Neutrophils are primary effector cells of innate immunity and fight infection by phagocytosis and degranulation. Activated neutrophils also release neutrophil extracellular traps (NETs) in response to a variety of stimuli. These NETs are net-like complexes composed of cell-free DNA, histones and neutrophil granule proteins. Besides the evolutionarily conserved mechanism to capture and eliminate pathogens, NETs are also associated with pathophysiological processes of various diseases. Here, we elucidate the mechanisms of NET formation and their different implications in disease. We focused on autoinflammatory and cardiovascular disorders as the leading cause of death. Neutrophil extracellular traps are not only present in various cardiovascular diseases but play an essential role in atherosclerotic plaque formation, arterial and venous thrombosis, as well as in the development and progression of abdominal aortic aneurysms. Furthermore, NETosis can be considered as a source of autoantigens and maintains an inflammatory milieu promoting autoimmune diseases. Indeed, there is further need for research into the balance between NET induction, inhibition, and degradation in order to pharmacologically target NETs and their compounds without impairing the patient’s immune defense. This review may be of interest to both basic scientists and clinicians to stimulate translational research and innovative clinical approaches.

Background: The analysis of angiogenesis factors in the blood of tumor patients has given diverse results on their prognostic or predictive value. Since mediators of angiogenesis are stored in platelets, their measurement in plasma is sensitive to inadvertent platelet activation during blood processing. Methods: Variants of blood withdrawal and plasma preparation were evaluated by ELISA for the detection of TSP-1, PF-4, VEGF and PD-ECGF. A total of 22 pancreatic cancer patients and 29 healthy volunteers were evaluated. Results: Plasma preparation with the anticoagulant mix of citrate, theophylline, adenosine, dipyridamole (CTAD) and immediate blood processing at 4°C was required for reproducible measurements of TSP-1, PF-4 and VEGF. Blood collection by venflon or inadvertent hemolysis during blood withdrawal caused significantly elevated TSP-1 and PF4 values. When optimized plasma preparation was applied, a significant increase of TSP-1 and VEGF in cancer patients was detected (P = 0.006; P < 0.001). Conclusion: The reliable plasma analysis of circulating platelet-stored angiogenesis factors requires preparation with CTAD at 4°C and blood collection by butterfly needle. Suboptimal procedures of plasma preparation are commonly applied in clinical monitoring of angiogenesis parameters which may account for the differences in reported plasma values and may have masked their predictive or prognostic marker potential.

Objective: Macrophages are immune cells, capable to remodel the extracellular matrix, which can harbor extracellular DNA incorporated into neutrophil extracellular traps (NETs). To study the breakdown of NETs we studied the capability of macrophage subsets to degrade these structures in vitro and in vivo in a murine thrombosis model. Furthermore, we analyzed human abdominal aortic aneurysm samples in support of our in vitro and in vivo results. Approach and Results: Macrophages were seeded onto blood clots or isolated NETs and polarized. All macrophages were capable to degrade NETs. For initial breakdown, macrophages relied on extracellular deoxyribonucleases. Proinflammatory polarization enhanced NET degradation. The boost in degradation was because of increased macropinocytosis, as inhibition by imipramine diminished their NET breakdown. Inhibition of macropinocytosis in a murine thrombosis model led to increased NET burden and reduced thrombus resolution in vivo. When analyzing abdominal aortic aneurysm samples, macrophage density furthermore corresponded negatively with the amount of local NETs in the intraluminal thrombi as well as in the vessel wall, as increased macrophage density was associated with a reduction in NET burden. Conclusions: We provide evidence that macrophages degrade NETs by extracellular predigestion and subsequent uptake. Furthermore, we show that proinflammatory macrophages increase NET degradation through enhanced macropinocytosis, priming them for NET engulfment. Based on our findings, that inhibition of macropinocytosis in mice corresponded to increased NET amounts in thrombi and that local macrophage density in human abdominal aortic aneurysm is negatively associated with surrounding NETs, we hypothesize, that macrophages are able to degrade NETs in vivo.