Publications
337 results found
Naish E, Wood AJ, Stewart AP, et al., 2023, The formation and function of the neutrophil phagosome., Immunological Reviews, Vol: 314, Pages: 158-180, ISSN: 0105-2896
Neutrophils are the most abundant circulating leukocyte and are crucial to the initial innate immune response to infection. One of their key pathogen-eliminating mechanisms is phagocytosis, the process of particle engulfment into a vacuole-like structure called the phagosome. The antimicrobial activity of the phagocytic process results from a collaboration of multiple systems and mechanisms within this organelle, where a complex interplay of ion fluxes, pH, reactive oxygen species, and antimicrobial proteins creates a dynamic antimicrobial environment. This complexity, combined with the difficulties of studying neutrophils ex vivo, has led to gaps in our knowledge of how the neutrophil phagosome optimizes pathogen killing. In particular, controversy has arisen regarding the relative contribution and integration of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-derived antimicrobial agents and granule-delivered antimicrobial proteins. Clinical syndromes arising from dysfunction in these systems in humans allow useful insight into these mechanisms, but their redundancy and synergy add to the complexity. In this article, we review the current knowledge regarding the formation and function of the neutrophil phagosome, examine new insights into the phagosomal environment that have been permitted by technological advances in recent years, and discuss aspects of the phagocytic process that are still under debate.
Pisacano N, McAdoo SP, Guck J, et al., 2022, Biomechanical Phenotype of Circulating Neutrophils Is Altered in ANCA Associated Vasculitis, Publisher: WILEY, Pages: 3359-3361, ISSN: 2326-5191
Stacey D, Chen L, Stanczyk PJ, et al., 2022, Elucidating mechanisms of genetic cross-disease associations at the <i>PROCR</i> vascular disease locus (vol 13, 1222, 2022), NATURE COMMUNICATIONS, Vol: 13
Stacey D, Chen L, Stanczyk P, et al., 2022, Elucidating mechanisms of genetic cross-disease associations at the PROCR vascular disease locus, Nature Communications, Vol: 13, ISSN: 2041-1723
Many individual genetic risk loci have been associated with multiple common human diseases. However, themolecular basis of this pleiotropy often remains unclear. We present an integrative approach to reveal themolecular mechanism underlying the PROCR locus, associated with lower coronary artery disease (CAD) riskbut higher venous thromboembolism (VTE) risk. We identify PROCR-p.Ser219Gly as the likely causal variantat the locus and protein C as a causal factor. Using genetic analyses, human recall-by-genotype and in vitroexperimentation, we demonstrate that PROCR-219Gly increases plasma levels of (activated) protein C throughendothelial protein C receptor (EPCR) ectodomain shedding in endothelial cells, attenuating leukocyte–endothelial cell adhesion and vascular inflammation. We also associate PROCR-219Gly with an increased pro-thrombotic state via coagulation factor VII, a ligand of EPCR. Our study, which links PROCR-219Gly to CADthrough anti-inflammatory mechanisms and to VTE through pro-thrombotic mechanisms, provides aframework to reveal the mechanisms underlying similar cross-phenotype associations.
Lodge K, Vassallo A, Liu B, et al., 2021, Hypoxia increases the potential for neutrophil-mediated endothelial damage in COPD, American Journal of Respiratory and Critical Care Medicine, Vol: 205, ISSN: 1073-449X
Rationale: Chronic obstructive pulmonary disease (COPD) patients experience excess cardiovascular morbidity and mortality, and exacerbations further increase the risk of such events. COPD is associated with persistent blood and airway neutrophilia, and systemic and tissue hypoxia. Hypoxia augments neutrophil elastase release, enhancing capacity for tissue injury.Objective: To determine whether hypoxia-driven neutrophil protein secretion contributes to endothelial damage in COPD.Methods: The healthy human neutrophil secretome generated under normoxic or hypoxic conditions was characterised by quantitative mass spectrometry, and the capacity for neutrophil-mediated endothelial damage assessed. Histotoxic protein levels were measured in normoxic versus hypoxic neutrophil supernatants and plasma from exacerbating COPD patients and healthy controls.Main results: Hypoxia promoted PI3Kγ-dependent neutrophil elastase secretion, with greater release seen in neutrophils from COPD patients. Supernatants from neutrophils incubated under hypoxia caused pulmonary endothelial cell damage and identical supernatants from COPD neutrophils increased neutrophil adherence to endothelial cells. Proteomics revealed differential neutrophil protein secretion under hypoxia and normoxia; hypoxia augmented secretion of a subset of histotoxic granule and cytosolic proteins, with significantly greater release seen in COPD neutrophils. The plasma of COPD patients had higher content of hypoxia-upregulated neutrophil-derived proteins and protease activity, and vascular injury markers.Conclusions: Hypoxia drives a destructive ‘hyper-secretory’ neutrophil phenotype conferring enhanced capacity for endothelial injury, with a corresponding signature of neutrophil degranulation and vascular injury identified in COPD patient plasma. Thus, hypoxic enhancement of neutrophil degranulation may contribute to increased cardiovascular risk in COPD. These insights may identify new therapeutic o
Johnson H, Piggin M, McKibben S, et al., 2021, Insight Report: Respiratory medicine research prioritisation online survey
Insight report summarising the findings from an online survey aiming to help shape respiratory medicine research priorities in North West London.
Wilson AM, Clark AB, Cahn A, et al., 2021, Co-trimoxazole to reduce mortality, transplant, or unplanned hospitalisation in people with moderate to very severe idiopathic pulmonary fibrosis: the EME-TIPAC RCT, Efficacy and Mechanism Evaluation, Vol: 8, Pages: 1-110, ISSN: 2050-4365
<jats:sec id="abs1-1"><jats:title>Background</jats:title><jats:p>Idiopathic pulmonary fibrosis is an irreversible fibrosing lung disorder with a poor prognosis. Current treatments slow the rate of decline in lung function and may influence survival, but they have a significant side-effect profile and so additional therapeutic options are required. People with idiopathic pulmonary fibrosis have altered innate immunity and altered lung microbiota, with the bacterial burden relating to mortality. Two randomised controlled trials have demonstrated beneficial effects with co-trimoxazole (SEPTRIN<jats:sup>®</jats:sup>; Essential Generics Ltd, Egham, UK; Chemidex Generics Ltd, Egham, UK), with the suggestion of an improvement in rates of survival.</jats:p></jats:sec><jats:sec id="abs1-2"><jats:title>Objectives</jats:title><jats:p>To determine the clinical efficacy of co-trimoxazole in people with moderate to severe idiopathic pulmonary fibrosis.</jats:p></jats:sec><jats:sec id="abs1-3"><jats:title>Design</jats:title><jats:p>A Phase II, double-blind, placebo-controlled, parallel-group, randomised multicentre study.</jats:p></jats:sec><jats:sec id="abs1-4"><jats:title>Setting</jats:title><jats:p>UK specialist interstitial lung disease centres.</jats:p></jats:sec><jats:sec id="abs1-5"><jats:title>Participants</jats:title><jats:p>Patients who were randomised had idiopathic pulmonary fibrosis diagnosed by a multidisciplinary team. In addition, patients had significant breathlessness (i.e. a Medical Research Council Dyspnoea Scale score of > 1) and impaired lung function (i.e. a forced vital capacity of < 75% predicted). Patients could be taking licensed medication for idiopathic pulmonary fibrosis, but were excluded if t
Li W, Long L, Yang X, et al., 2021, Circulating BMP9 protects the pulmonary endothelium during inflammation-induced lung injury in mice, American Journal of Respiratory and Critical Care Medicine, Vol: 11, Pages: 1419-1430, ISSN: 1073-449X
Rationale: Pulmonary endothelial permeability contributes to the high-permeability pulmonary edema that characterizes acute respiratory distress syndrome (ARDS). Circulating bone morphogeneic protein 9 (BMP9) is emerging as an important regulator of pulmonary vascular homeostasis. Objective: To determine whether endogenous BMP9 plays a role in preserving pulmonary endothelial integrity, and whether loss of endogenous BMP9 occurs during lipopolysacharride (LPS) challenge. Methods: A BMP9-neutralizing antibody was administrated to healthy adult mice and lung vasculature was examined. Potential mechanisms were delineated by transcript analysis in human lung endothelial cells. Impact of BMP9 administration was evaluated in a murine acute lung injury model induced by inhaled LPS. Levels of BMP9 were measured in plasma from patients with sepsis and endotoxemic mice. Main Results: Subacute neutralization of endogenous BMP9 in mice (N=12) resulted in increased lung vascular permeability (P=0.022), interstitial edema (P=0.0047) and neutrophil extravasation (P=0.029) compared with IgG control (N=6). In pulmonary endothelial cells, BMP9 regulated transcriptome pathways implicated in vascular permeability and cell membrane integrity. Augmentation of BMP9 signaling in mice (N=8) prevented inhaled LPS-induced lung injury (P=0.0027) and edema (P<0.0001). In endotoxemic mice (N=12), endogenous BMP9 levels were markedly reduced, due to a transient reduction in hepatic BMP9 mRNA expression and increased elastase activity in plasma. In human sepsis patients (N=10), circulating levels of BMP9 were also markedly reduced (P<0.0001). Conclusions: Endogenous circulating BMP9 is a pulmonary endothelial protective factor, down-regulated during inflammation. Exogenous BMP9 offers a potential therapy to prevent increased pulmonary endothelial permeability in lung injury.
Farahi N, Gillett D, Loutsios C, et al., 2021, Measurement of eosinophil kinetics In vivo, Methods in Molecular Biology, Vol: 2241, Pages: 183-191, ISSN: 1064-3745
Radiolabeled leukocyte scans are used in nuclear medicine to detect sites of infection and inflammation. We have previously demonstrated the use of clinical grade immunomagnetic beads to isolate autologous eosinophils and image their distribution in healthy volunteers. Here we describe the use of radiolabeled eosinophils coupled to single-photon emission computed tomography (SPECT) to quantify eosinophil uptake in the lungs of healthy volunteers, patients with asthma, and patients with focal eosinophilic inflammation.
Bashant KR, Aponte AM, Randazzo D, et al., 2021, Proteomic, biomechanical and functional analyses define neutrophil heterogeneity in systemic lupus erythematosus, Annals of the Rheumatic Diseases, Vol: 80, Pages: 209-218, ISSN: 0003-4967
OBJECTIVES: Low-density granulocytes (LDGs) are a distinct subset of proinflammatory and vasculopathic neutrophils expanded in systemic lupus erythematosus (SLE). Neutrophil trafficking and immune function are intimately linked to cellular biophysical properties. This study used proteomic, biomechanical and functional analyses to further define neutrophil heterogeneity in the context of SLE. METHODS: Proteomic/phosphoproteomic analyses were performed in healthy control (HC) normal density neutrophils (NDNs), SLE NDNs and autologous SLE LDGs. The biophysical properties of these neutrophil subsets were analysed by real-time deformability cytometry and lattice light-sheet microscopy. A two-dimensional endothelial flow system and a three-dimensional microfluidic microvasculature mimetic (MMM) were used to decouple the contributions of cell surface mediators and biophysical properties to neutrophil trafficking, respectively. RESULTS: Proteomic and phosphoproteomic differences were detected between HC and SLE neutrophils and between SLE NDNs and LDGs. Increased abundance of type 1 interferon-regulated proteins and differential phosphorylation of proteins associated with cytoskeletal organisation were identified in SLE LDGs relative to SLE NDNs. The cell surface of SLE LDGs was rougher than in SLE and HC NDNs, suggesting membrane perturbances. While SLE LDGs did not display increased binding to endothelial cells in the two-dimensional assay, they were increasingly retained/trapped in the narrow channels of the lung MMM. CONCLUSIONS: Modulation of the neutrophil proteome and distinct changes in biophysical properties are observed alongside differences in neutrophil trafficking. SLE LDGs may be increasingly retained in microvasculature networks, which has important pathogenic implications in the context of lupus organ damage and small vessel vasculopathy.
Lodge KM, Morrison T, Cowburn AS, et al., 2021, Leukocytes/Neutrophils, Encyclopedia of Respiratory Medicine, Second Edition, Pages: 200-206, ISBN: 9780081027233
The neutrophil is a phagocytic leukocyte, the most abundant in the human body and the most short-lived of all myeloid cells. This polymorphonuclear cell is packed with granules containing potent antimicrobial compounds and is absolutely essential for day-to-day successful defense against bacterial and fungal infections. Its recruitment and activation are carefully regulated, and control of its life span coupled with mechanisms allowing for a noninflammatory death pathway (apoptosis) reduce risks of tissue damage and disease arising from unwanted neutrophil activation. Given their destructive potential, and despite intensive regulation of their function, it is unsurprising that neutrophilic inflammatory diseases are prevalent; these include asthma, chronic obstructive pulmonary disease, the acute respiratory distress syndrome, and several vasculitides.
Wilson AM, Clark AB, Cahn T, et al., 2020, Effect of co-trimoxazole (Trimethoprim-Sulfamethoxazole) vs placebo on death, lung transplant, or hospital admission in patients with moderate and severe idiopathic pulmonary fibrosis the EME-TIPAC randomized clinical trial, JAMA: Journal of the American Medical Association, Vol: 324, Pages: 2282-2291, ISSN: 0098-7484
Importance Idiopathic pulmonary fibrosis (IPF) has a poor prognosis and limited treatment options. Patients with IPF have altered lung microbiota, with bacterial burden within the lungs associated with mortality; previous studies have suggested benefit with co-trimoxazole (trimethoprim-sulfamethoxazole).Objective To determine the efficacy of co-trimoxazole in patients with moderate and severe IPF.Design, Setting, and Participants Double-blind, placebo-controlled, parallel randomized trial of 342 patients with IPF, breathlessness (Medical Research Council dyspnea scale score >1), and impaired lung function (forced vital capacity ≤75% predicted) conducted in 39 UK specialist interstitial lung disease centers between April 2015 (first patient visit) and April 2019 (last patient follow-up).Interventions Study participants were randomized to receive 960 mg of oral co-trimoxazole twice daily (n = 170) or matched placebo (n = 172) for between 12 and 42 months. All patients received 5 mg of folic acid orally once daily.Main Outcomes and Measures The primary outcome was time to death (all causes), lung transplant, or first nonelective hospital admission. There were 15 secondary outcomes, including the individual components of the primary end point respiratory-related events, lung function (forced vital capacity and gas transfer), and patient-reported outcomes (Medical Research Council dyspnea scale, 5-level EuroQol 5-dimension questionnaire, cough severity, Leicester Cough Questionnaire, and King’s Brief Interstitial Lung Disease questionnaire scores).Results Among 342 individuals who were randomized (mean age, 71.3 years; 46 [13%] women), 283 (83%) completed the trial. The median (interquartile range) duration of follow-up was 1.02 (0.35-1.73) years. Events per person-year of follow-up among participants randomized to the co-trimoxazole and placebo groups were 0.45 (84/186) and 0.38 (80/209), respectively, with a hazard ratio of
Farahi N, Gillett D, Southwood M, et al., 2020, Lesson of the month: novel method to quantify neutrophil uptake in early lung cancer using SPECT-CT., Thorax, Vol: 75, Pages: 1020-1023, ISSN: 0040-6376
Neutrophils play an important role in the lung tumour microenvironment. We hypothesised that radiolabelled neutrophils coupled to single-photon emission CT (SPECT) may non-invasively quantify neutrophil uptake in tumours from patients with non-small cell lung cancer. We demonstrated increased uptake of radiolabelled neutrophils from the blood into tumours compared with non-specific uptake using radiolabelled transferrin. Moreover, indium-111-neutrophil activity in the tumour biopsies also correlated with myeloperoxidase (MPO)-positive neutrophils. Our data support the utility of imaging with In-111-labelled neutrophils and SPECT-CT to quantify neutrophil uptake in lung cancer.
Wood AJ, Vassallo AM, Ruchaud-Sparagano M-H, et al., 2020, C5a impairs phagosomal maturation in the neutrophil through phosphoproteomic remodeling., JCI Insight, Vol: 5, Pages: 1-17, ISSN: 2379-3708
Critical illness is accompanied by the release of large amounts of the anaphylotoxin, C5a. C5a suppresses antimicrobial functions of neutrophils which is associated with adverse outcomes. The signaling pathways that mediate C5a-induced neutrophil dysfunction are incompletely understood. Healthy donor neutrophils exposed to purified C5a demonstrated a prolonged defect (7 hours) in phagocytosis of Staphylococcus aureus. Phosphoproteomic profiling of 2712 phosphoproteins identified persistent C5a signaling and selective impairment of phagosomal protein phosphorylation on exposure to S. aureus. Notable proteins included early endosomal marker ZFYVE16 and V-ATPase proton channel component ATPV1G1. An assay of phagosomal acidification demonstrated C5a-induced impairment of phagosomal acidification, which was recapitulated in neutrophils from critically ill patients. Examination of the C5a-impaired protein phosphorylation indicated a role for the PI3K VPS34 in phagosomal maturation. Inhibition of VPS34 impaired neutrophil phagosomal acidification and killing of S. aureus. This study provides a phosphoproteomic assessment of human neutrophil signaling in response to S. aureus and its disruption by C5a, identifying a defect in phagosomal maturation and mechanisms of immune failure in critical illness.
Lau D, Garcon F, Chandra A, et al., 2020, Intravital imaging of adoptive T-Cell morphology, mobility and trafficking following immune checkpoint inhibition in a mouse melanoma model, Frontiers in Immunology, Vol: 11, Pages: 1-17, ISSN: 1664-3224
Efficient T-cell targeting, infiltration and activation within tumors is crucial for successful adoptive T-cell therapy. Intravital microscopy is a powerful tool for the visualization of T-cell behavior within tumors, as well as spatial and temporal heterogeneity in response to immunotherapy. Here we describe an experimental approach for intravital imaging of adoptive T-cell morphology, mobility and trafficking in a skin-flap tumor model, following immune modulation with immune checkpoint inhibitors (ICIs) targeting PD-L1 and CTLA-4. A syngeneic model of ovalbumin and mCherry-expressing amelanotic mouse melanoma was used in conjunction with adoptively transferred OT-1+ cytotoxic T-cells expressing GFP to image antigen-specific live T-cell behavior within the tumor microenvironment. Dynamic image analysis of T-cell motility showed distinct CD8+ T-cell migration patterns and morpho-dynamics within different tumor compartments in response to ICIs: this approach was used to cluster T-cell behavior into four groups based on velocity and meandering index. The results showed that most T-cells within the tumor periphery demonstrated Lévy-like trajectories, consistent with tumor cell searching strategies. T-cells adjacent to tumor cells had reduced velocity and appeared to probe the local environment, consistent with cell-cell interactions. An increased number of T-cells were detected following treatment, traveling at lower mean velocities than controls, and demonstrating reduced displacement consistent with target engagement. Histogram-based analysis of immunofluorescent images from harvested tumors showed that in the ICI-treated mice there was a higher density of CD31+ vessels compared to untreated controls and a greater infiltration of T-cells towards the tumor core, consistent with increased cellular trafficking post-treatment.
Pillay J, Tregay N, Juzenaite G, et al., 2020, Effect of the CXCR4 antagonist plerixafor on endogenous neutrophil dynamics in the bone marrow, lung and spleen, Journal of Leukocyte Biology, Vol: 107, Pages: 1175-1185, ISSN: 0741-5400
Treatment with the CXCR4 antagonist, plerixafor (AMD3100), has been proposed for clinical use in patients with WHIM (warts, hypogammaglobulinemia, infections, and myelokathexis) syndrome and in pulmonary fibrosis. However, there is controversy with respect to the impact of plerixafor on neutrophil dynamics in the lung, which may affect its safety profile. In this study, we investigated the kinetics of endogenous neutrophils by direct imaging, using confocal intravital microscopy in mouse bone marrow, spleen, and lungs. Neutrophils are observed increasing their velocity and exiting the bone marrow following plerixafor administration, with a concomitant increase in neutrophil numbers in the blood and spleen, while the marginated pool of neutrophils in the lung microvasculature remained unchanged in terms of numbers and cell velocity. Use of autologous radiolabeled neutrophils and SPECT/CT imaging in healthy volunteers showed that plerixafor did not affect GM‐CSF‐primed neutrophil entrapment or release in the lungs. Taken together, these data suggest that plerixafor causes neutrophil mobilization from the bone marrow but does not impact on lung marginated neutrophil dynamics and thus is unlikely to compromise respiratory host defense both in humans and mice.
Samanta R, Dunning J, Taylor A, et al., 2020, Severe Respiratory Failure in Adult Influenza Infection Is Characterised by Mechanisms Relating to Pulmonary Endothelial Leak and Interferon Gamma Induced Translational Silencing, International Conference of the American-Thoracic-Society (ATS), Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Bashant KR, Toepfner N, Day CJ, et al., 2020, The mechanics of myeloid cells, Biology of the Cell, Vol: 112, Pages: 103-112, ISSN: 0248-4900
The effects of cell size, shape and deformability on cellular function have long been a topic of interest. Recently, mechanical phenotyping technologies capable of analysing large numbers of cells in real time have become available. This has important implications for biology and medicine, especially haemato‐oncology and immunology, as immune cell mechanical phenotyping, immunologic function, and malignant cell transformation are closely linked and potentially exploitable to develop new diagnostics and therapeutics. In this review, we introduce the technologies used to analyse cellular mechanical properties and review emerging findings following the advent of high throughput deformability cytometry. We largely focus on cells from the myeloid lineage, which are derived from the bone marrow and include macrophages, granulocytes and erythrocytes. We highlight advances in mechanical phenotyping of cells in suspension that are revealing novel signatures of human blood diseases and providing new insights into pathogenesis of these diseases. The contributions of mechanical phenotyping of cells in suspension to our understanding of drug mechanisms, identification of novel therapeutics and monitoring of treatment efficacy particularly in instances of haematologic diseases are reviewed, and we suggest emerging topics of study to explore as high throughput deformability cytometers become prevalent in laboratories across the globe.
Thillai M, Ho J, Williams L, et al., 2020, Retrospective Analysis of 316 Cases of Cardiac Sarcoidosis Over Three Decades, Virtual International Conference of the American-Thoracic-Society, Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Wilson A, Hammond M, Clark A, et al., 2020, The Efficacy and Mechanism Evaluation of Treating Idiopathic Pulmonary Fibrosis with the Addition of Co-Trimoxazole - A Randomised Controlled Trial, Virtual International Conference of the American-Thoracic-Society, Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Lok LSC, Dennison TW, Mahbubani KM, et al., 2019, Phenotypically distinct neutrophils patrol uninfected human and mouse lymph nodes, Proceedings of the National Academy of Sciences of the United States of America, Vol: 116, Pages: 19083-19089, ISSN: 0027-8424
Neutrophils play a key role in innate immunity. As the dominant circulating phagocyte, they are rapidly recruited from the bloodstream to sites of infection or injury to internalize and destroy microbes. More recently, neutrophils have been identified in uninfected organs, challenging the classical view of their function. Here we showed that neutrophils were present in lymph nodes (LNs) in homeostasis. Using flow cytometry and confocal imaging, we identified neutrophils within LNs in naive, unchallenged mice, including LNs draining the skin, lungs and gastrointestinal tract. Neutrophils were enriched within specific anatomical regions, in the interfollicular zone, a site of T cell activation. Intravital two-photon microscopy demonstrated that LN neutrophils were motile, trafficked into LNs from both blood and tissues via high endothelial venules and afferent lymphatics respectively, and formed interactions with DCs in LNs. Murine and human LN neutrophils had a distinct phenotype compared to circulating neutrophils, with higher major histocompatibility complex II (MHCII) expression, suggesting a potential role in CD4 T cell activation. Upon ex vivo stimulation with IgG immune complex (IC), neutrophils upregulated expression of MHCII and co-stimulatory molecules, and increased T cell activation. In vivo, neutrophils were capable of delivering circulating IC to LNs, suggesting a broader functional remit. Overall, our data challenge the perception that neutrophil patrol is limited to the circulation in homeostasis, adding LNs to their routine surveillance territory
Vassallo A, Wood AJ, Subburayalu J, et al., 2019, The counter-intuitive role of the neutrophil in the acute respiratory distress syndrome, British Medical Bulletin, Vol: 131, Pages: 43-55, ISSN: 0007-1420
IntroductionNeutrophils are the primary effectors of the innate immune system but are profoundly histotoxic cells. The acute respiratory distress syndrome (ARDS) is considered to be a prime example of neutrophil-mediated tissue injury.Sources of dataThe information presented in this review is acquired from the published neutrophil cell biology literature and the longstanding interest of the senior authors in ARDS pathogenesis and clinical management.Areas of agreementInvestigators in the field would agree that neutrophils accumulate in high abundance in the pulmonary microcirculation, lung interstitium and alveolar airspace of patients with ARDS. ARDS is also associated with systemic neutrophil priming and delayed neutrophil apoptosis and clearance of neutrophils from the lungs. In animal models, reducing circulating neutrophil numbers ameliorates lung injury.Areas of controversyAreas of uncertainty include how neutrophils get stuck in the narrow pulmonary capillary network—whether this reflects changes in the mechanical properties of primed neutrophils alone or additional cell adhesion molecules, the role of neutrophil sub-sets or polarization states including pro-angiogenic and low-density neutrophils, whether neutrophil extracellular trap (NET) formation is beneficial (through bacterial capture) or harmful and the potential for neutrophils to participate in inflammatory resolution. The latter may involve the generation of specialized pro-resolving molecules (SPMs) and MMP-9, which is required for adequate matrix processing.Growing pointsDifferent and possibly stable endotypes of ARDS are increasingly being recognized, yet the relative contribution of the neutrophil to these endotypes is uncertain. There is renewed and intense interest in understanding the complex ‘new biology’ of the neutrophil, specifically whether this cell might be a valid therapeutic target in ARDS and other neutrophil-driven diseases and developing understanding of ways to
Hidalgo A, Chilvers ER, Summers C, et al., 2019, The neutrophil life cycle, Trends in Immunology, Vol: 40, Pages: 584-597, ISSN: 1471-4981
Neutrophils are recognized as an essential part of the innate immune response, but an active debate still exists regarding the life cycle of these cells. Neutrophils first differentiate in the bone marrow through progenitor intermediaries before entering the blood, in a process that gauges the extramedullary pool size. Once believed to be directly eliminated in the marrow, liver, and spleen, neutrophils, after circulating for less than 1 day, are now known to redistribute into multiple tissues with poorly understood kinetics. In this review, we provide an update on the dynamic distribution of neutrophils across tissues in health and disease, and emphasize differences between humans and model organisms. We further highlight issues to be addressed to exploit the unique features of neutrophils in the clinic.
Tregay N, Begg M, Cahn A, et al., 2019, Use of autologous 99mTechnetium-labelled neutrophils to quantify lung neutrophil clearance in COPD, Thorax, Vol: 74, Pages: 659-666, ISSN: 1468-3296
RATIONALE: There is a need to develop imaging protocols which assess neutrophilic inflammation in the lung. AIM: To quantify whole lung neutrophil accumulation in (1) healthy volunteers (HV) following inhaled lipopolysaccharide (LPS) or saline and (2) patients with COPD using radiolabelled autologous neutrophils and single-photon emission computed tomography/CT (SPECT/CT). METHODS: 20 patients with COPD (Global initiative for chronic obstructive lung disease (GOLD) stages 2-3) and 18 HVs were studied. HVs received inhaled saline (n=6) or LPS (50 µg, n=12) prior to the injection of radiolabelled cells. Neutrophils were isolated using dextran sedimentation and Percoll plasma gradients and labelled with 99mTechnetium (Tc)-hexamethylpropyleneamine oxime. SPECT was performed over the thorax/upper abdomen at 45 min, 2 hours, 4 hours and 6 hours. Circulating biomarkers were measured prechallenge and post challenge. Blood neutrophil clearance in the lung was determined using Patlak-Rutland graphical analysis. RESULTS: There was increased accumulation of 99mTc-neutrophils in the lungs of patients with COPD and LPS-challenged subjects compared with saline-challenged subjects (saline: 0.0006±0.0003 mL/min/mL lung blood distribution volume [mean ±1 SD]; COPD: 0.0022±0.0010 mL/min/mL [p<0.001]; LPS: 0.0025±0.0008 mL/min/mL [p<0.001]). The accumulation of labelled neutrophils in 10 patients with COPD who underwent repeat radiolabelling/imaging 7-10 days later was highly reproducible (0.0022±0.0010 mL/min/mL vs 0.0023±0.0009 mL/min/mL). Baseline interleukin (IL)-6 levels in patients with COPD were elevated compared with HVs (1.5±1.06 pg/mL [mean ±1 SD] vs 0.4±0.24 pg/mL). LPS challenge increased the circulating IL-6 levels (7.5±2.72 pg/mL) 9 hours post challenge. CONCLUSIONS: This study sho
De Alessandris S, Ferguson GJ, Dodd AJ, et al., 2019, Neutrophil GM-CSF receptor dynamics in acute lung injury, Journal of Leukocyte Biology, Vol: 105, Pages: 1183-1194, ISSN: 0741-5400
GM‐CSF is important in regulating acute, persistent neutrophilic inflammation in certain settings, including lung injury. Ligand binding induces rapid internalization of the GM‐CSF receptor (GM‐CSFRα) complex, a process essential for signaling. Whereas GM‐CSF controls many aspects of neutrophil biology, regulation of GM‐CSFRα expression is poorly understood, particularly the role of GM‐CSFRα in ligand clearance and whether signaling is sustained despite major down‐regulation of GM‐CSFRα surface expression. We established a quantitative assay of GM‐CSFRα surface expression and used this, together with selective anti‐GM‐CSFR antibodies, to define GM‐CSFRα kinetics in human neutrophils, and in murine blood and alveolar neutrophils in a lung injury model. Despite rapid sustained ligand‐induced GM‐CSFRα loss from the neutrophil surface, which persisted even following ligand removal, pro‐survival effects of GM‐CSF required ongoing ligand‐receptor interaction. Neutrophils recruited to the lungs following LPS challenge showed initially high mGM‐CSFRα expression, which along with mGM‐CSFRβ declined over 24 hr; this was associated with a transient increase in bronchoalveolar lavage fluid (BALF) mGM‐CSF concentration. Treating mice in an LPS challenge model with CAM‐3003, an anti‐mGM‐CSFRα mAb, inhibited inflammatory cell influx into the lung and maintained the level of BALF mGM‐CSF. Consistent with neutrophil consumption of GM‐CSF, human neutrophils depleted exogenous GM‐CSF, independent of protease activity. These data show that loss of membrane GM‐CSFRα following GM‐CSF exposure does not preclude sustained GM‐CSF/GM‐CSFRα signaling and that this receptor plays a key role in ligand clearance. Hence neutrophilic activation via GM‐CSFR may play an important role in neutrophilic lung inflammation even in the absence of high GM‐CSF levels or GM‐CSFRα expression.
Li W, Long L, Yang X, et al., 2019, Endogenous Circulating BMP9 Maintains Endothelial Cell Barrier Function, Publisher: SPRINGER, Pages: 269-270, ISSN: 0920-3206
Bashant KR, Vassallo A, Herold C, et al., 2019, Real-time deformability cytometry reveals sequential contraction and expansion during neutrophil priming, Journal of Leukocyte Biology, ISSN: 0741-5400
It has become increasingly apparent that the biomechanical properties of neutrophils impact on their trafficking through the circulation and in particularly through the pulmonary capillary bed. The retention of polarized or shape-changed neutrophils in the lungs was recently proposed to contribute to acute respiratory distress syndrome pathogenesis. Accordingly, this study tested the hypothesis that neutrophil priming is coupled to morpho-rheological (MORE) changes capable of altering cell function. We employ real-time deformability cytometry (RT-DC), a recently developed, rapid, and sensitive way to assess the distribution of size, shape, and deformability of thousands of cells within seconds. During RT-DC analysis, neutrophils can be easily identified within anticoagulated "whole blood" due to their unique granularity and size, thus avoiding the need for further isolation techniques, which affect biomechanical cell properties. Hence, RT-DC is uniquely suited to describe the kinetics of MORE cell changes. We reveal that, following activation or priming, neutrophils undergo a short period of cell shrinking and stiffening, followed by a phase of cell expansion and softening. In some contexts, neutrophils ultimately recover their un-primed mechanical phenotype. The mechanism(s) underlying changes in human neutrophil size are shown to be Na+ /H+ antiport-dependent and are predicted to have profound implications for neutrophil movement through the vascular system in health and disease.
Farahi N, Loutsios C, Tregay N, et al., 2019, Increased eosinophil uptake in the lungs of obese patients with asthma-to correct for obesity compared to obese controls Reply, JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY, Vol: 143, Pages: 1265-1266, ISSN: 0091-6749
Wood AJT, Vassallo A, Summers C, et al., 2018, C5a anaphylatoxin and its role in critical illness-induced organ dysfunction, European Journal of Clinical Investigation, Vol: 48, ISSN: 0014-2972
Critical illness is an aetiologically and clinically heterogeneous syndrome that is characterised by organ failure and immune dysfunction. Mortality in critically ill patients is driven by inflammation‐associated organ damage and a profound vulnerability to nosocomial infection. Both factors are influenced by the activated complement protein C5a, released by unbridled activation of the complement system during critical illness. C5a exerts deleterious effects on organ systems directly and suppresses antimicrobial functions of key immune cells. Whilst several recent reports have added key knowledge of the cellular signalling pathways triggered by C5a, there remain a number of areas that are incompletely understood and therapeutic opportunities are still being evaluated. In this review, we summarise the cellular basis for C5a‐induced vulnerability to nosocomial infection and organ dysfunction. We focus on cells of the innate immune system, highlighting the major areas in need of further research and potential avenues for targeted therapies.
Vignarajah M, Wood AJT, Nelmes E, et al., 2018, REGULATION OF INTERCELLULAR ADHESION MOLECULE-1 IN HUMAN NEUTROPHILS, Winter Meeting of the British-Thoracic-Society, Publisher: BMJ PUBLISHING GROUP, Pages: A52-A52, ISSN: 0040-6376
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