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1.
Cells ; 11(22)2022 Nov 16.
Article in English | MEDLINE | ID: covidwho-2142561

ABSTRACT

Alveolar macrophage (AM) proliferation and self-renewal play an important role in the lung tissue microenvironment. However, the impact of immune cells, especially the neutrophils, on AM homeostasis or function is not well characterized. In this study, we induced in vivo migration of neutrophils into bronchoalveolar lavage (BAL) fluid and lung using CXCL1, and then co-cultured these with AMs in vitro. Neutrophils in the BAL (BAL-neutrophils), rather than neutrophils of bone marrow (BM-neutrophils), were found to inhibit AM proliferation. Analysis of publicly available data showed high heterogeneity of lung neutrophils with distinct molecular signatures of BM- and blood-neutrophils. Unexpectedly, BAL-neutrophils from influenza virus PR8-infected mice (PR8-neutrophils) did not inhibit the proliferation of AMs. Bulk RNA sequencing further revealed that co-culture of AMs with PR8-neutrophils induced IFN-α and -γ responses and inflammatory response, and AMs co-cultured with BAL-neutrophils showed higher expression of metabolism- and ROS-associated genes; in addition, BAL-neutrophils from PR8-infected mice modulated AM polarization and phagocytosis. BAL-neutrophil-mediated suppression of AM proliferation was abrogated by a combination of inhibitors of different neutrophil death pathways. Collectively, our findings suggest that multiple cell death pathways of neutrophils regulate the proliferation of AMs. Targeting neutrophil death may represent a potential therapeutic strategy for improving AM homeostasis during respiratory diseases.


Subject(s)
Macrophages, Alveolar , Neutrophils , Mice , Animals , Macrophages, Alveolar/metabolism , Neutrophils/metabolism , Bronchoalveolar Lavage Fluid , Lung , Cell Proliferation
2.
Front Immunol ; 13: 993720, 2022.
Article in English | MEDLINE | ID: covidwho-2142018

ABSTRACT

Pathogenesis of lung injury in COVID-19 is not completely understood, leaving gaps in understanding how current treatments modulate the course of COVID-19. Neutrophil numbers and activation state in circulation have been found to correlate with COVID-19 severity, and neutrophil extracellular traps (NETs) have been found in the lung parenchyma of patients with acute respiratory distress syndrome (ARDS) in COVID-19. Targeting the pro-inflammatory functions of neutrophils may diminish lung injury in COVID-19 and ARDS. Neutrophils were isolated from peripheral blood of healthy donors, treated ex vivo with dexamethasone, tocilizumab and intravenous immunoglobulin (IVIG) and NET formation, oxidative burst, and phagocytosis were assessed. Plasma from critically ill COVID-19 patients before and after clinical treatment with IVIG and from healthy donors was assessed for neutrophil activation-related proteins. While dexamethasone and tocilizumab did not affect PMA- and nigericin-induced NET production ex vivo, IVIG induced a dose-dependent abrogation of NET production in both activation models. IVIG also reduced PMA-elicited reactive oxygen species production, but did not alter phagocytosis. COVID-19 patients were found to have elevated levels of cell-free DNA, neutrophil elastase and IL-8 as compared to healthy controls. Levels of both cell-free DNA and neutrophil elastase were lower 5 days after 4 days of daily treatment with IVIG. The lack of impact of dexamethasone or tocilizumab on these neutrophil functions suggests that these therapeutic agents may not act through suppression of neutrophil functions, indicating that the door might still be open for the addition of a neutrophil modulator to the COVID-19 therapeutic repertoire.


Subject(s)
COVID-19 , Cell-Free Nucleic Acids , Lung Injury , Respiratory Distress Syndrome , Humans , Neutrophils/metabolism , Immunoglobulins, Intravenous/therapeutic use , Immunoglobulins, Intravenous/pharmacology , COVID-19/drug therapy , Leukocyte Elastase/metabolism , Lung Injury/metabolism , Cell-Free Nucleic Acids/metabolism , Dexamethasone
3.
Front Immunol ; 13: 931388, 2022.
Article in English | MEDLINE | ID: covidwho-2141951

ABSTRACT

Intracranial inoculation of the neuroadapted JHM strain of mouse hepatitis virus (JHMV) into susceptible strains of mice results in acute encephalomyelitis followed by a cimmune-mediated demyelination similar to the human demyelinating disease multiple sclerosis (MS). JHMV infection of transgenic mice in which expression of the neutrophil chemoattractant chemokine CXCL1 is under the control of a tetracycline-inducible promoter active within GFAP-positive cells results in sustained neutrophil infiltration in the central nervous system (CNS) that correlates with an increase in spinal cord demyelination. We used single cell RNA sequencing (scRNAseq) and flow cytometry to characterize molecular and cellular changes within the CNS associated with increased demyelination in transgenic mice compared to control animals. These approaches revealed the presence of activated neutrophils as determined by expression of mRNA transcripts associated with neutrophil effector functions, including CD63, MMP9, S100a8, S100a9, and ASPRV1, as well as altered neutrophil morphology and protein expression. Collectively, these findings reveal insight into changes in the profile of neutrophils associated with increased white matter damage in mice persistently infected with a neurotropic coronavirus.


Subject(s)
Demyelinating Diseases , Multiple Sclerosis , Murine hepatitis virus , White Matter , Animals , Central Nervous System , Chemokine CXCL1/metabolism , Humans , Matrix Metalloproteinase 9/metabolism , Mice , Mice, Transgenic , Multiple Sclerosis/metabolism , Neutrophils/metabolism , RNA, Messenger , Tetracyclines , White Matter/metabolism
4.
J Am Soc Nephrol ; 33(5): 936-947, 2022 05.
Article in English | MEDLINE | ID: covidwho-2141044

ABSTRACT

BACKGROUND: The ANCA autoantigens proteinase 3 (PR3) and myeloperoxidase (MPO) are exclusively expressed by neutrophils and monocytes. ANCA-mediated activation of these cells is the key driver of the vascular injury process in ANCA-associated vasculitis (AAV), and neutrophil serine proteases (NSPs) are disease mediators. Cathepsin C (CatC) from zymogens activates the proteolytic function of NSPs, including PR3. Lack of NSP zymogen activation results in neutrophils with strongly reduced NSP proteins. METHODS: To explore AAV-relevant consequences of blocking NSP zymogen activation by CatC, we used myeloid cells from patients with Papillon-Lefèvre syndrome, a genetic deficiency of CatC, to assess NSPs and NSP-mediated endothelial cell injury. We also examined pharmacologic CatC inhibition in neutrophil-differentiated human hematopoietic stem cells, primary human umbilical vein cells, and primary glomerular microvascular endothelial cells. RESULTS: Patients with Papillon-Lefèvre syndrome showed strongly reduced NSPs in neutrophils and monocytes. Neutrophils from these patients produced a negative PR3-ANCA test, presented less PR3 on the surface of viable and apoptotic cells, and caused significantly less damage in human umbilical vein cells. These findings were recapitulated in human stem cells, in which a highly specific CatC inhibitor, but not prednisolone, reduced NSPs without affecting neutrophil differentiation, reduced membrane PR3, and diminished neutrophil activation upon PR3-ANCA but not MPO-ANCA stimulation. Compared with healthy controls, neutrophils from patients with Papillon-Lefèvre syndrome transferred less proteolytically active NSPs to glomerular microvascular endothelial cells, the cell type targeted in ANCA-induced necrotizing crescentic glomerulonephritis. Finally, both genetic CatC deficiency and pharmacologic inhibition, but not prednisolone, reduced neutrophil-induced glomerular microvascular endothelial cell damage. CONCLUSIONS: These findings may offer encouragement for clinical studies of adjunctive CatC inhibitor in patients with PR3-AAV.


Subject(s)
Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis , Papillon-Lefevre Disease , Antibodies, Antineutrophil Cytoplasmic , Cathepsin C/metabolism , Endothelial Cells/metabolism , Enzyme Precursors/metabolism , Humans , Myeloblastin/genetics , Neutrophils/metabolism , Papillon-Lefevre Disease/metabolism , Peroxidase
5.
PLoS One ; 17(10): e0276460, 2022.
Article in English | MEDLINE | ID: covidwho-2089429

ABSTRACT

Excessive neutrophil infiltration and dysfunction contribute to the progression and severity of hyper-inflammatory syndrome, such as in severe COVID19. In the current study, we re-analysed published scRNA-seq datasets of mouse and human neutrophils to classify and compare the transcriptional regulatory networks underlying neutrophil differentiation and inflammatory responses. Distinct sets of TF modules regulate neutrophil maturation, function, and inflammatory responses under the steady state and inflammatory conditions. In COVID19 patients, neutrophil activation was associated with the selective activation of inflammation-specific TF modules. SARS-CoV-2 RNA-positive neutrophils showed a higher expression of type I interferon response TF IRF7. Furthermore, IRF7 expression was abundant in neutrophils from severe patients in progression stage. Neutrophil-mediated inflammatory responses positively correlate with the expressional level of IRF7. Based on these results, we suggest that differential activation of activation-related TFs, such as IRF7 mediate neutrophil inflammatory responses during inflammation.


Subject(s)
COVID-19 , Neutrophils , Humans , COVID-19/genetics , COVID-19/metabolism , Inflammation/genetics , Interferon Type I/metabolism , Neutrophil Activation/genetics , Neutrophil Activation/physiology , Neutrophils/metabolism , RNA, Viral , RNA-Seq , SARS-CoV-2 , Single-Cell Analysis
6.
Int J Mol Sci ; 23(18)2022 Sep 09.
Article in English | MEDLINE | ID: covidwho-2071501

ABSTRACT

In SARS-CoV-2-infected humans, disease progression is often associated with acute respiratory distress syndrome involving severe lung injury, coagulopathy, and thrombosis of the alveolar capillaries. The pathogenesis of these pulmonary complications in COVID-19 patients has not been elucidated. Autopsy study of these patients showed SARS-CoV-2 virions in pulmonary vessels and sequestrated leukocytes infiltrates associated with endotheliopathy and microvascular thrombosis. Since SARS-CoV-2 enters and infects target cells by binding its spike (S) protein to cellular angiotensin-converting enzyme 2 (ACE2), and there is evidence that vascular endothelial cells and neutrophils express ACE2, we investigated the effect of S-proteins and cell-cell communication on primary human lung microvascular endothelial cells (HLMEC) and neutrophils expression of thrombogenic factors and the potential mechanisms. Using S-proteins of two different SARS-CoV-2 variants (Wuhan and Delta), we demonstrate that exposure of HLMEC or neutrophils to S-proteins, co-culture of HLMEC exposed to S-proteins with non-exposed neutrophils, or co-culture of neutrophils exposed to S-proteins with non-exposed HLMEC induced transcriptional upregulation of tissue factor (TF), significantly increased the expression and secretion of factor (F)-V, thrombin, and fibrinogen and inhibited tissue factor pathway inhibitor (TFPI), the primary regulator of the extrinsic pathway of blood coagulation, in both cell types. Recombinant (r)TFPI and a thiol blocker (5,5'-dithio-bis-(2-nitrobenzoic acid)) prevented S-protein-induced expression and secretion of Factor-V, thrombin, and fibrinogen. Thrombomodulin blocked S-protein-induced expression and secretion of fibrinogen but had no effect on S-protein-induced expression of Factor-V or thrombin. These results suggests that following SARS-CoV-2 contact with the pulmonary endothelium or neutrophils and endothelial-neutrophil interactions, viral S-proteins induce coagulopathy via the TF pathway and mechanisms involving functional thiol groups. These findings suggest that using rTFPI and/or thiol-based drugs could be a viable therapeutic strategy against SARS-CoV-2-induced coagulopathy and thrombosis.


Subject(s)
Blood Coagulation Disorders , COVID-19 , Thrombosis , Angiotensin-Converting Enzyme 2 , Cell Communication , Endothelial Cells/metabolism , Endothelium/metabolism , Fibrinogen , Humans , Lipoproteins , Lung/metabolism , Neutrophils/metabolism , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/metabolism , Sulfhydryl Compounds , Thrombin , Thrombomodulin , Thromboplastin , Thrombosis/etiology
7.
Front Immunol ; 13: 996637, 2022.
Article in English | MEDLINE | ID: covidwho-2043454

ABSTRACT

Increased neutrophils and elevated level of circulating calprotectin are hallmarks of severe COVID-19 and they contribute to the dysregulated immune responses and cytokine storm in susceptible patients. However, the precise mechanism controlling calprotectin production during SARS-CoV-2 infection remains elusive. In this study, we showed that Dok3 adaptor restrains calprotectin production by neutrophils in response to SARS-CoV-2 spike (S) protein engagement of TLR4. Dok3 recruits SHP-2 to mediate the de-phosphorylation of MyD88 at Y257, thereby attenuating downstream JAK2-STAT3 signaling and calprotectin production. Blocking of TLR4, JAK2 and STAT3 signaling could prevent excessive production of calprotectin by Dok3-/- neutrophils, revealing new targets for potential COVID-19 therapy. As S protein from SARS-CoV-2 Delta and Omicron variants can activate TLR4-driven calprotectin production in Dok3-/- neutrophils, our study suggests that targeting calprotectin production may be an effective strategy to combat severe COVID-19 manifestations associated with these emerging variants.


Subject(s)
Adaptor Proteins, Signal Transducing , COVID-19 , Spike Glycoprotein, Coronavirus , Adaptor Proteins, Signal Transducing/metabolism , Humans , Leukocyte L1 Antigen Complex , Myeloid Differentiation Factor 88/metabolism , Neutrophils/metabolism , SARS-CoV-2 , Toll-Like Receptor 4/metabolism
8.
Front Immunol ; 13: 964622, 2022.
Article in English | MEDLINE | ID: covidwho-2022748

ABSTRACT

Acute pancreatitis is a common critical and acute gastrointestinal disease worldwide, with an increasing percentage of morbidity. However, the gene expression pattern in peripheral blood has not been fully analyzed. In addition, the mechanism of coronavirus disease 2019 (COVID-19)-induced acute pancreatitis has not been investigated. Here, after bioinformatic analysis with machine-learning methods of the expression data of peripheral blood cells and validation in local patients, two functional gene modules in peripheral blood cells of acute pancreatitis were identified, and S100A6, S100A9, and S100A12 were validated as predictors of severe pancreatitis. Additionally, through a combination analysis of bulk sequencing and single-cell sequencing data of COVID-19 patients, a pivotal subtype of neutrophils with strong activation of the interferon-related pathway was identified as a pivotal peripheral blood cell subtype for COVID-19-induced acute pancreatitis. These results could facilitate the prognostic prediction of acute pancreatitis and research on COVID-19-induced acute pancreatitis.


Subject(s)
COVID-19 , Pancreatitis , Acute Disease , Biomarkers , COVID-19/genetics , Humans , Neutrophils/metabolism , Pancreatitis/metabolism
9.
Front Immunol ; 13: 953195, 2022.
Article in English | MEDLINE | ID: covidwho-1990285

ABSTRACT

Acute lung injury (ALI) is a heterogeneous inflammatory condition associated with high morbidity and mortality. Neutrophils play a key role in the development of different forms of ALI, and the release of neutrophil extracellular traps (NETs) is emerging as a common pathogenic mechanism. NETs are essential in controlling pathogens, and their defective release or increased degradation leads to a higher risk of infection. However, NETs also contain several pro-inflammatory and cytotoxic molecules than can exacerbate thromboinflammation and lung tissue injury. To reduce NET-mediated lung damage and inflammation, DNase is frequently used in preclinical models of ALI due to its capability of digesting NET DNA scaffold. Moreover, recent advances in neutrophil biology led to the development of selective NET inhibitors, which also appear to reduce ALI in experimental models. Here we provide an overview of the role of NETs in different forms of ALI discussing existing gaps in our knowledge and novel therapeutic approaches to modulate their impact on lung injury.


Subject(s)
Acute Lung Injury , Extracellular Traps , Thrombosis , Acute Lung Injury/pathology , Extracellular Traps/metabolism , Humans , Inflammation/metabolism , Neutrophils/metabolism , Thrombosis/metabolism
10.
Int J Mol Sci ; 23(15)2022 Jul 26.
Article in English | MEDLINE | ID: covidwho-1994077

ABSTRACT

The gut barrier is a single cell layer that separates gut micro-organisms from the host, and gut permeability defects result in the translocation of microbial molecules from the gut into the blood. Despite the silent clinical manifestation, gut translocation of microbial molecules can induce systemic inflammation that might be an endogenous exacerbating factor of systemic lupus erythematosus. In contrast, circulatory immune-complex deposition and the effect of medications on the gut, an organ with an extremely large surface area, of patients with active lupus might cause gut translocation of microbial molecules, which worsens lupus severity. Likewise, the imbalance of gut microbiota may initiate lupus and/or interfere with gut integrity which results in microbial translocation and lupus exacerbation. Moreover, immune hyper-responsiveness of innate immune cells (macrophages and neutrophils) is demonstrated in a lupus model from the loss of inhibitory Fc gamma receptor IIb (FcgRIIb), which induces prominent responses through the cross-link between activating-FcgRs and innate immune receptors. The immune hyper-responsiveness can cause cell death, especially apoptosis and neutrophil extracellular traps (NETosis), which possibly exacerbates lupus, partly through the enhanced exposure of the self-antigens. Leaky gut monitoring and treatments (such as probiotics) might be beneficial in lupus. Here, we discuss the current information on leaky gut in lupus.


Subject(s)
Autoimmune Diseases , Lupus Erythematosus, Systemic , Humans , Immunity, Innate , Macrophages/metabolism , Neutrophils/metabolism
11.
Cells ; 11(15)2022 08 08.
Article in English | MEDLINE | ID: covidwho-1993939

ABSTRACT

Innate immunity responds to infections and inflammatory stimuli through a carefully choreographed set of interactions between cells, stimuli and their specific receptors. Of particular importance are endogenous peptides, which assume roles as defensins or alarmins, growth factors or wound repair inducers. LL-37, a proteolytic fragment of cathelicidin, fulfills the roles of a defensin by inserting into the membranes of bacterial pathogens, functions as alarmin in stimulating chemotaxis of innate immune cells, and alters the structure and efficacy of various cytokines. Here, we draw attention to the direct effect of LL-37 on neutrophils and the release of extracellular traps (NETs), as NETs have been established as mediators of immune defense against pathogens but also as important contributors to chronic disease and tissue pathogenesis. We propose a specific structural basis for LL-37 function, in part by highlighting the structural flexibility of LL-37 and its ability to adapt to distinct microenvironments and interacting counterparts.


Subject(s)
Extracellular Traps , Bacteria , Chemotaxis , Immunity, Innate , Neutrophils/metabolism
12.
Expert Rev Hematol ; 15(8): 727-745, 2022 08.
Article in English | MEDLINE | ID: covidwho-1978157

ABSTRACT

INTRODUCTION: COVID-19 is associated to an increased risk of thrombosis, as a result of a complex process that involves the activation of vascular and circulating cells, the release of soluble inflammatory and thrombotic mediators and blood clotting activation. AREAS COVERED: This article reviews the pathophysiological role of platelets, neutrophils, and the endothelium, and of their interactions, in the thrombotic complications of COVID-19 patients, and the current and future therapeutic approaches targeting these cell types. EXPERT OPINION: Virus-induced platelet, neutrophil, and endothelial cell changes are crucial triggers of the thrombotic complications and of the adverse evolution of COVID-19. Both the direct interaction with the virus and the associated cytokine storm concur to trigger cell activation in a classical thromboinflammatory vicious circle. Although heparin has proven to be an effective prophylactic and therapeutic weapon for the prevention and treatment of COVID-19-associated thrombosis, it acts downstream of the cascade of events triggered by SARS-CoV-2. The identification of specific molecular targets interrupting the thromboinflammatory cascade upstream, and more specifically acting either on the interaction of SARS-CoV-2 with blood and vascular cells or on the specific signaling mechanisms associated with their COVID-19-associated activation, might theoretically offer greater protection with potentially lesser side effects.


Subject(s)
COVID-19 , Thrombosis , Blood Platelets/metabolism , COVID-19/complications , Endothelium/metabolism , Humans , Neutrophils/metabolism , SARS-CoV-2 , Thrombosis/etiology , Thrombosis/metabolism
13.
Biomolecules ; 12(8)2022 07 27.
Article in English | MEDLINE | ID: covidwho-1969083

ABSTRACT

Background: Neutrophil extracellular traps' (NETs') formation is a mechanism of defense that neutrophils deploy as an alternative to phagocytosis, to constrain the spread of microorganisms. Aim: The aim was to evaluate biomarkers of NETs' formation in a patient cohort admitted to intensive care unit (ICU) due to infection. Methods: Forty-six septic shock patients, 22 critical COVID-19 patients and 48 matched control subjects were recruited. Intact nucleosomes containing histone 3.1 (Nu.H3.1), or citrullinated histone H3R8 (Nu.Cit-H3R8), free citrullinated histone (Cit-H3), neutrophil elastase (NE) and myeloperoxidase (MPO) were measured. Results: Significant differences in Nu.H3.1 and NE levels were observed between septic shock and critical COVID-19 subjects as well as with controls (p-values < 0.05). The normalization of nucleosome levels according to the neutrophil count improved the discrimination between septic shock and critical COVID-19 patients. The ratio of Nu.Cit-H3R8 to Nu.H3.1 allowed the determination of nucleosome citrullination degree, presumably by PAD4. Conclusions: H3.1 and Cit-H3R8 nucleosomes appear to be interesting markers of global cell death and neutrophil activation when combined. Nu.H3.1 permits the evaluation of disease severity and differs between septic shock and critical COVID-19 patients, reflecting two distinct potential pathological processes in these conditions.


Subject(s)
COVID-19 , Extracellular Traps , Shock, Septic , Biomarkers/metabolism , Extracellular Traps/metabolism , Histones/metabolism , Humans , Neutrophils/metabolism , Nucleosomes/metabolism , Shock, Septic/metabolism
14.
Cell Rep Med ; 3(6): 100652, 2022 06 21.
Article in English | MEDLINE | ID: covidwho-1960088

ABSTRACT

Disease recovery dynamics are often difficult to assess, as patients display heterogeneous recovery courses. To model recovery dynamics, exemplified by severe COVID-19, we apply a computational scheme on longitudinally sampled blood transcriptomes, generating recovery states, which we then link to cellular and molecular mechanisms, presenting a framework for studying the kinetics of recovery compared with non-recovery over time and long-term effects of the disease. Specifically, a decrease in mature neutrophils is the strongest cellular effect during recovery, with direct implications on disease outcome. Furthermore, we present strong indications for global regulatory changes in gene programs, decoupled from cell compositional changes, including an early rise in T cell activation and differentiation, resulting in immune rebalancing between interferon and NF-κB activity and restoration of cell homeostasis. Overall, we present a clinically relevant computational framework for modeling disease recovery, paving the way for future studies of the recovery dynamics in other diseases and tissues.


Subject(s)
COVID-19 , NF-kappa B , Cell Differentiation , Humans , Interferons/metabolism , NF-kappa B/genetics , Neutrophils/metabolism , Signal Transduction
15.
Life Sci ; 305: 120782, 2022 Sep 15.
Article in English | MEDLINE | ID: covidwho-1956258

ABSTRACT

Acute lung injury (ALI) is characterized by diffuse pulmonary infiltrates and causes great mortality. ALI presents with overproduction of proinflammatory cytokines, cell death, destruction of alveoli-endothelial barriers, and neutrophil infiltration in lung tissues. Damage-associated molecular patterns (DAMPs) are molecules released from damaged cells due to infection, trauma, etc. DAMPs activate innate and adaptive immunity, trigger inflammatory responses, and are important in the initiation and development of ALI. We reviewed the literatures on DAMPs in ALI. Alveolar macrophages (AMs), neutrophils, and epithelial cells (AECs) are important in the pathogenesis of ALI. We comprehensively analyzed the interaction between DAMPs and AMs, alveolar neutrophils, and AECs. During the initial stage of ALI, ruptured cell membranes or destroyed mitochondria release DAMPs. DAMPs activate the inflammasome in nearby sentinel immune cells, such as AMs. AMs produce IL-1ß and other cytokines. These mediators upregulate adhesion molecules of the capillary endothelium that facilitate neutrophil recruitment. The recruited neutrophils detect DAMPs using formyl peptide receptors on the membrane, guiding their migration to the injured site. The accumulation of immune cells, cytokines, chemokines, proteases, etc., results in diffuse alveolar damage and pulmonary hyperpermeability with protein-rich fluid retention. Some clinical studies have shown that patients with ALI with higher circulating DAMPs have higher mortality rates. In conclusion, DAMPs are important in the initiation and progression of ALI. The interactions between DAMPs and AMs, neutrophils, and AECs are important in ALI. This review comprehensively addresses the mechanisms of DAMPs and their interactions in ALI.


Subject(s)
Acute Lung Injury , Acute Lung Injury/pathology , Alarmins/metabolism , Animals , Cytokines/metabolism , Humans , Lipopolysaccharides/metabolism , Lung/metabolism , Mice , Mice, Inbred C57BL , Neutrophil Infiltration , Neutrophils/metabolism
16.
Arterioscler Thromb Vasc Biol ; 42(9): 1103-1112, 2022 09.
Article in English | MEDLINE | ID: covidwho-1950525

ABSTRACT

The activating interplay of thrombosis and inflammation (thromboinflammation) has been established as a major underlying pathway, driving not only cardiovascular disease but also autoimmune disease and most recently, COVID-19. Throughout the years, innate immune cells have emerged as important modulators of this process. As the most abundant white blood cell in humans, neutrophils are well-positioned to propel thromboinflammation. This includes their ability to trigger an organized cell death pathway with the release of decondensed chromatin structures called neutrophil extracellular traps. Decorated with histones and cytoplasmic and granular proteins, neutrophil extracellular traps exert cytotoxic, immunogenic, and prothrombotic effects accelerating disease progression. Distinct steps leading to extracellular DNA release (NETosis) require the activities of PAD4 (protein arginine deiminase 4) catalyzing citrullination of histones and are supported by neutrophil inflammasome. By linking the immunologic function of neutrophils with the procoagulant and proinflammatory activities of monocytes and platelets, PAD4 activity holds important implications for understanding the processes that fuel thromboinflammation. We will also discuss mechanisms whereby vascular occlusion in thromboinflammation depends on the interaction of neutrophil extracellular traps with ultra-large VWF (von Willebrand Factor) and speculate on the importance of PAD4 in neutrophil inflammasome assembly and neutrophil extracellular traps in thromboinflammatory diseases including atherosclerosis and COVID-19.


Subject(s)
Atherosclerosis , COVID-19 , Extracellular Traps , Thrombosis , Atherosclerosis/metabolism , Extracellular Traps/metabolism , Histones/metabolism , Humans , Inflammasomes/metabolism , Inflammation/metabolism , Neutrophils/metabolism , Thromboinflammation , Thrombosis/etiology , Thrombosis/metabolism , von Willebrand Factor/metabolism
17.
Crit Care ; 26(1): 206, 2022 07 07.
Article in English | MEDLINE | ID: covidwho-1923570

ABSTRACT

BACKGROUND: The release of neutrophil extracellular traps (NETs) is associated with inflammation, coagulopathy, and organ damage found in severe cases of COVID-19. However, the molecular mechanisms underlying the release of NETs in COVID-19 remain unclear. OBJECTIVES: We aim to investigate the role of the Gasdermin-D (GSDMD) pathway on NETs release and the development of organ damage during COVID-19. METHODS: We performed a single-cell transcriptome analysis in public data of bronchoalveolar lavage. Then, we enrolled 63 hospitalized patients with moderate and severe COVID-19. We analyze in blood and lung tissue samples the expression of GSDMD, presence of NETs, and signaling pathways upstreaming. Furthermore, we analyzed the treatment with disulfiram in a mouse model of SARS-CoV-2 infection. RESULTS: We found that the SARS-CoV-2 virus directly activates the pore-forming protein GSDMD that triggers NET production and organ damage in COVID-19. Single-cell transcriptome analysis revealed that the expression of GSDMD and inflammasome-related genes were increased in COVID-19 patients. High expression of active GSDMD associated with NETs structures was found in the lung tissue of COVID-19 patients. Furthermore, we showed that activation of GSDMD in neutrophils requires active caspase1/4 and live SARS-CoV-2, which infects neutrophils. In a mouse model of SARS-CoV-2 infection, the treatment with disulfiram inhibited NETs release and reduced organ damage. CONCLUSION: These results demonstrated that GSDMD-dependent NETosis plays a critical role in COVID-19 immunopathology and suggests GSDMD as a novel potential target for improving the COVID-19 therapeutic strategy.


Subject(s)
COVID-19 , Extracellular Traps , Animals , COVID-19/drug therapy , Disulfiram/metabolism , Extracellular Traps/metabolism , Mice , Neutrophils/metabolism , SARS-CoV-2
18.
Ann Clin Lab Sci ; 52(3): 374-381, 2022 May.
Article in English | MEDLINE | ID: covidwho-1918736

ABSTRACT

OBJECTIVE: Exploration of biomarkers to predict the severity of COVID-19 is important to reduce mortality. Upon COVID-19 infection, neutrophil extracellular traps (NET) are formed, which leads to a cytokine storm and host damage. Hence, the extent of NET formation may reflect disease progression and predict mortality in COVID-19. METHODS: We measured 4 NET parameters - cell-free double stranded DNA (cell-free dsDNA), neutrophil elastase, citrullinated histone H3 (Cit-H3), and histone - DNA complex - in 188 COVID-19 patients and 20 healthy controls. Survivors (n=166) were hospitalized with or without oxygen supplementation, while non-survivors (n=22) expired during in-hospital treatment. RESULTS: Cell-free dsDNA was significantly elevated in non-survivors in comparison with survivors and controls. The survival rate of patients with high levels of cell-free dsDNA, neutrophil elastase, and Cit-H3 was significantly lower than that of patients with low levels. These three markers significantly correlated with inflammatory markers (absolute neutrophil count and C-reactive protein). CONCLUSION: Since the increase in NET parameters indicates the unfavourable course of COVID-19 infection, patients predisposed to poor outcome can be rapidly managed through risk stratification by using these NET parameters.


Subject(s)
COVID-19 , Extracellular Traps , Biomarkers/metabolism , COVID-19/diagnosis , Cell-Free Nucleic Acids/blood , Cell-Free Nucleic Acids/metabolism , Extracellular Traps/metabolism , Histones/blood , Histones/metabolism , Humans , Leukocyte Elastase/blood , Leukocyte Elastase/metabolism , Neutrophils/metabolism , Prognosis
19.
Sci Rep ; 12(1): 11078, 2022 06 30.
Article in English | MEDLINE | ID: covidwho-1908298

ABSTRACT

Immune cell chemotaxis to the sites of pathogen invasion is critical for fighting infection, but in life-threatening conditions such as sepsis and Covid-19, excess activation of the innate immune system is thought to cause a damaging invasion of immune cells into tissues and a consequent excessive release of cytokines, chemokines and neutrophil extracellular traps (NETs). In these circumstances, tempering excessive activation of the innate immune system may, paradoxically, promote recovery. Here we identify the antimalarial compound artemisinin as a potent and selective inhibitor of neutrophil and macrophage chemotaxis induced by a range of chemotactic agents. Artemisinin released calcium from intracellular stores in a similar way to thapsigargin, a known inhibitor of the Sarco/Endoplasmic Reticulum Calcium ATPase pump (SERCA), but unlike thapsigargin, artemisinin blocks only the SERCA3 isoform. Inhibition of SERCA3 by artemisinin was irreversible and was inhibited by iron chelation, suggesting iron-catalysed alkylation of a specific cysteine residue in SERCA3 as the mechanism by which artemisinin inhibits neutrophil motility. In murine infection models, artemisinin potently suppressed neutrophil invasion into both peritoneum and lung in vivo and inhibited the release of cytokines/chemokines and NETs. This work suggests that artemisinin may have value as a therapy in conditions such as sepsis and Covid-19 in which over-activation of the innate immune system causes tissue injury that can lead to death.


Subject(s)
Artemisinins , COVID-19 , Extracellular Traps , Macrophages , Neutrophils , Sepsis , Animals , Artemisinins/pharmacology , COVID-19/drug therapy , Calcium/metabolism , Calcium-Transporting ATPases/metabolism , Chemotaxis/drug effects , Cytokines/biosynthesis , Cytokines/metabolism , Extracellular Traps/metabolism , Macrophages/drug effects , Macrophages/metabolism , Mice , Neutrophils/drug effects , Neutrophils/metabolism , Thapsigargin/pharmacology
20.
Front Immunol ; 13: 879686, 2022.
Article in English | MEDLINE | ID: covidwho-1903014

ABSTRACT

Neutrophils play a significant role in determining disease severity following SARS-CoV-2 infection. Gene and protein expression defines several neutrophil clusters in COVID-19, including the emergence of low density neutrophils (LDN) that are associated with severe disease. The functional capabilities of these neutrophil clusters and correlation with gene and protein expression are unknown. To define host defense and immunosuppressive functions of normal density neutrophils (NDN) and LDN from COVID-19 patients, we recruited 64 patients with severe COVID-19 and 26 healthy donors (HD). Phagocytosis, respiratory burst activity, degranulation, neutrophil extracellular trap (NET) formation, and T-cell suppression in those neutrophil subsets were measured. NDN from severe/critical COVID-19 patients showed evidence of priming with enhanced phagocytosis, respiratory burst activity, and degranulation of secretory vesicles and gelatinase and specific granules, while NET formation was similar to HD NDN. COVID LDN response was impaired except for enhanced NET formation. A subset of COVID LDN with intermediate CD16 expression (CD16Int LDN) promoted T cell proliferation to a level similar to HD NDN, while COVID NDN and the CD16Hi LDN failed to stimulate T-cell activation. All 3 COVID-19 neutrophil populations suppressed stimulation of IFN-γ production, compared to HD NDN. We conclude that NDN and LDN from COVID-19 patients possess complementary functional capabilities that may act cooperatively to determine disease severity. We predict that global neutrophil responses that induce COVID-19 ARDS will vary depending on the proportion of neutrophil subsets.


Subject(s)
COVID-19 , Extracellular Traps , Extracellular Traps/metabolism , Humans , Neutrophils/metabolism , Respiratory Burst , SARS-CoV-2
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