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Sample records for circulating neutrophil numbers

  1. Differential uptake of grepafloxacin by human circulating blood neutrophils and those exudated into tissues.

    PubMed

    Niwa, M; Hotta, K; Kanamori, Y; Matsuno, H; Kozawa, O; Hirota, M; Uematsu, T

    2001-09-28

    The uptake of the antimicrobial quinolone agent, grepafloxacin, both by human circulating blood neutrophils and by those exudated into tissues, was evaluated in vitro by comparing the intracellular drug concentrations. In circulating blood neutrophils, the uptake of grepafloxacin was rapid and saturable at 37 degrees C. The uptake of grepafloxacin into circulating blood neutrophils was reduced by lowering the environmental temperature or by the presence of metabolic inhibitors, suggesting the involvement of an active transport mechanism. Furthermore, the uptake of grepafloxacin by tissue (salivary) neutrophils was also partially temperature-dependent and was significantly greater than that by circulating blood neutrophils, i.e. exudation of neutrophils into tissue results in a markedly enhanced transport mechanism for grepafloxacin. This phenomenon may be related to the higher defense activity against infection seen in exudated tissue neutrophils.

  2. Circulating platelet-neutrophil complexes are important for subsequent neutrophil activation and migration.

    PubMed

    Kornerup, Kristin N; Salmon, Gary P; Pitchford, Simon C; Liu, Wai L; Page, Clive P

    2010-09-01

    Previous studies in our laboratory have shown that platelets are essential for the migration of eosinophils into the lungs of allergic mice, and that this is dependent on the functional expression of platelet P-selectin. We sought to investigate whether the same is true for nonallergic, acute inflammatory stimuli administered to distinct anatomic compartments. Neutrophil trafficking was induced in two models, namely zymosan-induced peritonitis and LPS-induced lung inflammation, and the platelet dependence of these responses investigated utilizing mice rendered thrombocytopenic. The relative contribution of selectins was also investigated. The results presented herein clearly show that platelet depletion (>90%) significantly inhibits neutrophil recruitment in both models. In addition, we show that P-selectin glycoprotein ligand-1, but not P-selectin, is essential for neutrophil recruitment in mice in vivo, thus suggesting the existence of different regulatory mechanisms for the recruitment of leukocyte subsets in response to allergic and nonallergic stimuli. Further studies in human blood demonstrate that low-dose prothrombotic and pro-inflammatory stimuli (CCL17 or CCL22) synergize to induce platelet and neutrophil activation, as well as the formation of platelet-neutrophil conjugates. We conclude that adhesion between platelets and neutrophils in vivo is an important event in acute inflammatory responses. Targeting this interaction may be a successful strategy for inflammatory conditions where current therapy fails to provide adequate treatment.

  3. Circulating Neutrophil MicroRNAs as Biomarkers for the Detection of Lung Cancer

    PubMed Central

    Ma, Jie; Li, Ning; Lin, Yanli; Gupta, Chhavi; Jiang, Feng

    2016-01-01

    Peripheral neutrophils are the predominant circulating leukocytes and an important component of innate and adaptive immune systems, which is a primary defense against cancer. MicroRNAs (miRNAs) can modulate neutrophil functions and play important roles in cancer pathogenesis by regulating neutrophil gene expression. To investigate if assessment of differential miRNA levels of peripheral neutrophils has the potential for diagnosis of non-small-cell lung cancer (NSCLC), we examine neutrophils of 15 patients with stage I NSCLC and 15 smokers without cancer. We identify five neutrophil miRNAs that have an abnormal level in patients with NSCLC versus smokers without cancer. In a training set of 82 patients with lung cancer and 73 controls, a set of two genes (miRs-26a-2-3p and 574-3p) are developed, producing 77.8% sensitivity and 78.1% specificity for NSCLC detection. Furthermore, in a testing set of 60 patients with lung cancer and 58 smokers, the performance of analyzing the two miRNAs for lung cancer detection is confirmed. This study for the first time shows that a neutrophil miRNA profile may serve as a new category of circulating biomarkers for the detection of NSCLC. PMID:26823654

  4. Nerve growth factor induced hyperalgesia in the rat hind paw is dependent on circulating neutrophils.

    PubMed

    Bennett, G; al-Rashed, S; Hoult, J R; Brain, S D

    1998-09-01

    The mechanisms by which nerve growth factor (NGF) induces thermal hyperalgesia and neutrophil accumulation have been investigated in the rat. Thermal nociceptive thresholds in rat hind paw were measured as the time taken for paw withdrawal from a heat source and neutrophil accumulation was measured in hind paw and dorsal skin samples using a myeloperoxidase assay. NGF (23-80 pmol intraplantar (i.pl.) injection) induced a significant (P < 0.05, n = 6-16) thermal hyperalgesia at 5 h after injection and significant neutrophil accumulation (P < 0.05, n = 6) was observed with NGF (40 pmol). In dorsal skin, where multiple samples can be assessed, intradermal (i.d.) NGF was 10-30 times less potent than interleukin-1beta in inducing neutrophil accumulation. The 5-lipoxygenase inhibitor ZM230487 (10 nmol co-injected with NGF) significantly attenuated neutrophil accumulation and hyperalgesia induced by NGF; unlike the histamine and 5-hydroxytryptamine antagonists (mepyramine and methysergide) which were without effect at the times measured. Furthermore, depletion of circulating neutrophils (using a rabbit anti-rat neutrophil antibody) abolished NGF induced hyperalgesia. These results indicate that neutrophils, which accumulate in response to a 5-lipoxygenase product, play a crucial role in NGF-induced hyperalgesia.

  5. Insulin treatment prevents wounding associated changes in tissue and circulating neutrophil MMP-9 and NGAL in diabetic rats

    PubMed Central

    Abdollahi, Maryam; Ng, Taria Shin Yi; Rezaeizadeh, Alireza; Aamidor, Sarah; Twigg, Stephen M.; Min, Danqing; McLennan, Susan V.

    2017-01-01

    Neutrophils are important for wound repair, but their persistence can impair the healing process. Neutrophils express matrix metalloproteinases including MMP-9 and its regulator neutrophil gelatinase associated lipocalin (NGAL). Whether wounding affects neutrophil MMP-9 and NGAL in diabetic animals is not known. Skin wound tissue MMP-9 and NGAL was examined by qRT-PCR and immunohistochemistry in control, diabetic and insulin treated diabetic rats. The temporal expression of MMP-9 and NGAL mRNA, MMP-9 activity and the NGAL/MMP-9 complex was also investigated in an implant model and their circulating neutrophils. The cellular localisation of MMP-9 and NGAL was confirmed by immunofluorescence and the ability of glucose to regulate these factors was examined in isolated neutrophils. In skin wound tissue compared with control, diabetes increased neutrophil infiltration, NGAL mRNA and MMP-9 protein (P<0.05). Diabetes significantly increased implant neutrophil NGAL and MMP-9 protein as well as NGAL mRNA, wound fluid NGAL/MMP-9 complex and MMP-9 activity (all <0.05). Circulating neutrophil MMP-9 and NGAL was also increased in these diabetic animals (P<0.05). These changes were prevented by insulin treatment. Ex vivo, high glucose (25mM) increased neutrophil NGAL and MMP-9 (both by 2 fold, P<0.05). NGAL and MMP-9 are increased in wound and circulating neutrophils in diabetic rodents. These changes and the association between higher NGAL and increased wound fluid MMP-9 activity suggest that increased neutrophil NGAL may contribute to increased MMP-9 in poorly healing diabetic wounds. Whether targeting neutrophil NGAL or MMP-9 can improve diabetic wound healing remains to be investigated. PMID:28182694

  6. Neutrophil extracellular trap formation and circulating nucleosomes in patients with chronic myeloproliferative neoplasms

    PubMed Central

    Marin Oyarzún, Cecilia P.; Carestia, Agostina; Lev, Paola R.; Glembotsky, Ana C.; Castro Ríos, Miguel A.; Moiraghi, Beatriz; Molinas, Felisa C.; Marta, Rosana F.; Schattner, Mirta; Heller, Paula G.

    2016-01-01

    The mechanisms underlying increased thrombotic risk in chronic myeloproliferative neoplasms (MPN) are incompletely understood. We assessed whether neutrophil extracellular traps (NETs), which promote thrombosis, contribute to the procoagulant state in essential thrombocythemia, polycythemia vera and myelofibrosis (MF) patients. Although MPN neutrophils showed increased basal reactive oxygen species (ROS), enhanced NETosis by unstimulated neutrophils was an infrequent finding, whereas PMA-triggered NETosis was impaired, particularly in MF, due to decreased PMA-triggered ROS production. Elevated circulating nucleosomes were a prominent finding and were higher in patients with advanced disease, which may have potential prognostic implication. Histone-MPO complexes, proposed as specific NET biomarker, were seldomly detected, suggesting NETs may not be the main source of nucleosomes in most patients, whereas their correlation with high LDH points to increased cell turn-over as a plausible origin. Lack of association of nucleosomes or NETs with thrombosis or activation markers does not support their use as predictors of thrombosis although prospective studies in a larger cohort may help define their potential contribution to MPN thrombosis. These results do not provide evidence for relevant in vivo NETosis in MPN patients under steady state conditions, although availability of standardized NET biomarkers may contribute to further research in this field. PMID:27958278

  7. CIRCULATING CD11B EXPRESSION CORRELATES WITH THE NEUTROPHIL RESPONSE AND AIRWAY MCD-14 EXPRESSION IS ENHANCED FOLLOWING OZONE EXPOSURE IN HUMANS

    EPA Science Inventory

    We recently reported that baseline expression of circulating CD11b is associated with the magnitude of the neutrophil response following inhaled endotoxin. In this study, we examined whether circulating CD11b plays a similar role in the inflammatory response following inhaled ozo...

  8. Automated quantitation of circulating neutrophil and eosinophil activation in asthmatic patients

    PubMed Central

    Leckie, M.; Bryan, S.; Khan, J.; Dewar, A.; Aikman, S.; McGrath, J.; Okrongly, D.; Burman, J.; Barnes, P.; Hansel, T.

    2000-01-01

    BACKGROUND—Asthma has been associated with eosinophil activation, measured in serum, sputum, bronchoalveolar lavage (BAL) fluid, and urine. A whole blood automated method was developed to assess eosinophil and neutrophil activity in terms of peroxidase content and cell morphology using the Bayer haematology analyser. The method was applied to an in vitro stimulation model when fMLP was added to whole blood and the samples were then analysed for changes in granularity and shape. In addition, cells stimulated with interleukin (IL)-8 were examined by electron microscopy.
METHODS—A cross sectional analysis was performed on venous blood from non-atopic, non-asthmatic normal subjects (n = 37), mild (n= 46) and symptomatic (n = 22) asthmatic patients on inhaled β2 agonist only, and more severe asthmatic patients (n = 17) on inhaled and oral corticosteroid therapy. Samples were analysed by the haematology analyser and peroxidase leucograms gated using the WinMDI software program.
RESULTS—There were significant differences in the amount of light scatter by the neutrophil populations in the symptomatic (p = 0.007) and severe asthmatic (p = 0.0001) groups compared with the control group. However, abnormalities in eosinophil populations were not observed. In vitro activation of whole blood with fMLP caused similar changes in neutrophil light scatter, suggesting that neutrophil activation is present in peripheral blood of symptomatic asthmatic patients. IL-8 caused a change in shape of the neutrophils seen using transmission electron microscopy.
CONCLUSIONS—Evidence of neutrophil activation can be seen in whole blood from patients with asthma using a novel automated method. This may potentially be applied to other inflammatory diseases.

 PMID:10817795

  9. Expression of CD64 on Circulating Neutrophils Favoring Systemic Inflammatory Status in Erythema Nodosum Leprosum

    PubMed Central

    Prata, Rhana Berto da Silva; Barbosa, Mayara Garcia de Mattos; Mendes, Mayara Abud; Brandão, Sheila Santos; Amadeu, Thaís Porto; Rodrigues, Luciana Silva; Ferreira, Helen; Costa, Fabrício da Mota Ramalho; dos Santos, Jessica Brandão; Pacheco, Fabiana dos Santos; Machado, Alice de Miranda; Nery, José Augusto da Costa; Hacker, Mariana de Andrea; Sales, Anna Maria; Pinheiro, Roberta Olmo; Sarno, Euzenir Nunes

    2016-01-01

    Erythema Nodosum Leprosum (ENL) is an immune reaction in leprosy that aggravates the patient´s clinical condition. ENL presents systemic symptoms of an acute infectious syndrome with high leukocytosis and intense malaise clinically similar to sepsis. The treatment of ENL patients requires immunosuppression and thus needs to be early and efficient to prevent both disabilities and permanent nerve damage. Some patients experience multiple episodes of ENL and prolonged use of immunosuppressive drugs may lead to serious adverse effects. Thalidomide treatment is extremely effective at ameliorating ENL symptoms. Several mechanisms have been proposed to explain the efficacy of thalidomide in ENL, including the inhibition of TNF production. Given its teratogenicity, thalidomide is prohibitive for women of childbearing age. A rational search for molecular targets during ENL episodes is essential to better understand the disease mechanisms involved, which may also lead to the discovery of new drugs and diagnostic tests. Previous studies have demonstrated that IFN-γ and GM-CSF, involved in the induction of CD64 expression, increase during ENL. The aim of the present study was to investigate CD64 expression during ENL and whether thalidomide treatment modulated its expression. Leprosy patients were allocated to one of five groups: (1) Lepromatous leprosy, (2) Borderline leprosy, (3) Reversal reaction, (4) ENL, and (5) ENL 7 days after thalidomide treatment. The present study demonstrated that CD64 mRNA and protein were expressed in ENL lesions and that thalidomide treatment reduced CD64 expression and neutrophil infiltrates—a hallmark of ENL. We also showed that ENL blood neutrophils exclusively expressed CD64 on the cell surface and that thalidomide diminished overall expression. Patient classification based on clinical symptoms found that severe ENL presented high levels of neutrophil CD64. Collectively, these data revealed that ENL neutrophils express CD64, presumably

  10. BIIL 284 reduces neutrophils numbers but increases P. aeruginosa bacteraemia and inflammation in mouse lungs

    PubMed Central

    Döring, Gerd; Bragonzi, Alessandra; Paroni, Moira; Aktürk, Firdevs-Fatma; Cigana, Cristina; Schmidt, Annika; Gilpin, Deirdre; Heyder, Susanne; Born, Torsten; Smaczny, Christina; Kohlhäufl, Martin; Wagner, Thomas O. F.; Loebinger, Michael R.; Bilton, Diana; Tunney, Michael M.; Elborn, J. Stuart; Pier, Gerald B.; Konstan, Michael W.; Ulrich, Martina

    2014-01-01

    Background A clinical study to investigate the leukotriene B4 (LTB4)-receptor antagonist BIIL 284 in cystic fibrosis (CF) patients was prematurely terminated due to a significantly increased risk of adverse pulmonary events. We aimed to establish the effect of BIIL284 in models of Pseudomonas aeruginosa lung infection, thereby contributing to a better understanding of what could have led to adverse pulmonary events in CF patients. Methods P. aeruginosa DNA in the blood of CF patients during and after acute pulmonary exacerbations and in stable patients with non-CF bronchiectasis (NCFB) and healthy individuals was assessed by PCR. The effect of BIIL 284 treatment was tested in an agar beads murine model of Pseudomonas aeruginosa lung infection. Bacterial count and inflammation were evaluated in lung and other organs. Result Most CF patients (98%) and all patients with NCFB and healthy individuals had negative P. aeruginosa DNA in their blood. Similarly, the P. aeruginosa-infected mice showed bacterial counts in the lung but not blood or spleen. BIIL 284 treatment decreased pulmonary neutrophils and increased P. aeruginosa numbers in mouse lungs leading to significantly higher bacteremia rates and lung inflammation compared to placebo treated animals. Conclusions Decreased airway neutrophils induced lung proliferation and severe bacteraemia in a murine model of P. aeruginosa lung infection. These data suggest that caution should be taken when administering anti-inflammatory compounds to patients with bacterial infections. PMID:24183915

  11. Serum IL8 and mRNA level of CD11b in circulating neutrophils are increased in clinically amyopathic dermatomyositis with active interstitial lung disease.

    PubMed

    Zou, Jing; Chen, Jie; Yan, Qingran; Guo, Qiang; Bao, Chunde

    2016-01-01

    The objective of this study is to assess serum IL8 and the potential activity of circulating neutrophils on relative messenger RNA (mRNA) levels and their relationship with disease activity in clinically amyopathic dermatomyositis (CADM) associated with interstitial lung disease (ILD). We studied 18 CADM patients and compared them with 18 classic dermatomyositis (DM) patients and 18 healthy control subjects. Serum IL8 level and mRNA expressions of neutrophils (chemokine (C-X-C motif) receptor 1 (CXCR1), cluster of differentiation molecule 11b (CD11b), cluster of differentiation 64 (CD64), myeloid cell leukemia 1 (MCL1), interleukin-18 (IL18)) were detected. The overproduction of serum IL8 level was most significant in the CADM group with active period. The mRNA expressions of CD11b, IL18, and MCL1 were greatly increased in the neutrophils in patients with CADM compared with DM or healthy controls. Up-expressions of CD11b, IL18, and MCL1 were detected in the neutrophils in CADM patients of active period compared with remission period. A positive correlation was found between CD11b mRNA level and high-resolution computed tomography (HRCT) score, in CADM associated with ILD. Serum IL8 level and mRNA levels of CD11b, MCL1, and IL18 in circulating neutrophils are related with the disease activity of CADM-ILD. The mRNA level of CD11b is positively correlated with HRCT score in CADM-ILD.

  12. Decreased numbers of chemotactic factor receptors in chronic neutropenia with defective chemotaxis: spontaneous recovery from the neutrophil abnormalities during early childhood

    SciTech Connect

    Yasui, K.; Yamazaki, M.; Miyagawa, Y.; Komiyama, A.; Akabane, T.

    1987-05-01

    Childhood chronic neutropenia with decreased numbers of chemotactic factor receptors as well as defective chemotaxis was first demonstrated in an 8-month-old girl. Chemotactic factor receptors on neutrophils were assayed using tritiated N-formyl-methionyl-leucyl-phenylalanine (/sup 3/H-FMLP). The patient's neutrophils had decreased numbers of the receptors: numbers of the receptors were 20,000 (less than 3 SD) as compared with those of control cells of 52,000 +/- 6000 (mean +/- SD) (n = 10). The neutropenia disappeared spontaneously by 28 months of age parallel with the improvement of chemotaxis and increase in numbers of chemotactic factor receptors. These results demonstrate a transient decrease of neutrophil chemotactic factor receptors as one of the pathophysiological bases of a transient defect of neutrophil chemotaxis in this disorder.

  13. Prediction of cloud droplet number in a general circulation model

    SciTech Connect

    Ghan, S.J.; Leung, L.R.

    1996-04-01

    We have applied the Colorado State University Regional Atmospheric Modeling System (RAMS) bulk cloud microphysics parameterization to the treatment of stratiform clouds in the National Center for Atmospheric Research Community Climate Model (CCM2). The RAMS predicts mass concentrations of cloud water, cloud ice, rain and snow, and number concnetration of ice. We have introduced the droplet number conservation equation to predict droplet number and it`s dependence on aerosols.

  14. Dermal γδ T Cells Do Not Freely Re-Circulate Out of Skin and Produce IL-17 to Promote Neutrophil Infiltration during Primary Contact Hypersensitivity

    PubMed Central

    Jiang, Xiaodong; Park, Chang Ook; Geddes Sweeney, Jenna; Yoo, Min Jae; Gaide, Olivier; Kupper, Thomas Seth

    2017-01-01

    The role of mouse dermal γδ T cells in inflammatory skin disorders and host defense has been studied extensively. It is known that dendritic epidermal T cells (DETC) have a monomorphic γδ T cell receptor (TCR) and reside in murine epidermis from birth. We asked if dermal γδ cells freely re-circulated out of skin, or behaved more like dermal resident memory T cells (TRM) in mice. We found that, unlike epidermal γδ T cells (DETC), dermal γδ cells are not homogeneous with regard to TCR, express the tissue resident T cell markers CD69 and CD103, bear skin homing receptors, and produce IL-17 and IL-22. We created GFP+: GFP− parabiotic mice and found that dermal γδ T cells re-circulate very slowly—more rapidly than authentic αβ TCR TRM, but more slowly than the recently described dermal αβ TCR T migratory memory cells (TMM). Mice lacking the TCR δ gene (δ-/-) had a significant reduction of 2,4-dinitrofluorobenzene (DNFB)-induced contact hypersensitivity (CHS). We created mice deficient in dermal γδ T cells but not DETC, and these mice also showed a markedly reduced CHS response after DNFB challenge. The infiltration of effector T cells during CHS was not reduced in dermal γδ T cell-deficient mice; however, infiltration of Gr-1+CD11b+ neutrophils, as well as ear swelling, was reduced significantly. We next depleted Gr-1+ neutrophils in vivo, and demonstrated that neutrophils are required for ear swelling, the accepted metric for a CHS response. Depletion of IL-17-producing dermal Vγ4+ cells and neutralization of IL-17 in vivo, respectively, also led to a significantly reduced CHS response and diminished neutrophil infiltration. Our findings here suggest that dermal γδ T cells have an intermediate phenotype of T cell residence, and play an important role in primary CHS through producing IL-17 to promote neutrophil infiltration. PMID:28081153

  15. Testosterone replacement therapy can increase circulating endothelial progenitor cell number in men with late onset hypogonadism.

    PubMed

    Liao, C-H; Wu, Y-N; Lin, F-Y; Tsai, W-K; Liu, S-P; Chiang, H-S

    2013-07-01

    Circulating endothelial progenitor cells (EPCs) are bone marrow-derived cells required for endothelial repair. A low EPC number can be considered as an independent predictor of endothelial dysfunction and future cardiovascular events. Recent evidence shows that patients with hypogonadal symptoms without other confounding risk factors have a low number of circulating progenitor cells (PCs) and EPCs, thus highlighting the role of testosterone in the proliferation and differentiation of EPCs. Here, we investigate if testosterone replacement therapy (TRT) can increase circulating EPC number in men with late onset hypogonadism. Forty-six men (age range, 40-73 years; mean age, 58.3 years) with hypogonadal symptoms were recruited, and 29 men with serum total testosterone (TT) levels less than 350 ng/dL received TRT using transdermal testosterone gel (Androgel; 1% testosterone at 5 g/day) for 12 months. Circulating EPC numbers (per 100 000 monocytes) were calculated using flow cytometry. There was no significant association between serum TT levels and the number of circulating EPCs before TRT. Compared with the number of mean circulating EPCs at baseline (9.5 ± 6.2), the number was significantly higher after 3 months (16.6 ± 11.1, p = 0.027), 6 months (20.3 ± 15.3, p = 0.006) and 12 months (27.2 ± 15.5, p = 0.017) of TRT. Thus, we conclude that serum TT levels before TRT are not significantly associated with the number of circulating EPCs in men with late onset hypogonadism. However, TRT can increase the number of circulating EPCs, which implies the benefit of TRT on endothelial function in hypogonadal men.

  16. Neutrophil-derived microparticles are released into the coronary circulation following percutaneous coronary intervention in acute coronary syndrome patients

    PubMed Central

    Martínez, Gonzalo J.; Barraclough, Jennifer Y.; Nakhla, Shirley; Kienzle, Vivian; Robertson, Stacy; Mallat, Ziad; Celermajer, David S.

    2016-01-01

    To evaluate (i) local coronary and systemic levels of microparticles (MP) in acute coronary syndrome (ACS) and stable angina pectoris (SAP) patients and (ii) their release after plaque disruption with percutaneous coronary intervention (PCI). MP are small vesicles originating from plasma membranes of cells after activation or apoptosis and are implicated in the pathogenesis of atherosclerosis. Neutrophils play a role in plaque destabilization and shed neutrophil-derived MP that have the potential to drive significant proinflammatory and thrombotic downstream effects. Eight ACS and eight SAP patients were included. Coronary sinus (CS) samples pre-intervention (CS1), 45 s following balloon angioplasty (CS2) and at 45 s intervals following stent deployment (CS3, CS4 and CS5), together with peripheral vein samples, pre- and post-PCI were analysed for neutrophil-derived (CD66b+), endothelial-derived (CD144+), platelet-derived (CD41a+), monocyte-derived (CD14+) and apoptotic (Annexin V+) MP. ELISA for interleukin (IL)-6, myeloperoxidase (MPO) and P-selectin was also performed. CD66b+ MP levels were similar in both groups pre-intervention. Post-PCI, CS levels rose significantly in ACS but not SAP patients (ACS area under the curve (AUC): 549 ± 83, SAP AUC: 24 ± 29, P<0.01). CS CD41a+, CD144+, CD14+ and Annexin V+ MP levels did not differ between groups. Acute neutrophil-derived MP release post-PCI occurs in ACS compared with stable patients, likely to be reflective of plaque MP content in vulnerable lesions. PMID:27913753

  17. Short-term exercise training improves flow-mediated dilation and circulating angiogenic cell number in older sedentary adults.

    PubMed

    Landers-Ramos, Rian Q; Corrigan, Kelsey J; Guth, Lisa M; Altom, Christine N; Spangenburg, Espen E; Prior, Steven J; Hagberg, James M

    2016-08-01

    Cardiovascular disease risk increases with age due, in part, to impaired endothelial function and decreased circulating angiogenic cell (CAC) number and function. We sought to determine if 10 days of aerobic exercise training improves endothelial function, CAC number, and intracellular redox balance in older sedentary adults. Eleven healthy subjects (4 men, 7 women), 61 ± 2 years of age participated in 60 min of aerobic exercise at 70% maximal oxygen consumption for 10 consecutive days while maintaining body weight. Before and after training, endothelial function was measured as flow-mediated dilation of the brachial artery and fasting blood was drawn to enumerate 3 CAC subtypes. Intracellular reactive oxygen species (ROS) and nitric oxide (NO) in CD34+ CACs were measured using fluorescent probes and reinforced via real-time quantitative polymerase chain reaction. Flow-mediated dilation improved significantly following training (10% ± 1.3% before vs. 16% ± 1.4% after training; P < 0.05). Likewise, CD34+/KDR+ number increased 104% and KDR+ number increased 151% (P < 0.05 for both), although CD34+ number was not significantly altered (P > 0.05). Intracellular NO and ROS levels in CD34+ CACs were not different after training (P > 0.05 for both). Messenger RNA expression of SOD1, endothelial nitric oxide synthase, and NADPH oxidase 2 and neutrophil cytosolic factor 1 in CD34+ CACs was not significantly altered with training (P > 0.05). In conclusion, 10 consecutive days of aerobic exercise increased flow-mediated dilation and CAC number in older, previously sedentary adults, but did not affect intracellular redox balance in CD34+ CACs. Overall, these data indicate that even short-term aerobic exercise training can have a significant impact on cardiovascular disease risk factors.

  18. Circulating Level of Neutrophil Extracellular Traps Is Not a Useful Biomarker for Assessing Disease Activity in Antineutrophil Cytoplasmic Antibody-Associated Vasculitis

    PubMed Central

    Ma, Tian-Tian; Zhang, Lu-Xia; Chen, Min; Zhao, Ming-Hui

    2016-01-01

    Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a group of life-threatening disorders, and frequently affects the kidneys. This study investigated whether the circulating neutrophil extracellular traps (NETs) levels were associated with disease activity of AAV. We collected serum samples from 34 patients with AAV in active stage and 62 patients with AAV in remission. Cell free DNA in serum was quantified using the Quant-iT PicoGreen assay. NETs associated MPO-DNA complexes, citrullinated-histone H3-DNA (cit-H3-DNA) complexes and the concentration of deoxyribonuclease I (DNase I) were quantified using ELISA. The activity of DNase I was quantified using radial enzyme-diffusion method. Associations between circulating levels of NETs with clinico-pathological parameters were analyzed. Serum levels of NETs in active AAV patients were significantly higher than those in healthy controls, and the level of cell free DNA correlated with C-reactive protein (CRP). However, no correlation was found between MPO-DNA complexes or cit-H3-DNA complexes level and CRP. Also there was no significant correlation between NETs level and initial serum creatinine, estimated glomerular filtration rate (eGFR), crescents formation or Birmingham Vasculitis Activity Score (BVAS). Furthermore, there was no significant difference of serum levels of cell free DNA or MPO-DNA complexes between active stage and remission of AAV. In conclusion, circulating levels of NETs cannot be used as a biomarker to assess disease activity in AAV patients. PMID:26840412

  19. The relationship between circulating neutrophil gelatinase-associated lipocalin and early alteration of metabolic parameters is associated with dietary saturated fat intake in non-diabetic Korean women.

    PubMed

    Na, Ga Yoon; Yoon, So Ra; An, Juhyun; Yeo, Rimkyo; Song, Juhyun; Jo, Mi-Na; Han, Seongho; Kim, Oh Yoen

    2016-12-30

    Circulating neutrophil gelatinase-associated lipocalin (NGAL) is associated with obesity-related metabolic disorders. This study investigated the relationship between serum NGAL and early alteration of metabolic parameters in non-diabetic Korean women, particularly with respect to saturated fat (SFA) intake. Anthropometric parameters, fasting glycemic status, and levels of lipids, oxidative stress/inflammatory markers, and NGAL were measured in 82 non-diabetic Korean women [Super-healthy group (n=57) with 0 metabolic syndrome risk factor (MetS RF) and MetS-risk group (n=25) with MetS RF≥1]. Age, weight, waist circumference, blood pressure, fasting glucose, HbA1C, triglyceride, LDL and total-cholesterol, and NGAL levels were higher, and HDL-cholesterol was lower in the MetS-risk group than in the Super-healthy group. Age-adjusted serum NGAL levels were higher in the MetS-risk group than in the Super-healthy group. NGAL increased proportionally with increase in MetS RFs (p=0.038) and correlated positively with BMI, triglycerides, LDL- and total-cholesterol, interleukin-6, white blood cell count, and neutrophil%, and negatively with HDL-cholesterol and superoxide dismutase activity. Serum NGAL levels positively correlated with SFA intake before and after adjustment (age and BMI). Serum NGAL levels were higher in high-SFA consumers [≥7g/day, ≥7% of total calorie intake (TCI)] than in low-SFA consumers (<7g/day, <7% of TCI). Serum NGAL levels were highest in the MetS-risk group consuming higher SFA and lowest in the Super-healthy group consuming lower SFA. However, serum NGAL did not significantly differ between the low-SFA consuming MetS-risk and Super-healthy groups. The relationship between circulating NGAL and early alteration of metabolic parameters is associated with dietary SFA intake in non-diabetic Korean women.

  20. High-Reynolds Number Circulation Control Testing in the National Transonic Facility

    NASA Technical Reports Server (NTRS)

    Milholen, William E., II; Jones, Gregory S.; Chan, David T.; Goodliff, Scott L.

    2012-01-01

    A new capability to test active flow control concepts and propulsion simulations at high Reynolds numbers in the National Transonic Facility at the NASA Langley Research Center is being developed. The first active flow control experiment was completed using the new FAST-MAC semi-span model to study Reynolds number scaling effects for several circulation control concepts. Testing was conducted over a wide range of Mach numbers, up to chord Reynolds numbers of 30 million. The model was equipped with four onboard flow control valves allowing independent control of the circulation control plenums, which were directed over a 15% chord simple-hinged flap. Preliminary analysis of the uncorrected lift data showed that the circulation control increased the low-speed maximum lift coefficient by 33%. At transonic speeds, the circulation control was capable of positively altering the shockwave pattern on the upper wing surface and reducing flow separation. Furthermore, application of the technique to only the outboard portion of the wing demonstrated the feasibility of a pneumatic based roll control capability.

  1. G-CSF maintains controlled neutrophil mobilization during acute inflammation by negatively regulating CXCR2 signaling.

    PubMed

    Bajrami, Besnik; Zhu, Haiyan; Kwak, Hyun-Jeong; Mondal, Subhanjan; Hou, Qingming; Geng, Guangfeng; Karatepe, Kutay; Zhang, Yu C; Nombela-Arrieta, César; Park, Shin-Young; Loison, Fabien; Sakai, Jiro; Xu, Yuanfu; Silberstein, Leslie E; Luo, Hongbo R

    2016-09-19

    Cytokine-induced neutrophil mobilization from the bone marrow to circulation is a critical event in acute inflammation, but how it is accurately controlled remains poorly understood. In this study, we report that CXCR2 ligands are responsible for rapid neutrophil mobilization during early-stage acute inflammation. Nevertheless, although serum CXCR2 ligand concentrations increased during inflammation, neutrophil mobilization slowed after an initial acute fast phase, suggesting a suppression of neutrophil response to CXCR2 ligands after the acute phase. We demonstrate that granulocyte colony-stimulating factor (G-CSF), usually considered a prototypical neutrophil-mobilizing cytokine, was expressed later in the acute inflammatory response and unexpectedly impeded CXCR2-induced neutrophil mobilization by negatively regulating CXCR2-mediated intracellular signaling. Blocking G-CSF in vivo paradoxically elevated peripheral blood neutrophil counts in mice injected intraperitoneally with Escherichia coli and sequestered large numbers of neutrophils in the lungs, leading to sterile pulmonary inflammation. In a lipopolysaccharide-induced acute lung injury model, the homeostatic imbalance caused by G-CSF blockade enhanced neutrophil accumulation, edema, and inflammation in the lungs and ultimately led to significant lung damage. Thus, physiologically produced G-CSF not only acts as a neutrophil mobilizer at the relatively late stage of acute inflammation, but also prevents exaggerated neutrophil mobilization and the associated inflammation-induced tissue damage during early-phase infection and inflammation.

  2. Lipopolysaccharide: a p38 MAPK-dependent disrupter of neutrophil chemotaxis.

    PubMed

    Khan, Adil I; Heit, Bryan; Andonegui, Graciela; Colarusso, Pina; Kubes, Paul

    2005-01-01

    In sepsis, and in models of sepsis including endotoxemia, impaired neutrophil recruitment and chemotaxis have been reported. The inability of the endotoxemic neutrophil to chemotax could be attributed to the fact that intracellular signaling via LPS overrides signals from endogenous chemokines or, alternatively, that sequestration of neutrophils into lungs prevents access to peripheral tissues. Using both in vitro and in vivo chemotaxis assays the authors established that neutrophils from healthy mice chemotaxed in vivo toward MIP-2, whereas endotoxemic neutrophils did not. Since LPS activates leukocytes via the p38 MAPK pathway, SKF86002, a p38 MAPK inhibitor, was given to endotoxemic animals. SKF86002 significantly reversed the LPS-induced impairment in emigration of endotoxic neutrophils in response to MIP-2. Neutrophil chemotaxis in vitro was also impaired by LPS, via a p38 MAPK-dependent pathway, and this impairment could be reversed via p38 MAPK inhibition. Although neutrophil numbers dropped in the circulation and trapped in lungs during endotoxemia, SKF86002 did not reverse these parameters, demonstrating that p38 MAPK inhibition did not release trapped neutrophils from the lungs. In conclusion, the data suggest that the impaired emigration and chemotaxis of neutrophils at peripheral sites during endotoxemia may be partially due to a p38 MAPK-mediated inhibition of neutrophil responses to endogenous chemokines.

  3. G-CSF maintains controlled neutrophil mobilization during acute inflammation by negatively regulating CXCR2 signaling

    PubMed Central

    Bajrami, Besnik; Zhu, Haiyan; Zhang, Yu C.

    2016-01-01

    Cytokine-induced neutrophil mobilization from the bone marrow to circulation is a critical event in acute inflammation, but how it is accurately controlled remains poorly understood. In this study, we report that CXCR2 ligands are responsible for rapid neutrophil mobilization during early-stage acute inflammation. Nevertheless, although serum CXCR2 ligand concentrations increased during inflammation, neutrophil mobilization slowed after an initial acute fast phase, suggesting a suppression of neutrophil response to CXCR2 ligands after the acute phase. We demonstrate that granulocyte colony-stimulating factor (G-CSF), usually considered a prototypical neutrophil-mobilizing cytokine, was expressed later in the acute inflammatory response and unexpectedly impeded CXCR2-induced neutrophil mobilization by negatively regulating CXCR2-mediated intracellular signaling. Blocking G-CSF in vivo paradoxically elevated peripheral blood neutrophil counts in mice injected intraperitoneally with Escherichia coli and sequestered large numbers of neutrophils in the lungs, leading to sterile pulmonary inflammation. In a lipopolysaccharide-induced acute lung injury model, the homeostatic imbalance caused by G-CSF blockade enhanced neutrophil accumulation, edema, and inflammation in the lungs and ultimately led to significant lung damage. Thus, physiologically produced G-CSF not only acts as a neutrophil mobilizer at the relatively late stage of acute inflammation, but also prevents exaggerated neutrophil mobilization and the associated inflammation-induced tissue damage during early-phase infection and inflammation. PMID:27551153

  4. G-CSF Analogue Treatment Increases Peripheral Neutrophil Numbers in Pigs - a Potential Alternative for In-Feed Antibiotics

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Immunomodulators is a promising area for therapeutic, prophylactic, and metaphylactic use to prevent and combat infectious disease during periods of peak disease incidence. Granulocyte colony-stimulating factor (G-CSF) enhances neutrophil production and release from the bone marrow and is already li...

  5. Central Role of Conventional Dendritic Cells in Regulation of Bone Marrow Release and Survival of Neutrophils

    PubMed Central

    Jiao, Jingjing; Dragomir, Ana-Cristina; Kocabayoglu, Peri; Rahman, Adeeb H.; Chow, Andrew; Hashimoto, Daigo; Leboeuf, Marylene; Kraus, Thomas; Moran, Thomas; Carrasco-Avino, Gonzalo; Friedman, Scott L.; Merad, Miriam; Aloman, Costica

    2014-01-01

    Neutrophils are the most abundant cell type in the immune system and play an important role in the innate immune response. Using a diverse range of mouse models with either defective DC development or conditional DC depletion, we provide in vivo evidence indicating that conventional dendritic cells (cDC) play an important role in the regulation of neutrophil homeostasis. Flk2, Flt3L and Batf3 knockout mice, which have defects in DC development, have increased numbers of liver neutrophils in the steady state. Conversely, neutrophil frequency is reduced in DC-specific PTEN knockout mice, which have an expansion of CD8+ and CD103+ DCs. In chimeric CD11c-DTR mice, cDC depletion results in a systemic increase of neutrophils in peripheral organs in the absence of histological inflammation or an increase in pro-inflammatory cytokines. This effect is also present in splenectomized chimeric CD11c-DTR mice and is absent in chimeric mice with 50% normal bone marrow. In chimeric CD11c-DTR mice, DT treatment results in enhanced neutrophil trafficking from the bone marrow into circulation and increased neutrophil recruitment. Moreover, there is an increased expression of chemokines/cytokines involved in neutrophil homeostasis and reduced neutrophil apoptosis. These data underscore the role of the DC pool in regulating the neutrophil compartment in non-lymphoid organs. PMID:24591364

  6. Circulating dendritic cell number and intracellular TNF-α production in women with type 2 diabetes.

    PubMed

    Blank, Sally E; Johnson, Emily Carolyn; Weeks, Debra K; Wysham, Carol H

    2012-12-01

    Human dendritic cell (DC) subsets perform specialized functions for surveillance against bacterial and viral infections essential for the management of type 2 diabetes (T2D). Production of tumor necrosis factor-alpha (TNF-α) by DCs acts in autocrine fashion to regulate DC maturation and promotes the inflammatory response. This study was designed to compare circulating DC number and intracellular TNF-α production between post-menopausal women with T2D and healthy women. Blood samples were obtained (n = 21/group) and examined for plasma glucose and TNF-α concentrations, and dendritic cell subset immunophenotype (plasmacytoid, pDC, CD85k(ILT-3)(+)CD123(+)CD16(-)CD14(-) and myeloid, mDC, CD85k(ILT-3)(+)CD33(+)CD123(dim to neg)CD16(-)CD14(dim to neg)). Intracellular production of TNF-α was determined in unstimulated and stimulated DCs. Women with T2D had significantly (P < 0.05) greater plasma glucose and TNF-α concentrations when compared to healthy women. Women with T2D having poor glycemic control (T2D Poor Control, HbA1c ≥ 7%) had fewer circulating pDCs than women with T2D having good glycemic control (T2D Good Control, HbA1c < 7%) and healthy women. A significant interaction (P = 0.011) was observed between the effects of plasma glucose and group for intracellular expression of TNF-α in stimulated pDCs. Intracellular production of TNF-α in pDCs was significantly greater in healthy vs. T2D Poor Control (P < 0.0001) and T2D Good Control (P < 0.0001) but did not differ between T2D subgroups. The mDC number and intracellular production of TNF-α did not differ between groups. These findings indicate that TNF-α production by pDCs was reduced in women with T2D and circulating number of pDCs was associated with glycemic control.

  7. NASA High-Reynolds Number Circulation Control Research - Overview of CFD and Planned Experiments

    NASA Technical Reports Server (NTRS)

    Milholen, W. E., II; Jones, Greg S.; Cagle, Christopher M.

    2010-01-01

    A new capability to test active flow control concepts and propulsion simulations at high Reynolds numbers in the National Transonic Facility at the NASA Langley Research Center is being developed. This technique is focused on the use of semi-span models due to their increased model size and relative ease of routing high-pressure air to the model. A new dual flow-path high-pressure air delivery station has been designed, along with a new high performance transonic sem -si pan wing model. The modular wind tunnel model is designed for testing circulation control concepts at both transonic cruise and low-speed high-lift conditions. The ability of the model to test other active flow control techniques will be highlighted. In addition, a new higher capacity semi-span force and moment wind tunnel balance has been completed and calibrated to enable testing at transonic conditions.

  8. [The influence of consecutive application of radioprotector B-190 and interleukin-1beta on changes of number of peripheral blood leucocytes and functional status of neutrophils of irradiated mice].

    PubMed

    Grebeniuk, A N; Aksenova, N V; Zatsepin, V V; Nazarov, V B; Vlasenko, T N

    2013-01-01

    The estimation of efficiency of consecutive application of radioprotector B-190 and interleukin-1beta administered at early times after acute irradiation by studying the changes of the number of peripheral blood leukocytes, neutrophils and lymphocytes, as well as cytochemical research into the contents of glycogen, myeloperoxidase and alkaline phosphatase in neutrophils of the irradiated mice was carried out. It is established that administration of radioprotector B-190 at a doze of 50 mg/kg for 15 minutes before irradiation and interleukin-1beta at a doze of 50 mkg/kg for 15 minutes after irradiation reduces the expressiveness of post-irradiation leukopenia, accelerates restoration of the number of neutrophils and lymphocytes in peripheral blood, normalizes the functional status of neutrophils of irradiated mice to a greater degree than their isolated introduction.

  9. MECHANIZED CIRCULATION SYSTEM, LEHIGH UNIVERSITY LIBRARY. LIBRARY SYSTEMS ANALYSIS, REPORT NUMBER 4.

    ERIC Educational Resources Information Center

    FLANNERY, ANNE; MACK, JAMES D.

    A MECHANIZED CIRCULATION SYSTEM CURRENTLY IN OPERATION AT LEHIGH UNIVERSITY HAS PROVEN TO GIVE RELIABLE CONTROL OF CIRCULATION ALTHOUGH IT HAS NOT SAVED ON OPERATING COSTS. WHEN THE STUDY WAS UNDERTAKEN TO DETERMINE THE FEASIBILITY OF CHANGING FROM THE PREVIOUS MANUAL SYSTEM TO THE CURRENT ONE, THE LIBRARY WAS SERVING A STUDENT BODY OF 4500…

  10. Reproducible copy number variation patterns among single circulating tumor cells of lung cancer patients

    PubMed Central

    Ni, Xiaohui; Zhuo, Minglei; Su, Zhe; Duan, Jianchun; Gao, Yan; Wang, Zhijie; Zong, Chenghang; Bai, Hua; Chapman, Alec R.; Zhao, Jun; Xu, Liya; An, Tongtong; Ma, Qi; Wang, Yuyan; Wu, Meina; Sun, Yu; Wang, Shuhang; Li, Zhenxiang; Yang, Xiaodan; Yong, Jun; Su, Xiao-Dong; Lu, Youyong; Bai, Fan; Xie, X. Sunney; Wang, Jie

    2013-01-01

    Circulating tumor cells (CTCs) enter peripheral blood from primary tumors and seed metastases. The genome sequencing of CTCs could offer noninvasive prognosis or even diagnosis, but has been hampered by low single-cell genome coverage of scarce CTCs. Here, we report the use of the recently developed multiple annealing and looping-based amplification cycles for whole-genome amplification of single CTCs from lung cancer patients. We observed characteristic cancer-associated single-nucleotide variations and insertions/deletions in exomes of CTCs. These mutations provided information needed for individualized therapy, such as drug resistance and phenotypic transition, but were heterogeneous from cell to cell. In contrast, every CTC from an individual patient, regardless of the cancer subtypes, exhibited reproducible copy number variation (CNV) patterns, similar to those of the metastatic tumor of the same patient. Interestingly, different patients with the same lung cancer adenocarcinoma (ADC) shared similar CNV patterns in their CTCs. Even more interestingly, patients of small-cell lung cancer have CNV patterns distinctly different from those of ADC patients. Our finding suggests that CNVs at certain genomic loci are selected for the metastasis of cancer. The reproducibility of cancer-specific CNVs offers potential for CTC-based cancer diagnostics. PMID:24324171

  11. Vitamin D Status in Rheumatoid Arthritis: Inflammation, Arterial Stiffness and Circulating Progenitor Cell Number

    PubMed Central

    Bagnato, Gianluca; Aragona, Caterina Oriana; Imbalzano, Egidio; D’Ascola, Angela; Rotondo, Francesco; Cinquegrani, Antonella; Mormina, Enricomaria; Saitta, Carlo; Versace, Antonio Giovanni; Sardo, Maria Adriana; Lo Gullo, Renato; Loddo, Saverio; Saitta, Antonino

    2015-01-01

    Background and Aims Suboptimal vitamin D status was recently acknowledged as an independent predictor of cardiovascular diseases and all-cause mortality in several clinical settings, and its serum levels are commonly reduced in Rheumatoid Arthritis (RA). Patients affected by RA present accelerated atherosclerosis and increased cardiovascular morbidity and mortality with respect to the general population. In RA, it has been reported an impairment of the number and the activity of circulating proangiogenic haematopoietic cells (PHCs), including CD34+, that may play a role in endothelial homeostasis. The purpose of the study is to investigate the association between vitamin D levels and PHCs, inflammatory markers, and arterial stiffening in patients with RA. Methods and Results CD34+ cells were isolated from 27 RA patients and 41 controls. Vitamin D levels, C-reactive protein (CRP), fibrinogen, pulse wave velocity (PWV), and carotid intima-media thickness (cIMT) were also evaluated. CD34+ count and vitamin D levels were lower in RA patients as compared to controls, while fibrinogen, CRP, PWV and cIMT were higher in RA patients. CD34+ cell number appeared to be associated with vitamin D levels, and negatively correlated to fibrinogen and early atherosclerosis markers (PWV and cIMT); vitamin D levels appear also to be inversely associated to fibrinogen. Conclusions RA patients with moderate disease activity presented with low vitamin D levels, low CD34+ cell count, increased PWV and cIMT; we found that vitamin D deficiency is associated to CD34+ cell reduction in peripheral blood, and with fibrinogen levels. This suggests that vitamin D might contribute to endothelial homeostasis in patients with RA. PMID:26241902

  12. Reverse-migrated neutrophils regulated by JAM-C are involved in acute pancreatitis-associated lung injury.

    PubMed

    Wu, Deqing; Zeng, Yue; Fan, Yuting; Wu, Jianghong; Mulatibieke, Tunike; Ni, Jianbo; Yu, Ge; Wan, Rong; Wang, Xingpeng; Hu, Guoyong

    2016-02-04

    Junctional adhesion molecule-C (JAM-C) plays a key role in the promotion of the reverse transendothelial migration (rTEM) of neutrophils, which contributes to the dissemination of systemic inflammation and to secondary organ damage. During acute pancreatitis (AP), systemic inflammatory responses lead to distant organ damage and typically result in acute lung injury (ALI). Here, we investigated the role of rTEM neutrophils in AP-associated ALI and the molecular mechanisms by which JAM-C regulates neutrophil rTEM in this disorder. In this study, rTEM neutrophils were identified in the peripheral blood both in murine model of AP and human patients with AP, which elevated with increased severity of lung injury. Pancreatic JAM-C was downregulated during murine experimental pancreatitis, whose expression levels were inversely correlated with both increased neutrophil rTEM and severity of lung injury. Knockout of JAM-C resulted in more severe lung injury and systemic inflammation. Significantly greater numbers of rTEM neutrophils were present both in the circulation and pulmonary vascular washout in JAM-C knockout mice with AP. This study demonstrates that during AP, neutrophils that are recruited to the pancreas may migrate back into the circulation and then contribute to ALI. JAM-C downregulation may contribute to AP-associated ALI via promoting neutrophil rTEM.

  13. Reverse-migrated neutrophils regulated by JAM-C are involved in acute pancreatitis-associated lung injury

    PubMed Central

    Wu, Deqing; Zeng, Yue; Fan, Yuting; Wu, Jianghong; Mulatibieke, Tunike; Ni, Jianbo; Yu, Ge; Wan, Rong; Wang, Xingpeng; Hu, Guoyong

    2016-01-01

    Junctional adhesion molecule-C (JAM-C) plays a key role in the promotion of the reverse transendothelial migration (rTEM) of neutrophils, which contributes to the dissemination of systemic inflammation and to secondary organ damage. During acute pancreatitis (AP), systemic inflammatory responses lead to distant organ damage and typically result in acute lung injury (ALI). Here, we investigated the role of rTEM neutrophils in AP-associated ALI and the molecular mechanisms by which JAM-C regulates neutrophil rTEM in this disorder. In this study, rTEM neutrophils were identified in the peripheral blood both in murine model of AP and human patients with AP, which elevated with increased severity of lung injury. Pancreatic JAM-C was downregulated during murine experimental pancreatitis, whose expression levels were inversely correlated with both increased neutrophil rTEM and severity of lung injury. Knockout of JAM-C resulted in more severe lung injury and systemic inflammation. Significantly greater numbers of rTEM neutrophils were present both in the circulation and pulmonary vascular washout in JAM-C knockout mice with AP. This study demonstrates that during AP, neutrophils that are recruited to the pancreas may migrate back into the circulation and then contribute to ALI. JAM-C downregulation may contribute to AP-associated ALI via promoting neutrophil rTEM. PMID:26841848

  14. Propensity of crocin to offset Vipera russelli venom induced oxidative stress mediated neutrophil apoptosis: a biochemical insight.

    PubMed

    Santhosh, M Sebastin; Sundaram, M Shanmuga; Sunitha, K; Jnaneshwari, S; Devaraja, S; Kemparaju, K; Girish, K S

    2016-01-01

    Viper envenomation results in inflammation at the bitten site as well as target organs. Neutrophils and other polymorphonuclear leukocytes execute inflammation resolving mechanism and will undergo apoptosis after completing the task. However, the target specific toxins induce neutrophil apoptosis at the bitten site and in circulation prior to their function, thus reducing their number. Circulating activated neutrophils are major source of inflammatory cytokines and leakage of reactive oxygen species (ROS)/other toxic intermediates resulting in aggravation of inflammatory response at the bitten/target site. Therefore, neutralization of venom induced neutrophil apoptosis reduces inflammation besides increasing the functional neutrophil population. Therefore, the present study investigates the venom induced perturbances in isolated human neutrophils and its neutralization by crocin (Crocus sativus) a potent antioxidant carotenoid. Human neutrophils on treatment with venom resulted in altered ROS generation, intracellular Ca(2+) mobilization, mitochondrial membrane depolarization, cyt-c translocation, caspase activation, phosphatidylserine externalization and DNA damage. On the other hand significant protection against oxidative stress and apoptosis were evidenced in crocin pre-treated groups. In conclusion the viper venom induces neutrophil apoptosis and results in aggravation of inflammation and tissue damage. The present study demands the necessity of an auxiliary therapy in addition to antivenin therapy to treat secondary/overlooked complications of envenomation.

  15. A short-term extremely low frequency electromagnetic field exposure increases circulating leukocyte numbers and affects HPA-axis signaling in mice.

    PubMed

    de Kleijn, Stan; Ferwerda, Gerben; Wiese, Michelle; Trentelman, Jos; Cuppen, Jan; Kozicz, Tamas; de Jager, Linda; Hermans, Peter W M; Verburg-van Kemenade, B M Lidy

    2016-10-01

    There is still uncertainty whether extremely low frequency electromagnetic fields (ELF-EMF) can induce health effects like immunomodulation. Despite evidence obtained in vitro, an unambiguous association has not yet been established in vivo. Here, mice were exposed to ELF-EMF for 1, 4, and 24 h/day in a short-term (1 week) and long-term (15 weeks) set-up to investigate whole body effects on the level of stress regulation and immune response. ELF-EMF signal contained multiple frequencies (20-5000 Hz) and a magnetic flux density of 10 μT. After exposure, blood was analyzed for leukocyte numbers (short-term and long-term) and adrenocorticotropic hormone concentration (short-term only). Furthermore, in the short-term experiment, stress-related parameters, corticotropin-releasing hormone, proopiomelanocortin (POMC) and CYP11A1 gene-expression, respectively, were determined in the hypothalamic paraventricular nucleus, pituitary, and adrenal glands. In the short-term but not long-term experiment, leukocyte counts were significantly higher in the 24 h-exposed group compared with controls, mainly represented by increased neutrophils and CD4 ± lymphocytes. POMC expression and plasma adrenocorticotropic hormone were significantly lower compared with unexposed control mice. In conclusion, short-term ELF-EMF exposure may affect hypothalamic-pituitary-adrenal axis activation in mice. Changes in stress hormone release may explain changes in circulating leukocyte numbers and composition. Bioelectromagnetics. 37:433-443, 2016. © 2016 The Authors. Bioelectromagnetics Published by Wiley Periodicals, Inc.

  16. A short‐term extremely low frequency electromagnetic field exposure increases circulating leukocyte numbers and affects HPA‐axis signaling in mice

    PubMed Central

    de Kleijn, Stan; Ferwerda, Gerben; Wiese, Michelle; Trentelman, Jos; Cuppen, Jan; Kozicz, Tamas; de Jager, Linda; Hermans, Peter W. M.

    2016-01-01

    There is still uncertainty whether extremely low frequency electromagnetic fields (ELF‐EMF) can induce health effects like immunomodulation. Despite evidence obtained in vitro, an unambiguous association has not yet been established in vivo. Here, mice were exposed to ELF‐EMF for 1, 4, and 24 h/day in a short‐term (1 week) and long‐term (15 weeks) set‐up to investigate whole body effects on the level of stress regulation and immune response. ELF‐EMF signal contained multiple frequencies (20–5000 Hz) and a magnetic flux density of 10 μT. After exposure, blood was analyzed for leukocyte numbers (short‐term and long‐term) and adrenocorticotropic hormone concentration (short‐term only). Furthermore, in the short‐term experiment, stress‐related parameters, corticotropin‐releasing hormone, proopiomelanocortin (POMC) and CYP11A1 gene‐expression, respectively, were determined in the hypothalamic paraventricular nucleus, pituitary, and adrenal glands. In the short‐term but not long‐term experiment, leukocyte counts were significantly higher in the 24 h‐exposed group compared with controls, mainly represented by increased neutrophils and CD4 ± lymphocytes. POMC expression and plasma adrenocorticotropic hormone were significantly lower compared with unexposed control mice. In conclusion, short‐term ELF‐EMF exposure may affect hypothalamic‐pituitary‐adrenal axis activation in mice. Changes in stress hormone release may explain changes in circulating leukocyte numbers and composition. Bioelectromagnetics. 37:433–443, 2016. © 2016 The Authors. Bioelectromagnetics Published by Wiley Periodicals, Inc. PMID:27553635

  17. Organizational Change; The Centralization of a Divisional Circulation System. Technical Paper Number Five.

    ERIC Educational Resources Information Center

    Gherman, Paul

    The circulation systems of three divisional units of Wayne State University libraries (Education, Humanities, Social-Science) needed centralization because their separate operations had created many inconsistences and unnecessary inconveniences. This paper outlines the socio-technical decisions made in the organizational change. It describes the…

  18. Solar effects on circulation types over Europe: an analysis based on a large number of classifications

    NASA Astrophysics Data System (ADS)

    Huth, R.; Cahynová, M.; Kyselý, J.

    2010-09-01

    Recently, effects of the 11-year solar cycle on various aspects of tropospheric circulation in the Northern Hemisphere in winter have been recognized. One of our previous studies showed a significant solar effect on the frequency of synoptic types from the Hess-Brezowsky catalogue. Here, we use a large collection of varied classifications of circulation patterns, defined over central Europe, assembled within the COST733 Action "Harmonization and Applications of Weather Types Classifications for European Regions" to detect the solar effect on the frequency of synoptic types. The advantage of this multi-classification approach is that peculiarities or biases present in any single classification (catalogue) that might influence the detected solar signal are eliminated once a large ensemble of classifications is used. We divide winter months (December to March) into three groups according to the mean monthly solar activity, quantified by the solar 10.7 cm flux. The three groups correspond to the minima of the 11-year solar cycle, a moderate solar activity, and solar maxima. Within each group, frequencies of occurrence of individual circulation types are calculated. Differences in the occurrence of individual classes between solar activity groups indicate the presence of a solar activity effect on atmospheric circulation over Europe. Statistical significance of these differences is estimated by a block resampling method. An enhanced frequency under solar minima and a reduced frequency under solar maxima are observed almost exclusively for the types with easterly flow over central Europe. On the other hand, a reduced frequency under solar minima and an enhanced frequency under solar maxima are found for the types with westerly flow over central Europe. The research is supported by the Grant Agency of the Czech Academy of Sciences, project A300420805, and by the Ministry of Education, Youth, and Sports of the Czech Republic, contract OC115.

  19. APPLICATION OF PROTEOMICS TO NEUTROPHIL BIOLOGY

    PubMed Central

    Luerman, Gregory C.; Uriarte, Silvia M.; Rane, Madhavi J.; McLeish, Kenneth R.

    2009-01-01

    Polymorphonuclear leukocytes or neutrophils are a primary effector cell of the innate immune system and contribute to the development of adaptive immunity. Neutrophils participate in both the initiation and resolution of inflammatory responses through a series of highly coordinated molecular and phenotypic changes. To accomplish these changes, neutrophils express numerous receptors and use multiple overlapping and redundant signal transduction pathways. Dysregulation of the activation or resolution pathways plays a role in a number of human diseases. A comprehensive understanding of the regulation of neutrophil responses can be provided by high throughput proteomic technologies and sophisticated computational analysis. The first steps in the application of proteomics to understanding neutrophil biology have been taken. Here we review the application of expression, structural, and functional proteomic studies to neutrophils. Although defining the complex molecular events associated with neutrophil activation is in the early stages, the data generated to date suggest that proteomic technologies will dramatically enhance our understanding of neutrophil biology. PMID:19580889

  20. Lipopolysaccharide-Induced Neutrophil Dysfunction Following Transjugular Intrahepatic Portosystemic Stent Shunt (TIPSS) Insertion is Associated with Organ Failure and Mortality

    PubMed Central

    Macnaughtan, Jane; Mookerjee, Rajeshwar P.; Jalan, Rajiv

    2017-01-01

    Systemic lipopolysaccharide (LPS) is implicated in increasing mortality in patients with alcoholic hepatitis but the underlying mechanisms are not well characterised. The objective of this study was to characterise neutrophil function, LPS and cytokine concentrations within the splanchnic circulation of alcoholic cirrhotic patients undergoing TIPSS insertion for variceal haemorrhage and correlate this with outcome. 26 patients with alcoholic cirrhosis and variceal haemorrhage were studied prior to and 1-hour after TIPSS insertion. Neutrophil function, LPS and cytokine concentrations were determined in arterial, hepatic venous (HV) and portal venous blood (PV). Significantly higher LPS concentrations and neutrophil reactive oxidant species (ROS) production were observed in PV vs HV blood. Cross-incubation of HV plasma with PV neutrophils resulted in reduced ROS production. Insertion of TIPSS was associated with a significant increase in arterial LPS concentrations and deterioration in neutrophil phagocytosis. Number of organ failures and arterial IL-6 concentrations at presentation were associated with increased mortality. The portal circulation has a distinct immunological milieu characterised by a pathological neutrophil phenotype and an anti-inflammatory cytokine profile associated with heightened LPS levels. TIPSS insertion renders this neutrophil functional defect systemic, associated with an increase in arterial LPS and a susceptibility to sepsis. PMID:28051160

  1. The Circulation Analysis of Serial Use: Numbers Game or Key to Service?

    PubMed Central

    Raisig, L. Miles

    1967-01-01

    The conventionally erected and reported circulation analysis of serial use in the individual and the feeder library is found to be statistically invalid and misleading, since it measures neither the intellectual use of the serial's contents nor the physical handlings of serial units, and is nonrepresentative of the in-depth library use of serials. It fails utterly to report or even to suggest the relation of intralibrary and interlibrary serial resources. The actual mechanics of the serial use analysis, and the active variables in the library situation which affect serial use, are demonstrated in a simulated analysis and are explained at length. A positive design is offered for the objective gathering and reporting of data on the local intellectual use and physical handling of serials and the relating of resources. Data gathering in the feeder library, and implications for the extension of the feeder library's resources, are discussed. PMID:6055863

  2. Heterogeneity of the Mac-1 expression on peripheral blood neutrophils in patients with different types of epithelial ovarian cancer.

    PubMed

    Bednarska, Katarzyna; Klink, Magdalena; Wilczyński, Jacek R; Szyłło, Krzysztof; Malinowski, Andrzej; Sułowska, Zofia; Nowak, Marek

    2016-02-01

    The expression level of Mac-1 on the surface of neutrophils is an important indicator of neutrophil activation. Under pathological conditions, Mac-1 is believed a key adhesion molecule that facilitates cancer progression and mediates the adhesion of tumour cells to the endothelium of blood vessels. Our previous findings indicated that circulating peripheral blood neutrophils in patients with advanced epithelial ovarian cancer (EOC) expressed enhanced levels of Mac-1, which was functionally associated with an increased adhesive function of neutrophils. The objective of the current study was to analyse whether the value of individual components of the differential white cell count, including the neutrophil and lymphocyte ratios, which are markers of blood neutrophil activation, might be associated with certain types of ovarian cancer. We showed the increase in Mac-1 expression along with a parallel decrease of L-selectin and PSGL-1 on peripheral blood neutrophils of patients with EOC of early and advanced FIGO stages, which indicates an activated state of neutrophils in comparison to neutrophils of individuals without cancer. Despite a significant difference between Mac-1 expression in patients with and without cancer, a dramatic increase in Mac-1 expression was observed in the blood of patients with undifferentiated carcinomas compared with patients with other histological types of EOC. Moreover, the expression level of Mac-1 correlated with the number of neutrophils in patients with serous, endometrioid and undifferentiated EOC. The results of an ROC analysis demonstrated that the patients with the undifferentiated type of EOC form a distinct group with regard to Mac-1 expression on blood neutrophils. The results suggested a diverse biological cadre of immune cells in patients with undifferentiated ovarian carcinomas compared with patients with other histological types of EOC.

  3. A Bayesian Approach for Reconstructing the Past Ocean Circulation from a Limited Number of Sediment-Core Radiocarbon Measurements

    NASA Astrophysics Data System (ADS)

    Primeau, F.

    2014-12-01

    Paleoceanographers are faced with the problem of making inferences about the ventilation of the ocean in the past from localized benthic and planktonic radiocarbon measurements obtained from a small number of sediment cores, which leads to an underdetermined problem. With the goal of moving beyond testing the null hypothesis that the sediment core data are consistent with the modern circulation we seek to reconstruct the most probable paleocirculation based on our knowledge of ocean dynamics and available constraints from sediment-core radiocarbon records. We propose a Bayesian inversion approach in which we use a modern circulation estimate constrained by modern radiocarbon data to define the mean of the prior probability distribution for the unknown paleocirculation. The approach resolves the indeterminacy of the inverse problem by choosing a paleocirculation that is minimally different from the modern circulation while still being consistent with the available sediment-core radiocarbon records. In this talk we will present the general formulation of the method as well as various approximations to reduce the computational challenge.

  4. A chemotactic gradient sequestered on endothelial heparan sulfate induces directional intraluminal crawling of neutrophils.

    PubMed

    Massena, Sara; Christoffersson, Gustaf; Hjertström, Elina; Zcharia, Eyal; Vlodavsky, Israel; Ausmees, Nora; Rolny, Charlotte; Li, Jin-Ping; Phillipson, Mia

    2010-09-16

    During infection, chemokines sequestered on endothelium induce recruitment of circulating leukocytes into the tissue where they chemotax along chemokine gradients toward the afflicted site. The aim of this in vivo study was to determine whether a chemokine gradient was formed intravascularly and influenced intraluminal neutrophil crawling and transmigration. A chemokine gradient was induced by placing a macrophage inflammatory protein-2 (MIP-2)-containing (CXCL2) gel on the cremaster muscle of anesthetized wild-type mice or heparanase-overexpressing transgenic mice (hpa-tg) with truncated heparan sulfate (HS) side chains. Neutrophil-endothelial interactions were visualized by intravital microscopy and chemokine gradients detected by confocal microscopy. Localized extravascular chemokine release (MIP-2 gel) induced directed neutrophil crawling along a chemotactic gradient immobilized on the endothelium and accelerated their recruitment into the target tissue compared with homogeneous extravascular chemokine concentration (MIP-2 superfusion). Endothelial chemokine sequestration occurred exclusively in venules and was HS-dependent, and neutrophils in hpa-tg mice exhibited random crawling. Despite similar numbers of adherent neutrophils in hpa-tg and wild-type mice, the altered crawling in hpa-tg mice was translated into decreased number of emigrated neutrophils and ultimately decreased the ability to clear bacterial infections. In conclusion, an intravascular chemokine gradient sequestered by endothelial HS effectively directs crawling leukocytes toward transmigration loci close to the infection site.

  5. Circulating histamine and neutrophil chemotactic activity during allergen-induced asthma: the effect of inhaled antihistamines and anti-allergic compounds.

    PubMed

    Morgan, D J; Moodley, I; Cundell, D R; Sheinman, B D; Smart, W; Davies, R J

    1985-07-01

    Plasma histamine and serum neutrophil chemotactic activity (S-NCA) were measured in ten atopic asthmatic patients on four separate occasions after allergen bronchial provocation testing (BPT). Single doses of inhaled sodium cromoglycate (SCG; 20 mg), clemastine (0.5 mg), ketotifen (0.5 mg) and isotonic saline (0.9% NaCl) placebo were administered 30 min before bronchial provocation testing in random order and double-blind. The airflow obstruction after BPT was monitored by measurement of forced expiratory volume in 1 s (FEV1). Plasma histamine was measured by the double-isotope radioenzymatic assay and S-NCA by a modified Boyden chamber technique. A highly significant decrease in FEV1 after BPT occurred on the placebo pre-treatment visit (P less than 0.001). Prior administration of inhaled SCG, clemastine and ketotifen significantly reduced the decrease in airflow obstruction seen after BPT when compared with placebo treatment (P less than 0.01, P less than 0.02, P less than 0.05 respectively). No significant alteration in plasma histamine was detected during allergen-induced airflow obstruction. Levels of S-NCA were significantly higher 5, 10 and 15 min after BPT when compared with the pre-challenge level (P less than 0.01, P less than 0.01, P less than 0.001 respectively). These levels were not significantly decreased when airflow obstruction was inhibited by the prior inhalation of SCG, clemastine or ketotifen.

  6. The effect of cigarette smoking on neutrophil kinetics in human lungs (see comments

    SciTech Connect

    MacNee, W.; Wiggs, B.; Belzberg, A.S.; Hogg, J.C. )

    1989-10-05

    Neutrophils may play a part in the pathogenesis of the centrilobular emphysema associated with cigarette smoking. The capillary bed of the lungs concentrates neutrophils approximately 100-fold with respect to erythrocytes, producing a large pool of marginated cells. We examined the effect of cigarette smoking on the kinetics of this pool of cells, using 99mTc-labeled erythrocytes to measure regional blood velocity and 111In-labeled neutrophils to measure the removal of neutrophils during the first passage through the pulmonary circulation, their subsequent washout from the lungs, and the effect of local blood velocity on the number of neutrophils retained in each lung region. We observed no difference in these measurements between subjects who had never smoked (n = 6) and smokers who did not smoke during the study (n = 12). However, subjects who did smoke during the study (n = 12) had a significantly slower rate of washout of radiolabeled neutrophils from the lung (0.08 +/- 0.04 of the total per minute, as compared with 0.13 +/- 0.06 in smokers who did not smoke during the experiment and 0.14 +/- 0.08 in non-smokers) (P = 0.02). We also observed an increase in the regional retention of labeled neutrophils with respect to blood velocity in 5 of the 12 subjects who smoked during the study, but in none of the other subjects. We conclude that the presence of cigarette smoke in the lungs of some subjects increases the local concentration of neutrophils, and suggest that the lesions that characterize emphysema may be a result of the destruction of lung tissue by neutrophils that remain within pulmonary microvessels.

  7. Neutrophil Functions in Periodontal Homeostasis.

    PubMed

    Cortés-Vieyra, Ricarda; Rosales, Carlos; Uribe-Querol, Eileen

    2016-01-01

    Oral tissues are constantly exposed to damage from the mechanical effort of eating and to microorganisms, mostly bacteria. In healthy gingiva tissue remodeling and a balance between bacteria and innate immune cells are maintained. However, excess of bacteria biofilm (plaque) creates an inflammation state that recruits more immune cells, mainly neutrophils to the gingiva. Neutrophils create a barrier for bacteria to reach inside tissues. When neutrophils are insufficient, bacteria thrive causing more inflammation that has been associated with systemic effects on other conditions such as atherosclerosis, diabetes, and cancer. But paradoxically when neutrophils persist, they can also promote a chronic inflammatory state that leads to periodontitis, a condition that leads to damage of the bone-supporting tissues. In periodontitis, bone loss is a serious complication. How a neutrophil balance is needed for maintaining healthy oral tissues is the focus of this review. We present recent evidence on how alterations in neutrophil number and function can lead to inflammatory bone loss, and how some oral bacteria signal neutrophils to block their antimicrobial functions and promote an inflammatory state. Also, based on this new information, novel therapeutic approaches are discussed.

  8. Neutrophil Functions in Periodontal Homeostasis

    PubMed Central

    Cortés-Vieyra, Ricarda; Rosales, Carlos

    2016-01-01

    Oral tissues are constantly exposed to damage from the mechanical effort of eating and to microorganisms, mostly bacteria. In healthy gingiva tissue remodeling and a balance between bacteria and innate immune cells are maintained. However, excess of bacteria biofilm (plaque) creates an inflammation state that recruits more immune cells, mainly neutrophils to the gingiva. Neutrophils create a barrier for bacteria to reach inside tissues. When neutrophils are insufficient, bacteria thrive causing more inflammation that has been associated with systemic effects on other conditions such as atherosclerosis, diabetes, and cancer. But paradoxically when neutrophils persist, they can also promote a chronic inflammatory state that leads to periodontitis, a condition that leads to damage of the bone-supporting tissues. In periodontitis, bone loss is a serious complication. How a neutrophil balance is needed for maintaining healthy oral tissues is the focus of this review. We present recent evidence on how alterations in neutrophil number and function can lead to inflammatory bone loss, and how some oral bacteria signal neutrophils to block their antimicrobial functions and promote an inflammatory state. Also, based on this new information, novel therapeutic approaches are discussed. PMID:27019855

  9. Effects of endogenous and exogenous catecholamines on LPS-induced neutrophil trafficking and activation.

    PubMed

    Abraham, E; Kaneko, D J; Shenkar, R

    1999-01-01

    Endotoxemia produces elevations in catecholamine levels in the pulmonary and systemic circulation as well as rapid increases in neutrophil number and proinflammatory cytokine expression in the lungs. In the present experiments, we examined the effects of endogenous and exogenous adrenergic stimulation on endotoxin-induced lung neutrophil accumulation and activation. Levels of interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, and macrophage inflammatory protein (MIP)-2 mRNAs were increased in lung neutrophils from endotoxemic mice compared with those present in lung neutrophils from control mice or in peripheral blood neutrophils from endotoxemic or control mice. Treatment with the beta-adrenergic antagonist propranolol before endotoxin administration did not affect trafficking of neutrophils to the lungs or the expression of IL-1beta, TNF-alpha, or MIP-2 by lung neutrophils. Administration of the alpha-adrenergic antagonist phentolamine before endotoxemia did not alter lung neutrophil accumulation as measured by myeloperoxidase (MPO) levels but did result in significant increases in IL-1beta, TNF-alpha, and MIP-2 mRNA expression by lung neutrophils compared with endotoxemia alone. Administration of the alpha1-adrenergic agonist phenylephrine before endotoxin did not affect trafficking of neutrophils to the lungs but was associated with significantly increased expression of TNF-alpha and MIP-2 mRNAs by lung neutrophils compared with that found after endotoxin alone. In contrast, treatment with the alpha2-adrenergic agonist UK-14304 prevented endotoxin-induced increases in lung MPO and lung neutrophil cytokine mRNA levels. The suppressive effects of UK-14304 on endotoxin-induced increases in lung MPO were not affected by administration of the nitric oxide synthase inhibitor N-nitro-L-arginine methyl ester. These data demonstrate that the initial accumulation and activation of neutrophils in the lungs after endotoxemia can be significantly diminished by alpha

  10. Lung vaso-occlusion in sickle cell disease mediated by arteriolar neutrophil-platelet microemboli

    PubMed Central

    Bennewitz, Margaret F.; Jimenez, Maritza A.; Vats, Ravi; Tutuncuoglu, Egemen; Jonassaint, Jude; Kato, Gregory J.; Gladwin, Mark T.

    2017-01-01

    In patients with sickle cell disease (SCD), the polymerization of intraerythrocytic hemoglobin S promotes downstream vaso-occlusive events in the microvasculature. While vaso-occlusion is known to occur in the lung, often in the context of systemic vaso-occlusive crisis and the acute chest syndrome, the pathophysiological mechanisms that incite lung injury are unknown. We used intravital microscopy of the lung in transgenic humanized SCD mice to monitor acute vaso-occlusive events following an acute dose of systemic lipopolysaccharide sufficient to trigger events in SCD but not control mice. We observed cellular microembolism of precapillary pulmonary arteriolar bottlenecks by neutrophil-platelet aggregates. Blood from SCD patients was next studied under flow in an in vitro microfluidic system. Similar to the pulmonary circulation, circulating platelets nucleated around arrested neutrophils, translating to a greater number and duration of neutrophil-platelet interactions compared with normal human blood. Inhibition of platelet P-selectin with function-blocking antibody attenuated the neutrophil-platelet interactions in SCD patient blood in vitro and resolved pulmonary arteriole microembolism in SCD mice in vivo. These results establish the relevance of neutrophil-platelet aggregate formation in lung arterioles in promoting lung vaso-occlusion in SCD and highlight the therapeutic potential of targeting platelet adhesion molecules to prevent acute chest syndrome. PMID:28097236

  11. Change in number and size of circulating tumor cells with high telomerase activity during treatment of patients with gastric cancer

    PubMed Central

    Ito, Hiroaki; Yamaguchi, Noriko; Onimaru, Manabu; Kimura, Satoshi; Ohmori, Tohru; Ishikawa, Fumihiro; Sato, Jun; Ito, Shun; Inoue, Haruhiro

    2016-01-01

    Detection of circulating tumor cells (CTCs) in peripheral blood is useful for estimating the prognosis of patients with cancer. We previously reported the detection of CTCs by OBP-401, a telomerase-specific, replication-selective, oncolytic adenoviral agent carrying the green fluorescent protein (GFP) gene. We demonstrated that the number of large (L)-GFP+ cells (≥7.735 µm in diameter) in peripheral blood samples correlated significantly with the prognosis of treatment-naïve gastric cancer patients, whereas the number of small (S)-GFP+ cells (<7.735 µm in diameter) did not. In the present study, we studied the change in the number of GFP+ cells during treatment, and analyzed the association between the number of GFP+ cells in blood samples and the outcome of patients. Peripheral blood samples were obtained from 37 gastric patients prior and subsequent to surgery (three samples per time point). Upon infection of blood cells with OBP-401, GFP+ cells of different sizes were counted and measured. The association between the number of GFP+ cells and surgical outcome was determined by statistical analysis. The median follow-up period after surgery was 39 months. Although the difference was not significant, patients with ≥6 L-GFP+ cells in preoperative blood samples had a lower relapse-free survival rate than patients with 0–5 L-GFP+ cells. There was no significant correlation between the number of L-GFP+ cells in postoperative blood samples and the prognosis of patients receiving adjuvant therapy. Although the difference was not significant, the number of S-GFP+ cells in samples from patients who had received postoperative chemotherapy was higher than in those who had not. The number of L-GFP+ cells was not significantly correlated with the relapse-free survival rate in gastric cancer patients who underwent surgery. The number of S-GFP+ cells was relatively high in samples from patients who had received postoperative chemotherapy. PMID:28105179

  12. Influence of minor thermal injury on expression of complement receptor CR3 on human neutrophils.

    PubMed Central

    Nelson, R. D.; Hasslen, S. R.; Ahrenholz, D. H.; Haus, E.; Solem, L. D.

    1986-01-01

    Thermal injury is well known to inhibit functions of the circulating neutrophil related to its role in host defense against infection, but the mechanism(s) of this phenomenon are not fully understood. To gain further clues to these mechanisms, the authors have studied patients with thermal injury in terms of altered expression of neutrophil cell membrane receptors for the opsonic complement-derived ligand C3bi--complement receptor Type 3, or CR3. CR3 expression was selected for study because an increase in the number of receptors on the cell surface can be stimulated by products of complement activation known to accumulate after thermal injury and because of the role of CR3 in phagocytic and adherence functions of the neutrophil. Expression of CR3 was monitored semiquantitatively by flow cytometry with the use of a murine monoclonal antibody (OKM1) specific for an antigen (CD11) associated with this receptor. Patients evaluated were limited in this study to those with minor degrees of thermal injury (second-degree burn involving less than 20% of total body surface area) so that possible confounding effects of major injury and its complications could be eliminated. It was observed that patient neutrophil CR3 becomes significantly up-regulated during the first week, as early as 1 day after injury. The maximum level of expression of CR3 averaged greater than 150% (range, 70-314%) of the respective minimum level observed for each patient. The minimum levels of expression of CR3 on patient neutrophils, reached 11-37 days after injury for 7 of 8 patients, were comparable to the level of expression of CR3 on unstimulated control neutrophils. Such temporal up-regulation of patient neutrophil CR3 suggests the early generation of stimuli of CR3 mobilization in response to thermal injury. Increased numbers of CR3 on patient neutrophils may augment microbicidal function and enhance or inhibit delivery of cells to the burn site. PMID:3541642

  13. Ligation of the adhesion-GPCR EMR2 regulates human neutrophil function.

    PubMed

    Yona, Simon; Lin, Hsi-Hsien; Dri, Pietro; Davies, John Q; Hayhoe, Richard P G; Lewis, Sion M; Heinsbroek, Sigrid E M; Brown, K Alun; Perretti, Mauro; Hamann, Jörg; Treacher, David F; Gordon, Siamon; Stacey, Martin

    2008-03-01

    At present, approximately 150 different members of the adhesion-G protein-coupled receptor (GPCR) family have been identified in metazoans. Surprisingly, very little is known about their function, although they all possess large extracellular domains coupled to a seven-transmembrane domain, suggesting a potential role in cell adhesion and signaling. Here, we demonstrate how the human-restricted adhesion-GPCR, EMR2 (epidermal growth factor-like module-containing mucin-like hormone receptor), regulates neutrophil responses by potentiating the effects of a number of proinflammatory mediators and show that the transmembrane region is critical for adhesion-GPCR function. Using an anti-EMR2 antibody, ligation of EMR2 increases neutrophil adhesion and migration, and augments superoxide production and proteolytic enzyme degranulation. On neutrophil activation, EMR2 is rapidly translocated to membrane ruffles and the leading edge of the cell. Further supporting the role in neutrophil activation, EMR2 expression on circulating neutrophils is significantly increased in patients with systemic inflammation. These data illustrate a definitive function for a human adhesion-GPCR within the innate immune system and suggest an important role in potentiating the inflammatory response. Ligation of the adhesion-GPCR EMR2 regulates human neutrophil function.

  14. Circulating megakaryocytes: delivery of large numbers of intact, mature megakaryocytes to the lungs.

    PubMed

    Levine, R F; Eldor, A; Shoff, P K; Kirwin, S; Tenza, D; Cramer, E M

    1993-10-01

    To determine the locus of platelet production, we sought to determine if sufficient megakaryocytes reach the lungs in a state that could produce platelets. Elutriation was used to isolate megakaryocytes from blood reaching and leaving the lungs of 20 patients undergoing routine cardiac catheterizations. A mean of 5.0 intact megakaryocytes/ml were found in pulmonary artery blood, compared to only 0.5 megakaryocytes/ml, with partial cytoplasmic content, in aortic samples. The megakaryocytes in central venous and aortic samples were all mature. The identity of these cells as megakaryocytes, their maturity and normal morphology were confirmed by standard and immunoelectron microscopy. Cardiac outputs were obtained for each patient at the time of blood sampling, allowing an extrapolation that 40 x 10(6) intact, mature megakaryocytes were being delivered to the lungs every day in the average patient, compared to only 4.0 x 10(6) partially spent megakaryocytes exiting the lungs daily. About 98% of megakaryocyte cytoplasm reaching the lungs did not exit as recognizable megakaryocytes or fragments. The number and state of the megakaryocytes apparently filtered in the lungs is consistent with the hypothesis that megakaryocytes may shed platelets within the pulmonary microvasculature, which may be the primary site of platelet production.

  15. Circulating AR copy number and outcome to enzalutamide in docetaxel-treated metastatic castration-resistant prostate cancer.

    PubMed

    Salvi, Samanta; Casadio, Valentina; Conteduca, Vincenza; Lolli, Cristian; Gurioli, Giorgia; Martignano, Filippo; Schepisi, Giuseppe; Testoni, Sara; Scarpi, Emanuela; Amadori, Dino; Calistri, Daniele; Attard, Gerhardt; De Giorgi, Ugo

    2016-06-21

    In the present study, we aimed to evaluate the association of circulating AR copy number (CN) and outcome in a cohort of patients with advanced castration-resistant prostate cancer (CRPC) treated with enzalutamide after docetaxel. Fifty-nine CRPC patients were evaluated. AR CN was analyzed with real-time and digital PCR in the serum collected at starting of treatment. Progressive disease was defined on the basis of Prostate Cancer Working Group 2 criteria. AR CN gain was found in 21 of 59 (36%) patients. Median baseline PSA, alkaline phosphatase and lactate dehydrogenase levels were higher in the AR CN gained group (p = 0.007, p = 0.003, p = 0.0009, respectively). Median PFS of patients with AR CN gain was 2.4 (95%CI: 1.9-3.2) vs. 4.0 months (95%CI: 3.0-6.5) of those with no gain (p = 0.0004). Median OS of patients with AR CN gain was 6.1 (95%CI: 3.4-8.6) vs. 14.1 months (95%CI: 8.2-20.5) of those with no gain (p = 0.0003). At multivariate analysis, PSA decline ≥ 50% and AR CN showed a significant association with PFS (p = 0.008 and p = 0.002, respectively) and OS (p = 0.009 and p = 0.001, respectively). These findings indicate that the detection of circulating AR CN gain is a promising non-invasive biomarker for outcome prediction to enzalutamide treatment in CRPC patients.

  16. Achyrocline satureioides (Lam.) D.C. Hydroalcoholic Extract Inhibits Neutrophil Functions Related to Innate Host Defense

    PubMed Central

    Barioni, Eric Diego; Machado, Isabel Daufenback; Rodrigues, Stephen Fernandes de Paula; Ferraz-de-Paula, Viviane; Wagner, Theodoro Marcel; Cogliati, Bruno; Corrêa dos Santos, Matheus; Machado, Marina da Silva; de Andrade, Sérgio Faloni; Niero, Rivaldo; Farsky, Sandra Helena Poliselli

    2013-01-01

    Achyrocline satureioides (Lam.) D.C. is a herb native to South America, and its inflorescences are popularly employed to treat inflammatory diseases. Here, the effects of the in vivo actions of the hydroalcoholic extract obtained from inflorescences of A. satureioides on neutrophil trafficking into inflamed tissue were investigated. Male Wistar rats were orally treated with A. satureioides extract, and inflammation was induced one hour later by lipopolysaccharide injection into the subcutaneous tissue. The number of leukocytes and the amount of chemotactic mediators were quantified in the inflammatory exudate, and adhesion molecule and toll-like receptor 4 (TLR-4) expressions and phorbol-myristate-acetate- (PMA-) stimulated oxidative burst were quantified in circulating neutrophils. Leukocyte-endothelial interactions were quantified in the mesentery tissue. Enzymes and tissue morphology of the liver and kidney were evaluated. Treatment with A. satureioides extract reduced neutrophil influx and secretion of leukotriene B4 and CINC-1 in the exudates, the number of rolling and adhered leukocytes in the mesentery postcapillary venules, neutrophil L-selectin, β2-integrin and TLR-4 expression, and oxidative burst, but did not cause an alteration in the morphology and activities of liver and kidney. Together, the data show that A. satureioides extract inhibits neutrophil functions related to the innate response and does not cause systemic toxicity. PMID:23476704

  17. Isolation and Characterization of Neutrophils with Anti-Tumor Properties.

    PubMed

    Sionov, Ronit Vogt; Assi, Simaan; Gershkovitz, Maya; Sagiv, Jitka Y; Polyansky, Lola; Mishalian, Inbal; Fridlender, Zvi G; Granot, Zvi

    2015-06-19

    Neutrophils, the most abundant of all white blood cells in the human circulation, play an important role in the host defense against invading microorganisms. In addition, neutrophils play a central role in the immune surveillance of tumor cells. They have the ability to recognize tumor cells and induce tumor cell death either through a cell contact-dependent mechanism involving hydrogen peroxide or through antibody-dependent cell-mediated cytotoxicity (ADCC). Neutrophils with anti-tumor activity can be isolated from peripheral blood of cancer patients and of tumor-bearing mice. These neutrophils are termed tumor-entrained neutrophils (TEN) to distinguish them from neutrophils of healthy subjects or naïve mice that show no significant tumor cytotoxic activity. Compared with other white blood cells, neutrophils show different buoyancy making it feasible to obtain a > 98% pure neutrophil population when subjected to a density gradient. However, in addition to the normal high-density neutrophil population (HDN), in cancer patients, in tumor-bearing mice, as well as under chronic inflammatory conditions, distinct low-density neutrophil populations (LDN) appear in the circulation. LDN co-purify with the mononuclear fraction and can be separated from mononuclear cells using either positive or negative selection strategies. Once the purity of the isolated neutrophils is determined by flow cytometry, they can be used for in vitro and in vivo functional assays. We describe techniques for monitoring the anti-tumor activity of neutrophils, their ability to migrate and to produce reactive oxygen species, as well as monitoring their phagocytic capacity ex vivo. We further describe techniques to label the neutrophils for in vivo tracking, and to determine their anti-metastatic capacity in vivo. All these techniques are essential for understanding how to obtain and characterize neutrophils with anti-tumor function.

  18. Neutrophil-platelet adhesion: relative roles of platelet P-selectin and neutrophil beta2 (DC18) integrins.

    PubMed

    Brown, K K; Henson, P M; Maclouf, J; Moyle, M; Ely, J A; Worthen, G S

    1998-01-01

    Neutrophils and platelets interact both physically and metabolically during inflammation and thrombosis, but the mechanisms responsible for their adhesion remain incompletely understood. Neutrophil-platelet adhesion was measured after specific stimulation of neutrophils, platelets, or both and quantified by flow cytometry. Specific stimulation of either the neutrophil or the platelet led to a marked increase in the percentage of neutrophils that bound platelets, although platelet stimulation led to a large increase and neutrophil stimulation to only a small increase in the number of platelets per neutrophil. Stimulation of both cells further increased the number of neutrophil-platelet adhesive events and led to large numbers of platelets binding to each neutrophil. Confirming previous observations, blocking antibodies to platelet P-selectin (CD62P) partially inhibited adhesion. However, blockade of the neutrophil beta2 integrin CD11b/CD18 also inhibited the percentage of neutrophils that bound platelets. Combining P-selectin and CD11b/18 blockade further inhibited the stimulated increase in the percentage of neutrophils binding platelets and the increased number of platelets per neutrophil. Both cell adhesion molecules were active even when only a single cell type was primarily activated, supporting physiologically important transcellular activation. These data suggest that: (1) neutrophil-platelet adhesion can be initiated by specific activation of either the neutrophil or the platelet and that specific activation of either cell type leads to distinct patterns of adhesion, and (2) neutrophil-platelet adhesion uses both platelet P-selectin and the neutrophil beta2 integrin CD11b/CD18 when the cells are primarily or secondarily activated.

  19. Two neutrophilic dermatoses captured simultaneously on histology

    PubMed Central

    Wlodek, Christina; Bhatt, Nidhi; Kennedy, Cameron

    2016-01-01

    A number of neutrophilic dermatoses are associated with malignancies and their treatment. These rarely occur together in the same patient. A Caucasian 72-year-old male was treated for acute myeloid leukemia (AML) with chemotherapy including daunorubicin and cytarabine. Within 48 hours of commencing treatment, he developed pyrexia and, two days later, disseminated non-tender pink plaques on the limbs and trunk. A skin biopsy showed a dermal interstitial infiltrate of lymphocytes, histiocytoid cells and predominantly neutrophils. This extended into the subcutis, where a neutrophilic lobular panniculitis was seen. These findings are consistent with Sweet’s syndrome. In addition, a neutrophilic and lymphocytic infiltrate was also present around eccrine coils and lower ducts. The eccrine epithelium showed squamous metaplasia with dyskeratosis and sloughing into the lumen. These latter findings are consistent with neutrophilic eccrine hidradenitis (NEH). These two histologically distinct entities form part of the neutrophilic dermatoses that have been described in oncology patients with reports of concurrent or sequential occurrence of various neutrophilic dermatoses in the same patient. Ours, however, is only the second reported case of simultaneously captured Sweet’s and NEH in the setting of AML. The most likely explanation is that of an epiphenomenon, whereby the neutrophilic infiltrate extended around the sweat glands in the context of the neutrophilic dermatosis. PMID:27648385

  20. The hydrodynamic and ultrasound-induced forces on microbubbles under high Reynolds number flow representative of the human systemic circulation

    NASA Astrophysics Data System (ADS)

    Clark, Alicia; Aliseda, Alberto

    2016-11-01

    Ultrasound contrast agents (UCAs) are micron-sized bubbles that are used in conjunction with ultrasound (US) in medical applications such as thrombolysis and targeted intravenous drug delivery. Previous work has shown that the Bjerknes force, due to the phase difference between the incoming US pressure wave and the bubble volume oscillations, can be used to manipulate the trajectories of microbubbles. Our work explores the behavior of microbubbles in medium sized blood vessels under both uniform and pulsatile flows at a range of physiologically relevant Reynolds and Womersley numbers. High speed images were taken of the microbubbles in an in-vitro flow loop that replicates physiological flow conditions. During the imaging, the microbubbles were insonified at different diagnostic ultrasound settings (varying center frequency, PRF, etc.). An in-house Lagrangian particle tracking code was then used to determine the trajectories of the microbubbles and, thus, a dynamic model for the microbubbles including the Bjerknes forces acting on them, as well as drag, lift, and added mass. Preliminary work has also explored the behavior of the microbubbles in a patient-specific model of a carotid artery bifurcation to demonstrate the feasibility of preferential steering of microbubbles towards the intracranial circulation with US.

  1. Down-regulated resistin level in consequence of decreased neutrophil counts in untreated Grave's disease

    PubMed Central

    Huang, Fengjiao; Chen, Xinxin; Zhou, Yulin; Ye, Lei; Wang, Weiqing; Ning, Guang; Wang, Shu

    2016-01-01

    Resistin, belongs to cysteine-rich secretory protein, is mainly produced by circulating leukocytes, such as neutrophils monocytes and macrophages in humans. To date, few but controversial studies have reported about resistin concentrations in hyperthyroid patients, especially in Graves' disease (GD). We undertaked a controlled, prospective study to explore the serum resistin concentration in GD patients before and after -MMI treatment. In addition, we also investigated the main influencing factor on serum resistin level and discuessed the potential role of serum resistin plays in GD patients. 39 untreated GD (uGD) patients, including 8 males and 31 females, were enrolled in our investigation. All of these patients were prescribed with MMI treatment, in addition to 25 healthy controls. Anthropometric parameters and hormone assessment were measured. Enzyme-linked immunosorbent assay was used to detect serum resistin concentration in different stages of GD patients. Furthermore, neutrophil cell line NB4 with or without T3 treatment to detect the effect of thyroid hormones on resistin expression. The serum resistin level and neutrophil counts in untreated GD patients were significantly declined. And all of these parameters were recovered to normal after MMI treatment in ethyroid GD (eGD) and TRAb-negative conversion (nGD) patients. Resistin concentration exhibited a negative correlation with FT3 and FT4, but a positive correlation with absolute number of neutrophiles in uGD patients, whereas did not correlate with thyroid autoimmune antibodies and BMI. Neutrophile cell line, NB4, produced decreased expression of resistin when stimulated with T3. Our study showed a decrease of serum resistin level in GD patients and we suggested that the serum resistin might primarily secreted from circulating neutrophils and down-regulated by excessive thyroid hormones in GD patients. PMID:27637079

  2. [Ultrastructural changes of neutrophilic granulocytes in dilated cardiomyopathy and their dynamics after blood irradiation with Helium-Neon laser in vitro].

    PubMed

    Khomeriki, S G; Morozov, I A

    1998-01-01

    Venous blood from 10 patients with dilated cardiomyopathy was irradiated with a laser in vitro. The control group consisted of 20 healthy donors. The neutrophil granulocytes were separated at gradient centrifugation. Alterations of neutrophils manifested with an increase of specific cytoplasmic granules number, thickening of submembrane actin, cell configuration changes with a relative increase of their surface. Laser irradiation of the blood resulted in destruction of the altered (less resistant) cells while morphometric parameters of the remaining cells approaches those of donor cells. Thus, low-intensity laser irradiation results in the renewal of the neutrophil population in patients with dilated cardiomyopathy and normalization of structural-functional changes in the circulating neutrophil population.

  3. The use of spent renal dialysis membranes for the isolation of large numbers of human neutrophils for biochemical studies. Application to purification of the myeloid IgA receptor (Fc alpha R).

    PubMed

    Mazengera, R L; Kerr, M A; Todd, A S

    1992-01-21

    Human neutrophils (PMN) can be eluted from spent Cuprophan renal dialysis membranes in large numbers (10(9)-10(10) per dialyser cartridge) and in relatively high purity by washing the membranes with 0.35 M NaCl. This offers the possibility of isolating relatively large amounts (10(-4)-10(-3) g) of minor PMN proteins such as those expressed on the cell surface. Here the technique is applied to the purification of the neutrophil IgA receptor (Fc alpha R). Affinity chromatography on IgA-Sepharose of NP-40 extracts of 125I-labelled PMN isolated from fresh venous blood routinely gave a receptor preparation showing one diffuse band, Mr 50-70 kDa, upon analysis by SDS-PAGE and autoradiography. When the same method was used with larger numbers of unlabelled PMN from fresh venous blood or renal dialysis membranes a preparation was obtained which gave multiple bands upon analysis by SDS-PAGE silver stained gels due to contamination of the receptor with cytoplasmic proteins which bound non-specifically to the IgA-Sepharose. Most of these contaminants could be removed by chromatography of the IgA-Sepharose eluates on wheat germ agglutinin-Sepharose.

  4. Neutrophils: important contributors to tumor progression and metastasis.

    PubMed

    Swierczak, Agnieszka; Mouchemore, Kellie A; Hamilton, John A; Anderson, Robin L

    2015-12-01

    The presence of neutrophils in tumors has traditionally been considered to be indicative of a failed immune response against cancers. However, there is now evidence showing that neutrophils can promote tumor growth, and increasingly, the data support an active role for neutrophils in tumor progression to distant metastasis. Neutrophils have been implicated in promoting metastasis in cancer patients, where neutrophil numbers and neutrophil-related factors and functions have been associated with progressive disease. Nevertheless, the role of neutrophils in tumors, both at the primary and secondary sites, remains controversial, with some studies reporting their anti-tumor functions. This review will focus on the data demonstrating a role for neutrophils in both tumor growth and metastasis and will attempt to clarify the discrepancies in the literature.

  5. Neutrophils: critical components in experimental animal models of cancer

    PubMed Central

    Hagerling, Catharina; Werb, Zena

    2016-01-01

    Neutrophils have a crucial role in tumor development and metastatic progression. The contribution of neutrophils in tumor development is multifaceted and contradictory. On the one hand, neutrophils prompt tumor inception, promote tumor development by mediating the initial angiogenic switch and facilitate colonization of circulating tumor cells, and on the other hand, have cytotoxic and anti-metastatic capabilities. Our understanding of the role of neutrophils in tumor development has greatly depended on different experimental animal models of cancer. In this review we cover important findings that have been made about neutrophils in experimental animal models of cancer, point to their advantages and limitations, and discuss novel techniques that can be used to expand our knowledge of how neutrophils influence tumor progression. PMID:26976824

  6. Management of neutrophilic dermatoses.

    PubMed

    Schadt, Courtney R; Callen, Jeffrey P

    2012-01-01

    Neutrophilic dermatoses, including Sweet's syndrome, pyoderma gangrenosum, and rheumatoid neutrophilic dermatitis, are inflammatory conditions of the skin often associated with underlying systemic disease. These are characterized by the accumulation of neutrophils in the skin. The associated conditions, potential for systemic neutrophilic infiltration, and therapeutic management of these disorders can be similar. Sweet's syndrome can often be effectively treated with a brief course of systemic corticosteroids. Pyoderma gangrenosum, however, can be recurrent, and early initiation of a steroid-sparing agent is prudent. Second-line treatment for both of these conditions includes medications affecting neutrophil function, in addition to immunosuppressant medications.

  7. Novel Transgenic Mouse Model for Testing the Effect of Circulating IGF-I on Mammary Stem/Progenitor Cell Number and Tumorigenesis

    DTIC Science & Technology

    2007-08-01

    AD_________________ Award Number: W81XWH-06-1-0628 TITLE: Novel Transgenic Mouse Model for Testing ...TITLE AND SUBTITLE 5a. CONTRACT NUMBER Novel Transgenic Mouse Model for Testing the Effect of Circulating IGF-I on Mammary Stem/Progenitor Cell...tumorigenesis. We found no difference in time to tumor formation in ErbB2 vs. TTR-IGF-I/ErbB2 transgenic mice . Our conclusion is either that ErbB2

  8. Neutrophil Dysfunction in Sepsis

    PubMed Central

    Zhang, Fang; Liu, An-Lei; Gao, Shuang; Ma, Shui; Guo, Shu-Bin

    2016-01-01

    Objective: Sepsis is defined as life-threatening organ dysfunction due to a dysregulated host response to infection. In this article, we reviewed the correlation between neutrophil dysfunction and sepsis. Data Sources: Articles published up to May 31, 2016, were selected from the PubMed databases, with the keywords of “neutrophil function”, “neutrophil dysfunction”, and “sepsis”. Study Selection: Articles were obtained and reviewed to analyze the neutrophil function in infection and neutrophil dysfunction in sepsis. Results: We emphasized the diagnosis of sepsis and its limitations. Pathophysiological mechanisms involve a generalized circulatory, immune, coagulopathic, and/or neuroendocrine response to infection. Many studies focused on neutrophil burst or cytokines. Complement activation, impairment of neutrophil migration, and endothelial lesions are involved in this progress. Alterations of cytokines, chemokines, and other mediators contribute to neutrophil dysfunction in sepsis. Conclusions: Sepsis represents a severe derangement of the immune response to infection, resulting in neutrophil dysfunction. Neutrophil dysfunction promotes sepsis and even leads to organ failure. Mechanism studies, clinical practice, and strategies to interrupt dysregulated neutrophil function in sepsis are desperately needed. PMID:27824008

  9. Prevention of vascular inflammation by nanoparticle targeting of adherent neutrophils

    NASA Astrophysics Data System (ADS)

    Wang, Zhenjia; Li, Jing; Cho, Jaehyung; Malik, Asrar B.

    2014-03-01

    Inflammatory diseases such as acute lung injury and ischaemic tissue injury are caused by the adhesion of a type of white blood cell--polymorphonuclear neutrophils--to the lining of the circulatory system or vascular endothelium and unchecked neutrophil transmigration. Nanoparticle-mediated targeting of activated neutrophils on vascular endothelial cells at the site of injury may be a useful means of directly inactivating neutrophil transmigration and hence mitigating vascular inflammation. Here, we report a method employing drug-loaded albumin nanoparticles, which efficiently deliver drugs into neutrophils adherent to the surface of the inflamed endothelium. Using intravital microscopy of tumour necrosis factor-α-challenged mouse cremaster post-capillary venules, we demonstrate that fluorescently tagged albumin nanoparticles are largely internalized by neutrophils adherent to the activated endothelium via cell surface Fcɣ receptors. Administration of albumin nanoparticles loaded with the spleen tyrosine kinase inhibitor, piceatannol, which blocks `outside-in' β2 integrin signalling in leukocytes, detached the adherent neutrophils and elicited their release into the circulation. Thus, internalization of drug-loaded albumin nanoparticles into neutrophils inactivates the pro-inflammatory function of activated neutrophils, thereby offering a promising approach for treating inflammatory diseases resulting from inappropriate neutrophil sequestration and activation.

  10. Human neutrophils in auto-immunity.

    PubMed

    Thieblemont, Nathalie; Wright, Helen L; Edwards, Steven W; Witko-Sarsat, Véronique

    2016-04-01

    Human neutrophils have great capacity to cause tissue damage in inflammatory diseases via their inappropriate activation to release reactive oxygen species (ROS), proteases and other tissue-damaging molecules. Furthermore, activated neutrophils can release a wide variety of cytokines and chemokines that can regulate almost every element of the immune system. In addition to these important immuno-regulatory processes, activated neutrophils can also release, expose or generate neoepitopes that have the potential to break immune tolerance and result in the generation of autoantibodies, that characterise a number of human auto-immune diseases. For example, in vasculitis, anti-neutrophil cytoplasmic antibodies (ANCA) that are directed against proteinase 3 or myeloperoxidase are neutrophil-derived autoantigens and activated neutrophils are the main effector cells of vascular damage. In other auto-immune diseases, these neutrophil-derived neoepitopes may arise from a number of processes that include release of granule enzymes and ROS, changes in the properties of components of their plasma membrane as a result of activation or apoptosis, and via the release of Neutrophil Extracellular Traps (NETs). NETs are extracellular structures that contain chromatin that is decorated with granule enzymes (including citrullinated proteins) that can act as neo-epitopes to generate auto-immunity. This review therefore describes the processes that can result in neutrophil-mediated auto-immunity, and the role of neutrophils in the molecular pathologies of auto-immune diseases such as vasculitis, rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). We discuss the potential role of NETs in these processes and some of the debate in the literature regarding the role of this phenomenon in microbial killing, cell death and auto-immunity.

  11. Monoclonal B-cell lymphocytosis (MBL) with normal lymphocyte counts is associated with decreased numbers of normal circulating B-cell subsets.

    PubMed

    Hauswirth, Alexander W; Almeida, Julia; Nieto, Wendy G; Teodosio, Cristina; Rodriguez-Caballero, Arancha; Romero, Alfonso; López, Antonio; Fernandez-Navarro, Paulino; Vega, Tomas; Perez-Andres, Martin; Valent, Peter; Jäger, Ulrich; Orfao, Alberto

    2012-07-01

    Monoclonal B-cell lymphocytosis (MBL) with normal lymphocyte counts is associated with decreased numbers of normal circulating B-cell subsets.Little is known about the distribution of normal lymphoid cells and their subsets in the peripheral blood (PB) of subjects with monoclonal B-cell lymphocytosis (MBL). In our study, we compared the absolute number of PB lymphoid cells and their subpopulations in 95 MBL cases with normal lymphocyte counts vs. 617 age-/sex-matched non-MBL healthy subjects (controls), using highly sensitive flow cytometry. MBL cases showed significantly reduced numbers of normal circulating B-cells, at the expense of immature and naive B-cells; in addition, CD4+CD8+ double-positive T-cells and CD8+ T-cells were significantly lower and higher vs. controls, respectively. Moreover, most normal B-cell subsets were significantly decreased in PB at >1% MBL-counts, vs. "low-count" MBL cases, and lower amounts of immature/naive B-cells were detected in biclonal (particularly in cases with coexisting CLL-like- and non-CLL-like B-cell clones) vs. monoclonal MBL subjects. In summary, our results show imbalanced (reduced) absolute numbers of recently produced normal circulating B-cells (e.g., immature and naıve B-cells) in MBL, which becomes more pronounced as the MBL cell count increases.

  12. Neutrophilic dermatoses in children.

    PubMed

    Berk, David R; Bayliss, Susan J

    2008-01-01

    The neutrophilic dermatoses are rare disorders, especially in children, and are characterized by neutrophilic infiltrates in the skin and less commonly in extracutaneous tissue. The neutrophilic dermatoses share similar clinical appearances and associated conditions, including inflammatory bowel disease, malignancies, and medications. Overlap forms of disease demonstrating features of multiple neutrophilic dermatoses may be seen. The manuscript attempts to provide an up-to-date review of (i) classical neutrophilic dermatoses, focusing on distinctive features in children and (ii) neutrophilic dermatoses which may largely be pediatric or genodermatosis-associated (Majeed, SAPHO [synovitis, severe acne, sterile palmoplantar pustulosis, hyperostosis, and osteitis] syndrome, PAPA (pyogenic sterile arthritis, pyoderma gangrenosum, and acne), PFAPA (periodic fever with aphthous stomatitis, pharyngitis, and cervical adenopathy), and other periodic fever syndromes, and congenital erosive and vesicular dermatosis healing with reticulated supple scarring).

  13. Effect of clozapine on neutrophil kinetics in rabbits.

    PubMed

    Iverson, Suzanne; Kautiainen, Antti; Ip, Julia; Uetrecht, Jack P

    2010-07-19

    Clozapine is an atypical antipsychotic drug effective in the treatment of refractory schizophrenia; however, its use is limited due to its propensity to cause agranulocytosis in some patients. Little is known about the mechanism of idiosyncratic drug-induced agranulocytosis, in part because of the lack of a valid animal model. Clozapine is oxidized by activated human neutrophils and bone marrow cells to a reactive nitrenium ion by the myeloperoxidase-hydrogen peroxide system of neutrophils. This reactive metabolite has been shown in vitro to induce the apoptosis of neutrophils and bone marrow cells. While in vitro studies demonstrated the toxic potential of clozapine upon oxidation, it is not clear if similar conditions occur in vivo. In response to the difficulties encountered with detecting apoptotic neutrophils in vivo, we conducted a series of studies in rabbits using two fluorescent cell-labeling techniques to study the effect of clozapine treatment on neutrophil kinetics, that is, their rates of production and removal from circulation. The fluorescein dye, 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester (CFSE), was used as a general cell label to measure the half-life of neutrophils in blood. In addition, the thymidine analogue, 5-bromo-2-deoxyuridine (BrdU), was used to label dividing cells, thus enabling the measurement of the efflux of neutrophils from the bone marrow. Clozapine, indeed, increased the rate of both the release of neutrophils from the bone marrow and their subsequent disappearance from circulation. Failure of the bone marrow to compensate for a shorter neutrophil half-life could lead to agranulocytosis. Alternatively, the damage to neutrophils caused by clozapine could, in some patients, lead to an immune-mediated response against neutrophils resulting in agranulocytosis.

  14. Neutrophil extracellular traps in cancer progression.

    PubMed

    Cools-Lartigue, Jonathan; Spicer, Jonathan; Najmeh, Sara; Ferri, Lorenzo

    2014-11-01

    Neutrophils are being increasingly recognized as an important element in tumor progression. They have been shown to exert important effects at nearly every stage of tumor progression with a number of studies demonstrating that their presence is critical to tumor development. Novel aspects of neutrophil biology have recently been elucidated and its contribution to tumorigenesis is only beginning to be appreciated. Neutrophil extracellular traps (NETs) are neutrophil-derived structures composed of DNA decorated with antimicrobial peptides. They have been shown to trap and kill microorganisms, playing a critical role in host defense. However, their contribution to tumor development and metastasis has recently been demonstrated in a number of studies highlighting NETs as a potentially important therapeutic target. Here, studies implicating NETs as facilitators of tumor progression and metastasis are reviewed. In addition, potential mechanisms by which NETs may exert these effects are explored. Finally, the ability to target NETs therapeutically in human neoplastic disease is highlighted.

  15. The impact of trauma on neutrophil function.

    PubMed

    Hazeldine, Jon; Hampson, Peter; Lord, Janet M

    2014-12-01

    A well described consequence of traumatic injury is immune dysregulation, where an initial increase in immune activity is followed by a period of immune depression, the latter leaving hospitalised trauma patients at an increased risk of nosocomial infections. Here, we discuss the emerging role of the neutrophil, the most abundant leucocyte in human circulation and the first line of defence against microbial challenge, in the initiation and propagation of the inflammatory response to trauma. We review the findings of the most recent studies to have investigated the impact of trauma on neutrophil function and discuss how alterations in neutrophil biology are being investigated as potential biomarkers by which to predict the outcome of hospitalised trauma patients. Furthermore, with trauma-induced changes in neutrophil biology linked to the development of such post-traumatic complications as multiple organ failure and acute respiratory distress syndrome, we highlight an area of research within the field of trauma immunology that is gaining considerable interest: the manipulation of neutrophil function as a means by which to potentially improve patient outcome.

  16. Neutrophils in cancer.

    PubMed

    Treffers, Louise W; Hiemstra, Ida H; Kuijpers, Taco W; van den Berg, Timo K; Matlung, Hanke L

    2016-09-01

    Neutrophils play an important role in cancer. This does not only relate to the well-established prognostic value of the presence of neutrophils, either in the blood or in tumor tissue, in the context of cancer progression or for the monitoring of therapy, but also to their active role in the progression of cancer. In the current review, we describe what is known in general about the role of neutrophils in cancer. What is emerging is a complex, rather heterogeneous picture with both pro- and anti-tumorigenic roles, which apparently differs with cancer type and disease stage. Furthermore, we will discuss the well-known role of neutrophils as myeloid-derived suppressor cells (MDSC), and also on the role of neutrophils as important effector cells during antibody therapy in cancer. It is clear that neutrophils contribute substantially to cancer progression in multiple ways, and this includes both direct effects on the cancer cells and indirect effect on the tumor microenvironment. While in many cases neutrophils have been shown to promote tumor progression, for instance by acting as MDSC, there are also protective effects, particularly when antibody immunotherapy is performed. A better understanding of the role of neutrophils is likely to provide opportunities for immunomodulation and for improving the treatment of cancer patients.

  17. Enhancements to the FAST-MAC Circulation Control Model and Recent High-Reynolds Number Testing in the National Transonic Facility

    NASA Technical Reports Server (NTRS)

    Milholen, William E., II; Jones, Gregory S.; Chan, David T.; Goodliff, Scott L.; Anders, Scott G.; Melton, Latunia P.; Carter, Melissa B.; Allan, Brian G.; Capone, Francis J.

    2013-01-01

    A second wind tunnel test of the FAST-MAC circulation control model was recently completed in the National Transonic Facility at the NASA Langley Research Center. The model was equipped with four onboard flow control valves allowing independent control of the circulation control plenums, which were directed over a 15% chord simple-hinged flap. The model was configured for low-speed high-lift testing with flap deflections of 30 and 60 degrees, along with the transonic cruise configuration with zero degree flap deflection. Testing was again conducted over a wide range of Mach numbers up to 0.88, and Reynolds numbers up to 30 million based on the mean chord. The first wind tunnel test had poor transonic force and moment data repeatability at mild cryogenic conditions due to inadequate thermal conditioning of the balance. The second test demonstrated that an improvement to the balance heating system significantly improved the transonic data repeatability, but also indicated further improvements are still needed. The low-speed highlift performance of the model was improved by testing various blowing slot heights, and the circulation control was again demonstrated to be effective in re-attaching the flow over the wing at off-design transonic conditions. A new tailored spanwise blowing technique was also demonstrated to be effective at transonic conditions with the benefit of reduced mass flow requirements.

  18. Neutrophil paralysis in sepsis.

    PubMed

    Alves-Filho, José C; Spiller, Fernando; Cunha, Fernando Q

    2010-09-01

    Sepsis develops when the initial host response is unable to contain the primary infection, resulting in widespread inflammation and multiple organ dysfunction. The impairment of neutrophil migration into the infection site, also termed neutrophil paralysis, is a critical hallmark of sepsis, which is directly related to the severity of the disease. Although the precise mechanism of this phenomenon is not fully understood, there has been much advancement in the understanding of this field. In this review, we highlight the recent insights into the molecular mechanisms of neutrophil paralysis during sepsis.

  19. Sexy again: the renaissance of neutrophils in psoriasis.

    PubMed

    Schön, Michael P; Broekaert, Sigrid M C; Erpenbeck, Luise

    2017-04-01

    Notwithstanding their prominent presence in psoriatic skin, the functional role of neutrophilic granulocytes still remains somewhat enigmatic. Sparked by exciting scientific discoveries regarding neutrophil functions within the last years, the interest in these short-lived cells of the innate immune system has been boosted recently. While it had been known for some time that neutrophils produce and respond to a number of inflammatory mediators, recent research has linked neutrophils with the pathogenic functions of IL-17, possibly in conjunction with the formation of NETs (neutrophil extracellular traps). Antipsoriatic therapies exert their effects, at least in part, through interference with neutrophils. Neutrophils also appear to connect psoriasis with comorbid diseases. However, directly tampering with neutrophil functions is not trivial as evinced by the failure of therapeutic approaches targeting redundantly regulated cellular communication networks. It has also become apparent that neutrophils link important pathogenic functions of the innate and the adaptive immune system and that they are intricately involved in regulatory networks underlying the pathophysiology of psoriasis. In order to advocate intensified research into the role of this interesting cell population, we here highlight some features of neutrophils and put them into perspective with our current view of the pathophysiology of psoriasis.

  20. Tetramethylpyrazine inhibits neutrophil activation following permanent cerebral ischemia in rats.

    PubMed

    Chang, Cheng-Yi; Kao, Tsung-Kuei; Chen, Wen-Ying; Ou, Yen-Chuan; Li, Jian-Ri; Liao, Su-Lan; Raung, Shue-Ling; Chen, Chun-Jung

    2015-07-31

    Experimental studies have demonstrated the beneficial effects of tetramethylpyrazine (TMP) against ischemic stroke and highlighted its crucial role in anti-inflammatory activity. This study provides evidence of an alternative target for TMP and sheds light on the mechanism of its anti-inflammatory action against ischemic brain injury. We report a global inhibitory effect of TMP on inflammatory cell intracerebral activation and infiltration in a rat model of permanent cerebral ischemia. The results of immunohistochemistry, enzymatic assay, flow cytometric analysis, and cytological analysis revealed that intraperitoneal TMP administration reduced neuronal loss, macrophage/microglia activation, brain parenchyma infiltrative neutrophils, and circulating neutrophils after cerebral ischemia. Biochemical studies of cultured neutrophils further demonstrated that TMP attenuated neutrophil migration, endothelium adhesion, spontaneous nitric oxide (NO) production, and stimuli-activated NO production after cerebral ischemia. In parallel with these anti-neutrophil phenomena, TMP also attenuated the activities of ischemia-induced inflammation-associated signaling molecules, including plasma high-mobility group box-1 protein (HMGB1) and neutrophil toll-like receptor-4 (TLR4), Akt, extracellular signal-regulated kinase (ERK), and inducible nitric oxide synthase. Another finding in this study was that the anti-neutrophil effect of TMP was accompanied by a further elevated expression of NF-E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in neutrophils after cerebral ischemia. Taken together, our results suggest that both the promotion of endogenous anti-inflammatory defense capacity and the attenuation of pro-inflammatory responses via targeting of circulating neutrophils by elevating Nrf2/HO-1 expression and inhibiting HMGB1/TLR4, Akt, and ERK signaling might actively contribute to TMP-mediated neuroprotection against cerebral ischemia.

  1. Neutrophil depletion delays wound repair in aged mice

    PubMed Central

    Nishio, Naomi; Okawa, Yayoi; Sakurai, Hidetoshi

    2008-01-01

    One of the most important clinical problems in caring for elderly patients is treatment of pressure ulcers. One component of normal wound healing is the generation of an inflammatory reaction, which is characterized by the sequential infiltration of neutrophils, macrophages and lymphocytes. Neutrophils migrate early in the wound healing process. In aged C57BL/6 mice, wound healing is relatively inefficient. We examined the effects of neutrophil numbers on wound healing in both young and aged mice. We found that the depletion of neutrophils by anti-Gr-1 antibody dramatically delayed wound healing in aged mice. The depletion of neutrophils in young mice had less effect on the kinetics of wound healing. Intravenous G-CSF injection increased the migration of neutrophils to the wound site. While the rate of wound repair did not change significantly in young mice following G-CSF injection, it increased significantly in old mice. PMID:19424869

  2. Pulmonary lesions induced by Pasteurella haemolytica in neutrophil sufficient and neutrophil deficient calves.

    PubMed Central

    Breider, M A; Walker, R D; Hopkins, F M; Schultz, T W; Bowersock, T L

    1988-01-01

    The role of neutrophils in the development of peracute lung lesions of bovine pneumonic pasteurellosis was investigated. Eight calves were divided into two groups of four calves each. Group I was treated with intravenous phosphate-buffered saline and served as the neutrophil sufficient calves. Group II was treated with intravenous hydroxyurea which produced a state of neutropenia. When peripheral blood neutrophil numbers dropped below 300 cells/microL in group II, all calves were challenged with an intrabronchial bolus of Pasteurella haemolytica in the log phase of growth. An acute inflammatory process occurred in both groups of calves indicated by a rise in body temperature. While pulmonary lesions occurred in both groups by six hours postinoculation, they varied in pathological characteristics. Pulmonary lesions in the neutrophil sufficient calves consisted of fibrinopurulent alveolitis-bronchiolitis with associated alveolar septal necrosis, interlobular edema, and intravascular thrombi. The neutrophil deficient calves had extensive intra-alveolar edema, interlobular edema, intraalveolar hemorrhage, atelectasis, and focal areas of alveolar septal necrosis. These results show that P. haemolytica can induce severe pulmonary tissue damage through both neutrophil dependent and neutrophil independent mechanisms. Images Fig. 1. Fig. 2. PMID:3370555

  3. Neutrophil Interactions Stimulate Evasive Hyphal Branching by Aspergillus fumigatus

    PubMed Central

    Jorgensen, Julianne; Frydman, Galit H.; Jones, Caroline N.

    2017-01-01

    Invasive aspergillosis (IA), primarily caused by Aspergillus fumigatus, is an opportunistic fungal infection predominantly affecting immunocompromised and neutropenic patients that is difficult to treat and results in high mortality. Investigations of neutrophil-hypha interaction in vitro and in animal models of IA are limited by lack of temporal and spatial control over interactions. This study presents a new approach for studying neutrophil-hypha interaction at single cell resolution over time, which revealed an evasive fungal behavior triggered by interaction with neutrophils: Interacting hyphae performed de novo tip formation to generate new hyphal branches, allowing the fungi to avoid the interaction point and continue invasive growth. Induction of this mechanism was independent of neutrophil NADPH oxidase activity and neutrophil extracellular trap (NET) formation, but could be phenocopied by iron chelation and mechanical or physiological stalling of hyphal tip extension. The consequence of branch induction upon interaction outcome depends on the number and activity of neutrophils available: In the presence of sufficient neutrophils branching makes hyphae more vulnerable to destruction, while in the presence of limited neutrophils the interaction increases the number of hyphal tips, potentially making the infection more aggressive. This has direct implications for infections in neutrophil-deficient patients and opens new avenues for treatments targeting fungal branching. PMID:28076396

  4. Neutrophil mobilization via plerixafor-mediated CXCR4 inhibition arises from lung demargination and blockade of neutrophil homing to the bone marrow

    PubMed Central

    Devi, Sapna; Wang, Yilin; Chew, Weng Keong; Lima, Ronald; A-González, Noelia; Mattar, Citra N.Z.; Chong, Shu Zhen; Schlitzer, Andreas; Bakocevic, Nadja; Chew, Samantha; Keeble, Jo L.; Goh, Chi Ching; Li, Jackson L.Y.; Evrard, Maximilien; Malleret, Benoit; Larbi, Anis; Renia, Laurent; Haniffa, Muzlifah; Tan, Suet Mien; Chan, Jerry K.Y.; Balabanian, Karl; Nagasawa, Takashi; Bachelerie, Françoise; Hidalgo, Andrés; Ginhoux, Florent; Kubes, Paul

    2013-01-01

    Blood neutrophil homeostasis is essential for successful host defense against invading pathogens. Circulating neutrophil counts are positively regulated by CXCR2 signaling and negatively regulated by the CXCR4–CXCL12 axis. In particular, G-CSF, a known CXCR2 signaler, and plerixafor, a CXCR4 antagonist, have both been shown to correct neutropenia in human patients. G-CSF directly induces neutrophil mobilization from the bone marrow (BM) into the blood, but the mechanisms underlying plerixafor-induced neutrophilia remain poorly defined. Using a combination of intravital multiphoton microscopy, genetically modified mice and novel in vivo homing assays, we demonstrate that G-CSF and plerixafor work through distinct mechanisms. In contrast to G-CSF, CXCR4 inhibition via plerixafor does not result in neutrophil mobilization from the BM. Instead, plerixafor augments the frequency of circulating neutrophils through their release from the marginated pool present in the lung, while simultaneously preventing neutrophil return to the BM. Our study demonstrates for the first time that drastic changes in blood neutrophils can originate from alternative reservoirs other than the BM, while implicating a role for CXCR4–CXCL12 interactions in regulating lung neutrophil margination. Collectively, our data provides valuable insights into the fundamental regulation of neutrophil homeostasis, which may lead to the development of improved treatment regimens for neutropenic patients. PMID:24081949

  5. LES of High-Reynolds-Number Coanda Flow Separating from a Rounded Trailing Edge of a Circulation Control Airfoil

    NASA Technical Reports Server (NTRS)

    Nichino, Takafumi; Hahn, Seonghyeon; Shariff, Karim

    2010-01-01

    This slide presentation reviews the Large Eddy Simulation of a high reynolds number Coanda flow that is separated from a round trailing edge of a ciruclation control airfoil. The objectives of the study are: (1) To investigate detailed physics (flow structures and statistics) of the fully turbulent Coanda jet applied to a CC airfoil, by using LES (2) To compare LES and RANS results to figure out how to improve the performance of existing RANS models for this type of flow.

  6. A taurine-supplemented vegan diet may blunt the contribution of neutrophil activation to acute coronary events.

    PubMed

    McCarty, Mark F

    2004-01-01

    Neutrophils are activated in the coronary circulation during acute coronary events (unstable angina and myocardial infarction), often prior to the onset of ischemic damage. Moreover, neutrophils infiltrate coronary plaque in these circumstances, and may contribute to the rupture or erosion of this plaque, triggering thrombosis. Activated neutrophils secrete proteolytic enzymes in latent forms which are activated by the hypochlorous acid (HOCl) generated by myeloperoxidase. These phenomena may help to explain why an elevated white cell count has been found to be an independent coronary risk factor. Low-fat vegan diets can decrease circulating leukocytes--neutrophils and monocytes--possibly owing to down-regulation of systemic IGF-I activity. Thus, a relative neutropenia may contribute to the coronary protection afforded by such diets. However, vegetarian diets are devoid of taurine - the physiological antagonist of HOCl--and tissue levels of this nutrient are relatively low in vegetarians. Taurine has anti-atherosclerotic activity in animal models, possibly reflecting a role for macrophage-derived myeloperoxidase in the atherogenic process. Taurine also has platelet-stabilizing and anti-hypertensive effects that presumably could reduce coronary risk. Thus, it is proposed that a taurine-supplemented low-fat vegan diet represents a rational strategy for diminishing the contribution of activated neutrophils to acute coronary events; moreover, such a regimen would work in a number of other complementary ways to promote cardiovascular health. Moderate alcohol consumption, the well-tolerated drug pentoxifylline, and 5-lipoxygenase inhibitors--zileuton, boswellic acids, fish oil--may also have potential in this regard.

  7. Neutrophils scan for activated platelets to initiate inflammation

    PubMed Central

    Sreeramkumar, Vinatha; Adrover, José M.; Ballesteros, Ivan; Cuartero, Maria Isabel; Rossaint, Jan; Bilbao, Izaskun; Nácher, Maria; Pitaval, Christophe; Radovanovic, Irena; Fukui, Yoshinori; McEver, Rodger P.; Filippi, Marie-Dominique; Lizasoain, Ignacio; Ruiz-Cabello, Jesús; Zarbock, Alexander; Moro, María A.; Hidalgo, Andrés

    2014-01-01

    Immune and inflammatory responses require leukocytes to migrate within and through the vasculature, a process that is facilitated by their capacity to switch to a polarized morphology with asymmetric distribution of receptors. We report that neutrophil polarization within activated venules served to organize a protruding domain that engaged activated platelets present in the bloodstream. The selectin ligand PSGL-1 transduced signals emanating from these interactions, resulting in redistribution of receptors that drive neutrophil migration. Consequently, neutrophils unable to polarize or to transduce signals through PSGL-1 displayed aberrant crawling, and blockade of this domain protected mice against thrombo-inflammatory injury. These results reveal that recruited neutrophils scan for activated platelets, and suggest that their bipolarity allows integration of signals present at both the endothelium and the circulation before inflammation proceeds. PMID:25477463

  8. Fatty acids as modulators of neutrophil recruitment, function and survival.

    PubMed

    Rodrigues, Hosana G; Takeo Sato, Fabio; Curi, Rui; Vinolo, Marco A R

    2016-08-15

    Neutrophils are well-known to act in the destruction of invading microorganisms. They have also been implicated in the activation of other immune cells including B- and T-lymphocytes and in the resolution of inflammation and tissue regeneration. Neutrophils are produced in the bone marrow and released into the circulation from where they migrate to tissues to perform their effector functions. Neutrophils are in constant contact with fatty acids that can modulate their function, activation and fate (survival or cell death) through different mechanisms. In this review, the effects of fatty acids pertaining to five classes, namely, long-chain saturated fatty acids (LCSFAs), short-chain fatty acids (SCFAs), and omega-3 (n-3), omega-6 (n-6) and omega-9 (n-9) unsaturated fatty acids, on neutrophils and the relevance of these effects for disease development are discussed.

  9. Platelets enhance neutrophil transendothelial migration

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Platelets are increasingly recognized as important mediators of inflammation in addition to thrombosis. While platelets have been shown to promote neutrophil (PMN) adhesion to endothelium in various inflammatory models, it is unclear whether platelets enhance neutrophil transmigration across inflame...

  10. The Multifaceted Functions of Neutrophils

    PubMed Central

    Mayadas, Tanya N.; Cullere, Xavier; Lowell, Clifford A.

    2014-01-01

    Neutrophils and neutrophil-like cells are the major pathogen-fighting immune cells in organisms ranging from slime molds to mammals. Central to their function is their ability to be recruited to sites of infection, to recognize and phagocytose microbes, and then to kill pathogens through a combination of cytotoxic mechanisms. These include the production of reactive oxygen species, the release of antimicrobial peptides, and the recently discovered expulsion of their nuclear contents to form neutrophil extracellular traps. Here we discuss these primordial neutrophil functions, which also play key roles in tissue injury, by providing details of neutrophil cytotoxic functions and congenital disorders of neutrophils. In addition, we present more recent evidence that interactions between neutrophils and adaptive immune cells establish a feed-forward mechanism that amplifies pathologic inflammation. These newly appreciated contributions of neutrophils are described in the setting of several inflammatory and autoimmune diseases. PMID:24050624

  11. Changes in neutrophil functions in astronauts.

    PubMed

    Kaur, Indreshpal; Simons, Elizabeth R; Castro, Victoria A; Mark Ott, C; Pierson, Duane L

    2004-09-01

    Exploration class human spaceflight missions will require astronauts with robust immune systems. Innate immunity will be an essential element for the healthcare maintenance of astronauts during these lengthy expeditions. This study investigated neutrophil phagocytosis, oxidative burst, and degranulation of 25 astronauts after four space shuttle missions and in nine healthy control subjects. Space flight duration ranged from 5 to 11 days. Blood specimens were obtained 10 days before launch, immediately after landing, and 3 days after landing. The number of neutrophils increased by 85% at landing compared to preflight levels. The mean values for phagocytosis of Escherichia coli and oxidative burst capacity in neutrophils from astronauts on the 5-day mission were not significantly different from those observed in neutrophils from the control subjects. Before and after 9- to 11-day missions, however, phagocytosis and oxidative burst capacities were significantly lower than control mean values. No consistent changes in degranulation or expression of surface markers were observed before or after any of the space missions. This study indicates that neutrophil phagocytic and oxidative functions are affected by factors associated with space flight and this relationship may depend on mission duration.

  12. Neutrophil disorders and their management

    PubMed Central

    Lakshman, R; Finn, A

    2001-01-01

    Neutrophil disorders are an uncommon yet important cause of morbidity and mortality in infants and children. This article is an overview of these conditions, with emphasis on clinical recognition, rational investigation, and treatment. A comprehensive list of references is provided for further reading. Key Words: neutrophil disorders • chronic granulomatous disease • neutrophil chemotaxis • phagocytosis PMID:11271792

  13. Role of neutrophils in experimental septicemia and septic arthritis induced by Staphylococcus aureus.

    PubMed Central

    Verdrengh, M; Tarkowski, A

    1997-01-01

    We have previously described a murine model of hematogenously induced Staphylococcus aureus sepsis and arthritis. In this model, large numbers of granulocytes can be observed both in the circulation and locally in the inflamed synovium within 24 h after bacterial inoculation. To assess the role of neutrophils in this severe infection, mice were given granulocyte-depleting monoclonal antibody RB6-8C5 before being inoculated with S. aureus. All the control mice survived their intravenous injection with 3 x 10(7) CFU of S. aureus, whereas all the mice given RB6-8C5 antibody died of sepsis within 2 to 3 days. Even when the inoculum size was reduced sixfold (i.e., 6 x 10(6) CFU/mouse), 50% of the RB6-8C5-treated animals died within 6 days. The RB6-8C5-treated mice had a significantly higher burden of bacteria in their blood and kidneys 24 and 48 h after bacterial inoculation. In addition, when a suboptimal dose of bacteria was administered, the neutrophil-depleted animals displayed a higher frequency of arthritis than did the controls. The granulocyte-depleted animals exhibited increased levels of the proinflammatory cytokines tumor necrosis factor alpha, interleukin-6, and gamma interferon, reflecting the severity of their disease. This is the first direct demonstration of neutrophils playing a crucial protective role in the early phase of S. aureus infection. PMID:9199413

  14. Recapitulation of in vivo-like neutrophil transendothelial migration using a microfluidic platform.

    PubMed

    Wu, Xiaojie; Newbold, Molly A; Haynes, Christy L

    2015-08-07

    Neutrophil transendothelial migration (TEM) is an essential physiological process that regulates the recruitment of neutrophils in response to inflammatory signals. Herein, a versatile hydrogel scaffold is embedded in a microfluidic platform that supports an endothelial cell layer cultured in the vertical direction and highly stable chemical gradients; this construct is employed to mimic the in vivo neutrophil TEM process. We found that the number of neutrophils migrating across the endothelial cell layer is dependent on the presented chemoattractant concentration and the spatial profile of the chemical gradient. Endothelial cells play a critical role in neutrophil TEM by promoting neutrophil morphological changes as well as expressing surface receptor molecules that are indispensable for inducing neutrophil attachment and migration. Furthermore, the microfluidic device also supports competing chemoattractant gradients to facilitate neutrophil TEM studies in complex microenvironments that more accurately model the in vivo system than simplified microenvironments without the complexity of chemical gradients. This work demonstrates that combinations of any two different chemoattractants induce more significant neutrophil migration than a single chemoattractant in the same total amount, indicating synergistic effects between distinct chemoattractants. The in vitro reconstitution of neutrophil TEM successfully translates planar neutrophil movement into in vivo-like neutrophil recruitment and accelerates understanding of cellular interactions between neutrophils and endothelial cells within the complicated physiological milieu.

  15. The Neutrophil Nucleus and Its Role in Neutrophilic Function.

    PubMed

    Carvalho, Leonardo Olivieri; Aquino, Elaine Nascimento; Neves, Anne Caroline Dias; Fontes, Wagner

    2015-09-01

    The cell nucleus plays a key role in differentiation processes in eukaryotic cells. It is not the nucleus in particular, but the organization of the genes and their remodeling that provides the data for the adjustments to be made according to the medium. The neutrophil nucleus has a different morphology. It is a multi-lobed nucleus where some researchers argue no longer function. However, studies indicate that it is very probable the occurrence of chromatin remodeling during activation steps. It may be that the human neutrophil nucleus also contributes to the mobility of neutrophils through thin tissue spaces. Questions like these will be discussed in this small review. The topics include morphology of human neutrophil nucleus, maturation process and modifications of the neutrophil nucleus, neutrophil activation and chromatin modifications, causes and consequences of multi-lobulated segmented morphology, and importance of the nucleus in the formation of neutrophil extracellular traps (NETs).

  16. Reduced CD5(+) CD24(hi) CD38(hi) and interleukin-10(+) regulatory B cells in active anti-neutrophil cytoplasmic autoantibody-associated vasculitis permit increased circulating autoantibodies.

    PubMed

    Aybar, L T; McGregor, J G; Hogan, S L; Hu, Y; Mendoza, C E; Brant, E J; Poulton, C J; Henderson, C D; Falk, R J; Bunch, D O

    2015-05-01

    Pathogenesis of anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis is B cell-dependent, although how particular B cell subsets modulate immunopathogenesis remains unknown. Although their phenotype remains controversial, regulatory B cells (Bregs ), play a role in immunological tolerance via interleukin (IL)-10. Putative CD19(+) CD24(hi) CD38(hi) and CD19(+) CD24(hi) CD27(+) Bregs were evaluated in addition to their CD5(+) subsets in 69 patients with ANCA-associated vasculitis (AAV). B cell IL-10 was verified by flow cytometry following culture with CD40 ligand and cytosine-phosphate-guanosine (CpG) DNA. Patients with active disease had decreased levels of CD5(+) CD24(hi) CD38(hi) B cells and IL-10(+) B cells compared to patients in remission and healthy controls (HCs). As IL-10(+) and CD5(+) CD24(hi) CD38(hi) B cells normalized in remission within an individual, ANCA titres decreased. The CD5(+) subset of CD24(hi) CD38(hi) B cells decreases in active disease and rebounds during remission similarly to IL-10-producing B cells. Moreover, CD5(+) B cells are enriched in the ability to produce IL-10 compared to CD5(neg) B cells. Together these results suggest that CD5 may identify functional IL-10-producing Bregs . The malfunction of Bregs during active disease due to reduced IL-10 expression may thus permit ANCA production.

  17. Circulating progenitor cells in hypertensive subjects: Effectiveness of a treatment with olmesartan in improving cell number and miR profile in addition to expected pharmacological effects

    PubMed Central

    Aragona, Caterina Oriana; Cairo, Valentina; Scuruchi, Michele; Lo Gullo, Alberto; D’Ascola, Angela; Alibrandi, Angela; Loddo, Saverio; Quartuccio, Sebastiano; Morace, Carmela; Mormina, Enricomaria; Basile, Giorgio; Saitta, Antonino; Imbalzano, Egidio

    2017-01-01

    CD34+ circulating progenitor cells (CD34+CPCs) are a population of multipotent cells which can delay the development of atherosclerosis and cardiovascular disease (CVD) in conditions of increased CV risk. MicroRNAs (miRs) 221 and 222 modulate different genes regulating angiogenesis and inflammation; moreover, miR221/22 have beenshown to participate in differentiation and proliferation of CD34+CPCs, inhibiting cell migration and homing. miR221/222 in CD34+CPCs from hypertensive subjects are also increased and associated with CD34+cell number and reactive oxygen species (ROS). We evaluated CD34+CPC number, intracellular miR221/222 and ROS levels, arterial stiffness (AS)and echocardiography indices at baseline (T0).Then, after a six-month treatment with olmesartan, 20 mg/day (T1), in 57 hypertensive patients with left ventricular hypertrophy (LVH) and with no additional risk factor for CVD, and in 29 healthy controls (baseline),fibrinogen, C-reactive protein (CRP), glucose and lipid profiles were also evaluated.At T1, blood pressure values, CRP and fibrinogen levels, ROS and miR221/222 were significantly decreased (all p <0.001), as were AS indices and LV mass index (p<0.001), while cell number was increased (p<0.001). Olmesartan is effective in reducing miR and ROS levels in CD34+CPCs from hypertensive subjects, as well as in increasing CD34+CPC number, providing multilevel CV protection, in addition to its expected pharmacological effects. PMID:28301500

  18. A Wind Tunnel Experiment for Trailing Edge Circulation Control on a 6 Percent 2-D Airfoil up to Transonic Mach Numbers

    NASA Technical Reports Server (NTRS)

    Alexander, Michael G.; Anders, Scott G.; Johnson, Stuart K.

    2005-01-01

    A wind tunnel test was conducted on a six percent thick slightly cambered elliptical circulation control airfoil with both upper and lower surface blowing. Parametric evaluations of jet slot heights and Coanda surface shapes were conducted at mass flow coefficients (C(sub mu)) from 0.0 to 0.12. The test data was acquired in the NASA Langley Transonic Dynamics Tunnel at Mach numbers of 0.8 and 0.3 at Reynolds numbers per foot of 1.05 x 10(exp 6) and 2.43 x 10(exp 5) respectively. For the transonic condition, (Mach = 0.8 at alpha = +3 deg), it was generally found that the smaller slot and larger Coanda surface were more effective overall than other slot/Coanda surface combinations. Generally it was found at Mach = 0.3 at alpha = 6 deg that the smaller slot and smaller Coanda surface were more effective overall than other slot/Coanda surface combinations.

  19. Recent advances in understanding neutrophils

    PubMed Central

    Deniset, Justin F.; Kubes, Paul

    2016-01-01

    Neutrophils have long been regarded as key effectors of the innate immune response during acute inflammation. Recent evidence has revealed a greater functional diversity for these cells than previously appreciated, expanding roles for neutrophils in adaptive immunity and chronic pathologies. In this review, we summarize some of the evolving paradigms in the neutrophil field and highlight key advances that have contributed to our understanding of neutrophil behavior and function in vivo. We examine the concept of neutrophil subsets and polarization, we discuss novel immunomodulatory roles for neutrophils in shaping the immune response, and, finally, we identify technical advances that will further enhance our ability to track the function and fate of neutrophils. PMID:28105328

  20. Differential Use of Human Neutrophil Fcγ Receptors for Inducing Neutrophil Extracellular Trap Formation.

    PubMed

    Alemán, Omar Rafael; Mora, Nancy; Cortes-Vieyra, Ricarda; Uribe-Querol, Eileen; Rosales, Carlos

    2016-01-01

    Neutrophils (PMN) are the most abundant leukocytes in the blood. PMN migrate from the circulation to sites of infection, where they are responsible for antimicrobial functions. PMN use phagocytosis, degranulation, and formation of neutrophil extracellular traps (NETs) to kill microbes. NETs are fibers composed of chromatin and neutrophil-granule proteins. Several pathogens, including bacteria, fungi, and parasites, and also some pharmacological stimuli such as phorbol 12-myristate 13-acetate (PMA) are efficient inducers of NETs. Antigen-antibody complexes are also capable of inducing NET formation. However the particular Fcγ receptor involved in triggering this function is a matter of controversy. In order to provide some insight into what Fcγ receptor is responsible for NET formation, each of the two human Fcγ receptors was stimulated individually by specific monoclonal antibodies and NET formation was evaluated. FcγRIIa cross-linking did not promote NET formation. Cross-linking other receptors such as integrins also did not promote NET formation. In contrast FcγRIIIb cross-linking induced NET formation similarly to PMA stimulation. NET formation was dependent on NADPH-oxidase, PKC, and ERK activation. These data show that cross-linking FcγRIIIb is responsible for NET formation by the human neutrophil.

  1. Differential Use of Human Neutrophil Fcγ Receptors for Inducing Neutrophil Extracellular Trap Formation

    PubMed Central

    Alemán, Omar Rafael; Mora, Nancy; Cortes-Vieyra, Ricarda; Uribe-Querol, Eileen; Rosales, Carlos

    2016-01-01

    Neutrophils (PMN) are the most abundant leukocytes in the blood. PMN migrate from the circulation to sites of infection, where they are responsible for antimicrobial functions. PMN use phagocytosis, degranulation, and formation of neutrophil extracellular traps (NETs) to kill microbes. NETs are fibers composed of chromatin and neutrophil-granule proteins. Several pathogens, including bacteria, fungi, and parasites, and also some pharmacological stimuli such as phorbol 12-myristate 13-acetate (PMA) are efficient inducers of NETs. Antigen-antibody complexes are also capable of inducing NET formation. However the particular Fcγ receptor involved in triggering this function is a matter of controversy. In order to provide some insight into what Fcγ receptor is responsible for NET formation, each of the two human Fcγ receptors was stimulated individually by specific monoclonal antibodies and NET formation was evaluated. FcγRIIa cross-linking did not promote NET formation. Cross-linking other receptors such as integrins also did not promote NET formation. In contrast FcγRIIIb cross-linking induced NET formation similarly to PMA stimulation. NET formation was dependent on NADPH-oxidase, PKC, and ERK activation. These data show that cross-linking FcγRIIIb is responsible for NET formation by the human neutrophil. PMID:27034964

  2. Granule Protein Processing and Regulated Secretion in Neutrophils

    PubMed Central

    Sheshachalam, Avinash; Srivastava, Nutan; Mitchell, Troy; Lacy, Paige; Eitzen, Gary

    2014-01-01

    Neutrophils are part of a family of granulocytes that, together with eosinophils and basophils, play an essential role in innate immunity. Neutrophils are the most abundant circulating leukocytes and are vital for rapid immune responses, being recruited to sites of injury or infection within minutes, where they can act as specialized phagocytic cells. However, another prominent function of neutrophils is the release of pro-inflammatory compounds, including cytokines, chemokines, and digestive enzymes, which are stored in intracellular compartments and released through regulated exocytosis. Hence, an important feature that contributes to rapid immune responses is capacity of neutrophils to synthesize and store pre-formed pro-inflammatory mediators in specialized intracellular vesicles and thus no new synthesis is required. This review will focus on advancement in three topics relevant to neutrophil secretion. First, we will examine what is known about basal level pro-inflammatory mediator synthesis, trafficking, and storage in secretory compartments. Second, we will review recent advancements in the mechanisms that control vesicle mobilization and the release of pre-formed mediators. Third, we will examine the upregulation and de novo synthesis of pro-inflammatory mediators by neutrophils engaged at sites of infection. PMID:25285096

  3. Numerical simulation of distribution of neutrophils in a lattice alveolar capillary network.

    PubMed

    Shirai, Atsushi; Hayase, Toshiyuki

    2009-02-28

    Neutrophils are known to be retained in narrow pulmonary capillaries, even in normal lungs, due to their low deformability, resulting in a higher concentration than that in systemic circulation. In this study, to obtain a fundamental understanding of the behavior of neutrophils, we simplified an alveolar capillary network to a rectangular grid of short capillary segments and numerically investigated the flow of a suspension of neutrophils and plasma through the capillary network for various concentrations of the suspension, Csus, injected into the network. The cells traveled limited preferential paths in the network while Csus was low. Retention of a cell or cells induced plugging of the segment with a cessation of blood flow, and as the result of the changed plasma flow field caused by such plugging, the cells took various routes differing from the preferential paths. A low incidence of plugging helped to accelerate the cells flowing in the network with tight segments, resulting in a decrease in their mean transit time through the network as compared with the case of a single-cell transit. On the contrary, however, an increasing incidence of plugging induced backward motion of the cells and a resultant increase in the mean transit time. The time-averaged number of cells in the network increased with the increase in Csus, and the fractional residence time of cells in individual segments approached a constant. This means that a high concentration of neutrophils facilitates their uniform distribution in the network. However, the ratio between the time-averaged concentration of the cells in the network and Csus decreased and our numerical simulation did not reach the experimentally obtained value. This implies that, in a real alveolar capillary bed, plasma leaks through the plugged segments or that the capillary network has bypasses through which the plasma can flow.

  4. Proteinase 3 contributes to transendothelial migration of NB1-positive neutrophils.

    PubMed

    Kuckleburg, Christopher J; Tilkens, Sarah B; Santoso, Sentot; Newman, Peter J

    2012-03-01

    Neutrophil transmigration requires the localization of neutrophils to endothelial cell junctions, in which receptor-ligand interactions and the action of serine proteases promote leukocyte diapedesis. NB1 (CD177) is a neutrophil-expressed surface molecule that has been reported to bind proteinase 3 (PR3), a serine protease released from activated neutrophils. PR3 has demonstrated proteolytic activity on a number of substrates, including extracellular matrix proteins, although its role in neutrophil transmigration is unknown. Recently, NB1 has been shown to be a heterophilic binding partner for the endothelial cell junctional protein, PECAM-1. Disrupting the interaction between NB1 and PECAM-1 significantly inhibits neutrophil transendothelial cell migration on endothelial cell monolayers. Because NB1 interacts with endothelial cell PECAM-1 at cell junctions where transmigration occurs, we considered that NB1-PR3 interactions may play a role in aiding neutrophil diapedesis. Blocking Abs targeting the heterophilic binding domain of PECAM-1 significantly inhibited transmigration of NB1-positive neutrophils through IL-1β-stimulated endothelial cell monolayers. PR3 expression and activity were significantly increased on NB1-positive neutrophils following transmigration, whereas neutrophils lacking NB1 demonstrated no increase in PR3. Finally, using selective serine protease inhibitors, we determined that PR3 activity facilitated transmigration of NB1-positive neutrophils under both static and flow conditions. These data demonstrate that PR3 contributes in the selective recruitment of the NB1-positive neutrophil population.

  5. Commensal microbiota stimulate systemic neutrophil migration through induction of Serum amyloid A

    PubMed Central

    Kanther, Michelle; Tomkovich, Sarah; Sun, Xiaolun; Grosser, Melinda R.; Koo, Jaseol; Flynn, Edward J.; Jobin, Christian; Rawls, John F.

    2015-01-01

    Summary Neutrophils serve critical roles in inflammatory responses to infection and injury, and mechanisms governing their activity represent attractive targets for controlling inflammation. The commensal microbiota is known to regulate the activity of neutrophils and other leucocytes in the intestine, but the systemic impact of the microbiota on neutrophils remains unknown. Here we utilized in vivo imaging in gnotobiotic zebrafish to reveal diverse effects of microbiota colonization on systemic neutrophil development and function. The presence of a microbiota resulted in increased neutrophil number and myeloperoxidase expression, and altered neutrophil localization and migratory behaviours. These effects of the microbiota on neutrophil homeostasis were accompanied by an increased recruitment of neutrophils to injury. Genetic analysis identified the microbiota-induced acute phase protein serum amyloid A (Saa) as a host factor mediating microbial stimulation of tissue-specific neutrophil migratory behaviours. In vitro studies revealed that zebrafish cells respond to Saa exposure by activating NF-κB, and that Saa-dependent neutrophil migration requires NF-κB-dependent gene expression. These results implicate the commensal microbiota as an important environmental factor regulating diverse aspects of systemic neutrophil development and function, and reveal a critical role for a Saa-NF-κB signalling axis in mediating neutrophil migratory responses. PMID:24373309

  6. Characterization of Neutrophil Function in Human Cutaneous Leishmaniasis Caused by Leishmania braziliensis

    PubMed Central

    Conceição, Jacilara; Davis, Richard; Carneiro, Pedro Paulo; Giudice, Angela; Muniz, Aline C.; Wilson, Mary E.; Carvalho, Edgar M.; Bacellar, Olívia

    2016-01-01

    Infection with different Leishmania spp. protozoa can lead to a variety of clinical syndromes associated in many cases with inflammatory responses in the skin. Although macrophages harbor the majority of parasites throughout chronic infection, neutrophils are the first inflammatory cells to migrate to the site of infection. Whether neutrophils promote parasite clearance or exacerbate disease in murine models varies depending on the susceptible or resistant status of the host. Based on the hypothesis that neutrophils contribute to a systemic inflammatory state in humans with symptomatic L. braziliensis infection, we evaluated the phenotype of neutrophils from patients with cutaneous leishmaniasis (CL) during the course of L. braziliensis infection. After in vitro infection with L. braziliensis, CL patient neutrophils produced more reactive oxygen species (ROS) and higher levels of CXCL8 and CXCL9, chemokines associated with recruitment of neutrophils and Th1-type cells, than neutrophils from control healthy subjects (HS). Despite this, CL patient and HS neutrophils were equally capable of phagocytosis of L. braziliensis. There was no difference between the degree of activation of neutrophils from CL versus healthy subjects, assessed by CD66b and CD62L expression using flow cytometry. Of interest, these studies revealed that both parasite-infected and bystander neutrophils became activated during incubation with L. braziliensis. The enhanced ROS and chemokine production in neutrophils from CL patients reverted to baseline after treatment of disease. These data suggest that the circulating neutrophils during CL are not necessarily more microbicidal, but they have a more pro-inflammatory profile after parasite restimulation than neutrophils from healthy subjects. PMID:27167379

  7. Patrolling monocytes promote intravascular neutrophil activation and glomerular injury in the acutely inflamed glomerulus

    PubMed Central

    Finsterbusch, Michaela; Hall, Pam; Li, Anqi; Devi, Sapna; Westhorpe, Clare L. V.; Kitching, A. Richard

    2016-01-01

    Nonclassical monocytes undergo intravascular patrolling in blood vessels, positioning them ideally to coordinate responses to inflammatory stimuli. Under some circumstances, the actions of monocytes have been shown to involve promotion of neutrophil recruitment. However, the mechanisms whereby patrolling monocytes control the actions of neutrophils in the circulation are unclear. Here, we examined the contributions of monocytes to antibody- and neutrophil-dependent inflammation in a model of in situ immune complex-mediated glomerulonephritis. Multiphoton and spinning disk confocal intravital microscopy revealed that monocytes patrol both uninflamed and inflamed glomeruli using β2 and α4 integrins and CX3CR1. Monocyte depletion reduced glomerular injury, demonstrating that these cells promote inappropriate inflammation in this setting. Monocyte depletion also resulted in reductions in neutrophil recruitment and dwell time in glomerular capillaries and in reactive oxygen species (ROS) generation by neutrophils, suggesting a role for cross-talk between monocytes and neutrophils in induction of glomerulonephritis. Consistent with this hypothesis, patrolling monocytes and neutrophils underwent prolonged interactions in glomerular capillaries, with the duration of these interactions increasing during inflammation. Moreover, neutrophils that interacted with monocytes showed increased retention and a greater propensity for ROS generation in the glomerulus. Also, renal patrolling monocytes, but not neutrophils, produced TNF during inflammation, and TNF inhibition reduced neutrophil dwell time and ROS production, as well as renal injury. These findings show that monocytes and neutrophils undergo interactions within the glomerular microvasculature. Moreover, evidence indicates that, in response to an inflammatory stimulus, these interactions allow monocytes to promote neutrophil recruitment and activation within the glomerular microvasculature, leading to neutrophil

  8. Sulforaphane restores cellular glutathione levels and reduces chronic periodontitis neutrophil hyperactivity in vitro.

    PubMed

    Dias, Irundika H K; Chapple, Ian L C; Milward, Mike; Grant, Melissa M; Hill, Eric; Brown, James; Griffiths, Helen R

    2013-01-01

    The production of high levels of reactive oxygen species by neutrophils is associated with the local and systemic destructive phenotype found in the chronic inflammatory disease periodontitis. In the present study, we investigated the ability of sulforaphane (SFN) to restore cellular glutathione levels and reduce the hyperactivity of circulating neutrophils associated with chronic periodontitis. Using differentiated HL60 cells as a neutrophil model, here we show that generation of extracellular O2 (. -) by the nicotinamide adenine dinucleotide (NADPH) oxidase complex is increased by intracellular glutathione depletion. This may be attributed to the upregulation of thiol regulated acid sphingomyelinase driven lipid raft formation. Intracellular glutathione was also lower in primary neutrophils from periodontitis patients and, consistent with our previous findings, patients neutrophils were hyper-reactive to stimuli. The activity of nuclear factor erythroid-2-related factor 2 (Nrf2), a master regulator of the antioxidant response, is impaired in circulating neutrophils from chronic periodontitis patients. Although patients' neutrophils exhibit a low reduced glutathione (GSH)/oxidised glutathione (GSSG) ratio and a higher total Nrf2 level, the DNA-binding activity of nuclear Nrf2 remained unchanged relative to healthy controls and had reduced expression of glutamate cysteine ligase catalytic (GCLC), and modifier (GCLM) subunit mRNAs, compared to periodontally healthy subjects neutrophils. Pre-treatment with SFN increased expression of GCLC and GCM, improved intracellular GSH/GSSG ratios and reduced agonist-activated extracellular O2 (. -) production in both dHL60 and primary neutrophils from patients with periodontitis and controls. These findings suggest that a deficiency in Nrf2-dependent pathways may underpin susceptibility to hyper-reactivity in circulating primary neutrophils during chronic periodontitis.

  9. Changes in Neutrophil Functions in Astronauts

    NASA Technical Reports Server (NTRS)

    Kaur, Indreshpal; Simons, Elizabeth R.; Castro, Victoria; Pierson, Duane L.

    2002-01-01

    Neutrophil functions (phagocytosis, oxidative burst, degranulation) and expression of surface markers involved in these functions were studied in 25 astronauts before and after 4 space shuttle missions. Space flight duration ranged from 5 to 11 days. Blood specimens were obtained 10 days before launch (preflight or L-10), immediately after landing (landing or R+0), and again at 3 days after landing (postflight or R+3). Blood samples were also collected from 9 healthy low-stressed subjects at 3 time points simulating a 10-day shuttle mission. The number of neutrophils increased at landing by 85 percent when compared to the preflight numbers. Neutrophil functions were studied in whole blood using flow cytometric methods. Phagocytosis of E.coli-FITC and oxidative burst capacity of the neutrophils following the 9 to 11 day missions were lower at all three sampling points than the mean values for control subjects. Phagocytosis and oxidative burst capacity of the astronauts was decreased even 10-days before space flight. Mission duration appears to be a factor in phagocytic and oxidative functions. In contrast, following the short-duration (5-days) mission, these functions were unchanged from control values. No consistent changes in degranulation were observed following either short or medium length space missions. The expression of CD16, CD32, CD11a, CD11b, CD11c, L-selectin and CD36 was measured and found to be variable. Specifically, CD16 and CD32 did not correlate with the changes in oxidative burst and phagocytosis. We can conclude from this study that the stresses associated with space flight can alter the important functions of neutrophils.

  10. Sex Hormones Coordinate Neutrophil Immunity in the Vagina by Controlling Chemokine Gradients.

    PubMed

    Lasarte, Sandra; Samaniego, Rafael; Salinas-Muñoz, Laura; Guia-Gonzalez, Mauriel A; Weiss, Linnea A; Mercader, Enrique; Ceballos-García, Elena; Navarro-González, Teresa; Moreno-Ochoa, Laura; Perez-Millan, Federico; Pion, Marjorie; Sanchez-Mateos, Paloma; Hidalgo, Andres; Muñoz-Fernandez, Maria A; Relloso, Miguel

    2016-02-01

    Estradiol-based contraceptives and hormonal replacement therapy predispose women to Candida albicans infections. Moreover, during the ovulatory phase (high estradiol), neutrophil numbers decrease in the vaginal lumen and increase during the luteal phase (high progesterone). Vaginal secretions contain chemokines that drive neutrophil migration into the lumen. However, their expression during the ovarian cycle or in response to hormonal treatments are controversial and their role in vaginal defense remains unknown.To investigate the transepithelial migration of neutrophils, we used adoptive transfer of Cxcr2(-/-) neutrophils and chemokine immunofluorescence quantitative analysis in response to C. albicans vaginal infection in the presence of hormones.Our data show that the Cxcl1/Cxcr2 axis drives neutrophil transepithelial migration into the vagina. Progesterone promotes the Cxcl1 gradient to favor neutrophil migration. Estradiol disrupts the Cxcl1 gradient and favors neutrophil arrest in the vaginal stroma; as a result, the vagina becomes more vulnerable to pathogens.

  11. Characterization of neutrophil extracellular traps in cats naturally infected with feline leukemia virus.

    PubMed

    Wardini, Amanda B; Guimarães-Costa, Anderson B; Nascimento, Michelle T C; Nadaes, Natalia R; Danelli, Maria G M; Mazur, Carlos; Benjamim, Claudia F; Saraiva, Elvira M; Pinto-da-Silva, Lucia H

    2010-01-01

    Feline leukemia virus (FeLV), a common, naturally occurring gammaretrovirus in domestic cats, is associated with degenerative diseases of the haematopoietic system, immunodeficiency and neoplasia. FeLV infection causes an important suppression of neutrophil function, leading to opportunistic infections. Recently, a new microbicidal mechanism named NETosis was described in human, bovine and fish neutrophils, as well as in chicken heterophils. The purpose of the present study was to characterize NETosis in feline neutrophils, as well as to evaluate neutrophil function in FeLV naturally infected symptomatic and asymptomatic cats through the phagocytosis process, release of neutrophil extracellular traps (NETs) and myeloperoxidase (MPO) activity. The results showed that feline neutrophils stimulated with protozoa parasites released structures comprising DNA and histones, which were characterized as NETs by immunofluorescence. Quantification of NETs after neutrophil stimulation showed a significant increase in NET release by neutrophils from FeLV(-) and FeLV(+) asymptomatic cats compared with FeLV(+) symptomatic cats. Moreover, the number of released NETs and MPO activity in unstimulated neutrophils of FeLV(+) symptomatic cats were higher than those in unstimulated neutrophils from FeLV(-) and FeLV(+) asymptomatic cats. This study reports, for the first time, NET release by feline neutrophils, along with the fact that NET induction may be modulated by a viral infection. The results indicate that the NET mechanism appears to be overactivated in FeLV(+) cats and that this feature could be considered a marker of disease progression in FeLV infection.

  12. Inflammatory mechanisms and treatment of obstructive airway diseases with neutrophilic bronchitis.

    PubMed

    Simpson, Jodie L; Phipps, Simon; Gibson, Peter G

    2009-10-01

    Obstructive airway diseases such as asthma and chronic obstructive pulmonary disease (COPD) are major global health issues. Although considered as distinct diseases, airway inflammation is a key underlying pathophysiological process in asthma, COPD and bronchiectasis. Persistent neutrophilic airway inflammation (neutrophilic bronchitis) occurs with innate immune activation and is a feature of each of these airway diseases. Little is known about the mechanisms leading to neutrophilic bronchitis and few treatments are effective in reducing neutrophil accumulation in the airways. There is a similar pattern of inflammatory mediator release and toll like receptor 2 expression in asthma, COPD and bronchiectasis. We propose the existence of an active amplification mechanism, an effector arm of the innate immune system, involving toll like receptor 2, operating in persistent neutrophilic bronchitis. Neutrophil persistence in the airways can occur through a number of mechanisms such as impaired apoptosis, efferocytosis and mucus hypersecretion, all of which are impaired in airways disease. Impairment of neutrophil clearance results in a reduced ability to respond to bacterial infection. Persistent activation of airway neutrophils may result in the persistent activation of the innate immune system resulting in further airway insult. Current therapies are limited for the treatment of neutrophilic bronchitis; possible treatments being investigated include theophylline, statins, antagonists of pro-inflammatory cytokines and macrolide antibiotics. Macrolides have shown great promise in their ability to reduce airway inflammation, and can reduce airway neutrophils, levels of CXCL8 and neutrophil proteases in the airways. Studies also show improvements in quality of life and exacerbation rates in airways diseases.

  13. Neutrophil-related factors as biomarkers in EAE and MS

    PubMed Central

    Rumble, Julie M.; Huber, Amanda K.; Krishnamoorthy, Gurumoorthy; Srinivasan, Ashok; Giles, David A.; Zhang, Xu; Wang, Lu

    2015-01-01

    A major function of T helper (Th) 17 cells is to induce the production of factors that activate and mobilize neutrophils. Although Th17 cells have been implicated in the pathogenesis of multiple sclerosis (MS) and the animal model experimental autoimmune encephalomyelitis (EAE), little attention has been focused on the role of granulocytes in those disorders. We show that neutrophils, as well as monocytes, expand in the bone marrow and accumulate in the circulation before the clinical onset of EAE, in response to systemic up-regulation of granulocyte colony-stimulating factor (G-CSF) and the ELR+ CXC chemokine CXCL1. Neutrophils comprised a relatively high percentage of leukocytes infiltrating the central nervous system (CNS) early in disease development. G-CSF receptor deficiency and CXCL1 blockade suppressed myeloid cell accumulation in the blood and ameliorated the clinical course of mice that were injected with myelin-reactive Th17 cells. In relapsing MS patients, plasma levels of CXCL5, another ELR+ CXC chemokine, were elevated during acute lesion formation. Systemic expression of CXCL1, CXCL5, and neutrophil elastase correlated with measures of MS lesion burden and clinical disability. Based on these results, we advocate that neutrophil-related molecules be further investigated as novel biomarkers and therapeutic targets in MS. PMID:25559893

  14. Leukocyte subsets and neutrophil function after short-term spaceflight

    NASA Technical Reports Server (NTRS)

    Stowe, R. P.; Sams, C. F.; Mehta, S. K.; Kaur, I.; Jones, M. L.; Feeback, D. L.; Pierson, D. L.

    1999-01-01

    Changes in leukocyte subpopulations and function after spaceflight have been observed but the mechanisms underlying these changes are not well defined. This study investigated the effects of short-term spaceflight (8-15 days) on circulating leukocyte subsets, stress hormones, immunoglobulin levels, and neutrophil function. At landing, a 1.5-fold increase in neutrophils was observed compared with preflight values; lymphocytes were slightly decreased, whereas the results were variable for monocytes. No significant changes were observed in plasma levels of immunoglobulins, cortisol, or adrenocorticotropic hormone. In contrast, urinary epinephrine, norepinephrine, and cortisol were significantly elevated at landing. Band neutrophils were observed in 9 of 16 astronauts. Neutrophil chemotactic assays showed a 10-fold decrease in the optimal dose response after landing. Neutrophil adhesion to endothelial cells was increased both before and after spaceflight. At landing, the expression of MAC-1 was significantly decreased while L-selectin was significantly increased. These functional alterations may be of clinical significance on long-duration space missions.

  15. Circulation Systems on Microcomputers.

    ERIC Educational Resources Information Center

    Carlson, Gary

    1984-01-01

    Reports on the use of microcomputers in comprehensive library circulation systems. Topics covered include system requirements (reliability, completeness); determining circulation system needs (saving money, improving service, modernization); limitations of microcomputers (capacity, kinds of data stored, number of stations or terminals); system…

  16. Aged neutrophils contribute to the first line of defense in the acute inflammatory response

    PubMed Central

    Uhl, Bernd; Vadlau, Yannick; Zuchtriegel, Gabriele; Nekolla, Katharina; Sharaf, Kariem; Gaertner, Florian; Massberg, Steffen; Krombach, Fritz

    2016-01-01

    Under steady-state conditions, aged neutrophils are removed from the circulation in bone marrow, liver, and spleen, thereby maintaining myeloid cell homeostasis. The fate of these aged immune cells under inflammatory conditions, however, remains largely obscure. Here, we demonstrate that in the acute inflammatory response during endotoxemia, aged neutrophils cease returning to the bone marrow and instead rapidly migrate to the site of inflammation. Having arrived in inflamed tissue, aged neutrophils were found to exhibit a higher phagocytic activity as compared with the subsequently recruited nonaged neutrophils. This distinct behavior of aged neutrophils under inflammatory conditions is dependent on specific age-related changes in their molecular repertoire that enable these “experienced” immune cells to instantly translate inflammatory signals into immune responses. In particular, aged neutrophils engage Toll-like receptor-4- and p38 MAPK-dependent pathways to induce conformational changes in β2 integrins that allow these phagocytes to effectively accomplish their mission in the front line of the inflammatory response. Hence, ageing in the circulation might represent a critical process for neutrophils that enables these immune cells to properly unfold their functional properties for host defense. PMID:27609642

  17. The interaction of Acanthamoeba castellanii cysts with macrophages and neutrophils.

    PubMed

    Hurt, Michael; Proy, Vincent; Niederkorn, Jerry Y; Alizadeh, Hassan

    2003-06-01

    Acanthamoeba castellanii, a free-living amoeba, causes a sight-threatening form of keratitis. Even after extensive therapies, corneal damage can be severe, often requiring corneal transplantation to restore vision. However, A. castellanii cysts are not eliminated from the conjunctiva and stroma of humans and can excyst, resulting in infection of the corneal transplant. The aim of this study was to determine whether elements of the innate immune apparatus, neutrophils and macrophages, were capable of detecting and eliminating A. castellanii cysts and to examine the mechanism by which they kill the cysts. Results show that neither innate immune cell is attracted chemotactically to intact cysts, yet both were attracted to lysed cysts. Both macrophages and neutrophils were capable of killing significant numbers of cysts, yet neutrophils were 3-fold more efficient than macrophages. Activation of macrophages with lipopolysaccharide and interferon-gamma did not increase their cytolytic ability. Conditioned medium isolated from macrophages did not lyse the cysts; however, prevention of phagocytosis by cytochalasin D inhibited 100% of macrophage-mediated killing of the cysts. Conditioned medium from neutrophils did kill significant numbers of the cysts, and this killing was blocked by quercetin, a potent inhibitor of myeloperoxidase (MPO). These results indicate that neither macrophages nor neutrophils are chemoattracted to intact cysts, yet both are capable of killing the cysts. Macrophages killed the cysts by phagocytosis, whereas neutrophils killed cysts through the secretion of MPO.

  18. Two-dimensional numerical models of open-top hydrothermal convection at high Rayleigh and Nusselt numbers: Implications for mid-ocean ridge hydrothermal circulation

    NASA Astrophysics Data System (ADS)

    Fontaine, Fabrice J.; Wilcock, William S. D.

    2007-07-01

    Mid-ocean ridges host vigorous hydrothermal systems that remove large quantities of heat from the oceanic crust. Inferred Nusselt numbers (Nu), which are the ratios of the total heat flux to the heat flux that would be transported by conduction alone, range from 8 to several hundred. Such vigorous convection is not fully described by most numerical models of hydrothermal circulation. A major difficulty arises at high Nu from the numerical solution of the temperature equation. To avoid classical numerical artifacts such as nonphysical oscillatory behavior and artificial diffusion, we implement the Multidimensional Positive Definite Advection Transport Algorithm (MPDATA) technique, which solves the temperature equation using an iterated upwind corrected scheme. We first validate the method by comparing results for models with uniform fluid properties in closed- and open-top systems to existing solutions with Nu ≤ ˜20. We then incorporate realistic fluid properties and run models for Nu up to 50-60. Solutions are characterized by an unstable bottom thermal boundary layer where thermal instabilities arise locally. The pattern of heat extraction is periodic to chaotic. At any Nu > ˜13 the venting temperatures in a given plume are chaotic and oscillate from ˜350° to 450°C. Individual plumes can temporarily stop short of the surface for intervals ranging from tens to hundreds of years at times when other plumes vent with an increased flow rate. The solutions also display significant recirculation, and as a result large areas of downflow are relatively warm with temperatures commonly exceeding 150°C at middepths. Our results have important implications for mid-ocean ridge hydrothermal systems and suggest the following: (1) The reaction zones of mid-ocean ridge hydrothermal systems are enlarged by thermal instabilities that migrate laterally toward upflow zones. This will substantially increase the volume of rock involved in chemical reactions compared to steady

  19. Cytokine-induced neutrophil-derived interleukin-8.

    PubMed Central

    Strieter, R. M.; Kasahara, K.; Allen, R. M.; Standiford, T. J.; Rolfe, M. W.; Becker, F. S.; Chensue, S. W.; Kunkel, S. L.

    1992-01-01

    During acute inflammation, the first line of cellular response for host defense is the neutrophil. In addition to the historic role of the neutrophil as a phagocyte, recent studies have identified this cell as an important source of a number of cytokines. In this study, we provide evidence that the neutrophil is a significant source of interleukin-8 (IL-8). Neutrophils freshly isolated from whole blood were not found to constitutively express IL-8 mRNA. In contrast, when these leukocytes were cultured on plastic they were activated, leading to the significant expression of de novo steady-state levels of IL-8 mRNA. In addition, when neutrophils were treated with cycloheximide, there was evidence for "superinduction" of steady-state levels of IL-8 mRNA and inhibition of antigenic IL-8 production. Neutrophils were subsequently stimulated with lipopolysaccharide (LPS), tumor necrosis factor-alpha, or interleukin-1-beta and were found to express IL-8 mRNA and antigen in both a time- and dose-dependent manner. Furthermore, neutrophils stimulated with traditional chemotactic/activating factors, such as the split product of the fifth component of complement (C5a), formylmethionyleucylphenylalanine (fMLP), and leukotriene B4 (LTB4) in a dose-dependent manner did not produce significant antigenic IL-8, as compared with unstimulated controls. In contrast, when neutrophils were exposed to either of these neutrophil agonists in the presence of LPS, the production of antigenic IL-8 was significantly elevated, as compared with either of the stimuli alone, suggesting a synergistic response. These data would suggest that the neutrophil can no longer be viewed as only a phagocyte or warehouse for proteolytic enzymes, but is a pivotal effector cell that is able to respond to mediators in its environment and generate cytokines. This latter neutrophil response may be important for either the elicitation of additional neutrophils or to orchestrate the conventional immune response at

  20. Neutrophil Extracellular Traps in ANCA-Associated Vasculitis

    PubMed Central

    Söderberg, Daniel; Segelmark, Mårten

    2016-01-01

    A group of pauci-immune vasculitides, characterized by neutrophil-rich necrotizing inflammation of small vessels and the presence of antineutrophil cytoplasmic antibodies (ANCAs), is referred to as ANCA-associated vasculitis (AAV). ANCAs against proteinase 3 (PR3) (PR3-ANCA) or myeloperoxidase (MPO) (MPO-ANCA) are found in over 90% of patients with active disease, and these ANCAs are implicated in the pathogenesis of AAV. Dying neutrophils surrounding the walls of small vessels are a histological hallmark of AAV. Traditionally, it has been assumed that these neutrophils die by necrosis, but neutrophil extracellular traps (NETs) have recently been visualized at the sites of vasculitic lesions. AAV patients also possess elevated levels of NETs in the circulation. ANCAs are capable of inducing NETosis in neutrophils, and their potential to do so has been shown to be affinity dependent and to correlate with disease activity. Neutrophils from AAV patients are also more prone to release NETs spontaneously than neutrophils from healthy blood donors. NETs contain proinflammatory proteins and are thought to contribute to vessel inflammation directly by damaging endothelial cells and by activating the complement system and indirectly by acting as a link between the innate and adaptive immune system through the generation of PR3- and MPO-ANCA. Injection of NET-loaded myeloid dendritic cells into mice results in circulating PR3- and MPO-ANCA and the development of AAV-like disease. NETs have also been shown to be essential in a rodent model of drug-induced vasculitis. NETs induced by propylthiouracil could not be degraded by DNaseI, implying that disordered NETs might be important for the generation of ANCAs. NET degradation was also highlighted in another study showing that AAV patients have reduced DNaseI activity resulting in less NET degradation. With this in mind, it might be that prolonged exposure to proteins in the NETs due to the overproduction of NETs and/or reduced

  1. Role of osteopontin in hepatic neutrophil infiltration during alcoholic steatohepatitis

    SciTech Connect

    Apte, Udayan M.; Banerjee, Atrayee; McRee, Rachel; Wellberg, Elizabeth; Ramaiah, Shashi K. . E-mail: sramaiah@cvm.tamu.edu

    2005-08-22

    Alcoholic liver disease (ALD) is a major complication of heavy alcohol (EtOH) drinking and is characterized by three progressive stages of pathology: steatosis, steatohepatitis, and fibrosis/cirrhosis. Alcoholic steatosis (AS) is the initial stage of ALD and consists of fat accumulation in the liver accompanied by minimal liver injury. AS is known to render the hepatocytes increasingly sensitive to toxicants such as bacterial endotoxin (LPS). Alcoholic steatohepatitis (ASH), the second and rate-limiting step in the progression of ALD, is characterized by hepatic fat accumulation, neutrophil infiltration, and neutrophil-mediated parenchymal injury. However, the pathogenesis of ASH is poorly defined. It has been theorized that the pathogenesis of ASH involves interaction of increased circulating levels of LPS with hepatocytes being rendered highly sensitive to LPS due to heavy EtOH consumption. We hypothesize that osteopontin (OPN), a matricellular protein (MCP), plays an important role in the hepatic neutrophil recruitment due to its enhanced expression during the early phase of ALD (AS and ASH). To study the role of OPN in the pathogenesis of ASH, we induced AS in male Sprague-Dawley rats by feeding EtOH-containing Lieber-DeCarli liquid diet for 6 weeks. AS rats experienced extensive fat accumulation and minimal liver injury. Moderate induction in OPN was observed in AS group. ASH was induced by feeding male Sprague-Dawley rats EtOH-containing Lieber-DeCarli liquid diet for 6 weeks followed by LPS injection. The ASH rats had substantial neutrophil infiltration, coagulative oncotic necrosis, and developed higher liver injury. Significant increases in the hepatic and circulating levels of OPN was observed in the ASH rats. Higher levels of the active, thrombin-cleaved form of OPN in the liver in ASH group correlated remarkably with hepatic neutrophil infiltration. Finally, correlative studies between OPN and hepatic neutrophil infiltration was corroborated in a simple

  2. Type I Interferon Transcriptional Signature in Neutrophils and Low-Density Granulocytes Are Associated with Tissue Damage in Malaria.

    PubMed

    Rocha, Bruno Coelho; Marques, Pedro Elias; Leoratti, Fabiana Maria de Souza; Junqueira, Caroline; Pereira, Dhelio Batista; Antonelli, Lis Ribeiro do Valle; Menezes, Gustavo Batista; Golenbock, Douglas Taylor; Gazzinelli, Ricardo Tostes

    2015-12-29

    Neutrophils are the most abundant leukocyte population in the bloodstream, the primary compartment of Plasmodium sp. infection. However, the role of these polymorphonuclear cells in mediating either the resistance or the pathogenesis of malaria is poorly understood. We report that circulating neutrophils from malaria patients are highly activated, as indicated by a strong type I interferon transcriptional signature, increased expression of surface activation markers, enhanced release of reactive oxygen species and myeloperoxidase, and a high frequency of low-density granulocytes. The activation of neutrophils was associated with increased levels of serum alanine and aspartate aminotransferases, indicating liver damage. In a rodent malaria model, we observed intense recruitment of neutrophils to liver sinusoids. Neutrophil migration and IL-1β and chemokine expression as well as liver damage were all dependent on type I interferon signaling. The data suggest that type I interferon signaling has a central role in neutrophil activation and malaria pathogenesis.

  3. Solar and geomagnetic effects on the frequency of atmospheric circulation types over Europe: an analysis based on a large number of classifications

    NASA Astrophysics Data System (ADS)

    Huth, Radan; Cahynová, Monika; Kyselý, Jan

    2010-05-01

    Recently, effects of the 11-year solar cycle on various aspects of tropospheric circulation in the Northern Hemisphere in winter have been recognized. One of our previous studies showed a significant solar effect on the frequency of synoptic types from the Hess-Brezowsky catalogue. Here, we use a large collection of varied classifications of circulation patterns, assembled within the COST733 Action "Harmonization and Applications of Weather Types Classifications for European Regions" to detect the solar effect on the frequency of synoptic types. The collection contains both objective and subjective classifications. The advantage of this multi-classification approach is that peculiarities or biases of any single classification (catalogue) that might influence the detected solar signal vanish once a large ensemble of classifications is used. We divide winter months (December to March) into three groups according to the mean monthly solar activity, quantified by the F10.7 flux. The three groups correspond to the minima of the 11-year solar cycle, a moderate solar activity, and solar maxima. Within each group, frequencies of occurrence of individual circulation types are calculated. Differences in the occurrence of individual classes between solar activity groups indicate the presence of a solar activity effect on atmospheric circulation over Europe. Statistical significance of these differences is estimated by a block resampling method. The research is supported by the Grant Agency of the Czech Academy of Sciences, project A300420805, and by the Ministry of Education, Youth, and Sports of the Czech Republic, contract OC115.

  4. Mincle activation enhances neutrophil migration and resistance to polymicrobial septic peritonitis

    PubMed Central

    Lee, Wook-Bin; Yan, Ji-Jing; Kang, Ji-Seon; Zhang, Quanri; Choi, Won Young; Kim, Lark Kyun; Kim, Young-Joon

    2017-01-01

    Sepsis is a systemic inflammatory response to bacterial infection. The therapeutic options for treating sepsis are limited. Impaired neutrophil recruitment into the infection site is directly associated with severe sepsis, but the precise mechanism is unclear. Here, we show that Mincle plays a key role in neutrophil migration and resistance during polymicrobial sepsis. Mincle-deficient mice exhibited lower survival rates in experimental sepsis from cecal ligation and puncture and Escherichia coli–induced peritonitis. Mincle deficiency led to higher serum inflammatory cytokine levels and reduced bacterial clearance and neutrophil recruitment. Transcriptome analyses revealed that trehalose dimycolate, a Mincle ligand, reduced the expression of G protein–coupled receptor kinase 2 (GRK2) in neutrophils. Indeed, GRK2 expression was upregulated, but surface expression of the chemokine receptor CXCR2 was downregulated in blood neutrophils from Mincle-deficient mice with septic injury. Moreover, CXCL2-mediated adhesion, chemotactic responses, and F-actin polymerization were reduced in Mincle-deficient neutrophils. Finally, we found that fewer Mincle-deficient neutrophils infiltrated from the blood circulation into the peritoneal fluid in bacterial septic peritonitis compared with wild-type cells. Thus, our results indicate that Mincle plays an important role in neutrophil infiltration and suggest that Mincle signaling may provide a therapeutic target for treating sepsis. PMID:28112221

  5. Neutrophil recruitment by allergens contribute to allergic sensitization and allergic inflammation

    PubMed Central

    Hosoki, Koa; Boldogh, Istvan; Sur, Sanjiv

    2016-01-01

    Purpose of review To discuss the presence and role of neutrophils in asthma and allergic diseases, and outline importance of pollen and cat dander-induced innate neutrophil recruitment in induction of allergic sensitization and allergic inflammation. Recent findings Uncontrolled asthma is associated with elevated numbers of neutrophils, and levels of neutrophil-attracting chemokine IL-8 and IL-17 in BAL fluids. These parameters negatively correlate with lung function. Pollen allergens and cat dander recruit neutrophils to the airways in a TLR4, MD2 and CXCR2-dependent manner. Repeated recruitment of activated neutrophils by these allergens facilitates allergic sensitization and airway inflammation. Inhibition of neutrophil recruitment with CXCR2 inhibitor, disruption of TLR4, or siRNA against MD2 also inhibits allergic inflammation. The molecular mechanisms by which neutrophils shift the inflammatory response of the airways to inhaled allergens to an allergic phenotype is an area of active research. Summary Recent studies have revealed that neutrophil recruitment is important in development of allergic sensitization and inflammation. Inhibition of neutrophils recruitment may be strategy to control allergic inflammation. PMID:26694038

  6. Nitric oxide regulates neutrophil migration through microparticle formation.

    PubMed

    Nolan, Sarah; Dixon, Rachel; Norman, Keith; Hellewell, Paul; Ridger, Victoria

    2008-01-01

    The role of nitric oxide (NO) in regulating neutrophil migration has been investigated. Human neutrophil migration to interleukin (IL)-8 (1 nmol/L) was measured after a 1-hour incubation using a 96-well chemotaxis plate assay. The NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (L-NAME) significantly (P < 0.001) enhanced IL-8-induced migration by up to 45%. Anti-CD18 significantly (P < 0.001) inhibited both IL-8-induced and L-NAME enhanced migration. Antibodies to L-selectin or PSGL-1 had no effect on IL-8-induced migration but prevented the increased migration to IL-8 induced by L-NAME. L-NAME induced generation of neutrophil-derived microparticles that was significantly (P < 0.01) greater than untreated neutrophils or D-NAME. This microparticle formation was dependent on calpain activity and superoxide production. Only microparticles from L-NAME and not untreated or D-NAME-treated neutrophils induced a significant (P < 0.01) increase in IL-8-induced migration and transendothelial migration. Pretreatment of microparticles with antibodies to L-selectin (DREG-200) or PSGL-1 (PL-1) significantly (P < 0.001) inhibited this effect. The ability of L-NAME-induced microparticles to enhance migration was found to be dependent on the number of microparticles produced and not an increase in microparticle surface L-selectin or PSGL-1 expression. These data show that NO can modulate neutrophil migration by regulating microparticle formation.

  7. Tumor associated macrophages and neutrophils in cancer.

    PubMed

    Galdiero, Maria Rosaria; Bonavita, Eduardo; Barajon, Isabella; Garlanda, Cecilia; Mantovani, Alberto; Jaillon, Sébastien

    2013-11-01

    The tumor microenvironment is a complex framework, in which myeloid cells play important roles in sculpting cancer development from tumor initiation to metastasis. Immune cells are key participants of the tumor microenvironment where they can promote or inhibit cancer formation and development. Plasticity is a widely accepted hallmark of myeloid cells and in particular of the monocyte-macrophage lineage. It includes the ability to display a wide spectrum of activation states in response to distinct signals and classical M1 or alternative M2 macrophages represent a paradigm of this feature. Neutrophils have long been viewed as terminally differentiated effector cells, playing a major role during the acute phase of inflammation and resistance against microbes. Recent evidence questioned this limited point of view, indicating that neutrophils can interact with distinct cell populations and produce a wide number of cytokines and effector molecules. Therefore, macrophages and neutrophils are both integrated in the regulation of the innate and adaptive immune responses in various inflammatory situations, including cancer.

  8. Neutrophil in Viral Infections, Friend or Foe?

    PubMed Central

    Drescher, Brandon; Bai, Fengwei

    2012-01-01

    Polymorphonuclear leukocytes or neutrophils are the first immune cells to the site of injury and microbial infection. Neutrophils are crucial players in controlling bacterial and fungal infections, and in particular secondary infections, by phagocytosis, degranulation and neutrophil extracellular traps (NETs). While neutrophils have been shown to play important roles in viral pathogenesis, there is a lack of detailed investigation. In this article, we will review recent progresses toward understanding the role of neutrophils in viral pathogenesis. PMID:23178588

  9. The novel roles of neutrophils via opioid peptides: regulation of the estrous cycle and pain.

    PubMed

    Kobayashi, Yoshiro

    2013-06-01

    Neutrophils are excreted into the vaginal vault at metestrus during the estrous cycle, and this phenomenon has long been used to determine the phase of the estrous cycle. A much smaller number of neutrophils are also detected in the uterus and the ovary. Recently, we provided several lines of evidence supporting the notion that neutrophils infiltrate into the ovary to regulate the estrous cycle by opioid peptides. Upon inflammation, on the other hand, neutrophils infiltrate into the site of infection to suppress pain by opioid peptides. Thus, opioid peptides are key molecules by which neutrophils play a novel role in regulation of the pain and estrous cycle. In both cases, opioid peptides appear to be secreted by neutrophils stimulated with chemokines, such as MIP-2 and KC in mouse, corticotropin-releasing hormone and IL-1.

  10. Cell Intrinsic Galectin-3 Attenuates Neutrophil ROS-Dependent Killing of Candida by Modulating CR3 Downstream Syk Activation

    PubMed Central

    Wu, Sheng-Yang; Huang, Juin-Hua; Chen, Wen-Yu; Chan, Yi-Chen; Lin, Chun-Hung; Chen, Yee-Chun; Liu, Fu-Tong; Wu-Hsieh, Betty A.

    2017-01-01

    Invasive candidiasis is a leading cause of nosocomial bloodstream infection. Neutrophils are the important effector cells in host resistance to candidiasis. To investigate the modulation of neutrophil fungicidal function will advance our knowledge on the control of candidiasis. While recombinant galectin-3 enhances neutrophil phagocytosis of Candida, we found that intracellular galectin-3 downregulates neutrophil fungicidal functions. Co-immunoprecipitation and immunofluorescence staining reveal that cytosolic gal3 physically interacts with Syk in neutrophils after Candida stimulation. Gal3−/− neutrophils have higher level of Syk activation as well as greater abilities to generate reactive oxygen species (ROS) and kill Candida than gal3+/+ cells. While galectin-3 deficiency modulates neutrophil and macrophage activation and the recruitment of monocytes and dendritic cells, the deficiency does not affect the numbers of infiltrating neutrophils or macrophages. Galectin-3 deficiency ameliorates systemic candidiasis by reducing fungal burden, renal pathology, and mortality. Adoptive transfer experiments demonstrate that cell intrinsic galectin-3 negatively regulates neutrophil effector functions against candidiasis. Reducing galectin-3 expression or activity by siRNA or gal3 inhibitor TD139 enhances human neutrophil ROS production. Mice treated with TD139 have enhanced ability to clear the fungus. Our work unravels the mechanism by which galectin-3 regulates Syk-dependent neutrophil fungicidal functions and raises the possibility that blocking gal3 in neutrophils may be a promising therapeutic strategy for treating systemic candidiasis. PMID:28217127

  11. Comparative Efficiency and Impact on the Activity of Blood Neutrophils Isolated by Percoll, Ficoll and Spontaneous Sedimentation Methods.

    PubMed

    Mosca, Tainá; Forte, Wilma C N

    2016-01-01

    Studies on the role of cells in physiological and pathological processes generally require isolation of some populations, such as neutrophils. In the literature, several methods used for isolating neutrophils are described; however, there is no consensus on the best technique to be used in cell functional studies. The present study compares the efficiency and impact on the chemotactic and phagocytic activity of neutrophils isolated from blood by three different methods: Percoll and Ficoll density centrifugation gradients and spontaneous sedimentation technique. The neutrophil chemotaxis, stimulated with lipopolysaccharide (LPS), autologous serum or homologous serum, was determined by using Boyden chambers. The phagocytic capacity was assessed by ingestion of zimosan particles, and digestion phase was analyzed by nitroblue tetrazolium test (NBT). The results obtained from neutrophil isolation by Percoll and Ficoll density gradients, as compared to spontaneous sedimentation technique, showed similar degrees of cell yields and higher purity; however, these methods affected neutrophil responsiveness, accompanied by elevated chemotaxis and reduced chemotactic capacity to respond to subsequent stimulation. Neutrophil isolation by spontaneous sedimentation, in contrast, did not affect cellular activity and resulted in cell preparation with high number of neutrophils. Although neutrophil phagocytosis results were similar between the different methods, digestion phase of phagocytosis was significantly enhanced after LPS-stimulation, only in the neutrophils isolated by spontaneous sedimentation technique. In conclusion, the present study shows that isolation of blood neutrophils by the spontaneous sedimentation technique is appropriate for the assessment of cellular activity, since it neither primes or activates the neutrophils nor does it affect their functional responsiveness.

  12. AUTOINFLAMMATORY PUSTULAR NEUTROPHILIC DISEASES

    PubMed Central

    Naik, Haley B.; Cowen, Edward W.

    2013-01-01

    SYNOPSIS The inflammatory pustular dermatoses constitute a spectrum of non-infectious conditions ranging from localized involvement to generalized disease with associated acute systemic inflammation and multi-organ involvement. Despite the variability in extent and severity of cutaneous presentation, each of these diseases is characterized by non-infectious neutrophilic intra-epidermal microabscesses. Many share systemic findings including fever, elevated inflammatory markers, inflammatory bowel disease and/or osteoarticular involvement, suggesting potential common pathogenic links (Figure 1). The recent discoveries of several genes responsible for heritable pustular diseases have revealed a distinct link between pustular skin disease and regulation of innate immunity. These genetic advances have led to a deeper exploration of common pathways in pustular skin disease and offer the potential for a new era of biologic therapy which targets these shared pathways. This chapter provides a new categorization of inflammatory pustular dermatoses in the context of recent genetic and biologic insights. We will discuss recently-described monogenic diseases with pustular phenotypes, including deficiency of IL-1 receptor antagonist (DIRA), deficiency of the IL-36 receptor antagonist (DITRA), CARD14-associated pustular psoriasis (CAMPS), and pyogenic arthritis, pyoderma gangrenosum, acne (PAPA). We will then discuss how these new genetic advancements may inform how we view previously described pustular diseases, including pustular psoriasis and its clinical variants, with a focus on historical classification by clinical phenotype. PMID:23827244

  13. Nitrite is produced by elicited but not by circulating neutrophils

    PubMed Central

    Dusting, G. J.; Giarracca, R. G.; Harris, T.; Lim, Y.; Sobey, C. G.

    1993-01-01

    The generation of nitrite (NO2-) was used as an index of the production of nitric oxide by human and rat polymorphonuclear leukocytes (PMN) and rat peritoneal macrophages. Human peripheral blood PMN did not produce significant levels of NO2-. Attempts to induce NO2- generation in human PMN by incubation with GM–CSF (1 nM), TNFα (0.3 nM), endotoxin (1 μg/ml) or formyl-Met-Leu-Phe (100 nM) for up to 16 h were not successful. Addition of human PMN primed by GM–CSF (1 nM) to rabbit aortic ring preparations precontracted with phenylephrine had no effect on tone. In contrast to these observations, PMN, isolated from the peritoneum of oyster glycogen treated rats, generated NO2- via a pathway sensitive to inhibition by the nitric oxide synthase inhibitor, NG-monomethyl L-arginine. However, peripheral blood rat PMN obtained from the same animals did not produce NO2-, even during prolonged incubation for periods of up to 16 h. It is suggested that detectable NO production by PMN requires NO synthase activity to be induced either by the process of PMN migration or by exposure to certain cytokines produced locally at the site of inflammation. PMID:18475544

  14. Effect of exhaustive exercise on human neutrophils in athletes.

    PubMed

    Yamada, M; Suzuki, K; Kudo, S; Totsuka, M; Simoyama, T; Nakaji, S; Sugawara, K

    2000-01-01

    In order to investigate the effect of exercise on the capacity of neutrophils to produce reactive oxygen species (ROS), eight male cross-country skiers underwent maximal exercise. Peripheral blood samples were taken pre-exercise, 0 h, 1 h, and 2 h after finishing maximal exercise. Leukocyte counts significantly increased (p < 0. 05), particularly lymphocytes (p < 0.05), just after the exercise period (0 h) and significantly increased again (p < 0.05), particularly neutrophils (p < 0.05), 2 h after the exercise compared with pre-exercise values. The capacity of isolated neutrophils to produce ROS was assessed by lucigenin (Lg)-dependent chemiluminescence (CL) and luminol (Lm)-dependent CL on stimulation with opsonized zymosan (OZ) and phorbol myristate acetate (PMA). Just after exercise, the LgCL response was not affected, while the response of LmCL mixed with sodium azide, which inhibits catalase and myeloperoxidase (MPO) activity, was significantly enhanced (p < 0.05). In addition, just after exercise, the level of serum growth hormone increased significantly (p < 0.05). The serum cortisol level also increased significantly just after and 1 h after exercise (p < 0.05). These data indicated that maximal exercise not only mobilized neutrophils from marginated pools into the circulation, but also caused increased ROS generation.

  15. Neutrophil alkaline phosphatase score in chronic granulocytic leukaemia: effects of splenectomy and antileukaemic drugs.

    PubMed Central

    Spiers, A S; Liew, A; Baikie, A G

    1975-01-01

    Staining with naphthol AS phosphate and Fast Blue BB salt has been used for the estimation of neutrophil alkaline phosphatase (NAP) scores in patients with chronic granulocytic leukaemia (CGL). The very low scores found at diagnosis rise when the disease is treated, and there is some inverse correlation between the NAP score and the absolute neutrophil count. Patients treated intensively developed high NAP scores. Elective splenectomy performed during the chronic phase of CGL is followed by a pronounced but transient neutrophilia and a concurrent striking rise in the NAP score. Similar changes were observed in patients without CGL who underwent splenectomy. These observations can be explained by assuming that newly formed neutrophils in CGL have a normal content of NAP but are rapidly sequestered in non-circulating extramedullary pools, whereas the circulating neutrophil with a typically low NAP content is a relatively aged cell which has lost enzyme activity. In subjects with or without CGL, removal of the spleen, a major site of such pooling, temporarily permits the circulation of newly formed neutrophils but eventually other organs assume the sequestering functions of the spleen. Thus the aberrations of NAP score seen in CGL might be attributable not to an intrinsic cellular defect but to an exaggeration of the granulocyte storage phenomena which also occur in subjects without CGL. PMID:1056940

  16. Whole Blood Human Neutrophil Trafficking in a Microfluidic Model of Infection and Inflammation

    PubMed Central

    Hamza, Bashar; Irimia, Daniel

    2015-01-01

    Appropriate inflammatory responses to wounds and infections require adequate numbers of neutrophils arriving at injury sites. Both insufficient and excessive neutrophil recruitment can be detrimental, favouring systemic spread of microbes or triggering severe tissue damage. Despite its importance in health and disease, the trafficking of neutrophils through tissues remains difficult to control and the mechanisms regulating it are insufficiently understood. These mechanisms are also complex and difficult to isolate using traditional in vivo models. Here we designed a microfluidic model of tissue infection/inflammation, in which human neutrophils emerge from a droplet-size samples of whole blood and display bi-directional traffic between this and micro-chambers containing chemoattractant and microbe-like particles. Two geometrical barriers restrict the entrance of red blood cells from the blood to the micro-chambers and simulate the mechanical function of the endothelial barrier separating the cells in blood from cells in tissues. We found that in the presence of chemoattractant, the number of neutrophils departing the chambers by retrotaxis is in dynamic equilibrium with the neutrophils recruited by chemotaxis. We also found that in the presence of microbe-like particles, the number of neutrophils trapped in the chambers is proportional to the number of particles. Together, the dynamic equilibrium between migration, reversed-migration and trapping processes determine the optimal number of neutrophils at a site. These neutrophils are continuously refreshed and responsive to the number of microbes. Further studies using this infection-inflammation-on-a-chip-model could help study the processes of inflammation resolution. The new in vitro experimental tools may also eventually help testing new therapeutic strategies to limit neutrophil accumulation in tissues during chronic inflammation, without increasing the risk for infections. PMID:25987163

  17. Clostridium perfringens α-Toxin Impairs Innate Immunity via Inhibition of Neutrophil Differentiation

    PubMed Central

    Takehara, Masaya; Takagishi, Teruhisa; Seike, Soshi; Ohtani, Kaori; Kobayashi, Keiko; Miyamoto, Kazuaki; Shimizu, Tohru; Nagahama, Masahiro

    2016-01-01

    Although granulopoiesis is accelerated to suppress bacteria during infection, some bacteria can still cause life-threatening infections, but the mechanism behind this remains unclear. In this study, we found that mature neutrophils in bone marrow cells (BMCs) were decreased in C. perfringens-infected mice and also after injection of virulence factor α-toxin. C. perfringens infection interfered with the replenishment of mature neutrophils in the peripheral circulation and the accumulation of neutrophils at C. perfringens-infected sites in an α-toxin-dependent manner. Measurements of bacterial colony-forming units in C. perfringens-infected muscle revealed that α-toxin inhibited a reduction in the load of C. perfringens. In vitro treatment of isolated BMCs with α-toxin (phospholipase C) revealed that α-toxin directly decreased mature neutrophils. α-Toxin did not influence the viability of isolated mature neutrophils, while simultaneous treatment of BMCs with granulocyte colony-stimulating factor attenuated the reduction of mature neutrophils by α-toxin. Together, our results illustrate that impairment of the innate immune system by the inhibition of neutrophil differentiation is crucial for the pathogenesis of C. perfringens to promote disease to a life-threatening infection, which provides new insight to understand how pathogenic bacteria evade the host immune system. PMID:27306065

  18. Neutrophil Extracellular Traps Go Viral

    PubMed Central

    Schönrich, Günther; Raftery, Martin J.

    2016-01-01

    Neutrophils are the most numerous immune cells. Their importance as the first line of defense against bacterial and fungal pathogens is well described. In contrast, the role of neutrophils in controlling viral infections is less clear. Bacterial and fungal pathogens can stimulate neutrophils extracellular traps (NETs) in a process called NETosis. Although NETosis has previously been described as a special form of programmed cell death, there are forms of NET production that do not end with the demise of neutrophils. As an end result of NETosis, genomic DNA complexed with microbicidal proteins is expelled from neutrophils. These structures can kill pathogens or at least prevent their local spread within host tissue. On the other hand, disproportionate NET formation can cause local or systemic damage. Only recently, it was recognized that viruses can also induce NETosis. In this review, we discuss the mechanisms by which NETs are produced in the context of viral infection and how this may contribute to both antiviral immunity and immunopathology. Finally, we shed light on viral immune evasion mechanisms targeting NETs. PMID:27698656

  19. Characterization of canine neutrophil granules.

    PubMed Central

    O'Donnell, R T; Andersen, B R

    1982-01-01

    The purpose of this study was to isolate distinct populations of canine neutrophil granules and to compare them with neutrophil granules from other species. Size, shape, density, and content of canine neutrophil granules were determined. Neutrophils obtained by Ficoll-Hypaque sedimentation were homogenized, and granule populations were separated by isopycnic centrifugation on a linear sucrose gradient (rho, 1.14 to 1.22 g/ml). The most dense granule population (rho, 1.197 g/ml) contained all of the myeloperoxidase, beta-glucuronidase, and elastase, more than half of the acid beta-glycerophosphatase, and most of the lysozyme. The population with intermediate density (rho, 1.179 g/ml) contained lactoferrin, vitamin B12-binding protein, and the remainder of the acid beta-glycerophosphatase and lysozyme. The least dense granule population did not contain a major peak of any of the enzymes or binding proteins tested but was distinguished by density and morphology. The size and shape of the granules were determined from scanning electron micrographs and assessment of shape was aided by transmission electron micrographs. By these methods three populations of canine neutrophil granules were characterized and named: myeloperoxidase granules, vitamin B12-binding protein granules, and low-density granules. Images PMID:6292095

  20. Enhanced neutrophil longevity and recruitment contribute to the severity of oviduct pathology during Chlamydia muridarum infection.

    PubMed

    Frazer, Lauren C; O'Connell, Catherine M; Andrews, Charles W; Zurenski, Matthew A; Darville, Toni

    2011-10-01

    Our previous studies revealed that intravaginal infection of mice with a plasmid-deficient strain of Chlamydia muridarum, CM3.1, does not induce the development of oviduct pathology. In this study, we determined that infection with CM3.1 resulted in a significantly reduced frequency and absolute number of neutrophils in the oviducts during acute infection. This reduction in neutrophils was associated with significantly lower levels of neutrophil chemokines in the oviducts and decreased production of neutrophil chemokines by oviduct epithelial cells infected with CM3.1 in vitro. Infection with CM3.1 also resulted in an increased frequency of late apoptotic/dead neutrophils in the oviduct. Examination of the ability of Chlamydia trachomatis to prevent neutrophil apoptosis in vitro revealed that C. trachomatis strain D/UW-3/Cx exhibited an enhanced ability to prevent neutrophil apoptosis compared to plasmid-deficient CTD153, and this effect was dependent on the presence of CD14(high) monocytes. The presence of monocytes also resulted in enhanced neutrophil cytokine production and increased production of tissue-damaging molecules in response to D/UW-3/Cx relative to results with CTD153. Attempts to use antibody-mediated depletion to discern the specific role of neutrophils in infection control and pathology in vivo revealed that although Ly6G(high) neutrophils were eliminated from the blood and oviducts with this treatment, immature neutrophils and high levels of tissue-damaging molecules were still detectable in the upper genital tract. These data support the role of neutrophils in chlamydia-induced pathology and reveal that novel methods of depletion must be developed before their role can be specifically determined in vivo.

  1. Lutzomyia longipalpis saliva drives apoptosis and enhances parasite burden in neutrophils.

    PubMed

    Prates, Deboraci Brito; Araújo-Santos, Théo; Luz, Nívea Farias; Andrade, Bruno B; França-Costa, Jaqueline; Afonso, Lilian; Clarêncio, Jorge; Miranda, José Carlos; Bozza, Patrícia T; Dosreis, George A; Brodskyn, Cláudia; Barral-Netto, Manoel; Borges, Valéria Matos; Borges, Valéria de Matos; Barral, Aldina

    2011-09-01

    Neutrophils are considered the host's first line of defense against infections and have been implicated in the immunopathogenesis of Leishmaniasis. Leishmania parasites are inoculated alongside vectors' saliva, which is a rich source of pharmacologically active substances that interfere with host immune response. In the present study, we tested the hypothesis that salivary components from Lutzomyia longipalpis, an important vector of visceral Leishmaniasis, enhance neutrophil apoptosis. Murine inflammatory peritoneal neutrophils cultured in the presence of SGS presented increased surface expression of FasL and underwent caspase-dependent and FasL-mediated apoptosis. This proapoptosis effect of SGS on neutrophils was abrogated by pretreatment with protease as well as preincubation with antisaliva antibodies. Furthermore, in the presence of Leishmania chagasi, SGS also increased apoptosis on neutrophils and increased PGE(2) release and decreased ROS production by neutrophils, while enhancing parasite viability inside these cells. The increased parasite burden was abrogated by treatment with z-VAD, a pan caspase inhibitor, and NS-398, a COX-2 inhibitor. In the presence of SGS, Leishmania-infected neutrophils produced higher levels of MCP-1 and attracted a high number of macrophages by chemotaxis in vitro assays. Both of these events were abrogated by pretreatment of neutrophils with bindarit, an inhibitor of CCL2/MCP-1 expression. Taken together, our data support the hypothesis that vector salivary proteins trigger caspase-dependent and FasL-mediated apoptosis, thereby favoring Leishmania survival inside neutrophils, which may represent an important mechanism for the establishment of Leishmania infection.

  2. The interactions of human neutrophils with the constituents of an experimental dental biofilm.

    PubMed

    Shapira, L; Tepper, P; Steinberg, D

    2000-10-01

    Despite the antibacterial properties of neutrophils, their ability to prevent colonization of the dental biofilm by pathogenic bacteria is limited. The present study examined the ability of human neutrophils to attach to an experimental dental biofilm and tested their antibacterial functions following adhesion. Neutrophil adhesion was greatest to hydroxyapatite (HA) in the absence of biofilm. Among the biofilms, glucosyltransferase or fructosyltransferase adsorbed onto saliva-coated HA showed the highest adhesion of cells. The adhesion of neutrophils was directly related to their initial concentration in the solution and to the duration of incubation. Plasma was found to reduce neutrophil attachment significantly, while stimulation of the cells had no effect. Stimulation of attached neutrophils induced superoxide secretion with levels significantly lower than that secreted by suspended cells. The presence of neutrophils on the biofilm reduced the number and the viability of Streptococcus mutans attached to the beads. The present findings suggest that neutrophils are able to attach to dental biofilms and that the attached neutrophils retained their antibacterial activity.

  3. Role of platelets, neutrophils, and factor XII in spontaneous venous thrombosis in mice

    PubMed Central

    Heestermans, Marco; Salloum-Asfar, Salam; Salvatori, Daniela; Laghmani, El Houari; Luken, Brenda M.; Zeerleder, Sacha S.; Spronk, Henri M. H.; Korporaal, Suzanne J.; Wagenaar, Gerry T. M.; Reitsma, Pieter H.

    2016-01-01

    Recently, platelets, neutrophils, and factor XII (FXII) have been implicated as important players in the pathophysiology of venous thrombosis. Their role became evident in mouse models in which surgical handling was used to provoke thrombosis. Inhibiting anticoagulation in mice by using small interfering RNA (siRNA) targeting Serpinc1 and Proc also results in a thrombotic phenotype, which is spontaneous (no additional triggers) and reproducibly results in clots in the large veins of the head and fibrin deposition in the liver. This thrombotic phenotype is fatal but can be fully rescued by thrombin inhibition. The mouse model was used in this study to investigate the role of platelets, neutrophils, and FXII. After administration of siRNAs targeting Serpinc1 and Proc, antibody-mediated depletion of platelets fully abrogated the clinical features as well as microscopic aspects in the head. This was corroborated by strongly reduced fibrin deposition in the liver. Whereas neutrophils were abundant in siRNA-triggered thrombotic lesions, antibody-mediated depletion of circulating Ly6G-positive neutrophils did not affect onset, severity, or thrombus morphology. In addition, absence of circulating neutrophils did not affect quantitative liver fibrin deposition. Remarkably, siRNA-mediated depletion of plasma FXII accelerated the onset of the clinical phenotype; mice were affected with more severe thrombotic lesions. To summarize, in this study, onset and severity of the thrombotic phenotype are dependent on the presence of platelets but not circulating neutrophils. Unexpectedly, FXII has a protective effect. This study challenges the proposed roles of neutrophils and FXII in venous thrombosis pathophysiology. PMID:26932804

  4. Neutrophils and Granulocytic MDSC: The Janus God of Cancer Immunotherapy

    PubMed Central

    Zilio, Serena; Serafini, Paolo

    2016-01-01

    Neutrophils are the most abundant circulating blood cell type in humans, and are the first white blood cells recruited at the inflammation site where they orchestrate the initial immune response. Although their presence at the tumor site was recognized in the 1970s, until recently these cells have been neglected and considered to play just a neutral role in tumor progression. Indeed, in recent years neutrophils have been recognized to play a dual role in tumor development by either assisting the growth, angiogenesis, invasion, and metastasis or by exerting tumoricidal action directly via the secretion of antitumoral compounds, or indirectly via the orchestration of antitumor immunity. Understanding the biology of these cells and influencing their polarization in the tumor micro- and macro-environment may be the key for the development of new therapeutic strategies, which may finally hold the promise of an effective immunotherapy for cancer. PMID:27618112

  5. Cigarette smoke-induced damage-associated molecular pattern release from necrotic neutrophils triggers proinflammatory mediator release.

    PubMed

    Heijink, Irene H; Pouwels, Simon D; Leijendekker, Carin; de Bruin, Harold G; Zijlstra, G Jan; van der Vaart, Hester; ten Hacken, Nick H T; van Oosterhout, Antoon J M; Nawijn, Martijn C; van der Toorn, Marco

    2015-05-01

    Cigarette smoking, the major causative factor for the development of chronic obstructive pulmonary disease, is associated with neutrophilic airway inflammation. Cigarette smoke (CS) exposure can induce a switch from apoptotic to necrotic cell death in airway epithelium. Therefore, we hypothesized that CS promotes neutrophil necrosis with subsequent release of damage-associated molecular patterns (DAMPs), including high mobility group box 1 (HMGB1), alarming the innate immune system. We studied the effect of smoking two cigarettes on sputum neutrophils in healthy individuals and of 5-day CS or air exposure on neutrophil counts, myeloperoxidase, and HMGB1 levels in bronchoalveolar lavage fluid of BALB/c mice. In human peripheral blood neutrophils, mitochondrial membrane potential, apoptosis/necrosis markers, caspase activity, and DAMP release were studied after CS exposure. Finally, we assessed the effect of neutrophil-derived supernatants on the release of chemoattractant CXCL8 in normal human bronchial epithelial cells. Cigarette smoking caused a significant decrease in sputum neutrophil numbers after 3 hours. In mice, neutrophil counts were significantly increased 16 hours after repeated CS exposure but reduced 2 hours after an additional exposure. In vitro, CS induced necrotic neutrophil cell death, as indicated by mitochondrial dysfunction, inhibition of apoptosis, and DAMP release. Supernatants from CS-treated neutrophils significantly increased the release of CXCL8 in normal human bronchial epithelial cells. Together, these observations show, for the first time, that CS exposure induces neutrophil necrosis, leading to DAMP release, which may amplify CS-induced airway inflammation by promoting airway epithelial proinflammatory responses.

  6. Neutrophil adhesion and activation under flow

    PubMed Central

    Zarbock, Alexander; Ley, Klaus

    2009-01-01

    Neutrophil recruitment into inflamed tissue in response to injury or infection is tightly regulated. Reduced neutrophil recruitment can result in a reduced ability to fight invading microorganisms. During inflammation, neutrophils roll along the endothelial wall of postcapillary venules and integrate inflammatory signals. Neutrophil activation by selectins and chemokines regulates integrin adhesiveness. Binding of activated integrins to their counter-receptors on endothelial cells induces neutrophil arrest and firm adhesion. Adherent neutrophils can be further activated to undergo cytoskeletal rearrangement, crawling, transmigration, superoxide production and respiratory burst. Signaling through G-protein coupled receptors, selectin ligands, Fc receptors and outside-in signaling of integrins are all involved in neutrophil activation, but their interplay in the multistep process of recruitment are only beginning to emerge. This review provides an overview of signaling in rolling and adherent neutrophils. PMID:19037827

  7. Isolation and Functional Analysis of Human Neutrophils.

    PubMed

    Kuhns, Douglas B; Long Priel, Debra A; Chu, Jessica; Zarember, Kol A

    2015-11-02

    This unit describes the isolation of human polymorphonuclear neutrophils (PMN) from blood using dextran sedimentation and Percoll or Ficoll-Paque density gradients. Assays of neutrophil functions including respiratory burst activation, phagocytosis, and microbial killing are also described.

  8. Isolation and Functional Analysis of Human Neutrophils

    PubMed Central

    Kuhns, Douglas B.; Long Priel, Debra A.; Chu, Jessica; Zarember, Kol A.

    2015-01-01

    This unit describes the isolation of human polymorphonuclear neutrophils (PMN) from blood using dextran sedimentation and Percoll or Ficoll-Paque density gradients. Assays of neutrophil functions including respiratory burst activation, phagocytosis, and microbial killing are also described. PMID:26528633

  9. Neutrophil cell surface receptors and their intracellular signal transduction pathways☆

    PubMed Central

    Futosi, Krisztina; Fodor, Szabina; Mócsai, Attila

    2013-01-01

    Neutrophils play a critical role in the host defense against bacterial and fungal infections, but their inappropriate activation also contributes to tissue damage during autoimmune and inflammatory diseases. Neutrophils express a large number of cell surface receptors for the recognition of pathogen invasion and the inflammatory environment. Those include G-protein-coupled chemokine and chemoattractant receptors, Fc-receptors, adhesion receptors such as selectins/selectin ligands and integrins, various cytokine receptors, as well as innate immune receptors such as Toll-like receptors and C-type lectins. The various cell surface receptors trigger very diverse signal transduction pathways including activation of heterotrimeric and monomeric G-proteins, receptor-induced and store-operated Ca2 + signals, protein and lipid kinases, adapter proteins and cytoskeletal rearrangement. Here we provide an overview of the receptors involved in neutrophil activation and the intracellular signal transduction processes they trigger. This knowledge is crucial for understanding how neutrophils participate in antimicrobial host defense and inflammatory tissue damage and may also point to possible future targets of the pharmacological therapy of neutrophil-mediated autoimmune or inflammatory diseases. PMID:23994464

  10. Neutrophil biomarkers predict response to therapy with tumor necrosis factor inhibitors in rheumatoid arthritis.

    PubMed

    Wright, Helen L; Cox, Trevor; Moots, Robert J; Edwards, Steven W

    2017-03-01

    Neutrophils are implicated in the pathology of rheumatoid arthritis (RA), but the mechanisms regulating their activation are largely unknown. RA is a heterogeneous disease, and whereas many patients show clinical improvement during TNF inhibitor (TNFi) therapy, a significant proportion fails to respond. In vitro activation of neutrophils with agents, including TNF, results in rapid and selective changes in gene expression, but how neutrophils contribute to TNF signaling in RA and whether TNFi sensitivity involves differential neutrophil responses are unknown. With the use of RNA sequencing (RNA-Seq), we analyzed blood neutrophils from 20 RA patients, pre-TNFi therapy, to identify biomarkers of response, measured by a decrease in disease activity score based on 28 joint count (DAS28), 12 wk post-therapy. Biomarkers were validated by quantitative PCR (qPCR) of blood neutrophils from 2 further independent cohorts of RA patients: 16 pre-TNFi and 16 predisease-modifying anti-rheumatic drugs (DMARDs). Twenty-three neutrophil transcripts predicted a 12-wk response to TNFi: 10 (IFN-regulated) genes predicting a European League against Rheumatism (EULAR) good response and 13 different genes [neutrophil granule protein (NGP) genes] predicting a nonresponse. Statistical analysis indicated a predictive sensitivity and specificity of each gene in the panel of >80%, with some 100% specific. A combination of 3 genes [cytidine monophosphate kinase 2 (CMPK2), IFN-induced protein with tetratricopeptide repeats 1B (IFIT1B), and RNASE3] had the greatest predictive power [area under the curve (AUC) 0.94]. No correlation was found for a response to DMARDs. We conclude that this panel of genes is selective for predicting a response to TNFi and is not a surrogate marker for disease improvement. We also show that in RA, there is great plasticity in neutrophil phenotype, with circulating cells expressing genes normally only expressed in more immature cells.

  11. Neutrophil activation during acetaminophen hepatotoxicity and repair in mice and humans

    SciTech Connect

    Williams, C. David; Bajt, Mary Lynn; Sharpe, Matthew R.; McGill, Mitchell R.; Farhood, Anwar; Jaeschke, Hartmut

    2014-03-01

    Following acetaminophen (APAP) overdose there is an inflammatory response triggered by the release of cellular contents from necrotic hepatocytes into the systemic circulation which initiates the recruitment of neutrophils into the liver. It has been demonstrated that neutrophils do not contribute to APAP-induced liver injury, but their role and the role of NADPH oxidase in injury resolution are controversial. C57BL/6 mice were subjected to APAP overdose and neutrophil activation status was determined during liver injury and liver regeneration. Additionally, human APAP overdose patients (ALT: > 800 U/L) had serial blood draws during the injury and recovery phases for the determination of neutrophil activation. Neutrophils in the peripheral blood of mice showed an increasing activation status (CD11b expression and ROS priming) during and after the peak of injury but returned to baseline levels prior to complete injury resolution. Hepatic sequestered neutrophils showed an increased and sustained CD11b expression, but no ROS priming was observed. Confirming that NADPH oxidase is not critical to injury resolution, gp91{sup phox}−/− mice following APAP overdose displayed no alteration in injury resolution. Peripheral blood from APAP overdose patients also showed increased neutrophil activation status after the peak of liver injury and remained elevated until discharge from the hospital. In mice and humans, markers of activation, like ROS priming, were increased and sustained well after active liver injury had subsided. The similar findings between surviving patients and mice indicate that neutrophil activation may be a critical event for host defense or injury resolution following APAP overdose, but not a contributing factor to APAP-induced injury. - Highlights: • Neutrophil (PMN) function increases during liver repair after acetaminophen overdose. • Liver repair after acetaminophen (APAP)-overdose is not dependent on NADPH oxidase. • Human PMNs do not appear

  12. Neutrophil apoptosis in the lung after hemorrhage or endotoxemia: apoptosis and migration are independent of IL-1beta.

    PubMed

    Parsey, M V; Kaneko, D; Shenkar, R; Abraham, E

    1999-05-01

    Hemorrhage and endotoxemia are associated with neutrophil accumulation in the lungs and the development of acute inflammatory lung injury. Because alterations in the rate of apoptosis may affect the number and function of neutrophils in the lungs, we determined the percentage of neutrophils undergoing apoptosis in the lungs of control, hemorrhaged, or endotoxemic mice. In control mice, 18.5 +/- 1.2% of pulmonary neutrophils were apoptotic. The proportion of apoptotic neutrophils in the lungs was significantly decreased 1 h after hemorrhage (6.5 +/- 1.6%, P < 0.01 compared to control) or endotoxemia (7.0 +/- 0.9%, P < 0.01 compared to control). Between 1 and 24 h after endotoxemia or hemorrhage, the proportion of apoptotic neutrophils in the lung remained significantly depressed compared to that in control, unmanipulated mice. By 48 h, the proportion of apoptotic neutrophils returned to baseline levels in the lungs of hemorrhaged (21.4 +/- 1.4%) or endotoxemic (16.4 +/- 1. 6%) mice. Lung neutrophil IL-1beta mRNA was significantly increased from that of control mice [i.e., 0.12 +/- 0.06 relative absorbance units (RAU)] 1 h after hemorrhage (5.19 +/- 0.068 RAU, P < 0.05 compared to control) or endotoxemia (8.90 +/- 1.53 RAU, P < 0.01 compared to control). In IL-1beta-deficient mice, there was no significant difference in lung neutrophil apoptosis or neutrophil entry into the lung after hemorrhage or endotoxemia compared to wild-type mice. Our results show that apoptosis among lung neutrophils is decreased for more than 24 h after hemorrhage or endotoxemia. Although IL-1beta expression is increased in lung neutrophils under these conditions, IL-1beta is not responsible for either the influx of neutrophils into the lung or the reduction of apoptosis in neutrophil populations after hemorrhage or endotoxemia.

  13. Mechanisms of interferon-γ production by neutrophils and its function during Streptococcus pneumoniae pneumonia.

    PubMed

    Gomez, John C; Yamada, Mitsuhiro; Martin, Jessica R; Dang, Hong; Brickey, W June; Bergmeier, Wolfgang; Dinauer, Mary C; Doerschuk, Claire M

    2015-03-01

    Bacterial pneumonia is a common public health problem associated with significant mortality, morbidity, and cost. Neutrophils are usually the earliest leukocytes to respond to bacteria in the lungs. Neutrophils rapidly sequester in the pulmonary microvasculature and migrate into the lung parenchyma and alveolar spaces, where they perform numerous effector functions for host defense. Previous studies showed that migrated neutrophils produce IFN-γ early during pneumonia induced by Streptococcus pneumoniae and that early production of IFN-γ regulates bacterial clearance. IFN-γ production by neutrophils requires Rac2, Hck/Lyn/Fgr Src family tyrosine kinases, and NADPH oxidase. Our current studies examined the mechanisms that regulate IFN-γ production by lung neutrophils during acute S. pneumoniae pneumonia in mice and its function. We demonstrate that IFN-γ production by neutrophils is a tightly regulated process that does not require IL-12. The adaptor molecule MyD88 is critical for IFN-γ production by neutrophils. The guanine nucleotide exchange factor CalDAG-GEFI modulates IFN-γ production. The CD11/CD18 complex, CD44, Toll-like receptors 2 and 4, TRIF, and Nrf2 are not required for IFN-γ production by neutrophils. The recently described neutrophil-dendritic cell hybrid cell, identified by its expression of Ly6G and CD11c, is present at low numbers in pneumonic lungs and is not a source of IFN-γ. IFN-γ produced by neutrophils early during acute S. pneumoniae pneumonia induces transcription of target genes in the lungs, which are critical for host defense. These studies underline the complexity of the neutrophil responses during pneumonia in the acute inflammatory response and in subsequent resolution or initiation of immune responses.

  14. Type I interferon transcriptional signature in neutrophils and high frequency of low-density granulocytes are associated with tissue damage in malaria

    PubMed Central

    Rocha, Bruno Coelho; Marques, Pedro Elias; Leoratti, Fabiana Maria de Souza; Junqueira, Caroline; Pereira, Dhelio Batista; Antonelli, Lis Ribeiro do Valle; Menezes, Gustavo Batista

    2015-01-01

    SUMMARY Neutrophils are the most abundant leukocyte population in the bloodstream, the primary compartment of Plasmodium sp. infection. Yet, the role of these polymorphonuclear cells in mediating either resistance or pathogenesis of malaria is poorly understood. We report that circulating neutrophils from malaria patients are highly activated, as indicated by a strong type I interferon transcriptional signature, increased expression of surface activation markers, the enhanced release of reactive oxygen species and myeloperoxidase, as well as the high frequency of low-density granulocytes. The activation of neutrophils was associated with increased levels of serum alanine and aspartate aminotransferases, indicating liver damage. In a rodent malaria model, we observed an intense recruitment of neutrophils to liver sinusoids. Neutrophil migration, IL-1β and chemokine expression as well as liver damage were all dependent on type I interferon signaling. The data suggests that type I interferon signaling have a central role in neutrophil activation and malaria pathogenesis. PMID:26711347

  15. Neutrophils in cancer: neutral no more.

    PubMed

    Coffelt, Seth B; Wellenstein, Max D; de Visser, Karin E

    2016-07-01

    Neutrophils are indispensable antagonists of microbial infection and facilitators of wound healing. In the cancer setting, a newfound appreciation for neutrophils has come into view. The traditionally held belief that neutrophils are inert bystanders is being challenged by the recent literature. Emerging evidence indicates that tumours manipulate neutrophils, sometimes early in their differentiation process, to create diverse phenotypic and functional polarization states able to alter tumour behaviour. In this Review, we discuss the involvement of neutrophils in cancer initiation and progression, and their potential as clinical biomarkers and therapeutic targets.

  16. Application of Intracellular Alkaline Phosphatase Activity Measurement in Detection of Neutrophil Adherence In Vitro

    PubMed Central

    Bednarska, Katarzyna; Klink, Magdalena; Sulowska, Zofia

    2006-01-01

    We have proposed the use of the fluorimetric method with 4-methylumbelliferyl phosphate (4-MUP) specific substrate for the alkaline phosphatase determination in the neutrophil adhesion assay. We provide evidence that the endogenous neutrophil alkaline phosphatase (NAP) activity evaluation is reliable to quantify neutrophil adhesion at a wide range of cell numbers (104−106). The results obtained by fluorimetric NAP activity test correlate to the results of adherence evaluated using the MTT reduction assay. The fluorimetric NAP activity test may be applied for resting as well as activated neutrophils without the risk of the activators interferences into the test. The alkaline phosphatase survey with the use of 4-MUP substrate is recommended herein as a sensitive, repeatable, simple, and reliable method of the neutrophil adherence determination in vitro. PMID:17047286

  17. Neutrophils and neutrophil serine proteases are increased in the spleens of estrogen-treated C57BL/6 mice and several strains of spontaneous lupus-prone mice

    PubMed Central

    Dai, Rujuan; Cowan, Catharine; Heid, Bettina; Khan, Deena; Liang, Zhihong; Pham, Christine T. N.; Ahmed, S. Ansar

    2017-01-01

    Estrogen, a natural immunomodulator, regulates the development and function of diverse immune cell types. There is now renewed attention on neutrophils and neutrophil serine proteases (NSPs) such as neutrophil elastase (NE), proteinase 3 (PR3), and cathepsin G (CG) in inflammation and autoimmunity. In this study, we found that although estrogen treatment significantly reduced total splenocytes number, it markedly increased the splenic neutrophil absolute numbers in estrogen-treated C57BL/6 (B6) mice when compared to placebo controls. Concomitantly, the levels of NSPs and myeloperoxidase (MPO) were highly upregulated in the splenocytes from estrogen-treated mice. Despite the critical role of NSPs in the regulation of non-infectious inflammation, by employing NE-/-/PR3-/-/CG-/- triple knock out mice, we demonstrated that the absence of NSPs affected neither estrogen’s ability to increase splenic neutrophils nor the induction of inflammatory mediators (IFNγ, IL-1β, IL-6, TNFα, MCP-1, and NO) from ex vivo activated splenocytes. Depletion of neutrophils in vitro in splenocytes with anti-Ly6G antibody also had no obvious effect on NSP expression or LPS-induced IFNγ and MCP-1. These data suggest that estrogen augments NSPs, which appears to be independent of enhancing ex vivo inflammatory responses. Since estrogen has been implicated in regulating several experimental autoimmune diseases, we extended our observations in estrogen-treated B6 mice to spontaneous autoimmune-prone female MRL-lpr, B6-lpr and NZB/WF1 mice. There was a remarkable commonality with regards to the increase of neutrophils and concomitant increase of NSPs and MPO in the splenic cells of different strains of autoimmune-prone mice and estrogen-treated B6 mice. Collectively, since NSPs and neutrophils are involved in diverse pro-inflammatory activities, these data suggest a potential pathologic implication of increased neutrophils and NSPs that merits further investigation. PMID:28192517

  18. [Neutrophils and monocytes in gingival epithelium

    PubMed

    Meng, H X; Zheng, L P

    1994-06-01

    Neutrophils and monocytes of gingival epithellium in health gingiva(H),marginal gingivitis(MG),juvenile periodontitis(JP),adult periodontitis(AP) and subgingival bacteria were quantitated and analyzed,The results showed that the numbers of PMN within either pocket epithelium or oral gingival epithelium in JP were significantly lower than in AP and G.The amounts of PMN in AP were much larger than other three groups.Positive correlation between the number of PMN in sulcular pocket epitelium and the motile bacteri of subgingival plaque was demonstrated by correlation analysis.Monocytes mainly presented in deep pocket and junctional epithelum which were stained by NAE method,however very few Langhans cells were seen in these areas.

  19. Metabolic regulation of neutrophil spreading, membrane tubulovesicular extensions (cytonemes) formation and intracellular pH upon adhesion to fibronectin.

    PubMed

    Galkina, Svetlana I; Sud'ina, Galina F; Klein, Thomas

    2006-08-01

    Circulating leukocytes have a round cell shape and roll along vessel walls. However, metabolic disorders can lead them to adhere to the endothelium and spread (flatten). We studied the metabolic regulation of adhesion, spreading and intracellular pH (pHi) of neutrophils (polymorphonuclear leukocytes) upon adhesion to fibronectin-coated substrata. Resting neutrophils adhered and spread on fibronectin. An increase in pHi accompanied neutrophil spreading. Inhibition of oxidative phosphorylation or inhibition of P- and F-type ATPases affected neither neutrophil spreading nor pHi. Inhibition of glucose metabolism or V-ATPase impaired neutrophil spreading, blocked the increase in the pHi and induced extrusion of membrane tubulovesicular extensions (cytonemes), anchoring cells to substrata. Omission of extracellular Na(+) and inhibition of chloride channels caused a similar effect. We propose that these tubulovesicular extensions represent protrusions of exocytotic trafficking, supplying the plasma membrane of neutrophils with ion exchange mechanisms and additional membrane for spreading. Glucose metabolism and V-type ATPase could affect fusion of exocytotic trafficking with the plasma membrane, thus controlling neutrophil adhesive state and pHi. Cl(-) efflux through chloride channels and Na(+) influx seem to be involved in the regulation of the V-ATPase by carrying out charge compensation for the proton-pumping activity and through V-ATPase in regulation of neutrophil spreading and pHi.

  20. Microbe-specific unconventional T-cells induce human neutrophil differentiation into antigen cross-presenting cells

    PubMed Central

    Liuzzi, Anna Rita; Tyler, Christopher J.; Khan, Mohd Wajid A.; Szakmany, Tamas; Hall, Judith E.; Moser, Bernhard; Eberl, Matthias

    2014-01-01

    The early immune response to microbes is dominated by the recruitment of neutrophils whose primary function is to clear invading pathogens. However, there is emerging evidence that neutrophils play additional effector and regulatory roles. The present study demonstrates that human neutrophils assume antigen cross-presenting functions, and suggests a plausible scenario for the local generation of APC-like neutrophils through the mobilization of unconventional T-cells in response to microbial metabolites. Vγ9/Vδ2 T-cells and MAIT cells are abundant in blood, inflamed tissues and mucosal barriers. Here, both human cell types responded rapidly to neutrophils after phagocytosis of Gram-positive and Gram-negative bacteria producing the corresponding ligands, and in turn mediated the differentiation of neutrophils into APCs for both CD4+ and CD8+ T-cells through secretion of GM-CSF, IFN-γ and TNF-α. In patients with acute sepsis, circulating neutrophils displayed a similar APC-like phenotype and readily processed soluble proteins for cross-presentation of antigenic peptides to CD8+ T-cells, at a time when peripheral Vγ9/Vδ2 T-cells were highly activated. Our findings indicate that unconventional T-cells represent key controllers of neutrophil-driven innate and adaptive responses to a broad range of pathogens. PMID:25165152

  1. Real-time in vivo imaging reveals the ability of neutrophils to remove Cryptococcus neoformans directly from the brain vasculature.

    PubMed

    Zhang, Mingshun; Sun, Donglei; Liu, Gongguan; Wu, Hui; Zhou, Hong; Shi, Meiqing

    2016-03-01

    Although neutrophils are typically the first immune cells attracted to an infection site, little is known about how neutrophils dynamically interact with invading pathogens in vivo. Here, with the use of intravital microscopy, we demonstrate that neutrophils migrate to the arrested Cryptococcus neoformans, a leading agent to cause meningoencephalitis, in the brain microvasculature. Following interactions with C. neoformans, neutrophils were seen to internalize the organism and then circulate back into the bloodstream, resulting in a direct removal of the organism from the endothelial surface before its transmigration into the brain parenchyma. C. neoformans infection led to enhanced expression of adhesion molecules macrophage 1 antigen on neutrophils and ICAM-1 on brain endothelial cells. Depletion of neutrophils enhanced the brain fungal burden. Complement C3 was critically involved in the recognition of C. neoformans by neutrophils and subsequent clearance of the organism from the brain. Together, our finding of the direct removal of C. neoformans by neutrophils from its arrested site may represent a novel mechanism of host defense in the brain, in addition to the known, direct killing of microorganisms at the infection sites. These data are the first to characterize directly the dynamic interactions of leukocytes with a microbe in the brain of a living animal.

  2. Metabolic regulation of neutrophil spreading, membrane tubulovesicular extensions (cytonemes) formation and intracellular pH upon adhesion to fibronectin

    SciTech Connect

    Galkina, Svetlana I. . E-mail: galkina@genebee.msu.su; Sud'ina, Galina F.; Klein, Thomas

    2006-08-01

    Circulating leukocytes have a round cell shape and roll along vessel walls. However, metabolic disorders can lead them to adhere to the endothelium and spread (flatten). We studied the metabolic regulation of adhesion, spreading and intracellular pH (pHi) of neutrophils (polymorphonuclear leukocytes) upon adhesion to fibronectin-coated substrata. Resting neutrophils adhered and spread on fibronectin. An increase in pHi accompanied neutrophil spreading. Inhibition of oxidative phosphorylation or inhibition of P- and F-type ATPases affected neither neutrophil spreading nor pHi. Inhibition of glucose metabolism or V-ATPase impaired neutrophil spreading, blocked the increase in the pHi and induced extrusion of membrane tubulovesicular extensions (cytonemes), anchoring cells to substrata. Omission of extracellular Na{sup +} and inhibition of chloride channels caused a similar effect. We propose that these tubulovesicular extensions represent protrusions of exocytotic trafficking, supplying the plasma membrane of neutrophils with ion exchange mechanisms and additional membrane for spreading. Glucose metabolism and V-type ATPase could affect fusion of exocytotic trafficking with the plasma membrane, thus controlling neutrophil adhesive state and pHi. Cl{sup -} efflux through chloride channels and Na{sup +} influx seem to be involved in the regulation of the V-ATPase by carrying out charge compensation for the proton-pumping activity and through V-ATPase in regulation of neutrophil spreading and pHi.

  3. Neutrophils: Cinderella of innate immune system.

    PubMed

    Kumar, V; Sharma, A

    2010-11-01

    Neutrophils are the first line of innate immune defense against infectious diseases. However, since their discovery by Elie Metchnikoff, they have always been considered tissue-destructive cells responsible for inflammatory tissue damage occurring during acute infections. Now, extensive research in the field of neutrophil cell biology and their role skewing the immune response in various infections or inflammatory disorders revealed their importance in the regulation of immune response. Along with releasing various antimicrobial molecules, neutrophils also release neutrophil extracellular traps (NETs) for the containment of infection and inflammation. Activated neutrophils provide signals for the activation and maturation of macrophages as well as dendritic cells. Neutrophils are also involved in the regulation of T-cell immune response against various pathogens and tumor antigens. Thus, the present review is intended to highlight the emerging role of neutrophils in the regulation of both innate and adaptive immunity during acute infectious or inflammatory conditions.

  4. Neutrophils induce proangiogenic T cells with a regulatory phenotype in pregnancy

    PubMed Central

    Nadkarni, Suchita; Smith, Joanne; Sferruzzi-Perri, Amanda N.; Ledwozyw, Agata; Kishore, Madhav; Haas, Robert; Mauro, Claudio; Williams, David J.; Farsky, Sandra H. P.; Marelli-Berg, Federica M.; Perretti, Mauro

    2016-01-01

    Although neutrophils are known to be fundamental in controlling innate immune responses, their role in regulating adaptive immunity is just starting to be appreciated. We report that human neutrophils exposed to pregnancy hormones progesterone and estriol promote the establishment of maternal tolerance through the induction of a population of CD4+ T cells displaying a GARP+CD127loFOXP3+ phenotype following antigen activation. Neutrophil-induced T (niT) cells produce IL-10, IL-17, and VEGF and promote vessel growth in vitro. Neutrophil depletion during murine pregnancy leads to abnormal development of the fetal-maternal unit and reduced empbryo development, with placental architecture displaying poor trophoblast invasion and spiral artery development in the maternal decidua, accompanied by significantly attenuated niT cell numbers in draining lymph nodes. Using CD45 congenic cells, we show that induction of niT cells and their regulatory function occurs via transfer of apoptotic neutrophil-derived proteins, including forkhead box protein 1 (FOXO1), to T cells. Unlike in women with healthy pregnancies, neutrophils from blood and placental samples of preeclamptic women fail to induce niT cells as a direct consequence of their inability to transfer FOXO1 to T cells. Finally, neutrophil-selective FOXO1 knockdown leads to defective placentation and compromised embryo development, similar to that resulting from neutrophil depletion. These data define a nonredundant function of neutrophil–T cell interactions in the regulation of vascularization at the maternal–fetal interface. PMID:27956610

  5. The acute response of neutrophil function to a bout of judo training.

    PubMed

    Chinda, Daisuke; Umeda, Takashi; Shimoyama, Tadashi; Kojima, Arata; Tanabe, Masaru; Nakaji, Shigeyuki; Sugawara, Kazuo

    2003-01-01

    Intensive exercise training decreases neutrophil functions in athletes. However, no studies to date have investigated the effect of irregular-interval training, such as is associated with judo training programmes, on neutrophil functions. The purpose of this study was to examine such effects. Thirty-seven male college judoists participated in this study. Neutrophil oxidative burst activity, phagocytic activity and expression of CD11b and CD16 per cell were measured by fl ow cytometry before and after judo training. Total neutrophil counts increased significantly from 2.98 +/- 0.82 to 7.95 +/- 1.80 x 10(3)/ microL (p < 0.001). The proportion of neutrophils producing reactive oxygen species (ROS) was increased significantly (p < 0.001). On the other hand, the phagocytic activity decreased after training, as shown by a decrease in the amount of ingested opsonized zymosan per cell (p < 0.001), possibly as a compensatory effect for the increased numbers of ROS-producing neutrophils. Expression of CD11b and CD16 per cell decreased by 20% and 30%, respectively, after judo training. In conclusion, judo training induced a decrease in phagocytic activity through the lowered expression of CD11b and CD16 on the surface of neutrophils, and increased the oxidative burst activity of neutrophils.

  6. Effect of soy milk on circulating 17- β estradiol, number of neurons in cerebral cortex and hippocampus and determination of their ratio in neonatal ovariectomized rats

    PubMed Central

    Marzban Abbasabadi, Behrokh; Tadjalli, Mina

    2016-01-01

    This study was conducted to evaluate the effect of soy milk on serum 17- β estradiol level and number of neurons in cerebral cortex and hippocampus as well as determination of the ratio of neurons in cortical and hippocampal regions in neonatal ovariectomized rats. Thirty female rats (one day old) were divided into six groups of five. At day 7, ovariectomy surgery was performed in four groups and two other groups were assumed as sham and control groups. Three groups of ovareictomaized rats were fed with soy milk at the doses of 0.75, 1.50 and 3.00 mL kg-1 per day since they were 14. At day 60, the blood samples were collected to measure the17- β estradiol concentration, and then the brain of rats were prepared for histological studies. The serum 17- β estradiol level significantly increased in ovariectomized rats fed with soy milk compared to ovariectomized rats with no soy milk supplementation. In addition, the results showed that soy milk significantly increased the number of neurons in CA1, CA2 and dentate gyrus regions of hippocampus and granular layer of cerebral cortex in ovariectomized rats, whereas there was no significant change in number of neurons in CA3 zone of hippocampus and molecular, pyramidal and multiform layers of cerebral cortex in ovariectomized rats fed with soy milk. The ratio of cerebral cortex neurons to hippocampal neurons had no significant changes among the experimental groups. PMID:28144428

  7. Neutrophil extracellular traps - the dark side of neutrophils.

    PubMed

    Sørensen, Ole E; Borregaard, Niels

    2016-05-02

    Neutrophil extracellular traps (NETs) were discovered as extracellular strands of decondensed DNA in complex with histones and granule proteins, which were expelled from dying neutrophils to ensnare and kill microbes. NETs are formed during infection in vivo by mechanisms different from those originally described in vitro. Citrullination of histones by peptidyl arginine deiminase 4 (PAD4) is central for NET formation in vivo. NETs may spur formation of autoantibodies and may also serve as scaffolds for thrombosis, thereby providing a link among infection, autoimmunity, and thrombosis. In this review, we present the mechanisms by which NETs are formed and discuss the physiological and pathophysiological consequences of NET formation. We conclude that NETs may be of more importance in autoimmunity and thrombosis than in innate immune defense.

  8. The Effects of Pterostilbene on Neutrophil Activity in Experimental Model of Arthritis

    PubMed Central

    Drabikova, Katarina; Lojek, Antonin; Ciz, Milan; Ponist, Silvester; Bauerova, Katarina; Nosal, Radomir; Harmatha, Juraj; Jancinova, Viera

    2013-01-01

    It has been demonstrated that pterostilbene inhibits reactive oxygen species production in neutrophils in vitro. However, little is known about its effects on neutrophils during inflammation in vivo. In this study, the effect of pterostilbene on neutrophil activity was investigated in experimental arthritis model. Lewis rats were injected by a single intradermal injection of heat-killed Mycobacterium butyricum in Freund's adjuvant to develop arthritis. Another group of arthritic animals received pterostilbene 30 mg/kg, daily, p.o. The number and activity of neutrophils in blood were measured on a weekly basis during the whole experiment. Moreover, the total radical trapping potential in plasma was measured at the end of the experiment. In the pterostilbene treated arthritic group, the treatment significantly lowered the number of neutrophils in blood on days 14 and 21 without significant downregulation of neutrophil oxidative burst. Pterostilbene nonsignificantly increased total radical trapping potential in arthritic animals. These results indicate that the promising effects of pterostilbene on reactive oxygen species operate by different mechanisms in vitro and in the animal model of inflammation. In conclusion, the positive effects of pterostilbene in the model of arthritis may be attributed to regulation of neutrophil number. PMID:24195064

  9. The effect of free and carrier-bound cortisol on equine neutrophil function.

    PubMed

    Fratto, Melanie A; Hart, Kelsey A; Norton, Natalie A; Barton, Michelle H; Giguère, Steeve; Hurley, David J

    2017-01-01

    Cortisol is a key anti-inflammatory hormone that increases in bacterial sepsis and circulates predominantly bound to cortisol binding globulin (CBG). Only unbound cortisol was believed to be biologically active, but recent evidence suggests that CBG-bound cortisol also regulates inflammation. The objective of this study was to evaluate the effects of free and CBG-bound cortisol on equine neutrophil function ex vivo. We hypothesized that CBG would enhance cortisol-mediated suppression of neutrophil pro-inflammatory responses. Neutrophils isolated from 8 foals and 6 adult horses were exposed to Staphylococcus aureus antigen (SAA) alone and with hydrocortisone (HC), CBG, or both (CBG+HC). Inflammatory cytokine (TNF-α, IL-8) and reactive oxygen species (ROS) production were measured and compared among stimulants and between ages with linear mixed-effects models. CBG and CBG+HC inhibited ROS production induced by SAA in both foal and horse neutrophils, maintaining it at levels comparable to baseline production (P≤0.060-0.907). TNF-α production was not significantly different among stimulants (P=0.284). CBG+HC significantly (P≤0.016) increased IL-8 production by neutrophils in response to SAA in both foals and adults, although the response of foals was significantly greater than that of adults (P<0.001). These findings suggest that CBG directly modulates equine neutrophil responses, but the effects are cytokine- and age-specific.

  10. Cyclic GMP-dependent protein kinase and soluble guanylyl cyclase disappear in elicited rat neutrophils.

    PubMed

    Ciuman, Małgorzata; Siednienko, Jakub; Czyzyk, Rafał; Witwicka, Hanna; Kołosionek, Ewa; Kobiałka, Marcin; Gorczyca, Wojciech A

    2006-11-01

    The nitric oxide/soluble guanylyl cyclase/cGMP-dependent protein kinase (NO/sGC/PKG) cascade has been shown to affect important functions of circulating neutrophils. We demonstrate that neutrophils isolated from rats treated intraperitoneally with peptone protease cannot use this signaling pathway. Although PKG was detected at both the mRNA and protein levels in peripheral blood neutrophils (PBNs) of control rats, it was expressed neither in PBNs nor in peritoneal exudate neutrophils (PENs) of provoked rats. Also, mRNA of the alpha and beta chains of heterodimeric sGC was present in PBNs, but absent in PENs. Consistently, PBNs responded to activators of sGC with cGMP synthesis, while PENs did not. These results showed that neutrophils recruited by a provoking agent lost PKG and, in the case of PENs, also sGC and thus the capacity to respond to NO with cGMP signaling. We speculate that such downregulation of the sGC/PKG pathway is likely a result of the high activity of inducible NO synthase observed in inflammatory neutrophils.

  11. Equine neutrophil elastase in plasma, laminar tissue, and skin of horses administered black walnut heartwood extract.

    PubMed

    de la Rebière de Pouyade, Geoffroy; Riggs, Laura M; Moore, James N; Franck, Thierry; Deby-Dupont, Ginette; Hurley, David J; Serteyn, Didier

    2010-06-15

    Laminitis is a local manifestation of a systemic inflammatory response that is characterized by neutrophil activation and movement of neutrophils into the laminar tissues. Given the evidence for the involvement of neutrophils in the development of laminitis, we measured concentrations of neutrophil elastase, a serine protease released from the azurophilic granules of neutrophils, in plasma, skin and laminar tissues obtained from control horses and horses given black walnut heartwood extract (BWHE) to induce laminitis. Healthy horses (5-15 years old) were randomly assigned to 4 groups: 3 experimental groups given BWHE via nasogastric tube, and a control group given an equal volume of water. The experimental groups consisted of horses euthanized 1.5h (n=5), 3h (n=6) or 12h (n=10) after BWHE administration. Control horses (n=7) were euthanized 12h after intragastric administration of water. Plasma samples were collected in all horses of the control and 12h BWHE groups at 0, 1, 2, 3, 4, 6, 8, 10, and 12h after treatment, and laminar tissue and skin from the middle region of the neck were harvested at the time of euthanasia in all 1.5 and 3h BWHE horses, in 6 of the 12h BWHE horses and in 5 of the control horses. Plasma and tissue concentrations of neutrophil elastase were determined using an equine specific ELISA, and statistical significance was set at p<0.05. Plasma concentrations of neutrophil elastase in the BWHE group were significantly higher at 6 and 8h compared to the control group and at 8 and 10h compared to time 0. Concentrations of neutrophil elastase in skin and laminar tissue were significantly higher in the 3 and 12h BWHE groups compared to the control group. Concentrations of neutrophil elastase were significantly higher in the skin than in the lamina in the 12h BWHE horses. The administration of BWHE thus results in significant increases in the concentration of neutrophil elastase in the circulation, skin and laminar tissue. These results confirm a

  12. Treatment with selectin blocking antibodies after lengthening contractions of mouse muscle blunts neutrophil accumulation but does not reduce damage.

    PubMed

    Sloboda, Darcée D; Brooks, Susan V

    2016-01-01

    P- and E-selectins are expressed on the surface of endothelial cells and may contribute to neutrophil recruitment following injurious lengthening contractions of skeletal muscle. Blunting neutrophil, but not macrophage, accumulation after lengthening contractions may provide a therapeutic benefit as neutrophils exacerbate damage to muscle fibers, while macrophages promote repair. In this study, we tested the hypothesis that P- and E-selectins contribute to neutrophil, but not macrophage, accumulation in muscles after contraction-induced injury, and that reducing neutrophil accumulation by blocking the selectins would be sufficient to reduce damage to muscle fibers. To test our hypothesis, we treated mice with antibodies to block P- and E-selectin function and assessed leukocyte accumulation and damage in muscles 2 days after lengthening contractions. Treatment with P/E-selectin blocking antibodies reduced neutrophil content by about half in muscles subjected to lengthening contractions. In spite of the reduction in neutrophil accumulation, we did not detect a decrease in damage 2 days after lengthening contractions. We conclude that P- and/or E-selectin contribute to the neutrophil accumulation associated with contraction-induced muscle damage and that only a portion of the neutrophils that typically accumulate following injurious lengthening contractions is sufficient to induce muscle fiber damage and force deficits. Thus, therapeutic interventions based on blocking the selectins or other adhesion proteins will have to reduce neutrophil numbers by more than 50% in order to provide a benefit.

  13. Leishmania amazonensis Amastigotes Trigger Neutrophil Activation but Resist Neutrophil Microbicidal Mechanisms

    PubMed Central

    Carlsen, Eric D.; Hay, Christie; Henard, Calvin A.; Popov, Vsevolod; Garg, Nisha Jain

    2013-01-01

    Neutrophils are the first cells to infiltrate to the site of Leishmania promastigote infection, and these cells help to reduce parasite burden shortly after infection is initiated. Several clinical reports indicate that neutrophil recruitment is sustained over the course of leishmaniasis, and amastigote-laden neutrophils have been isolated from chronically infected patients and experimentally infected animals. The goal of this study was to compare how thioglycolate-elicited murine neutrophils respond to L. amazonensis metacyclic promastigotes and amastigotes derived from axenic cultures or from the lesions of infected mice. Neutrophils efficiently internalized both amastigote and promastigote forms of the parasite, and phagocytosis was enhanced in lipopolysaccharide (LPS)-activated neutrophils or when parasites were opsonized in serum from infected mice. Parasite uptake resulted in neutrophil activation, oxidative burst, and accelerated neutrophil death. While promastigotes triggered the release of tumor necrosis factor alpha (TNF-α), uptake of amastigotes preferentially resulted in the secretion of interleukin-10 (IL-10) from neutrophils. Finally, the majority of promastigotes were killed by neutrophils, while axenic culture- and lesion-derived amastigotes were highly resistant to neutrophil microbicidal mechanisms. This study indicates that neutrophils exhibit distinct responses to promastigote and amastigote infection. Our findings have important implications for determining the impact of sustained neutrophil recruitment and amastigote-neutrophil interactions during the late phase of cutaneous leishmaniasis. PMID:23918780

  14. Increased Nucleosomes and Neutrophil Activation Link to Disease Progression in Patients with Scrub Typhus but Not Murine Typhus in Laos.

    PubMed

    Paris, Daniel H; Stephan, Femke; Bulder, Ingrid; Wouters, Diana; van der Poll, Tom; Newton, Paul N; Day, Nicholas P J; Zeerleder, Sacha

    2015-01-01

    Cell-mediated immunity is essential in protection against rickettsial illnesses, but the role of neutrophils in these intracellular vasculotropic infections remains unclear. This study analyzed the plasma levels of nucleosomes, FSAP-activation (nucleosome-releasing factor), and neutrophil activation, as evidenced by neutrophil-elastase (ELA) complexes, in sympatric Lao patients with scrub typhus and murine typhus. In acute scrub typhus elevated nucleosome levels correlated with lower GCS scores, raised respiratory rate, jaundice and impaired liver function, whereas neutrophil activation correlated with fibrinolysis and high IL-8 plasma levels, a recently identified predictor of severe disease and mortality. Nucleosome and ELA complex levels were associated with a 4.8-fold and 4-fold increased risk of developing severe scrub typhus, beyond cut off values of 1,040 U/ml for nucleosomes and 275 U/ml for ELA complexes respectively. In murine typhus, nucleosome levels associated with pro-inflammatory cytokines and the duration of illness, while ELA complexes correlated strongly with inflammation markers, jaundice and increased respiratory rates. This study found strong correlations between circulating nucleosomes and neutrophil activation in patients with scrub typhus, but not murine typhus, providing indirect evidence that nucleosomes could originate from neutrophil extracellular trap (NET) degradation. High circulating plasma nucleosomes and ELA complexes represent independent risk factors for developing severe complications in scrub typhus. As nucleosomes and histones exposed on NETs are highly cytotoxic to endothelial cells and are strongly pro-coagulant, neutrophil-derived nucleosomes could contribute to vascular damage, the pro-coagulant state and exacerbation of disease in scrub typhus, thus indicating a detrimental role of neutrophil activation. The data suggest that increased neutrophil activation relates to disease progression and severe complications, and

  15. The effect of neutrophil migration and prolonged neutrophil contact on epithelial permeability.

    PubMed Central

    Parsons, P. E.; Sugahara, K.; Cott, G. R.; Mason, R. J.; Henson, P. M.

    1987-01-01

    The effect of neutrophil migration and prolonged neutrophil contact on epithelial permeability was examined. Although neutrophil migration was not associated with a change in epithelial permeability, prolonged neutrophil-epithelial contact following migration resulted in an increase in epithelial permeability. These results were not altered by catalase, a specific neutrophil elastase inhibitor, methoxysuccinyl-Ala-Ala-Pro-Val-chloromethyl ketone or cyclohexamide. This suggests that neutrophil migration does not occur via an H2O2-induced reversible mechanism of junctional opening, which we describe herein. PMID:3314530

  16. Fetal Circulation

    MedlinePlus

    ... Echocardiography/Your Unborn Baby's Heart - Fetal Echocardiogram Test - Detection of a Heart Defect - Fetal Circulation • Care & Treatment • Tools & Resources Popular Articles 1 Understanding Blood Pressure Readings 2 Sodium and Salt 3 Target Heart Rates 4 Heart Attack Symptoms in Women ...

  17. Altered Innate Immune Responses in Neutrophils from Patients with Well- and Suboptimally Controlled Asthma

    PubMed Central

    Tang, Francesca S. M.; Foxley, Gloria J.; Gibson, Peter G.; Burgess, Janette K.; Baines, Katherine J.; Oliver, Brian G.

    2015-01-01

    Background. Respiratory infections are a major cause of asthma exacerbations where neutrophilic inflammation dominates and is associated with steroid refractory asthma. Structural airway cells in asthma differ from nonasthmatics; however it is unknown if neutrophils differ. We investigated neutrophil immune responses in patients who have good (AGood) and suboptimal (ASubopt) asthma symptom control. Methods. Peripheral blood neutrophils from AGood (ACQ < 0.75, n = 11), ASubopt (ACQ > 0.75, n = 7), and healthy controls (HC) (n = 9) were stimulated with bacterial (LPS (1 μg/mL), fMLF (100 nM)), and viral (imiquimod (3 μg/mL), R848 (1.5 μg/mL), and poly I:C (10 μg/mL)) surrogates or live rhinovirus (RV) 16 (MOI1). Cell-free supernatant was collected after 1 h for neutrophil elastase (NE) and matrix metalloproteinase- (MMP-) 9 measurements or after 24 h for CXCL8 release. Results. Constitutive NE was enhanced in AGood neutrophils compared to HC. fMLF stimulated neutrophils from ASubopt but not AGood produced 50% of HC levels. fMLF induced MMP-9 was impaired in ASubopt and AGood compared to HC. fMLF stimulated CXCL8 but not MMP-9 was positively correlated with FEV1 and FEV1/FVC. ASubopt and AGood responded similarly to other stimuli. Conclusions. Circulating neutrophils are different in asthma; however, this is likely to be related to airflow limitation rather than asthma control. PMID:26663987

  18. Tracking neutrophil intraluminal crawling, transendothelial migration and chemotaxis in tissue by intravital video microscopy.

    PubMed

    Xu, Najia; Lei, Xi; Liu, Lixin

    2011-09-24

    The recruitment of circulating leukocytes from blood stream to the inflamed tissue is a crucial and complex process of inflammation(1,2). In the postcapillary venules of inflamed tissue, leukocytes initially tether and roll on the luminal surface of venular wall. Rolling leukocytes arrest on endothelium and undergo firm adhesion in response to chemokine or other chemoattractants on the venular surface. Many adherent leukocytes relocate from the initial site of adhesion to the junctional extravasation site in endothelium, a process termed intraluminal crawling(3). Following crawling, leukocytes move across endothelium (transmigration) and migrate in extravascular tissue toward the source of chemoattractant (chemotaxis)(4). Intravital microscopy is a powerful tool for visualizing leukocyte-endothelial cell interactions in vivo and revealing cellular and molecular mechanisms of leukocyte recruitment(2,5). In this report, we provide a comprehensive description of using brightfield intravital microscopy to visualize and determine the detailed processes of neutrophil recruitment in mouse cremaster muscle in response to the gradient of a neutrophil chemoattractant. To induce neutrophil recruitment, a small piece of agarose gel (~1-mm(3) size) containing neutrophil chemoattractant MIP-2 (CXCL2, a CXC chemokine) or WKYMVm (Trp-Lys-Tyr-Val-D-Met, a synthetic analog of bacterial peptide) is placed on the muscle tissue adjacent to the observed postcapillary venule. With time-lapsed video photography and computer software ImageJ, neutrophil intraluminal crawling on endothelium, neutrophil transendothelial migration and the migration and chemotaxis in tissue are visualized and tracked. This protocol allows reliable and quantitative analysis of many neutrophil recruitment parameters such as intraluminal crawling velocity, transmigration time, detachment time, migration velocity, chemotaxis velocity and chemotaxis index in tissue. We demonstrate that using this protocol, these

  19. Endothelial Domes Encapsulate Adherent Neutrophils and Minimize Increases in Vascular Permeability in Paracellular and Transcellular Emigration

    PubMed Central

    Phillipson, Mia; Kaur, Jaswinder; Colarusso, Pina; Ballantyne, Christie M.; Kubes, Paul

    2008-01-01

    Local edema, a cardinal sign of inflammation associates closely with neutrophil emigration. Neutrophil emigration has been described to occur primarily through endothelial junctions (paracellular) and more rarely directly through endothelial cells (transcellular). Recently, we reported that unlike in wild-type (wt) mice, Mac-1-/- (CD11b) neutrophils predominantly emigrated transcellularly and was significantly delayed taking 20–30 min longer than the paracellular emigration (wt). In the present study we noted significant anatomical disruption of the endothelium and hypothesized that transcellular emigration would greatly increase vascular permeability. Surprisingly, despite profound disruption of the endothelial barrier as the neutrophils moved through the cells, the changes in vascular permeability during transcellular emigration (Mac-1-/-) were not increased more than in wt mice. Instead increased vascular permeability completely tracked the number of emigrated cells and as such, permeability changes were delayed in Mac-1-/- mice. However, by 60 min neutrophils from both sets of mice were emigrating in large numbers. Electron-microscopy and spinning disk multichannel fluorescence confocal microscopy revealed endothelial docking structures that progressed to dome-like structures completely covering wt and Mac-1-/- neutrophils. These domes completely enveloped the emigrating neutrophils in both wt and Mac-1-/- mice making the mode of emigration underneath these structures extraneous to barrier function. In conclusion, predominantly paracellular versus predominantly transcellular emigration does not affect vascular barrier integrity as endothelial dome-like structures retain barrier function. PMID:18297135

  20. Endothelial domes encapsulate adherent neutrophils and minimize increases in vascular permeability in paracellular and transcellular emigration.

    PubMed

    Phillipson, Mia; Kaur, Jaswinder; Colarusso, Pina; Ballantyne, Christie M; Kubes, Paul

    2008-02-20

    Local edema, a cardinal sign of inflammation associates closely with neutrophil emigration. Neutrophil emigration has been described to occur primarily through endothelial junctions (paracellular) and more rarely directly through endothelial cells (transcellular). Recently, we reported that unlike in wild-type (wt) mice, Mac-1-/- (CD11b) neutrophils predominantly emigrated transcellularly and was significantly delayed taking 20-30 min longer than the paracellular emigration (wt). In the present study we noted significant anatomical disruption of the endothelium and hypothesized that transcellular emigration would greatly increase vascular permeability. Surprisingly, despite profound disruption of the endothelial barrier as the neutrophils moved through the cells, the changes in vascular permeability during transcellular emigration (Mac-1-/-) were not increased more than in wt mice. Instead increased vascular permeability completely tracked the number of emigrated cells and as such, permeability changes were delayed in Mac-1-/- mice. However, by 60 min neutrophils from both sets of mice were emigrating in large numbers. Electron-microscopy and spinning disk multichannel fluorescence confocal microscopy revealed endothelial docking structures that progressed to dome-like structures completely covering wt and Mac-1-/- neutrophils. These domes completely enveloped the emigrating neutrophils in both wt and Mac-1-/- mice making the mode of emigration underneath these structures extraneous to barrier function. In conclusion, predominantly paracellular versus predominantly transcellular emigration does not affect vascular barrier integrity as endothelial dome-like structures retain barrier function.

  1. Increased Numbers of Circulating CD8 Effector Memory T Cells before Transplantation Enhance the Risk of Acute Rejection in Lung Transplant Recipients

    PubMed Central

    San Segundo, David; Ballesteros, María Ángeles; Naranjo, Sara; Zurbano, Felipe; Miñambres, Eduardo; López-Hoyos, Marcos

    2013-01-01

    The effector and regulatory T cell subpopulations involved in the development of acute rejection episodes in lung transplantation remain to be elucidated. Twenty-seven lung transplant candidates were prospectively monitored before transplantation and within the first year post-transplantation. Regulatory, Th17, memory and naïve T cells were measured in peripheral blood of lung transplant recipients by flow cytometry. No association of acute rejection with number of peripheral regulatory T cells and Th17 cells was found. However, effector memory subsets in acute rejection patients were increased during the first two months post-transplant. Interestingly, patients waiting for lung transplant with levels of CD8+ effector memory T cells over 185 cells/mm3 had a significant increased risk of rejection [OR: 5.62 (95% CI: 1.08-29.37), p=0.04]. In multivariate analysis adjusted for age and gender the odds ratio for rejection was: OR: 5.89 (95% CI: 1.08-32.24), p=0.04. These data suggest a correlation between acute rejection and effector memory T cells in lung transplant recipients. The measurement of peripheral blood CD8+ effector memory T cells prior to lung transplant may define patients at high risk of acute lung rejection. PMID:24236187

  2. The effects and comparative differences of neutrophil specific chemokines on neutrophil chemotaxis of the neonate.

    PubMed

    Fox, Samuel E; Lu, Wenge; Maheshwari, Akhil; Christensen, Robert D; Calhoun, Darlene A

    2005-02-07

    Neutrophil specific chemokines are potent chemoattractants for neutrophils. IL-8/CXCL8 is the most extensively studied member of this group, and its concentrations increase during inflammatory conditions of the newborn infant including sepsis and chronic lung disease. A significant amount of information exists on the effects of IL-8/CXCL8 on neutrophil chemotaxis of neonates, but little is known about the other neutrophil specific chemokines. The aim of this study was to determine the relative potency of the neutrophil specific chemokines on chemotaxis of neonatal neutrophils and to compare this effect with the effect on adult neutrophils. Neutrophils were isolated from cord blood or healthy adult donors and incubated in a Neuroprobe chemotaxis chamber. Chemokine concentrations ranging from 1-1000 ng/mL were used. Differences in chemotactic potency existed among the seven neutrophil specific chemokines. Specifically, at 100 ng/mL, the order was IL-8/CXCL8>GRO-alpha/CXCL1>GCP-2/CXCL6>NAP-2/CXCL7>ENA-78/CXCL5>GRO-gamma/CXCL2>GRO-beta/CXCL3. This pattern was observed for adult and neonatal neutrophils. We conclude that (1) neutrophils from cord blood exhibit the same pattern of potency for each ELR chemokine as neutrophils from adults, and (2) migration of neonatal neutrophils is significantly less than that of adults at every concentration examined except the lowest (1 ng/mL).

  3. Mechanisms of Interferon-γ Production by Neutrophils and Its Function during Streptococcus pneumoniae Pneumonia

    PubMed Central

    Gomez, John C.; Yamada, Mitsuhiro; Martin, Jessica R.; Dang, Hong; Brickey, W. June; Bergmeier, Wolfgang; Dinauer, Mary C.

    2015-01-01

    Bacterial pneumonia is a common public health problem associated with significant mortality, morbidity, and cost. Neutrophils are usually the earliest leukocytes to respond to bacteria in the lungs. Neutrophils rapidly sequester in the pulmonary microvasculature and migrate into the lung parenchyma and alveolar spaces, where they perform numerous effector functions for host defense. Previous studies showed that migrated neutrophils produce IFN-γ early during pneumonia induced by Streptococcus pneumoniae and that early production of IFN-γ regulates bacterial clearance. IFN-γ production by neutrophils requires Rac2, Hck/Lyn/Fgr Src family tyrosine kinases, and NADPH oxidase. Our current studies examined the mechanisms that regulate IFN-γ production by lung neutrophils during acute S. pneumoniae pneumonia in mice and its function. We demonstrate that IFN-γ production by neutrophils is a tightly regulated process that does not require IL-12. The adaptor molecule MyD88 is critical for IFN-γ production by neutrophils. The guanine nucleotide exchange factor CalDAG-GEFI modulates IFN-γ production. The CD11/CD18 complex, CD44, Toll-like receptors 2 and 4, TRIF, and Nrf2 are not required for IFN-γ production by neutrophils. The recently described neutrophil–dendritic cell hybrid cell, identified by its expression of Ly6G and CD11c, is present at low numbers in pneumonic lungs and is not a source of IFN-γ. IFN-γ produced by neutrophils early during acute S. pneumoniae pneumonia induces transcription of target genes in the lungs, which are critical for host defense. These studies underline the complexity of the neutrophil responses during pneumonia in the acute inflammatory response and in subsequent resolution or initiation of immune responses. PMID:25100610

  4. N-acetyl-L-cysteine and cysteine increase intracellular calcium concentration in human neutrophils

    PubMed Central

    Hasan, Md. Ashraful; Ahn, Won-Gyun

    2016-01-01

    N-acetyl-L-cysteine (NAC) and cysteine have been implicated in a number of human neutrophils' functional responses. However, though Ca2+ signaling is one of the key signalings contributing to the functional responses of human neutrophils, effects of NAC and cysteine on intracellular calcium concentration ([Ca2+]i) in human neutrophils have not been investigated yet. Thus, this study was carried out with an objective to investigate the effects of NAC and cysteine on [Ca2+]i in human neutrophils. We observed that NAC (1 µM ~ 1 mM) and cysteine (10 µM ~ 1 mM) increased [Ca2+]i in human neutrophils in a concentration-dependent manner. In NAC pre-supplmented buffer, an additive effect on N-formyl-methionine-leucine-phenylalanine (fMLP)-induced increase in [Ca2+]i in human neutrophils was observed. In Ca2+-free buffer, NAC- and cysteine-induced [Ca2+]i increase in human neutrophils completely disappeared, suggesting that NAC- and cysteine-mediated increase in [Ca2+]i in human neutrophils occur through Ca2+ influx. NAC- and cysteine-induced [Ca2+]i increase was effectively inhibited by calcium channel inhibitors SKF96365 (10 µM) and ruthenium red (20 µM). In Na+-free HEPES, both NAC and cysteine induced a marked increase in [Ca2+]i in human neutrophils, arguing against the possibility that Na+-dependent intracellular uptake of NAC and cysteine is necessary for their [Ca2+]i increasing activity. Our results show that NAC and cysteine induce [Ca2+]i increase through Ca2+ influx in human neutrophils via SKF96365- and ruthenium red-dependent way. PMID:27610031

  5. G Protein-Coupled Receptor 43 Modulates Neutrophil Recruitment during Acute Inflammation

    PubMed Central

    Nicholls, Alyce J.; Oliveira, Ana Carolina; Mason, Linda J.; Binge, Lauren; Mackay, Charles R.; Wong, Connie H. Y.

    2016-01-01

    Fermentation of dietary fibre in the gut yields large amounts of short chain fatty acids (SCFAs). SCFAs can impart biological responses in cells through their engagement of ‘metabolite-sensing’ G protein-coupled receptors (GPCRs). One of the main SCFA receptors, GPR43, is highly expressed by neutrophils, which suggests that the actions of GPR43 and dietary fibre intake may affect neutrophil recruitment during inflammatory responses in vivo. Using intravital imaging of the small intestine, we found greater intravascular neutrophil rolling and adhesion in Gpr43−/−mice in response to LPS at 1 h. After 4 h of LPS challenge, the intravascular rolling velocity of GPR43-deficient neutrophils was reduced significantly and increased numbers of neutrophils were found in the lamina propria of Gpr43−/−mice. Additionally, GPR43-deficient leukocytes demonstrated exacerbated migration into the peritoneal cavity following fMLP challenge. The fMLP-induced neutrophil migration was significantly suppressed in wildtype mice that were treated with acetate, but not in Gpr43−/−mice, strongly suggesting a role for SCFAs in modulating neutrophil migration via GPR43. Indeed, neutrophils of no fibre-fed wildtype mice exhibited elevated migratory behaviour compared to normal chow-fed wildtype mice. Interestingly, this elevated migration could also be reproduced through simple transfer of a no fibre microbiota into germ-free mice, suggesting that the composition and function of microbiota stemming from a no fibre diet mediated the changes in neutrophil migration. Therefore, GPR43 and a microbiota composition that allows for SCFA production function to modulate neutrophil recruitment during inflammatory responses. PMID:27658303

  6. Differential neutrophil chemotactic response towards IL-8 and bacterial N-formyl peptides in term newborn infants

    PubMed Central

    Stålhammar, Maria E.; Douhan Håkansson, Lena; Jonzon, Anders; Sindelar, Richard

    2017-01-01

    Background A prerequisite for an effective innate immunity is the migrative ability of neutrophils to respond to inflammatory and infectious agents such as the intermediate interleukin (IL)-8 and the end-target formyl-methionyl-leucyl-phenylalanine (fMLP) chemoattractants. The aim was to study the chemotactic capacity of neutrophils from newborn infants and adults in response to IL-8 and the bacterial peptide fMLP. Methods In the under-agarose cell migration assay, isolated leukocytes from healthy adults and from cord blood of healthy term newborn infants were studied with dose responses towards IL-8 and fMLP. The same number of leukocytes (1 × 105 cells), with the same distribution of neutrophils and monocytes, were analyzed in neonates and adults. Chemotaxis was distinguished from randomly migrating neutrophils, and the neutrophil pattern of migration, i.e. the migration distance and the number of migrating neutrophils per distance, was evaluated. Results In comparison to adults, fewer neutrophils from newborn infants migrated towards IL-8 and for a shorter distance (P < .01, respectively). The number of neutrophils migrating to different gradients of fMLP, the distance they migrated, and the correlation between the number and the distance were the same for neonates and adults. Random migration did not differ in any instance. Conclusion Chemotaxis of neutrophils from newborn infants was as co-ordinated as neutrophils from adults in response to fMLP, whereas the response to IL-8 was reduced. The differential response of neutrophils from neonates to intermediate and end-target chemoattractants could indicate a reduced infectious response. PMID:27690722

  7. Annual maximum 5-day rainfall total and maximum number of consecutive dry days over Central America and the Caribbean in the late twenty-first century projected by an atmospheric general circulation model with three different horizontal resolutions

    NASA Astrophysics Data System (ADS)

    Nakaegawa, T.; Kitoh, A.; Murakami, H.; Kusunoki, S.

    2014-04-01

    We simulated changes in annual maximum 5-day rainfall (RX5D) and annual maximum number of consecutive dry days (CDD) in Central America, Mexico, and the Caribbean with three different horizontal resolution atmospheric global general circulation models (AGCMs) and quantified the uncertainty of the projections. The RX5Ds and CDDs were projected to increase in most areas in response to global warming. However, consistent changes were confined to small areas: for RX5D, both coastal zones of northern Mexico and the Yucatan Peninsula; for CDD, the Pacific coastal zone of Mexico, the Yucatan Peninsula, and Guatemala. All three AGCMs projected that RX5Ds and CDDs averaged over only the land area and over the entire area (land and ocean) would increase. The dependence of RX5D probability density functions on the horizontal resolutions was complex. Precipitation unrelated to tropical cyclones was primarily responsible for the projected increases in the frequency of RX5Ds greater than 300 mm.

  8. Plasma sE-selectin level is positively correlated with neutrophil count and diastolic blood pressure in Japanese men.

    PubMed

    Mochizuki, Kazuki; Inoue, Seiya; Miyauchi, Rie; Misaki, Yasumi; Shimada, Masaya; Kasezawa, Nobuhiko; Tohyama, Kazushige; Goda, Toshinao

    2013-01-01

    Increased levels of circulating soluble type of E-selectin (sE-selectin), neutrophil counts and blood pressure are associated with the development of cardiovascular diseases (CVD). In this study, we conducted a cross-sectional study of men who participated in health check-ups, and selected those who were not diagnosed with or being treated for metabolic diseases such as diabetes, hypertension and lipid abnormality according to the health check-ups. We measured their basic clinical parameters including blood pressure and neutrophil count, plasma sE-selectin concentration and lifestyle factors, and assessed their interrelations by multivariate linear regression (MLR) analysis. A total of 351 subjects aged 47.5±8.41 (range, 30-64) y were recruited. Significantly correlated with sE-selectin concentration were neutrophil count, diastolic blood pressure (DBP), and systolic blood pressure (SBP) (Pearson's correlation coefficient, 0.194, 0.220 and 0.175, respectively). MLR analysis showed that sE-selectin concentration was independently positively related with DBP and neutrophil count, whereas neutrophil count was positively associated with sE-selectin concentration but not DBP. DBP, but not SBP, was independently positively correlated with sE-selectin concentration but not neutrophil count. These results indicate that circulating sE-selectin concentration may be a biomarker for indicating subsequent development of metabolic diseases, in particular CVD, from a healthy state.

  9. Alarmins Link Neutrophils and Dendritic Cells

    PubMed Central

    Yang, De; de la Rosa, Gonzalo; Tewary, Poonam; Oppenheim, Joost J.

    2009-01-01

    Neutrophils are the first major population of leukocyte to infiltrate infected or injured tissues and are crucial for initiating host innate defense and adaptive immunity. Although the contribution of neutrophils to innate immune defense is mediated predominantly by phagocytosis and killing of microorganisms, neutrophils also participate in the induction of adaptive immune responses. At sites of infection and/or injury, neutrophils release numerous mediators upon degranulation or death, among these are alarmins which have a characteristic dual capacity to mobilize and activate antigen-presenting cells. We describe here how alarmins released by neutrophil degranulation and/or death can link neutrophils to dendritic cells by promoting their recruitment and activation, resulting in the augmentation of innate and adaptive immune responses. PMID:19699678

  10. The Role of Neutrophils in Transplanted Organs.

    PubMed

    Scozzi, D; Ibrahim, M; Menna, C; Krupnick, A S; Kreisel, D; Gelman, A E

    2017-02-01

    Neutrophils are often viewed as nonspecialized effector cells whose presence is a simple indicator of tissue inflammation. There is new evidence that neutrophils exist in subsets and have specialized effector functions that include extracellular trap generation and the stimulation of angiogenesis. The application of intravital imaging to transplanted organs has revealed novel requirements for neutrophil trafficking into graft tissue and has illuminated direct interactions between neutrophils and other leukocytes that promote alloimmunity. Paradoxically, retaining some neutrophilia may be important to induce or maintain tolerance. Neutrophils can stimulate anti-inflammatory signals in other phagocytes and release molecules that inhibit T cell activation. In this article, we will review the available evidence of how neutrophils regulate acute and chronic inflammation in transplanted organs and discuss the possibility of targeting these cells to promote tolerance.

  11. The Role of Neutrophils in Transplanted Organs

    PubMed Central

    Menna, Cecilia; Krupnick, Alexander S.; Kreisel, Daniel; Gelman, Andrew E.

    2016-01-01

    Neutrophils are often viewed as non-specialized effector cells whose presence is a simple indicator of tissue inflammation. There is new evidence that neutrophils exist in subsets and have specialized effector functions that include extracellular trap generation and the stimulation of angiogenesis. The application of intravital imaging to transplanted organs has revealed novel requirements for neutrophil trafficking into graft tissue and illuminated direct interactions between neutrophils and other leukocytes that promote alloimmunity. Paradoxically, retaining some neutrophilia may be important to induce or maintain tolerance. Neutrophils can stimulate anti-inflammatory signals in other phagocytes and release molecules that inhibit T cell activation. Here we will review the available evidence of how neutrophils regulate acute and chronic inflammation in transplanted organs and discuss the possibility of targeting these cells to promote tolerance. PMID:27344051

  12. A GCSFR/CSF3R zebrafish mutant models the persistent basal neutrophil deficiency of severe congenital neutropenia

    PubMed Central

    Pazhakh, Vahid; Clark, Sharon; Keightley, M. Cristina; Lieschke, Graham J.

    2017-01-01

    Granulocyte colony-stimulating factor (GCSF) and its receptor (GCSFR), also known as CSF3 and CSF3R, are required to maintain normal neutrophil numbers during basal and emergency granulopoiesis in humans, mice and zebrafish. Previous studies identified two zebrafish CSF3 ligands and a single CSF3 receptor. Transient antisense morpholino oligonucleotide knockdown of both these ligands and receptor reduces neutrophil numbers in zebrafish embryos, a technique widely used to evaluate neutrophil contributions to models of infection, inflammation and regeneration. We created an allelic series of zebrafish csf3r mutants by CRISPR/Cas9 mutagenesis targeting csf3r exon 2. Biallelic csf3r mutant embryos are viable and have normal early survival, despite a substantial reduction of their neutrophil population size, and normal macrophage abundance. Heterozygotes have a haploinsufficiency phenotype with an intermediate reduction in neutrophil numbers. csf3r mutants are viable as adults, with a 50% reduction in tissue neutrophil density and a substantial reduction in the number of myeloid cells in the kidney marrow. These csf3r mutants are a new animal model of human CSF3R-dependent congenital neutropenia. Furthermore, they will be valuable for studying the impact of neutrophil loss in the context of other zebrafish disease models by providing a genetically stable, persistent, reproducible neutrophil deficiency state throughout life. PMID:28281657

  13. Loss of Lung WWOX Expression Causes Neutrophilic Inflammation.

    PubMed

    Singla, Sunit; Chen, Jiwang; Sethuraman, Shruthi; Sysol, Justin R; Gampa, Amulya; Zhao, Shuangping; Machado, Roberto F

    2017-03-10

    The tumor suppressor, WWOX, exhibits regulatory interactions with an array of transcription factors and signaling molecules that are positioned at the well-known crossroads between inflammation and cancer. WWOX is also subject to downregulation by genotoxic environmental exposures, making it of potential interest to the study of lung pathobiology. Knockdown of lung WWOX expression in mice was observed to cause neutrophil influx, and accompanied by a corresponding vascular leak and inflammatory cytokine production. In cultured human alveolar epithelial cells, loss of WWOX expression resulted in increased c-Jun- and IL-8- dependent neutrophil chemotaxis towards cell monolayers. WWOX was observed to directly interact with c-Jun in these cells, and its absence resulted in increased nuclear translocation of c-Jun. Finally, inhibition of c-Jun activating kinase, JNK, abrogated the lung neutrophil influx observed during WWOX knockdown in mice. Altogether, these observations represent a novel mechanism of pulmonary neutrophil influx that is highly relevant to the pathobiology and potential treatment of a number of different lung inflammatory conditions.

  14. Endogenous Morphine Levels Are Increased in Sepsis: A Partial Implication of Neutrophils

    PubMed Central

    Lavaux, Thomas; Muller, Arnaud H.; Laux, Alexis; Zhang, Dan; Schmidt, Alexander R.; Delalande, François; Laventie, Benoît-Joseph; Dirrig-Grosch, Sylvie; Colin, Didier A.; Van Dorsselaer, Alain; Aunis, Dominique; Metz-Boutigue, Marie-Hélène; Schneider, Francis; Goumon, Yannick

    2010-01-01

    Background Mammalian cells synthesize morphine and the respective biosynthetic pathway has been elucidated. Human neutrophils release this alkaloid into the media after exposure to morphine precursors. However, the exact role of endogenous morphine in inflammatory processes remains unclear. We postulate that morphine is released during infection and can be determined in the serum of patients with severe infection such as sepsis. Methodology The presence and subcellular immunolocalization of endogenous morphine was investigated by ELISA, mass spectrometry analysis and laser confocal microscopy. Neutrophils were activated with Interleukin-8 (IL-8) or lipopolysaccharide (LPS). Morphine secretion was determined by a morphine-specific ELISA. μ opioid receptor expression was assessed with flow cytometry. Serum morphine concentrations of septic patients were determined with a morphine-specific ELISA and morphine identity was confirmed in human neutrophils and serum of septic patients by mass spectrometry analysis. The effects of the concentration of morphine found in serum of septic patients on LPS-induced release of IL-8 by human neutrophils were tested. Principal Findings We confirmed the presence of morphine in human neutrophil extracts and showed its colocalisation with lactoferrin within the secondary granules of neutrophils. Morphine secretion was quantified in the supernatant of activated human polymorphonuclear neutrophils in the presence and absence of Ca2+. LPS and IL-8 were able to induce a significant release of morphine only in presence of Ca2+. LPS treatment increased μ opioid receptor expression on neutrophils. Low concentration of morphine (8 nM) significantly inhibited the release of IL-8 from neutrophils when coincubated with LPS. This effect was reversed by naloxone. Patients with sepsis, severe sepsis and septic shock had significant higher circulating morphine levels compared to patients with systemic inflammatory response syndrome and healthy

  15. Proteolytic enzyme levels are increased during granulocyte colony-stimulating factor-induced hematopoietic stem cell mobilization in human donors but do not predict the number of mobilized stem cells.

    PubMed

    van Os, R; van Schie, M L J; Willemze, R; Fibbe, W E

    2002-06-01

    Previous studies from our laboratory indicate that functional, mature neutrophils are essential for interleukin-8 (IL-8)-induced stem cell mobilization. To study a possible role of neutrophils in granulocyte colony-stimulating factor (G-CSF) induced hematopoietic mobilization, we assessed the number of circulating CD34+ cells in healthy allogeneic stem cell donors on days 3, 4, and 5 of mobilization for comparison with the number of peripheral blood neutrophils and the plasma levels of IL-8, Flt3 ligand (FL), matrix metalloproteinase-9 (MMP-9), and human neutrophil elastase (HNE). Thirty-seven of 45 donors required 1 day of apheresis to obtain 5 x 10(6) CD34+/kg recipient body weight (high responders), the remaining 8 donors required 1 extra day of apheresis on day 6 (low responders). On day 5, CD34+ numbers in the blood were significantly highe in high responders (116 x 10(3) +/- 10.4/ml) than in low responders (54.1 x 10(3) +/- 10.3, p < 0.001). In all donors, MMP-9 and HNE levels were increased compared to nonmobilized individuals, but in high responders, plasma MMP-9 levels on days 3-5 of mobilization were substantially higher than in low responders (p < or = 0.02 for MMP-9 and p = 0.89, p = 0.05 and p = 0.52 for HNE on days 3, 4, and 5, respectively). These results are in accordance with the hypothesis that neutrophils play a role in G-CSF-induced mobilization through the release of proteases such as MMP-9 and elastase. No change in plasma levels of IL-8 or Flt3 ligand was observed, suggesting that these cytokines do not play a role in stem cell mobilization. However, because stem cell numbers could not be predicted by proteolytic enzyme levels and/or neutrophil numbers, other undefined factors may be more important.

  16. Neutrophil Migration into the Infected Uroepithelium Is Regulated by the Crosstalk between Resident and Helper Macrophages

    PubMed Central

    Zec, Kristina; Volke, Julia; Vijitha, Nirojah; Thiebes, Stephanie; Gunzer, Matthias; Kurts, Christian; Engel, Daniel Robert

    2016-01-01

    The antibacterial defense against infections depends on the cooperation between distinct phagocytes of the innate immune system, namely macrophages and neutrophils. However, the mechanisms driving this cooperation are incompletely understood. In this study we describe the crosstalk between Ly6C+ and Ly6C− macrophage-subtypes and neutrophils in the context of urinary tract infection (UTI) with uropathogenic E. coli (UPEC). Ly6C− macrophages acted as tissue resident sentinels and attracted circulating phagocytes by chemokines. Ly6C+ macrophages produced tumor necrosis factor (TNF) that licensed Ly6C− macrophages to release preformed CXCL2, which in turn caused matrix metalloproteinases (MMP-9) secretion by neutrophils to enable transepithelial migration. PMID:26861402

  17. Bordetella parapertussis Circumvents Neutrophil Extracellular Bactericidal Mechanisms

    PubMed Central

    Gorgojo, Juan; Scharrig, Emilia; Gómez, Ricardo M.; Harvill, Eric T.; Rodríguez, Maria Eugenia

    2017-01-01

    B. parapertussis is a whooping cough etiological agent with the ability to evade the immune response induced by pertussis vaccines. We previously demonstrated that in the absence of opsonic antibodies B. parapertussis hampers phagocytosis by neutrophils and macrophages and, when phagocytosed, blocks intracellular killing by interfering with phagolysosomal fusion. But neutrophils can kill and/or immobilize extracellular bacteria through non-phagocytic mechanisms such as degranulation and neutrophil extracellular traps (NETs). In this study we demonstrated that B. parapertussis also has the ability to circumvent these two neutrophil extracellular bactericidal activities. The lack of neutrophil degranulation was found dependent on the O antigen that targets the bacteria to cell lipid rafts, eventually avoiding the fusion of nascent phagosomes with specific and azurophilic granules. IgG opsonization overcame this inhibition of neutrophil degranulation. We further observed that B. parapertussis did not induce NETs release in resting neutrophils and inhibited NETs formation in response to phorbol myristate acetate (PMA) stimulation by a mechanism dependent on adenylate cyclase toxin (CyaA)-mediated inhibition of reactive oxygen species (ROS) generation. Thus, B. parapertussis modulates neutrophil bactericidal activity through two different mechanisms, one related to the lack of proper NETs-inducer stimuli and the other one related to an active inhibitory mechanism. Together with previous results these data suggest that B. parapertussis has the ability to subvert the main neutrophil bactericidal functions, inhibiting efficient clearance in non-immune hosts. PMID:28095485

  18. Plasticity of neutrophils reveals modulatory capacity

    PubMed Central

    Perobelli, S.M.; Galvani, R.G.; Gonçalves-Silva, T.; Xavier, C.R.; Nóbrega, A.; Bonomo, A.

    2015-01-01

    Neutrophils are widely known as proinflammatory cells associated with tissue damage and for their early arrival at sites of infection, where they exert their phagocytic activity, release their granule contents, and subsequently die. However, this view has been challenged by emerging evidence that neutrophils have other activities and are not so short-lived. Following activation, neutrophil effector functions include production and release of granule contents, reactive oxygen species (ROS), and neutrophil extracellular traps (NETs). Neutrophils have also been shown to produce a wide range of cytokines that have pro- or anti-inflammatory activity, adding a modulatory role for this cell, previously known as a suicide effector. The presence of cytokines almost always implies intercellular modulation, potentially unmasking interactions of neutrophils with other immune cells. In fact, neutrophils have been found to help B cells and to modulate dendritic cell (DC), macrophage, and T-cell activities. In this review, we describe some ways in which neutrophils influence the inflammatory environment in infection, cancer, and autoimmunity, regulating both innate and adaptive immune responses. These cells can switch phenotypes and exert functions beyond cytotoxicity against invading pathogens, extending the view of neutrophils beyond suicide effectors to include functions as regulatory and suppressor cells. PMID:26108096

  19. CFTR targeting during activation of human neutrophils.

    PubMed

    Ng, Hang Pong; Valentine, Vincent G; Wang, Guoshun

    2016-12-01

    Cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-activated chloride channel, plays critical roles in phagocytic host defense. However, how activated neutrophils regulate CFTR channel distribution subcellularly is not well defined. To investigate, we tested multiple Abs against different CFTR domains, to examine CFTR expression in human peripheral blood neutrophils by flow cytometry. The data confirmed that resting neutrophils had pronounced CFTR expression. Activation of neutrophils with soluble or particulate agonists did not significantly increase CFTR expression level, but induced CFTR redistribution to cell surface. Such CFTR mobilization correlated with cell-surface recruitment of formyl-peptide receptor during secretory vesicle exocytosis. Intriguingly, neutrophils from patients with ΔF508-CF, despite expression of the mutant CFTR, showed little cell-surface mobilization upon stimulation. Although normal neutrophils effectively targeted CFTR to their phagosomes, ΔF508-CF neutrophils had impairment in that process, resulting in deficient hypochlorous acid production. Taken together, activated neutrophils regulate CFTR distribution by targeting this chloride channel to the subcellular sites of activation, and ΔF508-CF neutrophils fail to achieve such targeting, thus undermining their host defense function.

  20. Coagulation induced by C3aR-dependent NETosis drives protumorigenic neutrophils during small intestinal tumorigenesis

    PubMed Central

    Guglietta, Silvia; Chiavelli, Andrea; Zagato, Elena; Krieg, Carsten; Gandini, Sara; Ravenda, Paola Simona; Bazolli, Barbara; Lu, Bao; Penna, Giuseppe; Rescigno, Maria

    2016-01-01

    Excessive activation of blood coagulation and neutrophil accumulation have been described in several human cancers. However, whether hypercoagulation and neutrophilia are linked and involved in cancer development is currently unknown. Here we show that spontaneous intestinal tumorigenesis correlates with the accumulation of low-density neutrophils with a pro-tumorigenic N2 phenotype and unprompted neutrophil extracellular traps (NET) formation. We find that increased circulating lipopolysaccharide induces upregulation of complement C3a receptor on neutrophils and activation of the complement cascade. This leads to NETosis, induction of coagulation and N2 polarization, which prompts tumorigenesis, showing a novel link between coagulation, neutrophilia and complement activation. Finally, in a cohort of patients with small but not large intestinal cancer, we find a correlation between neutrophilia and hypercoagulation. This study provides a mechanistic explanation for the tumour-promoting effects of hypercoagulation, which could be used as a new biomarker or as a therapeutic target. PMID:26996437

  1. Phenotype and function of tumor-associated neutrophils and their subsets in early-stage human lung cancer.

    PubMed

    Eruslanov, Evgeniy B

    2017-03-10

    Neutrophils accumulate in many types of human and murine tumors and represent a significant portion of tumor-infiltrating myeloid cells. Our current understanding of the role of neutrophils in tumor development has depended primarily on murine models of cancer. However, there are crucial species differences in the evolution of tumors, genetic diversity, immune and inflammatory responses, and intrinsic biology of neutrophils that might have a profound impact on the tumor development and function of neutrophils in mouse versus human tumors. To date, the majority of experimental approaches to study neutrophils in cancer patients have been limited to the examination of circulating blood neutrophils. The phenotype and function of tumor-associated neutrophils (TANs) in humans, particularly in the early stages of tumor development, have not been extensively investigated. Thus, the long-term goal of our work has been to characterize human TANs and determine their specific role in tumor development. Here, we summarize our findings on human TANs obtained from human early stage lung cancer patients. We will describe the phenotypes of different TAN subsets identified in early stage lung tumors, as well as their functional dialog with T cells.

  2. Inhibitor of in vitro neutrophil migration in sera of children with homozygous sickle cell gene during pain crisis.

    PubMed

    Akenzua, G I; Amiengheme, O R

    1981-03-01

    There is conflicting evidence for a causal relationship between infection and haematological crisis of sickle cell disease. To find out whether changes in leucotaxis occur during pain crisis, in-vitro neutrophil migration was determined in 38 children with Hb SS during steady state and during pain crisis. Migrations of neutrophils of sickle cell patients was 29 +/ 12 microns in steady state and 27.5 +/- 10.5 microns during pain crisis. These rates were comparable to migration of neutrophils of control children with normal haemoglobin of 34 +/- 9.6 microns. However, with addition of autologous serum to the cell suspension, neutrophil migration of patients in pain crisis was significantly retarded (16 +/- 13 microns) as compared to those in steady state (26 +/- 10.2 microns) and control children (28.7 +/- 10 microns). Sera of children in pain crisis also inhibited migration of neutrophils of healthy adults with normal Hb. Pooled normal plasma reversed inhibitory action of pain crisis serum on autologous and homologous neutrophil migration; but pain crisis plasma did not. Chemotactic effect of sera of Hb SS children in steady state or pain crisis and control children on neutrophils of eight adults with normal Hb were similar and comparable to that of pooled normal serum. Thus, children with sickle cell disease develop chemotactic inhibitor(s) in their circulation during pain crisis. They may lead to defective leucotaxis and enhanced susceptibility to infection.

  3. Phenotypic and functional characteristics of HLA-DR(+) neutrophils in Brazilians with cutaneous leishmaniasis.

    PubMed

    Davis, Richard E; Sharma, Smriti; Conceição, Jacilara; Carneiro, Pedro; Novais, Fernanda; Scott, Phillip; Sundar, Shyam; Bacellar, Olivia; Carvalho, Edgar M; Wilson, Mary E

    2017-03-01

    The protozoan Leishmania braziliensis causes cutaneous leishmaniasis (CL) in endemic regions. In murine models, neutrophils (PMNs) are recruited to the site of infection soon after parasite inoculation. However, the roles of neutrophils during chronic infection and in human disease remain undefined. We hypothesized that neutrophils help maintain a systemic inflammatory state in subjects with CL. Lesion biopsies from all patients with CL tested contained neutrophils expressing HLA-DR, a molecule thought to be restricted to professional antigen-presenting cells. Although CL is a localized disease, a subset of patients with CL also had circulating neutrophils expressing HLA-DR and the costimulatory molecules CD80, CD86, and CD40. PMNs isolated from a low-density leukocyte blood fraction (LD-PMNs) contained a higher percentage of HLA-DR(+) PMNs than did normal-density PMNs. In vitro coculture experiments suggested LD-PMNs do not suppress T cell responses, differentiating them from MDSCs. Flow-sorted HLA-DR(+) PMNs morphologically resembled conventional PMNs, and they exhibited functional properties of PMNs. Compared with conventional PMNs, HLA-DR(+) PMNs showed increased activation, degranulation, DHR123 oxidation, and phagocytic capacity. A few HLA-DR(+) PMNs were observed in healthy subjects, and that proportion could be increased by incubation in either inflammatory cytokines or in plasma from a patient with CL. This was accompanied by an increase in PMN hladrb1 mRNA, suggesting a possible connection between neutrophil "priming" and up-regulation of HLA-DR. These data suggest that PMNs that are primed for activation and that also express surface markers of antigen-presenting cells emerge in the circulation and infected tissue lesions of patients with CL.

  4. Serum and glucocorticoid-regulated kinase 1 regulates neutrophil clearance during inflammation resolution.

    PubMed

    Burgon, Joseph; Robertson, Anne L; Sadiku, Pranvera; Wang, Xingang; Hooper-Greenhill, Edward; Prince, Lynne R; Walker, Paul; Hoggett, Emily E; Ward, Jonathan R; Farrow, Stuart N; Zuercher, William J; Jeffrey, Philip; Savage, Caroline O; Ingham, Philip W; Hurlstone, Adam F; Whyte, Moira K B; Renshaw, Stephen A

    2014-02-15

    The inflammatory response is integral to maintaining health by functioning to resist microbial infection and repair tissue damage. Large numbers of neutrophils are recruited to inflammatory sites to neutralize invading bacteria through phagocytosis and the release of proteases and reactive oxygen species into the extracellular environment. Removal of the original inflammatory stimulus must be accompanied by resolution of the inflammatory response, including neutrophil clearance, to prevent inadvertent tissue damage. Neutrophil apoptosis and its temporary inhibition by survival signals provides a target for anti-inflammatory therapeutics, making it essential to better understand this process. GM-CSF, a neutrophil survival factor, causes a significant increase in mRNA levels for the known anti-apoptotic protein serum and glucocorticoid-regulated kinase 1 (SGK1). We have characterized the expression patterns and regulation of SGK family members in human neutrophils and shown that inhibition of SGK activity completely abrogates the antiapoptotic effect of GM-CSF. Using a transgenic zebrafish model, we have disrupted sgk1 gene function and shown this specifically delays inflammation resolution, without altering neutrophil recruitment to inflammatory sites in vivo. These data suggest SGK1 plays a key role in regulating neutrophil survival signaling and thus may prove a valuable therapeutic target for the treatment of inflammatory disease.

  5. Serum and Glucocorticoid Regulated Kinase 1 (SGK1) Regulates Neutrophil Clearance During Inflammation Resolution

    PubMed Central

    Burgon, Joseph; Robertson, Anne L.; Sadiku, Pranvera; Wang, Xingang; Hooper-Greenhill, Edward; Prince, Lynne R.; Walker, Paul; Hoggett, Emily E.; Ward, Jonathan R.; Farrow, Stuart N.; Zuercher, William J.; Jeffrey, Philip; Savage, Caroline O.; Ingham, Philip W.; Hurlstone, Adam F.; Whyte, Moira K. B.; Renshaw, Stephen A.

    2013-01-01

    The inflammatory response is integral to maintaining health, by functioning to resist microbial infection and repair tissue damage. Large numbers of neutrophils are recruited to inflammatory sites to neutralise invading bacteria through phagocytosis and the release of proteases and reactive oxygen species into the extracellular environment. Removal of the original inflammatory stimulus must be accompanied by resolution of the inflammatory response, including neutrophil clearance, to prevent inadvertent tissue damage. Neutrophil apoptosis and its temporary inhibition by survival signals provides a target for anti-inflammatory therapeutics, making it essential to better understand this process. GM-CSF, a neutrophil survival factor, causes a significant increase in mRNA levels for the known anti-apoptotic protein Serum and Glucocorticoid Regulated Kinase 1 (SGK1). We have characterised the expression patterns and regulation of SGK family members in human neutrophils, and shown that inhibition of SGK activity completely abrogates the anti-apoptotic effect of GM-CSF. Using a transgenic zebrafish model, we have disrupted sgk1 gene function and shown this specifically delays inflammation resolution, without altering neutrophil recruitment to inflammatory sites in vivo. These data suggest SGK1 plays a key role in regulating neutrophil survival signalling, and thus may prove a valuable therapeutic target for the treatment of inflammatory disease. PMID:24431232

  6. Galectin-3–null mice display defective neutrophil clearance during acute inflammation

    PubMed Central

    Wright, Rachael D; Souza, Patricia R.; Flak, Magdalena B.; Thedchanamoorthy, Prasheetha; Norling, Lucy V.; Cooper, Dianne

    2017-01-01

    Galectin-3 has been associated with a plethora of proinflammatory functions because of its ability, among others, to promote neutrophil activation and because of the reduction in neutrophil recruitment in models of infection in Gal-3-null mice. Conversely, it has also been linked to resolution of inflammation through its actions as an opsonin and its ability to promote efferocytosis of apoptotic neutrophils. Using a self-resolving model of peritonitis, we have addressed the modulation and role of Gal-3 in acute inflammation. We have shown that Gal-3 expression is increased in neutrophils that travel to the inflamed peritoneum and that cellular localization of this lectin is modulated during the course of the inflammatory response. Furthermore, neutrophil recruitment to the inflamed peritoneum is increased in Gal-3–null mice during the course of the response, and that correlates with reduced numbers of monocytes/macrophages in the cavities of those mice, as well as reduced apoptosis and efferocytosis of Gal-3–null neutrophils. These data indicate a role for endogenous Gal-3 in neutrophil clearance during acute inflammation. PMID:27733579

  7. Morphology and staining behavior of neutrophilic and eosinophilic granulocytes of the common marmoset (Callithrix jacchus).

    PubMed

    Bleyer, Martina; Curths, Christoph; Dahlmann, Franziska; Wichmann, Judy; Bauer, Natali; Moritz, Andreas; Braun, Armin; Knauf, Sascha; Kaup, Franz-Josef; Gruber-Dujardin, Eva

    2016-06-01

    Common marmosets (Callithrix jacchus) are frequently used as translational animal models for human diseases. However, a comparative study of cytological and histochemical detection methods as well as morphometric and ultrastructural characterization of neutrophils and eosinophils in this species is lacking. Blood samples of house dust mite sensitized and allergen challenged as well as lipopolysaccharide (LPS) challenged marmosets were analyzed with different cytological and histological staining methods. Furthermore, cell size and number of nuclear segments were compared between neutrophils and eosinophils. Electron microscopy was performed to characterize the ultrastructure of granulocytes. Of all applied cytological stains, three allowed differentiation of eosinophils and neutrophils and, thus, reliable quantification in blood smears: May-Grünwald-Giemsa stain, Congo Red and Naphthol AS-D Chloroacetate-Esterase. For histology, Hematoxylin-Eosin (H&E) could not demonstrate clear differences, whereas Sirius Red, Congo Red, and Naphthol AS-D Chloroacetate Esterase showed capable results for identification of eosinophils or neutrophils in lung tissue. Morphometry revealed that marmoset neutrophils have more nuclear segments and are slightly larger than eosinophils. Ultrastructurally, eosinophils presented with large homogeneous electron-dense granules without crystalloid cores, while neutrophils were characterized by heterogeneous granules of different size and density. Additionally, sombrero-like vesicles were detected in tissue eosinophils of atopic marmosets, indicative for hypersensitivity-related piecemeal degranulation. In conclusion, we provide a detailed overview of marmoset eosinophils and neutrophils, important for phenotypic characterization of marmoset models for human airway diseases.

  8. Neutrophil crawling in capillaries; a novel immune response to Staphylococcus aureus.

    PubMed

    Harding, Mark Geoffrey; Zhang, Kunyan; Conly, John; Kubes, Paul

    2014-10-01

    Methicillin-resistant Staphylococcus aureus (MRSA), particularly the USA300 strain, is a highly virulent pathogen responsible for an increasing number of skin and soft tissue infections globally. Furthermore, MRSA-induced soft tissue infections can rapidly progress into life-threatening conditions, such as sepsis and necrotizing fasciitis. The importance of neutrophils in these devastating soft tissue infections remains ambiguous, partly because of our incomplete understanding of their behaviour. Spinning disk confocal microscopy was used to visualize the behaviour of GR1-labelled neutrophils in subcutaneous tissue in response to GFP-expressing MRSA attached to a foreign particle (agarose bead). We observed significant directional neutrophil recruitment towards the S. aureus agarose bead but not a control agarose bead. A significant increase in neutrophil crawling within the capillaries surrounding the infectious nidus was noted, with impaired capillary perfusion in these vessels and increased parenchymal cell death. No neutrophils were able to emigrate from capillaries. The crawling within these capillaries was mediated by the β(2) and α(4) integrins and blocking these integrins 2 hours post infection eliminated neutrophil crawling, improved capillary perfusion, reduced cell death and reduced lesion size. Blocking prior to infection increased pathology. Neutrophil crawling within capillaries during MRSA soft tissue infections, while potentially contributing to walling off or preventing early dissemination of the pathogen, resulted in impaired perfusion and increased tissue injury with time.

  9. Neutrophil-derived JAML Inhibits Repair of Intestinal Epithelial Injury During Acute Inflammation

    PubMed Central

    Weber, Dominique A.; Sumagin, Ronen; McCall, Ingrid C.; Leoni, Giovanna; Neumann, Philipp A.; Andargachew, Rakieb; Brazil, Jennifer C.; Medina-Contreras, Oscar; Denning, Timothy L.; Nusrat, Asma; Parkos, Charles A.

    2014-01-01

    Neutrophil transepithelial migration (TEM) during acute inflammation is associated with mucosal injury. Using models of acute mucosal injury in-vitro and in-vivo, we describe a new mechanism by which neutrophils infiltrating the intestinal mucosa disrupt epithelial homeostasis. We report that junctional adhesion molecule-like protein (JAML) is cleaved from neutrophil surface by zinc-metalloproteases during TEM. Neutrophil-derived soluble JAML bound to the epithelial tight junction protein coxsackie-adenovirus receptor (CAR) resulting in compromised barrier and inhibition of wound repair, through decreased epithelial proliferation. The deleterious effects of JAML on barrier and wound repair were reversed with an anti-JAML mAb that inhibits JAML-CAR binding. Thus, JAML released from transmigrating neutrophils across inflamed epithelia can promote recruitment of leukocytes and aid in clearance of invading microorganisms. However, sustained release of JAML under pathologic conditions associated with persistence of large numbers of infiltrated neutrophil would compromise intestinal barrier and inhibit mucosal healing. Targeting JAML-CAR interactions may thus improve mucosal healing responses under conditions of dysregulated neutrophil recruitment. PMID:24621992

  10. Pathogen induced chemo-attractant hepoxilin A3 drives neutrophils, but not eosinophils across epithelial barriers.

    PubMed

    Kubala, S A; Patil, S U; Shreffler, W G; Hurley, B P

    2014-01-01

    Pathogen induced migration of neutrophils across mucosal epithelial barriers requires epithelial production of the chemotactic lipid mediator, hepoxilin A3 (HXA3). HXA3 is an eicosanoid derived from arachidonic acid. Although eosinophils are also capable of penetrating mucosal surfaces, eosinophilic infiltration occurs mainly during allergic processes whereas neutrophils dominate mucosal infection. Both neutrophils and eosinophils can respond to chemotactic gradients of certain eicosanoids, however, it is not known whether eosinophils respond to pathogen induced lipid mediators such as HXA3. In this study, neutrophils and eosinophils were isolated from human blood and placed on the basolateral side of polarized epithelial monolayers grown on permeable Transwell filters and challenged by various chemotactic gradients of distinct lipid mediators. We observed that both cell populations migrated across epithelial monolayers in response to a leukotriene B4 (LTB4) gradient, whereas only eosinophils migrated toward a prostaglandin D2 (PGD2) gradient. Interestingly, while pathogen induced neutrophil trans-epithelial migration was substantial, pathogen induced eosinophil trans-epithelial migration was not observed. Further, gradients of chemotactic lipids derived from pathogen infected epithelial cells known to be enriched for HXA3 as well as purified HXA3 drove significant numbers of neutrophils across epithelial barriers, whereas eosinophils failed to respond to these gradients. These data suggest that although the eicosanoid HXA3 serves as an important neutrophil chemo-attractant at mucosal surfaces during pathogenic infection, HXA3 does not appear to exhibit chemotactic activity toward eosinophils.

  11. Bacillary replication and macrophage necrosis are determinants of neutrophil recruitment in tuberculosis.

    PubMed

    Repasy, Teresa; Martinez, Nuria; Lee, Jinhee; West, Kim; Li, Wenjun; Kornfeld, Hardy

    2015-08-01

    We previously determined that burst size necrosis is the chief mode of mononuclear cell death in the lungs of mice with tuberculosis. The present study explored the link between infection-induced necrosis of mononuclear phagocytes and neutrophil accumulation in the lungs of mice challenged with one of four Mycobacterium tuberculosis strains of increasing virulence (RvΔphoPR mutant, H37Ra, H37Rv and Erdman). At all time points studied, Erdman produced the highest bacterial load and the highest proportion and number of M. tuberculosis-infected neutrophils. These parameters, and the proportion of TUNEL-positive cells, tracked with virulence across all strains tested. Differences in neutrophil infection were not reflected by levels of chemoattractant cytokines in bronchoalveolar lavage fluid, while interferon-γ (reported to suppress neutrophil trafficking to the lung in tuberculosis) was highest in Erdman-infected mice. Treating Erdman-infected mice with ethambutol decreased the proportion of mononuclear phagocytes with high bacterial burden and the ratio of infected neutrophils to infected mononuclear cells in a dose-dependent manner. We propose that faster replicating M. tuberculosis strains cause more necrosis which in turn promotes neutrophil recruitment. Neutrophils infected with M. tuberculosis constitute a biomarker for poorly controlled bacterial replication, infection-induced mononuclear cell death, and increased severity of immune pathology in tuberculosis.

  12. Transendothelial migration enhances integrin-dependent human neutrophil chemokinesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transendothelial migration of neutrophils induces phenotypic changes that influence the interactions of neutrophils with extravascular tissue components. To assess the influence of transmigration on neutrophil chemokinetic motility, we used polyethylene glycol hydrogels covalently modified with spec...

  13. Neutrophil-induced injury of rat pulmonary alveolar epithelial cells.

    PubMed Central

    Simon, R H; DeHart, P D; Todd, R F

    1986-01-01

    The damage to pulmonary alveolar epithelial cells that occurs in many inflammatory conditions is thought to be caused in part by phagocytic neutrophils. To investigate this process, we exposed monolayers of purified rat alveolar epithelial cells to stimulated human neutrophils and measured cytotoxicity using a 51Cr-release assay. We found that stimulated neutrophils killed epithelial cells by a process that did not require neutrophil-generated reactive oxygen metabolites. Pretreatment of neutrophils with an antibody (anti-Mo1) that reduced neutrophil adherence to epithelial cells limited killing. Although a variety of serine protease inhibitors partially inhibited cytotoxicity, we found that neutrophil cytoplasts, neutrophil lysates, neutrophil-conditioned medium, purified azurophilic or specific granule contents, and purified human neutrophil elastase did not duplicate the injury. We conclude that stimulated neutrophils can kill alveolar epithelial cells in an oxygen metabolite-independent manner. Tight adherence of stimulated neutrophils to epithelial cell monolayers appears to promote epithelial cell killing. Images PMID:3771800

  14. Bromelain treatment decreases neutrophil migration to sites of inflammation.

    PubMed

    Fitzhugh, David J; Shan, Siqing; Dewhirst, Mark W; Hale, Laura P

    2008-07-01

    Bromelain, a mixture of proteases derived from pineapple stem, has been reported to have therapeutic benefits in a variety of inflammatory diseases, including murine inflammatory bowel disease. The purpose of this work was to understand potential mechanisms for this anti-inflammatory activity. Exposure to bromelain in vitro has been shown to remove a number of cell surface molecules that are vital to leukocyte trafficking, including CD128a/CXCR1 and CD128b/CXCR2 that serve as receptors for the neutrophil chemoattractant IL-8 and its murine homologues. We hypothesized that specific proteolytic removal of CD128 molecules by bromelain would inhibit neutrophil migration to IL-8 and thus decrease acute responses to inflammatory stimuli. Using an in vitro chemotaxis assay, we demonstrated a 40% reduction in migration of bromelain- vs. sham-treated human neutrophils in response to rhIL-8. Migration to the bacterial peptide analog fMLP was unaffected, indicating that bromelain does not induce a global defect in leukocyte migration. In vivo bromelain treatment generated a 50-85% reduction in neutrophil migration in 3 different murine models of leukocyte migration into the inflamed peritoneal cavity. Intravital microscopy demonstrated that although in vivo bromelain treatment transiently decreased leukocyte rolling, its primary long-term effect was abrogation of firm adhesion of leukocytes to blood vessels at the site of inflammation. These changes in adhesion were correlated with rapid re-expression of the bromelain-sensitive CD62L/L-selectin molecules that mediate rolling following in vivo bromelain treatment and minimal re-expression of CD128 over the time period studied. Taken together, these studies demonstrate that bromelain can effectively decrease neutrophil migration to sites of acute inflammation and support the specific removal of the CD128 chemokine receptor as a potential mechanism of action.

  15. Bromelain Treatment Decreases Neutrophil Migration to Sites of Inflammation

    PubMed Central

    Fitzhugh, David J.; Shan, Siqing; Dewhirst, Mark W.; Hale, Laura P.

    2008-01-01

    Bromelain, a mixture of proteases derived from pineapple stem, has been reported to have therapeutic benefits in a variety of inflammatory diseases, including murine inflammatory bowel disease. The purpose of this work was to understand potential mechanisms for this anti-inflammatory activity. Exposure to bromelain in vitro has been shown to remove a number of cell surface molecules that are vital to leukocyte trafficking, including CD128a/CXCR1 and CD128b/CXCR2 that serve as receptors for the neutrophil chemoattractant IL-8 and its murine homologues. We hypothesized that specific proteolytic removal of CD128 molecules by bromelain would inhibit neutrophil migration to IL-8 and thus decrease acute responses to inflammatory stimuli. Using an in vitro chemotaxis assay, we demonstrated a 40% reduction in migration of bromelain- vs. sham-treated human neutrophils in response to rhIL-8. Migration to the bacterial peptide analog fMLP was unaffected, indicating that bromelain does not induce a global defect in leukocyte migration. In vivo bromelain treatment generated a 50 – 85% reduction in neutrophil migration in 3 different murine models of leukocyte migration into the inflamed peritoneal cavity. Intravital microscopy demonstrated that although in vivo bromelain treatment transiently decreased leukocyte rolling, its primary long-term effect was abrogation of firm adhesion of leukocytes to blood vessels at the site of inflammation. These changes in adhesion were correlated with rapid re-expression of the bromelain-sensitive CD62L/L-selectin molecules that mediate rolling following in vivo bromelain treatment and minimal re-expression of CD128 over the time period studied. Taken together, these studies demonstrate that bromelain can effectively decrease neutrophil migration to sites of acute inflammation and support the specific removal of the CD128 chemokine receptor as a potential mechanism of action. PMID:18482869

  16. On the maturation rate of the neutrophil.

    PubMed

    Zajicek, G; Shohat, M; Polliack, A

    1984-05-01

    Fifty-three maturing bone marrow cells of the granulocyte cell series stained with Giemsa stain and magnified 1,000 times were scanned by a "computerized microscope" consisting of a LSI-11/23 microprocessor and a black-and-white video camera attached to a "frame grabber ." Each sampled cell was digitized into 70 X 70 pixels, each pixel representing 0.04 micron of the real image. The pixel gray values ranged between 0 and 255. Zero stood for white, 255 represented black, while the numbers in between stood for the various shades of gray. The cells represented six different stages of granulocytic maturation: myeloblast, promyelocyte, myelocyte, metamyelocyte , band form, and polymorphonuclear granulocyte. A discriminant analysis program selected 19 features best distinguishing between the six different cell types and computed five canonical discriminant functions defining a Space in which maturation was studied. In the Space, distance between two cells serves as a measure of similarity. The closer two cells are, the more similar they are and vice versa. This measure was applied here to express the degree of similarity between the neutrophil maturation classes, and since they represent states in the neutrophil life history, it is applicable also as a yardstick for the quantitation of differentiation. In the Space, the life history of a cell is represented by a trajectory originating in the myeloblast and terminating in the granulocyte state. Displacement along the trajectory represents cell maturation that is expressed relatively to the least differentiated state of the myeloblast. The further a cell from this state the more mature it is. The same yardstick also serves for differentiation rate estimates represented in the Space by displacement velocities that are derived from the known "transit times" of a cell in each state. The methodology is also applied for cell production estimates. Unlike other "computerized microscopes" serving for cell classification, the

  17. IL-1 Coordinates the Neutrophil Response to C. albicans in the Oral Mucosa

    PubMed Central

    Altmeier, Simon; Toska, Albulena; Sparber, Florian; Teijeira, Alvaro; Halin, Cornelia; LeibundGut-Landmann, Salomé

    2016-01-01

    Mucosal infections with Candida albicans belong to the most frequent forms of fungal diseases. Host protection is conferred by cellular immunity; however, the induction of antifungal immunity is not well understood. Using a mouse model of oropharyngeal candidiasis (OPC) we show that interleukin-1 receptor (IL-1R) signaling is critical for fungal control at the onset of infection through its impact on neutrophils at two levels. We demonstrate that both the recruitment of circulating neutrophils to the site of infection and the mobilization of newly generated neutrophils from the bone marrow depended on IL-1R. Consistently, IL-1R-deficient mice displayed impaired chemokine production at the site of infection and defective secretion of granulocyte colony-stimulating factor (G-CSF) in the circulation in response to C. albicans. Strikingly, endothelial cells were identified as the primary cellular source of G-CSF during OPC, which responded to IL-1α that was released from keratinocytes in the infected tissue. The IL-1-dependent crosstalk between two different cellular subsets of the nonhematopoietic compartment was confirmed in vitro using a novel murine tongue-derived keratinocyte cell line and an established endothelial cell line. These data establish a new link between IL-1 and granulopoiesis in the context of fungal infection. Together, we identified two complementary mechanisms coordinating the neutrophil response in the oral mucosa, which is critical for preventing fungal growth and dissemination, and thus protects the host from disease. PMID:27632536

  18. Role of Transient Receptor Potential Vanilloid 4 in Neutrophil Activation and Acute Lung Injury.

    PubMed

    Yin, Jun; Michalick, Laura; Tang, Christine; Tabuchi, Arata; Goldenberg, Neil; Dan, Qinghong; Awwad, Khader; Wang, Liming; Erfinanda, Lasti; Nouailles, Geraldine; Witzenrath, Martin; Vogelzang, Alexis; Lv, Lu; Lee, Warren L; Zhang, Haibo; Rotstein, Ori; Kapus, Andras; Szaszi, Katalin; Fleming, Ingrid; Liedtke, Wolfgang B; Kuppe, Hermann; Kuebler, Wolfgang M

    2016-03-01

    The cation channel transient receptor potential vanilloid (TRPV) 4 is expressed in endothelial and immune cells; however, its role in acute lung injury (ALI) is unclear. The functional relevance of TRPV4 was assessed in vivo, in isolated murine lungs, and in isolated neutrophils. Genetic deficiency of TRPV4 attenuated the functional, histological, and inflammatory hallmarks of acid-induced ALI. Similar protection was obtained with prophylactic administration of the TRPV4 inhibitor, GSK2193874; however, therapeutic administration of the TRPV4 inhibitor, HC-067047, after ALI induction had no beneficial effect. In isolated lungs, platelet-activating factor (PAF) increased vascular permeability in lungs perfused with trpv4(+/+) more than with trpv4(-/-) blood, independent of lung genotype, suggesting a contribution of TRPV4 on blood cells to lung vascular barrier failure. In neutrophils, TRPV4 inhibition or deficiency attenuated the PAF-induced increase in intracellular calcium. PAF induced formation of epoxyeicosatrienoic acids by neutrophils, which, in turn, stimulated TRPV4-dependent Ca(2+) signaling, whereas inhibition of epoxyeicosatrienoic acid formation inhibited the Ca(2+) response to PAF. TRPV4 deficiency prevented neutrophil responses to proinflammatory stimuli, including the formation of reactive oxygen species, neutrophil adhesion, and chemotaxis, putatively due to reduced activation of Rac. In chimeric mice, however, the majority of protective effects in acid-induced ALI were attributable to genetic deficiency of TRPV4 in parenchymal tissue, whereas TRPV4 deficiency in circulating blood cells primarily reduced lung myeloperoxidase activity. Our findings identify TRPV4 as novel regulator of neutrophil activation and suggest contributions of both parenchymal and neutrophilic TRPV4 in the pathophysiology of ALI.

  19. Superoxide Induces Neutrophil Extracellular Trap Formation in a TLR-4 and NOX-Dependent Mechanism

    PubMed Central

    Al-Khafaji, Ahmed B; Tohme, Samer; Yazdani, Hamza Obaid; Miller, David; Huang, Hai; Tsung, Allan

    2016-01-01

    Neutrophils constitute the early innate immune response to perceived infectious and sterile threats. Neutrophil extracellular traps (NETs) are a novel mechanism to counter pathogenic invasion and sequelae of ischemia, including cell death and oxidative stress. Superoxide is a radical intermediate of oxygen metabolism produced by parenchymal and nonparenchymal hepatic cells, and is a hallmark of oxidative stress after liver ischemia-reperfusion (I/R). While extracellular superoxide recruits neutrophils to the liver and initiates sterile inflammatory injury, it is unknown whether superoxide induces the formation of NETs. We hypothesize that superoxide induces NET formation through a signaling cascade involving Toll-like receptor 4 (TLR-4) and neutrophil NADPH oxidase (NOX). We treated neutrophils with extracellular superoxide and observed NET DNA release, histone H3 citrullination and increased levels of MPO-DNA complexes occurring in a TLR-4–dependent manner. Inhibition of superoxide generation by allopurinol and inhibition of NOX by diphenyleneiodonium prevented NET formation. When mice were subjected to warm liver I/R, we found significant NET formation associated with liver necrosis and increased serum ALT in TLR-4 WT but not TLR-4 KO mice. To reduce circulating superoxide, we pretreated mice undergoing I/R with allopurinol and N-acetylcysteine, which resulted in decreased NETs and ameliorated liver injury. Our study demonstrates a requirement for TLR-4 and NOX in superoxide-induced NETs, and suggests involvement of superoxide-induced NETs in pathophysiologic settings. PMID:27453505

  20. Neutrophil heterogeneity in health and disease: a revitalized avenue in inflammation and immunity

    PubMed Central

    Beyrau, Martina; Bodkin, Jennifer Victoria; Nourshargh, Sussan

    2012-01-01

    Leucocytes form the principal cellular components of immunity and inflammation, existing as multiple subsets defined by distinct phenotypic and functional profiles. To date, this has most notably been documented for lymphocytes and monocytes. In contrast, as neutrophils are traditionally considered, to be short-lived, terminally differentiated cells that do not re-circulate, the potential existence of distinct neutrophil subsets with functional and phenotypic heterogeneity has not been widely considered or explored. A growing body of evidence is now challenging this scenario, and there is significant evidence for the existence of different neutrophil subsets under both physiological and pathological conditions. This review will summarize the key findings that have triggered a renewed interest in neutrophil phenotypic changes, both in terms of functional implications and consequences within disease models. Special emphasis will be placed on the potential pro- and anti-inflammatory roles of neutrophil subsets, as indicated by the recent works in models of ischaemia–reperfusion injury, trauma, cancer and sepsis. PMID:23226600

  1. Carbohydrate beverage ingestion and neutrophil degranulation responses following cycling to fatigue at 75% VO2 max.

    PubMed

    Bishop, N C; Blannin, A K; Walsh, N P; Gleeson, M

    2001-04-01

    Carbohydrate (CHO) beverage ingestion appears to influence neutrophil functional responses to prolonged exercise of a fixed duration. The aim of this randomised study was to examine the effect of CHO (5% w/v) beverage ingestion on lipopolysaccharide (LPS)-stimulated neutrophil degranulation responses in nine recreationally active males who cycled at 75% VO2 max until fatigue. On two separate occasions, subjects ingested either placebo (PLA) or CHO beverages before and at 15 min intervals during the exercise. Subjects exercised for 31% longer on the CHO trial compared with the PLA trial (P < 0.05). At fatigue plasma glucose concentration was significantly lower on the PLA trial compared with the CHO trial (P < 0.05). Plasma cortisol concentrations had increased similarly on both trials at this time. A marked neutrophilia was evident at fatigue and throughout the 4 h recovery period, the magnitude of which was similar on both trials. At fatigue LPS-stimulated elastase release per neutrophil had fallen similarly on both trials compared with pre-exercise values (47% and 50% on the PLA and CHO trials, respectively). In conclusion, our results suggest that CHO beverage ingestion has negligible influence on the hormonal, circulating neutrophil and LPS-stimulated neutrophil degranulation responses when exercise is performed to fatigue.

  2. Respiratory Syncytial Virus Fusion Protein Promotes TLR-4–Dependent Neutrophil Extracellular Trap Formation by Human Neutrophils

    PubMed Central

    Funchal, Giselle A.; Jaeger, Natália; Czepielewski, Rafael S.; Machado, Mileni S.; Muraro, Stéfanie P.; Stein, Renato T.; Bonorino, Cristina B. C.; Porto, Bárbara N.

    2015-01-01

    Acute viral bronchiolitis by Respiratory Syncytial Virus (RSV) is the most common respiratory illness in children in the first year of life. RSV bronchiolitis generates large numbers of hospitalizations and an important burden to health systems. Neutrophils and their products are present in the airways of RSV-infected patients who developed increased lung disease. Neutrophil Extracellular Traps (NETs) are formed by the release of granular and nuclear contents of neutrophils in the extracellular space in response to different stimuli and recent studies have proposed a role for NETs in viral infections. In this study, we show that RSV particles and RSV Fusion protein were both capable of inducing NET formation by human neutrophils. Moreover, we analyzed the mechanisms involved in RSV Fusion protein-induced NET formation. RSV F protein was able to induce NET release in a concentration-dependent fashion with both neutrophil elastase and myeloperoxidase expressed on DNA fibers and F protein-induced NETs was dismantled by DNase treatment, confirming that their backbone is chromatin. This viral protein caused the release of extracellular DNA dependent on TLR-4 activation, NADPH Oxidase-derived ROS production and ERK and p38 MAPK phosphorylation. Together, these results demonstrate a coordinated signaling pathway activated by F protein that led to NET production. The massive production of NETs in RSV infection could aggravate the inflammatory symptoms of the infection in young children and babies. We propose that targeting the binding of TLR-4 by F protein could potentially lead to novel therapeutic approaches to help control RSV-induced inflammatory consequences and pathology of viral bronchiolitis. PMID:25856628

  3. Myeloid-specific deletion of tumor suppressor PTEN augments neutrophil transendothelial migration during inflammation.

    PubMed

    Sarraj, Bara; Massberg, Steffen; Li, Yitang; Kasorn, Anongnard; Subramanian, Kulandayan; Loison, Fabien; Silberstein, Leslie E; von Andrian, Ulrich; Luo, Hongbo R

    2009-06-01

    Phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) is a second messenger that is involved in a number of cell activities including cell growth, proliferation, and motility. PIP(3) is produced by PI3K and regulated by PTEN (phosphatase and tensin homolog deleted on chromosome 10) and SHIP lipid phosphatases. Evidence from our experiments shows that enhanced PIP(3) production results in elevated neutrophil recruitment under inflammatory conditions. However, the mechanism of this elevation is not well understood. We used intravital video microscopy to investigate neutrophil recruitment in the cremaster venules of wild-type and PTEN knockout (KO) mice. Neutrophil transmigration was augmented in PTEN KO mice 4 h after TNF-alpha intrascrotal injection. PTEN KO neutrophils also showed significantly enhanced transmigration 2 h after MIP-2 intrascrotal injection, an effect that dramatically decreased when PI3K or Src kinase inhibitor treatments preceded MIP-2 stimulation. Similarly, fMLP superfusion of the cremaster muscle lead to enhanced emigration in PTEN KO mice. The observed elevation in neutrophil emigration was likely caused by increased speed of crawling, crossing the venular wall, and migrating through the muscular tissue in PTEN KO mice because the effect of PTEN depletion on neutrophil rolling or adhesion was minimal. Interestingly, chemoattractant-induced release of gelatinase and elastase was also elevated in PTEN null neutrophils, providing a potential mechanism for the enhanced neutrophil migration in the PTEN KO mice. Collectively, these results demonstrate that PTEN deletion in neutrophils enhances their invasivity and recruitment to inflamed sites more likely by raising the cell physical capability to cross the vascular and tissue barriers.

  4. Chronic neutrophilic leukaemia and plasma cell-related neutrophilic leukaemoid reactions.

    PubMed

    Bain, Barbara J; Ahmad, Shahzaib

    2015-11-01

    Many cases reported as 'chronic neutrophilic leukaemia' have had an associated plasma cell neoplasm. Recent evidence suggests that the great majority of such cases represent a neutrophilic leukaemoid reaction to the underlying multiple myeloma or monoclonal gammopathy of undetermined significance. We have analysed all accessible reported cases to clarify the likely diagnosis and to ascertain whether toxic granulation, Döhle bodies and an increased neutrophil alkaline phosphatase score were useful in making a distinction between chronic neutrophilic leukaemia and a neutrophilic leukaemoid reaction. We established that all these changes occur in both conditions. Toxic granulation and Döhle bodies are more consistently present in leukaemoid reactions but also occur quite frequently in chronic neutrophilic leukaemia. The neutrophil alkaline phosphatase score is increased in both conditions and is of no value in making a distinction.

  5. Alcohol-related changes in the intestinal microbiome influence neutrophil infiltration, inflammation and steatosis in early alcoholic hepatitis in mice

    PubMed Central

    Satishchandran, Abhishek; Iracheta-Vellve, Arvin; Ambade, Aditya; Kodys, Karen; Catalano, Donna; Ward, Doyle V.; Szabo, Gyongyi

    2017-01-01

    Background Alcohol-induced intestinal dysbiosis disrupts homeostatic gut-liver axis function and is essential in the development of alcoholic liver disease. Here, we investigate changes in enteric microbiome composition in a model of early alcoholic steatohepatitis and dissect the pathogenic role of intestinal microbes in alcohol-induced liver pathology. Materials and methods Wild type mice received a 10-day diet that was either 5% alcohol-containing or an isocaloric control diet plus a single binge. 16S rDNA sequencing defined the bacterial communities in the cecum of alcohol- and pair-fed animals. Some mice were treated with an antibiotic cocktail prior to and throughout alcohol feeding. Liver neutrophils, cytokines and steatosis were evaluated. Results Acute-on-chronic alcohol administration induced shifts in various bacterial phyla in the cecum, including increased Actinobacteria and a reduction in Verrucomicrobia driven entirely by a reduction in the genus Akkermansia. Antibiotic treatment reduced the gut bacterial load and circulating bacterial wall component lipopolysaccharide (LPS). We found that bacterial load suppression prevented alcohol-related increases in the number of myeloperoxidase- (MPO) positive infiltrating neutrophils in the liver. Expression of liver mRNA tumor necrosis factor alpha (Tnfα), C-X-C motif chemokine ligand 1 (Cxcl1) and circulating protein monocyte chemoattractant protein-1 (MCP-1) were also reduced in antibiotic-treated alcohol-fed mice. Alcohol-induced hepatic steatosis measured by Oil-Red O staining was significantly reduced in antibiotic treated mice. Genes regulating lipid production and storage were also altered by alcohol and antibiotic treatment. Interestingly, antibiotic treatment did not protect from alcohol-induced increases in serum aminotransferases (ALT/AST). Conclusions Our data indicate that acute-on-chronic alcohol feeding alters the microflora at multiple taxonomic levels and identifies loss of Akkermansia as an

  6. Evasion of Neutrophil Killing by Staphylococcus aureus

    PubMed Central

    McGuinness, Will A.; Kobayashi, Scott D.; DeLeo, Frank R.

    2016-01-01

    Staphylococcus aureus causes many types of infections, ranging from self-resolving skin infections to severe or fatal pneumonia. Human innate immune cells, called polymorphonuclear leukocytes (PMNs or neutrophils), are essential for defense against S. aureus infections. Neutrophils are the most prominent cell type of the innate immune system and are capable of producing non-specific antimicrobial molecules that are effective at eliminating bacteria. Although significant progress has been made over the past few decades, our knowledge of S. aureus-host innate immune system interactions is incomplete. Most notably, S. aureus has the capacity to produce numerous molecules that are directed to protect the bacterium from neutrophils. Here we review in brief the role played by neutrophils in defense against S. aureus infection, and correspondingly, highlight selected S. aureus molecules that target key neutrophil functions. PMID:26999220

  7. Neutrophil uptake of vaccinia virus in vitro

    SciTech Connect

    West, B.C.; Eschete, M.L.; Cox, M.E.; King, J.W.

    1987-10-01

    We studied human neutrophils for uptake of vaccinia virus. Uptake was determined radiometrically and by electron microscopy. Vaccinia virus was labeled with /sup 14/C or /sup 3/H, incubated with neutrophils, and quantified in neutrophil pellets in a new radiometric phagocytosis assay. Better results were obtained from assays of (/sup 3/H)thymidine-labeled virus; uptake increased through 1 hr and then plateaued. Phagocytosis of 3H-labeled Staphylococcus aureus was normal. Uptake of virus was serum dependent. Hexose monophosphate shunt activity was measured by two methods. No /sup 14/CO/sub 2/ from (/sup 14/C)1-glucose accompanied uptake of vaccinia virus, in contrast to the respiratory burst accompanying bacterial phagocytosis. Electron microscopy showed intact to slightly digested intraphagolysosomal vaccinia virus. Pock reduction assay showed a decrease in viral content due to neutrophils until 6 hr of incubation, when a modest but significant increase was observed. Thus, neutrophil uptake of vaccinia virus is distinguished from bacterial phagocytosis.

  8. Desensitisation of neutrophil responses by systemic interleukin 8 in cystic fibrosis.

    PubMed Central

    Dai, Y.; Dean, T. P.; Church, M. K.; Warner, J. O.; Shute, J. K.

    1994-01-01

    BACKGROUND--Inflammation associated with neutrophil infiltration is a commonly observed feature of children with cystic fibrosis. Production of the major neutrophil chemotactic cytokine interleukin 8 (IL-8) is potentially of great importance in the pathology of cystic fibrosis. Concentrations of IL-8 in both sputum and bronchoalveolar lavage fluid have been found to be higher in children with cystic fibrosis than in controls. The IL-8 induced chemotactic response and numbers of IL-8 receptors on peripheral neutrophils obtained from children with cystic fibrosis have been compared with a control group of children. METHODS--Cells were isolated from 18 patients with cystic fibrosis (aged 4-20 years) and 13 controls (aged 5-12 years) by dextran centrifugation followed by separation on Lymphoprep. Chemotaxis was assayed using multiwell microchemotaxis chambers and 5 microns polycarbonate filters. Filters were fixed and stained with Haema-Gurr for counting. Results were expressed as numbers of neutrophils per high power field (HPF). RESULTS--At the optimum concentration (1 x 10(-8) mol/l) the number of cells migrating were similar for controls (150 (12)/HPF) and for the cystic fibrosis group (140 (14)/HPF)). At lower concentrations the numbers of neutrophils migrating were lower for the cystic fibrosis group. Scatchard analysis of 125I-labelled IL-8 binding revealed lower numbers of receptors on neutrophils from patients with cystic fibrosis (22,000 per cell) than from controls (75,000 per cell). CONCLUSIONS--Reduced responsiveness to IL-8 of neutrophils from patients with cystic fibrosis is associated with receptor desensitisation as a result of exposure to high systemic levels of IL-8. PMID:7940424

  9. Clozapine induces oxidative stress and proapoptotic gene expression in neutrophils of schizophrenic patients.

    PubMed

    Fehsel, Karin; Loeffler, Stefan; Krieger, Klaus; Henning, Uwe; Agelink, Markus; Kolb-Bachofen, Victoria; Klimke, Ansgar

    2005-10-01

    The present study examined cellular effects of the atypical antipsychotic drug clozapine on blood cells of treated patients with and without clozapine-induced agranulocytosis (CA). Blood from one patient who commenced clozapine treatment was examined at weekly intervals for 128 days. Olanzapine-treated (n = 5) and polymedicated (n = 14) schizophrenic patients, as well as healthy subjects (n = 19) and septic shock patients (n = 8), were studied for comparison. We observed dramatically increased numbers of native neutrophils stained for superoxide anion production (P < or = 0.005, n = 10) and significantly elevated expression levels of the proapoptotic genes p53 (P < or = 0.020), bax alpha (P < or = 0.001), and bik (P < or = 0.002) in all tested non-CA patients (n = 19) and CA patients (n = 4). In non-CA patients, the expression of these genes did not correlate to the percentage of apoptotic neutrophils (2.0% +/- 1.3%), but in CA patients about 37% of the neutrophils show morphologic signs of apoptosis (P < or = 0.001). Under G-CSF therapy of CA, the number of apoptotic neutrophils and the expression of the proapoptotic genes decreased significantly. In conclusion, high production of reactive oxygen species in neutrophils of clozapine-treated patients, together with increased expression of proapoptotic genes, suggests that neutrophils are predisposed to apoptosis in schizophrenic patients under clozapine therapy. The correlation between drug and proapoptotic markers was highest for clozapine and bax alpha as well as superoxide anion radicals. This indicates oxidative mitochondrial stress in neutrophils of clozapine-treated patients which probably contributes to the induction of apoptosis and sudden loss of neutrophils and their precursors in CA patients.

  10. Neutrophil Extracellular Traps and Microcrystals

    PubMed Central

    2017-01-01

    Neutrophil extracellular traps represent a fascinating mechanism by which PMNs entrap extracellular microbes. The primary purpose of this innate immune mechanism is thought to localize the infection at an early stage. Interestingly, the ability of different microcrystals to induce NET formation has been recently described. Microcrystals are insoluble crystals with a size of 1–100 micrometers that have different composition and shape. Microcrystals have it in common that they irritate phagocytes including PMNs and typically trigger an inflammatory response. This review is the first to summarize observations with regard to PMN activation and NET release induced by microcrystals. Gout-causing monosodium urate crystals, pseudogout-causing calcium pyrophosphate dehydrate crystals, cholesterol crystals associated with atherosclerosis, silicosis-causing silica crystals, and adjuvant alum crystals are discussed. PMID:28373994

  11. Dynamic changes in circulating leukocytes during the induction of equine laminitis with black walnut extract.

    PubMed

    Hurley, David J; Parks, Robert J; Reber, Adrian J; Donovan, Douglas C; Okinaga, Tatsuyuki; Vandenplas, Michel L; Peroni, John F; Moore, James N

    2006-04-15

    Administration of black walnut heartwood extract (BWHE) via nasogastric tube induces acute laminitis in horses. However, the processes responsible for the development of laminitis, including laminitis induced with BWHE, remain unclear. The results of recent studies indicate that administration of BWHE initiates an inflammatory response in the laminar tissues and that this response may be due to extravasation of activated leukocytes from the circulation. This study examines the effects of BWHE administration on the dynamics of circulating neutrophils and monocytes, and the capacity of blood leukocytes to produce radical oxygen species (ROS) over the time period from administration of BWHE to the development of lameness consistent with Obel grade I laminitis. Individual horses, free of pre-existing musculoskeletal disease, were administered either 6l of BWHE or an equal volume of water at time 0 (T=0). Blood samples were collected prior to dosing and at 1, 2, 3, 4, 6, 8, 10 and 12h after dosing, or until the onset of Obel grade I laminitis. For each sample, total leukocyte counts were determined followed by collection of buffy coats and removal of erythrocytes by hypotonic lysis. Leukocytes were either fixed for flow cytometric assessment of differential counts or maintained in culture to measure endogenous and phorbol ester-induced production of ROS. At each sample time, the number of cells recovered and the flow cytometric differential counts were compared with corresponding total leukocyte counts determined by the Clinical Pathology laboratory. Horses administered BWHE had a significant reduction in circulating leukocytes at 3-4 h relative to values for horses administered the same volume of water. Horses that developed Obel grade I laminitis had a significant reduction in circulating leukocytes when compared to values for horses administered BWHE that did not become lame. Flow cytometric analysis revealed a consistent decrease in the total number of monocytes

  12. Fatty acid binding protein 4 in circulating leucocytes reflects atherosclerotic lesion progression in Apoe(-/-) mice.

    PubMed

    Agardh, Hanna E; Gertow, Karl; Salvado, Dolores M; Hermansson, Andreas; van Puijvelde, Gijs H; Hansson, Göran K; n-Berne, Gabrielle Paulsso; Gabrielsen, Anders

    2013-02-01

    Discovery of novel biomarkers for atherosclerosis is important to aid in early diagnosis of pre-symptomatic patients at high risk of cardiovascular events. The aim of the present study was therefore to identify potential biomarkers in circulating cells reflecting atherosclerotic lesion progression in the vessel wall. We performed gene arrays on circulating leucocytes from atherosclerosis prone Apoe(-/-) mice with increasing ages, using C57BL/6 mice as healthy controls. We identified fatty acid binding protein 4 (FABP4) mRNA to be augmented in mice with established disease compared with young Apoe(-/-) or controls. Interestingly, the transcript FABP4 correlated significantly with lesion size, further supporting a disease associated increase. In addition, validation of our finding on protein level showed augmented FABP4 in circulating leucocytes whereas, importantly, no change could be observed in plasma. Immunofluorescence analysis demonstrated FABP4 to be present mainly in circulating neutrophils and to some extent in monocytes. Moreover, FABP4-positive neutrophils and macrophages could be identified in the subintimal space in the plaque. Using human circulating leucocytes, we confirmed the presence of FABP4 protein in neutrophils and monocytes. In conclusion, we have showed that cellular levels of FABP4 in circulating leucocytes associate with lesion development in the experimental Apoe(-/-) model. The increased expression is primarily localized to neutrophils, but also in monocytes. We have identified FABP4 in leucocytes as a potential and easy accessible biomarker of atherosclerosis which could be of future clinical relevance.

  13. Emperipolesis of neutrophils by dysmorphic megakaryocytes.

    PubMed

    Parmley, R T; Kim, T H; Austin, R L; Alvarado, C S; Ragab, A H

    1982-12-01

    Neutrophil engulfment by megakaryocytes was observed within 20 to 30% of megakaryocytes from two children: one with metastatic rhabdomyosarcoma, the other with fever of unknown origin. Other cell types and neutrophil precursors were not observed within megakaryocytes. Only late megakaryocytes were involved in the process, and often these cells appeared vacuolated or degenerating at the light and electron microscope level. Ultrastructurally the engulfed neutrophils were intact and were within the open canalicular system of the megakaryocyte cytoplasm. No evidence of neutrophil granule exocytosis could be demonstrated in ultrastructural morphologic and peroxidase preparations; however, many neutrophils appeared to be endocytosing portions of the megakaryocyte cytoplasm. The phenomenon could not be transferred to normal marrow incubated with patient serum or plasma. Thus, our patients differ from previous observations of emperipolesis in: 1) the extreme frequency of the observation; 2) the selective involvement of neutrophils; and 3) the association of the anomaly with dysmorphic and/or disrupted megakaryocytes. These observations are consistent with a neutrophil response to altered and/or injured megakaryocytes.

  14. Characterization of arginase expression by equine neutrophils.

    PubMed

    Lavoie-Lamoureux, Anouk; Martin, James G; Lavoie, Jean-Pierre

    2014-02-15

    Neutrophils are the predominant cells recruited in the airways of horses suffering from heaves. These cells have been shown to express arginase in some species. The metabolism of l-arginine is thought to be involved in chronic inflammation, and airway obstruction and remodeling. The aim of this study was to assess the expression, regulation, activity, and functional role of arginase isoforms in equine neutrophils. Arginase I, arginase II, ornithine decarboxylase (ODC) and ornithine aminotransferase (OAT) expression were assessed in resting and stimulated (IL-4, LPS/fMLP, PMA; 5 and 18 h) blood neutrophils using quantitative PCR. Arginase expression was also studied by Western blot and enzyme activity assay. The effect of nor-NOHA (1mM), a specific arginase inhibitor, was assessed on arginase activity in vitro and ex vivo on neutrophil's inflammatory gene expression and viability. Results showed that equine neutrophils constitutively express arginase isoform 2, ODC and OAT. Neutrophil ex vivo stimulation did not induce arginase I or influence arginase II mRNA expression. Ex vivo inhibition of arginase activity by nor-NOHA had no effect on neutrophils inflammatory gene expression induced by LPS/fMLP (5h) but significantly reversed the cell loss observed after this stimulation.

  15. Exercise-induced extracellular 72 kDa heat shock protein (Hsp72) stimulates neutrophil phagocytic and fungicidal capacities via TLR-2.

    PubMed

    Giraldo, Esther; Martin-Cordero, Leticia; Garcia, Juan Jose; Gehrmann, Mathias; Gerhmann, Mathias; Multhoff, Gabriele; Ortega, Eduardo

    2010-01-01

    This study evaluated the role of toll like receptor 2 (TLR-2) in the interaction of 72 kDa extracellular heat shock protein (Hsp72, a stress-inducible protein) with neutrophils and the participation on TLR-2 in the stimulation of neutrophil phagocytic and fungicidal capacities by post-exercise physiological concentrations of Hsp72. Human peripheral blood neutrophils were incubated with fluorescein isothiocyanate-conjugated Hsp72, and were analyzed by immunofluorescence microscopy and flow cytometry. Both methods revealed an interaction of Hsp72 with neutrophils. In addition, when neutrophils were pre-incubated with an anti-TLR-2 antibody this interaction was clearly decreased. Post-exercise circulating concentration of Hsp72 (8.6 ng/ml) stimulated the phagocytic and fungicidal capacities of neutrophils and this effect could be also blocked using an antibody against TLR-2. Phosphatidylinositol-3-kinase (PI3K), extracellular signal-regulated kinase (ERK) and the nuclear transcription factor kappa beta (NF-kappabeta) were found to be involved in the signaling process, confirming the participation of TLR-2 in the stimulation of neutrophil function by Hsp72. In conclusion, TLR-2 is involved at least in part, in the stimulation of neutrophil phagocytic and fungicidal capacities induced by post-exercise physiological concentrations of Hsp72.

  16. Neutrophil infiltration increases matrix metalloproteinase-9 in the ischemic brain after occlusion/reperfusion of the middle cerebral artery in rats.

    PubMed

    Justicia, Carles; Panés, Julián; Solé, Sònia; Cervera, Alvaro; Deulofeu, Ramon; Chamorro, Angel; Planas, Anna M

    2003-12-01

    Matrix metalloproteinase-9 (MMP-9) activity increases in the brain during the first day after focal ischemia and might be involved in the pathogenesis of tissue damage. We previously showed MMP-9 in the extracellular space of brain parenchyma along with neutrophil recruitment after ischemia. In the present study, we tested whether neutrophils were a direct source of enhanced MMP-9 in the ischemic brain. Neutrophil infiltration was prevented either by injecting an antibody against ICAM-1, which abrogates neutrophil adhesion to the endothelial vessel wall, or by inducing neutropenia. One-hour intraluminal middle cerebral artery occlusion with reperfusion was induced, and studies were performed at 24 hours. Circulating neutrophils expressed 95-kDa MMP-9 and dimers, and infiltrated neutrophils stained positive for MMP-9. The expression of MMP-9 (mainly 95-kDa proform and dimers and, to a lesser extent, 88-kDa form) increased in brain after ischemia/reperfusion. Treatments preventing neutrophil infiltration failed to preclude the ischemia-induced increase in 88-kDa MMP-9 form and gelatinase activity in neurons and blood vessels. However, these treatments prevented the major increase in 95-kDa MMP-9 form and dimers. We conclude that neutrophil infiltration highly contributes to enhanced MMP-9 in the ischemic brain by releasing MMP-9 proform, which might participate in the tissular inflammatory reaction.

  17. Neutrophil-Mediated Phagocytosis of Staphylococcus aureus

    PubMed Central

    van Kessel, Kok P. M.; Bestebroer, Jovanka; van Strijp, Jos A. G.

    2014-01-01

    Initial elimination of invading Staphylococcus aureus from the body is mediated by professional phagocytes. The neutrophil is the major phagocyte of the innate immunity and plays a key role in the host defense against staphylococcal infections. Opsonization of the bacteria with immunoglobulins and complement factors enables efficient recognition by the neutrophil that subsequently leads to intracellular compartmentalization and killing. Here, we provide a review of the key processes evolved in neutrophil-mediated phagocytosis of S. aureus and briefly describe killing. As S. aureus is not helpless against the professional phagocytes, we will also highlight its immune evasion arsenal related to phagocytosis. PMID:25309547

  18. Platelet–neutrophil interactions under thromboinflammatory conditions

    PubMed Central

    Li, Jing; Kim, Kyungho; Barazia, Andrew; Tseng, Alan

    2015-01-01

    Platelets primarily mediate hemostasis and thrombosis, whereas leukocytes are responsible for immune responses. Since platelets interact with leukocytes at the site of vascular injury, thrombosis and vascular inflammation are closely intertwined and occur consecutively. Recent studies using real-time imaging technology demonstrated that platelet–neutrophil interactions on the activated endothelium are an important determinant of microvascular occlusion during thromboinflammatory disease in which inflammation is coupled to thrombosis. Although the major receptors and counter receptors have been identified, it remains poorly understood how heterotypic platelet–neutrophil interactions are regulated under disease conditions. This review discusses our current understanding of the regulatory mechanisms of platelet– neutrophil interactions in thromboinflammatory disease. PMID:25650236

  19. S. aureus blocks efferocytosis of neutrophils by macrophages through the activity of its virulence factor alpha toxin

    PubMed Central

    Cohen, Taylor S.; Jones-Nelson, Omari; Hotz, Meghan; Cheng, Lily; Miller, Lloyd S.; Suzich, JoAnn; Stover, C. Kendall; Sellman, Bret R.

    2016-01-01

    Bacterial pneumonia, such as those caused by Staphylococcus aureus, is associated with an influx of inflammatory neutrophils into the lung tissue and airways. Regulation and clearance of recruited neutrophils is essential for preventing tissue damage by “friendly fire”, a responsibility of macrophages in a process called efferocytosis. We hypothesized that S. aureus impairs efferocytosis by alveolar macrophages (AMs) through the activity of the secreted virulence factor alpha toxin (AT), which has been implicated in altering the antimicrobial function of AMs. Infection of mice lacking AMs resulted in significantly increased numbers of neutrophils in the lung, while clearance of neutrophils delivered intranasally into uninfected mice was reduced in AM depleted animals. In vitro, sublytic levels of AT impaired uptake of apoptotic neutrophils by purified AMs. In vivo, the presence of AT reduced uptake of neutrophils by AMs. Differential uptake of neutrophils was not due to changes in either the CD47/CD172 axis or CD36 levels. AT significantly reduced lung expression of CCN1 and altered AM surface localization of DD1α, two proteins known to influence efferocytosis. We conclude that AT may contribute to tissue damage during S. aureus pneumonia by inhibiting the ability of AM to clear neutrophils at the site of infection. PMID:27739519

  20. Protective effects of ethyl pyruvate on lipopolysaccharide-induced acute lung injury through inhibition of autophagy in neutrophils

    PubMed Central

    Zhu, Qingteng; Wang, Hui; Wang, Hairong; Luo, Yong; Yu, Yang; Du, Qirong; Fei, Aihua; Pan, Shuming

    2017-01-01

    Among a number of clinical factors, bacterial infection is one of the most common causes of acute lung injury (ALI), a serious complication that carries a high risk of mortality (~40%). During the process of ALI, intense local and systemic inflammation is elicited, which exacerbates the injury. Neutrophil infiltration into airspace is observed in early stage of ALI, and is required for the full development of ALI through an array of mechanisms, including the release of granule contents and the production of pro-inflammatory cytokines, due to the overactivation of complement and cytokines. The present study noted that ethyl pyruvate alleviated ALI in lipopolysaccharide (LPS)-induced ALI mice. Increased autophagy in neutrophils from ALI mice was observed, while ethyl pyruvate diminished autophagy in neutrophils and constrained granule release, and therefore myeloperoxidase (MPO) in bronchoalveolar lavage fluid and the production of proinflammatory cytokines. Using neutrophil cells, it was identified that autophagy was required for neutrophil activation and granule release, and that ethyl pyruvate caused neutrophil autophagy, leading to the restriction of granule release, and thus contributing to the mitigation of ALI. If autophagy was obviated through knockdown of key regulator of autophagy Atg5, the effects of ethyl pyruvate on granule release by neutrophils disappeared. Taken together, the results demonstrated that ethyl pyruvate alleviates ALI through inhibition of autophagy-induced granule release by neutrophils, and this mechanism suggested a novel potential therapeutic target in autophagy regulation for ALI. PMID:28098908

  1. Inhibition of Nicotinamide Phosphoribosyltransferase Reduces Neutrophil-Mediated Injury in Myocardial Infarction

    PubMed Central

    Bauer, Inga; Braunersreuther, Vincent; Bruzzone, Santina; Akhmedov, Alexander; Lüscher, Thomas F.; Speer, Timo; Poggi, Alessandro; Mannino, Elena; Pelli, Graziano; Galan, Katia; Bertolotto, Maria; Lenglet, Sébastien; Garuti, Anna; Montessuit, Christophe; Lerch, René; Pellieux, Corinne; Vuilleumier, Nicolas; Dallegri, Franco; Mage, Jacqueline; Sebastian, Carlos; Mostoslavsky, Raul; Gayet-Ageron, Angèle; Patrone, Franco; Mach, François; Nencioni, Alessio

    2013-01-01

    Abstract Aims: Nicotinamide phosphoribosyltransferase (Nampt) is a key enzyme for nicotinamide adenine dinucleotide (NAD+) biosynthesis, and recent evidence indicates its role in inflammatory processes. Here, we investigated the potential effects of pharmacological Nampt inhibition with FK866 in a mouse myocardial ischemia/reperfusion model. In vivo and ex vivo mouse myocardial ischemia/reperfusion procedures were performed. Results: Treatment with FK866 reduced myocardial infarct size, neutrophil infiltration, and reactive oxygen species (ROS) generation within infarcted hearts in vivo in a mouse model of ischemia and reperfusion. The benefit of FK866 was not shown in the Langendorff model (ex vivo model of working heart without circulating leukocytes), suggesting a direct involvement of these cells in cardiac injury. Sera from FK866-treated mice showed reduced circulating levels of the neutrophil chemoattractant CXCL2 and impaired capacity to prime migration of these cells in vitro. The release of CXCL8 (human homolog of murine chemokine CXCL2) by human peripheral blood mononuclear cells (PBMCs) and Jurkat cells was also reduced by FK866, as well as by sirtuin (SIRT) inhibitors and SIRT6 silencing, implying a pivotal role for this NAD+-dependent deacetylase in the production of this chemokine. Innovation: The pharmacological inhibition of Nampt might represent an effective approach to reduce neutrophilic inflammation- and oxidative stress-mediated tissue damage in early phases of reperfusion after a myocardial infarction. Conclusions: Nampt inhibition appears as a new strategy to dampen CXCL2-induced neutrophil recruitment and thereby reduce neutrophil-mediated tissue injury in mice. Antioxid. Redox Signal. 18, 630–641. PMID:22452634

  2. Vitamin C: A Novel Regulator of Neutrophil Extracellular Trap Formation

    PubMed Central

    Mohammed, Bassem M.; Fisher, Bernard J.; Kraskauskas, Donatas; Farkas, Daniela; Brophy, Donald F.; Fowler, Alpha A.; Natarajan, Ramesh

    2013-01-01

    Introduction: Neutrophil extracellular trap (NET) formation was recently identified as a novel mechanism to kill pathogens. However, excessive NET formation in sepsis can injure host tissues. We have recently shown that parenteral vitamin C (VitC) is protective in sepsis. Whether VitC alters NETosis is unknown. Methods: We used Gulo−/− mice as they lack the ability to synthesize VitC. Sepsis was induced by intraperitoneal infusion of a fecal stem solution (abdominal peritonitis, FIP). Some VitC deficient Gulo−/− mice received an infusion of ascorbic acid (AscA, 200 mg/kg) 30 min after induction of FIP. NETosis was assessed histologically and by quantification for circulating free DNA (cf-DNA) in serum. Autophagy, histone citrullination, endoplasmic reticulum (ER) stress, NFκB activation and apoptosis were investigated in peritoneal PMNs. Results: Sepsis produced significant NETs in the lungs of VitC deficient Gulo−/− mice and increased circulating cf-DNA. This was attenuated in the VitC sufficient Gulo−/− mice and in VitC deficient Gulo−/− mice infused with AscA. Polymorphonuclear neutrophils (PMNs) from VitC deficient Gulo−/− mice demonstrated increased activation of ER stress, autophagy, histone citrullination, and NFκB activation, while apoptosis was inhibited. VitC also significantly attenuated PMA induced NETosis in PMNs from healthy human volunteers. Conclusions: Our in vitro and in vivo findings identify VitC as a novel regulator of NET formation in sepsis. This study complements the notion that VitC is protective in sepsis settings. PMID:23939536

  3. CC chemokines induce neutrophils to chemotaxis, degranulation, and alpha-defensin release.

    PubMed

    Jan, Ming-Shiou; Huang, Yi-Hsien; Shieh, Biehuoy; Teng, Ru-Hsiu; Yan, Yao-Pei; Lee, Yuan-Ti; Liao, Ko-Kaung; Li, Ching

    2006-01-01

    We have previously shown that a Taiwanese cohort of HIV-uninfected individuals was associated with the significantly elevated levels of serum beta-chemokines, macrophage inflammatory protein (MIP-1)-alpha and MIP-beta, and RANTES. In the present study, we report that the members of this cohort have significantly greater numbers of lower buoyant-density neutrophils in their blood, which leads to further investigation of the effects of beta-chemokines on neutrophils. By electron and confocal microscopic techniques and FACScan, the results demonstrated that MIP-1alpha, MIP-beta, and/or RANTES readily activated the cells to release a large quantity of alpha-defensins in vitro through the degranulation process, which was the cause of low-buoyant-density neutrophil production. The purified neutrophils underwent chemotaxis and increased phagocytic capability when beta-chemokines were present. Only when using all 3 neutralizing antibodies for CCR1, CCR3, and CCR5 could the chemotaxis of neutrophils be inhibited completely, suggesting that these receptors are involved in transducing activating signals. Because neutrophils are the most abundant white blood cells that can be activated simultaneously to release alpha-defensins and because these proteins are antiviral, including anti-HIV, our results support the hypothesis that in addition to beta-chemokines, the innate immunity of the cohort plays a role in inhibiting the transmission of HIV.

  4. Capability of Neutrophils to Form NETs Is Not Directly Influenced by a CMA-Targeting Peptide

    PubMed Central

    Maueröder, Christian; Schall, Nicolas; Meyer, Frédéric; Mahajan, Aparna; Garnier, Benjamin; Hahn, Jonas; Kienhöfer, Deborah; Hoffmann, Markus H.; Muller, Sylviane

    2017-01-01

    During inflammatory reaction, neutrophils exhibit numerous cellular and immunological functions, notably the formation of neutrophil extracellular traps (NETs) and autophagy. NETs are composed of decondensed chromatin fibers coated with various antimicrobial molecules derived from neutrophil granules. NETs participate in antimicrobial defense and can also display detrimental roles and notably trigger some of the immune features of systemic lupus erythematosus (SLE) and other autoimmune diseases. Autophagy is a complex and finely regulated mechanism involved in the cell survival/death balance that may be connected to NET formation. To shed some light on the connection between autophagy and NET formation, we designed a number of experiments in human neutrophils and both in normal and lupus-prone MRL/lpr mice to determine whether the synthetic peptide P140, which is capable of selectively modulating chaperone-mediated autophagy (CMA) in lymphocytes, could alter NET formation. P140/Lupuzor™ is currently being evaluated in phase III clinical trials involving SLE patients. Overall our in vitro and in vivo studies established that P140 does not influence NET formation, cytokine/chemokine production, or CMA in neutrophils. Thus, the beneficial effect of P140/Lupuzor™ in SLE is apparently not directly related to modulation of neutrophil function. PMID:28191006

  5. 5-lipoxygenase-dependent recruitment of neutrophils and macrophages by eotaxin-stimulated murine eosinophils.

    PubMed

    Luz, Ricardo Alves; Xavier-Elsas, Pedro; de Luca, Bianca; Masid-de-Brito, Daniela; Cauduro, Priscila Soares; Arcanjo, Luiz Carlos Gondar; dos Santos, Ana Carolina Cordeiro Faria; de Oliveira, Ivi Cristina Maria; Gaspar-Elsas, Maria Ignez Capella

    2014-01-01

    The roles of eosinophils in antimicrobial defense remain incompletely understood. In ovalbumin-sensitized mice, eosinophils are selectively recruited to the peritoneal cavity by antigen, eotaxin, or leukotriene(LT)B4, a 5-lipoxygenase (5-LO) metabolite. 5-LO blockade prevents responses to both antigen and eotaxin. We examined responses to eotaxin in the absence of sensitization and their dependence on 5-LO. BALB/c or PAS mice and their mutants (5-LO-deficient ALOX; eosinophil-deficient GATA-1) were injected i.p. with eotaxin, eosinophils, or both, and leukocyte accumulation was quantified up to 24 h. Significant recruitment of eosinophils by eotaxin in BALB/c, up to 24 h, was accompanied by much larger numbers of recruited neutrophils and monocytes/macrophages. These effects were abolished by eotaxin neutralization and 5-LO-activating protein inhibitor MK886. In ALOX (but not PAS) mice, eotaxin recruitment was abolished for eosinophils and halved for neutrophils. In GATA-1 mutants, eotaxin recruited neither neutrophils nor macrophages. Transfer of eosinophils cultured from bone-marrow of BALB/c donors, or from ALOX donors, into GATA-1 mutant recipients, i.p., restored eotaxin recruitment of neutrophils and showed that the critical step dependent on 5-LO is the initial recruitment of eosinophils by eotaxin, not the secondary neutrophil accumulation. Eosinophil-dependent recruitment of neutrophils in naive BALB/c mice was associated with increased binding of bacteria.

  6. Pathogenic bacterial species associated with endodontic infection evade innate immune control by disabling neutrophils.

    PubMed

    Matsui, Aritsune; Jin, Jun-O; Johnston, Christopher D; Yamazaki, Hajime; Houri-Haddad, Yael; Rittling, Susan R

    2014-10-01

    Endodontic infections, in which oral bacteria access the tooth pulp chamber, are common and do not resolve once established. To investigate the effects of these infections on the innate immune response, we established a mouse subcutaneous chamber model, where a mixture of four oral pathogens commonly associated with these infections (endodontic pathogens [EP]), i.e., Fusobacterium nucleatum, Streptococcus intermedius, Parvimonas micra, and Prevotella intermedia, was inoculated into subcutaneously implanted titanium chambers. Cells that infiltrated the chamber after these infections were primarily neutrophils; however, these neutrophils were unable to control the infection. Infection with a nonpathogenic oral bacterial species, Streptococcus mitis, resulted in well-controlled infection, with bacterial numbers reduced by 4 to 5 log units after 7 days. Propidium iodide (PI) staining of the chamber neutrophils identified three distinct populations: neutrophils from EP-infected chambers were intermediate in PI staining, while cells in chambers from mice infected with S. mitis were PI positive (apoptotic) or negative (live). Strikingly, neutrophils from EP-infected chambers were severely impaired in their ability to phagocytose and to generate reactive oxygen species in vitro after removal from the chamber compared to cells from S. mitis-infected chambers. The mechanism of neutrophil impairment was necrotic cell death as determined by morphological analyses. P. intermedia alone could induce a similar neutrophil phenotype. We conclude that the endodontic pathogens, particularly P. intermedia, can efficiently disable and kill infiltrating neutrophils, allowing these infections to become established. These results can help explain the persistence of endodontic infections and demonstrate a new virulence mechanism associated with P. intermedia.

  7. Pathogenic Bacterial Species Associated with Endodontic Infection Evade Innate Immune Control by Disabling Neutrophils

    PubMed Central

    Matsui, Aritsune; Jin, Jun-O; Johnston, Christopher D.; Yamazaki, Hajime; Houri-Haddad, Yael

    2014-01-01

    Endodontic infections, in which oral bacteria access the tooth pulp chamber, are common and do not resolve once established. To investigate the effects of these infections on the innate immune response, we established a mouse subcutaneous chamber model, where a mixture of four oral pathogens commonly associated with these infections (endodontic pathogens [EP]), i.e., Fusobacterium nucleatum, Streptococcus intermedius, Parvimonas micra, and Prevotella intermedia, was inoculated into subcutaneously implanted titanium chambers. Cells that infiltrated the chamber after these infections were primarily neutrophils; however, these neutrophils were unable to control the infection. Infection with a nonpathogenic oral bacterial species, Streptococcus mitis, resulted in well-controlled infection, with bacterial numbers reduced by 4 to 5 log units after 7 days. Propidium iodide (PI) staining of the chamber neutrophils identified three distinct populations: neutrophils from EP-infected chambers were intermediate in PI staining, while cells in chambers from mice infected with S. mitis were PI positive (apoptotic) or negative (live). Strikingly, neutrophils from EP-infected chambers were severely impaired in their ability to phagocytose and to generate reactive oxygen species in vitro after removal from the chamber compared to cells from S. mitis-infected chambers. The mechanism of neutrophil impairment was necrotic cell death as determined by morphological analyses. P. intermedia alone could induce a similar neutrophil phenotype. We conclude that the endodontic pathogens, particularly P. intermedia, can efficiently disable and kill infiltrating neutrophils, allowing these infections to become established. These results can help explain the persistence of endodontic infections and demonstrate a new virulence mechanism associated with P. intermedia. PMID:25024367

  8. NETosis in Cancer – Platelet–Neutrophil Crosstalk Promotes Tumor-Associated Pathology

    PubMed Central

    Olsson, Anna-Karin; Cedervall, Jessica

    2016-01-01

    It has become increasingly clear that circulating immune cells in the body have a major impact on cancer development, progression, and outcome. The role of both platelets and neutrophils as independent regulators of various processes in cancer has been known for long, but it has quite recently emerged that the platelet–neutrophil interplay is yet a critical component to take into account during malignant disease. It was reported a few years ago that neutrophils in mice with cancer have increased propensity to form neutrophil extracellular traps (NETs) – web-like structures formed by externalized chromatin and secreted proteases. The initial finding describing this as a cell death-associated process has been followed by reports of additional mechanisms for NET formation (NETosis), and it has been shown that similar structures can be formed also without lysis and neutrophil cell death as a consequence. Furthermore, presence of NETs in humans with cancer has been verified in a few recent studies, indicating that tumor-induced NETosis is clinically relevant. Several reports have also described that NETs contribute to cancer-associated pathology, by promoting processes responsible for cancer-related death such as thrombosis, systemic inflammation, and relapse of the disease. This review summarizes current knowledge about NETosis in cancer, including the role of platelets as regulators of tumor-induced NETosis. It has been shown that platelets can serve as inducers of NETosis, and the platelet–neutrophil interface can therefore be an important issue to consider when designing therapies targeting cancer-associated pathology in the future. PMID:27708646

  9. Moesin regulates neutrophil rolling velocity in vivo.

    PubMed

    Matsumoto, Masanori; Hirata, Takako

    2016-01-01

    During inflammation, the selectin-induced slow rolling of neutrophils on venules cooperates with chemokine signaling to mediate neutrophil recruitment into tissues. Previous studies identified P-selectin glycoprotein ligand-1 (PSGL-1) and CD44 as E-selectin ligands that activate integrins to induce slow rolling. We show here that in TNF-α-treated cremaster muscle venules, slow leukocyte rolling was impaired in mice deficient in moesin, a member of the ezrin-radixin-moesin (ERM) family. Accordingly, neutrophil recruitment in a peritonitis model was decreased in moesin-deficient mice when chemokine signaling was blocked with pertussis toxin. These results suggest that moesin contributes to the slow rolling and subsequent recruitment of neutrophils during inflammation.

  10. Clinical and Laboratory Differences between Lymphocyte- and Neutrophil-Predominant Pleural Tuberculosis

    PubMed Central

    Kim, Kang; Kim, Sukyeon; Oh, Ki-Jong; Jeong, Suk Hyeon; Jung, Woo Jin; Shin, Beomsu; Jhun, Byung Woo; Lee, Hyun; Park, Hye Yun; Koh, Won-Jung

    2016-01-01

    Pleural tuberculosis (TB), a form of extrapulmonary TB, can be difficult to diagnose. High numbers of lymphocytes in pleural fluid have been considered part of the diagnostic criteria for pleural TB; however, in many cases, neutrophils rather than lymphocytes are the predominant cell type in pleural effusions, making diagnosis more complicated. Additionally, there is limited information on the clinical and laboratory characteristics of neutrophil-predominant pleural effusions caused by Mycobacterium tuberculosis (MTB). To investigate clinical and laboratory differences between lymphocyte- and neutrophil-predominant pleural TB, we retrospectively analyzed 200 patients with the two types of pleural TB. Of these patients, 9.5% had neutrophil-predominant pleural TB. Patients with lymphocyte-predominant and neutrophil-predominant pleural TB showed similar clinical signs and symptoms. However, neutrophil-predominant pleural TB was associated with significantly higher inflammatory serum markers, such as white blood cell count (P = 0.001) and C-reactive protein (P = 0.001). Moreover, MTB was more frequently detected in the pleural fluid from patients in the neutrophil-predominant group than the lymphocyte-predominant group, with the former group exhibiting significantly higher rates of positive results for acid-fast bacilli in sputum (36.8 versus 9.4%, P = 0.003), diagnostic yield of MTB culture (78.9% versus 22.7%, P < 0.001) and MTB detected by polymerase chain reaction (31.6% versus 5.0%, P = 0.001). Four of seven patients with repeated pleural fluid analyses revealed persistent neutrophil-predominant features, which does not support the traditional viewpoint that neutrophil-predominant pleural TB is a temporary form that rapidly develops into lymphocyte-predominant pleural TB. In conclusion, neutrophil-predominant pleural TB showed a more intense inflammatory response and a higher positive rate in microbiological testing compared to lymphocyte-predominant pleural TB

  11. Comprehensive multiplexed protein quantitation delineates eosinophilic and neutrophilic experimental asthma

    PubMed Central

    2014-01-01

    Background Improvements in asthma diagnosis and management require deeper understanding of the heterogeneity of the complex airway inflammation. We hypothesise that differences in the two major inflammatory phenotypes of asthma; eosinophilic and neutrophilic asthma, will be reflected in the lung protein expression profile of murine asthma models and can be delineated using proteomics of bronchoalveolar lavage (BAL). Methods BAL from mice challenged with ovalbumin (OVA/OVA) alone (standard model of asthma, here considered eosinophilic) or OVA in combination with endotoxin (OVA/LPS, model of neutrophilic asthma) was analysed using liquid chromatography coupled to high resolution mass spectrometry, and compared with steroid-treated animals and healthy controls. In addition, conventional inflammatory markers were analysed using multiplexed ELISA (Bio-Plex™ assay). Multivariate statistics was performed on integrative proteomic fingerprints using principal component analysis. Proteomic data were complemented with lung mechanics and BAL cell counts. Results Several of the analysed proteins displayed significant differences between the controls and either or both of the two models reflecting eosinophilic and neutrophilic asthma. Most of the proteins found with mass spectrometry analysis displayed a considerable increase in neutrophilic asthma compared with the other groups. Conversely, the larger number of the inflammatory markers analysed with Bio-Plex™ analysis were found to be increased in the eosinophilic model. In addition, major inflammation markers were correlated to peripheral airway closure, while commonly used asthma biomarkers only reflect central inflammation. Conclusion Our data suggest that the commercial markers we are currently relying on to diagnose asthma subtypes are not giving us comprehensive or specific enough information. The analysed protein profiles allowed to discriminate the two models and may add useful information for characterization of

  12. Neutrophilic inflammatory response and oxidative stress in premenopausal women chronically exposed to indoor air pollution from biomass burning.

    PubMed

    Banerjee, Anirban; Mondal, Nandan Kumar; Das, Debangshu; Ray, Manas Ranjan

    2012-04-01

    The possibility of inflammation and neutrophil activation in response to indoor air pollution (IAP) from biomass fuel use has been investigated. For this, 142 premenopausal, never-smoking women (median age, 34 years) who cook exclusively with biomass (wood, dung, crop wastes) and 126 age-matched control women who cook with cleaner fuel liquefied petroleum gas (LPG) were enrolled. The neutrophil count in blood and sputum was significantly higher (p < 0.05) in biomass users than the control group. Flow cytometric analysis revealed marked increase in the surface expression of CD35 (complement receptor-1), CD16 (F(C)γ receptor III), and β(2) Mac-1 integrin (CD11b/CD18) on circulating neutrophils of biomass users. Besides, enzyme-linked immunosorbent assay showed that they had 72%, 67%, and 54% higher plasma levels of the proinflammatory cytokines tumor necrosis factor-alpha, interleukin-6, and interleukin-12, respectively, and doubled neutrophil chemoattractant interleukin-8. Immunocytochemical study revealed significantly higher percentage of airway neutrophils expressing inducible nitric oxide synthase, while the serum level of nitric oxide was doubled in women who cooked with biomass. Spectrophotometric analysis documented higher myeloperoxidase activity in circulating neutrophils of biomass users, suggesting neutrophil activation. Flow cytometry showed excess generation of reactive oxygen species (ROS) by leukocytes of biomass-using women, whereas their erythrocytes contained a depleted level of antioxidant enzyme superoxide dismutase (SOD). Indoor air of biomass-using households had two to four times more particulate matter with diameters of <10 μm (PM(10)) and <2.5 μm (PM(2.5)) as measured by real-time laser photometer. After controlling potential confounders, rise in proinflammatory mediators among biomass users were positively associated with PM(10) and PM(2.5) in indoor air, suggesting a close relationship between IAP and neutrophil activation. Besides

  13. Neutrophil function in pregnancy and rheumatoid arthritis

    PubMed Central

    Crocker, I; Baker, P; Fletcher, J

    2000-01-01

    BACKGROUND—Pregnancy exerts suppressive effects on rheumatoid arthritis (RA). An attenuation in neutrophil function in late pregnancy which may explain this amelioration has previously been reported.
OBJECTIVE—A longitudinal investigation of neutrophil activity in healthy pregnant women (n=9) and pregnant patients with RA (n=9), compared with age matched non-pregnant patients with RA (n=12) and healthy controls (n=22).
METHODS—Neutrophil activation was measured in response to the physiological receptor agonists, n-formyl-methionyl-leucyl-phenylalanine (fMLP) and zymosan activated serum (ZAS). Superoxide anion production (respiratory burst) was determined by lucigenin enhanced chemiluminescence (LUCL); secondary granule lactoferrin release by enzyme linked immunosorbent assay (ELISA); and CD11b, CD18, and CD62L expression by flow cytometric analysis.
RESULTS—Stimulated neutrophil LUCL was significantly reduced in both pregnant women with RA and healthy pregnant women in the second (fMLP 43% and 69%, ZAS 43% and 59%, respectively) and third trimesters (fMLP 24% and 44%, ZAS 32% and 38%, respectively). Responses returned to normal within eight weeks of delivery and unstimulated levels remained unchanged throughout pregnancy. Basal and stimulated CD11b, CD18, and CD62L expression showed no variations throughout gestation for both pregnancy groups. Likewise, stimulated lactoferrin release and plasma lactoferrin remained unchanged. Certain morphological differences in RA neutrophils were highlighted by the flow cytometric analysis. Moreover, resting neutrophils and stimulated cells from patients with RA, including pregnant subjects, showed a marked increase in LUCL, but a reduction in CD11b, CD18, and CD62L. Low dose prednisolone and methylprednisolone had no effect on neutrophil parameters over the period of treatment with non-steroidal anti-inflammatory drugs.
CONCLUSION—The attenuation to neutrophil respiratory burst in both healthy and RA

  14. Photothermal image cytometry of human neutrophils

    NASA Astrophysics Data System (ADS)

    Lapotko, Dmitry

    2001-07-01

    Photothermal imaging, when being applied to the study of living cells, provides morpho-functional information about the cell populations. In technical terms, the method is complementary to optical microscopy. The photothermal method was used for cell imaging and quantitative studies. Preliminary results of the studies on living human neutrophils are presented. Differences between normal and pathological neutrophil populations from blood of healthy donors and patients with saracoidosis and pleuritis are demonstrated.

  15. PEGylated single-walled carbon nanotubes activate neutrophils to increase production of hypochlorous acid, the oxidant capable of degrading nanotubes.

    PubMed

    Vlasova, Irina I; Vakhrusheva, Tatyana V; Sokolov, Alexey V; Kostevich, Valeria A; Gusev, Alexandr A; Gusev, Sergey A; Melnikova, Viktoriya I; Lobach, Anatolii S

    2012-10-01

    Perspectives for the use of carbon nanotubes in biomedical applications depend largely on their ability to degrade in the body into products that can be easily cleared out. Carboxylated single-walled carbon nanotubes (c-SWCNTs) were shown to be degraded by oxidants generated by peroxidases in the presence of hydrogen peroxide. In the present study we demonstrated that conjugation of poly(ethylene glycol) (PEG) to c-SWCNTs does not interfere with their degradation by peroxidase/H(2)O(2) system or by hypochlorite. Comparison of different heme-containing proteins for their ability to degrade PEG-SWCNTs has led us to conclude that the myeloperoxidase (MPO) product hypochlorous acid (HOCl) is the major oxidant that may be responsible for biodegradation of PEG-SWCNTs in vivo. MPO is secreted mainly by neutrophils upon activation. We hypothesize that SWCNTs may enhance neutrophil activation and therefore stimulate their own biodegradation due to MPO-generated HOCl. PEG-SWCNTs at concentrations similar to those commonly used in in vivo studies were found to activate isolated human neutrophils to produce HOCl. Both PEG-SWCNTs and c-SWCNTs enhanced HOCl generation from isolated neutrophils upon serum-opsonized zymosan stimulation. Both types of nanotubes were also found to activate neutrophils in whole blood samples. Intraperitoneal injection of a low dose of PEG-SWCNTs into mice induced an increase in percentage of circulating neutrophils and activation of neutrophils and macrophages in the peritoneal cavity, suggesting the evolution of an inflammatory response. Activated neutrophils can produce high local concentrations of HOCl, thereby creating the conditions favorable for degradation of the nanotubes.

  16. Dictyostelium amoebae and neutrophils can swim.

    PubMed

    Barry, Nicholas P; Bretscher, Mark S

    2010-06-22

    Animal cells migrating over a substratum crawl in amoeboid fashion; how the force against the substratum is achieved remains uncertain. We find that amoebae and neutrophils, cells traditionally used to study cell migration on a solid surface, move toward a chemotactic source while suspended in solution. They can swim and do so with speeds similar to those on a solid substrate. Based on the surprisingly rapidly changing shape of amoebae as they swim and earlier theoretical schemes for how suspended microorganisms can migrate (Purcell EM (1977) Life at low Reynolds number. Am J Phys 45:3-11), we suggest the general features these cells use to gain traction with the medium. This motion requires either the movement of the cell's surface from the cell's front toward its rear or protrusions that move down the length of the elongated cell. Our results indicate that a solid substratum is not a prerequisite for these cells to produce a forward thrust during movement and suggest that crawling and swimming are similar processes, a comparison we think is helpful in understanding how cells migrate.

  17. Isolation of healthy individuals' and rheumatoid arthritis patients' peripheral blood neutrophils by the gelatin and Ficoll-Hypaque methods: comparative efficiency and impact on the neutrophil oxidative metabolism and Fcγ receptor expression.

    PubMed

    Paoliello-Paschoalato, A B; Azzolini, A E C S; Cruz, M F C; Marchi, L F; Kabeya, L M; Donadi, E A; Lucisano-Valim, Y M

    2014-10-01

    In vitro assessment of the functional responses of leukocytes sometimes requires their isolation from blood, joint and tissues. In this study, we compared the efficiency of two procedures - the gelatin method and Ficoll-Hypaque density centrifugation gradient - to isolate peripheral blood neutrophils of healthy individuals and patients with active rheumatoid arthritis (RA). We also assessed whether these procedures affect the neutrophil activation status. Both purification procedures were concluded in 90min, and yielded cell populations with similar degrees of purity (80-90%), number of neutrophils (1-2×10(6) cells per mL of blood), and viability (97-100%). In vitro neutrophil priming with granulocyte-macrophage colony-stimulating factor (GM-CSF) significantly increased the reactive oxygen species producing ability of the cells stimulated with n-formyl-methionyl-leucyl-phenylalanine (n-fMLP), soluble immune complexes (s-ICs), and insoluble immune complexes (i-ICs). Isolated neutrophils not treated with GM-CSF responded to n-fMLP and i-IC, but not to s-IC. Almost all of the neutrophils (98-100%) purified by both methods expressed FcγRII/CD32 and FcγRIII/CD16, but they did not express significant levels of FcγRI/CD64. Similar results were obtained for healthy individuals' and RA patients' neutrophils. In summary, the gelatin method was comparable to Ficoll-Hypaque gradient in terms of purity, yield, and viability of the neutrophil preparations. Both methods neither primed or activated the neutrophils, nor affected their functional responsiveness. Therefore, both methods are suitable to isolate peripheral blood neutrophils of healthy individuals and RA patients.

  18. Stability analysis of micropipette aspiration of neutrophils.

    PubMed Central

    Derganc, J; Bozic, B; Svetina, S; Zeks, B

    2000-01-01

    During micropipette aspiration, neutrophil leukocytes exhibit a liquid-drop behavior, i.e., if a neutrophil is aspirated by a pressure larger than a certain threshold pressure, it flows continuously into the pipette. The point of the largest aspiration pressure at which the neutrophil can still be held in a stable equilibrium is called the critical point of aspiration. Here, we present a theoretical analysis of the equilibrium behavior and stability of a neutrophil during micropipette aspiration with the aim to rigorously characterize the critical point. We take the energy minimization approach, in which the critical point is well defined as the point of the stability breakdown. We use the basic liquid-drop model of neutrophil rheology extended by considering also the neutrophil elastic area expansivity. Our analysis predicts that the behavior at large pipette radii or small elastic area expansivity is close to the one predicted by the basic liquid-drop model, where the critical point is attained slightly before the projection length reaches the pipette radius. The effect of elastic area expansivity is qualitatively different at smaller pipette radii, where our analysis predicts that the critical point is attained at the projection lengths that may significantly exceed the pipette radius. PMID:10866944

  19. Circulating Progenitor Cells and Scleroderma

    PubMed Central

    2010-01-01

    Scleroderma (systemic sclerosis) is a disease of unknown origins that involves tissue ischemia and fibrosis in the skin and internal organs such as the lungs. The tissue ischemia is due to a lack of functional blood vessels and an inability to form new blood vessels. Bone marrow–derived circulating endothelial progenitor cells play a key role in blood vessel repair and neovascularization. Scleroderma patients appear to have defects in the number and function of circulating endothelial progenitor cells. Scleroderma patients also develop fibrotic lesions, possibly as the result of tissue ischemia. Fibroblast-like cells called fibrocytes that differentiate from a different pool of bone marrow–derived circulating progenitor cells seem to be involved in this process. Manipulating the production, function, and differentiation of circulating progenitor cells represents an exciting new possibility for treating scleroderma. PMID:18638425

  20. Early enhanced local neutrophil recruitment in peritonitis-induced sepsis improves bacterial clearance and survival.

    PubMed

    Craciun, Florin L; Schuller, Elizabeth R; Remick, Daniel G

    2010-12-01

    Neutrophils are critical for the rapid eradication of bacterial pathogens, but they also contribute to the development of multiple organ failure in sepsis. We hypothesized that increasing early recruitment of neutrophils to the focus of infection will increase bacterial clearance and improve survival. Sepsis was induced in mice, using cecal ligation and puncture (CLP); blood samples were collected at 6 and 24 h; and survival was followed for 28 d. In separate experiments, peritoneal bacteria and inflammatory cells were measured. Septic mice predicted to die based on IL-6 levels (Die-P) had higher concentrations of CXCL1 and CXCL2 in the peritoneum and plasma compared with those predicted to live (Live-P). At 6 h, Live-P and Die-P had equivalent numbers of peritoneal neutrophils and bacteria. In Die-P mice the number of peritoneal bacteria increased between 6 and 24 h post-CLP, whereas in Live-P it decreased. The i.p. injection of CXCL1 and CXCL2 in naive mice resulted in local neutrophil recruitment. When given immediately after CLP, CXC chemokines increased peritoneal neutrophil recruitment at 6 h after CLP. This early increase in neutrophils induced by exogenous chemokines resulted in significantly fewer peritoneal bacteria by 24 h [CFU (log) = 6.04 versus 4.99 for vehicle versus chemokine treatment; p < 0.05]. Chemokine treatment significantly improved survival at both 5 d (40 versus 72%) and 28 d (27 versus 52%; p < 0.02 vehicle versus chemokines). These data demonstrate that early, local treatment with CXC chemokines enhances neutrophil recruitment and clearance of bacteria as well as improves survival in the CLP model of sepsis.

  1. Inhibition of Ras signalling reduces neutrophil infiltration and tissue damage in severe acute pancreatitis.

    PubMed

    Yu, Changhui; Merza, Mohammed; Luo, Lingtao; Thorlacius, Henrik

    2015-01-05

    Neutrophil recruitment is known to be a rate-limiting step in mediating tissue injury in severe acute pancreatitis (AP). However, the signalling mechanisms controlling inflammation and organ damage in AP remain elusive. Herein, we examined the role of Ras signalling in AP. Male C57BL/6 mice were treated with a Ras inhibitor (farnesylthiosalicylic acid, FTS) before infusion of taurocholate into the pancreatic duct. Pancreatic and lung tissues as well as blood were collected 24 h after pancreatitis induction. Pretreatment with FTS decreased serum amylase levels by 82% and significantly attenuated acinar cell necrosis, tissue haemorrhage and oedema formation in taurocholate-induced pancreatitis. Inhibition of Ras signalling reduced myeloperoxidase (MPO) levels in the inflamed pancreas by 42%. In addition, administration of FTS decreased pancreatic levels of CXC chemokines as well as circulating levels of interleukin-6 and high-mobility group box 1 in animals exposed to taurocholate. Moreover, treatment with FTS reduced taurocholate-induced MPO levels in the lung. Inhibition of Ras signalling had no effect on neutrophil expression of Mac-1 in mice with pancreatitis. Moreover, FTS had no direct impact on trypsin activation in isolated pancreatic acinar cells. These results indicate that Ras signalling controls CXC chemokine formation, neutrophil recruitment and tissue injury in severe AP. Thus, our findings highlight a new signalling mechanism regulating neutrophil recruitment in the pancreas and suggest that inhibition of Ras signalling might be a useful strategy to attenuate local and systemic inflammation in severe AP.

  2. PEGylated single-walled carbon nanotubes activate neutrophils to increase production of hypochlorous acid, the oxidant capable of degrading nanotubes

    SciTech Connect

    Vlasova, Irina I.; Vakhrusheva, Tatyana V.; Sokolov, Alexey V.; Kostevich, Valeria A.; Gusev, Alexandr A.; Gusev, Sergey A.; Melnikova, Viktoriya I.; Lobach, Anatolii S.

    2012-10-01

    Perspectives for the use of carbon nanotubes in biomedical applications depend largely on their ability to degrade in the body into products that can be easily cleared out. Carboxylated single-walled carbon nanotubes (c-SWCNTs) were shown to be degraded by oxidants generated by peroxidases in the presence of hydrogen peroxide. In the present study we demonstrated that conjugation of poly(ethylene glycol) (PEG) to c-SWCNTs does not interfere with their degradation by peroxidase/H{sub 2}O{sub 2} system or by hypochlorite. Comparison of different heme-containing proteins for their ability to degrade PEG-SWCNTs has led us to conclude that the myeloperoxidase (MPO) product hypochlorous acid (HOCl) is the major oxidant that may be responsible for biodegradation of PEG-SWCNTs in vivo. MPO is secreted mainly by neutrophils upon activation. We hypothesize that SWCNTs may enhance neutrophil activation and therefore stimulate their own biodegradation due to MPO-generated HOCl. PEG-SWCNTs at concentrations similar to those commonly used in in vivo studies were found to activate isolated human neutrophils to produce HOCl. Both PEG-SWCNTs and c-SWCNTs enhanced HOCl generation from isolated neutrophils upon serum-opsonized zymosan stimulation. Both types of nanotubes were also found to activate neutrophils in whole blood samples. Intraperitoneal injection of a low dose of PEG-SWCNTs into mice induced an increase in percentage of circulating neutrophils and activation of neutrophils and macrophages in the peritoneal cavity, suggesting the evolution of an inflammatory response. Activated neutrophils can produce high local concentrations of HOCl, thereby creating the conditions favorable for degradation of the nanotubes. -- Highlights: ► Myeloperoxidase (MPO) product hypochlorous acid is able to degrade CNTs. ► PEGylated SWCNTs stimulate isolated neutrophils to produce hypochlorous acid. ► SWCNTs are capable of activating neutrophils in blood samples. ► Activation of

  3. Gene Expression during the Generation and Activation of Mouse Neutrophils: Implication of Novel Functional and Regulatory Pathways

    PubMed Central

    Ericson, Jeffrey A.; Duffau, Pierre; Yasuda, Kei; Ortiz-Lopez, Adriana; Rothamel, Katherine; Rifkin, Ian R.; Monach, Paul A.

    2014-01-01

    As part of the Immunological Genome Project (ImmGen), gene expression was determined in unstimulated (circulating) mouse neutrophils and three populations of neutrophils activated in vivo, with comparison among these populations and to other leukocytes. Activation conditions included serum-transfer arthritis (mediated by immune complexes), thioglycollate-induced peritonitis, and uric acid-induced peritonitis. Neutrophils expressed fewer genes than any other leukocyte population studied in ImmGen, and down-regulation of genes related to translation was particularly striking. However, genes with expression relatively specific to neutrophils were also identified, particularly three genes of unknown function: Stfa2l1, Mrgpr2a and Mrgpr2b. Comparison of genes up-regulated in activated neutrophils led to several novel findings: increased expression of genes related to synthesis and use of glutathione and of genes related to uptake and metabolism of modified lipoproteins, particularly in neutrophils elicited by thioglycollate; increased expression of genes for transcription factors in the Nr4a family, only in neutrophils elicited by serum-transfer arthritis; and increased expression of genes important in synthesis of prostaglandins and response to leukotrienes, particularly in neutrophils elicited by uric acid. Up-regulation of genes related to apoptosis, response to microbial products, NFkB family members and their regulators, and MHC class II expression was also seen, in agreement with previous studies. A regulatory model developed from the ImmGen data was used to infer regulatory genes involved in the changes in gene expression during neutrophil activation. Among 64, mostly novel, regulatory genes predicted to influence these changes in gene expression, Irf5 was shown to be important for optimal secretion of IL-10, IP-10, MIP-1α, MIP-1β, and TNF-α by mouse neutrophils in vitro after stimulation through TLR9. This data-set and its analysis using the ImmGen regulatory

  4. Recombinant gamma interferon causes neutrophil migration mediated by the release of a macrophage neutrophil chemotactic factor.

    PubMed Central

    Ribeiro, R. A.; Cunha, F. Q.; Ferreira, S. H.

    1990-01-01

    A dose-dependent neutrophil migration was observed following the injection of purified (Hu IFN-gamma) or recombinant (rIFN-gamma) human gamma interferon into rat peritoneal cavities. This finding contrasts with their inability to cause chemotaxis in vitro in the Boyden chamber. Neutrophil migration into peritoneal cavities and subcutaneous air pouches induced by both preparations of interferon was abolished by pretreatment of the animals with dexamethasone. IFN-gamma-induced neutrophil migration was enhanced when the macrophage population of the peritoneal cavities was increased by previous injection of thioglycollate and reduced by peritoneal lavage. Macrophage monolayers pretreated either with rIFN-gamma or with lipopolysaccharide from E. coli release into the supernatant a factor that stimulates neutrophil recruitment in animals treated with dexamethasone. Dexamethasone blocked this release but did not affect the neutrophil recruitment induced by this factor. These results suggest that IFN-gamma-induced neutrophil migration in vivo may be mediated by the release from resident macrophages of a neutrophil chemotactic factor and that dexamethasone blockade of neutrophil recruitment by IFN-gamma is due to inhibition of the release of this factor. PMID:2119790

  5. Neutrophils in Cancer: Two Sides of the Same Coin.

    PubMed

    Uribe-Querol, Eileen; Rosales, Carlos

    2015-01-01

    Neutrophils are the most abundant leukocytes in blood and are considered to be the first line of defense during inflammation and infections. In addition, neutrophils are also found infiltrating many types of tumors. Tumor-associated neutrophils (TANs) have relevant roles in malignant disease. Indeed neutrophils may be potent antitumor effector cells. However, increasing clinical evidence shows TANs correlate with poor prognosis. The tumor microenvironment controls neutrophil recruitment and in turn TANs help tumor progression. Hence, TANs can be beneficial or detrimental to the host. It is the purpose of this review to highlight these two sides of the neutrophil coin in cancer and to describe recent studies that provide some light on the mechanisms for neutrophil recruitment to the tumor, for neutrophils supporting tumor progression, and for neutrophil activation to enhance their antitumor functions.

  6. Superoxide anion production by human neutrophils activated by Trichomonas vaginalis.

    PubMed

    Song, Hyun-Ouk; Ryu, Jae-Sook

    2013-08-01

    Neutrophils are the predominant inflammatory cells found in vaginal discharges of patients infected with Trichomonas vaginalis. In this study, we examined superoxide anion (O2 (.-)) production by neutrophils activated by T. vaginalis. Human neutrophils produced superoxide anions when stimulated with either a lysate of T. vaginalis, its membrane component (MC), or excretory-secretory product (ESP). To assess the role of trichomonad protease in production of superoxide anions by neutrophils, T. vaginalis lysate, ESP, and MC were each pretreated with a protease inhibitor cocktail before incubation with neutrophils. Superoxide anion production was significantly decreased by this treatment. Trichomonad growth was inhibited by preincubation with supernatants of neutrophils incubated for 3 hr with T. vaginalis lysate. Furthermore, myeloperoxidase (MPO) production by neutrophils was stimulated by live trichomonads. These results indicate that the production of superoxide anions and MPO by neutrophils stimulated with T. vaginalis may be a part of defense mechanisms of neutrophils in trichomoniasis.

  7. Neutrophils in Cancer: Two Sides of the Same Coin

    PubMed Central

    Uribe-Querol, Eileen; Rosales, Carlos

    2015-01-01

    Neutrophils are the most abundant leukocytes in blood and are considered to be the first line of defense during inflammation and infections. In addition, neutrophils are also found infiltrating many types of tumors. Tumor-associated neutrophils (TANs) have relevant roles in malignant disease. Indeed neutrophils may be potent antitumor effector cells. However, increasing clinical evidence shows TANs correlate with poor prognosis. The tumor microenvironment controls neutrophil recruitment and in turn TANs help tumor progression. Hence, TANs can be beneficial or detrimental to the host. It is the purpose of this review to highlight these two sides of the neutrophil coin in cancer and to describe recent studies that provide some light on the mechanisms for neutrophil recruitment to the tumor, for neutrophils supporting tumor progression, and for neutrophil activation to enhance their antitumor functions. PMID:26819959

  8. [Neutrophils expression of adhesion molecules in diabetic nephropaty patients].

    PubMed

    Shcherban', T D

    2013-01-01

    CD11b and CD54 expression on neutrophils in patients with diabetic nephropathy (DN), arterial hypertension patients and healthy donors were examined. Development of DN associates with an increase of the number of CD11b and CD54 positive cells and violation of cellular co-operation. In the conditions of diabetic microenvironment expression of adhesion molecules rises substantially, what may characterized the mechanism of connection between hyperglycemia and vascular and tissues injury at DN. Authentication of morphological and biochemical markers of intercellular co-operation must in a prospect assist the deeper understanding of pathogenic mechanisms of DN.

  9. IL-4 induces neutrophilic maturation of HL-60 cells and activation of human peripheral blood neutrophils.

    PubMed Central

    Bober, L A; Waters, T A; Pugliese-Sivo, C C; Sullivan, L M; Narula, S K; Grace, M J

    1995-01-01

    IL-4 is a T-helper cell derived cytokine that has effects on myelomonocytic cell maturation and activation. We have studied the effect of IL-4 on neutrophilic maturation using the cell line HL-60 and found that it has a profound effect on the maturation and activation of the cell line. The treatment of HL-60 cells with recombinant hu IL-4 (0.15 to 15.0 ng/ml) induced a shift in the percentage of HL-60 cells staining positive for chloroacetate esterase enzyme activity (indicating commitment to the neutrophilic lineage). IL-4 increased surface expression of the neutrophil-lineage antigen WEM G11, the complement receptors CR3 (CD11b) and CR1 (CD35), but not for the monocyte differentiation antigen CD14. IL-4 treated HL-60 cells demonstrated enhanced Fc- and complement-mediated phagocytic capacity and increased hexose-monophosphate shunt activity. In addition, IL-4 was capable of sustaining the neutrophil maturation of HL-60 cells that had been pre-treated for 24 h with DMSO. To investigate the effect of IL-4 on the mature neutrophil, we studied freshly isolated and rested human peripheral blood neutrophils. In the absence of other stimuli, neutrophils were induced by IL-4 to have significantly elevated phagocytic responses. The response was specific since treatment with anti-human IL-4 abolished phagocytic stimulation. Finally, IL-4 treatment also stimulated resting neutrophils to migrate toward zymosan-activated serum (ZAS) and human IL-5. The results demonstrate that IL-4 is a potent maturation factor for myelocytes to become neutrophils and that IL-4 can stimulate resting mature neutrophils. PMID:7529148

  10. Stimulation of neutrophils by tumor necrosis factor

    SciTech Connect

    Klebanoff, S.J.; Vadas, M.A.; Harlan, J.M.; Sparks, L.H.; Gamble, J.R.; Agosti, J.M.; Waltersdorph, A.M.

    1986-06-01

    Human recombinant tumor necrosis factor (TNF) was shown to be a weak direct stimulus of the neutrophil respiratory burst and degranulation. The stimulation, as measured by iodination, H/sub 2/O/sub 2/ production, and lysozyme release, was considerably increased by the presence of unopsonized zymosan in the reaction mixture, an effect which was associated with the increased ingestion of the zymosan. TNF does not act as an opsonin but, rather, reacts with the neutrophil to increase its phagocytic activity. TNF-dependent phagocytosis, as measured indirectly by iodination, is inhibited by monoclonal antibodies (Mab) 60.1 and 60.3, which recognize different epitopes on the C3bi receptor/adherence-promoting surface glycoprotein of neutrophils. Other neutrophil stimulants, namely N-formyl-methionyl-leucyl-phenylalanine, the Ca2+ ionophore A23187, and phorbol myristic acetate, also increase iodination in the presence of zymosan; as with TNF, the effect of these stimulants is inhibited by Mab 60.1 and 60.3, whereas, in contrast to that of TNF, their stimulation of iodination is unaffected by an Mab directed against TNF. TNF may be a natural stimulant of neutrophils which promotes adherence to endothelial cells and to particles, leading to increased phagocytosis, respiratory burst activity, and degranulation.

  11. Blocking neutrophil diapedesis prevents hemorrhage during thrombocytopenia.

    PubMed

    Hillgruber, Carina; Pöppelmann, Birgit; Weishaupt, Carsten; Steingräber, Annika Kathrin; Wessel, Florian; Berdel, Wolfgang E; Gessner, J Engelbert; Ho-Tin-Noé, Benoît; Vestweber, Dietmar; Goerge, Tobias

    2015-07-27

    Spontaneous organ hemorrhage is the major complication in thrombocytopenia with a potential fatal outcome. However, the exact mechanisms regulating vascular integrity are still unknown. Here, we demonstrate that neutrophils recruited to inflammatory sites are the cellular culprits inducing thrombocytopenic tissue hemorrhage. Exposure of thrombocytopenic mice to UVB light provokes cutaneous petechial bleeding. This phenomenon is also observed in immune-thrombocytopenic patients when tested for UVB tolerance. Mechanistically, we show, analyzing several inflammatory models, that it is neutrophil diapedesis through the endothelial barrier that is responsible for the bleeding defect. First, bleeding is triggered by neutrophil-mediated mechanisms, which act downstream of capturing, adhesion, and crawling on the blood vessel wall and require Gαi signaling in neutrophils. Second, mutating Y731 in the cytoplasmic tail of VE-cadherin, known to selectively affect leukocyte diapedesis, but not the induction of vascular permeability, attenuates bleeding. Third, and in line with this, simply destabilizing endothelial junctions by histamine did not trigger bleeding. We conclude that specifically targeting neutrophil diapedesis through the endothelial barrier may represent a new therapeutic avenue to prevent fatal bleeding in immune-thrombocytopenic patients.

  12. Neutrophil Leukocyte: Combustive Microbicidal Action and Chemiluminescence.

    PubMed

    Allen, Robert C

    2015-01-01

    Neutrophil leukocytes protect against a varied and complex array of microbes by providing microbicidal action that is simple, potent, and focused. Neutrophils provide such action via redox reactions that change the frontier orbitals of oxygen (O2) facilitating combustion. The spin conservation rules define the symmetry barrier that prevents direct reaction of diradical O2 with nonradical molecules, explaining why combustion is not spontaneous. In burning, the spin barrier is overcome when energy causes homolytic bond cleavage producing radicals capable of reacting with diradical O2 to yield oxygenated radical products that further participate in reactive propagation. Neutrophil mediated combustion is by a different pathway. Changing the spin quantum state of O2 removes the symmetry restriction to reaction. Electronically excited singlet molecular oxygen ((1)O2(*)) is a potent electrophilic reactant with a finite lifetime that restricts its radius of reactivity and focuses combustive action on the target microbe. The resulting exergonic dioxygenation reactions produce electronically excited carbonyls that relax by light emission, that is, chemiluminescence. This overview of neutrophil combustive microbicidal action takes the perspectives of spin conservation and bosonic-fermionic frontier orbital considerations. The necessary principles of particle physics and quantum mechanics are developed and integrated into a fundamental explanation of neutrophil microbicidal metabolism.

  13. Neutrophil Leukocyte: Combustive Microbicidal Action and Chemiluminescence

    PubMed Central

    Allen, Robert C.

    2015-01-01

    Neutrophil leukocytes protect against a varied and complex array of microbes by providing microbicidal action that is simple, potent, and focused. Neutrophils provide such action via redox reactions that change the frontier orbitals of oxygen (O2) facilitating combustion. The spin conservation rules define the symmetry barrier that prevents direct reaction of diradical O2 with nonradical molecules, explaining why combustion is not spontaneous. In burning, the spin barrier is overcome when energy causes homolytic bond cleavage producing radicals capable of reacting with diradical O2 to yield oxygenated radical products that further participate in reactive propagation. Neutrophil mediated combustion is by a different pathway. Changing the spin quantum state of O2 removes the symmetry restriction to reaction. Electronically excited singlet molecular oxygen (1O2*) is a potent electrophilic reactant with a finite lifetime that restricts its radius of reactivity and focuses combustive action on the target microbe. The resulting exergonic dioxygenation reactions produce electronically excited carbonyls that relax by light emission, that is, chemiluminescence. This overview of neutrophil combustive microbicidal action takes the perspectives of spin conservation and bosonic-fermionic frontier orbital considerations. The necessary principles of particle physics and quantum mechanics are developed and integrated into a fundamental explanation of neutrophil microbicidal metabolism. PMID:26783542

  14. Distinct cellular sources of hepoxilin A3 and leukotriene B4 are used to coordinate bacterial-induced neutrophil transepithelial migration.

    PubMed

    Pazos, Michael A; Pirzai, Waheed; Yonker, Lael M; Morisseau, Christophe; Gronert, Karsten; Hurley, Bryan P

    2015-02-01

    Neutrophilic infiltration is a leading contributor to pathology in a number of pulmonary disease states, including cystic fibrosis. Hepoxilin A3 (HXA3) is a chemotactic eicosanoid shown to mediate the transepithelial passage of neutrophils in response to infection in several model systems and at multiple mucosal surfaces. Another well-known eicosanoid mediating general neutrophil chemotaxis is leukotriene B4 (LTB4). We sought to distinguish the roles of each eicosanoid in the context of infection of lung epithelial monolayers by Pseudomonas aeruginosa. Using human and mouse in vitro transwell model systems, we used a combination of biosynthetic inhibitors, receptor antagonists, as well as mutant sources of neutrophils to assess the contribution of each chemoattractant in driving neutrophil transepithelial migration. We found that following chemotaxis to epithelial-derived HXA3 signals, neutrophil-derived LTB4 is required to amplify the magnitude of neutrophil migration. LTB4 signaling is not required for migration to HXA3 signals, but LTB4 generation by migrated neutrophils plays a significant role in augmenting the initial HXA3-mediated migration. We conclude that HXA3 and LTB4 serve independent roles to collectively coordinate an effective neutrophilic transepithelial migratory response.

  15. Genetic and pharmacological inhibition of CDK9 drives neutrophil apoptosis to resolve inflammation in zebrafish in vivo

    PubMed Central

    Hoodless, Laura J.; Lucas, Christopher D.; Duffin, Rodger; Denvir, Martin A.; Haslett, Christopher; Tucker, Carl S.; Rossi, Adriano G.

    2016-01-01

    Neutrophilic inflammation is tightly regulated and subsequently resolves to limit tissue damage and promote repair. When the timely resolution of inflammation is dysregulated, tissue damage and disease results. One key control mechanism is neutrophil apoptosis, followed by apoptotic cell clearance by phagocytes such as macrophages. Cyclin-dependent kinase (CDK) inhibitor drugs induce neutrophil apoptosis in vitro and promote resolution of inflammation in rodent models. Here we present the first in vivo evidence, using pharmacological and genetic approaches, that CDK9 is involved in the resolution of neutrophil-dependent inflammation. Using live cell imaging in zebrafish with labelled neutrophils and macrophages, we show that pharmacological inhibition, morpholino-mediated knockdown and CRISPR/cas9-mediated knockout of CDK9 enhances inflammation resolution by reducing neutrophil numbers via induction of apoptosis after tailfin injury. Importantly, knockdown of the negative regulator La-related protein 7 (LaRP7) increased neutrophilic inflammation. Our data show that CDK9 is a possible target for controlling resolution of inflammation. PMID:27833165

  16. Characterization of a receptor for human monocyte-derived neutrophil chemotactic factor/interleukin-8

    SciTech Connect

    Grob, P.M.; David, E.; Warren, T.C.; DeLeon, R.P.; Farina, P.R.; Homon, C.A. )

    1990-05-15

    Monocyte-derived neutrophil chemotactic factor/interleukin-8 (MDNCF/IL-8) is an 8,000-dalton protein produced by monocytes which exhibits activity as a chemoattractant for neutrophils with maximal activity achieved at a concentration of 50 ng/ml. This polypeptide has been iodinated by chloramine-T methodology (350 Ci/mM), and specific receptors for MDNCF/IL-8 have been detected on human neutrophils, U937 cells, THP-1 cells, and dimethyl sulfoxide-differentiated HL-60 cells. The binding of MDNCF/IL-8 to human neutrophils is not inhibited by interleukin-1 alpha, tumor necrosis factor-alpha, insulin, or epidermal growth factor. In addition, chemoattractants such as C5a, fMet-Leu-Phe, leukotriene B4, and platelet-activating factor fail to inhibit binding, suggesting that MDNCF/IL-8 utilizes a unique receptor. The receptor for MDNCF/IL-8 is apparently glycosylated since ligand binding is inhibited by the presence of wheat germ agglutinin, a lectin with a binding specificity for N-acetylglucosamine and neuraminic acid. Steady state binding experiments indicate Kd values of 4 and 0.5 nM and receptor numbers of 75,000 and 7,400 for human neutrophils and differentiated HL-60 cells, respectively. 125I-MDNCF/IL-8 bound to human neutrophils is rapidly internalized and subsequently released from cells as trichloroacetic acid-soluble radioactivity. Affinity labeling experiments suggest that the human neutrophil MDNCF/IL-8 receptor exhibits a mass of approximately 58,000 daltons.

  17. hMSCs suppress neutrophil-dominant airway inflammation in a murine model of asthma

    PubMed Central

    Hong, Gyong Hwa; Kwon, Hyouk-Soo; Lee, Kyoung Young; Ha, Eun Hee; Moon, Keun-Ai; Kim, Seong Who; Oh, Wonil; Kim, Tae-Bum; Moon, Hee-Bom; Cho, You Sook

    2017-01-01

    Although chronic eosinophilic inflammation is a common feature in patients with asthma, some patients have neutrophil-dominant inflammation, which is known to be associated with severe asthma.Human mesenchymal stem cells (hMSCs) have shown promise in treating various refractory immunological diseases. Thus, hMSCs may represent an alternative therapeutic option for asthma patients with neutrophil-dominant inflammation, in whom current treatments are ineffective. BALB/c mice exposed to ovalbumin and polyinosinic:polycytidylic acid (Poly I:C) to induce neutrophilic airway inflammation were systemically treated with hMSCs to examine whether the hMSCs can modulate neutrophilic airway inflammation. In addition, cytokine production was evaluated in co-cultures of hMSCs with either anti-CD3/CD28-stimulated peripheral blood mononuclear cells (PBMCs) obtained from asthmatic patients or cells of the human bronchial epithelial cell line BEAS-2B to assess the response to hMSC treatment. The total number of immune cells in bronchoalveolar lavage fluid (BALF) showed a dramatic decrease in hMSC-treated asthmatic mice, and, in particular, neutrophilic infiltration was significantly attenuated. This phenomenon was accompanied by reduced CXCL15 production in the BALF. BEAS-2B cells co-cultured with hMSCs showed reduced secretion of IL-8. Moreover, decreased secretion of IL-4, IL-13 and IFN-γ was observed when human PBMCs were cultured with hMSCs, whereas IL-10 production was greatly enhanced. Our data imply that hMSCs may have a role in reducing neutrophilic airway inflammation by downregulating neutrophil chemokine production and modulating T-cell responses. PMID:28127050

  18. Enhanced neutrophil activity is associated with shorter time to tumor progression in glioblastoma patients

    PubMed Central

    Rahbar, Afsar; Cederarv, Madeleine; Wolmer-Solberg, Nina; Tammik, Charlotte; Stragliotto, Giuseppe; Peredo, Inti; Fornara, Olesja; Xu, Xinling; Dzabic, Mensur; Taher, Chato; Skarman, Petra; Söderberg-Nauclér, Cecilia

    2016-01-01

    ABSTRACT Glioblastoma multiforme (GBM) is a highly malignant tumor with a poor outcome that is often positive for human cytomegalovirus (HCMV). GBM patients often have excessive numbers of neutrophils and macrophages near and within the tumor. Here, we characterized the cytokine patterns in the blood of GBM patients with and without Valganciclovir treatment. Furthermore, we determined whether neutrophil activation is related to HCMV status and patient outcome. Blood samples for analyses of cytokines and growth factors were collected from 42 GBM patients at the time of diagnosis (n = 42) and at weeks 12 and 24 after surgery. Blood neutrophils of 28 GBM patients were examined for CD11b expression. The levels of pro- and anti-inflammatory cytokines and chemokines—including interleukin (IL)-1β, IL-2, IL-6, IL-8, IL-10, IL-12p70, IL-17A, transforming growth factor (TGF)-β1, interferon-γ, interferon-α, tumor necrosis factor α, and monocyte chemoattractant protein (MCP)-1were analyzed with a bead-based flow cytometry assay. During the first six months after surgery, neutrophil activity was increased in 12 patients and was unchanged or decreased in 16. Patients with increased neutrophil activity had enhanced IL-12p70, high grade HCMV and a shorter time to tumor progression (TTP) than patients without or decreased neutrophil activity (median TTP; 5.4 vs. 12 months, 95% confidence interval; 1.6–10 vs. 0.1–0.6, hazard ratio = 3 vs. 0.4, p = 0.004). The levels of IL-12p70 were significantly decreased in Valganciclovir treated patients (n = 22, T 12W vs. T 24W, p = 0.03). In conclusion, our findings suggest that neutrophil activation is an early sign of tumor progression in GBM patients. PMID:27057448

  19. CXCL8((3-73))K11R/G31P antagonizes the neutrophil chemoattractants present in pasteurellosis and mastitis lesions and abrogates neutrophil influx into intradermal endotoxin challenge sites in vivo.

    PubMed

    Li, Fang; Zhang, Xiaobei; Mizzi, Chris; Gordon, John R

    2002-11-01

    The ELR(+) CXC chemokines are critical for protective neutrophil responses to most bacterial infections, but nevertheless can contribute importantly to the pathogenic effects of many inflammatory responses. We recently engineered a series of high affinity CXCL8/IL-8 antagonists, one of which, CXCL8((3-73))K11R/G31P, binds very strongly to neutrophils via the CXCR1 and CXCR2. Herein we show in competitive 125I-ligand binding assays that bovine CXCL8((3-73))K11R/G31P has an affinity for neutrophils that is 2-3 orders of magnitude higher than that of CXCL8/IL-8. Furthermore, when used at approximately 0.5 nM, CXCL8((3-73))K11R/G31P inhibited by 50% the chemotactic responses of neutrophils to 129 nM CXCL8/IL-8, but it also blocked chemotactic responses to the alternate ELR-CXC chemokines CXCL1/GRO alpha and CXCL5/ENA-78. Furthermore, CXCL8((3-73))K11R/G31P could inhibit by 93-97% the spectrum of neutrophil chemotactic activities present within wash fluids from clinical bacterial pneumonia or experimental endotoxin-induced mastitis lesions. Finally, intramuscular or subcutaneous application of CXCL8((3-73))K11R/G31P (75 micro g/kg) reduced by up to 97% neutrophil infiltration into intradermal endotoxin challenge sites in cattle, and prevented their circulating neutrophils from responding to CXCL8/IL-8 or ENA-78 in vitro. This data thus encourages further investigation of the potential impact of this novel antagonist on ELR-CXC chemokine-driven inflammatory disorders.

  20. Neutrophil maturation rate determines the effects of dipeptidyl peptidase 1 inhibition on neutrophil serine protease activity

    PubMed Central

    Wikell, C; Clifton, S; Shearer, J; Benjamin, A; Peters, S A

    2016-01-01

    Background and Purpose Neutrophil serine proteases (NSPs) are activated by dipeptidyl peptidase 1 (DPP1) during neutrophil maturation. The effects of neutrophil turnover rate on NSP activity following DPP1 inhibition was studied in a rat pharmacokinetic/pharmacodynamic model. Experimental Approach Rats were treated with a DPP1 inhibitor twice daily for up to 14 days; NSP activity was measured in onset or recovery studies, and an indirect response model was fitted to the data to estimate the turnover rate of the response. Key Results Maximum NSP inhibition was achieved after 8 days of treatment and a reduction of around 75% NSP activity was achieved at 75% in vitro DPP1 inhibition. Both the rate of inhibition and recovery of NSP activity were consistent with a neutrophil turnover rate of between 4–6 days. Using human neutrophil turnover rate, it is predicted that maximum NSP inhibition following DPP1 inhibition takes around 20 days in human. Conclusions and Implications Following inhibition of DPP1 in the rat, the NSP activity was determined by the amount of DPP1 inhibition and the turnover of neutrophils and is thus supportive of the role of neutrophil maturation in the activation of NSPs. Clinical trials to monitor the effect of a DPP1 inhibitor on NSPs should take into account the delay in maximal response on the one hand as well as the potential delay in a return to baseline NSP levels following cessation of treatment. PMID:27186823

  1. GROUP B STREPTOCOCCUS CIRCUMVENTS NEUTROPHILS AND NEUTROPHIL EXTRACELLULAR TRAPS DURING AMNIOTIC CAVITY INVASION AND PRETERM LABOR

    PubMed Central

    Boldenow, Erica; Gendrin, Claire; Ngo, Lisa; Bierle, Craig; Vornhagen, Jay; Coleman, Michelle; Merillat, Sean; Armistead, Blair; Whidbey, Christopher; Alishetti, Varchita; Santana-Ufret, Veronica; Ogle, Jason; Gough, Michael; Srinouanprachanh, Sengkeo; MacDonald, James W; Bammler, Theo K; Bansal, Aasthaa; Liggitt, H. Denny; Rajagopal, Lakshmi; Waldorf, Kristina M Adams

    2016-01-01

    Preterm birth is a leading cause of neonatal morbidity and mortality. Although microbial invasion of the amniotic cavity (MIAC) is associated with the majority of early preterm births, the temporal events that occur during MIAC and preterm labor are not known. Group B Streptococci (GBS) are β-hemolytic, gram-positive bacteria, which commonly colonize the vagina but have been recovered from the amniotic fluid in preterm birth cases. To understand temporal events that occur during MIAC, we utilized a unique chronically catheterized nonhuman primate model that closely emulates human pregnancy. This model allows monitoring of uterine contractions, timing of MIAC and immune responses during pregnancy-associated infections. Here, we show that adverse outcomes such as preterm labor, MIAC, and fetal sepsis were observed more frequently during infection with hemolytic GBS when compared to nonhemolytic GBS. Although MIAC was associated with systematic progression in chorioamnionitis beginning with chorionic vasculitis and progressing to neutrophilic infiltration, the ability of the GBS hemolytic pigment toxin to induce neutrophil cell death and subvert killing by neutrophil extracellular traps (NETs) in placental membranes in vivo facilitated MIAC and fetal injury. Furthermore, compared to maternal neutrophils, fetal neutrophils exhibit decreased neutrophil elastase activity and impaired phagocytic functions to GBS. Collectively, our studies demonstrate how a unique bacterial hemolytic lipid toxin enables GBS to circumvent neutrophils and NETs in placental membranes to induce fetal injury and preterm labor. PMID:27819066

  2. Attachment and ingestion of gonococci human neutrophils.

    PubMed

    Dilworth, J A; Hendley, J O; Mandell, G L

    1975-03-01

    Previous studies have indirectly shown that type 1 gonococci are more resistant to phagocytosis by human neutrophils (PMN) than type 3 gonococci. Using phase contrast, fluorescent, and light microscopy, we directly quantitated PMN-gonococcal interaction, with emphasis on separating ingestion from attachment. PMN monolayers were incubated on slides with type 1 or type 3 gonococcal fluorescent antibody (FA). After methanol fixation, the FA-stained gonococci associated with PMN were cointed. Since the live PMN excludes FA, the FA-stained gonococci represent only extracellular gonococci. Methylene blue was then added to the smae slide to stain both ingested and surface attached gonococci. Using these methods, intracellular and extracellular cell-associated gonococci were quantitated under varying conditions. The numbers of methylene blue-stained cell-associated gonococci that were ingested were: with normal serum, 3.7 plus or minus 4.1 per cent for type 1 and 56.2 plus or minus 3.7 percent for type 3 (P smaller than 0.001); with heat-inactivated serum, 1.0 plus or minus 3.0 per cent for type 1 and 52.6 plus or minus 3.7 per cent for type 3 (P smaller than 0.001); with higher-titer anti-gonococcal antibody serum, 4.8 plus or minus 4.3 percent for type 1 and 64.0 plus or minus 1.6 per cent for type 3 (P smaller than 0.001). Thus, most type 3 organisms were ingested, but most type 1 gonococci were bound on the PMN surface.

  3. Attachment and ingestion of gonococci human neutrophils.

    PubMed Central

    Dilworth, J A; Hendley, J O; Mandell, G L

    1975-01-01

    Previous studies have indirectly shown that type 1 gonococci are more resistant to phagocytosis by human neutrophils (PMN) than type 3 gonococci. Using phase contrast, fluorescent, and light microscopy, we directly quantitated PMN-gonococcal interaction, with emphasis on separating ingestion from attachment. PMN monolayers were incubated on slides with type 1 or type 3 gonococcal fluorescent antibody (FA). After methanol fixation, the FA-stained gonococci associated with PMN were cointed. Since the live PMN excludes FA, the FA-stained gonococci represent only extracellular gonococci. Methylene blue was then added to the smae slide to stain both ingested and surface attached gonococci. Using these methods, intracellular and extracellular cell-associated gonococci were quantitated under varying conditions. The numbers of methylene blue-stained cell-associated gonococci that were ingested were: with normal serum, 3.7 plus or minus 4.1 per cent for type 1 and 56.2 plus or minus 3.7 percent for type 3 (P smaller than 0.001); with heat-inactivated serum, 1.0 plus or minus 3.0 per cent for type 1 and 52.6 plus or minus 3.7 per cent for type 3 (P smaller than 0.001); with higher-titer anti-gonococcal antibody serum, 4.8 plus or minus 4.3 percent for type 1 and 64.0 plus or minus 1.6 per cent for type 3 (P smaller than 0.001). Thus, most type 3 organisms were ingested, but most type 1 gonococci were bound on the PMN surface. Images PMID:46842

  4. Metabolic requirements for neutrophil extracellular traps formation

    PubMed Central

    Rodríguez-Espinosa, Oscar; Rojas-Espinosa, Oscar; Moreno-Altamirano, María Maximina Bertha; López-Villegas, Edgar Oliver; Sánchez-García, Francisco Javier

    2015-01-01

    As part of the innate immune response, neutrophils are at the forefront of defence against infection, resolution of inflammation and wound healing. They are the most abundant leucocytes in the peripheral blood, have a short lifespan and an estimated turnover of 1010 to 1011 cells per day. Neutrophils efficiently clear microbial infections by phagocytosis and by oxygen-dependent and oxygen-independent mechanisms. In 2004, a new neutrophil anti-microbial mechanism was described, the release of neutrophil extracellular traps (NETs) composed of DNA, histones and anti-microbial peptides. Several microorganisms, bacterial products, as well as pharmacological stimuli such as PMA, were shown to induce NETs. Neutrophils contain relatively few mitochondria, and derive most of their energy from glycolysis. In this scenario we aimed to analyse some of the metabolic requirements for NET formation. Here it is shown that NETs formation is strictly dependent on glucose and to a lesser extent on glutamine, that Glut-1, glucose uptake, and glycolysis rate increase upon PMA stimulation, and that NET formation is inhibited by the glycolysis inhibitor, 2-deoxy-glucose, and to a lesser extent by the ATP synthase inhibitor oligomycin. Moreover, when neutrophils were exposed to PMA in glucose-free medium for 3 hr, they lost their characteristic polymorphic nuclei but did not release NETs. However, if glucose (but not pyruvate) was added at this time, NET release took place within minutes, suggesting that NET formation could be metabolically divided into two phases; the first, independent from exogenous glucose (chromatin decondensation) and, the second (NET release), strictly dependent on exogenous glucose and glycolysis. PMID:25545227

  5. Adaptation to Resistance Training Is Associated with Higher Phagocytic (but Not Oxidative) Activity in Neutrophils of Older Women

    PubMed Central

    Bartholomeu-Neto, João; Brito, Ciro José; Nóbrega, Otávio Toledo; Sousa, Vinícius Carolino; Oliveira Toledo, Juliana; Silva Paula, Roberta; Alves, David Junger Fonseca; Ferreira, Aparecido Pimentel; Franco Moraes, Clayton; Córdova, Cláudio

    2015-01-01

    Failure in antimicrobial activity contributes to high morbidity and mortality in the geriatric population. Little is known about the potential effect of resistance training (RT) on the functional properties of the innate immunity. This study aimed to investigate the influence of long-term RT on the endocytic and oxidative activities of neutrophils and monocytes in healthy older women. Our results indicate that the phagocytosis index (PhI) of neutrophils (but not of monocytes) in the RT-adapted group was significantly higher (P < 0.001; effect size, (d) = 0.90, 95% CI: [0.75–1.04]) compared to that in sedentary subjects. In contrast, the oxidative activity of either neutrophils or monocytes was not significantly influenced by RT. Also, total energy and carbohydrate intake as well as serum IL6 levels had a significant influence on the phagocytic activity of neutrophils (P = 0.04), being considered in the model. Multivariate regression identified the physical condition of the subject (β = 0.425; P = 0.01) as a significant predictor of PhI. In conclusion, circulating neutrophils of older women adapted to a long-term RT program expressed higher phagocytic activity. PMID:26524964

  6. A neutrophil intrinsic impairment affecting Rab27a and degranulation in cystic fibrosis is corrected by CFTR potentiator therapy

    PubMed Central

    Pohl, Kerstin; Hayes, Elaine; Keenan, Joanne; Henry, Michael; Meleady, Paula; Molloy, Kevin; Jundi, Bakr; Bergin, David A.; McCarthy, Cormac; McElvaney, Oliver J.; White, Michelle M.; Clynes, Martin; McElvaney, Noel G.

    2014-01-01

    Studies have endeavored to reconcile whether dysfunction of neutrophils in people with cystic fibrosis (CF) is a result of the genetic defect or is secondary due to infection and inflammation. In this study, we illustrate that disrupted function of the CF transmembrane conductance regulator (CFTR), such as that which occurs in patients with ∆F508 and/or G551D mutations, correlates with impaired degranulation of antimicrobial proteins. We demonstrate that CF blood neutrophils release less secondary and tertiary granule components compared with control cells and that activation of the low-molecular-mass GTP-binding protein Rab27a, involved in the regulation of granule trafficking, is defective. The mechanism leading to impaired degranulation involves altered ion homeostasis caused by defective CFTR function with increased cytosolic levels of chloride and sodium, yet decreased magnesium measured in CF neutrophils. Decreased magnesium concentration in vivo and in vitro resulted in significantly decreased levels of GTP-bound Rab27a. Treatment of G551D patients with the ion channel potentiator ivacaftor resulted in normalized neutrophil cytosolic ion levels and activation of Rab27a, thereby leading to increased degranulation and bacterial killing. Our results confirm that intrinsic alterations of circulating neutrophils from patients with CF are corrected by ivacaftor, thus illustrating additional clinical benefits for CFTR modulator therapy. PMID:24934256

  7. Fatty acid binding protein 4 in circulating leucocytes reflects atherosclerotic lesion progression in Apoe−/− mice

    PubMed Central

    Agardh, Hanna E; Gertow, Karl; Salvado, Dolores M; Hermansson, Andreas; Puijvelde, Gijs H; Hansson, Göran K; n-Berne, Gabrielle Paulsso; Gabrielsen, Anders

    2013-01-01

    Discovery of novel biomarkers for atherosclerosis is important to aid in early diagnosis of pre-symptomatic patients at high risk of cardiovascular events. The aim of the present study was therefore to identify potential biomarkers in circulating cells reflecting atherosclerotic lesion progression in the vessel wall. We performed gene arrays on circulating leucocytes from atherosclerosis prone Apoe−/− mice with increasing ages, using C57BL/6 mice as healthy controls. We identified fatty acid binding protein 4 (FABP4) mRNA to be augmented in mice with established disease compared with young Apoe−/− or controls. Interestingly, the transcript FABP4 correlated significantly with lesion size, further supporting a disease associated increase. In addition, validation of our finding on protein level showed augmented FABP4 in circulating leucocytes whereas, importantly, no change could be observed in plasma. Immunofluorescence analysis demonstrated FABP4 to be present mainly in circulating neutrophils and to some extent in monocytes. Moreover, FABP4-positive neutrophils and macrophages could be identified in the subintimal space in the plaque. Using human circulating leucocytes, we confirmed the presence of FABP4 protein in neutrophils and monocytes. In conclusion, we have showed that cellular levels of FABP4 in circulating leucocytes associate with lesion development in the experimental Apoe−/− model. The increased expression is primarily localized to neutrophils, but also in monocytes. We have identified FABP4 in leucocytes as a potential and easy accessible biomarker of atherosclerosis which could be of future clinical relevance. PMID:23387955

  8. P-selectin must extend a sufficient length from the plasma membrane to mediate rolling of neutrophils

    PubMed Central

    1995-01-01

    Under physiological shear stress, neutrophils roll on P-selectin on activated endothelial cells or platelets through interactions with P- selectin glycoprotein ligand-1 (PSGL-1). Both P-selectin and PSGL-1 are extended molecules. Human P-selectin contains an NH2-terminal lectin domain, an EGF domain, nine consensus repeats (CRs), a transmembrane domain, and a cytoplasmic tail. To determine whether the length of P- selectin affected its interactions with PSGL-1, we examined the adhesion of neutrophils to CHO cells expressing membrane-anchored P- selectin constructs in which various numbers of CRs were deleted. Under static conditions, neutrophils attached equivalently to wild-type P- selectin and to constructs containing from 2-6 CRs. Under shear stress, neutrophils attached equivalently to wild-type and 6 CR P-selectin and nearly as well to 5 CR P-selectin. However, fewer neutrophils attached to the 4 CR construct, and those that did attach rolled faster and were more readily detached by increasing shear stress. Flowing neutrophils failed to attach to the 3 CR and 2 CR constructs. Neutrophils attached and rolled more efficiently on 4 CR P-selectin expressed on glycosylation-defective Lec8 CHO cells, which have less glycocalyx. We conclude that P-selectin must project its lectin domain well above the membrane to mediate optimal attachment of neutrophils under shear forces. The length of P-selectin may: (a) facilitate interactions with PSGL-1 on flowing neutrophils, and (b) increase the intermembrane distance where specific bonds form, minimizing contacts between the glycocalyces that result in cell-cell repulsion. PMID:8557755

  9. Characterisation of Neutropenia-Associated Neutrophil Elastase Mutations in a Murine Differentiation Model In Vitro and In Vivo

    PubMed Central

    Wiesmeier, Michael; Gautam, Sanjivan

    2016-01-01

    Severe congenital neutropenia (SCN) is characterised by a differentiation block in the bone marrow and low neutrophil numbers in the peripheral blood, which correlates with increased risk of bacterial infections. Several underlying gene defects have been identified in SCN patients. Mutations in the neutrophil elastase (ELANE) gene are frequently found in SCN and cyclic neutropenia. Both mislocalization and misfolding of mutant neutrophil elastase protein resulting in ER stress and subsequent induction of the unfolded protein response (UPR) have been proposed to be responsible for neutrophil survival and maturation defects. However, the detailed molecular mechanisms still remain unclear, in part due to the lack of appropriate in vitro and in vivo models. Here we used a system of neutrophil differentiation from immortalised progenitor lines by conditional expression of Hoxb8, permitting the generation of mature near-primary neutrophils in vitro and in vivo. NE-deficient Hoxb8 progenitors were reconstituted with murine and human forms of typical NE mutants representative of SCN and cyclic neutropenia, and differentiation of the cells was analysed in vitro and in vivo. ER stress induction by NE mutations could be recapitulated during neutrophil differentiation in all NE mutant-reconstituted Hoxb8 cells. Despite ER stress induction, no change in survival, maturation or function of differentiating cells expressing either murine or human NE mutants was observed. Further analysis of in vivo differentiation of Hoxb8 cells in a murine model of adoptive transfer did not reveal any defects in survival or differentiation in the mouse. Although the Hoxb8 system has been found to be useful for dissection of defects in neutrophil development, our findings indicate that the use of murine systems for analysis of NE-mutation-associated pathogenesis is complicated by differences between humans and mice in the physiology of granulopoiesis, which may go beyond possible differences in

  10. Characterisation of Neutropenia-Associated Neutrophil Elastase Mutations in a Murine Differentiation Model In Vitro and In Vivo.

    PubMed

    Wiesmeier, Michael; Gautam, Sanjivan; Kirschnek, Susanne; Häcker, Georg

    2016-01-01

    Severe congenital neutropenia (SCN) is characterised by a differentiation block in the bone marrow and low neutrophil numbers in the peripheral blood, which correlates with increased risk of bacterial infections. Several underlying gene defects have been identified in SCN patients. Mutations in the neutrophil elastase (ELANE) gene are frequently found in SCN and cyclic neutropenia. Both mislocalization and misfolding of mutant neutrophil elastase protein resulting in ER stress and subsequent induction of the unfolded protein response (UPR) have been proposed to be responsible for neutrophil survival and maturation defects. However, the detailed molecular mechanisms still remain unclear, in part due to the lack of appropriate in vitro and in vivo models. Here we used a system of neutrophil differentiation from immortalised progenitor lines by conditional expression of Hoxb8, permitting the generation of mature near-primary neutrophils in vitro and in vivo. NE-deficient Hoxb8 progenitors were reconstituted with murine and human forms of typical NE mutants representative of SCN and cyclic neutropenia, and differentiation of the cells was analysed in vitro and in vivo. ER stress induction by NE mutations could be recapitulated during neutrophil differentiation in all NE mutant-reconstituted Hoxb8 cells. Despite ER stress induction, no change in survival, maturation or function of differentiating cells expressing either murine or human NE mutants was observed. Further analysis of in vivo differentiation of Hoxb8 cells in a murine model of adoptive transfer did not reveal any defects in survival or differentiation in the mouse. Although the Hoxb8 system has been found to be useful for dissection of defects in neutrophil development, our findings indicate that the use of murine systems for analysis of NE-mutation-associated pathogenesis is complicated by differences between humans and mice in the physiology of granulopoiesis, which may go beyond possible differences in

  11. A Morphological and Cytochemical Study of the Interaction between Paracoccidiodes brasiliensis and Neutrophils

    NASA Astrophysics Data System (ADS)

    Dias, Maria Fernanda R. G.; Filgueira, Absalom L.; de Souza, Wanderley

    2004-04-01

    Paracoccidioidomycosis is a systemic granulomatous disease caused by the dimorphic fungus Paracoccidioides brasiliensis. It is the most prevalent systemic mycosis of Latin America and 80% of the reported cases are from Brazil. Because of the great number of neutrophils found in the P. brasiliensis granuloma, studies have been done to evaluate the role of these cells during the development of the infection. Scanning and transmission electron microscopy of thin sections showed that the neutrophils ingest yeast cells through a typical phagocytic process with the formation of pseudopodes. The pseudopodes even disrupt the connection established between the mother and the bud cells. Neutrophils also associate to each other, forming a kind of extracellular vacuole where large yeast cells are encapsulated. Cytochemical studies showed that once P. brasiliensis attaches to the neutrophil surface, it triggers a respiratory burst with release of oxygen-derived products. Attachment also triggers neutrophils' degranulation, with release of endogenous peroxidase localized in cytoplasmic granules. Together, these processes lead to killing of both ingested and extracellular P. brasiliensis.

  12. Decreased phagocytic function in neutrophils and monocytes from peripheral blood in periodontal disease

    PubMed Central

    CARNEIRO, Valéria Martins Araújo; BEZERRA, Ana Cristina Barreto; GUIMARÃES, Maria do Carmo Machado; MUNIZ-JUNQUEIRA, Maria Imaculada

    2012-01-01

    Phagocytosis by neutrophils and monocytes constitutes the main defense mechanism against bacterial challenges in periodontitis. Phagocytosis by neutrophils has already been evaluated, whereas phagocytic function of monocytes has hardly been addressed so far. Objectives The aim of this study was to assess phagocytosis by neutrophils and monocytes in periodontitis. Material and Methods The sample included 30 subjects with severe periodontitis and 27 control subjects without periodontal disease. The phagocytic index (PhI) was calculated as the mean number of adhered/ingested Saccharomyces cerevisiae per phagocytozing monocyte or neutrophil multiplied by the percentage of phagocytes involved in phagocytosis. Results A significant reduction in phagocyte functions was observed in individuals with periodontitis. The median of PhI of neutrophils using non-sensitized S. cerevisiae was 3 for the control group, and 1.5 for the periodontitis group (p=0.01, Mann-Whitney test). The median of PhI of monocytes with non-sensitized S. cerevisiae was 26.13 for the control group, and 13.23 for the periodontitis group (p=0.03, Mann Whitney test). The median of PhI of monocytes assessed with sensitized S. cerevisiae was 97.92 for the control group and 60.1 for the periodontitis group (p=0.005, t-test). Conclusion The data demonstrated a reduction in the function of phagocytes, suggesting a decrease in immune defenses in periodontitis. PMID:23138734

  13. Immunostimulation of sugar cane extract on neutrophils to Salmonella typhimurium infection in mice.

    PubMed

    Chen, Ming-Hua; Lo, Dan-Yuan; Liao, Jiunn-Wang; Hsuan, Shih-Ling; Chien, Maw-Sheng; Lin, Cheng-Chung; Chen, Ter-Hsin; Lee, Wei-Cheng

    2012-07-01

    The aim of this study was to evaluate the immunomodulatory effects of sugar cane extract (SCE) on the biological activities of neutrophils in mice. Six-week-old BALB/c mice were fed 1250 mg/kg of SCE once. The generation, migration and biological functions of neutrophils and the survival rates of the mice in response to Salmonella typhimurium infection were evaluated. The results show that the numbers of both bone marrow cells and neutrophils were significantly increased in response to SCE administration (p < 0.05) compared with controls. The migration, phagocytosis and H₂O₂ generation of neutrophils were all significantly enhanced in SCE-treated mice (p < 0.05). After challenge with S. typhimurium (lethal dose, 50% (LD₅₀), SCE-treated mice had a 19.2% higher survival rate and milder hepatic lesions than the controls. Additionally, fewer invasive bacteria were recovered from the spleens of SCE-treated mice. In conclusion, our results suggest that SCE has a positive regulatory effect on the biological function of mouse neutrophils that may increase host resistance against bacterial infections.

  14. Mycobacterium tuberculosis inhibits neutrophil apoptosis, leading to delayed activation of naive CD4 T cells.

    PubMed

    Blomgran, Robert; Desvignes, Ludovic; Briken, Volker; Ernst, Joel D

    2012-01-19

    Mycobacterium tuberculosis promotes its replication by inhibiting the apoptosis of infected macrophages. A proapoptotic M. tuberculosis mutant lacking nuoG, a subunit of the type I NADH dehydrogenase complex, exhibits attenuated growth in vivo, indicating that this virulence mechanism is essential. We show that M. tuberculosis also suppresses neutrophil apoptosis. Compared to wild-type, the nuoG mutant spread to a larger number of lung phagocytic cells. Consistent with the shorter lifespan of infected neutrophils, infection with the nuoG mutant resulted in fewer bacteria per infected neutrophil, accelerated bacterial acquisition by dendritic cells, earlier trafficking of these dendritic cells to lymph nodes, and faster CD4 T cell priming. Neutrophil depletion abrogated accelerated CD4 T cell priming by the nuoG mutant, suggesting that inhibiting neutrophil apoptosis delays adaptive immunity in tuberculosis. Thus, pathogen modulation of apoptosis is beneficial at multiple levels, and enhancing phagocyte apoptosis promotes CD4 as well as CD8 T cell responses.

  15. Altered neutrophil immunophenotypes in childhood B-cell precursor acute lymphoblastic leukemia

    PubMed Central

    Oliveira, Elen; Bacelar, Thiago S.; Ciudad, Juana; Ribeiro, Maria Cecília M.; Garcia, Daniela R.N.; Sedek, Lukasz; Maia, Simone F.; Aranha, Daniel B.; Machado, Indyara C.; Ikeda, Arissa; Baglioli, Bianca F.; Lopez-Duarte, Nathalia; Teixeira, Lisandra A. C.; Szczepanski, Tomasz; Silva, Maria Luiza M.; Land, Marcelo G.P.

    2016-01-01

    An increasing number of evidences suggest a genetic predisposition in acute lymphoblastic leukemia (ALL) that might favor the occurrence of the driver genetic alterations. Such genetic background might also translate into phenotypic alterations of residual hematopoietic cells. Whether such phenotypic alterations are present in bone marrow (BM) cells from childhood B-cell precursor (BCP)-ALL remains to be investigated. Here we analyzed the immunophenotypic profile of BM and peripheral blood (PB) maturing/matured neutrophils from 118 children with BCP-ALL and their relationship with the features of the disease. Our results showed altered neutrophil phenotypes in most (77%) BCP-ALL cases. The most frequently altered marker was CD10 (53%), followed by CD33 (34%), CD13 (15%), CD15/CD65 (10%) and CD123 (7%). Of note, patients with altered neutrophil phenotypes had younger age (p = 0.03) and lower percentages of BM maturing neutrophils (p = 0.004) together with greater BM lymphocyte (p = 0.04), and mature B-cell (p = 0.03) counts. No significant association was found between an altered neutrophil phenotype and other disease features. These findings point out the potential existence of an altered residual hematopoiesis in most childhood BCP-ALL cases. PMID:27028865

  16. Primed neutrophil infiltrations into multiple organs in child physical abuse cases: A preliminary study.

    PubMed

    Hayashi, Takahito; Nakamae, Takuma; Higo, Eri; Ikematsu, Kazuya; Ogata, Mamoru

    2017-01-01

    Physical abuse of the elderly induces a massive primed neutrophil infiltration into the lung and liver through chemotaxis by interleukin (IL)-8, similar to cases of traumatic or hemorrhagic shock. Here, we used immunohistochemical analyses to investigate this infiltration in cases of physically abused children. In addition, we examined the expression of neutrophil elastase (NE) as the inflammatory mediator and α1-antitrypsin (AAT) as the elastase inhibitor. The number of neutrophils in the abuse cases was increased significantly in the heart, lung, liver, and kidney, compared with that of control cases. IL-8-positive cells and NE-positive cells in all organs of abuse cases were significantly greater than those in control cases. Large quantities of oxidized AAT, which fails to inactivate NE and results in tissue damage, was detected in the liver of abuse cases. Neutrophil infiltration showed positive correlation with the degree of systemic accumulation of non-fatal injuries caused by repetitive abusive behavior. Although further investigation using more autopsy samples is necessary, results of our preliminary study indicate that massive neutrophil infiltration induced by IL-8 in multiple organs is a new complementary diagnostic indicator of physical abuse in children. Moreover, the demonstration of NE-positive cells and oxidized AAT provides firm evidence of tissue damage.

  17. Detection of Circulating Tumor Cells

    PubMed Central

    Terstappen, Leon W. M. M.

    2014-01-01

    The increasing number of treatment options for patients with metastatic carcinomas has created an accompanying need for methods to determine if the tumor will be responsive to the intended therapy and to monitor its effectiveness. Ideally, these methods would be noninvasive and provide quantitative real-time analysis of tumor activity in a variety of carcinomas. Assessment of circulating tumor cells shed into the blood during metastasis may satisfy this need. Here we review the CellSearch technology used for the detection of circulating tumor cells and discuss potential future directions for improvements. PMID:25133014

  18. A bacterial siren song: intimate interactions between neutrophils and pathogenic Neisseria

    PubMed Central

    Criss, Alison K.; Seifert, H. Steven

    2012-01-01

    Preface Neisseria gonorrhoeae and Neisseria meningitidis are Gram-negative bacterial pathogens that are exquisitely adapted for growth at human mucosal surfaces and for efficient transmission between hosts. One factor that is essential to neisserial pathogenesis is the interaction between the bacteria and neutrophils, which are recruited in high numbers during infection. Although this vigorous host response could simply reflect effective immune recognition of the bacteria, there is mounting evidence that in fact these obligate human pathogens manipulate the innate immune response to promote infectious processes. This Review summarizes the mechanisms used by pathogenic neisseriae to resist and modulate the antimicrobial activities of neutrophils. It also details some of the major outstanding questions about the Neisseria–neutrophil relationship and proposes potential benefits of this relationship for the pathogen. PMID:22290508

  19. Aberrant "Barbed-Wire" Nuclear Projections of Neutrophils in Trisomy 18 (Edwards Syndrome).

    PubMed

    Kahwash, Basil M; Nowacki, Nicholas B; Kahwash, Samir B

    2015-01-01

    We discuss the significance of neutrophils with increased, aberrant nuclear projections mimicking "barbed-wire" in a newborn child with trisomy 18 (T18). Increased, aberrant nuclear projections have been previously reported in trisomy of the D group of chromosomes (chromosomes 13, 14, and 15), and we report similar findings in a patient with T18. The peripheral blood smear showed relative neutrophilia with the majority (37%) of neutrophils showing two or more thin, rod-shaped or spike-shaped, and often pedunculated aberrant nuclear projections. The number of projections ranged from 2 to 6 per cell, averaged 2 per affected neutrophil, and ranged in length from 0.22 μm to 0.83 μm. This case confirms that the morphologic finding described is not restricted to trisomy of one of the chromosomes in group D, as implied in the literature.

  20. Neutrophil homeostasis and its regulation by danger signaling.

    PubMed

    Wirths, Stefan; Bugl, Stefanie; Kopp, Hans-Georg

    2014-06-05

    Hematopoiesis in general is demand driven and adaptive, but in contrast to erythropoiesis or thrombocytopoiesis, our knowledge on how neutrophil production is adapted to individual needs remains incomplete. Recently, neutrophil homeostasis has been shown to depend on danger receptors, macrophages, and even circadian rhythms. Puzzle pieces for a broader view of neutrophil homeostasis accumulate, and we will herein try to put seemingly contradictory evidence in a perspective of neutrophil homeostasis and emergency granulopoiesis determined by innate immunologic signaling.

  1. Analysis of Human and Mouse Neutrophil Phagocytosis by Flow Cytometry.

    PubMed

    Fine, Noah; Barzilay, Oriyah; Glogauer, Michael

    2017-01-01

    Neutrophils are primary phagocytes that recognize their targets through surface chemistry, either through Pattern Recognition Receptor (PPR) interaction with Pathogen-Associated Molecular Patterns (PAMPs) or through immunoglobulin (Ig) or complement mediated recognition. Opsonization can be important for target recognition, and phagocytosis by neutrophils in whole blood can be greatly enhanced due to the presence of blood serum components and platelets. Powerful and sensitive flow cytometry based methods are presented to measure phagocytosis by human blood neutrophils and mouse peritoneal neutrophils.

  2. Antimicrobial Decapeptide KSL-W Enhances Neutrophil Chemotaxis and Function

    DTIC Science & Technology

    2011-12-16

    its antimicrobial activity [25]. Because of the known multifunctional activities associated with many antimicrobial peptides, we became interested in...stated. 2.5. Neutrophil treatment and measuring actin polymerization Purified human neutrophils were treated with HBSS, FMLP (10−7 M and 10−10 M), or...control neutrophils were resuspended in 1 ml of 1× DPBS. 2.7. Actin polymerization F- actin content in unstimulated and FMLP-/KSLW-treated neutrophils

  3. CD66b Overexpression and Loss of C5a Receptors as Surface Markers for Staphylococcus aureus-Induced Neutrophil Dysfunction

    PubMed Central

    Schnitzler, Norbert; Grüger, Thomas; Brandenburg, Kerstin; Zinserling, Jörg; Zündorf, Josef

    2015-01-01

    Neutrophil granulocytes constitute the main component of innate immunity in the clearance of bacterial infections. However, during systemic inflammation, immunoparalysis may occur resulting in neutrophil dysfunction. This study presents a new in vitro model for analyzing the dysfunction of human peripheral blood neutrophils resulting from the interaction with Staphylococcus aureus components in whole blood. After induction of a massive complement activation by S. aureus supernatant, the neutrophils exhibit a reduced phagocytic capacity resulting in a dramatic reduction of the antibacterial activity similar to that of neutrophils isolated from septic patients. The number of phagocytozing neutrophils is drastically reduced as well as the phagocytic capacity designated by a significantly lower number of ingested microbes. This dysfunction correlates with the loss of complement component 5a receptor 1 from the neutrophil cell surface and can be further characterized by a C5a-induced CD66b overexpression. The presented in vitro model offers a new platform for preclinical testing of immunosuppressive drugs and delivers new information for the understanding of neutrophil dysfunctions under the conditions described. PMID:26176669

  4. Neutrophil Emigration in the Skin, Lungs, and Peritoneum: Different Requirements for CD11/CD18 Revealed by CD18-deficient Mice

    PubMed Central

    Mizgerd, Joseph P.; Kubo, Hiroshi; Kutkoski, Gregory J.; Bhagwan, Sabrina D.; Scharffetter-Kochanek, Karin; Beaudet, Arthur L.; Doerschuk, Claire M.

    1997-01-01

    To determine the role of CD11/CD18 complexes in neutrophil emigration, inflammation was induced in the skin, lungs, or peritoneum of mutant mice deficient in CD18 (CD18−/− mutants). Peripheral blood of CD18−/− mutants contained 11-fold more neutrophils than did blood of wild-type (WT) mice. During irritant dermatitis induced by topical application of croton oil, the number of emigrated neutrophils in histological sections of dermis was 98% less in CD18−/− mutants than in WT mice. During Streptococcus pneumoniae pneumonia, neutrophil emigration in CD18−/− mutants was not reduced. These data are consistent with expectations based on studies using blocking antibodies to inhibit CD11/CD18 complexes, and on observations of humans lacking CD11/CD18 complexes. The number of emigrated neutrophils in lung sections during Escherichia coli pneumonia, or in peritoneal lavage fluid after 4 h of S. pneumoniae peritonitis, was not reduced in CD18−/− mutants, but rather was greater than the WT values (240 ± 30 and 220 ± 30% WT, respectively). Also, there was no inhibition of neutrophil emigration during sterile peritonitis induced by intraperitoneal injection of thioglycollate (90 ± 20% WT). These data contrast with expectations. Whereas CD11/CD18 complexes are essential to the dermal emigration of neutrophils during acute dermatitis, CD18−/− mutant mice demonstrate surprising alternative pathways for neutrophil emigration during pneumonia or peritonitis. PMID:9334375

  5. Evaluation of altitude-appropriate reference ranges for neutrophils in diagnosis of sepsis in very low birth weight infants: A multicenter retrospective study

    PubMed Central

    Yu, Jialin; Fan, Juan; He, Yu; Dong, Wenhui; Wang, Zhengli

    2017-01-01

    Background The circulating neutrophil count was commonly believed to be influenced by altitude. At present, neutrophil reference ranges (RRs) for very low birth weight (VLBW) neonates are only available from the sea level and from high altitude. This study aimed to construct RRs for neutrophils in VLBW infants in an intermediate-altitude area and assess its usefulness in evaluation for sepsis. Methods This was a multicenter retrospective study of 2173 VLBW infants admitted to 20 hospitals in Chongqing in southwest of China with altitude from 1000 to 2600 feet. The RRs for absolute total neutrophils (ATN), absolute total immature neutrophils (ATI), and immature: total neutrophil proportion were constructed based on “normal” neonates (unlikely infection). Values of neutrophil from septic and uninfected neonates were respectively assessed using the revised RRs and the previous Mouzinho’s and Schmutz’s RRs. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were compared using the McNemar’s test or χ2 test. Results The upper limits for ATN and ATI using the revised RRs were much higher than those using Mouzinho’s RRs, but lower than those using Schmutz’s RRs. The revised RRs and Mouzinho’s RRs had higher sensitivities than Schmutz’s RRs at 73–672 h. The revised RRs had a higher specificity than Mouzinho’s RRs at both 0–72 h and 73–672 h. The NPV for any abnormality in neutrophil values was high at both 0–72 h and 73–672 h irrespective of the RRs used. Conclusion Altitude-appropriate RRs for neutrophils is more suitable to guide the diagnosis and management of neonatal sepsis in VLBW infants. PMID:28182674

  6. In Vitro Oxidation of Collagen Promotes the Formation of Advanced Oxidation Protein Products and the Activation of Human Neutrophils.

    PubMed

    Bochi, Guilherme Vargas; Torbitz, Vanessa Dorneles; de Campos, Luízi Prestes; Sangoi, Manuela Borges; Fernandes, Natieli Flores; Gomes, Patrícia; Moretto, Maria Beatriz; Barbisan, Fernanda; da Cruz, Ivana Beatrice Mânica; Moresco, Rafael Noal

    2016-04-01

    The accumulation of advanced oxidation protein products (AOPPs) has been linked to several pathological conditions. Here, we investigated collagen as a potential source for AOPP formation and determined the effects of hypochlorous acid (HOCl)-treated collagen (collagen-AOPPs) on human neutrophil activity. We also assessed whether alpha-tocopherol could counteract these effects. Exposure to HOCl increased the levels of collagen-AOPPs. Collagen-AOPPs also stimulated the production of AOPPs, nitric oxide (NO), superoxide radicals (O2(-)), and HOCl by neutrophils. Collagen-AOPPs induced apoptosis and decreased the number of viable cells. Alpha-tocopherol prevented the formation of collagen-AOPPs, strongly inhibited the collagen-AOPP-induced production of O2(-) and HOCl, and increased the viability of neutrophils. Our results suggest that collagen is an important protein that interacts with HOCl to form AOPPs, and consequently, collagen-AOPP formation is related to human neutrophil activation and cell death.

  7. Staphylococcus epidermidis and biofilm-associated neutrophils in chronic rhinosinusitis. A pilot study.

    PubMed

    Marcinkiewicz, Janusz; Stręk, Paweł; Strus, Magdalena; Głowacki, Roman; Ciszek-Lenda, Marta; Zagórska-Świeży, Katarzyna; Gawda, Anna; Tomusiak, Anna

    2015-12-01

    A key role of bacterial biofilm in the pathogenesis of chronic rhinosinusitis (CRS) with (CRSwNP) and without nasal polyps (CRSsNP) is commonly accepted. However, the impact of some bacterial species isolated from inflamed sinus mucosa on biofilm formation is unclear. In particular, the role of Staphylococcus epidermidis as aetiological agents of CRS is controversial. Moreover, the effect of biofilm formation on neutrophil infiltration and activity in CRSwNP calls for explanation. In this study, biofilms were found in three of 10 patients (mean age = 46 ± 14) with CRS undergoing endoscopic sinus surgery by means of scanning electron microscopy. Unexpectedly, S. epidermidis was the primary isolated bacteria and was also found to be present in all biofilm-positive mucosa specimens, indicating its pivotal role in the pathogenesis of severe chronic infections associated with biofilm formation. We have also measured the activity of myeloperoxidase (MPO), the most abundant neutrophil enzyme, to demonstrate the presence of neutrophils in the samples tested. Our present results show that the level of MPO in CRS associated with biofilm is lower than that without biofilm. It may suggest either a low number of neutrophils or the presence of a type of neutrophils with compromised antimicrobial activity, described as biofilm-associated neutrophils (BAN). Finally, we conclude that further studies with a large number of CRS cases should be performed to establish the association between S. epidermidis and other frequently isolated bacterial species from paranasal sinuses, with the severity of CRS, biofilm formation and the infiltration of BAN.

  8. Restraint stress alters neutrophil and macrophage phenotypes during wound healing.

    PubMed

    Tymen, Stéphanie D; Rojas, Isolde G; Zhou, Xiaofeng; Fang, Zong Juan; Zhao, Yan; Marucha, Phillip T

    2013-02-01

    Previous studies reported that stress delays wound healing, impairs bacterial clearance, and elevates the risk for opportunistic infection. Neutrophils and macrophages are responsible for the removal of bacteria present at the wound site. The appropriate recruitment and functions of these cells are necessary for efficient bacterial clearance. In our current study we found that restraint stress induced an excessive recruitment of neutrophils extending the inflammatory phase of healing, and the gene expression of neutrophil attracting chemokines MIP-2 and KC. However, restraint stress did not affect macrophage infiltration. Stress decreased the phagocytic abilities of phagocytic cells ex vivo, yet it did not affect superoxide production. The cell surface expression of adhesion molecules CD11b and TLR4 were decreased in peripheral blood monocytes in stressed mice. The phenotype of macrophages present at the wound site was also altered. Gene expression of markers of pro-inflammatory classically activated macrophages, CXCL10 and CCL5, were down-regulated; as were markers associated with wound healing macrophages, CCL22, IGF-1, RELMα; and the regulatory macrophage marker, chemokine CCL1. Restraint stress also induced up-regulation of IL10 gene expression. In summary, our study has shown that restraint stress suppresses the phenotype shift of the macrophage population, as compared to the changes observed during normal wound healing, while the number of macrophages remains constant. We also observed a general suppression of chemokine gene expression. Modulation of the macrophage phenotype could provide a new therapeutic approach in the treatment of wounds under stress conditions in the clinical setting.

  9. Wegener's granulomatosis and autoantibodies to neutrophil antigens

    PubMed Central

    McCluskey, D R; Maxwell, A P; Watt, L

    1988-01-01

    We report five cases of Wegener's granulomatosis all of whom had clinical and histological evidence of disease activity at presentation and in whom autoantibodies to neutrophil antigens were detected. This test may prove useful for the diagnosis of this serious condition and help to monitor disease activity during treatment. PMID:3068870

  10. Extracellular proton release by stimulated neutrophils

    SciTech Connect

    van Zwieten, R.; Wever, R.; Hamers, M.N.; Weening, R.S.; Roos, D.

    1981-07-01

    We have tried to elucidate the mechanism of phagosome acidification in human neutrophils. Assuming that phenomena occurring at the plasma membrane reflect reactions in the phagocytic vacuoles, we have stimulated human neutrophils with agents that induce a ''respiratory burst,'' and we have measured the release of protons into the extracellular medium. Phorbol myristate acetate, N-formyl-methionyl-leucyl-phenylalanine and serum-opsonized zymosan particles each caused a rapid release of protons, concomitant with the increase in oxygen consumption. The stimulated release of protons was strictly coupled to the increase respiration of the cells, because inhibition of the respiration of either anaerobiosis, chlorpromazine, or glycolytic inhibitors also inhibited the release of protons. Also, in the presence of the above-mentioned stimulating agents, neutrophils from three patients with chronic granulomatous disease enhanced neither respiration not proton release. In normal cells, the ratio of deltaH+/-deltaO2 was 1.04 +/- 0.19 (mean +/ SD, n . 13). The mechanism of this proton release is not clear. The amount of lactic and carbonic acid produced by stimulated neutrophils was inadequate to explain the amount of protons released. Perhydroxyl radicals were also ruled out as the source of the protons. Because the cells did not release measurable amounts of phosphate ions, a phosphate-hydroxyl-ion antiport was also excluded. Finally, the lack of any effect of uncouplers renders it unlikely that a respiration-driven proton gradient is built up across the plasma membrane.

  11. [Congenital neutrophil defects and periodontal diseases].

    PubMed

    Del Fabbro, M; Francetti, L; Pizzoni, L; Weinstein, R L

    2000-06-01

    An alteration of the immune system function is one of the main factors involved in the development of periodontal disease. Polymorpho-nuclear neutrophil leukocytes (PMN) play a crucial role in the cell-mediated immune response against bacterial challenge. The mechanism of neutralization of pathogen microorganisms by PMNs involves many different steps: adhesion to capillary endothelium in the inflamed region, trans-endothelial migration, chemotaxis, phagocytosis and, ultimately, bacterial killing by oxidative and non-oxidative mechanisms. A defect in one of these steps leads to altered neutrophil function and, consequently, to a higher host susceptibility to periodontal tissue infection. The main intrinsic neutrophil diseases such as neutropenia, leukocyte adhesion deficiency (LAD-1), Chediak-Higashi syndrome, Papillon-Lefèvre syndrome, chronic granulomatous disease (CGD), are often related to severe and early-onset forms of periodontitis, as described by many evidences in the literature. Therefore PMN dysfunctions, both intrinsic and extrinsic, represent an important risk factor for periodontal disease. Studies on the basic molecular mechanisms of such dysfunctions, also in terms of genetic polymorphisms, recently allowed to identify some specific markers related to a higher susceptibility to the development of disease. Many researches have yet to be performed aiming to gain insight on the dynamics of PMN activation and interaction with other cells, in order to improve and modulate neutrophil function and to develop specific approaches for care and prevention of periodontal diseases.

  12. Leukotriene B4 binding to human neutrophils

    SciTech Connect

    Lin, A.H.; Ruppel, P.L.; Gorman, R.R.

    1984-12-01

    (/sup 3/H) Leukotriene B4 (LTB4) binds concentration dependently to intact human polymorphonuclear leukocytes (PMN's). The binding is saturable, reaches equilibrium in 10 min at 4 degrees C, and is readily reversible. Mathematical modeling analysis reveals biphasic binding of (/sup 3/H) LTB4 indicating two discrete populations of binding sites. The high affinity binding sites have a dissociation constant of 0.46 X 10(-9)M and Bmax of 1.96 X 10(4) sites per neutrophil; the low affinity binding sites have a dissociation constant of 541 X 10(-9)M and a Bmax of 45.16 X 10(4) sites per neutrophil. Competitive binding experiments with structural analogues of LTB4 demonstrate that the interaction between LTB4 and the binding site is stereospecific, and correlates with the relative biological activity of the analogs. At 25 degrees C (/sup 3/H) LTB4 is rapidly dissociated from the binding site and metabolized to 20-OH and 20-COOH-LTB4. Purification of neutrophils in the presence of 5-lipoxygenase inhibitors significantly increases specific (/sup 3/H) LTB4 binding, suggesting that LTB4 is biosynthesized during the purification procedure. These data suggest that stereospecific binding and metabolism of LTB4 in neutrophils are tightly coupled processes.

  13. Association of epithelial damage and signs of neutrophil mobilization in the airways during acute exacerbations of paediatric asthma

    PubMed Central

    Yoshihara, S; Yamada, Y; Abe, T; Lindén, A; Arisaka, O

    2006-01-01

    We examined whether epithelial damage is associated with mobilization of neutrophils or eosinophils in the airway lumen during acute exacerbations of paediatric asthma. Aspirated sputum samples were harvested from 65 paediatric patients (mean age 3·4 ± 0·4 years) during acute exacerbations of asthma. Patients with signs of infection were excluded. The presence of conglomerates of epithelial cells (i.e. ‘Creola bodies) in the aspirated sputum was utilized as a marker of epithelial damage. Among the paediatric asthma patients, 60% displayed Creola bodies (CrB+: n = 39) in their sputum samples whereas the remaining patients did not (CrB–: n = 26). CrB+ patients displayed more than a 20-fold increase in the concentration of the neutrophil-mobilizing cytokine interleukin (IL)-8 (pg/ml) and of the neutrophil product neutrophil elastase (NE, g/l), respectively, compared with CrB– patients (IL-8: 7468·2 ± 1953·6 versus 347·9 ± 72·6, P < 0·01; NE: 2072·4 ± 419·0 versus 438·5 ± 125·7, P < 0·01). Even though not statistically significant, a corresponding trend was observed for the relative number of sputum neutrophils. In contrast, the concentration of the eosinophil-mobilizing cytokine IL-5 and the esoinophil product ECP tended to be lower in CrB+ than in CrB– patients (P > 0·05). In conclusion, as indicated by the analysis of aspirated sputum, epithelial damage is associated with a locally enhanced chemotactic signal for and activity of neutrophils, but not eosinophils, during acute exacerbations of paediatric asthma. It remains to be determined whether these indirect signs of neutrophil mobilization in the airway lumen mirror an increased number of neutrophils in the surrounding airway tissue. PMID:16634793

  14. Endothelial CD99 supports arrest of mouse neutrophils in venules and binds to neutrophil PILRs.

    PubMed

    Goswami, Debashree; März, Sigrid; Li, Yu-Tung; Artz, Annette; Schäfer, Kerstin; Seelige, Ruth; Pacheco-Blanco, Mariana; Jing, Ding; Bixel, Maria Gabriele; Araki, Masatake; Araki, Kimi; Yamamura, Ken-Ichi; Vestweber, Dietmar

    2017-03-30

    CD99 is a crucial regulator of the transmigration (diapedesis) of leukocytes through the blood vessel wall. Here, we report that CD99 acts at 2 different steps in the extravasation process. In agreement with previous antibody-blocking experiments, we found that CD99 gene inactivation caused neutrophil accumulation between venular endothelial cells and the basement membrane in the inflamed cremaster. Unexpectedly, we additionally found that leukocyte attachment to the luminal surface of the venular endothelium was impaired in the absence of CD99. Intravital video microscopy revealed that CD99 supported rapid chemokine-induced leukocyte arrest. Inhibition of leukocyte attachment and extravasation were both solely due to the absence of CD99 on endothelial cells, whereas CD99 on leukocytes was irrelevant. Therefore, we searched for heterophilic ligands of endothelial CD99 on neutrophils. We found that endothelial cells bind to the paired immunoglobulinlike receptors (PILRs) in a strictly CD99-dependent way. In addition, endothelial CD99 was coprecipitated with PILRs from neutrophils that adhered to endothelial cells. Furthermore, soluble CD99 carrying a transferable biotin tag could transfer this tag covalently to PILR when incubated with intact neutrophils. Binding of neutrophils under flow to a surface coated with P-selectin fragment crystallizable (Fc) and intercellular adhesion molecule 1 (ICAM-1) Fc became more shear resistant if CD99 Fc was coimmobilized. This increased shear resistance was lost if neutrophils were preincubated with anti-PILR antibodies. We concluded that endothelial CD99 promotes leukocyte attachment to endothelium in inflamed vessels by a heterophilic ligand. In addition, CD99 binds to PILRs on neutrophils, an interaction that leads to increased shear resistance of the neutrophil attachment to ICAM-1.

  15. Live Brugia malayi microfilariae inhibit transendothelial migration of neutrophils and monocytes.

    PubMed

    Schroeder, Jan-Hendrik; Simbi, Bigboy H; Ford, Louise; Cole, Sara R; Taylor, Mark J; Lawson, Charlotte; Lawrence, Rachel A

    2012-01-01

    Lymphatic filariasis is a major tropical disease caused by the parasite Brugia malayi. Microfilariae (Mf) circulate in the peripheral blood for 2-3 hours in synchronisation with maximal feeding of the mosquito vector. When absent from the peripheral blood, Mf sequester in the capillaries of the lungs. Mf are therefore in close contact with vascular endothelial cells (EC) and may induce EC immune function and/or wound repair mechanisms such as angiogenesis. In this study, Mf were co-cultured with human umbilical vein EC (HUVEC) or human lung microvascular EC (HLMVEC) and the transendothelial migration of leukocyte subsets was analysed. In addition, the protein and/or mRNA expression of chemokine, cytokine and angiogenic mediators in endothelial cells in the presence of live microfilariae were measured by a combination of cDNA arrays, protein arrays, ELISA and fluorescence antibody tests.Surprisingly, our findings indicate that Mf presence partially blocked transendothelial migration of monocytes and neutrophils, but not lymphocytes. However, Mf exposure did not result in altered vascular EC expression of key mediators of the tethering stage of extravasation, such as ICAM-1, VCAM-1 and various chemokines. To further analyse the immunological function of vascular EC in the presence of Mf, we measured the mRNA and/or protein expression of a number of pro-inflammatory mediators. We found that expression levels of the mediators tested were predominantly unaltered upon B. malayi Mf exposure. In addition, a comparison of angiogenic mediators induced by intact Mf and Wolbachia-depleted Mf revealed that even intact Mf induce the expression of remarkably few angiogenic mediators in vascular EC. Our study suggests that live microfilariae are remarkably inert in their induction and/or activation of vascular cells in their immediate local environment. Overall, this work presents important insights into the immunological function of the vascular endothelium during an infection

  16. Periodontal Ligament Stem Cells Regulate Apoptosis of Neutrophils

    PubMed Central

    Wang, Qing; Ding, Gang; Xu, Xin

    2017-01-01

    Abstract Periodontal ligament stem cells (PDLSCs) are promising cell resource for the cell-based therapy for periodontitis and regeneration of bio-root. In this study, we investigated the effect of PDLSCs on neutrophil, a critical constituent of innate immunity, and the underlying mechanisms. The effect of PDLSCs on the proliferation and apoptosis of resting neutrophils and IL-8 activated neutrophils was tested under cell-cell contact culture and Transwell culture, with or without anti-IL-6 neutralizing antibody. We found that PDLSCs could promote the proliferation and reduce the apoptosis of neutrophils whether under cell-cell contact or Transwell culture. Anti-IL-6 antibody reduced PDLSCs-mediated inhibition of neutrophil apoptosis. IL-6 at the concentration of 10ng/ml and 20ng/ml could inhibit neutrophil apoptosis statistically. Collectively, PDLSCs could reduce the apoptosis of neutrophils via IL-6.

  17. Effects of Space Flight on Neutrophil Functions in Astronauts

    NASA Technical Reports Server (NTRS)

    Kaur, Indreshpal; Valadez, Victoria A.; Simons, Elizabeth R.; Pierson, Duane L.

    2000-01-01

    Neutrophil phagocytosis, oxidative burst, degranulation, and the expression of selected surface markers were studied in 25 astronauts following 4 space shuttle missions. Space flight duration ranged from 5 to 11 days. Blood specimens were obtained 10 days before launch, immediately after landing, and again at 3 days after landing. The number of neutrophils increased at landing by 85%. Phagocytosis of Escherichia coli (E. coli) and oxidative burst following the medium length (9 to 11 days) missions were lower than the control mean values. Whereas, following the short-duration (5 days) mission, these functions were unchanged from control values. No consistent changes in degranulation were observed following either short or medium length space missions. The expression of CD16, CD32, CD11a, CD11b, CD11c, L-selectin and CD36 were measured and found to be variable. Specifically, CD16 and CD32 did not correlate with the changes in oxidative burst. Mission duration appears to be a factor in phagocytic and oxidative functions.

  18. Regulators and Effectors of Arf GTPases in Neutrophils.

    PubMed

    Gamara, Jouda; Chouinard, François; Davis, Lynn; Aoudjit, Fawzi; Bourgoin, Sylvain G

    2015-01-01

    Polymorphonuclear neutrophils (PMNs) are key innate immune cells that represent the first line of defence against infection. They are the first leukocytes to migrate from the blood to injured or infected sites. This process involves molecular mechanisms that coordinate cell polarization, delivery of receptors, and activation of integrins at the leading edge of migrating PMNs. These phagocytes actively engulf microorganisms or form neutrophil extracellular traps (NETs) to trap and kill pathogens with bactericidal compounds. Association of the NADPH oxidase complex at the phagosomal membrane for production of reactive oxygen species (ROS) and delivery of proteolytic enzymes into the phagosome initiate pathogen killing and removal. G protein-dependent signalling pathways tightly control PMN functions. In this review, we will focus on the small monomeric GTPases of the Arf family and their guanine exchange factors (GEFs) and GTPase activating proteins (GAPs) as components of signalling cascades regulating PMN responses. GEFs and GAPs are multidomain proteins that control cellular events in time and space through interaction with other proteins and lipids inside the cells. The number of Arf GAPs identified in PMNs is expanding, and dissecting their functions will provide important insights into the role of these proteins in PMN physiology.

  19. Regulators and Effectors of Arf GTPases in Neutrophils

    PubMed Central

    Gamara, Jouda; Chouinard, François; Davis, Lynn; Aoudjit, Fawzi; Bourgoin, Sylvain G.

    2015-01-01

    Polymorphonuclear neutrophils (PMNs) are key innate immune cells that represent the first line of defence against infection. They are the first leukocytes to migrate from the blood to injured or infected sites. This process involves molecular mechanisms that coordinate cell polarization, delivery of receptors, and activation of integrins at the leading edge of migrating PMNs. These phagocytes actively engulf microorganisms or form neutrophil extracellular traps (NETs) to trap and kill pathogens with bactericidal compounds. Association of the NADPH oxidase complex at the phagosomal membrane for production of reactive oxygen species (ROS) and delivery of proteolytic enzymes into the phagosome initiate pathogen killing and removal. G protein-dependent signalling pathways tightly control PMN functions. In this review, we will focus on the small monomeric GTPases of the Arf family and their guanine exchange factors (GEFs) and GTPase activating proteins (GAPs) as components of signalling cascades regulating PMN responses. GEFs and GAPs are multidomain proteins that control cellular events in time and space through interaction with other proteins and lipids inside the cells. The number of Arf GAPs identified in PMNs is expanding, and dissecting their functions will provide important insights into the role of these proteins in PMN physiology. PMID:26609537

  20. Neutrophil extracellular traps involvement in corneal fungal infection

    PubMed Central

    Zhao, Yingying; Zhang, Fan; Wan, Ting; Fan, Fangli; Xie, Xin; Lin, Zhenyun

    2016-01-01

    Purpose Neutrophils release neutrophil extracellular traps (NETs) when defending against invading microorganisms. We investigated the existence of NETs in fungal keratitis. Methods Fourteen patients with unilateral fungal keratitis were included. Detailed information about each patient was recorded, including (1) patient history (onset of symptoms and previous therapy), (2) ocular examination findings by slit-lamp biomicroscopy, (3) laboratory findings from direct smear examination and culture of corneal scrapings, (4) NET formation, and (5) treatment strategy and prognosis. Immunofluorescence staining was used to evaluate the existence of NETs on corneal scrapings. The relationship between the quantification of NETs and the clinical character of the fungal keratitis was identified. Results NETs were identified in all 14 patients. Patients with a higher grade of NET formation and fewer fungal hyphae always showed a good treatment response and a short course of infection. NETs were consistently found mixed with fungal hyphae in the corneal scrapings from infected patients. No statistical significance was found between the grade of NETs formed and the course of infection before presentation, and no relationship between the quantification of NETs and the size of the ulcer was found. Conclusions The results suggest that NETs are involved in fungal keratitis. The number of NETs in infected corneas may provide a tool for evaluating the prognosis for fungal keratitis. PMID:27559290

  1. Circulating glioma biomarkers

    PubMed Central

    Kros, Johan M.; Mustafa, Dana M.; Dekker, Lennard J.M.; Sillevis Smitt, Peter A.E.; Luider, Theo M.; Zheng, Ping-Pin

    2015-01-01

    Validated biomarkers for patients suffering from gliomas are urgently needed for standardizing measurements of the effects of treatment in daily clinical practice and trials. Circulating body fluids offer easily accessible sources for such markers. This review highlights various categories of tumor-associated circulating biomarkers identified in blood and cerebrospinal fluid of glioma patients, including circulating tumor cells, exosomes, nucleic acids, proteins, and oncometabolites. The validation and potential clinical utility of these biomarkers is briefly discussed. Although many candidate circulating protein biomarkers were reported, none of these have reached the required validation to be introduced for clinical practice. Recent developments in tracing circulating tumor cells and their derivatives as exosomes and circulating nuclear acids may become more successful in providing useful biomarkers. It is to be expected that current technical developments will contribute to the finding and validation of circulating biomarkers. PMID:25253418

  2. Neutrophil elastase processing of Gelatinase A is mediated by extracellular matrix

    SciTech Connect

    Rice, A.; Banda, M.J.

    1995-07-18

    Gelatinase A (72-kDa type IV collagenase) is a metalloproteinase that is expressed by many cells in culture and is overexpressed by some tumor cells. It has been suggested that the serine proteinase neutrophil elastase might play a role iii the posttranslational processing of gelatinase A and that noncatalytic interactions between gelatinase A and components of the extracellular matrix might alter potential processing pathways. These questions were addressed with the use of gelatin substrate zymography, gelatinolytic activity assays, and amino acid sequence analysis. We found that neutrophil elastase does proteolytically modify gelatinase A by cleaving at a number of sites within gelatinase A. Sequential treatment of gelatinase A with 4-aminophenylmercuric acetate (APMA) and neutrophil elastase yielded an active gelatinase with a 4-fold increase in gelatinolytic activity. The increased gelatinolytic activity correlated with that of a 40-kDa fragment of gelatinase A. Matrix components altered the proteolytic modifications in gelatinase A that were mediated by neutrophil elastase. In the absence of gelatin, neutrophil elastase destructively degraded gelatinase A by hydrolyzing at least two bonds within the fibronectin-like gelatin-binding domain of gelatinase A. In the presence of gelatin, these two inactivating cleavage sites were protected, and cleavage at a site within the hemopexin-like carboxyl-terminal domain resulted in a truncated yet active gelatinase. The results suggest a regulatory role for extracellular matrix molecules in stabilizing gelatinase A fragments and in altering the availability of sites susceptible to destructive proteolysis by neutrophil elastase. 32 refs., 10 figs.

  3. Neutrophil Functions and Cytokines Expression Profile in Buffaloes with Impending Postpartum Reproductive Disorders

    PubMed Central

    Patra, Manas Kumar; Kumar, Harendra; Nandi, Sukdeb

    2013-01-01

    The study was conducted to correlate the periparturient immune status in terms of neutrophil functions and cytokine expression in peripheral blood mononuclear cell culture with impending postpartum reproductive disorders in buffaloes. Forty pregnant buffaloes were observed for occurrence of postpartum reproductive disorders (PRD), i.e., metritis, endometritis and delayed uterine involution etc., during one week prepartum to four weeks postpartum period. A representative number (n = 6) of buffaloes that did not develop any PRD were included in group I (healthy, control), while the animals which experienced PRD were assigned into group II (PRD, n = 8). The blood samples were collected at weekly interval from one week prepartum to four weeks postpartum period considering the day of calving as ‘d 0’. Differential leucocytes counts, superoxide and hydrogen peroxide production activity in isolated neutrophils and the mRNA expression profile of cytokines i.e., IL-2, IL-4 and IFN-γ in PBMC culture were studied in all the samples. A higher total leucocytes, neutrophil and band cells count along with impaired neutrophil functions i.e., lowered level of production of superoxide and hydrogen peroxide before parturition and during early postpartum period were observed in buffaloes developing PRD. Further, a lower expression of IL-2, IFN-γ and IL-4 mRNA in PBMC culture was observed at calving in buffaloes that subsequently developed PRD at later postpartum. Thus, suppression in neutrophil function and cytokine expression at prepartum to early postpartum period predisposes the buffaloes to develop postpartum reproductive disorders. Hence, monitoring of neutrophils function and cytokine expression profile would be effective to predict certain reproductive disorders at late pregnancy or immediately after parturition in buffaloes. In future, this may be a novel approach for determining suitable management and therapeutic decisions for prevention of commonly occurring

  4. Prevention of in vitro neutrophil-endothelial attachment through shedding of L-selectin by nonsteroidal antiinflammatory drugs.

    PubMed Central

    Díaz-González, F; González-Alvaro, I; Campanero, M R; Mollinedo, F; del Pozo, M A; Muñoz, C; Pivel, J P; Sánchez-Madrid, F

    1995-01-01

    The activation of the endothelial cells by extravascular stimuli is the key event in the extravasation of circulating leukocytes to target tissues. L-selectin, a member of the selectin family, is constitutively expressed by white cells, and is the molecule involved in the initial binding of leukocytes to activated endothelium. After activation, leukocytes rapidly release L-selectin from the cell surface, suggesting that the functional activity of this molecule is controlled in large part by its appearance and disappearance from cell surface. We have studied in a neutrophil-activated endothelial cell binding assay, the effect of different antiinflammatory drugs (steroidal and nonsteroidal) in the L-selectin-mediated interaction of neutrophils with activated endothelial cells. Some nonsteroidal antiinflammatory drugs (NSAIDs), such as indomethacin, diclofenac, ketoprofen, and aspirin, but not steroids, strongly inhibited the neutrophil-endothelial cell attachment. Furthermore, we also investigated the underlying mechanism of this functional effect. The expression of L-selectin on the neutrophil surface rapidly decreased in the presence of different NSAIDs, in a dose- and time-dependent manner, whereas no changes in the expression of other adhesion molecules such as CD11a, CD11b, CD31, or ICAM-3 (CD50) were observed. Interestingly, studies in vivo on healthy volunteers treated with physiological doses of indomethacin showed a significant decrease of L-selectin neutrophil expression. Only diclofenac induced an upregulation of CD11b expression, suggesting an activating effect on neutrophils. No enzyme release was observed upon treatment of neutrophils with different NSAIDs, indicating a lack of degranulatory activity of NSAIDs, with the exception of diclofenac. The downregulation of L-selectin expression was due to the rapid cleavage and shedding of the membrane L-selectin, as determined by both immunoprecipitation from 125I-labeled neutrophils, and quantitative

  5. The role of neutrophils and monocytic cells in controlling the initiation of Clostridium perfringens gas gangrene.

    PubMed

    O'Brien, David K; Therit, Blair H; Woodman, Michael E; Melville, Stephen B

    2007-06-01

    Clostridium perfringens is a common cause of the fatal disease gas gangrene (myonecrosis). Established gas gangrene is notable for a profound absence of neutrophils and monocytic cells (phagocytes), and it has been suggested that the bactericidal activities of these cells play an insignificant role in controlling the progression of the infection. However, large inocula of bacteria are needed to establish an infection in experimental animals, suggesting phagocytes may play a role in inhibiting the initiation of gangrene. Examination of tissue sections of mice infected with a lethal (1 x 10(9)) or sublethal (1 x 10(6)) inoculum of C. perfringens revealed that phagocyte infiltration in the first 3 h postinfection was inhibited with a lethal dose but not with a sublethal dose, indicating that exclusion of phagocytes begins very early in the infection cycle. Experiments in which mice were depleted of either circulating monocytes or neutrophils before infection with C. perfringens showed that monocytes play a role in inhibiting the onset of gas gangrene at intermediate inocula but, although neutrophils can slow the onset of the infection, they are not protective. These results suggest that treatments designed to increase monocyte infiltration and activate macrophages may lead to increased resistance to the initiation of gas gangrene.

  6. β2 integrin mediates hantavirus-induced release of neutrophil extracellular traps.

    PubMed

    Raftery, Martin J; Lalwani, Pritesh; Krautkrӓmer, Ellen; Peters, Thorsten; Scharffetter-Kochanek, Karin; Krüger, Renate; Hofmann, Jörg; Seeger, Karl; Krüger, Detlev H; Schönrich, Günther

    2014-06-30

    Rodent-borne hantaviruses are emerging human pathogens that cause severe human disease. The underlying mechanisms are not well understood, as hantaviruses replicate in endothelial and epithelial cells without causing any cytopathic effect. We demonstrate that hantaviruses strongly stimulated neutrophils to release neutrophil extracellular traps (NETs). Hantavirus infection induced high systemic levels of circulating NETs in patients and this systemic NET overflow was accompanied by production of autoantibodies to nuclear antigens. Analysis of the responsible mechanism using neutrophils from β2 null mice identified β2 integrin receptors as a master switch for NET induction. Further experiments suggested that β2 integrin receptors such as complement receptor 3 (CR3) and 4 (CR4) may act as novel hantavirus entry receptors. Using adenoviruses, we confirmed that viral interaction with β2 integrin induced strong NET formation. Collectively, β2 integrin-mediated systemic NET overflow is a novel viral mechanism of immunopathology that may be responsible for characteristic aspects of hantavirus-associated disease such as kidney and lung damage.

  7. Depletion of Neutrophils Exacerbates the Early Inflammatory Immune Response in Lungs of Mice Infected with Paracoccidioides brasiliensis

    PubMed Central

    Lopera, Damaris; Urán-Jiménez, Martha Eugenia

    2016-01-01

    Neutrophils predominate during the acute phase of the Paracoccidioides brasiliensis infection. Herein, we determined the role of the neutrophil during the early stages of experimental pulmonary paracoccidioidomycosis using a monoclonal antibody (mAb) specific for neutrophils. Male BALB/c mice were inoculated intranasally with 1.5 × 106 or 2 × 106 P. brasiliensis yeast cells. The mAb was administered 24 h before infection, followed by doses every 48 h until mice were sacrificed. Survival time was evaluated and mice were sacrificed at 48 h and 96 h after inoculation to assess cellularity, fungal load, cytokine/chemokine levels, and histopathological analysis. Neutrophils from mAb-treated mice were efficiently depleted (99.04%). Eighty percent of the mice treated with the mAb and infected with 1.5 × 106 yeast cells died during the first two weeks after infection. When mice were treated and infected with 2 × 106 yeast cells, 100% of them succumbed by the first week after infection. During the acute inflammatory response significant increases in numbers of eosinophils, fungal load and levels of proinflammatory cytokines/chemokines were observed in the mAb-treated mice. We also confirmed that neutrophils are an important source of IFN-γ and IL-17. These results indicate that neutrophils are essential for protection as well as being important for regulating the early inflammatory immune response in experimental pulmonary paracoccidioidomycosis. PMID:27642235

  8. NOD2 dependent neutrophil recruitment is required for early protective immune responses against infectious Litomosoides sigmodontis L3 larvae

    PubMed Central

    Ajendra, Jesuthas; Specht, Sabine; Ziewer, Sebastian; Schiefer, Andrea; Pfarr, Kenneth; Parčina, Marijo; Kufer, Thomas A.; Hoerauf, Achim; Hübner, Marc P.

    2016-01-01

    Nucleotide-binding oligomerization domain-containing protein 2 (NOD2) recognizes muramyl dipeptide (MDP) of bacterial cell walls, triggering NFκB-induced pro-inflammation. As most human pathogenic filariae contain Wolbachia endobacteria that synthesize the MDP-containing cell wall precursor lipid II, NOD2’s role during infection with the rodent filaria Litomosoides sigmodontis was investigated. In NFκB reporter-cells, worm-extract containing Wolbachia induced NOD2 and NOD1. NOD2-deficient mice infected with L. sigmodontis had significantly more worms than wildtype controls early in infection. Increased worm burden was not observed after subcutaneous infection, suggesting that protective NOD2-dependent immune responses occur within the skin. Flow cytometry demonstrated that neutrophil recruitment to the skin was impaired in NOD2−/− mice after intradermal injection of third stage larvae (L3), and blood neutrophil numbers were reduced after L. sigmodontis infection. PCR array supported the requirement of NOD2 for recruitment of neutrophils to the skin, as genes associated with neutrophil recruitment and activation were downregulated in NOD2−/− mice after intradermal L3 injection. Neutrophil depletion before L. sigmodontis infection increased worm recovery in wildtype mice, confirming that neutrophils are essential against invading L3 larvae. This study indicates that NOD-like receptors are implemented in first-line protective immune responses against filarial nematodes. PMID:28004792

  9. Neutrophil recruitment is critical for 5-fluorouracil-induced diarrhea and the decrease in aquaporins in the colon.

    PubMed

    Sakai, Hiroyasu; Sagara, Atsunobu; Matsumoto, Kenjiro; Jo, Ara; Hirosaki, Akiko; Takase, Kazuhide; Sugiyama, Ryoto; Sato, Ken; Ikegami, Daigo; Horie, Syunji; Matoba, Motohiro; Narita, Minoru

    2014-09-01

    Diarrhea is a common side effect experienced by cancer patients undergoing clinical chemotherapy, such as with 5-fluorouracil (5-FU). However, the precise mechanisms underlying 5-FU-induced diarrhea remain unclear. In the present study, we examined the role of neutrophil in 5-FU-induced diarrhea. Mice were given 5-FU (50mg/kg, i.p.) daily for 4 days. Sivelestat sodium (100 or 300 mg/kg, i.p., neutorophil elastase inhibitor) or SB225002 (3 or 9 mg/kg, i.p., CXCR2 antagonist) was administered before the administration of 5-FU. Gene expression levels of aquaporin (AQP) 4 and 8, CXCL1, CXCL2, CXCL3, neutrophil elastase (Elane) and myeloperoxidase (MPO) in the colon were examined by real-time RT-PCR. The neutrophil (Ly-6G positive cell) number in the mucosa of colon was measured by flow-cytometric analysis. Administration of 5-FU induced diarrhea and decreased the expression levels of AQP 4 and 8 in the colon. Under the present conditions, the expression levels of CXCL1, CXCL2, CXCL3, the neutrophil markers Elane and MPO, as well as Ly-6G-positive neutrophils, in the colon were significantly increased by 5-FU. Neutrophil recruitment with decreased levels of AQP 4 and 8 were dramatically inhibited by either sivelestat sodium or SB225002. Furthermore, these reagents reduced the 5-FU-induced body weight loss and diarrhea. These findings provide evidence that neutrophil recruitment and neutrophil elastase may decrease the levels of AQP 4 and 8 in the colon of mice treated with 5-FU and contribute to the pathophysiology of 5-FU-induced body weight loss and diarrhea.

  10. Peripheral and intrauterine neutrophil function in the cow: the influence of endogenous and exogenous sex steroid hormones.

    PubMed

    Subandrio, A L; Sheldon, I M; Noakes, D E

    2000-05-01

    It has been accepted for many years that the susceptibility of the genital tract to infection is reduced during the follicular phase compared with the luteal phase of the estrous cycle. Since the role of intrauterine neutrophils is paramount in the elimination of bacteria, it can be hypothesized that these differences in resistance to infection could be mediated by differences in uterine-derived neutrophil function. In order to test this hypothesis two groups of cows were used in this study. Group 1 cows (n=5) were studied at estrus, diestrus, after ovariectomy, after exogenous estradiol and after progesterone treatment, at which time they underwent intrauterine infusion with 1% oyster glycogen (OG) and a bacterial-free filtrate (BFF) of Actinomyces genes (BFF), the latter having been recovered from a clinical case of endometritis; neutrophils were harvested by flushing from the lumen 15 to 18 h later. A peripheral blood sample was collected at the time of flushing for the assay of estradiol and progesterone for a WBC and differential count and for the harvesting of neutrophils using a Percoll single-stage discontinuous gradient. After the recovery of the cells they were re-suspended in HBSS. Group 2 (n=4) were infused with BFF during during all reproductive states as Group 1, but with OG only after ovariectomy and after treatment with progesterone and estradiol. Neutrophil chemotaxis was assessed by measuring their migration using a modified Boyden chamber and Zymogen-activated serum as a chemoattractant. Phagocytic activity was measured by determining the number of Candida albicans ingested by each neutrophil after incubation. The percentage of kill was determined using a radiometric assay in which C. albicans was labeled with L-(5-3H) Proline. Peripheral WBC concentration was not influenced by the reproductive state of the cow; however, the mean neutrophil concentration was significantly different between the reproductive states (P<0.001) and between individual

  11. Circulating tumor cells

    PubMed Central

    Raimondi, Cristina; Nicolazzo, Chiara; Gradilone, Angela; Giannini, Giuseppe; De Falco, Elena; Chimenti, Isotta; Varriale, Elisa; Hauch, Siegfried; Plappert, Linda; Cortesi, Enrico; Gazzaniga, Paola

    2014-01-01

    The hypothesis of the “liquid biopsy” using circulating tumor cells (CTCs) emerged as a minimally invasive alternative to traditional tissue biopsy to determine cancer therapy. Discordance for biomarkers expression between primary tumor tissue and circulating tumor cells (CTCs) has been widely reported, thus rendering the biological characterization of CTCs an attractive tool for biomarkers assessment and treatment selection. Studies performed in metastatic colorectal cancer (mCRC) patients using CellSearch, the only FDA-cleared test for CTCs assessment, demonstrated a much lower yield of CTCs in this tumor type compared with breast and prostate cancer, both at baseline and during the course of treatment. Thus, although attractive, the possibility to use CTCs as therapy-related biomarker for colorectal cancer patients is still limited by a number of technical issues mainly due to the low sensitivity of the CellSearch method. In the present study we found a significant discordance between CellSearch and AdnaTest in the detection of CTCs from mCRC patients. We then investigated KRAS pathway activating mutations in CTCs and determined the degree of heterogeneity for KRAS oncogenic mutations between CTCs and tumor tissues. Whether KRAS gene amplification may represent an alternative pathway responsible for KRAS activation was further explored. KRAS gene amplification emerged as a functionally equivalent and mutually exclusive mechanism of KRAS pathway activation in CTCs, possibly related to transcriptional activation. The serial assessment of CTCs may represent an early biomarker of treatment response, able to overcome the intrinsic limit of current molecular biomarkers represented by intratumor heterogeneity. PMID:24521660

  12. Photoimmunological properties of borage in bovine neutrophil in vitro model.

    PubMed

    Asadollahi, Firouzeh; Mehrzad, Jalil; Chaichi, Mohammad Javad; Taghavi Razavizadeh, Alireza

    2015-10-01

    Borage (Echium amoenum fisch) is one of the most commonly used medicinal plants, and has long been used as a traditional herbal medicine for many (non)infectious diseases in Iran. Study on photoredox and photoimmunology of borage is little. Natural immunomodulatory plants with minimal adverse/toxic effects could help boost animal health and, ultimately, public health. To determine the effect of borage on the functions of key circulating innate immune cells, effects of borage extract (BE) on bovine neutrophils (PMN) photoredox and phagocytosis events were evaluated using an in vitro model system. Blood PMN isolated from healthy high yielding dairy cows (n = 8/treatment) were pre-incubated with BE and the impact on phagocytosis-dependent-and-independent cellular chemiluminescence (CL), phagocytosis, killing of Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli), fluorescence-based PMN H2O2 production and necrosis were assessed. Relative to control (no BE) PMN, treatment with BE significantly increased phagocytosis-dependent-and-independent PMN CL (>10-15% increase). While BE also led to increased PMN H2O2 production, necrosis was also surprisingly higher in these cells. Phagocytosis and killing of both E. coli and S. aureus by PMN treated with BE was substantially higher than that by control PMN. The increased photoimmunobiological events especially intracellular CL, intracellular H2O2 formation, and phagocytic capacity of BE-treated PMN support the potential immunotherapeutic implications of borage and its components for particularly immunocompromised animals and humans.

  13. Neutrophil myeloperoxidase destruction by ultraviolet irradiation

    SciTech Connect

    Hanker, J.; Giammara, B.; Strauss, G.

    1988-01-01

    The peroxidase activity of enriched leukocyte preparations on coverslips was determined cytochemically with a newly developed method. The techniques utilizes diaminobenzidine medium and cupric nitrate intensification and is suitable for analysis with light microscopy, SEM, and TEM. Blood specimens from control individuals were studied with and without in vitro UV irradiation and compared with those from psoriasis patients exposed therapeutically to various types of UV in phototherapy. All UV irradiated samples showed diminished neutrophil myeloperoxidase (MP) activity although that of the principal eosinophil peroxidase was unaffected. The SEMs supported the contention that decreased neutrophil MP activity might be related to UV induced degranulation. It is believed to be possible, eventually, to equate the observed MP degranulation effect after UV irradiation with diminished ability to fight bacterial infections.

  14. Neutrophil extracellular traps in tissue pathology.

    PubMed

    Nakazawa, Daigo; Kumar, Santosh; Desai, Jyaysi; Anders, Hans-Joachim

    2017-03-01

    Neutrophil extracellular traps (NETs) are innate immune systems against invading pathogens. NETs are characterized as released DNA mixed with cytoplasmic antimicrobial proteins such as myeloperoxidase, proteinase3 and neutrophil elastase. While NETs are thought to have an important role in host defense, recent work has suggested that NETs contribute to tissue injury in non-infectious disease states. Uncontrolled NET formation in autoimmune diseases, metabolic disorders, cancers and thrombotic diseases can exacerbate a disease or even be a major initiator of tissue injury. But spotting NETs in tissues is not easy. Here we review the available histopathological evidence on the presence of NETs in a variety of diseases. We discuss technical difficulties and potential sources of misinterpretation while trying to detect NETs in tissue samples.

  15. Neutrophil adherence induced by lipopolysaccharide in vitro. Role of plasma component interaction with lipopolysaccharide.

    PubMed Central

    Worthen, G S; Avdi, N; Vukajlovich, S; Tobias, P S

    1992-01-01

    Endotoxemia results in neutrophil localization within a number of microcirculatory beds, reflecting in part an adhesive interaction between neutrophils and the vascular endothelial cell. In previous studies, endotoxin or lipopolysaccharide (LPS) treatment of rabbits resulted in neutrophil sequestration at LPS concentrations well below those effective at increasing neutrophil adherence in vitro. We hypothesized that LPS-induced neutrophil adherence involved a plasma component. In the absence of plasma, high concentrations of LPS (10 micrograms/ml) were required to increase human neutrophil adherence to endothelial cells in vitro. With the inclusion of as little as 1% plasma or serum, however, the LPS dose-response curve was markedly shifted, resulting in increments in adherence at 10 ng/ml, and the time course of enhanced adherence was accelerated. Pretreatment studies suggested that the effect of LPS was on the neutrophil rather than the endothelial cell. Immunoprecipitation of 0111:B4 LPS paralleled the loss of functional activity, suggesting that LPS was an integral part of the active complex, rather than altering a plasma component to make it active. The incubation of plasma with LPS decreased the apparent molecular mass of LPS from 500-1,000 kD to approximately 100 kD. The disaggregated 0111:B4 LPS eluted in the range of albumin and was able to increase adherence in the absence of additional plasma. Plasma depleted of lipoproteins or heat treated retained activity, suggesting that the interaction of LPS with HDL or complement did not account for the observed findings. An LPS-binding protein isolated from rabbit serum enhanced the adherence-inducing effects of both 0111:B4 and Re595 LPS. Furthermore, the activity of rabbit serum was abolished after incubation with an antibody directed against this LPS-binding protein (LBP). An antibody directed against CD14, the putative receptor of the LPS-LBP complex, prevented the adhesive response to LPS. These data suggest

  16. In Vitro Evaluation of the Link Between Cell Activation State and Its Rheological Impact on the Microscale Flow of Neutrophil Suspensions.

    PubMed

    Akenhead, Michael L; Horrall, Nolan M; Rowe, Dylan; Sethu, Palaniappan; Shin, Hainsworth Y

    2015-09-01

    Activated neutrophils have been reported to affect peripheral resistance, for example, by plugging capillaries or adhering to the microvasculature. In vivo and ex vivo data indicate that activated neutrophils circulating in the blood also influence peripheral resistance. We used viscometry and microvascular mimics for in vitro corroboration. The rheological impact of differentiated neutrophil-like HL-60 promyelocytes (dHL60s) or human neutrophil suspensions stimulated with 10 nM fMet-Leu-Phe (fMLP) was quantified using a cone-plate rheometer (450 s(-1) shear rate). To evaluate their impact on microscale flow resistance, we used 10-μm Isopore® membranes to model capillaries as well as single 200 × 50 μm microchannels and networks of twenty 20 × 50 μm microfluidic channels to mimic noncapillary microvasculature. Stimulation of dHL60 and neutrophil populations significantly altered their flow behavior as evidenced by their impact on suspension viscosity. Notably, hematocrit abrogated the impact of leukocyte activation on blood cell suspension viscosity. In micropore filters, activated cell suspensions enhanced flow resistance. This effect was further enhanced by the presence of erythrocytes. The resistance of our noncapillary microvascular mimics to flow of activated neutrophil suspensions was significantly increased only with hematocrit. Notably, it was elevated to a higher extent within the micronetwork chambers compared to the single-channel chambers. Collectively, our findings provide supportive evidence that activated neutrophils passing through the microcirculation may alter hemodynamic resistance due to their altered rheology in the noncapillary microvasculature. This effect is another way neutrophil activation due to chronic inflammation may, at least in part, contribute to the elevated hemodynamic resistance associated with cardiovascular diseases (e.g., hypertension and hypercholesterolemia).

  17. In Vitro Evaluation of the Link Between Cell Activation State and Its Rheological Impact on the Microscale Flow of Neutrophil Suspensions

    PubMed Central

    Akenhead, Michael L.; Horrall, Nolan M.; Rowe, Dylan; Sethu, Palaniappan; Shin, Hainsworth Y.

    2015-01-01

    Activated neutrophils have been reported to affect peripheral resistance, for example, by plugging capillaries or adhering to the microvasculature. In vivo and ex vivo data indicate that activated neutrophils circulating in the blood also influence peripheral resistance. We used viscometry and microvascular mimics for in vitro corroboration. The rheological impact of differentiated neutrophil-like HL-60 promyelocytes (dHL60s) or human neutrophil suspensions stimulated with 10 nM fMet-Leu-Phe (fMLP) was quantified using a cone-plate rheometer (450 s−1 shear rate). To evaluate their impact on microscale flow resistance, we used 10-μm Isopore® membranes to model capillaries as well as single 200 × 50 μm microchannels and networks of twenty 20 × 50 μm microfluidic channels to mimic noncapillary microvasculature. Stimulation of dHL60 and neutrophil populations significantly altered their flow behavior as evidenced by their impact on suspension viscosity. Notably, hematocrit abrogated the impact of leukocyte activation on blood cell suspension viscosity. In micropore filters, activated cell suspensions enhanced flow resistance. This effect was further enhanced by the presence of erythrocytes. The resistance of our noncapillary microvascular mimics to flow of activated neutrophil suspensions was significantly increased only with hematocrit. Notably, it was elevated to a higher extent within the micronetwork chambers compared to the single-channel chambers. Collectively, our findings provide supportive evidence that activated neutrophils passing through the microcirculation may alter hemodynamic resistance due to their altered rheology in the noncapillary microvasculature. This effect is another way neutrophil activation due to chronic inflammation may, at least in part, contribute to the elevated hemodynamic resistance associated with cardiovascular diseases (e.g., hypertension and hypercholesterolemia). PMID:26065495

  18. Leukotriene B4-Neutrophil Elastase Axis Drives Neutrophil Reverse Transendothelial Cell Migration In Vivo

    PubMed Central

    Colom, Bartomeu; Bodkin, Jennifer V.; Beyrau, Martina; Woodfin, Abigail; Ody, Christiane; Rourke, Claire; Chavakis, Triantafyllos; Brohi, Karim; Imhof, Beat A.; Nourshargh, Sussan

    2015-01-01

    Summary Breaching endothelial cells (ECs) is a decisive step in the migration of leukocytes from the vascular lumen to the extravascular tissue, but fundamental aspects of this response remain largely unknown. We have previously shown that neutrophils can exhibit abluminal-to-luminal migration through EC junctions within mouse cremasteric venules and that this response is elicited following reduced expression and/or functionality of the EC junctional adhesion molecule-C (JAM-C). Here we demonstrate that the lipid chemoattractant leukotriene B4 (LTB4) was efficacious at causing loss of venular JAM-C and promoting neutrophil reverse transendothelial cell migration (rTEM) in vivo. Local proteolytic cleavage of EC JAM-C by neutrophil elastase (NE) drove this cascade of events as supported by presentation of NE to JAM-C via the neutrophil adhesion molecule Mac-1. The results identify local LTB4-NE axis as a promoter of neutrophil rTEM and provide evidence that this pathway can propagate a local sterile inflammatory response to become systemic. PMID:26047922

  19. Leukotriene B4-Neutrophil Elastase Axis Drives Neutrophil Reverse Transendothelial Cell Migration In Vivo.

    PubMed

    Colom, Bartomeu; Bodkin, Jennifer V; Beyrau, Martina; Woodfin, Abigail; Ody, Christiane; Rourke, Claire; Chavakis, Triantafyllos; Brohi, Karim; Imhof, Beat A; Nourshargh, Sussan

    2015-06-16

    Breaching endothelial cells (ECs) is a decisive step in the migration of leukocytes from the vascular lumen to the extravascular tissue, but fundamental aspects of this response remain largely unknown. We have previously shown that neutrophils can exhibit abluminal-to-luminal migration through EC junctions within mouse cremasteric venules and that this response is elicited following reduced expression and/or functionality of the EC junctional adhesion molecule-C (JAM-C). Here we demonstrate that the lipid chemoattractant leukotriene B4 (LTB4) was efficacious at causing loss of venular JAM-C and promoting neutrophil reverse transendothelial cell migration (rTEM) in vivo. Local proteolytic cleavage of EC JAM-C by neutrophil elastase (NE) drove this cascade of events as supported by presentation of NE to JAM-C via the neutrophil adhesion molecule Mac-1. The results identify local LTB4-NE axis as a promoter of neutrophil rTEM and provide evidence that this pathway can propagate a local sterile inflammatory response to become systemic.

  20. Autophagy is induced by anti-neutrophil cytoplasmic Abs and promotes neutrophil extracellular traps formation.

    PubMed

    Sha, Li-Li; Wang, Huan; Wang, Chen; Peng, Hong-Ying; Chen, Min; Zhao, Ming-Hui

    2016-11-01

    Dysregulated neutrophil extracellular traps (NETs) formation contributes to the pathogenesis of anti-neutrophil cytoplasmic Ab (ANCA)-associated vasculitis (AAV). Increasing evidence indicates that autophagy is involved in the process of NETs formation. In this study, we aimed to investigate whether ANCA could induce autophagy in the process of NETs formation. Autophagy was detected using live cell imaging, microtubule-associated protein light chain 3B (LC3B) accumulation and Western blotting. The results showed that autophagy vacuolization was detected in neutrophils treated with ANCA-positive IgG by live cell imaging. This effect was enhanced by rapamycin, the autophagy inducer, and weakened by 3-methyladenine (3-MA), the autophagy inhibitor. In line with these results, the autophagy marker, LC3B, showed a punctate distribution pattern in the neutrophils stimulated with ANCA-positive IgG. In the presence of rapamycin, LC3B accumulation was further increased; however, this effect was attenuated by 3-MA. Moreover, incubated with ANCA-positive IgG, the NETosis rate significantly increased compared with the unstimulated group. And, the rate significantly increased or decreased in the neutrophils pretreated with rapamycin or 3-MA, respectively, as compared with the cells incubated with ANCA-positive IgG. Overall, this study demonstrates that autophagy is induced by ANCA and promotes ANCA-induced NETs formation.

  1. Lupus Erythematosus and Neutrophilic Urticarial Dermatosis

    PubMed Central

    Gusdorf, Laurence; Bessis, Didier; Lipsker, Dan

    2014-01-01

    Abstract Neutrophilic urticarial dermatosis (NUD) resembles urticaria clinically but is a neutrophilic dermatosis histopathologically. The majority of patients with NUD have an underlying systemic condition, mainly, autoinflammatory disorders such as cryopyrin-associated periodic syndromes, Schnitzler syndrome, and adult-onset Still disease, but a few also have systemic lupus erythematosus (LE). Here, we confirm these data and we report relevant clinical and histopathological data of 7 patients with LE and NUD. We retrospectively retrieved the medical records of all patients with LE in whom skin biopsy showed NUD in registers of Strasbourg and Montpellier University hospitals since 2000. All were female and aged between 13 and 45 years. Skin lesions were typically rose or red macules or slightly elevated papules occurring in a wide distribution. Individual lesions resolved within 24 hours and were not or only slightly itchy. Every patient had associated signs, most of the time polyarthritis and/or fever. NUD was the presenting mode of LE in 2 patients. NUD was misdiagnosed as a classic lupus flare and led to therapeutic intensification with the introduction of immunosuppressive drugs in 4 patients. Histopathological findings consisted of intense neutrophilic interstitial and perivascular infiltrate with leukocytoclasia and without fibrinoid necrosis of vessel walls. Direct immunofluorescence testing showed a lupus band in 4 patients. Antinuclear antibodies were always positive, anti-dsDNA antibodies were positive in 5 patients, and anti-Ro/SSA antibodies in 6 patients. Immunosuppressive drugs such as prednisone, hydroxychloroquine, mycophenolate mofetil, and methotrexate were never effective to treat NUD. Antihistamines were effective in 1 patient and dapsone or colchicine was effective in 5 patients. NUD is not exceptional in patients with systemic LE and is easily misdiagnosed as an acute LE flare. Furthermore, we show that conventional immunosuppressive LE

  2. Treadmill exercise induces neutrophil recruitment into muscle tissue in a reactive oxygen species-dependent manner. An intravital microscopy study.

    PubMed

    Nunes-Silva, Albená; Bernardes, Priscila T T; Rezende, Bárbara M; Lopes, Fernando; Gomes, Elisa C; Marques, Pedro E; Lima, Paulo M A; Coimbra, Cândido C; Menezes, Gustavo B; Teixeira, Mauro M; Pinho, Vanessa

    2014-01-01

    Intense exercise is a physiological stress capable of inducing the interaction of neutrophils with muscle endothelial cells and their transmigration into tissue. Mechanisms driving this physiological inflammatory response are not known. Here, we investigate whether production of reactive oxygen species is relevant for neutrophil interaction with endothelial cells and recruitment into the quadriceps muscle in mice subjected to the treadmill fatiguing exercise protocol. Mice exercised until fatigue by running for 56.3±6.8 min on an electric treadmill. Skeletal muscle was evaluated by intravital microscopy at different time points after exercise, and then removed to assess local oxidative stress and histopathological analysis. We observed an increase in plasma lactate and creatine kinase (CK) concentrations after exercise. The numbers of monocytes, neutrophils, and lymphocytes in blood increased 12 and 24 hours after the exercise. Numbers of rolling and adherent leukocytes increased 3, 6, 12, and 24 hours post-exercise, as assessed by intravital microscopy. Using LysM-eGFP mice and confocal intravital microscopy technology, we show that the number of transmigrating neutrophils increased 12 hours post-exercise. Mutant gp91phox-/- (non-functional NADPH oxidase) mice and mice treated with apocynin showed diminished neutrophil recruitment. SOD treatment promoted further adhesion and transmigration of leukocytes 12 hours after the exercise. These findings confirm our hypothesis that treadmill exercise increases the recruitment of leukocytes to the postcapillary venules, and NADPH oxidase-induced ROS plays an important role in this process.

  3. Treadmill Exercise Induces Neutrophil Recruitment into Muscle Tissue in a Reactive Oxygen Species-Dependent Manner. An Intravital Microscopy Study

    PubMed Central

    Nunes-Silva, Albená; Bernardes, Priscila T. T.; Rezende, Bárbara M.; Lopes, Fernando; Gomes, Elisa C.; Marques, Pedro E.; Lima, Paulo M. A.; Coimbra, Cândido C.; Menezes, Gustavo B.; Teixeira, Mauro M.; Pinho, Vanessa

    2014-01-01

    Intense exercise is a physiological stress capable of inducing the interaction of neutrophils with muscle endothelial cells and their transmigration into tissue. Mechanisms driving this physiological inflammatory response are not known. Here, we investigate whether production of reactive oxygen species is relevant for neutrophil interaction with endothelial cells and recruitment into the quadriceps muscle in mice subjected to the treadmill fatiguing exercise protocol. Mice exercised until fatigue by running for 56.3±6.8 min on an electric treadmill. Skeletal muscle was evaluated by intravital microscopy at different time points after exercise, and then removed to assess local oxidative stress and histopathological analysis. We observed an increase in plasma lactate and creatine kinase (CK) concentrations after exercise. The numbers of monocytes, neutrophils, and lymphocytes in blood increased 12 and 24 hours after the exercise. Numbers of rolling and adherent leukocytes increased 3, 6, 12, and 24 hours post-exercise, as assessed by intravital microscopy. Using LysM-eGFP mice and confocal intravital microscopy technology, we show that the number of transmigrating neutrophils increased 12 hours post-exercise. Mutant gp91phox-/- (non-functional NADPH oxidase) mice and mice treated with apocynin showed diminished neutrophil recruitment. SOD treatment promoted further adhesion and transmigration of leukocytes 12 hours after the exercise. These findings confirm our hypothesis that treadmill exercise increases the recruitment of leukocytes to the postcapillary venules, and NADPH oxidase-induced ROS plays an important role in this process. PMID:24798414

  4. Role of neutrophilic inflammation in ozone-induced epithelial alterations in the nasal airways of rats

    NASA Astrophysics Data System (ADS)

    Cho, Hye Youn

    Ozone is a principal oxidant air pollutant in photochemical smog. Epithelial cells lining the centriacinar region of lung and the proximal aspects of nasal passage are primary target sites for ozone-induced injury in laboratory animals. Acute exposure of rats to high ambient concentrations of ozone (e.g., 0.5 ppm) results in neutrophilic inflammation, epithelial hyperplasia and mucous cell metaplasia (MCM) in the nasal transitional epithelium (NTE) lining the proximal nasal airways. The principal purpose of the present study was to investigate the role of pre-metaplastic cellular responses, especially neutrophilic inflammation, in the pathogenesis of ozone-induced MCM in rat NTE. For this purpose, three specific hypotheses-based whole-animal inhalation studies were conducted. Male F344/N rats were exposed in whole-body inhalation chambers to 0 (filtered air) or 0.5 ppm ozone for 1-3 days (8 h/day). Histochemical, immunochemical, molecular and morphometric techniques were used to investigate the ozone-induced cellular and molecular events in the NTE. Two in vitro studies were also conducted to examine the effects of ozone-inducible cytokines (i.e., tumor necrosis factor-alpha; TNF- a, and interleukin-6; IL-6) on mucin gene (rMuc-5AC) expression. Ozone induced a rapid increase of rMuc-5AC mRNA in nasal tissues within hours after the start of exposure. It preceded the appearance of MCM, and persisted with MCM. Ozone-induced neutrophilic inflammation accompanied the mucin gene upregulation, but was resolved when MCM first appeared in the NTE. Antibody-mediated depletion of circulating neutrophils attenuated ozone-induced MCM, although it did not affect the ozone-induced epithelial hyperplasia and mucin mRNA upregulation. In another study, it was found that preexisting neutrophilic rhinitis induced by endotoxin augmented the ozone-induced MCM. However, pre-existing rhinitis did not alter the severity of ozone-induced epithelial hyperplasia and mucin gene upregulation

  5. Transendothelial migration enables subsequent transmigration of neutrophils through underlying pericytes.

    PubMed

    Ayres-Sander, Chantal E; Lauridsen, Holly; Maier, Cheryl L; Sava, Parid; Pober, Jordan S; Gonzalez, Anjelica L

    2013-01-01

    During acute inflammation, neutrophil recruitment into extravascular tissue requires neutrophil tethering and rolling on cytokine-activated endothelial cells (ECs), tight adhesion, crawling towards EC junctions and transendothelial migration (TEM). Following TEM, neutrophils must still traverse the subendothelial basement membrane and network of pericytes (PCs). Until recently, the contribution of the PC layer to neutrophil recruitment was largely ignored. Here we analyze human neutrophil interactions with interleukin (IL)-1β-activated human EC monolayers, PC monolayers and EC/PC bilayers in vitro. Compared to EC, PC support much lower levels of neutrophil binding (54.6% vs. 7.1%, respectively) and transmigration (63.7 vs. 8.8%, respectively) despite comparable levels of IL-8 (CXCL8) synthesis and display. Remarkably, EC/PC bilayers support intermediate levels of transmigration (37.7%). Neutrophil adhesion to both cell types is Mac-1-dependent and while ICAM-1 transduction of PCs increases neutrophil adhesion to (41.4%), it does not increase transmigration through PC monolayers. TEM, which increases neutrophil Mac-1 surface expression, concomitantly increases the ability of neutrophils to traverse PCs (19.2%). These data indicate that contributions from both PCs and ECs must be considered in evaluation of microvasculature function in acute inflammation.

  6. Targeting neutrophils in ischemic stroke: translational insights from experimental studies

    PubMed Central

    Jickling, Glen C; Liu, DaZhi; Ander, Bradley P; Stamova, Boryana; Zhan, Xinhua; Sharp, Frank R

    2015-01-01

    Neutrophils have key roles in ischemic brain injury, thrombosis, and atherosclerosis. As such, neutrophils are of great interest as targets to treat and prevent ischemic stroke. After stroke, neutrophils respond rapidly promoting blood–brain barrier disruption, cerebral edema, and brain injury. A surge of neutrophil-derived reactive oxygen species, proteases, and cytokines are released as neutrophils interact with cerebral endothelium. Neutrophils also are linked to the major processes that cause ischemic stroke, thrombosis, and atherosclerosis. Thrombosis is promoted through interactions with platelets, clotting factors, and release of prothrombotic molecules. In atherosclerosis, neutrophils promote plaque formation and rupture by generating oxidized-low density lipoprotein, enhancing monocyte infiltration, and degrading the fibrous cap. In experimental studies targeting neutrophils can improve stroke. However, early human studies have been met with challenges, and suggest that selective targeting of neutrophils may be required. Several properties of neutrophil are beneficial and thus may important to preserve in patients with stroke including antimicrobial, antiinflammatory, and neuroprotective functions. PMID:25806703

  7. Differentiating neutrophils using the optical coulter counter

    NASA Astrophysics Data System (ADS)

    Schonbrun, Ethan; Di Caprio, Giuseppe

    2015-11-01

    We present an optofluidic measurement system that quantifies cell volume, dry mass, and nuclear morphology of neutrophils in high-throughput. While current clinical hematology analyzers can differentiate neutrophils from a blood sample, they do not give other quantitative information beyond their count. In order to better understand the distribution of neutrophil phenotypes in a blood sample, we perform two distinct multivariate measurements. In both measurements, white blood cells are driven through a microfluidic channel and imaged while in flow onto a color camera using a single exposure. In the first measurement, we quantify cell volume, scattering strength, and cell dry mass by combining quantitative phase imaging with dye exclusion cell volumetric imaging. In the second measurement, we quantify cell volume and nuclear morphology using a nucleic acid fluorescent stain. In this way, we can correlate cell volume to other cellular characteristics, which would not be possible using an electrical coulter counter. Unlike phase imaging or cell scattering analysis, the optical coulter counter is capable of quantifying cell volume virtually independent of the cell's refractive index and unlike optical tomography, measurements are possible on quickly flowing cells, enabling high-throughput.

  8. Differentiating neutrophils using the optical coulter counter

    NASA Astrophysics Data System (ADS)

    Schonbrun, E.; Di Caprio, G.

    2015-03-01

    We present an opto-fluidic measurement system that quantifies cell volume, dry mass and nuclear morphology of neutrophils in high-throughput. While current clinical hematology analyzers can differentiate neutrophils from a blood sample, they do not give other quantitative information beyond their count. In order to better understand the distribution of neutrophil phenotypes in a blood sample, we perform two distinct multivariate measurements. In both measurements, white blood cells are driven through a microfluidic channel and imaged while in flow onto a color camera using a single exposure. In the first measurement, we quantify cell volume, scattering strength, and cell dry mass by combining quantitative phase imaging with dye exclusion cell volumetric imaging. In the second measurement, we quantify cell volume and nuclear morphology using a nucleic acid fluorescent stain. In this way, we can correlate cell volume to other cellular characteristics, which would not be possible using an electrical coulter counter. Unlike phase imaging or cell scattering analysis, the optical coulter counter is capable of quantifying cell volume virtually independent of the cell's refractive index and unlike optical tomography, measurements are possible on quickly flowing cells, enabling high-throughput.

  9. Galectin-1 promotes human neutrophil migration.

    PubMed

    Auvynet, Constance; Moreno, Samadhi; Melchy, Erika; Coronado-Martínez, Iris; Montiel, Jose Luis; Aguilar-Delfin, Irma; Rosenstein, Yvonne

    2013-01-01

    An important step of innate immune response is the recruitment of polymorphonuclear leukocytes (PMN) to injured tissues through chemotactic molecules. Galectins, a family of endogenous lectins, participate in numerous functions such as lymphoid cell migration, homing, cell-cell and cell-matrix interactions. Particularly, galectin-3 (Gal-3) and -9 have been implicated in the modulation of acute and chronic inflammation by inducing the directional migration of monocytes/macrophages and eosinophils, whereas Gal-1 is considered to function as an anti-inflammatory molecule, capable of inhibiting the influx of PMN to the site of injury. In this study, we assessed the effect of Gal-1 on neutrophil recruitment, in the absence of additional inflammatory insults. Contrasting with its capacity to inhibit cell trafficking and modulate the release of mediators described in models of acute inflammation and autoimmunity, we evidenced that Gal-1 has the capacity to induce neutrophil migration both in vitro and in vivo. This effect is not mediated through a G-protein-coupled receptor but potentially through the sialoglycoprotein CD43, via carbohydrate binding and through the p38 mitogen-activated protein kinase pathway. These results suggest a novel biological function for CD43 on neutrophils and highlight that depending on the environment, Gal-1 can act either as chemoattractant or, as a molecule that negatively regulates migration under acute inflammatory conditions, underscoring the potential of Gal-1 as a target for innovative drug development.

  10. Neutrophil activator of matrix metalloproteinase-2 (NAM).

    PubMed

    Rollo, Ellen E; Hymowitz, Michelle; Schmidt, Cathleen E; Montana, Steve; Foda, Hussein; Zucker, Stanley

    2006-01-01

    We have isolated a novel soluble factor(s), neutrophil activator of matrix metalloproteinases (NAM), secreted by unstimulated normal human peripheral blood neutrophils that causes the activation of cell secreted promatrix metalloproteinase-2 (proMMP-2). Partially purified preparations of NAM have been isolated from the conditioned media of neutrophils employing gelatin-Sepharose chromatography and differential membrane filter centrifugation. NAM activity, as assessed by exposing primary human umbilical vein endothelial cells (HUVEC) or HT1080 cells to NAM followed by gelatin zymography, was seen within one hour. Tissue inhibitor of metalloproteinase-2 (TIMP-2) and hydroxamic acid derived inhibitors of MMPs (CT1746 and BB94) abrogated the activation of proMMP-2 by NAM, while inhibitors of serine and cysteine proteases showed no effect. NAM also produced an increase in TIMP-2 binding to HUVEC and HT1080 cell surfaces that was inhibited by TIMP-2, CT1746, and BB94. Time-dependent increases in MT1-MMP protein and mRNA were seen following the addition of NAM to cells. These data support a role for NAM in cancer dissemination.

  11. 'Slings' enable neutrophil rolling at high shear.

    PubMed

    Sundd, Prithu; Gutierrez, Edgar; Koltsova, Ekaterina K; Kuwano, Yoshihiro; Fukuda, Satoru; Pospieszalska, Maria K; Groisman, Alex; Ley, Klaus

    2012-08-16

    Most leukocytes can roll along the walls of venules at low shear stress (1 dyn cm−2), but neutrophils have the ability to roll at tenfold higher shear stress in microvessels in vivo. The mechanisms involved in this shear-resistant rolling are known to involve cell flattening and pulling of long membrane tethers at the rear. Here we show that these long tethers do not retract as postulated, but instead persist and appear as 'slings' at the front of rolling cells. We demonstrate slings in a model of acute inflammation in vivo and on P-selectin in vitro, where P-selectin-glycoprotein-ligand-1 (PSGL-1) is found in discrete sticky patches whereas LFA-1 is expressed over the entire length on slings. As neutrophils roll forward, slings wrap around the rolling cells and undergo a step-wise peeling from the P-selectin substrate enabled by the failure of PSGL-1 patches under hydrodynamic forces. The 'step-wise peeling of slings' is distinct from the 'pulling of tethers' reported previously. Each sling effectively lays out a cell-autonomous adhesive substrate in front of neutrophils rolling at high shear stress during inflammation.

  12. Human neutrophil elastase in RSV bronchiolitis.

    PubMed

    Emboriadou, M; Hatzistilianou, Maria; Magnisali, Ch; Sakelaropoulou, A; Exintari, M; Conti, Pio; Aivazis, V

    2007-01-01

    Acute bronchiolitis is the most common lower respiratory tract infection in young children and may be life-threatening in those with underlying cardiac or respiratory conditions. We evaluated the nasal and serum levels of human neutrophil elastase (HNE) in patients with acute respiratory syncytial virus (RSV) bronchiolitis and investigated the correlation of these levels with illness severity. Fifty-one patients (28 boys, 23 girls) with acute bronchiolitis positive for RSV by direct immunoenzyme assay in nasal secretions (Group A) were studied. Thirty healthy children (17 boys, 13 girls) constituted the control group (Group B). Subjects in both groups were matched for age and gender. The ages (mean+/-SE) in Groups A and B were 4.5+/-0.41 and 5.0+/-0.65 mo, respectively. Venous blood and nasal secretions were taken from patients in group A on 1, 5, and 15 days after admission and once from controls (Group B) for determinations of HNE in nasal lavage and serum, as well as white blood counts (WBC). The peripheral blood eosinophil and neutrophil counts were elevated in 22/51 patients (43.1%) and 15/51 patients (29.4%), respectively. In nasal lavage specimens, neutrophils represented>or=75% and eosinophils>2% of all cells in 42/51 (82.0%) patients and 11/51 (21.5%) patients, respectively. There was strong correlation between the level of HNE and the percentage of neutrophils in nasal lavage (r=0.92). The mean nasal HNE concentrations of the patients on 1, 5, and 15 days after admission were higher than those of Group B (p<0.0001, p<0.001, p<0.001, respectively). Mean serum HNE concentrations on 1, 5, and 15 days after admission were higher in Group A than in Group B (p<0.0001, p<0.0001, p<0.0001, respectively). Nasal and serum HNE concentrations showed no correlations with the clinical score of disease severity (r=0.28 and r=0.29, respectively). This study shows that (a) serum and nasal HNE concentrations were significantly higher in RSV bronchiolitis patients than in

  13. Force-dependent calcium signaling and its pathway of human neutrophils on P-selectin in flow.

    PubMed

    Huang, Bing; Ling, Yingchen; Lin, Jiangguo; Du, Xin; Fang, Ying; Wu, Jianhua

    2017-02-01

    P-selectin engagement of P-selectin glycoprotein ligand-1 (PSGL-1) causes circulating leukocytes to roll on and adhere to the vascular surface, and mediates intracellular calcium flux, a key but unclear event for subsequent arresting firmly at and migrating into the infection or injured tissue. Using a parallel plate flow chamber technique and intracellular calcium ion detector (Fluo-4 AM), the intracellular calcium flux of firmly adhered neutrophils on immobilized P-selectin in the absence of chemokines at various wall shear stresses was investigated here in real time by fluorescence microscopy. The results demonstrated that P-selectin engagement of PSGL-1 induced the intracellular calcium flux of firmly adhered neutrophils in flow, increasing P-selectin concentration enhanced cellular calcium signaling, and, force triggered, enhanced and quickened the cytoplasmic calcium bursting of neutrophils on immobilized P-selectin. This P-selectin-induced calcium signaling should come from intracellular calcium release rather than extracellular calcium influx, and be along the mechano-chemical signal pathway involving the cytoskeleton, moesin and Spleen tyrosine kinase (Syk). These results provide a novel insight into the mechano-chemical regulation mechanism for P-selectin-induced calcium signaling of neutrophils in flow.

  14. Circulating Mitochondrial DAMPs Cause Inflammatory Responses to Injury

    PubMed Central

    Zhang, Qin; Raoof, Mustafa; Chen, Yu; Sumi, Yuka; Sursal, Tolga; Junger, Wolfgang; Brohi, Karim; Itagaki, Kiyoshi; Hauser, Carl J.

    2009-01-01

    Injury causes a systemic inflammatory response syndrome (SIRS) clinically much like sepsis 1. Microbial pathogen-associated molecular patterns (PAMPs) activate innate immunocytes through pattern recognition receptors 2. Similarly, cellular injury can release endogenous damage-associated molecular patterns (DAMPs) that activate innate immunity 3. Mitochondria are evolutionary endosymbionts that were derived from bacteria 4 and so might bear bacterial molecular motifs. We show here that injury releases mitochondrial DAMPs (MTD) into the circulation with functionally important immune consequences. MTD include formyl peptides and mitochondrial DNA. These activate human neutrophils (PMN) through formyl peptide receptor-1 and TLR9 respectively. MTD promote PMN Ca2+ flux and phosphorylation of MAP kinases, thus leading to PMN migration and degranulation in vitro and in vivo. Circulating MTD can elicit neutrophil-mediated organ injury. Cellular disruption by trauma releases mitochondrial DAMPs with evolutionarily conserved similarities to bacterial PAMPs into the circulation. These can then signal through identical innate immune pathways to create a sepsis-like state. The release of such mitochondrial ‘enemies within’ by cellular injury is a key link between trauma, inflammation and SIRS. PMID:20203610

  15. Human neutrophil kinetics: modeling of stable isotope labeling data supports short blood neutrophil half-lives

    PubMed Central

    Lahoz-Beneytez, Julio; Elemans, Marjet; Zhang, Yan; Ahmed, Raya; Salam, Arafa; Block, Michael; Niederalt, Christoph; Macallan, Derek

    2016-01-01

    Human neutrophils have traditionally been thought to have a short half-life in blood; estimates vary from 4 to 18 hours. This dogma was recently challenged by stable isotope labeling studies with heavy water, which yielded estimates in excess of 3 days. To investigate this disparity, we generated new stable isotope labeling data in healthy adult subjects using both heavy water (n = 4) and deuterium-labeled glucose (n = 9), a compound with more rapid labeling kinetics. To interpret results, we developed a novel mechanistic model and applied it to previously published (n = 5) and newly generated data. We initially constrained the ratio of the blood neutrophil pool to the marrow precursor pool (ratio = 0.26; from published values). Analysis of heavy water data sets yielded turnover rates consistent with a short blood half-life, but parameters, particularly marrow transit time, were poorly defined. Analysis of glucose-labeling data yielded more precise estimates of half-life (0.79 ± 0.25 days; 19 hours) and marrow transit time (5.80 ± 0.42 days). Substitution of this marrow transit time in the heavy water analysis gave a better-defined blood half-life of 0.77 ± 0.14 days (18.5 hours), close to glucose-derived values. Allowing the ratio of blood neutrophils to mitotic neutrophil precursors (R) to vary yielded a best-fit value of 0.19. Reanalysis of the previously published model and data also revealed the origin of their long estimates for neutrophil half-life: an implicit assumption that R is very large, which is physiologically untenable. We conclude that stable isotope labeling in healthy humans is consistent with a blood neutrophil half-life of less than 1 day. PMID:27136946

  16. Neutrophils in host defense: new insights from zebrafish

    PubMed Central

    Harvie, Elizabeth A.; Huttenlocher, Anna

    2015-01-01

    Neutrophils are highly motile phagocytic cells that play a critical role in the immune response to infection. Zebrafish (Danio rerio) are increasingly used to study neutrophil function and host-pathogen interactions. The generation of transgenic zebrafish lines with fluorescently labeled leukocytes has made it possible to visualize the neutrophil response to infection in real time by use of optically transparent zebrafish larvae. In addition, the genetic tractability of zebrafish has allowed for the generation of models of inherited neutrophil disorders. In this review, we discuss several zebrafish models of infectious disease, both in the context of immunocompetent, as well as neutrophil-deficient hosts and how these models have shed light on neutrophil behavior during infection. PMID:25717145

  17. Perivascular macrophages mediate neutrophil recruitment during bacterial skin infection

    PubMed Central

    Abtin, Arby; Jain, Rohit; Mitchell, Andrew J.; Roediger, Ben; Brzoska, Anthony J.; Tikoo, Shweta; Cheng, Qiang; Ng, Lai Guan; Cavanagh, Lois L.; von Andrian, Ulrich H.; Hickey, Michael J.; Firth, Neville; Weninger, Wolfgang

    2014-01-01

    Transendothelial migration of neutrophils in post-capillary venules is a key event in the inflammatory response against pathogens and tissue damage. The precise regulation of this process is incompletely understood. We report that perivascular macrophages are critical for neutrophil migration into skin infected with the pathogen Staphylococcus aureus. Using multiphoton intravital microscopy we show that neutrophils extravasate from inflamed dermal venules in close proximity to perivascular macrophages, which are a major source of neutrophil chemoattractants. The virulence factor alpha-hemolysin lyses perivascular macrophages leading to decreased neutrophil transmigration. Our data illustrate a previously unrecognized role for perivascular macrophages in neutrophil recruitment to inflamed skin, and indicate that Staphylococcus aureus uses hemolysin-dependent killing of these cells as an immune evasion strategy. PMID:24270515

  18. Neutrophils and Macrophages: the Main Partners of Phagocyte Cell Systems

    PubMed Central

    Silva, Manuel T.; Correia-Neves, Margarida

    2012-01-01

    Biological cellular systems are groups of cells sharing a set of characteristics, mainly key function and origin. Phagocytes are crucial in the host defense against microbial infection. The previously proposed phagocyte cell systems including the most recent and presently prevailing one, the mononuclear phagocyte system (MPS), grouped mononuclear cells but excluded neutrophils, creating an unacceptable situation. As neutrophils are archetypical phagocytes that must be members of comprehensive phagocyte systems, Silva recently proposed the creation of a myeloid phagocyte system (MYPS) that adds neutrophils to the MPS. The phagocytes grouped in the MYPS include the leukocytes neutrophils, inflammatory monocytes, macrophages, and immature myeloid DCs. Here the justifications behind the inclusion of neutrophils in a phagocyte system is expanded and the MYPS are further characterized as a group of dedicated phagocytic cells that function in an interacting and cooperative way in the host defense against microbial infection. Neutrophils and macrophages are considered the main arms of this system. PMID:22783254

  19. Neutrophil function is preserved in a pooled granulocyte component prepared from whole blood donations.

    PubMed

    Bashir, Saber; Stanworth, Simon; Massey, Edwin; Goddard, Fred; Cardigan, Rebecca

    2008-03-01

    Whole blood-derived granulocytes (buffy coats) are issued as an alternative to apheresis donations, but are heavily contaminated with red cells and platelets and there is minimal in vitro data describing their functionality. We developed a purer pooled granulocyte component (PGC) from whole blood donations by pooling 10 ABO-matched buffy coats with 400 ml of platelet additive solution (SSP+) and re-centrifuging. The PGC was irradiated (25-50 Gy) and neutrophil viability, chemotaxis, phagocytosis and respiratory burst activity were determined by flow cytometry. Results from 13 PGC at 16-20 h following donation were compared with those obtained from 20 standard individual buffy coats and with fresh whole blood. The PGC contained similar numbers of neutrophils (approximately 0.9 x 10(10)) with a reduced volume and haemoglobin content when compared with 10 individual buffy coats. Neutrophils in the PGC maintained >90% viability, oxidative burst and phagocytic activity and their ability to migrate towards a chemoattractant 16-20 h following donation, which is similar to results obtained with either fresh whole blood or standard buffy coats. Therefore, neutrophil function in the PGC was preserved 16-20 h following donation, but this product had significantly lower red cell contamination compared with 10 buffy coats, which are currently transfused.

  20. A neutrophil elastase inhibitor prevents bleomycin-induced pulmonary fibrosis in mice.

    PubMed

    Takemasa, Akihiro; Ishii, Yoshiki; Fukuda, Takeshi

    2012-12-01

    Neutrophil elastase plays pivotal roles in the pathogenesis of pulmonary fibrosis. The neutrophil elastase inhibitor, sivelestat, could alleviate pulmonary fibrosis; however, the antifibrotic mechanisms have not yet been clarified. We examined the antifibrotic mechanisms, mainly focusing on a key fibrotic cytokine, transforming growth factor (TGF)-β1, in this study. To elucidate the antifibrotic mechanisms of sivelestat, we examined a murine model of bleomycin-induced early-stage pulmonary fibrosis. After intratracheal instillation of bleomycin, sivelestat was administered intraperitoneally once a day for 7 or 14 days. Bronchoalveolar lavage fluid and lung samples were examined on day 7 or day 14 after bleomycin instillation. In the bleomycin-induced early-stage pulmonary fibrosis model, the neutrophil elastase level was increased in the lungs. Sivelestat significantly inhibited the increase in lung collagen content, fibrotic changes, the numbers of total cells (including macrophages, neutrophils and lymphocytes), the levels of the active form of TGF-β1 and phospho-Smad2 in bleomycin-induced early-stage pulmonary fibrosis. The total TGF-β1 levels and relative changes of TGF-β1 mRNA expression, however, were not decreased significantly by sivelestat. These results suggest that sivelestat alleviated bleomycin-induced pulmonary fibrosis via inhibition of both TGF-β activation and inflammatory cell recruitment in the lung.

  1. Neutrophil activity in chronic venous leg ulcers—A target for therapy?

    PubMed Central

    McDaniel, Jodi C.; Roy, Sashwati; Wilgus, Traci A.

    2013-01-01

    Chronic venous leg ulcers (CVLUs) affect approximately 600,000 people annually in the United States and accrue yearly treatment costs of US$2.5–5 billion. As the population ages, demands on health care resources for CVLU treatments are predicted to drastically increase because the incidence of CVLUs is highest in those ≥65 years of age. Furthermore, regardless of current standards of care, healing complications and high recurrence rates prevail. Thus, it is critical that factors leading to or exacerbating CVLUs be discerned and more effective, adjuvant, evidence-based treatment strategies be utilized. Previous studies have suggested that CVLUs’ pathogenesis is related to the prolonged presence of high numbers of activated neutrophils secreting proteases in the wound bed that destroy growth factors, receptors, and the extracellular matrix that are essential for healing. These events are believed to contribute to a chronically inflamed wound that fails to heal. Therefore, the purpose of this project was to review studies from the past 15 years (1996–2011) that characterized neutrophil activity in the microenvironment of human CVLUs for new evidence that could explicate the proposed relationship between excessive, sustained neutrophil activity and CVLUs. We also appraised the strength of evidence for current and potential therapeutics that target excessive neutrophil activity. PMID:23551462

  2. Technical note: proteomic approaches to fundamental questions about neutrophil biology.

    PubMed

    McLeish, Kenneth R; Merchant, Michael L; Klein, Jon B; Ward, Richard A

    2013-10-01

    Proteomics is one of a group of technologies that generates high-throughput, large-scale datasets that can be used to understand cell or organ functions at a systems level. This review will focus on the application of proteomics to the understanding of neutrophil biology. The strengths and weaknesses of common proteomic methods and their application to neutrophils are reviewed, with the goal of evaluating whether the technology is ready to advance our understanding of neutrophil biology.

  3. Propagation of thrombosis by neutrophils and extracellular nucleosome networks

    PubMed Central

    Pfeiler, Susanne; Stark, Konstantin; Massberg, Steffen; Engelmann, Bernd

    2017-01-01

    Neutrophils, early mediators of the innate immune defense, are recruited to developing thrombi in different types of thrombosis. They amplify intravascular coagulation by stimulating the tissue factor-dependent extrinsic pathway via inactivation of endogenous anticoagulants, enhancing factor XII activation or decreasing plasmin generation. Neutrophil-dependent prothrombotic mechanisms are supported by the externalization of decondensed nucleosomes and granule proteins that together form neutrophil extracellular traps. These traps, either in intact or fragmented form, are causally involved in various forms of experimental thrombosis as first indicated by their role in the enhancement of both microvascular thrombosis during bacterial infection and carotid artery thrombosis. Neutrophil extracellular traps can be induced by interactions of neutrophils with activated platelets; vice versa, these traps enhance adhesion of platelets via von Willebrand factor. Neutrophil-induced microvascular thrombus formation can restrict the dissemination and survival of blood-borne bacteria and thereby sustain intravascular immunity. Dysregulation of this innate immune pathway may support sepsis-associated coagulopathies. Notably, neutrophils and extracellular nucleosomes, together with platelets, critically promote fibrin formation during flow restriction-induced deep vein thrombosis. Neutrophil extracellular traps/extracellular nucleosomes are increased in thrombi and in the blood of patients with different vaso-occlusive pathologies and could be therapeutically targeted for the prevention of thrombosis. Thus, during infections and in response to blood vessel damage, neutrophils and externalized nucleosomes are major promoters of intravascular blood coagulation and thrombosis. PMID:27927771

  4. Regulation of the estrous cycle by neutrophils via opioid peptides.

    PubMed

    Sasaki, Soichiro; Tamaki, Yutaka; Nagata, Kisaburo; Kobayashi, Yoshiro

    2011-07-15

    We found previously that neutrophil-depleted mice exhibited significant blockading of both the regular estrous cycle and cyclic changes of steroid hormone levels. In this study, we aimed at elucidation of the underlying mechanism. To examine the possibility that an increase in bacteria in the vaginal vault of neutrophil-depleted mice causes blockading of the estrous cycle, we treated neutrophil-depleted mice with antibiotics but failed to restore the estrous cycle. We then examined another possibility that neutrophils regulate the estrous cycle via opioid peptides, because opioid peptides regulate steroidogenesis in theca and granulosa cells in the ovaries, and because neutrophils contain opioid peptides. In support of this possibility, naloxone, an opioid antagonist, blocked the estrous cycle and a μ opioid receptor agonist restored the estrous cycle in neutrophil-depleted mice. Pro-opiomelanocortin was immunohistochemically detected in peripheral blood neutrophils but not in ones that had infiltrated into the ovaries. i.v. injection of anti-MIP-2 polyclonal Ab caused blockading of the estrous cycle, whereas MIP-2 was detected in the ovaries, suggesting a role of MIP-2 in the regulation of the estrous cycle. Moreover, i.v. injection of MIP-2 decreased the pro-opiomelanocortin signal in peripheral blood neutrophils and caused blockading of the estrous cycle. Together, these results suggest that neutrophils maintain the estrous cycle via opioid peptides.

  5. Neutrophils: Between Host Defence, Immune Modulation, and Tissue Injury

    PubMed Central

    Kruger, Philipp; Saffarzadeh, Mona; Weber, Alexander N. R.; Rieber, Nikolaus; Radsak, Markus; von Bernuth, Horst; Benarafa, Charaf; Roos, Dirk; Skokowa, Julia; Hartl, Dominik

    2015-01-01

    Neutrophils, the most abundant human immune cells, are rapidly recruited to sites of infection, where they fulfill their life-saving antimicrobial functions. While traditionally regarded as short-lived phagocytes, recent findings on long-term survival, neutrophil extracellular trap (NET) formation, heterogeneity and plasticity, suppressive functions, and tissue injury have expanded our understanding of their diverse role in infection and inflammation. This review summarises our current understanding of neutrophils in host-pathogen interactions and disease involvement, illustrating the versatility and plasticity of the neutrophil, moving between host defence, immune modulation, and tissue damage. PMID:25764063

  6. Exploring inflammatory disease drug effects on neutrophil function.

    PubMed

    Wu, Xiaojie; Kim, Donghyuk; Young, Ashlyn T; Haynes, Christy L

    2014-08-21

    Neutrophils are critical inflammatory cells; thus, it is important to characterize the effects of drugs on neutrophil function in the context of inflammatory diseases. Herein, chemically guided neutrophil migration, known as chemotaxis, is studied in the context of drug treatment at the single cell level using a microfluidic platform, complemented by cell viability assays and calcium imaging. Three representative drugs known to inhibit surface receptor expression, signaling enzyme activity, and the elevation of intracellular Ca(2+) levels, each playing a significant role in neutrophil chemotactic pathways, are used to examine the in vitro drug effects on cellular behaviors. The microfluidic device establishes a stable concentration gradient of chemokines across a cell culture chamber so that neutrophil migration can be monitored under various drug-exposure conditions. Different time- and concentration-dependent regulatory effects were observed by comparing the motility, polarization, and effectiveness of neutrophil chemotaxis in response to the three drugs. Viability assays revealed distinct drug capabilities in reducing neutrophil viability while calcium imaging clarified the role of Ca(2+) in the neutrophil chemotaxis. This study provides mechanistic insight into the drug effects on neutrophil function, facilitating comparison of current and potential pharmaceutical approaches.

  7. Neutrophils and Immunity: From Bactericidal Action to Being Conquered

    PubMed Central

    Teng, Tie-Shan

    2017-01-01

    The neutrophil is the major phagocyte and the final effector cell of the innate immunity, with a primary role in the clearance of extracellular pathogens. Using the broad array of cytokines, extracellular traps, and effector molecules as the humoral arm, neutrophils play a crucial role in the host defense against pathogen infections. On the other hand, the pathogen has the capacity to overcome neutrophil-mediated host defense to establish infection causing human disease. Pathogens, such as S. aureus, have the potential to thwart neutrophil chemotaxis and phagocytosis and thereby succeed in evading killing by neutrophils. Furthermore, S. aureus surviving within neutrophils promotes neutrophil cytolysis, resulting in the release of host-derived molecules that promote local inflammation. Here, we provide a detailed overview of the mechanisms by which neutrophils kill the extracellular pathogens and how pathogens evade neutrophils degradation. This review will provide insights that might be useful for the development of novel therapies against infections caused by antibiotic resistant pathogens. PMID:28299345

  8. Human filarial Wolbachia lipopeptide directly activates human neutrophils in vitro.

    PubMed

    Tamarozzi, F; Wright, H L; Johnston, K L; Edwards, S W; Turner, J D; Taylor, M J

    2014-10-01

    The host inflammatory response to the Onchocerca volvulus endosymbiont, Wolbachia, is a major contributing factor in the development of chronic pathology in humans (onchocerciasis/river blindness). Recently, the toll-like pattern recognition receptor motif of the major inflammatory ligands of filarial Wolbachia, membrane-associated diacylated lipoproteins, was functionally defined in murine models of pathology, including mediation of neutrophil recruitment to the cornea. However, the extent to which human neutrophils can be activated in response to this Wolbachia pattern recognition motif is not known. Therefore, the responses of purified peripheral blood human neutrophils to a synthetic N-terminal diacylated lipopeptide (WoLP) of filarial Wolbachia peptidoglycan-associated lipoprotein (PAL) were characterized. WoLP exposure led to a dose-dependent activation of healthy, human neutrophils that included gross morphological alterations and modulation of surface expressed integrins involved in tethering, rolling and extravasation. WoLP exposure induced chemotaxis but not chemokinesis of neutrophils, and secretion of the major neutrophil chemokine, interleukin 8. WoLP also induced and primed the respiratory burst, and enhanced neutrophil survival by delay of apoptosis. These results indicate that the major inflammatory motif of filarial Wolbachia lipoproteins directly activates human neutrophils in vitro and promotes a molecular pathway by which human neutrophils are recruited to sites of Onchocerca parasitism.

  9. Neutrophilic Skin Lesions in Autoimmune Connective Tissue Diseases

    PubMed Central

    Hau, Estelle; Vignon Pennamen, Marie-Dominique; Battistella, Maxime; Saussine, Anne; Bergis, Maud; Cavelier-Balloy, Benedicte; Janier, Michel; Cordoliani, Florence; Bagot, Martine; Rybojad, Michel; Bouaziz, Jean-David

    2014-01-01

    Abstract The pathophysiology of neutrophilic dermatoses (NDs) and autoimmune connective tissue diseases (AICTDs) is incompletely understood. The association between NDs and AICTDs is rare; recently, however, a distinctive subset of cutaneous lupus erythematosus (LE, the prototypical AICTD) with neutrophilic histological features has been proposed to be included in the spectrum of lupus. The aim of our study was to test the validity of such a classification. We conducted a monocentric retrospective study of 7028 AICTDs patients. Among these 7028 patients, a skin biopsy was performed in 932 cases with mainly neutrophilic infiltrate on histology in 9 cases. Combining our 9 cases and an exhaustive literature review, pyoderma gangrenosum, Sweet syndrome (n = 49), Sweet-like ND (n = 13), neutrophilic urticarial dermatosis (n = 6), palisaded neutrophilic granulomatous dermatitis (n = 12), and histiocytoid neutrophilic dermatitis (n = 2) were likely to occur both in AICTDs and autoinflammatory diseases. Other NDs were specifically encountered in AICTDs: bullous LE (n = 71), amicrobial pustulosis of the folds (n = 28), autoimmunity-related ND (n = 24), ND resembling erythema gyratum repens (n = 1), and neutrophilic annular erythema (n = 1). The improvement of AICTDS neutrophilic lesions under neutrophil targeting therapy suggests possible common physiopathological pathways between NDs and AICTDs. PMID:25546688

  10. Physiology of the fetal circulation.

    PubMed

    Kiserud, Torvid

    2005-12-01

    Our understanding of fetal circulatory physiology is based on experimental animal data, and this continues to be an important source of new insight into developmental mechanisms. A growing number of human studies have investigated the human physiology, with results that are similar but not identical to those from animal studies. It is time to appreciate these differences and base more of our clinical approach on human physiology. Accordingly, the present review focuses on distributional patterns and adaptational mechanisms that were mainly discovered by human studies. These include cardiac output, pulmonary and placental circulation, fetal brain and liver, venous return to the heart, and the fetal shunts (ductus venosus, foramen ovale and ductus arteriosus). Placental compromise induces a set of adaptational and compensational mechanisms reflecting the plasticity of the developing circulation, with both short- and long-term implications. Some of these aspects have become part of the clinical physiology of today with consequences for surveillance and treatment.

  11. Temporal adaptation of neutrophil oxidative responsiveness to n-formyl-methionyl-leucyl-phenylalanine. Acceleration by granulocyte-macrophage colony stimulating factor.

    PubMed

    English, D; Broxmeyer, H E; Gabig, T G; Akard, L P; Williams, D E; Hoffman, R

    1988-10-01

    This investigation was undertaken to clarify the mechanism by which purified recombinant human granulocyte-macrophage colony stimulating factor (GM-CSF) potentiates neutrophil oxidative responses triggered by the chemotactic peptide, FMLP. Previous studies have shown that GM-CSF priming of neutrophil responses to FMLP is induced relatively slowly, requiring 90 to 120 min of incubation in vitro, is not associated with increased levels of cytoplasmic free Ca2+, but is associated with up-regulation of cell-surface FMLP receptors. We have confirmed these findings and further characterized the process of GM-CSF priming. We found that the effect of GM-CSF on neutrophil oxidative responsiveness was induced in a temperature-dependent manner and was not reversed when the cells were washed extensively to remove the growth factor before stimulation with FMLP. Extracellular Ca2+ was not required for functional enhancement by GM-CSF and GM-CSF alone effected no detectable alteration in the 32P-labeled phospholipid content of neutrophils during incubation in vitro. Our data indicate that GM-CSF exerts its influence on neutrophils by accelerating a process that occurs spontaneously and results in up-regulation of both cell-surface FMLP receptors and oxidative responsiveness to FMLP. Thus, the results demonstrate that, with respect to oxidative activation, circulating endstage polymorphonuclear leukocytes are nonresponsive or hyporesponsive to FMLP; functional responsiveness increases dramatically as surface FMLP receptors are gradually deployed after the cells leave the circulation. Thus, as neutrophils mature, their responsiveness to FMLP changes in a manner which may be crucial for efficient host defense. At 37 degrees C, this process is markedly potentiated by GM-CSF. We conclude that endogenous GM-CSF, released systemically or at sites of infection and inflammation, potentially plays an important role in host defense by accelerating functional maturation of responding

  12. Phenotypic differences of human neutrophils of carriers of the PSGL-1 A and B-allele in binding to immobilised P-selectin under flow conditions.

    PubMed

    Meyer dos Santos, Sascha; Klinkhardt, Ute; Lang, Katharina; Parisius, Jeannine; Kuczka, Karina; Harder, Sebastian

    2011-02-01

    P-selectin glycoprotein ligand-1 (PSGL-1) interacts with P-selectin expressed on endothelial cells and platelets. PSGL-1 extracellular mucin-like domain displays a variable number of tandem repeats (VNTRs) polymorphism. The wildtype consists of 16 decameric repeats (designated A isoforms) and variants with 15 (B allele) and 14 (C allele) repeats that are assumed to be associated with reduced risk of vascular disease. We investigated the adhesion of these natural variants to P-selectin in native human neutrophils. Healthy volunteers were genotyped and the adhesion of neutrophils expressing the PSGL-1 isoforms A/A, A/B and B/B were studied under static and physiologic flow conditions. Homozygous B/B neutrophils attached significantly weaker to P-selectin at elevated shear rates from 24 up to 64 dyn/cm(2) than A/A and A/B neutrophils. No difference in adhesion rate was found under static conditions and shear stress below 24 dyn/cm(2), but B/B neutrophils rolled significantly faster than A/A neutrophils at shear stress ≥ 12 dyn/cm(2). There was no difference in the adhesive capacity between A/A an A/B neutrophils. These data support the view that the role of the decamers is to extend the ligand binding domain far above the cell surface to support stable leukocyte adhesion and rolling.

  13. Energy Metabolism of Human Neutrophils during Phagocytosis

    PubMed Central

    Borregaard, Niels; Herlin, Troels

    1982-01-01

    Detailed quantitative studies were performed on the generation and utilization of energy by resting and phagocytosing human neutrophils. The ATP content was 1.9 fmol/cell, was constant during rest, and was not influenced by the presence or absence of glucose in the medium. The intracellular content of phosphocreatine was less than 0.2 fmol/cell. In the presence of glucose, ATP was generated almost exclusively from lactate produced from glucose taken up from the surrounding medium. The amount of lactate produced could account for 85% of the glucose taken up by the cells, and the intracellular glycosyl store, glycogen, was not drawn upon. The rate of ATP generation as calculated from the rate of lactate production was 1.3 fmol/cell/min. During phagocytosis, there was no measurable increase in glucose consumption or lactate production, and the ATP content fell rapidly to 0.8 fmol/cell. This disappearance of ATP was apparently irreversible since no corresponding increase in ADP or AMP was observed. It therefore appears that this phagocytosis-induced fall in ATP concentration represents all the extra energy utilized in human neutrophils in the presence of glucose. In the absence of glucose, the rate of ATP generation in the resting cell was considerably smaller, 0.75 fmol/cell per min, as calculated from the rate of glycolysis, which is sustained exclusively by glycogenolysis. Under this condition, however, phagocytosis induces significant enhancement of glycogenolysis and the rate of lactate production is increased by 60%, raising the rate of ATP generation to 1.2 fmol/cell per min. Nonetheless, the ATP content drops significantly from 1.9 to 1.0 fmol/cell. Neutrophils from patients with chronic granulomatous disease have the same rate of glycolysis and the same ATP content as normal cells, thus confirming that the defective respiration of these cells does not affect their energy metabolism. PMID:7107894

  14. Cationic liposomes evoke proinflammatory mediator release and neutrophil extracellular traps (NETs) toward human neutrophils.

    PubMed

    Hwang, Tsong-Long; Hsu, Ching-Yun; Aljuffali, Ibrahim A; Chen, Chun-Han; Chang, Yuan-Ting; Fang, Jia-You

    2015-04-01

    Cationic liposomes are widely used as nanocarriers for therapeutic and diagnostic purposes. The cationic components of liposomes can induce inflammatory responses. This study examined the effect of cationic liposomes on human neutrophil activation. Cetyltrimethylammonium bromide (CTAB) or soyaethyl morpholinium ethosulfate (SME) was incorporated into liposomes as the cationic additive. The liposomes' cytotoxicity and their induction of proinflammatory mediators, intracellular calcium, and neutrophil extracellular traps (NETs) were investigated. The interaction of the liposomes with the plasma membrane triggered the stimulation of neutrophils. CTAB liposomes induced complete leakage of lactate dehydrogenase (LDH) at all concentrations tested, whereas SME liposomes released LDH in a concentration-dependent manner. CTAB liposomes proved to more effectively activate neutrophils compared with SME liposomes, as indicated by increased superoxide anion and elastase levels. Calcium influx increased 9-fold after treatment with CTAB liposomes. This influx was not changed by SME liposomes compared with the untreated control. Scanning electron microscopy (SEM) and immunofluorescence images indicated the presence of NETs after treatment with cationic liposomes. NETs could be quickly formed, within minutes, after CTAB liposomal treatment. In contrast to this result, NET formation was slowly and gradually increased by SME liposomes, within 4h. Based on the data presented here, it is important to consider the toxicity of cationic liposomes during administration in the body. This is the first report providing evidence of NET production induced by cationic liposomes.

  15. Neutrophil extracellular traps in neuropathy with anti-neutrophil cytoplasmic autoantibody-associated microscopic polyangiitis.

    PubMed

    Takeuchi, Hiroki; Kawasaki, Teruaki; Shigematsu, Kazuo; Kawamura, Kazuyuki; Oka, Nobuyuki

    2017-04-01

    To clarify the roles of neutrophils in anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitic neuropathy, we studied neutrophil extracellular traps (NETs) in peripheral nerve vasculitis. Stored nerve samples from 17 patients with microscopic polyangiitis (MPA) were immunohistochemically analyzed using antibodies for citrullinated histone H3 (citH3) and various neutrophil enzymes. We defined merged citH3 and extracellularly released myeloperoxidase (MPO) as NET formation. We also compared NET formation between MPO-ANCA-positive/negative MPA and rheumatoid arthritis (RA)-associated vasculitic neuropathy. NETs were identified mostly in vasculitic small arterioles of 6 of 12 MPO-ANCA-positive MPA patients, and their frequency was higher (p < 0.05) than in ANCA-negative patients. NETs were not found in vasculitic neuropathy with RA or patients with chronic inflammatory demyelinating polyradiculoneuropathy. NETs were also observed in the peripheral nervous system of MPA patients as well as in the lung and kidney. These results suggest that NETs may be involved in the pathogenesis of MPA neuropathy.

  16. Intravital Imaging of Neutrophil Recruitment Reveals the Efficacy of FPR1 Blockade in Hepatic Ischemia-Reperfusion Injury.

    PubMed

    Honda, Masaki; Takeichi, Takayuki; Hashimoto, Shintaro; Yoshii, Daiki; Isono, Kaori; Hayashida, Shintaro; Ohya, Yuki; Yamamoto, Hidekazu; Sugawara, Yasuhiko; Inomata, Yukihiro

    2017-02-15

    Neutrophils are considered responsible for the pathophysiological changes resulting from hepatic ischemia-reperfusion (I/R) injury, which is a complication of trauma, shock, liver resection, and transplantation. Recently, evidence is accumulating that formyl-peptide receptor (FPR) signaling constitutes an important danger signal that guides neutrophils to sites of inflammation. This study aimed to investigate dynamic neutrophil recruitment using two-photon laser-scanning microscopy (TPLSM) in response to FPR1 blockade during hepatic I/R. LysM-eGFP mice were subjected to partial warm hepatic I/R. They were pretreated with an FPR1 antagonist, cyclosporine H (CsH), or formyl peptide, fMLF. Liver was imaged after hepatic laser irradiation or I/R using the TPLSM technique. CsH treatment alleviated hepatic I/R injury, as evidenced by decreased serum transaminase levels, reduced hepatocyte necrosis/apoptosis, and diminished inflammatory cytokine, chemokine, and oxidative stress. In contrast, systemic administration of fMLF showed few effects. Time-lapse TPLSM showed that FPR1 blockade inhibited the accumulation of neutrophils in the necrotic area induced by laser irradiation in vivo. In the CsH-treated I/R group, the number and crawling velocity of neutrophils in the nonperfused area were lower than those in the control group. Meanwhile, FPR1 blockade did not affect monocyte/macrophage recruitment. Hepatic I/R promoted the retention of neutrophils and their active behavior in the spleen, whereas CsH treatment prevented their changes. Intravital TPLSM revealed that formyl-peptide-FPR1 signaling is responsible for regulating neutrophil chemotaxis to allow migration into the necrotic area in hepatic I/R. Our findings suggest effective approaches for elucidating the mechanisms of immune cell responses in hepatic I/R.

  17. Osteopontin Undergoes Polymerization in Vivo and Gains Chemotactic Activity for Neutrophils Mediated by Integrin α9β1*

    PubMed Central

    Nishimichi, Norihisa; Hayashita-Kinoh, Hiromi; Chen, Chun; Matsuda, Haruo; Sheppard, Dean; Yokosaki, Yasuyuki

    2011-01-01

    Osteopontin (OPN) is an integrin-binding inflammatory cytokine that undergoes polymerization catalyzed by transglutaminase 2. We have previously reported that polymeric OPN (polyOPN), but not unpolymerized OPN (OPN*), attracts neutrophils in vitro by presenting an acquired binding site for integrin α9β1. Among many in vitro substrates for transglutaminase 2, only a few have evidence for in vivo polymerization and concomitant function. Although polyOPN has been identified in bone and aorta, the in vivo functional significance of polyOPN is unknown. To determine whether OPN polymerization contributes to neutrophil recruitment in vivo, we injected OPN* into the peritoneal space of mice. Polymeric OPN was detected by immunoblotting in the peritoneal wash of mice injected with OPN*, and both intraperitoneal and plasma OPN* levels were higher in mice injected with a polymerization-incompetent mutant, confirming that OPN* polymerizes in vivo. OPN* injection induced neutrophil accumulation, which was significantly less following injection of a mutant OPN that was incapable of polymerization. The importance of in vivo polymerization was further confirmed with cystamine, a transglutaminase inhibitor, which blocked the polymerization and attenuated OPN*-mediated neutrophil recruitment. The thrombin-cleaved N-terminal fragment of OPN, another ligand for α9β1, was not responsible for neutrophil accumulation because a thrombin cleavage-incompetent mutant recruited similar numbers of neutrophils as wild type OPN*. Neutrophil accumulation in response to both wild type and thrombin cleavage-incompetent OPN* was reduced in mice lacking the integrin α9 subunit in leukocytes, indicating that α9β1 is required for polymerization-induced recruitment. We have illustrated a physiological role of molecular polymerization by demonstrating acquired chemotactic properties for OPN. PMID:21321126

  18. Are Neutrophil Extracellular Traps Playing a Role in the Parasite Control in Active American Tegumentary Leishmaniasis Lesions?

    PubMed Central

    Morgado, Fernanda Nazaré; Nascimento, Michelle T. C.; Saraiva, Elvira M.; de Oliveira-Ribeiro, Carla; Madeira, Maria de Fátima; da Costa-Santos, Marcela; Vasconcellos, Erica C. F.; F. Pimentel, Maria Ines; Rosandiski Lyra, Marcelo; Schubach, Armando de Oliveira; Conceição-Silva, Fátima

    2015-01-01

    Neutrophil extracellular traps (NETs) have been described as a network of extracellular fibers composed by DNA, histones and various proteins/enzymes. Studies have demonstrated that NETs could be responsible for the trapping and elimination of a variety of infectious agents. In order to verify the presence of NETs in American tegumentary leishmaniasis (ATL) and their relationship with the presence of amastigotes we evaluated active cutaneous lesions of 35 patients before treatment by the detection of parasites, neutrophils (neutrophil elastase) and histones through immunohistochemistry and confocal immunofluorescence. Intact neutrophils could be detected in all ATL lesions. NETs were present in 27 patients (median 1.1; range from 0.1 to 23.5/mm2) with lesion duration ranging from one to seven months. NETs were in close proximity with neutrophils (r = 0.586; p = 0.0001) and amastigotes (r = 0.710; p = 0.0001). Two patterns of NET formation were detected: small homogeneously distributed networks observed in all lesions; and large structures that could be visualized at a lower magnification in lesions presenting at least 20% of neutrophils. Lesions presenting the larger NET formation showed high parasite detection. A correlation between NET size and the number of intact amastigotes was observed (p=0.02). As we detected an association between NET and amastigotes, our results suggest that neutrophil migration and NET formation could be stimulated and maintained by stimuli derived from the parasite burden/parasite antigen in the extracellular environment. The observation of areas containing only antigens not intermingled with NETs (elastase and histone) suggests that the involvement of these structures in the control of parasite burden is a dynamic process in which the formation of NETs is exhausted with the destruction of the parasites. Since NETs were also associated with granulomas, this trapping would favor the activity of macrophages in order to control the parasite

  19. LFA-1 and Mac-1 define characteristically different intralumenal crawling and emigration patterns for monocytes and neutrophils in situ.

    PubMed

    Sumagin, Ronen; Prizant, Hen; Lomakina, Elena; Waugh, Richard E; Sarelius, Ingrid H

    2010-12-01

    To exit blood vessels, most (∼80%) of the lumenally adhered monocytes and neutrophils crawl toward locations that support transmigration. Using intravital confocal microscopy of anesthetized mouse cremaster muscle, we separately examined the crawling and emigration patterns of monocytes and neutrophils in blood-perfused unstimulated or TNF-α-activated venules. Most of the interacting cells in microvessels are neutrophils; however, in unstimulated venules, a greater percentage of the total monocyte population is adherent compared with neutrophils (58.2 ± 6.1% versus 13.6 ± 0.9%, adhered/total interacting), and they crawl for significantly longer distances (147.3 ± 13.4 versus 61.8 ± 5.4 μm). Intriguingly, after TNF-α activation, monocytes crawled for significantly shorter distances (67.4 ± 9.6 μm), resembling neutrophil crawling. Using function-blocking Abs, we show that these different crawling patterns were due to CD11a/CD18 (LFA-1)- versus CD11b/CD18 (Mac-1)-mediated crawling. Blockade of either Mac-1 or LFA-1 revealed that both LFA-1 and Mac-1 contribute to monocyte crawling; however, the LFA-1-dependent crawling in unstimulated venules becomes Mac-1 dependent upon inflammation, likely due to increased expression of Mac-1. Mac-1 alone was responsible for neutrophil crawling in both unstimulated and TNF-α-activated venules. Consistent with the role of Mac-1 in crawling, Mac-1 block (compared with LFA-1) was also significantly more efficient in blocking TNF-α-induced extravasation of both monocytes and neutrophils in cremaster tissue and the peritoneal cavity. Thus, mechanisms underlying leukocyte crawling are important in regulating the inflammatory responses by regulating the numbers of leukocytes that transmigrate.

  20. Monocyte and neutrophil isolation and migration assays.

    PubMed

    Yona, Simon; Hayhoe, Richard; Avraham-Davidi, Inbal

    2010-02-01

    This unit describes methods for isolating mouse monocytes and neutrophils, as well as in vitro protocols for measuring cell migration and polarization. The method employed here for the isolation of naïve phagocytes overcomes many of the difficulties previously encountered concerning phagocyte activation. Three in vitro protocols are provided for the analysis of cell migration, one requiring no specialized equipment, one requiring the modified Boyden chamber, and the other employing a flow chamber, which measures cell adhesion, rolling, and migration. Finally, a method is provided for imaging polarized cells by confocal microscopy.

  1. Neutrophils Discriminate between Lipopolysaccharides of Different Bacterial Sources and Selectively Release Neutrophil Extracellular Traps

    PubMed Central

    Pieterse, Elmar; Rother, Nils; Yanginlar, Cansu; Hilbrands, Luuk B.; van der Vlag, Johan

    2016-01-01

    The release of neutrophil extracellular traps (NETs), either during “suicidal” or “vital” NETosis, represents an important strategy of neutrophils to combat Gram-negative bacteria. Lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria, is a reported stimulus for NET formation. Although it is widely acknowledged that the structural diversity in LPS structures can elicit heterogeneous immune responses, species- and serotype-specific differences in the capacity of LPS to trigger NET formation have not yet been investigated. In the present study, we compared the NET-inducing potential of LPS derived from Escherichia coli (serotypes O55:B5, O127:B8, O128:B12, O111:B4, and O26:B6), Salmonella enterica (serotype enteritidis), and Pseudomonas aeruginosa (serotype 10), under platelet-free and platelet-rich conditions in vitro, and in whole blood ex vivo. Here, we demonstrate that under serum- and platelet-free conditions, mimicking tissue circumstances, neutrophils discriminate between LPS of different bacterial sources and selectively release NETs only in response to LPS derived from E. coli O128:B12 and P. aeruginosa 10, which both induced “suicidal” NETosis in an autophagy- and reactive oxygen species (ROS)-dependent, but TLR4-independent manner. Intriguingly, in whole blood cultures ex vivo, or in vitro in the presence of platelets, all LPS serotypes induced “vital” NET formation. This platelet-dependent release of NETs occurred rapidly without neutrophil cell death and was independent from ROS formation and autophagy but required platelet TLR4 and CD62P-dependent platelet–neutrophil interactions. Taken together, our data reveal a complex interplay between neutrophils and LPS, which can induce both “suicidal” and “vital” NETosis, depending on the bacterial origin of LPS and the presence or absence of platelets. Our findings suggest that LPS sensing by neutrophils may be a critical determinant for

  2. The number of circulating CD14+ cells is related to infarct size and postinfarct volumes in ST segment elevation myocardial infarction but not non-ST segment elevation myocardial infarction

    PubMed Central

    Montange, Damien; Davani, Siamak; Deschaseaux, Frédéric; Séronde, Marie France; Chopard, Romain; Schiele, François; Jehl, Jérome; Bassand, Jean Pierre; Kantelip, Jean-Pierre; Meneveau, Nicolas

    2012-01-01

    OBJECTIVE: To determine the relationship between the number of CD14+ cells, myocardial infarct (MI) size and left ventricular (LV) volumes in ST segment elevation MI (STEMI) and non-ST segment elevation MI (NSTEMI) patients. METHODS: A total of 62 patients with STEMI (n=34) or NSTEMI (n=28) were enrolled. The number of CD14+ cells was assessed at admission. Infarct size, left ventricular ejection fraction (LVEF) and LV volumes were measured using magnetic resonance imaging five days after MI and six months after MI. Results: In STEMI patients, the number of CD14+ cells was positively and significantly correlated with infarct size at day 5 (r=0.40; P=0.016) and after six months (r=0.34; P=0.047), negatively correlated with LVEF at day 5 (r=−0.50; P=0.002) and after six months (r=−0.46; P=0.005) and positively correlated with end-diastolic (r=0.38; P=0.02) and end-systolic (r=0.49; P=0.002) volumes after six months. In NSTEMI patients, no significant correlation was found between the number of CD14+ cells and infarct size, LVEF or LV volumes at day 5 or after six months. CONCLUSIONS: The number of CD14+ cells at admission was associated with infarct size and LV remodelling in STEMI patients with large infarct size, whereas in NSTEMI patients, no relationship was observed between numbers of CD14+ cells and LV remodelling. PMID:23620701

  3. Characterization of C1 inhibitor binding to neutrophils.

    PubMed Central

    Chang, N S; Boackle, R J; Leu, R W

    1991-01-01

    In a previous study we have isolated neutrophil membrane proteins that non-covalently bind to native C1-INH (105,000 MW) and a non-functional, degraded C1-INH (88,000 MW; C1-INH-88). To further characterize the binding nature, we have designed a novel kinetic C1 titration assay which enables not only a quantification of the removal of fluid-phase C1-INH by neutrophils, but also a concomitant measure of residual C1-INH function. Native C1-INH, when adsorbed to EDTA-pretreated neutrophils, lost its function in the inhibition of fluid-phase C1. The non-functional C1-INH-88, which is probably devoid of a reactive centre, was found to block the binding of native C1-INH to neutrophils. Pretreatment of neutrophils with serine esterase inhibitors did not abrogate binding capacity of the cells for C1-INH, whereas the binding affinity for C1-INH was lost when the cells were pretreated with trypsin. An array of human peripheral blood leucocytes and several lymphoid cell lines has surface binding sites for C1-INH, but not on human erythrocytes and U937 cells. Binding was further confirmed using (i) C1-INH-microsphere beads to neutrophils, in which the binding was blocked when pretreating neutrophils with excess C1-INH or with trypsin, and (ii) radiolabelled C1-INH to neutrophils, which was competitively blocked by unlabelled non-functional C1-INH-88. Desialylation of C1-INH significantly reduced its binding affinity for neutrophils, indicating that the membrane receptor sites on neutrophils could be specific for the binding of sialic acid residues on C1-INH. Overall, our studies indicate that neutrophils or other leucocytes possess specific surface binding sites for the sialic acid-containing portion of C1-INH. PMID:2045131

  4. Nucleosomes and neutrophil activation in sickle cell disease painful crisis.

    PubMed

    Schimmel, Marein; Nur, Erfan; Biemond, Bart J; van Mierlo, Gerard J; Solati, Shabnam; Brandjes, Dees P; Otten, Hans-Martin; Schnog, John-John; Zeerleder, Sacha

    2013-11-01

    Activated polymorphonuclear neutrophils play an important role in the pathogenesis of vaso-occlusive painful sickle cell crisis. Upon activation, polymorphonuclear neutrophils can form neutrophil extracellular traps. Neutrophil extracellular traps consist of a meshwork of extracellular DNA, nucleosomes, histones and neutrophil proteases. Neutrophil extracellular traps have been demonstrated to be toxic to endothelial and parenchymal cells. This prospective cohort study was conducted to determine neutrophil extracellular trap formation in sickle cell patients during steady state and painful crisis. As a measure of neutrophil extracellular traps, plasma nucleosomes levels were determined and polymorphonuclear neutrophil activation was assessed measuring plasma levels of elastase-α1-antitrypsin complexes in 74 patients in steady state, 70 patients during painful crisis, and 24 race-matched controls using Enzyme Linked Immunosorbent Assay. Nucleosome levels in steady state sickle cell patients were significantly higher than levels in controls. During painful crisis levels of both nucleosomes and elastase-α1-antitrypsin complexes increased significantly. Levels of nucleosomes correlated significantly to elastase-α1-antitrypsin complex levels during painful crisis, (Sr = 0.654, P<0.001). This was seen in both HbSS/HbSβ(0)-thalassemia (Sr=0.55, P<0.001) and HbSC/HbSβ(+-)thalassemia patients (Sr=0.90, P<0.001) during painful crisis. Levels of nucleosomes showed a correlation with length of hospital stay and were highest in patients with acute chest syndrome. These data support the concept that neutrophil extracellular trap formation and neutrophil activation may play a role in the pathogenesis of painful sickle cell crisis and acute chest syndrome.

  5. tPA-MMP-9 Axis Plays a Pivotal Role in Mobilization of Endothelial Progenitor Cells from Bone Marrow to Circulation and Ischemic Region for Angiogenesis

    PubMed Central

    Day, Yuan-Ji

    2016-01-01

    We examined the role of tissue plasminogen activator- (tPA-) matrix metalloproteinase- (MMP-) 9 in mobilizing endothelial progenitor cells (EPCs) from bone marrow to circulation and critical limb ischemia (CLI) region. Male C57BL/6J mice having been irradiated were categorized into wild-type mice (WT) receiving WT bone marrow cell (BMC) transfusion (group 1), WT mice receiving MMP-9 knockout (MMP-9−/−) BMC (group 2), MMP-9−/− receiving MMP-9−/− BMC (group 3), and MMP-9−/− receiving WT BMC (group 4), each of which was subdivided into sham control (SC), CLI, SC-tPA, and CLI-tPA. In groups 1 and 4, by post-CLI 18 h and day 14, circulating EPC (C-kit+/CD31+, Sca-1+/KDR+) levels were highest in CLI-tPA subgroup. In groups 2 and 3, EPC levels did not differ among all subgroups. The EPC levels in bone marrow were higher in groups 2 and 3 than those in groups 1 and 4. By day 14, in animals with CLI, expression levels of proangiogenic factors (CXCR4, SDF-1α, and VEGF) showed similar trends as circulating EPC levels. Moreover, the number of infiltrated neutrophils and macrophages in quadriceps was higher in groups 1 and 4 than groups in 2 and 3. In conclusion, tPA-MMP-9 axis plays a crucial role in EPC mobilization and angiogenesis in experimental CLI. PMID:27610138

  6. Circulation Scenarios: Or, Where Have All the New Books Gone?

    ERIC Educational Resources Information Center

    Johnson, Mary E.

    1986-01-01

    This analysis of the circulation records of books added to a psychiatric library from July 1983 through May 1985 reports the total number of books circulated, their number and percentage in specific subject areas, missing titles, and usage by in-house and off-site borrowers. Implications for future collection development are explained. (Author/EM)

  7. Shielding of a lipooligosaccharide IgM epitope allows evasion of neutrophil-mediated killing of an invasive strain of nontypeable Haemophilus influenzae.

    PubMed

    Langereis, Jeroen D; Weiser, Jeffrey N

    2014-07-22

    Nontypeable Haemophilus influenzae is a frequent cause of noninvasive mucosal inflammatory diseases but may also cause invasive diseases, such as sepsis and meningitis, especially in children and the elderly. Infection by nontypeable Haemophilus influenzae is characterized by recruitment of neutrophilic granulocytes. Despite the presence of a large number of neutrophils, infections with nontypeable Haemophilus influenzae are often not cleared effectively by the antimicrobial activity of these immune cells. Herein, we examined how nontypeable Haemophilus influenzae evades neutrophil-mediated killing. Transposon sequencing (Tn-seq) was used on an isolate resistant to neutrophil-mediated killing to identify genes required for its survival in the presence of human neutrophils and serum, which provided a source of complement and antibodies. Results show that nontypeable Haemophilus influenzae prevents complement-dependent neutrophil-mediated killing by expression of surface galactose-containing oligosaccharide structures. These outer-core structures block recognition of an inner-core lipooligosaccharide epitope containing glucose attached to heptose HepIII-β1,2-Glc by replacement with galactose attached to HepIII or through shielding HepIII-β1,2-Glc by phase-variable attachment of oligosaccharide chain extensions. When the HepIII-β1,2-Glc-containing epitope is expressed and exposed, nontypeable Haemophilus influenzae is opsonized by naturally acquired IgM generally present in human serum and subsequently phagocytosed and killed by human neutrophils. Clinical nontypeable Haemophilus influenzae isolates containing galactose attached to HepIII that are not recognized by this IgM are more often found to cause invasive infections. Importance: Neutrophils are white blood cells that specialize in killing pathogens and are recruited to sites of inflammation. However, despite the presence of large numbers of neutrophils in the middle ear cavity and lungs of patients with

  8. Simulation model for flow of neutrophils in pulmonary capillary network.

    PubMed

    Shirai, Atsushi; Fujita, Ryo; Hayase, Toshiyuki

    2005-01-01

    The concentration of neutrophils in the pulmonary microvasculature is higher than in systemic large vessels. It is thought that the high concentration of neutrophils facilitates their effective recruitment to sites of inflammation. Thus, in order to understand the role of neutrophils in the immune system, it is important to clarify their flow characteristics in the pulmonary microvasculature. In previous studies, we numerically investigated the motion of a neutrophil through a single capillary segment modeled by a moderate axisymmetric constriction in a straight pipe, developing a mathematical model for the prediction of the transit time of the cell through the segment. In the present study, this model was extended for application to network simulation of the motion of neutrophils. First, we numerically investigated shape recovery of a neutrophil after expulsion from a narrow capillary segment. This process was modeled in two different phases: elastic recovery and viscous recovery. The resulting model was combined with the previously developed models to simulate motion of the cells and plasma flow in a capillary network. A numerical simulation of the motion of neutrophils and plasma flow in a simple lattice capillary network showed that neutrophils were widely dispersed in the network with an increased concentration.

  9. Intergrin-dependent neutrophil migration in the injured mouse cornea

    Technology Transfer Automated Retrieval System (TEKTRAN)

    As an early responder to an inflammatory stimulus, neutrophils must exit the vasculature and migrate through the extravascular tissue to the site of insult, which is often remote from the point of extravasation. Following a central epithelial corneal abrasion, neutrophils recruited from the peripher...

  10. Promoting metastasis: neutrophils and T cells join forces.

    PubMed

    Fridlender, Zvi G; Albelda, Steven M; Granot, Zvi

    2015-07-01

    The role neutrophils play in cancer is a matter of debate as both pro- and anti-tumor functions have been documented. In a recent publication in Nature, Coffelt et al. identify a new mechanism where neutrophils and T cells cooperate to generate metastasis-supporting immune suppression.

  11. Transepithelial migration of neutrophils into the lung requires TREM-1

    PubMed Central

    Klesney-Tait, Julia; Keck, Kathy; Li, Xiaopeng; Gilfillan, Susan; Otero, Karel; Baruah, Sankar; Meyerholz, David K.; Varga, Steven M.; Knudson, Cory J.; Moninger, Thomas O.; Moreland, Jessica; Zabner, Joseph; Colonna, Marco

    2012-01-01

    Acute respiratory infections are responsible for more than 4 million deaths each year. Neutrophils play an essential role in the innate immune response to lung infection. These cells have an armamentarium of pattern recognition molecules and antimicrobial agents that identify and eliminate pathogens. In the setting of infection, neutrophil triggering receptor expressed on myeloid cells 1 (TREM-1) amplifies inflammatory signaling. Here we demonstrate for the first time that TREM-1 also plays an important role in transepithelial migration of neutrophils into the airspace. We developed a TREM-1/3–deficient mouse model of pneumonia and found that absence of TREM-1/3 markedly increased mortality following Pseudomonas aeruginosa challenge. Unexpectedly, TREM-1/3 deficiency resulted in increased local and systemic cytokine production. TREM-1/3–deficient neutrophils demonstrated intact bacterial killing, phagocytosis, and chemotaxis; however, histologic examination of TREM-1/3–deficient lungs revealed decreased neutrophil infiltration of the airways. TREM-1/3–deficient neutrophils effectively migrated across primary endothelial cell monolayers but failed to migrate across primary airway epithelia grown at the air-liquid interface. These data define a new function for TREM-1 in neutrophil migration across airway epithelial cells and suggest that it amplifies inflammation through targeted neutrophil migration into the lung. PMID:23241959

  12. Impaired neutrophil directional chemotactic accuracy in chronic periodontitis patients

    PubMed Central

    Roberts, Helen M; Ling, Martin R; Insall, Robert; Kalna, Gabriela; Spengler, Julia; Grant, Melissa M; Chapple, Iain LC

    2015-01-01

    Aim To investigate the chemotactic accuracy of peripheral blood neutrophils from patients with chronic periodontitis compared with matched healthy controls, before and after non-surgical periodontal therapy. Material & Methods Neutrophils were isolated from patients and controls (n = 18) by density centrifugation. Using the Insall chamber and video microscopy, neutrophils were analysed for directional chemotaxis towards N-formyl-methionyl-leucyl-phenylalanine [fMLP (10 nM), or CXCL8 (200 ng/ml)]. Circular statistics were utilized for the analysis of cell movement. Results Prior to treatment, neutrophils from patients with chronic periodontitis had significantly reduced speed, velocity and chemotactic accuracy compared to healthy controls for both chemoattractants. Following periodontal treatment, patient neutrophils continued to display reduced speed in response to both chemoattractants. However, velocity and accuracy were normalized for the weak chemoattractant CXCL8 while they remained significantly reduced for fMLP. Conclusions Chronic periodontitis is associated with reduced neutrophil chemotaxis, and this is only partially restored by successful treatment. Dysfunctional neutrophil chemotaxis may predispose patients with periodontitis to their disease by increasing tissue transit times, thus exacerbating neutrophil-mediated collateral host tissue damage. PMID:25360483

  13. Transepithelial migration of neutrophils into the lung requires TREM-1.

    PubMed

    Klesney-Tait, Julia; Keck, Kathy; Li, Xiaopeng; Gilfillan, Susan; Otero, Karel; Baruah, Sankar; Meyerholz, David K; Varga, Steven M; Knudson, Cory J; Moninger, Thomas O; Moreland, Jessica; Zabner, Joseph; Colonna, Marco

    2013-01-01

    Acute respiratory infections are responsible for more than 4 million deaths each year. Neutrophils play an essential role in the innate immune response to lung infection. These cells have an armamentarium of pattern recognition molecules and antimicrobial agents that identify and eliminate pathogens. In the setting of infection, neutrophil triggering receptor expressed on myeloid cells 1 (TREM-1) amplifies inflammatory signaling. Here we demonstrate for the first time that TREM-1 also plays an important role in transepithelial migration of neutrophils into the airspace. We developed a TREM-1/3-deficient mouse model of pneumonia and found that absence of TREM-1/3 markedly increased mortality following Pseudomonas aeruginosa challenge. Unexpectedly, TREM-1/3 deficiency resulted in increased local and systemic cytokine production. TREM-1/3-deficient neutrophils demonstrated intact bacterial killing, phagocytosis, and chemotaxis; however, histologic examination of TREM-1/3-deficient lungs revealed decreased neutrophil infiltration of the airways. TREM-1/3-deficient neutrophils effectively migrated across primary endothelial cell monolayers but failed to migrate across primary airway epithelia grown at the air-liquid interface. These data define a new function for TREM-1 in neutrophil migration across airway epithelial cells and suggest that it amplifies inflammation through targeted neutrophil migration into the lung.

  14. Human neutrophil leukocyte elastase activity is inhibited by Phenol Red

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Neutrophil elastase (NE) activity in urine, sputum and nasal mucous is used as an indicator of inflammation due to viral or bacterial infection. However, bovine nasal mucous neutrophils collected, lysed and stored in Dulbecco's minimal medium containing Phenol Red, showed no NE activity with methox...

  15. How Neutrophil Extracellular Traps Become Visible

    PubMed Central

    2016-01-01

    Neutrophil extracellular traps (NETs) have been identified as a fundamental innate immune defense mechanism against different pathogens. NETs are characterized as released nuclear DNA associated with histones and granule proteins, which form an extracellular web-like structure that is able to entrap and occasionally kill certain microbes. Furthermore, NETs have been shown to contribute to several noninfectious disease conditions when released by activated neutrophils during inflammation. The identification of NETs has mainly been succeeded by various microscopy techniques, for example, immunofluorescence microscopy, transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Since the last years the development and improvement of new immunofluorescence-based techniques enabled optimized visualization and quantification of NETs. On the one hand in vitro live-cell imaging led to profound new ideas about the mechanisms involved in the formation and functionality of NETs. On the other hand different intravital, in vivo, and in situ microscopy techniques led to deeper insights into the role of NET formation during health and disease. This paper presents an overview of the main used microscopy techniques to visualize NETs and describes their advantages as well as disadvantages. PMID:27294157

  16. Toxic oxygen metabolite production by circulating phagocytic cells in inflammatory bowel disease.

    PubMed Central

    Williams, J G; Hughes, L E; Hallett, M B

    1990-01-01

    To investigate the possibility that the oxidative capacity of phagocytic cells may be defective in inflammatory bowel disease, toxic oxygen metabolite production by circulating neutrophils and monocytes has been measured by luminol dependent chemiluminescence. Neutrophils from patients with Crohn's disease and ulcerative colitis produced significantly lower chemiluminescent responses after chemotactic stimulation with formylmethionylleucylphenylalanine (fMLP) than neutrophils from control patients, p = 0.018 and 0.043 respectively. Chemiluminescent responses of neutrophils from patients with inflammatory bowel disease, however, were similar to control responses when cells were stimulated with latex beads or phorbol myristate acetate. Monocytes from patients with Crohn's disease produced significantly greater levels of chemiluminescence than control monocytes when stimulated with either fMLP (p less than 0.002), phorbol myristate acetate (p less than 0.0005) or latex beads (p less than 0.002). Monocytes from patients with ulcerative colitis also produced significantly greater levels of chemiluminescence than controls when stimulated with latex beads (p less than 0.5) or phorbol myristate acetate (p less than 0.0005), although there was no difference in the level of chemiluminescence in response to fMLP. These results exclude a generalised defect in phagocytic cell oxidase activity in inflammatory bowel disease and suggest that circulating monocytes are 'activated'. PMID:2311976

  17. Interaction of natural killer cells with neutrophils exerts a significant antitumor immunity in hematopoietic stem cell transplantation recipients.

    PubMed

    Ueda, Ryosuke; Narumi, Kenta; Hashimoto, Hisayoshi; Miyakawa, Reina; Okusaka, Takuji; Aoki, Kazunori

    2016-01-01

    Autologous hematopoietic stem cell transplantation (HSCT) can induce a strong antitumor immunity by homeostatic proliferation (HP) of T cells and suppression of regulatory T cells following preconditioning-induced lymphopenia. However, the role of innate immunity including natural killer (NK) cells is still not understood. Here, first, we examined whether NK cells exert an antitumor effect after syngeneic HSCT in a murine colon cancer model. Flow cytometry showed that NK cells as well as T cells rapidly proliferated after HSCT, and the frequency of mature NK cells was increased in tumor during HP. Furthermore, NK cells undergoing HP were highly activated, which contributed to substantial tumor suppression. Then, we found that a large number of neutrophils accumulated in tumor early after syngeneic HSCT. It was recently reported that neutrophil-derived mediators modulate NK cell effector functions, and so we examined whether the neutrophils infiltrated in tumor are associated with NK cell-mediated antitumor effect. The depletion of neutrophils significantly impaired an activation of NK cells in tumor and increased the fraction of proliferative NK cells accompanied by a decrease in NK cell survival. The results suggested that neutrophils in tumor prevent NK cells from activation-induced cell death during HP, thus leading to a significant antitumor effect by NK cells. This study revealed a novel aspect of antitumor immunity induced by HSCT and may contribute to the development of an effective therapeutic strategy for cancer using HSCT.

  18. Human neutrophil elastase detection with fluorescent peptide sensors conjugated to cellulosic and nanocellulosic materials: part II, structure/function analysis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Human neutrophil elastase (HNE) is one of a number of proteases that is receiving increased attention as a marker for inflammatory diseases and sensor-based point of care diagnostics. Integral to sensor-based detection is the transducer surface which is the platform of the sensor's signal transmitta...

  19. Circulating serotonin in vertebrates.

    PubMed

    Maurer-Spurej, E

    2005-08-01

    The role of circulating serotonin is unclear and whether or not serotonin is present in the blood of non-mammalian species is not known. This study provides the first evidence for the presence of serotonin in thrombocytes of birds and three reptilian species, the endothermic leatherback sea turtle, the green sea turtle and the partially endothermic American alligator. Thrombocytes from a fresh water turtle, American bullfrog, Yellowfin tuna, and Chinook salmon did not contain serotonin. Serotonin is a vasoactive substance that regulates skin blood flow, a major mechanism for endothermic body temperature regulation, which could explain why circulating serotonin is present in warm-blooded species. The temperature sensitivity of human blood platelets with concomitant changes in serotonin content further supports a link between circulating serotonin and thermoregulation. Phylogenetic comparison of the presence of circulating serotonin indicated an evolutionary divergence within reptilian species that might coincide with the emergence of endothermy.

  20. Structural Determination of Circulation.

    ERIC Educational Resources Information Center

    Blankenburg, William B.

    1981-01-01

    Analyzes the effects of both structural factors (demographics, economic conditions, and competition) and discretionary factors (content, design, and marketing techniques) and concludes that it is the former that determine a newspaper's circulation. (FL)

  1. Exosomes Mediate LTB4 Release during Neutrophil Chemotaxis

    PubMed Central

    Majumdar, Ritankar; Tavakoli Tameh, Aidin; Parent, Carole A.

    2016-01-01

    Leukotriene B4 (LTB4) is secreted by chemotactic neutrophils, forming a secondary gradient that amplifies the reach of primary chemoattractants. This strategy increases the recruitment range for neutrophils and is important during inflammation. Here, we show that LTB4 and its synthesizing enzymes localize to intracellular multivesicular bodies that, upon stimulation, release their content as exosomes. Purified exosomes can activate resting neutrophils and elicit chemotactic activity in a LTB4 receptor-dependent manner. Inhibition of exosome release leads to loss of directional motility with concomitant loss of LTB4 release. Our findings establish that the exosomal pool of LTB4 acts in an autocrine fashion to sensitize neutrophils towards the primary chemoattractant, and in a paracrine fashion to mediate the recruitment of neighboring neutrophils in trans. We envision that this mechanism is used by other signals to foster communication between cells in harsh extracellular environments. PMID:26741884

  2. Swell activated chloride channel function in human neutrophils

    SciTech Connect

    Salmon, Michael D.; Ahluwalia, Jatinder

    2009-04-17

    Non-excitable cells such as neutrophil granulocytes are the archetypal inflammatory immune cell involved in critical functions of the innate immune system. The electron current generated (I{sub e}) by the neutrophil NADPH oxidase is electrogenic and rapidly depolarises the membrane potential. For continuous function of the NADPH oxidase, I{sub e} has to be balanced to preserve electroneutrality, if not; sufficient depolarisation would prevent electrons from leaving the cell and neutrophil function would be abrogated. Subsequently, the depolarisation generated by the neutrophil NADPH oxidase I{sub e} must be counteracted by ion transport. The finding that depolarisation required counter-ions to compensate electron transport was followed by the observation that chloride channels activated by swell can counteract the NADPH oxidase membrane depolarisation. In this mini review, we discuss the research findings that revealed the essential role of swell activated chloride channels in human neutrophil function.

  3. Coexistence of chronic neutrophilic leukemia with multiple myeloma.

    PubMed

    Dinçol, Günçağ; Nalçaci, Meliha; Doğan, Oner; Aktan, Melih; Küçükkaya, Reyhan; Ağan, Mehmet; Dinçol, Koray

    2002-03-01

    A case report of simultaneous presentation of chronic neutrophilic leukemia and multiple myeloma (IgG kappa) in a 71-year-old male is described. The patient showed mature neutrophilic leukocytosis, hepatosplenomegaly, high neutrophil alkaline phosphatase score, hyperuricemia, neutrophils with toxic granulation and Döhle bodies, absence of Philadelphia chromosome and of the bcr-abl fusion gene. Moreover, a monoclonal IgG kappa paraproteinemia (36.93 g l(-1)) was detected. Bence-Jones proteinuria was 3.84 g l(-1). The bone marrow was grossly hypercellular with marked myeloid hyperplasia and aggregates of plasma cells. The patient died of severe bronchopneumonia after the transformation of chronic neutrophilic leukemia to acute myelomonocytic leukemia, 1.5 years following diagnosis.

  4. Structural divergence of GPI-80 in activated human neutrophils.

    PubMed

    Nitto, Takeaki; Takeda, Yuji; Yoshitake, Hiroshi; Sendo, Fujiro; Araki, Yoshihiko

    2007-07-27

    GPI-80 is a glycosylphosphatidylinositol (GPI)-anchored protein that is mainly expressed in human neutrophils. Previous studies using 3H9, a monoclonal antibody (mAb) against GPI-80, suggested that GPI-80 regulates leukocyte adherence and migration through Mac-1. GPI-80, which is anchored at the plasma membrane in resting neutrophils, moves into the pseudopodia and is released from activated human neutrophils. Here, we demonstrate that neutrophil activation affects GPI-80 dynamics using a new anti-GPI-80 mAb, designated 4D4, which is directed against the form of GPI-80 found on resting human neutrophils. Similar to 3H9, 4D4 influences Mac-1-dependent neutrophil adhesion. Treatment of purified GPI-80 with periodic acid and trypsin indicated that 3H9 and 4D4 recognize peptide and carbohydrate moieties, respectively. Stimulation with fMLP decreased the binding of 4D4 to GPI-80 on the neutrophil surface but increased the overall expression of GPI-80, as visualized by the 3H9 signal. Confocal laser microscopy revealed the 4D4 signal mainly on cell bodies and at a low level on pseudopodia during migration toward increasing concentrations of fMLP, whereas the 3H9 signal was observed in both areas. In addition, soluble GPI-80 released from activated neutrophils did not bind 4D4. These results suggest that there are two populations of GPI-80 that differ in the ability to bind 4D4. The 4D4-recognized form may regulate Mac-1-dependent neutrophil adhesion, and may subsequently be converted to a 4D4-unrecognized form during neutrophil activation.

  5. Role of the endothelial surface layer in neutrophil recruitment.

    PubMed

    Marki, Alex; Esko, Jeffrey D; Pries, Axel R; Ley, Klaus

    2015-10-01

    Neutrophil recruitment in most tissues is limited to postcapillary venules, where E- and P-selectins are inducibly expressed by venular endothelial cells. These molecules support neutrophil rolling via binding of PSGL-1 and other ligands on neutrophils. Selectins extend ≤ 38 nm above the endothelial plasma membrane, and PSGL-1 extends to 50 nm above the neutrophil plasma membrane. However, endothelial cells are covered with an ESL composed of glycosaminoglycans that is ≥ 500 nm thick and has measurable resistance against compression. The neutrophil surface is also covered with a surface layer. These surface layers would be expected to completely shield adhesion molecules; thus, neutrophils should not be able to roll and adhere. However, in the cremaster muscle and in many other models investigated using intravital microscopy, neutrophils clearly roll, and their rolling is easily and quickly induced. This conundrum was thought to be resolved by the observation that the induction of selectins is accompanied by ESL shedding; however, ESL shedding only partially reduces the ESL thickness (to 200 nm) and thus is insufficient to expose adhesion molecules. In addition to its antiadhesive functions, the ESL also presents neutrophil arrest-inducing chemokines. ESL heparan sulfate can also bind L-selectin expressed by the neutrophils, which contributes to rolling and arrest. We conclude that ESL has both proadhesive and antiadhesive functions. However, most previous studies considered either only the proadhesive or only the antiadhesive effects of the ESL. An integrated model for the role of the ESL in neutrophil rolling, arrest, and transmigration is needed.

  6. Visceral Leishmaniasis Patients Display Altered Composition and Maturity of Neutrophils as well as Impaired Neutrophil Effector Functions

    PubMed Central

    Yizengaw, Endalew; Getahun, Mulusew; Tajebe, Fitsumbrhan; Cruz Cervera, Edward; Adem, Emebet; Mesfin, Getnet; Hailu, Asrat; Van der Auwera, Gert; Yardley, Vanessa; Lemma, Mulualem; Skhedy, Ziv; Diro, Ermias; Yeshanew, Arega; Melkamu, Roma; Mengesha, Bewketu; Modolell, Manuel; Munder, Markus; Müller, Ingrid; Takele, Yegnasew; Kropf, Pascale

    2016-01-01

    Immunologically, active visceral leishmaniasis (VL) is characterized by profound immunosuppression, severe systemic inflammatory responses, and an impaired capacity to control parasite replication. Neutrophils are highly versatile cells, which play a crucial role in the induction as well as the resolution of inflammation, the control of pathogen replication, and the regulation of immune responses. Neutrophil functions have been investigated in human cutaneous leishmaniasis; however, their role in human VL is poorly understood. In the present study we evaluated the activation status and effector functions of neutrophils in patients with active VL and after successful anti-leishmanial treatment. Our results show that neutrophils are highly activated and have degranulated; high levels of arginase, myeloperoxidase, and elastase, all contained in neutrophils’ granules, were found in the plasma of VL patients. In addition, we show that a large proportion of these cells are immature. We also analyzed effector functions of neutrophils that are essential for pathogen clearance and show that neutrophils have an impaired capacity to release neutrophil extracellular traps, produce reactive oxygen species, and phagocytose bacterial particles, but not Leishmania parasites. Our results suggest that impaired effector functions, increased activation, and immaturity of neutrophils play a key role in the pathogenesis of VL. PMID:27965662

  7. CXC Receptor 1 and 2 and Neutrophil Elastase Inhibitors Alter Radiation-induced Lung Disease in the Mouse

    SciTech Connect

    Fox, Jessica; Haston, Christina K.

    2013-01-01

    Purpose: We previously reported increased numbers of neutrophils to be associated with the development of the radiation-induced lung responses of alveolitis (pneumonitis) and fibrosis in mice. In the present study we investigated whether CXC receptor 1 and 2 antagonism with DF2156A, a small molecule inhibitor of neutrophil chemotaxis, or the neutrophil elastase inhibitor sivelestat decreases the lung response to irradiation. Methods and Materials: KK/HIJ mice received 14 Gy whole-thorax irradiation, and a subset of them received drug treatment 3 times per week from the day of irradiation until they were killed because of respiratory distress symptoms. Results: Irradiated mice receiving sivelestat survived 18% longer than did mice receiving radiation alone (73 vs 60 days for female mice, 91 vs 79 days for male mice), whereas postirradiation survival times did not differ between the group of mice receiving DF2156A and the radiation-only group. The numbers of neutrophils in lung tissue and in bronchoalveolar lavage fluid did not differ among groups of irradiated mice, but they significantly exceeded the levels in unirradiated control mice. The extent of alveolitis, assessed histologically, did not differ between irradiated mice treated with either drug and those receiving radiation alone, when assessed at the end of the experiment, but it was significantly reduced, as were the neutrophil measures, in sivelestat-treated mice at the common kill time of 60 days after irradiation. Mice treated with radiation and DF2156A developed significantly less fibrosis than did mice receiving radiation alone, and this difference was associated with decreased expression of interleukin-13 in lung tissue. Conclusions: We conclude that neutrophil elastase inhibition affects alveolitis and prolongs survival, whereas CXCR1/2 antagonism reduces radiation-induced fibrotic lung disease in mice without affecting the onset of distress.

  8. Circulating Histones Are Mediators of Trauma-associated Lung Injury

    PubMed Central

    Abrams, Simon T.; Zhang, Nan; Manson, Joanna; Liu, Tingting; Dart, Caroline; Baluwa, Florence; Wang, Susan Siyu; Brohi, Karim; Kipar, Anja; Yu, Weiping

    2013-01-01

    Rationale: Acute lung injury is a common complication after severe trauma, which predisposes patients to multiple organ failure. This syndrome largely accounts for the late mortality that arises and despite many theories, the pathological mechanism is not fully understood. Discovery of histone-induced toxicity in mice presents a new dimension for elucidating the underlying pathophysiology. Objectives: To investigate the pathological roles of circulating histones in trauma-induced lung injury. Methods: Circulating histone levels in patients with severe trauma were determined and correlated with respiratory failure and Sequential Organ Failure Assessment (SOFA) scores. Their cause–effect relationship was studied using cells and mouse models. Measurements and Main Results: In a cohort of 52 patients with severe nonthoracic blunt trauma, circulating histones surged immediately after trauma to levels that were toxic to cultured endothelial cells. The high levels were significantly associated with the incidence of acute lung injury and SOFA scores, as well as markers of endothelial damage and coagulation activation. In in vitro systems, histones damaged endothelial cells, stimulated cytokine release, and induced neutrophil extracellular trap formation and myeloperoxidase release. Cellular toxicity resulted from their direct membrane interaction and resultant calcium influx. In mouse models, cytokines and markers for endothelial damage and coagulation activation significantly increased immediately after trauma or histone infusion. Pathological examinations showed that lungs were the predominantly affected organ with edema, hemorrhage, microvascular thrombosis, and neutrophil congestion. An anti-histone antibody could reduce these changes and protect mice from histone-induced lethality. Conclusions: This study elucidates a new mechanism for acute lung injury after severe trauma and proposes that circulating histones are viable therapeutic targets for improving survival

  9. Neutrophils Infiltrate the Spinal Cord Parenchyma of Rats with Experimental Diabetic Neuropathy

    PubMed Central

    Newton, Victoria L.; Guck, Jonathan D.; Cotter, Mary A.

    2017-01-01

    Spinal glial cell activation and cytokine secretion have been implicated in the etiology of neuropathic pain in a number of experimental models, including diabetic neuropathy. In this study, streptozotocin- (STZ-) induced diabetic rats were either untreated or treated with gabapentin (50 mg/kg/day by gavage for 2 weeks, from 6 weeks after STZ). At 8 weeks after STZ, hypersensitivity was confirmed in the untreated diabetic rats as a reduced response threshold to touch, whilst mechanical thresholds in gabapentin-treated diabetic rats were no different from controls. Diabetes-associated thermal hypersensitivity was also ameliorated by gabapentin. We performed a cytokine profiling array in lumbar spinal cord samples from control and diabetic rats. This revealed an increase in L-selectin, an adhesion molecule important for neutrophil transmigration, in the spinal cord of diabetic rats but not diabetic rats treated with gabapentin. Furthermore, we found an increase in the number of neutrophils present in the parenchyma of the spinal cord, which was again ameliorated in gabapentin-treated diabetic rats. Therefore, we suggest that dysregulated spinal L-selectin and neutrophil infiltration into the spinal cord could contribute to the pathogenesis of painful diabetic neuropathy. PMID:28293643

  10. Science review: Cell membrane expression (connectivity) regulates neutrophil delivery, function and clearance

    PubMed Central

    Seely, Andrew JE; Pascual, José L; Christou, Nicolas V

    2003-01-01

    As the principal cellular component of the inflammatory host defense and contributor to host injury after severe physiologic insult, the neutrophil is inherently coupled to patient outcome in both health and disease. Extensive research has focused on the mechanisms that regulate neutrophil delivery, function, and clearance from the inflammatory microenvironment. The neutrophil cell membrane mediates the interaction of the neutrophil with the extracellular environment; it expresses a complex array of adhesion molecules and receptors for various ligands, including mediators, cytokines, immunoglobulins, and membrane molecules on other cells. This article presents a review and analysis of the evidence that the neutrophil membrane plays a central role in regulating neutrophil delivery (production, rolling, adhesion, diapedesis, and chemotaxis), function (priming and activation, microbicidal activity, and neutrophil-mediated host injury), and clearance (apoptosis and necrosis). In addition, we review how change in neutrophil membrane expression is synonymous with change in neutrophil function in vivo. Employing a complementary analysis of the neutrophil as a complex system, neutrophil membrane expression may be regarded as a measure of neutrophil connectivity, with altered patterns of connectivity representing functionally distinct neutrophil states. Thus, not only does the neutrophil membrane mediate the processes that characterize the neutrophil lifecycle, but characterization of neutrophil membrane expression represents a technology with which to evaluate neutrophil function. PMID:12930553

  11. Inactivated pepsin inhibits neutrophil activation by Fcgamma-receptor-dependent and independent stimuli.

    PubMed

    Kustiawan, Iwan; Derksen, Ninotska; Rispens, Theo

    2016-08-01

    Pepsin is widely used to produce F(ab')2 fragments of immunoglobulin G (IgG). In many cases, at least part of the pepsin will remain present in the F(ab')2 preparation, albeit in (irreversibly) inactivated form. Here we report on a potent immunomodulatory effect of irreversibly inactivated pepsin on activated human neutrophils. Degranulation, induced by coated IgG or via cytochalasin B/N-formyl-Met-Leu-Phe, was measured by quantifying elastase release, and was found to be inhibited in a dose-dependent manner by inactivated pepsin. Since a number of intravenous immunoglobulin (IVIg) products are also treated by limited digestion with pepsin, we investigated if pepsin would be present in quantities large enough to inhibit neutrophil activation. The amounts of pepsin detected in three different pepsin-treated IVIg products were found to be too low to induce an effect, at least in an in vitro setting.

  12. Interaction of Bacterial Exotoxins with Neutrophil Extracellular Traps: Impact for the Infected Host

    PubMed Central

    von Köckritz-Blickwede, Maren; Blodkamp, Stefanie; Nizet, Victor

    2016-01-01

    Since their discovery in 2004, neutrophil extracellular traps (NETs) have been characterized as a fundamental host innate immune defense against various pathogens. Released in response to infectious and pro-inflammatory stimuli, NETs can immobilize invading pathogens within a fibrous matrix consisting of DNA, histones, and antimicrobial peptides. Conversely, excessive or dysregulated NET release may hold a variety of detrimental consequences for the host. A fine balance between NET formation and elimination is necessary to sustain a protective effect during infectious challenge. In recent years, a number of microbial virulence factors have been shown to modulate formation of NETs, thereby facilitating colonization or spread within the host. In this mini-review we summarize the contemporary research on the interaction of bacterial exotoxins with neutrophils that modulate NET production, focusing particular attention on consequences for the host. Understanding host–pathogen dynamics in this extracellular battlefield of innate immunity may provide novel therapeutic approaches for infectious and inflammatory disorders. PMID:27064864

  13. Borrelia burgdorferi upregulates expression of adhesion molecules on endothelial cells and promotes transendothelial migration of neutrophils in vitro.

    PubMed Central

    Sellati, T J; Burns, M J; Ficazzola, M A; Furie, M B

    1995-01-01

    The accumulation of leukocytic infiltrates in perivascular tissues is a key step in the pathogenesis of Lyme disease, a chronic inflammatory disorder caused by Borrelia burgdorferi. During an inflammatory response, endothelial cell adhesion molecules mediate the attachment of circulating leukocytes to the blood vessel wall and their subsequent extravasation into perivascular tissues. Using cultured human umbilical vein endothelial cells (HUVEC) in a whole-cell enzyme-linked immunosorbent assay, we demonstrated that B. burgdorferi activated endothelium in a dose- and time-dependent fashion as measured by upregulation of the adhesion molecules E-selectin, vascular cell adhesion molecule 1 (VCAM-1), and intercellular adhesion molecule 1 (ICAM-1). As few as one spirochete per endothelial cell stimulated increased expression of these molecules. Expression of E-selectin peaked after spirochetes and HUVEC were coincubated for 4 h and returned to near-basal levels by 24 h. In contrast, expression of VCAM-1 and ICAM-1 peaked at 12 h and remained elevated at 24 h. HUVEC monolayers cultured on acellular amniotic tissue were used to investigate the consequences of endothelial cell activation by spirochetes. After incubation of HUVEC-amnion cultures with B. burgdorferi, subsequently added neutrophils migrated across the endothelial monolayers. This process was mediated by E-selectin and by CD11/CD18 leukocytic integrins. The extent of migration depended on both the number of spirochetes used to stimulate the HUVEC and the length of the coincubation period. These results raise the possibility that B. burgdorferi induces a host inflammatory response and accompanying perivascular damage through activation of vascular endothelium. PMID:7591083

  14. Elevated fecal calprotectin levels during necrotizing enterocolitis are associated with activated neutrophils extruding neutrophil extracellular traps

    PubMed Central

    MacQueen, BC; Christensen, RD; Yost, CC; Lambert, DK; Baer, VL; Sheffield, MJ; Gordon, PV; Cody, MJ; Gerday, E; Schlaberg, R; Lowe, J; Shepherd, JG

    2016-01-01

    BACKGROUND Neonates with necrotizing enterocolitis (NEC) have higher calprotectin levels in stool than do healthy neonates. However, it is not known whether high stool calprotectin at the onset of bowel symptoms identifies neonates who truly have NEC vs. other bowel disorders. STUDY DESIGN Neonates were eligible for this study when an x-ray was ordered to “rule-out NEC”. Stool calprotectin was quantified at that time and in a follow-up stool. Each episode was later categorized as NEC or not NEC. The location of calprotectin in the bowel was determined by immunohistochemistry. RESULTS Neonates with NEC had higher initial and follow-up stool calprotectin levels than did neonates without NEC. Calprotectin in bowel from neonates with NEC was within neutrophil extracellular traps (NETs). CONCLUSION At the onset of signs concerning for NEC, fecal calprotectin is likely to be higher in neonates with NEC. Calprotectin in their stools is exported from neutrophils via NETs. PMID:27388941

  15. Matrix-free constructions of circulant and block circulant preconditioners

    SciTech Connect

    Yang, Chao; Ng, Esmond G.; Penczek, Pawel A.

    2001-12-01

    A framework for constructing circulant and block circulant preconditioners (C) for a symmetric linear system Ax=b arising from certain signal and image processing applications is presented in this paper. The proposed scheme does not make explicit use of matrix elements of A. It is ideal for applications in which A only exists in the form of a matrix vector multiplication routine, and in which the process of extracting matrix elements of A is costly. The proposed algorithm takes advantage of the fact that for many linear systems arising from signal or image processing applications, eigenvectors of A can be well represented by a small number of Fourier modes. Therefore, the construction of C can be carried out in the frequency domain by carefully choosing its eigenvalues so that the condition number of C{sup T} AC can be reduced significantly. We illustrate how to construct the spectrum of C in a way such that the smallest eigenvalues of C{sup T} AC overlaps with those of A extremely well while the largest eigenvalues of C{sup T} AC are smaller than those of A by several orders of magnitude. Numerical examples are provided to demonstrate the effectiveness of the preconditioner on accelerating the solution of linear systems arising from image reconstruction application.

  16. Diversity of Planetary Atmospheric Circulations and Climates in a Simplified General Circulation Model

    NASA Astrophysics Data System (ADS)

    Wang, Yixiong; Read, Peter

    2014-04-01

    The parametric dependence of terrestrial planetary atmospheric circulations and climates on characteristic parameters is studied. A simplified general circulation model-PUMA is employed to investigate the dynamic effects of planetary rotation rate and equator-to-pole temperature difference on the circulation and climate of terrestrial planetary atmospheres. Five different types of circulation regime are identified by mapping the experimental results in a 2-D parameter space defined by thermal Rossby number and frictional Taylor number. The effect of the transfer and redistribution of radiative energy is studied by building up a new two-band semi-gray radiative-convective scheme, which is capable of modelling greenhouse and anti-greenhouse effects while keeping the tunable parameters as few as possible. The results will provide insights into predicting the habitability of terrestrial exoplanets.

  17. Neutrophil-to-lymphocyte ratio: an inflammation marker related to cardiovascular risk in children.

    PubMed

    Prats-Puig, Anna; Gispert-Saüch, Montserrat; Díaz-Roldán, Ferran; Carreras-Badosa, Gemma; Osiniri, Inés; Planella-Colomer, Montserrat; Mayol, Lluís; de Zegher, Francis; Ibánez, Lourdes; Bassols, Judit; López-Bermejo, Abel

    2015-10-01

    Low-grade chronic inflammation plays a pathogenic role in cardiovascular disease. An increase in the ratio of circulating neutrophils to lymphocytes (N/L ratio) may serve as a marker of cardiovascular risk in adults. It was the study objective to study whether N/L ratio associates with vascular parameters in children. Subjects were 501 prepubertal and early pubertal Caucasian children (mean age 8.0 years; mean body mass index (BMI) Z-score 0.2 ± 0.9; 266 boys and 235 girls) recruited within an ongoing population-based study. The subjects were stratified into three groups according to age. Neutrophil, lymphocyte, BMI, waist circumference, systolic blood pressure (SBP) and carotid intima-media thickness (cIMT), assessed in all children. The N/L ratio, derived from the absolute neutrophil and lymphocyte counts. In children aged < 7 years (n=190, all prepubertal), no associations were observed between N/L ratio and either anthropometric or cardiovascular parameters. In children aged 7-9 years (n=171, 1.7% early pubertal), higher N/L ratio associated with higher BMI Z-score and waist circumference (p=0.008 to p < 0.0001). In children aged >9 years (n=140, 29.2% early pubertal), N/L ratio associated again with BMI Z-score and waist circumference and also positively with SBP and cIMT (all p=0.008 to p<0.0001). These associations remained significant in linear regression models following adjustment for possible confounding variables such as age, gender, fasting triglycerides, C-reactive protein and puberty (and for SBP and cIMT, adjustment also for BMI). In conclusion, our results provide the first evidence that a higher N/L ratio is associated with a less favourable cardiovascular profile in children and delineate the development of these associations from late childhood onwards.

  18. Dermal Neutrophil, Macrophage and Dendritic Cell Responses to Yersinia pestis Transmitted by Fleas

    PubMed Central

    Shannon, Jeffrey G.; Bosio, Christopher F.; Hinnebusch, B. Joseph

    2015-01-01

    Yersinia pestis, the causative agent of plague, is typically transmitted by the bite of an infected flea. Many aspects of mammalian innate immune response early after Y. pestis infection remain poorly understood. A previous study by our lab showed that neutrophils are the most prominent cell type recruited to the injection site after intradermal needle inoculation of Y. pestis, suggesting that neutrophil interactions with Y. pestis may be important in bubonic plague pathogenesis. In the present study, we developed new tools allowing for intravital microscopy of Y. pestis in the dermis of an infected mouse after transmission by its natural route of infection, the bite of an infected flea. We found that uninfected flea bites typically induced minimal neutrophil recruitment. The magnitude of neutrophil response to flea-transmitted Y. pestis varied considerably and appeared to correspond to the number of bacteria deposited at the bite site. Macrophages migrated towards flea bite sites and interacted with small numbers of flea-transmitted bacteria. Consistent with a previous study, we observed minimal interaction between Y. pestis and dendritic cells; however, dendritic cells did consistently migrate towards flea bite sites containing Y. pestis. Interestingly, we often recovered viable Y. pestis from the draining lymph node (dLN) 1 h after flea feeding, indicating that the migration of bacteria from the dermis to the dLN may be more rapid than previously reported. Overall, the innate cellular host responses to flea-transmitted Y. pestis differed from and were more variable than responses to needle-inoculated bacteria. This work highlights the importance of studying the interactions between fleas, Y. pestis and the mammalian host to gain a better understanding of the early events in plague pathogenesis. PMID:25781984

  19. Dermal neutrophil, macrophage and dendritic cell responses to Yersinia pestis transmitted by fleas.

    PubMed

    Shannon, Jeffrey G; Bosio, Christopher F; Hinnebusch, B Joseph

    2015-03-01

    Yersinia pestis, the causative agent of plague, is typically transmitted by the bite of an infected flea. Many aspects of mammalian innate immune response early after Y. pestis infection remain poorly understood. A previous study by our lab showed that neutrophils are the most prominent cell type recruited to the injection site after intradermal needle inoculation of Y. pestis, suggesting that neutrophil interactions with Y. pestis may be important in bubonic plague pathogenesis. In the present study, we developed new tools allowing for intravital microscopy of Y. pestis in the dermis of an infected mouse after transmission by its natural route of infection, the bite of an infected flea. We found that uninfected flea bites typically induced minimal neutrophil recruitment. The magnitude of neutrophil response to flea-transmitted Y. pestis varied considerably and appeared to correspond to the number of bacteria deposited at the bite site. Macrophages migrated towards flea bite sites and interacted with small numbers of flea-transmitted bacteria. Consistent with a previous study, we observed minimal interaction between Y. pestis and dendritic cells; however, dendritic cells did consistently migrate towards flea bite sites containing Y. pestis. Interestingly, we often recovered viable Y. pestis from the draining lymph node (dLN) 1 h after flea feeding, indicating that the migration of bacteria from the dermis to the dLN may be more rapid than previously reported. Overall, the innate cellular host responses to flea-transmitted Y. pestis differed from and were more variable than responses to needle-inoculated bacteria. This work highlights the importance of studying the interactions between fleas, Y. pestis and the mammalian host to gain a better understanding of the early events in plague pathogenesis.

  20. Ocean circulation using altimetry

    NASA Technical Reports Server (NTRS)

    Minster, Jean-Francois; Brossier, C.; Gennero, M. C.; Mazzega, P.; Remy, F.; Letraon, P. Y.; Blanc, F.

    1991-01-01

    Our group has been very actively involved in promoting satellite altimetry as a unique tool for observing ocean circulation and its variability. TOPEX/POSEIDON is particularly interesting as it is optimized for this purpose. It will probably be the first instrument really capable of observing the seasonal and interannual variability of subtropical and polar gyres and the first to eventually document the corresponding variability of their heat flux transport. The studies of these phenomena require data of the best quality, unbiased extraction of the signal, mixing of these satellite data with in situ measurements, and assimilation of the whole set into a dynamic description of ocean circulation. Our group intends to develop responses to all these requirements. We will concentrate mostly on the circulation of the South Atlantic and Indian Oceans: This will be done in close connection with other groups involved in the study of circulation of the tropical Atlantic Ocean, in the altimetry measurements (in particular, those of the tidal issue), and in the techniques of data assimilation in ocean circulation models.

  1. Chemotactic and Phagocytic Activity of Blood Neutrophils in Allergic Asthma.

    PubMed

    Mosca, Tainá; Menezes, Maria C S; Silva, Ademir Veras; Stirbulov, Roberto; Forte, Wilma C N

    2015-01-01

    Allergic asthma is a chronic inflammatory airway disease, and has been considered a T helper-2-biased response. Studies suggest that neutrophils may be associated with exacerbation and asthma severity. We sought to evaluate the chemotactic activity and phagocytic capacity by peripheral blood neutrophils from individuals with controlled and uncontrolled allergic asthma, and compare the results with non-asthmatic controls groups. Blood neutrophils were isolated from 95 patients: 24 with controlled asthma, 24 uncontrolled asthma, 24 healthy subjects and 23 patients with IgE-mediated allergies other than asthma. The neutrophil chemotaxis, stimulated with LPS, autologous serum or homologous serum, was determined using Boyden chambers. The phagocytic capacity was assessed by ingestion of zimosan particles, and digestion phase was analyzed by NBT test. The phagocytic digestion phase and chemotaxis by neutrophils from asthmatic patients was higher than in non-asthmatic controls (p  < 0.05). Autologous serum-induced neutrophil chemotaxis in patients with uncontrolled asthma was greater (p  < 0.05) than in other study groups. The ingestion phase of phagocytosis showed similar values in asthmatics and non-asthmatics. We conclude that the blood neutrophil from controlled and uncontrolled asthmatic patients exhibit activation markers, particularly phagocytic digestion and chemotactic activities.

  2. Potentiation and inhibition of migration of human neutrophils by auranofin.

    PubMed Central

    Elferink, J G; de Koster, B M

    1993-01-01

    OBJECTIVES--As auranofin resembles some neutrophil activating sulphur containing compounds, it was decided to investigate whether it had activating effects on neutrophil migration in addition to the published inhibitory effects. METHODS--The Boyden chamber assay was used to determine the migration velocity of human neutrophils. The difference between chemotaxis and chemokinesis was established with a chequerboard assay. RESULTS--Low concentrations of auranofin stimulated human neutrophil migration; concentrations of auranofin higher than 1 mumol/l were inhibitory. Inhibitors of leukotriene formation, or of protein kinase C, had the same effect on auranofin induced potentiation of migration as on fMLP activated migration. Auranofin, at a concentration of 100 nmol/l, caused a transient increase in the cGMP level of neutrophils. The auranofin induced increase in migration was strongly inhibited by methylene blue and by LY83583, two inhibitors of cGMP accumulation. CONCLUSIONS--The auranofin induced enhancement of migration is partly due to a chemokinetic effect, but mainly due to a chemotactic effect. The potentiating effect of auranofin on migration is not specifically due to the ability of the drug to inhibit protein kinase C activity or to generate leukotrienes. These results suggest that the enhancement of neutrophil migration by low levels of auranofin is related to the enhancement of cGMP levels in neutrophils. PMID:8215623

  3. Flow cytometric study of in vitro neutrophil activation by biomaterials.

    PubMed

    Gorbet, M B; Yeo, E L; Sefton, M V

    1999-03-05

    Neutrophil activation for adherent and nonadherent cells, as measured by flow cytometry, was not strongly dependent on material surface chemistry. We had hypothesized that material-induced neutrophil activation was an important parameter associated with material failure. All materials tested [cellophane, an acrylonitrile copolymer (AN69), Pellethane, nylon, polyethylene terephthalate, low density polyethylene, and polydimethylsiloxane] activated isolated human neutrophils, which were resuspended in plasma or serum, to similar extents based on L-selectin shedding, CD11b upregulation, and stimulation of the oxidative burst after 30-min exposure. Inhibition of complement activation by sCR1 unexpectedly had little effect if any on nonadherent neutrophils. However, neutrophil adhesion, but not the level of activation of the adherent cells, was strongly dependent on complement activation. Pretreatment with albumin did not inhibit adhesion or reduce neutrophil activation, but plasma pretreatment resulted in increased activation for nonadherent and adherent cells. More adhesion and a higher level of activation of adherent cells was observed following pretreatment with fibrinogen, a ligand of CD11b. Taken together these results suggest that upon contact with a material, neutrophil activation may occur though mechanisms that are not mediated by complement. For example, the presence of plasma proteins such as fibrinogen at the interface may trigger activation and the release of other activating agents. Although the material differences are small, the extent of activation may be significant and warrant further study of the mechanism and consequences of that activation.

  4. Constitutive apoptosis in equine peripheral blood neutrophils in vitro

    PubMed Central

    Brazil, Timothy J.; Dixon, Padraic M.; Haslett, Christopher; Murray, Joanna; McGorum, Bruce C.

    2014-01-01

    The aim of this study was to characterise constitutive apoptosis in equine peripheral blood neutrophils, including assessment of factors that potentially modulate neutrophil survival through alteration of the rate of constitutive apoptosis. Cells underwent spontaneous time-dependent constitutive apoptosis when aged in culture for up to 36 h, developing the structural and functional features of apoptosis observed in many cell types, including human neutrophils. Neutrophils undergoing apoptosis also had diminished zymosan activated serum (ZAS)-stimulated chemiluminescence, but maintained responsiveness to phorbol myristate acetate (PMA). The constitutive rate of equine neutrophil apoptosis was promoted by lipopolysaccharide (LPS), tumour necrosis factor α and phagocytosis of opsonised ovine erythrocytes, while it was inhibited by dexamethasone and ZAS (a source of C5a). Formyl-Met-Leu-Phe, leukotriene B4, platelet activating factor and PMA had no demonstrable effect on equine neutrophil apoptosis. There was a difference between equine and human neutrophil apoptosis in response to LPS and the time-dependence of the response to dexamethasone. PMID:25239298

  5. Constitutive apoptosis in equine peripheral blood neutrophils in vitro.

    PubMed

    Brazil, Timothy J; Dixon, Padraic M; Haslett, Christopher; Murray, Joanna; McGorum, Bruce C

    2014-12-01

    The aim of this study was to characterise constitutive apoptosis in equine peripheral blood neutrophils, including assessment of factors that potentially modulate neutrophil survival through alteration of the rate of constitutive apoptosis. Cells underwent spontaneous time-dependent constitutive apoptosis when aged in culture for up to 36 h, developing the structural and functional features of apoptosis observed in many cell types, including human neutrophils. Neutrophils undergoing apoptosis also had diminished zymosan activated serum (ZAS)-stimulated chemiluminescence, but maintained responsiveness to phorbol myristate acetate (PMA). The constitutive rate of equine neutrophil apoptosis was promoted by lipopolysaccharide (LPS), tumour necrosis factor α and phagocytosis of opsonised ovine erythrocytes, while it was inhibited by dexamethasone and ZAS (a source of C5a). Formyl-Met-Leu-Phe, leukotriene B4, platelet activating factor and PMA had no demonstrable effect on equine neutrophil apoptosis. There was a difference between equine and human neutrophil apoptosis in response to LPS and the time-dependence of the response to dexamethasone.

  6. Review of the neutrophil response to Bordetella pertussis infection.

    PubMed

    Eby, Joshua C; Hoffman, Casandra L; Gonyar, Laura A; Hewlett, Erik L

    2015-12-01

    The nature and timing of the neutrophil response to infection with Bordetella pertussis is influenced by multiple virulence factors expressed by the bacterium. After inoculation of the host airway, the recruitment of neutrophils signaled by B. pertussis lipooligosaccharide (LOS) is suppressed by pertussis toxin (PTX). Over the next week, the combined activities of PTX, LOS and adenylate cyclase toxin (ACT) result in production of cytokines that generate an IL-17 response, promoting neutrophil recruitment which peaks at 10-14 days after inoculation in mice. Arriving at the site of infection, neutrophils encounter the powerful local inhibitory activity of ACT, in conjunction with filamentous hemagglutinin. With the help of antibodies, neutrophils contribute to clearance of B. pertussis, but only after 28-35 days in a naïve mouse. Studies of the lasting, antigen-specific IL-17 response to infection in mice and baboons has led to progress in vaccine development and understanding of pathogenesis. Questions remain about the mediators that coordinate neutrophil recruitment and the mechanisms by which neutrophils overcome B. pertussis virulence factors.

  7. Review of the neutrophil response to Bordetella pertussis infection

    PubMed Central

    Eby, Joshua C.; Hoffman, Casandra L.; Gonyar, Laura A.; Hewlett, Erik L.

    2015-01-01

    The nature and timing of the neutrophil response to infection with Bordetella pertussis is influenced by multiple virulence factors expressed by the bacterium. After inoculation of the host airway, the recruitment of neutrophils signaled by B. pertussis lipooligosaccharide (LOS) is suppressed by pertussis toxin (PTX). Over the next week, the combined activities of PTX, LOS and adenylate cyclase toxin (ACT) result in production of cytokines that generate an IL-17 response, promoting neutrophil recruitment which peaks at 10–14 days after inoculation in mice. Arriving at the site of infection, neutrophils encounter the powerful local inhibitory activity of ACT, in conjunction with filamentous hemagglutinin. With the help of antibodies, neutrophils contribute to clearance of B. pertussis, but only after 28–35 days in a naïve mouse. Studies of the lasting, antigen-specific IL-17 response to infection in mice and baboons has led to progress in vaccine development and understanding of pathogenesis. Questions remain about the mediators that coordinate neutrophil recruitment and the mechanisms by which neutrophils overcome B. pertussis virulence factors. PMID:26432818

  8. Age-Appropriate Functions and Dysfunctions of the Neonatal Neutrophil

    PubMed Central

    Lawrence, Shelley Melissa; Corriden, Ross; Nizet, Victor

    2017-01-01

    Neonatal and adult neutrophils are distinctly different from one another due to well-defined and documented deficiencies in neonatal cells, including impaired functions, reduced concentrations of microbicidal proteins and enzymes necessary for pathogen destruction, and variances in cell surface receptors. Neutrophil maturation is clearly demonstrated throughout pregnancy from the earliest hematopoietic precursors in the yolk sac to the well-developed myeloid progenitor cells in the bone marrow around the seventh month of gestation. Notable deficiencies of neonatal neutrophils are generally correlated with gestational age and clinical condition, so that the least functional neutrophils are found in the youngest, sickest neonates. Interruption of normal gestation secondary to preterm birth exposes these shortcomings and places the neonate at an exceptionally high rate of infection and sepsis-related mortality. Because the fetus develops in a sterile environment, neonatal adaptive immune responses are deficient from lack of antigen exposure in utero. Newborns must therefore rely on innate immunity to protect against early infection. Neutrophils are a vital component of innate immunity since they are the first cells to respond to and defend against bacterial, viral, and fungal infections. However, notable phenotypic and functional disparities exist between neonatal and adult cells. Below is review of neutrophil ontogeny, as well as a discussion regarding known differences between preterm and term neonatal and adult neutrophils with respect to cell membrane receptors and functions. Our analysis will also explain how these variations decrease with postnatal age. PMID:28293548

  9. Alterations in Circulating Immune Cells in Neovascular Age-Related Macular Degeneration.

    PubMed

    Lechner, Judith; Chen, Mei; Hogg, Ruth E; Toth, Levente; Silvestri, Giuliana; Chakravarthy, Usha; Xu, Heping

    2015-11-17

    Neovascular age-related macular degeneration (nAMD) is the leading cause of irreversible blindness in developed countries. Recent advances have highlighted the essential role of inflammation in the development of the disease. In addition to local retinal chronic inflammatory response, systemic immune alterations have also been observed in AMD patients. In this study we investigated the association between the frequency of circulating leukocyte populations and the prevalence as well as clinical presentations of nAMD. Leukocyte subsets of 103 nAMD patients (most of them were receiving anti-VEGF therapy prior to enrolment) and 26 controls were analysed by flow cytometry by relative cell size, granularity and surface markers. Circulating CD11b(+) cells and CD16(hi)HLA-DR(-) neutrophils were significantly increased (P = 0.015 and 0.009 respectively) in nAMD when compared to controls. The percentage of circulating CD4(+) T-cells was reduced in nAMD patients without subretinal fibrosis (P = 0.026) compared to patients with subretinal fibrosis. There was no correlation between the percentage of circulating leukocytes and the responsiveness to anti-VEGF therapy in nAMD patients. Our results suggest that higher levels of circulating CD11b(+) cells and neutrophils are associated with nAMD and that reduced levels of CD4(+) T-cells are associated with the absence of subretinal fibrosis in nAMD.

  10. Chorionic plate vessels as an origin of amniotic fluid neutrophils.

    PubMed

    Lee, Soong Deok; Kim, Mi Ran; Hwang, Pil Gyu; Shim, Soon-Sup; Yoon, Bo Hyun; Kim, Chong Jai

    2004-07-01

    The present study was conducted to investigate the potential anatomical source of amniotic fluid neutrophils. Microdissection of neutrophils from the chorioamnion of the fetal membranes and the amnion of the chorionic plates of 10 preterm placentas with acute chorioamnionitis was performed and the genotypes of the neutrophils were compared with those of the mother and fetus using polymerase chain reaction of nine autosomal STR loci. In separate analyses, we reviewed eight cases of fetal autopsies with increased amniotic fluid neutrophils for the presence of neutrophils in the alveoli, and also analyzed the relationship between the amniotic fluid white blood cell (WBC) count and the histological pattern of placental inflammation. The genotypes of all of the neutrophils found in the chorioamnion of the fetal membrane matched those of the mother (n = 10). The genotypes of neutrophils found in the chorionic plate were of mixed maternal and fetal origin (n = 4). In the autopsy series of the fetuses with amniotic fluid WBC (n = 8), only five cases showed neutrophils in the alveolar space, while all the placentas had chorioamnionitis. There was no significant difference in amniotic fluid WBC count between the cases with or without acute membranitis, while among the cases with placental inflammation, those with inflammation of the chorionic plate had a significantly higher amniotic fluid WBC count than both the membranitis-only cases (P < 0.001) and the membranitis and funisitis cases (P < 0.05). These results imply that fetal vasculature at the chorionic plate is the main source of amniotic fluid neutrophils, especially in the cases without funisitis.

  11. Evidence for chemokine synergy during neutrophil migration in ARDS

    PubMed Central

    Williams, Andrew E; José, Ricardo J; Mercer, Paul F; Brealey, David; Parekh, Dhruv; Thickett, David R; O'Kane, Cecelia; McAuley, Danny F; Chambers, Rachel C

    2017-01-01

    Background Acute respiratory distress syndrome (ARDS) is a life-threatening condition characterised by pulmonary oedema, respiratory failure and severe inflammation. ARDS is further characterised by the recruitment of neutrophils into the lung interstitium and alveolar space. Objectives The factors that regulate neutrophil infiltration into the inflamed lung and our understanding of the pathomechanisms in ARDS remain incomplete. This study aimed at determining the role of the chemokine (C-C motif) ligand (CCL)2 and CCL7 in ARDS. Methods CCL2 and CCL7 protein levels were measured in bronchoalveolar lavage (BAL) fluid obtained from lipopolysaccharide(LPS)-challenged human volunteers and two separate cohorts of patients with ARDS. Neutrophil chemotaxis to ARDS BAL fluid was evaluated and the contribution of each was assessed and compared with chemokine (C-X-C motif) ligand 8 (CXCL8). Chemokine receptor expression on neutrophils from blood or BAL fluid of patients with ARDS was analysed by flow cytometry. Results CCL2 and CCL7 were significantly elevated in BAL fluid recovered from LPS-challenged volunteers and patients with ARDS. BAL fluid from patients with ARDS was highly chemotactic for human neutrophils and neutralising either CCL2 or CCL7 attenuated the neutrophil chemotactic response. Moreover, CCL2 and CCL7 synergised with CXCL8 to promote neutrophil migration. Furthermore, neutrophils isolated from the blood or BAL fluid differentially regulated the cell surface expression of chemokine (C-X-C motif) receptor 1 and C-C chemokine receptor type 2 during ARDS. Conclusion This study highlights important inflammatory chemokines involved in regulating neutrophil migration, which may have potential value as therapeutic targets for the treatment of ARDS. PMID:27496101

  12. Neutrophil adhesion and chemotaxis depend on substrate mechanics

    NASA Astrophysics Data System (ADS)

    Jannat, Risat A.; Robbins, Gregory P.; Ricart, Brendon G.; Dembo, Micah; Hammer, Daniel A.

    2010-05-01

    Neutrophil adhesion to the vasculature and chemotaxis within tissues play critical roles in the inflammatory response to injury and pathogens. Unregulated neutrophil activity has been implicated in the progression of numerous chronic and acute diseases such as rheumatoid arthritis, asthma and sepsis. Cell migration of anchorage-dependent cells is known to depend on both chemical and mechanical interactions. Although neutrophil responses to chemical cues have been well characterized, little is known about the effect of underlying tissue mechanics on neutrophil adhesion and migration. To address this question, we quantified neutrophil migration and traction stresses on compliant hydrogel substrates with varying elasticity in a micromachined gradient chamber in which we could apply either a uniform concentration or a precise gradient of the bacterial chemoattractant fMLP. Neutrophils spread more extensively on substrates of greater stiffness. In addition, increasing the stiffness of the substrate leads to a significant increase in the chemotactic index for each fMLP gradient tested. As the substrate becomes stiffer, neutrophils generate higher traction forces without significant changes in cell speed. These forces are often displayed in pairs and focused in the uropod. Increases in the mean fMLP concentration beyond the KD of the receptor lead to a decrease in chemotactic index on all surfaces. Blocking with an antibody against β2-integrins leads to a significant reduction, but not an elimination, of directed motility on stiff materials, but no change in motility on soft materials, suggesting neutrophils can display both integrin-dependent and integrin-independent motility. These findings are critical for understanding how neutrophil migration may change in different mechanical environments in vivo and can be used to guide the design of migration inhibitors that more efficiently target inflammation.

  13. Comparison of neutrophil:lymphocyte ratios following coronary artery bypass surgery with or without cardiopulmonary bypass

    PubMed Central

    Aldemir, Mustafa; Adalı, Fahri; Çarşanba, Görkem; Tecer, Evren; Bakı, Elif Doğan; Taş, Hanife Uzel

    2015-01-01

    Objective Coronary artery bypass graft (CABG) surgery may induce postoperative systemic changes in leukocyte counts, including leukocytosis, neutrophilia or lymphopenia. This retrospective clinical study investigated whether offpump coronary artery bypass (OPCAB) surgery working on the beating heart without extracorporeal circulation could favourably affect leukocyte counts, including neutrophil-tolymphocyte (N:L) ratio, after CABG. Methods In this study, 30 patients who underwent isolated CABG with cardiopulmonary bypass (CPB), and another 30 patients who underwent the same operation without CPB between May 2010 and May 2013, were screened from the computerised database of our hospital. Pre-operative, and first and fifth postoperative day differential counts of leukocytes with the N:L ratio of peripheral blood were obtained. Results A significant increase in total leukocyte and neutrophil counts and N:L ratio, and a decrease in lymphocyte counts were observed at all time points after surgery in both groups. N:L ratio was significantly higher in the CPB group compared with the OPCAB group on the first postoperative day (20.73 ± 13.85 vs 10.19 ± 4.55, p < 0.001), but this difference disappeared on the fifth postoperative day. Conclusion CPB results in transient but significant changes in leukocyte counts in the peripheral blood stream in terms of N:L ratio compared with the off-pump technique of CABG. PMID:25903477

  14. Oxidative stress, superoxide production, and apoptosis of neutrophils in dogs with chronic kidney disease.

    PubMed

    Silva, Adriana Carolina Rodrigues Almeida; de Almeida, Breno Fernando Martins; Soeiro, Carolina Soares; Ferreira, Wagner Luis; de Lima, Valéria Marçal Félix; Ciarlini, Paulo César

    2013-04-01

    Oxidative stress is a key component in the immunosuppression of chronic kidney disease (CKD), and neutrophil function may be impaired by oxidative stress. To test the hypothesis that in uremic dogs with CKD, oxidative stress is increased and neutrophils become less viable and functional, 18 adult dogs with CKD were compared with 15 healthy adult dogs. Blood count and urinalysis were done, and the serum biochemical profile and plasma lipid peroxidation (measurement of thiobarbituric acid reactive substances) were determined with the use of commercial reagents. Plasma total antioxidant capacity (TAC) was measured with a spectrophotometer and commercial reagents, superoxide production with a hydroethidine probe, and the viability and apoptosis of neutrophils with capillary flow cytometry and the annexin V-PE system. The plasma concentrations of cholesterol (P = 0.0415), creatinine (P < 0.0001), and urea (P < 0.0001) were significantly greater in the uremic dogs than in the control dogs. The hematocrit (P = 0.0004), urine specific gravity (P = 0.015), and plasma lipid peroxidation (P < 0.0001) were significantly lower in the dogs that were in late stages of CKD than in the control group. Compared with those isolated from the control group, neutrophils isolated from the CKD group showed a higher rate of spontaneous (0.10 ± 0.05 versus 0.49 ± 0.09; P = 0.0033; median ± standard error of mean) and camptothecin-induced (18.53 ± 4.06 versus 44.67 ± 4.85; P = 0.0066) apoptosis and lower levels of superoxide production in the presence (1278.8 ± 372.8 versus 75.65 ± 86.6; P = 0.0022) and absence (135.29 ± 51.74 versus 41.29 ± 8.38; P = 0.0138) of phorbol-12-myristate-13-acetate stimulation. Thus, oxidative stress and acceleration of apoptosis occurs in dogs with CKD, the apoptosis diminishing the number of viable neutrophils and neutrophil superoxide production.

  15. Oxidative stress, superoxide production, and apoptosis of neutrophils in dogs with chronic kidney disease

    PubMed Central

    Silva, Adriana Carolina Rodrigues Almeida; de Almeida, Breno Fernando Martins; Soeiro, Carolina Soares; Ferreira, Wagner Luis; de Lima, Valéria Marçal Félix; Ciarlini, Paulo César

    2013-01-01

    Oxidative stress is a key component in the immunosuppression of chronic kidney disease (CKD), and neutrophil function may be impaired by oxidative stress. To test the hypothesis that in uremic dogs with CKD, oxidative stress is increased and neutrophils become less viable and functional, 18 adult dogs with CKD were compared with 15 healthy adult dogs. Blood count and urinalysis were done, and the serum biochemical profile and plasma lipid peroxidation (measurement of thiobarbituric acid reactive substances) were determined with the use of commercial reagents. Plasma total antioxidant capacity (TAC) was measured with a spectrophotometer and commercial reagents, superoxide production with a hydroethidine probe, and the viability and apoptosis of neutrophils with capillary flow cytometry and the annexin V-PE system. The plasma concentrations of cholesterol (P = 0.0415), creatinine (P < 0.0001), and urea (P < 0.0001) were significantly greater in the uremic dogs than in the control dogs. The hematocrit (P = 0.0004), urine specific gravity (P = 0.015), and plasma lipid peroxidation (P < 0.0001) were significantly lower in the dogs that were in late stages of CKD than in the control group. Compared with those isolated from the control group, neutrophils isolated from the CKD group showed a higher rate of spontaneous (0.10 ± 0.05 versus 0.49 ± 0.09; P = 0.0033; median ± standard error of mean) and camptothecin-induced (18.53 ± 4.06 versus 44.67 ± 4.85; P = 0.0066) apoptosis and lower levels of superoxide production in the presence (1278.8 ± 372.8 versus 75.65 ± 86.6; P = 0.0022) and absence (135.29 ± 51.74 versus 41.29 ± 8.38; P = 0.0138) of phorbol-12-myristate-13-acetate stimulation. Thus, oxidative stress and acceleration of apoptosis occurs in dogs with CKD, the apoptosis diminishing the number of viable neutrophils and neutrophil superoxide production. PMID:24082406

  16. On the North Atlantic circulation

    SciTech Connect

    Schmitz, W.J. Jr.; McCartney, M.S. )

    1993-02-01

    A summary for North Atlantic circulation is proposed to replace the circulation scheme hypothesized by Worthington in 1976. Divergences from the previous model are in thermohaline circulation, cross-equatorical transport and Florida Current sources, flow in the eastern Atlantic, circulation in the Newfoundland Basin, slope water currents, and flow pattern near the Bahamas. The circulation patterns presented here are consistent with the majority of of published accounts of flow components. 77 refs., 14 figs., 3 tabs.

  17. Measurement of Phenotype and Absolute Number of Circulating Heparin-Binding Hemagglutinin, ESAT-6 and CFP-10, and Purified Protein Derivative Antigen-Specific CD4 T Cells Can Discriminate Active from Latent Tuberculosis Infection

    PubMed Central

    Barkham, Timothy M. S.; Tang, Wenying; Kemeny, David M.; Chee, Cynthia Bin-Eng; Wang, Yee T.

    2014-01-01

    The tuberculin skin test (TST) and interferon gamma (IFN-γ) release assays (IGRAs) are used as adjunctive tests for the evaluation of suspected cases of active tuberculosis (TB). However, a positive test does not differentiate latent from active TB. We investigated whether flow cytometric measurement of novel combinations of intracellular cytokines and surface makers on CD4 T cells could differentiate between active and latent TB after stimulation with Mycobacterium tuberculosis-specific proteins. Blood samples from 60 patients referred to the Singapore Tuberculosis Control Unit for evaluation for active TB or as TB contacts were stimulated with purified protein derivative (PPD), ESAT-6 and CFP-10, or heparin-binding hemagglutinin (HBHA). The CD4 T cell cytokine response (IFN-γ, interleukin-2 [IL-2], interleukin-17A [IL-17A], interleukin-22 [IL-22], granulocyte-macrophage colony-stimulating factor [GM-CSF], and tumor necrosis factor alpha [TNF-α]) and surface marker expression (CD27, CXCR3, and CD154) were then measured. We found that the proportion of PPD-specific CD4 T cells, defined as CD154+ TNF-α+ cells that were negative for CD27 and positive for GM-CSF, gave the strongest discrimination between subjects with latent and those with active TB (area under the receiver operator characteristic [ROC] curve of 0.9277; P < 0.0001). Also, the proportions and absolute numbers of HBHA-specific CD4 T cells were significantly higher in those with latent TB infection, particularly CD154+ TNF-α+ IFN-γ+ IL-2+ and CD154+ TNF-α+ CXCR3+. Finally, we found that the ratio of ESAT-6- and CFP-10-responding to HBHA-responding CD4 T cells was significantly different between the two study populations. In conclusion, we found novel markers of M. tuberculosis-specific CD4 cells which differentiate between active and latent TB. PMID:25520147

  18. Quantifying and localizing actin-free barbed ends in neutrophils.

    PubMed

    Glogauer, Michael

    2007-01-01

    We describe here a permeablization method that retains coupling between N-formylmethionyl-leucyl-phenylalanine (fMLP) receptor stimulation and barbed-end actin nucleation in neutrophils. Using fluorescently-tagged actin monomers, we are able to quantify and localize actin-free barbed ends generated downstream of chemoattractant receptors. Partial permeabilization of the neutrophils with the mild detergent n-octyl-beta-glucopyranoside maintains signaling from membrane receptor to the actin cytoskeleton while allowing for the introduction of inhibitors and activators of signal transduction pathways implicated in regulating actin cytoskeleton dynamics. This is a useful assay for studying signal transduction to the actin cytoskeleton in neutrophils.

  19. Haem Biosynthesis and Antioxidant Enzymes in Circulating Cells of Acute Intermittent Porphyria Patients

    PubMed Central

    Ferrer, Miguel D.; Mestre-Alfaro, Antonia; Martínez-Tomé, Magdalena; Carrera-Quintanar, Lucrecia; Capó, Xavier; Jiménez-Monreal, Antonia M.; García-Diz, Luis; Roche, Enrique; Murcia, María A.; Tur, Josep A.

    2016-01-01

    The aims of the present study were to explore the expression pattern of haem biosynthesis enzymes in circulating cells of patients affected by two types of porphyria (acute intermittent, AIP, and variegate porphyria, VP), together with the antioxidant enzyme pattern in AIP in order to identify a possible situation of oxidative stress. Sixteen and twelve patients affected by AIP and VP, respectively, were analysed with the same numbers of healthy matched controls. Erythrocytes, neutrophils and peripheral blood mononuclear cells (PBMCs) were purified from blood, and RNA and proteins were extracted for quantitative real time PCR (qRT-PCR) and Western-blot analysis, respectively. Porhobilinogen deaminase (PBGD) and protoporphyrinogen oxidase (PPOX) gene and protein expression was analysed. Antioxidant enzyme activity and gene expression were additionally determined in blood cells, together with protein carbonyl content in plasma. PBMCs isolated from AIP patients presented low mRNA levels of PBGD when compared to controls, while PBMCs isolated from VP patients presented a decrease in PPOX mRNA. PPOX protein content was higher in AIP patients and lower in VP patients, compared to healthy controls. Regarding antioxidant enzymes, PBMCs and erythrocyte superoxide dismutase (SOD) presented statistically significant higher activity in AIP patients compared to controls, while catalase activity tended to be lower in these patients. No differences were observed regarding antioxidant gene expression in white blood cells. Circulating cells in AIP and VP patients present altered expression of haem biosynthetic enzymes, which could be useful for the differential diagnosis of these two types of porphyria in certain difficult cases. AIP patients present a condition of potential oxidative stress similar to VP patients, evidenced by the post-transcriptional activation of SOD and possible catalase impairment. PMID:27788171

  20. Depletion of Neutrophils Promotes the Resolution of Pulmonary Inflammation and Fibrosis in Mice Infected with Paracoccidioides brasiliensis

    PubMed Central

    Arango, Julián Camilo

    2016-01-01

    Chronic stages of paracoccidioidomycosis (PCM) are characterized by granulomatous lesions which promote the development of pulmonary fibrosis leading to the loss of respiratory function in 50% of patients; in addition, it has been observed that neutrophils predominate during these chronic stages of P. brasiliensis infection. The goal of this study was to evaluate the role of the neutrophil during the chronic stages of experimental pulmonary PCM and during the fibrosis development and tissue repair using a monoclonal specific to this phagocytic cell. Male BALB/c mice were inoculated intranasally with 1.5x106 P. brasiliensis yeast cells. A monoclonal antibody specific to neutrophils was administered at 4 weeks post-inoculation followed by doses every 48h during two weeks. Mice were sacrificed at 8 and 12 weeks post-inoculation to assess cellularity, fungal load, cytokine/chemokine levels, histopathological analysis, collagen and expression of genes related to fibrosis development. Depletion of neutrophils was associated with a significant decrease in the number of eosinophils, dendritic cells, B cells, CD4-T cells, MDSCs and Treg cells, fungal load and levels of most of the pro-inflammatory cytokines/chemokines evaluated, including IL-17, TNF-α and TGF-β1. Recovery of lung architecture was also associated with reduced levels of collagen, high expression of TGF-β3, matrix metalloproteinase (MMP)-12 and -14, and decreased expression of tissue inhibitor metalloproteinase (TIMP)-2, and MMP-8. Depletion of neutrophils might attenuate lung fibrosis and inflammation through down-regulating TGF-β1, TNF-α, IL-17, MMP-8 and TIMP-2. These results suggest that neutrophil could be considered as a therapeutic target in pulmonary fibrosis induced by P. brasiliensis. PMID:27690127

  1. Low molecular weight heparins prevent the induction of autophagy of activated neutrophils and the formation of neutrophil extracellular traps.

    PubMed

    Manfredi, Angelo A; Rovere-Querini, Patrizia; D'Angelo, Armando; Maugeri, Norma

    2017-02-01

    The protection exerted by neutrophils against invading microbes is partially mediated via the generation of neutrophil extracellular traps (NETs). In sterile conditions NETs are damaging species, enriched in autoantigens and endowed with the ability to damage the vessel wall and bystander tissues, to promote thrombogenesis, and to impair wound healing. To identify and reposition agents that can be used to modulate the formation of NETs is a priority in the research agenda. Low molecular weight heparins (LMWH) are currently used, mostly on an empirical basis, in conditions in which NETs play a critical role, such as pregnancy complications associated to autoimmune disease. Here we report that LMWHs induce a profound change in the ability of human neutrophils to generate NETs and to mobilize the content of the primary granules in response to unrelated inflammatory stimuli, such as IL-8, PMA and HMGB1. Autophagy consistently accompanies NET generation in our system and autophagy inhibitors, 3-MA and wortmannin, prevent NET generation. Pretreatment with LMWH in vitro critically jeopardizes neutrophil ability to activate autophagy, a mechanism that might contribute to neutrophil unresponsiveness. Finally, we verified that treatment of healthy volunteers with a single prophylactic dose of parnaparin abrogated the ability of neutrophils to activate autophagy and to generate NETs. Together, these results support the contention that neutrophils, and NET generation in particular, might represent a preferential target of the anti-inflammatory action of LMWH.

  2. 3,4-methylenedioxymethamphetamine (MDMA--Ecstasy) decreases neutrophil activity through the glucocorticoid pathway and impairs host resistance to Listeria monocytogenes infection in mice.

    PubMed

    Ferraz-de-Paula, V; Ribeiro, A; Souza-Queiroz, J; Pinheiro, M L; Vecina, J F; Souza, D P M; Quinteiro-Filho, W M; Moreau, R L M; Queiroz, M L S; Palermo-Neto, J

    2014-12-01

    Ecstasy is the popular name of the abuse drug 3,4-methylenedioxymethamphetamine (MDMA) that decreases immunity in animals. The mechanisms that generate such alterations are still controversial. Seven independent pharmacological approaches were performed in mice to identify the possible mechanisms underlying the decrease of neutrophil activity induced by MDMA and the possible effects of MDMA on host resistance to Listeria monocytogenes. Our data showed that MDMA (10 mg kg(-1)) administration decreases NFκB expression in circulating neutrophils. Metyrapone or RU-486 administration prior to MDMA treatment abrogated MDMA effects on neutrophil activity and NFκB expression, while 6-OHDA or ICI-118,551 administration did not. As MDMA treatment increased the plasmatic levels of adrenaline and noradrenaline, propranolol pre-treatment effects were also evaluated. Propranolol suppressed both MDMA-induced increase in corticosterone serum levels and its effects on neutrophil activity. In a L. monocytogenes experimental infection context, we showed that MDMA: induced myelosuppression by decreasing granulocyte-macrophage hematopoietic progenitors (CFU-GM) in the bone marrow but increased CFU-GM in the spleen; decreased circulating leukocytes and bone marrow cellularity and increased spleen cellularity; decreased pro-inflammatory cytokine (IL-12p70, TNF, IFN-γ, IL-6) and chemokine (MCP-1) production 24 h after the infection; increased the production of pro-inflammatory cytokines and chemokines 72 h after infection and decreased IL-10 levels at all time points analyzed. It was proposed that MDMA immunosuppressive effects on neutrophil activity and host resistance to L monocytogenes rely on NFκB signaling, being mediated by HPA axis activity and corticosterone.

  3. Neutrophil surface presentation of the anti-neutrophil cytoplasmic antibody-antigen proteinase 3 depends on N-terminal processing

    PubMed Central

    von Vietinghoff, S; Eulenberg, C; Wellner, M; Luft, F C; Kettritz, R

    2008-01-01

    The neutrophil serine protease proteinase 3 (PR3) is a main autoantigen in anti-neutrophil cytoplasmic antibody-associated vasculitis. PR3 surface presentation on neutrophilic granulocytes, the main effector cells, is pathogenically important. PR3 is presented by the NB1 (CD177) glycoprotein, but how the presentation develops during neutrophil differentiation is not known. An N-terminally unprocessed PR3 (proPR3) is produced early during neutrophil development and promotes myeloid cell differentiation. We therefore investigated if PR3 presentation depended on NB1 during neutrophil differentiation and if PR3 and proPR3 could both be presented by NB1. In contrast to mature neutrophils, differentiating neutrophils showed an early NB1-independent PR3 surface display that was recognized by only two of four monoclonal anti-PR3 antibodies and occurred in parallel with proPR3, but not PR3 secretion, suggesting that the NB1-independent surface PR3 was proPR3. PR3 gene expression preceeded NB1. When the NB1 receptor was detected on the surface, a mode of PR3 surface display similar to mature neutrophils developed together with the degranulation system. Ectopic expression studies showed that NB1 was a sufficient receptor for PR3 but not proPR3. ProPR3 display on the plasma membrane may influence the bone marrow microenvironment. NB1-mediated PR3 presentation depended on PR3 N-terminal processing implicating the PR3–N-terminus as NB1-binding site. PMID:18462208

  4. Improving Circulation Services through Staff Involvement

    ERIC Educational Resources Information Center

    Kisby, Cynthia M.; Kilman, Marcus D.

    2007-01-01

    The Circulation Services Department at the University of Central Florida Libraries reports on leadership and training initiatives that resulted in a number of service-enhancing projects implemented by a highly motivated and involved staff. Key elements in reinvigorating the department included a change in leadership philosophy, increased…

  5. Mitochondrial DNA is released by shock and activates neutrophils via p38 map kinase.

    PubMed

    Zhang, Qin; Itagaki, Kiyoshi; Hauser, Carl J

    2010-07-01

    Bacterial DNA (bDNA) can activate an innate-immune stimulatory "danger" response via toll-like receptor 9 (TLR9). Mitochondrial DNA (mtDNA) is unique among endogenous molecules in that mitochondria evolved from prokaryotic ancestors. Thus, mtDNA retains molecular motifs similar to bDNA. It is unknown, however, whether mtDNA is released by shock or is capable of eliciting immune responses like bDNA. We hypothesized shock-injured tissues might release mtDNA and that mtDNA might act as a danger-associated molecular pattern (or "alarmin") that can activate neutrophils (PMNs) and contribute to systemic inflammatory response syndrome. Standardized trauma/hemorrhagic shock caused circulation of mtDNA as well as nuclear DNA. Human PMNs were incubated in vitro with purified mtDNA or nuclear DNA, with or without pretreatment by chloroquine (an inhibitor of endosomal receptors like TLR9). Neutrophil activation was assessed as matrix metalloproteinase (MMP) 8 and MMP-9 release as well as p38 and p44/42 mitogen-activated protein kinase (MAPK) phosphorylation. Mitochondrial DNA induced PMN MMP-8/MMP-9 release and p38 phosphorylation but did not activate p44/42. Responses were inhibited by chloroquine. Nuclear DNA did not induce PMN activation. Intravenous injection of disrupted mitochondria (mitochondrial debris) into rats induced p38 MAPK activation and IL-6 and TNF-alpha accumulation in the liver. In summary, mtDNA is released into the circulation by shock. Mitochondrial DNA activates PMN p38 MAPK, probably via TLR9, inducing an inflammatory phenotype. Mitochondrial DNA may act as a danger-associated molecular pattern or alarmin after shock, contributing to the initiation of systemic inflammatory response syndrome.

  6. PhagoSight: An Open-Source MATLAB® Package for the Analysis of Fluorescent Neutrophil and Macrophage Migration in a Zebrafish Model

    PubMed Central

    Henry, Katherine M.; Pase, Luke; Ramos-Lopez, Carlos Fernando; Lieschke, Graham J.; Renshaw, Stephen A.; Reyes-Aldasoro, Constantino Carlos

    2013-01-01

    Neutrophil migration in zebrafish larvae is increasingly used as a model to study the response of these leukocytes to different determinants of the cellular inflammatory response. However, it remains challenging to extract comprehensive information describing the behaviour of neutrophils from the multi-dimensional data sets acquired with widefield or confocal microscopes. Here, we describe PhagoSight, an open-source software package for the segmentation, tracking and visualisation of migrating phagocytes in three dimensions. The algorithms in PhagoSight extract a large number of measurements that summarise the behaviour of neutrophils, but that could potentially be applied to any moving fluorescent cells. To derive a useful panel of variables quantifying aspects of neutrophil migratory behaviour, and to demonstrate the utility of PhagoSight, we evaluated changes in the volume of migrating neutrophils. Cell volume increased as neutrophils migrated towards the wound region of injured zebrafish. PhagoSight is openly available as MATLAB® m-files under the GNU General Public License. Synthetic data sets and a comprehensive user manual are available from http://www.phagosight.org. PMID:24023630

  7. Thalidomide enhances both primary and secondary host resistances to Listeria monocytogenes infection by a neutrophil-related mechanism in female B6C3F1 mice

    SciTech Connect

    Guo, Tai L. . E-mail: tlguo@hsc.vcu.edu; Chi, Rui P.; Karrow, Niel A.; Zhang, Ling X.; Pruett, Stephen B.; Germolec, Dori R.; White, Kimber L.

    2005-12-15

    Previously, we have reported that thalidomide can modulate the immune responses in female B6C3F1 mice. Furthermore, thalidomide immunomodulation increased primary host resistance to intravenously infected Listeria monocytogenes. The present study was intended to evaluate the mechanisms underlying the enhanced host resistance to L. monocytogenes by focusing on the neutrophils. Female B6C3F1 mice were treated intraperitoneally with thalidomide (100 mg/kg) for 15 days. Exposure to thalidomide increased the numbers of neutrophils in the spleens and livers of L. monocytogenes-infected mice when compared to the L. monocytogenes-infected control mice. Additionally, the percentage of neutrophils was also significantly increased after Thd treatment in L. monocytogenes-infected mice. Further studies using antibodies to deplete corresponding cells indicated that thalidomide-mediated increase in primary host resistance (both the moribundity and colony counts in the liver and spleen) to L. monocytogenes infection was due to its effect on neutrophils but not CD8{sup +} T cells or NK cells. Finally, Thd exposure also increased host resistance to secondary host resistance to L. monocytogenes infection, and depletion of neutrophils abolished the protective effect. In conclusion, thalidomide enhanced host resistance to both primary and secondary L. monocytogenes infections by a neutrophil-related mechanism in female B6C3F1 mice.

  8. Neutropenic Mice Provide Insight into the Role of Skin-Infiltrating Neutrophils in the Host Protective Immunity against Filarial Infective Larvae

    PubMed Central

    Pionnier, Nicolas; Brotin, Emilie; Karadjian, Gregory; Hemon, Patrice; Gaudin-Nomé, Françoise; Vallarino-Lhermitte, Nathaly; Nieguitsila, Adélaïde; Fercoq, Frédéric; Aknin, Marie-Laure; Marin-Esteban, Viviana; Chollet-Martin, Sylvie; Schlecht-Louf, Géraldine

    2016-01-01

    Our knowledge and control of the pathogenesis induced by the filariae remain limited due to experimental obstacles presented by parasitic nematode biology and the lack of selective prophylactic or curative drugs. Here we thought to investigate the role of neutrophils in the host innate immune response to the infection caused by the Litomosoides sigmodontis murine model of human filariasis using mice harboring a gain-of-function mutation of the chemokine receptor CXCR4 and characterized by a profound blood neutropenia (Cxcr4+/1013). We provided manifold evidence emphasizing the major role of neutrophils in the control of the early stages of infection occurring in the skin. Firstly, we uncovered that the filarial parasitic success was dramatically decreased in Cxcr4+/1013 mice upon subcutaneous delivery of the infective stages of filariae (infective larvae, L3). This protection was linked to a larger number of neutrophils constitutively present in the skin of the mutant mice herein characterized as compared to wild type (wt) mice. Indeed, the parasitic success in Cxcr4+/1013 mice was normalized either upon depleting neutrophils, including the pool in the skin, or bypassing the skin via the intravenous infection of L3. Second, extending these observations to wt mice we found that subcutaneous delivery of L3 elicited an increase of neutrophils in the skin. Finally, living L3 larvae were able to promote in both wt and mutant mice, an oxidative burst response and the release of neutrophil extracellular traps (NET). This response of neutrophils, which is adapted to the large size of the L3 infective stages, likely directly contributes to the anti-parasitic strategies implemented by the host. Collectively, our results are demonstrating the contribution of neutrophils in early anti-filarial host responses through their capacity to undertake different anti-filarial strategies such as oxidative burst, degranulation and NETosis. PMID:27111140

  9. Overhauser-Enhanced MRI of Elastase Activity from In Vitro Human Neutrophil Degranulation

    PubMed Central

    Parzy, Elodie; Bouchaud, Véronique; Massot, Philippe; Voisin, Pierre; Koonjoo, Neha; Moncelet, Damien; Franconi, Jean-Michel; Thiaudière, Eric; Mellet, Philippe

    2013-01-01

    Background Magnetic resonance imaging can reveal exquisite anatomical details. However several diseases would benefit from an imaging technique able to specifically detect biochemical alterations. In this context protease activity imaging is one of the most promising areas of research. Methodology/Principal Findings We designed an elastase substrate by grafting stable nitroxide free radicals on soluble elastin. This substrate generates a high Overhauser magnetic resonance imaging (OMRI) contrast upon digestion by the target proteases through the modulation of its rotational correlation time. The sensitivity is sufficient to generate contrasted images of the degranulation of neutrophils induced by a calcium ionophore from 2×104 cells per milliliter, well under the physiological neutrophils concentrations. Conclusions/Significance These ex-vivo experiments give evidence that OMRI is suitable for imaging elastase activity from neutrophil degranulation. Provided that a fast protease-substrate is used these results open the door to better diagnoses of a number of important pathologies (cystic fibrosis, inflammation, pancreatitis) by OMRI or Electron Paramagnetic Resonance Imaging in vivo. It also provides a long-expected method to monitor anti-protease treatments efficiency and help pharmaceutical research. PMID:23469112

  10. Short communication: Relationship between urinary neutrophil gelatinase-associated lipocalin and noninfectious pyuria in dogs.

    PubMed

    Proverbio, D; Spada, E; Baggiani, L; Bagnagatti De Giorgi, G; Ferro, E; Martino, P A; Perego, R

    2015-01-01

    Neutrophil gelatinase-associated lipocalin (NGAL) is a neutrophil-derived protein whose concentration increases in plasma and urine with ongoing renal damage. Urinary leucocytes can be a potential source of urinary NGAL. The aim of this study is to investigate the effects of urinary neutrophil count and other urinary parameters on urinary NGAL values in urine with negative culture. Urinalysis, urine culture, and determination of urinary NGAL were performed on 33 clinically healthy nonproteinuric dogs with negative urinoculture. The median uNGAL