatory environment) leads to thrombotic complications (red path, tissue damage) in microcirculation (E) or large vessels (F). DIC, disseminated intravascular coagulation.

Apart from neutrophils, extracellular traps (ETs) formation has also been described in other types of granulocytes, such as eosinophils and mast cells.[31] [32] Very recently data implicate macrophages, mast cells, and eosinophils through ETs generation in atherosclerotic plaque formation and thrombosis.[33] [34] However, ETs formation in macrophages is controversial and remains unclear whether it is distinct from pyroptosis.[35]

In view of the above, NETs could be perceived as a double-edged sword during disease processes. They may be beneficial by enhancing the antimicrobial potential in numerous infectious diseases or contributing to normal hemostasis and pathogen curbing in neutrophil clots lattice, but also harmful by amplifying systemic or local inflammation leading to tissue damage and thrombosis[15] [36] ([Fig. 2]). Therefore, NETs and their components emerge today as novel candidate for diagnostic and therapeutic targets of thrombosis in many clinical settings.[12] [15] [37] [38]

^* These authors contributed equally to this work.

• References

• 1 Jackson SP. Arterial thrombosis--insidious, unpredictable and deadly. Nat Med 2011; 17 (11) 1423-1436
• 2 Massberg S, Grahl L, von Bruehl M-L. , et al. Reciprocal coupling of coagulation and innate immunity via neutrophil serine proteases. Nat Med 2010; 16 (08) 887-896
• 3 Kambas K, Mitroulis I, Ritis K. The emerging role of neutrophils in thrombosis-the journey of TF through NETs. Front Immunol 2012; 3: 385
• 4 Jackson SP, Darbousset R, Schoenwaelder SM. Thromboinflammation: challenges of therapeutically targeting coagulation and other host defense mechanisms. Blood 2019; 133 (09) 906-918
• 5 Burzynski LC, Humphry M, Pyrillou K. , et al. The coagulation and immune systems are directly linked through the activation of interleukin-1α by thrombin. Immunity 2019; 50 (04) 1033-1042
• 6 Ridker PM, Everett BM, Thuren T. , et al; CANTOS Trial Group. Antiinflammatory therapy with canakinumab for atherosclerotic disease. N Engl J Med 2017; 377 (12) 1119-1131
• 7 Mócsai A. Diverse novel functions of neutrophils in immunity, inflammation, and beyond. J Exp Med 2013; 210 (07) 1283-1299
• 8 Gonyea LM, Kallsen RA, Marlow AA. The occurrence of the “L. E.” cell in clotted blood. J Invest Dermatol 1950; 15 (01) 11-12
• 9 Ritis K, Doumas M, Mastellos D. , et al. A novel C5a receptor-tissue factor cross-talk in neutrophils links innate immunity to coagulation pathways. J Immunol 2006; 177 (07) 4794-4802
• 10 Maugeri N, Brambilla M, Camera M. , et al. Human polymorphonuclear leukocytes produce and express functional tissue factor upon stimulation. J Thromb Haemost 2006; 4 (06) 1323-1330
• 11 Kapoor S, Opneja A, Nayak L. The role of neutrophils in thrombosis. Thromb Res 2018; 170: 87-96
• 12 Bonaventura A, Montecucco F, Dallegri F. , et al. Novel findings in neutrophil biology and their impact on cardiovascular disease. Cardiovasc Res 2019; 115 (08) 1266-1285
• 13 von Brühl M-L, Stark K, Steinhart A. , et al. Monocytes, neutrophils, and platelets cooperate to initiate and propagate venous thrombosis in mice in vivo. J Exp Med 2012; 209 (04) 819-835
• 14 Darbousset R, Thomas GM, Mezouar S. , et al. Tissue factor-positive neutrophils bind to injured endothelial wall and initiate thrombus formation. Blood 2012; 120 (10) 2133-2143
• 15 Mitsios A, Arampatzioglou A, Arelaki S, Mitroulis I, Ritis K. NETopathies? Unraveling the dark side of old diseases through neutrophils. Front Immunol 2017; 7: 678
• 16 Brinkmann V, Reichard U, Goosmann C. , et al. Neutrophil extracellular traps kill bacteria. Science 2004; 303 (5663): 1532-1535
• 17 Jorch SK, Kubes P. An emerging role for neutrophil extracellular traps in noninfectious disease. Nat Med 2017; 23 (03) 279-287
• 18 Jiménez-Alcázar M, Rangaswamy C, Panda R. , et al. Host DNases prevent vascular occlusion by neutrophil extracellular traps. Science 2017; 358 (6367): 1202-1206
• 19 Stakos DA, Kambas K, Konstantinidis T. , et al. Expression of functional tissue factor by neutrophil extracellular traps in culprit artery of acute myocardial infarction. Eur Heart J 2015; 36 (22) 1405-1414
• 20 Caudrillier A, Kessenbrock K, Gilliss BM. , et al. Platelets induce neutrophil extracellular traps in transfusion-related acute lung injury. J Clin Invest 2012; 122 (07) 2661-2671
• 21 Clark SR, Ma AC, Tavener SA. , et al. Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood. Nat Med 2007; 13 (04) 463-469
• 22 Maugeri N, Campana L, Gavina M. , et al. Activated platelets present high mobility group box 1 to neutrophils, inducing autophagy and promoting the extrusion of neutrophil extracellular traps. J Thromb Haemost 2014; 12 (12) 2074-2088
• 23 Rossaint J, Herter JM, Van Aken H. , et al. Synchronized integrin engagement and chemokine activation is crucial in neutrophil extracellular trap-mediated sterile inflammation. Blood 2014; 123 (16) 2573-2584
• 24 Chrysanthopoulou A, Kambas K, Stakos D. , et al. Interferon lambda1/IL-29 and inorganic polyphosphate are novel regulators of neutrophil-driven thromboinflammation. J Pathol 2017; 243 (01) 111-122
• 25 Martinod K, Wagner DD. Thrombosis: tangled up in NETs. Blood 2014; 123 (18) 2768-2776
• 26 Xu J, Zhang X, Pelayo R. , et al. Extracellular histones are major mediators of death in sepsis. Nat Med 2009; 15 (11) 1318-1321
• 27 Fuchs TA, Brill A, Duerschmied D. , et al. Extracellular DNA traps promote thrombosis. Proc Natl Acad Sci U S A 2010; 107 (36) 15880-15885
• 28 Iwanaga S, Lee BL. Recent advances in the innate immunity of invertebrate animals. J Biochem Mol Biol 2005; 38 (02) 128-150
• 29 Engelmann B, Massberg S. Thrombosis as an intravascular effector of innate immunity. Nat Rev Immunol 2013; 13 (01) 34-45
• 30 Loof TG, Mörgelin M, Johansson L. , et al. Coagulation, an ancestral serine protease cascade, exerts a novel function in early immune defense. Blood 2011; 118 (09) 2589-2598
• 31 Yousefi S, Gold JA, Andina N. , et al. Catapult-like release of mitochondrial DNA by eosinophils contributes to antibacterial defense. Nat Med 2008; 14 (09) 949-953
• 32 von Köckritz-Blickwede M, Goldmann O, Thulin P. , et al. Phagocytosis-independent antimicrobial activity of mast cells by means of extracellular trap formation. Blood 2008; 111 (06) 3070-3080
• 33 Pertiwi KR, de Boer OJ, Mackaaij C. , et al. Extracellular traps derived from macrophages, mast cells, eosinophils and neutrophils are generated in a time-dependent manner during atherothrombosis. J Pathol 2019; 247 (04) 505-512
• 34 Marx C, Novotny J, Salbeck D. , et al. Eosinophil-platelet interactions promote atherosclerosis and stabilize thrombosis with eosinophil extracellular traps. Blood 2019; 134 (21) 1859-1872
• 35 Boeltz S, Amini P, Anders H-J. , et al. To NET or not to NET:current opinions and state of the science regarding the formation of neutrophil extracellular traps. Cell Death Differ 2019; 26 (03) 395-408
• 36 Papayannopoulos V. Neutrophil extracellular traps in immunity and disease. Nat Rev Immunol 2018; 18 (02) 134-147
• 37 Mauracher L-M, Posch F, Martinod K. , et al. Citrullinated histone H3, a biomarker of neutrophil extracellular trap formation, predicts the risk of venous thromboembolism in cancer patients. J Thromb Haemost 2018; 16 (03) 508-518
• 38 Barnado A, Crofford LJ, Oates JC. At the Bedside: neutrophil extracellular traps (NETs) as targets for biomarkers and therapies in autoimmune diseases. J Leukoc Biol 2016; 99 (02) 265-278
• 39 Vogel S, Bodenstein R, Chen Q. , et al. Platelet-derived HMGB1 is a critical mediator of thrombosis. J Clin Invest 2015; 125 (12) 4638-4654
• 40 Sollberger G, Tilley DO, Zychlinsky A. Neutrophil extracellular traps: the biology of chromatin externalization. Dev Cell 2018; 44 (05) 542-553
• 41 Chen KW, Monteleone M, Boucher D. , et al. Noncanonical inflammasome signaling elicits gasdermin D-dependent neutrophil extracellular traps. Sci Immunol 2018; 3 (26) 3
• 42 Farrera C, Fadeel B. Macrophage clearance of neutrophil extracellular traps is a silent process. J Immunol 2013; 191 (05) 2647-2656
• 43 Apostolidou E, Skendros P, Kambas K. , et al. Neutrophil extracellular traps regulate IL-1β-mediated inflammation in familial Mediterranean fever. Ann Rheum Dis 2016; 75 (01) 269-277
• 44 Schauer C, Janko C, Munoz LE. , et al. Aggregated neutrophil extracellular traps limit inflammation by degrading cytokines and chemokines. Nat Med 2014; 20 (05) 511-517
• 45 Kolaczkowska E, Jenne CN, Surewaard BGJ. , et al. Molecular mechanisms of NET formation and degradation revealed by intravital imaging in the liver vasculature. Nat Commun 2015; 6: 6673
• 46 Chrysanthopoulou A, Mitroulis I, Apostolidou E. , et al. Neutrophil extracellular traps promote differentiation and function of fibroblasts. J Pathol 2014; 233 (03) 294-307
• 47 Kambas K, Mitroulis I, Apostolidou E. , et al. Autophagy mediates the delivery of thrombogenic tissue factor to neutrophil extracellular traps in human sepsis. PLoS One 2012; 7 (09) e45427
• 48 Angelidou I, Chrysanthopoulou A, Mitsios A. , et al. REDD1/autophagy pathway is associated with neutrophil-driven IL-1β inflammatory response in active ulcerative colitis. J Immunol 2018; 200 (12) 3950-3961
• 49 Skendros P, Chrysanthopoulou A, Rousset F. , et al. Regulated in development and DNA damage responses 1 (REDD1) links stress with IL-1β-mediated familial Mediterranean fever attack through autophagy-driven neutrophil extracellular traps. J Allergy Clin Immunol 2017; 140 (05) 1378-1387
• 50 Lande R, Ganguly D, Facchinetti V. , et al. Neutrophils activate plasmacytoid dendritic cells by releasing self-DNA-peptide complexes in systemic lupus erythematosus. Sci Transl Med 2011; 3 (73) 73ra19
• 51 Arampatzioglou A, Papazoglou D, Konstantinidis T. , et al. Clarithromycin enhances the antibacterial activity and wound healing capacity in type 2 diabetes mellitus by increasing LL-37 load on neutrophil extracellular traps. Front Immunol 2018; 9: 2064
• 52 Frangou E, Chrysanthopoulou A, Mitsios A. , et al. REDD1/autophagy pathway promotes thromboinflammation and fibrosis in human systemic lupus erythematosus (SLE) through NETs decorated with tissue factor (TF) and interleukin-17A (IL-17A). Ann Rheum Dis 2019; 78 (02) 238-248
• 53 Oklu R, Albadawi H, Watkins MT, Monestier M, Sillesen M, Wicky S. Detection of extracellular genomic DNA scaffold in human thrombus: implications for the use of deoxyribonuclease enzymes in thrombolysis. J Vasc Interv Radiol 2012; 23 (05) 712-718
• 54 De Meyer SF, Suidan GL, Fuchs TA, Monestier M, Wagner DD. Extracellular chromatin is an important mediator of ischemic stroke in mice. Arterioscler Thromb Vasc Biol 2012; 32 (08) 1884-1891
• 55 Ge L, Zhou X, Ji W-J. , et al. Neutrophil extracellular traps in ischemia-reperfusion injury-induced myocardial no-reflow: therapeutic potential of DNase-based reperfusion strategy. Am J Physiol Heart Circ Physiol 2015; 308 (05) H500-H509
• 56 Silvestre-Roig C, Braster Q, Wichapong K. , et al. Externalized histone H4 orchestrates chronic inflammation by inducing lytic cell death. Nature 2019; 569 (7755): 236-240
• 57 Folco EJ, Mawson TL, Vromman A. , et al. Neutrophil extracellular traps induce endothelial cell activation and tissue factor production through interleukin-1α and cathepsin G. Arterioscler Thromb Vasc Biol 2018; 38 (08) 1901-1912
• 58 Quillard T, Araújo HA, Franck G, Shvartz E, Sukhova G, Libby P. TLR2 and neutrophils potentiate endothelial stress, apoptosis and detachment: implications for superficial erosion. Eur Heart J 2015; 36 (22) 1394-1404
• 59 Franck G, Mawson T, Sausen G. , et al. Flow perturbation mediates neutrophil recruitment and potentiates endothelial injury via TLR2 in mice: implications for superficial erosion. Circ Res 2017; 121 (01) 31-42
• 60 Megens RTA, Vijayan S, Lievens D. , et al. Presence of luminal neutrophil extracellular traps in atherosclerosis. Thromb Haemost 2012; 107 (03) 597-598
• 61 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 (07) 1182-1192
• 62 Riegger J, Byrne RA, Joner M. , et al; Prevention of Late Stent Thrombosis by an Interdisciplinary Global European Effort (PRESTIGE) Investigators. 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. Eur Heart J 2016; 37 (19) 1538-1549
• 63 de Boer OJ, Li X, Teeling P. , et al. Neutrophils, neutrophil extracellular traps and interleukin-17 associate with the organisation of thrombi in acute myocardial infarction. Thromb Haemost 2013; 109 (02) 290-297
• 64 Maione F, Cicala C, Liverani E, Mascolo N, Perretti M, D'Acquisto F. IL-17A increases ADP-induced platelet aggregation. Biochem Biophys Res Commun 2011; 408 (04) 658-662
• 65 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 (08) 2032-2040
• 66 Geiger S, Holdenrieder S, Stieber P. , et al. Nucleosomes in serum of patients with early cerebral stroke. Cerebrovasc Dis 2006; 21 (1-2): 32-37
• 67 Lam NY-L, Rainer TH, Wong LK-S, Lam W, Lo YM. Plasma DNA as a prognostic marker for stroke patients with negative neuroimaging within the first 24 h of symptom onset. Resuscitation 2006; 68 (01) 71-78
• 68 Vallés J, Lago A, Santos MT. , et al. Neutrophil extracellular traps are increased in patients with acute ischemic stroke: prognostic significance. Thromb Haemost 2017; 117 (10) 1919-1929
• 69 Ducroux C, Di Meglio L, Loyau S. , et al. Thrombus neutrophil extracellular traps content impair tPA-induced thrombolysis in acute ischemic stroke. Stroke 2018; 49 (03) 754-757
• 70 Rangé H, Labreuche J, Louedec L. , et al. Periodontal bacteria in human carotid atherothrombosis as a potential trigger for neutrophil activation. Atherosclerosis 2014; 236 (02) 448-455
• 71 Delbosc S, Alsac J-M, Journe C. , et al. Porphyromonas gingivalis participates in pathogenesis of human abdominal aortic aneurysm by neutrophil activation. Proof of concept in rats. PLoS One 2011; 6 (04) e18679
• 72 Brill A, Fuchs TA, Savchenko AS. , et al. Neutrophil extracellular traps promote deep vein thrombosis in mice. J Thromb Haemost 2012; 10 (01) 136-144
• 73 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 (01) 147-151
• 74 Diaz JA, Fuchs TA, Jackson TO. , et al; for the Michigan Research Venous Group*. Plasma DNA is elevated in patients with deep vein thrombosis. J Vasc Surg Venous Lymphat Disord 2013; 1 (04) 1
• 75 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 (06) 860-870
• 76 Meier TR, Myers Jr DD, Wrobleski SK. , et al. Prophylactic P-selectin inhibition with PSI-421 promotes resolution of venous thrombosis without anticoagulation. Thromb Haemost 2008; 99 (02) 343-351
• 77 Das R, Burke T, Plow EF. Histone H2B as a functionally important plasminogen receptor on macrophages. Blood 2007; 110 (10) 3763-3772
• 78 Varjú I, Longstaff C, Szabó L. , et al. DNA, histones and neutrophil extracellular traps exert anti-fibrinolytic effects in a plasma environment. Thromb Haemost 2015; 113 (06) 1289-1298
• 79 Longstaff C, Varjú I, Sótonyi P. , et al. Mechanical stability and fibrinolytic resistance of clots containing fibrin, DNA, and histones. J Biol Chem 2013; 288 (10) 6946-6956
• 80 Lang IM, Madani M. Update on chronic thromboembolic pulmonary hypertension. Circulation 2014; 130 (06) 508-518
• 81 Aldabbous L, Abdul-Salam V, McKinnon T. , et al. Neutrophil extracellular traps promote angiogenesis: evidence from vascular pathology in pulmonary hypertension. Arterioscler Thromb Vasc Biol 2016; 36 (10) 2078-2087
• 82 Knight JS, Carmona-Rivera C, Kaplan MJ. Proteins derived from neutrophil extracellular traps may serve as self-antigens and mediate organ damage in autoimmune diseases. Front Immunol 2012; 3: 380
• 83 Nakazawa D, Tomaru U, Yamamoto C, Jodo S, Ishizu A. Abundant neutrophil extracellular traps in thrombus of patient with microscopic polyangiitis. Front Immunol 2012; 3: 333
• 84 Imamoto T, Nakazawa D, Shida H. , et al. Possible linkage between microscopic polyangiitis and thrombosis via neutrophil extracellular traps. Clin Exp Rheumatol 2014; 32 (01) 149-150
• 85 Kambas K, Chrysanthopoulou A, Vassilopoulos D. , et al. Tissue factor expression in neutrophil extracellular traps and neutrophil derived microparticles in antineutrophil cytoplasmic antibody associated vasculitis may promote thromboinflammation and the thrombophilic state associated with the disease. Ann Rheum Dis 2014; 73 (10) 1854-1863
• 86 Huang Y-M, Wang H, Wang C, Chen M, Zhao MH. Promotion of hypercoagulability in antineutrophil cytoplasmic antibody-associated vasculitis by C5a-induced tissue factor-expressing microparticles and neutrophil extracellular traps. Arthritis Rheumatol 2015; 67 (10) 2780-2790
• 87 Mitroulis I, Kambas K, Anyfanti P, Doumas M, Ritis K. The multivalent activity of the tissue factor-thrombin pathway in thrombotic and non-thrombotic disorders as a target for therapeutic intervention. Expert Opin Ther Targets 2011; 15 (01) 75-89
• 88 Posma JJ, Grover SP, Hisada Y. , et al. Roles of coagulation proteases and PARs (protease-activated receptors) in mouse models of inflammatory diseases. Arterioscler Thromb Vasc Biol 2019; 39 (01) 13-24
• 89 Leffler J, Stojanovich L, Shoenfeld Y, Bogdanovic G, Hesselstrand R, Blom AM. Degradation of neutrophil extracellular traps is decreased in patients with antiphospholipid syndrome. Clin Exp Rheumatol 2014; 32 (01) 66-70
• 90 Yalavarthi S, Gould TJ, Rao AN. , et al. Release of neutrophil extracellular traps by neutrophils stimulated with antiphospholipid antibodies: a newly identified mechanism of thrombosis in the antiphospholipid syndrome. Arthritis Rheumatol 2015; 67 (11) 2990-3003
• 91 Meng H, Yalavarthi S, Kanthi Y. , et al. In vivo role of neutrophil extracellular traps in antiphospholipid antibody-mediated venous thrombosis. Arthritis Rheumatol 2017; 69 (03) 655-667
• 92 Gollomp K, Kim M, Johnston I. , et al. Neutrophil accumulation and NET release contribute to thrombosis in HIT. JCI Insight 2018; 3 (18) 3
• 93 Perdomo J, Leung HHL, Ahmadi Z. , et al. Neutrophil activation and NETosis are the major drivers of thrombosis in heparin-induced thrombocytopenia. Nat Commun 2019; 10 (01) 1322
• 94 Jiménez-Alcázar M, Napirei M, Panda R. , et al. Impaired DNase1-mediated degradation of neutrophil extracellular traps is associated with acute thrombotic microangiopathies. J Thromb Haemost 2015; 13 (05) 732-742
• 95 Wang Y, Luo L, Braun OÖ. , et al. Neutrophil extracellular trap-microparticle complexes enhance thrombin generation via the intrinsic pathway of coagulation in mice. Sci Rep 2018; 8 (01) 4020
• 96 He Z, Si Y, Jiang T. , et al. Phosphotidylserine exposure and neutrophil extracellular traps enhance procoagulant activity in patients with inflammatory bowel disease. Thromb Haemost 2016; 115 (04) 738-751
• 97 Maugeri N, Capobianco A, Rovere-Querini P. , et al. Platelet microparticles sustain autophagy-associated activation of neutrophils in systemic sclerosis. Sci Transl Med 2018; 10 (451) 10
• 98 Qi H, Yang S, Zhang L. Neutrophil extracellular traps and endothelial dysfunction in atherosclerosis and thrombosis. Front Immunol 2017; 8: 928
• 99 Wiseman SJ, Ralston SH, Wardlaw JM. Cerebrovascular disease in rheumatic diseases: a systematic review and meta-analysis. Stroke 2016; 47 (04) 943-950
• 100 Agca R, Heslinga SC, Rollefstad S. , et al. EULAR recommendations for cardiovascular disease risk management in patients with rheumatoid arthritis and other forms of inflammatory joint disorders: 2015/2016 update. Ann Rheum Dis 2017; 76 (01) 17-28
• 101 Gu M-M, Wang X-P, Cheng Q-Y. , et al. A meta-analysis of cardiovascular events in systemic lupus erythematosus. Immunol Invest 2019; 48 (05) 505-520
• 102 Knight JS, Zhao W, Luo W. , et al. Peptidylarginine deiminase inhibition is immunomodulatory and vasculoprotective in murine lupus. J Clin Invest 2013; 123 (07) 2981-2993
• 103 Grayson PC, Carmona-Rivera C, Xu L. , et al; Rituximab in ANCA-Associated Vasculitis-Immune Tolerance Network Research Group. Neutrophil-related gene expression and low-density granulocytes associated with disease activity and response to treatment in antineutrophil cytoplasmic antibody-associated vasculitis. Arthritis Rheumatol 2015; 67 (07) 1922-1932
• 104 Knight JS, Meng H, Coit P. , et al. Activated signature of antiphospholipid syndrome neutrophils reveals potential therapeutic target. JCI Insight 2017; 2 (18) 2
• 105 Weeding E, Coit P, Yalavarthi S, Kaplan MJ, Knight JS, Sawalha AH. Genome-wide DNA methylation analysis in primary antiphospholipid syndrome neutrophils. Clin Immunol 2018; 196: 110-116
• 106 Ali RA, Gandhi AA, Meng H. , et al. Adenosine receptor agonism protects against NETosis and thrombosis in antiphospholipid syndrome. Nat Commun 2019; 10 (01) 1916
• 107 White C, Noble SIR, Watson M. , et al. Prevalence, symptom burden, and natural history of deep vein thrombosis in people with advanced cancer in specialist palliative care units (HIDDen): a prospective longitudinal observational study. Lancet Haematol 2019; 6 (02) e79-e88
• 108 Spek CA, Versteeg HH, Borensztajn KS. Anticoagulant therapy of cancer patients: Will patient selection increase overall survival?. Thromb Haemost 2015; 114 (03) 530-536
• 109 Demers M, Krause DS, Schatzberg D. , et al. Cancers predispose neutrophils to release extracellular DNA traps that contribute to cancer-associated thrombosis. Proc Natl Acad Sci U S A 2012; 109 (32) 13076-13081
• 110 Alfaro C, Teijeira A, Oñate C. , et al. Tumor-produced interleukin-8 attracts human myeloid-derived suppressor cells and elicits extrusion of neutrophil extracellular traps (NETs). Clin Cancer Res 2016; 22 (15) 3924-3936
• 111 Leal AC, Mizurini DM, Gomes T. , et al. Tumor-derived exosomes induce the formation of neutrophil extracellular traps: implications for the establishment of cancer-associated thrombosis. Sci Rep 2017; 7 (01) 6438
• 112 Cools-Lartigue J, Spicer J, McDonald B. , et al. Neutrophil extracellular traps sequester circulating tumor cells and promote metastasis. J Clin Invest 2013; 67484
• 113 Cedervall J, Zhang Y, Huang H. , et al. Neutrophil extracellular traps accumulate in peripheral blood vessels and compromise organ function in tumor-bearing animals. Cancer Res 2015; 75 (13) 2653-2662
• 114 Park J, Wysocki RW, Amoozgar Z. , et al. Cancer cells induce metastasis-supporting neutrophil extracellular DNA traps. Sci Transl Med 2016; 8 (361) 361ra138
• 115 Wolach O, Sellar RS, Martinod K. , et al. Increased neutrophil extracellular trap formation promotes thrombosis in myeloproliferative neoplasms. Sci Transl Med 2018; 10 (436) 10
• 116 Thålin 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
• 117 Hisada Y, Grover SP, Maqsood A. , et al. Neutrophils and neutrophil extracellular traps enhance venous thrombosis in mice bearing human pancreatic tumors. Haematologica 2019; haematol.2019.217083
• 118 Arelaki S, Arampatzioglou A, Kambas K. , et al. Gradient infiltration of neutrophil extracellular traps in colon cancer and evidence for their involvement in tumour growth. PLoS One 2016; 11 (05) e0154484
• 119 Bauer AT, Suckau J, Frank K. , et al. von Willebrand factor fibers promote cancer-associated platelet aggregation in malignant melanoma of mice and humans. Blood 2015; 125 (20) 3153-3163
• 120 Guglietta S, Chiavelli A, Zagato E. , et al. Coagulation induced by C3aR-dependent NETosis drives protumorigenic neutrophils during small intestinal tumorigenesis. Nat Commun 2016; 7: 11037
• 121 Grilz E, Mauracher L-M, Posch F. , et al. Citrullinated histone H3, a biomarker for neutrophil extracellular trap formation, predicts the risk of mortality in patients with cancer. Br J Haematol 2019; 186 (02) 311-320
• 122 McDonald B, Davis RP, Kim S-J. , et al. Platelets and neutrophil extracellular traps collaborate to promote intravascular coagulation during sepsis in mice. Blood 2017; 129 (10) 1357-1367
• 123 Iba T, Miki T, Hashiguchi N, Tabe Y, Nagaoka I. Is the neutrophil a ‘prima donna’ in the procoagulant process during sepsis?. Crit Care 2014; 18 (04) 230
• 124 Liaw PC, Ito T, Iba T, Thachil J, Zeerleder S. DAMP and DIC: the role of extracellular DNA and DNA-binding proteins in the pathogenesis of DIC. Blood Rev 2016; 30 (04) 257-261
• 125 Gould TJ, Vu TT, Stafford AR. , et al. Cell-free DNA modulates clot structure and impairs fibrinolysis in sepsis. Arterioscler Thromb Vasc Biol 2015; 35 (12) 2544-2553
• 126 Yang S, Qi H, Kan K. , et al. Neutrophil extracellular traps promote hypercoagulability in patients with sepsis. Shock 2017; 47 (02) 132-139
• 127 Delabranche X, Stiel L, Severac F. , et al. Evidence of netosis in septic shock-induced disseminated intravascular coagulation. Shock 2017; 47 (03) 313-317
• 128 Sharma A, McCann K, Tripathi JK. , et al. Tamoxifen restores extracellular trap formation in neutrophils from patients with chronic granulomatous disease in a reactive oxygen species-independent manner. J Allergy Clin Immunol 2019; 144 (02) 597-600.e3
• 129 Rockel JS, Kapoor M. Autophagy: controlling cell fate in rheumatic diseases. Nat Rev Rheumatol 2016; 12 (09) 517-531
• 130 Papagoras C, Chrysanthopoulou A, Mitsios A, Arampatzioglou A, Ritis K, Skendros P. Autophagy inhibition in adult-onset Still's disease: still more space for hydroxychloroquine?. Clin Exp Rheumatol 2017; 35 (06) (Suppl. 108) 133-134
• 131 Belizna C, Pregnolato F, Abad S. , et al. HIBISCUS: hydroxychloroquine for the secondary prevention of thrombotic and obstetrical events in primary antiphospholipid syndrome. Autoimmun Rev 2018; 17 (12) 1153-1168
• 132 Manfredi AA, Rovere-Querini P, D'Angelo A, Maugeri N. Low molecular weight heparins prevent the induction of autophagy of activated neutrophils and the formation of neutrophil extracellular traps. Pharmacol Res 2017; 123: 146-156
• 133 Van Avondt K, Maegdefessel L, Soehnlein O. Therapeutic targeting of neutrophil extracellular traps in atherogenic inflammation. Thromb Haemost 2019; 119 (04) 542-552
• 134 Dubois AV, Gauthier A, Bréa D. , et al. Influence of DNA on the activities and inhibition of neutrophil serine proteases in cystic fibrosis sputum. Am J Respir Cell Mol Biol 2012; 47 (01) 80-86
• 135 Mastellos DC, Reis ES, Ricklin D, Smith RJ, Lambris JD. Complement C3-targeted therapy: replacing long-held assertions with evidence-based discovery. Trends Immunol 2017; 38 (06) 383-394
• 136 Boone BA, Murthy P, Miller-Ocuin J. , et al. Chloroquine reduces hypercoagulability in pancreatic cancer through inhibition of neutrophil extracellular traps. BMC Cancer 2018; 18 (01) 678
• 137 Mitroulis I, Kambas K, Chrysanthopoulou A. , et al. Neutrophil extracellular trap formation is associated with IL-1β and autophagy-related signaling in gout. PLoS One 2011; 6 (12) e29318
• 138 Healy LD, Puy C, Itakura A. , et al. Colocalization of neutrophils, extracellular DNA and coagulation factors during NETosis: development and utility of an immunofluorescence-based microscopy platform. J Immunol Methods 2016; 435: 77-84
• 139 Abrams ST, Morton B, Alhamdi Y. , et al. A novel assay for neutrophil extracellular traps (NETs) formation independently predicts disseminated intravascular coagulation and mortality in critically ill patients. Am J Respir Crit Care Med 2019; 200 (07) 869-880