Neutrophils have traditionally been thought of as simple foot soldiers of the innate immune system with a restricted set of pro-inflammatory functions. More recently, it has become apparent that ...neutrophils are, in fact, complex cells capable of a vast array of specialized functions. Although neutrophils are undoubtedly major effectors of acute inflammation, several lines of evidence indicate that they also contribute to chronic inflammatory conditions and adaptive immune responses. Here, we discuss the key features of the life of a neutrophil, from its release from bone marrow to its death. We discuss the possible existence of different neutrophil subsets and their putative anti-inflammatory roles. We focus on how neutrophils are recruited to infected or injured tissues and describe differences in neutrophil recruitment between different tissues. Finally, we explain the mechanisms that are used by neutrophils to promote protective or pathological immune responses at different sites.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Neutrophil extracellular traps (NETs; webs of DNA coated in antimicrobial proteins) are released into the vasculature during sepsis where they contribute to host defense, but also cause tissue damage ...and organ dysfunction. Various components of NETs have also been implicated as activators of coagulation. Using multicolor confocal intravital microscopy in mouse models of sepsis, we observed profound platelet aggregation, thrombin activation, and fibrin clot formation within (and downstream of) NETs in vivo. NETs were critical for the development of sepsis-induced intravascular coagulation regardless of the inciting bacterial stimulus (gram-negative, gram-positive, or bacterial products). Removal of NETs via DNase infusion, or in peptidylarginine deiminase-4–deficient mice (which have impaired NET production), resulted in significantly lower quantities of intravascular thrombin activity, reduced platelet aggregation, and improved microvascular perfusion. NET-induced intravascular coagulation was dependent on a collaborative interaction between histone H4 in NETs, platelets, and the release of inorganic polyphosphate. Real-time perfusion imaging revealed markedly improved microvascular perfusion in response to the blockade of NET-induced coagulation, which correlated with reduced markers of systemic intravascular coagulation and end-organ damage in septic mice. Together, these data demonstrate, for the first time in an in vivo model of infection, a dynamic NET–platelet–thrombin axis that promotes intravascular coagulation and microvascular dysfunction in sepsis.
•In vivo imaging reveals a NET–platelet–thrombin axis that promotes intravascular coagulation in sepsis.•Inhibition of NETs during sepsis reduces intravascular coagulation, improves microvascular perfusion, and reduces organ damage.
Neutrophil extracellular traps (NETs) composed of DNA decorated with histones and proteases trap and kill bacteria but also injure host tissue. Here we show that during a bloodstream infection with ...methicillin-resistant Staphylococcus aureus, the majority of bacteria are sequestered immediately by hepatic Kupffer cells, resulting in transient increases in liver enzymes, focal ischaemic areas and a robust neutrophil infiltration into the liver. The neutrophils release NETs into the liver vasculature, which remain anchored to the vascular wall via von Willebrand factor and reveal significant neutrophil elastase (NE) proteolytic activity. Importantly, DNase although very effective at DNA removal, and somewhat effective at inhibiting NE proteolytic activity, fails to remove the majority of histones from the vessel wall and only partly reduces injury. By contrast, inhibition of NET production as modelled by PAD4-deficiency, or prevention of NET formation and proteolytic activity as modelled in NE(-/-) mice prevent collateral host tissue damage.
Obesity manifests itself with low-grade chronic inflammation that shapes immune responses during infection. Albeit obese individuals are at risk of higher mortality due to comorbidities, they are ...better protected from systemic inflammation. Recently, we showed that in the vasculature of obese mice kept on high-fat diet (HFD), neutrophils produce less neutrophil extracellular traps (NETs) than in lean controls (normal diet, ND). NETs are used by neutrophils to counteract severe infection, but they also cause collateral damage. Hardly anything is known about metabolic requirements for their formation, especially in the context of obesity and/or sepsis. Thus, we aimed to study the immunometabolism of NET formation by application of ex vivo neutrophil analyses (Seahorse analyzer, selective inhibitors, confocal imaging) and intravital microscopy. The obtained data show that glycolysis and/or pentose phosphate pathway are involved in NETs release by ND neutrophils in both physiological and inflammatory conditions. In contrast, such cells of septic HFD mice utilize these routes only to spontaneously cast NETs, while after secondary ex vivo activation they exhibit so called “exhausted phenotype”, which manifests itself in diminished NET release despite high glycolytic potential and flexibility to oxidize fatty acids. Moreover, impact of ATP synthase inhibition on NET formation is revealed. Overall, the study shows that the neutrophil potential to cast NETs depends on both the metabolic and inflammatory state of the individual.
SARS-CoV-2 infection poses a major threat to the lungs and multiple other organs, occasionally causing death. Until effective vaccines are developed to curb the pandemic, it is paramount to define ...the mechanisms and develop protective therapies to prevent organ dysfunction in patients with COVID-19. Individuals that develop severe manifestations have signs of dysregulated innate and adaptive immune responses. Emerging evidence implicates neutrophils and the disbalance between neutrophil extracellular trap (NET) formation and degradation plays a central role in the pathophysiology of inflammation, coagulopathy, organ damage, and immunothrombosis that characterize severe cases of COVID-19. Here, we discuss the evidence supporting a role for NETs in COVID-19 manifestations and present putative mechanisms, by which NETs promote tissue injury and immunothrombosis. We present therapeutic strategies, which have been successful in the treatment of immunο-inflammatory disorders and which target dysregulated NET formation or degradation, as potential approaches that may benefit patients with severe COVID-19.
Although neutrophil extracellular traps (NETs) were discovered only 16 years ago, they have already taken us from heaven to hell as we learned that apart from beneficial trapping of pathogens, they ...cause, or contribute to, numerous disorders. The latter is connected to their persistent presence in the blood or tissue, and we hardly know how they are removed in mild pathophysiological conditions and why their removal is impaired in multiple severe pathological conditions. Herein, we bring together all data available up till now on how NETs are cleared-from engaged cells, their phenotypes, to involved enzymes and molecules. Moreover, we hypothesize on why NET removal is challenged in multiple disorders and propose further directions for studies on NET removal as well as possible therapeutic strategies to have them cleared.
Since the discovery and definition of neutrophil extracellular traps (NETs) 14 years ago, numerous characteristics and physiological functions of NETs have been uncovered. Nowadays, the field ...continues to expand and novel mechanisms that orchestrate formation of NETs, their previously unknown properties, and novel implications in disease continue to emerge. The abundance of available data has also led to some confusion in the NET research community due to contradictory results and divergent scientific concepts, such as pro- and anti-inflammatory roles in pathologic conditions, demarcation from other forms of cell death, or the origin of the DNA that forms the NET scaffold. Here, we present prevailing concepts and state of the science in NET-related research and elaborate on open questions and areas of dispute.
Monocytes are recruited from the blood to sites of inflammation, where they contribute to wound healing and tissue repair. There are at least two subsets of monocytes: classical or proinflammatory ...(CCR2(hi)CX3CR1(low)) and nonclassical, patrolling, or alternative (CCR2(low)CX3CR1(hi)) monocytes. Using spinning-disk confocal intravital microscopy and mice with fluorescent reporters for each of these subsets, we were able to track the dynamic spectrum of monocytes that enter a site of sterile hepatic injury in vivo. We observed that the CCR2(hi)CX3CR1(low) monocytes were recruited early and persisted for at least 48 h, forming a ringlike structure around the injured area. These monocytes transitioned, in situ, from CCR2(hi)Cx3CR1(low) to CX3CR1(hi)CCR2(low) within the ringlike structure and then entered the injury site. This phenotypic conversion was essential for optimal repair. These results demonstrate a local, cytokine driven reprogramming of classic, proinflammatory monocytes into nonclassical or alternative monocytes to facilitate proper wound-healing.
Neutrophil extracellular traps (NETs) immobilize pathogens during early stages of systemic inflammation but as the reaction progresses they become detrimental to endothelial cells and the ...organ-specific cells. For this reason it would be of importance to control their formation by either physiological or pharmacological means. Endogenously, formation of NETs is under control of cellular and whole organism metabolism as shown previously in the course of bacterial systemic inflammation, obesity or the combination of the two. Numerous leukocytes are subjected to immunometabolic regulation and in macrophages exposure to lipopolysaccharide (LPS) leads to two breaks in the Krebs cycle that impact this cell functioning. As a consequence of the first break, anti-microbial itaconic acid (itaconate) is produced whereas the second break activates hypoxia-inducible factor-1α (Hif-1α). In turn, itaconate activates transcription of the anti-inflammatory nuclear factor erythroid 2-related factor 2 (Nrf2) which upregulates cyto-protective heme oxygenase (HO-1). Here we report that exogenously added derivative of the itaconic acid, 4-octyl itaconate (4-OI), diminishes formation of NETs by neutrophils of either normal (lean) or obese mice, and independently of the age of the animals or immunoaging. Elucidating the mechanism of this inhibition we unravel that although Nrf2/HO-1 expression itself is not altered by 4-OI, it is up-regulated when compared against the NET formation while Hif-1α is downregulated in 4-OI-pre-treated LPS-stimulated neutrophils in either way. We further show that blockage of Hif-1α by its specific inhibitor diminishes NET release as does inhibition by 4-OI. Also inhibition of HO-1 activity correlates with diminished LPS-induced NET release upon pre-treatment with 4-OI albeit LPS alone induced NETs are not HO-1-dependent. In summary, we unravel that 4-OI inhibits NET formation by murine neutrophils independently of their origin (health vs. metabolically challenged animals) and the age of individuals/immunosenescence
via
inhibition of Hif-1α and induction of HO-1.
Although platelets have been extensively studied in hemostasis and inflammation, their role is not well understood in sterile liver injury and repair. Using a thermally induced focal liver injury and ...repair model and multichannel spinning disk confocal microscopy allowed visualization of the dynamic behavior of platelets and neutrophils in this insult. Platelets instantaneously adhered to molecularly altered sinusoidal endothelium adjacent to the afflicted area, paving approximately 200 µm abutting the injury. Platelets remained adherent for at least 4 hours, but dissipated by 8 hours. The early recruitment occurred by GPIIbIIIa (CD41) and the later recruitment was dependent upon both GPIIbIIIa and GPIb (CD42B). Platelets did not occlude the vessels, but rather paved the altered endothelium. Endothelin‐induced vasoconstriction by hepatic stellate cells, and not platelet accumulation or coagulation, was responsible for temporarily restricted perfusion around the injury. Neutrophils crawled into the injury from significant distances through the sinusoids. The crawling neutrophils required the platelet‐paved endothelium given that very little neutrophil recruitment was noted in thrombocytopenic or CD41‐deficient mice. As platelets slowly dissipated, neutrophil recruitment was also halted. Previous work suggested that platelets binding to immobilized neutrophils induced neutrophil extracellular trap (NET) formation in response to infection as well as during thrombosis and other forms of sterile injury. In this model of neutrophils crawling on immobilized platelets, very few NETs were observed and no additional injury was noted. In fact, GPIIbIIIa‐deficient mice had delayed repair. Conclusion: In a liver model of sterile injury and repair, platelets play a critical role in forming a substratum and pave the way for neutrophils to enter the injured site for subsequent repair. (Hepatology 2015;62:1593–1605)
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