PhD Scientific Days 2023

Translational Medicine III.

Quantitative determination of neutrophil extracellular trap components in the systemic circulation of patients with arterial thrombosis and malignacy

Text of the abstract

Background: Neutrophil extracellular traps (NETs) are formed in tumor tissues, arterial and venous thrombi and influence the outcome of these pathologies. The main components of NETs are DNA and attached histones, in which the arginine residues are partially converted to citrulline, as a result of the NET-related enzymatic deimination.
Aims: 1) To develop methods for NET-marker determination in plasma samples. 2) To quantify systemic NET-markers in patients with arterial thrombosis (acute myocardial infarction and peripheral arterial thrombosis) with (Tu-AMI; Tu-PAT) or without (AMI; PAT) accompanying malignant tumor (Tu). 3) To investigate the forming and lysability of plasma clots in relation to their NET-marker content.
Methods: Citrullinated H3 histone (H3cit), double-starnded DNA (dsDNA), H3cit-DNA complex and plasma levels of 47 (26 AMI, 3 Tu-AMI, 12 PAT, 6 Tu-PAT) patients and 8 healthy volunteers were determined by ELISA, and Invitrogen Quant-iT Picogreen DNA reagent. The kinetics of formation and lysis of PAT and Tu-PATplasma clots was investigated by turbidimerty.
Results: The median plasma level of dsDNA was 40% higher in AMI [30;60], Tu-AMI [31;44] and PAT [32;47] patients and 70%[47;81] higher in Tu-PAT patients than in healthy subjects (median[bottom;top quartile]). The H3cit elevation was pronounced in AMI (2.2-fold [1.1;2.9]) and Tu-PAT (2.9-fold [2.1;4.0]). H3cit-DNA was elevated in AMI (3.6[0.3;7.8] ng/ml) and PAT (2.2[0;8.6] ng/ml) patients. The clotting of recalcified plasma was accelerated in all patients (clotting time was reduced by 50%[48;60]), while lysis time of plasma clots by tissue type plasminogen activator was increased (up to 80%[61;100] in Tu-PAT patients).
Conclusion: Only one out of the three systemic NET-markers, the dsDNA increased in all examined thrombotic states. The most specific NET-biomarker, the H3cit-DNA complex was elevated in non-tumor patients. Increased stability of plasma clots containing histone and DNA implies that the examined NET-components are not only biomarkers, but may act as pathogenic factors contributing to a systemic prothrombotic state.