THE EFFECTS OF DNASE I AND LOW-MOLECULAR-WEIGHT HEPARIN IN A MURINE MODEL OF POLYMICROBIAL ABDOMINAL SEPSIS.

ABSTRACT: Introduction: Cell-free DNA (CFDNA) has emerged as a prognostic biomarker in patients with sepsis. Circulating CFDNA is hypothesized to be associated with histones in the form of nucleosomes. In vitro, DNA activates coagulation and inhibits fibrinolysis, whereas histones activate platelets and are cytotoxic to endothelial cells. Previous studies have targeted CFDNA or histones in animal models of sepsis using DNase I or heparins, respectively, which has reduced inflammatory and thrombosis markers, thereby improving survival. In this study, we explored the possibility that the combination of DNase I and a low-molecular weight heparin (LMWH) may be a better therapeutic approach than monotherapy in a murine model of abdominal sepsis. Methods: C57Bl/6 mice (8-12 weeks old, both sexes) were subjected to either cecal ligation and puncture or sham surgery. Mice were given antibiotics, fluids, and either saline, DNase I (intraperitoneally, 20 mg/kg/8 h), LMWH (dalteparin, subcutaneously 500 IU/kg/12 h), or a combination of both (n = 12-31). Mice were monitored over 72 h for survival. Organs and blood were harvested for analysis. Levels of LMWH, CFDNA, IL-6, citrullinated histone-H3, thrombin-antithrombin complexes, and protein C were measured in plasma. Results: Administration of either DNase I (81.8%) or LMWH (83.3%, prophylactic range of 0.12 ± 0.07 IU/mL achieved) improved the survival of septic mice compared with saline- (38.7%) and combination-treated mice (48.8%, P < 0.05). Combination-treated mice also showed a small but insignificant improvement in survival compared with saline-treated cecal ligation and puncture mice. Monotherapies may be improving survival by reducing blood bacterial loads, citrullinated histone-H3, and thrombin-antithrombin complexes, and improving protein C levels. Conclusions: Compared with saline- and combination-treated mice, administration of monotherapies to septic mice improved survival. These findings suggest that there may be a negative drug-drug interaction between DNase I and LMWH when DNase I is administered intraperitoneally in a murine model of polymicrobial abdominal sepsis.

Does cell-free DNA promote coagulation and inhibit fibrinolysis in patients with unprovoked venous thromboembolism?

INTRODUCTION: Cell-free DNA (CFDNA) is the major structural component of neutrophil extracellular traps (NETs). CFDNA contributes to the prothrombotic potential of NETs by promoting thrombin generation and inhibiting fibrinolysis. Patients with venous thromboembolism (VTE) have elevated circulating nucleosomes (i.e. DNA-histone complexes). In this study, we investigated if CFDNA contributes to a procoagulant and an antifibrinolytic state in patients with unprovoked VTE. MATERIALS AND METHODS: Plasma samples from patients with a first episode of unprovoked VTE were obtained from the D-Dimer Optimal Duration Study (DODS). We measured CFDNA plasma levels in 263 patients while on warfarin and 1-month after stopping. Thrombin generation assays and clot lysis assays were measured in patients after stopping warfarin. Comparisons were made with healthy controls. RESULTS: CFDNA levels in VTE patients who stopped warfarin (5.53 µg/mL; 95%CI: 5.34-5.72) were higher than during warfarin therapy (3.11 µg/mL; 95%CI: 2.98-3.25; p < .001), and higher than in healthy controls (2.77 µg/mL; 95%CI: 2.42-3.11; p < .001). VTE patients had a procoagulant state as evidenced by a shorter lag time (30.8 min; 95%CI: 29.2-32.4) compared to controls (48.2 min; 95%CI :41.0-55.5; p < .001) and a greater endogenous thrombin potential (2656 nM∗min; 95%CI: 2479-2836) compared to healthy controls (1198 nM ∗ min; 95%CI: 793-1603). There was a higher proportion of clots generated from patient plasma that were resistant to lysis (43.7%) compared to healthy controls (46.3%; p < .05). CFDNA levels were not associated with enhanced thrombin generation or impaired fibrinolysis in VTE patients. CONCLUSION: CFDNA levels are elevated in patients with unprovoked VTE but do not correlate with the procoagulant or anti-fibrinolytic properties of patient plasma. This study suggests that additional factors in addition to CFDNA may contribute to the pathogenesis of VTE.