Markers of neutrophil activation and neutrophil extracellular traps in diagnosing patients with acute venous thromboembolism: A feasibility study based on two VTE cohorts

Abstract

Background: Venous thromboembolism (VTE) diagnosis would greatly benefit from the identification of novel biomarkers to complement D-dimer, a marker limited by low specificity. Neutrophil extracellular traps (NETs) have been shown to promote thrombosis and could hypothetically be used for diagnosis of acute VTE.

Objectives: To assess the levels of specific markers of neutrophil activation and NETs and compare their diagnostic accuracy to D-dimer.

Methods: We measured plasma levels of neutrophil activation marker neutrophil elastase (NE), the NET marker nucleosomal citrullinated histone H3 (H3Cit-DNA) and cell-free DNA in patients (n = 294) with suspected VTE (pulmonary embolism and deep vein thrombosis) as well as healthy controls (n = 30). A total of 112 VTE positive and 182 VTE negative patients from two prospective cohort studies were included.

Results: Higher levels of H3Cit-DNA and NE, but not cell-free DNA, were associated with VTE. Area under receiver operating curves (AUC) were 0.90 and 0.93 for D-dimer, 0.65 and 0.68 for NE and 0.60 and 0.67 for H3Cit-DNA in the respective cohorts. Adding NE and H3Cit-DNA to a D-dimer based risk model did not improve AUC.

Conclusions: Our study demonstrates the presence of neutrophil activation and NET formation in VTE using specific markers. However, the addition of NE or H3Cit-DNA to D-dimer did not improve the discrimination compared to D-dimer alone. This study provides information on the feasibility of using markers of NETs as diagnostic tools in acute VTE. Based on our findings, we believe the potential of these markers are limited in this setting.

Fig 1. Flow chart describing inclusion of study participants.

Fig 2. Plasma levels of investigated markers.

Increased plasma levels of H3Cit-DNA and NE but not cfDNA was observed in VTE positive study particants compared to VTE negative study participants in both the VEBIOS ER cohort (A-C) and the DFW-VTE cohort (D-F). In the healthy control group (G-I), H3Cit-DNA and NE levels were generally lower compared to the VTE cohorts. However, no difference in plasma levels of cfDNA was observed when compared to VTE negative or VTE positive participants. Lines represent medians with IQR. NS P > 0.05, * P < 0.05, ** P < 0.01, ***P < 0.001, **** P < 0.0001.