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Sample records for bovine blood neutrophils

  1. Evaluation of endotoxin (LPS) activity in bovine blood using neutrophil dependent chemiluminescence

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    The purpose of this study was to evaluate the applicability of a neutrophil chemiluminescence-based assay for the measurement of LPS stimulatory activity in bovine whole blood. The assay is based on the capacity for LPS to trigger the respiratory oxidative burst activity (RBA) of autologous neutroph...

  2. Effector responses of bovine blood neutrophils against Escherichia coli: Role of NOD1/NF-κB signalling pathway.

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    Tan, Xun; Wei, Liang-Jun; Fan, Guo-Juan; Jiang, Ya-Nan; Yu, Xu-Ping

    2015-11-15

    Neutrophils use a broad array of pattern recognition receptors to sense and respond to invading pathogens and are important in the early control of acute bacterial infections. Nucleotide-binding oligomerizing domain-1 (NOD1) is a cytoplasmic receptor involved in recognizing bacterial peptidoglycan. Reduced neutrophil NOD1 expression has been reported in periparturient dairy cows. The aim of this study was to investigate the role of NOD1 signalling in the early responses of bovine neutrophils to bacterial infections. Blood neutrophils from healthy heifers were preincubated for 2h with ML130, a selective inhibitor of NOD1-dependent nuclear factor-κB (NF-κB) activation. Thereafter, cells were cultured with live Escherichia coli for additional 30 min or subjected to Boyden chamber cell migration assay with E. coli in the lower chamber. Results showed that ML130 inhibited E. coli-induced NF-κB nuclear translocation. There was an indication, although not significant, that ML130 down-regulated gene expression of proinflammatory cytokines interleukin (IL)-1β and tumour necrosis factor (TNF)-α, chemokines IL-8 and C-X-C motif ligand 2 (CXCL2), and adhesion molecule CD62L, in E. coli-challenged neutrophils. Flow cytometry-based Annexin V staining revealed a considerable increase in neutrophil survival upon E. coli infection, an effect that was attenuated in the presence of ML130. Additionally, inhibition of NOD1/NF-κB signalling resulted in reduced migration of neutrophils to E. coli, and impaired phagocytosis, intracellular bacterial killing and reactive oxygen species production by neutrophils. These results indicate that NOD1/NF-κB pathway plays a crucial role in modulating neutrophil responses that are important for early control of infections. Approaches aiming at restoring neutrophil NOD1 function could be beneficial for prevention or treatment of coliform mastitis.

  3. Staphylococcus aureus capsular polysaccharide types 5 and 8 reduce killing by bovine neutrophils in vitro.

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    Kampen, Annette H; Tollersrud, Tore; Lund, Arve

    2005-03-01

    Isogenic variants of Staphylococcus aureus strain Reynolds expressing either no capsule or capsular polysaccharide (CP) type 5 (CP5) or type 8 (CP8) were used to assess the effect of CP on bacterial killing and the respiratory burst of bovine neutrophils. The effects of antisera specific for CP5 and CP8 were also evaluated. The killing of live bacteria by isolated neutrophils was quantified in a bactericidal assay, while the respiratory burst after stimulation with live bacteria in whole blood was measured by flow cytometry. The expression of a CP5 or CP8 capsule protected the bacteria from being killed by bovine neutrophils in vitro (P killing of the capsule-expressing bacteria and enhanced their stimulating effect in the respiratory burst assay (P killing and prevents the bacteria from inducing respiratory burst of bovine neutrophils in vitro and that these effects can be reversed by the addition of serotype-specific antisera.

  4. Investigation of the effect of Mycobacterium bovis infection on bovine neutrophils functions.

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    Wang, Jin; Zhou, Xiangmei; Pan, Bo; Yang, Lifeng; Yin, Xiaomin; Xu, Binrui; Zhao, Deming

    2013-11-01

    Bovine tuberculosis is a disease in cattle caused by infection with Mycobacterium bovis. The disease has posed significant economic losses and remains a public health hazard worldwide. Interactions between M. bovis and bovine macrophages have been extensively characterized in various studies, while similar analyses in neutrophils, which are one of the other types of white blood cells in mammals, were often overlooked. Neutrophils provide defense against all microbes and can present a diverse collection of antimicrobial molecules, which play an important role in the control of tuberculosis progression. Much of the available data about the involvement of neutrophils in the killing M. bovis is controversial. In this study, we assessed the effect of in vitro infection with M. bovis on some parameters of neutrophils functions including phenotypic changes, apoptosis rate and inflammatory cytokines production. Our results demonstrated that phagocytosis of M. bovis activated and enhanced bovine neutrophils functions as well as initialed their defense mechanism, but failed to eliminate the mycobacteria. Moreover, autophagy might get involved in the defense infection process functioning as a protective mechanism, and inducible-autophagy by lipopolysaccharides stimulation and starvation treatment could efficiently reverse the inability of neutrophils for killing M. bovis, suggesting a potential target for anti-mycobacterial drug-therapy.

  5. Apoptosis of bovine neutrophils following diapedesis through a monolayer of endothelial and mammary epithelial cells.

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    Van Oostveldt, K; Paape, Max; Burvenich, Christian

    2002-01-01

    In a two-chamber system, isolated blood polymorphonuclear neutrophil leukocytes (PMN) were allowed to migrate (5 h, 37 C) in response to bovine complement component C5a across calfskin and rat-tail type I collagen-coated micropore membranes, arterial endothelial, or mammary epithelial cell monolayer on calfskin and rat-tail collagen-coated membranes, respectively. Migration through calfskin collagen-coated membranes resulted in 14.5% +/- 3.4% apoptotic PMN, which was significantly higher than...

  6. [Ultrastructural location of enzymes in peripheral blood neutrophils and in cerebrospinal fluid neutrophils in neuroinfections].

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    Skotarczak, B

    1993-01-01

    Using cytochemical methods the location and activity were determined of alkaline phosphatase, ATP-ase and succinate dehydrogenase as representative enzymes for the metabolic processes in neutrophils isolated from blood and cerebrospinal fluid (CSF) of patients with meningococcal meningoencephalitis as compared with peripheral blood neutrophils in a control group. The study showed presence of phosphatase on the membranes of many intracellular structures. The activity of the enzymes was higher than in the control group in the membranes of neutrophils in blood and CSF. This is explained as an effect of action of the chemotactic factor on the cell membrane and activation of the cell to movements and phagocytosis. ATP-ase activity in peripheral blood neutrophils in controls was found in all membranous structures in the cell. However, in peripheral blood neutrophils and CSF neutrophils in the acute stage of the disease the active enzyme was noted, in the first place, in cell membranes and digesting vacuoles, which reflected probably the direction of metabolic processes for phagocytosis and destroying of bacteria. The activity of succinate dehydrogenase was found in mitochondrial membranes. Peripheral blood and CSF neutrophils showed a high activity of the enzyme. In the CSF cells in acute phase atypical sites of succinate dehydrogenase activity were noted, which was explained as a sign of cell destruction.

  7. Neutrophil-Mediated Delivery of Therapeutic Nanoparticles across Blood Vessel Barrier for Treatment of Inflammation and Infection.

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    Chu, Dafeng; Gao, Jin; Wang, Zhenjia

    2015-12-22

    Endothelial cells form a monolayer in lumen of blood vessels presenting a great barrier for delivery of therapeutic nanoparticles (NPs) into extravascular tissues where most diseases occur, such as inflammation disorders and infection. Here, we report a strategy for delivering therapeutic NPs across this blood vessel barrier by nanoparticle in situ hitchhiking activated neutrophils. Using intravital microscopy of TNF-α-induced inflammation of mouse cremaster venules and a mouse model of acute lung inflammation, we demonstrated that intravenously (iv) infused NPs made from denatured bovine serum albumin (BSA) were specifically internalized by activated neutrophils, and subsequently, the neutrophils containing NPs migrated across blood vessels into inflammatory tissues. When neutrophils were depleted using anti-Gr-1 in a mouse, the transport of albumin NPs across blood vessel walls was robustly abolished. Furthermore, it was found that albumin nanoparticle internalization did not affect neutrophil mobility and functions. Administration of drug-loaded albumin NPs markedly mitigated the lung inflammation induced by LPS (lipopolysaccharide) or infection by Pseudomonas aeruginosa. These results demonstrate the use of an albumin nanoparticle platform for in situ targeting of activated neutrophils for delivery of therapeutics across the blood vessel barriers into diseased sites. This study demonstrates our ability to hijack neutrophils to deliver nanoparticles to targeted diseased sites.

  8. Advanced oxidation protein products are generated by bovine neutrophils and inhibit free radical production in vitro.

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    Bordignon, Milena; Da Dalt, Laura; Marinelli, Lieta; Gabai, Gianfranco

    2014-01-01

    Despite the recognised importance of oxidative stress in the health and immune function of dairy cows, protein oxidation markers have been poorly studied in this species. The current study aimed to characterise markers of protein oxidation generated by activated bovine neutrophils and investigate the biological effects of advanced oxidation protein products (AOPP) on bovine neutrophils. Markers of protein oxidation (AOPP, dityrosines and carbonyls) were measured in culture medium containing bovine serum albumin (BSA) exposed to neutrophils. The effect of AOPP-BSA on generation of reactive oxygen species (ROS) was assessed by chemiluminescence. Activation of caspases-3, -8 and -9 and the presence of DNA laddering were used as apoptosis markers. Greater amounts of AOPP were generated by phorbol myristate acetate (PMA)-activated than non-activated neutrophils (1.46 ± 0.13 vs. 0.75 ± 0.13 nmol/mg protein, respectively; P<0.05). Activated neutrophils and hypochlorous acid generated slightly different patterns of oxidized protein markers. Exposure to AOPP-BSA did not stimulate ROS production. Activated neutrophils generated a lesser amount of ROS when incubated with AOPP-BSA (P<0.001). Activation with PMA induced a loss of viable neutrophils after 3h, which was greater with AOPP-BSA incubation (P<0.05). Detectable amounts of active caspases-3, -8 and -9 were found in nearly all samples but differences in caspase activation or DNA laddering were not observed comparing treatment groups. Apoptosis was unlikely to be responsible for the greater loss of PMA-activated neutrophils cultured in AOPP-BSA and it is possible that primary necrosis occurred. The results suggest that accumulation of oxidized proteins at an inflammatory site might result in a progressive reduction of neutrophil viability.

  9. Differential neutrophil gene expression in early bovine pregnancy

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    Kizaki Keiichiro

    2013-02-01

    Full Text Available Abstract Background In food production animals, especially cattle, the diagnosis of gestation is important because the timing of gestation directly affects the running of farms. Various methods have been used to detect gestation, but none of them are ideal because of problems with the timing of detection or the accuracy, simplicity, or cost of the method. A new method for detecting gestation, which involves assessing interferon-tau (IFNT-stimulated gene expression in peripheral blood leukocytes (PBL, was recently proposed. PBL fractionation methods were used to examine whether the expression profiles of various PBL populations could be used as reliable diagnostic markers of bovine gestation. Methods PBL were collected on days 0 (just before artificial insemination, 7, 14, 17, 21, and 28 of gestation. The gene expression levels of the PBL were assessed with microarray analysis and/or quantitative real-time reverse transcription (q PCR. PBL fractions were collected by flow cytometry or density gradient cell separation using Histopaque 1083 or Ficoll-Conray solutions. The expression levels of four IFNT-stimulated genes, interferon-stimulated protein 15 kDa (ISG15, myxovirus-resistance (MX 1 and 2, and 2′-5′-oligoadenylate synthetase (OAS1, were then analyzed in each fraction through day 28 of gestation using qPCR. Results Microarray analysis detected 72 and 28 genes in whole PBL that were significantly higher on days 14 and 21 of gestation, respectively, than on day 0. The upregulated genes included IFNT-stimulated genes. The expression levels of these genes increased with the progression of gestation until day 21. In flow cytometry experiments, on day 14 the expression levels of all of the genes were significantly higher in the granulocyte fraction than in the other fractions. Their expression gradually decreased through day 28 of gestation. Strong correlations were observed between the expression levels of the four genes in the granulocyte

  10. Peripheral blood neutrophil cytokine hyper-reactivity in chronic periodontitis.

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    Ling, Martin R; Chapple, Iain L C; Matthews, John B

    2015-10-01

    Pro-inflammatory cytokine release (IL-8, IL-6, TNF-α, IL-1β) by peripheral blood neutrophils, isolated from periodontitis patients (before/after therapy) and matched controls, was determined after 18 h culture in the presence/absence of Escherichia coli LPS, opsonised Staphylococcus aureus, heat-killed Fusobacterium nucleatum and Porphyromonas gingivalis. All cultures demonstrated differences in the amounts of each cytokine detected (P IL-6 > TNF-α = IL-1β). Median cytokine release from unstimulated patient neutrophils was consistently, but non-significantly, higher than from control cells. Stimulated cytokine release from untreated patient neutrophils was also consistently higher than from control cells. This hyper-reactivity was significant for all tested cytokines when data for all stimuli were combined (P TNF-α), opsonised S. aureus (IL-8, TNF-α, IL-1β) and P. gingivalis (IL-8, IL-1β). Cytokine production by patient neutrophils did not reduce following successful non-surgical periodontal therapy and, except for responses to F. nucleatum, the cytokine hyper-reactivity detected pre-therapy was retained. These data demonstrate that chronic periodontitis is characterised by neutrophils that constitutively exhibit cytokine hyper-reactivity, the effects of which could modulate local and systemic inflammatory-immune responses and influence the risk and severity of periodontitis-associated systemic inflammatory diseases.

  11. Generation of lipid neutrophil chemoattractant by irradiated bovine aortic endothelial cells.

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    Matzner, Y; Cohn, M; Hyam, E; Razin, E; Fuks, Z; Buchanan, M R; Haas, T A; Vlodavsky, I; Eldor, A

    1988-04-15

    Radiation injury to blood vessels is associated with an acute inflammatory process. We investigated the capacity of cultured bovine aortic endothelial cells (BAEC) to produce chemotactic factors after radiation injury. BAEC in serum-free media were irradiated with a cobalt-60 Gammacell 220 and the cell supernatants were assayed for chemotactic activity for human neutrophils in a Boyden chamber. There was a rapid release of chemotactic activity into the BAEC supernatants which was dependent both on the dose of radiation (5 to 40 Gy) and the time between irradiation and sample collection. In contrast, isolation of BAEC lysates by freeze-thawing was not associated with the presence of similar chemotactic activity. The chemotactic activity released from the irradiated BAEC was not destroyed by boiling nor by treatment with trypsin. The release of the chemotactic activity was, however, inhibited by the addition of a lipoxygenase inhibitor but not by the addition of a cyclooxygenase inhibitor before the irradiation. The chemotactic activity was recovered from the cell supernatants in the lipid phase after extraction with chloroform/methanol. Furthermore, the chloroform/methanol extracts co-eluted with authentic leukotriene B4 when the BAEC were prelabeled with [14C] arachidonic acid. However, we were unable to detect endogenous leukotriene B4 with RIA. Instead, the only detectable endogenous lipid present in the supernatants was 13-hydroxyoctadecadienoic acid which is derived from linoleic acid via the lipoxygenase pathway. 13-Hydroxyoctadecadienoic acid, however, had no chemotactic activity. These findings suggest that endothelial cells rapidly release a chemotactic agent after irradiation, the release of which is associated with a lipoxygenase pathway. The release of this chemotactic activity may account in part for the acute inflammatory response that is observed after ionizing irradiation.

  12. Interaction of bovine peripheral blood polymorphonuclear cells and Leptospira species; innate responses in the natural bovine reservoir host.

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    Jennifer H Wilson-Welder

    2016-07-01

    Full Text Available Cattle are the reservoir hosts of Leptospira borgpetersenii serovar Hardjo, and can also be reservoir hosts of other Leptospira species such as L. kirschneri, and L. interrogans. As a reservoir host, cattle shed Leptospira, infecting other animals, including humans. Previous studies with human and murine neutrophils have shown activation of neutrophil extracellular trap or NET formation, and upregulation of inflammatory mediators by neutrophils in the presence of Leptospira. Humans, companion animals and most widely studied models of Leptospirosis are of acute infection, hallmarked by systemic inflammatory response, neutrophilia and septicemia. In contrast, cattle exhibit chronic infection with few outward clinical signs aside from reproductive failure. Taking into consideration that there is host species variation in innate immunity, especially in pathogen recognition and response, the interaction of bovine peripheral blood polymorphonuclear cells (PMNs and several Leptospira strains was evaluated. Studies including bovine-adapted strains, human pathogen strains, a saprophyte and inactivated organisms. Incubation of PMNs with Leptospira did induce slight activation of neutrophil NETs, greater than unstimulated cells but less than the quantity from E. coli P4 stimulated PMNs. Very low but significant from non-stimulated, levels of reactive oxygen peroxides were produced in the presence of all Leptospira strains and E. coli P4. Similarly, significant levels of reactive nitrogen intermediaries (NO2 was produced from PMNs when incubated with the Leptospira strains and greater quantities in the presence of E. coli P4. PMNs incubated with Leptospira induced RNA transcripts of IL-1β, MIP-1α, and TNF-α, with greater amounts induced by live organisms when compared to heat-inactivated leptospires. Transcript for inflammatory cytokine IL-8 was also induced, at similar levels regardless of Leptospira strain or viability. However, incubation of

  13. Absolute counting of neutrophils in whole blood using flow cytometry.

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    Brunck, Marion E G; Andersen, Stacey B; Timmins, Nicholas E; Osborne, Geoffrey W; Nielsen, Lars K

    2014-12-01

    Absolute neutrophil count (ANC) is used clinically to monitor physiological dysfunctions such as myelosuppression or infection. In the research laboratory, ANC is a valuable measure to monitor the evolution of a wide range of disease states in disease models. Flow cytometry (FCM) is a fast, widely used approach to confidently identify thousands of cells within minutes. FCM can be optimised for absolute counting using spiked-in beads or by measuring the sample volume analysed. Here we combine the 1A8 antibody, specific for the mouse granulocyte protein Ly6G, with flow cytometric counting in straightforward FCM assays for mouse ANC, easily implementable in the research laboratory. Volumetric and Trucount™ bead assays were optimized for mouse neutrophils, and ANC values obtained with these protocols were compared to ANC measured by a dual-platform assay using the Orphee Mythic 18 veterinary haematology analyser. The single platform assays were more precise with decreased intra-assay variability compared with ANC obtained using the dual protocol. Defining ANC based on Ly6G expression produces a 15% higher estimate than the dual protocol. Allowing for this difference in ANC definition, the flow cytometry counting assays using Ly6G can be used reliably in the research laboratory to quantify mouse ANC from a small volume of blood. We demonstrate the utility of the volumetric protocol in a time-course study of chemotherapy induced neutropenia using four drug regimens.

  14. Comparative in vitro phagocytosis and F-actin polymerization of bovine neonatal neutrophils.

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    Moiola, F; Spycher, M; Wyder-Walther, M; Zwahlen, R D

    1994-04-01

    Analysis of neonatal neutrophil (PMN) functions should help to reveal factors which could contribute to the impaired host defense system of neonates. We analysed functional parameters of PMN from newborn calves (N-PMN) and adult bovines (A-PMN): cellular volume and F-actin content upon stimulation with complement factors, by cytofluorometry and phagocytosis of E. coli 78:80B with a colorimetric assay. Polymerization of F-actin was rapid in both N- and A-PMN, but reached higher levels in N-PMN. N-PMN are significantly smaller than A-PMN throughout the whole activation time. Percentage of phagocytosing PMN, the rate of phagocytosis, and the rate of killing are similar between A- and N-PMN after opsonization of bacteria with adult serum (AS). Opsonization with newborn serum (NS) reduced all three examined parameters: in A-PMN more (P dexamethasone) and non-steroidal (phenylbutazone) anti-inflammatory drugs inhibited phagocytosis by N-PMN less than by A-PMN. Higher relative F-actin content of N-PMN can be correlated with the documented functional hyperactivity of bovine N-PMN. However, the exaggerated impairment of phagocytosis in calves observed after age-matched opsonization of bacteria could potentially indicate a specific host defence defect.

  15. Venous levels of shear support neutrophil-platelet adhesion and neutrophil aggregation in blood via P-selectin and beta2-integrin

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    Konstantopoulos, K.; Neelamegham, S.; Burns, A. R.; Hentzen, E.; Kansas, G. S.; Snapp, K. R.; Berg, E. L.; Hellums, J. D.; Smith, C. W.; McIntire, L. V.; Simon, S. I.

    1998-01-01

    BACKGROUND: After activation, platelets adhere to neutrophils via P-selectin and beta2-integrin. The molecular mechanisms and adhesion events in whole blood exposed to venous levels of hydrodynamic shear in the absence of exogenous activation remain unknown. METHODS AND RESULTS: Whole blood was sheared at approximately 100 s(-1). The kinetics of neutrophil-platelet adhesion and neutrophil aggregation were measured in real time by flow cytometry. P-selectin was upregulated to the platelet surface in response to shear and was the primary factor mediating neutrophil-platelet adhesion. The extent of neutrophil aggregation increased linearly with platelet adhesion to neutrophils. Blocking either P-selectin, its glycoprotein ligand PSGL-1, or both simultaneously by preincubation with a monoclonal antibody resulted in equivalent inhibition of neutrophil-platelet adhesion (approximately 30%) and neutrophil aggregation (approximately 70%). The residual amount of neutrophil adhesion was blocked with anti-CD11b/CD18. Treatment of blood with prostacyclin analogue ZK36374, which raises cAMP levels in platelets, blocked P-selectin upregulation and neutrophil aggregation to baseline. Complete abrogation of platelet-neutrophil adhesion required both ZK36374 and anti-CD18. Electron microscopic observations of fixed blood specimens revealed that platelets augmented neutrophil aggregation both by forming bridges between neutrophils and through contact-mediated activation. CONCLUSIONS: The results are consistent with a model in which venous levels of shear support platelet adherence to neutrophils via P-selectin binding PSGL-1. This interaction alone is sufficient to mediate neutrophil aggregation. Abrogation of platelet adhesion and aggregation requires blocking Mac-1 in addition to PSGL-1 or P-selectin. The described mechanisms are likely of key importance in the pathogenesis and progression of thrombotic disorders that are exacerbated by leukocyte-platelet aggregation.

  16. Response of Neutrophils to Extracellular Haemoglobin and LTA in Human Blood System

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    Sae-Kyung Lee

    2015-03-01

    Interpretation: metHb and metHb + LTA complex are ligands of TLR2, inducing an unconventional TLR signalling pathway. Neutrophils are a highly sensitive cell type to metHb + LTA complex. During a haemolytic infection, white blood cells in the vicinity crosstalk to modulate neutrophil TLR-signalling induced by metHb and LTA.

  17. [Effects of periodontitis and opsonized zymosan on chemiluminescence of blood neutrophils].

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    Zekonis, Gediminas; Ivanauskiene, Egle

    2002-01-01

    The objective of the present investigation was to explore oxidative function of parodontitis patient's blood neutrophils stimulated with opsonized zymosan by method of luminol- and lucigenin-dependent chemiluminescence. The leukocytes for this study were obtained from peripheral venous blood of 16 parodontitis patients and 10 healthy subjects. The luminol-dependent chemiluminescence of stimulated neutrophils of parodontitis patients did not differ (p > 0.05) from control subjects (66,849 +/- 6372 cpm and 61,243 +/- 5240 cpm, respectively). Lucigenin-dependent chemiluminescence of stimulated neutrophils of parodontitis patients was increased (p parodontitis patients was significantly higher (p parodontitis patients possibly can affect parodontal health.

  18. Cathepsin G-dependent modulation of platelet thrombus formation in vivo by blood neutrophils.

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    Nauder Faraday

    Full Text Available Neutrophils are consistently associated with arterial thrombotic morbidity in human clinical studies but the causal basis for this association is unclear. We tested the hypothesis that neutrophils modulate platelet activation and thrombus formation in vivo in a cathepsin G-dependent manner. Neutrophils enhanced aggregation of human platelets in vitro in dose-dependent fashion and this effect was diminished by pharmacologic inhibition of cathepsin G activity and knockdown of cathepsin G expression. Tail bleeding time in the mouse was prolonged by a cathepsin G inhibitor and in cathepsin G knockout mice, and formation of neutrophil-platelet conjugates in blood that was shed from transected tails was reduced in the absence of cathepsin G. Bleeding time was highly correlated with blood neutrophil count in wildtype but not cathepsin G deficient mice. In the presence of elevated blood neutrophil counts, the anti-thrombotic effect of cathepsin G inhibition was greater than that of aspirin and additive to it when administered in combination. Both pharmacologic inhibition of cathepsin G and its congenital absence prolonged the time for platelet thrombus to form in ferric chloride-injured mouse mesenteric arterioles. In a vaso-occlusive model of ischemic stroke, inhibition of cathepsin G and its congenital absence improved cerebral blood flow, reduced histologic brain injury, and improved neurobehavioral outcome. These experiments demonstrate that neutrophil cathepsin G is a physiologic modulator of platelet thrombus formation in vivo and has potential as a target for novel anti-thrombotic therapies.

  19. Multicenter Systems Analysis of Human Blood Reveals Immature Neutrophils in Males and During Pregnancy

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    Blazkova, Jana; Gupta, Sarthak; Liu, Yudong; Gaudilliere, Brice; Ganio, Edward A.; Bolen, Christopher R.; Saar-Dover, Ron; Fragiadakis, Gabriela K.; Angst, Martin S.; Hasni, Sarfaraz; Aghaeepour, Nima; Stevenson, David; Baldwin, Nicole; Anguiano, Esperanza; Chaussabel, Damien; Altman, Matthew C.; Kaplan, Mariana J.; Davis, Mark M.

    2017-01-01

    Despite clear differences in immune system responses and in the prevalence of autoimmune diseases between males and females, there is little understanding of the processes involved. In this study, we identified a gene signature of immature-like neutrophils, characterized by the overexpression of genes encoding for several granule-containing proteins, which was found at higher levels (up to 3-fold) in young (20–30 y old) but not older (60 to >89 y old) males compared with females. Functional and phenotypic characterization of peripheral blood neutrophils revealed more mature and responsive neutrophils in young females, which also exhibited an elevated capacity in neutrophil extracellular trap formation at baseline and upon microbial or sterile autoimmune stimuli. The expression levels of the immature-like neutrophil signature increased linearly with pregnancy, an immune state of increased susceptibility to certain infections. Using mass cytometry, we also find increased frequencies of immature forms of neutrophils in the blood of women during late pregnancy. Thus, our findings show novel sex differences in innate immunity and identify a common neutrophil signature in males and in pregnant women. PMID:28179497

  20. Reactive Oxygen Species Production in Peripheral Blood Neutrophils of Obstructive Sleep Apnea Patients

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    Guoda Pilkauskaite; Skaidrius Miliauskas; Raimundas Sakalauskas

    2013-01-01

    Obstructive sleep apnea (OSA) as well as obesity is associated with increased production of reactive oxygen species (ROS). Neutrophils produce great amounts of ROS. The aim was to evaluate peripheral blood neutrophils ROS production in men with OSA and to establish relations with disease severity and obesity. Methods. Forty-six men with OSA and 10 controls were investigated. OSA was confirmed by polysomnography (PSG), when apnea/hypopnea index was >5/h. Body mass index (BMI) was evaluated. Ne...

  1. Identification of highly active flocculant proteins in bovine blood.

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    Piazza, George J; Nuñez, Alberto; Garcia, Rafael A

    2012-03-01

    Synthetic polymeric flocculants are used extensively for wastewater remediation, soil stabilization, and reduction in water leakage from unlined canals. Sources of highly active, inexpensive, renewable flocculants are needed to replace synthetic flocculants. High kaolin flocculant activity was documented for bovine blood (BB) and blood plasma with several anticoagulant treatments. BB serum also had high flocculant activity. To address the hypothesis that some blood proteins have strong flocculating activity, the BB proteins were separated by SEC. Then, the major proteins of the flocculant-active fractions were separated by SDS-PAGE. Identity of the major protein components was determined by tryptic digestion and peptide analysis by MALDI TOF MS. The sequence of selected peptides was confirmed using TOF/TOF-MS/MS fragmentation. Hemoglobin dimer (subunits α and β) was identified as the major protein component of the active fraction in BB; its high flocculation activity was confirmed by testing a commercial sample of hemoglobin. In the same manner, three proteins from blood plasma (fibrinogen, γ-globulin, α-2-macroglobulin) were found to be highly active flocculants, but bovine serum albumin, α-globulin, and β-globulin were not flocculants. On a mass basis, hemoglobin, γ-globulin, α-2-macroglobulin were as effective as anionic polyacrylamide (PAM), a widely used synthetic flocculant. The blood proteins acted faster than PAM, and unlike PAM, the blood proteins flocculants did not require calcium salts for their activity.

  2. Adsorbed plasma proteins modulate the effects of single-walled carbon nanotubes on neutrophils in blood.

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    Vlasova, Irina I; Mikhalchik, Elena V; Barinov, Nikolay A; Kostevich, Valeria A; Smolina, Natalia V; Klinov, Dmitry V; Sokolov, Alexey V

    2016-08-01

    Proteins adsorbed on a surface may affect the interaction of this surface with cells. Here, we studied the binding of human serum albumin (HSA), fibrinogen (FBG) and immunoglobulin G (IgG) to PEGylated single-walled carbon nanotubes (PEG-SWCNTs) and evaluated the impact of PEG-SWCNT treated by these proteins on neutrophils in whole blood samples. Measurements of adsorption parameters revealed tight binding of proteins to PEG-SWCNTs. AFM was employed to directly observe protein binding to sidewalls of PEG-SWCNTs. Fluorescein-labeled IgG was used to ascertain the stability of PEG-SWCNT-IgG complexes in plasma. In blood samples, all plasma proteins mitigated damage of neutrophils observed just after blood exposure to PEG-SWCNTs, while only treatment of PEG-SWCNTs with IgG resulted in dose- and time-dependent enhancement of CNT-induced neutrophil activation and in potentiation of oxidative stress. Our study demonstrates the ability of adsorbed plasma proteins to influence neutrophil response caused by PEG-SWCNTs in whole blood.

  3. Effects of maternal protein-energy malnutrition and cold stress on neutrophil function of bovine neonates.

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    Woodard, L F; Eckblad, W P; Olson, D P; Bull, R C; Everson, D O

    1980-08-01

    The effects of maternal protein or calorie deprivation (or both) on the bactericidal activity of neutrophils and sera from newborn calves subjected to cold stress were studied. Nutritional deficiencies in the dam had little effect on in vitro bactericidal activity of neutrophils and base-line sera taken at birth. Neutrophils obtained at birth destroyed Staphylococcus aureus but not Escherichia coli when incubated with either unheated or heated autologous base-line sera. Heat treatment of base-line sera to inactivate complement did not alter bacterial growth. When incubated in the presence of autologous base-line sera, neutrophils from 3-day-old calves were no more active in the destruction of either bacterium than were neutrophils from newborn calves. However, addition of day 3 (immunoglobulin-containing) sera enabled day 3 neutrophils to destroy E coli (P heat treatment of the sera. Maternal protein deficiency significantly increased (P calves exposed to 1 C or 21 C environmental chambers for 3 days. Also, cold stress-nutritional stress interactions were not detected.

  4. The influence of bovine neutrophils on in vitro phagocytosis and killing of Staphylococcus aureus in heifers supplemented with selenium and vitamin E

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    Chalong Wachirapakorn

    2007-05-01

    Full Text Available An experiment was performed to determine the influence of bovine nutrophils on in vitro phagocytosis and killing of Staphylococcus aureus after selenium (Se, selenium yeast and vitamin E (vit E wassupplemented in heifers. Twelve healthycrossbred 75% Holstein-Friesian x Sahiwal heifers were divided into 4 groups in a 2x2 factorial arrangement in CRD. Heifers were supplemented organic selenium (seleniumyeast and vitamin E powder. The treatments were as follows; treatment 1 3 mg Se 2,000 IU vit E /hd/d (3Se2E, treatment 2 3 mg Se 4,000 IUvit E/hd/d (3Se 4E, treatment 3 6 mg Se 2,000 IU vit E/hd/d (6Se2E,and treatment 4 6 mg Se 4,000 IU vit E/hd/d (6Se 4E. The experiment comprised 3 periods: pre-supplementation (8 days, supplementation (8 days and post-supplementation (8 days periods. All heiferswere offered concentrate (15% CP at 4 kg/hd/d and rice straw hay ad libitum. Blood neutrophils were isolated from each heifer. Phagocytosis was determined by direct ingested count and killing of S. aureus byNBT reducting test. Phagocytosis and killing of S. aureus had a greater non-specific immune response during the supplementation period than in the pre-supplementation period in all treatments (P<0.05. Supplementationof 3Se4E resulted in a greater number of white blood cells (54,900 cells/cu.mm and neutrophils (11,398 cells/cu.mm. and improved phagocytosis (85% and killing of S. aureus (47% as compared to the pre-supplementation period.

  5. Expression of CD14 and toll-like receptors 2 and 4 by milk neutrophils in bovine mammary glands infected with Corynebacterium bovis

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    Maiara G. Blagitz

    2015-01-01

    Full Text Available This study evaluated the expression of CD14, toll-like receptor (TLR 2 and TLR4 on the surface of milk neutrophils in bovine mammary glands infected with Corynebacterium bovis. Here, we used 23 culture-negative control quarters with no abnormal secretion on the strip cup test and milk somatic cell count lower than 1x105 cells/mL, and 14 C. bovis infected quarters. The identification of neutrophils, as well as, the percentage of neutrophils that expressed CD14, TLR2 and TLR4 were analyzed by flow cytometry using monoclonal antibodies. The present study encountered no significant difference in the percentages of milk neutrophils that expressed TLR2 and TLR4 or in the expression of TLR4 by milk neutrophils. Conversely, a lower median fluorescence intensity of TLR2 in milk neutrophils was observed in C. bovis-infected quarters. The percentage of neutrophils that expressed CD14 and the median fluorescence intensity of CD14 in milk neutrophils was also lower in C. bovis-infected quarters.

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

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    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.

  7. Investigation of some hematological and blood biochemical parameters in cattle spontaneously infected with bovine leukosis virus

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    Sandev Nikolay

    2013-09-01

    Full Text Available The aim of the present study was to follow out the alterations in some haematological and blood biochemical parameters in cattle spontaneously infected with enzootic bovine leukosis virus with regard to the invivodifferentiation of bovine leukosis stages. The experiment included 76 cows at various ages and body weight. Serological leukosis tests were done by agar-gel immunodiffusion test with a commercial kit of Synbiotiсs (France, containing standardised gp 51 antigen and positive serum approved by the EU. On the basis of haematological results, the cows were divided into three groups: first group – EBL-seropositive with normal haemogramme; second group – EBL seropositive with altered haemogramme and third group – controls. In cows from the first and the second group, a statistically significantly increased blood cell counts was established compared to healthy controls. The total WBC were increased in the second group (leukocytosis up to 33.21×109/l vs reference range of 5-10×109/l as well as lymphocyte percentages (lymphocytosis – 81.89% (reference 40–63%. A reduction in the proportion of neutrophils to 12.78% (relative neutropenia vs the reference range of 22-49% and monocytes (monocytopenia to 1.78% (reference range 2–6% was observed. A statistically significant reduction in Ca concentrations (4.41 mg/dl and higher inorganic phosphate levels (5.28 mg/dl were established in cows from the second group. Also, ASAT activity was considerably lower – 47.03 U/l, while alkaline phosphatase increased slightly within the reference range up to 167.68 U/l and 165.81 U/l in groups one and two, respectively. The present haematological and whole blood/serum biochemical results in cows spontaneously infected with EBL virus could be used as prognostic markers of the course of the disease, to distinguish the stages of infection with regard to alive diagnostics.

  8. White blood cells, neutrophils, and reactive oxygen metabolites among asymptomatic subjects

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    Kazuhiko Kotani

    2012-01-01

    Full Text Available Objectives: Chronic inflammation and oxidative stress are associated with health and the disease status. The objective of the present study was to investigate the association among white blood cell (WBC counts, neutrophil counts as a WBC subpopulation, and diacron reactive oxygen metabolites (d-ROMs levels in an asymptomatic population. Methods: The clinical data, including general cardiovascular risk variables and high-sensitivity C-reactive protein (hs-CRP, were collected from 100 female subjects (mean age, 62 years in outpatient clinics. The correlation of the d-ROMs with hs-CRP, WBC, and neutrophil counts was examined. Results: The mean/median levels were WBC counts 5.9 × 10 9 /L, neutrophil counts 3.6 × 10 9 /L, hs-CRP 0.06 mg/dL, and d-ROMs 359 CURR U. A simple correlation analysis showed a significant positive correlation of the d-ROMs with the WBC counts, neutrophil counts, or hs-CRP levels. The correlation between d-ROMs and neutrophil counts (β = 0.22, P < 0.05, as well as that between d-ROMs and hs-CRP (β = 0.28, P < 0.01, remained significant and independent in a multiple linear regression analysis adjusted for other variables. A multiple linear regression analysis showed that WBC counts had only a positive correlation tendency to the d-ROMs. Conclusions: Neutrophils may be slightly but more involved in the oxidative stress status, as assessed by d-ROMs, in comparison to the overall WBC. Further studies are needed to clarify the biologic mechanism(s of the observed relationship.

  9. Platelet-neutrophil complex formation-a detailed in vitro analysis of murine and human blood samples.

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    Mauler, Maximilian; Seyfert, Julia; Haenel, David; Seeba, Hannah; Guenther, Janine; Stallmann, Daniela; Schoenichen, Claudia; Hilgendorf, Ingo; Bode, Christoph; Ahrens, Ingo; Duerschmied, Daniel

    2016-05-01

    Platelets form complexes with neutrophils during inflammatory processes. These aggregates migrate into affected tissues and also circulate within the organism. Several studies have evaluated platelet-neutrophil complexes as a marker of cardiovascular diseases in human and mouse. Although multiple publications have reported platelet-neutrophil complex counts, we noticed that different methods were used to analyze platelet-neutrophil complex formation, resulting in significant differences, even in baseline values. We established a protocol for platelet-neutrophil complex measurement with flow cytometry in murine and human whole blood samples. In vitro platelet-neutrophil complex formation was stimulated with ADP or PMA. We tested the effect of different sample preparation steps and cytometer settings on platelet-neutrophil complex detection and noticed false-positive counts with increasing acquisition speed. Platelet-neutrophil complex formation depends on platelet P-selectin expression, and antibody blocking of P-selectin consequently prevented ADP-induced platelet-neutrophil complex formation. These findings may help generating more comparable data among different research groups that examine platelet-neutrophil complexes as a marker for cardiovascular disease and novel therapeutic interventions.

  10. Recruitment of neutrophils across the blood-brain barrier: the role of posttraumatic hepatic ischemia

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    Mantovani Mario

    2003-01-01

    Full Text Available PURPOSE: To study the effects of total hepatic ischemia, and reperfusion on the accumulation of neutrophils in the brain of rats submitted to normovolemic conditions as well as to controlled hemorrhagic shock state. METHODS: Thirty two adult male Wistar rats, were divided into four groups: the Control group, was submitted to the standard procedures for a period of 60 min of observation; Shock group, was submitted to controlled hemorrhagic shock (mean arterial blood pressure=40mmHg, 20min followed by volemic resuscitation (lactated Ringer's solution + blood, 3:1 and reperfusion for 60min; Pringle group, was submitted to total hepatic ischemia for 15min and reperfusion for 60min. The total group was submitted to controlled hemorrhagic shock for 20min followed by volemic resuscitation (lactated Ringer's solution + blood, 3:1, total hepatic ischemia for 15min and reperfusion for 60min. Measurements of serum lactate and base excess were used to characterize the hemorrhagic shock state with low tissue perfusion. The counting of neutrophils on the brain was performed after the euthanasia of animals. RESULTS: The values for the counting of neutrophils on the brain indicate that did not occur difference among studied groups (p=0.196 (Control 0.12± 0.11, Shock 0.12± 0.13, Pringle 0.02± 0.04, Total 0.14± 0.16. CONCLUSION: Hemorrhagic shock associated to total hepatic ischemia for 15 minutes, followed by 60 minutes of reperfusion, did not causes significant neutrophils accumulation in the brain of rats.

  11. Phenotypic, ultra-structural and functional characterization of bovine peripheral blood dendritic cell subsets

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    Dendritic cells (DC) are multifunctional cells that bridge the gap between innate and adaptive immune systems. In bovine, significant information is lacking on the precise identity and role of peripheral blood DC subsets. In this study, we identify and characterize bovine peripheral blood DC subsets...

  12. Gene networks in skeletal muscle following endurance exercise are co-expressed in blood neutrophils and linked with blood inflammation markers.

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    Broadbent, James A; Sampson, Dayle; Sabapathy, Surendran; Haseler, Luke J; Wagner, Karl-Heinz; Bulmer, Andrew Cameron; Peake, Jonathan M; Neubauer, Oliver

    2017-01-19

    It remains incompletely understood whether there is an association between the transcriptome profiles of skeletal muscle and blood leukocytes in response to exercise or other physiological stressors. We have previously analyzed the changes in the muscle and blood neutrophil transcriptome in eight trained men before and 3 h, 48 h and 96 h after 2 h cycling and running. Because we collected muscle and blood in the same individuals and under the same conditions, we were able to directly compare gene expression between the muscle and blood neutrophils. Applying weighted gene co-expression network analysis (WGCNA) as an advanced network-driven method to these original datasets enabled us to compare the muscle and neutrophil transcriptomes in a rigorous and systematic manner. Two gene networks were identified that were preserved between skeletal muscle and blood neutrophils, functionally related to mitochondria and post-translational processes. Strong preservation measures (Zsummary > 10) for both muscle-neutrophil gene networks were evident within the post-exercise recovery period. Muscle and neutrophil gene co-expression was strongly correlated in the mitochondria-related network (r = 0.97; p = 3.17E-2). We also identified multiple correlations between muscular gene sub-networks and exercise-induced changes in blood leukocyte counts, inflammation and muscle damage markers. These data reveal previously unidentified gene co-expression between skeletal muscle and blood neutrophils following exercise, showing the value of WGCNA to understand exercise physiology. Furthermore, these findings provide preliminary evidence in support of the notion that blood neutrophil gene networks may potentially help us to track physiological and pathophysiological changes in the muscle.

  13. CYTOKINE REGULATION OF RESPIRATORY BURST IN BLOOD NEUTROPHILS FOR PREDICTION OF ABDOMINAL SEPSIS IN PATIENTS WITH EXTENDED PURULENT PERITONITIS

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    A. A. Savchenko

    2016-01-01

    Full Text Available The aim of this study was to evaluate correlations between cytokine concentrations and parameters of respiratory burst in blood neutrophils, in order to predict potential development of abdominal sepsis in the patients with extended purulent peritonitis (EPP. The study involved fifteen patients with EPP. Peripheral blood samples were taken on the first day post-admissiona. Abdominal sepsis was diagnosed in thirty-five patients (70% within 5…10 days after surgical intervention. Clinical complications were absent in fifteen patients (30%. Sixty-seven healthy individuals were examined as a control group. Evaluation of the cytokine concentrations was performed by ELISA technique. The degree of respiratory burst in blood neutrophils was measured by means of chemiluminescence assay.It is revealed that the EPP patients exhibited increased levels of serum proinflammatory cytokines and IFNγ, along with higher intensity of respiratory burst in blood neutrophils, due to activated synthesis of of both primary and secondary reactive oxygen species (ROS. The EPP patients who later developed abdominal sepsis showed reduced spontaneous synthesis of primary ROS and increased spontaneous synthesis of secondary ROS. Upon zymosan induction of neutrophils, both primary and secondary ROS levels proved to be similar in the EPP subgroups with or without subsequent sepsis. EPP patients with uncomplicated post-surgical period still exhibited a predominant regulation of respiratory burst of neutrophils by IFNγ activity. Meanwhile, the neutrophil respiratory burst was correlated with TNFα and IL-6 in those patients who further developed abdominal sepsis. A stimulatory effect of IFNγ and a presumably inhibitory action of TNFα and IL-6 upon respiratory burst of blood neutrophils in EPP patients are associated with a release of large cytokine amounts during acute immune inflammatory events, and migration of activated cells to the inflammatory focus. In particular, the

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

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    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.

  15. Effect of acute and chronic excesses of dietary nitrogen on blood neutrophil functions in cattle.

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    Raboisson, D; Caubet, C; Tasca, C; De Marchi, L; Ferraton, J M; Gannac, S; Millet, A; Enjalbert, F; Schelcher, F; Foucras, G

    2014-12-01

    Excess dietary nitrogen (EDN) is commonly expected in dairy herds, but no data are available regarding its consequences on cattle immunity. In this study neutrophil functions were assessed during EDN in steers. In experiment 1, 4 one-month periods, 4 diets [16% crude protein (CP; DM basis), 20% CP based on soybean meal, 20% CP based on urea, and 24% CP based on urea and soybean meal], and 4 steers were included in a crossover design to determine the effects of a chronic excess. In experiment 2, the repercussions of an acute excess were assessed with 2 periods of 10 d, the same 4 steers, and 2 diets containing 14 and 20% CP. Sampling was done during the fourth week of each period in experiment 1, and on d 0, 1, 2, 3, 7, and 9 of each period in experiment 2. Individual blood biochemistry parameters were measured and neutrophil factors, such as counts, recovery after isolation, surface expression of CD11b and CD62L, phagocytosis, diapedesis, reactive oxygen species (ROS) production, and bacteria killing, were determined. Data were analyzed by general linear models of R, with period, diet or biochemical component, and animal as explanatory variables. The outcome variables were biochemical or immune variables. The variables diet, period, and animal were forced as fixed effects. Data collected over the entire period of experiment 2 were pooled. Several multiples linear regressions or ANOVA were performed and a Bonferroni correction was applied. In experiment 2 (acute EDN), neutrophil counts were negatively associated with nitrogen intake, conversely to CD62L expression. The observed relative neutropenia may be due to neutrophil margination because CD62L-expressing neutrophils are more likely to stick to endothelium. Interestingly, ROS production was changed by EDN: chronic EDN (experiment 1) was negatively associated with opsonized zymozan (OZ)-induced ROS production and acute EDN (experiment 2) with spontaneous ROS production. For chronic EDN, ROS production upon

  16. Oral antibiotics increase blood neutrophil maturation and reduce bacteremia and necrotizing enterocolitis in the immediate postnatal period of preterm pigs.

    Science.gov (United States)

    Nguyen, Duc Ninh; Fuglsang, Eva; Jiang, Pingping; Birck, Malene M; Pan, Xiaoyu; Kamal, Shamrulazhar B S; Pors, Susanne E; Gammelgaard, Pernille L; Nielsen, Dennis S; Thymann, Thomas; Levy, Ofer; Frøkiær, Hanne; Sangild, Per T

    2016-01-01

    Immature immunity may predispose preterm neonates to infections and necrotizing enterocolitis (NEC). Intravenous antibiotics are frequently given to prevent and treat sepsis, while oral antibiotics are seldom used. We hypothesized that oral antibiotics promote maturation of systemic immunity and delay gut bacterial colonization and thereby protect preterm neonates against both NEC and bacteremia in the immediate postnatal period. Preterm pigs were given formula and administered saline (CON) or broad-spectrum antibiotics orally (ORA) or systemically (SYS) for 5 d after birth. Temporal changes in blood parameters and bacterial composition in the intestine, blood and immune organs were analyzed. Newborn preterm pigs had few blood neutrophils and a high frequency of progenitor cells. Neutrophils gradually matured after preterm birth with increasing CD14 and decreasing CD172a expressions. Preterm neutrophil and monocyte TLR2 expression and TLR2-mediated blood cytokine responses were low relative to adults. ORA pigs showed enhanced blood neutrophil maturation with reduced cell size and CD172a expression. Only ORA pigs, but not SYS pigs, were protected from a high density of gut Gram-positive bacteria, high gut permeability, Gram-positive bacteremia and NEC. Neonatal oral antibiotics may benefit mucosal and systemic immunity via delayed gut colonization and enhanced blood neutrophil maturation just after preterm birth.

  17. PARTICIPATION OF TLR4 IN ENGULFMENT OF ESCHERICHIA COLI BY HUMAN BLOOD NEUTROPHILS IN PRESENCE OF LIPOPOLYSACCHARIDES

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    S. V. Zubova

    2012-01-01

    Full Text Available Abstract. TLR4 is a key player in signaling system of host cells. Possible role of TLR4 is actively discussed, e.g. its significance for phagocytosis. A capacity of neutrophils to engulf FITC-labeled E. coli bacteria upon activation with LPS of different origin was studied in presence of anti-TLR4 Mab’s (HTA125 clone. It was shown that, in whole blood, TLR4 does not play any essential role in engulfment of bacteria by the neutrophils. Phagocytic activity of neutrophils in blood increases increased after their priming with E. coli endotoxins. LPS from Rb. сapsulatus did not affect phagocytosis. In presence of endotoxins, the degree of TLR4 involvement in neutrophil phagocytosis depends on LPS structure.

  18. Interleukin 17 expression in peripheral blood neutrophils from fungal keratitis patients and healthy cohorts in southern India.

    Science.gov (United States)

    Karthikeyan, Rajapandian Sivaganesa; Vareechon, Chairut; Prajna, Namperumalsamy Venkatesh; Dharmalingam, Kuppamuthu; Pearlman, Eric; Lalitha, Prajna

    2015-01-01

    Interleukin 17A (IL-17) production by peripheral blood neutrophils was examined in patients with fungal keratitis and in uninfected individuals in southern India, which has high levels of airborne Aspergillus and Fusarium conidia. Il17a gene expression and intracellular IL-17 were detected in all groups, although levels were significantly elevated in neutrophils from patients with keratitis. There were no significant differences in plasma IL-17 and IL-23 between patients with keratitis and uninfected individuals; however, combined data from all groups showed a correlation between the percentage IL-17 producing neutrophils and plasma IL-23, and between plasma IL-17 and IL-6 and IL-23.

  19. DNA damage in peripheral blood mononuclear cells and neutrophils of dairy cows during the transition period

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    S. Oikawa

    2012-06-01

    Full Text Available This study was designed to investigate the apoptotic process in peripheral blood mononuclear cells (PBMC and polymorphonuclear neutrophil leukocytes (PMN in dairy cattle during the transition period. Blood samples were collected from 4 dairy cattle at 3 weeks before the expected parturition (wk -3, parturition (wk 0 and 3 weeks after parturition (wk +3. The DNA damage of PBMC and PMN was evaluated based on the comet assay using visual scoring (arbitrary units. Undamaged DNA remained within the core (score 0 and the broken DNA migrated from the core towards the anode forming the tail of a comet (scores 1-4. Significantly higher scores in PBMC at wk 0 and wk +3 were observed compared with those in PMN although there were no significant changes of scores in either cell type during the experimental period. It is suggested that the apoptotic rate of PBMC is accelerated compared with that of PMC during the transition period.

  20. The influence of different anticoagulants and sample preparation methods on measurement of mCD14 on bovine monocytes and polymorphonuclear neutrophil leukocytes

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    Ibeagha-Awemu Eveline M

    2012-02-01

    Full Text Available Abstract Background Membrane-CD14 (mCD14 is expressed on the surface of monocytes, macrophages and polymorphonuclear neutrophil leukocytes (PMN. mCD14 acts as a co-receptor along with Toll like receptor 4 (TLR 4 and MD-2 for the detection of lipopolysaccharide (LPS. However, studies using different sample preparation methods and anticoagulants have reported different levels of mCD14 on the surface of monocytes and neutrophils. In this study, the influence of various anticoagulants and processing methods on measurement of mCD14 on monocytes and neutrophils was examined. Results Whole blood samples were collected in vacutainer tubes containing either sodium heparin (HEPARIN, ethylenediaminetetraacetic acid (EDTA or sodium citrate (CITRATE. mCD14 on neutrophils and monocytes in whole blood samples or isolated cells was measured by the method of flow cytometry using fluorescein isothiocyanate (FITC-labeled monoclonal antibody. There was a significant difference (p p Conclusion From these results, it is suggested that sodium heparin should be the preferred anticoagulant for use in the reliable quantification of the surface expression of mCD14. Furthermore, measurement of mCD14 is best carried out in whole blood samples, both for neutrophils and monocytes.

  1. IFN-gamma is produced by polymorphonuclear neutrophils in human uterine endometrium and by cultured peripheral blood polymorphonuclear neutrophils.

    Science.gov (United States)

    Yeaman, G R; Collins, J E; Currie, J K; Guyre, P M; Wira, C R; Fanger, M W

    1998-05-15

    Cytokines present in the human uterus play an important role both in modulating immune responses to infectious challenge and in the establishment and maintenance of pregnancy. In particular, successful implantation and pregnancy is thought to require the establishment of a Th2 environment, while Th1 cytokines are associated with pregnancy loss and infertility. On the other hand, a Th1 response appears to be required for the resolution of acute infection. Using novel confocal microscopic analysis of fresh sections of human tissue, we have investigated the production of IFN-gamma, a Th1 cytokine, in human endometria. Extracellular IFN-gamma, mostly associated with matrix components, was located immediately beneath the luminal epithelium and along the glandular epithelium proximal to the lumen. As evidenced by intracellular staining, IFN-gamma is produced by both stromal cells and intraepithelial lymphocytes through all stages of the menstrual cycle. Surprisingly, the stromal cell containing intracellular IFN-gamma was identified as a polymorphonuclear neutrophil on the basis of its reactivity with a panel of mAbs and its nuclear morphology. We further found that polymorphonuclear neutrophils isolated from normal donors produce IFN-gamma in response to stimulation with LPS, IL-12, and TNF-alpha. Taken together, these findings suggest that polymorphonuclear neutrophils are capable of producing IFN-gamma both in vitro and in vivo, indicating that their role in shaping immune responses may be more extensive than previously thought. Furthermore, these studies strongly suggest that polymorphonuclear neutrophils play an important role in determining immune responsiveness within the female reproductive tract.

  2. Effect of bovine lactoferrin on functions of activated feline peripheral blood mononuclear cells during chronic feline immunodeficiency virus infection.

    Science.gov (United States)

    Kobayashi, Saori; Sato, Reeko; Aoki, Takako; Omoe, Katsuhiko; Inanami, Osamu; Hankanga, Careen; Yamada, Yuichi; Tomizawa, Nobuyuki; Yasuda, Jun; Sasaki, Juso

    2008-05-01

    Feline immunodeficiency virus (FIV) infection is characterized by chronic overactivation of immune and inflammatory system, resulting in anergic state and dysfunction of immune cells. Lactoferrin (LF), a glycoprotein present in exocrine secretions and neutrophils, plays an important role in host defense system. Our previous study showed that oral administration of bovine LF (bLF) suppressed oral inflammation, improved the clinical symptoms and decreased serum gamma-globulin as a marker of inflammation in FIV-infected cats with intractable stomatitis. The anti-inflammatory effect was partly involved in regulation of neutrophil function by bLF. In this study, to clarify the relationship between anti-inflammatory effects of bLF and peripheral blood mononuclear cells (PBMC), we examined the effect of bLF on proliferation, cell cycle progression and cytokine expression in mitogen-activated PBMC. MTT [3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl tetrazolium bromide] assay showed that bLF inhibited the concanavalin A (ConA)-induced cell proliferation in FIV-infected cats with the asymptomatic carrier and AIDS-related complex (ARC) phase. Bovine LF restored ConA-induced cell cycle progression and resulted in suppression of the induced apoptosis in feline PBMC. Real-time RT-PCR showed that bLF suppressed ConA-induced expression of interferon-gamma and interleukin-2 in cells of the ARC group regardless of the time of its addition to the medium. These results suggest the hypothesis that therapy with bLF may have the potential to improve and protect functions of overactivated lymphocytes by modulating the cell proliferation, cell cycle and cytokines expression in cats in terminal stage of FIV infection.

  3. Arbutin and decrease of potentially toxic substances generated in human blood neutrophils.

    Science.gov (United States)

    Pečivová, Jana; Nosál', Radomír; Sviteková, Klára; Mačičková, Tatiana

    2014-12-01

    Neutrophils, highly motile phagocytic cells, constitute the first line of host defense and simultaneously they are considered to be central cells of chronic inflammation. In combination with standard therapeutic procedures, natural substances are gaining interest as an option for enhancing the effectiveness of treatment of inflammatory diseases. We investigated the effect of arbutin and carvedilol and of their combination on 4β-phorbol-12β-myristate-13α-acetate- stimulated functions of human isolated neutrophils. Cells were preincubated with the drugs tested and subsequently stimulated. Superoxide (with or without blood platelets, in the rate close to physiological conditions [1:50]) and HOCl generation, elastase and myeloperoxidase release were determined spectrophotometrically and phospholipase D activation spectrofluorometrically. The combined effect of arbutin and carvedilol was found to be more effective than the effect of each compound alone. Our study provided evidence supporting the potential beneficial effect of arbutin alone or in combination with carvedilol in diminishing tissue damage by decreasing phospholipase D, myeloperoxidase and elastase activity and by attenuating the generation of superoxide and the subsequently derived reactive oxygen species. The presented data indicate the ability of arbutin to suppress the onset and progression of inflammation.

  4. Enhancement of neutrophil-mediated killing of Plasmodium falciparum asexual blood forms by fatty acids: importance of fatty acid structure.

    Science.gov (United States)

    Kumaratilake, L M; Ferrante, A; Robinson, B S; Jaeger, T; Poulos, A

    1997-10-01

    Effects of fatty acids on human neutrophil-mediated killing of Plasmodium falciparum asexual blood forms were investigated by using a quantitative radiometric assay. The results showed that the antiparasitic activity of neutrophils can be greatly increased (>threefold) by short-term treatment with fatty acids with 20 to 24 carbon atoms and at least three double bonds. In particular, the n-3 polyenoic fatty acids, eicosapentaenoic and docosahexaenoic acids, and the n-6 fatty acid, arachidonic acid, significantly enhanced neutrophil antiparasitic activity. This effect was >1.5-fold higher than that induced by an optical concentration of the known agonist cytokine tumor necrosis factor alpha (TNF-alpha). At suboptimal concentrations, the combination of arachidonic acid and TNF-alpha caused a synergistic increase in neutrophil-mediated parasite killing. The fatty acid-induced effect was independent of the availability of serum opsonins but dependent on the structure of the fatty acids. The length of the carbon chain, degree of unsaturation, and availability of a free carboxyl group were important determinants of fatty acid activity. The fatty acids which increased neutrophil-mediated killing primed the enhanced superoxide radical generation of neutrophils in response to P. falciparum as detected by chemiluminescence. Scavengers of oxygen radicals significantly reduced the fatty acid-enhanced parasite killing, but cyclooxygenase and lipoxygenase inhibitors had no effect. These findings have identified a new class of immunoenhancers that could be exploited to increase resistance against Plasmodium species.

  5. Epinephrine enhances platelet-neutrophil adhesion in whole blood in vitro.

    NARCIS (Netherlands)

    Horn, N.A.; Anastase, D.M.; Hecker, K.E.; Baumert, J.H.; Robitzsch, T.; Rossaint, R.

    2005-01-01

    Previous studies showed that alpha- or beta-adrenoceptor stimulation by catecholamines influenced neutrophil function, cytokine liberation, and platelet aggregability. We investigated whether adrenergic stimulation with epinephrine also alters platelet-neutrophil adhesion. This might be of specific

  6. Impact of total body irradiation on successful neutrophil engraftment in unrelated bone marrow or cord blood transplantation.

    Science.gov (United States)

    Nakasone, Hideki; Fuji, Shigeo; Yakushijin, Kimikazu; Onizuka, Makoto; Shinohara, Akihito; Ohashi, Kazuteru; Miyamura, Koichi; Uchida, Naoyuki; Takanashi, Minoko; Ichinohe, Tatsuo; Atsuta, Yoshiko; Fukuda, Takahiro; Ogata, Masao

    2017-02-01

    Total body irradiation (TBI) has been thought to promote donor cell engraftment in allogeneic hematopoietic cell transplantation (HCT) from alternative donors. However, recent progress in HCT strategies may affect the clinical significance of TBI on neutrophil engraftment. With the use of a Japanese transplant registry database, we analyzed 3933 adult recipients (>15 y.o.) who underwent HCT between 2006 and 2013 from an 8/8 HLA-matched unrelated bone marrow donor (MUD, n = 1367), an HLA-mismatched unrelated bone marrow donor (MMUD, n = 1102), or unrelated cord blood (CBT, n = 1464). Conditioning regimens were divided into five groups: High-TBI-(>8Gy), Low-TBI- (≤8Gy), and no-TBI-myeloablative conditioning (MAC), and Low-TBI- and no-TBI-reduced-intensity conditioning (RIC). In both MUD and MMUD, neutrophil engraftment rate was >90% in each of the five conditioning groups, and TBI was not associated with prompt neutrophil engraftment in multivariate analyses. Conversely, in CBT, TBI regimens had a higher rate of day-30 neutrophil engraftment than no-TBI-regimens: 78% in High-TBI-MAC, 83% in Low-TBI-MAC, and 76% in Low-TBI-RIC versus 65% in No-TBI-MAC, and 68% in No-TBI-RIC (P < .001). Multivariate analyses in CBT demonstrated that TBI-regimens were significantly associated with a higher rate of neutrophil engraftment. Subsequently focusing on CBT patients alone, TBI-regimens were significantly associated with a higher rate of neutrophil engraftment in patients who received CBT with a 4/6 or less HLA allele-match, or who had anti-HLA antibodies. In summary, TBI-regimens had no impact on neutrophil engraftment in the current practice of unrelated bone marrow transplantation. However, in CBT, TBI is still necessary to enhance engraftment.

  7. The subcellular particulate NADPH-dependent O2.(-)-generating oxidase from human blood monocytes: comparison to the neutrophil system.

    Science.gov (United States)

    Chaudhry, A N; Santinga, J T; Gabig, T G

    1982-10-01

    Highly purified preparations of normal human monocytes obtained from peripheral blood were shown to contain a subcellular particulate O2.(-)-generating oxidase system. This O2.(-)-generating activity was present in particulate preparations from monocytes that had been previously stimulated with phorbol myristate acetate but was low or absent in control preparations from unstimulated monocytes or stimulated monocytes from a patient with chronic granulomatous disease. In the stimulated preparations from normal monocytes, O2.(-)-generation was linearly proportional to cell protein concentration, insensitive to inhibition by azide, and dependent on NADPH as substrate. These characteristics are similar to the O2.(-)-generating oxidase system from human neutrophils. A significant difference in the apparent Km for NADPH was shown between preparations from stimulated monocytes and neutrophils (monocyte 83 +/- 16 microM, neutrophil 31 +/- 5 microM, mean +/- SE). Additionally, affinity of the stimulated monocyte particulate preparation for NADH was unmeasurably low.

  8. Blood Level of Polymorphonuclear Neutrophil Leukocytes and Bronchial Hyperreactivity in Chronic Obstructive Pulmonary Disease

    Science.gov (United States)

    Cukic, Vesna

    2015-01-01

    Introduction: Polymorphonuclear neutrophil leukocytes (PMNL) have an important defensive role against various microorganisms and other agents, but by liberating various substances, first of all the superoxide anion (O 2¯), they can damage the bronchial mucosa and influence the development of bronchial inflammation which is the fundamental of bronchial hyperreactivity (BHR). Objective: to show the role of the PMNL for development and level of BHR in patients with chronic obstructive pulmonary disease (COPD). Material and methods: We observed 160 patients with COPD treated in Clinic for Pulmonary Diseases and TB “Podhrastovi” Sarajevo during three years :from 2012 to 2014. They were divided into groups and subgroups according to the first registration of BHR in the course of illness and to the number of exacerbations of the disease in one year. The number of blood PMNL was measured in a stable state of disease at the begging and at the end of investigation. Results: The number of blood PMNL was significantly greater in patients with 3 or more exacerbations per one year (p <0.01). Patients with BHR had significantly greater number blood PMNL than patients without BHR (p< 0.05). Patients with 3 exacerbations per year had a statistically significant increase of number of PMNL between first and last examination (p<0.01). Conclusion: There is statistically significant correlation between the number of blood PMNL and the level of BHR in COPD, but future examination need to be done to determine real role and mode of action of PMNL for these processes. PMID:26543311

  9. Greater expression of TLR2, TLR4, and IL6 due to negative energy balance is associated with lower expression of HLA-DRA and HLA-A in bovine blood neutrophils after intramammary mastitis challenge with Streptococcus uberis

    DEFF Research Database (Denmark)

    Moyes, Kasey; Drackley, James K; Morin, Dawn E;

    2010-01-01

    response and metabolism. A total of 12 genes were differentially expressed in blood PMN in NEB versus PEB cows. Upregulated genes by NEB were ALOX5AP, CPNE3, IL1R2, IL6, TLR2, TLR4, and THY1, and downregulated genes were HLA-DRA, HLA-A, IRAK1, SOD1, and TNF. Network analysis revealed that TNF...... with NEB and the corresponding inhibition of immune response in lactating dairy cows....

  10. β2 integrin-mediated crawling on endothelial ICAM-1 and ICAM-2 is a prerequisite for transcellular neutrophil diapedesis across the inflamed blood-brain barrier.

    Science.gov (United States)

    Gorina, Roser; Lyck, Ruth; Vestweber, Dietmar; Engelhardt, Britta

    2014-01-01

    In acute neuroinflammatory states such as meningitis, neutrophils cross the blood-brain barrier (BBB) and contribute to pathological alterations of cerebral function. The mechanisms that govern neutrophil migration across the BBB are ill defined. Using live-cell imaging, we show that LPS-stimulated BBB endothelium supports neutrophil arrest, crawling, and diapedesis under physiological flow in vitro. Investigating the interactions of neutrophils from wild-type, CD11a(-/-), CD11b(-/-), and CD18(null) mice with wild-type, junctional adhesion molecule-A(-/-), ICAM-1(null), ICAM-2(-/-), or ICAM-1(null)/ICAM-2(-/-) primary mouse brain microvascular endothelial cells, we demonstrate that neutrophil arrest, polarization, and crawling required G-protein-coupled receptor-dependent activation of β2 integrins and binding to endothelial ICAM-1. LFA-1 was the prevailing ligand for endothelial ICAM-1 in mediating neutrophil shear resistant arrest, whereas Mac-1 was dominant over LFA-1 in mediating neutrophil polarization on the BBB in vitro. Neutrophil crawling was mediated by endothelial ICAM-1 and ICAM-2 and neutrophil LFA-1 and Mac-1. In the absence of crawling, few neutrophils maintained adhesive interactions with the BBB endothelium by remaining either stationary on endothelial junctions or displaying transient adhesive interactions characterized by a fast displacement on the endothelium along the direction of flow. Diapedesis of stationary neutrophils was unchanged by the lack of endothelial ICAM-1 and ICAM-2 and occurred exclusively via the paracellular pathway. Crawling neutrophils, although preferentially crossing the BBB through the endothelial junctions, could additionally breach the BBB via the transcellular route. Thus, β2 integrin-mediated neutrophil crawling on endothelial ICAM-1 and ICAM-2 is a prerequisite for transcellular neutrophil diapedesis across the inflamed BBB.

  11. [The participation of neutrophilic mechanisms in the pathogenesis of chronic elevated blood immune complexes].

    Science.gov (United States)

    Bidiuk, M M; Chop'iak, V V; Liubinets', L A; Pavlovich, S I; Nykytiuk, H P; Porokhovs'ka, Z S

    1997-01-01

    For reproduction of chronic hyperimmunocomplexemia model the classic Cochrane C. G., 1973 elaboration was used. The circulating immune complexes (CIC) were identified by precipitation methods, and fixated--by immunofluorescent in endothelium reproduction of aorta, and their clearance organs--liver and spleen. The estimation of neutrophil status was carried out according to rosette-forming neutrophils ability--by latex, zimosane, mieloperoxidasa, NST tests. The growth of large and middle CIC in vessel bed was estimated, their fixation in aorta bifurcation, and also on liver and spleen calls, but their intensivity is less, comparing to aorta endothelium. The neutrophil status was characterised by O-neutrophils growth, strengthening of capturing macrophages function, considerable activation of fermentative polynuclear systems, although reserve possibilities in them grew in less degree. Morphologic investigations allow to speak mediatingly about neutrophils participation in endotheliocytes damage, and also about their ability to secure the mononuclear cells further participation in damaging of aorta walls and partially of liver and spleen.

  12. Role of gastric blood flow, neutrophil infiltration, and mucosal cell proliferation in gastric adaptation to aspirin in the rat.

    Science.gov (United States)

    Konturek, S J; Brzozowski, T; Stachura, J; Dembinski, A; Majka, J

    1994-01-01

    Gastric mucosa exhibits the ability to adapt to ulcerogenic action of aspirin but the mechanism of this phenomenon is unknown. In this study, acute gastric lesions were produced by single or repeated doses of acidified aspirin in rats with intact or resected salivary glands and with intact or suppressed synthase of nitric oxide. A single oral dose of aspirin produced a dose dependent increase in gastric lesions accompanied by considerable blood neutrophilia and mucosal neutrophil infiltration, significant reduction in gastric blood flow, and almost complete suppression of biosynthesis of prostaglandins. After rechallenge with aspirin, the mucosal damage became smaller and progressively declined with repeated aspirin insults. Gastric adaptation to aspirin was accompanied by a significant rise in gastric blood flow, reduction in both blood neutrophilia and mucosal neutrophil infiltration, and a remarkable increase in mucosal cell regeneration and mucosal content of epidermal growth factor. Salivectomy, which reduced the mucosal content of epidermal growth factor, aggravated the initial mucosal damage induced by the first exposure to acidified aspirin but did not prevent the adaptation of this mucosa to repeated aspirin insults. Pretreatment with NG-nitro-L-arginine (L-NNA), a specific inhibitor of nitric oxide synthase, eliminated the hyperaemic response to repeated aspirin but did not abolish the development of adaptation to aspirin showing that the maintenance of the gastric blood flow plays little part in this adaptation. In conclusion, the stomach adapts readily to repeated aspirin insults and this is accompanied by a considerable reduction in blood neutrophilia and the severity of neutrophil infiltration and by an extensive proliferation of mucosal cells possibly involving epidermal growth factor. PMID:7525421

  13. Reduced expression of C5a receptors on neutrophils from cord blood

    DEFF Research Database (Denmark)

    Nybo, Mads; Sørensen, O; Leslie, R;

    1998-01-01

    MLP was tested by measuring migration and exocytosis of myeloperoxidase and lactoferrin. RESULTS: C5a mean fluorescence on neutrophils from neonates was significantly lower (22.4 (SD 3.5)) than in adult controls (31.5 (3.1)). Neutrophils from neonates migrated poorly towards both C5a and fMLP compared with those...... from adult controls. Exocytosis of myeloperoxidase, but not lactoferrin from neonatal neutrophils stimulated with C5a, was significantly lower than in adult controls. fMLP stimulation, on the other hand, resulted in significantly higher exocytosis in neonates. CONCLUSION: The lower expression of C5a...... receptors on neutrophils from neonates could be related to reduced C5a mediated exocytosis of myeloperoxidase....

  14. Rapid Detection of Neutrophil Oxidative Burst Capacity is Predictive of Whole Blood Cytokine Responses.

    Directory of Open Access Journals (Sweden)

    Philip J Vernon

    Full Text Available Maladaptive immune responses, particularly cytokine and chemokine-driven, are a significant contributor to the deleterious inflammation present in many types of injury and infection. Widely available applications to rapidly assess individual inflammatory capacity could permit identification of patients at risk for exacerbated immune responses and guide therapy. Here we evaluate neutrophil oxidative burst (NOX capacity measured by plate reader to immuno-type Rhesus Macaques as an acute strategy to rapidly detect inflammatory capacity and predict maladaptive immune responses as assayed by cytokine array.Whole blood was collected from anesthetized Rhesus Macaques (n = 25 and analyzed for plasma cytokine secretion (23-plex Luminex assay and NOX capacity. For cytokine secretion, paired samples were either unstimulated or ex-vivo lipopolysaccharide (LPS-stimulated (100μg/mL/24h. NOX capacity was measured in dihydrorhodamine-123 loaded samples following phorbol 12-myristate 13-acetate (PMA/ionomycin treatment. Pearson's test was utilized to correlate NOX capacity with cytokine secretion, p<0.05 considered significant.LPS stimulation induced secretion of the inflammatory molecules G-CSF, IL-1β, IL-1RA, IL-6, IL-10, IL-12/23(p40, IL-18, MIP-1α, MIP-1β, and TNFα. Although values were variable, several cytokines correlated with NOX capacity, p-values≤0.0001. Specifically, IL-1β (r = 0.66, IL-6 (r = 0.74, the Th1-polarizing cytokine IL-12/23(p40 (r = 0.78, and TNFα (r = 0.76 were strongly associated with NOX.NOX capacity correlated with Th1-polarizing cytokine secretion, indicating its ability to rapidly predict inflammatory responses. These data suggest that NOX capacity may quickly identify patients at risk for maladaptive immune responses and who may benefit from immuno-modulatory therapies. Future studies will assess the in-vivo predictive value of NOX in animal models of immune-mediated pathologies.

  15. The effect of citrus-derived oil on bovine blood neutrophil response in vitro

    Science.gov (United States)

    Research on the use of natural products to treat or prevent microbial invasion as alternatives to antibiotic use is growing.Polymorphonuclear leukocytes (PMNL) play a vital role with regard to the innate immune response that affects severity and or duration of mastitis. To our knowledge, effect of c...

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

    Science.gov (United States)

    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.

  17. Systemic inflammation in COPD visualised by gene profiling in peripheral blood neutrophils

    NARCIS (Netherlands)

    Oudijk, EJD; Nijhuis, EHJ; Zwank, MD; van de Graaf, EA; Mager, HJ; Coffer, PJ; Lammers, JWJ; Koenderman, L

    2005-01-01

    Background: The inflammatory process in chronic obstructive pulmonary disease ( COPD) is characterised by the presence of neutrophils in the lung that are able to synthesise de novo several inflammatory mediators. The local chronic persistent inflammatory response is accompanied by systemic effects

  18. Phenotypic, ultra-structural, and functional characterization of bovine peripheral blood dendritic cell subsets.

    Directory of Open Access Journals (Sweden)

    Janet J Sei

    Full Text Available Dendritic cells (DC are multi-functional cells that bridge the gap between innate and adaptive immune systems. In bovine, significant information is lacking on the precise identity and role of peripheral blood DC subsets. In this study, we identify and characterize bovine peripheral blood DC subsets directly ex vivo, without further in vitro manipulation. Multi-color flow cytometric analysis revealed that three DC subsets could be identified. Bovine plasmacytoid DC were phenotypically identified by a unique pattern of cell surface protein expression including CD4, exhibited an extensive endoplasmic reticulum and Golgi apparatus, efficiently internalized and degraded exogenous antigen, and were the only peripheral blood cells specialized in the production of type I IFN following activation with Toll-like receptor (TLR agonists. Conventional DC were identified by expression of a different pattern of cell surface proteins including CD11c, MHC class II, and CD80, among others, the display of extensive dendritic protrusions on their plasma membrane, expression of very high levels of MHC class II and co-stimulatory molecules, efficient internalization and degradation of exogenous antigen, and ready production of detectable levels of TNF-alpha in response to TLR activation. Our investigations also revealed a third novel DC subset that may be a precursor of conventional DC that were MHC class II+ and CD11c-. These cells exhibited a smooth plasma membrane with a rounded nucleus, produced TNF-alpha in response to TLR-activation (albeit lower than CD11c+ DC, and were the least efficient in internalization/degradation of exogenous antigen. These studies define three bovine blood DC subsets with distinct phenotypic and functional characteristics which can be analyzed during immune responses to pathogens and vaccinations of cattle.

  19. Neutrophils, from marrow to microbes

    DEFF Research Database (Denmark)

    Borregaard, Niels

    2010-01-01

    . Neutrophils circulate in the blood as dormant cells. At sites of infection, endothelial cells capture bypassing neutrophils and guide them through the endothelial cell lining whereby the neutrophils are activated and tuned for the subsequent interaction with microbes. Once in tissues, neutrophils kill...... microorganisms by microbicidal agents liberated from granules or generated by metabolic activation. As a final act, neutrophils can extrude stands of DNA with bactericidal proteins attached that act as extracellular traps for microorganisms....

  20. Aldehyde dehydrogenase-independent bioactivation of nitroglycerin in porcine and bovine blood vessels.

    Science.gov (United States)

    Neubauer, Regina; Wölkart, Gerald; Opelt, Marissa; Schwarzenegger, Christine; Hofinger, Marielies; Neubauer, Andrea; Kollau, Alexander; Schmidt, Kurt; Schrammel, Astrid; Mayer, Bernd

    2015-02-15

    The vascular bioactivation of the antianginal drug nitroglycerin (GTN), yielding 1,2-glycerol dinitrate and nitric oxide or a related activator of soluble guanylate cyclase, is catalyzed by aldehyde dehydrogenase-2 (ALDH2) in rodent and human blood vessels. The essential role of ALDH2 has been confirmed in many studies and is considered as general principle of GTN-induced vasodilation in mammals. However, this view is challenged by an early report showing that diphenyleneiodonium, which we recently characterized as potent ALDH2 inhibitor, has no effect on GTN-induced relaxation of bovine coronary arteries (De La Lande et al., 1996). We investigated this issue and found that inhibition of ALDH2 attenuates GTN-induced coronary vasodilation in isolated perfused rat hearts but has no effect on relaxation to GTN of bovine and porcine coronary arteries. This observation is explained by low levels of ALDH2 protein expression in bovine coronary arteries and several types of porcine blood vessels. ALDH2 mRNA expression and the rates of GTN denitration were similarly low, excluding a significant contribution of ALDH2 to the bioactivation of GTN in these vessels. Attempts to identify the responsible pathway with enzyme inhibitors did not provide conclusive evidence for the involvement of ALDH3A1, cytochrome P450, or GSH-S-transferase. Thus, the present manuscript describes a hitherto unrecognized pathway of GTN bioactivation in bovine and porcine blood vessels. If present in the human vasculature, this pathway might contribute to the therapeutic effects of organic nitrates that are not metabolized by ALDH2.

  1. СHANGES IN PARAMETERS OF LUMINOL-DEPENDENT AND LUCIGENIN-DEPENDENT CHEMILUMINESCENCE OF PERIPHERAL BLOOD NEUTROPHILS IN PATIENTS WITH BLADDER CANCER IN THE DISEASE DYNAMICS

    Directory of Open Access Journals (Sweden)

    L. M. Kurtasova

    2015-01-01

    Full Text Available The study deals with parameters of luminol-dependent and lucigenin-dependent chemiluminescence (CL of peripheral blood neutrophils from patients with bladder cancer (BC prior to surgical treatment. We examined sixty patients (45 to 55 years old with advanced bladder cancer (TNM prior to the operation, and forty-six patients at 10 days after surgical treatment. A control group consisted of 56 healthy donors. Luminol-dependent and lucigenin-dependent chemiluminescence of blood neutrophils was assessed according to De Sole et al. (1983. Chemiluminescence assays of peripheral blood neutrophils from the patients with bladder cancer revealed changes in production of reactive oxygen species (ROS, both for initial stage of oxidation reaction, and total level of active oxygen radicals. We have found disturbed values of primary-to-secondary ROS ratio in the cells. In the patients with bladder cancer, some changes in oxidative metabolism of the blood neutrophils have been registered. These alterations may play an important role in promotion of potential effector cell functions, thus, probably, affecting the whole-scale development of a cytopathic effect exerted by neutrophilic granulocytes. 

  2. Blood Level of Polymorphonuclear Neutrophil Leukocytes and Bronchial Hyperreactivity in Chronic Obstructive Pulmonary Disease

    OpenAIRE

    Cukic, Vesna

    2015-01-01

    Introduction: Polymorphonuclear neutrophil leukocytes (PMNL) have an important defensive role against various microorganisms and other agents, but by liberating various substances, first of all the superoxide anion (O 2¯), they can damage the bronchial mucosa and influence the development of bronchial inflammation which is the fundamental of bronchial hyperreactivity (BHR). Objective: to show the role of the PMNL for development and level of BHR in patients with chronic obstructive pulmonary ...

  3. Mucosal adaptation to aspirin induced gastric damage in humans. Studies on blood flow, gastric mucosal growth, and neutrophil activation.

    Science.gov (United States)

    Konturek, J W; Dembinski, A; Stoll, R; Domschke, W; Konturek, S J

    1994-01-01

    The gastropathy associated with the ingestion of non-steroidal anti-inflammatory drugs (NSAIDs) such as aspirin is a common side effect of this class of drugs, but the precise mechanisms by which they cause mucosal damage have not been fully explained. During continued use of an injurious substance, such as aspirin, the extent of gastric mucosal damage decreases and this phenomenon is named gastric adaptation. To assess the extent of mucosal damage by aspirin and subsequent adaptation the effects of 14 days of continuous, oral administration of aspirin (2 g per day) to eight healthy male volunteers was studied. To estimate the rate of mucosal damage, gastroscopy was performed before (day 0) and at days 3, 7, 14 of aspirin treatment. Gastric microbleeding and gastric mucosal blood flow were measured using laser Doppler flowmeter and mucosal biopsy specimens were taken for the estimation of tissue DNA synthesis and RNA and DNA concentration. In addition, the activation of neutrophils in peripheral blood was assessed by measuring their ability to associate with platelets. Aspirin induced acute damage mainly in gastric corpus, reaching at day 3 about 3.5 on the endoscopic Lanza score but lessened to about 1.5 at day 14 pointing to the occurrence of gastric adaptation. Mucosal blood flow increased at day 3 by about 50% in the gastric corpus and by 88% in the antrum. The in vitro DNA synthesis and RNA concentration, an index of mucosal growth, were reduced at day 3 but then increased to reach about 150% of initial value at the end of aspirin treatment. It is concluded that the treatment with aspirin in humans induces gastric adaptation to this agent, which entails the increase in mucosal blood flow, the rise in neutrophil activation, and the enhancement in mucosal growth. PMID:7959223

  4. Glucose supplementation has minimal effects on blood neutrophil functionand gene expression in vitro

    Science.gov (United States)

    During early lactation, glucose availability is low and the effect of glucose supply on bovine polymorphonuclear leukocyte (PMNL) function is poorly understood. The objective of this study was to determine the effect of glucose supplementation on the function and transcriptomic inflammatory respons...

  5. The Numerical Predominance and Large Transcriptome Differences of Neutrophils in Peripheral Blood Together Inevitably Account for a Reported Pulmonary Tuberculosis Signature

    Science.gov (United States)

    Deng, Wang-Long; Zhang, Hao

    2017-01-01

    Previous transcriptomic analysis revealed a 393-transcript signature (PTBsig), which is dominated by interferon inducible genes, in whole blood of pulmonary tuberculosis (PTB) patients. Comparisons with a limited set of interferon-driven genes among separated monocytes, CD4+ T cells, CD8+ T cells, and neutrophils indicated that the signature is due to changes in neutrophils, the overwhelmingly predominant cell type. By extending the analysis to the entire 393 transcripts of PTBsig and by switching the cell proportions between separated monocytes, CD4+ T cells, CD8+ T cells, and neutrophils, we create putative PTBsig for whole blood (pPTBsig) in which CD4+ or CD8+ T cells or monocytes predominated or in which the cell proportions were unchanged. These putative signatures are then compared to the actual reported PTBsig. We show that, because of their predominance in peripheral blood and their larger transcriptional responses, neutrophils were indeed almost exclusively responsible for PTBsig. We caution that the functional significance of changes in other cell types might escape notice in transcriptome analysis that is based upon whole blood.

  6. Isolation of Mouse Neutrophils.

    Science.gov (United States)

    Swamydas, Muthulekha; Luo, Yi; Dorf, Martin E; Lionakis, Michail S

    2015-08-03

    Neutrophils represent the first line of defense against bacterial and fungal pathogens. Indeed, patients with inherited and acquired qualitative and quantitative neutrophil defects are at high risk for developing bacterial and fungal infections and suffering adverse outcomes from these infections. Therefore, research aiming at defining the molecular factors that modulate neutrophil effector function under homeostatic conditions and during infection is essential for devising strategies to augment neutrophil function and improve the outcome of infected individuals. This unit describes a reproducible density gradient centrifugation-based protocol that can be applied in any laboratory to harvest large numbers of highly enriched and highly viable neutrophils from the bone marrow of mice both at the steady state and following infection with Candida albicans as described in UNIT. In another protocol, we also present a method that combines gentle enzymatic tissue digestion with a positive immunomagnetic selection technique or Fluorescence-activated cell sorting (FACS) to harvest highly pure and highly viable preparations of neutrophils directly from mouse tissues such as the kidney, the liver or the spleen. Finally, methods for isolating neutrophils from mouse peritoneal fluid and peripheral blood are included. Mouse neutrophils isolated by these protocols can be used for examining several aspects of cellular function ex vivo including pathogen binding, phagocytosis and killing, neutrophil chemotaxis, oxidative burst, degranulation and cytokine production, and for performing neutrophil adoptive transfer experiments.

  7. Health status of birds fed diets containing three differently processed discarded vegetable-bovine blood-rumen content mixtures.

    Science.gov (United States)

    Ekunseitan, D A; Balogun, O O; Sogunle, O M; Yusuf, A O; Ayoola, A A; Egbeyale, L T; Adeyemi, O A; Allison, I B; Iyanda, A I

    2013-04-01

    This study was conducted to determine the effect of feeding three differently processed mixtures on health status of broilers. A total of 1080 day-old Marshal broilers were fed; discarded vegetable-fresh bovine blood-fresh rumen digesta (P1), discarded vegetable-ensiled bovine blood-fresh rumen digesta (P2) and discarded vegetable-fresh bovine blood-ensiled rumen digesta (P3) at three levels of inclusion (0, 3 and 6%). Data on blood parameters was taken and were subjected to 3 x 3 factorial arrangements in a completely randomized design. Birds fed P1 had least values (p cell volume, Haemoglobin, White blood cell and Red blood cell values. However, those fed at 0% level of inclusion recorded the highest albumin value. At finisher phase, birds fed P2 and P3 had the highest glucose, uric acid and creatinine values. 6% level of inclusion significantly (p vegetable-fresh bovine blood-ensiled rumen digesta (P3) up to 6% level of inclusion.

  8. Functional neutrophils from human ES cells

    OpenAIRE

    Sweeney, Colin L; Malech, Harry L.

    2009-01-01

    In this issue of Blood, Yokoyama and colleagues demonstrate in vitro differentiation of hESCs into mature neutrophils with functional capabilities (chemotaxis, phagocytosis, microbicidal oxidase activity, and bacterial killing) approaching or equal to that of normal peripheral blood neutrophils.

  9. Evaluation of a simple Theileria annulata culture protocol from experimentally infected bovine whole blood

    Directory of Open Access Journals (Sweden)

    Gharbi M.

    2012-08-01

    Full Text Available We have evaluated a new simple technique using whole blood from experimentally infected cattle for the isolation and cultivation of Theileria annulata. The study was carried out on 20 Holstein-Frisian bovines that had been experimentally infected with a virulent lethal dose of Theileria annulata. This technique has been compared to the classical peripheral blood monocyte isolation with Ficoll carried out on 22 experimentally infected Holstein-Friesian calves. The effectiveness of the reference technique was estimated to 86.4%, whilst the effectiveness of the new technique was 100%. Moreover, this new technique leads to time and money saving estimated to € 3.06 per sample. It decreases the contamination risks by reducing the steps of sample manipulation.

  10. Engineered stealth porous silicon nanoparticles via surface encapsulation of bovine serum albumin for prolonging blood circulation in vivo.

    Science.gov (United States)

    Xia, Bing; Zhang, Wenyi; Shi, Jisen; Xiao, Shou-jun

    2013-11-27

    Luminescent porous silicon nanoparticles (PSiNPs) have been widely used as drug delivery. However, fast biodegradation and short blood circulation have been major challenges for their biomedical applications. Herein, bovine serum albumin was readily encapsulated onto alkyl-terminated PSiNPs surfaces via hydrophobic interaction, which could significantly improve their water-dispersibility and long-term stability under physiological conditions. Furthermore, compared with PSiNPs alone, PSiNPs coated with bovine serum albumin remarkably reduced nonspecific cellular uptake in vitro and prolonged blood circulation in vivo.

  11. Evaluation of Toll-like, chemokine, and integrin receptors on monocytes and neutrophils from peripheral blood of septic patients and their correlation with clinical outcomes

    Energy Technology Data Exchange (ETDEWEB)

    Silva, S.C.; Baggio-Zappia, G.L.; Brunialti, M.K.C. [Universidade Federal de São Paulo, Escola Paulista de Medicina, Hospital São Paulo, Disciplina de Infectologia, Departamento de Medicina, São Paulo, SP, Brasil, Disciplina de Infectologia, Departamento de Medicina, Hospital São Paulo, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Assunçao, M.S.C. [Hospital Israelita Albert Einstein, Unidade de Terapia Intensiva, São Paulo, SP, Brasil, Unidade de Terapia Intensiva, Hospital Israelita Albert Einstein, São Paulo, SP (Brazil); Azevedo, L.C.P. [Hospital Sírio Libanês, Unidade de Terapia Intensiva, São Paulo, SP, Brasil, Unidade de Terapia Intensiva, Hospital Sírio Libanês, São Paulo, SP (Brazil); Machado, F.R. [Universidade Federal de São Paulo, Escola Paulista de Medicina, Hospital São Paulo, Disciplina de Anestesiologia, Departamento de Cirurgia, São Paulo, SP, Brasil, Disciplina de Anestesiologia, Departamento de Cirurgia, Hospital São Paulo, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Salomao, R. [Universidade Federal de São Paulo, Escola Paulista de Medicina, Hospital São Paulo, Disciplina de Infectologia, Departamento de Medicina, São Paulo, SP, Brasil, Disciplina de Infectologia, Departamento de Medicina, Hospital São Paulo, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil)

    2014-04-11

    Recognition of pathogens is performed by specific receptors in cells of the innate immune system, which may undergo modulation during the continuum of clinical manifestations of sepsis. Monocytes and neutrophils play a key role in host defense by sensing and destroying microorganisms. This study aimed to evaluate the expression of CD14 receptors on monocytes; CD66b and CXCR2 receptors on neutrophils; and TLR2, TLR4, TLR5, TLR9, and CD11b receptors on both cell types of septic patients. Seventy-seven septic patients (SP) and 40 healthy volunteers (HV) were included in the study, and blood samples were collected on day zero (D0) and after 7 days of therapy (D7). Evaluation of the cellular receptors was carried out by flow cytometry. Expression of CD14 on monocytes and of CD11b and CXCR2 on neutrophils from SP was lower than that from HV. Conversely, expression of TLR5 on monocytes and neutrophils was higher in SP compared with HV. Expression of TLR2 on the surface of neutrophils and that of TLR5 on monocytes and neutrophils of SP was lower at D7 than at D0. In addition, SP who survived showed reduced expression of TLR2 and TLR4 on the surface of neutrophils at D7 compared to D0. Expression of CXCR2 for surviving patients was higher at follow-up compared to baseline. We conclude that expression of recognition and cell signaling receptors is differentially regulated between SP and HV depending on the receptor being evaluated.

  12. Suppression of blood monocyte and neutrophil chemotaxis in acute human malaria

    DEFF Research Database (Denmark)

    Nielsen, H; Kharazmi, A; Theander, T G

    1986-01-01

    The host response to Plasmodia includes the production of enlarged populations of peripheral blood monocytes and tissue macrophages in the spleen and the liver. Since the hyperplasia of the mononuclear phagocyte system is believed to arise as a consequence of an enhanced blood monocyte influx, we....... In conclusion, not all cell functions were altered in concert, and the previously unreported suppression of chemotactic migration might reflect a change in blood leucocyte subpopulations, deactivation in vivo or a direct suppressive effect of plasmodia induced products....

  13. Identification of bovine material in porcine spray-dried blood derivatives using the Polymerase Chain Reaction technique

    Directory of Open Access Journals (Sweden)

    Sánchez A.

    2004-01-01

    Full Text Available Due to the widely supported theory of bovine spongiform encephalopathy (BSE spread in cattle by contaminated animal feeds, screening of feed products has become essential. For many years, manufacturers have used blood and plasma proteins as high quality ingredients of foods for both pets and farm animals. However, in Europe, the Commission Regulation 1234/2003/EC temporally bans the use of processed animal proteins, including blood-derivative products, in feedstuffs for all farm animals which are fattened or bred for the production of food. This regulation has some exceptions, such as the use of non ruminant blood products into the feed of farm fish. Authorization of the re-introduction of these proteins into animal feed formulations, especially non ruminant proteins into the feed for non ruminant farm animals, is expected when adequate control methods to discriminate ruminant proteins exist. Currently, the number of validated methods to differentiate the species of origin for most of the animal by-products is limited. Here we report the development of a rapid and sensitive polymerase chain reaction (PCR-based assay, which allows detection of bovine or porcine specific mitochondrial DNAfrom spray-dried blood derivate products (plasma, whole blood and red cells, as a marker for bovine contamination in porcine products. Sample extracts, suitable for PCR, were easily and quickly obtained with the commercial PrepManTM Ultra reagent (Applied Biosystems. To confirm the porcine origin of the samples, primers targeting a specific region of 134 bp of the porcine cytochrome b coding sequence were designed (cytbporc1-F and cytbporc2-R. Previously published PCR primers (L8129 and H8357, specific for a 271 bp fragment of the bovine mitochondrial ATPase 8-ATPase 6 genes, were chosen to accomplish amplification of bovine DNA. The limit of detection (LOD of the bovine PCR assay was at least of 0.05% (v/v of bovine inclusion in spray-dried porcine plasma or red

  14. Molecular detection of bovine leukemia virus in peripheral blood of Iranian cattle, camel and sheep.

    Science.gov (United States)

    Nekoei, S; Hafshejani, T Taktaz; Doosti, A; Khamesipour, F

    2015-01-01

    Bovine leukemia virus (BLV) is a deltaretrovirus which infects and induces proliferation of B-lymphocytes in the peripheral blood circulation and in lymphoid organs primarily of cattle, leading to leukemia/lymphoma. This study was carried out to investigate the presence of BLV in cattle, sheep and camels from the Chaharmahal va Bakhtiary and Isfahan provinces in Iran. A total of 874 blood samples collected from cattle, sheep and camels were used in this study to detect BLV using a nested-PCR. The results from this study indicated that 17.2% (n=874) of all blood samples collected were positive for BLV. The percentages of blood samples positive for BLV from cattle, sheep and camels were 22.1 (n=657), 5.3 (n=95) and 0 (n=122) respectively. The results from this study showed that BLV infected cattle and sheep. Camels seemed to be resistant to BLV infection. This study contributes to the nationwide effort to obtain baseline information on the prevalence of BLV, which will assist in planning the control strategy for the disease in Iran.

  15. Intravaginal Administration of Sildenafil Citrate Increases Blood Flow in the Bovine Uterus

    Directory of Open Access Journals (Sweden)

    Dzięcioł Michał

    2015-04-01

    Full Text Available The aim of the study was to evaluate the influence of sildenafil citrate administrated intravaginaly on the blood flow in the bovine uterus during dioestrus. Uterine blood flow was examined in six healthy adult cows. Sildenafil was administrated intravaginaly to each co w between the 6th and 8th d of the ovarian cycle, in the form of vaginal suppositories containing 100 mg of active substance at a dose of 100, 200, or 300 mg per animal. Uterine perfusion was estimated by the colour Doppler examination, and obtained results were analysed with the Pixel Flux Software (Chameleon, Germany. Moreover, cardiovascular parameters were also evaluated. Animals were examined before and five times after drug application (two times at 15 min intervals, and three times at 2 h intervals. A placebo suppository was also given to the cows. The analysis of the intensity and velocity of blood flow in the uterus proved that sildenafil administrated intravaginaly significantly increased blood flow in the uterus and the effect of increased perfusion was observed for 4 h and 30 min after administration. The effect of increased uterine perfusion was observed after low as well as high doses of sildenafil. Significant changes in the cardio-vascular parameters were not detected. There were no changes in the uterine perfusion as well as in cardiovascular parameters after placebo administration.

  16. Protein adsorption to monosodium urate crystals: differential responses of human peripheral blood neutrophils. [Etiology of acute gouty arthritis

    Energy Technology Data Exchange (ETDEWEB)

    Skosey, J.L.; Kozin, F.; Ginsberg, M.

    1976-01-01

    In order for acute gouty arthritis to occur, neutrophils must interact with monosodium urate (MSU) crystals. As a result of this interaction, enzymes, chemotactic factors, and other mediators of the inflammatory response are released from neutrophil lysosomes. It was observed that MSU crystals adsorb gamma globulin, albumin, and other proteins found in serum and joint fluid. Results are reported from a study designed to demonstrate the effects of coating of MSU crystals with proteins on the phlogistic responses of neutrophils to crystals.

  17. Effect of hemolysis on nonesterified fatty acid and beta-hydroxybutyrate concentrations in bovine blood.

    Science.gov (United States)

    Stokol, Tracy; Nydam, Daryl V

    2006-09-01

    Nonesterified fatty acid (NEFA) and beta-hydroxybutyrate (BHB) assays are used for evaluating dairy herds for negative energy balance and subclinical ketosis, respectively. Hemolysis is a common artifact in samples submitted to diagnostic laboratories. The effect of hemolysis on NEFA and BHB in bovine serum was determined. Hemolysis was introduced into 26 serum samples by adding serial dilutions of a red cell hemolysate, prepared by repeated freeze-thawing of EDTA-anticoagulated bovine blood. NEFA, BHB, and degree of hemolysis (hemolytic index) were measured by an automated chemistry analyzer. Two endpoint assays that differed by inclusion of a sample blank were used for NEFA measurement. A kinetic enzymatic assay with 2 reagent sources was used for BHB measurement. The assessed methods yielded similar NEFA or BHB results in baseline, nonhemolyzed samples (median NEFA: 0.25 mEq/L, median BHB: 3 mg/dL, median hemolytic index: 8 units). NEFA results were adversely affected by hemolysis, with values increasing significantly with higher degrees of hemolysis. Median values increased above a critical medical decision limit (0.40 mEq/L) at a hemolytic index of 506 units (marked hemolysis). This increase was prevented by inclusion of a sample blank. Result interpretation was affected in individual animals when samples were moderately hemolyzed (median hemolytic index: 258 units). In contrast, BHB results were unaffected by hemolysis with either reagent source. Thus, assays for measuring NEFAs should include a sample blank and NEFA results should not be interpreted in moderately to markedly hemolyzed bovine samples, because result accuracy cannot be assured.

  18. Neutrophils, from marrow to microbes

    DEFF Research Database (Denmark)

    Borregaard, Niels

    2010-01-01

    . Neutrophils circulate in the blood as dormant cells. At sites of infection, endothelial cells capture bypassing neutrophils and guide them through the endothelial cell lining whereby the neutrophils are activated and tuned for the subsequent interaction with microbes. Once in tissues, neutrophils kill......Neutrophils are produced in the bone marrow from stem cells that proliferate and differentiate to mature neutrophils fully equipped with an armory of granules. These contain proteins that enable the neutrophil to deliver lethal hits against microorganisms, but also to cause great tissue damage...... microorganisms by microbicidal agents liberated from granules or generated by metabolic activation. As a final act, neutrophils can extrude stands of DNA with bactericidal proteins attached that act as extracellular traps for microorganisms....

  19. Biodegradation and bioabsorption innovation of the functionally graded bovine bone-originated apatite with blood permeability.

    Science.gov (United States)

    Akazawa, Toshiyuki; Murata, Masaru; Sasaki, Tomoya; Tazaki, Junichi; Kobayashi, Masayoshi; Kanno, Tohru; Nakamura, Katsuo; Arisue, Makoto

    2006-01-01

    Bioabsorbable and functionally graded apatite (fg-HAp) ceramics were designed using bovine bone by the calcination and partial dissolution-precipitation methods. The fg-HAp ceramics that were developed had gradual distributions of the degree of crystallinity and the grain size of single-phase hydroxyapatite from the surface layer of the pore wall to the bulk structure region. Calcination at 1073 K gave a specific surface area of 30 m2 x g-1 and porosities of 60-80%. The pore structure of the fg-HAp was classified into two regions: a macro-pore region (100-600 microm) originating from spongy bone and a micro-pore region (10-160 nm) related to body fluid permeation and blood permeability. By implantation in subcutaneous tissue of rat, it was confirmed that body fluid permeated the bulk region of the fg-HAp ceramics through the micro-pores. The volumetric populations occupied by body fluid were 60% at 4 weeks and 68% at 8 weeks in the ceramics explants, indicating drastic bioabsorption, although the body fluid was found to be immunopositive for an albumin as the main serum protein in blood. On the fg-HAp ceramics developed here, the bioabsorption rate could be controlled by careful selection of the calcination temperature. These ceramics can be applied as new biomimetic ceramics exhibiting surface and bulk degradations and cellular absorption by giant cells.

  20. The influence of Rubus idaeus and Rubus caesius leaf extracts on platelet aggregation in whole blood. Cross-talk of platelets and neutrophils.

    Science.gov (United States)

    Dudzinska, Dominika; Bednarska, Katarzyna; Boncler, Magdalena; Luzak, Boguslawa; Watala, Cezary

    2016-07-01

    Recently, polyphenols have gained attention as potential natural cardioprotective therapeutics, due to their antiplatelet, anti-inflammatory and anticoagulant activity. Species belonging to the genus Rubus sp. have been reported to be a source of polyphenolic compounds with antioxidative proprieties and beneficial biological activities. This study investigates the effects of leaf extracts obtained from red raspberry (Rubus idaeus L.) and European dewberry (Rubus caesius L.) on the reactivity of blood platelets. In ADP-stimulated blood, raspberry and dewberry extracts (15 µg/ml) markedly decreased platelet surface membrane expression of activated GPIIbIIIa receptor by 16% and 21%, respectively (P raspberry and by 38-55% for dewberry, P raspberry and dewberry leaf extracts considerably modulated blood platelet reactivity in whole blood: they influenced blood platelet aggregation, possibly via the modulation of the redox status dependent on the oxidative activity of neutrophils.

  1. A combretastatin-mediated decrease in neutrophil concentration in peripheral blood and the impact on the anti-tumor activity of this drug in two different murine tumor models.

    Directory of Open Access Journals (Sweden)

    Anja Bille Bohn

    Full Text Available The vascular disrupting agent combretastatin A-4 disodium phosphate (CA4P induces fluctuations in peripheral blood neutrophil concentration. Because neutrophils have the potential to induce both vascular damage and angiogenesis we analyzed neutrophil involvement in the anti-tumoral effects of CA4P in C3H mammary carcinomas in CDF1 mice and in SCCVII squamous cell carcinomas in C3H/HeN mice. Flow cytometry analyses of peripheral blood before and up to 144 h after CA4P administration (25 and 250 mg/kg revealed a decrease 1 h after treatment, followed by an early (3-6 h and a late (>72 h increase in the granulocyte concentration. We suggest that the early increase (3-6 h in granulocyte concentration was caused by the initial decrease at 1 h and found that the late increase was associated with tumor size, and hence independent of CA4P. No alterations in neutrophil infiltration into the C3H tumor after CA4P treatment (25 and 250 mg/kg were found. Correspondingly, neutrophil depletion in vivo, using an anti-neutrophil antibody, followed by CA4P treatment (25 mg/kg did not increase the necrotic fraction in C3H tumors significantly. However, by increasing the CA4P dose to 250 mg/kg we found a significant increase of 359% in necrotic fraction when compared to neutrophil-depleted mice; in mice with no neutrophil depletion CA4P induced an 89% change indicating that the presence of neutrophils reduced the effect of CA4P. In contrast, neither CA4P nor 1A8 affected the necrotic fraction in the SCCVII tumors significantly. Hence, we suggest that the initial decrease in granulocyte concentration was caused by non-tumor-specific recruitment of neutrophils and that neutrophils may attenuate CA4P-mediated anti-tumor effect in some tumor models.

  2. Whole Blood Gene Expression Profiling in Preclinical and Clinical Cattle Infected with Atypical Bovine Spongiform Encephalopathy

    Science.gov (United States)

    Xerxa, Elena; Barbisin, Maura; Chieppa, Maria Novella; Krmac, Helena; Vallino Costassa, Elena; Vatta, Paolo; Simmons, Marion; Caramelli, Maria; Casalone, Cristina; Corona, Cristiano

    2016-01-01

    Prion diseases, such as bovine spongiform encephalopathies (BSE), are transmissible neurodegenerative disorders affecting humans and a wide variety of mammals. Variant Creutzfeldt-Jakob disease (vCJD), a prion disease in humans, has been linked to exposure to BSE prions. This classical BSE (cBSE) is now rapidly disappearing as a result of appropriate measures to control animal feeding. Besides cBSE, two atypical forms (named H- and L-type BSE) have recently been described in Europe, Japan, and North America. Here we describe the first wide-spectrum microarray analysis in whole blood of atypical BSE-infected cattle. Transcriptome changes in infected animals were analyzed prior to and after the onset of clinical signs. The microarray analysis revealed gene expression changes in blood prior to the appearance of the clinical signs and during the progression of the disease. A set of 32 differentially expressed genes was found to be in common between clinical and preclinical stages and showed a very similar expression pattern in the two phases. A 22-gene signature showed an oscillating pattern of expression, being differentially expressed in the preclinical stage and then going back to control levels in the symptomatic phase. One gene, SEL1L3, was downregulated during the progression of the disease. Most of the studies performed up to date utilized various tissues, which are not suitable for a rapid analysis of infected animals and patients. Our findings suggest the intriguing possibility to take advantage of whole blood RNA transcriptional profiling for the preclinical identification of prion infection. Further, this study highlighted several pathways, such as immune response and metabolism that may play an important role in peripheral prion pathogenesis. Finally, the gene expression changes identified in the present study may be further investigated as a fingerprint for monitoring the progression of disease and for developing targeted therapeutic interventions. PMID

  3. Whole Blood Gene Expression Profiling in Preclinical and Clinical Cattle Infected with Atypical Bovine Spongiform Encephalopathy.

    Science.gov (United States)

    Xerxa, Elena; Barbisin, Maura; Chieppa, Maria Novella; Krmac, Helena; Vallino Costassa, Elena; Vatta, Paolo; Simmons, Marion; Caramelli, Maria; Casalone, Cristina; Corona, Cristiano; Legname, Giuseppe

    2016-01-01

    Prion diseases, such as bovine spongiform encephalopathies (BSE), are transmissible neurodegenerative disorders affecting humans and a wide variety of mammals. Variant Creutzfeldt-Jakob disease (vCJD), a prion disease in humans, has been linked to exposure to BSE prions. This classical BSE (cBSE) is now rapidly disappearing as a result of appropriate measures to control animal feeding. Besides cBSE, two atypical forms (named H- and L-type BSE) have recently been described in Europe, Japan, and North America. Here we describe the first wide-spectrum microarray analysis in whole blood of atypical BSE-infected cattle. Transcriptome changes in infected animals were analyzed prior to and after the onset of clinical signs. The microarray analysis revealed gene expression changes in blood prior to the appearance of the clinical signs and during the progression of the disease. A set of 32 differentially expressed genes was found to be in common between clinical and preclinical stages and showed a very similar expression pattern in the two phases. A 22-gene signature showed an oscillating pattern of expression, being differentially expressed in the preclinical stage and then going back to control levels in the symptomatic phase. One gene, SEL1L3, was downregulated during the progression of the disease. Most of the studies performed up to date utilized various tissues, which are not suitable for a rapid analysis of infected animals and patients. Our findings suggest the intriguing possibility to take advantage of whole blood RNA transcriptional profiling for the preclinical identification of prion infection. Further, this study highlighted several pathways, such as immune response and metabolism that may play an important role in peripheral prion pathogenesis. Finally, the gene expression changes identified in the present study may be further investigated as a fingerprint for monitoring the progression of disease and for developing targeted therapeutic interventions.

  4. Airborne particulate matter PM2.5 from Mexico City affects the generation of reactive oxygen species by blood neutrophils from asthmatics: an in vitro approach

    Directory of Open Access Journals (Sweden)

    Ceballos Guillermo

    2009-06-01

    Full Text Available Abstract Background The Mexico City Metropolitan Area is densely populated, and toxic air pollutants are generated and concentrated at a higher rate because of its geographic characteristics. It is well known that exposure to particulate matter, especially to fine and ultra-fine particles, enhances the risk of cardio-respiratory diseases, especially in populations susceptible to oxidative stress. The aim of this study was to evaluate the effect of fine particles on the respiratory burst of circulating neutrophils from asthmatic patients living in Mexico City. Methods In total, 6 subjects diagnosed with mild asthma and 11 healthy volunteers were asked to participate. Neutrophils were isolated from peripheral venous blood and incubated with fine particles, and the generation of reactive oxygen species was recorded by chemiluminescence. We also measured plasma lipoperoxidation susceptibility and plasma myeloperoxidase and paraoxonase activities by spectrophotometry. Results Asthmatic patients showed significantly lower plasma paraoxonase activity, higher susceptibility to plasma lipoperoxidation and an increase in myeloperoxidase activity that differed significantly from the control group. In the presence of fine particles, neutrophils from asthmatic patients showed an increased tendency to generate reactive oxygen species after stimulation with fine particles (PM2.5. Conclusion These findings suggest that asthmatic patients have higher oxidation of plasmatic lipids due to reduced antioxidant defense. Furthermore, fine particles tended to increase the respiratory burst of blood human neutrophils from the asthmatic group. On the whole, increased myeloperoxidase activity and susceptibility to lipoperoxidation with a concomitant decrease in paraoxonase activity in asthmatic patients could favor lung infection and hence disrupt the control of asthmatic crises.

  5. Blood neutrophil bactericidal activity against methicillin-resistant and methicillin-sensitive Staphylococcus aureus during cardiac surgery.

    Science.gov (United States)

    Mekontso-Dessap, Armand; Honoré, Stéphanie; Kirsch, Matthias; Plonquet, Anne; Fernandez, Eric; Touqui, Lhousseine; Farcet, Jean-Pierre; Soussy, Claude-James; Loisance, Daniel; Delclaux, Christophe

    2005-08-01

    Whether methicillin-resistant Staphylococcus aureus (MRSA) constitutes per se an independent risk factor for morbidity and mortality after surgery as compared with methicillin-sensitive Staphylococcus aureus (MSSA) remains a subject of debate. The aim of this study was to assess whether innate defenses against MRSA and MSSA strains are similarly impaired after cardiac surgery. Both intracellular (isolated neutrophil functions) and extracellular (plasma) defenses of 12 patients undergoing scheduled cardiac surgery were evaluated preoperatively (day 0) and postoperatively (day 3) against two MSSA strains with a low level of catalase secretion and two MRSA strains with a high level of catalase secretion, inasmuch as SA killing by neutrophils relies on oxygen-dependent mechanisms. After surgery, an increase in plasma concentration of IL-10, an anti-inflammatory cytokine able to inhibit reactive oxygen species secretion and bactericidal activity of neutrophils, was evidenced. Despite the fact that univariate analysis suggested a specific impairment of neutrophil functions against MRSA strains, two-way repeated-measures ANOVA failed to demonstrate that the effect of S. aureus phenotype was significant. On the other hand, an increase in type-II secretory phospholipase A2 activity, a circulating enzyme involved in SA lysis, was evidenced and was associated with an enhancement of extracellular defenses (bactericidal activity of plasma) against MRSA. Overall, cardiac surgery and S. aureus phenotype had a significant effect on plasma bactericidal activity. Cardiac surgery was characterized by enhanced antibacterial defenses of plasma, whereas neutrophil killing properties were reduced. The overall effect of S. aureus phenotype on neutrophil functions did not seem significant.

  6. Human Neutrophil Elastase Induce Interleukin-10 Expression in Peripheral Blood Mononuclear Cells through Protein Kinase C Theta/Delta and Phospholipase Pathways

    Directory of Open Access Journals (Sweden)

    Jin Kawata

    2016-02-01

    Full Text Available Objective: Neutrophils have an important role in the rapid innate immune response, and the release or active secretion of elastase from neutrophils is linked to various inflammatory responses. Purpose of this study was to determine how the human neutrophil elastase affects the interleukin-10 (IL-10 response in peripheral blood mononuclear cells (PBMC. Materials and Methods: In this prospective study, changes in IL-10 messenger RNA (mRNA and protein expression levels in monocytes derived from human PBMCs were investigated after stimulation with human neutrophil elastase (HNE. A set of inhibitors was used for examining the pathways for IL-10 production induced by HNE. Results: Reverse transcription polymerase chain reaction (RT-PCR showed that stimulation with HNE upregulated IL-10 mRNA expression by monocytes, while the enzyme-linked immunosorbent assay (ELISA revealed an increase of IL-10 protein level in the culture medium. A phospholipase C inhibitor (U73122 partially blunted the induction of IL-10 mRNA expression by HNE, while IL-10 mRNA expression was significantly reduced by a protein kinase C (PKC inhibitor (Rottlerin. A calcium chelator (3,4,5-trimethoxybenzoic acid 8-(diethylaminooctyl ester: TMB-8 inhibited the response of IL-10 mRNA to stimulation by HNE. In addition, pretreatment with a broad-spectrum PKC inhibitor (Ro-318425 partly blocked the response to HNE. Finally, an inhibitor of PKC theta/delta abolished the increased level of IL-10 mRNA expression. Conclusion: These results indicate that HNE mainly upregulates IL-10 mRNA expression and protein production in moncytes via a novel PKC theta/delta, although partially via the conventional PKC pathway.

  7. In vitro replication activity of bovine viral diarrhea virus in an epithelial cell line and in bovine peripheral blood mononuclear cells.

    Science.gov (United States)

    Turin, Lauretta; Lucchini, Barbara; Bronzo, Valerio; Luzzago, Camilla

    2012-11-01

    The present study focused on the in vitro infection of Madin-Darby bovine kidney (MDBK) cells and bovine peripheral blood mononuclear cells (PBMCs) from naÏve animals with non-cytopathic (ncp, BVDV-1b NY-1) and cytopathic (cp, BVDV-1a NADL) strains. Infections with 0.1 and 1 multiplicity of infections (MOI) and incubation times of 18 and 36 hr were compared. Twelve BVDV naÏve heifers were enrolled to collect PBMCs. The viral loads in MDBK cells and in PBMCs after in vitro infections were measured by real-time polymerase chain reaction (PCR) assays. The highest viral loads were measured at 1 MOI and 36 hr post infection in both cell systems and the lowest at 0.1 MOI and 18 hr with the exception of the cp strain NADL in PBMCs, for which the highest viral load was observed at 0.1 MOI and 36 hr. Viral load mean values were higher for the cp strain than the ncp strain irrespective of the extent of the infection period and MOI. The models of infection studied uncovered different replication activities respectively according to the biotype of virus, the cell substrate and the duration of infection. Replication tends to be higher in PBMCs, particularly at low MOIs and for the ncp strain.

  8. Supplemental Smartamine M or MetaSmart during the transition period benefits postpartal cow performance and blood neutrophil function.

    Science.gov (United States)

    Osorio, J S; Ji, P; Drackley, J K; Luchini, D; Loor, J J

    2013-10-01

    source increased milk yield, milk protein percentage, energy-corrected milk, and milk fat yield by 3.4 kg/d, 0.18% units, 3.9 kg/d, and 0.18 kg/d, respectively. Those responses were associated with greater postpartum concentration of growth hormone but not insulin-like growth factor 1. There was a diet × time effect for nonesterified fatty acid concentration due to greater values on d 7 for MS; however, liver concentration of triacylglycerol was not affected by diet or diet × time but increased postpartum. Blood neutrophil phagocytosis at 21 d was greater with Met supplementation, suggesting better immune function. Supplemental MS or SM resulted in a tendency for lower incidence of ketosis postpartum. Although supplemental MS or SM did not decrease liver triacylglycerol, it improved milk production-related traits by enhancing voluntary DMI.

  9. Ultrastructural characterization of bovine umbilical cord blood cells Caracterização ultra-estrutural das células sanguíneas do cordão umbilical bovino

    Directory of Open Access Journals (Sweden)

    Gustavo C Rodrigues

    2010-10-01

    Full Text Available The umbilical cord blood (UCB is an important source of pluripotent stem cells, which motivated researches on ontogeny and transplantation. The morphological characterization of umbilical cord cells is the first step to establish subsequent experiments on these areas. Although some information on humans can be found, no data on UCB is available for bovines. Therefore, this work is the first attempt to conduct an ultrastructural characterization of bovine umbilical cord blood. Blood was collected from the umbilical cord of twenty fetuses by punction of the umbilical vein. Samples were processed for whole leucocytes observation by centrifugation and the buffy coat was collected. Cells were washed and pelleted and prepared according to the standard protocol of the transmission electron microscopy. The presence of cells with morphologic characteristics compatible with the precursors from the erythrocytic, neutrophilic, eosinophilic, basophilic, and lymphocytic lineages was observed. Atypical cells with peculiar morphological features, strongly similar to apoptotic cells, were seen. Bovine neutrophils with three types of cytoplasmic granules were also found in the blood. The ultrastructural characteristics of observed bovine UCB cells where similar to those found in other species, suggesting that bovines could possibly constitute an experimental model for approaches on UCB cells research.O sangue de cordão umbilical (SCU é uma importante fonte de células progenitoras pluripotentes, que motiva pesquisas em ontogenia e transplantes. A caracterização morfológica das células de cordão umbilical é o primeiro passo para se estabelecer experimentos subsequentes nessas áreas. Embora algumas informações sobre SCU em humanos possam ser encontradas, não existe nenhuma informação disponível sobre elas em bovinos. Portanto, este trabalho é a primeira tentativa de se conduzir uma caracterização ultra-estrutural do sangue de cordão umbilical

  10. Investigations of presence of antibodies against bovine herpesvirus-1 in blood serum of calves prior to colostrum diet

    Directory of Open Access Journals (Sweden)

    Lazić Sava

    2010-01-01

    Full Text Available The paper presents the results of investigations of the presence of the bovine herpesvirus-1 (BHV-1 in samples of blood serum from 106 cows and 107 of their calves (one cow had twins. Blood was sampled from the cows immediately after parturition, and from the calves before feeding on colostrum. The examined cows and their calves originated from 5 herds in which previous investigations had shown infection with the bovine herpesvirus-1. The determination of antibodies against BHV-1 was performed using the method of virus neutralization in culture of MDBK cells with 100 TCID/50 viruses (BHV-1, TN-41 Am. Bio Research, USA. Antibodies against BHV-1 were determined in all blood serum samples of cows and in 16 samples of precolostral blood serums of calves. The antibody titer values in cows ranged from 1:4 to 1:512, and in calves the determined values were from 1:2 to 1:16. The results indicate that cows that are seropositive to BHV-1 can deliver calves seropositive to BHV-1 in about 15% cases. This must be kept in mind in selecting cows for the production of breeding material, in particular bulls for reproduction centers, as well as in making a programme for the immunoprophylaxis of calves against BHV-1. .

  11. The ultrastructure of camel blood platelets: a comparative study with human, bovine, and equine cells.

    Science.gov (United States)

    Gader, Abdel Galil M Abdel; Ghumlas, Abeer K Al; Hussain, Mansour F; Haidari, Ahmed Al; White, James G

    2008-02-01

    Previous studies indicated that the camel has a very active haemostatic mechanism with a short bleeding time and thrombocytosis. However, platelet function, when tested by agonist-induced aggregation and PFA 100 closure time, showed marked inhibition compared to humans. Since camels are also far more resistant to long exposure to excessive heat and high body temperature than humans, it seemed worthwhile to explore fundamental morphological differences between human and camel platelets and those from other species. The present study has examined the ultrastructure of camel platelets and compared them with the fine structures of human, bovine and equine thrombocytes. Camel platelets, like bovine and equine cells, are discoid in shape and about two-thirds the size of human platelets. A circumferential coil of microtubular supports the disk-like form of camel platelets. Their cytoplasm, like bovine and equine platelets, is filled with alpha granule twice as large as those in human platelets, but lacking the organized matrix of equine alpha granules. Dense bodies are present in camel platelets with whip-like extensions not present on bovine or equine thrombocytes, but found on occasional human platelet dense bodies. Camel platelets, like bovine and equine thrombocytes, lack an open canalicular system (OCS) and must secrete granule products by fusion with the cell wall rather than an OCS. Future studies will determine if the differences in ultrastructural anatomy protect camel platelets from heat more than human thrombocytes.

  12. Development of a preliminary diagnostic measure for bovine leukosis in dairy cows using peripheral white blood cell and lymphocyte counts

    Science.gov (United States)

    NISHIIKE, Masao; HAOKA, Michiyo; DOI, Takashi; KOHDA, Tomoko; MUKAMOTO, Masafumi

    2016-01-01

    Analysis of the association between antibodies against bovine leukemia virus (BLV), BLV proviral load, and white blood cell (WBC) and lymphocyte counts was performed with 774 dairy cows. The average age, WBC counts and lymphoid cell counts tended to be higher in BLV antibody-positive cows than in antibody-negative cows. There was a similar trend in levels of proviral DNA. We analyzed age, WBC counts and lymphocyte counts by principal component analyses to create a distribution chart of the principle component scores. Using the chart, we categorized cows into four quadrants based on additional information, such as the presence of antibody and the levels of proviral DNA. Antibody-positive cows and cows with high BLV proviral load were found mostly in one quadrant of the chart, indicating that it is possible to predict the risk of infection without any knowledge on antibody status by using information, such as WBC counts as a biomarker. When only antibody-positive cows were included in the analysis, a characteristic distribution of different levels of proviral DNA was seen in the quadrants, suggesting that it is possible to estimate the extent of bovine leukosis infection by using this analysis. For this analysis and categorization of the cows into quadrants, we computed a mathematical formulation using discriminant analysis based on age and WBC and lymphocyte counts. This mathematical formulation for the hematological preliminary diagnosis of the disease is recommended as a screening tool to monitor bovine leukosis. PMID:27064146

  13. Neutrophil biology

    OpenAIRE

    Kobayashi, Yoshiro

    2015-01-01

    Neutrophil extracellular traps (NETs) are involved in bacterial killing as well as autoimmunity, because NETs contain proteases, bactericidal peptides, DNA and ribonucleoprotein. NETs are formed via a novel type of cell death called NETosis. NETosis is distinct from apoptosis, but it resembles necrosis in that both membranes are not intact so that they allow intracellular proteins to leak outside of the cells. Removal of NETs and neutrophils undergoing NETosis by phagocytes and its subsequent...

  14. Advances in Comprehensive Exploitation and Utilization of Bovine Blood%牛血资源综合开发利用研究进展

    Institute of Scientific and Technical Information of China (English)

    张玉斌; 曹晖; 郭兆斌; 余群力

    2011-01-01

    介绍了牛血的成分、营养特性及开发利用价值和综合利用现状,同时对牛血资源在食品工业、生化制药工业以及饲料工业的研究进展进行了概述,并讨论了牛血资源综合利用存在的问题和解决措施。%The composition, nutritional characteristics, exploitation value and current utilization situation of bovine blood are introduced in this paper. Meanwhile, the progress of applications of bovine blood in the food, biopharmaceutical and feed industries are overviewed, and problems present in the comprehensive utilization of bovine blood and solutions are explored.

  15. Biochemical identification of the bovine blood group M' antigen as a major histocompatibility complex class I-like molecule

    DEFF Research Database (Denmark)

    Hønberg, L S; Larsen, B; Koch, C;

    1995-01-01

    . To elucidate the structural relationship between BoLA-A16 and blood group antigen M', immunoprecipitation experiments on red and white cell lysates isolated from M'-A16 positive and negative cattle were carried out. These results showed that M(r) 44,000 and M(r) 12000 polypeptides can be precipitated from both......Absorption and elution experiments showed that it was impossible to separate antibodies against blood group factor M' from antibodies against bovine lymphocyte antigen (BoLA) A16 in an antiserum showing haemolytic activity against M' as well as lymphocytotoxic activity against BoLA-A16...... red and white cells isolated from M'-A16 positive animals, whereas no bands were seen in M'-A16 negative animals in precipitations with the same antibody. Precipitation with a crossreacting human beta 2-microglobulin (beta 2-m) specific antibody confirmed a class-I-like structure associated with beta...

  16. [Nitroblue tetrazolium reduction by the neutrophils of the cerebrospinal fluid and peripheral blood and by the monocytic-reticular cells of the cerebrospinal fluid in neuroinfections].

    Science.gov (United States)

    Kucharska-Demczuk, K

    1980-01-01

    Using the method of Park et al. the author studied spontaneous and stimulated NBT reduction by neutrophil granulocytes in the cerebrospinal fluid and blood, and by monocytic-reticular cells in the cerebrospinal fluid of the patients with bacterial and viral meningitis and meningismus. The author performed 333 investigations in 74 patients. Significantly higher mean values of the index of spontaneous and stimulated NBT reduction by the granulocytes and cerebrospinal fluid were observed in cases of bacterial meningitis as compared with the granulocytes of the peripheral blood in healthy subjects. It was demonstrated that in patients with bacterial meningitis blood and fluid granulocytes showed a similar phagocytic acitivty independent of the humoral environment. In the patients with bacterial and viral meningitis the monocytic-reticular cells the cerebrospinal fluid showed a similar, sometimes high, phagocytic activity depending on the phase and severity of the disease. On the otherhand, in most cases of meningismus these cells failed to manifest any phagocytic and bactericidal activity. In only few isolated cells in the fluid weak NBT reduction was observed. The obtained results of investigations showed the usefulness of the NBT test not only for the differential diagnosis of the aetiology of neuroinfections but also for the assessment of immune processes taking place in the nervous system.

  17. Concentrations of procalcitonin and C-reactive protein, white blood cell count, and the immature-to-total neutrophil ratio in the blood of neonates with nosocomial infections: Gram-negative bacilli vs coagulase-negative staphylococci.

    Science.gov (United States)

    Kordek, A

    2011-03-01

    This study was undertaken to determine whether concentrations of procalcitonin in the blood of neonates with nosocomial infections depend on the type of pathogen. Qualification for the study group was based on the clinical signs of infection. We found that infections with Gram-positive (chiefly coagulase-negative staphylococci) and Gram-negative bacteria are accompanied by elevated concentrations of procalcitonin. In the case of Gram-positive bacteria, other laboratory signs of infection studied by us (concentration of C-reactive protein, white blood cell count, immature-to-total neutrophil ratio) were not discriminatory, confirming the diagnostic usefulness of procalcitonin measurements in nosocomial infections of the neonate with Gram-negative or Gram-positive bacteria.

  18. Analysis of gene expression in white blood cells of cattle orally challenged with bovine amyloidotic spongiform encephalopathy.

    Science.gov (United States)

    Panelli, Simona; Strozzi, Francesco; Capoferri, Rossana; Barbieri, Ilaria; Martinelli, Nicola; Capucci, Lorenzo; Lombardi, Guerino; Williams, John L

    2011-01-01

    Bovine amyloidotic spongiform encephalopathy (BASE) is one of the recently discovered atypical forms of BSE, which is transmissible to primates, and may be the bovine equivalent of sporadic Creutzfeldt-Jacob disease (CJD) in humans. Although it is transmissible, it is unknown whether BASE is acquired through infection or arises spontaneously. In the present study, the gene expression of white blood cells (WBCs) from 5 cattle at 1 yr after oral BASE challenge was compared with negative controls using a custom microarray containing 43,768 unique gene probes. In total, 56 genes were found to be differentially expressed between BASE and control animals with a log fold change of 2 or greater. Of these, 39 were upregulated in BASE animals, while 17 were downregulated. The majority of these genes are related to immune function. In particular, BASE animals appeared to have significantly modified expression of genes linked to T- and B-cell development and activation, and to inflammatory responses. The potential impacts of these gene expression changes are described.

  19. Bovine colostrum modulates immune activation cascades in human peripheral blood mononuclear cells in vitro

    DEFF Research Database (Denmark)

    Jenny, Marcel; Pedersen, Ninfa R; Hidayat, Budi J

    2010-01-01

    factors and has a long history of use in traditional medicine. In an approach to evaluate the effects of bovine colostrum (BC) on the T-cell/macrophage interplay, we investigated and compared the capacity of BC containing low and high amounts of lactose and lactoferrin to modulate tryptophan degradation...... of lactose present in BC seems to diminish the activity of BC in our test system, since BC with higher amounts of lactose attenuated the stimulatory as well as the suppressive activity of BC....

  20. Synthesis and evaluation of the potential deleterious effects of ZnO nanomaterials (nanoneedles and nanoflowers) on blood components, including albumin, erythrocytes and human isolated primary neutrophils

    Science.gov (United States)

    Pastrello, Bruna; Paracatu, Luana Chiquetto; de Carvalho Bertozo, Luiza; Paino, Iêda Maria Martinez; Lisboa-Filho, Paulo Noronha; Ximenes, Valdecir Farias

    2016-07-01

    The application of zinc oxide (ZnO) nanoparticles in biomaterials has increased significantly in the recent years. Here, we aimed to study the potential deleterious effects of ZnO on blood components, including human serum albumin (HSA), erythrocytes and human isolated primary neutrophils. To test the influence of the morphology of the nanomaterials, ZnO nanoneedles (ZnO-nn) and nanoflowers (ZnO-nf) were synthesized. The zeta potential and mean size of ZnO-nf and ZnO-nn suspensions in phosphate-buffered saline were -10.73 mV and 3.81 nm and -5.27 mV and 18.26 nm, respectively. The incubation of ZnO with HSA did not cause its denaturation as verified by the absence of significant alterations in the intrinsic and extrinsic fluorescence and in the circular dichroism spectrum of the protein. The capacity of HSA as a drug carrier was not affected as verified by employing site I and II fluorescent markers. Neither type of ZnO was able to provoke the activation of neutrophils, as verified by lucigenin- and luminol-dependent chemiluminescence and by the extracellular release of hydrogen peroxide. ZnO-nf, but not ZnO-nn, induced the haemolysis of erythrocytes. In conclusion, our results reinforce the concept that ZnO nanomaterials are relatively safe for usage in biomaterials. A potential exception is the capacity of ZnO-nf to promote the lysis of erythrocytes, a discovery that shows the importance of the morphology in the toxicity of nanoparticles.

  1. Effect of coumaphos on cholinesterase activity, hematology, and biochemical blood parameters of bovines in tropical regions of Mexico.

    Science.gov (United States)

    Pardío, Violeta T; Ibarra, Nelly De J; Waliszewski, Krzysztof N; López, Karla M

    2007-05-01

    To assess the effect of coumaphos [O-(3-chloro-4-methyl-2-oxo-2H-1-benzopyran-7-yl) O,O-diethyl phosphorothioate] exposure on physiological responses during bovine production, acetylcolinesterase (AChE) and butyrylcholinesterase (BuChE) activities were measured in whole blood, erythrocytes, and plasma of healthy male steers (Bos Taurus x Bos indicus) sprayed with coumaphos at a non-lethal dose of 1 mg kg(- 1) body weight per day once every 14 (in vivo group) or 21 days (southern and central groups). Coumaphos topically administered at 1 mg/kg body weight per day to cattle under normal management practices in tropical areas produced a significant inhibition in erythrocyte (RBC) AChE and BuAChE activities when compared to baseline levels. RBC-AChE activity for the in vivo group decreased 71.3% (P < 0.05) and BuChE activity 59.1% (P < 0.05); RBC-AChE activity decreased 55.1% (P < 0.05) (southern group) and 43.4% (P < 0.05) (central group). Compared to the control specimens, steers from in vivo, southern, and central groups after 150 days of exposure had lower (P < 0.05) leukocyte count, absolute lymphocyte, erythrocyte, and platelet counts. Decreases in RBC-AChE activities correlated with decreased lymphocyte (r = 1.000, p = 0.01), erythrocyte (r = 1.000, p = 0.003), and platelet counts (r = 0.841, p = 0.036). Significantly increased BUN levels (P < 0.05) correlated with the decrease in RBC-AChE activities (r = - 0.997, p = 0.047) and with the decrease in absolute red blood cell (r = - 0.883, p = 0.020) and lymphocyte (r = - 0.825, p = 0.043) counts; increased (P < 0.05) total plasma protein levels correlated with the decrease in RBC-AChE activities (r = -0.998, p = 0.043), absolute red blood cell (r = - 0.998, p = 0.040), lymphocyte (r = - 0.893, p = 0.017), and platelet (r = -0.855, p = 0.030) counts. The physiological responses correlated with the erythrocyte acetylcholinesterase inhibition could be considered as early indicators or warning responses of bovine

  2. Effect of sevoflurane on human neutrophil apoptosis.

    LENUS (Irish Health Repository)

    Tyther, R

    2012-02-03

    BACKGROUND AND OBJECTIVE: Both chronic occupational exposure to volatile anaesthetic agents and acute in vitro exposure of neutrophils to isoflurane have been shown to inhibit the rate of apoptosis of human neutrophils. It is possible that inhibition of neutrophil apoptosis arises through delaying mitochondrial membrane potential collapse. We assessed mitochondrial depolarization and apoptosis in unexposed neutrophils and neutrophils exposed to sevoflurane in vivo. METHODS: A total of 20 mL venous blood was withdrawn pre- and postinduction of anaesthesia, the neutrophils isolated and maintained in culture. At 1, 12 and 24 h in culture, the percentage of neutrophil apoptosis was assessed by dual staining with annexin V-FITC and propidium iodide. Mitochondrial depolarization was measured using the dual emission styryl dye JC-1. RESULTS: Apoptosis was significantly inhibited in neutrophils exposed to sevoflurane in vivo at 24 (exposed: 38 (12)% versus control: 28 (11)%, P = 0.001), but not at 1 or 12 h, in culture. Mitochondrial depolarization was not delayed in neutrophils exposed to sevoflurane. CONCLUSIONS: The most important findings are that sevoflurane inhibits neutrophil apoptosis in vivo and that inhibition is not mediated primarily by an effect on mitochondrial depolarization.

  3. Mechanisms of Degranulation in Neutrophils

    Directory of Open Access Journals (Sweden)

    Lacy Paige

    2006-09-01

    Full Text Available Abstract Neutrophils are critical inflammatory cells that cause tissue damage in a range of diseases and disorders. Being bone marrow-derived white blood cells, they migrate from the bloodstream to sites of tissue inflammation in response to chemotactic signals and induce inflammation by undergoing receptor-mediated respiratory burst and degranulation. Degranulation from neutrophils has been implicated as a major causative factor in pulmonary disorders, including severe asphyxic episodes of asthma. However, the mechanisms that control neutrophil degranulation are not well understood. Recent observations indicate that granule release from neutrophils depends on activation of intracellular signalling pathways, including β-arrestins, the Rho guanosine triphosphatase Rac2, soluble NSF attachment protein (SNAP receptors, the src family of tyrosine kinases, and the tyrosine phosphatase MEG2. Some of these observations suggest that degranulation from neutrophils is selective and depends on nonredundant signalling pathways. This review focuses on new findings from the literature on the mechanisms that control the release of granule-derived mediators from neutrophils.

  4. Diabetic microangiopathy and peripheral blood neutrophil count%糖尿病微血管病变与外周血中性粒细胞计数

    Institute of Scientific and Technical Information of China (English)

    王静; 何方平; 娜丽玛; 伊力哈木江·依马木

    2015-01-01

    BACKGROUND:Course of diabetes and hypertension are independent risk factors for diabetic microvascular complications, and fasting C-peptide level is a protective factor. There are many studies concerning glycated hemoglobin, C-reactive protein, white blood cel count and diabetic microvascular complications, but studies about neutrophils are rarely reported. OBJECTIVE:To investigate the relationship of peripheral blood neutrophil count with diabetic microvascular complications. METHODS:Totaly 112 patients with type 2 diabetes melitus were selected and divided into two groups according to the presence or absence of microvascular complications: diabetes group and diabetes+ microvascular complication group. The gender, age, ethnic group, disease history and laboratory examination parameters were colected and analyzed with univariate analysis and multivariate analysis. RESULTS AND CONCLUSION:(1) Univariate analysis: Patients in the diabetes+microvascular complication group had significantly higher age, course of diabetes, globulin, creatinine, the percentage of neutrophile granulocytes, nertrophil count, rate of hypertension than those in the diabetes group (P < 0.05), while the lymphocyte percentage, glomerular filtration rate, and triglyceride level were significantly lower than those in the diabetes group (P < 0.05). (2) Logistic regression analysis showed that age, course of diabetes and nertrophil count were the independent risk factors of microvascular complications in patients with type 2 diabetes melitus (odds ratio=1.155, 2.145, 2.275, alP < 0.01). Based on the adjustment of the age factor, the partial correlation analysis showed that the nertrophil count was positively correlated to the levels of creatinine, low-density lipoprotein cholesterol, globulin and hemoglobin, but negatively correlated to the levels of serum bile acid and glomerular filtration rate. Chronic subclinical inflammations are closely correlated to the occurrence of microvascular

  5. Postpartal immunometabolic gene network expression and function in blood neutrophils are altered in response to prepartal energy intake and postpartal intramammary inflammatory challenge.

    Science.gov (United States)

    Moyes, K M; Graugnard, D E; Khan, M J; Mukesh, M; Loor, J J

    2014-01-01

    The effect of over-feeding energy prepartum on blood polymorphonuclear neutrophil (PMN) response remains unclear. Cows fed controlled (CON; 1.34Mcal/kg of dry matter) or excess energy (OVE; 1.62Mcal/kg dry matter) during the dry period (~45d before expected calving date) received an intramammary (IM) challenge with Escherichia coli lipopolysaccharide (LPS) during the postpartal period to determine the effects of IM LPS and prepartal diet on the expression of key genes associated with immunometabolic response in blood PMN. Feed intake and daily milk yield were recorded throughout the study period. At 7d in milk (DIM), all cows received LPS (200µg) into 1 rear mammary quarter. Blood PMN were isolated at 7, 14, and 30 DIM, as well as before (0h) and after (12h) IM LPS challenge for gene expression analysis using quantitative real time PCR. Phagocytosis capabilities in vitro were assessed at 7, 14, and 30 DIM. Data were analyzed using the MIXED procedure of SAS with repeated measures. No differences in feed intake and milk yield were observed between OVE- and CON-fed cows. As expected, IM LPS challenge altered the expression of genes associated with the immune response (e.g., 1.9- and 1.8-fold for SELL and TLR2, respectively), metabolism (e.g., 1.8- and -1.8-fold for LDHA and SLC2A1, respectively), and transcription (e.g., 1.1- and 1.7-fold for NCOR1 and PPARD, respectively). At 12h postchallenge, an upregulation of TLR2 (1.8-fold), HIF1A (1.9-fold), and NFKB1 (1.5-fold) was observed for OVE rather than CON. At 7 DIM, S100A9 tended (2.2-fold) to be upregulated for OVE rather than CON. At 14 DIM, OVE resulted in lower PMN phagocytosis and an upregulation of NCOR2 (1.6-fold) and RXRA (1.9-fold) compared with CON-fed cows. At 30 DIM, an upregulation of MPO (3.5-fold) and PLA2G4A (1.5-fold) and a tendency for RXRA (1.7-fold) was observed for OVE- rather than CON-fed cows. Our results suggest that IM LPS challenge altered gene expression associated with metabolism in PMN

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

    Directory of Open Access Journals (Sweden)

    Eileen Uribe-Querol

    2015-01-01

    Full Text Available 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.

  7. Peripheral blood-derived bovine dendritic cells promote IgG1-restricted B cell responses in vitro.

    Science.gov (United States)

    Bajer, Anna A; Garcia-Tapia, David; Jordan, Kimberly R; Haas, Karen M; Werling, Dirk; Howard, Chris J; Estes, D Mark

    2003-01-01

    Regulation of humoral responses involves multiple cell types including the requirements for cognate interactions between T and B cells to drive CD40-dependent responses to T-dependent antigens. A third cell type has also been shown to play an essential role, the dendritic cell (DC). We demonstrate that bovine peripheral blood-derived (PB)-DC are similar in function to features described for human interstitial DC including the production of signature type 2 cytokines [interleukin (IL)-13, IL-10]. PB-DC express moderate-to-high costimulatory molecule expression, and major histocompatibility complex class II is negative for CD14 expression and has low or no expression of CD11c. Consistent with the interstitial phenotype is the ability of PB-DC to influence B cell activation and differentiation via direct expression of CD40L and type 2 cytokines. Collectively, these results suggest that direct B cell-DC interactions may promote an immunoglobulin-isotype expression pattern consistent with type 2 responses, independent of direct T cell involvement.

  8. Peripheral blood mononuclear cells from field cattle immune to bovine viral diarrhea virus (BVDV) are permissive in vitro to BVDV.

    Science.gov (United States)

    Gupta, V; Mishra, N; Pateriya, A; Behera, S P; Rajukumar, K

    2014-01-01

    The aim of this study was to determine the in vitro permissivity of peripheral blood mononuclear cells (PBMCs) from bovine viral diarrhea virus (BVDV)-immune field cattle to homologous and heterologous BVDVs. PBMCs from seventeen BVDV-naïve and sixteen BVDV-immune animals were infected with noncytopathic BVDV-1 or BVDV-2. The immune status of cattle was indicated by the presence of virus neutralizing antibodies, while viral load of PBMCs was determined by real-time RT-PCR. The results revealed that the PBMCs from naïve or immune animals were permissive to either BVDV-1 or BVDV-2, but the viral load was significantly higher for the naïve than for the immune animals. Furthermore, the load of homologous virus in PBMCs from immune animals was lower than that of heterologous virus. Our results provide evidence that the PBMCs from BVDV-immune cattle in field are susceptible to reinfection with homologous or heterologous BVDV, albeit to a lower extent in the former case.

  9. Clinical Microfluidics for Neutrophil Genomics and Proteomics

    OpenAIRE

    2010-01-01

    Neutrophils play critical roles in modulating the immune response. We present a robust methodology for rapidly isolating neutrophils directly from whole blood and develop ‘on-chip’ processing for mRNA and protein isolation for genomics and proteomics. We validate this device with an ex vivo stimulation experiment and by comparison with standard bulk isolation methodologies. Lastly, we implement this tool as part of a near patient blood processing system within a multi-center clinical study of...

  10. The influence of porcine prophenin on neutrophils isolated from rabbit blood during implantation of calcium sulphate graft material into bone tissue

    Directory of Open Access Journals (Sweden)

    Joanna Wessely-Szponder

    2012-10-01

    Full Text Available Immune dysfunction induced by surgical trauma may comprise either an inappropriately exaggerated inflammatory response or a profound suppression of cell- mediated immunity. Neutrophils are the leading cells in the first response to trauma. It is known that they mediate initial resistance to bacterial infection. Activated neutrophils can degranulate and release some enzymes such as elastase and myeloperoxidase (MPO. The function of elastase is, among others, to kill bacterial, whereas MPO is a specific enzyme of primary granules of neutrophils and a marker of in vivo neutrophil activation. Previous reports estimated that some cathelicidins could act to increase or diminish an innate immune response in which neutrophils participate. The aim of this study was to evaluate prophenins (PF isolated from porcine leukocytes in respect to neutrophil activity and survival during implantation of calcium sulphate bone grafts substitution in rabbits. Obtained results pointed out that neutrophils responded to  PF  depending upon concentration. Thirty min from implantation of calcium sulphate graft, we observed the greatest release of elastase (57.01±0.49% of maximal release in cultures stimulated with 10 mg/ml of PF, at 0 mg/ml was 51.15±0.23%, while after 24 h of incubation the greatest response was at a concentration of 20 mg/ml.  MPO release after 30 min from surgery decreased significantly at 10 mg/ml. In higher concentrations, the inhibition was less pronounced. Moreover, we estimated that PF causes cytotoxicity in the highest concentration as well as the apoptosis of neutrophils.

  11. 中国北方人初乳、牛初乳、牛常乳、牛血中胰岛素样生长因子-1和神经生长因子含量的比较%Comparison of IGF-1 and NGF in human colostrum, bovine colostrum,bovine milk and bovine blood serum in northern China

    Institute of Scientific and Technical Information of China (English)

    王洋; 生庆海; 张玉梅; 赵艾; 贾梦; 薛勇; 王培玉

    2011-01-01

    Objective To quantify and compare the levels of insulin-like growth factor 1 ( IGF-1 ) and nerve growth factor (NGF) in human colostrum, bovine colostrum, bovine milk and bovine blood serum. Methods Radioimmunoassay was used in the quantification of IGF-1 and NGF. Data were analyzed by using SPSS 13. 0. Results The levels of IGF-1 in human colostrum, bovine colostrum, bovine milk and bovine blood serum were 26. 91, 38.40, 20. 14 and 37.35 μg/L,respectively. The levels of NGF in human colostrum, bovine colostrum, milk and bovine blood serum were 300. 47,69.82, 110. 37 and 9. 63 ng/L, respectively. The differences between the levels of IGF-1 in human colostrum and bovine colostrum were not significant, but the levels of IGF-1 in bovine colostrum were obviously higher than that in bovine milk (P <0. 05), and there was no significant difference between the levels of IGF-1 in bovine milk and bovine blood serum.The level of NGF in human colostrum was obviously higher than that in bovine colostrum ( P < 0.05 ). There was no significant difference between the levels of NGF in bovine colostrum and bovine milk. The level of NGF in bovine blood serum was obviously lower than that in bovine milk ( P < 0. 05 ). Conclusion The level of IGF-1 in human colostrum was not significantly different from that in bovine colostrum, but the level of NGF in human colostrum was much higher than that in bovine colostrum.%目的 检测人初乳、集中饲养的新西兰进口荷斯坦乳牛的初乳、常乳和血液中胰岛素样生长因子-1(IGF-1)和神经生长因子(NGF)的含量,比较人初乳和牛初乳、牛初乳和牛常乳以及牛乳和同期采集的血液中IGF-1、NGF的含量.方法 采用放射免疫试剂盒测定人初乳、牛初乳、牛常乳和血中IGF-1、NGF的含量,使用SPSS 13.0进行统计分析.结果 本研究中测定的人初乳中IGF-1的含量是26.91μg/L,荷斯坦乳牛的初乳、常乳和血清中IGF-1的含量分别是38.40、20.14和37.35

  12. Effect of a Korean traditional formulation, Hwaotang, on superoxide generation in human neutrophils, platelet aggregation in human blood, and nitric oxide, prostaglandin E2 production and paw oedema induced by carrageenan in mice.

    Science.gov (United States)

    Park, Won-Hwan; Park, Soo-Young; Kim, Hyung-Min; Kim, Cheorl-Ho

    2004-02-01

    Hwaotang, a traditional Korean medicinal formulation, is a dried decoctum of a mixture of 7 herbal medicines, consisting of Angelica gigantis Radix, Rehmanniae radix, Paeoniae radix, Ciniamomi cortex, Cnidii rhizoma, Persicae semen and Carthami flos. We have investigated that Hwaotang water extract (HOT) has various effects on stimulus-induced superoxide generation in human neutrophils. The effects of HOT on superoxide generation in human neutrophils were investigated. HOT significantly inhibited N-formyl-methionyl-leucyl-phenylalanine (fMLP)-induced superoxide generation in a concentration-dependent manner, but not that induced by arachidonic acid (AA). On the other hand, HOT enhanced superoxide generation induced by phorbol 12-myristate 13-acetate (PMA) in a concentration-dependent manner. The superoxide generation induced by PMA with HOT was suppressed by staurosporine, an inhibitor of protein kinase C, but was not suppressed by genistein, an inhibitor of protein tyrosine kinase. Tyrosyl phosphorylation of a 58 kDa protein, which was increased by fMLP, was inhibited by HOT. HOT also inhibited the generation of a 47 kDa protein and platelet aggregation in human blood. The results suggest that protein tyrosine kinase participates in fMLP-mediated superoxide generation by HOT-treated human neutrophils. HOT inhibited neutrophil functions, including degranulation, superoxide generation, and leukotriene B4 production, without any effect on 5-lipoxygenase activity. HOT reduced nitric oxide (NO) and prostaglandin E2 production in mouse peritoneal macrophages stimulated with lipopolysaccharide, whereas no influence on the activity of iNOS, COX-2 or COX-1 was observed. HOT significantly reduced mouse paw oedema induced by carrageenan. Western blot analysis showed that HOT reduced the expression of iNOS and COX-2. The results indicate that HOT exerts anti-inflammatory effects related to the inhibition of neutrophil functions and of NO and prostaglandin E2 production, which

  13. Meningitic Escherichia coli K1 penetration and neutrophil transmigration across the blood-brain barrier are modulated by alpha7 nicotinic receptor.

    Directory of Open Access Journals (Sweden)

    Feng Chi

    Full Text Available Alpha7 nicotinic acetylcholine receptor (nAChR, an essential regulator of inflammation, is abundantly expressed in hippocampal neurons, which are vulnerable to bacterial meningitis. However, it is unknown whether α7 nAChR contributes to the regulation of these events. In this report, an aggravating role of α7 nAChR in host defense against meningitic E. coli infection was demonstrated by using α7-deficient (α7(-/- mouse brain microvascular endothelial cells (BMEC and animal model systems. As shown in our in vitro and in vivo studies, E. coli K1 invasion and polymorphonuclear neutrophil (PMN transmigration across the blood-brain barrier (BBB were significantly reduced in α7(-/- BMEC and α7(-/- mice. Stimulation by nicotine was abolished in the α7(-/- cells and animals. The same blocking effect was achieved by methyllycaconitine (α7 antagonist. The tight junction molecules occludin and ZO-1 were significantly reduced in the brain cortex of wildtype mice infected with E. coli and treated with nicotine, compared to α7(-/- cells and animals. Decreased neuronal injury in the hippocampal dentate gyrus was observed in α7(-/- mice with meningitis. Proinflammatory cytokines (IL-1β, IL-6, TNFα, MCP-1, MIP-1alpha, and RANTES and adhesion molecules (CD44 and ICAM-1 were significantly reduced in the cerebrospinal fluids of the α7(-/- mice with E. coli meningitis. Furthermore, α7 nAChR is the major calcium channel for nicotine- and E. coli K1-increased intracellular calcium concentrations of mouse BMEC. Taken together, our data suggest that α7 nAChR plays a detrimental role in the host defense against meningitic infection by modulation of pathogen invasion, PMN recruitment, calcium signaling and neuronal inflammation.

  14. CFTR targeting during activation of human neutrophils.

    Science.gov (United States)

    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.

  15. In vitro effects of Escherichia coli lipopolysaccharide on the function and gene expression of neutrophils isolated from the blood of dairy cows.

    Science.gov (United States)

    Revelo, X S; Waldron, M R

    2012-05-01

    The objectives of this study were to investigate the effect of Escherichia coli lipopolysaccharide (LPS) on the function of bovine neutrophils (PMNL) collected from mid lactation cows and determine the differential effects of LPS on gene expression of PMNL purified from early and mid lactation cows. The PMNL from mid lactation cows (187±13 d postpartum) were incubated with 0, 1, 25, and 50 μg/mL of LPS for 120 min, and the generation of reactive oxygen species (ROS), PMNL extracellular traps (NET), chemotaxis, and killing of Staphylococcus aureus were determined. Incubation of PMNL with 25 μg/mL of LPS increased intracellular ROS by 79% in mitogen-stimulated PMNL. Addition of 50 μg/mL of LPS enhanced intracellular ROS by nonstimulated and stimulated PMNL by 184 and 154%, respectively. Nonstimulated PMNL incubated with 25 and 50 μg/mL of LPS had a 105% increase in NET. Addition of LPS had no effect on subsequent PMNL chemotaxis or killing of Staph. aureus. To examine the effect of LPS on the expression of genes involved in PMNL function and cytokine production, mRNA was purified from PMNL isolated from mid lactation (146±2 postpartum; n=10) and early lactation cows (7 d postpartum; n=10), after a 120-min incubation with 0 or 50 μg/mL of LPS. Amounts of interleukin-8 (IL-8), tumor necrosis factor (TNF), bactericidal/permeability-increasing protein (BPI), myeloperoxidase (MPO), superoxide dismutase 2 (SOD2), NADPH oxidase 4 (NOX4), Cytochrome b-245, α polypeptide (CYBA), histone H2A/1 (H2A/1), and histone H2B-like (H2B) mRNA were determined relative to that of β-actin by real-time quantitative PCR. Regardless of stage of lactation, PMNL incubated with 50 μg/mL of LPS had 537 and 45% higher mRNA contents of IL-8 and SOD2 compared with 0 μg/mL LPS, respectively. In addition, LPS augmented the expression of TNF, BPI, and CYBA (2,908, 59, and 158% compared with controls, respectively) only in PMNL from mid lactation cows. Addition of LPS did not affect m

  16. Modulation of polymorphonuclear neutrophil functions by astrocytes

    Directory of Open Access Journals (Sweden)

    Xie Luokun

    2010-09-01

    Full Text Available Abstract Background Neuroinflammation is a complex process involving cells from the immune system and the central nerve system (CNS. Polymorphonuclear neutrophils (PMN are the most abundant class of white blood cells, and typically the first type of leukocyte recruited to sites of inflammation. In the CNS, astrocytes are the most abundant glial cell population and participate in the local innate immune response triggered by a variety of insults. In the present study, we investigated the impacts of astrocytes on PMN function. Methods Primary astrocyte cultures were derived from postnatal C57BL/6 mice and primary neutrophils were isolated from 8 to 12 weeks old C57BL/6 mice. PMNs respiratory burst was analyzed by H2DCFDA assay. For phagocytosis assay, neutrophils were incubated with FITC-labeled E. coli and the phagocytosis of E coli was determined by flow cytometer. PMNs degranulation was determined by myeloperoxidase assay. Cytokine expression was determined by real-time PCR. To determine the involvement of different signaling pathway, protein lysates were prepared and western blots were conducted to assess the activation of Akt, Erk1/2, and p38. Results Using ex vivo neutrophils and primary astrocyte cultures, our study demonstrated that astrocytes differentially regulate neutrophil functions, depending upon whether the interactions between the two cell types are direct or indirect. Upon direct cell-cell contact, astrocytes attenuate neutrophil apoptosis, respiratory bust, and degranulation, while enhancing neutrophil phagocytic capability and pro-inflammatory cytokine expression. Through indirect interaction with neutrophils, astrocytes attenuate apoptosis and enhance necrosis in neutrophils, augment neutrophil phagocytosis and respiratory burst, and inhibit neutrophil degranulation. In addition, astrocytes could augment Akt, Erk1/2, and p38 activation in neutrophils. Conclusions Astrocytes differentially regulate neutrophil functions through

  17. Labeling of rabbit neutrophils with (/sup 111/In)oxine

    Energy Technology Data Exchange (ETDEWEB)

    Lane, T.A. (Veterans Administration Medical Center, San Diego, CA (USA). Dept. of Pathology); Bergum, P.W.; Lichter, J.P.; Spragg, R.G. (California Univ., San Diego, La Jolla (USA). School of Medicine)

    1982-06-25

    The successful labeling of rabbit peripheral blood neutrophils with (/sup 111/In)oxine is reported here. Standard techniques for preparation of rabbit neutrophils, while acceptable for maintenance of in vitro function, rendered the neutrophils ineffective for in vivo use after labeling with /sup 111/In. Specifically, rabbit neutrophils were sensitive to the use of hypotonic shock for red cell elimination, centrifugation into a button during preparation, and the presence of oxine during chemotaxis in vitro. Using a carefully modified method of neutrophil preparation and labeling, it was found that /sup 111/In-labeled rabbit neutrophils retained normal in vitro function, including chemotaxis. In addition, using this method, 34% +- 5% of labeled neutrophils were recoverable in peripheral blood 5 min after intravenous injection. The half-life of circulating radiolabeled neutrophils was 5.6 +- 2 h. Continuous external imaging of radiolabeled neutrophils after intravenous injection showed initial lung uptake, followed by rapid clearance of radioactivity in the lungs (50% clearance in 10.5 +- 3.3 min.). Hepatic radioactivity was maximal by 30 min after injection and thereafter slowly declined. Finally, it was found that /sup 111/In-labeled rabbit neutrophils migrated to sites of artificially induced inflammation. These findings indicate that /sup 111/In-labeled rabbit neutrophils, if prepared under optimal conditions, should provide a useful tool for investigating the fate of neutrophils in experimental inflammatory conditions in this animal.

  18. Lyophilized bovine hemoglobin as a possible reference material for the determination of hemoglobin derivatives in human blood

    NARCIS (Netherlands)

    Maas, BHA; Buursma, A; Ernst, RAJ; Maas, AHJ; Zijlstra, WG

    1998-01-01

    We investigated the suitability of a lyophilized bovine hemoglobin (LBH) preparation containing various fractions of oxyhemoglobin (O(2)Hb), carboxyhemoglobin (COHb), and methemoglobin (MetHb) for quality assessment in multicomponent analysis (MCA) of hemoglobin derivatives. It was demonstrated that

  19. Calcium and magnesium concentrations in uterine fluid and blood serum during the estrous cycle in the bovine

    Directory of Open Access Journals (Sweden)

    Sayed Mortaza Alavi-Shoushtari

    2012-06-01

    Full Text Available To investigate uterine and serum Ca++ and Mg++ variations during the estrous cycle in the bovine, 66 genital tracts and blood samples were collected from Urmia abattoir, Urmia, Iran. The phase of the estrous cycle was determined by examination of the structures present on ovaries and uterine tonicity. Of the collected samples, 17 were pro-estrus, 12 estrus, 14 metestrus and 23 diestrus. The uterine fluid was collected by gentle scraping of the uterine mucosa with a curette. The mean ± SEM concentration of serum Ca++ in pro-estrus, estrus, metestrus and diestrus was 5.77 ± 0.69, 8.87 ± 1.83, 10.95 ± 1.52, 11.09 ± 1.08 mg dL-1, and the mean concentration of uterine fluid Ca++ was 4.40 ± 0.72, 3.15 ± 0.67, 5.89 ± 0.88, 8.63 ± 0.97 mg dL-1, respectively. The mean concentration of serum Mg++ in pro-estrus, estrus, metestrus and diestrus was 3.53 ± 0.30, 4.20 ± 0.52, 3.49 ± 0.38, 3.39 ± 0.29 mg dL-1, and mean concentration of uterine fluid Mg++ was 5.27 ± 0.42, 4.92 ± 0.60, 5.56 ± 0.30, 5.88 ± 0.36 mg dL-1, respectively. The serum and uterine fluid Ca++ in pro-estrus were significantly different from those of the metestrus and diestrus. In all stages of estrous cycle the mean concentration of serum Ca++ was higher than that in the uterine fluid. The difference between serum and uterine fluid Ca++ in estrus, metestrus and diestrus was significant. There was no significant difference between serum Mg++ content nor was it different from uterine fluid Mg++ content at any stages of estrous cycle. In all stages of estrous cycle the uterine fluid Mg++ was higher than that of the serum. These results suggest that during the estrous cycle in the cow, Ca++ is passively secreted in uterine fluids and is mostly dependent on blood serum Ca++ variations but Mg++ is secreted independently and does not follow variations in the serum concentrations.

  20. Supplementing Zn, Mn, and Cu from amino acid complexes and Co from cobalt glucoheptonate during the peripartal period benefits postpartal cow performance and blood neutrophil function.

    Science.gov (United States)

    Osorio, J S; Trevisi, E; Li, C; Drackley, J K; Socha, M T; Loor, J J

    2016-03-01

    /d more protein during the first 30 DIM. Although blood glucose, fatty acids, and liver triacylglycerol were not affected by diet, the Precision Xtra ketones (1.44 vs. 2.18 mmol/L) and γ-glutamyltransferase (liver function biomarker) were lower in AAC than INO. Furthermore, feeding AAC increased (D × T) polymorphonuclear neutrophilic lymphocyte phagocytosis, antioxidant capacity postpartum, and overall concentration of liver tissue Co and Cu. Overall, the positive response in milk yield and milk protein in AAC cows might be partly explained by the beneficial effects of AAC on postpartal DM intake driven at least in part by better liver and immune function as a result of improved antioxidant status.

  1. Photothermal image cytometry of human neutrophils

    Science.gov (United States)

    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.

  2. Neutrophils at work

    DEFF Research Database (Denmark)

    Nauseef, William M; Borregaard, Niels

    2014-01-01

    In this Review we discuss data demonstrating recently recognized aspects of neutrophil homeostasis in the steady state, granulopoiesis in 'emergency' conditions and interactions of neutrophils with the adaptive immune system. We explore in vivo observations of the recruitment of neutrophils from ...

  3. The effect of maternal antibodies on the detection of bovine virus diarrhoea virus in peripheral blood samples

    NARCIS (Netherlands)

    Zimmer, G.M.; Maanen, van C.; Goey, de I.; Brinkhof, J.; Wentink, G.H.

    2004-01-01

    Persistently infected animals (PI animals), that is those animals born after an intrauterine infection of the dam during the first 120 days of gestation, are the main source of bovine virus diarrhoea virus (BVD virus) in a cattle population. The success of any BVD virus eradication programme depends

  4. A study to assess and compare the peripheral blood neutrophil chemotaxis in smokers and non smokers with healthy periodontium, gingivitis, and chronic periodontitis

    OpenAIRE

    M Srinivas; Chethana, K. C.; Padma, R.; Suragimath, Girish; Anil, M.; Pai, B. S. Jagadish; Walvekar, Amit

    2012-01-01

    Background: Chronic periodontitis is the inflammation within the supporting tissues of the teeth resulting in attachment loss and bone loss. There are certain environmental factors such as smoking that can modify the host response to plaque organisms; hence can account for the aggressive progression of the disease. Smokers show a decreased expression of clinical inflammation even in the presence of abundant plaque accumulation. Neutrophils are the predominant host defense cells which protect ...

  5. Short communication: amino acid supplementation and stage of lactation alter apparent utilization of nutrients by blood neutrophils from lactating dairy cows in vitro

    Science.gov (United States)

    Glutamine is the preferred AA used by polymorphonuclear leukocytes (PMN) during the inflammatory response. However, the effect of other AA on bovine PMN response during inflammation and how this is altered by stage of lactation has not been fully elucidated. The objective of this study was to dete...

  6. Low-Chlorinated Non-Dioxin-like Polychlorinated Biphenyls Present in Blood and Breast Milk Induce Higher Levels of Reactive Oxygen Species in Neutrophil Granulocytes than High-Chlorinated Congeners.

    Science.gov (United States)

    Berntsen, Hanne Friis; Fonnum, Frode; Walaas, Sven Ivar; Bogen, Inger Lise

    2016-12-01

    Despite their ban several decades ago, polychlorinated biphenyls (PCBs) still pose a health threat to human beings due to their persistent and accumulative nature and continued presence in the environment. Non-dioxin-like (NDL)-PCBs have earlier been found to have effects on the immune system, including human neutrophil granulocytes. The aim of this study was to investigate the differences between ortho-chlorinated NDL-PCBs with a low or high degree of chlorination in their capability to induce the production of reactive oxygen species (ROS) in human neutrophil granulocytes in vitro. We used some of the congeners occurring at the highest levels in blood, breast milk and food: PCB 52 representing the low-chlorinated congeners and PCB 180 the high-chlorinated congeners. In addition, the extensively studied PCB 153 was included as a reference compound. ROS production was assessed with the luminol-amplified chemiluminescence and DCF fluorescence assays. The involvement of intracellular signalling mechanisms was investigated using different pharmacological substances. At high concentrations (10-20 μM), PCB 52 induced more ROS than PCB 153 and PCB 180. The role of extracellular signal-regulated kinase (ERK) 1/2 and/or ERK 5 signalling in PCB-induced ROS production was implicated through the reduction in ROS in the presence of the specific inhibitor U0126, whereas reduced ROS production after the use of SB203580 and SP600125 indicated the involvement of the p38 mitogen-activated protein kinase (MAPK) and c-Jun amino-terminal kinase (JNK) pathways, respectively. In addition, the calcineurin inhibitor FK-506, the intracellular calcium chelator BAPTA-AM and the antioxidant vitamin E reduced the levels of ROS. The intracellular signalling mechanisms involved in ROS production in human neutrophil granulocytes appeared to be similar for PCB 52, PCB 153 and PCB 180. Based on the results from the present and previous studies, we conclude that for abundant ortho-chlorinated PCBs

  7. Functional differentiation of normal human neutrophils.

    Science.gov (United States)

    Glasser, L; Fiederlein, R L

    1987-03-01

    In the past differentiation of human neutrophils has been defined by morphology, cytochemistry, or surface markers. In our experiments we have sequenced the various events that occur during the functional differentiation of the normal human neutrophil and have also examined some of the functional properties in relationship to surface markers and biochemical events. Granulocytes were obtained from the bone marrow and blood of hematologically normal individuals. Cells were separated into different stages of maturation by their physical properties using counterflow centrifugal elutriation and density gradient separation. Three cell fractions were obtained that were enriched for either immature myeloid cells, band neutrophils, or segmented neutrophils. Since the enriched fractions were not entirely pure, methodologies for functional assays were chosen that allowed cytologic evaluation of the functional capacity of each cell type. The criteria used to classify the stages of differentiation included both morphology by light microscopy and DNA labeling with tritiated thymidine. Various neutrophilic properties were studied: Fc receptors, complement receptors (CR1, CR3), phagocytosis of both live and dead opsonized Staphylococcus aureus, microbial killing of S aureus, NBT dye reduction after cellular stimulation with endotoxin, and chemotaxis. Our results indicate that the functional properties of the neutrophil appear in a distinct order. The sequence for the functional differentiation of the human neutrophil appears to be the following: Fc receptors----immune phagocytosis----complement receptors----oxygen-independent microbial killing----oxygen-dependent microbial killing----chemotaxis.

  8. Transcriptome kinetics of circulating neutrophils during human experimental endotoxemia.

    Directory of Open Access Journals (Sweden)

    Stan de Kleijn

    Full Text Available Polymorphonuclear cells (neutrophils play an important role in the systemic inflammatory response syndrome and the development of sepsis. These cells are essential for the defense against microorganisms, but may also cause tissue damage. Therefore, neutrophil numbers and activity are considered to be tightly regulated. Previous studies have investigated gene transcription during experimental endotoxemia in whole blood and peripheral blood mononuclear cells. However, the gene transcription response of the circulating pool of neutrophils to systemic inflammatory stimulation in vivo is currently unclear. We examined neutrophil gene transcription kinetics in healthy human subjects (n = 4 administered a single dose of endotoxin (LPS, 2 ng/kg iv. In addition, freshly isolated neutrophils were stimulated ex vivo with LPS, TNFα, G-CSF and GM-CSF to identify stimulus-specific gene transcription responses. Whole transcriptome microarray analysis of circulating neutrophils at 2, 4 and 6 hours after LPS infusion revealed activation of inflammatory networks which are involved in signaling of TNFα and IL-1α and IL-1β. The transcriptome profile of inflammatory activated neutrophils in vivo reflects extended survival and regulation of inflammatory responses. These changes in neutrophil transcriptome suggest a combination of early activation of circulating neutrophils by TNFα and G-CSF and a mobilization of young neutrophils from the bone marrow.

  9. Mycotic bovine nasal granuloma.

    Science.gov (United States)

    Conti Díaz, Ismael Alejandro; Vargas, Roberto; Apolo, Ada; Moraña, José Antonio; Pedrana, Graciela; Cardozo, Elena; Almeida, Edgardo

    2003-01-01

    A case of mycotic bovine nasal granuloma in a 10 year-old Jersey cow, produced by Drechslera halodes is presented. Histopathological sections showed abundant hyaline and pigmented extra and intracellular fungal structures together with a polymorphic cellular granuloma formed by neutrophils, lymphocytes, plasmocytes, histiocytes and giant cells of the Langhans type. It is the first case of mycotic bovine nasal granuloma recognized in Uruguay although this disease seems to be frequent according to the opinion of veterinarian specialists. Another similar clinical case also in a Jersey cow from the same dairy house with an intense cellular infiltrate rich in eosinophils without granulomatous image, together with extracellular hyaline and fuliginous fungal forms, is also referred for comparative purposes. Geotrichum sp. was isolated. The need of an early diagnosis and treatment of the disease is stressed.

  10. Separating the roles of nitrogen and oxygen in high pressure-induced blood-borne microparticle elevations, neutrophil activation, and vascular injury in mice.

    Science.gov (United States)

    Yang, Ming; Bhopale, Veena M; Thom, Stephen R

    2015-08-01

    An elevation in levels of circulating microparticles (MPs) due to high air pressure exposure and the associated inflammatory changes and vascular injury that occur with it may be due to oxidative stress. We hypothesized that these responses arise due to elevated partial pressures of N2 and not because of high-pressure O2. A comparison was made among high-pressure air, normoxic high-pressure N2, and high-pressure O2 in causing an elevation in circulating annexin V-positive MPs, neutrophil activation, and vascular injury by assessing the leakage of high-molecular-weight dextran in a murine model. After mice were exposed for 2 h to 790 kPa air, there were over 3-fold elevations in total circulating MPs as well as subgroups bearing Ly6G, CD41, Ter119, CD31, and CD142 surface proteins-evidence of neutrophil activation; platelet-neutrophil interaction; and vascular injury to brain, omentum, psoas, and skeletal muscles. Similar changes were found in mice exposed to high-pressure N2 using a gas mixture so that O2 partial pressure was the same as that of ambient air, whereas none of these changes occurred after exposures to 166 kPa O2, the same partial pressure that occurs during high-pressure air exposures. We conclude that N2 plays a central role in intra- and perivascular changes associated with exposure to high air pressure and that these responses appear to be a novel form of oxidative stress.

  11. Elevated manganese levels in blood and central nervous system occur before onset of clinical signs in scrapie and bovine spongiform encephalopathy.

    Science.gov (United States)

    Hesketh, S; Sassoon, J; Knight, R; Hopkins, J; Brown, D R

    2007-06-01

    Prion diseases, or transmissible spongiform encephalopathies, are neurodegenerative diseases that can only be accurately diagnosed by analysis of central nervous system tissue for the presence of an abnormal isoform of the prion protein known as PrP(Sc). Furthermore, these diseases have long incubation periods during which there are no clear symptoms but where the infectious agent could still be present in the tissues. Therefore, the development of diagnostic assays to detect a surrogate marker for the presence of prion disease is essential. Previous studies on mice experimentally infected with scrapie, an ovine spongiform encephalopathy, suggested that changes in the levels of Mn occur in the blood and brain before the onset of symptoms of the disease. To assess whether these findings have relevance to the animal diseases scrapie and bovine spongiform encephalopathy, tissues from bovine spongiform encephalopathy- and scrapie-infected cattle and sheep were analyzed for their metal content and compared with values for noninfected animals. In field cases and experimentally infected animals, elevated Mn was associated with prion infection. Although some central nervous system regions showed elevated Mn, other regions did not. The most consistent finding was an elevation of Mn in blood. This change was present in experimentally infected animals before the onset of symptoms. In scrapie-infected sheep, elevated Mn levels occurred regardless of the genotype of the sheep and were even detected in scrapie-resistant sheep in which no symptoms of disease were detected. These findings suggest that elevated blood Mn could be a potential diagnostic marker for prion infection even in the absence of apparent clinical disease.

  12. Elevated neutrophil and monocyte counts in peripheral blood are associated with poor survival in patients with metastatic melanoma: a prognostic model

    DEFF Research Database (Denmark)

    Schmidt, Henrik; Bastholt, Lars; Geertsen, Poul;

    2005-01-01

          treated as part of several phase II protocols and the majority received       treatment with intermediate dose subcutaneous IL-2 and interferon-alpha.       Neutrophil and monocyte counts, lactate dehydrogenase (LDH), number of       metastatic sites, location of metastases and performance status were all......       statistically significant prognostic factors in univariate analyses.       Subsequently, a multivariate Cox's regression analysis identified elevated       LDH (P

  13. Neutrophilic dermatoses in children.

    Science.gov (United States)

    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).

  14. Inorganic arsenic can be potent granulotoxin in mammalian neutrophils in vitro.

    Science.gov (United States)

    Taheri, Masumeh; Mehrzad, Jalil; Afshari, Reza; Saleh-Moghaddam, Massoud; Mahmudy Gharaie, Mohamad Hosein

    2016-09-01

    An important outcome arising out of occupational/environmental exposure to arsenic (As) is immunotoxicity. To determine the impact of inorganic As on innate immune cells, effects of a low dose of NaAsO2 (i.e. 20 ng As/ml) on select parameters associated with human and bovine neutrophils (PMN) were evaluated in vitro. PMN isolated from the blood of healthy individuals and cows (n = 8/treatment) were pre-incubated with NaAsO2 for 12 h before effects on PMN phagocytosis, transcription of TLR2, TLR4 and CD64 in human PMN - as well as on phagocytosis-dependent/-independent cell chemiluminescence (CL), phagocytosis and killing of Staphylococcus aureus and Escherichia coli, PMN H2O2 production and necrosis and TLR4 transcription in bovine PMN - were assessed. Relative to control (no As) PMN, treatment with As significantly decreased phagocytic capacity and CD64 mRNA, but increased TLR2 and TLR4 mRNA, in human PMN. In bovine PMN, while As also led to increased TLR4 mRNA abundance, it resulted in decreases in phagocytosis-dependent and -independent CL, PMN H2O2 production, PMN phagocytosis and killing of both E. coli and S. aureus by PMN. Considering the broad roles of PMN in immunology, the results of these studies increase our understanding of functional consequences of As exposure in inducing immunotoxicity and increasing susceptibility to (infectious) diseases in mammals.

  15. IL-1α-induced microvascular endothelial cells promote neutrophil killing by increasing MMP-9 concentration and lysozyme activity.

    Science.gov (United States)

    Liu, Xiaoye; Dong, Hong; Wang, Mingming; Gao, Ying; Zhang, Tao; Hu, Ge; Duan, Huiqing; Mu, Xiang

    2016-02-01

    The recruitment of neutrophils by endothelial cells during infection has been extensively studied, but little is known about the regulation of neutrophils activity by endothelial cells. To examine the role of microvascular endothelial cells in neutrophil killing, we established a transmigration model using rat intestinal microvascular endothelial cells (RIMVECs) and measured the extracellular and intracellular killing of Escherichia coli, Lactobacillus acidophilus, and Staphylococcus aureus by transendothelial neutrophils. We observed that blood neutrophils engulfed bacteria but did not kill them, and lipopolysaccharide- or hemolysin-injured RIMVECs inhibited the extracellular and intracellular bactericidal activity of transendothelial neutrophils. In comparison, interleukin-1α-induced RIMVECs promoted the extracellular and intracellular killing activity of transendothelial neutrophils and significantly increased MMP-9 concentration and lysozyme activity in transendothelial neutrophils (p neutrophils and bacterial toxin damage of endothelial cells led to reduction in bactericidal activity of transendothelial neutrophils. These findings offered new insight into the role of endothelial cells in the bactericidal activity of neutrophils.

  16. Clinical microfluidics for neutrophil genomics and proteomics.

    Science.gov (United States)

    Kotz, Kenneth T; Xiao, Wenzong; Miller-Graziano, Carol; Qian, Wei-Jun; Russom, Aman; Warner, Elizabeth A; Moldawer, Lyle L; De, Asit; Bankey, Paul E; Petritis, Brianne O; Camp, David G; Rosenbach, Alan E; Goverman, Jeremy; Fagan, Shawn P; Brownstein, Bernard H; Irimia, Daniel; Xu, Weihong; Wilhelmy, Julie; Mindrinos, Michael N; Smith, Richard D; Davis, Ronald W; Tompkins, Ronald G; Toner, Mehmet

    2010-09-01

    Neutrophils have key roles in modulating the immune response. We present a robust methodology for rapidly isolating neutrophils directly from whole blood with 'on-chip' processing for mRNA and protein isolation for genomics and proteomics. We validate this device with an ex vivo stimulation experiment and by comparison with standard bulk isolation methodologies. Last, we implement this tool as part of a near-patient blood processing system within a multi-center clinical study of the immune response to severe trauma and burn injury. The preliminary results from a small cohort of subjects in our study and healthy controls show a unique time-dependent gene expression pattern clearly demonstrating the ability of this tool to discriminate temporal transcriptional events of neutrophils within a clinical setting.

  17. The permeation of dynorphin A 1-6 across the blood brain barrier and its effect on bovine brain microvessel endothelial cell monolayer permeability.

    Science.gov (United States)

    Sloan, Courtney D Kuhnline; Audus, Kenneth L; Aldrich, Jane V; Lunte, Susan M

    2012-12-01

    Dynorphin A 1-17 (Dyn A 1-17) is an endogenous neuropeptide known to act at the kappa opioid receptor; it has been implicated in a number of neurological disorders, including neuropathic pain, stress, depression, and Alzheimer's and Parkinson's diseases. The investigation of Dyn A 1-17 metabolism at the blood-brain barrier (BBB) is important since the metabolites exhibit unique biological functions compared to the parent compound. In this work, Dyn A 1-6 is identified as a metabolite of Dyn A 1-17 in the presence of bovine brain microvessel endhothelial cells (BBMECs), using LC-MS/MS. The transport of Dyn A 1-6 at the BBB was examined using this in vitro cell culture model of the BBB. Furthermore, the permeation of the BBB by the low molecular weight permeability marker fluorescein was characterized in the presence and absences of Dyn A 1-6.

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  14. File list: ALL.Bld.05.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  15. File list: Unc.Bld.10.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  16. File list: ALL.Bld.10.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  17. File list: Unc.Bld.50.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Bld.50.AllAg.Neutrophils hg19 Unclassified Blood Neutrophils SRX956546,SRX95655...2,SRX956549 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Unc.Bld.50.AllAg.Neutrophils.bed ...

  18. Kinetics of cytokine expression in bovine PBMCs and whole blood after in vitro stimulation with foot-and-mouth disease virus (FMDV) antigen.

    Science.gov (United States)

    Dar, Pervaiz A; Hajam, Irshad A; Suryanarayana, Velavurthy S; Kishore, Subodh; Kondabattula, Ganesh

    2015-03-01

    The interest in analysing antigen-specific cytokine responses has substantially increased in recent years, in part due to their use in assessing vaccine efficacy. In the present study, the kinetics of IL-2, IL-4 and IFN-γ expression was determined in bovine PBMCs by real-time PCR and in whole blood by cytokine-release assay after in vitro stimulation with recall foot-and-mouth disease virus (FMDV) antigen. The results showed that the cytokine mRNA of IL-2 and IFN-γ in PBMCs were induced early (peak induction at 6 h), whereas the IL-4 mRNA showed delayed induction (peaked at 24 h). In contrast, the kinetics of cytokine proteins in whole blood was different and required the accumulation of the proteins before being optimally detected. The peak accumulation of cytokine protein in whole blood was recorded at 72 h for IL-2 and IL-4, and 96 h for IFN-γ. The findings of this study are of importance when selecting an optimal time points for measuring antigen-specific cytokine expression in cattle.

  19. A Lectin Purified from Blood Red Bracket Mushroom, Pycnoporus sanguineus (Agaricomycetidae), Mycelium Displayed Affinity Toward Bovine Transferrin.

    Science.gov (United States)

    Albores, Silvana; Moros, Maria; Cerdeiras, Maria Pia; de la Fuente, Jesus Martinez; Grazu, Valeria; Fraguas, Laura Franco

    2016-01-01

    Fungal lectins constitute excellent ligands for development of affinity adsorbents useful in affinity chromatography. In this work, a lectin was purified from Pycnoporus sanguineus (PSL) mycelium using 3 procedures: by affinity chromatography, using magnetic galactosyl-nanoparticles or galactose coupled to Sepharose, and by ionic exchange chromatography (IEC). The highest lectin yield was achieved by IEC (55%); SDS-PAGE of PSL showed 2 bands with molecular mass of 68.7 and 55.2 kDa and IEC displayed 2 bands at pi 5.5 and 5.2. The lectin agglutinates rat erythrocytes, exhibiting broad specificity toward several monosaccharides, including galactose. The agglutination was also inhibited by the glycoproteins fetal calf fetuin, bovine lactoferrin, bovine transferrin, and horseradish peroxidase. The lectin was then used to synthesize an affinity adsorbent (PSL-Sepharose) and the interaction with glycoproteins was evaluated by analyzing their chromatographic behaviors. The strongest interaction with the PSL-derivative was observed with transferrin, although lower interactions were also displayed toward fetuin and lactoferrin. These results indicate that the purified PSL constitutes an interesting ligand for the design of affinity adsorbents to be used (i.e., in glycoprotein purification).

  20. Evaluation of the interferon-γ assay on blood collected at exsanguination of cattle under field conditions for surveillance of bovine tuberculosis.

    Science.gov (United States)

    Okafor, C C; Grooms, D L; Bolin, S R; Averill, J J; Kaneene, J B

    2014-12-01

    Development of point of concentration (POC) surveillance strategies for bovine tuberculosis (bTB) would facilitate global efforts to eradicate bTB. The interferon-gamma (IFNγ) assay can detect IFNγ responses to Mycobacterium bovis in blood collected at commencement of exsanguination (COE) of experimentally challenged cattle but has not been evaluated under field conditions. The current study was aimed at determining (i) whether blood collected at COE of cattle at slaughter, under field conditions, is practical to obtain and useful for identifying cattle as IFNγ positive for bTB, (ii) whether the results of the IFNγ assay obtained at COE reliably compare with results obtained from live animals in the field, and (iii) whether the identified animal(s) originated from bTB-infected or bTB-exposed herds. Cattle from three risk groups were used: the highest risk group consisted of 49 cattle from 3 bTB-infected herds; the medium risk group consisted of 24 cattle from a potentially exposed herd; and the lowest risk group consisted of 60 cattle from herds with no known history of bTB exposure. The IFNγ assay was performed on blood collected both before stunning and at COE of cattle at slaughter. An enhanced slaughter inspection for gross lesions consistent with bTB was performed on all cattle. In addition, lymph nodes were cultured for M. bovis for cattle that tested positive for bTB via the IFNγ assay and for most cattle that tested negative for bTB. Cattle, both with and without lesions consistent with bTB, were identified as positive for bTB by the IFNγ assay using blood collected at COE, but none of the positive cattle originated from the lowest risk group. The current study demonstrates that blood collected at COE of cattle is both a practical and moderately reliable sample for accessing bTB infection using the IFNγ assay.

  1. Myeloperoxidase Stimulates Neutrophil Degranulation.

    Science.gov (United States)

    Grigorieva, D V; Gorudko, I V; Sokolov, A V; Kostevich, V A; Vasilyev, V B; Cherenkevich, S N; Panasenko, O M

    2016-08-01

    Myeloperoxidase, heme enzyme of azurophilic granules in neutrophils, is released into the extracellular space in the inflammation foci. In neutrophils, it stimulates a dose-dependent release of lactoferrin (a protein of specific granules), lysozyme (a protein of specific and azurophilic granules), and elastase (a protein of azurophilic granules). 4-Aminobenzoic acid hydrazide, a potent inhibitor of peroxidase activity of myeloperoxidase, produced no effect on neutrophil degranulation. Using signal transduction inhibitors (genistein, methoxyverapamil, wortmannin, and NiCl2), we demonstrated that myeloperoxidase-induced degranulation of neutrophils resulted from enzyme interaction with the plasma membrane and depends on activation of tyrosine kinases, phosphatidylinositol 3-kinases (PI3K), and calcium signaling. Myeloperoxidase modified by oxidative/halogenation stress (chlorinated and monomeric forms of the enzyme) lost the potency to activate neutrophil degranulation.

  2. Quantitative proteomics reveals differential biological processes in healthy neonatal cord neutrophils and adult neutrophils

    KAUST Repository

    Zhu, Jiang

    2014-06-11

    Neonatal neutrophils are characterized by the immaturity of bactericidal mechanisms that contributes largely to neonatal mortality. However, underlying molecular mechanism associated with the immaturity remains incompletely understood. In this study, we performed comparative proteomic analysis on neonatal neutrophils derived from human cord blood and adult peripheral neutrophils. A total of 1332 proteins were identified and quantified, and 127 proteins were characterized as differentially expressed between adult and cord neutrophils. The differentially expressed proteins are mapped in KEGG pathways into five clusters and indicated impaired functions of neonatal neutrophils in proteasome, lysosome, phagosome, and leukocyte transendothelial migration. In particular, many proteins associated with NETosis, a critical mechanism for antimicrobial process and auto-clearance, were also found to be downregulated in cord neutrophils. This study represents a first comparative proteome profiling of neonatal and adult neutrophils, and provides a global view of differentially expressed proteome for enhancing our understanding of their various functional difference. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Ranitidine improves postoperative monocyte and neutrophil function

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Nielsen, H; Jensen, S;

    1994-01-01

    BACKGROUND: The histamine H2-receptor antagonist ranitidine hydrochloride has been shown to improve trauma-, blood transfusion-, and sepsis-induced immunosuppression. OBJECTIVE: To evaluate the effect of ranitidine on postoperative impairment in monocyte and neutrophil function. METHODS: Twenty...... difference (P detected. There were no infectious complications in ranitidine-treated patients. CONCLUSION: These results support previous studies...

  4. N-Formyl-Perosamine Surface Homopolysaccharides Hinder the Recognition of Brucella abortus by Mouse Neutrophils.

    Science.gov (United States)

    Mora-Cartín, Ricardo; Chacón-Díaz, Carlos; Gutiérrez-Jiménez, Cristina; Gurdián-Murillo, Stephany; Lomonte, Bruno; Chaves-Olarte, Esteban; Barquero-Calvo, Elías; Moreno, Edgardo

    2016-06-01

    Brucella abortus is an intracellular pathogen of monocytes, macrophages, dendritic cells, and placental trophoblasts. This bacterium causes a chronic disease in bovines and in humans. In these hosts, the bacterium also invades neutrophils; however, it fails to replicate and just resists the killing action of these leukocytes without inducing significant activation or neutrophilia. Moreover, B. abortus causes the premature cell death of human neutrophils. In the murine model, the bacterium is found within macrophages and dendritic cells at early times of infection but seldom in neutrophils. Based on this observation, we explored the interaction of mouse neutrophils with B. abortus In contrast to human, dog, and bovine neutrophils, naive mouse neutrophils fail to recognize smooth B. abortus bacteria at early stages of infection. Murine normal serum components do not opsonize smooth Brucella strains, and neutrophil phagocytosis is achieved only after the appearance of antibodies. Alternatively, mouse normal serum is capable of opsonizing rough Brucella mutants. Despite this, neutrophils still fail to kill Brucella, and the bacterium induces cell death of murine leukocytes. In addition, mouse serum does not opsonize Yersinia enterocolitica O:9, a bacterium displaying the same surface polysaccharide antigen as smooth B. abortus Therefore, the lack of murine serum opsonization and absence of murine neutrophil recognition are specific, and the molecules responsible for the Brucella camouflage are N-formyl-perosamine surface homopolysaccharides. Although the mouse is a valuable model for understanding the immunobiology of brucellosis, direct extrapolation from one animal system to another has to be undertaken with caution.

  5. Olfactomedin 4 defines a subset of human neutrophils

    DEFF Research Database (Denmark)

    Clemmensen, Stine N; Bohr, Christina T; Rørvig, Sara;

    2012-01-01

    OLFM4 was identified initially as a gene highly induced in myeloid stem cells by G-CSF treatment. A bioinformatics method using a global meta-analysis of microarray data predicted that OLFM4 would be associated with specific granules in human neutrophils. Subcellular fractionation of peripheral...... blood neutrophils demonstrated complete colocalization of OLFM4 with the specific granule protein NGAL, and stimulation of neutrophils with PMA resulted in corelease of NGAL and OLFM4, proving that OLFM4 is a genuine constituent of neutrophil-specific granules. In accordance with this, OLFM4 mRNA peaked...... at the MY/MM stage of maturation. OLFM4 was, however, present in only 20-25% of peripheral blood neutrophils, as determined by immunocytochemistry and flow cytometry, whereas mRNA for OLFM4 was present in all MY/MM, indicating post-transcriptional regulation as a basis for the heterogeneous expression...

  6. A serine proteinase inhibitor isolated from Tamarindus indica seeds and its effects on the release of human neutrophil elastase.

    Science.gov (United States)

    Fook, J M S L L; Macedo, L L P; Moura, G E D D; Teixeira, F M; Oliveira, A S; Queiroz, A F S; Sales, M P

    2005-05-01

    Proteinaceous inhibitors with high inhibitory activities against human neutrophil elastase (HNE) were found in seeds of the Tamarind tree (Tamarindus indica). A serine proteinase inhibitor denoted PG50 was purified using ammonium sulphate and acetone precipitation followed by Sephacryl S-300 and Sephadex G-50 gel filtration chromatographies. Inhibitor PG50 showed a Mr of 14.9 K on Sephadex G-50 calibrated column and a Mr of 11.6 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. PG50 had selective activity while cysteine proteinases (papain and bromelain) and serine proteinases (porcine pancreatic elastase and bovine chymotrypsin) were not inhibited, it was strongly effective against serine proteinases such as bovine trypsin and isolated human neutrophil elastase. The IC50 value was determined to be 55.96 microg.mL-1. PG50 showed neither cytotoxic nor haemolytic activity on human blood cells. After pre-incubation of PG50 with cytochalasin B, the exocytosis of elastase was initiated using PAF and fMLP. PG50 exhibited different inhibition on elastase release by PAF, at 44.6% and on release by fMLP, at 28.4%. These results showed that PG50 preferentially affected elastase release by PAF stimuli and this may indicate selective inhibition on PAF receptors.

  7. Bovine milk exosome proteome

    Science.gov (United States)

    Exosomes are 40-100 nm membrane vesicles of endocytic origin and are found in blood, urine, amniotic fluid, bronchoalveolar lavage (BAL) fluid, as well as human and bovine milk. Exosomes are extracellular organelles important in intracellular communication/signaling, immune function, and biomarkers ...

  8. Inducible nitric oxide synthase (NOS II) is constitutive in human neutrophils.

    Science.gov (United States)

    Cedergren, Jan; Follin, Per; Forslund, Tony; Lindmark, Maria; Sundqvist, Tommy; Skogh, Thomas

    2003-10-01

    The objective was to study the expression of inducible nitric oxide synthase (NOS II) in and NO production by human blood neutrophils and in in vivo exudated neutrophils. Cellular expression of NOS II was evaluated by flow cytometry in whole blood, in isolated blood neutrophils, and in neutrophils obtained by exudation in vivo into skin chambers. Neutrophil NOS II was also demonstrated by Western blotting. Uptake of 3H-labelled L-arginine was studied in vitro and NOS activity measured in a whole cell assay by the conversion of 3H-arginine to 3H-citrulline. In contrast to unseparated blood cells, NOS II was demonstrable both in isolated blood neutrophils and exudated cells. The failure to detect NOS II by flow cytometry in whole blood cells thus proved to be due to the quenching effect of hemoglobin. Western blotting revealed a 130 kD band corresponding to NOS II in isolated blood neutrophils, but detection was dependent on diisopropylfluorophosphate for proteinase inhibition. L-arginine was taken up by neutrophils, but enzymatic activity could not be demonstrated. We conclude that human neutrophils constitutively express NOS II, but that its demonstration by FITC-labelling is inhibited by hemoglobin-mediated quenching in whole blood samples.

  9. High affinity capture and concentration of quinacrine in polymorphonuclear neutrophils via vacuolar ATPase-mediated ion trapping: Comparison with other peripheral blood leukocytes and implications for the distribution of cationic drugs

    Energy Technology Data Exchange (ETDEWEB)

    Roy, Caroline; Gagné, Valérie; Fernandes, Maria J.G.; Marceau, François, E-mail: francois.marceau@crchul.ulaval.ca

    2013-07-15

    Many cationic drugs are concentrated in acidic cell compartments due to low retro-diffusion of the protonated molecule (ion trapping), with an ensuing vacuolar and autophagic cytopathology. In solid tissues, there is evidence that phagocytic cells, e.g., histiocytes, preferentially concentrate cationic drugs. We hypothesized that peripheral blood leukocytes could differentially take up a fluorescent model cation, quinacrine, depending on their phagocytic competence. Quinacrine transport parameters were determined in purified or total leukocyte suspensions at 37 °C. Purified polymorphonuclear leukocytes (PMNLs, essentially neutrophils) exhibited a quinacrine uptake velocity inferior to that of lymphocytes, but a consistently higher affinity (apparent K{sub M} 1.1 vs. 6.3 μM, respectively). However, the vacuolar (V)-ATPase inhibitor bafilomycin A1 prevented quinacrine transport or initiated its release in either cell type. PMNLs capture most of the quinacrine added at low concentrations to fresh peripheral blood leukocytes compared with lymphocytes and monocytes (cytofluorometry). Accumulation of the autophagy marker LC3-II occurred rapidly and at low drug concentrations in quinacrine-treated PMNLs (significant at ≥ 2.5 μM, ≥ 2 h). Lymphocytes contained more LAMP1 than PMNLs, suggesting that the mass of lysosomes and late endosomes is a determinant of quinacrine uptake V{sub max}. PMNLs, however, exhibited the highest capacity for pinocytosis (uptake of fluorescent dextran into endosomes). The selectivity of quinacrine distribution in peripheral blood leukocytes may be determined by the collaboration of a non-concentrating plasma membrane transport mechanism, tentatively identified as pinocytosis in PMNLs, with V-ATPase-mediated concentration. Intracellular reservoirs of cationic drugs are a potential source of toxicity (e.g., loss of lysosomal function in phagocytes). - Highlights: • Quinacrine is concentrated in acidic organelles via V-ATPase-mediated ion

  10. Human filarial Wolbachia lipopeptide directly activates human neutrophils in vitro.

    Science.gov (United States)

    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.

  11. Bayesian estimation of sensitivity and specificity of Coxiella burnetii antibody ELISA tests in bovine blood and milk

    DEFF Research Database (Denmark)

    Paul, Suman; Toft, Nils; Agerholm, Jørgen S.;

    2013-01-01

    Serological tests for Coxiella burnetii (the causative agent of Q fever) antibodies are usually based on enzyme linked immunosorbent assay (ELISA) although this method is not thoroughly evaluated. The objective of this study was to determine the sensitivity and specificity of an ELISA for detection...... of the ELISA methods on milk and blood were equal at 0.99. No conditional dependence was observed between the specificity estimates of the two test methods. However, the sensitivity estimates of both tests were significantly reduced when conditional covariances ≥40 were used. Collection of milk samples from...... to positive (S/P) cut-off of 40 for both blood and milk ELISAs. At this cut-off, sensitivity of milk ELISA was 0.86 (95% posterior credibility interval [PCI] [0.76; 0.96]). This was slightly but insignificantly higher than sensitivity of blood ELISA (0.84; 95% PCI [0.75; 0.93]). The specificity estimates...

  12. Ultra-sensitive detection of prion protein fibrils by flow cytometry in blood from cattle affected with bovine spongiform encephalopathy

    Directory of Open Access Journals (Sweden)

    Maas Elke

    2005-10-01

    Full Text Available Abstract Background The definite diagnosis of prion diseases such as Creutzfeldt-Jakob disease (CJD in humans or bovine spongiform encephalopathy (BSE in cattle currently relies on the post mortem detection of the pathological form of the prion protein (PrPSc in brain tissue. Infectivity studies indicate that PrPSc may also be present in body fluids, even at presymptomatic stages of the disease, albeit at concentrations well below the detection limits of currently available analytical methods. Results We developed a highly sensitive method for detecting prion protein aggregates that takes advantage of kinetic differences between seeded and unseeded polymerization of prion protein monomers. Detection of the aggregates was carried out by flow cytometry. In the presence of prion seeds, the association of labelled recombinant PrP monomers in plasma and serum proceeds much more efficiently than in the absence of seeds. In a diagnostic model system, synthetic PrP aggregates were detected down to a concentration of approximately 10-8 nM [0.24 fg/ml]. A specific signal was detected in six out of six available serum samples from BSE-positive cattle. Conclusion We have developed a method based on seed-dependent PrP fibril formation that shows promising results in differentiating a small number of BSE-positive serum samples from healthy controls. This method may provide the basis for an ante mortem diagnostic test for prion diseases.

  13. Neutrophil biology: an update

    OpenAIRE

    Kobayashi, Yoshiro

    2015-01-01

    Neutrophil extracellular traps (NETs) are involved in bacterial killing as well as autoimmunity, because NETs contain proteases, bactericidal peptides, DNA and ribonucleoprotein. NETs are formed via a novel type of cell death called NETosis. NETosis is distinct from apoptosis, but it resembles necrosis in that both membranes are not intact so that they allow intracellular proteins to leak outside of the cells. Removal of NETs and neutrophils undergoing NETosis by phagocytes and its subsequent...

  14. [The phagocytosis of polymorphonuclear neutrophilic granulocytes in progressive periodontitis].

    Science.gov (United States)

    Konopka, T; Zietek, M

    1995-01-01

    The aim of this paper was the evaluation of the phagocytic activity of neutrophils in blood and in gingival pocket fluid in patients suffering from rapidly progressive periodontitis (RPP) and postjuvenile periodontitis (PJP). Prior to periodontal treatment the authors evaluated the capacity to phagocytose latex particles of peripheral blood neutrophils from 21 patients with RPP, 51 with PJP and 59 healthy subjects (control group) as well as the phagocytic activity of neutrophils in pocket fluid from 21 patients with RPP, 14 with PJP and from 20 healthy subjects. This phagocytic activity was significantly lower in all examined groups in comparison with the control group. A similar evaluation executed 3 months after treatment revealed normal phagocytosis of blood neutrophils from patients with RPP. In patients receiving complementary pharmacotherapy (spiramycine combined with metronidazol), a better improvement of phagocytosis was noted, than that observed in patients treated only surgically.

  15. Human Umbilical Cord Blood Serum: Effective Substitute of Fetal Bovine Serum for Culturing of Human Multipotent Mesenchymal Stromal Cells.

    Science.gov (United States)

    Romanov, Yu A; Balashova, E E; Volgina, N E; Kabaeva, N V; Dugina, T N; Sukhikh, G T

    2017-02-01

    Optimal conditions for culturing of multipotent mesenchymal stromal cells in the presence of pooled umbilical cord blood serum were determined. It was found that umbilical cord blood serum in a concentration range of 1-10% effectively supported high viability and proliferative activity of cells with unaltered phenotype and preserved multilineage differentiation capacity. The proposed approach allows avoiding the use of xenogenic animal sera for culturing of multipotent mesenchymal stromal cells and creates prerequisites for designing and manufacturing safe cellular and/or acellular products for medical purposes.

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

    Science.gov (United States)

    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.

  17. FUNCTIONAL AND METABOLIC ACTIVITY OF NEUTROPHILIC GRANULOCYTES IN CASE OF ACUTE BACTERIAL RHINOSINUSITIS

    Directory of Open Access Journals (Sweden)

    O. A. Kolenchukova

    2013-01-01

    Full Text Available Abstract. The functional and metabolic activities of neutrophilic granulocytes in patients with acute bacterial rhinosinusitis (ABRS have been studied. Characteristics of the indices of chemiluminescence and bioluminescence for neutrophils, extracted from venous blood and maxillary sinus were compared. It was demonstrated the decrease of intensity of APK production in neutrophils, extracted from inflammation point, with simultaneous decrease of intensity of plastic processes and increasing of energy processes in compare with the same indices in blood cells.

  18. Genetic characterization of bovine viral diarrhea virus strains in Beijing, China and innate immune responses of peripheral blood mononuclear cells in persistently infected dairy cattle.

    Science.gov (United States)

    Weng, Xiao Gang; Song, Quan Jiang; Wu, Qiong; Liu, Ming Chao; Wang, Meng Ling; Wang, Jiu Feng

    2015-01-01

    To acquire epidemiological data on the bovine viral diarrhea virus (BVDV) and identify cattle persistently infected (PI) with this virus, 4,327 samples from Holstein dairy cows were screened over a four-year period in Beijing, China. Eighteen BVD viruses were isolated, 12 from PI cattle. Based on genetic analysis of their 5'-untranslated region (5'-UTR), the 18 isolates were assigned to subgenotype BVDV-1m, 1a, 1d, 1q, and 1b. To investigate the innate immune responses in the peripheral-blood mononuclear cells of PI cattle, the expression of Toll-like receptors (TLRs), RIG-I-like receptors, interferon-α (IFN-α), IFN-β, myxovirus (influenza virus) resistance 1 (MX1), and interferon stimulatory gene 15 (ISG15) was assessed by qPCR. When compared with healthy cattle, the expression of TLR-7, IFN-α, and IFN-β mRNA was downregulated, but the expression of MX1 and ISG-15 mRNA was upregulated in PI cattle. Immunoblotting analysis revealed that the expression of interferon regulatory factor 3 (IRF-3) and IRF-7 was lower in PI cattle than in healthy cattle. Thus, BVDV-1m and 1a are the predominant subgenotypes in the Beijing region, and the strains are highly divergent. Our findings also suggest that the TLR-7/IRF-7 signaling pathway plays a role in evasion of host restriction by BVDV.

  19. Metabolic requirements for neutrophil extracellular traps formation

    Science.gov (United States)

    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

  20. A large-scale electrophoresis- and chromatography-based determination of gene expression profiles in bovine brain capillary endothelial cells after the re-induction of blood-brain barrier properties

    Directory of Open Access Journals (Sweden)

    Duban-Deweer Sophie

    2010-11-01

    Full Text Available Abstract Background Brain capillary endothelial cells (BCECs form the physiological basis of the blood-brain barrier (BBB. The barrier function is (at least in part due to well-known proteins such as transporters, tight junctions and metabolic barrier proteins (e.g. monoamine oxidase, gamma glutamyltranspeptidase and P-glycoprotein. Our previous 2-dimensional gel proteome analysis had identified a large number of proteins and revealed the major role of dynamic cytoskeletal remodelling in the differentiation of bovine BCECs. The aim of the present study was to elaborate a reference proteome of Triton X-100-soluble species from bovine BCECs cultured in the well-established in vitro BBB model developed in our laboratory. Results A total of 215 protein spots (corresponding to 130 distinct proteins were identified by 2-dimensional gel electrophoresis, whereas over 350 proteins were identified by a shotgun approach. We classified around 430 distinct proteins expressed by bovine BCECs. Our large-scale gene expression analysis enabled the correction of mistakes referenced into protein databases (e.g. bovine vinculin and constitutes valuable evidence for predictions based on genome annotation. Conclusions Elaboration of a reference proteome constitutes the first step in creating a gene expression database dedicated to capillary endothelial cells displaying BBB characteristics. It improves of our knowledge of the BBB and the key proteins in cell structures, cytoskeleton organization, metabolism, detoxification and drug resistance. Moreover, our results emphasize the need for both appropriate experimental design and correct interpretation of proteome datasets.

  1. Alteration of the exDNA profile in blood serum of LLC-bearing mice under the decrease of tumour invasion potential by bovine pancreatic DNase I treatment

    Science.gov (United States)

    Brenner, Evgenyi V.; Kurilshikov, Alexander M.; Patutina, Olga A.; Zenkova, Marina A.

    2017-01-01

    Taking into account recently obtained data indicating the participation of circulating extracellular DNA (exDNA) in tumorigenesis, enzymes with deoxyribonucleic activity have again been considered as potential antitumour and antimetastatic drugs. Previously, using murine Lewis lung carcinoma and hepatocellular carcinoma A1 tumour models, we have shown the antimetastatic activity of bovine DNase I, which correlates with an increase of DNase activity and a decrease of exDNA concentration in the blood serum of tumour-bearing mice. In this work, using next-generation sequencing on the ABS SOLiD™ 5.500 platform, we performed a search for molecular targets of DNase I by comparing the exDNA profiles of healthy animals, untreated animals with Lewis lung carcinoma (LLC) and those with LLC treated with DNase I. We found that upon DNase I treatment of LLC-bearing mice, together with inhibition of metastasis, a number of strong alterations in the patterns of exDNA were observed. The major differences in exDNA profiles between groups were: i) the level of GC-poor sequences increased during tumour development was reduced to that of healthy mice; ii) levels of sequences corresponding to tumour-associated genes Hmga2, Myc and Jun were reduced in the DNase I-treated group in comparison with non-treated mice; iii) 224 types of tandem repeat over-presented in untreated LLC-bearing mice were significantly reduced after DNase I treatment. The most important result obtained in the work is that DNase I decreased the level of B-subfamily repeats having homology to human ALU repeats, known as markers of carcinogenesis, to the level of healthy animals. Thus, the obtained data lead us to suppose that circulating exDNA plays a role in tumour dissemination, and alteration of multiple molecular targets in the bloodstream by DNase I reduces the invasive potential of tumours. PMID:28222152

  2. Changes in some pro-and anti-inflammatory cytokines produced by bovine peripheral blood mononuclear cells following foot and mouth disease vaccination

    Directory of Open Access Journals (Sweden)

    N. Delirezh

    2016-09-01

    Full Text Available Interleukin (IL-17 is exclusively produced by CD4 helper T-cells upon activation. It most often acts as a pro-inflammatory cytokine, which stimulates the release of pro-inflammatory cytokines IL-6, IL-8, TNF-α, and granulocyte-macrophage colony-stimulating factor (GM-CSF. In this study, we studied the in-vitro IL-17 response to specific antigens and a variety of mitogens and compared the IL-17 response to IL-2, IL-4, IL-5, IL-6, IL-10, and IFN-γ responses. We used a foot and mouth disease (FMD vaccine as specific antigens and mitogens (phytohemagglutinin [PHA], pokeweed mitogen [PWM], and concanavalin A [Con A] to stimulate peripheral blood mononuclear cells (PBMCs of vaccinated calves. Cell culture supernatant was harvested and analyzed for cytokines, using commercially available bovine ELISA kits. The mitogens induced a significant increase in IL-17 production. IL-17 was produced at high levels in response to the T cell-stimulated mitogens, PHA, and Con A, and at low levels in response to PWM mitogens. In contrast, level of the produced IL-17 cytokines in response to the FMDV antigens was lower as compared to those produced by mitogens. The FMDV antigens and mitogens significantly increased IL-17 production. There was not a correlation between IL-17 production and type-1 cytokine, IFN-γ, and IL-2, while there was a correlation between type-2 cytokine, IL-4, and IL-5 at either cytokine level produced by PBMCs stimulated by FMDV antigens. Moreover, there was an interaction between IL-17 and IL-6, that is, as IL-6 cytokine level elevated or diminished, IL-17 cytokine level increased or decreased, as well.

  3. Ultrasensitive detection of PrP(Sc) in the cerebrospinal fluid and blood of macaques infected with bovine spongiform encephalopathy prion.

    Science.gov (United States)

    Murayama, Yuichi; Masujin, Kentaro; Imamura, Morikazu; Ono, Fumiko; Shibata, Hiroaki; Tobiume, Minoru; Yamamura, Tomoaki; Shimozaki, Noriko; Terao, Keiji; Yamakawa, Yoshio; Sata, Tetsutaro

    2014-11-01

    Prion diseases are characterized by the prominent accumulation of the misfolded form of a normal cellular protein (PrP(Sc)) in the central nervous system. The pathological features and biochemical properties of PrP(Sc) in macaque monkeys infected with the bovine spongiform encephalopathy (BSE) prion have been found to be similar to those of human subjects with variant Creutzfeldt-Jakob disease (vCJD). Non-human primate models are thus ideally suited for performing valid diagnostic tests and determining the efficacy of potential therapeutic agents. In the current study, we developed a highly efficient method for in vitro amplification of cynomolgus macaque BSE PrP(Sc). This method involves amplifying PrP(Sc) by protein misfolding cyclic amplification (PMCA) using mouse brain homogenate as a PrP(C) substrate in the presence of sulfated dextran compounds. This method is capable of amplifying very small amounts of PrP(Sc) contained in the cerebrospinal fluid (CSF) and white blood cells (WBCs), as well as in the peripheral tissues of macaques that have been intracerebrally inoculated with the BSE prion. After clinical signs of the disease appeared in three macaques, we detected PrP(Sc) in the CSF by serial PMCA, and the CSF levels of PrP(Sc) tended to increase with disease progression. In addition, PrP(Sc) was detectable in WBCs at the clinical phases of the disease in two of the three macaques. Thus, our highly sensitive, novel method may be useful for furthering the understanding of the tissue distribution of PrP(Sc) in non-human primate models of CJD.

  4. Genetic polymorphism of Babesia bovis merozoite surface antigens-2 (MSA-2) isolates from bovine blood and Rhipicephalus annulatus ticks in Israel.

    Science.gov (United States)

    Molad, T; Fleiderovitz, L; Leibovich, B; Wolkomirsky, R; Erster, O; Roth, A; Mazuz, M L; Markovics, A; Shkap, V

    2014-09-15

    This study demonstrated the genetic diversity among MSA-2c, MSA-2a1 and MSA-2b proteins of Babesia bovis isolates obtained from bovine blood and Rhipicephalus annulatus tick samples. The least identities that were observed among the deduced amino acid sequences of MSA-2c, MSA-2a1 and MSA-2b were 55, 63, and 71%, respectively. During the study four B. bovis calves, aged about 1 month, were found to be infected with virulent field strains and developed babesiosis. Probably, the calves had received insufficient antibodies, or the antibodies raised against the vaccine strain did not cross-protect against virulent field isolates. The complete msa-2 locus from the Israeli B. bovis vaccine strain and two field isolates were characterized. Similarly to the Australian strains and isolates, the msa-2 loci of the examined Israeli strain and isolates had only two msa-2 genes - msa-2c and msa-2a/b - located between msa-2c and orfB. Several of the examined samples, contained different MSA-2 genotypes concurrently. No obvious geographical relationships among isolates from various regions of Israel were established. Moreover, in the phylogenetic analyses, the Israeli deduced MSA-2 amino acid sequences of the three examined genes were clustered together with sequences derived from other countries, proving that the msa-2 gene sequences of B. bovis shared the same genetic characters worldwide. The present study clearly showed that the MSA-2 proteins of B. bovis isolates from Israel were genetically distinct from the vaccine strains. Thus, further research will be needed in order to understand the genetic diversity mechanisms of B. bovis, and the immunological responses of the infected animals.

  5. Ultrastructural observation of human neutrophils during apoptotic cell death triggered by Entamoeba histolytica.

    Science.gov (United States)

    Sim, Seobo; Kim, Kyeong Ah; Yong, Tai-Soon; Park, Soon-Jung; Im, Kyung-il; Shin, Myeong Heon

    2004-12-01

    Neutrophils are important effector cells against protozoan extracellular parasite Entamoeba histolytica, which causes amoebic colitis and liver abscess in human beings. Apoptotic cell death of neutrophils is an important event in the resolution of inflammation and parasite's survival in vivo. This study was undertaken to investigate the ultrastructural aspects of apoptotic cells during neutrophil death triggered by Entamoeba histolytica. Isolated human neutrophils from the peripheral blood were incubated with or without live trophozoites of E. histolytica and examined by transmission electron microscopy (TEM). Neutrophils incubated with E. histolytica were observed to show apoptotic characteristics, such as compaction of the nuclear chromatin and swelling of the nuclear envelop. In contrast, neutrophils incubated in the absence of the amoeba had many protrusions of irregular cell surfaces and heterogenous nuclear chromatin. Therefore, it is suggested that Entamoeba-induced neutrophil apoptosis contribute to prevent unwanted tissue inflammation and damage in the amoeba-invaded lesions in vivo.

  6. Age associated variations in human neutrophil and sperm functioning

    Institute of Scientific and Technical Information of China (English)

    Kaveri Purandhar; Sriram Seshadri

    2013-01-01

    Objective: To determine the functional and biochemical variations in sperm and the neutrophil with the progression of age. Methods: Ninety healthy male subjects were selected in the age group 26-40 for the collection of semen and blood samples were collected. Basic semen analysis, hematogram, differential count serum analysis, seminal plasma and serum biochemistry was performed. Mitochondrial isolation from sperm and neutrophil was done to ascertain mitochondrial markers. Results: Our data shows a significant age-dependent reduction in the levels of mitochondrial Superoxide Dismutase (SOD) and Catalase (CAT) in sperm and the neutrophil. The functional attributes of sperm and neutrophil did not show any specific trend.Conclusion:The decreasing trend of the mitochondrial antioxidants enzymes in the sperm and the neutrophil is an indicative of the reduction in the functioning of sperm and the neutrophil. The antioxidants enzymes of sperm and neutrophil shows similar declining trend with the progression of age suggesting its possible role as a prognostic marker for age related deformities and even in male fertility.

  7. Characterization of neutrophil adhesion to different titanium surfaces

    Indian Academy of Sciences (India)

    V Campos; R C N Melo; L P Silva; E N Aquino; M S Castro; W Fontes

    2014-02-01

    Although titanium (Ti) is known to elicit a foreign body response when implanted into humans, Ti implant healing resembles normal wound healing in terms of inflammatory cell recruitment and inflammation persistence. Rough implant surfaces may present better conditions for protein adsorption and for the adhesion of platelets and inflammatory cells such as neutrophils. Implanted biomedical devices initially interact with coagulating blood; however, direct contact between the oxide layer of the implant and neutrophils has not been completely described. The aim of the present study is to compare the behaviours of neutrophils in direct contact with different Ti surfaces. Isolated human neutrophils were placed into contact with Ti discs, which had been rendered as `smooth' or `rough', following different surface treatments. Scanning electron microscopy and flow cytometry were used to measure cell adhesion to the surfaces and exposure of membrane proteins such as CD62L and CD11b. Topographic roughness was demonstrated as higher for SLA treated surfaces, measured by atomic force microscopy and elemental analysis was performed by energy dispersive X-ray, showing a similar composition for both surfaces. The adhesion of neutrophils to the `rough' Ti surface was initially stronger than adhesion to the `smooth' surface. The cell morphology and adhesion marker results revealed clear signs of neutrophil activation by either surface, with different neutrophil morphological characteristics being observed between the two surface types. Understanding the cellular mechanisms regulating cell–implant interactions should help researchers to improve the surface topography of biomedical implant devices.

  8. Antimicrobial peptides and nitric oxide production by neutrophils from periodontitis subjects.

    Science.gov (United States)

    Mariano, F S; Campanelli, A P; Nociti Jr, F H; Mattos-Graner, R O; Gonçalves, R B

    2012-11-01

    Neutrophils play an important role in periodontitis by producing nitric oxide (NO) and antimicrobial peptides, molecules with microbicidal activity via oxygen-dependent and -independent mechanisms, respectively. It is unknown whether variation in the production of antimicrobial peptides such as LL-37, human neutrophil peptides (HNP) 1-3, and NO by neutrophils influences the pathogenesis of periodontal diseases. We compared the production of these peptides and NO by lipopolysaccharide (LPS)-stimulated neutrophils isolated from healthy subjects and from patients with periodontitis. Peripheral blood neutrophils were cultured with or without Aggregatibacter actinomycetemcomitans-LPS (Aa-LPS), Porphyromonas gingivalis-LPS (Pg-LPS) and Escherichia coli-LPS (Ec-LPS). qRT-PCR was used to determine quantities of HNP 1-3 and LL-37 mRNA in neutrophils. Amounts of HNP 1-3 and LL-37 proteins in the cell culture supernatants were also determined by ELISA. In addition, NO levels in neutrophil culture supernatants were quantitated by the Griess reaction. Neutrophils from periodontitis patients cultured with Aa-LPS, Pg-LPS and Ec-LPS expressed higher HNP 1-3 mRNA than neutrophils from healthy subjects. LL-37 mRNA expression was higher in neutrophils from patients stimulated with Aa-LPS. Neutrophils from periodontitis patients produced significantly higher LL-37 protein levels than neutrophils from healthy subjects when stimulated with Pg-LPS and Ec-LPS, but no difference was observed in HNP 1-3 production. Neutrophils from periodontitis patients cultured or not with Pg-LPS and Ec-LPS produced significantly lower NO levels than neutrophils from healthy subjects. The significant differences in the production of LL-37 and NO between neutrophils from healthy and periodontitis subjects indicate that production of these molecules might influence individual susceptibility to important periodontal pathogens.

  9. Antimicrobial peptides and nitric oxide production by neutrophils from periodontitis subjects

    Directory of Open Access Journals (Sweden)

    F.S. Mariano

    2012-11-01

    Full Text Available Neutrophils play an important role in periodontitis by producing nitric oxide (NO and antimicrobial peptides, molecules with microbicidal activity via oxygen-dependent and -independent mechanisms, respectively. It is unknown whether variation in the production of antimicrobial peptides such as LL-37, human neutrophil peptides (HNP 1-3, and NO by neutrophils influences the pathogenesis of periodontal diseases. We compared the production of these peptides and NO by lipopolysaccharide (LPS-stimulated neutrophils isolated from healthy subjects and from patients with periodontitis. Peripheral blood neutrophils were cultured with or without Aggregatibacter actinomycetemcomitans-LPS (Aa-LPS, Porphyromonas gingivalis-LPS (Pg-LPS and Escherichia coli-LPS (Ec-LPS. qRT-PCR was used to determine quantities of HNP 1-3 and LL-37 mRNA in neutrophils. Amounts of HNP 1-3 and LL-37 proteins in the cell culture supernatants were also determined by ELISA. In addition, NO levels in neutrophil culture supernatants were quantitated by the Griess reaction. Neutrophils from periodontitis patients cultured with Aa-LPS, Pg-LPS and Ec-LPS expressed higher HNP 1-3 mRNA than neutrophils from healthy subjects. LL-37 mRNA expression was higher in neutrophils from patients stimulated with Aa-LPS. Neutrophils from periodontitis patients produced significantly higher LL-37 protein levels than neutrophils from healthy subjects when stimulated with Pg-LPS and Ec-LPS, but no difference was observed in HNP 1-3 production. Neutrophils from periodontitis patients cultured or not with Pg-LPS and Ec-LPS produced significantly lower NO levels than neutrophils from healthy subjects. The significant differences in the production of LL-37 and NO between neutrophils from healthy and periodontitis subjects indicate that production of these molecules might influence individual susceptibility to important periodontal pathogens.

  10. Blood

    Science.gov (United States)

    ... Also, blood is either Rh-positive or Rh-negative. So if you have type A blood, it's either A positive or A negative. Which type you are is important if you need a blood transfusion. And your Rh factor could be important ...

  11. Oxidative burst of neutrophils against melanoma B16-F10.

    Science.gov (United States)

    Zivkovic, Morana; Poljak-Blazi, Marija; Zarkovic, Kamelija; Mihaljevic, Danijela; Schaur, Rudolf Joerg; Zarkovic, Neven

    2007-02-08

    Intensive oxidative burst was determined by chemiluminescence of peripheral blood neutrophils of mice that were intramuscularly injected with melanoma B16-F10 and/or subcutaneously with Sephadex G-200. The neutrophils from papula developed at the site of Sephadex injection were cytotoxic for the B16-F10 cells in vitro. However, survival of Sephadex injected tumour-bearing mice was lower than of control animals bearing B16-F10, while their tumours grew faster and were less necrotic. Thus, it is likely that injection of Sephadex distracted the neutrophils from the tumour allowing faster progression of the tumour, indicating that neutrophils may have an important role in the host defence against malignant cells in the early stage of tumour development.

  12. Neutrophil extracellular traps in dermatology: Caught in the NET.

    Science.gov (United States)

    Hoffmann, Jochen H O; Enk, Alexander H

    2016-10-01

    Neutrophil, or polymorphonuclear granulocytes (PMN) constitute the most abundant type of leucocytes in peripheral human blood. One of the major advances in the last decade was the discovery of neutrophil extracellular trap (NET) formation: a process by which neutrophils externalize web-like chromatin strands decorated with antimicrobial peptides. These structures were soon implicated in immune defense and auto-immunity alike and now link neutrophils to the pathogenesis of a variety of diseases of dermatological relevance. Currently, NET formation is mainly subdivided into suicidal and vital NETosis. Controversy exists regarding the capacity of NETs to kill pathogens, and little is known about the way NETs are formed in vivo. Here, we discuss the current terminology, methods for NET quantification, pathways leading to NET formation, and the role of NETs in systemic and cutaneous immune defense and auto-immunity, with a focus on psoriasis and systemic lupus erythematosus.

  13. Neutrophil myeloperoxidase destruction by ultraviolet irradiation

    Energy Technology Data Exchange (ETDEWEB)

    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. Concentrations of procalcitonin and C-reactive protein, white blood cell count, and the immature-to-total neutrophil ratio in the blood of neonates with nosocomial infections: Gram-negative bacilli vs coagulase-negative staphylococci

    OpenAIRE

    Kordekag, A.

    2011-01-01

    Abstract This study was undertaken to determine whether concentrations of procalcitonin in the blood of neonates with nosocomial infections depend on the type of pathogen. Qualification for the study group was based on the clinical signs of infection. We found that infections with Gram-positive (chiefly coagulase-negative staphylococci) and Gram-negative bacteria are accompanied by elevated concentrations of procalcitonin. In the case of Gram-positive bacteria, other laboratory sig...

  15. LukMF′ is the major secreted leukocidin of bovine Staphylococcus aureus and is produced in vivo during bovine mastitis

    NARCIS (Netherlands)

    Vrieling, Manouk; Boerhout, Eveline M.; Wigcheren, Van Glenn F.; Koymans, Kirsten J.; Mols-Vorstermans, Tanja G.; Haas, de Carla J.C.; Aerts, Piet C.; Daemen, Ineke J.J.M.; Kessel, Van Kok P.M.; Koets, Ad P.; Rutten, Victor P.M.G.; Nuijten, Piet J.M.; Strijp, van Jos A.G.; Benedictus, Lindert

    2016-01-01

    Staphylococcus aureus is a major human and animal pathogen and a common cause of mastitis in cattle. S. aureus secretes several leukocidins that target bovine neutrophils, crucial effector cells in the defence against bacterial pathogens. In this study, we investigated the role of staphylococcal leu

  16. Effects of whole-body X-radiation on the neutrophils of the peripheral blood of the primate Cebus apella (weeping capuchin); Acao dos raios X corpo total sobre os neutrofilos do sangue periferico em primata Cebus apella (macaco prego)

    Energy Technology Data Exchange (ETDEWEB)

    Egami, Mizue Imoto; Silva, Maria Regina Regis; Paiva, Elias Rodrigues de; Segreto, Camilo [Escola Paulista de Medicina, Sao Paulo, SP (Brazil); Diniz, Lilian Munao [Fundacao Parque Zoologico de Sao Paulo, SP (Brazil)

    1994-12-31

    The effects of ionizing radiation on the neutrophils of Primate Cebus apella were studied after whole-body x-radiation to a single exposure of 25.8 m C/kg (100 R 0), Wright`s stained preparations showed changes in the nucleus and the cytoplasm of neutrophils at 1,3 and 6 days after irradiation. during this period of time, the cytochemical methods revealed a considerable variation in the pattern of distribution of glycogen, sudanophilic and myeloperoxidase positive granules. Under these same experimental conditions the number of caryoschizes increased on the first and third day. On the ninetieth day post exposure, the morphological and cytochemical appearances of neutrophils as well as the number of caryoschized were similar to the controls. (author) 14 refs., 5 figs., 1 tab.

  17. Measurement of neutrophil membrane CD64 and HLA-Dr in a patient with abdominal sepsis.

    NARCIS (Netherlands)

    Meer, W. van der; Scott, C.S.; Verlaat, C.; Klein Gunnewiek, J.M.T.; Warris, A.

    2006-01-01

    A patient with abdominal sepsis, had both intra and extracellular bacteria in a blood smear, and high levels of neutrophil membrane CD64 and HLA-Dr. Intracellular bacteria are only observed in the terminal phase of a sepsis. Our patient recovered, suggesting that a high expression of neutrophil CD64

  18. Neutrophil-to-lymphocyte ratio: A novel and simple prognostic marker for infective endocarditis.

    Science.gov (United States)

    Bozbay, Mehmet; Uyarel, Huseyin

    2015-08-01

    Infective endocarditis is a life-threatining infectious disease characterized by high morbidity and mortality. Leukocytes play a main role in infectious diseases. Neutrophils and lymphocytes are subgroup of leukocytes, and they are routinely measured as a part of automated complete blood count test. The neutrophil-to-lymphocyte ratio is an independent predictor of unfavorable clinical outcomes in infectious and cardiovascular diseases.

  19. Neutrophil, TLR2, and TLR4 expression in newborns at risk of sepsis

    Directory of Open Access Journals (Sweden)

    Ari Yunanto

    2013-06-01

    Conclusion There are significant increases in neutrophils, as well as neutrophil expression of TLR2 and TLR4 in the saliva and blood from newborns with sepsis risk factors compared to those of healthy newborns. [Paediatr Indones. 2013;53:132-7.

  20. Update on Comparative Studies for Diagnosis of Bovine Tuberculosis Using The Interferon Gamma (IFN-gamma) Assay with Tuberculins or Alternative Antigens for Whole Blood Stimulation

    Science.gov (United States)

    Bovine Tuberculosis is a respiratory disease caused by Mycobacterium bovis (M. bovis). It is a major infectious disease found worldwide in domestic animals, particularly cattle, as well as in certain wildlife populations. Although the disease has been effectively eliminated in many countries and reg...

  1. Interaction of bovine serum albumin and human blood plasma with PEO-tethered surfaces : influence of PEO chain length, grafting density and temperature

    NARCIS (Netherlands)

    Norde, W.; Gage, R.A.

    2004-01-01

    Solid surfaces are modified by grafting poly(ethylene oxide), PEO, to influence their interaction with indwelling particles, in particular molecules of bovine serum albumin and human plasma proteins. As a rule, the grafted PEO layers suppress protein adsorption. The suppression is most effective whe

  2. Chemokine receptor Ccr1 drives neutrophil-mediated kidney immunopathology and mortality in invasive candidiasis.

    Science.gov (United States)

    Lionakis, Michail S; Fischer, Brett G; Lim, Jean K; Swamydas, Muthulekha; Wan, Wuzhou; Richard Lee, Chyi-Chia; Cohen, Jeffrey I; Scheinberg, Phillip; Gao, Ji-Liang; Murphy, Philip M

    2012-01-01

    Invasive candidiasis is the 4(th) leading cause of nosocomial bloodstream infection in the US with mortality that exceeds 40% despite administration of antifungal therapy; neutropenia is a major risk factor for poor outcome after invasive candidiasis. In a fatal mouse model of invasive candidiasis that mimics human bloodstream-derived invasive candidiasis, the most highly infected organ is the kidney and neutrophils are the major cellular mediators of host defense; however, factors regulating neutrophil recruitment have not been previously defined. Here we show that mice lacking chemokine receptor Ccr1, which is widely expressed on leukocytes, had selectively impaired accumulation of neutrophils in the kidney limited to the late phase of the time course of the model; surprisingly, this was associated with improved renal function and survival without affecting tissue fungal burden. Consistent with this, neutrophils from wild-type mice in blood and kidney switched from Ccr1(lo) to Ccr1(high) at late time-points post-infection, when Ccr1 ligands were produced at high levels in the kidney and were chemotactic for kidney neutrophils ex vivo. Further, when a 1∶1 mixture of Ccr1(+/+) and Ccr1(-/-) donor neutrophils was adoptively transferred intravenously into Candida-infected Ccr1(+/+) recipient mice, neutrophil trafficking into the kidney was significantly skewed toward Ccr1(+/+) cells. Thus, neutrophil Ccr1 amplifies late renal immunopathology and increases mortality in invasive candidiasis by mediating excessive recruitment of neutrophils from the blood to the target organ.

  3. Endomorphins delay constitutive apoptosis and alter the innate host defense functions of neutrophils.

    Science.gov (United States)

    Azuma, Yasutaka; Ohura, Kiyoshi; Wang, Pao-Li; Shinohara, Mitsuko

    2002-04-01

    Recent studies have shown that opioid peptides are released from cells of the immune system during inflammation and stress, and are associated with altered immune responses. Moreover, concentrations of opioid peptides are increased in peripheral blood and at the sites of inflammatory reactions. The aim of this study was to evaluate immunological effects of opioid peptides endomorphins 1 and 2 on constitutive apoptosis, superoxide anion production, hydrogen peroxide production, adhesion, phagocytosis, and chemotaxis of neutrophils. Neutrophils were isolated by peritoneal lavage from rats. Endomorphins 1 and 2 significantly delayed constitutive neutrophil apoptosis. The delay of neutrophil apoptosis was markedly attenuated by LY294002, a phosphoinositide 3-kinase inhibitor. Moreover, endomorphins 1 and 2 activated the phosphoinositide 3-kinase pathway as determined by phosphorylation of BAD. In contrast, endomorphins 1 and 2 blocked the production of superoxide anion and hydrogen peroxide by PMA-stimulated neutrophils. In addition, endomorphins 1 and 2 inhibited neutrophil adhesion to fibronectin. Moreover, endomorphins 1 and 2 potentiated neutrophil chemotaxis toward zymosan-activated serum and IL-8, respectively. However, endomorphins 1 and 2 did not alter phagocytosis of Escherichia coli by neutrophils. These results suggest that endomorphins 1 and 2 may act to delay neutrophil apoptosis and alter the natural immune functions of neutrophils.

  4. Suppressed neutrophil function in children with acute lymphoblastic leukemia.

    Science.gov (United States)

    Tanaka, Fumiko; Goto, Hiroaki; Yokosuka, Tomoko; Yanagimachi, Masakatsu; Kajiwara, Ryosuke; Naruto, Takuya; Nishimaki, Shigeru; Yokota, Shumpei

    2009-10-01

    Infection is a major obstacle in cancer chemotherapy. Neutropenia has been considered to be the most important risk factor for severe infection; however, other factors, such as impaired neutrophil function, may be involved in susceptibility to infection in patients undergoing chemotherapy. In this study, we analyzed neutrophil function in children with acute lymphoblastic leukemia (ALL). Whole blood samples were obtained from 16 children with ALL at diagnosis, after induction chemotherapy, and after consolidation chemotherapy. Oxidative burst and phagocytic activity of neutrophils were analyzed by flow cytometry. Oxidative burst of neutrophils was impaired in ALL patients. The percentage of neutrophils with normal oxidative burst after PMA stimulation was 59.0 +/- 13.2 or 70.0 +/- 21.0% at diagnosis or after induction chemotherapy, respectively, which was significantly lower compared with 93.8 +/- 6.1% in healthy control subjects (P = 0.00004, or 0.002, respectively); however, this value was normal after consolidation chemotherapy. No significant differences were noted in phagocytic activity in children with ALL compared with healthy control subjects. Impaired oxidative burst of neutrophils may be one risk factor for infections in children with ALL, especially in the initial periods of treatment.

  5. Neutrophil activator of matrix metalloproteinase-2 (NAM).

    Science.gov (United States)

    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.

  6. Human Neutrophils Kill Bacillus anthracis.

    Directory of Open Access Journals (Sweden)

    2005-11-01

    Full Text Available Bacillus anthracis spores cause natural infections and are used as biological weapons. Inhalation infection with B. anthracis, the etiological agent of anthrax, is almost always lethal, yet cutaneous infections usually remain localized and resolve spontaneously. Neutrophils are typically recruited to cutaneous but seldom to other forms of anthrax infections, raising the possibility that neutrophils kill B. anthracis. In this study we infected human neutrophils with either spores or vegetative bacteria of a wild-type strain, or strains, expressing only one of the two major virulence factors. The human neutrophils engulfed B. anthracis spores, which germinated intracellularly and were then efficiently killed. Interestingly, neutrophil killing was independent of reactive oxygen species production. We fractionated a human neutrophil granule extract by high-performance liquid chromatography and identified alpha-defensins as the component responsible for B. anthracis killing. These data suggest that the timely recruitment of neutrophils can control cutaneous infections and possibly other forms of B. anthracis infections, and that alpha-defensins play an important role in the potent anti-B. anthracis activity of neutrophils.

  7. Human neutrophils kill Bacillus anthracis.

    Directory of Open Access Journals (Sweden)

    Anne Mayer-Scholl

    2005-11-01

    Full Text Available Bacillus anthracis spores cause natural infections and are used as biological weapons. Inhalation infection with B. anthracis, the etiological agent of anthrax, is almost always lethal, yet cutaneous infections usually remain localized and resolve spontaneously. Neutrophils are typically recruited to cutaneous but seldom to other forms of anthrax infections, raising the possibility that neutrophils kill B. anthracis. In this study we infected human neutrophils with either spores or vegetative bacteria of a wild-type strain, or strains, expressing only one of the two major virulence factors. The human neutrophils engulfed B. anthracis spores, which germinated intracellularly and were then efficiently killed. Interestingly, neutrophil killing was independent of reactive oxygen species production. We fractionated a human neutrophil granule extract by high-performance liquid chromatography and identified alpha-defensins as the component responsible for B. anthracis killing. These data suggest that the timely recruitment of neutrophils can control cutaneous infections and possibly other forms of B. anthracis infections, and that alpha-defensins play an important role in the potent anti-B. anthracis activity of neutrophils.

  8. Human neutrophils kill Bacillus anthracis.

    Science.gov (United States)

    Mayer-Scholl, Anne; Hurwitz, Robert; Brinkmann, Volker; Schmid, Monika; Jungblut, Peter; Weinrauch, Yvette; Zychlinsky, Arturo

    2005-11-01

    Bacillus anthracis spores cause natural infections and are used as biological weapons. Inhalation infection with B. anthracis, the etiological agent of anthrax, is almost always lethal, yet cutaneous infections usually remain localized and resolve spontaneously. Neutrophils are typically recruited to cutaneous but seldom to other forms of anthrax infections, raising the possibility that neutrophils kill B. anthracis. In this study we infected human neutrophils with either spores or vegetative bacteria of a wild-type strain, or strains, expressing only one of the two major virulence factors. The human neutrophils engulfed B. anthracis spores, which germinated intracellularly and were then efficiently killed. Interestingly, neutrophil killing was independent of reactive oxygen species production. We fractionated a human neutrophil granule extract by high-performance liquid chromatography and identified alpha-defensins as the component responsible for B. anthracis killing. These data suggest that the timely recruitment of neutrophils can control cutaneous infections and possibly other forms of B. anthracis infections, and that alpha-defensins play an important role in the potent anti-B. anthracis activity of neutrophils.

  9. Immunosenescence of Polymorphonuclear Neutrophils

    Directory of Open Access Journals (Sweden)

    Inga Wessels

    2010-01-01

    Full Text Available All immune cells are affected by aging, contributing to the high susceptibility to infections and increased mortality observed in the elderly. The effect of aging on cells of the adaptive immune system is well documented. In contrast, knowledge concerning age-related defects of polymorphonuclear neutrophils (PMN is limited. During the past decade, it has become evident that in addition to their traditional role as phagocytes, neutrophils are able to secrete a wide array of immunomodulating molecules. Their importance is underlined by the finding that genetic defects that lead to neutropenia increase susceptibility to infections. Whereas there is consistence about the constant circulating number of PMN throughout aging, the abilities of tissue infiltration, phagocytosis, and oxidative burst of PMN from aged donors are discussed controversially. Furthermore, there are numerous discrepancies between in vivo and in vitro results, as well as between results for murine and human PMN. Most of the reported functional changes can be explained by defective signaling pathways, but further research is required to get a detailed insight into the underlying molecular mechanisms. This could form the basis for drug development in order to prevent or treat age-related diseases, and thus to unburden the public health systems.

  10. Occupational Neutrophilic Asthma

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    Richard Leigh

    1999-01-01

    Full Text Available Occupational asthma is typically associated with an eosinophilic bronchitis. The case of a 41-year-old woman who developed symptoms of asthma after occupational exposure to metal working fluids is reported. The diagnosis of asthma was confirmed by an forced expiratory volume in 1 s (FEV1 of 1.7 (59% predicted, with 11% reversibility after inhaled bronchodilator and a provocation concentration of methacholine to cause a fall in FEV1 of 20% (PC20 of 0.4 mg/mL. Induced sputum examination showed a marked neutrophilia. Over the next six months, serial sputum analyses confirmed the presence of a marked sterile neutrophilic bronchitis during periods of occupational exposure to metal working fluids, which resolved when the patient was away from work and recurred when she returned to work. The sputum findings were mirrored by corresponding changes in spirometry and PC20 methacholine. The findings indicate the occurrence of occupational asthma associated with an intense, sterile neutrophilic bronchitis after exposure to metal working fluids.

  11. Human neutrophil antimicrobial activity.

    Science.gov (United States)

    Thomas, E L; Lehrer, R I; Rest, R F

    1988-01-01

    Polymorphonuclear neutrophilic leukocytes (PMNs) take up opsonized microorganisms into phagosomes that fuse with secretory granules in the PMN cytoplasm to form phagolysosomes. Killing and digestion of microorganisms take place within phagolysosomes. Antimicrobial activities in phagolysosomes are divided into two classes. Oxygen (O2)-dependent mechanisms are expressed when PMNs undergo the "respiratory burst." An NADPH oxidase in the phagolysosome membrane is activated and reduces O2 to superoxide (O2-). O2 reduction is the first step in a series of reactions that produce toxic oxidants. For example, .O2- dismutases to hydrogen peroxide (H2O2), and the azurophil granule enzyme myeloperoxidase catalyzes the oxidation of Cl- by H2O2 to yield hypochlorous acid (HOCl). The reaction of HOCl with ammonia and amines modulates the toxicity of this oxidant. O2-independent antimicrobial mechanisms include the activities of lysosomal proteases, other hydrolytic enzymes, and proteins and peptides that bind to microorganisms and disrupt essential processes or structural components. For example, the bactericidal/permeability-increasing protein, cathepsin G, and the defensins are released into phagolysosomes from the azurophil granules. Proposed mechanisms of action of neutrophil antimicrobial agents, their range of microbial targets, and their possible interactions within phagolysosomes are discussed.

  12. Neutrophil function and metabolism in individuals with diabetes mellitus

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    T.C. Alba-Loureiro

    2007-08-01

    Full Text Available Neutrophils act as first-line-of-defense cells and the reduction of their functional activity contributes to the high susceptibilityto and severity of infections in diabetes mellitus. Clinical investigations in diabetic patients and experimental studies in diabetic rats and mice clearly demonstrated consistent defects of neutrophil chemotactic, phagocytic and microbicidal activities. Other alterations that have been reported to occur during inflammation in diabetes mellitus include: decreased microvascular responses to inflammatory mediators such as histamine and bradykinin, reduced protein leakage and edema formation, reduced mast cell degranulation, impairment of neutrophil adhesionto the endothelium and migration to the site of inflammation, production of reactive oxygen species and reduced release of cytokines and prostaglandin by neutrophils, increased leukocyte apoptosis, and reduction in lymph node retention capacity. Since neutrophil function requires energy, metabolic changes (i.e., glycolytic and glutaminolytic pathways may be involved in the reduction of neutrophil function observed in diabetic states. Metabolic routes by which hyperglycemia is linked to neutrophil dysfunction include the advanced protein glycosylation reaction, the polyol pathway, oxygen-free radical formation, the nitric oxide-cyclic guanosine-3'-5'monophosphate pathway, and the glycolytic and glutaminolytic pathways. Lowering of blood glucose levels by insulin treatment of diabetic patients or experimental animals has been reported to have significant correlation with improvement of neutrophil functional activity. Therefore, changes might be primarily linked to a continuing insulin deficiency or to secondary hyperglycemia occurring in the diabetic individual. Accordingly, effective control with insulin treatment is likely to be relevant during infection in diabetic patients.

  13. Ranitidine improves postoperative monocyte and neutrophil function

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Nielsen, H; Jensen, S;

    1994-01-01

    BACKGROUND: The histamine H2-receptor antagonist ranitidine hydrochloride has been shown to improve trauma-, blood transfusion-, and sepsis-induced immunosuppression. OBJECTIVE: To evaluate the effect of ranitidine on postoperative impairment in monocyte and neutrophil function. METHODS: Twenty......-four patients undergoing major elective abdominal surgery were randomized to receive adjuvant treatment with ranitidine hydrochloride (100 mg) administered twice a day intravenously from skin incision for 4 days, followed by oral ranitidine hydrochloride (150 mg) administered twice a day for 5 days (n = 11...

  14. MPLA inhibits release of cytotoxic mediators from human neutrophils while preserving efficient bacterial killing.

    Science.gov (United States)

    Ruchaud-Sparagano, Marie-Hélène; Mills, Ross; Scott, Jonathan; Simpson, A John

    2014-10-01

    Monophosphoryl lipid A (MPLA) is a lipopolysaccharides (LPS) derivative associated with neutrophil-dependent anti-inflammatory outcomes in animal models of sepsis. Little is known about the effect of MPLA on neutrophil function. This study sought to test the hypothesis that MPLA would reduce release of cytotoxic mediators from neutrophils without impairing bacterial clearance. Neutrophils were isolated from whole blood of healthy volunteers. The effects of MPLA and LPS on autologous serum-opsonised Pseudomonas aeruginosa killing by neutrophils and phagocytosis of autologous serum-opsonised zymosan were examined. Neutrophil oxidative burst, chemotaxis, enzyme and cytokine release as well as Toll-like receptor 4 (TLR4) expression were assessed following exposure to LPS or MPLA. LPS, but not MPLA, induced significant release of superoxide and myeloperoxidase from neutrophils. However, MPLA did not impair neutrophil capacity to ingest microbial particles and kill P. aeruginosa efficiently. MPLA was directly chemotactic for neutrophils, involving TLR4, p38 mitogen-activated protein kinase and tyrosine and alkaline phosphatases. LPS, but not MPLA, impaired N-formyl-methionyl-leucyl phenylalanine-directed migration of neutrophils, increased surface expression of TLR4, increased interleukin-8 release and strongly activated the myeloid differentiation primary response 88 pathway. Phosphoinositide 3-kinase inhibition significantly augmented IL-8 release from MPLA-treated neutrophils. The addition of MPLA to LPS-preincubated neutrophils led to a significant reduction in LPS-mediated superoxide release and TLR4 surface expression. Collectively, these findings suggest that MPLA directs efficient chemotaxis and bacterial killing in human neutrophils without inducing extracellular release of cytotoxic mediators and suggest that MPLA warrants further attention as a potential therapeutic in human sepsis.

  15. Activated Neutrophils Are Associated with Pediatric Cerebral Malaria Vasculopathy in Malawian Children

    Science.gov (United States)

    Feintuch, Catherine Manix; Saidi, Alex; Seydel, Karl; Chen, Grace; Goldman-Yassen, Adam; Mita-Mendoza, Neida K.; Kim, Ryung S.; Frenette, Paul S.; Taylor, Terrie

    2016-01-01

    ABSTRACT Most patients with cerebral malaria (CM) sustain cerebral microvascular sequestration of Plasmodium falciparum-infected red blood cells (iRBCs). Although many young children are infected with P. falciparum, CM remains a rare outcome; thus, we hypothesized that specific host conditions facilitate iRBC cerebral sequestration. To identify these host factors, we compared the peripheral whole-blood transcriptomes of Malawian children with iRBC cerebral sequestration, identified as malarial-retinopathy-positive CM (Ret+CM), to the transcriptomes of children with CM and no cerebral iRBC sequestration, defined as malarial-retinopathy-negative CM (Ret-CM). Ret+CM was associated with upregulation of 103 gene set pathways, including cytokine, blood coagulation, and extracellular matrix (ECM) pathways (P < 0.01; false-discovery rate [FDR] of <0.05). Neutrophil transcripts were the most highly upregulated individual transcripts in Ret+CM patients. Activated neutrophils can modulate diverse host processes, including the ECM, inflammation, and platelet biology to potentially facilitate parasite sequestration. Therefore, we compared plasma neutrophil proteins and neutrophil chemotaxis between Ret+CM and Ret-CM patients. Plasma levels of human neutrophil elastase, myeloperoxidase, and proteinase 3, but not lactoferrin or lipocalin, were elevated in Ret+CM patients, and neutrophil chemotaxis was impaired, possibly related to increased plasma heme. Neutrophils were rarely seen in CM brain microvasculature autopsy samples, and no neutrophil extracellular traps were found, suggesting that a putative neutrophil effect on endothelial cell biology results from neutrophil soluble factors rather than direct neutrophil cellular tissue effects. Meanwhile, children with Ret-CM had lower levels of inflammation, higher levels of alpha interferon, and upregulation of Toll-like receptor pathways and other host transcriptional pathways, which may represent responses that do not favor

  16. Light scattering by neutrophils: Model, simulation, and experiment

    NARCIS (Netherlands)

    Orlova, D.Y.; Yurkin, M.A.; Hoekstra, A.G.; Maltsev, V.P.

    2008-01-01

    We studied the elastic light-scattering properties of human blood neutrophils, both experimentally and theoretically. The experimental study was performed with a scanning flow cytometer measuring the light-scattering patterns (LSPs) of individual cells over an angular range of 5-60 deg. We determine

  17. Hidden truth of circulating neutrophils (polymorphonuclear neutrophil function in periodontally healthy smoker subjects

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    Chitra Agarwal

    2016-01-01

    Full Text Available Context: Tobacco smoking is considered to be a major risk factor associated with periodontal disease. Smoking exerts a major effect on the protective elements of the immune response, resulting in an increase in the extent and severity of periodontal destruction. Aims: The aim of the present study was to assess viability and phagocytic function of neutrophils in circulating blood of the smokers and nonsmokers who are periodontally healthy. Settings and Design: Two hundred subjects in the mean range of 20–30 years of age were included in the study population. It was a retrospective study carried out for 6 months. Materials and Methods: Two hundred subjects were divided into four groups: 50 nonsmokers, 50 light smokers (15 cigarettes/day. Full mouth plaque index, sulcus bleeding index, and probing depths were measured. Percentage viability of circulating neutrophils and average number of phagocytosed Candida albicans were recorded. Statistical Analysis Used: Means and standard deviations were calculated from data obtained within the groups. Comparison between the smokers and nonsmokers was performed by Kruskal–Wallis ANOVA analysis. Comparison between smoker groups was performed using Mann–Whitney–Wilcoxon test. Results: Percentage viability of neutrophils was significantly less in heavy smokers (66.9 ± 4.0, moderate (76.6 ± 4.2, light smokers (83.1 ± 2.5 as compared to nonsmokers (92.3 ± 2.6 (P < 0.01. The ability of neutrophils to phagocytose, i.e., mean particle number was significantly less in light smokers (3.5 ± 0.5, moderate smokers (2.3 ± 0.5, and heavy smokers (1.4 ± 0.5 compared to nonsmokers (4.9 ± 0.7 (P < 0.01 with evidence of dose-response effect. Conclusions: Smoking significantly affects neutrophils viability and phagocytic function in periodontally healthy population.

  18. Neutrophil Functions in Periodontal Homeostasis

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    Ricarda Cortés-Vieyra

    2016-01-01

    Full Text Available 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.

  19. Neutrophilic dermatosis of dorsal hands

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    S Kaur

    2015-01-01

    Full Text Available Sweet′s syndrome is characterized by erythematous tender nodules and plaques over face and extremities. Fever, leukocytosis with neutrophilia, and a neutrophilic infiltrate in the dermis are characteristic features. Neutrophilic dermatosis of dorsal hands is a rare localized variant of Sweet′s syndrome occurring predominantly over dorsa of hands. Various degrees of vascular damage may be observed on histopathology of these lesions. Both Sweet′s syndrome and its dorsal hand variant have been reported in association with malignancies, inflammatory bowel diseases, and drugs. We report a patient with neutrophilic dermatoses of dorsal hands associated with erythema nodosum. He showed an excellent response to corticosteroids and dapsone.

  20. A unique protein profile of peripheral neutrophils from COPD patients does not reflect cytokine-induced protein profiles of neutrophils in vitro

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    Koenderman Leo

    2011-09-01

    Full Text Available Abstract Background Inflammation, both local and systemic, is a hallmark of chronic obstructive pulmonary disease (COPD. Inflammatory mediators such as TNFα and GM-CSF are secreted by lung epithelium, alveolar macrophages and other inflammatory cells and are thought to be important contributors in the pathogenesis of COPD. Indeed, neutrophils are activated by these cytokines and these cells are one of the major inflammatory cell types recruited to the pulmonary compartment of COPD patients. Furthermore, these inflammatory mediators are found in the peripheral blood of COPD patients and, therefore, we hypothesized that TNFα/GM-CSF-induced protein profiles can be found in peripheral neutrophils of COPD patients. Methods Using fluorescence 2-dimensional difference gel electrophoresis we investigated differentially regulated proteins in peripheral neutrophils from COPD patients and healthy age-matched control subjects. Furthermore, protein profiles from COPD patients were compared with those of neutrophils of healthy age-matched controls that were stimulated with TNFα and/or GM-CSF in vitro. Protein gels were compared using DeCyder 7.0 software. Results We identified 7 significantly regulated protein spots between peripheral neutrophils from COPD patients and age-matched healthy control subjects. Stimulation of peripheral neutrophils with TNFα, GM-CSF or TNFα + GM-CSF in vitro resulted in 13, 20 and 22 regulated protein spots, respectively. However, these cytokine-induced protein differences did not correspond with the protein differences found in neutrophils from COPD patients. Conclusion These results show that neutrophils from COPD patients have a unique protein profile compared to neutrophils from healthy age-matched controls. Furthermore, the neutrophil profiles of COPD patients do not reflect putative dominant signals induced by TNFα, GM-CSF or their combination. Our results indicate that systemic neutrophil responses in COPD patients

  1. Anti-neutrophil cytoplasmic antibodies stimulate release of neutrophil microparticles.

    LENUS (Irish Health Repository)

    Hong, Ying

    2012-01-01

    The mechanisms by which anti-neutrophil cytoplasmic antibodies (ANCAs) may contribute to the pathogenesis of ANCA-associated vasculitis are not well understood. In this study, both polyclonal ANCAs isolated from patients and chimeric proteinase 3-ANCA induced the release of neutrophil microparticles from primed neutrophils. These microparticles expressed a variety of markers, including the ANCA autoantigens proteinase 3 and myeloperoxidase. They bound endothelial cells via a CD18-mediated mechanism and induced an increase in endothelial intercellular adhesion molecule-1 expression, production of endothelial reactive oxygen species, and release of endothelial IL-6 and IL-8. Removal of the neutrophil microparticles by filtration or inhibition of reactive oxygen species production with antioxidants abolished microparticle-mediated endothelial activation. In addition, these microparticles promoted the generation of thrombin. In vivo, we detected more neutrophil microparticles in the plasma of children with ANCA-associated vasculitis compared with that in healthy controls or those with inactive vasculitis. Taken together, these results support a role for neutrophil microparticles in the pathogenesis of ANCA-associated vasculitis, potentially providing a target for future therapeutics.

  2. High Throughput Measurement of Extracellular DNA Release and Quantitative NET Formation in Human Neutrophils In Vitro.

    Science.gov (United States)

    Sil, Payel; Yoo, Dae-Goon; Floyd, Madison; Gingerich, Aaron; Rada, Balazs

    2016-06-18

    Neutrophil granulocytes are the most abundant leukocytes in the human blood. Neutrophils are the first to arrive at the site of infection. Neutrophils developed several antimicrobial mechanisms including phagocytosis, degranulation and formation of neutrophil extracellular traps (NETs). NETs consist of a DNA scaffold decorated with histones and several granule markers including myeloperoxidase (MPO) and human neutrophil elastase (HNE). NET release is an active process involving characteristic morphological changes of neutrophils leading to expulsion of their DNA into the extracellular space. NETs are essential to fight microbes, but uncontrolled release of NETs has been associated with several disorders. To learn more about the clinical relevance and the mechanism of NET formation, there is a need to have reliable tools capable of NET quantitation. Here three methods are presented that can assess NET release from human neutrophils in vitro. The first one is a high throughput assay to measure extracellular DNA release from human neutrophils using a membrane impermeable DNA-binding dye. In addition, two other methods are described capable of quantitating NET formation by measuring levels of NET-specific MPO-DNA and HNE-DNA complexes. These microplate-based methods in combination provide great tools to efficiently study the mechanism and regulation of NET formation of human neutrophils.

  3. Neutrophils are dispensable in the modulation of T cell immunity against cutaneous HSV-1 infection

    Science.gov (United States)

    Hor, Jyh Liang; Heath, William R.; Mueller, Scott N.

    2017-01-01

    Neutrophils rapidly infiltrate sites of inflammation during peripheral infection or tissue injury. In addition to their well described roles as pro-inflammatory phagocytes responsible for pathogen clearance, recent studies have demonstrated a broader functional repertoire including mediating crosstalk between innate and adaptive arms of the immune system. Specifically, neutrophils have been proposed to mediate antigen transport to lymph nodes (LN) to modulate T cell priming and to influence T cell migration to infected tissues. Using a mouse model of cutaneous herpes simplex virus type 1 (HSV-1) infection we explored potential contributions of neutrophils toward anti-viral immunity. While a transient, early influx of neutrophils was triggered by dermal scarification, we did not detect migration of neutrophils from the skin to LN. Furthermore, despite recruitment of neutrophils into LN from the blood, priming and expansion of CD4+ and CD8+ T cells was unaffected following neutrophil depletion. Finally, we found that neutrophils were dispensable for the migration of effector T cells into infected skin. Our study suggests that the immunomodulatory roles of neutrophils toward adaptive immunity may be context-dependent, and are likely determined by the type of pathogen and anatomical site of infection. PMID:28112242

  4. Ageing exacerbates damage of systemic and salivary neutrophils from patients presenting Candida-related denture stomatitis

    Directory of Open Access Journals (Sweden)

    Porto Vinicius

    2009-03-01

    Full Text Available Abstract Background Ageing leads to a decline in the function of the immune system, increasing the body's susceptibility to infections through the impairment of T-cells, macrophages, neutrophils and dendritic cells Denture stomatitis is a primary oral disease affecting elderly denture wearers. The major etiologic factor involved in this pathology is the infection by Candida albicans, an opportunistic pathogen that causes local and disseminated diseases in immunosuppressed humans. Neutrophils play a critical role in the immune response against C. albicans and are continually present in the salivary fluid and in the blood. The aim of this study was to determine ageing-related changes in salivary and blood neutrophils and their potential implications in Candida-related denture stomatitis. Results Our results showed a lower number of neutrophils in the saliva from patients presenting Candida-related denture stomatitis in comparison to their matched controls. Furthermore, fewer neutrophils were isolated from the saliva of aged control individuals in comparison to matched younger subjects. CXCR1, CD62L and CD11b expression were significantly greater on systemic neutrophils from younger control individuals. Elderly individuals showed more apoptotic salivary neutrophils and lower GM-CSF levels than younger ones, regardless of the occurrence of Candida infection. On the other hand, CXCL-8 concentrations were higher in the saliva from elderly individuals. Besides, TNF-α was detected at elevated levels in the saliva from infected elderly subjects. Salivary neutrophils from elderly and young patients presented impaired phagocytic activity against C. albicans. However, just systemic neutrophils from elderly showed decreased phagocytosis when compared to the younger ones, regardless of the occurrence of infection. In addition, neutrophils from aged individuals and young patients presented low fungicidal activity. Conclusion The data suggests that the Candida

  5. Effects of Aggregatibacter actinomycetemcomitans leukotoxin on neutrophil migration and extracellular trap formation

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    Josefine Hirschfeld

    2016-11-01

    Full Text Available Background: Aggressive periodontitis is associated with the presence of Aggregatibacter actinomycetemcomitans, a leukotoxin (Ltx-producing periodontal pathogen. Ltx has the ability to lyse white blood cells including neutrophils. Objectives: This study was aimed at investigating the interactions between neutrophils and Ltx with regard to the chemotactic properties of Ltx and the release of neutrophil extracellular traps (NETs. Methods: Neutrophils from healthy blood donors were isolated and incubated for 30 min and 3 h with increasing concentrations of Ltx (1, 10, and 100 ng/mL as well as with A. actinomycetemcomitans strains (NCTC 9710 and HK 1651 producing different levels of Ltx. Formation of NETs and cell lysis were assessed by microscopy, fluorescence-based assays, and measurement of released lactate dehydrogenase. Neutrophil migration in response to different Ltx gradients was monitored by real-time video microscopy, and image analysis was performed using ImageJ software. Results: Although Ltx (10 and 100 ng/mL and the leukotoxic A. actinomycetemcomitans strain HK 1651 lysed some neutrophils, other cells were still capable of performing NETosis in a concentration-dependent manner. Low doses of Ltx and the weakly leukotoxic strain NCTC 9710 did not lead to neutrophil lysis, but did induce some NETosis. Furthermore, all three concentrations of Ltx enhanced random neutrophil movement; however, low directional accuracy was observed compared with the positive control (fMLP. Conclusions: The results indicate that Ltx acts both as a neutrophil activator and also causes cell death. In addition, Ltx directly induces NETosis in neutrophils prior to cell lysis. In future studies, the underlying pathways involved in Ltx-meditated neutrophil activation and NETosis need to be investigated further.

  6. Effects of Aggregatibacter actinomycetemcomitans leukotoxin on neutrophil migration and extracellular trap formation

    Science.gov (United States)

    Hirschfeld, Josefine; Roberts, Helen M.; Chapple, Iain L. C.; Parčina, Marijo; Jepsen, Søren; Johansson, Anders; Claesson, Rolf

    2016-01-01

    Background Aggressive periodontitis is associated with the presence of Aggregatibacter actinomycetemcomitans, a leukotoxin (Ltx)-producing periodontal pathogen. Ltx has the ability to lyse white blood cells including neutrophils. Objectives This study was aimed at investigating the interactions between neutrophils and Ltx with regard to the chemotactic properties of Ltx and the release of neutrophil extracellular traps (NETs). Methods Neutrophils from healthy blood donors were isolated and incubated for 30 min and 3 h with increasing concentrations of Ltx (1, 10, and 100 ng/mL) as well as with A. actinomycetemcomitans strains (NCTC 9710 and HK 1651) producing different levels of Ltx. Formation of NETs and cell lysis were assessed by microscopy, fluorescence-based assays, and measurement of released lactate dehydrogenase. Neutrophil migration in response to different Ltx gradients was monitored by real-time video microscopy, and image analysis was performed using ImageJ software. Results Although Ltx (10 and 100 ng/mL) and the leukotoxic A. actinomycetemcomitans strain HK 1651 lysed some neutrophils, other cells were still capable of performing NETosis in a concentration-dependent manner. Low doses of Ltx and the weakly leukotoxic strain NCTC 9710 did not lead to neutrophil lysis, but did induce some NETosis. Furthermore, all three concentrations of Ltx enhanced random neutrophil movement; however, low directional accuracy was observed compared with the positive control (fMLP). Conclusions The results indicate that Ltx acts both as a neutrophil activator and also causes cell death. In addition, Ltx directly induces NETosis in neutrophils prior to cell lysis. In future studies, the underlying pathways involved in Ltx-meditated neutrophil activation and NETosis need to be investigated further. PMID:27834173

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

    Science.gov (United States)

    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

  8. Light scattering by neutrophils: model, simulation, and experiment.

    Science.gov (United States)

    Orlova, Darya Yu; Yurkin, Maxim A; Hoekstra, Alfons G; Maltsev, Valeri P

    2008-01-01

    We studied the elastic light-scattering properties of human blood neutrophils, both experimentally and theoretically. The experimental study was performed with a scanning flow cytometer measuring the light-scattering patterns (LSPs) of individual cells over an angular range of 5-60 deg. We determined the absolute differential light-scattering cross sections of neutrophils. We also proposed an optical model for a neutrophil as a sphere filled by small spheres and prolate spheroids that correspond to granules and segmented nucleus, respectively. This model was used in simulations of LSPs using the discrete dipole approximation and different compositions of internal organelles. A comparison of experimentally measured and simulated LSPs gives a good qualitative agreement in LSP shape and quantitative agreement in overall magnitude of the differential light-scattering cross section.

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

    Science.gov (United States)

    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.

  10. Inhibition of neutrophil activity improves cardiac function after cardiopulmonary bypass

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    Grünwald Frank

    2007-10-01

    Full Text Available Abstract Background The arterial in line application of the leukocyte inhibition module (LIM in the cardiopulmonary bypass (CPB limits overshooting leukocyte activity during cardiac surgery. We studied in a porcine model whether LIM may have beneficial effects on cardiac function after CPB. Methods German landrace pigs underwent CPB (60 min myocardial ischemia; 30 min reperfusion without (group I; n = 6 or with LIM (group II; n = 6. The cardiac indices (CI and cardiac function were analyzed pre and post CPB with a Swan-Ganz catheter and the cardiac function analyzer. Neutrophil labeling with technetium, scintigraphy, and histological analyses were done to track activated neutrophils within the organs. Results LIM prevented CPB-associated increase of neutrophil counts in peripheral blood. In group I, the CI significantly declined post CPB (post: 3.26 ± 0.31; pre: 4.05 ± 0.45 l/min/m2; p 2; p = 0.23. Post CPB, the intergroup difference showed significantly higher CI values in the LIM group (p Conclusion Our data provides strong evidence that LIM improves perioperative hemodynamics and cardiac function after CPB by limiting neutrophil activity and inducing accelerated sequestration of neutrophils in the spleen.

  11. Neutrophil biology and the next generation of myeloid growth factors.

    Science.gov (United States)

    Dale, David C

    2009-01-01

    Neutrophils are the body's critical phagocytic cells for defense against bacterial and fungal infections; bone marrow must produce approximately 10 x 10(9) neutrophils/kg/d to maintain normal blood neutrophil counts. Production of neutrophils depends on myeloid growth factors, particularly granulocyte colony-stimulating factor (G-CSF). After the original phase of development, researchers modified these growth factors to increase their size and delay renal clearance, increase their biologic potency, and create unique molecules for business purposes. Pegylated G-CSF is a successful product of these efforts. Researchers have also tried to identify small molecules to serve as oral agents that mimic the parent molecules, but these programs have been less successful. In 2006, the European Medicines Agency established guidelines for the introduction of new biologic medicinal products claimed to be similar to reference products that had previously been granted marketing authorization in the European community, called bio-similars. Globally, new and copied versions of G-CSF and other myeloid growth factors are now appearing. Some properties of the myeloid growth factors are similar to other agents, offering opportunities for the development of alternative drugs and treatments. For example, recent research shows that hematopoietic progenitor cells can be mobilized with a chemokine receptor antagonist, chemotherapy, G-CSF, and granulocyte macrophage colony-stimulating factor. Advances in neutrophil biology coupled with better understanding and development of myeloid growth factors offer great promise for improving the care of patients with cancer and many other disorders.

  12. AUTOINFLAMMATORY PUSTULAR NEUTROPHILIC DISEASES

    Science.gov (United States)

    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. Low White Blood Cell Count

    Science.gov (United States)

    Symptoms Low white blood cell count By Mayo Clinic Staff A low white blood cell count (leukopenia) is a decrease in disease-fighting cells ( ... a decrease in a certain type of white blood cell (neutrophil). The definition of low white blood cell ...

  14. Artificial blood.

    OpenAIRE

    1983-01-01

    #Blood substitutes have been developed for almost a century. The various type of artificial blood was continuously available on the market. The theme of this report is to identify the best substitute in emergency situation for some patients and science students. The definition of best is given; thus, as the vital part of the report, the comparison between them is described and discussed. Modified hemoglobin, bovine-based hemoglobin and PFCs are three basic types. In terms of the perfor...

  15. 77 FR 29914 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products

    Science.gov (United States)

    2012-05-21

    ... RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products AGENCY... live bovines and products derived from bovines with regard to bovine spongiform encephalopathy. This... with regard to bovine spongiform encephalopathy. Comments on the proposed rule were required to......

  16. Proinflammatory mediators stimulate neutrophil-directed angiogenesis.

    LENUS (Irish Health Repository)

    McCourt, M

    2012-02-03

    BACKGROUND: Vascular endothelial growth factor (VEGF; vascular permeability factor) is one of the most potent proangiogenic cytokines, and it plays a central role in mediating the process of angiogenesis or new blood vessel formation. Neutrophils (PMNs) recently have been shown to produce VEGF. HYPOTHESIS: The acute inflammatory response is a potent stimulus for PMN-directed angiogenesis. METHODS: Neutrophils were isolated from healthy volunteers and stimulated with lipopolysaccharide (LPS), tumor necrosis factor alpha (TNF-alpha), interleukin 6 (IL-6), and anti-human Fas monoclonal antibody. Culture supernatants were assayed for VEGF using enzyme-linked immunosorbent assays. Culture supernatants from LPS- and TNF-alpha-stimulated PMNs were then added to human umbilical vein endothelial cells and human microvessel endothelial cells and assessed for endothelial cell proliferation using 5-bromodeoxyuridine labeling. Tubule formation was also assessed on MATRIGEL basement membrane matrix. Neutrophils were lysed to measure total VEGF release, and VEGF expression was detected using Western blot analysis. RESULTS: Lipopolysaccharide and TNF-alpha stimulation resulted in significantly increased release of PMN VEGF (532+\\/-49 and 484+\\/-80 pg\\/mL, respectively; for all, presented as mean +\\/- SEM) compared with control experiments (32+\\/-4 pg\\/mL). Interleukin 6 and Fas had no effect. Culture supernatants from LPS- and TNF-alpha-stimulated PMNs also resulted in significant increases (P<.005) in macrovascular and microvascular endothelial cell proliferation and tubule formation. Adding anti-human VEGF-neutralizing polyclonal antibody to stimulated PMN supernatant inhibited these effects. Total VEGF release following cell lysis and Western blot analysis suggests that the VEGF is released from an intracellular store. CONCLUSION: Activated human PMNs are directly angiogenic by releasing VEGF, and this has important implications for inflammation, capillary leak syndrome

  17. Avaliação do efeito da hipotermia por crioimersão corporal, nos neutrófilos e linfócitos sanguíneos de ratos submetidos ao exercício físico agudo Evaluation of the effect of hypothermia by cold water immersion on blood neutrophils and lymphocytes of rats submitted to acute exercise

    Directory of Open Access Journals (Sweden)

    José A. Bachur

    2008-12-01

    Full Text Available O estresse sistêmico induzido pelo exercício libera substâncias bioativas determinantes da mobilização neutrofílica. A crioterapia diminui a reação inflamatória e atenua a elevação da perfusão sanguínea induzida pelo exercício. O objetivo deste trabalho foi analisar a influência da hipotermia decorrente da crioimersão corporal (CIC imediata ao esforço físico agudo nas concentrações neutrofílicas e linfocíticas no sangue. Os ratos do grupo controle (AI foram mantidos em repouso enquanto os do grupo AII foram submetidos ao protocolo de CIC a 10ºC por 10 minutos. Enquanto os animais dos grupos BI, BII, BIII e BIV realizaram o esforço físico agudo (EFA em água a 31ºC durante 100 minutos com sobrecarga corpórea de 5% do peso corporal, os dos grupos CI, CII, CIII e CIV foram submetidos ao EFA seguido imediatamente de CIC. Nos grupos B e C, os animais foram sacrificados nos períodos de 06 (I, 12 (II, 24 (III e 48 (IV horas posteriores ao EFA. Através da microscopia óptica realizou-se a contagem dos neutrófilos e linfócitos. Utilizou-se do Teste T Student para análise estatística considerando-se nível de significância p Systemic stress induced by exercise increases bioactive substances in plasma which leads to neutrophilic mobilization. Cryotherapy causes a decrease in the inflammatory reaction and attenuates high blood perfusion after exercise. The objective of this work was to analyze the influence of cold water immersion (CWI after acute exercise on neutrophil and lymphocyte mobilization. A control group of rats (AI was kept at rest and a second group (AII was submitted to CWI at 10º C for 10 minutes. The animals of Groups BI, BII, BIII and BIV were submitted to acute exercise which consisted in swimming in water at 31º C for 100 minutes with a load equivalent to 5% of the body weight. Groups CI, CII, CIII and CIV were submitted to CWI immediately after acute exercise. The animals were sacrificed at 6 (I, 12 (II

  18. Relationship of neutrophils lymphocyte ratio and red blood cell distribution width with idiopathic facial palsy%中性粒细胞淋巴细胞比及红细胞分布宽度与特发性面神经麻痹的关系

    Institute of Scientific and Technical Information of China (English)

    舒湘宁; 马跃文

    2016-01-01

    背景:红细胞分布宽度与C-反应蛋白一样,是机体炎症水平的一个标志,红细胞表面可能存在多种炎症因子受体,推测红细胞参与了炎症过程,炎症导致红细胞分布宽度增加。目的:分析中性粒细胞淋巴细胞比和红细胞分布宽度与特发性面神经麻痹的发病机制及发病时疾病的严重程度之间关系。方法:特发性面神经麻痹患者30例(实验组),使用HB(House and Brackmann Facial Grading System)评分和SB(Sunnybrook System)评分评估患者病情严重程度,将实验组按SB评分分为一般瘫痪组(30-100分)和严重瘫痪组(0-29分);按照HB评估结果分为轻度(Ⅱ/Ⅲ)、中度(Ⅳ)、重度瘫痪组(Ⅴ/Ⅵ)。同时选健康者32人做对照。对比各组间中性粒细胞淋巴细胞比、红细胞分布宽度指标的差异;分析SB评分和HB评估结果与中性粒细胞淋巴细胞比、红细胞分布宽度之间的相关性。结果与结论:实验组中性粒细胞淋巴细胞比值明显高于对照组(P<0.01);红细胞分布宽度在实验组不同疾病程度组间均存在显著差异,且与SB评分呈显著负相关(P <0.01),与HB评估等级呈显著正相关(P<0.01)。结果说明,实验组中性粒细胞淋巴细胞比值显著高于对照组,红细胞分布宽度与疾病严重程度呈正相关,提示特发性面神经麻痹发病机制与炎症相关,红细胞分布宽度指标可能对特发性面神经麻痹发病时疾病严重程度评估有参考意义。%BACKGROUND:Similarly with C-reactive protein, red blood cel distribution width can reflect the inflammatory process. Red blood cels involved in inflammatory process leads to the increase in red blood cel distribution width due to various inflammation factor receptors existingon the surface of red blood cel s. OBJECTIVE:To analyze the relationship between the neutrophils lymphocyte ratio, red blood cel distribution width and the pathogenesis andseverity

  19. Ability of Staphylococcus aureus coagulase genotypes to resist neutrophil bactericidal activity and phagocytosis

    DEFF Research Database (Denmark)

    Aarestrup, Frank Møller; Scott, N. L.; Sordillo, L. M.

    1994-01-01

    genotype. The interaction between bacteria and neutrophils was measured by phagocytosis and bactericidal effect. The average percent killing of bacteria was lowest (40.0%) with strains belonging to the most common genotype, medium (50%) with strains belonging to the intermediate type, and highest (64......; rare type, 10.5/cell). These findings suggest that one of the reasons for the variation in prevalence of different genotypes of S. aureus in the mammary gland is due to the superior ability of some types to resist phagocytosis and/or killing by bovine neutrophils......This study investigated the functional capabilities of neutrophils against different Staphylococcus aureus genotypes isolated from cows with mastitis. Six strains of S. aureus were chosen for use in the study, two with a common genotype, two with an intermediate genotype, and two with a rare...

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

    Directory of Open Access Journals (Sweden)

    Francesca S. M. Tang

    2015-01-01

    Full Text Available 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=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.

  1. Endogenous TNFα orchestrates the trafficking of neutrophils into and within lymphatic vessels during acute inflammation

    Science.gov (United States)

    Arokiasamy, Samantha; Zakian, Christian; Dilliway, Jessica; Wang, Wen; Nourshargh, Sussan; Voisin, Mathieu-Benoit

    2017-01-01

    Neutrophils are recognised to play a pivotal role at the interface between innate and acquired immunities following their recruitment to inflamed tissues and lymphoid organs. While neutrophil trafficking through blood vessels has been extensively studied, the molecular mechanisms regulating their migration into the lymphatic system are still poorly understood. Here, we have analysed neutrophil-lymphatic vessel interactions in real time and in vivo using intravital confocal microscopy applied to inflamed cremaster muscles. We show that antigen sensitisation of the tissues induces a rapid but transient entry of tissue-infiltrated neutrophils into lymphatic vessels and subsequent crawling along the luminal side of the lymphatic endothelium. Interestingly, using mice deficient in both TNF receptors p55 and p75, chimeric animals and anti-TNFα antibody blockade we demonstrate that tissue-release of TNFα governs both neutrophil migration through the lymphatic endothelium and luminal crawling. Mechanistically, we show that TNFα primes directly the neutrophils to enter the lymphatic vessels in a strictly CCR7-dependent manner; and induces ICAM-1 up-regulation on lymphatic vessels, allowing neutrophils to crawl along the lumen of the lymphatic endothelium in an ICAM-1/MAC-1-dependent manner. Collectively, our findings demonstrate a new role for TNFα as a key regulator of neutrophil trafficking into and within lymphatic system in vivo. PMID:28287124

  2. GPR55 regulates cannabinoid 2 receptor-mediated responses in human neutrophils

    Institute of Scientific and Technical Information of China (English)

    Nariman A B Balenga; Maria Waldhoer; Elma Aflaki; Julia Kargl; Wolfgang Platzer; Ralf Schr(o)der; Stefanie Bl(a)ttermann; Evi Kostenis; Andrew J Brown; Akos Heinemann

    2011-01-01

    The directional migration of neutrophils towards inflammatory mediators,such as chemokines and cannabinoids,occurs via the activation of seven transmembrane G protein coupled receptors (7TM/GPCRs) and is a highly organized process.A crucial role for controlling neutrophil migration has been ascribed to the cannabinoid CB2 receptor (CB2R),but additional modulatory sites distinct from CB2R have recently been suggested to impact CB2R-mediated effector functions in neutrophils.Here,we provide evidence that the recently de-orphanized 7TM/GPCR GPR55potently modulates CB2R-mediated responses.We show that GPR55 is expressed in human blood neutrophils and its activation augments the migratory response towards the CB2R agonist 2-arachidonoylglycerol (2-AG),while inhibiting neutrophil degranulation and reactive oxygen species (ROS) production.Using HEK293 and HL60 cell lines,along with primary neutrophils,we show that GPR55 and CB2R interfere with each other's signaling pathways at the level of small GTPases,such as Rac2 and Cdc42.This ultimately leads to cellular polarization and efficient migration as well as abrogation of degranulation and ROS formation in neutrophils.Therefore,GPR55 limits the tissueinjuring inflammatory responses mediated by CB2R,while it synergizes with CB2R in recruiting neutrophils to sites of inflammation.

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

    Energy Technology Data Exchange (ETDEWEB)

    Williams, C. David; Bajt, Mary Lynn [Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Sharpe, Matthew R. [Department of Internal Medicine, University of Kansas Hospital, Kansas City, KS (United States); McGill, Mitchell R. [Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Farhood, Anwar [Department of Pathology, St. David' s North Austin Medical Center, Austin, TX 78756 (United States); Jaeschke, Hartmut, E-mail: hjaeschke@kumc.edu [Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States)

    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

  4. A possible role for neutrophils in allergic rhinitis revealed after cellular subclassification

    Science.gov (United States)

    Arebro, Julia; Ekstedt, Sandra; Hjalmarsson, Eric; Winqvist, Ola; Kumlien Georén, Susanna; Cardell, Lars-Olaf

    2017-01-01

    A re-examination of former concepts is required to meet today’s medical challenges in allergic rhinitis. Previously, neutrophils have been treated as a relatively homogenous cell population found in the nose both when the patient is suffering at the height of the allergic season as well as when the patient report no symptoms. However, new data indicates that neutrophils can be divided into different subsets with diverse roles in inflammation. We showed increased levels of neutrophils in peripheral blood, nasal biopsies and nasal lavage fluid (NAL) from allergic patients during the pollen season compared to healthy controls. A closer examination revealed that the activated subset of neutrophils, CD16high CD62Ldim, outweighed the normal form CD16high CD62Lhigh in nasal tissue among these patients. This skewed distribution was not seen in controls. The normal subset prevailed in peripheral blood from patients as well as controls, whereas CD16high CD62Ldim and CD16dim CD62Ldim subsets, the latter considered “end state” neutrophils before apoptosis, were elevated in NAL. Functional in vitro experiments revealed that activated neutrophils exhibit a T cell priming capacity and an ability to enhance eosinophil migration. Activated neutrophils may thus contribute to allergic inflammation seen in allergic rhinitis by priming T cells and attracting eosinophils. PMID:28272395

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

    Science.gov (United States)

    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 genes p53 (P 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 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.

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

    Science.gov (United States)

    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.

  7. Alpha-1-antitrypsin is produced by human neutrophil granulocytes and their precursors and liberated during granule exocytosis

    DEFF Research Database (Denmark)

    Clemmensen, Stine N; Jacobsen, Lars C; Rørvig, Sara;

    2011-01-01

    . Neutrophils from patients with A1AT-deficiency carrying the (PI)ZZ mutation in the A1AT gene appeared structurally and functionally normal, but A1AT produced in leukocytes of these patients lacked the ability to bind proteases efficiently. We conclude that A1AT generation and release from neutrophils add......Alpha-1-antitrypsin (A1AT) is an important inhibitor of neutrophil proteases including elastase, cathepsin G, and proteinase 3. Transcription profiling data suggest that A1AT is expressed by human neutrophil granulocytes during all developmental stages. A1AT has hitherto only been found associated......1AT is produced at all stages of myeloid maturation in the bone marrow. The production increases as neutrophils enter circulation and increases further upon migration to tissues as observed in skin windows and when blood neutrophils are incubated with granulocyte colony-stimulating factor...

  8. General versus regional anaesthesia for cataract surgery: effects on neutrophil apoptosis and the postoperative pro-inflammatory state.

    LENUS (Irish Health Repository)

    Goto, Y

    2012-02-03

    At clinically relevant concentrations, volatile anaesthetic agents influence neutrophil function. Our hypothesis was that sevoflurane would inhibit neutrophil apoptosis and consequently influence the postoperative pro-inflammatory state. In order to identify selectively the effect of the anaesthetic agent sevoflurane, we studied patients undergoing minimally stimulating (cataract) surgery randomly allocated to receive either sevoflurane (n = 11) or local anaesthesia (n = 12). Venous blood samples were taken immediately prior to anaesthesia and at 1, 8 and 24 h thereafter. The rate of neutrophil apoptosis, plasma concentration of cytokines and differential white cell count were measured. The rates of neutrophil apoptosis and plasma concentrations of IL-1beta, TNF-alpha and IL-8 at each time point were similar in the two groups. IL-6 concentrations increased significantly and to a similar extent compared to preanaesthetic levels at 8 and 24 h. This study demonstrates that sevoflurane does not influence the rate of neutrophil apoptosis, cytokine concentrations and neutrophil count following cataract surgery.

  9. Cytobiologic and clinical aspects in a patient with chronic neutrophilic leukemia after thorotrast exposure

    Energy Technology Data Exchange (ETDEWEB)

    Boggs, D.R.; Kaplan, S.S.

    1986-11-01

    A patient with fairly typical chronic neutrophilic leukemia, as represented by some two dozen such reported cases, had been given Thorotrast more than 20 years before. Typical myeloblastic crisis developed with remarkable terminal leukocytosis. Mature blood neutrophils had normal function with respect to phagocytosis, bacterial killing, metabolic activation, and chemotactic response. The number of cells producing colonies of neutrophils and monocytes in in vitro semisolid cultures was normal in the blood and increased in marrow. Colony size was smaller than is usually observed in normal patients or in typical patients with chronic myeloid leukemia. Termination in blast crisis, also seen in a few other patients with chronic neutrophilic leukemia, indicates that this is indeed a form of leukemia and not a leukemoid reaction of obscure cause. The differential diagnosis of extreme neutrophilia is discussed.

  10. Spontaneous neutrophil activation in HTLV-1 infected patients

    Directory of Open Access Journals (Sweden)

    Jaqueline B. Guerreiro

    2005-12-01

    Full Text Available Human T cell lymphotropic Virus type-1 (HTLV-1 induces lymphocyte activation and proliferation, but little is known about the innate immune response due to HTLV-1 infection. We evaluated the percentage of neutrophils that metabolize Nitroblue tetrazolium (NBT to formazan in HTLV-1 infected subjects and the association between neutrophil activation and IFN-gamma and TNF-alpha levels. Blood was collected from 35 HTLV-1 carriers, from 8 patients with HAM/TSP (HTLV-1- associated myelopathy; 22 healthy individuals were evaluated for spontaneous and lipopolysaccharide (LPS-stimulated neutrophil activity (reduction of NBT to formazan. The production of IFN-gamma and TNF-alpha by unstimulated mononuclear cells was determined by ELISA. Spontaneous NBT levels, as well as spontaneous IFN-gamma and TNF-alpha production, were significantly higher (p<0.001 in HTLV-1 infected subjects than in healthy individuals. A trend towards a positive correlation was noted, with increasing percentage of NBT positive neutrophils and levels of IFN-gamma. The high IFN-gamma producing HTLV-1 patient group had significantly greater NBT than healthy controls, 43±24% and 17±4.8% respectively (p< 0.001, while no significant difference was observed between healthy controls and the low IFN-gamma-producing HTLV-1 patient group (30±20%. Spontaneous neutrophil activation is another marker of immune perturbation resulting from HTLV-1 infection. In vivo activation of neutrophils observed in HTLV-1 infected subjects is likely to be the same process that causes spontaneous IFN-gamma production, or it may partially result from direct IFN-gamma stimulation.

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

    Directory of Open Access Journals (Sweden)

    Serena Zilio

    2016-09-01

    Full Text Available 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.

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

    Science.gov (United States)

    Zilio, Serena; Serafini, Paolo

    2016-09-09

    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.

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

    Science.gov (United States)

    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

  14. Neutrophil Function in 8 Cases of Papillon-Lefevre Syndrome

    Directory of Open Access Journals (Sweden)

    M.Lotfazar

    2004-03-01

    Full Text Available Statement of Problem: Papillon Lefevre syndrome (PLS is a rate autosomal recessive disorder, which is characterized by palmar- plantar hyperkeratosis and rapid periodontal destruction of primary and permanent dentitions.Purpose: The aim of the present study was to evaluate the peripheral blood neutrophil function including random locomotion, chemotaxis and oxidative mechanism of killing in a group of patients with PLS.Materials and Methods: Peripheral blood was obtained from 8 PLS patients and 92 healthy control subjects. PMN mobility was measured by a modification of the micromethod of Addison and Babbage. Latex-Stimulated NBT reduction test described by Park et al was followed. Data were analyzed by Mann Whitney U test.Results: The chemotactic activity in the PLS group was significantly lower than control group (89.5±21.6 vs 113±16 mm, P0.05.Conclusion: The present study indicated an impaired neutrophil chemotaxis in PLS patients.

  15. Characterization of neutrophil subsets in healthy human pregnancies.

    Directory of Open Access Journals (Sweden)

    Aloysius Ssemaganda

    Full Text Available We have previously shown that in successful pregnancies increased arginase activity is a mechanism that contributes to the suppression of the maternal immune system. We identified the main type of arginase-expressing cells as a population of activated low-density granulocytes (LDGs in peripheral blood mononuclear cells and in term placentae. In the present study, we analyzed the phenotype of LDGs and compared it to the phenotype of normal density granulocytes (NDGs in maternal peripheral blood, placental biopsies and cord blood. Our data reveal that only LDGs but no NDGs could be detected in placental biopsies. Phenotypically, NDGs and LDGs from both maternal and cord blood expressed different levels of maturation, activation and degranulation markers. NDGs from the maternal and cord blood were phenotypically similar, while maternal, cord and placental LDGs showed different expression levels of CD66b. LDGs present in cord blood expressed higher levels of arginase compared to maternal and placental LDGs. In summary, our results show that in maternal and cord blood, two phenotypically different populations of neutrophils can be identified, whereas in term placentae, only activated neutrophils are present.

  16. Ensemble models of neutrophil trafficking in severe sepsis.

    Directory of Open Access Journals (Sweden)

    Sang Ok Song

    Full Text Available A hallmark of severe sepsis is systemic inflammation which activates leukocytes and can result in their misdirection. This leads to both impaired migration to the locus of infection and increased infiltration into healthy tissues. In order to better understand the pathophysiologic mechanisms involved, we developed a coarse-grained phenomenological model of the acute inflammatory response in CLP (cecal ligation and puncture-induced sepsis in rats. This model incorporates distinct neutrophil kinetic responses to the inflammatory stimulus and the dynamic interactions between components of a compartmentalized inflammatory response. Ensembles of model parameter sets consistent with experimental observations were statistically generated using a Markov-Chain Monte Carlo sampling. Prediction uncertainty in the model states was quantified over the resulting ensemble parameter sets. Forward simulation of the parameter ensembles successfully captured experimental features and predicted that systemically activated circulating neutrophils display impaired migration to the tissue and neutrophil sequestration in the lung, consequently contributing to tissue damage and mortality. Principal component and multiple regression analyses of the parameter ensembles estimated from survivor and non-survivor cohorts provide insight into pathologic mechanisms dictating outcome in sepsis. Furthermore, the model was extended to incorporate hypothetical mechanisms by which immune modulation using extracorporeal blood purification results in improved outcome in septic rats. Simulations identified a sub-population (about 18% of the treated population that benefited from blood purification. Survivors displayed enhanced neutrophil migration to tissue and reduced sequestration of lung neutrophils, contributing to improved outcome. The model ensemble presented herein provides a platform for generating and testing hypotheses in silico, as well as motivating further experimental

  17. Standardization and validation of simple PCR, duplex PCR and RAPD in comparison to blood smear examination for diagnosing bovine tropical theileriosis.

    Science.gov (United States)

    Sudan, Vikrant; Shanker, Daya; Jaiswal, Amit; Singh, Amit; Pandey, Vijay

    2017-03-01

    Bovine Tropical Theileriosis (BTT) is an important vector-borne protozoan disease that imposing serious constraints on the health and productivity of domestic cattle. It is matter of common fact that following recovery from primary infection, cattle become persistent carriers and act as reservoirs of infection thereby, playing a critical role in disease epidemiology. The present study describes the comparative diagnostic efficiency of simplex PCR, duplex PCR and RAPD assays for detection of Theileria annulata in cattle. An optimized simple PCR and duplex PCR assay were established using TAMS F/R as primer sets encoding for 721 bp amplicon alongside a RAPD with arbitrary primer coding for 963 bp product of T. annulata. The simple PCR and duplex PCR detected pathogen with almost same level of sensitivity, irrespective of whether its DNA was amplified in isolation or together with DNA of another pathogen without nonspecific amplifications. RAPD failed to give comparable results and suffered from limitations of sensitivity as well as specificity. The developed assays may be seen as a good tool for epidemiological studies aiming at assessing the burden of chronic infections and improving control of the associated diseases in endemic regions.

  18. Somatotropina bovina recombinante (rBST nos aspectos hematológicos e metabólitos do sangue de novilhas (½ Nelore x ½ Red Angus em confinamento Recombinant bovine somatotropin (rBST on hematologic aspects and metabolites of heifers (½ Nellore x ½ Red Angus blood, in feedlot

    Directory of Open Access Journals (Sweden)

    Ivanor Nunes do Prado

    2003-04-01

    Full Text Available O objetivo deste trabalho foi avaliar o efeito da somatotropina bovina recombinante (rBST - análogo do BST, obtido comercialmente pela técnica do DNA recombinante, sobre os aspectos hematológicos (hematócrito, eritrócitos, hemoglobina, leucócitos, neutrófilos, eosinófilos, linfócitos e monócitos e metabólitos (glicose, insulina, IGF-I, triglicérides, colesterol total e uréia do sangue de novilhas confinadas. Foram utilizadas 24 novilhas mestiças (½ Nelore x ½ Red Angus, com aproximadamente 18 meses de idade e peso médio 255 kg. Os animais foram alimentados com uma dieta contendo silagem de milho como volumoso e polpa de citrus peletizada e farelo de soja, como concentrado, durante 84 dias. Essa dieta foi utilizada para os três tratamentos, que se diferenciaram pela aplicação de 250 mg de rBST, por via subcutânea, na fossa ísqueo-retal, onde: 1 controle (aplicação de dois mL de solução salina; 2 dose única e 3 uma dose a cada 14 dias. Os animais foram distribuídos em delineamento inteiramente casualizado, com três tratamentos e oito repetições. Coleta de sangue foi realizada no início do experimento e nos dias 28, 56 e 84 do experimento, para as determinações dos aspectos hematológicos e metabólitos. Não houve influência da aplicação de rBST sobre os aspectos hematológicos do sangue no início e final do experimento. Todavia, independentemente do tratamento, o dia de coleta teve um efeito linear positivo sobre os níveis de hematócrito, linear negativo sobre os níveis de glicose e triglicérides e quadrático positivo sobre os níveis de IGF-I e uréia. Ainda, não foi observado efeito do dia de coleta sobre os níveis de insulina e colesterol total.This work was carried out to evaluate the recombinant bovine somatotropin effect (rBST - BST analogous, commercially obtained by recombinant DNA technique, on hematological aspects (hematocrit, erythrocyte, hemoglobin, leukocyte, neutrophil, eosinophil

  19. NSP4, an elastase-related protease in human neutrophils with arginine specificity

    Science.gov (United States)

    Perera, Natascha C.; Schilling, Oliver; Kittel, Heike; Back, Walter; Kremmer, Elisabeth; Jenne, Dieter E.

    2012-01-01

    Neutrophil serine proteases (NSPs) in cytoplasmic granules of neutrophils are regarded as important antimicrobial defense weapons after engulfment and exposure of pathogens to the content of primary granules. Despite intensive studies on neutrophils during the last three decades, only three active serine proteases, neutrophil elastase (NE), cathepsin G (CG), and proteinase 3 (PR3) have been identified in these short-lived cells. Here, we report on the identification of a fourth serine protease (NSP4) with 39% identity to NE and PR3, but arginine specificity, yet sharing features like propeptide processing by dipeptidyl peptidase I, storage, and release as an active enzyme with the three active proteases. We established monoclonal antibodies against NSP4, excluded cross-reactivity to human granzymes, NE, CG, PR3, and azurocidin, and screened for NSP4 protein expression in various human tissues and blood leukocyte populations. Only granulocyte precursors and neutrophil populations from peripheral blood were positive. The content of NSP4 in neutrophil lysates, however, was about 20-fold lower compared with CG. Upon neutrophil activation, NSP4 was released into the supernatant. Profiling its specificity with peptide libraries from Escherichia coli revealed a preference for arginine in P1; it cleaved Tyr-Arg-Phe-Arg-AMC and Ala-Pro-Nva-thiobenzyl esters. NSP4 was inhibited by α1-proteinase inhibitor (α1–antitrypsin), C1 inhibitor, and most efficiently by antithrombin-heparin, but not by elafin, secretory leukocyte protease inhibitor, α1–antichymotrypsin, and monocyte-neutrophil elastase inhibitor. Functional specialization and preferred natural substrates of NSP4 remain to be determined to understand the biological interplay of all four NSPs during neutrophil responses. PMID:22474388

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

    Science.gov (United States)

    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.

  1. Enhancement by platelets of oxygen radical responses of human neutrophils

    Energy Technology Data Exchange (ETDEWEB)

    McCulloch, K.K.; Powell, J.; Johnson, K.J.; Ward, P.A.

    1986-03-01

    When human blood neutrophils were incubated with immune complexes (consisting of IgG antibody) in the presence of platelets, there was a 2 to 10 fold enhancement in the generation of O-/sub 2/ and H/sub 2/O/sub 2/. This enhancement phenomenon was proportional to the dose of immune complex added and the number of platelets present. The response was not agonist specific since similar enhancement also occurred with the following agonists: phorbol myristate acetate, opsonized zymosan particles and the chemotactic peptide N-formyl-met-leu-phe. The platelet related phenomenon of enhanced O-/sub 2/ generation could not be reproduced by the addition of serotonin, histamine or platelet-derived growth factor and was not affected by prior treatment of platelets with cyclooxygenase inhibitors (indomethacin, piroxicam) or lipoxygenase inhibitors (nafazatrom, BW755C or nordihydroguaiaretic acid). However, activation of platelets by thrombin caused release into the platelet supernatant fluid of a factor that, only in the presence of immune complexes, caused enhanced O-/sub 2/ responses to neutrophils. These data indicate that platelets potentiate oxygen radical responses of human neutrophils and suggest a mechanisms by which platelets may participate in tissue injury which is mediated by oxygen radical products from activated neutrophils.

  2. Transport of monocarboxylic acids at the blood-brain barrier: Studies with monolayers of primary cultured bovine brain capillary endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Terasaki, T.; Takakuwa, S.; Moritani, S.; Tsuji, A. (Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Kanazawa University (Japan))

    1991-09-01

    The kinetics and mechanism of the transport of monocarboxylic acids (MCAs) were studied by using primary cultured bovine brain capillary endothelial cells. Concentration-dependent uptake of acetic acid was observed, and the kinetic parameters were estimated as follows: the Michaelis constant, Kt, was 3.41 {plus minus} 1.87 mM, the maximum uptake rate, Jmax, was 144.7 {plus minus} 55.7 nmol/mg of protein/min and the nonsaturable first-order rate constant, Kd, was 6.66 {plus minus} 1.98 microliters/mg of protein/min. At medium pH below 7.0, the uptake rate of (3H)acetic acid increased markedly with decreasing medium pH, whereas pH-independent uptake was observed in the presence of 10 mM acetic acid. An energy requirement for (3H)acetic acid uptake was also demonstrated, because metabolic inhibitors (2,4-dinitrophenol and rotenone) reduced significantly the uptake rate (P less than .05). Carbonylcyanide-p-trifluoro-methoxyphenylhydrazone, a protonophore, inhibited significantly the uptake of (3H)acetic acid at medium pH of 5.0 and 6.0, whereas 4,4{prime}-diisothiocyanostilben-2,2{prime}-disulfonic acid did not. Several MCAs inhibited significantly the uptake rate of (3H)acetic acid, whereas di- and tricarboxylic acids did not. The uptake of (3H)acetic acid was competitively inhibited by salicylic acid, with an inhibition constant, Ki, of 3.60 mM, suggesting a common transport system between acetic acid and salicylic acid. Moreover, at the medium pH of 7.4, salicylic acid and valproic acid inhibited significantly the uptake of (3H)acetic acid, demonstrating that the transport of MCA drugs could also be ascribed to the MCA transport system at the physiologic pH.

  3. Risk factors against bovine respiratory diseade in suckling calves from Argentina

    OpenAIRE

    García-Bocanegra, Ignacio; Arenas-Montes, Antonio José; Perea-Remujo, J.A.; Arenas-Casas, Antonio; Torralbo, A.; Borge-Rodríguez, Carmen; Carbonero-Martínez, Alfonso; Maldonado García, Alfonso

    2011-01-01

    An observacional cross-sectional study was performed to determine the risk factors associated to the main viral agents of the bovine respiratory disease: bovine herpesvirus type 1 (HVB1), bovine viral diarrhoea virus (VDVB), bovine respiratory syncytial virus (VRSB) and parainfluenza 3 virus (VPI3). Blood samples from dairy calves in the provinces of Cordova and Santa Fe (Argentina) were obtained, and an epidemiological ques-tionnaire was filled. Antibodies against studied viruses were detect...

  4. The endothelin B receptor plays a crucial role for the adhesion of neutrophils to the endothelium in sickle cell disease.

    Science.gov (United States)

    Koehl, Bérengère; Nivoit, Pierre; El Nemer, Wassim; Lenoir, Olivia; Hermand, Patricia; Pereira, Catia; Brousse, Valentine; Guyonnet, Léa; Ghinatti, Giulia; Benkerrou, Malika; Colin, Yves; Le Van Kim, Caroline; Tharaux, Pierre-Louis

    2017-04-06

    Although the primary origin of sickle cell disease is a hemoglobin disorder, several cell types contribute considerably to the physiopathology of the disease. The adhesion of neutrophils to activated endothelium is critical in sickle cell disease pathophysiology and the targeting of neutrophils and their interactions with endothelium represent important opportunities for new therapeutics. We focused on endothelin-1, a mediator involved in neutrophil activation and recruitment in tissues, and we investigated the involvement of the endothelin receptors in interaction of neutrophils with endothelial cells. We used fluorescence intravital microscopy analyses of the microcirculation in sickle mice and quantitative microfluidic fluorescence microscopy of human blood. Both experiments on mouse model and patients indicate that blocking endothelin receptors, particularly ETB receptor highly influences neutrophils recruitment under inflammatory conditions in sickle cell disease. We show that human neutrophils display functional ETB receptors with calcium signaling capability leading to increased adhesion to the endothelium, through action on both endothelial cells and neutrophils. ETB intact function was also found to be required for TNFα-dependent upregulation of CD11b on neutrophils. Furthermore, we confirmed that human neutrophils synthesize endothelin-1 that may be involved in autocrine and paracrine pathophysiological actions. Thus, the endothelin-ETB axis should be considered as a cytokine-like potent pro-inflammatory pathway in sickle cell disease. Blockade of endothelin receptor, including ETB, may provide major benefits for preventing or treating vaso-occlusive crises of sickle cell patients.

  5. The effects of extracellular matrix proteins on neutrophil-endothelial interaction--a roadway to multiple therapeutic opportunities.

    Science.gov (United States)

    Padmanabhan, Jagannath; Gonzalez, Anjelica L

    2012-06-01

    Polymorphoneuclear leukocytes or neutrophils, a major component of white blood cells, contribute to the innate immune response in humans. Upon sensing changes in the microenvironment, neutrophils adhere to the vascular wall, migrate through the endothelial cell (EC)-pericyte bilayer, and subsequently through the extracellular matrix to reach the site of inflammation. These cells are capable of destroying microbes, cell debris, and foreign proteins by oxidative and non-oxidative processes. While primarily mediators of tissue homeostasis, there are an increasing number of studies indicating that neutrophil recruitment and transmigration can also lead to host-tissue injury and subsequently inflammation-related diseases. Neutrophil-induced tissue injury is highly regulated by the microenvironment of the infiltrated tissue, which includes cytokines, chemokines, and the provisional extracellular matrix, remodeled through increased vascular permeability and other cellular infiltrates. Thus, investigation of the effects of matrix proteins on neutrophil-EC interaction and neutrophil transmigration may help identify the proteins that induce pro- or anti-inflammatory responses. This area of research presents an opportunity to identify therapeutic targets in inflammation-related diseases. This review will summarize recent literature on the role of neutrophils and the effects of matrix proteins on neutrophil-EC interactions, with focus on three different disease models: 1) atherosclerosis, 2) COPD, and 3) tumor growth and progression. For each disease model, inflammatory molecules released by neutrophils, important regulatory matrix proteins, current anti-inflammatory treatments, and the scope for further research will be summarized.

  6. Effects of chronic occupational exposure to anaesthetic gases on the rate of neutrophil apoptosis among anaesthetists.

    LENUS (Irish Health Repository)

    Tyther, R

    2012-02-03

    BACKGROUND AND OBJECTIVE: Volatile anaesthetic agents are known to influence neutrophil function. The aim was to determine the effect of chronic occupational exposure to volatile anaesthetic agents on the rate of neutrophil apoptosis among anaesthetists. To test this hypothesis, we compared the rate of neutrophil apoptosis in anaesthetists who had been chronically exposed to volatile anaesthetic agents with that in unexposed volunteers. METHODS: Venous blood (20 mL) was withdrawn from 24 ASA I-II volunteers, from which neutrophils were isolated, and maintained in culture. At 1, 12 and 24 h in culture, the percentage of neutrophil apoptosis was assessed by dual staining with annexin V-FITC and propidium iodide. RESULTS: At 1 h (but not at 12 and 24 h) in culture, the rate of neutrophil apoptosis was significantly less in the anaesthetists--13.8 (12.9%) versus 34.4 (12.1%) (P = 0.001). CONCLUSIONS: Chronic occupational exposure to volatile anaesthetic agents may inhibit neutrophil apoptosis. This may have implications for anaesthetists and similarly exposed healthcare workers in terms of the adequacy of their inflammatory response.

  7. Rapid and label-free microfluidic neutrophil purification and phenotyping in diabetes mellitus

    Science.gov (United States)

    Hou, Han Wei; Petchakup, Chayakorn; Tay, Hui Min; Tam, Zhi Yang; Dalan, Rinkoo; Chew, Daniel Ek Kwang; Li, King Ho Holden; Boehm, Bernhard O.

    2016-07-01

    Advanced management of dysmetabolic syndromes such as diabetes will benefit from a timely mechanistic insight enabling personalized medicine approaches. Herein, we present a rapid microfluidic neutrophil sorting and functional phenotyping strategy for type 2 diabetes mellitus (T2DM) patients using small blood volumes (fingerprick ~100 μL). The developed inertial microfluidics technology enables single-step neutrophil isolation (>90% purity) without immuno-labeling and sorted neutrophils are used to characterize their rolling behavior on E-selectin, a critical step in leukocyte recruitment during inflammation. The integrated microfluidics testing methodology facilitates high throughput single-cell quantification of neutrophil rolling to detect subtle differences in speed distribution. Higher rolling speed was observed in T2DM patients (P < 0.01) which strongly correlated with neutrophil activation, rolling ligand P-selectin glycoprotein ligand 1 (PSGL-1) expression, as well as established cardiovascular risk factors (cholesterol, high-sensitive C-reactive protein (CRP) and HbA1c). Rolling phenotype can be modulated by common disease risk modifiers (metformin and pravastatin). Receiver operating characteristics (ROC) and principal component analysis (PCA) revealed neutrophil rolling as an important functional phenotype in T2DM diagnostics. These results suggest a new point-of-care testing methodology, and neutrophil rolling speed as a functional biomarker for rapid profiling of dysmetabolic subjects in clinical and patient-oriented settings.

  8. Endothelial paxillin and focal adhesion kinase (FAK) play a critical role in neutrophil transmigration.

    Science.gov (United States)

    Parsons, Sean A; Sharma, Ritu; Roccamatisi, Dawn L; Zhang, Hong; Petri, Björn; Kubes, Paul; Colarusso, Pina; Patel, Kamala D

    2012-02-01

    During an inflammatory response, endothelial cells undergo morphological changes to allow for the passage of neutrophils from the blood vessel to the site of injury or infection. Although endothelial cell junctions and the cytoskeleton undergo reorganization during inflammation, little is known about another class of cellular structures, the focal adhesions. In this study, we examined several focal adhesion proteins during an inflammatory response. We found that there was selective loss of paxillin and focal adhesion kinase (FAK) from focal adhesions in proximity to transmigrating neutrophils; in contrast the levels of the focal adhesion proteins β1-integrin and vinculin were unaffected. Paxillin was lost from focal adhesions during neutrophil transmigration both under static and flow conditions. Down-regulating endothelial paxillin with siRNA blocked neutrophil transmigration while having no effect on rolling or adhesion. As paxillin dynamics are regulated partly by FAK, the role of FAK in neutrophil transmigration was examined using two complementary methods. siRNA was used to down-regulate total FAK protein while dominant-negative, kinase-deficient FAK was expressed to block FAK signaling. Disruption of the FAK protein or FAK signaling decreased neutrophil transmigration. Collectively, these findings reveal a novel role for endothelial focal adhesion proteins paxillin and FAK in regulating neutrophil transmigration.

  9. Rapid and label-free microfluidic neutrophil purification and phenotyping in diabetes mellitus

    Science.gov (United States)

    Hou, Han Wei; Petchakup, Chayakorn; Tay, Hui Min; Tam, Zhi Yang; Dalan, Rinkoo; Chew, Daniel Ek Kwang; Li, King Ho Holden; Boehm, Bernhard O.

    2016-01-01

    Advanced management of dysmetabolic syndromes such as diabetes will benefit from a timely mechanistic insight enabling personalized medicine approaches. Herein, we present a rapid microfluidic neutrophil sorting and functional phenotyping strategy for type 2 diabetes mellitus (T2DM) patients using small blood volumes (fingerprick ~100 μL). The developed inertial microfluidics technology enables single-step neutrophil isolation (>90% purity) without immuno-labeling and sorted neutrophils are used to characterize their rolling behavior on E-selectin, a critical step in leukocyte recruitment during inflammation. The integrated microfluidics testing methodology facilitates high throughput single-cell quantification of neutrophil rolling to detect subtle differences in speed distribution. Higher rolling speed was observed in T2DM patients (P < 0.01) which strongly correlated with neutrophil activation, rolling ligand P-selectin glycoprotein ligand 1 (PSGL-1) expression, as well as established cardiovascular risk factors (cholesterol, high-sensitive C-reactive protein (CRP) and HbA1c). Rolling phenotype can be modulated by common disease risk modifiers (metformin and pravastatin). Receiver operating characteristics (ROC) and principal component analysis (PCA) revealed neutrophil rolling as an important functional phenotype in T2DM diagnostics. These results suggest a new point-of-care testing methodology, and neutrophil rolling speed as a functional biomarker for rapid profiling of dysmetabolic subjects in clinical and patient-oriented settings. PMID:27381673

  10. Incorporation of radio-labelled nucleic acid precursors by Theileria parva in bovine blood and salivary glands of Rhipicephalus appendiculatus ticks

    Energy Technology Data Exchange (ETDEWEB)

    Irvin, A.D.; Boarer, C.D.H.; Kurtti, T.J.; Ocama, J.G.R. (International Lab. for Research on Animal Diseases, Nairobi (Kenya))

    1981-12-01

    The uptake of radio-labelled nucleic acid precursors by blood and tick salivary gland forms of Theileria parva was studied. Piroplasms took up tritiated purines, particularly hypoxanthine, but not pyrimidines. Similar uptake was recorded by T. parva, both in tick saliva and in salivary glands maintained in vitro. Intermediate parasite stages were those most readily labelled in glands; this reflected active nucleic acid synthesis associated with rapid parasite division. Radio-labelling of T. parva in tick salivary glands could be of value in procedures used for concentrating and purifying theilerial sporozoites.

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

    Science.gov (United States)

    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

  12. Tumor-Derived CXCL1 Promotes Lung Cancer Growth via Recruitment of Tumor-Associated Neutrophils

    Directory of Open Access Journals (Sweden)

    Ming Yuan

    2016-01-01

    Full Text Available Neutrophils have a traditional role in inflammatory process and act as the first line of defense against infections. Although their contribution to tumorigenesis and progression is still controversial, accumulating evidence recently has demonstrated that tumor-associated neutrophils (TANs play a key role in multiple aspects of cancer biology. Here, we detected that chemokine CXCL1 was dramatically elevated in serum from 3LL tumor-bearing mice. In vitro, 3LL cells constitutively expressed and secreted higher level of CXCL1. Furthermore, knocking down CXCL1 expression in 3LL cells significantly hindered tumor growth by inhibiting recruitment of neutrophils from peripheral blood into tumor tissues. Additionally, tumor-infiltrated neutrophils expressed higher levels of MPO and Fas/FasL, which may be involved in TAN-mediated inhibition of CD4+ and CD8+ T cells. These results demonstrate that tumor-derived CXCL1 contributes to TANs infiltration in lung cancer which promotes tumor growth.

  13. The axonal repellent, Slit2, inhibits directional migration of circulating neutrophils.

    Science.gov (United States)

    Tole, Soumitra; Mukovozov, Ilya M; Huang, Yi-Wei; Magalhaes, Marco A O; Yan, Ming; Crow, Min Rui; Liu, Guang Ying; Sun, Chun Xiang; Durocher, Yves; Glogauer, Michael; Robinson, Lisa A

    2009-12-01

    In inflammatory diseases, circulating neutrophils are recruited to sites of injury. Attractant signals are provided by many different chemotactic molecules, such that blockade of one may not prevent neutrophil recruitment effectively. The Slit family of secreted proteins and their transmembrane receptor, Robo, repel axonal migration during CNS development. Emerging evidence shows that by inhibiting the activation of Rho-family GTPases, Slit2/Robo also inhibit migration of other cell types toward a variety of chemotactic factors in vitro and in vivo. The role of Slit2 in inflammation, however, has been largely unexplored. We isolated primary neutrophils from human peripheral blood and mouse bone marrow and detected Robo-1 expression. Using video-microscopic live cell tracking, we found that Slit2 selectively impaired directional migration but not random movement of neutrophils toward fMLP. Slit2 also inhibited neutrophil migration toward other chemoattractants, namely C5a and IL-8. Slit2 inhibited neutrophil chemotaxis by preventing chemoattractant-induced actin barbed end formation and cell polarization. Slit2 mediated these effects by suppressing inducible activation of Cdc42 and Rac2 but did not impair activation of other major kinase pathways involved in neutrophil migration. We further tested the effects of Slit2 in vivo using mouse models of peritoneal inflammation induced by sodium periodate, C5a, and MIP-2. In all instances, Slit2 reduced neutrophil recruitment effectively (PSlit2 potently inhibits chemotaxis but not random motion of circulating neutrophils and point to Slit2 as a potential new therapeutic for preventing localized inflammation.

  14. Vav1 is essential for mechanotactic crawling and migration of neutrophils out of the inflamed microvasculature.

    Science.gov (United States)

    Phillipson, Mia; Heit, Bryan; Parsons, Sean A; Petri, Björn; Mullaly, Sarah C; Colarusso, Pina; Gower, R Michael; Neely, Gregory; Simon, Scott I; Kubes, Paul

    2009-06-01

    Mac-1-dependent crawling is a new step in the leukocyte recruitment cascade that follows LFA-1-dependent adhesion and precedes emigration. Neutrophil adhesion via LFA-1 has been shown to induce cytoskeletal reorganization through Vav1-dependent signaling, and the current study investigates the role of Vav1 in the leukocyte recruitment process in vivo with particular attention to the events immediately downstream of LFA-1-dependent adhesion. Intravital and spinning-disk-confocal microscopy was used to investigate intravascular crawling in relation to endothelial junctions in vivo in wild-type and Vav1(-/-) mice. Adherent wild-type neutrophils almost immediately began crawling perpendicular to blood flow via Mac-1 until they reached an endothelial junction where they often changed direction. This pattern of perpendicular, mechanotactic crawling was recapitulated in vitro when shear was applied. In sharp contrast, the movement of Vav1(-/-) neutrophils was always in the direction of flow and appeared more passive as if the cells were dragged in the direction of flow in vivo and in vitro. More than 80% of Vav1(-/-) neutrophils moved independent of Mac-1 and could be detached with LFA-1 Abs. An inability to release the uropod was frequently noted for Vav1(-/-) neutrophils, leading to greatly elongated tails. The Vav1(-/-) neutrophils failed to stop or follow junctions and ultimately detached, leading to fewer emigrated neutrophils. The Vav1(-/-) phenotype resulted in fewer neutrophils recruited in a relevant model of infectious peritonitis. Clearly, Vav1 is critical for the complex interplay between LFA-1 and Mac-1 that underlies the programmed intravascular crawling of neutrophils.

  15. Interleukin-18 delays neutrophil apoptosis following alcohol intoxication and burn injury.

    Science.gov (United States)

    Akhtar, Suhail; Li, Xiaoling; Kovacs, Elizabeth J; Gamelli, Richard L; Choudhry, Mashkoor A

    2011-01-01

    Studies have shown that burn patients who are intoxicated at the time of injury are more susceptible to infection and have a higher incidence of mortality. A major cause of death in burn and trauma patients regardless of their alcohol (EtOH) exposure is multiple organ dysfunction, which is driven in part by the systemic inflammatory response and activated neutrophils. Neutrophils are short lived and undergo apoptosis to maintain homeostasis and resolution of inflammation. A delay in apoptosis of neutrophils is one important mechanism which allows for their prolonged presence and the release of potentially harmful enzymes. The purpose of this study was to examine whether EtOH intoxication combined with burn injury influences neutrophil apoptosis and whether IL-18 plays any role in this setting. To accomplish this investigation, rats were gavaged with EtOH (3.2 g/kg) 4 h before being subjected to sham or burn injury of ~12.5% of the total body surface area, and then killed on d 1 after injury. Peripheral blood neutrophils were isolated and lysed. The lysates were analyzed for pro- and antiapoptotic proteins. We found that EtOH combined with burn injury prolonged neutrophil survival. This prolonged neutrophil survival was accompanied by a decrease in the levels of the neutrophil proapoptotic protein Bax, and an increase in antiapoptotic proteins Mcl-1 and Bcl-xl. Administration of IL-18 antibody following burn injury normalized the levels of Bax, Mcl-1 and Bcl-xl. The decrease in caspase-3 and DNA fragmentation observed following EtOH and burn injury was also normalized in rats treated with anti-IL-18 antibody. These findings suggest that IL-18 delays neutrophil apoptosis following EtOH and burn injury by modulating the pro- and antiapoptotic proteins.

  16. 乳酸盐与丙酮酸盐腹膜透析液对外周血中性粒细胞凋亡的影响%Effects of glucose-based lactate and pyruvate peritoneal dialysates on peripheral blood neutrophils apoptosis

    Institute of Scientific and Technical Information of China (English)

    金惠敏; 叶志斌

    2000-01-01

    目的:比较研究乳酸盐与丙酮酸盐腹膜透析(腹透)液对外周血中性粒细胞(PMN)凋亡以及吞噬和杀菌能力的影响。方法:含有不同浓度(1.5%,4.25%)葡萄糖的乳酸盐或丙酮酸盐腹透液,与外周血PMN孵育不同时间(10,30min)后,利用流式细胞术及荧光素标记的Annexin-V检测细胞凋亡情况;通过菌落计数方法观察PMN对金黄色葡萄球菌(金葡菌)吞噬和杀菌功能的改变,分别以细胞外和细胞内活菌下降的百分数表示。结果:PMN与1.5%乳酸盐腹透液(pH6.7)孵育10min,收集细胞,培养72h后细胞凋亡率与正常对照组比较无差异,而孵育时间延长至30min后,培养的PMN在72h凋亡率[(66.90±2.35)%]明显增加,与4.25%乳酸盐腹透液(pH7.2)10min组结果相似[(65.36±2.60)%];1.5%葡萄糖乳酸盐[(66.90±2.35)%]明显增加,与4.25%乳酸盐腹透液(pH7.2)10min组结果相似[(65.36±2.60)%];1.5%葡萄糖乳酸盐腹透孵育30min及4.25%葡萄糖乳酸盐腹透液孵育10min组,PMN的吞噬及杀菌功能均较对照组显著降低。无论是含1.5%还是4.25%葡萄糖的丙酮酸盐腹透液,PMN凋亡率均与对照组无显著差异;对PMN吞噬和杀金葡菌功能也无显著影响。结论:丙酮酸盐腹透液对外周血PMN凋亡以及吞噬和杀菌功能的影响均较小,具有较好的临床应用前景。%Objective:To compare the effects of variable glucose-basedlactate and pyruvate peritoneal dialysates on apoptosis,phagocytosis and bacteral killing of staphylococcus aureus in peripheral blood neutrophils.Methods:Cell apoptosis was analyzed by flow cytometry (FCM) to detect phosphatidylserine (PS) exposure on early apoptotic neutrophils using fluorescein labeled Annexin-V.Phagocytosis and intracellular bactericidal assays were expressed with the decrease in the number of viable extracellular and intracellular bacteria.Results:After exposure to 1.5% glucose lactate

  17. Stress-sensitive organs and blood corticosterone after immobilization of active and passive rats immunized with glutamate-bovine serum albumin conjugate.

    Science.gov (United States)

    Umryukhin, A E; Sotnikov, S V; Chekmareva, N Yu; Vetrile, L A; Zakharova, I A

    2014-12-01

    We studied stress-induced organ and hormonal responses in behaviorally active and passive rats against the background of immunization with glutamate-BSA conjugate. The relative weight of the adrenal glands after immobilization was lower in rats immunized with the conjugate in comparison with non-immunized animals. The weight of the adrenal glands in behaviorally active rats decreased in parallel with the decrease in blood corticosterone. In behaviorally active and passive rats immunized with the conjugate, ulcer formation in the stomach was slightly intensified after immobilization. It was hypothesized that immunization with glutamate-BSA conjugate suppresses activity of the hypothalamic-pituitary-adrenal feedback mechanism underlying the production of glucocorticoid hormones, which is manifested in slightly increased ulceration due to attenuation of the gastroprotective action of corticosterone under stress.

  18. The natural stilbenoid pinosylvin and activated neutrophils: effects on oxidative burst, protein kinase C, apoptosis and efficiency in adjuvant arthritis

    Institute of Scientific and Technical Information of China (English)

    Viera JAN(C)INOV(A); Tomá(s) PERE(C)KO; Rado NOS(A)(L); Juraj HARMATHA; Jan (S)MIDRKAL; Katarína DR(A)BIKOV(A)

    2012-01-01

    Aim:To investigate the effects of the naturally occurring stilbenoid pinosylvin on neutrophil activity in vitro and in experimental arthritis,and to examine whether protein kinase C (PKC) activation served as an assumed target of pinosylvin action.Methods:Fresh human blood neutrophils were isolated.The oxidative burst of neutrophils was evaluated on the basis of enhanced chemiluminescence.Neutrophil viability was evaluated with flow cytometry,and PKC phosphorylation was assessed by Western blotting analysis.Adjuvant arthritis was induced in Lewis rats with heat-killed Mycobacterium butyricum,and the animals were administered with pinosylvin (30 mg/kg,po) daily for 21 d after arthritis induction.Results:In isolated human neutrophils,pinosylvin (10 and 100 μmol/L) significantly decreased the formation of oxidants,both extraand intracellularly,and effectively inhibited PKC activation stimulated by phorbol myristate acetate (0.05 μmol/L).The inhibition was not due to neutrophil damage or increased apoptosis.In arthritic rats,the number of neutrophils in blood was dramatically increased,and whole blood chemiluminescence (spontaneous and PMA-stimulated) was markedly enhanced.Pinosylvin administration decreased the number of neutrophils (from 69 671±5588/μL to 51 293±3947/μL,P=0.0198) and significantly reduced the amount of reactive oxygen species in blood.Conclusion:Pinosylvin is an effective inhibitor of neutrophil activity,and is potentially useful as a complementary medicine in states associated with persistent inflammation.

  19. 牛病毒性腹泻病毒感染牛外周血单核细胞对CD80和CD86 mRNA转录的影响%Effects of bovine viral diarrhea viruses in vitro on transcription of CD80 and CD86 mRNA in bovine peripheral blood mononuclear cells

    Institute of Scientific and Technical Information of China (English)

    韩猛立; 黄新; 钟发刚

    2012-01-01

    The clinically healthy Holstein bovine tested negative for bovine viral diarrhea virus(BVDV) in peripheral blood mononuclear cells(PBMC) were infected with noncytopathic(NCP) and cytopathic(CP) BVDV.The changes levels of mRNA of CD80 and CD86 genes were analyzed using a real-time fluorescent quantitative PCR(real-time FQ-PCR).The results showed that the transcription of CD80 mRNA exhibited two transcription high at 4 h(P0.05) and 12,24 h(P0.01) post-inoculation(PI),and CD86 mRNA reached the highest level at 6 h(P0.05)PI in NCP BVDV-infected PBMC;then the mRNAs transcriptions of CD80 peaked at 24 h(P0.05)and CD86 peaked at 6 h(P0.05) in CP BVDV-infected PBMC with signifficant differences compared to that of the other PI.While the transcription on CD80 mRNAs witnessed more kinetic changes,it indicats NCP and CP BVDV could significantly supress the transcription of CD80-CD86 genes early during the infection,and the situation might weaken the antigen presentation of PBMC in the inoculated bovine.%用非致细胞病变(noncytopathic,NCP)和致细胞病变(cytopathic,CP)型牛病毒性腹泻病毒(BVDV)感染临床健康BVDV检测阴性的荷斯坦奶牛外周血单核细胞(PBMC),利用实时荧光定量PCR技术对感染后共刺激分子CD80和CD86mRNA转录水平的变化进行定量分析。结果表明,在NCP型BVDV感染牛PBMC后CD80在4h(P〈0.05)和12,24h(P〈0.01)出现2次转录高峰,CD86在6h(P〈0.05)出现转录高峰;CP型BVDV感染后,CD80在24h(P〈0.05)出现转录高峰,CD86在6h(P〈0.05)出现转录高峰。尽管CD80在NCP型BVDV感染后呈现较复杂的动态变化,但结果提示NCP型和CP型BVDV感染均可导致牛PBMC的共刺激分子CD80和CD86基因转录在感染早期明显受到抑制,PBMC的抗原呈递能力受到影响。

  20. Protein Quality of Rice Drinks Fortified with Bovine and Porcine Blood Plasma / Calidad Proteica de Bebidas de Arroz Fortificadas con Plasma Sanguíneo de Bovino y Porcino

    Directory of Open Access Journals (Sweden)

    Piedad Margarita Montero Castillo

    2014-12-01

    Full Text Available Abstract. The future of nutrition in Colombia, and perhaps inother developing countries, will depend in large part on the abilityof food technology to take full advantage of the food sourcesavailable in the country and to adapt and develop new productsthat will vary and complement the diets of the majority of thepopulation at a low cost. The objective of this study was to evaluatethe protein quality of rice-based drinks fortified with bovine andporcine blood plasma. Six treatments were prepared with differentlevels of fortification (14.5%, 18.5% and 29%. The effects of theplasma type and the addition levels on the protein content, theamino acid profile, and the in vitro digestibility of the drinks wereobserved. The AOAC method was employed for the determinationof the protein content; the amino acid profile was created usingHPLC. The protein digestibility was determined by subjecting adispersion of the drink to the action of a multi-enzymatic solution.The protein content increased with the level of fortification. Thedrinks fortified with bovine plasma (104% and porcine plasma(89% presented a better protein quality index than the unfortifieddrink. The digestibility of the fortified drinks did not demonstratesignificant improvements in comparison with the unfortified drink.The chemical score of the drinks fortified with porcine plasma(71.6 and bovine plasma (78.5 showed that the latter had thebest nutritional quality. / Resumen. El futuro de la alimentación en Colombia y quizás deotros países en desarrollo va a depender en gran parte de quela tecnología de alimentos sea capaz de aprovechar las fuentesdisponibles de alimentos en el país y de adaptar y desarrollarnuevos productos que permitan variar y complementar la dietade la mayoría de la población a bajo costo. El objetivo de estetrabajo fue evaluar la calidad proteica de bebidas a base dearroz fortificadas con plasma sanguíneo de bovino y porcino. Seprepararon seis tratamientos con

  1. Neutrophil Reverse Migration Becomes Transparent with Zebrafish

    Directory of Open Access Journals (Sweden)

    Taylor W. Starnes

    2012-01-01

    Full Text Available The precise control of neutrophil-mediated inflammation is critical for both host defense and the prevention of immunopathology. In vivo imaging studies in zebrafish, and more recently in mice, have made the novel observation that neutrophils leave a site of inflammation through a process called neutrophil reverse migration. The application of advanced imaging techniques to the genetically tractable, optically transparent zebrafish larvae was critical for these advances. Still, the mechanisms underlying neutrophil reverse migration and its effects on the resolution or priming of immune responses remain unclear. Here, we review the current knowledge of neutrophil reverse migration, its potential roles in host immunity, and the live imaging tools that make zebrafish a valuable model for increasing our knowledge of neutrophil behavior in vivo.

  2. Neutrophil Responses to Sterile Implant Materials.

    Directory of Open Access Journals (Sweden)

    Siddharth Jhunjhunwala

    Full Text Available In vivo implantation of sterile materials and devices results in a foreign body immune response leading to fibrosis of implanted material. Neutrophils, one of the first immune cells to be recruited to implantation sites, have been suggested to contribute to the establishment of the inflammatory microenvironment that initiates the fibrotic response. However, the precise numbers and roles of neutrophils in response to implanted devices remains unclear. Using a mouse model of peritoneal microcapsule implantation, we show 30-500 fold increased neutrophil presence in the peritoneal exudates in response to implants. We demonstrate that these neutrophils secrete increased amounts of a variety of inflammatory cytokines and chemokines. Further, we observe that they participate in the foreign body response through the formation of neutrophil extracellular traps (NETs on implant surfaces. Our results provide new insight into neutrophil function during a foreign body response to peritoneal implants which has implications for the development of biologically compatible medical devices.

  3. Ca2+ response in neutrophils after exposure to bacterial N-formyl-methionyl-leucyl-phenylalanine: delayed response in ulcerative colitis

    DEFF Research Database (Denmark)

    Vainer, Ben; Lamberth, Kasper; Brimnes, Jens;

    2003-01-01

    In acute stages of ulcerative colitis (UC), neutrophils migrate from the circulation into inflamed colonic tissue, initiated by yet unknown stimuli. The bacterial peptide N-formyl-methionyl-leucyl-phenylalanine (FMLP) is a component of the surface membrane of colonic bacteria such as Escherichia...... coli and stimulates Ca2+ influx into neutrophils, reflecting the fact that ionized calcium is an important secondary messenger for several neutrophil functions, including locomotion, phagocytosis and free oxygen radical production. Recent studies have revealed that Ca2+ dependent ICAM-1/beta 2-integrin...... mediated neutrophil migration is impaired in UC patients. The aim of the present work was to study the influx of Ca2+ into peripheral blood neutrophils of UC patients after exposure to FMLP and after binding of either beta 2-integrins or intercellular adhesion molecule-1 (ICAM-1)....

  4. Cryptococcus Neoformans Modulates Extracellular Killing by Neutrophils

    OpenAIRE

    Qureshi, Asfia; Grey, Angus; Rose, Kristie L; Schey, Kevin L.; Del Poeta, Maurizio

    2011-01-01

    We recently established a key role for host sphingomyelin synthase (SMS) in regulating the killing activity of neutrophils against Cryptococcus neoformans. In this paper, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and natural killer (NK) cells (Tgε26 mice). To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in...

  5. Viral and Bacterial Pathogens in Bovine Respiratory Disease in Finland

    Directory of Open Access Journals (Sweden)

    Soveri T

    2004-12-01

    Full Text Available Pathogens causing bovine respiratory tract disease in Finland were investigated. Eighteen cattle herds with bovine respiratory disease were included. Five diseased calves from each farm were chosen for closer examination and tracheobronchial lavage. Blood samples were taken from the calves at the time of the investigation and from 86 calves 3–4 weeks later. In addition, 6–10 blood samples from animals of different ages were collected from each herd, resulting in 169 samples. Serum samples were tested for antibodies to bovine parainfluenza virus-3 (PIV-3, bovine respiratory syncytial virus (BRSV, bovine coronavirus (BCV, bovine adenovirus-3 (BAV-3 and bovine adenovirus-7 (BAV-7. About one third of the samples were also tested for antibodies to bovine virus diarrhoea virus (BVDV with negative results. Bacteria were cultured from lavage fluid and in vitro susceptibility to selected antimicrobials was tested. According to serological findings, PIV-3, BAV-7, BAV-3, BCV and BRSV are common pathogens in Finnish cattle with respiratory problems. A titre rise especially for BAV-7 and BAV-3, the dual growth of Mycoplasma dispar and Pasteurella multocida, were typical findings in diseased calves. Pasteurella sp. strains showed no resistance to tested antimicrobials. Mycoplasma bovis and Mannheimia haemolytica were not found.

  6. Presence of Gumprecht shadows (smudge cells) in bovine leukemia virus-positive cattle.

    Science.gov (United States)

    Panei, Carlos Javier; Larsen, Alejandra; González, Ester Teresa; Echeverría, María Gabriela

    2013-11-01

    Enzootic Bovine Leukosis is a chronic disease caused by the bovine leukemia virus (BLV). Smudge cells, also known as Gumprecht shadows, are not simple artifacts of slide preparation, but ragged lymphoid cells found mainly in peripheral blood smears from human patients with chronic lymphocytic leukemia. In this study, we report the presence of Gumprecht shadows in peripheral blood from BLV-positive cattle.

  7. Bordetella parapertussis Circumvents Neutrophil Extracellular Bactericidal Mechanisms

    Science.gov (United States)

    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

  8. Fungal and bacterial killing by neutrophils.

    Science.gov (United States)

    Ermert, David; Zychlinsky, Arturo; Urban, Constantin

    2009-01-01

    Neutrophils are professional phagocytes of the innate immune system that are essential to control bacterial and fungal infections. These cells engulf and kill invading microbes. Additionally, activated neutrophils are able to release neutrophil extracellular traps (NETs). These fibers consist of chromatin decorated with antimicrobial proteins to trap and kill microbes. Appropriate quantitative methods are required to understand the nature of interactions of neutrophils with pathogens. Here we present assays to measure killing mediated by phagocytosis, by NETs, by a combination of both, and by granular extract. As examples, we use Candida albicans for fungal and Shigella flexneri for bacterial pathogens.

  9. Tumor Necrosis Factor, but Not Neutrophils, Alters the Metabolic Profile in Acute Experimental Arthritis.

    Directory of Open Access Journals (Sweden)

    Marina C Oliveira

    Full Text Available Metabolic alterations are associated with arthritis apart from obesity. However, it is still unclear which is the underlying process behind these metabolic changes. Here, we investigate the role of tumor necrosis factor (TNF in this process in an acute model of antigen-induced arthritis (AIA. Immunized male BALB/c mice received an intra-articular injection of PBS (control or methylated bovine serum albumin (mBSA into their knees, and were also pre-treated with different drugs: Etanercept, an anti-TNF drug, DF2156A, a CXCR1/2 receptor antagonist, or a monoclonal antibody RB6-8C5 to deplete neutrophils. Local challenge with mBSA evoked an acute neutrophil influx into the knee joint, and enhanced the joint nociception, along with a transient systemic metabolic alteration (higher levels of glucose and lipids, and altered adipocytokines. Pre-treatment with the conventional biological Etanercept, an inhibitor of TNF action, ameliorated the nociception and the acute joint inflammation dominated by neutrophils, and markedly improved many of the altered systemic metabolites (glucose and lipids, adipocytokines and PTX3. However, the lessening of metabolic changes was not due to diminished accumulation of neutrophils in the joint by Etanercept. Reduction of neutrophil recruitment by pre-treating AIA mice with DF2156A, or even the depletion of these cells by using RB6-8C5 reduced all of the inflammatory parameters and hypernociception developed after AIA challenge, but could not prevent the metabolic changes. Therefore, the induction of joint inflammation provoked acute metabolic alterations which were involved with TNF. We suggest that the role of TNF in arthritis-associated metabolic changes is not due to local neutrophils, which are the major cells present in this model, but rather due to cytokines.

  10. Quality and safety of bovine clones and their products.

    Science.gov (United States)

    Heyman, Y; Chavatte-Palmer, P; Fromentin, G; Berthelot, V; Jurie, C; Bas, P; Dubarry, M; Mialot, J P; Remy, D; Richard, C; Martignat, L; Vignon, X; Renard, J P

    2007-08-01

    A multidisciplinary research programme was developed to get a scientific expertise for the quality assessment of products obtained from cloned livestock. Thirty-seven bovine Holstein female clones of five different genotypes and their products were analysed in comparison with 38 control animals obtained by conventional artificial insemination and raised under the same conditions at the same experimental farm. Animal evaluation included over 150 criteria and more than 10 000 measurements to check the physiological status and health over a 3-year period. All the parameters studied were in the normal range for age and breed, but some significant differences were detected between clone and control groups in terms of delayed onset of puberty in clones, higher neutrophil counts in haematology or lower biochemical plasma concentrations of gamma glutamyl transferase. Milk and meat analyses were conformable to expected values. We, however, found some differences in fatty acid (FA) composition of milk and muscle suggesting a possible deviation in lipid metabolism as assessed by higher delta-9 desaturase activity indexes in both milk and muscles from clones compared with controls. Repeated muscle biopsies in the semitendinosus muscle of the same animals demonstrated a higher oxidative activity in muscle of young clones (8 months of age) compared with controls, suggesting a delayed muscle maturation in clones. Nutritional evaluation of milk and meat using the rat feeding trials did not show any difference between clone and control products for food intake, growth rate, body composition of the rats, nor for possible allergenicity. Possible reactivation of bovine endogenous retroviruses (BERVs) was analysed and compared between normal and cloned cattle. As expected, these BERV sequences are not transcribed and no RNA was detected in the blood of clones, donor animals or controls; therefore, it may be assumed that the sanitary risk associated with BERV sequences is not higher in

  11. 77 FR 20319 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products

    Science.gov (United States)

    2012-04-04

    ...; ] DEPARTMENT OF AGRICULTURE Animal and Plant Health Inspection Service 9 CFR Part 93 RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products Correction In proposed rule...

  12. 78 FR 73993 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products

    Science.gov (United States)

    2013-12-10

    ... Health Inspection Service 9 CFR Parts 92, 93, 94, 95, 96, and 98 RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products Corrections In rule document 2013-28228 appearing...

  13. Neutrophils are immuno-modulatory in rhinovirus infections

    NARCIS (Netherlands)

    Tang, Francesca; Hansbro, Philip; Burgess, Janette; Baines, Katherine; Oliver, Brian

    2015-01-01

    Background: Neutrophils are important in controlling bacterial infections however; their role in viral infections remains unclear. Previously, we found that neutrophils respond to viral mimetics but not replication competent rhinovirus (RV). Aim: To investigate if neutrophils are activated when expo

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

    Science.gov (United States)

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

    1986-11-01

    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.

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

    Science.gov (United States)

    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.

  16. Bovine Peripheral Blood Monocyte Derived Dendritic Cell Culture and Identification in Vitro%奶牛外周血树突状细胞体外诱导培养与鉴定

    Institute of Scientific and Technical Information of China (English)

    詹康; 赵倩明; 隋雁南; 封飞飞; 占今舜; 赵国琦

    2016-01-01

    通过粒-巨噬细胞集落刺激因子( GM⁃CSF)和白细胞介素-4( IL⁃4)体外诱导外周血单核细胞为树突状细胞,为利用树突状细胞免疫疗法治疗奶牛乳房炎奠定基础和提供细胞模型。利用淋巴细胞分离液分离获得奶牛外周血单核细胞,在6孔板内培养2h后,弃掉含有大量的T细胞和B细胞上清液,贴壁的基本上是单核细胞,磷酸盐缓冲液清洗5次,加入含有GM⁃CSF和IL⁃4的2 mL培养基进行3 d诱导。之后,从培养基顶部小心吸弃1.4 mL的培养基,然后再补加含有GM⁃CSF和IL⁃4的1.8 mL培养基继续诱导3 d。每天通过显微镜观察细胞形态。第7天经流式检测细胞表面抗原 CD11c、CD14、主要组织相容性复合体Ⅱ( MHCⅡ)、CD40、CD80、CD86的表达。结果表明:1)第2天,一些细胞表面可以生长出刺突并伴随着伪足的生长。第3天,细胞表面的刺突和伪足越来越多。第4、5天,一些带有刺突和伪足的细胞开始聚集和融合。第6天,单核细胞基本被诱导为树突状细胞,细胞表面含有大量清晰可见的刺突和伪足。2)经流式检测,CD14、CD11c、MHCⅡ阳性表达细胞分别占诱导细胞的6.8%、65.0%、75.9%,CD80和CD86阳性表达细胞分别占诱导细胞的2.0%和1.2%。综上所述,采用奶牛外周血单核细胞经体外诱导能够获得一定纯度的奶牛树突状细胞。%This study aimed to induce bovine peripheral blood monocyte derived dendritic cell by granulocyte⁃macrophage colony stimulating factor ( GM⁃CSF) and interleukin⁃4 ( IL⁃4) cytokines, which could lay founda⁃tion and provide cell model for dairy cow mastitis treatment using cell immunotherapy. The bovine peripheral blood monocyte was acquired by lymphocyte separation medium and seeded in 6⁃proe plate to culture for 2 h. Then, suspended cells containing an amount of B and T cells were discarded, and

  17. The innate defense regulator peptides IDR-HH2, IDR-1002, and IDR-1018 modulate human neutrophil functions.

    Science.gov (United States)

    Niyonsaba, François; Madera, Laurence; Afacan, Nicole; Okumura, Ko; Ogawa, Hideoki; Hancock, Robert E W

    2013-07-01

    Although HDPs were originally hypothesized to act as antimicrobial agents, they also have been shown to broadly modulate the immune response through the activation of different cell types. We recently developed a series of novel, synthetic peptides, termed IDRs, which are conceptually based on a natural HDP, bovine bactenecin. We showed that IDR-1 and IDR-1002 protect the host against bacterial infections through the induction of chemokines. The objective of this study was to investigate the effects of the IDRs on various functions of human neutrophils. Here, we demonstrated that IDR-HH2, IDR-1002, and IDR-1018 modulated the expression of neutrophil adhesion and activation markers. Moreover, these IDRs enhanced neutrophil adhesion to endothelial cells in a β₂ integrin-dependent manner and induced neutrophil migration and chemokine production. The IDR peptides also increased the release of the neutrophil-generated HDPs (antimicrobial), human α-defensins, and LL-37 and augmented neutrophil-mediated killing of Escherichia coli. Notably, the IDRs significantly suppressed LPS-mediated neutrophil degranulation, the release of ROS, and the production of the inflammatory cytokines TNF-α and IL-10, consistent with their ability to dampen inflammation. As evidenced by the inhibitory effects of MAPK-specific inhibitors, IDRs activated the MAPK pathway that was required for chemokine production. In conclusion, our study provides novel evidence regarding the contribution of the IDR peptides to the innate immune response through the modulation of neutrophil functions. The results described here may aid in the development of IDRs as novel, anti-infective and immunomodulatory agents.

  18. IL-17 attenuates the anti-apoptotic effects of GM-CSF in human neutrophils.

    Science.gov (United States)

    Dragon, Stéphane; Saffar, Arash Shoja; Shan, Lianyu; Gounni, Abdelilah Soussi

    2008-01-01

    Interleukin (IL)-17A is a pleiotropic, pro-inflammatory cytokine that is implicated in chronic inflammatory and degenerative disorders. IL-17 has been demonstrated to link activated T-lymphocyte with the recruitment of neutrophils at sites of inflammation, however whether IL-17 can mediate neutrophil survival and subsequently affect inflammatory responses has not fully been elucidated. In our study, we demonstrate that human peripheral blood and HL-60 differentiated neutrophils express mRNA and cell surface IL-17A receptor. IL-17A does not affect the rate of spontaneous neutrophil apoptosis, however significantly decreased granulocyte macrophage-colony stimulating factor (GM-CSF)-mediated survival by antagonizing the signal transduction pathways of p38, Erk1/2 and signal transducer and activator of transcription (STAT) 5B. These events were associated with reduced myeloid cell lymphoma-1 (Mcl-1) protein levels, increased translocation and aggregation of Bax to mitochondria, decreased mitochondrial transmembrane potential and in an increase in caspase-3/7 activity. These events were independent of increased Fas or soluble Fas ligand expression levels. Taken together, our findings suggest that IL-17 may regulate neutrophil homeostasis and favor the resolution of inflamed tissues by attenuating the delay in neutrophil apoptosis induced by inflammatory cytokines.

  19. Value of peripheral blood neutrophil to lymphocyte ratio for the early diagnosis of acute coronary syndrome%外周血中性粒细胞与淋巴细胞比值对急性冠脉综合征早期诊断的价值

    Institute of Scientific and Technical Information of China (English)

    尹炳坚; 傅强; 严全能; 陈若峰; 郭建浩; 李志樑

    2015-01-01

    目的:探讨外周血中性粒细胞与淋巴细胞比值(NLR)对急性冠脉综合征(ACS)早期诊断的价值。方法选择疑似ACS的胸痛患者247例,包括急性ST段抬高型心肌梗死(STEMI)51例,急性非ST 段抬高型心肌梗死(NSTEMI)42例,不稳定型心绞痛(UA)87例,非心源性胸痛(NCCP)67例。根据患者入院时白细胞计数及分类计数结果,计算并分析NLR对ACS的诊断灵敏度、特异度、阳性预测值、阴性预测值,同时分析NLR的受试者工作特征(ROC)曲线。结果所有患者中,UA患者所占比例最高,其余依次为 NCCP、STEMI和 NSTEMI。各组中性粒细胞百分比、白细胞计数变化趋势一致,由 NCCP组、UA 组、NSTEMI组到STEMI组依次升高,但中淋巴细胞百分比呈相反的变化趋势。NLR对ACS的诊断灵敏度、特异度、准确性、阳性预测值、阴性预测值高于白细胞计数。结论 NLR具有费用低廉、简便易行、结果稳定、易于重复及动态观察等优点,对ACS早期诊断、病情评估及预后判断有重要指导意义。%Objective To investigate the peripheral blood neutrophil to lymphocyte ratio(NLR) for the early diagnosis of acute coronary syndrome(ACS) .Methods A total of 247 patients with suspected ACS and chest pain ,including 51 cases with acute ST segment elevation myocardial infarction(STEMI) ,42 cases with acute non‐ST segment elevation myocardial infarction(NSTEMI) , 87 cases with unstable angina pectoris(UA) and 67 cases with non‐cardiogenic chest pain(NCCP)were enrolled and detected for white blood cells count and classification .The sensitivity ,specificity ,positive predictive value ,negative predictive value ,receiver op‐erating characteristic(ROC) curve of NLR were analyzed .Results Among all patients ,the most common was UA ,followed by NC‐CP ,STEMI and NSTEMI .Level of neutrophil proportion and white blood cell count were lowest in NCCP group

  20. Unlocking the bovine genome

    Directory of Open Access Journals (Sweden)

    Worley Kim C

    2009-04-01

    Full Text Available Abstract The draft genome sequence of cattle (Bos taurus has now been analyzed by the Bovine Genome Sequencing and Analysis Consortium and the Bovine HapMap Consortium, which together represent an extensive collaboration involving more than 300 scientists from 25 different countries.

  1. Elevated Neutrophil Lymphocyte Ratio in Recurrent Optic Neuritis

    Directory of Open Access Journals (Sweden)

    Hande Guclu

    2015-01-01

    Full Text Available Purpose. To demonstrate the relation between optic neuritis (ON and systemic inflammation markers as neutrophil lymphocyte ratio (N/L ratio, platelet count, mean platelet volume (MPV, and red cell distribution width (RDW and furthermore to evaluate the utilization of these markers to predict the frequency of the ON episodes. Methods. Forty-two patients with acute ON and forty healthy subjects were enrolled into the study. The medical records were reviewed for age, sex, hemoglobin (Hb, Haematocrit (Htc, RDW, platelet count, MPV, white blood cell count (WBC, neutrophil and lymphocyte count, and neutrophil lymphocyte ratio (N/L ratio. Results. The mean N/L ratio, platelet counts, and RDW were significantly higher in ON group (p=0.000, p=0.048, and p=0.002. There was a significant relation between N/L ratio and number of episodes (r=0.492, p=0.001. There was a statistically significant difference for MPV between one episode group and recurrent ON group (p=0.035. Conclusions. Simple and inexpensive laboratory methods could help us show systemic inflammation and monitor ON patients. Higher N/L ratio can be a useful marker for predicting recurrent attacks.

  2. Yersinia enterocolitica-mediated degradation of neutrophil extracellular traps (NETs).

    Science.gov (United States)

    Möllerherm, Helene; Neumann, Ariane; Schilcher, Katrin; Blodkamp, Stefanie; Zeitouni, Nathalie E; Dersch, Petra; Lüthje, Petra; Naim, Hassan Y; Zinkernagel, Annelies S; von Köckritz-Blickwede, Maren

    2015-12-01

    Neutrophil extracellular trap (NET) formation is described as a tool of the innate host defence to fight against invading pathogens. Fibre-like DNA structures associated with proteins such as histones, cell-specific enzymes and antimicrobial peptides are released, thereby entrapping invading pathogens. It has been reported that several bacteria are able to degrade NETs by nucleases and thus evade the NET-mediated entrapment. Here we studied the ability of three different Yersinia serotypes to induce and degrade NETs. We found that the common Yersinia enterocolitica serotypes O:3, O:8 and O:9 were able to induce NETs in human blood-derived neutrophils during the first hour of co-incubation. At later time points, the NET amount was reduced, suggesting that degradation of NETs has occurred. This was confirmed by NET degradation assays with phorbol-myristate-acetate-pre-stimulated neutrophils. In addition, we found that the Yersinia supernatants were able to degrade purified plasmid DNA. The absence of Ca(2+) and Mg(2+) ions, but not that of a protease inhibitor cocktail, completely abolished NET degradation. We therefore postulate that Y. enterocolitica produces Ca(2+)/Mg(2+)-dependent NET-degrading nucleases as shown for some Gram-positive pathogens.

  3. Neutrophil granules in health and disease

    DEFF Research Database (Denmark)

    Häger, M; Cowland, J B; Borregaard, N

    2010-01-01

    Neutrophil granules store proteins that are critically important for the neutrophil to move from the vascular bed to tissues and to kill microorganisms. This is illustrated in nature when individual proteins are deleted due to inherited mutations of their cognate genes, and such deficiencies resu...

  4. Neutrophils: potential therapeutic targets in tularemia?

    Directory of Open Access Journals (Sweden)

    Lee-Ann H Allen

    2013-12-01

    Full Text Available The central role of neutrophils in innate immunity and host defense has long been recognized, and the ability of these cells to efficiently engulf and kill invading bacteria has been extensively studied, as has the role of neutrophil apoptosis in resolution of the inflammatory response. In the past few years additional immunoregulatory properties of neutrophils were discovered, and it is now clear that these cells play a much greater role in control of the immune response than was previously appreciated. In this regard, it is noteworthy that Francisella tularensis is one of relatively few pathogens that can successfully parasitize neutrophils as well as macrophages, DC and epithelial cells. Herein we will review the mechanisms used by F. tularensis to evade elimination by neutrophils. We will also reprise effects of this pathogen on neutrophil migration and lifespan as compared with other infectious and inflammatory disease states. In addition, we will discuss the evidence which suggests that neutrophils contribute to disease progression rather than effective defense during tularemia, and consider whether manipulation of neutrophil migration or turnover may be suitable adjunctive therapeutic strategies.

  5. Proteases, neutrophils, and periodontitis: the NET effect.

    Science.gov (United States)

    Nauseef, William M

    2014-10-01

    Neutrophils exert potent antimicrobial activities in their role as first-line cellular defenders against infection. The synergistic and collective actions of oxidants and granule proteins, including serine proteases, support the microbial killing in phagosomes, where most neutrophil-mediated antimicrobial action occurs. In addition to phagocytosis, specific stimuli prompt neutrophils to extrude a matrix of DNA, histones, and granule proteins to produce neutrophil extracellular traps (NETs), which can trap microbes. Mice lacking the serine proteases necessary for NET production are more susceptible to infection, an observation suggesting that functional NETs are required for host protection. In this issue of the JCI, Sørensen and colleagues characterize neutrophils from a patient with Papillon-Lefèvre syndrome. The patient has an inactivating mutation in the gene encoding dipeptidyl peptidase I, resulting in neutrophils lacking elastase, a serine protease required for NET production. Despite the inability to form NETS, neutrophils from this patient killed pathogens in vitro, and the patient did not exhibit evidence of an increased propensity toward bacterial infections. Together, these results suggest that proteases in human neutrophils are dispensable for protection against bacterial infection and that the ability to generate NETs in vitro does not compromise host defense.

  6. Camel and bovine chymosin

    DEFF Research Database (Denmark)

    Langholm Jensen, Jesper; Mølgaard, Anne; Navarro Poulsen, Jens Christian;

    2013-01-01

    Bovine and camel chymosin are aspartic peptidases that are used industrially in cheese production. They cleave the Phe105-Met106 bond of the milk protein κ-casein, releasing its predominantly negatively charged C-terminus, which leads to the separation of the milk into curds and whey. Despite...... having 85% sequence identity, camel chymosin shows a 70% higher milk-clotting activity than bovine chymosin towards bovine milk. The activities, structures, thermal stabilities and glycosylation patterns of bovine and camel chymosin obtained by fermentation in Aspergillus niger have been examined...... interactions arising from variation in the surface charges and the greater malleability both in domain movements and substrate binding contribute to the better milk-clotting activity of camel chymosin towards bovine milk....

  7. Iron-chelating agent, deferasirox, inhibits neutrophil activation and extracellular trap formation.

    Science.gov (United States)

    Kono, Mari; Saigo, Katsuyasu; Yamamoto, Shiori; Shirai, Kohei; Iwamoto, Shuta; Uematsu, Tomoko; Takahashi, Takayuki; Imoto, Shion; Hashimoto, Makoto; Minami, Yosuke; Wada, Atsushi; Takenokuchi, Mariko; Kawano, Seiji

    2016-10-01

    Iron-chelating agents, which are frequently prescribed to transfusion-dependent patients, have various useful biological effects in addition to chelation. Reactive oxygen species (ROS) produced by neutrophils can cause pulmonary endothelial cell damage, which can lead to acute lung injury (ALI). We previously reported that deferasirox (DFS), an iron-chelating agent, inhibits phorbol myristate acetate (PMA) or formyl-methionyl-leucyl-phenylalanine (fMLP)-induced ROS production in neutrophils, in vitro. Here, we investigate whether DFS inhibits vacuolization in neutrophils and neutrophil extracellular trap (NET) formation. Human neutrophils were incubated with DFS and stimulated with PMA or fMLP. Human neutrophils were separated from heparinized peripheral blood using density gradient centrifugation, and subsequently incubated with DFS. After 10 minutes, neutrophils were stimulated by PMA or fMLP. Vacuole formation was observed by electron microscopy. For observing NET formations using microscopes, immunohistological analyses using citrullinated histone H3 and myeloperoxidase antibodies, and SYTOX Green (an impermeable DNA detection dye) staining, were conducted. NET formation was measured as the quantity of double-stranded DNA (dsDNA), using the AccuBlue Broad Range dsDNA Quantitation Kit. DFS (50 μmol/L) inhibited vacuole formation in the cytoplasm and NET formation. Additionally, 5-100 μmol/L concentration of DFS inhibited the release of dsDNA in a dose-independent manner. We demonstrate that DFS inhibits not only ROS production but also vacuolization and NET formation in neutrophils. These results suggest the possibility of protective effects of DFS against NET-related adverse effects, including ALI and thrombosis.

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

    Science.gov (United States)

    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

  9. Trace of antibody to myeloperoxidase with nanocrystal quantum dot labeled antibody recognizing activating neutrophils

    Science.gov (United States)

    Hoshino, Akiyoshi; Nagao, Tomokazu; Yamamoto, Kenji; Suzuki, Kazuo

    2006-02-01

    It is assumed that activated neutrophils contribute to the development of anti-neutrophil cytoplasmic auto-antibody (ANCA)-associated vasculitis due to the association of myelopeoxidase(MPO)-ANCA with MPO expressed on the surface of activated neutrophils. FITC-labeled antibody (Ab) used widely are not suitable for neutrophil examination because of the labile fluorescence emission of FITC. Therefore, it is necessary to develop specific fluorescent probes for MPO detection in neutrophils in vivo. Recently, fluorescent nanocrystal quantum dots (QDs) have been used for biotechnological and medical applications because of their greater and far longer fluorescence in. QDs have several advantages over organic fluorophores: high luminescence, far longer stability against photobleaching, and a range of fluorescence wavelengths from blue to infrared, depending on particle size. Thus, we examined the role of MPO and the Ab to MPO in the pathogenesis of glomerulonephritis associated with MPO-ANCA in experimental glomerulonephritis mice using QDs. We demonstrated the QD-conjugated anti-MPO Ab visualized the expression of MPO on the neutrophil surface after stimulation with proinflammatory cytokines. In addition, QD immuno-conjugates with anti-recombinant murine MPO (rmMPO) Ab revealed the trafficking of MPO-ANCA in vivo. Deceleration of blood flow in kidney vessels occurred in model mice, in which serum proteins including anti-rmMPO Ab were leaked out from collapsed glomeruli into the proximal tubule. Thus, sustained MPO expression on the neutrophil surface was significantly related to glomerulonephritis. These results indicate that the expressed MPO on the activated neutrophils with anti-MPO Ab may coordinately play essential roles in the initial steps for the development of glomerulonephritis.

  10. RT-PCR em pools de soros sangüíneos para o diagnóstico da infecção aguda e de animais persistentemente infectados pelo vírus da diarréia viral bovina RT-PCR in pools of bovine blood serum to detect acute infection and persistently infected animals with bovine viral diarrhea virus

    Directory of Open Access Journals (Sweden)

    D. Pilz

    2007-02-01

    Full Text Available Utilizou-se a técnica da RT-PCR para a detecção da região 5' UTR do genoma do vírus da diarréia viral bovina (BVDV em pools de soros sangüíneos provenientes de um rebanho, constituído por 226 animais, que apresentava distúrbios da reprodução. A partir das amostras individuais de soro e de acordo com a categoria dos animais e o número de animais por categoria foram formados 10 pools (A a J de soros. A primeira avaliação revelou a amplificação de um produto com 290pb nas reações referentes aos grupos D (35 vacas e H (25 bezerros lactentes que, após o desmembramento em amostras individuais, resultou na identificação de 11 vacas lactantes e 12 bezerros em amamentação positivos. Para a identificação de animais persistentemente infectados (PI entre os 23 positivos na primeira avaliação, realizou-se a segunda colheita de soros sangüíneos, três meses após. A RT-PCR das amostras individuais de soro revelou resultado positivo em cinco bezerros. Em dois, foi possível isolar o BVDV em cultivo de células MDBK. A especificidade das reações da RT-PCR foi confirmada pelo seqüenciamento dos produtos amplificados a partir do soro de uma vaca com infecção aguda, de um bezerro PI e das duas amostras do BVDV isoladas em cultivo celular. A utilização da RT-PCR em pools de soros sangüíneos demonstrou ser uma estratégia rápida de diagnóstico etiológico e de baixo custo tanto para a detecção de infecção aguda quanto de animais PI.The 5' untranslated region of the bovine viral diarrhea virus (BVDV genome was detected by RT-PCR assay in pools of blood sera samples collected from a cattle herd (n=226 animals with reproductive failures. Based on the classes of animal and the number of animals per class, the individual blood serum samples were distributed in 10 sera pools (A to J. During the first evaluation a 290bp amplicon was amplified in reactions from groups D (35 cows and H (25 sucking calves. The individual analysis

  11. Evasion of Neutrophil Killing by Staphylococcus aureus

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    Will A. McGuinness

    2016-03-01

    Full Text Available 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.

  12. Breast cancer associated a2 isoform vacuolar ATPase immunomodulates neutrophils: potential role in tumor progression.

    Science.gov (United States)

    Ibrahim, Safaa A; Katara, Gajendra K; Kulshrestha, Arpita; Jaiswal, Mukesh K; Amin, Magdy A; Beaman, Kenneth D

    2015-10-20

    In invasive breast cancer, tumor associated neutrophils (TAN) represent a significant portion of the tumor mass and are associated with increased angiogenesis and metastasis. Identifying the regulatory factors that control TAN behavior will help in developing ideal immunotherapies. Vacuolar ATPases (V-ATPases), multi-subunit proton pumps, are highly expressed in metastatic breast cancer cells. A cleaved peptide from a2 isoform V-ATPase (a2NTD) has immunomodulatory role in tumor microenvironment. Here, we report for the first time the role of V-ATPase in neutrophils modulation. In invasive breast cancer cells, a2NTD was detected and a2V was highly expressed on the surface. Immunohistochemical analysis of invasive breast cancer tissues revealed that increased neutrophil recruitment and blood vessel density correlated with increased a2NTD levels. In order to determine the direct regulatory role of a2NTD on neutrophils, recombinant a2NTD was used for the treatment of neutrophils isolated from the peripheral blood of healthy volunteers. Neutrophils treated with a2NTD (a2Neuɸ) showed increased secretion of IL-1RA, IL-10, CCL-2 and IL-6 that are important mediators in cancer related inflammation. Moreover, a2Neuɸ exhibited an increased production of protumorigenic factors including IL-8, matrix metaloprotinase-9 and vascular endothelial growth factor. Further, functional characterization of a2Neuɸ revealed that a2Neuɸ derived products induce in vitro angiogenesis as well as increase the invasiveness of breast cancer cells. This study establishes the modulatory effect of breast cancer associated a2V on neutrophils, by the action of a2NTD, which has a positive impact on tumor progression, supporting that a2V can be a potential selective target for breast cancer therapy.

  13. Ubiquitin-proteasome-rich cytoplasmic structures in neutrophils of patients with Shwachman-Diamond syndrome

    Science.gov (United States)

    Necchi, Vittorio; Minelli, Antonella; Sommi, Patrizia; Vitali, Agostina; Caruso, Roberta; Longoni, Daniela; Frau, Maria Rita; Nasi, Cristina; De Gregorio, Fabiola; Zecca, Marco; Ricci, Vittorio; Danesino, Cesare; Solcia, Enrico

    2012-01-01

    Background Shwachman–Diamond syndrome is an autosomal recessive disorder in which severe bone marrow dysfunction causes neutropenia and an increased risk of leukemia. Recently, novel particulate cytoplasmic structures, rich in ubiquitinated and proteasomal proteins, have been detected in epithelial cells and neutrophils from patients with Helicobacter pylori gastritis and several epithelial neoplasms. Design and Methods Blood neutrophils from 13 cases of Shwachman–Diamond syndrome – ten with and three without SBDS gene mutation – and ten controls were investigated by confocal microscopy and ultrastructural immunocytochemistry using antibodies against ubiquitinated proteins, proteasomes, p62 protein, and Helicobacter pylori VacA, urease and outer membrane proteins. Results Many extensively disseminated particulate cytoplasmic structures, accounting for 22.78±5.57% (mean ± standard deviation) of the total cytoplasm, were found in blood neutrophils from mutated Shwachman–Diamond syndrome patients. The particulate cytoplasmic structures showed immunoreactivity for polyubiquitinated proteins and proteasomes, but no reactivity for Helicobacter pylori products, which are present in particulate cytoplasmic structures of Helicobacter pylori-positive gastritis. Neutrophils from patients with Shwachman–Diamond syndrome frequently showed p62-positive autophagic vacuoles and apoptotic changes in 5% of cells. No particulate cytoplasmic structures were observed in most control neutrophils; however, in a few cells from two cases we noted focal development of minute particulate cytoplasmic structures, accounting for 0.74±0.56% of the total cytoplasm (P<0.001 versus particulate cytoplasmic structures from mutated Shwachman–Diamond syndrome patients). Neutrophils from non-mutated Shwachman–Diamond-syndrome-like patients resembled controls in two cases, and a third case showed particulate cytoplasmic structure patterns intermediate between those in controls and

  14. Dynamic interactions of neutrophils and biofilms

    Directory of Open Access Journals (Sweden)

    Josefine Hirschfeld

    2014-12-01

    Full Text Available Background: The majority of microbial infections in humans are biofilm-associated and difficult to treat, as biofilms are highly resistant to antimicrobial agents and protect themselves from external threats in various ways. Biofilms are tenaciously attached to surfaces and impede the ability of host defense molecules and cells to penetrate them. On the other hand, some biofilms are beneficial for the host and contain protective microorganisms. Microbes in biofilms express pathogen-associated molecular patterns and epitopes that can be recognized by innate immune cells and opsonins, leading to activation of neutrophils and other leukocytes. Neutrophils are part of the first line of defense and have multiple antimicrobial strategies allowing them to attack pathogenic biofilms. Objective/design: In this paper, interaction modes of neutrophils with biofilms are reviewed. Antimicrobial strategies of neutrophils and the counteractions of the biofilm communities, with special attention to oral biofilms, are presented. Moreover, possible adverse effects of neutrophil activity and their biofilm-promoting side effects are discussed. Results/conclusion: Biofilms are partially, but not entirely, protected against neutrophil assault, which include the processes of phagocytosis, degranulation, and formation of neutrophil extracellular traps. However, virulence factors of microorganisms, microbial composition, and properties of the extracellular matrix determine whether a biofilm and subsequent microbial spread can be controlled by neutrophils and other host defense factors. Besides, neutrophils may inadvertently contribute to the physical and ecological stability of biofilms by promoting selection of more resistant strains. Moreover, neutrophil enzymes can degrade collagen and other proteins and, as a result, cause harm to the host tissues. These parameters could be crucial factors in the onset of periodontal inflammation and the subsequent tissue breakdown.

  15. Effects of bovine viral diarrhea viruses in vitro on transcription of interferon-al- pha, beta, gamma mRNA in bovine peripheral blood mononuclear cells%牛病毒性腹泻病毒感染牛外周血单核细胞对IFN-α、β、γmRNA转录的影响

    Institute of Scientific and Technical Information of China (English)

    韩猛立; 黄新; 钟发刚

    2012-01-01

    The study was done to survery the interferon-alpha, beta and gamma mRNA transcription profiles of bovine viral diarrfea viruse(BVDV) infection,and to investigate the host-BVDV interaction. The clinically healthy Holstein cows tested negative for bovine viral diarrhea virus(BVDV) in peripheral blood mononuclear cells(PBMC) were in- fected with noncytopathic(NCP) and cytopathic(NCP) BVDV. The mRNA levels of IFN-α,β and γ genes were ana lyzed using a reaPtime fluorescent quantitative PCR(reaPtime FQ-PCR). The results indicated that the transcription of I type(IFN-α,β) mRNA showed a different increasing levels (P〈0.01) ,after infected CP- and NCP BVDV in PBMC;only IFN-α decreased at 4,12 h(P〈0. 05) after infected CP-BVDV. And IFN-γ was increased throughout the infection process of CP and NCP BVDV in PBMC (P〈0. 05). The transcription levels of IFN mRNA were in- creased when two biotype of BVDV infected in PBMC.%为了解牛病毒性腹泻病毒(BVDV)感染对干扰素(IFN)mRNA转录时相的影响,探讨宿主-病毒之间的相互关系,用非致细胞病变(noncytopathic,NCP)和致细胞病变(cytopathic,CP)型BVDV感染临床健康BVDV检测阴性的荷斯坦奶牛外周血单核细胞(PBMC),利用实时荧光定量PCR技术对感染后IFN-α、β、γmRNA转录水平的变化进行定量分析。结果表明,CP型和NCP型BVDV感染PBMC后,Ⅰ型IFN(IFN-α、β)均呈现出不同程度的转录水平上调,且差异极显著(P〈0.01);只有IFN-α在CP型BVDV感染后4,12h(P〈0.5)出现转录下调。IFN-γ在整个感染过程中均呈现出不同程度的转录水平上调,且差异显著(P〈0.05)。这表明2种生物型BVDV感染可引起PBMC中IFN mRNA转录水平升高。

  16. Comparison of the neutrophil proteome in trauma patients and normal controls

    DEFF Research Database (Denmark)

    Teles, Liz M B; Aquino, Elaine N; Neves, Anne C D;

    2012-01-01

    Neutrophils have an impressive array of microbicidal weapons, and in the presence of a pathogen, progress from a quiescent state in the bloodstream to a completely activated state. Failure to regulate this activation, for example, when the blood is flooded with cytokines after severe trauma, causes...

  17. Monocytic cell differentiation from band-stage neutrophils under inflammatory conditions via MKK6 activation

    NARCIS (Netherlands)

    Koffel, R.; Meshcheryakova, A.; Warszawska, J.; Hennig, A.; Wagner, K.; Jorgl, A.; Gubi, D.; Moser, D.; Hladik, A.; Hoffmann, U.; Fischer, M.B.; Berg, W.B. van den; Koenders, M.I.; Scheinecker, C.; Gesslbauer, B.; Knapp, S.; Strobl, H.

    2014-01-01

    During inflammation, neutrophils are rapidly mobilized from the bone marrow storage pool into peripheral blood (PB) to enter lesional sites, where most rapidly undergo apoptosis. Monocytes constitute a second wave of inflammatory immigrates, giving rise to long-lived macrophages and dendritic cell s

  18. Modulation of human immune responses by bovine interleukin-10.

    Directory of Open Access Journals (Sweden)

    Gerco den Hartog

    Full Text Available Cytokines can be functionally active across species barriers. Bovine IL-10 has an amino acid sequence identity with human IL-10 of 76.8%. Therefore, the aim of this study was to evaluate whether bovine IL-10 has immunomodulatory activities on human monocytes and dendritic cells. Peripheral blood monocytes were isolated from healthy donors, and used directly or allowed to differentiate to dendritic cells under the influence of IL-4 and GM-CSF. Recombinant bovine IL-10 inhibited TLR induced activation of monocytes, and dose-dependently inhibited LPS-induced activation of monocyte-derived DCs comparable to human IL-10. By using blocking antibodies to either bovine IL-10 or the human IL-10 receptor it was demonstrated that inhibition of monocyte activation by bovine IL-10 was dependent on binding of bovine IL-10 to the human IL-10R. These data demonstrate that bovine IL-10 potently inhibits the activation of human myeloid cells in response to TLR activation. Bovine IL-10 present in dairy products may thus potentially contribute to the prevention of necrotizing enterocolitis and allergy, enhance mucosal tolerance induction and decrease intestinal inflammation and may therefore be applicable in infant foods and in immunomodulatory diets.

  19. Bovine Herpesvirus 4 infections and bovine mastitis

    NARCIS (Netherlands)

    Wellenberg, Gerardus Johannus

    2002-01-01

    Mastitis is an often occurring disease in dairy cattle with an enormous economic impact for milk producers worldwide. Despite intensive research, which is historically based on the detection of bacterial udder pathogens, still around 20-35% of clinical cases of bovine mastitis have an unknown aetiol

  20. Neutrophil Extracellular Traps and Microcrystals

    Science.gov (United States)

    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

  1. Combining Immature and Total Neutrophil Counts to Predict Early Onset Sepsis in Term and Late Preterm Newborns: Use of the I/T2

    Science.gov (United States)

    Newman, Thomas B.; Draper, David; Puopolo, Karen M.; Wi, Soora; Escobar, Gabriel J.

    2014-01-01

    Background The absolute neutrophil count and the immature/total neutrophil ratio (I/T) provide information about the risk of early-onset sepsis in newborns. However, it is not clear how to combine their potentially overlapping information into a single likelihood ratio. Methods We obtained electronic records of blood cultures and of complete blood counts with manual differentials drawn 4 hours of age and when the pretest probability of infection is close to the treatment threshold. PMID:24503598

  2. Candida albicans escapes from mouse neutrophils.

    Science.gov (United States)

    Ermert, David; Niemiec, Maria J; Röhm, Marc; Glenthøj, Andreas; Borregaard, Niels; Urban, Constantin F

    2013-08-01

    Candida albicans, the most commonly isolated human fungal pathogen, is able to grow as budding yeasts or filamentous forms, such as hyphae. The ability to switch morphology has been attributed a crucial role for the pathogenesis of C. albicans. To mimic disseminated candidiasis in humans, the mouse is the most widely used model organism. Neutrophils are essential immune cells to prevent opportunistic mycoses. To explore potential differences between the rodent infection model and the human host, we compared the interactions of C. albicans with neutrophil granulocytes from mice and humans. We revealed that murine neutrophils exhibited a significantly lower ability to kill C. albicans than their human counterparts. Strikingly, C. albicans yeast cells formed germ tubes upon internalization by murine neutrophils, eventually rupturing the neutrophil membrane and thereby, killing the phagocyte. On the contrary, growth and subsequent escape of C. albicans are blocked inside human neutrophils. According to our findings, this blockage in human neutrophils might be a result of higher levels of MPO activity and the presence of α-defensins. We therefore outline differences in antifungal immune defense between humans and mouse strains, which facilitates a more accurate interpretation of in vivo results.

  3. 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.

    Science.gov (United States)

    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.

  4. Utilización del Método de Elisa en la detección directa de antígeno de virus diarrea viral bovina en muestras de suero sanguíneo de bovinos Use of an ELISA test in the direct diagnosis of viral antigens of Bovine Viral Diarrhea Virus (BVDV in bovine blood serum samples

    Directory of Open Access Journals (Sweden)

    G. REINHARDT

    2003-01-01

    Full Text Available El virus de la Diarrea Viral Bovina (VDVB es un agente infeccioso importante del ganado bovino y está distribuido ampliamente en el mundo, produciendo pérdidas económicas sustanciales en la producción pecuaria. La principal fuente de contagio de los animales susceptibles está en las secreciones y excreciones de los animales infectados persistentes e inmunotolerantes (PI, condición que se produce en la etapa gestacional, específicamente antes de los 120 días de preñez, período en que el sistema inmune del embrión aún no se desarrolla adecuadamente. El propósito de este estudio fue aplicar la utilización de un método inmunoenzimático (ELISA-antígeno para detectar la presencia de animales PI en planteles lecheros de la Xª Región de Chile, a partir de muestras de suero sanguíneo. Para ello se examinaron 335 sueros de bovinos provenientes de 9 predios. Los resultados obtenidos indican que el 33.3% de los planteles analizados presentaron algún animal PI y que a nivel de prevalencia intrapredial, ella varió entre 0.7 y 1.0%. Se concluyó que el método utilizado permite detectar animales PI en forma rápida y sencilla, pudiendo utilizarse en gran cantidad de muestrasBVDV is an important virus of cattle worldwide that induces to substantial economic losses in dairy farms. The major source of infection are secretions and excretions of immunotolerant and persistent infected cattle. That condition is adquired during the early gestational period. The scope of this communication is to inform the use of an ELISA test to detect BVDV persistent infected bovine using blood serum samples in cattle of 9 dairy farms from the Xth. Region of Chile. The results indicated that 0.3% of the serum samples were positive to the ELISA test, and 33.3% of the dairy herds with persistently infected animals. It is concluded that this method diagnose persistently infected cattle, and is very easy to manipulate therefore, is possible to test many animals in

  5. Neutrophil granules in health and disease

    DEFF Research Database (Denmark)

    Häger, M; Cowland, J B; Borregaard, N

    2010-01-01

    Neutrophil granules store proteins that are critically important for the neutrophil to move from the vascular bed to tissues and to kill microorganisms. This is illustrated in nature when individual proteins are deleted due to inherited mutations of their cognate genes, and such deficiencies result...... in the conditions leucocyte adhesion deficiency and chronic granulomatous disease. The granules of the neutrophil have traditionally been divided into two or three major types but are instead a continuum where several subtypes can be identified with differences in protein content and propensity for mobilization...

  6. The Predictive Value of Total Neutrophil Count and Neutrophil/ Lymphocyte Ratio in Predicting In-hospital Mortality and Complications after STEMI

    Directory of Open Access Journals (Sweden)

    Samad Ghaffari

    2014-03-01

    Full Text Available Introduction: Leukocytosis, predominantly neutrophilia, has previously been described following ST elevation myocardial infarction (STEMI. The exact contribution of this phenomenon to the clinical outcome of STEMI is yet to be shown. We examined cellular inflammatory response to STEMI in the blood and its association with in-hospital mortality and/or adverse clinical events.Methods: In this cross-sectional study, 404 patients who were admitted with the diagnosis of acute STEMI at Madani Heart Hospital from March 2010 to March 2012 were studied. The complete blood cell count (CBC was obtained from all patientswithin12-24 hours of the onset of symptoms. Total leukocytes were counted and differential count was obtained for neutrophils, lymphocytes and neutrophil/lymphocyte ratio (NLR were evaluated. Association of cellular response with the incidence of post-MI mortality/complications was assessed by multiple logistic regression analyses.Results: In-hospital mortality and post-STEMI complication rate were 3.7% and 43.6%, respectively. Higher age (P=0.04, female gender (0.002, lower ejection fraction (P<0.001 and absolute neutrophil count (P=0.04 were predictors of mortality. Pump failure in the form of acute pulmonary edema or cardiogenic shock occurred in 35 (8.9% of patients. Higher leukocyte (P<0.03 and neutrophil counts (P<0.03 and higher NLR (P=0.01 were predictors of failure. The frequency of ventricular tachyarrhythmias (VT/VF at the first day was associated with higher neutrophil count (P<0.001 and higher NLR level (P<0.001. In multivariate analysis neutrophil count was an independent predictor of mortality (OR=2.94; 1.1-8.4, P=0.04, and neutrophil count [OR=1.1, CI (1.01-1.20, P=0.02], female gender [OR=2.34, CI (1.02-4.88, P=0.04] and diabetes [OR=2.52, CI (1.21-5.2, P=0.003] were independent predictors of heart failure.Conclusion: A single CBC analysis may help to identify STEMI patients at risk for mortality and heart failure, and total

  7. Neutrophils recruited to sites of infection protect from virus challenge by releasing neutrophil extracellular traps.

    Science.gov (United States)

    Jenne, Craig N; Wong, Connie H Y; Zemp, Franz J; McDonald, Braedon; Rahman, Masmudur M; Forsyth, Peter A; McFadden, Grant; Kubes, Paul

    2013-02-13

    Neutrophils mediate bacterial clearance through various mechanisms, including the release of mesh-like DNA structures or neutrophil extracellular traps (NETs) that capture bacteria. Although neutrophils are also recruited to sites of viral infection, their role in antiviral innate immunity is less clear. We show that systemic administration of virus analogs or poxvirus infection induces neutrophil recruitment to the liver microvasculature and the release of NETs that protect host cells from virus infection. After systemic intravenous poxvirus challenge, mice exhibit thrombocytopenia and the recruitment of both neutrophils and platelets to the liver vasculature. Circulating platelets interact with, roll along, and adhere to the surface of adherent neutrophils, forming large, dynamic aggregates. These interactions facilitate the release of NETs within the liver vasculature that are able to protect host cells from poxvirus infection. These findings highlight the role of NETs and early tissue-wide responses in preventing viral infection.

  8. Flow Cytometric Evaluation of Human Neutrophil Apoptosis During Nitric Oxide Generation In Vitro: The Role of Exogenous Antioxidants

    Directory of Open Access Journals (Sweden)

    Zofia Sulowska

    2005-01-01

    in vitro. The effect of exogenous supply of NO donors such as SNP, SIN-1, and GEA-3162 on the course of human neutrophil apoptosis and the role of extracellular antioxidants in this process was investigated. Isolated from peripheral blood, neutrophils were cultured in the presence or absence of NO donor compounds and antioxidants for 8, 12, and 20 hours. Apoptosis of neutrophils was determined in vitro by flow cytometric analysis of cellular DNA content and Annexin V protein binding to the cell surface. Exposure of human neutrophils to GEA-3162 and SIN-1 significantly accelerates and enhances their apoptosis in vitro in a time-dependent fashion. In the presence of SNP, intensification of apoptosis has not been revealed until 12 hours after the culture. The inhibition of GEA-3162- and SIN-1-mediated neutrophil apoptosis by superoxide dismutase (SOD but not by catalase (CAT was observed. Our results show that SOD and CAT can protect neutrophils against NO-donors-induced apoptosis and suggest that the interaction of NO and oxygen metabolites signals may determine the destructive or protective role of NO donor compounds during apoptotic neutrophil death.

  9. DETECTION OF A NEUTROPHIL CHEMOTACTIC FACTOR IN JAPANESE ENCEPHALITIS PATIENTS

    Directory of Open Access Journals (Sweden)

    Aditi Singh

    2012-12-01

    Full Text Available Japanese encephalitis (JE one of the most common cause of acute encephalitis in tropical regions, has generated much public anxiety in India. An early influx of macrophages followed by neutrophils at the site of injury in different organs in humans and mice has previously been reported. It correlated with production of a neutrophil chemotactic protein derived from macrophages. In the present study out of a total of 324 acute encephalitic patients, admitted in Gandhi memorial and associated hospitals, Lucknow, 121 patients with one or more indicators of JE virus infection were included. Significant pleocytosis (mean TLC value of 126+52 cells / mm3 in CSF and leucocytosis (>11,000 cells/mm3 in peripheral blood was observed at the time of admission. The leucocytosis increased significantly during second week in 67% of patients. The peripheral blood mononuclear cells culture done on alternate days was tested for chemotactic activity (hMDF, which was observed to be highest in second week of illness. The direct detection of hMDF in circulation by dot blot was positive in 92% of acute serum samples, with negligible (12.5% reactivity for convalescent sera. A correlation between the hMDF levels and severity of illness has also been observed.

  10. SEXUAL DIMORPHISM IN MORPHOLOGY AND MORPHOMETRY OF NEUTROPHIL DRUMSTICKS

    Directory of Open Access Journals (Sweden)

    Sharmila Tupakula

    2014-12-01

    Full Text Available Introduction: Sex chromatin is a chromatin mass of 1 micron size usually seen at the periphery of nucleus in females. In the literature majority reported its absence in males while few reported its low incidence in males. The term ‘sex chromatin’ comprises two superficially dissimilar structures the “Barr body” present in epithelial and other tissue cells and the “Drumstick” of the polymorphonuclear leucocytes. Materials and methods: The present study was conducted to observe the morphology, morphometry and percentage incidence of Drumsticks in the blood neutrophils of 110 individuals ranging from 17-30 age group and both sexes using a calibrated ocular/eye piece micrometer. Results: The percentage incidence of drumsticks including non-specific appendages as well as the total number of true drumsticks in females exceeds that in males. Four different types of nonspecific appendages-sessile nodules, racket structures, minor lobes and small clubs were found in the blood neutrophils along with the drumsticks. A higher percentage of non-specific appendages i.e. minor lobes (46.2%, racket structures (42.3%, and small clubs (11.5% were observed in males and sessile nodules were found only in females. Conclusion: Observations on morphology, morphometry and percentage incidence of polymorphonuclear drumsticks presented a valuable data on sex differences.

  11. [Toxinology of bovine paraplegic syndrome].

    Science.gov (United States)

    Sevcik, C; Brito, J C; D'Suze, G; Mijares, A J; Domínguez, M G

    1993-01-01

    A clinical entity named "Bovine Paraplegic Syndrome" ("Síndrome Parapléjico de los Bovinos") has spread alarmingly, in the cattle growing areas of the central and eastern plains of Venezuela. Approximately four million cattle are bread in the area were the disease occurs. The mortality index due to the disease ranges 5 to 25% of the animals at risk, mostly cows, pregnant or lactating. The principal characteristic of the bovine paraplegic syndrome is decubitus, ventral or sternal, in animals that make vane efforts to stand when stimulated. The diagnosis is established ruling out, clinically and with laboratory findings, that the animals are suffering known diseases with similar symptoms such as paralytic rabies, botulism and blood parasites such Trypanosoma sp., Babesia sp., and Anaplasma sp.. Death occurs always, usually after few days, and to this date there is no known treatment able to save the sick cows. In this work, we describe results that suggest the presence of a toxin in the cattle suffering and prone to suffer the syndrome; it is a natural toxin produced by ruminal bacteria. In squid giant axons under voltage clamp conditions, this toxin is very specific to block sodium current during nerve electrical activity.

  12. Omega-3 Fatty acids and inflammation: novel interactions reveal a new step in neutrophil recruitment.

    Directory of Open Access Journals (Sweden)

    Samantha P Tull

    2009-08-01

    Full Text Available Inflammation is a physiological response to tissue trauma or infection, but leukocytes, which are the effector cells of the inflammatory process, have powerful tissue remodelling capabilities. Thus, to ensure their precise localisation, passage of leukocytes from the blood into inflamed tissue is tightly regulated. Recruitment of blood borne neutrophils to the tissue stroma occurs during early inflammation. In this process, peptide agonists of the chemokine family are assumed to provide a chemotactic stimulus capable of supporting the migration of neutrophils across vascular endothelial cells, through the basement membrane of the vessel wall, and out into the tissue stroma. Here, we show that, although an initial chemokine stimulus is essential for the recruitment of flowing neutrophils by endothelial cells stimulated with the inflammatory cytokine tumour necrosis factor-alpha, transit of the endothelial monolayer is regulated by an additional and downstream stimulus. This signal is supplied by the metabolism of the omega-6-polyunsaturated fatty acid (n-6-PUFA, arachidonic acid, into the eicosanoid prostaglandin-D(2 (PGD(2 by cyclooxygenase (COX enzymes. This new step in the neutrophil recruitment process was revealed when the dietary n-3-PUFA, eicosapentaenoic acid (EPA, was utilised as an alternative substrate for COX enzymes, leading to the generation of PGD(3. This alternative series eicosanoid inhibited the migration of neutrophils across endothelial cells by antagonising the PGD(2 receptor. Here, we describe a new step in the neutrophil recruitment process that relies upon a lipid-mediated signal to regulate the migration of neutrophils across endothelial cells. PGD(2 signalling is subordinate to the chemokine-mediated activation of neutrophils, but without the sequential delivery of this signal, neutrophils fail to penetrate the endothelial cell monolayer. Importantly, the ability of the dietary n-3-PUFA, EPA, to inhibit this process not

  13. Phagocytic and oxidative burst activity of neutrophils in the end stage of liver cirrhosis

    Institute of Scientific and Technical Information of China (English)

    Anatol Panasiuk; Jolanta Wysocka; Elzbieta Maciorkowska; Bozena Panasiuk; Danuta Prokopowicz; Janusz Zak; Karol Radomski

    2005-01-01

    AIM: To evaluate the phagocytic activity and neutrophil oxidative burst in liver cirrhosis.METHODS: In 45 patients with advanced postalcoholic liver cirrhosis (aged 45±14 years) and in 25 healthy volunteers (aged 38±5 years), the percentage of phagocytizing cells after in vitro incubation with E. coli (Phagotest Kit), phagocytic activity (mean intensity of fluorescence, MIF) and the percentage of neutrophil oxidative burst (Bursttest Kit), and the level of free oxygen radical production (MIF of Rodamine 123)were analyzed by flow cytometry. The levels of soluble sICAM-1, sVCAM-1, sP-selectin, sE-selectin, sL-selectin,and TNF-α were determined in blood serum.RESULTS: The percentage of E. coli phagocytizing neutrophils in liver cirrhosis patients was comparable to that in healthy subjects. MIF of neutrophil - ingested E. coli was higher in patients with liver cirrhosis. The oxidative burst in E. coli phagocytizing neutrophils generated less amount of active oxygen compounds in liver cirrhosis patients (MIF of R123:24.7±7.1 and 29.7±6.6 in healthy,P<0.01). Phorbol myristate acetate (PMA) - stimulated neutrophilsproduced less reactive oxidants in liver cirrhosis patients than in healthy subjects (MIF of R123: 42.7±14.6 vs 50.2±13.3, P<0.01). A negative correlation was observed between oxidative burst MIF of PMA-stimulated neutrophils and ALT and AST levels (r -0.35, P<0.05;r-0.4, P<0.03). sVCAM-1, sICAM-1, sE-selectin concentrations correlated negatively with the oxygen free radical production (MIF of R123) in neutrophils after PMA stimulation in liver cirrhosis patients (r-0.45, P<0.05;r-0.41, P<0.05; r-0.39, P<0.05, respectively).CONCLUSION: Neutrophil metabolic activity diminishes together with the intensification of liver failure. The metabolic potential of phagocytizing neutrophils is significantly lower in liver cirrhosis patients, which can be one of the causes of immune mechanism damage. The evaluation of oxygen metabolism of E. coli

  14. Neutrophil-mediated phagocytosis of Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Jos A.G. Van Strijp

    2014-09-01

    Full Text Available For invading staphylococci, phagocytosis an killing bij human neutrophils is the biggest threat. Neutrophils are the only cells that can effectively kill staphylococci by engulfment and subsequent bombardment with proteases, amidases, antimicrobial peptides and proteins in concert with reactive oxygen species that are generated during the metabolic burst.Both complement and antibodies are crucial for effective uptake and neutrophil activation. S. aureus is not an innocent bystander in this process. It actively secretes several proteins to impair every single step in this process from receptor modulation, to complement inhibition to neutrophil lysis to protease, antimicrobial peptide inhibition and resistance to reactive oxygen species. For the design of future novel antimicrobial strategies: therapeutic antibodies, vaccines, novel antibiotics, all this should be taken into account. Still the best way to treat diseases is to help to enhance the natural defence mechanism that are already in place.

  15. Store-operated calcium signaling in neutrophils.

    Science.gov (United States)

    Clemens, Regina A; Lowell, Clifford A

    2015-10-01

    Calcium signals in neutrophils are initiated by a variety of cell-surface receptors, including formyl peptide and other GPCRs, FcRs, and integrins. The predominant pathway by which calcium enters immune cells is termed SOCE, whereby plasma membrane CRAC channels allow influx of extracellular calcium into the cytoplasm when intracellular ER stores are depleted. The identification of 2 key families of SOCE regulators, STIM calcium "sensors" and ORAI calcium channels, has allowed for genetic manipulation of SOCE pathways and provided valuable insight into the molecular mechanism of calcium signaling in immune cells, including neutrophils. This review focuses on our current knowledge of the molecules involved in neutrophil SOCE and how study of these molecules has further informed our understanding of the role of calcium signaling in neutrophil activation.

  16. Involvement of BLT1 endocytosis and Yes kinase activation in leukotriene B4-induced neutrophil degranulation.

    Science.gov (United States)

    Gaudreault, Eric; Thompson, Charles; Stankova, Jana; Rola-Pleszczynski, Marek

    2005-03-15

    One of the important biological activities of human neutrophils is degranulation, which can be induced by leukotriene B4 (LTB4). Here we investigated the intracellular signaling events involved in neutrophil degranulation mediated by the high affinity LTB4 receptor, BLT1. Peripheral blood neutrophils as well as the promyeloid PLB-985 cell line, stably transfected with BLT1 cDNA and differentiated into a neutrophil-like cell phenotype, were used throughout this study. LTB4-induced enzyme release was inhibited by 50-80% when cells were pretreated with the pharmacological inhibitors of endocytosis sucrose, Con A and NH4Cl. In addition, transient transfection with a dominant negative form of dynamin (K44A) resulted in approximately 70% inhibition of ligand-induced degranulation. Pretreating neutrophils or BLT1-expressing PLB-985 cells with the Src family kinase inhibitor PP1 resulted in a 30-60% inhibition in BLT1-mediated degranulation. Yes kinase, but not c-Src, Fgr, Hck, or Lyn, was found to exhibit up-regulated kinase activity after LTB4 stimulation. Moreover, BLT1 endocytosis was found to be necessary for Yes kinase activation in neutrophils. LTB4-induced degranulation was also sensitive to inhibition of PI3K. In contrast, it was not affected by inhibition of the mitogen-activated protein kinase MEK kinase, the Janus kinases, or the receptor tyrosine kinase epidermal growth factor receptor or platelet-derived growth factor receptor. Taken together, our results suggest an essential role for BLT1 endocytosis and Yes kinase activation in LTB4-mediated degranulation of human neutrophils.

  17. Aqueous extract of Rosmarinus officinalis L. inhibits neutrophil influx and cytokine secretion.

    Science.gov (United States)

    Silva, Ana Mara de Oliveira E; Machado, Isabel Daufenback; Santin, José Roberto; de Melo, Illana Louise Pereira; Pedrosa, Gabriela Vieira; Genovese, Maria Ines; Farsky, Sandra Helena Poliselli; Mancini-Filho, Jorge

    2015-01-01

    Rosmarinus officinalis L. phenolic compounds have attracted considerable attention because of their antioxidant and antimicrobial properties, including its ability to treat inflammatory disorders. In this work, we investigated the in vivo and in vitro effects of R. officinalis aqueous extract on neutrophil trafficking from the blood into an inflamed tissue, on cell-derived secretion of chemical mediators, and on oxidative stress. Anti-inflammatory activity was investigated using carrageenan-induced inflammation in the subcutaneous tissue of male Wistar rats orally treated with the R. officinalis extract (100, 200, or 400 mg/kg). The leukocyte influx (optical microscopy), secretion of chemical mediators (prostaglandin E2 (PGE2), TNF-α, interleukin 6 (IL-6), leukotriene B4 (LTB4), and cytokine-induced neutrophil chemoattractant 1 by enzyme-linked immunosorbent assay), and the anti-oxidative profile (super oxide dismutase (SOD), glutathione peroxidase, and thiobarbituric acid reactive substance (TBARS) spectrophotometry) were quantified in the inflamed exudate. N-Formyl-methionine-leucine-phenylalanine-induced chemotaxis, lipopolysaccharide-induced NO2 (-) production (Greiss reaction), and adhesion molecule expression (flow cytometry) were in vitro quantified using oyster glycogen recruited peritoneal neutrophils previous treated with the extract (1, 10, or 100 µg/mL). Animals orally treated with phosphate-buffered saline and neutrophils incubated with Hank's balanced salt solution were used as control. R. officinalis extract oral treatment caused a dose-dependent reduction in the neutrophil migration as well as decreased SOD, TBARS, LTB4, PGE2, IL-6, and TNF-α levels in the inflamed exudate. In vitro treatment with R. officinalis decreased neutrophil chemotaxis, NO2 (-) production, and shedding of L-selectin and β2 integrin expressions. Results here presented show that R. officinalis aqueous extract displays important in vivo and in vitro anti

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

    Science.gov (United States)

    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

  19. GM-CSF Differentially Regulates Eosinophil and Neutrophil Adhesive Interactions with Vascular Endothelium in Vivo

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    Nooshin Sheikh Bahaie

    2010-12-01

    Full Text Available Allergic airway inflammation is characterized by elaboration of cytokines and chemokines leading to recruitment of inflammatory leukocytes, predominantly eosinophils, to the airways. Granulocyte macrophage colony stimulating factor (GM-CSF is generated in the lungs of human subjects with asthma in response to allergen challenge and is necessary for the development of allergen-induced bronchial eosinophilia in mice. The effect of GM-CSF on human eosinophil and neutrophil interactions with the vascular endothelium under conditions of blood flow was investigated in post-capillary venules of the rabbit mesentery by intravital microscopy.While GM-CSF significantly reduced the rolling fraction of neutrophils in vivo and induced consistent shedding of neutrophil L-selectin in vitro, its effect on eosinophil rolling was variable. Eosinophils from 57% of the donors demonstrated inhibition of rolling, while eosinophils from the remaining 43% of donors demonstrated no inhibition or increased rolling. The variable effect of GM-CSF on inhibition of eosinophil rolling was associated with variable shedding of L-selectin in vitro. In contrast to the differential effect of GM-CSF on neutrophils versus eosinophils, stimulation with phorbol myristate acetate demonstrated a similar degree of inhibition of rolling and L-selectin shedding by neutrophils and eosinophils suggesting that there was no defect in L-selectin shedding in the eosinophil donors who did not respond to GM-CSF. Overall, these studies demonstrate that GM-CSF consistently inhibits interaction of neutrophils with endothelium in vivo, whereas its effect on eosinophil-endothelial interactions is variable. GM-CSF may thus be one factor accounting for the varying percentage of eosinophils and neutrophils recruited to sites of allergic inflammation in different individuals.

  20. Aberrant “Barbed-Wire” Nuclear Projections of Neutrophils in Trisomy 18 (Edwards Syndrome

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    Basil M. Kahwash

    2015-01-01

    Full Text Available 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.

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

    Science.gov (United States)

    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.

  2. Mechanisms of lung neutrophil activation after hemorrhage or endotoxemia: roles of reactive oxygen intermediates, NF-kappa B, and cyclic AMP response element binding protein.

    Science.gov (United States)

    Shenkar, R; Abraham, E

    1999-07-15

    Acute inflammatory lung injury occurs frequently in the setting of severe infection or blood loss. Accumulation of activated neutrophils in the lungs and increased pulmonary proinflammatory cytokine levels are major characteristics of acute lung injury. In the present experiments, we examined mechanisms leading to neutrophil accumulation and activation in the lungs after endotoxemia or hemorrhage. Levels of IL-1 beta, TNF-alpha, and macrophage inflammatory protein-2 mRNA were increased in lung neutrophils from endotoxemic or hemorrhaged mice compared with those present in lung neutrophils from control mice or in peripheral blood neutrophils from endotoxemic, hemorrhaged, or control mice. The transcriptional regulatory factors NF-kappa B and cAMP response element binding protein were activated in lung but not blood neutrophils after hemorrhage or endotoxemia. Xanthine oxidase inhibition, achieved by feeding allopurinol or tungsten-containing diets, did not affect neutrophil trafficking to the lungs after hemorrhage or endotoxemia. Xanthine oxidase inhibition did prevent hemorrhage- but not endotoxemia-induced increases in proinflammatory cytokine expression among lung neutrophils. Hemorrhage- or endotoxemia-associated activation of NF-kappa B in lung neutrophils was not affected by inhibition of xanthine oxidase. cAMP response element binding protein activation was increased after hemorrhage, but not endotoxemia, in mice fed xanthine oxidase-inhibiting diets. Our results indicate that xanthine oxidase modulates cAMP response element binding protein activation and proinflammatory cytokine expression in lung neutrophils after hemorrhage, but not endotoxemia. These findings suggest that the mechanisms leading to acute inflammatory lung injury after hemorrhage differ from those associated with endotoxemia.

  3. Subproteome analysis of the neutrophil cytoskeleton

    OpenAIRE

    Xu, Ping; Crawford, Mark; Way, Michael; Godovac-Zimmermann, Jasminka; Segal, Anthony W.; Radulovic, Marko

    2009-01-01

    Neutrophils play a key role in the early host-defense mechanisms due to their capacity to migrate into inflamed tissues and phagocytose microorganisms. The cytoskeleton has an essential role in these neutrophil functions, however, its composition is still poorly understood. We separately analyzed different cytoskeletal compartments: cytosolic skeleton, phagosome membrane skeleton, and plasma membrane skeleton. Using a proteomic approach, 138 nonredundant proteins were identified. Proteins not...

  4. Cryptococcus neoformans modulates extracellular killing by neutrophils

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    Asfia eQureshi

    2011-09-01

    Full Text Available We recently established a key role for host sphingomyelin synthase (SMS in the regulation of the killing activity of neutrophils against Cryptococcus neoformans. In this work, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and NK cells (Tgε26 mice. To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in vitro and in vivo. We show that unlike C. albicans, neither the presence nor the capsule size of C. neoformans cells have any effect on neutrophil viability. Interestingly, melanized C. neoformans cells totally abrogated the killing activity of neutrophils. Next, we monitored how exposure of neutrophils to C. neoformans cells would interfere with any further killing activity of the medium and found that pre-incubation with live but not heat-killed fungal cells significantly inhibits further killing activity of the medium. We next studied whether activation of SMS at the site of C. neoformans infection is dependent on T and NK cells. Using matrix-assisted laser desorption-ionization (MALDI tissue imaging in infected lung we found that similarly to previous observations in the isogenic wild type CBA/J mice, SM 16:0 levels are significantly elevated at the site of infection in mice lacking T and NK cells but only at early time points. This study highlights that C. neoformans may negatively regulate the killing activity of neutrophils and that SMS activation in neutrophils appears to be partially independent of T and/or NK cells.

  5. Cryptococcus neoformans modulates extracellular killing by neutrophils.

    Science.gov (United States)

    Qureshi, Asfia; Grey, Angus; Rose, Kristie L; Schey, Kevin L; Del Poeta, Maurizio

    2011-01-01

    We recently established a key role for host sphingomyelin synthase (SMS) in regulating the killing activity of neutrophils against Cryptococcus neoformans. In this paper, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and natural killer (NK) cells (Tgε26 mice). To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in vitro and in vivo. We show that unlike Candida albicans, neither the presence nor the capsule size of C. neoformans cells have any effect on neutrophil viability. Interestingly, melanized C. neoformans cells totally abrogated the killing activity of neutrophils. We monitored how exposure of neutrophils to C. neoformans cells would interfere with any further killing activity of the conditioned medium and found that pre-incubation with live but not "heat-killed" fungal cells significantly inhibits further killing activity of the medium. We then studied whether activation of SMS at the site of C. neoformans infection is dependent on T and NK cells. Using matrix-assisted laser desorption-ionization tissue imaging in infected lung we found that similar to previous observations in the isogenic wild-type CBA/J mice, SM 16:0 levels are significantly elevated at the site of infection in mice lacking T and NK cells, but only at early time points. This study highlights that C. neoformans may negatively regulate the killing activity of neutrophils and that SMS activation in neutrophils appears to be partially independent of T and/or NK cells.

  6. Neutrophils in asthma--a review.

    Science.gov (United States)

    Ciepiela, Olga; Ostafin, Magdalena; Demkow, Urszula

    2015-04-01

    Asthma is a chronic inflammatory disease, with an array of cells involved in the pathogenesis of the disease. The role of neutrophils in the development of bronchial asthma is found to be complex, as they may trigger activation of immunocompetent cells and are a potent source of free oxygen radicals and enzymes participating in airway remodeling. The review highlights the role of neutrophils in bronchial asthma.

  7. Neutrophil Responses to Sterile Implant Materials

    OpenAIRE

    Siddharth Jhunjhunwala; Stephanie Aresta-DaSilva; Katherine Tang; David Alvarez; Webber, Matthew J.; Tang, Benjamin C.; Lavin, Danya M.; Omid Veiseh; Doloff, Joshua C; Suman Bose; Arturo Vegas; Minglin Ma; Gaurav Sahay; Alan Chiu; Andrew Bader

    2015-01-01

    In vivo implantation of sterile materials and devices results in a foreign body immune response leading to fibrosis of implanted material. Neutrophils, one of the first immune cells to be recruited to implantation sites, have been suggested to contribute to the establishment of the inflammatory microenvironment that initiates the fibrotic response. However, the precise numbers and roles of neutrophils in response to implanted devices remains unclear. Using a mouse model of peritoneal microcap...

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

    Energy Technology Data Exchange (ETDEWEB)

    Vlasova, Irina I., E-mail: irina.vlasova@yahoo.com [Research Institute for Physico-Chemical Medicine, Federal Medico-Biological Agency, Moscow (Russian Federation); Vakhrusheva, Tatyana V. [Research Institute for Physico-Chemical Medicine, Federal Medico-Biological Agency, Moscow (Russian Federation); Sokolov, Alexey V.; Kostevich, Valeria A. [Research Institute for Physico-Chemical Medicine, Federal Medico-Biological Agency, Moscow (Russian Federation); Research Institute for Experimental Medicine, Russian Academy of Medical Science, Saint Petersburg (Russian Federation); Gusev, Alexandr A.; Gusev, Sergey A. [Research Institute for Physico-Chemical Medicine, Federal Medico-Biological Agency, Moscow (Russian Federation); Melnikova, Viktoriya I. [Institute of Developmental Biology, Russian Academy of Science, Moscow (Russian Federation); Lobach, Anatolii S. [Institute of Problems of Chemical Physics, Russian Academy of Science, Chernogolovka (Russian Federation)

    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

  9. Weaning management of newly received beef calves with or without continuous exposure to a persistently infected bovine viral diarrhea virus pen mate: Effects on rectal temperature, peripheral blood leukocytes and serum

    Science.gov (United States)

    Exposure to animals persistently infected (PI) with bovine viral diarrhea virus (BVDV) results in immunomodulation in cohorts. It is hypothesized that the extent of modulation differs for preconditioned (PC) vs. auction market (AM) cattle. Our objective was to compare immune responses of PC or AM ca...

  10. Functional activity of neutrophils in diabetes mellitus and coronary heart disease: role of myeloperoxidase in the development of oxidative stress.

    Science.gov (United States)

    Gorudko, I V; Kostevich, V A; Sokolov, A V; Shamova, E V; Buko, I V; Konstantinova, E E; Vasiliev, V B; Cherenkevich, S N; Panasenko, O M

    2012-11-01

    We performed a comparative analysis of functional activity of neutrophils in patients with type 2 diabetes mellitus with and without symptoms of CHD. Enhanced H2O2 production by neutrophils in response to N-formyl-Met-Leu-Phe (fMLP) was found in patients with type 2 diabetes mellitus. In patients with type 2 diabetes mellitus associated with CHD, fMLP-induced release of myeloperoxidase from azurophilic granules of neutrophils was reduced and plasma myeloperoxidase level was elevated. Increased peroxidase activity of myeloperoxidase, reduced plasma catalase activity, and increased levels of TBA-reactive lipid peroxidation products and oxidized glutathione were detected in patients of both groups. Since myeloperoxidase is an important neutrophilic mediator of oxidative stress, its increased activity in the blood can be an additional marker of oxidative stress and cardiovascular risk in patients with diabetes mellitus.

  11. NET amyloidogenic backbone in human activated neutrophils.

    Science.gov (United States)

    Pulze, L; Bassani, B; Gini, E; D'Antona, P; Grimaldi, A; Luini, A; Marino, F; Noonan, D M; Tettamanti, G; Valvassori, R; de Eguileor, M

    2016-03-01

    Activated human neutrophils produce a fibrillar DNA network [neutrophil extracellular traps (NETs)] for entrapping and killing bacteria, fungi, protozoa and viruses. Our results suggest that the neutrophil extracellular traps show a resistant amyloidogenic backbone utilized for addressing reputed proteins and DNA against the non-self. The formation of amyloid fibrils in neutrophils is regulated by the imbalance of reactive oxygen species (ROS) in the cytoplasm. The intensity and source of the ROS signal is determinant for promoting stress-associated responses such as amyloidogenesis and closely related events: autophagy, exosome release, activation of the adrenocorticotrophin hormone/α-melanocyte-stimulating hormone (ACTH/α-MSH) loop and synthesis of specific cytokines. These interconnected responses in human activated neutrophils, that have been evaluated from a morphofunctional and quantitative viewpoint, represent primitive, but potent, innate defence mechanisms. In invertebrates, circulating phagocytic immune cells, when activated, show responses similar to those described previously for activated human neutrophils. Invertebrate cells within endoplasmic reticulum cisternae produce a fibrillar material which is then assembled into an amyloidogenic scaffold utilized to convey melanin close to the invader. These findings, in consideration to the critical role played by NET in the development of several pathologies, could explain the structural resistance of these scaffolds and could provide the basis for developing new diagnostic and therapeutic approaches in immunomediated diseases in which the innate branch of the immune system has a pivotal role.

  12. A3R Phage and Staphylococcus aureus Lysate Do Not Induce Neutrophil Degranulation

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    Jan Borysowski

    2017-02-01

    Full Text Available The objective of this study was to evaluate the effects of A3R phage and Staphylococcus aureus lysate obtained after phage infection on neutrophil degranulation. The exocytosis of primary and secondary granules from neutrophils was investigated in vitro in whole blood specimens by flow cytometry based on the expression of specific markers of exocytosis (CD63 for primary granules and CD66b for secondary granules. We found that both A3R and S. aureus lysate had no significant effect on the exocytosis of primary and secondary granules. These data suggest that neither A3R virions nor any products of phage-induced lysis of S. aureus are likely to induce neutrophil degranulation in patients who are treated with phage preparations. Since neutrophil granules contain some potentially toxic proteins, our results provide an important argument for the safety of phage therapy. Moreover, these data indicate that the induction of neutrophil degranulation is not likely to contribute to antibacterial effects of phages.

  13. Arachidonic acid metabolism in the platelets and neutrophils of diabetic rabbit and human subjects

    Energy Technology Data Exchange (ETDEWEB)

    Greco, N.J.

    1985-01-01

    An alteration of arachidonic acid metabolism to prostaglandins and leukotrienes from platelets and polymorphonuclear leukocytes respectively is evident in subjects with diabetes mellitus. There is evidence of altered platelet/vascular wall interactions in diabetes mellitus and evidence that polymorphonuclear leukocytes influence the vascular walls. Theories on the pathogenesis of atherosclerosis include both blood cells. Platelet hypersensitivity is evident in those platelets from the alloxan-induced diabetic rabbit either suspended in plasma or buffer. Arachidonic acid- and collagen-induced platelet aggregation, release of /sup 14/serotonin, and T x B/sub 2/ and 12-HETE production is enhanced when responses of diabetic platelets are compared to control platelets. Control rabbit neutrophils produce more LTB/sub 4/, LTB/sub 4/ isomers and 5-HETE than diabetic rabbits neutrophils. Decreased synthesis from diabetic rabbit neutrophils is not explained by increased catabolism of LTB/sub 4/, reesterification of 5-HETE, or increased eicosanoid formation. These experiments demonstrate both platelet and neutrophil dysfunction in diabetic subjects. Because of the involvement of these cells in regulating circulatory homeostatis, abnormal behavior could aggravate the atherosclerotic process. Platelet and neutrophil dysfunctions are noted before macroscopic vascular lesions are apparent suggesting an important role in the pathogenesis of atherosclerosis.

  14. A3R Phage and Staphylococcus aureus Lysate Do Not Induce Neutrophil Degranulation

    Science.gov (United States)

    Borysowski, Jan; Międzybrodzki, Ryszard; Wierzbicki, Piotr; Kłosowska, Danuta; Korczak-Kowalska, Grażyna; Weber-Dąbrowska, Beata; Górski, Andrzej

    2017-01-01

    The objective of this study was to evaluate the effects of A3R phage and Staphylococcus aureus lysate obtained after phage infection on neutrophil degranulation. The exocytosis of primary and secondary granules from neutrophils was investigated in vitro in whole blood specimens by flow cytometry based on the expression of specific markers of exocytosis (CD63 for primary granules and CD66b for secondary granules). We found that both A3R and S. aureus lysate had no significant effect on the exocytosis of primary and secondary granules. These data suggest that neither A3R virions nor any products of phage-induced lysis of S. aureus are likely to induce neutrophil degranulation in patients who are treated with phage preparations. Since neutrophil granules contain some potentially toxic proteins, our results provide an important argument for the safety of phage therapy. Moreover, these data indicate that the induction of neutrophil degranulation is not likely to contribute to antibacterial effects of phages. PMID:28230780

  15. Expression and role of a2 vacuolar-ATPase (a2V) in trafficking of human neutrophil granules and exocytosis.

    Science.gov (United States)

    Gilman-Sachs, Alice; Tikoo, Anjali; Akman-Anderson, Leyla; Jaiswal, Mukesh; Ntrivalas, Evangelos; Beaman, Kenneth

    2015-06-01

    Neutrophils kill microorganisms by inducing exocytosis of granules with antibacterial properties. Four isoforms of the "a" subunit of V-ATPase-a1V, a2V, a3V, and a4V-have been identified. a2V is expressed in white blood cells, that is, on the surface of monocytes or activated lymphocytes. Neutrophil associated-a2V was found on membranes of primary (azurophilic) granules and less often on secondary (specific) granules, tertiary (gelatinase granules), and secretory vesicles. However, it was not found on the surface of resting neutrophils. Following stimulation of neutrophils, primary granules containing a2V as well as CD63 translocated to the surface of the cell because of exocytosis. a2V was also found on the cell surface when the neutrophils were incubated in ammonium chloride buffer (pH 7.4) a weak base. The intracellular pH (cytosol) became alkaline within 5 min after stimulation, and the pH increased from 7.2 to 7.8; this pH change correlated with intragranular acidification of the neutrophil granules. Upon translocation and exocytosis, a2V on the membrane of primary granules remained on the cell surface, but myeloperoxidase was secreted. V-ATPase may have a role in the fusion of the granule membrane with the cell surface membrane before exocytosis. These findings suggest that the granule-associated a2V isoform has a role in maintaining a pH gradient within the cell between the cytosol and granules in neutrophils and also in fusion between the surface and the granules before exocytosis. Because a2V is not found on the surface of resting neutrophils, surface a2V may be useful as a biomarker for activated neutrophils.

  16. Simvastatin antagonizes CD40L secretion, CXC chemokine formation, and pulmonary infiltration of neutrophils in abdominal sepsis.

    Science.gov (United States)

    Zhang, Su; Rahman, Milladur; Zhang, Songen; Qi, Zhongquan; Thorlacius, Henrik

    2011-05-01

    Statins have been reported to exert anti-inflammatory actions and protect against septic organ dysfunction. Herein, we hypothesized that simvastatin may attenuate neutrophil activation and lung damage in abdominal sepsis. Male C57BL/6 mice were pretreated with simvastatin (0.5 or 10 mg/kg) before CLP. In separate groups, mice received an anti-CD40L antibody or a CXCR2 antagonist (SB225002) prior to CLP. BALF and lung tissue were harvested for analysis of neutrophil infiltration, as well as edema and CXC chemokine formation. Blood was collected for analysis of Mac-1 and CD40L expression on neutrophils and platelets, as well as soluble CD40L in plasma. Simvastatin decreased CLP-induced neutrophil infiltration and edema formation in the lung. Moreover, Mac-1 expression increased on septic neutrophils, which was significantly attenuated by simvastatin. Inhibition of CD40L reduced CLP-induced up-regulation of Mac-1 on neutrophils. Simvastatin prevented CD40L shedding from the surface of platelets and reduced circulating levels of CD40L in septic mice. CXC chemokine-induced migration of neutrophils in vitro was decreased greatly by simvastatin. Moreover, simvastatin abolished CLP-evoked formation of CXC chemokines in the lung, and a CXCR2 antagonist attenuated pulmonary accumulation of neutrophils. Our data suggest that the inhibitory effect of simvastatin on pulmonary accumulation of neutrophils may be related to a reduction of CD40L secretion into the circulation, as well as a decrease in CXC chemokine formation in the lung. Thus, these protective mechanisms help to explain the beneficial actions exerted by statins, such as simvastatin, in sepsis.

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

    Science.gov (United States)

    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 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, the levels of neutrophil activation and inflammation markers were positively associated with generation of ROS and negatively with SOD, indicating a role of oxidative stress in mediating neutrophilic inflammatory response following chronic inhalation of biomass smoke.

  18. Modulation and Apoptosis of Neutrophil Granulocytes by Extracorporeal Photopheresis in the Treatment of Chronic Graft-Versus-Host Disease.

    Directory of Open Access Journals (Sweden)

    Cindy Franklin

    Full Text Available Chronic graft-versus-host disease (cGVHD is a common side effect of allogeneic stem cell transplantation and a major cause of morbidity and mortality in affected patients. Especially skin, eyes and oral mucosa are affected. This can lead to pain and functional impairment. Extracorporeal photopheresis (ECP is an effective immunomodulatory therapy with minimal side effects but its mode of action is still largely unknown. The objective of the present study was to examine the effects of ECP on neutrophil granulocytes in patients with cGVHD. Analysis of leukocytes from cGVHD patients obtained from the ECP device during treatment showed that neutrophil granulocytes account for the majority of cells treated during ECP. Neutrophils from healthy donors treated in vitro with 8-methoxypsoralen and UVA light as well as neutrophils from buffy coats of patients with cGVHD treated by ECP showed increased apoptosis and decreased half-life. In remaining non-apoptotic cells chemoirradiation resulted in loss of activation markers and reduced effector functions. This was accompanied by an increase in extracellular arginase-1 activity. Additional comparison of neutrophils isolated from blood of cGVHD patients before and 24h after ECP revealed a decreased half-life and reduction of effector functions of post-ECP neutrophils ex vivo. These observations strongly suggest that ECP induces both apoptosis and physiological changes in neutrophils and that these changes also take place in vivo. This study is the first to show that ECP modulates apoptosis and inflammatory activity in neutrophil granulocytes, indicating that neutrophils may significantly contribute to the overall immunomodulatory effects attributed to this treatment.

  19. DETECTION OF THE BOVINE VIRAL DIARRHEA ANTIBODIES

    Directory of Open Access Journals (Sweden)

    I. V. Goraichuk

    2013-06-01

    Full Text Available Bovine viral diarrhea is a widespread infection of cattle that has a wide range of clinical symptoms in domestic and wild ruminants. It is a major problem in cattle and causes significant economic losses in the cattle industry. The virus infects bovines of all ages and causes both immunosuppression and reproductive, respiratory and digestive disorders. Persistently infected cattle are the main factor in transmission of the disease between and among herds. Comparative results of antibodies presence received by two methods of enzymoimmunoassay and virus neutralization test are given in the paper. During the work, 1010 samples of blood serum of cattle from three farms in the Kharkiv region were selected and analyzed. Bovine viral diarrhea virus concerning antibodies were found by enzymoimmunoassay in 704 samples (69.7% using commercial kit and in 690 samples (68.3% using in house method. After results clarification by virus neutralization test, bovine viral diarrhea antibodies were found in 712 samples (70.5%. Immunoenzyme analysis is recommended for mass screening of cattle for viral diarrhea occurrence. The results confirm that the sensitivity immunoenzyme analysis satisfies the requirements of the diagnostic methods. Using the neutralization reaction of viruses as the «gold standard» of serological methods, it is appropriate to clarify the results of immunoenzyme analysis. Since the results contain a signi ficant number of false positive results, it is necessary to carry out comprehensive studies using both serological and molecular genetics methods.

  20. Neutrophil extracellular trap formation is associated with IL-1β and autophagy-related signaling in gout.

    Directory of Open Access Journals (Sweden)

    Ioannis Mitroulis

    Full Text Available BACKGROUND: Gout is a prevalent inflammatory arthritis affecting 1-2% of adults characterized by activation of innate immune cells by monosodium urate (MSU crystals resulting in the secretion of interleukin-1β (IL-1β. Since neutrophils play a major role in gout we sought to determine whether their activation may involve the formation of proinflammatory neutrophil extracellular traps (NETs in relation to autophagy and IL-1β. METHODOLOGY/PRINCIPAL FINDINGS: Synovial fluid neutrophils from six patients with gout crisis and peripheral blood neutrophils from six patients with acute gout and six control subjects were isolated. MSU crystals, as well as synovial fluid or serum obtained from patients with acute gout, were used for the treatment of control neutrophils. NET formation was assessed using immunofluorescence microscopy. MSU crystals or synovial fluid or serum from patients induced NET formation in control neutrophils. Importantly, NET production was observed in neutrophils isolated from synovial fluid or peripheral blood from patients with acute gout. NETs contained the alarmin high mobility group box 1 (HMGB1 supporting their pro-inflammatory potential. Inhibition of phosphatidylinositol 3-kinase signaling or phagolysosomal fusion prevented NET formation, implicating autophagy in this process. NET formation was driven at least in part by IL-1β as demonstrated by experiments involving IL-1β and its inhibitor anakinra. CONCLUSIONS/SIGNIFICANCE: These findings document for the first time that activation of neutrophils in gout is associated with the formation of proinflammatory NETs and links this process to both autophagy and IL-1β. Modulation of the autophagic machinery may represent an additional therapeutic study in crystalline arthritides.

  1. Differential Ability of Bovine Antimicrobial Cathelicidins to Mediate Nucleic Acid Sensing by Epithelial Cells

    Science.gov (United States)

    Baumann, Arnaud; Kiener, Mirjam Susanna; Haigh, Brendan; Perreten, Vincent; Summerfield, Artur

    2017-01-01

    Cathelicidins encompass a family of cationic peptides characterized by antimicrobial activity and other functions, such as the ability to enhance the sensing of nucleic acids by the innate immune system. The present study aimed to investigate the ability of the bovine cathelicidins indolicidin, bactenecin (Bac)1, Bac5, bovine myeloid antimicrobial peptide (BMAP)-27, BMAP-28, and BMAP-34 to inhibit the growth of bacteria and to enhance the sensing of nucleic acid by the host’s immune system. BMAP-27 was the most effective at killing Staphylococcus aureus, Streptococcus uberis, and Escherichia coli, and this was dependent on its amphipathic structure and cationic charge. Although most cathelicidins possessed DNA complexing activity, only the alpha-helical BMAP cathelicidins and the cysteine-rich disulfide-bridged Bac1 were able to enhance the sensing of nucleic acids by primary epithelial cells. We also compared these responses with those mediated by neutrophils. Activation of neutrophils with phorbol myristate acetate resulted in degranulation and release of cathelicidins as well as bactericidal activity in the supernatants. However, only supernatants from unstimulated neutrophils were able to promote nucleic acid sensing in epithelial cells. Collectively, the present data support a role for certain bovine cathelicidins in helping the innate immune system to sense nucleic acids. The latter effect is observed at concentrations clearly below those required for direct antimicrobial functions. These findings are relevant in development of future strategies to promote protection at mucosal surfaces against pathogen invasion. PMID:28203238

  2. Sexing Bovine Embryos Using PCR Amplification of Bovine SRY Sequence

    Institute of Scientific and Technical Information of China (English)

    曾溢滔; 张美兰; 陈美珏; 周霞娣; 黄英; 任兆瑞; 黄淑帧; 胡明信; 吴学清; 高建明; 张斌; 徐慧如

    1994-01-01

    This study analyses the bovine SRY DNA sequence by direct sequencing procedure, followed by the designation of the PCR primers specific for bovine SRY. Using PCR amplification of bovine SRY gene, the embryo sex was determined. The results of the embryo sex identification were confirmed after the embryo transfer and pregnancies.

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

    Science.gov (United States)

    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.

  4. Effect of Weightlessness on Neutrophils and Lymphocytes of Rats

    Directory of Open Access Journals (Sweden)

    Khusi Muhammad Saqib, Zia-ur-Rahman1 and Saeed Ahmad Nagra2

    2012-01-01

    Full Text Available In the present study, two hundreds and forty healthy albino young (n=120 and old (n=120 rats were used during winter and summer season. Rats were divided into four groups in each season i.e. young and old, consisting of male (n=30 and female (n=30 in each age category. In each age  sex matched rats, three subgroups were made and have been given the name as cage control (CC group, horizontal restrained group (HR and head down suspended (HDS group. For winter season, the room temperature of experimental period ranged from 20 to 23°C and for summer season, the experimental room temperature ranged from 30 to 33°C. A 12 hours light/12 hours dark cycle with ad libitum food offered each day to an individual rats as well as fresh water (at normal temperature were provided every day from 9-10 h (morning Rats were decapitated on day 7th (n=5 and day 28th (n=5 of experimental period from all groups to collected the blood in a hepranized tubes for the estimation of lymphocytes and neutrophils. Appropriate statistical analysis was performed to estimate the difference between age, days, treatments and their possible interactions during each season. During winter and summer seasons, male and female rats did show a significant decrease in lymphocytes, however a significant increase in the neutrophils percent was also observed in the HR and HDS groups. During summer, a significant increase in neutrophils and a decrease in lymphocytes were observed in male and female rats of HR and HDS groups.

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

    Science.gov (United States)

    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.

  6. Neutrophil superoxide-anion generating capacity in chronic smoking: effect of long-term -tocopherol therapy

    Indian Academy of Sciences (India)

    Lambertus J Hvan Tits; Frouwkje De Waart; Heidi L M Hak-Lemmers; Jacqueline De Graaf; Pierre N M Demacker; Anton F H Stalenhoef

    2003-02-01

    We investigated whether long-term -tocopherol therapy in chronic smoking affects superoxide generating capacity of neutrophils ex vivo. To this purpose, we randomly assigned 128 male chronic smokers (37 ± 21 pack years of smoking) to treatment with placebo ( = 64) or -tocopherol (400 IU dL--tocopherol daily, = 64). After two years of therapy, we measured phorbol 12-myristate 13-acetate-induced superoxide production of isolated neutrophils and of diluted whole blood by monitoring reduction of ferricytochrome and luminolenhanced peroxidase-catalyzed chemiluminescence. Plasma lipids and lipoproteins were not different between the two treatment groups. As expected, concentrations of -tocopherol in plasma and in low-density lipoproteins were markedly elevated in the supplemented group compared to the placebo group (+ 120%, P < 0.0001 and + 83%, < 0.0001, respectively). Consequently, resistance to in vitro oxidation of low-density lipoproteins (reflected by lag time of conjugated diene formation) was higher in the supplemented group than in the placebo group (+ 22%, < 0.0001). Superoxide generating capacity of neutrophils and superoxide production in diluted whole blood did not differ between -tocopherol and placebo group. It is concluded that in chronic smoking long-term supranormal -tocopherol intake does not reduce neutrophil superoxide-anion generating capacity, despite large increases in the concentrations of -tocopherol in plasma and in low-density lipoproteins.

  7. Neutrophilic airways inflammation in lung cancer: the role of exhaled LTB-4 and IL-8

    Directory of Open Access Journals (Sweden)

    Orlando Silvio

    2011-06-01

    Full Text Available Abstract Background Recent advances in lung cancer biology presuppose its inflammatory origin. In this regard, LTB-4 and IL-8 are recognized to play a crucial role in neutrophil recruitment into airways during lung cancer. Notwithstanding the intriguing hypothesis, the exact role of neutrophilic inflammation in tumour biology remains complex and not completely known. The aim of this study was to give our contribution in this field by investigating LTB-4 and IL-8 in the breath condensate of NSCLC patients and verifying their role in cancer development and progression. Method We enrolled 50 NSCLC patients and 35 controls. LTB-4 and IL-8 concentrations were measured in the breath condensate and the blood of all the subjects under study using EIA kits. Thirty NSCLC patients and ten controls underwent induced sputum collection and analysis. Results LTB-4 and IL-8 resulted higher in breath condensate and the blood of NSCLC patients compared to controls. Significantly higher concentrations were found as the cancer stages progressed. A positive correlation was observed between exhaled IL-8 and LTB-4 and the percentage of neutrophils in the induced sputum. Conclusion The high concentrations of exhaled LTB-4 and IL-8 showed the presence of a neutrophilic inflammation in the airways of NSCLC patients and gave a further support to the inflammatory signalling in lung cancer. These exhaled proteins could represent a suitable non-invasive marker in the diagnosis and monitoring of lung cancer.

  8. Influence of neopterin on generation of reactive species by myeloperoxidase in human neutrophils.

    Science.gov (United States)

    Razumovitch, Julia A; Fuchs, Dietmar; Semenkova, Galina N; Cherenkevich, Sergei N

    2004-04-07

    Increased neopterin concentrations in human serum indicate activation of cell-mediated immune response. Earlier we have shown that neopterin enhanced generation of singlet oxygen, hydroxyl radical and nitric oxide in human peripheral blood neutrophils by NADPH-independent pathways. To further investigate a participation of neopterin in reactive species production by neutrophils, we studied its influence on myeloperoxidase (MPO) activity. MPO was isolated from human peripheral blood neutrophils from healthy donors. Generation of reactive species by MPO/H(2)O(2) in Earl's solution (pH=7.2) at 37 degrees C was investigated by monitoring of chemiluminescence using luminol as light emitter. In the MPO/H(2)O(2) system, neopterin increased singlet oxygen in a concentration-dependent manner, but it decreased formation of other oxidizing species. Comparing several oxygen scavengers, formation of reactive species was totally blocked by sodium azide (NaN(3)), both in the presence and in the absence of neopterin. Superoxide dismutase (SOD) and d-mannitol insignificantly decreased chemiluminescence of this reaction, but diazabicyclo[2.2.2]octane (DABCO) strongly inhibited it. We conclude that the effects of neopterin on neutrophils' MPO are directed to increase singlet oxygen and to decrease other reactive species via inhibition of MPO and/or scavenging of reactive species.

  9. Enzootic bovine leukosis and Bovine leukemia virus

    OpenAIRE

    Amauri Alcindo Alfieri; Alice Fernandes Alfieri; Luis Álvaro Leuzzi Junior

    2004-01-01

    All over de World the Enzootic Bovine Leukosis is a important viral infection in cattle herds. This revision points out topics relative to the etiological agent, clinical signals, diagnosis methods, control and prophylaxis of the infection.A Leucose Enzoótica Bovina é uma infecção viral amplamente disseminada em rebanhos bovinos de todo o mundo. Esta revisão tem por objetivo apresentar tópicos relacionados ao agente etiológico, à doença clínica e aos métodos de diagnóstico, controle e profila...

  10. A Comparison of Human Neutrophils Acquired from Four Experimental Models of Inflammation

    Science.gov (United States)

    Motwani, Madhur P.; Day, Richard M.; Gilroy, Derek W.; O’Brien, Alastair J.

    2016-01-01

    Defects in neutrophil function have been implicated in a wide spectrum of clinical conditions. Several models are employed to study activated human neutrophils akin to those found at a site of inflammation. These include whole blood (WB) ex vivo stimulation with lipopolysaccharide (LPS) and in vivo techniques: cantharidin blister, skin windows and intra-dermal injection of UV-killed E.coli (UVKEc). Neutrophils obtained from these have never been compared. We compared the activation status of neutrophils from each technique in order to inform the optimal model for use in human studies. Healthy male volunteers were randomised to undergo one of the four techniques (n = 5/group). LPS: WB stimulated with 1ng/ml of LPS for 4 hours. Cantharidin: 12.5μl of 0.1% cantharidin elicited a single blister, aspirated at 24 hours. Skin windows: four 6mm mechanical-suction blisters created, de-roofed and an exudate-collection chamber placed over the windows for 4 hours before aspiration. UVKEc: 1.5 x 107 UVKEc injected intra-dermally. A single 10mm mechanical-suction blister formed and aspirated at 4 hours. Unstimulated WB used as the control. Flow cytometry was used to determine activation status using CD16, CD11b, CD54, CD62L and CD88. Functional status was assessed with a phagocytosis assay. The pattern of neutrophil activation was similar in all models. Neutrophil CD11b was elevated in all models, most markedly in UVKEc (p<0.0001), and CD54 was also elevated but only significant in the LPS model (p = 0.001). CD62L was significantly reduced in all 4 models (p<0.0001) and CD88 was also suppressed in all. There were no changes in CD16 in any model, neither was there any significant difference in the phagocytic capacity of the neutrophils. In summary, there are no significant differences in activation marker expression or phagocytic capacity in the neutrophils obtained from each technique. Therefore we believe whole blood stimulation is the best model in experimentally challenging

  11. High glucose impairs superoxide production from isolated blood neutrophils

    DEFF Research Database (Denmark)

    Perner, A; Nielsen, S E; Rask-Madsen, J

    2003-01-01

    Superoxide (O(2)(-)), a key antimicrobial agent in phagocytes, is produced by the activity of NADPH oxidase. High glucose concentrations may, however, impair the production of O(2)(-) through inhibition of glucose-6-phosphate dehydrogenase (G6PD), which catalyzes the formation of NADPH. This study...

  12. Dihydroxyoctadecamonoenoate esters inhibit the neutrophil respiratory burst

    Indian Academy of Sciences (India)

    David Alan Thompson; Bruce D Hammock

    2007-03-01

    The leukotoxins [9(10)- and 12(13)-EpOME] are produced by activated inflammatory leukocytes such as neutrophils. High EpOME levels are observed in disorders such as acute respiratory distress syndrome and in patients with extensive burns. Although the physiological significance of the EpOMEs remains poorly understood, in some systems, the EpOMEs act as a protoxin, with their corresponding epoxide hydrolase metabolites, 9,10- and 12,13-DiHOME, specifically exerting toxicity. Both the EpOMEs and the DiHOMEs were also recently shown to have neutrophil chemotactic activity. We evaluated whether the neutrophil respiratory burst, a surge of oxidant production thought to play an important role in limiting certain bacterial and fungal infections, is modulated by members of the EpOME metabolic pathway. We present evidence that the DiHOMEs suppress the neutrophil respiratory burst by a mechanism distinct from that of respiratory burst inhibitors such as cyclosporin H or lipoxin A4, which inhibit multiple aspects of neutrophil activation.

  13. TEMPO DE VIABILIDADE DE AMOSTRAS DE SANGUE VENOSO BOVINO DESTINADAS AO EXAME HEMOGASOMÉTRICO, QUANDO MANTIDAS SOB CONSERVAÇÃO EM ÁGUA GELADA VIABILITY TIME OF BLOOD GAS ANALYSIS IN BOVINE VENOUS BLOOD SAMPLES STORED IN ICE WATER BATH

    Directory of Open Access Journals (Sweden)

    Júlio Augusto Naylor Lisbôa

    2001-04-01

    adequadamente conservadas em banho de água gelada, mantendo, assim, o seu valor diagnóstico.In order to verify the viability of blood gas analysis in bovine venous blood stored on ice water bath, two samples (10ml each were taken from the jugular vein of 14 healthy animals (7 males and 7 females, 1- to 5-year-old, using plastic syringes and attached needles filled with sodium heparin (1,000IU. The blood samples were obtained anaerobically, the air bubbles observed were immediately removed, and the needle was maintened capped with a rubber stopper. Each syringe of the pair was distinctally stored at room temperature (23-30ºC or in ice water bath (0-4ºC during the experimental period. Values of pH, carbon dioxide (PvCO2 and oxigen (PvO2 tensions, bicarbonate (HCO3-, total carbon dioxide (TCO2, base excess (BE, standard bicarbonate (StB, oxigen saturation (SatO2, and oxigen content (O2 were determined soon after sampling and after 1, 2, 3, 4, 5, 6, 8, 10, 12 and 24 hours. According to the type of storage temperature, the results were analysed through repeated measurements ANOVA, considering the contrast between the mean value of each time and the initial one. On the storage at room temperature, the in vitro changes were characterized from continuous decreases in pH, PvO2, BE, StB, SatO2, and O2 values, and gradual increase in PvCO2, starting at 2- or 3-hour after the collection. In the samples stored at 0-4ºC, on the other hand, the changes in pH occurred only at the 4th hour, and the stability of the PvCO2, BE, and StB values were maintened for up to the 6th hour. These results indicated that the diagnostic utility of blood gas analysis is conserved in bovine venous blood samples adequately stored up to 6 hours in ice water bath, at 0-4ºC.

  14. Regulators and Effectors of Arf GTPases in Neutrophils.

    Science.gov (United States)

    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.

  15. Regulators and Effectors of Arf GTPases in Neutrophils

    Directory of Open Access Journals (Sweden)

    Jouda Gamara

    2015-01-01

    Full Text Available 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.

  16. Neutrophil, TLR2, and TLR4 expression in newborns at risk of sepsis

    Directory of Open Access Journals (Sweden)

    Ari Yunanto

    2013-05-01

    Full Text Available Background There is increasing evidence that toll-like receptors (TLR play a key role in the mediation of systemic responses to invading pathogens during sepsis. Saliva is an important body fluid for detecting physiological and pathological conditions of the human body. Neutrophils are participants in the acute response against pathogens in many tissues, and their influx into the oral cavity may occur at any time. Objective To compare mean neutrophils and the expression of TLR2 and TLR4 in saliva and blood of newborns at risk for sepsis to those of healthy newborns. Methods This cross-sectional study was conducted from July to December 2011 in the Division of Neonatology, Department of Child Health, Ulin General Hospital, Lambung Mangkurat University Medical School, Banjarmasin. Case subjects were newborns with sepsis risk factors (30 infants, while 30 healthy infants were in the control group. Saliva and blood specimen examinations were performed in the Biomedical Laboratory of Brawijaya University Medical School, Malang. We used T-test for statistical analyses. Results From saliva specimens, mean neutrophils were significantly higher in the case group than in the control group [14.43 (SD 12.21 % vs. 5.63 (SD 6.78 %, respectively, (P=0.021]. In addition, mean TLR2 and mean TLR4 saliva levels were significantly higher in the case group than in the control group [TLR2: 64.97 (SD 26.42 % vs. 40.06 (SD 6.23 %, respectively, (P=0.011; TLR4: 1.5 (SD 1.61 % vs. 0.57 (SD 0.53 %, respectively, (P=0.044]. From blood specimens, mean neutrophils were also significantly higher in the case group than in the control group [1.09 (SD 0.61% vs. 0.21 (SD 0.09%, respectively, (P=0.000]. Similarly, mean blood TLR2 and TLR4 levels were significantly higher in the case group than in the control group [TLR2: 92.51 (SD 5.51 % vs. 81.74 (SD 11.79 %, respectively, (P=0.003; TLR4: 0.71 (SD 1.42 % vs. 0.12 (SD 0.06 %, respectively, (P=0.000]. Conclusion There are

  17. Neutrophil, TLR2, and TLR4 expression in newborns at risk of sepsis

    Directory of Open Access Journals (Sweden)

    Ari Yunanto

    2013-01-01

    Full Text Available Background There is increasing evidence that toll-like receptors (TLR play a key role in the mediation of systemic responses to invading pathogens during sepsis. Saliva is an important body fluid for detecting physiological and pathological conditions of the human body. Neutrophils are participants in the acute response against pathogens in many tissues, and their influx into the oral cavity may occur at any time.Objective To compare mean neutrophils and the expression of TLR2 and TLR4 in saliva and blood of newborns at risk for sepsis to those of healthy newborns.Methods This cross-sectional study was conducted from July to December 2011 in the Division of Neonatology, Department of Child Health, Ulin General Hospital, Lambung Mangkurat University Medical School, Banjarmasin. Case subjects were newborns with sepsis risk factors (30 infants, while 30 healthy infants were in the control group. Saliva and blood specimen examinations were performed in the Biomedical Laboratory of Brawijaya University Medical School, Malang. We used T-test for statistical analyses.Results From saliva specimens, mean neutrophils were significantly higher in the case group than in the control group [14.43 (SD 12.21 % vs. 5.63 (SD 6.78 %, respectively, (P=0.021]. In addition, mean TLR2 and mean TLR4 saliva levels were significantly higher in the case group than in the control group [TLR2: 64.97 (SD 26.42 % vs. 40.06 (SD 6.23 %, respectively, (P=0.011; TLR4: 1.5 (SD 1.61 % vs. 0.57 (SD 0.53 %, respectively, (P=0.044]. From blood specimens, mean neutrophils were also significantly higher in the case group than in the control group [1.09 (SD 0.61% vs. 0.21 (SD 0.09%, respectively, (P=0.000]. Similarly, mean blood TLR2 and TLR4 levels were significantly higher in the case group than in the control group [TLR2: 92.51 (SD 5.51 % vs. 81.74 (SD 11.79 %, respectively, (P=0.003; TLR4: 0.71 (SD 1.42 % vs. 0.12 (SD 0.06 %, respectively, (P=0.000].Conclusion There are significant

  18. Granulopoiesis and granules of human neutrophils

    DEFF Research Database (Denmark)

    Cowland, Jack B; Borregaard, Niels

    2016-01-01

    Granules are essential for the ability of neutrophils to fulfill their role in innate immunity. Granule membranes contain proteins that react to environmental cues directing neutrophils to sites of infection and initiate generation of bactericidal oxygen species. Granules are densely packed...... with proteins that contribute to microbial killing when liberated to the phagosome or extracellularly. Granules are, however, highly heterogeneous and are traditionally subdivided into azurophil granules, specific granules, and gelatinase granules in addition to secretory vesicles. This review will address...... issues pertinent to formation of granules, which is a process intimately connected to maturation of neutrophils from their precursors in the bone marrow. We further discuss possible mechanisms by which decisions are made regarding sorting of proteins to constitutive secretion or storage in granules...

  19. Major neutrophil functions subverted by Porphyromonas gingivalis

    Directory of Open Access Journals (Sweden)

    Ingar Olsen

    2016-03-01

    Full Text Available Polymorphonuclear leukocytes (neutrophils constitute an integrated component of the innate host defense in the gingival sulcus/periodontal pocket. However, the keystone periodontal pathogen Porphyromonas gingivalis has in the course of evolution developed a number of capacities to subvert this defense to its own advantage. The present review describes the major mechanisms that P. gingivalis uses to subvert neutrophil homeostasis, such as impaired recruitment and chemotaxis, resistance to granule-derived antimicrobial agents and to the oxidative burst, inhibition of phagocytic killing while promoting a nutritionally favorable inflammatory response, and delay of neutrophil apoptosis. Studies in animal models have shown that at least some of these mechanisms promote the dysbiotic transformation of the periodontal polymicrobial community, thereby leading to inflammation and bone loss. It is apparent that neutrophil–P. gingivalis interactions and subversion of innate immunity are key contributing factors to the pathogenesis of periodontal disease.

  20. Manejo de Bovinos en Sistemas Productivos: Caracterización de dos estilos de manejo y niveles sanguíneos de cortisol - Productive Systems in Bovine Management. The Characterization in Two Styles and Blood Cortisol Levels

    Directory of Open Access Journals (Sweden)

    Costa, Alejandro

    2007-12-01

    , de que este incremento comparado con el registrado para el tratamiento con pautas de bienestar, no fuesignificativo (P= 0,2603, el valor hallado evidenciaría el modo en el cual los bovinos recuerdan el maltrato.En ambos tratamientos se registraron diferencias en el comportamiento. Con el tratamientosegún pautas de bienestar el manejo se efectuó sin que los animales presentaran signos de intranquilidad, en tanto que en el tratamiento convencional, signos como una marcada polipnea y fuerte excitación, se manifestaron con intensidad.AbstractThe animal welfare problematic, has gained International relevance and in this frame exists recommendations about its significance over de International Commerce. At the same time, in Argentina exists laws about this question; laws that have placed the discussion in theConsorcio de Exportadores de Carnes Argentinas (ABC.The main goal of this paper is the characterization of two different styles of bovine handling: the first one considers that the animals are dominated by force and the other one, based in the knowledge of the bovinebehavior; each of them has its hormonal correspondence which is measured through blood’s cortisol as an indicator of stress.In this research, an 80-young bull share was divided, in a random way, in two sub-shares of same number of animals. Each sub-share was submitted toone of the two different styles. At the same time, 10 young bulls of each sub-share, were selected in a random way. Two blood samples were taken from each animal and the blood cortisol was measured. The first sample informed about the cortisol level under both treatments. The second one, was taken after the animal was calmed, looking forward to clear the sympathetic alarm. It was possible to see the existence of significant differences in the cortisol levels between the two sub-shares (P 0,00413, p<0,01.The second examination,show a significant increase in the level of cortisol (P 0,0017, p<0,001; increase that appears as results of

  1. Platelets augment respiratory burst in neutrophils activated by selected species of gram-positive or gram-negative bacteria.

    Directory of Open Access Journals (Sweden)

    Kamila Pytel

    2008-12-01

    Full Text Available Neutrophils and platelets circulate in blood system and play important physiological roles as part of immunological system. Neutrophils are the first line of host defense against various intruders, and platelets are satellite cells cooperating with other components of defense system. Recent studies report about the cooperation among these types of cells. We analyzed the effect of platelets on oxygen burst in neutrophils triggered by Staphylococcus aureus and Escherichia coli bacteria in vitro. The effect of platelets on oxygen burst in neutrophils was measured by luminol enhanced chemiluminescence. Opsonized and non-opsonized bacteria were used as activators. Activation of neutrophils with live non-opsonized and opsonized bacteria in the presence of platelets increased the oxygen burst as compared to the same system without platelets. The gram-positive bacteria (Staphylococcus aureus were causing higher activation than gram-negative bacteria (Escherichia coli. This work demonstrate that platelets potentate the response of neutrophils augmenting their respiratory burst in vitro when triggered by bacteria.

  2. NOD2 dependent neutrophil recruitment is required for early protective immune responses against infectious Litomosoides sigmodontis L3 larvae

    Science.gov (United States)

    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

  3. Low molecular weight heparin alters porcine neutrophil responses to platelet-activating factor.

    Science.gov (United States)

    Kruse-Elliott, K T; Chaban, K; Grossman, J E; Tomasko, S; Kamke, C; Darien, B

    1998-09-01

    Because platelet-activating factor (PAF) is an important mediator of inflammation and heparin has anti-inflammatory effects, we hypothesized that low molecular weight heparin (LMWH) would inhibit PAF-induced activation and chemotaxis in porcine neutrophils. Citrated blood was obtained from pentobarbital-anesthetized pigs, and neutrophils were isolated over a 55%/65% Percoll gradient. The effect of LMWH on basal phorbol myristate acetate (PMA)-induced superoxide (SO) release, as well as its effect on PAF priming for PMA-induced SO release, were investigated. Additionally, the effect of LMWH on PAF-induced chemotaxis of neutrophils across transwell membranes was evaluated. Baseline SO release in response to PMA was .351+/-.046 nmol/10(6) cells/min, and this was decreased to .289+/-.034 nmol/10(6) cells/min by pretreatment with 50 U/mL LMWH. PMA-induced SO production was increased by .240+/-.042 nmol/10(6) cells/min when cells were primed with 10 microM PAF. This priming effect of PAF was reduced significantly by pretreatment of neutrophils with LMWH at 10 and 50 U/mL. Chemotaxis of neutrophils in response to 100 microM PAF was significantly decreased to 70.02+/-6.4% (n = 8) of the control response by pretreatment of cells with 50 U/mL LMWH. We conclude that LMWH has anti-inflammatory effects on porcine neutrophils, which includes attenuation of cell activation and chemotaxis in response to the lipid-derived inflammatory mediator, PAF.

  4. Bovine milk glycome.

    Science.gov (United States)

    Tao, N; DePeters, E J; Freeman, S; German, J B; Grimm, R; Lebrilla, C B

    2008-10-01

    Bovine milk oligosaccharides have several potentially important biological activities including the prevention of pathogen binding to the intestinal epithelial and as nutrients for beneficial bacteria. It has been suggested that milk oligosaccharides are an important source of complex carbohydrates as supplements for the food and the pharmaceutical industries. However, only a small number of structures of bovine milk oligosaccharides (bMO) are known. There have been no systematic studies on bMO. High-performance mass spectrometry and separation methods are used to evaluate bMO, and nearly 40 oligosaccharides are present in bovine milk. Bovine milk oligosaccharides are composed of shorter oligomeric chains than are those in human milk. They are significantly more anionic with nearly 70%, measured abundances, being sialylated. Additionally, bMO are built not only on the lactose core (as are nearly all human milk oligosaccharides), but also on lactose amines. Sialic acid residues include both N-acetyl and N-glycolylneuraminic acid, although the former is significantly more abundant.

  5. Intervet Symposium: bovine neosporosis

    NARCIS (Netherlands)

    Schetters, T.; Dubey, J.P.; Adrianarivo, A.; Frankena, K.; Romero, J.J.; Pérez, E.; Heuer, C.; Nicholson, C.; Russell, D.; Weston, J.

    2004-01-01

    This article summarises the most relevant data of presentations delivered at the 19th International Conference of the World Association for the Advancement of Veterinary Parasitology (WAAVP) held in New Orleans, LA, USA, from 10 to 14 August 2003) in a symposium session on bovine neosporosis. The sy

  6. Bovine Spongiform Encephalopathy

    Science.gov (United States)

    Bovine spongiform encephalopathy (BSE) is caused by a novel contagion, known to as a prion. Prions are proteins capable of converting a normal cellular protein into a prion, thereby propagating an infection. BSE is the first known prion zoonotic. As such it has attracted broad scientific and, to a r...

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

    Science.gov (United States)

    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.

  8. Anti-neutrophil cytoplasm autoantibodies (ANCA) in autoimmune liver diseases

    NARCIS (Netherlands)

    Roozendaal, C.; Kallenberg, Cees

    1999-01-01

    Anti-neutrophil cytoplasm antibodies (ANCA) are autoantibodies directed against cytoplasmic constituents of neutrophil granulocytes and monocytes. ANCA have been detected in serum from patients with inflammatory bowel diseases (mainly ulcerative colitis) and autoimmune mediated liver diseases (mainl

  9. Bovine Mastitis: Frontiers in Immunogenetics

    Directory of Open Access Journals (Sweden)

    Kathleen eThompson-Crispi

    2014-10-01

    Full Text Available Mastitis is one of the most prevalent and costly diseases in the dairy industry with losses attributable to reduced milk production, discarded milk, early culling, veterinary services, and labor costs. Typically, mastitis is an inflammation of the mammary gland most often, but not limited to, bacterial infection, and is characterized by the movement of leukocytes and serum proteins from the blood to the site of infection. It contributes to compromised milk quality and the potential spread of antimicrobial resistance if antibiotic treatment is not astutely applied. Despite the implementation of management practises and genetic selection approaches, bovine mastitis control continues to be inadequate. However, some novel genetic strategies have recently been demonstrated to reduce mastitis incidence by taking advantage of a cow’s natural ability to make appropriate immune responses against invading pathogens. Specifically, dairy cattle with enhanced and balanced immune responses have a lower occurrence of disease, including mastitis, and they can be identified and selected for using the High Immune Response (HIR technology. Enhanced immune responsiveness is also associated with improved response to vaccination, increased milk and colostrum quality. Since immunity is an important fitness trait, beneficial associations with longevity and reproduction are also often noted. This review highlights the genetic regulation of the bovine immune system and its vital contributions to disease resistance. Genetic selection approaches currently used in the dairy industry to reduce the incidence of disease are reviewed, including the HIR technology, genomics to improve disease resistance or immune response, as well as the Immunity+TM sire line. Improving the overall immune responsiveness of cattle is expected to provide superior disease resistance, increasing animal welfare and food quality while maintaining favourable production levels to feed a growing

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

    Science.gov (United States)

    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.

  11. Blood Types

    Science.gov (United States)

    ... maternity. Learn About Blood Blood Facts and Statistics Blood Components Whole Blood and Red Blood Cells Platelets Plasma ... About Blood Blood Facts and Statistics Blood Types Blood Components What Happens to Donated Blood Blood and Diversity ...

  12. A comparison of neutrophil gelatinase-associated lipocalin and immature to total neutrophil ratio for diagnosing early-onset neonatal sepsis

    Directory of Open Access Journals (Sweden)

    Rocky Wilar

    2016-07-01

    Full Text Available Background Neonatal sepsis is a clinical syndrome caused by the invasion of microorganisms into the bloodstream. Early diagnosis of early-onset neonatal sepsis (EONS is difficult. Laboratory tests with high sensitivity and specificity are needed in order to make early diagnoses in newborns.Objective To compare the sensitivity and specificity of neutrophil gelatinase-associated lipocalin (NGAL and immature to total (IT neutrophil ratio for the diagnosis of early-onset neonatal sepsis.Methods This observational study with cross-sectional design was conducted in the Neonatology Division, Prof. R. D. Kandou General Hospital from November 2012 to April 2014. Consecutive sampling was applied. There were 103 newborns with suspected EONS who fulfilled the inclusion criteria. Complete blood counts, blood cultures, as well as NGAL and IT ratio measurements were performed.Results NGAL was not significantly more sensitive than IT ratio [80.4% vs. 67.3%, respectively; (P=0.058]. However, NGAL had lower specificity than IT ratio (27.7% vs. 50.0%, respectively; P=0.016. The positive predictive values (57.0% vs. 64.9%, respectively; P=0.176, and negative predictive values (54.2% vs. 52.6%, respectively; P=0.451 were similar in both diagnostic tests.Conclusion Immature to total neutrophil (IT ratio has higher specificity compared to NGAL for early diagnosis of EONS. However, the difference in sensitivity between the two test is not statistically significant.

  13. The Molecular Mechanisms of Glucocorticoids-Mediated Neutrophil Survival

    OpenAIRE

    2011-01-01

    Neutrophil-dominated inflammation plays an important role in many airway diseases including asthma, chronic obstructive pulmonary disease (COPD), bronchiolitis and cystic fibrosis. In cases of asthma where neutrophil-dominated inflammation is a major contributing factor to the disease, treatment with corticosteroids can be problematic as corticosteroids have been shown to promote neutrophil survival which, in turn, accentuates neutrophilic inflammation. In light of such cases, novel targeted ...

  14. Phagocytosis and Killing of Staphylococcus aureus by Human Neutrophils

    OpenAIRE

    Lu, Thea; Porter, Adeline R.; Kennedy, Adam D.; Kobayashi, Scott D.; Frank R DeLeo

    2014-01-01

    Neutrophils are essential for host defense against Staphylococcus aureus infections. Although significant progress has been made, our understanding of neutrophil interactions with S. aureus remains incomplete. To provide a more comprehensive view of this process, we investigated phagocytosis and killing of S. aureus by human neutrophils using varied assay conditions in vitro. A greater percentage of bacteria were internalized by adherent neutrophils compared to those in suspension, and unexpe...

  15. Yersinia pseudotuberculosis is resistant to killing by human neutrophils.

    Science.gov (United States)

    Laws, Thomas R; Davey, Martin S; Green, Christopher; Cooper, Ian A M; Titball, Richard W; Lukaszewski, Roman A

    2011-06-01

    The interaction between human neutrophils and the Gram negative gastrointestinal pathogen Yersinia pseudotuberculosis was investigated in vitro. Despite the wealth of data describing how Yersinia can affect the function of neutrophils, there are no published studies describing if neutrophil cells can affect the viability of Y. pseudotuberculosis. The wild-type IP32953 strain of Y. pseudotuberculosis was found to be resistant to killing by human neutrophils. Confocal examination and flow-cytometric analysis of this interaction revealed that bacteria were taken up.

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

    OpenAIRE

    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 neut...

  17. LFA-1 and Mac-1 define characteristically different intralumenal crawling and emigration patterns for monocytes and neutrophils in situ.

    Science.gov (United States)

    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.

  18. Expression of CD11c and EMR2 on neutrophils: potential diagnostic biomarkers for sepsis and systemic inflammation.

    Science.gov (United States)

    Lewis, S M; Treacher, D F; Edgeworth, J; Mahalingam, G; Brown, C S; Mare, T A; Stacey, M; Beale, R; Brown, K A

    2015-11-01

    There is a need for cellular biomarkers to differentiate patients with sepsis from those with the non-infectious systemic inflammatory response syndrome (SIRS). In this double-blind study we determined whether the expression of known (CD11a/b/c, CD62L) and putative adhesion molecules [CD64, CD97 and epidermal growth factor (EGF)-like molecule containing mucin-like hormone receptor (EMR2)] on blood neutrophils could serve as useful biomarkers of infection and of non-infectious SIRS in critically ill patients. We studied 103 patients with SIRS, 83 of whom had sepsis, and 50 healthy normal subjects, using flow cytometry to characterize neutrophils phenotypically in whole blood samples. Patients with SIRS had an increased prevalence of neutrophils expressing CD11c, CD64 and EMR2 in comparison with healthy subjects (P sepsis, EMR2 with SIRS and CD64 with sepsis and SIRS. Neutrophils expressing CD11c had the highest sensitivity (81%) and specificity (80%) for the detection of sepsis, and there was an association between the percentage of neutrophils expressing EMR2 and the extent of organ failure (P sepsis and SIRS, respectively.

  19. Regulatory T cells, dendritic cells and neutrophils in patients with renal cell carcinoma.

    Science.gov (United States)

    Minárik, Ivo; Lašťovička, Jan; Budinský, Vít; Kayserová, Jana; Spíšek, Radek; Jarolím, Ladislav; Fialová, Anna; Babjuk, Marek; Bartůňková, Jiřina

    2013-05-01

    We evaluated dendritic cells (DC), regulatory T lymphocytes (Treg) and neutrophils in 37 patients with newly diagnosed renal cell carcinoma (RCC) in the tumor and peripheral blood (PB) and correlated these parameters with tumor staging (early-T1, 2, late-T3, 4 and metastatic disease). The number of myeloid and plasmacytoid DC in blood of RCC patients was higher than in healthy controls. The percentage of myeloid dendritic cells (mDC) from CD45+ cells in tumors was higher in comparison with peripheral blood irrespective of disease stage. Higher percentage of these cells expressed a maturation marker in the periphery in the early stage (CD83 expressing cells). The number of plasmacytoid dendritic cells (pDC) in PB was similar in both early and late stage groups, but the early group displayed a significantly higher percentage of pDC in tumor cell suspension. Neutrophil counts in the peripheral blood of RCC patients were higher than in healthy controls, but the counts in both tumor stage groups were similar. The proportion of neutrophils from CD45+ cells was higher in late stage tumors. Higher percentage of Treg from CD4+ cells was detected in renal carcinoma tissue in comparison to PB with no difference between stages of the disease. Our results reflect the complex interplay between various cells of the immune system and the tumor microenvironment. Activation of dendritic cell subpopulations at early stages of the disease course is counterbalanced by the early appearance of T regulatory cells both in the periphery and tumor tissue. Later stages are characterized by the accumulation of neutrophils in the tumor. Appropriate timing of anticancer strategies, especially immunotherapy, should take these dynamics of the immune response in RCC patients into account.

  20. Exposure to Leishmania braziliensis triggers neutrophil activation and apoptosis.

    Directory of Open Access Journals (Sweden)

    Sarah A C Falcão

    2015-03-01

    Full Text Available BACKGROUND: Neutrophils are the first line of defense against invading pathogens and are rapidly recruited to the sites of Leishmania inoculation. During Leishmania braziliensis infection, depletion of inflammatory cells significantly increases the parasite load whereas co-inoculation of neutrophils plus L. braziliensis had an opposite effect. Moreover, the co-culture of infected macrophages and neutrophils also induced parasite killing leading us to ask how neutrophils alone respond to an L. braziliensis exposure. Herein we focused on understanding the interaction between neutrophils and L. braziliensis, exploring cell activation and apoptotic fate. METHODS AND FINDINGS: Inoculation of serum-opsonized L. braziliensis promastigotes in mice induced neutrophil accumulation in vivo, peaking at 24 h. In vitro, exposure of thyoglycollate-elicited inflammatory or bone marrow neutrophils to L. braziliensis modulated the expression of surface molecules such as CD18 and CD62L, and induced the oxidative burst. Using mCherry-expressing L. braziliensis, we determined that such effects were mainly observed in infected and not in bystander cells. Neutrophil activation following contact with L. braziliensis was also confirmed by the release of TNF-α and neutrophil elastase. Lastly, neutrophils infected with L. braziliensis but not with L. major displayed markers of early apoptosis. CONCLUSIONS: We show that L. braziliensis induces neutrophil recruitment in vivo and that neutrophils exposed to the parasite in vitro respond through activation and release of inflammatory mediators. This outcome may impact on parasite elimination, particularly at the early stages of infection.

  1. Peripheral and intrauterine neutrophil function in the cow: the influence of endogenous and exogenous sex steroid hormones.

    Science.gov (United States)

    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

  2. Neutrophilic dermatoses and inflammatory bowel diseases.

    Science.gov (United States)

    Marzano, A V; Menicanti, C; Crosti, C; Trevisan, V

    2013-04-01

    Pyoderma gangrenosum (PG) and Sweet's Syndrome (SS) are inflammatory skin diseases caused by the accumulation of neutrophils in the skin and, rarely, in internal organs, which led to coining the term of neutrophilic dermatoses (ND) to define these conditions. Recently, ND have been included among the autoinflammatory diseases, which are forms due to mutations of genes regulating the innate immune responses. Both PG and SS are frequently associated with inflammatory bowel diseases (IBD), a group of chronic intestinal disorders which comprises ulcerative colitis and Crohn's disease and whose pathogenesis involves both the innate and adaptive immunity in genetically prone individuals. Patients with IBD develop PG in 1-3% of cases, while SS is rarer. PG presents with deep erythematous-to-violaceous painful ulcers with undermined borders, but bullous, pustular, and vegetative variants can also occur. SS, also known as acute febrile neutrophilic dermatosis, is characterized by the abrupt onset of fever, peripheral neutrophilia, tender erythematous skin lesions and a diffuse neutrophilic dermal infiltrate. In this review that will be focused on PG and SS, we will describe also the aseptic abscesses syndrome, a new entity within the spectrum of ND which frequently occurs in association with IBD and is characterized by deep abscesses mainly involving the spleen and skin and by polymorphic cutaneous manifestations including PG- and SS-like lesions.

  3. Modulation of neutrophil apoptosis by antimicrobial peptides.

    Science.gov (United States)

    Nagaoka, Isao; Suzuki, Kaori; Niyonsaba, François; Tamura, Hiroshi; Hirata, Michimasa

    2012-01-01

    Peptide antibiotics possess the potent antimicrobial activities against invading microorganisms and contribute to the innate host defense. Human antimicrobial peptides, α-defensins (human neutrophil peptides, HNPs), human β-defensins (hBDs), and cathelicidin (LL-37) not only exhibit potent bactericidal activities against Gram-negative and Gram-positive bacteria, but also function as immunomodulatory molecules by inducing cytokine and chemokine production, and inflammatory and immune cell activation. Neutrophil is a critical effector cell in host defense against microbial infection, and its lifespan is regulated by various pathogen- and host-derived substances. Here, we provided the evidence that HNP-1, hBD-3, and LL-37 cannot only destroy bacteria but also potently modulate (suppress) neutrophil apoptosis, accompanied with the phosphorylation of ERK-1/-2, the downregulation of tBid (an proapoptotic protein) and upregulation of Bcl-xL (an antiapoptotic protein), and the inhibition of mitochondrial membrane potential change and caspase 3 activity, possibly via the actions on the distinct receptors, the P2Y6 nucleotide receptor, the chemokine receptor CCR6, and the low-affinity formyl-peptide receptor FPRL1/the nucleotide receptor P2X7, respectively. Suppression of neutrophil apoptosis results in the prolongation of their lifespan and may be advantageous for the host defense against bacterial invasion.

  4. On the mechanism of oscillations in neutrophils

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Barington, Torben; Olsen, Lars Folke

    2010-01-01

    We have investigated the regulation of the oscillatory generation of H(2)O(2) and oscillations in shape and size in neutrophils in suspension. The oscillations are independent of cell density and hence do not represent a collective phenomena. Furthermore, the oscillations are independent of the e...

  5. Effect of post-exercise protein-leucine feeding on neutrophil function, immunomodulatory plasma metabolites and cortisol during a 6-day block of intense cycling.

    Science.gov (United States)

    Nelson, Andre R; Jackson, Lara; Clarke, Jim; Stellingwerff, Trent; Broadbent, Suzanne; Rowlands, David S

    2013-09-01

    Whey protein and leucine ingestion following exercise increases muscle protein synthesis and could influence neutrophil function during recovery from prolonged intense exercise. We examined the effects of whey protein and leucine ingestion post-exercise on neutrophil function and immunomodulators during a period of intense cycling. In a randomized double-blind crossover, 12 male cyclists ingested protein/leucine/carbohydrate/fat (LEUPRO 20/7.5/89/22 g h(-1), respectively) or isocaloric carbohydrate/fat control (CON 119/22 g h(-1)) beverages for 1-3 h post-exercise during 6 days of high-intensity training. Blood was taken pre- and post-exercise on days 1, 2, 4 and 6 for phorbol myristate acetate (PMA)-stimulated neutrophil superoxide (O2 (-)) production, immune cell counts, amino acid and lipid metabolism via metabolomics, hormones (cortisol, testosterone) and cytokines (interleukin-6, interleukin-10). During recovery on day 1, LEUPRO ingestion increased mean concentrations of plasma amino acids (glycine, arginine, glutamine, leucine) and myristic acid metabolites (acylcarnitines C14, myristoylcarnitine; and C14:1-OH, hydroxymyristoleylcarnitine) with neutrophil priming capacity, and reduced neutrophil O2 production (15-17 mmol O2 (-) cell(-1) ± 90 % confidence limits 20 mmol O2 (-) cell(-1)). On day 2, LEUPRO increased pre-exercise plasma volume (6.6 ± 3.8 %) but haematological effects were trivial. LEUPRO supplementation did not substantially alter neutrophil elastase, testosterone, or cytokine concentrations. By day 6, however, LEUPRO reduced pre-exercise cortisol 21 % (±15 %) and acylcarnitine C16 (palmitoylcarnitine) during exercise, and increased post-exercise neutrophil O2 (-) (33 ± 20 mmol O2 (-) cell(-1)), relative to control. Altered plasma amino acid and acylcarnitine concentrations with protein-leucine feeding might partly explain the acute post-exercise reduction in neutrophil function and increased exercise-stimulated neutrophil oxidative burst on

  6. Biologic therapy improves psoriasis by decreasing the activity of monocytes and neutrophils.

    Science.gov (United States)

    Yamanaka, Keiichi; Umezawa, Yoshinori; Yamagiwa, Akisa; Saeki, Hidehisa; Kondo, Makoto; Gabazza, Esteban C; Nakagawa, Hidemi; Mizutani, Hitoshi

    2014-08-01

    Therapy with monoclonal antibodies to tumor necrosis factor (TNF)-α and the interleukin (IL)-12/23 p40 subunit has significantly improved the clinical outcome of patients with psoriasis. These antibodies inhibit the effects of the target cytokines and thus the major concern during their use is the induction of excessive immunosuppression. Recent studies evaluating the long-term efficacy and safety of biologic therapy in psoriasis have shown no significant appearance of serious adverse effects including infections and malignancies. However, the immunological consequence and the mechanism by which the blockade of a single cytokine by biologics can successfully control the activity of psoriasis remain unclear. In the current study, we investigated the effect of biologic therapy on cytokine production of various lymphocytes and on the activity of monocytes and neutrophils in psoriatic patients. Neutrophils, monocytes and T cells were purified from heparinized peripheral venous blood by Ficoll density gradient centrifugation, and γ-interferon, TNF-α and IL-17 production from lymphocytes was measured by flow cytometer. The activation maker of neutrophils and the activated subsets of monocytes were also analyzed. Biologic therapy induced no significant changes in the cytokine production by lymphocytes from the skin and gut-homing T cells. However, neutrophil activity and the ratio of activated monocyte population increased in severely psoriatic patients were normalized in psoriatic patients receiving biologic therapy. The present study showed that biologic therapy ameliorates clinical symptoms and controls the immune response in patients with psoriasis.

  7. T(H)2 cytokines modulate the IL-9R expression on human neutrophils.

    Science.gov (United States)

    Dragon, Stéphane; Takhar, Manrit Kaur; Shan, Lianyu; Hayglass, Kent T; Simons, F Estelle; Gounni, Abdelilah S

    2009-06-26

    Interleukin (IL)-9 is associated with key pathological features of asthma such as airway hyperresponsiveness, bronchoconstriction and mucus production. Inflammatory responses mediated by IL-9 rely on the expression of the IL-9R which has been reported on lung epithelial cells, T lymphocytes and recently on airway granulocyte infiltrates. In this study, we assessed the regulatory and constitutive cell surface expression of the IL-9Ralpha in unfractionated and purified human neutrophils from atopic asthmatics, atopic non-asthmatics and healthy normal controls. We demonstrate that T(H)2 cytokines (IL-4 or IL-13) and granulocyte macrophage-colony stimulating factor (GM-CSF) up-regulated mRNA and cell surface expression levels of the IL-9Ralpha in primary human and HL-60 differentiated neutrophils. Pharmacological inhibition of NF-kappaB did not affect T(H)2-mediated IL-9Ralpha expression in human neutrophils although IFN-gamma and IL-10 down-regulated IL-9Ralpha expression when co-incubated with IL-4, IL-13 or GM-CSF. Collectively, our results reveal a regulatory function for IFN-gamma and IL-10 on modulating the inducible IL-9Ralpha expression levels on peripheral blood neutrophils by T(H)2 cytokines.

  8. The effect of the anaesthetic agent isoflurane on the rate of neutrophil apoptosis in vitro.

    LENUS (Irish Health Repository)

    Tyther, R

    2012-02-03

    BACKGROUND: Volatile anaesthetic agents influence neutrophil function, and potentially, the inflammatory response to surgery. AIM: The objective of this study was to determine the effect of isoflurane (1-4%) on human polymorphonuclear neutrophil apoptosis in vitro. METHODS: Venous blood from 12 healthy volunteers was exposed to 0, 1, and 4% isoflurane delivered via a 14G Wallace flexihub internal jugular cannula, at a fresh gas flow of 0.51\\/min for 5 minutes. Isolated neutrophils were assessed for apoptosis at 1, 12, and 24 hours in culture using dual staining with annexin V-FITC and propidium iodide (Annexin-V FITC assay). Data were analysed using paired, one-tailed Student\\'s t-tests. p<0.05 was considered significant. RESULTS: At 1 hour apoptosis was inhibited in the 1% (5.1 [6.8]%; p=0.017) and 4% (4.8 [4.5]%; p=0.008) isoflurane groups compared to control (11.3 [6.9]%). At 12 and 24 hours, a dose-dependent inhibition of apoptosis was demonstrated, i.e. 4% > 1% > 0%. CONCLUSION: Human neutrophil apoptosis is inhibited in a concentration-dependent manner in vitro by isoflurane in clinical concentrations.

  9. Selenium in bovine spermatozoa.

    Science.gov (United States)

    Niemi, S M; Kuzan, F B; Senger, P L

    1981-05-01

    This study investigated the association of selenium with ejaculated bovine spermatozoa. Over 75% of the radioactive spermatozoa. Over 75% of the radioactive selenium-75 was released after 30 min of incubation in 2 X 10(-3) dithiothreitol. Of the selenium-75 released by dithiothreitol, 85% was associated with spermatozoal protein. Protein containing selenium-75 was found predominantly in a single band after polyacrylamide gel electrophoresis. Molecular weight was approximately 21,500 daltons.

  10. Infectious bovine keratoconjunctivitis (pinkeye)

    OpenAIRE

    Angelos, JA

    2015-01-01

    Copyright © 2015 Elsevier Inc. All rights reserved. As is the case for controlling other infectious livestock diseases, the most successful efforts to control infectious bovine keratoconjunctivitis (IBK) will include consideration of the host, the environment, herd management, and ongoing surveillance even after the immediate crisis has passed. Research over many years has led to the discovery of a variety of antibiotic treatments and antibiotic regimens that can be effective against IBK. The...

  11. Bovine leukocyte adhesion deficiency (BLAD): a review.

    Science.gov (United States)

    Nagahata, Hajime

    2004-12-01

    Bovine leukocyte adhesion deficiency (BLAD) in Holstein cattle is an autosomal recessive congenital disease characterized by recurrent bacterial infections, delayed wound healing and stunted growth, and is also associated with persistent marked neutrophilia. The molecular basis of BLAD is a single point mutation (adenine to guanine) at position 383 of the CD18 gene, which caused an aspartic acid to glycine substitution at amino acid 128 (D128G) in the adhesion molecule CD18. Neutrophils from BLAD cattle have impaired expression of the beta2 integrin (CD11a,b,c/CD18) of the leukocyte adhesion molecule. Abnormalities in a wide spectrum of adherence dependent functions of leukocytes have been fully characterized. Cattle affected with BLAD have severe ulcers on oral mucous membranes, severe periodontitis, loss of teeth, chronic pneumonia and recurrent or chronic diarrhea. Affected cattle die at an early age due to the infectious complications. Holstein bulls, including carrier sires that had a mutant BLAD gene in heterozygote were controlled from dairy cattle for a decade. The control of BLAD in Holstein cattle by publishing the genotypes and avoiding the mating between BLAD carriers was found to be successful. This paper provides an overview of the genetic disease BLAD with reference to the disease in Holstein cattle.

  12. Survival and function of phagocytes in blood culture media

    DEFF Research Database (Denmark)

    Fischer, T K; Prag, J; Kharazmi, A

    1999-01-01

    The survival and function of human phagocytes in sterile aerobic and anaerobic blood culture media were investigated using neutrophil morphology, white blood cell count in a haemoanalyser, flow cytometry, oxidative burst response, and bactericidal effect in Colorbact and Septi-Chek blood culture...

  13. Periodontal Ligament Stem Cells Regulate Apoptosis of Neutrophils

    Science.gov (United States)

    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.

  14. Escape of Mycobacterium tuberculosis from oxidative killing by neutrophils.

    Science.gov (United States)

    Corleis, Björn; Korbel, Daniel; Wilson, Robert; Bylund, Johan; Chee, Ronnie; Schaible, Ulrich E

    2012-07-01

    Neutrophils enter sites of infection, where they can eliminate pathogenic bacteria in an oxidative manner. Despite their predominance in active tuberculosis lesions, the function of neutrophils in this important human infection is still highly controversial. We observed that virulent Mycobacterium tuberculosis survived inside human neutrophils despite prompt activation of these defence cells' microbicidal effectors. Survival of M. tuberculosis was accompanied by necrotic cell death of infected neutrophils. Necrotic cell death entirely depended on radical oxygen species production since chronic granulomatous disease neutrophils were protected from M. tuberculosis-triggered necrosis. More, importantly, the M. tuberculosis ΔRD1 mutant failed to induce neutrophil necrosis rendering this strain susceptible to radical oxygen species-mediated killing. We conclude that this virulence function is instrumental for M. tuberculosis to escape killing by neutrophils and contributes to pathogenesis in tuberculosis.

  15. Phagocytosis and killing of Streptococcus suis by porcine neutrophils.

    Science.gov (United States)

    Chabot-Roy, Geneviève; Willson, Philip; Segura, Mariela; Lacouture, Sonia; Gottschalk, Marcelo

    2006-07-01

    Streptococcus suis serotype 2 is an important swine pathogen responsible for diverse infections, mainly meningitis. Virulence factors and the pathogenesis of infection are not well understood. Neutrophils may play an important role in the pathogenesis of infection given that infiltration by neutrophils and mononuclear cells are frequently observed in lesions caused by S. suis. The objective of this work was to study the interactions between S. suis serotype 2 and porcine neutrophils. Results showed that suilysin is toxic to neutrophils and this could help S. suis evade innate immunity. Moreover, suilysin appears to affect complement-dependent killing by decreasing the opsonization of S. suis and the bactericidal capacity of neutrophils. Our results confirm that capsule polysaccharide protects S. suis against killing and phagocytosis by neutrophils. We also showed that the presence of specific IgG against S. suis serotype 2 promoted killing by neutrophils, indicating that the induction of a strong humoral response is beneficial for clearance of this pathogen.

  16. The cystic fibrosis neutrophil: a specialized yet potentially defective cell.

    LENUS (Irish Health Repository)

    Hayes, Elaine

    2012-02-01

    Cystic fibrosis (CF) is one of the commonest genetically inherited diseases in the world. It is characterized by recurrent respiratory tract infections eventually leading to respiratory failure. One of the hallmarks of this disease is a persistent and predominantly neutrophil driven inflammation. Neutrophils provide the first line of defence by killing and digesting phagocytosed bacteria and fungi, yet despite advances in our understanding of the molecular and cellular basis of CF, there remains a paradox of why recruited CF neutrophils fail to eradicate bacterial infections in the lung. This review describes mechanisms involved in neutrophil migration, microbial killing and apoptosis leading to inflammatory resolution. We discuss dysregulated neutrophil activity and consider genetic versus inflammatory neutrophil reprogramming in CF and ultimately pharmacological modulation of the CF neutrophil for therapeutic intervention.

  17. The cystic fibrosis neutrophil: a specialized yet potentially defective cell.

    LENUS (Irish Health Repository)

    Hayes, Elaine

    2011-04-01

    Cystic fibrosis (CF) is one of the commonest genetically inherited diseases in the world. It is characterized by recurrent respiratory tract infections eventually leading to respiratory failure. One of the hallmarks of this disease is a persistent and predominantly neutrophil driven inflammation. Neutrophils provide the first line of defence by killing and digesting phagocytosed bacteria and fungi, yet despite advances in our understanding of the molecular and cellular basis of CF, there remains a paradox of why recruited CF neutrophils fail to eradicate bacterial infections in the lung. This review describes mechanisms involved in neutrophil migration, microbial killing and apoptosis leading to inflammatory resolution. We discuss dysregulated neutrophil activity and consider genetic versus inflammatory neutrophil reprogramming in CF and ultimately pharmacological modulation of the CF neutrophil for therapeutic intervention.

  18. Flow cytometric detection of neutrophil-associated immunoglobulin in patients with or without neutropenia and establishment of the reference interval.

    Science.gov (United States)

    Hwang, Keumrock; Park, Chan-Jeoung; Huh, Hee Jin; Han, Sang Hee; Jang, Seongsoo; Chi, Hyun-Sook

    2011-01-01

    We measured neutrophil-associated immunoglobulin (NAIg) levels using flow cytometry to establish the reference interval for NAIg and to estimate NAIg in patients with or without neutropenia. Peripheral blood from 152 individuals was analyzed for NAIg detection by flow cytometry. Using fluorescescent-conjugated anti-CD10 monoclonal antibody and anti-human immunoglobulins, proportions of NAIgG, NAIgA, and NAIgM bound to neutrophils were measured. Reference intervals for NAIg were set as NAIgG reference intervals defined herein, patients with neutropenia or adverse transfusion reactions may be evaluated in a clinically relevant manner.

  19. Diagnostic imaging in bovine orthopedics.

    Science.gov (United States)

    Kofler, Johann; Geissbühler, Urs; Steiner, Adrian

    2014-03-01

    Although a radiographic unit is not standard equipment for bovine practitioners in hospital or field situations, ultrasound machines with 7.5-MHz linear transducers have been used in bovine reproduction for many years, and are eminently suitable for evaluation of orthopedic disorders. The goal of this article is to encourage veterinarians to use radiology and ultrasonography for the evaluation of bovine orthopedic disorders. These diagnostic imaging techniques improve the likelihood of a definitive diagnosis in every bovine patient but especially in highly valuable cattle, whose owners demand increasingly more diagnostic and surgical interventions that require high-level specialized techniques.

  20. Detection of monoclonal integration of bovine leukemia virus proviral DNA as a malignant marker in two enzootic bovine leukosis cases with difficult clinical diagnosis

    OpenAIRE

    Miura, Saori; HORIUCHI, Noriyuki; Matsumoto, Kotaro; KOBAYASHI, Yoshiyasu; Kawazu, Shin-ichiro; INOKUMA, Hisashi

    2015-01-01

    Monoclonal integration of bovine leukemia virus (BLV) proviral DNA into bovine genomes was detected in peripheral blood from two clinical cases of enzootic bovine leukosis (EBL) without enlargement of superficial lymph nodes. A BLV-specific probe hybridized with 1 to 3 EcoRI and HindIII fragments in these 2 atypical EBL cattle by Southern blotting and hybridization, as well as in 3 typical EBL cattle. The probe also hybridized to a large number of EcoRI and HindIII fragments in 5 cattle with ...

  1. Severe exercise and exercise training exert opposite effects on human neutrophil apoptosis via altering the redox status.

    Directory of Open Access Journals (Sweden)

    Guan-Da Syu

    Full Text Available Neutrophil spontaneous apoptosis, a process crucial for immune regulation, is mainly controlled by alterations in reactive oxygen species (ROS and mitochondria integrity. Exercise has been proposed to be a physiological way to modulate immunity; while acute severe exercise (ASE usually impedes immunity, chronic moderate exercise (CME improves it. This study aimed to investigate whether and how ASE and CME oppositely regulate human neutrophil apoptosis. Thirteen sedentary young males underwent an initial ASE and were subsequently divided into exercise and control groups. The exercise group (n = 8 underwent 2 months of CME followed by 2 months of detraining. Additional ASE paradigms were performed at the end of each month. Neutrophils were isolated from blood specimens drawn at rest and immediately after each ASE for assaying neutrophil spontaneous apoptosis (annexin-V binding on the outer surface along with redox-related parameters and mitochondria-related parameters. Our results showed that i the initial ASE immediately increased the oxidative stress (cytosolic ROS and glutathione oxidation, and sequentially accelerated the reduction of mitochondrial membrane potential, the surface binding of annexin-V, and the generation of mitochondrial ROS; ii CME upregulated glutathione level, retarded spontaneous apoptosis and delayed mitochondria deterioration; iii most effects of CME were unchanged after detraining; and iv CME blocked ASE effects and this capability remained intact even after detraining. Furthermore, the ASE effects on neutrophil spontaneous apoptosis were mimicked by adding exogenous H(2O(2, but not by suppressing mitochondrial membrane potential. In conclusion, while ASE induced an oxidative state and resulted in acceleration of human neutrophil apoptosis, CME delayed neutrophil apoptosis by maintaining a reduced state for long periods of time even after detraining.

  2. Expression of IL-1ra mRNA and dysfunction of phagocytosis and intraceilular killing of peripheral blood neutrophils in HD patients%维持性血透患者外周血中性粒细胞的IL-1ra基因表达和吞噬杀菌功能

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    目的:对维持性血液透析(HD)患者外周血中性粒细胞(PMN)白介素-1受体拮抗物(IL-1ra)基因表达和吞噬及杀菌功能进行研究,并探讨二者的相关性。方法:用RT-PCR方法检测HD患者外周血PMNicIL-1ra(胞内型)和sIL-1ra(分泌型)基因表达;通过菌落计数方法观察PMN对金黄色葡萄球菌(金葡菌)吞噬和杀菌功能的改变。结果:HD患者PMN中可检测到512bp(icIL-1ra)和146bp(sIL-1ra)的RT-PCR产物;HD患者PMN对金葡菌的吞噬和胞内杀菌能力均较正常对照明显下降(P<0.01或P<0.05)。结论:HD患者PMN可自发表达icIL-1ra和sIL-1ra,这种预激活状态与其对金葡菌的吞噬和杀灭作用下降相平行。%Objective:To investigate the function and IL-1ra gene expressionof neutrophils in hemodialysis (HD) patients.Methods:Gene expression was studied by reverse transcribe polymerase chain reaction (RT-PCR) from untreated neutrophils in healthy controls and HD patients.Phagocytosis and bactericidal assays decreased in the number of viable extracellular and intracellular bacteria by colony counts.Results:Freshly isolated neutrophils spontaneously expressed icIL-1ra,sIL-1ra mRNA in HD patients.Phagocytosis and intracellular killing of neutrophils in HD patients impaired obviously when exposed to staphylococcus aureus in comparison with healthy controls.Conclusion:Untreated neutrophils in HD patients spontaneously express iclL-1ra and sIL-1ra mRNA,which is consistent with the suppressed phagocytosis and intracellular killing capacity.

  3. Neutrophil extracellular traps in tissue pathology.

    Science.gov (United States)

    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.

  4. Leukotriene B4-Neutrophil Elastase Axis Drives Neutrophil Reverse Transendothelial Cell Migration In Vivo.

    Science.gov (United States)

    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.

  5. Leukotriene B4-Neutrophil Elastase Axis Drives Neutrophil Reverse Transendothelial Cell Migration In Vivo

    Science.gov (United States)

    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

  6. Autophagy is induced by anti-neutrophil cytoplasmic Abs and promotes neutrophil extracellular traps formation.

    Science.gov (United States)

    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.

  7. Phagocytosis and killing of Staphylococcus aureus by human neutrophils.

    Science.gov (United States)

    Lu, Thea; Porter, Adeline R; Kennedy, Adam D; Kobayashi, Scott D; DeLeo, Frank R

    2014-01-01

    Neutrophils are essential for host defense against Staphylococcus aureus infections. Although significant progress has been made, our understanding of neutrophil interactions with S. aureus remains incomplete. To provide a more comprehensive view of this process, we investigated phagocytosis and killing of S. aureus by human neutrophils using varied assay conditions in vitro. A greater percentage of bacteria were internalized by adherent neutrophils compared to those in suspension, and, unexpectedly, uptake of S. aureus by adherent neutrophils occurred efficiently in the absence of opsonins. An antibody specific for S. aureus promoted uptake of unopsonized bacteria in suspension, but had little or no capacity to enhance phagocytosis of S. aureus opsonized with normal human serum or by adherent neutrophils. Collectively, these results indicate that assay conditions can have a significant influence on the phagocytosis and killing of S. aureus by neutrophils. More importantly, the results suggest a vaccine approach directed to enhance opsonophagocytosis alone is not sufficient to promote increased killing of S. aureus by human neutrophils. With the emergence and reemergence of antibiotic-resistant microorganisms, establishing parameters that are optimal for studying neutrophil-S. aureus interactions will pave the way towards developing immune-directed strategies for anti-staphylococcal therapies.

  8. Human exposure to bovine polyomavirus: a zoonosis

    Energy Technology Data Exchange (ETDEWEB)

    Parry, J.V.; Gardner, S.D.

    1986-01-01

    A competitive-type solid phase radioimmunoassay (RIA) was developed for the detection of antibody to bovine polyomavirus. Comparison of RIA and counter-immunoelectrophoresis (CIE) results on 273 cattle sera indicated that both techniques were detecting antibody of like specificity. Human sera from 256 blood donors, 219 people recently vaccinated against polio, rubella or rabies, 50 immunosuppressed patients and 472 people with various occupational exposure to cattle were tested for antibody to bovine polyomavirus, the foetal rhesus monkey kidney strain, (anti-FRKV) by RIA. Apart from one blood donor and one of 108 rabies vaccinees only those in close contact with cattle possessed anti-FRKV. Compared with 62 per cent seropositive in the natural hosts, cattle, 71 per cent of veterinary surgeons, 50 per cent of cattle farmers, 40 per cent of abattoir workers, 16 per cent of veterinary institute technical staff and 10 per cent of veterinary students were anti-FRKV positive. Our findings indicate that the theoretical hazard of FRKV infection from undetected contamination of current tissue culture derived vaccines may, in practice, be remote. Proposed wider use of primate kidney cells as substrates for new vaccines may increase this risk.

  9. Calprotectin mRNA (MRP8/MRP14 expression in neutrophils of periodontitis patients with type 2 diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Ahmad Syaify

    2009-09-01

    Full Text Available Background: Calprotectin, a major cytosolic protein of leukocytes, is detected in neutrophils and monocytes/machrophages. This protein is known to be a marker for several inflammatory diseases including periodontitis. In type 2 diabetes mellitus patients, the severity of periodontitis was strongly thought to be caused by decreasing of leukocytes function such as neutrophils. Previous research found that the calprotectin level in serum of periodontitis patients with type 2 DM is higher than periodontits patients non DM. Purpose: The aim of this study was to determine calprotectin mRNA (MRP8/MRP14 expression in human neutrophils of periodontitis patients with type 2 diabetes mellitus. Methods: Neutrophils were isolated from the peripheral blood of periodontitis patients with uncontrolled type 2 DM, controlled type 2 DM, and non DM. The expression of calprotectin mRNA (MRP8 and MRP14 were detected by RTPCR. Result: The result showed that the value of mRNA calprotectin expression in DM patients were higher than non DM, and the highest expression was on the uncontrolled type 2 DM. Conclusion: The basal level of calprotectin mRNA MRP8/MRP14 expression increased in neutrophil of periodontitis patient with type 2 DM compared non diabetic subjects. It was suggested that high basal level of calprotectin mRNA has a role in the regulation of periodontitis severity with diabetes mellitus patients.

  10. Effect of academic psychological stress in post-graduate students: the modulatory role of cortisol on superoxide release by neutrophils.

    Science.gov (United States)

    Ignacchiti, M D C; Sesti-Costa, R; Marchi, L F; Chedraoui-Silva, S; Mantovani, B

    2011-05-01

    Experimental and clinical evidence shows that neutrophils play an important role in the mechanism of tissue injury in immune complex diseases through the generation of reactive oxygen species. In this study, we examined the influence of academic psychological stress in post-graduate students on the capacity of their blood neutrophils to release superoxide when stimulated by immune complexes bound to nonphagocytosable surfaces and investigated the modulatory effect of cortisol on this immune function. The tests were performed on the day before the final examination. The state-trait anxiety inventory questionnaire was used to examine whether this stressful event caused emotional distress. In our study, the psychological stress not only increased plasma cortisol concentration, but it also provoked a reduction in superoxide release by neutrophils. This decrease in superoxide release was accompanied by diminished mRNA expression for subunit p47(phox) of the phagocyte superoxide-generating nicotinamide adenine dinucleotide phosphate-oxidase. These inhibitory effects were also observed by in vitro exposure of neutrophils from control volunteers to 10(- 7) M hydrocortisone, and could be prevented by the glucocorticoid receptor antagonist RU-486. These results show that in a situation of psychological stress, the increased levels of cortisol could inhibit superoxide release by neutrophils stimulated by IgG immune complexes bound to nonphagocytosable surfaces, which could attenuate the inflammatory state.

  11. Treadmill exercise induces neutrophil recruitment into muscle tissue in a reactive oxygen species-dependent manner. An intravital microscopy study.

    Directory of Open Access Journals (Sweden)

    Albená Nunes-Silva

    Full Text Available 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.

  12. Scavenging properties of neutrophil 4-hydroxyphenylpyruvate dioxygenase are based on a hypothesis that does not stand up to scrutiny.

    Science.gov (United States)

    Salerno, Costantino; Zicari, Alessandra; Mari, Emanuela; D'Eufemia, Patrizia

    2014-10-01

    It was previously reported by D'Eufemia et al. [9] that neutrophil preparations from a patient with tyrosinemia type III, i.e. with inherited deficiency of 4-hydroxyphenylpyruvate dioxygenase (HPPD), exhibited a far higher NO release than controls, when NO was estimated in terms of nitrite content in the suspending media. It was hypothesized that HPPD might participate to NO sequestration in neutrophils and that excessive NO release might reflect the lack of the scavenging action in defective cells. In recent control experiments, we found that HPPD activity in neutrophils preparations from healthy subjects is below the detection limit of the enzymatic assay (less than 3nmol product/h per mg protein). This indicates that HPPD concentration in neutrophils is very low, if any, confirming what was already suggested in literature, and rules out the possibility of a prominent role of HPPD as NO scavenger in these cells. Moreover, we found that 500μM l-tyrosine increases nitrite release and accumulation in suspending media of U-937 cells, a human monoblast-like lymphoma cell line which displays many characteristics of macrophages, including the expression of inducible and endothelial nitric oxide synthases. We hypothesize that the increase of nitrite release by patient's neutrophils might be related to the presence of high l-tyrosine concentrations in the blood samples (426μmol/L instead of 52.1±10.9μmol/L as healthy subjects), rather than to HPPD deficiency of in these cells.

  13. Flow-cytochemical differential leukocyte analysis with quantitation of neutrophil left shift. An evaluation of the Cobas-Helios analyzer.

    Science.gov (United States)

    Bentley, S A; Johnson, T S; Sohier, C H; Bishop, C A

    1994-08-01

    The Cobas-Helios (Roche Diagnostic Systems, Inc., Branchburg, NJ) is a new, fully automated hematology analyzer that performs a complete blood count and differential leukocyte count (DLC), classifying leukocytes by flow-cytochemical technology. The DLC component of the Cobas-Helios was evaluated according to the National Committee for Clinical Laboratory Standards H20-A protocol. Instrument performance was acceptable with respect to all parameters investigated, including imprecision, inaccuracy and clinical sensitivity for the identification of quantitative and qualitative leukocyte abnormalities. In a minority of samples with neutrophil left shift, neutrophils tended to overlap the monocyte domain, resulting in overestimation of monocytes and underestimation of neutrophils. This problem did not affect clinical sensitivity and was generally associated with a positive instrumental left-shift flag. Flags for the identification of specific qualitative abnormalities of the leukocyte population (atypical lymphoid cells, nucleated red cells, blast cells, immature granulocytes and neutrophil left shift) performed well. In addition to a conventional five-part DLC, the Cobas-Helios also identifies and quantitates atypical lymphoid cells and "large immature cells," the latter corresponding to bands and immature granulocytes. Counts of atypical lymphoid cells and large immature cells correlated well with the equivalent cell classes as enumerated by the reference method of the National Committee for Clinical Laboratory Standards. The Cobas-Helios offers the most reliable quantitative index of neutrophil left shift currently available in a commercial automated DLC analyzer.

  14. Bovine coronavirus hemagglutinin protein.

    Science.gov (United States)

    King, B; Potts, B J; Brian, D A

    1985-02-01

    Treatment of purified bovine coronavirus (Mebus strain) with pronase destroyed the integrity of virion surface glycoproteins gp140, gp120, gp100, reduced the amount of gp26 and destroyed the hemagglutinating activity of the virus. Bromelain, on the other hand, destroyed the integrity of gp120, gp100 and gp26 but failed to remove gp140 and failed to destroy viral hemagglutinating activity. These experiments suggest that gp140 is the virion hemagglutinin. Immunoblotting studies using monospecific antiserum demonstrate that gp140 is a disulfide-linked dimeric structure reducible to monomers of 65 kDa.

  15. Infectious bovine keratoconjunctivitis (pinkeye).

    Science.gov (United States)

    Angelos, John A

    2015-03-01

    As is the case for controlling other infectious livestock diseases, the most successful efforts to control infectious bovine keratoconjunctivitis (IBK) will include consideration of the host, the environment, herd management, and ongoing surveillance even after the immediate crisis has passed. Research over many years has led to the discovery of a variety of antibiotic treatments and antibiotic regimens that can be effective against IBK. The discoveries of Mor bovoculi and reports of IBK associated with Mycoplasma spp without concurrent Mor bovis or Mor bovoculi have raised new questions into the roles that other organisms may play in IBK pathogenesis.

  16. Proteomic Analysis of Bovine Nucleolus

    Institute of Scientific and Technical Information of China (English)

    Amrutlal K.Patel; Doug Olson; Suresh K. Tikoo

    2010-01-01

    Nucleolus is the most prominent subnuclear structure, which performs a wide variety of functions in the eu-karyotic cellular processes. In order to understand the structural and functional role of the nucleoli in bovine cells,we analyzed the proteomie composition of the bovine nueleoli. The nucleoli were isolated from Madin Darby bo-vine kidney cells and subjected to proteomie analysis by LC-MS/MS after fractionation by SDS-PAGE and strongcation exchange chromatography. Analysis of the data using the Mascot database search and the GPM databasesearch identified 311 proteins in the bovine nucleoli, which contained 22 proteins previously not identified in theproteomic analysis of human nucleoli. Analysis of the identified proteins using the GoMiner software suggestedthat the bovine nueleoli contained proteins involved in ribosomal biogenesis, cell cycle control, transcriptional,translational and post-translational regulation, transport, and structural organization.

  17. Combined anti CXC receptors 1 and 2 therapy is a promising anti-inflammatory treatment for respiratory diseases by reducing neutrophil migration and activation.

    Science.gov (United States)

    Planagumà, A; Domènech, T; Pont, M; Calama, E; García-González, V; López, R; Aulí, M; López, M; Fonquerna, S; Ramos, I; de Alba, J; Nueda, A; Prats, N; Segarra, V; Miralpeix, M; Lehner, M D

    2015-10-01

    Neutrophil infiltration and activation in the lung are important pathophysiological features in COPD, severe asthma and bronchiectasis mostly mediated by CXCL8 and CXCL1 via CXCR1 and CXCR2. No thorough study to date has been performed to compare the anti-inflammatory effect profile of dual CXCR1/2 vs. selective CXCR2 antagonists in relevant human neutrophil assays and pulmonary inflammation models. Dual CXCR1/2 (SCH527123, diaminocyclobutandione-1) and selective CXCR2 (SB265610, thiopyrimidine-1) antagonist activity and receptor residence time were determined by [(35)S]GTPγS binding in human (h)- and guinea pig (gp)-CXCR1 and CXCR2 overexpressing membranes. h-neutrophil chemotaxis, degranulation and ROS production were established using CXCL8 or CXCL1 to evaluate dual CXCR1/2- or selective CXCR2-dependent activities. LPS-induced lung inflammation in gp was selected to assess in vivo potency. Dual CXCR1/2 antagonists blocked both CXCL8 and CXCL1-induced h-neutrophil functions and [(35)S]GTPγS binding. In contrary, selective CXCR2 antagonists displayed significantly reduced potency in CXCL8 -mediated h-neutrophil responses despite being active in CXCR2 assays. Upon LPS challenge in gp, administration of SCH527123 inhibited the increase of neutrophils in BALF, modestly reduced blood neutrophils and induced minor neutrophil accumulation in bone marrow. Differentiation of CXCR1/2 vs. CXCR2 antagonists could not be extended to in vivo due to differences in CXCR1 receptor homology between h and gp. Dual CXCR1/2 therapy may represent a promising anti-inflammatory treatment for respiratory diseases reducing more effectively neutrophil migration and activation in the lung than a CXCR2 selective treatment. However, the in vivo confirmation of this claim is still missing due to species differences in CXCR1.

  18. Comparison of Neutrophil Apoptosis by the Pseudomonas Aeruginosa Exotoxins between Healthy Individuals and Term Infants

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    Soheila Khazaei

    2013-04-01

    Full Text Available Background: Pseudomonas aeruginosa may be colonized in different human tissues and result in some infections potentially. Thus, considering that these bacteria are resistance to most of the current antibiotics, an examination on pathogenesis mechanisms of such bacteria can be effective in controlling the infections developed by it.Materials and Methods: In this project, among 40 blood samples (20 healthy persons, 20 infants, an amount of 5 ml (2 ml in the infants heparinized blood was collected form each and then neutrophils were isolated by a standard method and were counted by neubauer lam. After culturing Pseudomonas bacteria in broth medium, some tubes with densities of 1, 2, 3 and 4 McFarland were prepared and the bacteria were isolated by centrifuge method with 3000rpm for 10 minutes and then its exotoxin were exposed to neutrophils of the groups under study. The effect of time and the bacteria count on the amount of the secreted toxin and in adjacency to neutrophils was measured.Results: There were 11 men and 9 women in the health group and the infants group consisted of 12 boys and 8 girls. Death cell percentage of neutrophils was 100% in the health group and 8.90% in the infants group. Percentage of bacterial growth in the medium 1 and 2 McFarland was zero; in the medium 3 McFarland, it was 12.5% in the healthy group and 1% in the infants group (p<0.10. The average rate of cell death in the minute 15th was different in two groups (68.5% in health group vs. 92.5% in the infants (p<0.0005. Conclusion: This study showed the effect of Pseudomonas bacteria on the development of early cell death in the infants very well. As it was shown, this effect is time-dependent and this cell death (apoptosis is occurred in the infants earlier than health people.

  19. Viral infections and bovine mastitis: a review

    NARCIS (Netherlands)

    Wellenberg, G.J.; Poel, van der W.H.M.; Oirschot, van J.T.

    2002-01-01

    This review deals with the role of viruses in the aetiology of bovine mastitis. Bovine herpesvirus 1, bovine herpesvirus 4, foot-and-mouth disease virus, and parainfluenza 3 virus have been isolated from milk from cows with clinical mastitis. Intramammary inoculations of bovine herpesvirus 1 or para

  20. Fast detection and determination of bovine viral diarrhea virus in blood samples%进口奶牛病毒性腹泻病毒快速检测鉴定

    Institute of Scientific and Technical Information of China (English)

    赵秀玲; 闻伟刚; 黄素文; 张吉红; 杜华宏

    2007-01-01

    采用ELISA方法对3 073头奶牛全血进行牛病毒性腹泻病毒(Bovine viral diarrhea virus,BVDV)检测,结果发现编号a889的血样呈阳性反应,另外有3份血样(a126、a803、b1277)呈可疑反应;进一步的白细胞抽提物ELISA反应表明,a889血样呈阳性反应,其余呈阴性反应.对a889血样进行RT-PCR扩增,得到1条310 bp大小的特异性条带,与预期片段大小吻合;经序列同源性比较分析,确定a889血样中存在牛病毒性腹泻病毒.

  1. Mechanism of neutrophil recruitment to the lung after pulmonary contusion.

    Science.gov (United States)

    Hoth, J Jason; Wells, Jonathan D; Hiltbold, Elizabeth M; McCall, Charles E; Yoza, Barbara K

    2011-06-01

    Blunt chest trauma resulting in pulmonary contusion is a common but poorly understood injury. We previously demonstrated that lung contusion activates localized and systemic innate immune mechanisms and recruits neutrophils to the injured lung. We hypothesized that the innate immune and inflammatory activation of neutrophils may figure prominently in the response to lung injury. To investigate this, we used a model of pulmonary contusion in the mouse that is similar to that observed clinically in humans and evaluated postinjury lung function and pulmonary neutrophil recruitment. Comparisons were made between injured mice with and without neutrophil depletion. We further examined the role of chemokines and adhesion receptors in neutrophil recruitment to the injured lung. We found that lung injury and resultant physiological dysfunction after contusion were dependent on the presence of neutrophils in the alveolar space. We show that CXCL1, CXCL2/3, and CXCR2 are involved in neutrophil recruitment to the lung after injury and that intercellular adhesion molecule 1 is locally expressed and actively participates in this process. Injured gp91-deficient mice showed improved lung function, indicating that oxidant production by neutrophil NADPH oxidase mediates lung dysfunction after contusion. These data suggest that both neutrophil presence and function are required for lung injury after lung contusion.

  2. Specificity and Effector Functions of Human RSV-Specific IgG from Bovine Milk.

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    Gerco den Hartog

    Full Text Available BACKGROUND: Respiratory syncytial virus (RSV infection is the second most important cause of death in the first year of life, and early RSV infections are associated with the development of asthma. Breastfeeding and serum IgG have been shown to protect against RSV infection. Yet, many infants depend on bovine milk-based nutrition, which at present lacks intact immunoglobulins. OBJECTIVE: To investigate whether IgG purified from bovine milk (bIgG can modulate immune responses against human RSV. METHODS: ELISAs were performed to analyse binding of bIgG to human respiratory pathogens. bIgG or hRSV was coated to plates to assess dose-dependent binding of bIgG to human Fcγ receptors (FcγR or bIgG-mediated binding of myeloid cells to hRSV respectively. S. Epidermidis and RSV were used to test bIgG-mediated binding and internalisation of pathogens by myeloid cells. Finally, the ability of bIgG to neutralise infection of HEp2 cells by hRSV was evaluated. RESULTS: bIgG recognised human RSV, influenza haemagglutinin and Haemophilus influenza. bIgG bound to FcγRII on neutrophils, monocytes and macrophages, but not to FcγRI and FcγRIII, and could bind simultaneously to hRSV and human FcγRII on neutrophils. In addition, human neutrophils and dendritic cells internalised pathogens that were opsonised with bIgG. Finally, bIgG could prevent infection of HEp2 cells by hRSV. CONCLUSIONS: The data presented here show that bIgG binds to hRSV and other human respiratory pathogens and induces effector functions through binding to human FcγRII on phagocytes. Thus bovine IgG may contribute to immune protection against RSV.

  3. Mast cell activation and neutrophil recruitment promotes early and robust inflammation in the meninges in EAE.

    Science.gov (United States)

    Christy, Alison L; Walker, Margaret E; Hessner, Martin J; Brown, Melissa A

    2013-05-01

    The meninges are often considered inert tissues that house the CSF and provide protection for the brain and spinal cord. Yet emerging data demonstrates that they are also active sites of immune responses. Furthermore, the blood-CSF barrier surrounding meningeal blood vessels, together with the blood-brain barrier (BBB), is postulated to serve as a gateway for the pathological infiltration of immune cells into the CNS in multiple sclerosis (MS). Our previous studies using mast cell-deficient (Kit(W/Wv)) mice demonstrated that mast cells resident in the dura mater and pia mater exacerbate experimental autoimmune encephalomyelitis (EAE), a rodent model of MS, by facilitating CNS inflammatory cell influx. Here we examined the underlying mechanisms that mediate these effects. We demonstrate that there are dramatic alterations in immune associated gene expression in the meninges in pre-clinical disease, including those associated with mast cell and neutrophil function. Meningeal mast cells are activated within 24 h of disease induction, but do not directly compromise CNS vascular integrity. Rather, through production of TNF, mast cells elicit an early influx of neutrophils, cells known to alter vascular permeability, into the meninges. These data add to the growing evidence that inflammation in the meninges precedes CNS immune cell infiltration and establish that mast cells are among the earliest participants in these disease-initiating events. We hypothesize that mast cell-dependent neutrophil recruitment and activation in the meninges promotes early breakdown of the local BBB and CSF-blood barrier allowing initial immune cell access to the CNS.

  4. Effect of lemon verbena supplementation on muscular damage markers, proinflammatory cytokines release and neutrophils' oxidative stress in chronic exercise.

    Science.gov (United States)

    Funes, Lorena; Carrera-Quintanar, Lucrecia; Cerdán-Calero, Manuela; Ferrer, Miguel D; Drobnic, Franchek; Pons, Antoni; Roche, Enrique; Micol, Vicente

    2011-04-01

    Intense exercise is directly related to muscular damage and oxidative stress due to excessive reactive oxygen species (ROS) in both, plasma and white blood cells. Nevertheless, exercise-derived ROS are essential to regulate cellular adaptation to exercise. Studies on antioxidant supplements have provided controversial results. The purpose of this study was to determine the effect of moderate antioxidant supplementation (lemon verbena extract) in healthy male volunteers that followed a 90-min running eccentric exercise protocol for 21 days. Antioxidant enzymes activities and oxidative stress markers were measured in neutrophils. Besides, inflammatory cytokines and muscular damage were determined in whole blood and serum samples, respectively. Intense running exercise for 21 days induced antioxidant response in neutrophils of trained male through the increase of the antioxidant enzymes catalase, glutathione peroxidase and glutathione reductase. Supplementation with moderate levels of an antioxidant lemon verbena extract did not block this cellular adaptive response and also reduced exercise-induced oxidative damage of proteins and lipids in neutrophils and decreased myeloperoxidase activity. Moreover, lemon verbena supplementation maintained or decreased the level of serum transaminases activity indicating a protection of muscular tissue. Exercise induced a decrease of interleukin-6 and interleukin-1β levels after 21 days measured in basal conditions, which was not inhibited by antioxidant supplementation. Therefore, moderate antioxidant supplementation with lemon verbena extract protects neutrophils against oxidative damage, decreases the signs of muscular damage in chronic running exercise without blocking the cellular adaptation to exercise.

  5. Shielding of a lipooligosaccharide IgM epitope allows evasion of neutrophil-mediated killing of an invasive strain of nontypeable Haemophilus influenzae.

    Science.gov (United States)

    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

  6. Gene deletion of P-Selectin and ICAM-1 does not inhibit neutrophil infiltration into peritoneal cavity following cecal ligation-puncture

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    Hess Karen

    2004-07-01

    Full Text Available Abstract Background Neutrophil infiltration is one of the critical cellular components of an inflammatory response during peritonitis. The adhesion molecules, P-selectin and intercellular adhesion molecule (ICAM-1, mediate neutrophil-endothelial cell interactions and the subsequent neutrophil transendothelial migration during the inflammatory response. Despite very strong preclinical data, recent clinical trials failed to show a protective effect of anti-adhesion therapy, suggesting that the length of injury might be a critical factor in neutrophil infiltration. Therefore, the objective of this study was to determine the role of P-selectin and ICAM-1 in neutrophil infiltration into the peritoneal cavity during early and late phases of peritonitis. Methods Peritonitis was induced in both male wild-type and P-selectin/ICAM-1 double deficient (P/I null mice by cecal ligation-puncture (CLP. Peripheral blood and peritoneal lavage were collected at 6 and 24 hours after CLP. The total leukocyte and neutrophil contents were determined, and neutrophils were identified with the aid of in situ immunohistochemical staining. Comparisons between groups were made by applying ANOVA and student t-test analysis. Results CLP induced a severe inflammatory response associated with a significant leukopenia in both wild-type and P/I null mice. Additionally, CLP caused a significant neutrophil infiltration into the peritoneal cavity that was detected in both groups of mice. However, neutrophil infiltration in the P/I null mice at 6 hours of CLP was significantly lower than the corresponding wild-type mice, which reached a similar magnitude at 24 hours of CLP. In contrast, in peritonitis induced by intraperitoneal inoculation of 2% glycogen, no significant difference in neutrophil infiltration was observed between the P/I null and wild-type mice at 6 hours of peritonitis. Conclusions The data suggest that alternative adhesion pathway(s independent of P-selectin and ICAM

  7. Systemic signs of neutrophil mobilization during clinically stable periods and during exacerbations in smokers with obstructive pulmonary disease

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    Andelid K

    2015-06-01

    Full Text Available Kristina Andelid,1 Anders Andersson,1 Shigemi Yoshihara,2 Christina Åhrén,4 Pernilla Jirholt,3 Ann Ekberg-Jansson,1 Anders Lindén1,51Department of Internal Medicine and Clinical Nutrition, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden; 2Department of Pediatrics, Dokkyo Medical University, Japan; 3Department of Rheumatology and Inflammation Research, Institute of Medicine, 4Department of Bacteriology, Institute of Laboratory Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg; 5Unit for Lung and Airway Research, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, SwedenBackground: It is still unclear whether signs of neutrophil mobilization in the blood of patients with chronic obstructive pulmonary disease represent true systemic events and how these relate to bacterial colonization in the airways. In this study, we evaluated these issues during clinically stable periods and during exacerbations in smokers with obstructive pulmonary disease and chronic bronchitis (OPD-CB.Methods: Over a period of 60 weeks for each subject, blood samples were repeatedly collected from 60 smokers with OPD-CB during clinically stable periods, as well as during and after exacerbations. Myeloperoxidase (MPO and neutrophil elastase (NE protein and mRNA, growth of bacteria in sputum, and clinical parameters were analyzed. Ten asymptomatic smokers and ten never-smokers were included as controls.Results: We found that, during clinically stable periods, neutrophil and NE protein concentrations were increased in smokers with OPD-CB and in the asymptomatic smokers when compared with never-smokers. During exacerbations, neutrophil and MPO protein concentrations were further increased in smokers with OPD-CB, without a detectable increase in the corresponding mRNA during exacerbations. However, MPO and NE protein and mRNA displayed positive correlations. During exacerbations, only increased neutrophil

  8. Immune response to bovine viral diarrhea virus--looking at newly defined targets.

    Science.gov (United States)

    Chase, Christopher C L; Thakur, Neelu; Darweesh, Mahmoud F; Morarie-Kane, Susan E; Rajput, Mrigendra K

    2015-06-01

    Bovine viral diarrhea virus (BVDV) has long been associated with a wide variety of clinical syndromes and immune dysregulation, many which result in secondary bacterial infections. Current understanding of immune cell interactions that result in activation and tolerance are explored in light of BVDV infection including: depletion of lymphocytes, effects on neutrophils, natural killer cells, and the role of receptors and cytokines. In addition, we review some new information on the effect of BVDV on immune development in the fetal liver, the role of resident macrophages, and greater implications for persistent infection.

  9. Espectrofotometria de proteínas totais em plasma de sangue bovino por análise em fluxo Spectrophotometry of total protein in bovine blood plasma by flow analysis

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    Gilmara Caseri de Luca

    2002-06-01

    handling during operation. In this work an automated flow procedure for total protein determination in bovine serum samples, employing the Biuret method, was developed. The system, including an automatic injector commutator and a three way solenoid valve computer controlled, was designed in order to permit on line sample dilution. Since protein standard solutions were introduced with and without dilution. A dilution factor was estimated and used to calculate protein concentration obtained after on-line dilution of the samples. Solutions samples were inserted through a three way solenoid valve that allows precise dilution minimizing hand operations. The analytical range for total protein determination was 2.5 to 20.0 g L-1, and considering the dilution employed, bovine plasma samples with 12.5 to 100.0 g L-1 of total protein were analysed. The procedure presented a 2.8% rsd and analytical frequency of 76 determinations per hour. Results were in good agreement with the traditional method (Biuret and no significant difference was verified at 95%.

  10. Bovine herpesvirus 1 infection and infectious bovine rhinotracheitis

    OpenAIRE

    2007-01-01

    International audience; Bovine herpesvirus 1 (BoHV-1), classified as an alphaherpesvirus, is a major pathogen of cattle. Primary infection is accompanied by various clinical manifestations such as infectious bovine rhinotracheitis, abortion, infectious pustular vulvovaginitis, and systemic infection in neonates. When animals survive, a life-long latent infection is established in nervous sensory ganglia. Several reactivation stimuli can lead to viral re-excretion, which is responsible for the...

  11. Blood sugar test - blood

    Science.gov (United States)

    ... blood glucose level ( hypoglycemia ) may be due to: Hypopituitarism (a pituitary gland disorder) Underactive thyroid gland or ... tonic-clonic seizure Glucagon blood test Glucagonoma Hyperthyroidism Hypopituitarism Hypothyroidism Insulinoma Low blood sugar Multiple endocrine neoplasia ( ...

  12. Intervet symposium: bovine neosporosis.

    Science.gov (United States)

    Schetters, T; Dubey, J P; Adrianarivo, A; Frankena, K; Romero, J J; Pérez, E; Heuer, C; Nicholson, C; Russell, D; Weston, J

    2004-10-28

    This article summarises the most relevant data of presentations delivered at the 19th International Conference of the World Association for the Advancement of Veterinary Parasitology (WAAVP)held in New Orleans, LA, USA, from 10 to 14 August 2003) in a symposium session on bovine neosporosis. The symposium was organised by Juan Muñoz-Bielsa,Wicher Holland, Enzo Foccoliand Theo Schetters (chairman). The focus was on the present state of knowledge of the biology, epidemiology(presented by J.P. Dubey) and immunology of Neospora infection (presented by A. Adrianarivo),with special emphasis on the prospects of vaccination of cattle against Neospora-induced abortion (presentations of K. Frankena (Costa Rican trial) and C. Heuer (New Zealand trial)).

  13. Polymorphonuclear neutrophils: an effective antimicrobial force.

    Science.gov (United States)

    Sawyer, D W; Donowitz, G R; Mandell, G L

    1989-01-01

    The production and deployment of polymorphonuclear neutrophils (PMNs) are under close regulation. PMNs interact through cytokines with a number of cell types, including macrophages, lymphocytes, and endothelial cells. PMNs are guided by bacterial products and cytokines to target sites, where microbes are recognized and killed. Killing occurs through oxygen-dependent and oxygen-independent mechanisms. The frequent and severe infections seen in patients with defects (either congenital or acquired) in PMN function demonstrate the importance of PMNs in host defense against infection. PMNs are potent inflammatory cells and can exacerbate disease states such as myocardial ischemia, gram-negative bacterial sepsis, and the adult respiratory distress syndrome.

  14. A scabies mite serpin interferes with complement-mediated neutrophil functions and promotes staphylococcal growth.

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    Pearl M Swe

    2014-06-01

    Full Text Available BACKGROUND: Scabies is a contagious skin disease caused by the parasitic mite Sarcoptes scabiei. The disease is highly prevalent worldwide and known to predispose to secondary bacterial infections, in particular by Streptococcus pyogenes and Staphylococcus aureus. Reports of scabies patients co-infected with methicillin resistant S. aureus (MRSA pose a major concern for serious down-stream complications. We previously reported that a range of complement inhibitors secreted by the mites promoted the growth of S. pyogenes. Here, we show that a recently characterized mite serine protease inhibitor (SMSB4 inhibits the complement-mediated blood killing of S. aureus. METHODOLOGY/PRINCIPAL FINDINGS: Blood killing of S. aureus was measured in whole blood bactericidal assays, counting viable bacteria recovered after treatment in fresh blood containing active complement and phagocytes, treated with recombinant SMSB4. SMSB4 inhibited the blood killing of various strains of S. aureus including methicillin-resistant and methicillin-sensitive isolates. Staphylococcal growth was promoted in a dose-dependent manner. We investigated the effect of SMSB4 on the complement-mediated neutrophil functions, namely phagocytosis, opsonization and anaphylatoxin release, by flow cytometry and in enzyme linked immuno sorbent assays (ELISA. SMSB4 reduced phagocytosis of S. aureus by neutrophils. It inhibited the deposition of C3b, C4b and properdin on the bacteria surface, but did not affect the depositions of C1q and MBL. SMSB4 also inhibited C5 cleavage as indicated by a reduced C5b-9 deposition. CONCLUSIONS/SIGNIFICANCE: We postulate that SMSB4 interferes with the activation of all three complement pathways by reducing the amount of C3 convertase formed. We conclude that SMSB4 interferes with the complement-dependent killing function of neutrophils, thereby reducing opsonization, phagocytosis and further recruitment of neutrophils to the site of infection. As a

  15. Respiratory syncytial virus fusion protein promotes TLR-4-dependent neutrophil extracellular trap formation by human neutrophils.

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    Giselle A Funchal

    Full Text Available 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.

  16. Gr-1dimCD11b+ immature myeloid-derived suppressor cells but not neutrophils are markers of lethal tuberculosis infection in mice.

    Science.gov (United States)

    Tsiganov, Evgeny N; Verbina, Elena M; Radaeva, Tatiana V; Sosunov, Vasily V; Kosmiadi, George A; Nikitina, Irina Yu; Lyadova, Irina V

    2014-05-15

    Tuberculosis (TB) disease may progress at different rates and have different outcomes. Neutrophils have been implicated in TB progression; however, data on their role during TB are controversial. In this study, we show that in mice, TB progression is associated with the accumulation of cells that express neutrophilic markers Gr-1 and Ly-6G but do not belong to conventional neutrophils. The cells exhibit unsegmented nuclei, have Gr-1(dim)Ly-6G(dim)CD11b(+) phenotype, and express F4/80, CD49d, Ly-6C, CD117, and CD135 markers characteristic not of neutrophils but of immature myeloid cells. The cells accumulate in the lungs, bone marrow, spleen, and blood at the advanced (prelethal) stage of Mycobacterium tuberculosis infection and represent a heterogeneous population of myeloid cells at different stages of their differentiation. The accumulation of Gr-1(dim)CD11b(+) cells is accompanied by the disappearance of conventional neutrophils (Gr-1(hi)Ly-6G(hi)-expressing cells). The Gr-1(dim)CD11b(+) cells suppress T cell proliferation and IFN-γ production in vitro via NO-dependent mechanisms, that is, they exhibit characteristics of myeloid-derived suppressor cells. These results document the generation of myeloid-derived suppressor cells during TB, suggesting their role in TB pathogenesis, and arguing that neutrophils do not contribute to TB pathology at the advanced disease stage.

  17. Purification and characterization of a lipid thiobis ester from human neutrophil cytosol that reversibly deactivates the O2- -generating NADPH oxidase.

    Science.gov (United States)

    Eklund, E A; Gabig, T G

    1990-05-25

    Intact neutrophils possess a cellular mechanism that efficiently deactivates the microbicidal O2-generating NADPH oxidase during the respiratory burst (Akard, L. P., English, D., and Gabig, T. G. (1988) Blood 72, 322-327). The present studies directed at identifying the molecular mechanism(s) involved in NADPH oxidase deactivation showed that a heat- and trypsin-insensitive species in the cytosolic fraction from normal unstimulated neutrophils was capable of deactivating the membrane-associated NADPH oxidase isolated from opsonized zymosan- or phorbol 12-myristate 13-acetate-stimulated neutrophils. This cytosolic species also deactivated the cell-free-activated oxidase. Deactivation by this cytosolic species occurred in the absence of NADPH-dependent catalytic turnover and was reversible, since NADPH oxidase activity could be subsequently reactivated in the cell-free system. The sedimentable particulate fraction from unstimulated neutrophils did not demonstrate deactivator activity. Deactivator activity was demonstrated in the neutral lipid fraction of neutrophil cytosol extracted with chloroform:methanol. Following complete purification of cytosolic deactivator activity by thin layer chromatography and reversed phase high performance liquid chromatography, the deactivator species was shown to be a lipid thiobis ester compound by mass spectroscopy. Cellular metabolism of this compound in human neutrophils may reveal a unique mechanism for enzymatic control of the NADPH oxidase system and thereby play an important role in regulation of the inflammatory response.

  18. Induction of autoimmune diseases by radiation exposure. Changes in molecular structures of neutrophil myeloperoxidase

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Kazuo; Ogawara, Akiko; Hashimoto, Yuki; Yamagoe, Satoshi; Borregaad, Niels; Mizuno, Satoshi [National Inst. of Health, Tokyo (Japan)

    1999-02-01

    It has been suggested that functional abnormalities of leucocyte induced by radiation exposure might be related to development of autoimmune diseases. The authors demonstrated that fragmentation of MPO, a lysozyme enzyme of neutrophil caused by high dose exposure resulted in a lowering of its activity and an enhancement of its degradation. It was assumed that release of MPO fragment, 30 kDa might be involved in development of autoimmune diseases. In this study, 4 peptides in neutrophil; defensin 3, hCAP-18, FMLP-R and CD35 were selected as the indicators for activation of neutrophils and the effects of {gamma}-ray exposure on their mRNA expressions were investigated. A suspension of neutrophils from healthy peripheral blood were exposed to {gamma}-ray at 0, 10 or 30 Gy and incubated after addition of cytochalasin B and FMLP for 10 min to release MPO. The activities of MPO in the cell and in the extracellular fluid were assayed with tetramethylbenzidine as the substrate. The MPO activity released from the neutrophils was significantly increased by {gamma}-ray exposure at 10 or 30 Gy. The increase was slightly higher for exposure at 10 Gy than that at 30 Gy. Next, the effects of exposure were investigated on mRNA expressions of 4 different peptides of neutrophil. From the exposed cells, whole RNA were extracted by AGPC method and used as a primer to produce the respective cDNAs and mRNA assay was conducted by PCR method. The exposure to {gamma}-ray at 10 Gy increased mRNA expressions of all four peptides. The increase was marked for CD35, moderate for defensin 3 and hCAP-18, and slight for FMLP-R. These results indicate that MPO fragments can be produced through radiation exposure as well as heat treatment. Therefore, it was suggested that anti-MPO antibody, MPO-ANCA might be produced by MPO fragments produced by radiation and/or lymphatic functional disorders. (M.N.)

  19. [Lymphocyte transformation test following stimulation with a protein factor from neutrophilic granulocytes (PMNL) in psoriasis patients].

    Science.gov (United States)

    Ruszczak, Z; Ciborska, L; Kaszuba, A

    1988-12-01

    The lymphocyte transformation test (LTT) was given to 20 healthy subjects and 43 patients with generalized psoriasis vulgaris: it was given right after stimulation with PHA (spontaneous) and after stimulation with allogenic and autogenic protein factor (NPF). NPF was isolated from secondary lysosome granules of peripheral blood neutrophils. The results were analyzed using computer statistic tests. No distinct differences were noticed between the spontaneous transformation test in psoriatic patients compared to the controls. After stimulation with PHA, the percentage of blast cells was significantly lower in patients with psoriasis. When allogenic and autogenic NPF was used for stimulation, the LTT values were significantly higher in the psoriasis group than in the control subjects. This fact points out the increase in sensitivity of lymphocytes to NPF in active psoriasis and the possibility of abnormal neutrophil-lymphocyte interactions in vivo. This phenomenon may be intensified when under the influence of bacterial or viral agents, or medicaments; the degranulation of secondary lysosome granules of neutrophils occurs, causing the release of NPF. These investigations support our opinion that psoriasis is a systemic disease and that NPF plays a considerable role in the psoriatic reaction.

  20. Chronic Inflammation and Neutrophil Activation as Possible Causes of Joint Diseases in Ballet Dancers

    Science.gov (United States)

    Borges, Leandro da Silva; Santos, Vinicius Coneglian; de Moura, Nivaldo Ribeiro; Dermargos, Alexandre; Cury-Boaventura, Maria Fernanda; Gorjão, Renata; Pithon-Curi, Tania Cristina; Hatanaka, Elaine

    2014-01-01

    Herein, we investigated the effects of a ballet class on the kinetic profiles of creatine kinase (CK) and lactate dehydrogenase (LDH) activities, cytokines, complement component 3 (C3), and the concentrations of immunoglobulin (Ig), IgA and IgM, in ballerinas. We also verified neutrophil death and ROS release. Blood samples were taken from 13 dancers before, immediately after, and 18 hours after a ballet class. The ballet class increased the plasma activities of CK-total (2.0-fold) immediately after class, while the activities of CK-cardiac muscle (1.0-fold) and LDH (3.0-fold) were observed to increase 18 hours after the class. Levels of the TNF-α, IL-1β, IgG, and IgA were not affected under the study conditions. The exercise was found to induce neutrophil apoptosis (6.0-fold) 18 hours after the ballet class. Additionally, immediately after the ballet class, the neutrophils from the ballerinas were found to be less responsive to PMA stimulus. Conclusion. Ballet class was found to result in inflammation in dancers. The inflammation caused by the ballet class remained for 18 hours after the exercise. These findings are important in preventing the development of chronic lesions that are commonly observed in dancers, such as those with arthritis and synovitis. PMID:24701035

  1. Chronic Inflammation and Neutrophil Activation as Possible Causes of Joint Diseases in Ballet Dancers

    Directory of Open Access Journals (Sweden)

    Leandro da Silva Borges

    2014-01-01

    Full Text Available Herein, we investigated the effects of a ballet class on the kinetic profiles of creatine kinase (CK and lactate dehydrogenase (LDH activities, cytokines, complement component 3 (C3, and the concentrations of immunoglobulin (Ig, IgA and IgM, in ballerinas. We also verified neutrophil death and ROS release. Blood samples were taken from 13 dancers before, immediately after, and 18 hours after a ballet class. The ballet class increased the plasma activities of CK-total (2.0-fold immediately after class, while the activities of CK-cardiac muscle (1.0-fold and LDH (3.0-fold were observed to increase 18 hours after the class. Levels of the TNF-α, IL-1β, IgG, and IgA were not affected under the study conditions. The exercise was found to induce neutrophil apoptosis (6.0-fold 18 hours after the ballet class. Additionally, immediately after the ballet class, the neutrophils from the ballerinas were found to be less responsive to PMA stimulus. Conclusion. Ballet class was found to result in inflammation in dancers. The inflammation caused by the ballet class remained for 18 hours after the exercise. These findings are important in preventing the development of chronic lesions that are commonly observed in dancers, such as those with arthritis and synovitis.

  2. Effects of systemic glucocorticosteroids on peripheral neutrophil functions in asthmatic subjects: an ex vivo study

    Directory of Open Access Journals (Sweden)

    P. L. Paggiaro

    1995-01-01

    Full Text Available In 21 asthmatic subjects, several functions of isolated peripheral neutrophils (chemokinesis and chemotaxis toward 10% E. coli; superoxide anion generation after PMA; leukotriene B4 (LTB4 release from whole blood and isolated neutrophtls, before and after different stimuli were evaluated during an acute exacerbation of asthma, and after 14 – 54 days of treatment with systemic glucocorticosteroids (GCS. During acute exacerbation, superoxide anion generation was higher in asthmatics than in eleven normal subjects (39.2 ± 14.1 vs. 25.2 ± 7.3 nmol, p 20% after GCS treatment (from 131 ± 18 to 117 ± 21 μm, p = 0.005. Chemokinesis sicantly decreased in all subjects, and the changes significantly correlated with an arbitrary score of the total administered dose of GCS (r = 0.57, p < 0.05. These data suggest that neutrophil activation plays a minor role in asthma, and that treatment with GCS is not able to modify most functions of peripheral neutrophils in asthmatic subjects; chemotaxis seems to be related only to the severity of the asthma and it could reflect the improvement of the disease.

  3. A Potential Role for Acrolein in Neutrophil-Mediated Chronic Inflammation.

    Science.gov (United States)

    Noerager, Brett D; Xu, Xin; Davis, Virginia A; Jones, Caleb W; Okafor, Svetlana; Whitehead, Alicia; Blalock, J Edwin; Jackson, Patricia L

    2015-12-01

    Neutrophils (PMNs) are key mediators of inflammatory processes throughout the body. In this study, we investigated the role of acrolein, a highly reactive aldehyde that is ubiquitously present in the environment and produced endogenously at sites of inflammation, in mediating PMN-mediated degradation of collagen facilitating proline-glycine-proline (PGP) production. We treated peripheral blood neutrophils with acrolein and analyzed cell supernatants and lysates for matrix metalloproteinase-9 (MMP-9) and prolyl endopeptidase (PE), assessed their ability to break down collagen and release PGP, and assayed for the presence of leukotriene A4 hydrolase (LTA4H) and its ability to degrade PGP. Acrolein treatment induced elevated production and functionality of collagen-degrading enzymes and generation of PGP fragments. Meanwhile, LTA4H levels and triaminopeptidase activity declined with increasing concentrations of acrolein thereby sparing PGP from enzymatic destruction. These findings suggest that acrolein exacerbates the acute inflammatory response mediated by neutrophils and sets the stage for chronic pulmonary and systemic inflammation.

  4. Abundant neutrophil extracellular traps in thrombus of patient with microscopic polyangiitis

    Directory of Open Access Journals (Sweden)

    Daigo eNakazawa

    2012-11-01

    Full Text Available This is a case study of a patient diagnosed with microscopic polyangiitis (MPA and complicated with deep vein thrombosis (DVT, who died of respiratory failure despite treatment. Autopsy revealed severe crescentic glomerulonephritis and massive alveolar hemorrhage. The thrombus contained abundant neutrophils. Although it is reported that patients with ANCA-associated vasculitis (AAV have an increased risk of DVT, it remains elusive why they are prone to thrombosis. A recent study has demonstrated the presence of neutrophil extracellular traps (NETs, a newly recognized mode of neutrophil cell-death, in glomerular crescents of MPA patients. Interestingly, NETs were identified in the thrombus as well as in the glomerular crescents in the present case. When compared to other thrombi unrelated to MPA, the amount of NETs was significantly greater in the MPA patient. On the other hand, NETs are critically involved in thrombogenesis because histones within NETs can bind platelets and blood coagulants. Although this is important in regard to containment of microbes within NETs, excessive NETs could cause thrombosis. The collective findings suggest the possibility that thrombosis could be critically associated with MPA via NETs, and that NETs could be a therapeutic target in MPA patients.

  5. In vivo characterization of neutrophil extracellular traps in various organs of a murine sepsis model.

    Directory of Open Access Journals (Sweden)

    Koji Tanaka

    Full Text Available Neutrophil extracellular traps (NETs represent extracellular microbial trapping and killing. Recently, it has been implicated in thrombogenesis, autoimmune disease, and cancer progression. The aim of this study was to characterize NETs in various organs of a murine sepsis model in vivo and to investigate their associations with platelets, leukocytes, or vascular endothelium. NETs were classified as two distinct forms; cell-free NETs that were released away from neutrophils and anchored NETs that were anchored to neutrophils. Circulating cell-free NETs were characterized as fragmented or cotton-like structures, while anchored NETs were characterized as linear, reticular, membranous, or spot-like structures. In septic mice, both anchored and cell-free NETs were significantly increased in postcapillary venules of the cecum and hepatic sinusoids with increased leukocyte-endothelial interactions. NETs were also observed in both alveolar space and pulmonary capillaries of the lung. The interactions of NETs with platelet aggregates, leukocyte-platelet aggregates or vascular endothelium of arterioles and venules were observed in the microcirculation of septic mice. Microvessel occlusions which may be caused by platelet aggregates or leukocyte-platelet aggregates and heterogeneously decreased blood flow were also observed in septic mice. NETs appeared to be associated with the formation of platelet aggregates or leukocyte-platelet aggregates. These observational findings may suggest the adverse effect of intravascular NETs on the host during a sepsis.

  6. Desempenho da tilápia-do-Nilo arraçoada com dietas contendo farinha de sangue bovino atomizado ou convencional = Performance of nile tilapia fed with spray-dried or vat-dries bovine blood meal

    Directory of Open Access Journals (Sweden)

    Willian Vicente Narváez-Solarte

    2011-07-01

    Full Text Available Foi avaliado o desempenho e os índices de rendimento da tilápia-do-Nilo (Oreochromis niloticus alimentada com níveis crescentes de farinha de sangue atomizado (FSA ou de farinha de sangue convencional (FSC em dietas formuladas com base em aminoácidos digestíveis. Foram utilizados 252 alevinos, distribuídos num delineamento inteiramente casualizado, em esquema fatorial (2 x 4 + 1, duas classes de farinha de sangue com quatro níveis de inclusão de cada farinha na dieta, e uma dieta-controle, com quatro repetições. Os tratamentos consistiram em uma dieta-controle à base de farelo de soja, contendo 34% de proteína digestível (PD e 3.200 kcal de energia digestível kg-1 (ED, mais quatro rações formuladas com FSA e quatro rações com FSC, com inclusões de 5, 10, 15 e 20% de cada farinha na ração, mantendo-se os níveis de PD, ED, fósforo, cálcio, lisina, metionina, treonina e triptofano idênticos aos da dieta-controle. Concluiu-se que é possível utilizar até 15% da FSC em rações para tilápia-do-Nilo na fase de 5 a 150 g de peso vivo.The study evaluated the performance and carcass composition index of Nile tilapia (Oreochromis niloticus fed with diets containing increasing levels of spray-dried blood meal (SDBM and vat-dried blood meal (VDBM and formulated based on digestible amino acids. Two hundred and fifty-two fingerlings were distributed in a completelyrandomized design, in a (2 x 4 + 1 factorial model, two types of blood meal with four levels of each blood meal in the diet, and a control diet (without blood meal, with four replications. The treatments consisted of soybean meal-based control diet, with 34%digestible protein (DP and 3,200 kcal of digestible energy kg-1 (DE, plus four diets formulated with SDBM and four diets with VDBM, containing 5, 10, 15 and 20% of each meal in feed, maintaining identical DP, DE, phosphorus, calcium, lysine, methionine, threonine and tryptophan levels as those of the control diet. The

  7. A novel immune regulatory function of neutrophils in rhinovirus infections

    NARCIS (Netherlands)

    Tang, Francesca; Hansbro, Phil; Burgess, Janette; Baines, Katherine; Oliver, Brian

    2015-01-01

    Rationale: Rhinovirus (RV) is a major precipitant of asthma exacerbations. During lung infections there is elevated neutrophilic inflammation which is associated with more severe asthma symptoms. Previously, we found that neutrophils respond to viral mimetics but not live RV. Here we investigated if

  8. The Role of Neutrophil Collagenase in Endotoxic Acute Lung Injury

    Institute of Scientific and Technical Information of China (English)

    徐涛; 曾邦雄; 李兴旺

    2004-01-01

    The aim of this study was to determine the role of neutrophil collagenase in the pathogenesis of acute lung injury induced by endotoxin. 28 Sprague-Dawley were randomized into control group and LPS-enduced groups. Samples of left lung were obtained in 2 h (group L1 ), 6 h (group L2), 12 h (group L3 ) after intravenous LPS. Immunohistochemsitry was employed for detection of expression of neutrophil collagenase. Pathological scores, lung wet/dry weight ratio and the number of neutrophils were measured. The results showed that the concentration of neutrophil collagenase in LPS-enduced groups (group L1, L2, L3 ) were significantly higher than that of control group (P<0.01). Pathological scores, lung wet/dry weight ratio and the number of neutrophils in LPS-enduced groups (group L1, L2, L3 ) were also significantly higher than that of control group (P<0.01).Moreover, among group L1, L2 and L3, there were significant correlations in concentration of neutrophil collagenase and pathological scores, lung wet/dry weight ratio, the number of neutrophils (P<0.05). The present study showed that neutrophil collagenase play an important role in the pathogenesis and progress of endotoxic acute lung injury.

  9. Early diagnostic method for sepsis based on neutrophil MR imaging

    Directory of Open Access Journals (Sweden)

    Shanhua Han

    2015-06-01

    Conclusion: Mouse and human neutrophils could be more effectively labelled by Mannan-coated SPION in vitro than Feridex. Sepsis analog neutrophils labelled by Mannan-coated SPIONs could be efficiently detected on MR images, which may serve as an early diagnostic method for sepsis.

  10. Ascorbate recycling in human neutrophils: Induction by bacteria

    OpenAIRE

    Wang, Yaohui; Russo, Thomas A.; Kwon, Oran; Chanock, Stephen; Rumsey, Steven C.; Levine, Mark

    1997-01-01

    Ascorbate (vitamin C) recycling occurs when extracellular ascorbate is oxidized, transported as dehydroascorbic acid, and reduced intracellularly to ascorbate. We investigated microorganism induction of ascorbate recycling in human neutrophils and in microorganisms themselves. Ascorbate recycling was determined by measuring intracellular ascorbate accumulation. Ascorbate recycling in neutrophils was induced by both Gram-positive and Gram-negative pathogenic bacteria, and the fungal pathogen C...

  11. Intracellular localization of VAMP-1 protein in human neutrophils.

    Science.gov (United States)

    Nabokina, S M

    2001-02-01

    We studied the intracellular localization of vesicle-associated membrane protein VAMP-1 in human neutrophils. VAMP-1 was associated with membranes of gelatinase and specific secretory granules rapidly mobilized during exocytosis. VAMP-1 probably acts as a component of the SNARE complex during exocytosis of gelatinase and specific granules in human neutrophils.

  12. Killing of Mycobacterium tuberculosis by neutrophils: a nonoxidative process.

    Science.gov (United States)

    Jones, G S; Amirault, H J; Andersen, B R

    1990-09-01

    To determine the role of oxygen radicals in the killing of Mycobacterium tuberculosis by neutrophils, the effects of free-radical inhibitors and enzymes, catalase, superoxide dismutase, taurine, deferoxamine, and histidine were evaluated. Changes in the viability of M. tuberculosis were determined by agar plate colony counts and a radiometric assay. No impairment in killing was seen with any of the inhibitors or enzymes. Patients with chronic granulomatous disease (CGD) have a defect in the NADPH oxidase pathway, causing their neutrophils to be unable to generate oxygen radicals. If these radicals are involved in killing, then CGD neutrophils should be less effective killers of M. tuberculosis than normal neutrophils. There was no evidence by either measure of M. tuberculosis viability that CGD neutrophils were less bactericidal than normal neutrophils. Killing by normal neutrophils was also effective in the absence of serum. These results lead to the conclusion that the mechanism by which M. tuberculosis is killed by neutrophils is independent of the oxygen metabolic burst.

  13. Quercetin inhibits degranulation and superoxide generation in PMA stimulated neutrophils

    OpenAIRE

    2012-01-01

    Activated neutrophils represent the main source of myeloperoxidase (MPO), superoxide (SO) and subsequently derived oxygen metabolites. They have important microbicidal activities, however in inflammatory conditions they may secondarily attack surrounding tissues. Overproduction of reactive oxygen species, prolonged or excessive liberation of MPO and other effective yet also toxic substances from neutrophils may participate in disturbed apoptosis, intensify the inflammatory processes and resul...

  14. Intergrin-dependent neutrophil migration in the injured mouse cornea

    Science.gov (United States)

    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...

  15. Synchronisation of glycolytic oscillations in a suspension of human neutrophils

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Poulsen, Allan K.; Olsen, Lars Folke

    Neutrophils are known to be able to synchronize their production of superoxide. We show that glycolysis is also synchronized in human neutrophils being in suspension and suggest that oscillations in glycolysis are driving the pulsatile production of superoxide. The synchronising agent remains so...

  16. Activation of the Small GTPase Rap1 in Human Neutrophils

    NARCIS (Netherlands)

    M'Rabet, Laura; Coffer, P.J.; Zwartkruis, G.J.T.; Franke, Barbara; Segal, Anthony W.; Koenderman, L.; Bos, J.L.

    2002-01-01

    The small GTPase Rap1 is highly expressed in human neutrophils, but its function is largely unknown. Using the Rap1- binding domain of RalGDS (RalGDS-RBD) as an activationspecific probe for Rap1, we have investigated the regulation of Rap1 activity in primary human neutrophils. We found that a varie

  17. Resistance of Neisseria gonorrhoeae to neutrophils

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    M. Brittany eJohnson

    2011-04-01

    Full Text Available Infection with the human-specific bacterial pathogen Neisseria gonorrhoeae triggers a potent, local inflammatory response driven by polymorphonuclear leukocytes (neutrophils or PMNs. PMNs are terminally differentiated phagocytic cells that are a vital component of the host innate immune response and are the first responders to bacterial and fungal infections. PMNs possess a diverse arsenal of components to combat microorganisms, including the production of reactive oxygen species and release of degradative enzymes and antimicrobial peptides. Despite numerous PMNs at the site of gonococcal infection, N. gonorrhoeae can be cultured from the PMN-rich exudates of individuals with acute gonorrhea, indicating that some bacteria resist killing by neutrophils. The contribution of PMNs to gonorrheal pathogenesis has been modeled in vivo by human male urethral challenge and murine female genital inoculation and in vitro using isolated primary PMNs or PMN-derived cell lines. These systems reveal that some gonococci survive and replicate within PMNs and suggest that gonococci defend themselves against PMNs in two ways: they express virulence factors that defend against PMNs’ oxidative and non-oxidative antimicrobial components, and they modulate the ability of PMNs to phagocytose gonococci and to release antimicrobial components. In this review, we will highlight the varied and complementary approaches used by N. gonorrhoeae to resist clearance by human PMNs, with an emphasis on gonococcal gene products that modulate bacterial-PMN interactions. Understanding how some gonococci survive exposure to PMNs will help guide future initiatives for combating gonorrheal disease.

  18. Toxicology of a bovine paraplegic syndrome.

    Science.gov (United States)

    Sevcik, C; Brito, J C; D'Suze, G; Domínguez-Bello, M G; Lovera, M; Mijares, A J; Bónoli, S

    1993-12-01

    A clinical entity named 'bovine paraplegic syndrome' ('síndrome parapléjico de los bovinos') has spread alarmingly in the cattle-growing areas of the central and eastern plains of Venezuela. It is estimated that four million cattle are bred in the area where the disease occurs. The mortality ranges from 5 to 25% of the animals at risk, mostly pregnant or lactating cows. The principal characteristic of the bovine paraplegic syndrome is ventral or sternal decubitus, in animals that make vain efforts to stand when stimulated. The diagnosis is established when all other possible causes (e.g. paralytic rabies, botulism and blood parasites such as Anaplasma marginal, Babesia bovis, B. bigemina, and Trypanosoma vivax) have been ruled out clinically and by laboratory tests. Death always occurs, usually after a few days, and there is no known treatment. In this work, we describe results that show the presence of a toxin in the cattle suffering from, or liable to suffer from the syndrome. The toxin is produced by ruminal bacteria. In squid giant axons under voltage clamp conditions, the toxin blocks the sodium current. We detected the toxin analytically by absorbance measurements at 340 nm after reacting with picrylsulfonic acid. We obtained a good separation of the toxin with isocratic high pressure liquid chromatography, using 40% methanol in water on phenylborasil columns.

  19. The effect of lidocaine on in vitro neutrophil and endothelial adhesion molecule expression induced by plasma obtained during tourniquet-induced ischaemia and reperfusion.

    LENUS (Irish Health Repository)

    Lan, W

    2012-02-03

    BACKGROUND: Changes in neutrophil and endothelial adhesion molecule expression occur during perioperative ischaemia and reperfusion (I\\/R) injury. We investigated the effects of lidocaine on neutrophil-independent changes in neutrophil and endothelial adhesion molecule expression associated with tourniquet-induced I\\/R. METHODS: Plasma was obtained from venous blood samples (tourniquet arm) taken before (baseline), during, 15 min, 2 and 24 h following tourniquet release in seven patients undergoing elective upper limb surgery with tourniquet application. Isolated neutrophils from healthy volunteers (n = 7) were pretreated in the presence or absence of lidocaine (0.005, 0.05 and 0.5 mg mL(-1) for 1 h, and then incubated with I\\/R plasma for 2 h. Human umbilical vein endothelial cells (HUVECs) were pretreated in the presence or absence of lidocaine (0.005, 0.05 and 0.5 mg mL(-1)) for 1 h, and then incubated with the plasma for 4 h. Adhesion molecule expression was estimated using flow cytometry. Data were analysed using ANOVA and post hoc Student-Newman-Keuls tests. RESULTS: I\\/R plasma (withdrawn 15 min following tourniquet release) increased isolated neutrophil CD11b (P = 0.03), CD18 (P = 0.01) and endothelial intercellular adhesion molecule-1 (ICAM-1) (P = 0.008) expression compared to baseline. CD11b, CD18 and ICAM-1 expression on lidocaine (0.005 mg mL(-1)) treated neutrophils was similar to control. CD11b (P < 0.001), CD18 (P = 0.03) and ICAM-1 (P = 0.002) expression on lidocaine (0.05 mg mL(-1)) treated neutrophils and HUVECs was less than that on controls. CONCLUSION: Increased in vitro neutrophil and endothelial cell adhesion molecule expression on exposure to plasma obtained during the early reperfusion phase is diminished by lidocaine at greater than clinically relevant plasma concentrations.

  20. Neutrophil adhesion to E-selectin under shear promotes the redistribution and co-clustering of ADAM17 and its proteolytic substrate L-selectin.

    Science.gov (United States)

    Schaff, Ulrich; Mattila, Polly E; Simon, Scott I; Walcheck, Bruce

    2008-01-01

    E-selectin is expressed by the vascular endothelium and binds flowing neutrophils in the blood to facilitate their recruitment into the underlying tissue at sites of inflammation. L-selectin on neutrophils is engaged by E-selectin and undergoes rapid clustering and then coalescence in the trailing edge of polarizing cells. These processes are believed to increase the valency and capacity of L-selectin to signal CD18 integrin activity. Neutrophils, upon exiting the microvasculature, down-regulate their surface L-selectin through ectodomain shedding by a disintegrin and metalloprotease 17 (ADAM17). We reasoned that neutrophil tethering and rolling on E-selectin might initiate a coordinate change in the membrane distribution of ADAM17 as well. We found that ADAM17 indeed underwent a dramatic cell surface redistribution to the trailing edge of neutrophils rolling on purified E-selectin when activated by a chemoattractant under shear flow; however, its lateral migration occurred at a slower rate than L-selectin. ADAM17 and L-selectin also redistributed in the same manner in neutrophils attached to IL-1beta-stimulated HUVEC under shear flow. In contrast, the coalescence of L-selectin on the surface of neutrophils by antibody cross-linking did not promote the redistribution of ADAM17, suggesting that these molecules do not constitutively associate in the plasma membrane. Together, our findings reveal that neutrophil activation upon E-selectin adhesion initiates active transport of ADAM17 and L-selectin to the cell uropod, thus providing additional insight into the molecular mechanisms that regulate L-selectin during leukocyte extravasation.

  1. Neutropenic Mice Provide Insight into the Role of Skin-Infiltrating Neutrophils in the Host Protective Immunity against Filarial Infective Larvae

    Science.gov (United States)

    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

  2. Neutrophil extracellular trap (NET-mediated killing of Pseudomonas aeruginosa: evidence of acquired resistance within the CF airway, independent of CFTR.

    Directory of Open Access Journals (Sweden)

    Robert L Young

    Full Text Available The inability of neutrophils to eradicate Pseudomonas aeruginosa within the cystic fibrosis (CF airway eventually results in chronic infection by the bacteria in nearly 80 percent of patients. Phagocytic killing of P. aeruginosa by CF neutrophils is impaired due to decreased cystic fibrosis transmembrane conductance regulator (CFTR function and virulence factors acquired by the bacteria. Recently, neutrophil extracellular traps (NETs, extracellular structures composed of neutrophil chromatin complexed with granule contents, were identified as an alternative mechanism of pathogen killing. The hypothesis that NET-mediated killing of P. aeruginosa is impaired in the context of the CF airway was tested. P. aeruginosa induced NET formation by neutrophils from healthy donors in a bacterial density dependent fashion. When maintained in suspension through continuous rotation, P. aeruginosa became physically associated with NETs. Under these conditions, NETs were the predominant mechanism of killing, across a wide range of bacterial densities. Peripheral blood neutrophils isolated from CF patients demonstrated no impairment in NET formation or function against P. aeruginosa. However, isogenic clinical isolates of P. aeruginosa obtained from CF patients early and later in the course of infection demonstrated an acquired capacity to withstand NET-mediated killing in 8 of 9 isolates tested. This resistance correlated with development of the mucoid phenotype, but was not a direct result of the excess alginate production that is characteristic of mucoidy. Together, these results demonstrate that neutrophils can kill P. aeruginosa via NETs, and in vitro this response is most effective under non-stationary conditions with a low ratio of bacteria to neutrophils. NET-mediated killing is independent of CFTR function or bacterial opsonization. Failure of this response in the context of the CF airway may occur, in part, due to an acquired resistance against NET

  3. Neutrophil extracellular trap (NET)-mediated killing of Pseudomonas aeruginosa: evidence of acquired resistance within the CF airway, independent of CFTR.

    Science.gov (United States)

    Young, Robert L; Malcolm, Kenneth C; Kret, Jennifer E; Caceres, Silvia M; Poch, Katie R; Nichols, David P; Taylor-Cousar, Jennifer L; Saavedra, Milene T; Randell, Scott H; Vasil, Michael L; Burns, Jane L; Moskowitz, Samuel M; Nick, Jerry A

    2011-01-01

    The inability of neutrophils to eradicate Pseudomonas aeruginosa within the cystic fibrosis (CF) airway eventually results in chronic infection by the bacteria in nearly 80 percent of patients. Phagocytic killing of P. aeruginosa by CF neutrophils is impaired due to decreased cystic fibrosis transmembrane conductance regulator (CFTR) function and virulence factors acquired by the bacteria. Recently, neutrophil extracellular traps (NETs), extracellular structures composed of neutrophil chromatin complexed with granule contents, were identified as an alternative mechanism of pathogen killing. The hypothesis that NET-mediated killing of P. aeruginosa is impaired in the context of the CF airway was tested. P. aerug