Background: Neutrophil extracellular traps (NETs) containing DNA and histones are expelled from neutrophils in infection and thrombosis. Heparins, anticoagulant polyanions, can neutralize histones with a potential therapeutic advantage in sepsis. Polyphosphates, procoagulant polyanions, are released by platelets and microorganisms. Objectives: To characterize the combined effects of NET components and polyanions on clot structure, mechanical properties and lytic susceptibility. Methods: Scanning electron microscopy, pressure-driven permeation, turbidimetry, and oscillation rheometry were used for the characterization of the structure, viscoelasticity, and kinetics of formation and lysis of fibrin and plasma clots containing histones+/-DNA in combination with unfractionated heparin, its desulfated derivatives, low molecular weight heparin (LMWH), pentasaccharide, and polyphosphates of different sizes. Results: Histones and DNA inhibited fibrin lysis by plasmin, but this behavior was not neutralized by negatively charged heparins or short polyphosphates. Rather, fibrin lysis was further inhibited by added polyanions. Histones inhibited plasma clot lysis by tissue plasminogen activator and the response to added heparin was size dependent. Unfractionated heparin, LMWH, and pentasaccharide had no effect, exacerbated, or reversed histone inhibition, respectively. Histones increased the mechanical strength of fibrin, which was exacerbated by smaller heparin and polyphosphate molecules. Histones increased fibrin diameter and pore size of fibrin clots and this effect was neutralized by all heparin variants but enhanced by polyphosphates. Conclusions: Despite their common polyanionic character, heparins and polyphosphates exert distinct effects on fibrin mechanical and fibrinolytic stability. Anti-fibrinolytic effects of histones were more often enhanced by polyanions not counteracted. Careful selection of anti-histone strategies is required if they are to be combined with thrombolytic therapy.
Bibliographical noteFunding Information:
This work was supported by the Hungarian National Research, Development and Innovation Office (NKFIH; 129528 to KK, 131657 to AB), the Higher Education Institutional Excellence Programme of the Ministry of Human Capacities in Hungary for the Molecular Biology thematic program of Semmelweis University (KK), STIA‐2020‐OTKA/137209 (KK) and the Excellence Program BIONANO_GINOP‐2.3.2‐15‐2016‐00017 (AB). The authors are grateful to John Hogwood and Elaine Gray for providing the heparin derivatives and to Györgyi Oravecz and Krisztián Bálint for technical assistance.
© 2021 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals LLC on behalf of International Society on Thrombosis and Haemostasis