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Sample records for macrophages rapid killing

  1. Kinetics of killing Listeria monocytogenes by macrophages: rapid killing accompanying phagocytosis

    SciTech Connect

    Davies, W.A.

    1983-08-01

    The kinetics of bactericidal activity of activated macrophages can be precisely described by a mathematical model in which phagocytosis, killing, digestion, and release of degraded bacterial material are considered to occur continuously. To gain a better understanding of these events, I have determined the period of time between first contact of bacteria with macrophages and the onset of killing. Activated rat peritoneal macrophages were incubated for various times up to 15 min with Listeria monocytogenes previously labeled with /sup 3/H-thymidine and the unassociated bacteria removed by two centrifugations through a density interface. Both cell-associated radioactivity and cell-associated viable bacteria, determined as colony forming units after sonication of the cell pellet, increased with time of incubation. However, the specific viability of these bacteria, expressed as the ratio of number of viable bacteria per unit radioactivity declined with time, as an approximate inverse exponential, after a lag period of 2.9 +/- 0.8 min. Evidence is given that other possible causes for this decline in specific viability, other than death of the bacteria, such as preferential ingestion of dead Listeria, clumping of bacteria, variations in autolytic activity, or release of Listericidins are unlikely. I conclude therefore that activated macrophages kill Listeria approximately 3 min after the cell and the bacterium first make contact.

  2. On the killing of mycobacteria by macrophages.

    PubMed

    Jordao, Luisa; Bleck, Christopher K E; Mayorga, Luis; Griffiths, Gareth; Anes, Elsa

    2008-02-01

    Both pathogenic and non-pathogenic mycobacteria are internalized into macrophage phagosomes. Whereas the non-pathogenic types are invariably killed by all macrophages, the pathogens generally survive and grow. Here, we addressed the survival, production of nitrogen intermediates (RNI) and intracellular trafficking of the non-pathogenic Mycobacterium smegmatis, the pathogen-like, BCG and the pathogenic M. bovis in different mouse, human and bovine macrophages. The bacteriocidal effects of RNI were restricted for all bacterial species to the early stages of infection. EM analysis showed clearly that all the mycobacteria remained within phagosomes even at late times of infection. The fraction of BCG and M. bovis found in mature phagolysosomes rarely exceeded 10% of total, irrespective of whether bacteria were growing, latent or being killed, with little correlation between the extent of phagosome maturation and the degree of killing. Theoretical modelling of our data identified two different potential sets of explanations that are consistent with our results. The model we favour is one in which a small but significant fraction of BCG is killed in an early phagosome, then maturation of a small fraction of phagosomes with both live and killed bacteria, followed by extremely rapid killing and digestion of the bacteria in phago-lysosomes.

  3. Inability to sustain intraphagolysosomal killing of Staphylococcus aureus predisposes to bacterial persistence in macrophages.

    PubMed

    Jubrail, Jamil; Morris, Paul; Bewley, Martin A; Stoneham, Simon; Johnston, Simon A; Foster, Simon J; Peden, Andrew A; Read, Robert C; Marriott, Helen M; Dockrell, David H

    2016-01-01

    Macrophages are critical effectors of the early innate response to bacteria in tissues. Phagocytosis and killing of bacteria are interrelated functions essential for bacterial clearance but the rate-limiting step when macrophages are challenged with large numbers of the major medical pathogen Staphylococcus aureus is unknown. We show that macrophages have a finite capacity for intracellular killing and fail to match sustained phagocytosis with sustained microbial killing when exposed to large inocula of S. aureus (Newman, SH1000 and USA300 strains). S. aureus ingestion by macrophages is associated with a rapid decline in bacterial viability immediately after phagocytosis. However, not all bacteria are killed in the phagolysosome, and we demonstrate reduced acidification of the phagolysosome, associated with failure of phagolysosomal maturation and reduced activation of cathepsin D. This results in accumulation of viable intracellular bacteria in macrophages. We show macrophages fail to engage apoptosis-associated bacterial killing. Ultittop mately macrophages with viable bacteria undergo cell lysis, and viable bacteria are released and can be internalized by other macrophages. We show that cycles of lysis and reuptake maintain a pool of viable intracellular bacteria over time when killing is overwhelmed and demonstrate intracellular persistence in alveolar macrophages in the lungs in a murine model.

  4. Inability to sustain intraphagolysosomal killing of Staphylococcus aureus predisposes to bacterial persistence in macrophages

    PubMed Central

    Jubrail, Jamil; Morris, Paul; Bewley, Martin A.; Stoneham, Simon; Johnston, Simon A.; Foster, Simon J.; Peden, Andrew A.; Read, Robert C.; Marriott, Helen M.

    2015-01-01

    Summary Macrophages are critical effectors of the early innate response to bacteria in tissues. Phagocytosis and killing of bacteria are interrelated functions essential for bacterial clearance but the rate‐limiting step when macrophages are challenged with large numbers of the major medical pathogen Staphylococcus aureus is unknown. We show that macrophages have a finite capacity for intracellular killing and fail to match sustained phagocytosis with sustained microbial killing when exposed to large inocula of S. aureus (Newman, SH1000 and USA300 strains). S. aureus ingestion by macrophages is associated with a rapid decline in bacterial viability immediately after phagocytosis. However, not all bacteria are killed in the phagolysosome, and we demonstrate reduced acidification of the phagolysosome, associated with failure of phagolysosomal maturation and reduced activation of cathepsin D. This results in accumulation of viable intracellular bacteria in macrophages. We show macrophages fail to engage apoptosis‐associated bacterial killing. Ultittop mately macrophages with viable bacteria undergo cell lysis, and viable bacteria are released and can be internalized by other macrophages. We show that cycles of lysis and reuptake maintain a pool of viable intracellular bacteria over time when killing is overwhelmed and demonstrate intracellular persistence in alveolar macrophages in the lungs in a murine model. PMID:26248337

  5. Ultrastructural studies of the killing of schistosomula of Schistosoma mansoni by activated macrophages in vitro.

    PubMed

    McLaren, D J; James, S L

    1985-05-01

    Immunologically activated murine macrophages have been shown elsewhere to kill skin stage schistosomula of Schistosoma mansoni in vitro, in a manner analogous to the extracellular killing of tumour cell targets. In this study, the kinetics of the interaction between activated macrophages and larval targets and the resultant ultrastructural changes in parasite morphology that culminated in death have been analysed in detail. Unlike granulocyte-mediated schistosomular killing, macrophage-mediated cytotoxicity did not appear to be directed against the surface tissues of the parasite. Macrophages adhered only transiently following initiation of the cultures, yet changes in the subtegumental mitochondria and muscle cells of the larva were detected within the first hour of incubation. Progressive internal disorganisation followed rapidly, but the tegument and tegumental outer membrane remained intact, to form a 'shell' that maintained the general shape of the parasite. Such changes were recognised irrespective of whether the effector cell population comprised peritoneal macrophages activated by lymphokine treatment in vitro, or by infection with Mycobacterium bovis (strain BCG), or S. mansoni in vivo. That macrophages rather than contaminating granulocytes or lymphocytes, had mediated the observed damage was demonstrated by the use of a lymphokine treated macrophage cell line, IC-21. The observation that macrophage cytotoxicity is directed against internal organelles rather than the tegumental outer membrane of this multicellular target, may help to elucidate the general mechanism of extracellular killing by these cells. PMID:3892433

  6. The killing of macrophages by Corynebacterium ulcerans.

    PubMed

    Hacker, Elena; Ott, Lisa; Schulze-Luehrmann, Jan; Lührmann, Anja; Wiesmann, Veit; Wittenberg, Thomas; Burkovski, Andreas

    2016-01-01

    Corynebacterium ulcerans is an emerging pathogen transmitted by a zoonotic pathway with a very broad host spectrum to humans. Despite rising numbers of infections and potentially fatal outcomes, data on the molecular basis of pathogenicity are scarce. In this study, the interaction of 2 C. ulcerans isolates - one from an asymptomatic dog, one from a fatal case of human infection - with human macrophages was investigated. C. ulcerans strains were able to survive in macrophages for at least 20 hours. Uptake led to delay of phagolysosome maturation and detrimental effects on the macrophages as deduced from cytotoxicity measurements and FACS analyses. The data presented here indicate a high infectious potential of this emerging pathogen.

  7. Measurement of bacterial ingestion and killing by macrophages.

    PubMed

    Campbell, P A; Canono, B P; Drevets, D A

    2001-05-01

    This unit presents fairly simple assays for measuring the binding of bacteria to macrophages, internalization of bacteria (also called ingestion or phagocytosis), and bacterial killing by macrophages. The first basic protocol describes how to measure the ability of macrophages to ingest bacteria. Because it is critical to remove residual extracellular organisms, the protocol presents two alternative steps to accomplish this: a washing procedure and a more stringent method in which cells are sedimented through sucrose. In addition, it is important to distinguish those bacteria truly ingested by a macrophage from those that are bound to, but not internalized by, the cell. A simple but effective way to do this is described in an alternate protocol. The unit also presents two ways to measure the ability of a macrophage to kill bacteria it has internalized. The first is a straightforward assay in which bacterial colonies are enumerated before and after a killing period; a subsequent colony count will indicate whether the bacteria grew within or were killed by the macrophage. The second protocol describes a way to measure bacterial viability based on bacterial metabolism, in which the ability of bacterial dehydrogenases to mediate the reduction of a tetrazolium salt to purple formazan is monitored by measuring absorbance spectrophotometrically.

  8. Interactions between neutrophils and macrophages promote macrophage killing of rat muscle cells in vitro

    NASA Technical Reports Server (NTRS)

    Nguyen, Hal X.; Tidball, James G.

    2003-01-01

    Current evidence indicates that the physiological functions of inflammatory cells are highly sensitive to their microenvironment, which is partially determined by the inflammatory cells and their potential targets. In the present investigation, interactions between neutrophils, macrophages and muscle cells that may influence muscle cell death are examined. Findings show that in the absence of macrophages, neutrophils kill muscle cells in vitro by superoxide-dependent mechanisms, and that low concentrations of nitric oxide (NO) protect against neutrophil-mediated killing. In the absence of neutrophils, macrophages kill muscle cells through a NO-dependent mechanism, and the presence of target muscle cells causes a three-fold increase in NO production by macrophages, with no change in the concentration of inducible nitric oxide synthase. Muscle cells that are co-cultured with both neutrophils and macrophages in proportions that are observed in injured muscle show cytotoxicity through a NO-dependent, superoxide-independent mechanism. Furthermore, the concentration of myeloid cells that is necessary for muscle killing is greatly reduced in assays that use mixed myeloid cell populations, rather than uniform populations of neutrophils or macrophages. These findings collectively show that the magnitude and mechanism of muscle cell killing by myeloid cells are modified by interactions between muscle cells and neutrophils, between muscle cells and macrophages and between macrophages and neutrophils.

  9. The Tuberculosis Necrotizing Toxin kills macrophages by hydrolyzing NAD

    PubMed Central

    Sun, Jim; Siroy, Axel; Lokareddy, Ravi K.; Speer, Alexander; Doornbos, Kathryn S.; Cingolani, Gino; Niederweis, Michael

    2015-01-01

    Mycobacterium tuberculosis (Mtb) induces necrosis of infected cells to evade immune responses. Recently, we found that Mtb utilizes the protein CpnT to kill human macrophages by secreting its C-terminal domain, named tuberculosis necrotizing toxin (TNT) that induces necrosis by an unknown mechanism. Here we show that TNT gains access to the cytosol of Mtb-infected macrophages, where it hydrolyzes the essential co-enzyme nicotinamide adenine dinucleotide (NAD+). Expression or injection of a non-catalytic TNT mutant showed no cytotoxicity in macrophages or zebrafish zygotes, respectively, demonstrating that the NAD+-glycohydrolase activity is required for TNT-induced cell death. To prevent self-poisoning, Mtb produces an immunity factor for TNT (IFT) that binds TNT and inhibits its activity. The crystal structure of the TNT-IFT complex revealed a novel NAD+-glycohydrolase fold of TNT, which constitutes the founding member of a toxin family wide-spread in pathogenic microorganisms. PMID:26237511

  10. Interaction of human leukocytes and Entamoeba histolytica. Killing of virulent amebae by the activated macrophage.

    PubMed Central

    Salata, R A; Pearson, R D; Ravdin, J I

    1985-01-01

    Capable effector mechanisms in the human immune response against the cytolytic, protozoan parasite Entamoeba histolytica have not been described. To identify a competent human effector cell, we studied the in vitro interactions of normal human polymorphonuclear neutrophils, peripheral blood mononuclear cells (PBMC), monocytes (MC), and MC-derived macrophages with virulent axenic amebae (strain HMI-IMSS). Amebae killed neutrophils, PBMC, MC, and MC-derived macrophages (P less than 0.001), without loss of parasite viability. The addition of heat-inactivated immune serum did not enable leukocytes to kill amebae, nor did it protect these host cells from amebae. MC-derived macrophages, activated with lymphokine elicited by the mitogens conconavalin A, phytohemagglutinin, or an amebic soluble protein preparation (strain HK9), killed 55% of amebae by 3 h in a trypan blue exclusion assay (P less than 0.001); during this time, 40% of the activated macrophages died. Lysis of amebae was confirmed using 111Indium oxine radiolabeled parasites and was antibody independent. Macrophage death appeared to be due to the deleterious effect of lysed amebae rather than the contact-dependent effector mechanisms of E. histolytica. Adherence between activated macrophages and amebae was greater than that between other leukocytes and amebae (P less than 0.001). Microscopic observations, kinetic analysis of the killing of amebae by activated macrophages, and suspension of amebae with adherent activated macrophages in a 10% dextran solution indicated that contact by activated macrophages was necessary to initiate the killing of amebae. Catalase but not superoxide dismutase inhibited the amebicidal capacity of activated macrophages (P less than 0.001). However, activated macrophages from an individual with chronic granulomatous disease were able to kill amebae, but not as effectively as normal cells (P less than 0.01). In summary, activated MC-derived macrophages killed virulent E. histolytica

  11. Interaction of human leukocytes and Entamoeba histolytica. Killing of virulent amebae by the activated macrophage.

    PubMed

    Salata, R A; Pearson, R D; Ravdin, J I

    1985-08-01

    Capable effector mechanisms in the human immune response against the cytolytic, protozoan parasite Entamoeba histolytica have not been described. To identify a competent human effector cell, we studied the in vitro interactions of normal human polymorphonuclear neutrophils, peripheral blood mononuclear cells (PBMC), monocytes (MC), and MC-derived macrophages with virulent axenic amebae (strain HMI-IMSS). Amebae killed neutrophils, PBMC, MC, and MC-derived macrophages (P less than 0.001), without loss of parasite viability. The addition of heat-inactivated immune serum did not enable leukocytes to kill amebae, nor did it protect these host cells from amebae. MC-derived macrophages, activated with lymphokine elicited by the mitogens conconavalin A, phytohemagglutinin, or an amebic soluble protein preparation (strain HK9), killed 55% of amebae by 3 h in a trypan blue exclusion assay (P less than 0.001); during this time, 40% of the activated macrophages died. Lysis of amebae was confirmed using 111Indium oxine radiolabeled parasites and was antibody independent. Macrophage death appeared to be due to the deleterious effect of lysed amebae rather than the contact-dependent effector mechanisms of E. histolytica. Adherence between activated macrophages and amebae was greater than that between other leukocytes and amebae (P less than 0.001). Microscopic observations, kinetic analysis of the killing of amebae by activated macrophages, and suspension of amebae with adherent activated macrophages in a 10% dextran solution indicated that contact by activated macrophages was necessary to initiate the killing of amebae. Catalase but not superoxide dismutase inhibited the amebicidal capacity of activated macrophages (P less than 0.001). However, activated macrophages from an individual with chronic granulomatous disease were able to kill amebae, but not as effectively as normal cells (P less than 0.01). In summary, activated MC-derived macrophages killed virulent E. histolytica

  12. Bactericidal Activity of Human Macrophages: Analysis of Factors Influencing the Killing of Listeria monocytogenes

    PubMed Central

    Cline, Martin J.

    1970-01-01

    A technique is described for the measurement of listericidal activity of human macrophages grown from blood monocytes. Phagocytosis of Listeria monocytogenes was inhibited by a glycolytic poison (NaF) but was unaffected by anaerobic conditions, cyanide, or 2,4-dinitrophenol (DNP). Killing by macrophages was slower than that by neutrophils, and Listeria phagocytized by macrophages began to synthesize deoxyribonucleic acid within 3 hr of the time of ingestion. Differentiated macrophages ingested and killed more organisms per cell than newly isolated monocytes. Maximal killing of Listeria required oxygen but was unaffected by cyanide or DNP. Macrophages isolated from patients with chronic intracellular infection (leprosy, tuberculosis, fungal diseases) and from patients with active Hodgkin's disease were more bactericidal than macrophages from normal subjects. Images PMID:16557814

  13. Increases in Calmodulin Abundance and Stabilization of Activated iNOS Mediate Bacterial Killing in RAW 264.7 Macrophages

    SciTech Connect

    Smallwood, Heather S.; Shi, Liang; Squier, Thomas C.

    2006-08-01

    The rapid activation of macrophages in response to bacterial antigens is central to the innate immune system that permits the recognition and killing of pathogens to limit infection. To understand regulatory mechanisms underlying macrophage activation, we have investigated changes in the abundance of calmodulin (CaM) and iNOS in response to the bacterial cell wall component lipopolysaccharide (LPS) using RAW 264.7 macrophages. Critical to these measurements was the ability to differentiate free iNOS from the CaM-bound (active) form of iNOS associated with nitric oxide generation. We observe a rapid two-fold increase in CaM abundance during the first 30 minutes that is blocked by inhibition of NF?B nuclear translocation or protein synthesis. A similar two-fold increase in the abundance of the complex between CaM and iNOS is observed with the same time dependence. In contrast, there are no detectable increases in the CaM-free (i.e., inactive) form of iNOS within the first hour; it remains at a very low abundance during the initial phase of macrophage activation. Increasing cellular CaM levels in stably transfected cells results in a corresponding increase in the abundance of the CaM/iNOS complex that promotes effective bacterial killing following challenge by Salmonella typhimurium. Thus, LPS-dependent increases in CaM abundance function in the stabilization and activation of iNOS on the rapid time-scale associated with macrophage activation and bacterial killing. These results explain how CaM and iNOS coordinately function to form a stable complex that is part of a rapid host-response that functions within the first 30 minutes following bacterial infection to up-regulate the innate immune system involving macrophage activation.

  14. Reactive-oxygen-species-mediated P. aeruginosa killing is functional in human cystic fibrosis macrophages.

    PubMed

    Cifani, Noemi; Pompili, Barbara; Anile, Marco; Patella, Miriam; Diso, Daniele; Venuta, Federico; Cimino, Giuseppe; Quattrucci, Serena; Di Domenico, Enea Gino; Ascenzioni, Fiorentina; Del Porto, Paola

    2013-01-01

    Pseudomonas aeruginosa is the most common pathogen for chronic lung infection in cystic fibrosis (CF) patients. About 80% of adult CF patients have chronic P. aeruginosa infection, which accounts for much of the morbidity and most of the mortality. Both bacterial genetic adaptations and defective innate immune responses contribute to the bacteria persistence. It is well accepted that CF transmembrane conductance regulator (CFTR) dysfunction impairs the airways-epithelium-mediated lung defence; however, other innate immune cells also appear to be affected, such as neutrophils and macrophages, which thus contribute to this infectious pathology in the CF lung. In macrophages, the absence of CFTR has been linked to defective P. aeruginosa killing, increased pro-inflammatory cytokine secretion, and reduced reactive oxygen species (ROS) production. To learn more about macrophage dysfunction in CF patients, we investigated the generation of the oxidative burst and its impact on bacterial killing in CF macrophages isolated from peripheral blood or lung parenchyma of CF patients, after P. aeruginosa infection. Our data demonstrate that CF macrophages show an oxidative response of similar intensity to that of non-CF macrophages. Intracellular ROS are recognized as one of the earliest microbicidal mechanisms against engulfed pathogens that are activated by macrophages. Accordingly, NADPH inhibition resulted in a significant increase in the intracellular bacteria survival in CF and non-CF macrophages, both as monocyte-derived macrophages and as lung macrophages. These data strongly suggest that the contribution of ROS to P. aeruginosa killing is not affected by CFTR mutations.

  15. Effects of inhibitors of tumoricidal activity upon schistosomulum killing by activated macrophages.

    PubMed

    James, S L; Glaven, J A

    1987-12-01

    Larvae of the helminth parasite Schistosoma mansoni are efficiently killed in vitro by lymphokine-activated macrophages, leading to the hypothesis that these cells may participate in the effector mechanism of protective immunity against schistosomiasis. Larvacidal activity has also been demonstrated in the IC-21 macrophage cell line in the absence of a demonstrable respiratory burst, indicating that macrophages possess nonoxidative mechanisms of schistosomulum killing. In this study, we demonstrated that IC-21 larval killing was most effective when contact was allowed between cells and target. Nonoxidative larvacidal activity was prevented by protein synthesis inhibitors, by the inhibition of microtubule polymerization, and by tosyllysylchloromethylketone but not by other inhibitors or substrates of tryptic or chymotryptic protease activity. The addition of excess iron to the culture also prevented IC-21-mediated larval killing, suggesting that the production of an iron-binding molecule may be involved. In contrast, the addition of excess thymidine or arginine did not reverse macrophage larvacidal activity, nor did lysosomotropic agents that depress the activity of acid hydrolases. Under appropriate conditions of activation and surface membrane stimulation, IC-21 cells could be induced to release soluble cytotoxic factors retaining larvacidal activity. These observations provide insight into the mechanism of macrophage-mediated schistosome killing, in comparison to the cytotoxic mechanisms described in the better-studied tumoricidal models, and supply a basis for further biochemical investigation of macrophage function against a multicellular target. PMID:3119500

  16. Effects of inhibitors of tumoricidal activity upon schistosomulum killing by activated macrophages.

    PubMed Central

    James, S L; Glaven, J A

    1987-01-01

    Larvae of the helminth parasite Schistosoma mansoni are efficiently killed in vitro by lymphokine-activated macrophages, leading to the hypothesis that these cells may participate in the effector mechanism of protective immunity against schistosomiasis. Larvacidal activity has also been demonstrated in the IC-21 macrophage cell line in the absence of a demonstrable respiratory burst, indicating that macrophages possess nonoxidative mechanisms of schistosomulum killing. In this study, we demonstrated that IC-21 larval killing was most effective when contact was allowed between cells and target. Nonoxidative larvacidal activity was prevented by protein synthesis inhibitors, by the inhibition of microtubule polymerization, and by tosyllysylchloromethylketone but not by other inhibitors or substrates of tryptic or chymotryptic protease activity. The addition of excess iron to the culture also prevented IC-21-mediated larval killing, suggesting that the production of an iron-binding molecule may be involved. In contrast, the addition of excess thymidine or arginine did not reverse macrophage larvacidal activity, nor did lysosomotropic agents that depress the activity of acid hydrolases. Under appropriate conditions of activation and surface membrane stimulation, IC-21 cells could be induced to release soluble cytotoxic factors retaining larvacidal activity. These observations provide insight into the mechanism of macrophage-mediated schistosome killing, in comparison to the cytotoxic mechanisms described in the better-studied tumoricidal models, and supply a basis for further biochemical investigation of macrophage function against a multicellular target. PMID:3119500

  17. The role of macrophages in the cytotoxic killing of tumour cells in vitro

    PubMed Central

    Zembala, M.; Ptak, W.; Hanczakowska, Maria

    1973-01-01

    Lymph node and spleen cells from normal mice were cultured for 3 days with polyoma virus-induced tumour, Ehrlich's ascites tumour or leukaemia L 1210 cells. This resulted in in vitro immunization of the lymphocytes, which were then transferred to irradiated target cells labelled with 51Cr. Normal, i.e. non-immune thioglycollate-stimulated peritoneal macrophages were also added to some tubes. Non-immune macrophages mixed with immunized lymphocytes showed a significantly increased ability to destroy tumour cells as compared with macrophages in the absence of immunized lymphocytes. The immunized lymphocytes were almost entirely inactive alone. When the number of macrophages was kept constant the cytotoxicity was dependent on the number of viable immunized lymphocytes placed on the target cells. Immunized lymphocytes, in the presence of macrophages, only exhibited strong killing of the target cells against which they had been immunized; some lysis of `bystander' cells was, however, seen provided specific target cells were present. Macrophage monolayers exposed to immunized lymphocytes upon contact with specific antigen became `armed' and showed a significant cytotoxicity for specific target cells. When immunized lymphocytes and normal macrophages were treated with actinomycin D and puromycin, cytotoxicity was inhibited in the immunized lymphocytes but not in the macrophages. The possible mechanism of normal macrophage cooperation with immunized lymphocytes in the cytotoxic killing reaction is discussed. Results presented in this paper favour the view that immunologically specific cytophilic factor (presumptive cytophilic antibody) is involved in the macrophage-mediated cytotoxicity in the system studied. PMID:4356674

  18. Phagocytosis, germination and killing of Bacillus subtilis spores presenting heterologous antigens in human macrophages.

    PubMed

    Ceragioli, Mara; Cangiano, Giuseppina; Esin, Semih; Ghelardi, Emilia; Ricca, Ezio; Senesi, Sonia

    2009-02-01

    Bacillus subtilis is a Gram-positive spore-bearing bacterium long used as a probiotic product and more recently regarded as an attractive vehicle for delivering heterologous antigens to be used for mucosal vaccination. This report describes the in vitro interaction between human macrophages and B. subtilis spores displaying the tetanus toxin fragment C or the B subunit of the heat-labile toxin of Escherichia coli on their surface in comparison to spores of the parental strain. Recombinant and parental B. subtilis spores were similarly internalized by human macrophages, at a frequency lower than 2.5%. Inside macrophages, nearly all spores germinated and were killed within 6 h. Using germination-defective spores and inhibiting spore germination inside macrophages, evidence was produced that only germinated spores were killed by human macrophages and that intracellular spore germination was mediated by an alanine-dependent pathway. The germinated spores were killed by macrophages before any round of cell duplication, as estimated by fluorescence microscopy analysis of macrophages infected with spores carrying the gfp gene fused to abrB, a B. subtilis gene shown here to be expressed at the transition between outgrowth and vegetative growth. Monitoring of macrophage infection never revealed cytotoxic effects being exerted by B. subtilis spores. These in vitro data support the hypothesis that B. subtilis spores may potentially be used as a suitable and safe vehicle for administering heterologous antigens to humans.

  19. Macrophages sense and kill bacteria through carbon monoxide–dependent inflammasome activation

    PubMed Central

    Wegiel, Barbara; Larsen, Rasmus; Gallo, David; Chin, Beek Yoke; Harris, Clair; Mannam, Praveen; Kaczmarek, Elzbieta; Lee, Patty J.; Zuckerbraun, Brian S.; Flavell, Richard; Soares, Miguel P.; Otterbein, Leo E.

    2014-01-01

    Microbial clearance by eukaryotes relies on complex and coordinated processes that remain poorly understood. The gasotransmitter carbon monoxide (CO) is generated by the stress-responsive enzyme heme oxygenase-1 (HO-1, encoded by Hmox1), which is highly induced in macrophages in response to bacterial infection. HO-1 deficiency results in inadequate pathogen clearance, exaggerated tissue damage, and increased mortality. Here, we determined that macrophage-generated CO promotes ATP production and release by bacteria, which then activates the Nacht, LRR, and PYD domains-containing protein 3 (NALP3) inflammasome, intensifying bacterial killing. Bacterial killing defects in HO-1–deficient murine macrophages were restored by administration of CO. Moreover, increased CO levels enhanced the bacterial clearance capacity of human macrophages and WT murine macrophages. CO-dependent bacterial clearance required the NALP3 inflammasome, as CO did not increase bacterial killing in macrophages isolated from NALP3-deficient or caspase-1–deficient mice. IL-1β cleavage and secretion were impaired in HO-1–deficient macrophages, and CO-dependent processing of IL-1β required the presence of bacteria-derived ATP. We found that bacteria remained viable to generate and release ATP in response to CO. The ATP then bound to macrophage nucleotide P2 receptors, resulting in activation of the NALP3/IL-1β inflammasome to amplify bacterial phagocytosis by macrophages. Taken together, our results indicate that macrophage-derived CO permits efficient and coordinated regulation of the host innate response to invading microbes. PMID:25295542

  20. Killing Is Not Enough: How Apoptosis Hijacks Tumor-Associated Macrophages to Promote Cancer Progression.

    PubMed

    Weigert, Andreas; Mora, Javier; Sekar, Divya; Syed, Shahzad; Brüne, Bernhard

    2016-01-01

    Macrophages are a group of heterogeneous cells of the innate immune system that are crucial to the initiation, progression, and resolution of inflammation. Moreover, they control tissue homeostasis in healthy tissue and command a broad sensory arsenal to detect disturbances in tissue integrity. Macrophages possess a remarkable functional plasticity to respond to irregularities and to initiate programs that allow overcoming them in order to return back to normal. Thus, macrophages kill malignant or transformed cells, rearrange extracellular matrix, take up and recycle cellular as well as molecular debris, initiate cellular growth cascades, and favor directed migration of cells. As an example, apoptotic death of bystander cells is sensed by macrophages, initiating functional responses that support all hallmarks of cancer. In this chapter, we describe how tumor cell apoptosis hijacks tumor-associated macrophages to promote tumor growth. We propose that tumor therapy should not only kill malignant cells but also target the interaction of the host with apoptotic cancer cells, as this might be efficient to limit the protumor action of apoptotic cells and boost the antitumor potential of macrophages. Leaving the apoptotic cell/macrophage interaction untouched might also limit the benefit of conventional tumor cell apoptosis-focused therapy since surviving tumor cells might receive overwhelming support by the wound healing response that apoptotic tumor cells will trigger in local macrophages, thereby enhancing tumor recurrence. PMID:27558823

  1. Alveolar macrophages in pulmonary host defence – the unrecognized role of apoptosis as a mechanism of intracellular bacterial killing

    PubMed Central

    Aberdein, J D; Cole, J; Bewley, M A; Marriott, H M; Dockrell, D H

    2013-01-01

    Alveolar macrophages play an essential role in clearing bacteria from the lower airway, as the resident phagocyte alveolar macrophages must both phagocytose and kill bacteria, and if unable to do this completely must co-ordinate an inflammatory response. The decision to escalate the inflammatory response represents the transition between subclinical infection and the development of pneumonia. Alveolar macrophages are well equipped to phagocytose bacteria and have a large phagolysosomal capacity in which ingested bacteria are killed. The rate-limiting step in control of extracellular bacteria, such as Streptococcus pneumoniae, is the capacity of alveolar macrophages to kill ingested bacteria. Therefore, alveolar macrophages complement canonical microbicidal strategies with an additional level of apoptosis-associated killing to help kill ingested bacteria. PMID:23841514

  2. Rapid effects of androgens in macrophages.

    PubMed

    Benten, W Peter M; Guo, Z; Krücken, J; Wunderlich, F

    2004-08-01

    We investigated the existence of membrane receptors for testosterone (mAR) in mouse macrophages of the cell lines IC-21 and RAW 264.7 as well as their roles in nongenomic pathways, gene expression and cell functioning. Both cell lines lack intracellular androgen receptors (iARs) and respond to testosterone with rapid rises in [Ca2+]i. These rises in [Ca2+]i can neither be inhibited by iAR- nor by iER blockers, but are rather mediated through mAR. Pharmacological approaches suggest that the mAR belongs to the class of membrane receptors which are coupled to phospholipase C via pertussis toxin (PTX) sensitive G-proteins. The mAR can be localized as specific surface binding sites for testosterone-BSA-FITC by confocal laser scanning microscopy (CLSM)and flow cytometry, and are characterized by their agonist-sequestrability. In order to examine a possible role of the testosterone-induced rise in [Ca2+]i on gene expression, a c-fos promoter reporter gene construct was transfected into RAW 264.7 macrophages. The increase in [Ca2+]i induced by testosterone cannot significantly activate the c-fos promoter directly. Also, no significant activation of ERK1/2, JNK/SAPK and p38 can be observed following testosterone-stimulation alone. However, testosterone-induced rises in [Ca2+]i do have specific effects on gene expression in context with lipopolysaccharide (LPS)-induced genotropic signaling: testosterone specifically down-regulates LPS-induced activation of c-fos promoter, p38 MAPK and NO production. In fetal calf serum (FCS)-induced genotropic signaling, the situation is reversed, i.e. testosterone augments the activation of c-fos promoter and ERK1/2. Our studies demonstrate a cross-talk between the testosterone-induced nongenomic Ca2+ signaling and the genotropic signaling induced by LPS and FCS in macrophages. PMID:15288774

  3. Phenylbutyrate induces LL-37-dependent autophagy and intracellular killing of Mycobacterium tuberculosis in human macrophages.

    PubMed

    Rekha, Rokeya Sultana; Rao Muvva, S S V Jagadeeswara; Wan, Min; Raqib, Rubhana; Bergman, Peter; Brighenti, Susanna; Gudmundsson, Gudmundur H; Agerberth, Birgitta

    2015-01-01

    LL-37 is a human antimicrobial peptide (AMP) of the cathelicidin family with multiple activities including a mediator of vitamin D-induced autophagy in human macrophages, resulting in intracellular killing of Mycobacterium tuberculosis (Mtb). In a previous trial in healthy volunteers, we have shown that LL-37 expression and subsequent Mtb-killing can be further enhanced by 4-phenylbutyrate (PBA), also an inducer of LL-37 expression. Here, we explore a potential mechanism(s) behind PBA and LL-37-induced autophagy and intracellular killing of Mtb. Mtb infection of macrophages downregulated the expression of both the CAMP transcript and LL-37 peptide as well as certain autophagy-related genes (BECN1 and ATG5) at both the mRNA and protein levels. In addition, activation of LC3-II in primary macrophages and THP-1 cells was not detected. PBA and the active form of vitamin D3 (1,25[OH]2D3), separately or particularly in combination, were able to overcome Mtb-induced suppression of LL-37 expression. Notably, reactivation of autophagy occurred by stimulation of macrophages with PBA and promoted colocalization of LL-37 and LC3-II in autophagosomes. Importantly, PBA treatment failed to induce autophagy in Mtb-infected THP-1 cells, when the expression of LL-37 was silenced. However, PBA-induced autophagy was restored when the LL-37 knockdown cells were supplemented with synthetic LL-37. Interestingly, we have found that LL-37-induced autophagy was mediated via P2RX7 receptor followed by enhanced cytosolic free Ca(2+), and activation of AMPK and PtdIns3K pathways. Altogether, these results suggest a novel activity for PBA as an inducer of autophagy, which is LL-37-dependent and promotes intracellular killing of Mtb in human macrophages.

  4. L-arginine-dependent reactive nitrogen intermediates as mediators of tumor cell killing by activated macrophages.

    PubMed

    Keller, R; Geiges, M; Keist, R

    1990-03-01

    The capacities of lymphokines and of various microbes to induce in a pure population of bone marrow-derived mononuclear phagocytes tumoricidal activity and/or the production of L-arginine-dependent reactive nitrogen intermediates, measured by the release of nitrite, were comparatively assessed. These parameters were found to be closely correlated in a variety of experimental situations, i.e., enhanced by a surplus of L-arginine and abrogated by N-monomethyl-L-arginine, a selective inhibitor of L-arginine-dependent effector mechanisms. In other macrophage/tumor cell combinations, such correlation was less obvious or not at all detectable, suggesting that, in these models, L-arginine-dependent reactive nitrogen intermediates are not or not alone responsible for the mediation of tumoricidal activity by activated macrophages. Collectively, the present findings suggest that the mechanism of tumor cell killing by activated macrophages may differ, depending on the tumor cell type and the pathway of macrophage activation. Among the various effector mechanisms considered to be involved in tumor cell killing by activated macrophages, L-arginine-dependent reactive nitrogen intermediates appear to hold a major role.

  5. Mannosylated lipoarabinomannans from Mycobacterium avium subsp. paratuberculosis alters the inflammatory response by bovine macrophages and suppresses killing of Mycobacterium avium subsp. avium organisms.

    PubMed

    Souza, Cleverson; Davis, William C; Eckstein, Torsten M; Sreevatsan, Srinand; Weiss, Douglas J

    2013-01-01

    Analysis of the mechanisms through which pathogenic mycobacteria interfere with macrophage activation and phagosome maturation have shown that engagement of specific membrane receptors with bacterial ligands is the initiating event. Mannosylated lipoarabinomannan (Man-LAM) has been identified as one of the ligands that modulates macrophage function. We evaluated the effects of Man-LAM derived from Mycobacterium avium subsp. paratuberculosis (MAP) on bovine macrophages. Man-LAM induced a rapid and prolonged expression of IL-10 message as well as transient expression of TNF-α. Preincubation with Man-LAM for up to 16 h did not suppress expression of IL-12 in response to interferon-γ. Evaluation of the effect of Man-LAM on phagosome acidification, phagosome maturation, and killing of Mycobacterium avium subsp. avium (MAA) showed that preincubation of macrophages with Man-LAM before addition of MAA inhibited phagosome acidification, phagolysosome fusion, and reduced killing. Analysis of signaling pathways provided indirect evidence that inhibition of killing was associated with activation of the MAPK-p38 signaling pathway but not the pathway involved in regulation of expression of IL-10. These results support the hypothesis that MAP Man-LAM is one of the virulence factors facilitating survival of MAP in macrophages.

  6. Ursolic Acid Activates Intracellular Killing Effect of Macrophages During Mycobacterium tuberculosis Infection.

    PubMed

    Podder, Biswajit; Jang, Woong Sik; Nam, Kung-Woo; Lee, Byung-Eui; Song, Ho-Yeon

    2015-05-01

    Tuberculosis is one of the most threatening infectious diseases to public health all over the world, for which Mycobacterium tuberculosis (MTB) is the etiological agent of pathogenesis. Ursolic acid (UA) has immunomodulatory function and exhibits antimycobacterial activity. However, the intracellular killing effect of UA has yet to be elucidated. The aim of this study was to evaluate the intracellular killing effect of UA during mycobacterial infection. The intracellular killing activity of UA was evaluated in the macrophage cell line THP-1 by the MGIT 960 system as well as by CFU count. The production of reactive oxygen species (ROS) and the level of nitric oxide (NO) were measured using DCF-DA and Griess reagent, respectively. Phagocytosis was observed by a fluorescence-based staining method, and the colony forming units were enumerated on 7H11 agar medium following infection. In addition, MRP8 mRNA expression was measured by qRT-PCR. UA significantly decreased the number of intracellular Mycobacterium through generation of ROS and NO. In addition, it profoundly activated the phagocytosis process of THP-1 cells during MTB-infection. Furthermore, our data demonstrated that UA activated the phagocytosis process in human monocyte cells through MRP8 induction. These data suggest that UA firmly contributes to the intracellular killing effect of macrophages during mycobacterial infection.

  7. P2X7 receptor-mediated killing of an intracellular parasite, Toxoplasma gondii, by human and murine macrophages1

    PubMed Central

    Lees, Michael P.; Fuller, Stephen J.; McLeod, Rima; Boulter, Nicola R.; Miller, Catherine M.; Zakrzewski, Alana M.; Mui, Ernest J.; Witola, William H.; Coyne, Jessica J.; Hargrave, Aubrey C.; Jamieson, Sarra E.; Blackwell, Jenefer M.; Wiley, James S.; Smith, Nicholas C.

    2010-01-01

    The P2X7 receptor (P2X7R)4 is highly expressed on the macrophage cell surface and activation of infected cells by extracellular ATP has been shown to kill intracellular bacteria and parasites. Furthermore, single nucleotide polymorphisms (SNPs) that decrease receptor function reduce the ability of human macrophages to kill Mycobacterium tuberculosis and are associated with extrapulmonary tuberculosis. In this paper we show that macrophages from people with the 1513C (rs3751143) loss-of-function P2X7R SNP are less effective in killing intracellular Toxoplasma gondii after exposure to ATP compared with macrophages from people with the 1513A wild-type allele. Supporting a P2X7R-specific effect on T. gondii, macrophages from P2X7R knock-out mice (P2X7R−/−) are unable to kill T. gondii as effectively as macrophages from wild-type mice. We show that P2X7R-mediated T. gondii killing occurs in parallel with host cell apoptosis and is independent of NO production. PMID:20488797

  8. Bacterial killing in macrophages and amoeba: do they all use a brass dagger?

    PubMed

    German, Nadezhda; Doyscher, Dominik; Rensing, Christopher

    2013-10-01

    Macrophages are immune cells that are known to engulf pathogens and destroy them by employing several mechanisms, including oxidative burst, induction of Fe(II) and Mn(II) efflux, and through elevation of Cu(I) and Zn(II) concentrations in the phagosome ('brass dagger'). The importance of the latter mechanism is supported by the presence of multiple counteracting efflux systems in bacteria, responsible for the efflux of toxic metals. We hypothesize that similar bacteria-killing mechanisms are found in predatory protozoa/amoeba species. Here, we present a brief summary of soft metal-related mechanisms used by macrophages, and perhaps amoeba, to inactivate and destroy bacteria. Based on this, we think it is likely that copper resistance is also selected for by protozoan grazing in the environment.

  9. Post lung-stage schistosomula of Schistosoma mansoni exhibit transient susceptibility to macrophage-mediated cytotoxicity in vitro that may relate to late phase killing in vivo.

    PubMed

    Pearce, E J; James, S L

    1986-09-01

    Studies of protective immunity against Schistosoma mansoni in immunized mice suggest that a proportion of challenge parasites may be eliminated after they have passed through the lungs of the host several days after infection; however, no potential immune effector mechanism of resistance against this stage of the parasite has yet been identified, since schistosomes have been shown to rapidly become resistant to antibody-dependent killing mechanisms. In this study, different development stages of S. mansoni were examined for their susceptibility to in vitro cytotoxicity by lymphokine-activated macrophages. As previously shown, newly transformed larvae were readily killed by lymphokine-treated peritoneal macrophages or the macrophage cell line IC-21 (80% mortality over 48 h in vitro), whereas 7 and 10 day old lung-stage parasites had become refractory to macrophage effects. However, after 2 to 2 1/2 weeks of development in vivo, juvenile parasites recovered from the liver were again susceptible to activated macrophage-mediated cytotoxicity (25-65% mortality). Ultrastructural studies of 2 1/2 week old parasites co-cultured with activated IC-21 cells revealed that damage was largely restricted to the areas beneath the parasite surface and gut syncitia; surface membrane disruption was not evident. This late stage of susceptibility was transient and by 4 to 6 weeks liver-stage worms had again become refractory to macrophage killing. The interaction of post lung-stage parasites with activated macrophages was antibody independent. Furthermore, schistosomes isolated from the portal circulation 2 1/2 weeks after infection showed no evidence of surface-bound immunoglobulin in a quantitative immunofluorescence assay, nor did antisera from chronically infected mice (CIS) or mice vaccinated with irradiated cercariae (VS) react with the surface of these parasites in vitro, making the possibility of direct antibody-dependent killing mechanisms unlikely. However, both CIS and VS

  10. Mannose-binding lectin enhances phagocytosis and killing of Neisseria meningitidis by human macrophages.

    PubMed

    Jack, Dominic L; Lee, Margaret E; Turner, Malcolm W; Klein, Nigel J; Read, Robert C

    2005-03-01

    Deficiency of mannose-binding lectin (MBL) is probably the most common human immunodeficiency and is associated with an increased risk of mucosally acquired infections including meningococcal disease. Tissue macrophages are an important component of mucosal defense, and so we determined the effect of MBL on uptake of meningococci by human monocyte-derived macrophages. Opsonization with MBL significantly increased the capture and doubled the amount of internalization of Neisseria meningitidis. Inhibition of f-actin polymerization indicated that MBL exerted this effect by a dose-dependent acceleration of uptake into phagosomes, which was maximal within the normal physiological concentration of MBL (1.5 microg/ml) and was independent of scavenger receptors. MBL accelerated the acquisition and subsequent loss of the early endosome marker, early endosomal antigen-1, and enhanced the acquisition of the late endosomal marker, lysosome-associated membrane protein-1. MBL reduced the survival of meningococci within macrophages by more than half, despite the increased uptake of organisms, and significantly reduced the number of viable extracellular bacteria by 80%. We conclude that MBL is a dependent opsonin able to accelerate microbial uptake and killing. These results suggest that MBL could modify disease susceptibility by modulating macrophage interactions with mucosal organisms at the site of initial acquisition.

  11. Antibodies Against Sporothrix schenckii Enhance TNF-α Production and Killing by Macrophages.

    PubMed

    Franco, D de Lima; Nascimento, R C; Ferreira, K S; Almeida, S R

    2012-02-01

    Sporotrichosis is a chronic granulomatous mycosis caused by the dimorphic fungus Sporothrix schenckii. The immunological mechanisms involved in the prevention and control of sporotrichosis suggest that cell-mediated immunity plays an important role in protecting the host against S. schenckii. Nonetheless, recent data strongly support the existence of protective Abs against this pathogenic fungus. In a previous study, we showed that passive Ab therapy led to a significant reduction in the number of colony forming unit in the organs of mice when the MAb was injected before and during S. schenckii infection. The ability of opsonization to enhance macrophage damage to S. schenckii and subsequent cytokine production was investigated in this work. Here we show that the fungicidal characteristics of macrophages are increased when the fungus is phagocytosed in the presence of inactivated serum from mice infected with S. schenckii or mAb anti-gp70. Additionally, we show an increase in the levels of pro-inflammatory cytokines such as TNF-α and IL-1β. This study provides additional support for the importance of antibodies in protecting against S. schenckii and concludes that opsonization is an important process to increase TNF-α production and fungus killing by macrophages in experimental sporotrichosis.

  12. Phagocytic and chemiluminescent responses of mouse peritoneal macrophages to living and killed Salmonella typhimurium and other bacteria.

    PubMed Central

    Tomita, T; Blumenstock, E; Kanegasaki, S

    1981-01-01

    In the presence of luminol, resident as well as thioglycolate-induced and immunized macrophages emitted chemiluminescence more efficiently when the cells were exposed to living Salmonella typhimurium than when they were exposed to the same bacterium killed by ultraviolet light or heat. This phenomenon was observed whether or not the bacterium was opsonized. The different response to living and killed bacteria was also found with Escherichia coli, Pseudomonas aeruginosa, Proteus morganii, and Enterobacter aerogenes, but not with Shigella sonnei, Klebsiella pneumoniae, and Propionibacterium acnes. The results suggest that macrophages respond better to living, motile bacteria than to nonmotile or killed bacteria. The experimental results obtained with motility mutants of S. typhimurium, E. coli, and P. aeruginosa confirm that macrophages exposed to the motile bacteria emit chemiluminescence more efficiently and ingest the motile bacteria at a much faster rate than the nonmotile bacteria. Images PMID:6788707

  13. Phagocytic and chemiluminescent responses of mouse peritoneal macrophages to living and killed Salmonella typhimurium and other bacteria

    SciTech Connect

    Tomita, T.; Blumenstock, E.; Kanegasaki, S.

    1981-06-01

    In the presence of luminol, resident as well as thioglycolate-induced and immunized macrophages emitted chemiluminescence more efficiently when the cells were exposed to living Salmonella typhimurium than when they were exposed to the same bacterium killed by ultraviolet light or heat. This phenomenon was observed whether or not the bacterium was opsonized. The different response to living and killed bacteria was also found with Escherichia coli, Pseudomonas aeruginosa, Proteus morganii, and Enterobacter aerogenes, but not with Shigella sonnei, Klebsiella pneumoniae, and Propionibacterium acnes. The results suggest that macrophages respond better to living, motile bacteria than to nonmotile or killed bacteria. The experimental results obtained with motility mutants of S. typhimurium, E. coli, and P. aeruginosa confirm that macrophages exposed to the motile bacteria emit chemiluminescence more efficiently and ingest the motile bacteria at a much faster rate than the nonmotile bacteria.

  14. Comparison between three adjuvants for a vaccine against canine leishmaniasis: In vitro evaluation of macrophage killing ability.

    PubMed

    Trotta, T; Fasanella, A; Scaltrito, D; Gradoni, L; Mitolo, V; Brandonisio, O; Acquafredda, A; Panaro, M A

    2010-03-01

    The aim of this study was to evaluate, in terms of dog macrophage killing ability in vitro, a vaccine based on Leishmania infantum promastigote soluble antigen (LSA) formulated with three different adjuvants (BCG, AdjuPrime, MPL/TDM/CWS). A significant increase of the macrophage killing ability was observed in dogs vaccinated with LSA+MPL/TDM/CWS after 1 month from vaccination. A similar increase of macrophage parasitocidal ability was present only after 5 months in dogs vaccinated with LSA+BCG or LSA+AdjuPrime. In all dogs the augmented killing percentage was still present after 12 months from vaccination. Therefore, in particular LSA+MPL/TDM/CWS vaccine seems promising for further studies in dogs.

  15. Delineating the importance of serum opsonins and the bacterial capsule in affecting the uptake and killing of Burkholderia pseudomallei by murine neutrophils and macrophages.

    PubMed

    Mulye, Minal; Bechill, Michael P; Grose, William; Ferreira, Viviana P; Lafontaine, Eric R; Wooten, R Mark

    2014-08-01

    Infection of susceptible hosts by the encapsulated Gram-negative bacterium Burkholderia pseudomallei (Bp) causes melioidosis, with septic patients attaining mortality rates ≥ 40%. Due to its high infectivity through inhalation and limited effective therapies, Bp is considered a potential bioweapon. Thus, there is great interest in identifying immune effectors that effectively kill Bp. Our goal is to compare the relative abilities of murine macrophages and neutrophils to clear Bp, as well as determine the importance of serum opsonins and bacterial capsule. Our findings indicate that murine macrophages and neutrophils are inherently unable to clear either unopsonized Bp or the relatively-avirulent acapsular bacterium B. thailandensis (Bt). Opsonization of Bp and Bt with complement or pathogen-specific antibodies increases macrophage-uptake, but does not promote clearance, although antibody-binding enhances complement deposition. In contrast, complement opsonization of Bp and Bt causes enhanced uptake and killing by neutrophils, which is linked with rapid ROS induction against bacteria exhibiting a threshold level of complement deposition. Addition of bacteria-specific antibodies enhances complement deposition, but antibody-binding alone cannot elicit neutrophil clearance. Bp capsule provides some resistance to complement deposition, but is not anti-phagocytic or protective against reactive oxygen species (ROS)-killing. Macrophages were observed to efficiently clear Bp only after pre-activation with IFNγ, which is independent of serum- and/or antibody-opsonization. These studies indicate that antibody-enhanced complement activation is sufficient for neutrophil-clearance of Bp, whereas macrophages are ineffective at clearing serum-opsonized Bp unless pre-activated with IFNγ. This suggests that effective immune therapies would need to elicit both antibodies and Th1-adaptive responses for successful prevention/eradication of melioidosis.

  16. Delineating the Importance of Serum Opsonins and the Bacterial Capsule in Affecting the Uptake and Killing of Burkholderia pseudomallei by Murine Neutrophils and Macrophages

    PubMed Central

    Mulye, Minal; Bechill, Michael P.; Grose, William; Ferreira, Viviana P.; Lafontaine, Eric R.; Wooten, R. Mark

    2014-01-01

    Infection of susceptible hosts by the encapsulated Gram-negative bacterium Burkholderia pseudomallei (Bp) causes melioidosis, with septic patients attaining mortality rates ≥40%. Due to its high infectivity through inhalation and limited effective therapies, Bp is considered a potential bioweapon. Thus, there is great interest in identifying immune effectors that effectively kill Bp. Our goal is to compare the relative abilities of murine macrophages and neutrophils to clear Bp, as well as determine the importance of serum opsonins and bacterial capsule. Our findings indicate that murine macrophages and neutrophils are inherently unable to clear either unopsonized Bp or the relatively-avirulent acapsular bacterium B. thailandensis (Bt). Opsonization of Bp and Bt with complement or pathogen-specific antibodies increases macrophage-uptake, but does not promote clearance, although antibody-binding enhances complement deposition. In contrast, complement opsonization of Bp and Bt causes enhanced uptake and killing by neutrophils, which is linked with rapid ROS induction against bacteria exhibiting a threshold level of complement deposition. Addition of bacteria-specific antibodies enhances complement deposition, but antibody-binding alone cannot elicit neutrophil clearance. Bp capsule provides some resistance to complement deposition, but is not anti-phagocytic or protective against reactive oxygen species (ROS)-killing. Macrophages were observed to efficiently clear Bp only after pre-activation with IFNγ, which is independent of serum- and/or antibody-opsonization. These studies indicate that antibody-enhanced complement activation is sufficient for neutrophil-clearance of Bp, whereas macrophages are ineffective at clearing serum-opsonized Bp unless pre-activated with IFNγ. This suggests that effective immune therapies would need to elicit both antibodies and Th1-adaptive responses for successful prevention/eradication of melioidosis. PMID:25144195

  17. Autophagy-Related Proteins Target Ubiquitin-Free Mycobacterial Compartment to Promote Killing in Macrophages

    PubMed Central

    Bah, Aïcha; Lacarrière, Camille; Vergne, Isabelle

    2016-01-01

    Autophagy is a lysosomal degradative process that plays essential functions in innate immunity, particularly, in the clearance of intracellular bacteria such as Mycobacterium tuberculosis. The molecular mechanisms involved in autophagy activation and targeting of mycobacteria, in innate immune responses of macrophages, are only partially characterized. Autophagy targets pathogenic M. tuberculosis via a cytosolic DNA recognition- and an ubiquitin-dependent pathway. In this report, we show that non-pathogenic M. smegmatis induces a robust autophagic response in THP-1 macrophages with an up regulation of several autophagy-related genes. Autophagy activation relies in part on recognition of mycobacteria by Toll-like receptor 2 (TLR2). Notably, LC3 targeting of M. smegmatis does not rely on membrane damage, ubiquitination, or autophagy receptor recruitment. Lastly, M. smegmatis promotes recruitment of several autophagy proteins, which are required for mycobacterial killing. In conclusion, our study uncovered an alternative autophagic pathway triggered by mycobacteria which involves cell surface recognition but not bacterial ubiquitination. PMID:27242971

  18. Nitric oxide not apoptosis mediates differential killing of Mycobacterium bovis in bovine macrophages.

    PubMed

    Esquivel-Solís, Hugo; Vallecillo, Antonio J; Benítez-Guzmán, Alejandro; Adams, L Garry; López-Vidal, Yolanda; Gutiérrez-Pabello, José A

    2013-01-01

    To identify the resistance phenotype against Mycobacterium bovis in cattle, we used a bactericidal assay that has been considered a marker of this trait. Three of 24 cows (12.5%) were phenotyped as resistant and 21 as susceptible. Resistance of bovine macrophages (MΦ) to BCG challenge was evaluated for its association with SLC11A1 GT microsatellite polymorphisms within 3'UTR region. Twenty-three cows (95.8%) had a GT13 genotype, reported as resistant, consequently the SLC11A1 polymorphism was not in agreement with our bactericidal assay results. MΦ of cows with resistant or susceptible phenotype were challenged in vitro with virulent M. bovis field strain or BCG, and nitric oxide production, bacterial killing and apoptosis induction were measured in resting and LPS-primed states. M. bovis field strain induced more apoptosis than BCG, although the difference was not significant. Resistant MΦ controlled better the replication of M. bovis (P<0.01), produced more nitric oxide (P<0.05) and were slightly more prone to undergo apoptosis than susceptible cells. LPS pretreatment of MΦ enhanced all the functional parameters analyzed. Inhibition of nitric oxide production with n (G)-monomethyl-L-arginine monoacetate enhanced replication of M. bovis but did not modify apoptosis rates in both resistant and susceptible MΦ. We conclude that nitric oxide production not apoptosis is a major determinant of macrophage resistance to M. bovis infection in cattle and that the influence of SLC11A1 gene 3'UTR polymorphism is not associated with this event.

  19. Nitric Oxide Not Apoptosis Mediates Differential Killing of Mycobacterium bovis in Bovine Macrophages

    PubMed Central

    Esquivel-Solís, Hugo; Vallecillo, Antonio J.; Benítez-Guzmán, Alejandro; Adams, L. Garry; López-Vidal, Yolanda; Gutiérrez-Pabello, José A.

    2013-01-01

    To identify the resistance phenotype against Mycobacterium bovis in cattle, we used a bactericidal assay that has been considered a marker of this trait. Three of 24 cows (12.5%) were phenotyped as resistant and 21 as susceptible. Resistance of bovine macrophages (MΦ) to BCG challenge was evaluated for its association with SLC11A1 GT microsatellite polymorphisms within 3′UTR region. Twenty-three cows (95.8%) had a GT13 genotype, reported as resistant, consequently the SLC11A1polymorphism was not in agreement with our bactericidal assay results. MΦ of cows with resistant or susceptible phenotype were challenged in vitro with virulent M. bovis field strain or BCG, and nitric oxide production, bacterial killing and apoptosis induction were measured in resting and LPS-primed states. M. bovis field strain induced more apoptosis than BCG, although the difference was not significant. Resistant MΦ controlled better the replication of M. bovis (P<0.01), produced more nitric oxide (P<0.05) and were slightly more prone to undergo apoptosis than susceptible cells. LPS pretreatment of MΦ enhanced all the functional parameters analyzed. Inhibition of nitric oxide production with nG-monomethyl-L-arginine monoacetate enhanced replication of M. bovis but did not modify apoptosis rates in both resistant and susceptible MΦ. We conclude that nitric oxide production not apoptosis is a major determinant of macrophage resistance to M. bovis infection in cattle and that the influence of SLC11A1 gene 3′UTR polymorphism is not associated with this event. PMID:23691050

  20. Killing of Brucella antigen-sensitized macrophages by T lymphocytes in bovine brucellosis.

    PubMed

    Wyckoff, John H; Potts, Richard D

    2007-12-15

    The present study was an investigation into the role of T lymphocytes in the killing of antigen-sensitized macrophages (M Phi) in bovine brucellosis. Following confirmation of bovine T lymphocyte cell lines derived from Brucella abortus Strain 19 vaccinated steers as antigen-specific in proliferation studies using various antigens, we adapted an apoptosis assay for evaluation of cytotoxicity by these bovine T cells against autologous monocyte-derived macrophages (MDM Phi) as target cells. Various B. abortus antigen preparations were tested including whole gamma-irradiated B. abortus bacteria (gamma BA), a soluble cytosolic protein fraction and a membrane-associated protein fraction. Both polyclonal and cloned T lymphocyte cell lines exhibited cytotoxicity against MDM Phi targets in an antigen-specific fashion. Polyclonal and cloned T lymphocyte cell lines demonstrated cytotoxic responses to varying degrees against B. abortus antigens regardless of whether the antigen used was whole nonviable bacteria, a soluble protein extract or a membrane-associated fraction of extracted bacteria. To further develop correlation of these responses to an in vivo host defense mechanism, cytotoxicity was evaluated using target cells that had been infected with live B. abortus S19 or B. abortus Strain 2308. Cytotoxic responses were also demonstrated consistently against infected targets with either strain of B. abortus although in most cases, cytotoxicity was higher against target cells sensitized with gamma BA compared to those infected with live bacteria. Cloned T lymphocyte cell lines were all CD4+, CD8(-) cells indicating that the observed cytotoxic responses were most likely due to an inflammatory Th1 response and may represent an important host defense mechanism induced by vaccination with live attenuated strains of B. abortus in cattle.

  1. The respiratory burst is not required for killing of intracellular and extracellular parasites by a lymphokine-activated macrophage cell line.

    PubMed

    Scott, P; James, S; Sher, A

    1985-06-01

    The macrophage cell line, IC-21, was found to be incapable of producing the oxygen products associated with the respiratory burst. However, IC-21 cells were activated by lymphokine (LK) to kill intracellular (Leishmania donovani amastigotes) and extracellular (Schistosoma mansoni larvae) parasites, as well as tumor cells. In each case, the cytotoxicity exhibited by activated IC-21 cells and activated peritoneal macrophages was indistinguishable. However, nonactivated IC-21 cells were unable to kill L. donovani log-growth phase promastigotes, while nonactivated peritoneal macrophages destroyed greater than 90% of the initial infection. These results indicate that amastigotes and schistosome larvae are susceptible to killing by nonoxidative cytotoxic mechanism induced by lymphokine activation but, on the other hand, support the concept that the killing of log-growth phase promastigotes by nonactivated cells is dependent upon the respiratory burst. We propose that the IC-21 cell line may be a useful model for studying nonoxidative killing functions of activated macrophages. PMID:2988973

  2. Ion Channel Blockers as Antimicrobial Agents, Efflux Inhibitors, and Enhancers of Macrophage Killing Activity against Drug Resistant Mycobacterium tuberculosis

    PubMed Central

    Perdigão, João; Couto, Isabel; Portugal, Isabel; Martins, Marta; Amaral, Leonard; Anes, Elsa; Viveiros, Miguel

    2016-01-01

    Given the ability of M. tuberculosis to survive as an intracellular pathogen and its propensity to develop resistance to the existing antituberculosis drugs, its treatment requires new approaches. Here the antimycobacterial properties of verapamil, thioridazine, chlorpromazine, flupenthixol and haloperidol were investigated against a panel of drug resistant M. tuberculosis strains, both in vitro and on human-infected macrophages. These compounds are efflux inhibitors that share among them the characteristic of being ion channel blockers. In vitro, all compounds exhibited synergistic inhibitory activities when combined with isoniazid and rifampicin, and were able to inhibit active efflux, demonstrating their role as efflux inhibitors. Gene expression analysis showed that M. tuberculosis efflux genes were overexpressed in response to antibiotic exposure, in vitro and within macrophages, irrespective of their resistance pattern. These compounds displayed a rapid and high killing activity against M. tuberculosis, associated with a decrease in intracellular ATP levels demonstrating that the bactericidal action of the ion channel blockers against M. tuberculosis clinical strains is associated with their interference with energy metabolism. The compounds led to a decrease in the intracellular mycobacterial load by increasing phagosome acidification and activating lysosomal hydrolases. The results presented in this study enable us to propose the following mechanism of action for these compounds: a) in the bacteria, the compounds generate a cascade of events involving the inhibition of the respiratory chain complexes and energy production for efflux activity. Indirectly, this reduce the resistance level to antituberculosis drugs potentiating their activity; b) on the host cell, the treatment with the ion channel blockers increases phagosome acidification and induces the expression of phagosomal hydrolases, leading to bacterial growth restriction irrespective of their

  3. Ion Channel Blockers as Antimicrobial Agents, Efflux Inhibitors, and Enhancers of Macrophage Killing Activity against Drug Resistant Mycobacterium tuberculosis.

    PubMed

    Machado, Diana; Pires, David; Perdigão, João; Couto, Isabel; Portugal, Isabel; Martins, Marta; Amaral, Leonard; Anes, Elsa; Viveiros, Miguel

    2016-01-01

    Given the ability of M. tuberculosis to survive as an intracellular pathogen and its propensity to develop resistance to the existing antituberculosis drugs, its treatment requires new approaches. Here the antimycobacterial properties of verapamil, thioridazine, chlorpromazine, flupenthixol and haloperidol were investigated against a panel of drug resistant M. tuberculosis strains, both in vitro and on human-infected macrophages. These compounds are efflux inhibitors that share among them the characteristic of being ion channel blockers. In vitro, all compounds exhibited synergistic inhibitory activities when combined with isoniazid and rifampicin, and were able to inhibit active efflux, demonstrating their role as efflux inhibitors. Gene expression analysis showed that M. tuberculosis efflux genes were overexpressed in response to antibiotic exposure, in vitro and within macrophages, irrespective of their resistance pattern. These compounds displayed a rapid and high killing activity against M. tuberculosis, associated with a decrease in intracellular ATP levels demonstrating that the bactericidal action of the ion channel blockers against M. tuberculosis clinical strains is associated with their interference with energy metabolism. The compounds led to a decrease in the intracellular mycobacterial load by increasing phagosome acidification and activating lysosomal hydrolases. The results presented in this study enable us to propose the following mechanism of action for these compounds: a) in the bacteria, the compounds generate a cascade of events involving the inhibition of the respiratory chain complexes and energy production for efflux activity. Indirectly, this reduce the resistance level to antituberculosis drugs potentiating their activity; b) on the host cell, the treatment with the ion channel blockers increases phagosome acidification and induces the expression of phagosomal hydrolases, leading to bacterial growth restriction irrespective of their

  4. Antibodies against invasive phenotype-specific antigens increase Mycobacterium avium subspecies paratuberculosis translocation across a polarized epithelial cell model and enhance killing by bovine macrophages.

    PubMed

    Everman, Jamie L; Bermudez, Luiz E

    2015-01-01

    Johne's disease, caused by Mycobacterium avium subspecies paratuberculosis (MAP), is a severe chronic enteritis which affects large populations of ruminants globally. Prevention strategies to combat the spread of Johne's disease among cattle herds involve adhering to strict calving practices to ensure young susceptible animals do not come in contact with MAP-contaminated colostrum, milk, or fecal material. Unfortunately, the current vaccination options available are associated with high cost and suboptimal efficacy. To more successfully combat the spread of Johne's disease to young calves, an efficient method of protection is needed. In this study, we examined passive immunization as a mode of introducing protective antibodies against MAP to prevent the passage of the bacterium to young animals via colostrum and milk. Utilizing the infectious MAP phenotype developed after bacterial exposure to milk, we demonstrate that in vitro opsonization with serum from Johne's-positive cattle results in enhanced translocation across a bovine MDBK polarized epithelial cell monolayer. Furthermore, immune serum opsonization of MAP results in a rapid host cell-mediated killing by bovine macrophages in an oxidative-, nitrosative-, and extracellular DNA trap-independent manner. This study illustrates that antibody opsonization of MAP expressing an infectious phenotype leads to the killing of the bacterium during the initial stage of macrophage infection.

  5. Antibodies against invasive phenotype-specific antigens increase Mycobacterium avium subspecies paratuberculosis translocation across a polarized epithelial cell model and enhance killing by bovine macrophages

    PubMed Central

    Everman, Jamie L.; Bermudez, Luiz E.

    2015-01-01

    Johne's disease, caused by Mycobacterium avium subspecies paratuberculosis (MAP), is a severe chronic enteritis which affects large populations of ruminants globally. Prevention strategies to combat the spread of Johne's disease among cattle herds involve adhering to strict calving practices to ensure young susceptible animals do not come in contact with MAP-contaminated colostrum, milk, or fecal material. Unfortunately, the current vaccination options available are associated with high cost and suboptimal efficacy. To more successfully combat the spread of Johne's disease to young calves, an efficient method of protection is needed. In this study, we examined passive immunization as a mode of introducing protective antibodies against MAP to prevent the passage of the bacterium to young animals via colostrum and milk. Utilizing the infectious MAP phenotype developed after bacterial exposure to milk, we demonstrate that in vitro opsonization with serum from Johne's-positive cattle results in enhanced translocation across a bovine MDBK polarized epithelial cell monolayer. Furthermore, immune serum opsonization of MAP results in a rapid host cell-mediated killing by bovine macrophages in an oxidative-, nitrosative-, and extracellular DNA trap-independent manner. This study illustrates that antibody opsonization of MAP expressing an infectious phenotype leads to the killing of the bacterium during the initial stage of macrophage infection. PMID:26301206

  6. Antibodies against invasive phenotype-specific antigens increase Mycobacterium avium subspecies paratuberculosis translocation across a polarized epithelial cell model and enhance killing by bovine macrophages.

    PubMed

    Everman, Jamie L; Bermudez, Luiz E

    2015-01-01

    Johne's disease, caused by Mycobacterium avium subspecies paratuberculosis (MAP), is a severe chronic enteritis which affects large populations of ruminants globally. Prevention strategies to combat the spread of Johne's disease among cattle herds involve adhering to strict calving practices to ensure young susceptible animals do not come in contact with MAP-contaminated colostrum, milk, or fecal material. Unfortunately, the current vaccination options available are associated with high cost and suboptimal efficacy. To more successfully combat the spread of Johne's disease to young calves, an efficient method of protection is needed. In this study, we examined passive immunization as a mode of introducing protective antibodies against MAP to prevent the passage of the bacterium to young animals via colostrum and milk. Utilizing the infectious MAP phenotype developed after bacterial exposure to milk, we demonstrate that in vitro opsonization with serum from Johne's-positive cattle results in enhanced translocation across a bovine MDBK polarized epithelial cell monolayer. Furthermore, immune serum opsonization of MAP results in a rapid host cell-mediated killing by bovine macrophages in an oxidative-, nitrosative-, and extracellular DNA trap-independent manner. This study illustrates that antibody opsonization of MAP expressing an infectious phenotype leads to the killing of the bacterium during the initial stage of macrophage infection. PMID:26301206

  7. Hypercapnia Inhibits Autophagy and Bacterial Killing in Human Macrophages by Increasing Expression of Bcl-2 and Bcl-xL

    PubMed Central

    Casalino-Matsuda, S. Marina; Nair, Aisha; Beitel, Greg J.; Gates, Khalilah L.; Sporn, Peter H. S.

    2015-01-01

    Hypercapnia, the elevation of CO2 in blood and tissue, commonly develops in patients with advanced lung disease and severe pulmonary infections, and is associated with high mortality. We previously reported that hypercapnia alters expression of host defense genes, inhibits phagocytosis, and increases the mortality of Pseudomonas pneumonia in mice. However, the effect of hypercapnia on autophagy, a conserved process by which cells sequester and degrade proteins and damaged organelles that also plays a key role in antimicrobial host defense and pathogen clearance, has not previously been examined. In the present study we show that hypercapnia inhibits autophagy induced by starvation, rapamycin, LPS, heat-killed and live bacteria in the human macrophage. Inhibition of autophagy by elevated CO2 was not attributable to acidosis. Hypercapnia also reduced macrophage killing of Pseudomonas aeruginosa. Moreover, elevated CO2 induced the expression of Bcl-2 and Bcl-xL, anti-apoptotic factors that negatively regulate autophagy by blocking Beclin 1, an essential component of the autophagy initiation complex. Furthermore, siRNA targeting Bcl-2 and Bcl-xL and the small molecule Z36, which blocks Bcl-2 and Bcl-xL binding to Beclin 1, prevented hypercapnic inhibition of autophagy and bacterial killing. These results suggest that targeting the Bcl-2/Bcl-xL-Beclin 1 interaction may hold promise for ameliorating hypercapnia-induced immunosuppression and improving resistance to infection in patients with advanced lung disease and hypercapnia. PMID:25895534

  8. Bactericidal activity of juvenile chinook salmon macrophages against Aeromonas salmonicida after exposure to live or heat-killed Renibacterium salmoninarum or to soluble proteins produced by R. salmoninarum

    USGS Publications Warehouse

    Siegel, D.C.; Congleton, J.L.

    1997-01-01

    Macrophages isolated from the anterior kidney of juvenile chinook salmon Oncorhynchus tshawytscha in 96-well microtiter plates were exposed for 72 h to 0, 105, or 106 live or heat-killed Renibacterium salmoninarum cells per well or to 0, 0.1, 1.0, or 10 ??g/mL of R. salmoninarum soluble proteins. After treatment, the bactericidal activity of the macrophages against Aerornonas salmonicida was determined by a colorimetric assay based on the reduction of the tetrazolium dye MTT to formazan by viable bacteria. The MTT assay was modified to allow estimation of the percentage of bacteria killed by reference to a standard curve relating the number of bacteria added to microtiter wells to absorbance by formazan at 600 nm. The live and heat-killed R. salmoninarum treatments significantly (P < 0.001) increased killing of A. salmonicida by chinook salmon macrophages. In each of the five trials, significantly (P < 0.05) greater increases in killing occurred after exposure to 105 R. salmoninarum cells than to 106 R. salmoninarum cells per well. In contrast, treatment of macrophages with 10 ??g/mL R. salmoninarum soluble proteins significantly (P < 0.001) decreased killing of A. salmonicida, but treatment with lower doses did not. These results show that the bactericidal activity of chinook salmon macrophages is stimulated by exposure to R. salmoninarum cells at lower dose levels but inhibited by exposure to R. salmoninarum cells or soluble proteins at higher dose levels.

  9. Role of intracellular free calcium in killing Penicillium marneffei within human macrophages.

    PubMed

    Chen, Renqiong; Ji, Guangquan; Ma, Tuan; Huang, Xiaowen; Ren, Hong; Xi, Liyan

    2015-01-01

    Increases in cytosolic Ca(2+) concentration ([Ca(2+)]c) promote phagocyte antimicrobial responses. Here, we investigated macrophages stimulated by Penicillium marneffei (P. marneffei). [Ca(2+)]c was determined in macrophages loaded with the fluorescent calcium probe Fura 2/AM as they were stimulated by P. marneffei. We found that P. marneffei induced an increase in [Ca(2+)]c in human macrophages. Further, increased [Ca(2+)]c with the ionophore A23187 promoted phagosomal acidification and maturation and reduced intracellular replication of P. marneffei in P. marneffei-infected human macrophages, whereas decreased [Ca(2+)]c with the chelation MAPTAM decreased TNF-α production, inhibited phagosomal acidification and maturation and increased intracellular replication of P. marneffei. These data indicate that Ca(2+) signaling may play an important role in controlling the replication of P. marneffei within macrophages.

  10. Killing of Leishmania parasites in activated murine macrophages is based on an L-arginine-dependent process that produces nitrogen derivatives

    SciTech Connect

    Maul, J.R.; Ransijn, A.; Buchmueller-Rouiller, Y. )

    1991-01-01

    The experiments described in this report were aimed at determining whether L-arginine (L-arg)-derived nitrogen oxidation products (nitric oxide, nitrous acid, nitrites) are involved in the intracellular killing of Leishmania parasites by activated murine macrophages in vitro. Peritoneal or bone marrow-derived macrophages were infected with L. enriettii or L. major, then activated by exposure to recombinant murine interferon-gamma or to macrophage activating factor (MAF)-rich media in the presence of lipopolysaccharide. Activation of macrophages in regular (i.e., arginine-containing) culture medium led to complete destruction of the microorganisms within 24 h (L. enriettii) or 48 h (L. major), concomitant with accumulation of nitrites (NO2-) in the culture fluids. When macrophage activation was carried out in L-arg-free medium, however, neither parasite killing nor NO2- production was obtained. A similar inhibition of macrophage leishmanicidal activity and of NO2- release was observed using media treated with arginase (which converts L-arg to urea and ornithine), or supplemented with NG-monomethyl-L-arg or guanidine (which inhibit the conversion of L-arg to nitrogen oxidation products). In all these situations, an excellent correlation between the levels of NO2- production by macrophages and intracellular killing of Leishmania was observed, whereas no strict correlation was detectable between leishmanicidal activity and superoxide production. Intracellular parasite killing by activated macrophages could be prevented by addition of iron salts to the incubation fluids. Incubation of free parasites with NaNO2 at acid pH led to immobilisation, multiplication arrest, and morphological degeneration of the microorganisms. Similarly, exposure of infected cells to NaNO2 led to killing of the intracellular parasite without affecting macrophage viability.

  11. Novel water-based antiseptic lotion demonstrates rapid, broad-spectrum kill compared with alcohol antiseptic.

    PubMed

    Czerwinski, Steven E; Cozean, Jesse; Cozean, Colette

    2014-01-01

    A novel alcohol-based antiseptic and a novel water-based antiseptic lotion, both with a synergistic combination of antimicrobial ingredients containing 0.2% benzethonium chloride, were evaluated using the standard time-kill method against 25 FDA-specified challenge microorganisms. The purpose of the testing was to determine whether a non-alcohol product could have equivalent rapid and broad-spectrum kill to a traditional alcohol sanitizer. Both the alcohol- and water-based products showed rapid and broad-spectrum antimicrobial activity. The average 15-s kill was 99.999% of the challenge organism for the alcohol-based antiseptic and 99.971% for the water-based antiseptic. The alcohol-based product demonstrated 100% of peak efficacy (60s) within the first 15s, whereas the water-based product showed 99.97%. The novel alcohol-based antiseptic reduced concentrations of 100% of organisms by 99.999%, whereas the water-based antiseptic lotion showed the same reduction for 96% of organisms. A novel water-based antiseptic product demonstrated equivalent rapid, broad-spectrum antimicrobial activity to an alcohol-based sanitizer and provided additional benefits of reduced irritation, persistent effect, and greater efficacy against common viruses. The combination of rapid, broad-spectrum immediate kill and persistent efficacy against pathogens may have significant clinical benefit in limiting the spread of disease.

  12. Live and Heat-Killed Lactobacillus rhamnosus ATCC 7469 May Induce Modulatory Cytokines Profiles on Macrophages RAW 264.7

    PubMed Central

    Jorjão, Adeline Lacerda; de Oliveira, Felipe Eduardo; Leão, Mariella Vieira Pereira; Carvalho, Cláudio Antonio Talge; Jorge, Antonio Olavo Cardoso; de Oliveira, Luciane Dias

    2015-01-01

    This study aimed to evaluate the capacity of Lactobacillus rhamnosus and/or its products to induce the synthesis of cytokines (TNF-α, IL-1β, IL-4, IL-6, IL-10, and IL-12) by mouse macrophages (RAW 264.7). Three microorganism preparations were used: live L. rhamnosus (LLR) suspension, heat-killed L. rhamnosus (HKLR) suspension, and the supernatant of a heat-killed L. rhamnosus (SHKLR) suspension, which were cultured with macrophages (37°C, 5% CO2) for 2 h and 30 min. After that, cells were cultured for 16 h. The supernatants were used for the quantitation of cytokines, by ELISA. The results were compared with the synthesis induced by lipopolysaccharide (LPS) and analysed, using ANOVA and Tukey test, 5%. LLR and HKLR groups were able to significantly increase the production of TNF-α, IL-6, and IL-10 (P < 0.05). SHKLR also significantly increased the production of TNF-α and IL-10 (P < 0.05) but not IL-6 (P > 0.05). All the L. rhamnosus suspensions were not able to produce detectable levels of IL-1β or significant levels of IL-4 and IL-12 (P > 0.05). In conclusion, live and heat-killed L. rhamnosus suspensions were able to induce the synthesis of different cytokines with proinflammatory (TNF-α and IL-6) or regulatory (IL-10) functions, suggesting the role of strain L. rhamnosus ATCC 7469 in the modulation or in the stimulation of immune responses. PMID:26649329

  13. Autophagic Killing Effects against Mycobacterium tuberculosis by Alveolar Macrophages from Young and Aged Rhesus Macaques

    PubMed Central

    Pacheco, Sophia A.; Powers, Katelyn M.; Engelmann, Flora; Messaoudi, Ilhem; Purdy, Georgiana E.

    2013-01-01

    Non-human primates, notably rhesus macaques (Macaca mulatta, RM), provide a robust experimental model to investigate the immune response to and effective control of Mycobacterium tuberculosis infections. Changes in the function of immune cells and immunosenescence may contribute to the increased susceptibility of the elderly to tuberculosis. The goal of this study was to examine the impact of age on M. tuberculosis host-pathogen interactions following infection of primary alveolar macrophages derived from young and aged rhesus macaques. Of specific interest to us was whether the mycobactericidal capacity of autophagic macrophages was reduced in older animals since decreased autophagosome formation and autophagolysosomal fusion has been observed in other cells types of aged animals. Our data demonstrate that alveolar macrophages from old RM are as competent as those from young animals for autophagic clearance of M. tuberculosis infection and controlling mycobacterial replication. While our data do not reveal significant differences between alveolar macrophage responses to M. tuberculosis by young and old animals, these studies are the first to functionally characterize autophagic clearance of M. tuberculosis by alveolar macrophages from RM. PMID:23825603

  14. Nerve growth factor promotes killing of Leishmania donovani by macrophages through the induction of hydrogen peroxide.

    PubMed

    Chiba, Rieko; Amagai, Yosuke; Tanaka, Akane; Katakura, Ken; Matsuda, Hiroshi

    2014-08-01

    Visceral leishmaniasis is protozoonosis that occurs worldwide and still requires effective therapies with less toxicity. In this study, we examined the antileishmanial effect of nerve growth factor (NGF) using a murine infection model. NGF blocked the infection of macrophages by Leishmania donovani, which was completely cancelled by a hydrogen peroxide inhibitor. In vivo, not only did NGF show antileishmanial effects, but combination therapy of NGF and sodium stibogluconate synergistically exhibited the activity more potently than each monotherapy. These results indicate that NGF exerts antileishmanial effect by stimulating hydrogen peroxide production in macrophages and can be a novel therapy for leishmaniasis.

  15. Chronic Iron Overload Results in Impaired Bacterial Killing of THP-1 Derived Macrophage through the Inhibition of Lysosomal Acidification

    PubMed Central

    Kao, Jun-Kai; Wang, Shih-Chung; Ho, Li-Wei; Huang, Shi-Wei; Chang, Shu-Hao; Yang, Rei-Cheng; Ke, Yu-Yuan; Wu, Chun-Ying; Wang, Jiu-Yao; Shieh, Jeng-Jer

    2016-01-01

    Iron is essential for living organisms and the disturbance of iron homeostasis is associated with altered immune function. Additionally, bacterial infections can cause major complications in instances of chronic iron overload, such as patients with transfusion-dependent thalassemia. Monocytes and macrophages play important roles in maintaining systemic iron homoeostasis and in defense against invading pathogens. However, the effect of iron overload on the function of monocytes and macrophages is unclear. We elucidated the effects of chronic iron overload on human monocytic cell line (THP-1) and THP-1 derived macrophages (TDM) by continuously exposing them to high levels of iron (100 μM) to create I-THP-1 and I-TDM, respectively. Our results show that iron overload did not affect morphology or granularity of I-THP-1, but increased the granularity of I-TDM. Bactericidal assays for non-pathogenic E. coli DH5α, JM109 and pathogenic P. aeruginosa all revealed decreased efficiency with increasing iron concentration in I-TDM. The impaired P. aeruginosa killing ability of human primary monocyte derived macrophages (hMDM) was also found when cells are cultured in iron contained medium. Further studies on the bactericidal activity of I-TDM revealed lysosomal dysfunction associated with the inhibition of lysosomal acidification resulting in increasing lysosomal pH, the impairment of post-translational processing of cathepsins (especially cathepsin D), and decreased autophagic flux. These findings may explain the impaired innate immunity of thalassemic patients with chronic iron overload, suggesting the manipulation of lysosomal function as a novel therapeutic approach. PMID:27244448

  16. Innate Immune Memory: Activation of Macrophage Killing Ability by Developmental Duties.

    PubMed

    Schneider, David; Tate, Ann Thomas

    2016-06-20

    Innate immune systems in many taxa exhibit hallmarks of memory in response to previous microbial exposure. A new study demonstrates that innate immune memory in Drosophila embryonic macrophages can also be induced by the successful engulfment of apoptotic cells, highlighting the importance of early exposure events for developing responsive immune systems.

  17. IgE Mediates Killing of Intracellular Toxoplasma gondii by Human Macrophages through CD23-Dependent, Interleukin-10 Sensitive Pathway

    PubMed Central

    Vouldoukis, Ioannis; Mazier, Dominique; Moynet, Daniel; Thiolat, Denis; Malvy, Denis; Mossalayi, M. Djavad

    2011-01-01

    Background In addition to helminthic infections, elevated serum IgE levels were observed in many protozoal infections, while their contribution during immune response to these pathogens remained unclear. As IgE/antigen immune complexes (IgE-IC) bind to human cells through FcεRI or FcεRII/CD23 surface molecules, the present study aimed to identify which functional receptor may be involved in IgE-IC interaction with human macrophages, the major effector cell during parasite infection. Methodology/Principal Findings Human monocyte-derived macrophages were infected with Toxoplasma gondii before being incubated with IgE-IC. IgE receptors were then identified using appropriate blocking antibodies. The activation of cells and parasiticidal activity were evaluated by mediator quantification and direct counting of infected macrophages. RNAs were extracted and cell supernatants were also collected for their content in tumor necrosis factor (TNF)-α, interleukin-10 (IL-10) and nitrites. Sera from symptomatic infected patients were also tested for their content of IgE, IL-10 and nitrites, and compared to values found in healthy donors. Results showed that IgE-IC induced intracellular elimination of parasites by human macrophages. IgE-mediated effect was FcεRI-independent, but required cross-linking of surface FcεRII/CD23, cell activation and the generation of nitric oxide (NO). Although TNF-α was shown to be produced during cell activation, this cytokine had minor contribution in this phenomenon while endogenous and exogenous IL-10 down-regulated parasite killing. Inverse relationship was found between IL-10 and NO expression by infected human macrophages at both mRNA and mediator levels. The relationship between these in vitro data and in vivo levels of various factors in T. gondii infected patients supports the involvement of CD23 antigen and IL-10 expression in disease control. Conclusion Thus, IgE may be considered as immune mediator during antiprotozoal activity of

  18. Role of protein kinase R in the killing of Leishmania major by macrophages in response to neutrophil elastase and TLR4 via TNFα and IFNβ.

    PubMed

    Faria, Marilia S; Calegari-Silva, Tereza C; de Carvalho Vivarini, Aislan; Mottram, Jeremy C; Lopes, Ulisses Gazos; Lima, Ana Paula C A

    2014-07-01

    In cutaneous leishmaniasis, Leishmania amazonensis activates macrophage double-stranded, RNA-activated protein kinase R (PKR) to promote parasite growth. In our study, Leishmania major grew normally in RAW cells, RAW-expressing dominant-negative PKR (PKR-DN) cells, and macrophages of PKR-knockout mice, revealing that PKR is dispensable for L. major growth in macrophages. PKR activation in infected macrophages with poly I:C resulted in parasite death. Fifty percent of L. major-knockout lines for the ecotin-like serine peptidase inhibitor (ISP2; Δisp2/isp3), an inhibitor of neutrophil elastase (NE), died in RAW cells or macrophages from 129Sv mice, as a result of PKR activation. Inhibition of PKR or NE or neutralization of Toll-like receptor 4 or 2(TLR4 or TLR2) prevented the death of Δisp2/isp3. Δisp2/isp3 grew normally in RAW-PKR-DN cells or macrophages from 129Sv pkr(-/-), tlr2(-/-), trif(-/-), and myd88(-/-) mice, associating NE activity, PKR, and TLR responses with parasite death. Δisp2/isp3 increased the expression of mRNA for TNF-α by 2-fold and of interferon β (IFNβ) in a PKR-dependent manner. Antibodies to TNF-α reversed the 95% killing by Δisp2/isp3, whereas they grew normally in macrophages from IFN receptor-knockout mice. We propose that ISP2 prevents the activation of PKR via an NE-TLR4-TLR2 axis to control innate responses that contribute to the killing of L. major.-Faria, M. S., Calegari-Silva, T. C., de Carvalho Vivarini, A., Mottram, J. C., Lopes, U. G., Lima, A. P. C. A. Role of protein kinase R in the killing of Leishmania major by macrophages in response to neutrophil elastase and TLR4 via TNFα and IFNβ. PMID:24732131

  19. SseL, a Salmonella deubiquitinase required for macrophage killing and virulence

    PubMed Central

    Rytkönen, Anne; Poh, John; Garmendia, Junkal; Boyle, Cliona; Thompson, Arthur; Liu, Mei; Freemont, Paul; Hinton, Jay C. D.; Holden, David W.

    2007-01-01

    Expression of the Salmonella enterica serovar Typhimurium pathogenicity island 2 (SPI-2) type III secretion system is controlled by the two-component regulatory system SsrA-SsrB. We used a transcriptomic approach to help define the SsrA-SsrB regulon. We identified a gene encoding an uncharacterized effector (SseL) whose translocation into host cells depends on the SPI-2 secretion system. SseL has similarities to cysteine proteases with deubiquitinating activity. A GST-SseL fusion protein specifically hydrolyzed mono- and polyubiquitin substrates in vitro with a preference for K63-linked ubiquitin chains. Ubiquitin-modified proteins accumulated in macrophages infected with Salmonella sseL mutant strains but to a lesser extent when infected with bacteria expressing active protein, demonstrating that SseL functions as a deubiquitinase in vivo. Salmonella sseL mutant strains did not show a replication defect or induce altered levels of cytokine production upon infection of macrophages but were defective for a delayed cytotoxic effect and were attenuated for virulence in mice. PMID:17360673

  20. Enhancement of macrophage candidacidal activity by interferon-gamma. Increased phagocytosis, killing, and calcium signal mediated by a decreased number of mannose receptors.

    PubMed Central

    Maródi, L; Schreiber, S; Anderson, D C; MacDermott, R P; Korchak, H M; Johnston, R B

    1993-01-01

    In contrast to its macrophage-activating capacity, IFN-gamma downregulates expression of the macrophage mannose receptor (MMR), which mediates uptake of Candida and other microorganisms. We found that IFN-gamma induced a concentration-dependent increase in the capacity of human monocyte-derived macrophages to ingest and kill both opsonized and unopsonized Candida albicans and to release superoxide anion upon stimulation with Candida. Mannan or mannosylated albumin inhibited this activated uptake of unopsonized Candida, but glucan did not. Addition of mAb to complement receptor (CR) 3 did not inhibit ingestion; macrophages that lacked CR3 (leukocyte adhesion defect) showed normal upregulation of ingestion by IFN-gamma. The increased candidacidal activity of IFN-gamma-activated macrophages was associated with reduced expression of MMR by a mean of 79% and decreased pinocytic uptake of 125I-mannosylated BSA by 73%; K(uptake) of pinocytosis was not changed. Exposure of resident macrophages to unopsonized Candida did not elicit a transient increase in intracellular free Ca2+ ([Ca2+]i); macrophages activated by IFN-gamma expressed a brisk increase in [Ca2+]i on exposure to Candida. These data suggest that macrophage activation by IFN-gamma can enhance resistance to C. albicans infection in spite of downregulation of the MMR, perhaps through enhanced coupling of the MMR to microbicidal functions. PMID:8390485

  1. Small heat-shock proteins, IbpAB, protect non-pathogenic Escherichia coli from killing by macrophage-derived reactive oxygen species.

    PubMed

    Goeser, Laura; Fan, Ting-Jia; Tchaptchet, Sandrine; Stasulli, Nikolas; Goldman, William E; Sartor, R Balfour; Hansen, Jonathan J

    2015-01-01

    Many intracellular bacterial pathogens possess virulence factors that prevent detection and killing by macrophages. However, similar virulence factors in non-pathogenic bacteria are less well-characterized and may contribute to the pathogenesis of chronic inflammatory conditions such as Crohn's disease. We hypothesize that the small heat shock proteins IbpAB, which have previously been shown to reduce oxidative damage to proteins in vitro and be upregulated in luminal non-pathogenic Escherichia strain NC101 during experimental colitis in vivo, protect commensal E. coli from killing by macrophage-derived reactive oxygen species (ROS). Using real-time PCR, we measured ibpAB expression in commensal E. coli NC101 within wild-type (wt) and ROS-deficient (gp91phox(-/-)) macrophages and in NC101 treated with the ROS generator paraquat. We also quantified survival of NC101 and isogenic mutants in wt and gp91phox(-/-) macrophages using gentamicin protection assays. Similar assays were performed using a pathogenic E. coli strain O157:H7. We show that non-pathogenic E. coli NC101inside macrophages upregulate ibpAB within 2 hrs of phagocytosis in a ROS-dependent manner and that ibpAB protect E. coli from killing by macrophage-derived ROS. Moreover, we demonstrate that ROS-induced ibpAB expression is mediated by the small E. coli regulatory RNA, oxyS. IbpAB are not upregulated in pathogenic E. coli O157:H7 and do not affect its survival within macrophages. Together, these findings indicate that ibpAB may be novel virulence factors for certain non-pathogenic E. coli strains. PMID:25798870

  2. Kinetics of killing Listeria monocytogenes by macrophages: correlation of /sup 3/H-DNA release from labeled bacteria and changes in numbers of viable organisms by mathematical model

    SciTech Connect

    Davies, W.A.

    1982-12-01

    Conventional methods of assessing antibacterial activities of macrophages by viable counting are limited by the precision of the statistics and are difficult to interpret quantitatively because of unrestrained extracellular growth of bacteria. An alternative technique based on the release of radioactive DNA from labeled bacteria has been offered as overcoming these drawbacks. To assess it for use with macrophages I have made a correlation with the conventional viable counting method using a mathematical model. Opsonized Listeria monocytogenes labeled with /sup 3/H-thymidine were exposed to rat macrophages for periods up to 4 hr. Numbers of viable bacteria determined after sonication increased exponentially in the absence of live cells and this growth rate was progressively inhibited by increasing numbers of macrophages. After a lag period of 30-60 min soluble /sup 3/H appeared in the supernatant, the amount increasing with time and numbers of macrophages. To correlate these data I developed a mathematical model that considered that changes in numbers of viable organisms were due to the difference between rates of 1) growth of extracellular bacteria and 2) killing within the macrophage. On the basis of this model curves of best fit to the viable counts data were used to predict the release of radioactivity, assuming that death of a bacterium led to the total release of its label. These predictions and the experimental data agreed well, the lag period of 30-60 min between death of the bacterium and release of radioactivity being consistent with intracellular digestion. Release of soluble radioactivity appears to be an accurate reflection of the number of bacteria killed within the macrophage.

  3. Extracellular killing of inhaled pneumococci in rats

    SciTech Connect

    Coonrod, J.D.; Marple, S.; Holmes, G.P.; Rehm, S.R.

    1987-12-01

    Early clearance of inhaled Staphylococcus aureus is believed to be caused by phagocytosis by alveolar macrophages. In murine models inhaled pneumococci are cleared even more rapidly than S. aureus. Conventional opsonins appear to play no role in this clearance, and recently it has been shown that murine alveolar lining material contains free fatty acids and other soluble factors that are directly bactericidal for pneumococci. To determine whether non-phagocytic factors are involved in pneumococcal clearance, we compared the site of killing of inhaled pneumococci and S. aureus in rats using histologic methods and bronchoalveolar lavage. Spontaneous lysis of pneumococci was prevented by use of autolysin-defective pneumococci or by substitution of ethanolamine for choline in the cell wall. Histologic studies showed that the percent of inhaled staphylococci associated with alveolar macrophages always exceeded the percent of staphylococci cleared, whereas there was little association of pneumococci with macrophages during clearance. Analysis of the intracellular or extracellular location of iron 59 in bronchoalveolar lavage fluid of rats that had inhaled aerosols of /sup 59/Fe-labeled bacteria suggested that staphylococci were killed predominantly in macrophages and pneumococci in the extracellular space. When /sup 59/Fe-labeled pneumococci or staphylococci were ingested and killed by macrophages in vitro, the /sup 59/Fe remained with the macrophages, suggesting that the extracellular location of /sup 59/Fe during pneumococcal killing in vivo was not caused by rapid turnover of /sup 59/Fe in macrophages. Studies of the site of killing of inhaled type 25 pneumococci labeled exclusively in the cell wall with carbon 14-ethanolamine confirmed the results obtained with /sup 59/Fe-labeled pneumococci. Thus, early killing of inhaled pneumococci, unlike staphylococci, appears to take place outside of macrophages.

  4. Rapid recruitment of macrophages in interleukin-12-mediated tumour regression.

    PubMed Central

    Ha, S J; Lee, S B; Kim, C M; Shin, H S; Sung, Y C

    1998-01-01

    In order to study the mechanism of interleukin-12 (IL-12) antitumour activity, RH7777 rat hepatoma cells were engineered to express mouse IL-12 (mIL-12) (RH7777/mIL-12) under the tight control of doxycycline (dox). The production of the mIL-12 protein was regulated by the concentration of dox that was present in the culture medium. RH7777/mIL-12 cells appeared to have the same tumorigenic activity as did parental RH7777 cells, when subcutaneously injected into syngeneic rat (BUF/N) in the absence of dox. However, the tumorigenicity of RH7777/mIL-12, but not RH7777, cells were significantly decreased when dox was administrated to the animals. In addition, established tumours of RH7777/mIL-12 cells gradually disappeared upon the induction of mIL-12 by dox. To elucidate the kinetic profile of immune cells involved in the mIL-12-induced tumour regression, both histological and immunohistochemical analyses were performed 1, 3 and 14 days after the dox treatment on rats bearing tumours that were approximately 0. 5 cm in diameter. Tumour-infiltrating macrophages began to appear at the tumour site one day after dox treatment. As time elapsed, the number of tumour infiltrates including CD4+, CD8+, natural killer (NK) cells and macrophages gradually increased. In particular, CD8+ and NK cells constituted the major population of the tumour-infiltrated cells. Furthermore, it was found that resting peritoneal macrophages (PM) from rats were chemoattracted in response to mIL-12. The effects of mIL-12 on PM chemotaxis were reproducibly observed in concentrations as low as 0.1 ng/ml. These findings suggest that IL-12 can directly recruit macrophages into tumour sites which, in turn, leads to a broad and intense immunological response against tumour. Images Figure 3 Figure 4 PMID:9767471

  5. Treatment of murine macrophages with murine interferon-gamma and tumour necrosis factor-alpha enhances uptake and intracellular killing of Pseudomonas aeruginosa.

    PubMed Central

    Pierangeli, S S; Sonnenfeld, G

    1993-01-01

    Murine interferon-gamma (MuIFN-gamma) and murine tumour necrosis-alpha (MuTNF-alpha) are known to be potent immunomodulators of several aspects of the immune response, and they have been shown to exert profound effects on macrophages and monocytes. The purpose of this study was to determine the effects of MuIFN-gamma and MuTNF-alpha on the phagocytosis (uptake and intracellular killing) of opsonized Pseudomonas aeruginosa. Unstimulated peritoneal macrophages obtained from CBA/c mice were exposed to different concentrations of recombinant forms of the cytokines (rMuIFN-gamma and rMuTNF-alpha) for different periods of time. Phagocytosis was assayed using different concentrations of opsonized Ps. aeruginosa. In all cases the pretreatment of the cells with the cytokines increased significantly the uptake and the intracellular killing of bacteria in a dose-dependent manner. rMuTNF-alpha was effective only at 1000 U/ml. Combined treatment with the cytokines showed a less than additive effect with rMuIFN-gamma and rMuTNF-alpha at concentrations of 10 U/ml and 100 U/ml. In the in vivo experiments, peritoneal macrophages obtained from rMuIFN-gamma- or rMuTNF-alpha-treated mice showed enhancement of the intracellular killing of opsonized bacteria in a dose-dependent manner. PMID:8348741

  6. Gold Nanopopcorn Attached Single-Walled Carbon Nanotube Hybrid for Rapid Detection and Killing of Bacteria

    PubMed Central

    Ondera, Thomas. J.

    2014-01-01

    We report a strategy to fabricate a rapid and stable surface-enhanced Raman scattering (SERS)-based hybrid nanomaterial using gold nanopopcorns attached single-walled carbon nanotubes (AuNP@f3-SWCNTs) for label-free detection and photothermal killing of bacteria. Herein, previously ester-functionalized single-walled carbon nanotubes (f1-SWCNTs) undergo 1,3-dipolar cycloaddition reaction with in-situ generated nitrile imine under Microwave (MW) irradiation to form a doubly ester terminated SWCNTs cycloadduct (f2-SWCNTs). The ester terminals are further modified with 4-aminothiophenol (4-ATP) under MW-irradiation to form thiol-terminated SWCNTs templates (f3-SWCNTs) that allow gold nanopopcorns (AuNPs) to covalently and uniformly attach at a minimum inter-particle distance thus yielding a hybrid nanomaterial (AuNP@f3-SWCNT) with good aqueous stability and abundant ‘hotspots’. Consequently, monoclonal E. coli antibody-conjugated bioassays fabricated with our AuNP@f3-SWCNT substrates (mAb-AuNP@f3-SWCNT) rapidly detect E. coli in water with good selectivity and impressive SERS sensitivity. The detection limit of E. coli 49979, selected as a model to establish proof of principle, was found to be 1.0×102 CFU/mL. Furthermore, the AuNP@f3-SWCNT hybrid nanomaterial offers impressive photothermal pathogen killing effects. The synergy-type enhancement effect arising from the inherent noble properties of the respective components of the hybrid nanomaterial indicate that our AuNP@f3-SWCNT has the potential for further application in multiplex detection in samples. PMID:25414794

  7. Bovine SLC11A1 3' UTR SSCP genotype evaluated by a macrophage in vitro killing assay employing a Brucella abortus strain.

    PubMed

    Martinez, R; Toro, R; Montoya, F; Burbano, M; Tobón, J; Gallego, J; Dunner, S; Cañón, J

    2008-08-01

    The 3' untranslated region (3' UTR) of the bovine natural resistance-associated macrophage gene (NRAMP1 or SLC11A1) was genotyped in Colombian Creole Blanco Orejinegro (BON) (Bos taurus) (n = 140) and Zebu Brahman (Bos indicus) (Z) (n = 20) cattle and their crosses (BON x Zebu Brahman [B x Z] [n = 10]; Zebu Brahman x BON [Z x B] [n = 10]), and in animals from a Holstein x BON (H x B) (n = 10) cross. Direct sequencing and single-strand conformation polymorphism analysis (SSCP) helped in detecting the polymorphic behaviour. The association between resistance to brucellosis infection and SSCP genotype was evaluated using a macrophage in vitro killing assay employing a virulent Brucella abortus strain. The 3' UTR (GT) repeated polymorphism was gentoyped and its association with resistance to brucellosis was evaluated. When all breeds were grouped, a high frequency in the homozygote GT(12) (AA genotype) (0.823) and a very low frequency in the homozygote GT(10) (BB genotype) (0.047) were detected. The BON (0.963), Z x B (0.60) and H x B (1.00) cattle showed high GT(12) allele frequencies, unlike that seen for the B x Z and Zebu cattle (0.3002 and 0.218, respectively). The GT(10) allele was only found in the Zebu cattle (0.391). A significant association (p < 0.001) was found between the B. abortus macrophage in vitro killing assay phenotypes and the bovine SLC11A1 3' UTR genotypes, which suggests that the A allele may be associated with resistance. Because only nine animals had the BB genotype, the results require some confirmation in more extensive populations.

  8. Bovine SLC11A1 3' UTR SSCP genotype evaluated by a macrophage in vitro killing assay employing a Brucella abortus strain.

    PubMed

    Martinez, R; Toro, R; Montoya, F; Burbano, M; Tobón, J; Gallego, J; Dunner, S; Cañón, J

    2008-08-01

    The 3' untranslated region (3' UTR) of the bovine natural resistance-associated macrophage gene (NRAMP1 or SLC11A1) was genotyped in Colombian Creole Blanco Orejinegro (BON) (Bos taurus) (n = 140) and Zebu Brahman (Bos indicus) (Z) (n = 20) cattle and their crosses (BON x Zebu Brahman [B x Z] [n = 10]; Zebu Brahman x BON [Z x B] [n = 10]), and in animals from a Holstein x BON (H x B) (n = 10) cross. Direct sequencing and single-strand conformation polymorphism analysis (SSCP) helped in detecting the polymorphic behaviour. The association between resistance to brucellosis infection and SSCP genotype was evaluated using a macrophage in vitro killing assay employing a virulent Brucella abortus strain. The 3' UTR (GT) repeated polymorphism was gentoyped and its association with resistance to brucellosis was evaluated. When all breeds were grouped, a high frequency in the homozygote GT(12) (AA genotype) (0.823) and a very low frequency in the homozygote GT(10) (BB genotype) (0.047) were detected. The BON (0.963), Z x B (0.60) and H x B (1.00) cattle showed high GT(12) allele frequencies, unlike that seen for the B x Z and Zebu cattle (0.3002 and 0.218, respectively). The GT(10) allele was only found in the Zebu cattle (0.391). A significant association (p < 0.001) was found between the B. abortus macrophage in vitro killing assay phenotypes and the bovine SLC11A1 3' UTR genotypes, which suggests that the A allele may be associated with resistance. Because only nine animals had the BB genotype, the results require some confirmation in more extensive populations. PMID:18717968

  9. Toll-like receptor 5 (TLR5), IL-1β secretion, and asparagine endopeptidase are critical factors for alveolar macrophage phagocytosis and bacterial killing.

    PubMed

    Descamps, Delphyne; Le Gars, Mathieu; Balloy, Viviane; Barbier, Diane; Maschalidi, Sophia; Tohme, Mira; Chignard, Michel; Ramphal, Reuben; Manoury, Bénédicte; Sallenave, Jean-Michel

    2012-01-31

    A deficit in early clearance of Pseudomonas aeruginosa (P. aeruginosa) is crucial in nosocomial pneumonia and in chronic lung infections. Few studies have addressed the role of Toll-like receptors (TLRs), which are early pathogen associated molecular pattern receptors, in pathogen uptake and clearance by alveolar macrophages (AMs). Here, we report that TLR5 engagement is crucial for bacterial clearance by AMs in vitro and in vivo because unflagellated P. aeruginosa or different mutants defective in TLR5 activation were resistant to AM phagocytosis and killing. In addition, the clearance of PAK (a wild-type P. aeruginosa strain) by primary AMs was causally associated with increased IL-1β release, which was dramatically reduced with PAK mutants or in WT PAK-infected primary TLR5(-/-) AMs, demonstrating the dependence of IL-1β production on TLR5. We showed that this IL-1β production was important in endosomal pH acidification and in inducing the killing of bacteria by AMs through asparagine endopeptidase (AEP), a key endosomal cysteine protease. In agreement, AMs from IL-1R1(-/-) and AEP(-/-) mice were unable to kill P. aeruginosa. Altogether, these findings demonstrate that TLR5 engagement plays a major role in P. aeruginosa internalization and in triggering IL-1β formation. PMID:22307620

  10. Toll-like receptor 5 (TLR5), IL-1β secretion, and asparagine endopeptidase are critical factors for alveolar macrophage phagocytosis and bacterial killing

    PubMed Central

    Descamps, Delphyne; Le Gars, Mathieu; Balloy, Viviane; Barbier, Diane; Maschalidi, Sophia; Tohme, Mira; Chignard, Michel; Ramphal, Reuben; Manoury, Bénédicte; Sallenave, Jean-Michel

    2012-01-01

    A deficit in early clearance of Pseudomonas aeruginosa (P. aeruginosa) is crucial in nosocomial pneumonia and in chronic lung infections. Few studies have addressed the role of Toll-like receptors (TLRs), which are early pathogen associated molecular pattern receptors, in pathogen uptake and clearance by alveolar macrophages (AMs). Here, we report that TLR5 engagement is crucial for bacterial clearance by AMs in vitro and in vivo because unflagellated P. aeruginosa or different mutants defective in TLR5 activation were resistant to AM phagocytosis and killing. In addition, the clearance of PAK (a wild-type P. aeruginosa strain) by primary AMs was causally associated with increased IL-1β release, which was dramatically reduced with PAK mutants or in WT PAK-infected primary TLR5−/− AMs, demonstrating the dependence of IL-1β production on TLR5. We showed that this IL-1β production was important in endosomal pH acidification and in inducing the killing of bacteria by AMs through asparagine endopeptidase (AEP), a key endosomal cysteine protease. In agreement, AMs from IL-1R1−/− and AEP−/− mice were unable to kill P. aeruginosa. Altogether, these findings demonstrate that TLR5 engagement plays a major role in P. aeruginosa internalization and in triggering IL-1β formation. PMID:22307620

  11. Evaluation of methods of rapid mass killing of segregated early weaned piglets

    PubMed Central

    Whiting, Terry L.; Steele, Gregory G.; Wamnes, Steinar; Green, Chris

    2011-01-01

    The operational logistics of mass killing of healthy, surplus piglets by manual blunt force trauma, controlled blunt force trauma, intraperitoneal injection of barbiturate, and free bullet were recorded. Objective performance variables evaluated were, speed of application, human resource and input cost, animal restraint required, and failure rate. Subjective evaluation of esthetics and difficulty of application indicated manual blunt force trauma is an unacceptable technique. Under field conditions, physical methods of killing were superior to intraperitoneal injection of concentrated pentobarbital. Considering animal welfare metrics in isolation, controlled blunt force trauma was superior to all other techniques attempted. PMID:22210939

  12. Short communication: rapid preparation of preventive and therapeutic whole-killed retroviral vaccines using the microbicide taurine chloramine.

    PubMed

    Dudani, A K; Martyres, A; Fliss, H

    2008-04-01

    A current urgent priority is to develop microbicides and vaccines to combat retroviruses like human immunodeficiency virus (HIV). We show that the cysteine-selective natural compound, taurine chloramine (T-NCl), can be effective in this task. A number of proteins in all retroviruses contain highly conserved cysteine-rich regions that are essential for infection and replication. Our data show that by targeting these essential cysteine residues, T-NCl (2 or 5 mM) acts as a highly effective and safe microbicide that fully blocks the infectivity of high HIV-1 titers (10(6) TCID(50) units/ml) but is not injurious to eukaryotic cells. We also demonstrate that T-NCl can be used to prepare a highly effective whole-killed vaccine against murine AIDS (MAIDS) that shows both preventive and therapeutic efficacy. The vaccine consists of a T-NCl-inactivated retrovirus suspension in host cell lysate. The novelty of our approach lies in the ease and speed of vaccine preparation and its avoidance of harsh inactivation or purification steps that can alter native viral conformation. Our approach is therefore likely to overcome a number of intractable obstacles to the preparation of an effective whole-killed HIV vaccine, such as surviving infective viral particles, rapid viral mutation rates, numerous viral strains, and harsh purification steps. Our approach may also permit the rapid preparation of autologous, or custom-made, vaccines for individual patients.

  13. Rapid and efficient cancer cell killing mediated by high-affinity death receptor homotrimerizing TRAIL variants

    PubMed Central

    Reis, C R; van der Sloot, A M; Natoni, A; Szegezdi, E; Setroikromo, R; Meijer, M; Sjollema, K; Stricher, F; Cool, R H; Samali, A; Serrano, L; Quax, W J

    2010-01-01

    The tumour necrosis factor family member TNF-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in a variety of cancer cells through the activation of death receptors 4 (DR4) and 5 (DR5) and is considered a promising anticancer therapeutic agent. As apoptosis seems to occur primarily via only one of the two death receptors in many cancer cells, the introduction of DR selectivity is thought to create more potent TRAIL agonists with superior therapeutic properties. By use of a computer-aided structure-based design followed by rational combination of mutations, we obtained variants that signal exclusively via DR4. Besides an enhanced selectivity, these TRAIL-DR4 agonists show superior affinity to DR4, and a high apoptosis-inducing activity against several TRAIL-sensitive and -resistant cancer cell lines in vitro. Intriguingly, combined treatment of the DR4-selective variant and a DR5-selective TRAIL variant in cancer cell lines signalling by both death receptors leads to a significant increase in activity when compared with wild-type rhTRAIL or each single rhTRAIL variant. Our results suggest that TRAIL induced apoptosis via high-affinity and rapid-selective homotrimerization of each DR represent an important step towards an efficient cancer treatment. PMID:21368856

  14. Selective delivery of rifampicin incorporated into poly(DL-lactic-co-glycolic) acid microspheres after phagocytotic uptake by alveolar macrophages, and the killing effect against intracellular Mycobacterium bovis Calmette-Guérin.

    PubMed

    Yoshida, Aya; Matumoto, Makoto; Hshizume, Hiroyuki; Oba, Yoshiro; Tomishige, Tatuo; Inagawa, Hiroyuki; Kohchi, Chie; Hino, Mami; Ito, Fuminori; Tomoda, Keishiro; Nakajima, Takehisa; Makino, Kimiko; Terada, Hiroshi; Hori, Hitoshi; Soma, Gen-Ichiro

    2006-08-01

    Macrophages and their phagocytotic abilities play a dominant role for defense against infected organisms. However, Mycobacterium tuberculosis can survive in the phagosomes of macrophages. In this study, the effective delivery of a drug and the killing effect of tubercle bacilli within macrophages were investigated utilizing the phagocytotic uptake of rifampicin (RFP) that had been incorporated into poly(DL-lactic-co-glycolic) acid (PLGA) microspheres. The microspheres were composed of PLGA that had a monomer ratio (lactic acid/glycolic acid) of either 50/50 or 75/25. They had molecular weights from 5000 to 20,000, and diameters of 1.5, 3.5, 6.2 and 8.9 microm. The most significant factor for phagocytotic activity of macrophages was the diameter of the microspheres. By contrast, molecular weight and monomer ratio of PLGA did not influence phagocytosis. The amount of RFP delivered into cells was also investigated. RFP-PLGA microspheres composed of PLGA with a molecular weight of 20,000 and monomer ratio of 75/25 showed the highest amount of delivery (4 microg/1 x 10(6) cells). Fourteen days after infection, the survival rate of treated intracellular bacilli was 1% when compared with untreated cells. There was almost no killing effect of free RFP (4 or 15 microg/ml) on intracellular bacilli. In vivo efficacy of RFP-PLGA was also examined in rats infected with M. tuberculosis Kurono. Intratracheal administration of RFP-PLGA microspheres was shown to be superior to free RFP for killing of intracellular bacilli and preventing granuloma formation in some lobes. These results suggest that phagocytotic activity could be part of a new drug delivery system that selectively targeted macrophages.

  15. Control of Rhagoletis indifferents using Thiamethoxam and Spinosad baits under external fly pressure and its relation to rapidity of kill and residual bait activity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Control of western cherry fruit fly (Rhagoletis indifferens Curran) using thiamethoxam in sucrose bait and spinosad bait in cherry orchards under external fly pressure and its relation to rapidity of kill and residual bait activity were studied in Washington and Utah in 2010 and 2011. Thiamethoxam ...

  16. Phagocyte roulette in Salmonella killing.

    PubMed

    Fenlon, Luke A; Slauch, James M

    2014-01-15

    Salmonella propagates in macrophages to cause life-threatening infections, but the role of neutrophils in combating Salmonella has been controversial. In this issue, Burton et al. (2014) use single cell analyses and modeling to explain the ability of Salmonella to survive in macrophages while being killed by neutrophils.

  17. Killing of intracellular Mycobacterium tuberculosis by receptor-mediated drug delivery

    SciTech Connect

    Majumdar, S.; Basu, S.K. )

    1991-01-01

    p-Aminosalicylic acid (PAS) conjugated to maleylated bovine serum albumin (MBSA) was taken up efficiently through high-affinity MBSA-binding sites on macrophages. Binding of the radiolabeled conjugate to cultured mouse peritoneal macrophages at 4 degrees C was competed for by MBSA but not by PAS. At 37 degrees C, the radiolabeled conjugate was rapidly degraded by the macrophages, leading to release of acid-soluble degradation products in the medium. The drug conjugate was nearly 100 times as effective as free PAS in killing the intracellular mycobacteria in mouse peritoneal macrophages infected in culture with Mycobacterium tuberculosis. The killing of intracellular mycobacteria mediated by the drug conjugate was effectively prevented by simultaneous addition of excess MBSA (100 micrograms/ml) or chloroquine (3 microM) to the medium, whereas these agents did not affect the microbicidal action of free PAS. These results suggest that (i) uptake of the PAS-MBSA conjugate was mediated by cell surface receptors on macrophages which recognize MBSA and (ii) lysosomal hydrolysis of the internalized conjugate resulted in intracellular release of a pharmacologically active form of the drug, which led to selective killing of the M. tuberculosis harbored by mouse macrophages infected in culture. This receptor-mediated modality of delivering drugs to macrophages could contribute to greater therapeutic efficacy and minimization of toxic side effects in the management of tuberculosis and other intracellular mycobacterial infections.

  18. Treatment of Mycobacterium tuberculosis-Infected Macrophages with Poly(Lactic-Co-Glycolic Acid) Microparticles Drives NFκB and Autophagy Dependent Bacillary Killing.

    PubMed

    Lawlor, Ciaran; O'Connor, Gemma; O'Leary, Seonadh; Gallagher, Paul J; Cryan, Sally-Ann; Keane, Joseph; O'Sullivan, Mary P

    2016-01-01

    The emergence of multiple-drug-resistant tuberculosis (MDR-TB) has pushed our available repertoire of anti-TB therapies to the limit of effectiveness. This has increased the urgency to develop novel treatment modalities, and inhalable microparticle (MP) formulations are a promising option to target the site of infection. We have engineered poly(lactic-co-glycolic acid) (PLGA) MPs which can carry a payload of anti-TB agents, and are successfully taken up by human alveolar macrophages. Even without a drug cargo, MPs can be potent immunogens; yet little is known about how they influence macrophage function in the setting of Mycobacterium tuberculosis (Mtb) infection. To address this issue we infected THP-1 macrophages with Mtb H37Ra or H37Rv and treated with MPs. In controlled experiments we saw a reproducible reduction in bacillary viability when THP-1 macrophages were treated with drug-free MPs. NFκB activity was increased in MP-treated macrophages, although cytokine secretion was unaltered. Confocal microscopy of immortalized murine bone marrow-derived macrophages expressing GFP-tagged LC3 demonstrated induction of autophagy. Inhibition of caspases did not influence the MP-induced restriction of bacillary growth, however, blockade of NFκB or autophagy with pharmacological inhibitors reversed this MP effect on macrophage function. These data support harnessing inhaled PLGA MP-drug delivery systems as an immunotherapeutic in addition to serving as a vehicle for targeted drug delivery. Such "added value" could be exploited in the generation of inhaled vaccines as well as inhaled MDR-TB therapeutics when used as an adjunct to existing treatments. PMID:26894562

  19. Treatment of Mycobacterium tuberculosis-Infected Macrophages with Poly(Lactic-Co-Glycolic Acid) Microparticles Drives NFκB and Autophagy Dependent Bacillary Killing

    PubMed Central

    Lawlor, Ciaran; O’Connor, Gemma; O’Leary, Seonadh; Gallagher, Paul J.

    2016-01-01

    The emergence of multiple-drug-resistant tuberculosis (MDR-TB) has pushed our available repertoire of anti-TB therapies to the limit of effectiveness. This has increased the urgency to develop novel treatment modalities, and inhalable microparticle (MP) formulations are a promising option to target the site of infection. We have engineered poly(lactic-co-glycolic acid) (PLGA) MPs which can carry a payload of anti-TB agents, and are successfully taken up by human alveolar macrophages. Even without a drug cargo, MPs can be potent immunogens; yet little is known about how they influence macrophage function in the setting of Mycobacterium tuberculosis (Mtb) infection. To address this issue we infected THP-1 macrophages with Mtb H37Ra or H37Rv and treated with MPs. In controlled experiments we saw a reproducible reduction in bacillary viability when THP-1 macrophages were treated with drug-free MPs. NFκB activity was increased in MP-treated macrophages, although cytokine secretion was unaltered. Confocal microscopy of immortalized murine bone marrow-derived macrophages expressing GFP-tagged LC3 demonstrated induction of autophagy. Inhibition of caspases did not influence the MP-induced restriction of bacillary growth, however, blockade of NFκB or autophagy with pharmacological inhibitors reversed this MP effect on macrophage function. These data support harnessing inhaled PLGA MP-drug delivery systems as an immunotherapeutic in addition to serving as a vehicle for targeted drug delivery. Such “added value” could be exploited in the generation of inhaled vaccines as well as inhaled MDR-TB therapeutics when used as an adjunct to existing treatments. PMID:26894562

  20. Treatment of Mycobacterium tuberculosis-Infected Macrophages with Poly(Lactic-Co-Glycolic Acid) Microparticles Drives NFκB and Autophagy Dependent Bacillary Killing.

    PubMed

    Lawlor, Ciaran; O'Connor, Gemma; O'Leary, Seonadh; Gallagher, Paul J; Cryan, Sally-Ann; Keane, Joseph; O'Sullivan, Mary P

    2016-01-01

    The emergence of multiple-drug-resistant tuberculosis (MDR-TB) has pushed our available repertoire of anti-TB therapies to the limit of effectiveness. This has increased the urgency to develop novel treatment modalities, and inhalable microparticle (MP) formulations are a promising option to target the site of infection. We have engineered poly(lactic-co-glycolic acid) (PLGA) MPs which can carry a payload of anti-TB agents, and are successfully taken up by human alveolar macrophages. Even without a drug cargo, MPs can be potent immunogens; yet little is known about how they influence macrophage function in the setting of Mycobacterium tuberculosis (Mtb) infection. To address this issue we infected THP-1 macrophages with Mtb H37Ra or H37Rv and treated with MPs. In controlled experiments we saw a reproducible reduction in bacillary viability when THP-1 macrophages were treated with drug-free MPs. NFκB activity was increased in MP-treated macrophages, although cytokine secretion was unaltered. Confocal microscopy of immortalized murine bone marrow-derived macrophages expressing GFP-tagged LC3 demonstrated induction of autophagy. Inhibition of caspases did not influence the MP-induced restriction of bacillary growth, however, blockade of NFκB or autophagy with pharmacological inhibitors reversed this MP effect on macrophage function. These data support harnessing inhaled PLGA MP-drug delivery systems as an immunotherapeutic in addition to serving as a vehicle for targeted drug delivery. Such "added value" could be exploited in the generation of inhaled vaccines as well as inhaled MDR-TB therapeutics when used as an adjunct to existing treatments.

  1. Murine macrophage-lymphocyte interactions: scanning electron microscopic study.

    PubMed Central

    Albrecht, R M; Hinsdill, R D; Sandok, P L; Horowitz, S D

    1978-01-01

    Light and scanning electron microscopic observations revealed murine macrophage-lymphocyte interactions involving the initial contact of peritoneal, spleen, or thymus lymphocytes with peritoneal macrophage processes or microprocesses followed by clustering of lymphocytes over the central nuclear area of the macrophages. Lymphocyte-lymphocyte clustering was not observed in the absence of macrophages. Attachment and subsequent clustering appeared not to require the presence of serum or antigen; the attachment of allogeneic or xenogeneic lymphocytes was comparable to that seen in the syngeneic system, but central clustering of these lymphocytes failed to occur. No attachment or clustering was observed when thymic lymphocytes were cultured with thymus derived fibroblasts rather than with peritoneal macrophages. Lymphocyte attachment to immune, antigen-activated, syngeneic macrophages occurred more rapidly than that to normal unstimulated syngeneic macrophages; however, lymphocytes attached to the "activated" macrophages appeared to be killed by a nonphagocytic mechanism. A similar increase in the rate of lymphocyte attachment to macrophages occurred in the presence of migration inhibitory factor. Subsequent lymphocyte clustering on macrophages was observed in the migration inhibitory factor-stimulated cultures. In addition, lymphocyte-macrophage interactions similar to those in vitro were observed to occur in vivo on intraperitoneally implanted cover slips. Images PMID:101458

  2. Cytotoxic macrophage-released tumour necrosis factor-alpha (TNF-α) as a killing mechanism for cancer cell death after cold plasma activation

    NASA Astrophysics Data System (ADS)

    Kaushik, Nagendra Kumar; Kaushik, Neha; Min, Booki; Choi, Ki Hong; Hong, Young June; Miller, Vandana; Fridman, Alexander; Choi, Eun Ha

    2016-03-01

    The present study aims at studying the anticancer role of cold plasma-activated immune cells. The direct anti-cancer activity of plasma-activated immune cells against human solid cancers has not been described so far. Hence, we assessed the effect of plasma-treated RAW264.7 macrophages on cancer cell growth after co-culture. In particular, flow cytometer analysis revealed that plasma did not induce any cell death in RAW264.7 macrophages. Interestingly, immunofluorescence and western blot analysis confirmed that TNF-α released from plasma-activated macrophages acts as a tumour cell death inducer. In support of these findings, activated macrophages down-regulated the cell growth in solid cancer cell lines and induced cell death in vitro. Together our findings suggest plasma-induced reactive species recruit cytotoxic macrophages to release TNF-α, which blocks cancer cell growth and can have the potential to contribute to reducing tumour growth in vivo in the near future.

  3. Beyond killing

    PubMed Central

    Vale, Pedro F.; McNally, Luke; Doeschl-Wilson, Andrea; King, Kayla C.; Popat, Roman; Domingo-Sananes, Maria R.; Allen, Judith E.; Soares, Miguel P.; Kümmerli, Rolf

    2016-01-01

    The antibiotic pipeline is running dry and infectious disease remains a major threat to public health. An efficient strategy to stay ahead of rapidly adapting pathogens should include approaches that replace, complement or enhance the effect of both current and novel antimicrobial compounds. In recent years, a number of innovative approaches to manage disease without the aid of traditional antibiotics and without eliminating the pathogens directly have emerged. These include disabling pathogen virulence-factors, increasing host tissue damage control or altering the microbiota to provide colonization resistance, immune resistance or disease tolerance against pathogens. We discuss the therapeutic potential of these approaches and examine their possible consequences for pathogen evolution. To guarantee a longer half-life of these alternatives to directly killing pathogens, and to gain a full understanding of their population-level consequences, we encourage future work to incorporate evolutionary perspectives into the development of these treatments. PMID:27016341

  4. Surface expression and rapid internalization of macrosialin (mouse CD68) on elicited mouse peritoneal macrophages.

    PubMed

    Kurushima, H; Ramprasad, M; Kondratenko, N; Foster, D M; Quehenberger, O; Steinberg, D

    2000-01-01

    Macrosialin, the mouse homolog of human CD68, is a heavily glycosylated transmembrane protein found almost exclusively in macrophages. Its function remains uncertain. It has a high affinity for oxidized low-density lipoprotein (LDL) in ligand blots and antibodies against the human homolog, CD68, inhibit the binding of oxidized LDL to a human monocyte-derived cell line (THP-1). However, there is still controversy as to whether macrosialin, found predominantly in late endosomes, is expressed at all on the plasma membrane. The present studies, done in thioglycollate-elicited peritoneal macrophages, confirm that macrosialin is predominantly intracellular but show clearly that 10-15% of it is expressed on the cell surface. Exchange with intracellular pools occurs at an extremely high rate. The results are compatible with a surface function, including internalization of bound ligands or adhesion to surfaces.

  5. A Highly Conserved Toxo1 Haplotype Directs Resistance to Toxoplasmosis and Its Associated Caspase-1 Dependent Killing of Parasite and Host Macrophage

    PubMed Central

    Bisanz, Cordelia; Lagrange, Dominique; Pilloux, Ludovic; Massera, Céline; Cristinelli, Sara; Jublot, Delphine; Bastien, Olivier; Loeuillet, Corinne; Aldebert, Delphine; Touquet, Bastien; Fournié, Gilbert J.; Cesbron-Delauw, Marie France

    2014-01-01

    Natural immunity or resistance to pathogens most often relies on the genetic make-up of the host. In a LEW rat model of refractoriness to toxoplasmosis, we previously identified on chromosome 10 the Toxo1 locus that directs toxoplasmosis outcome and controls parasite spreading by a macrophage-dependent mechanism. Now, we narrowed down Toxo1 to a 891 kb interval containing 29 genes syntenic to human 17p13 region. Strikingly, Toxo1 is included in a haplotype block strictly conserved among all refractory rat strains. The sequencing of Toxo1 in nine rat strains (5 refractory and 4 susceptible) revealed resistant-restricted conserved polymorphisms displaying a distribution gradient that peaks at the bottom border of Toxo1, and highlighting the NOD-like receptor, Nlrp1a, as a major candidate. The Nlrp1 inflammasome is known to trigger, upon pathogen intracellular sensing, pyroptosis programmed-cell death involving caspase-1 activation and cleavage of IL-1β. Functional studies demonstrated that the Toxo1-dependent refractoriness in vivo correlated with both the ability of macrophages to restrict T. gondii growth and a T. gondii-induced death of intracellular parasites and its host macrophages. The parasite-induced cell death of infected macrophages bearing the LEW-Toxo1 alleles was found to exhibit pyroptosis-like features with ROS production, the activation of caspase-1 and IL1-β secretion. The pharmacological inactivation of caspase-1 using YVAD and Z-VAD inhibitors prevented the death of both intravacuolar parasites and host non-permissive macrophages but failed to restore parasite proliferation. These findings demonstrated that the Toxo1-dependent response of rat macrophages to T. gondii infection may trigger two pathways leading to the control of parasite proliferation and the death of parasites and host macrophages. The NOD-like receptor NLRP1a/Caspase-1 pathway is the best candidate to mediate the parasite-induced cell death. These data represent new insights

  6. A highly conserved Toxo1 haplotype directs resistance to toxoplasmosis and its associated caspase-1 dependent killing of parasite and host macrophage.

    PubMed

    Cavailles, Pierre; Flori, Pierre; Papapietro, Olivier; Bisanz, Cordelia; Lagrange, Dominique; Pilloux, Ludovic; Massera, Céline; Cristinelli, Sara; Jublot, Delphine; Bastien, Olivier; Loeuillet, Corinne; Aldebert, Delphine; Touquet, Bastien; Fournié, Gilbert J; Cesbron-Delauw, Marie France

    2014-04-01

    Natural immunity or resistance to pathogens most often relies on the genetic make-up of the host. In a LEW rat model of refractoriness to toxoplasmosis, we previously identified on chromosome 10 the Toxo1 locus that directs toxoplasmosis outcome and controls parasite spreading by a macrophage-dependent mechanism. Now, we narrowed down Toxo1 to a 891 kb interval containing 29 genes syntenic to human 17p13 region. Strikingly, Toxo1 is included in a haplotype block strictly conserved among all refractory rat strains. The sequencing of Toxo1 in nine rat strains (5 refractory and 4 susceptible) revealed resistant-restricted conserved polymorphisms displaying a distribution gradient that peaks at the bottom border of Toxo1, and highlighting the NOD-like receptor, Nlrp1a, as a major candidate. The Nlrp1 inflammasome is known to trigger, upon pathogen intracellular sensing, pyroptosis programmed-cell death involving caspase-1 activation and cleavage of IL-1β. Functional studies demonstrated that the Toxo1-dependent refractoriness in vivo correlated with both the ability of macrophages to restrict T. gondii growth and a T. gondii-induced death of intracellular parasites and its host macrophages. The parasite-induced cell death of infected macrophages bearing the LEW-Toxo1 alleles was found to exhibit pyroptosis-like features with ROS production, the activation of caspase-1 and IL1-β secretion. The pharmacological inactivation of caspase-1 using YVAD and Z-VAD inhibitors prevented the death of both intravacuolar parasites and host non-permissive macrophages but failed to restore parasite proliferation. These findings demonstrated that the Toxo1-dependent response of rat macrophages to T. gondii infection may trigger two pathways leading to the control of parasite proliferation and the death of parasites and host macrophages. The NOD-like receptor NLRP1a/Caspase-1 pathway is the best candidate to mediate the parasite-induced cell death. These data represent new insights

  7. Guanylate binding proteins enable rapid activation of canonical and noncanonical inflammasomes in Chlamydia-infected macrophages.

    PubMed

    Finethy, Ryan; Jorgensen, Ine; Haldar, Arun K; de Zoete, Marcel R; Strowig, Till; Flavell, Richard A; Yamamoto, Masahiro; Nagarajan, Uma M; Miao, Edward A; Coers, Jörn

    2015-12-01

    Interferon (IFN)-inducible guanylate binding proteins (GBPs) mediate cell-autonomous host resistance to bacterial pathogens and promote inflammasome activation. The prevailing model postulates that these two GBP-controlled activities are directly linked through GBP-dependent vacuolar lysis. It was proposed that the rupture of pathogen-containing vacuoles (PVs) by GBPs destroyed the microbial refuge and simultaneously contaminated the host cell cytosol with microbial activators of inflammasomes. Here, we demonstrate that GBP-mediated host resistance and GBP-mediated inflammatory responses can be uncoupled. We show that PVs formed by the rodent pathogen Chlamydia muridarum, so-called inclusions, remain free of GBPs and that C. muridarum is impervious to GBP-mediated restrictions on bacterial growth. Although GBPs neither bind to C. muridarum inclusions nor restrict C. muridarum growth, we find that GBPs promote inflammasome activation in C. muridarum-infected macrophages. We demonstrate that C. muridarum infections induce GBP-dependent pyroptosis through both caspase-11-dependent noncanonical and caspase-1-dependent canonical inflammasomes. Among canonical inflammasomes, we find that C. muridarum and the human pathogen Chlamydia trachomatis activate not only NLRP3 but also AIM2. Our data show that GBPs support fast-kinetics processing and secretion of interleukin-1β (IL-1β) and IL-18 by the NLRP3 inflammasome but are dispensable for the secretion of the same cytokines at later times postinfection. Because IFN-γ fails to induce IL-1β transcription, GBP-dependent fast-kinetics inflammasome activation can drive the preferential processing of constitutively expressed IL-18 in IFN-γ-primed macrophages in the absence of prior Toll-like receptor stimulation. Together, our results reveal that GBPs control the kinetics of inflammasome activation and thereby shape macrophage responses to Chlamydia infections.

  8. Guanylate Binding Proteins Enable Rapid Activation of Canonical and Noncanonical Inflammasomes in Chlamydia-Infected Macrophages

    PubMed Central

    Finethy, Ryan; Jorgensen, Ine; Haldar, Arun K.; de Zoete, Marcel R.; Strowig, Till; Flavell, Richard A.; Yamamoto, Masahiro; Nagarajan, Uma M.; Miao, Edward A.

    2015-01-01

    Interferon (IFN)-inducible guanylate binding proteins (GBPs) mediate cell-autonomous host resistance to bacterial pathogens and promote inflammasome activation. The prevailing model postulates that these two GBP-controlled activities are directly linked through GBP-dependent vacuolar lysis. It was proposed that the rupture of pathogen-containing vacuoles (PVs) by GBPs destroyed the microbial refuge and simultaneously contaminated the host cell cytosol with microbial activators of inflammasomes. Here, we demonstrate that GBP-mediated host resistance and GBP-mediated inflammatory responses can be uncoupled. We show that PVs formed by the rodent pathogen Chlamydia muridarum, so-called inclusions, remain free of GBPs and that C. muridarum is impervious to GBP-mediated restrictions on bacterial growth. Although GBPs neither bind to C. muridarum inclusions nor restrict C. muridarum growth, we find that GBPs promote inflammasome activation in C. muridarum-infected macrophages. We demonstrate that C. muridarum infections induce GBP-dependent pyroptosis through both caspase-11-dependent noncanonical and caspase-1-dependent canonical inflammasomes. Among canonical inflammasomes, we find that C. muridarum and the human pathogen Chlamydia trachomatis activate not only NLRP3 but also AIM2. Our data show that GBPs support fast-kinetics processing and secretion of interleukin-1β (IL-1β) and IL-18 by the NLRP3 inflammasome but are dispensable for the secretion of the same cytokines at later times postinfection. Because IFN-γ fails to induce IL-1β transcription, GBP-dependent fast-kinetics inflammasome activation can drive the preferential processing of constitutively expressed IL-18 in IFN-γ-primed macrophages in the absence of prior Toll-like receptor stimulation. Together, our results reveal that GBPs control the kinetics of inflammasome activation and thereby shape macrophage responses to Chlamydia infections. PMID:26416908

  9. A deficiency in the B cell response of C57BL/6 mice correlates with loss of macrophage-mediated killing of Leishmania amazonensis

    PubMed Central

    Gibson-Corley, Katherine N.; Boggiatto, Paola M.; Mukbel, Rami M.; Petersen, Christine A.; Jones, Douglas E.

    2010-01-01

    Infection of C3HeB/FeJ and C57BL/6 mice with Leishmania major stimulates a healing cell-mediated immune response, while Leishmania amazonensis infection leads to chronic disease. Here we show C3HeB/FeJ mice co-infected with both species of Leishmania heal, while co-infected C57BL/6 mice do not. Using an in vitro killing assay we determined B cells from infected C57BL/6 mice are ineffective in promoting parasite killing compared with B cells from infected C3HeB/FeJ mice. Furthermore, infected C57BL/6 mice produce less antigen-specific antibodies compared with infected C3HeB/FeJ mice. These findings suggest B cells play a required role in the cell-mediated immune response against L. amazonensis. PMID:20004204

  10. Killing and conformal Killing tensors

    NASA Astrophysics Data System (ADS)

    Heil, Konstantin; Moroianu, Andrei; Semmelmann, Uwe

    2016-08-01

    We introduce an appropriate formalism in order to study conformal Killing (symmetric) tensors on Riemannian manifolds. We reprove in a simple way some known results in the field and obtain several new results, like the classification of conformal Killing 2-tensors on Riemannian products of compact manifolds, Weitzenböck formulas leading to non-existence results, and construct various examples of manifolds with conformal Killing tensors.

  11. Killing Coyotes.

    ERIC Educational Resources Information Center

    Beasley, Conger, Jr.

    1993-01-01

    Presents different viewpoints concerning the federal government's Animal Damage Control (ADC) Program cited as responsible for killing millions of predators. Critics provide evidence of outdated and inhumane methods exemplified in the coyote killings. The ADC emphasizes new, nonlethal methods of controlling animals cited as "noxious." (MCO)

  12. Depletion of Spleen Macrophages Delays AA Amyloid Development: A Study Performed in the Rapid Mouse Model of AA Amyloidosis

    PubMed Central

    Lundmark, Katarzyna; Vahdat Shariatpanahi, Aida; Westermark, Gunilla T.

    2013-01-01

    AA amyloidosis is a systemic disease that develops secondary to chronic inflammatory diseases Macrophages are often found in the vicinity of amyloid deposits and considered to play a role in both formation and degradation of amyloid fibrils. In spleen reside at least three types of macrophages, red pulp macrophages (RPM), marginal zone macrophages (MZM), metallophilic marginal zone macrophages (MMZM). MMZM and MZM are located in the marginal zone and express a unique collection of scavenger receptors that are involved in the uptake of blood-born particles. The murine AA amyloid model that resembles the human form of the disease has been used to study amyloid effects on different macrophage populations. Amyloid was induced by intravenous injection of amyloid enhancing factor and subcutaneous injections of silver nitrate and macrophages were identified with specific antibodies. We show that MZMs are highly sensitive to amyloid and decrease in number progressively with increasing amyloid load. Total area of MMZMs is unaffected by amyloid but cells are activated and migrate into the white pulp. In a group of mice spleen macrophages were depleted by an intravenous injection of clodronate filled liposomes. Subsequent injections of AEF and silver nitrate showed a sustained amyloid development. RPMs that constitute the majority of macrophages in spleen, appear insensitive to amyloid and do not participate in amyloid formation. PMID:24236094

  13. Rapid and transient activation of γδ T cells to IFN-γ production, NK cell-like killing, and antigen processing during acute virus infection.

    PubMed

    Toka, Felix N; Kenney, Mary A; Golde, William T

    2011-04-15

    γδ T cells are the majority peripheral blood T cells in young cattle. The role of γδ T cells in innate responses against infection with foot-and-mouth disease virus was analyzed on consecutive 5 d following infection. Before infection, bovine WC1(+) γδ T cells expressed a nonactivated phenotype relative to CD62L, CD45RO, and CD25 expression and did not produce IFN-γ ex vivo. Additionally, CD335 expression was lacking and no spontaneous target cell lysis could be detected in vitro, although perforin was detectable at a very low level. MHC class II and CD13 expression were also lacking. Following infection with foot-and-mouth disease virus, expression of CD62L and CD45RO was greatly reduced on WC1(+) γδ T cells, and unexpectedly, CD45RO expression did not recover. A transient increase in expression of CD25 correlated with production of IFN-γ. Expression of CD335 and production of perforin were detected on a subset of γδ T cells, and this correlated with an increased spontaneous killing of xenogeneic target cells. Furthermore, increased MHC class II expression was detected on WC1(+) γδ T cells, and these cells processed protein Ags. These activities are rapidly induced, within 3 d, and wane by 5 d following infection. All of these functions, NK-like killing, Ag processing, and IFN-γ production, have been demonstrated for these cells in various species. However, these results are unique in that all these functions are detected in the same samples of WC1(+) γδ T cells, suggesting a pivotal role of these cells in controlling virus infection.

  14. The contribution of both oxygen and nitrogen intermediates to the intracellular killing mechanisms of C1q-opsonized Listeria monocytogenes by the macrophage-like IC-21 cell line.

    PubMed

    Alvarez-Domínguez, C; Carrasco-Marín, E; López-Mato, P; Leyva-Cobián, F

    2000-09-01

    Listeria monocytogenes is a facultative intracellular pathogen which is internalized by host mammalian cells upon binding to their surface. Further listerial growth occurs in the cytosol after escape from the phagosomal-endosomal compartment. We have previously reported that C1q is able to potentiate L. monocytogenes phagocytosis upon bacterial opsonization by ingestion through C1q-binding structures. In this report, we analysed the post-phagocytic events upon internalization of C1q-opsonized L. monocytogenes and found an induction of macrophage (Mphi)-like IC-21 cell bactericidal mechanisms displayed by the production of oxygen and nitrogen metabolites. Both types of molecules are effective in L. monocytogenes killing. Further analysis of the cellular responses promoted by interaction of C1q with its surface binding structures, leads us to consider C1q as a collaborative molecule involved in Mphi activation. Upon interaction with surface binding structures, C1q was able to trigger and/or amplify the production of reactive oxygen and nitrogen intermediates induced by stimuli such as interferon-gamma and L. monocytogenes phagocytosis. PMID:11012757

  15. The contribution of both oxygen and nitrogen intermediates to the intracellular killing mechanisms of C1q-opsonized Listeria monocytogenes by the macrophage-like IC-21 cell line

    PubMed Central

    Álvarez-Domínguez, C; Carrasco-Marín, E; López-Mato, P; Leyva-Cobián, F

    2000-01-01

    Listeria monocytogenes is a facultative intracellular pathogen which is internalized by host mammalian cells upon binding to their surface. Further listerial growth occurs in the cytosol after escape from the phagosomal–endosomal compartment. We have previously reported that C1q is able to potentiate L. monocytogenes phagocytosis upon bacterial opsonization by ingestion through C1q-binding structures. In this report, we analysed the post-phagocytic events upon internalization of C1q-opsonized L. monocytogenes and found an induction of macrophage (Mφ)-like IC-21 cell bactericidal mechanisms displayed by the production of oxygen and nitrogen metabolites. Both types of molecules are effective in L. monocytogenes killing. Further analysis of the cellular responses promoted by interaction of C1q with its surface binding structures, leads us to consider C1q as a collaborative molecule involved in Mφ activation. Upon interaction with surface binding structures, C1q was able to trigger and/or amplify the production of reactive oxygen and nitrogen intermediates induced by stimuli such as interferon-γ and L. monocytogenes phagocytosis. PMID:11012757

  16. Macrophage proliferation, provenance, and plasticity in macroparasite infection

    PubMed Central

    Rückerl, Dominik; Allen, Judith E

    2014-01-01

    Summary: Macrophages have long been center stage in the host response to microbial infection, but only in the past 10–15 years has there been a growing appreciation for their role in helminth infection and the associated type 2 response. Through the actions of the IL-4 receptor α (IL-4Rα), type 2 cytokines result in the accumulation of macrophages with a distinctive activation phenotype. Although our knowledge of IL-4Rα-induced genes is growing rapidly, the specific functions of these macrophages have yet to be established in most disease settings. Understanding the interplay between IL-4Rα-activated macrophages and the other cellular players is confounded by the enormous transcriptional heterogeneity within the macrophage population and by their highly plastic nature. Another level of complexity is added by the new knowledge that tissue macrophages can be derived either from a resident prenatal population or from blood monocyte recruitment and that IL-4 can increase macrophage numbers through proliferative expansion. Here, we review current knowledge on the contribution of macrophages to helminth killing and wound repair, with specific attention paid to distinct cellular origins and plasticity potential. PMID:25319331

  17. Killing Range

    PubMed Central

    Asal, Victor; Rethemeyer, R. Karl; Horgan, John

    2015-01-01

    This paper presents an analysis of the Provisional Irish Republican Army's (PIRA) brigade level behavior during the Northern Ireland Conflict (1970-1998) and identifies the organizational factors that impact a brigade's lethality as measured via terrorist attacks. Key independent variables include levels of technical expertise, cadre age, counter-terrorism policies experienced, brigade size, and IED components and delivery methods. We find that technical expertise within a brigade allows for careful IED usage, which significantly minimizes civilian casualties (a specific strategic goal of PIRA) while increasing the ability to kill more high value targets with IEDs. Lethal counter-terrorism events also significantly affect a brigade's likelihood of killing both civilians and high-value targets but in different ways. Killing PIRA members significantly decreases IED fatalities but also significantly decreases the possibility of zero civilian IED-related deaths in a given year. Killing innocent Catholics in a Brigade's county significantly increases total and civilian IED fatalities. Together the results suggest the necessity to analyze dynamic situational variables that impact terrorist group behavior at the sub-unit level. PMID:25838603

  18. High viral burden and rapid CD4+ cell depletion in human immunodeficiency virus type 1-infected SCID-hu mice suggest direct viral killing of thymocytes in vivo.

    PubMed Central

    Jamieson, B D; Uittenbogaart, C H; Schmid, I; Zack, J A

    1997-01-01

    The mechanism of CD4+ cell loss in lymphoid organs is unknown. In this study, human immunodeficiency virus (HIV) infection of human fetal thymus/liver implants in severe combined immunodeficient mice was used to investigate the mechanism of HIV-induced depletion of CD4-bearing cells in vivo. The implants were assessed for depletion of CD4+ thymocytes, apoptosis, and viral burden. We detected two phases of CD4 cell depletion, an initial rapid phase and a more gradual later phase. Compared to mock-infected implants, HIV-infected implants did not demonstrate detectable increases in the levels of apoptosis while severe depletion of CD4-bearing cells was ongoing. During peak loss of CD4+ cells, high viral burden was observed, suggesting that loss of CD4+ cells in this in vivo system is due to direct killing of infected thymocytes. Increased levels of apoptosis were observed during the later phase of thymocyte depletion; however, these apoptotic cells lacked CD4. This finding suggests that a second indirect mechanism may be responsible for the destruction of CD4- CD8+ thymocytes in vivo. Taken together, these results suggest that CD4+ and CD4- cells may die by different mechanism(s). PMID:9343176

  19. AN EXAMINATION OF THE CYTOTOXIC EFFECTS OF SILICA ON MACROPHAGES

    PubMed Central

    Allison, A. C.; Harington, J. S.; Birbeck, M.

    1966-01-01

    Effects of silica, diamond dust, and carrageenan on mouse macrophages were studied by phase-contrast cine-micrography, electron microscopy, histochemical techniques for lysosomal enzymes and measurements of the release of lysosomal enzymes into the culture medium. All added materials were rapidly taken up into phagosomes, to which lysosomes became attached. In all cases lysosomal enzymes were discharged into the phagosomes to form secondary lysosomes. Within 24 hr most of the silica particles and enzyme had escaped from the secondary lysosomes and lysosomal enzymes were found in the culture media. Most macrophages were killed by this time. With nontoxic particles (diamond dust, aluminium-coated silica, or silica in the presence of the protective agent polyvinyl-pyridine-N-oxide, PVPNO) ingested particles and lysosomal enzymes were retained within the secondary lysosomes for a much longer time, and cytotoxic effects were considerably delayed or absent altogether. It is concluded that silica particles are toxic because they are efficiently taken up by macrophages and can then react relatively rapidly with the membranes surrounding the secondary lysosomes. The particles and lytic enzymes can then escape into the cytoplasm, producing general damage, and thence into the culture medium. It is suggested that hydrogen bonding of silicic acid with lipid and protein constituents of the membrane accounts for the induced permeability. Protective agents such as PVPNO are retamed in lysosomes and preferentially form hydrogen bonds with silicic acid. Carrageenan is demonstrable within macrophages by its metachromatic reaction. It brings about release of enzymes from secondary lysosomes, but much more slowly than does silica. Silica released from killed macrophages is as cytotoxic as the original preparation. It is suggested that repeated cycles of macrophage killing in vivo leads to the mobilization of fibroblasts and fibrogenesis characterizing the disease silicosis. PMID

  20. How microglia kill neurons.

    PubMed

    Brown, Guy C; Vilalta, Anna

    2015-12-01

    Microglia are resident brain macrophages that become inflammatory activated in most brain pathologies. Microglia normally protect neurons, but may accidentally kill neurons when attempting to limit infections or damage, and this may be more common with degenerative disease as there was no significant selection pressure on the aged brain in the past. A number of mechanisms by which activated microglia kill neurons have been identified, including: (i) stimulation of the phagocyte NADPH oxidase (PHOX) to produce superoxide and derivative oxidants, (ii) expression of inducible nitric oxide synthase (iNOS) producing NO and derivative oxidants, (iii) release of glutamate and glutaminase, (iv) release of TNFα, (v) release of cathepsin B, (vi) phagocytosis of stressed neurons, and (vii) decreased release of nutritive BDNF and IGF-1. PHOX stimulation contributes to microglial activation, but is not directly neurotoxic unless NO is present. NO is normally neuroprotective, but can react with superoxide to produce neurotoxic peroxynitrite, or in the presence of hypoxia inhibit mitochondrial respiration. Glutamate can be released by glia or neurons, but is neurotoxic only if the neurons are depolarised, for example as a result of mitochondrial inhibition. TNFα is normally neuroprotective, but can become toxic if caspase-8 or NF-κB activation are inhibited. If the above mechanisms do not kill neurons, they may still stress the neurons sufficiently to make them susceptible to phagocytosis by activated microglia. We review here whether microglial killing of neurons is an artefact, makes evolutionary sense or contributes in common neuropathologies and by what mechanisms. This article is part of a Special Issue entitled SI: Neuroprotection.

  1. Effects of lipopolysaccharide on the catabolic activity of macrophages

    SciTech Connect

    Cluff, C.; Ziegler, H.K.

    1986-03-05

    The ability of macrophages to degrade and catabolize antigens is of relevance both as a means to process complex antigens prior to presentation to T cells, as well as a way to down regulate immune responses by destroying the antigenicity of polypeptides. With these considerations, the authors have investigated the regulation of macrophage catabolic activity by lipopolysaccharide (LPS). Catabolic activity was quantitated by following the distribution and molecular form of /sup 125/-I labelled surface components of heat-killed Listeria monocytogenes (HKLM) subsequent to their uptake by macrophages. They have compared the catabolic activity of macrophages from peritoneal exudates of mice injected i.p. with saline or LPS and have found that LPS-elicited macrophages display a greatly enhanced (3 fold) rate of catabolism. This increase in catabolic activity peaks 3 days after LPS injection and steadily declines thereafter, approaching a baseline level after 3 weeks. The enhancement of catabolic activity is under LPS gene control. LPS-elicited macrophages rapidly destroy the antigenicity of bacterial antigens and function poorly as antigen presenting cells in vitro. These results suggest that LPS elicits a macrophage population specialized for antigen degradation functions with negative regulatory effects on the induction of specific immune responses.

  2. Bioluminescent Mycobacterium aurum expressing firefly luciferase for rapid and high throughput screening of antimycobacterial drugs in vitro and in infected macrophages.

    PubMed

    Deb, D K; Srivastava, K K; Srivastava, R; Srivastava, B S

    2000-12-20

    The slow growth and highly infectious nature of Mycobacterium tuberculosis is a limiting factor in its use as test organism in high throughput screening for inhibitory compounds. To overcome these problems, use of surrogate strains and reporter genes have been considered. In this study, we have investigated the application of a fast growing nonpathogenic M. aurum expressing firefly luciferase in rapid screening of antituberculosis compounds in vitro and in infected macrophages using bioluminescence assay. The assay is based on luminescence determination using luciferin as substrate. Inhibition of bioluminescence was obtained with frontline antimycobacterial drugs like streptomycin, rifampicin, isoniazid, ethambutol, ofloxacin, and sparfloxacin at their reported MICs. Inhibition could be observed as early as 2 h in vitro and within 24 h in infected macrophages. The system can reliably be used in high throughput screening.

  3. Selective Killing of Nonreplicating Mycobacteria

    PubMed Central

    Bryk, Ruslana; Gold, Benjamin; Venugopal, Aditya; Singh, Jasbir; Samy, Raghu; Pupek, Krzysztof; Cao, Hua; Popescu, Carmen; Gurney, Mark; Hotha, Srinivas; Cherian, Joseph; Rhee, Kyu; Ly, Lan; Converse, Paul J.; Ehrt, Sabine; Vandal, Omar; Jiang, Xiuju; Schneider, Jean; Lin, Gang; Nathan, Carl

    2008-01-01

    SUMMARY Antibiotics are typically more effective against replicating rather than nonreplicating bacteria. However, a major need in global health is to eradicate persistent or nonreplicating subpopulations of bacteria such as Mycobacterium tuberculosis (Mtb). Hence, identifying chemical inhibitors that selectively kill bacteria that are not replicating is of practical importance. To address this, we screened for inhibitors of dihydrolipoamide acyltransferase (DlaT), an enzyme required by Mtb to cause tuberculosis in guinea pigs and used by the bacterium to resist nitric oxide-derived reactive nitrogen intermediates, a stress encountered in the host. Chemical screening for inhibitors of Mtb DlaT identified select rhodanines as compounds that almost exclusively kill nonreplicating mycobacteria in synergy with products of host immunity, such as nitric oxide and hypoxia, and are effective on bacteria within macrophages, a cellular reservoir for latent Mtb. Compounds that kill nonreplicating pathogens in cooperation with host immunity could complement the conventional chemotherapy of infectious disease. PMID:18329613

  4. Why is particulate matter produced by wildfires toxic to lung macrophages?

    SciTech Connect

    Franzi, Lisa M.; Bratt, Jennifer M.; Williams, Keisha M.; Last, Jerold A.

    2011-12-15

    The mechanistic basis of the high toxicity to lung macrophages of coarse PM from the California wildfires of 2008 was examined in cell culture experiments with mouse macrophages. Wildfire PM directly killed macrophages very rapidly in cell culture at relatively low doses. The wildfire coarse PM is about four times more toxic to macrophages on an equal weight basis than the same sized PM collected from normal ambient air (no wildfires) from the same region and season. There was a good correlation between the extent of cytotoxicity and the amount of oxidative stress observed at a given dose of wildfire PM in vitro. Our data implicate NF-{kappa}B signaling in the response of macrophages to wildfire PM, and suggest that most, if not all, of the cytotoxicity of wildfire PM to lung macrophages is the result of oxidative stress. The relative ratio of toxicity and of expression of biomarkers of oxidant stress between wildfire PM and 'normal' PM collected from ambient air is consistent with our previous results in mice in vivo, also suggesting that most, if not all, of the cytotoxicity of wildfire PM to lung macrophages is the result of oxidative stress. Our findings from this and earlier studies suggest that the active components of coarse PM from the wildfire are heat-labile organic compounds. While we cannot rule out a minor role for endotoxin in coarse PM preparations from the collected wildfire PM in our observed results both in vitro and in vivo, based on experiments using the inhibitor Polymyxin B most of the oxidant stress and pro-inflammatory activity observed was not due to endotoxin. -- Highlights: Black-Right-Pointing-Pointer Wildfire coarse PM kills macrophages at lower doses than coarse. Black-Right-Pointing-Pointer Wildfire coarse PM activates the NF-kB pathway at lower doses than ambient. Black-Right-Pointing-Pointer Wildfire coarse PM in vitro and in vivo kill macrophages by oxidative stress.

  5. M1 and M2 Macrophages: The Chicken and the Egg of Immunity

    PubMed Central

    Mills, Charles D.; Ley, Klaus

    2015-01-01

    The purpose of this perspective is to describe a critical advance in understanding how immune responses work. Macrophages are required for all animal life: ‘Inhibit’ type macrophages in all animals (called M1) can rapidly kill pathogens, and are thus the primary host defense, and ‘Heal’ type macrophages (M2) routinely repair and maintain tissue integrity. Macrophages perform these activities in all animals without T cells, and also in T cell-deficient vertebrates. Although adaptive immunity can amplify macrophage polarization, the long-held notion that macrophages need to be ‘activated’ or ‘alternatively activated’ by T cells is incorrect; indeed, immunology has had it backward. M1/M2-type macrophages necessarily direct T cells toward Th1- or Th2-like activities, respectively. That such macrophage-innate activities are the central directing element in immune responses is a dramatic change in understanding how immune systems operate. Most important, this revelation is opening up whole new approaches to immunotherapy. For example, many modern diseases, such as cancer and atherosclerosis, may not display ‘foreign’ antigens. However, there are clear imbalances in M1/M2-type responses. Correcting such innate imbalances can result in better health. Macrophages are the chicken and the egg of immunity. PMID:25138714

  6. Macrophage Activation Redirects Yersinia-Infected Host Cell Death from Apoptosis to Caspase-1-Dependent Pyroptosis

    PubMed Central

    Bergsbaken, Tessa; Cookson, Brad T

    2007-01-01

    Infection of macrophages by Yersinia species results in YopJ-dependent apoptosis, and naïve macrophages are highly susceptible to this form of cell death. Previous studies have demonstrated that macrophages activated with lipopolysaccharide (LPS) prior to infection are resistant to YopJ-dependent cell death; we found this simultaneously renders macrophages susceptible to killing by YopJ− Yersinia pseudotuberculosis (Yptb). YopJ− Yptb-induced macrophage death was dependent on caspase-1 activation, resulting in rapid permeability to small molecules, followed by membrane breakdown and DNA damage, and accompanied by cleavage and release of proinflammatory interleukin-18. Induction of caspase-1-dependent death, or pyroptosis, required the bacterial type III translocon but none of its known translocated proteins. Wild-type Yptb infection also triggered pyroptosis: YopJ-dependent activation of proapoptotic caspase-3 was significantly delayed in activated macrophages and resulted in caspase-1-dependent pyroptosis. The transition to susceptibility was not limited to LPS activation; it was also seen in macrophages activated with other Toll-like receptor (TLR) ligands and intact nonviable bacteria. Yptb infection triggered macrophage activation and activation of caspase-1 in vivo. Y. pestis infection of activated macrophages also stimulated caspase-1 activation. These results indicate that host signaling triggered by TLR and other activating ligands during the course of Yersinia infection redirects both the mechanism of host cell death and the downstream consequences of death by shifting from noninflammatory apoptosis to inflammatory pyroptosis. PMID:17983266

  7. CD4 Depletion in SIV-Infected Macaques Results in Macrophage and Microglia Infection with Rapid Turnover of Infected Cells

    PubMed Central

    Ortiz, Alexandra M.; Ryan, Emily S.; McGary, Colleen S.; Deleage, Claire; McAtee, Brigitte B.; He, Tianyu; Apetrei, Cristian; Easley, Kirk; Pahwa, Savita; Collman, Ronald G.; Derdeyn, Cynthia A.; Davenport, Miles P.; Estes, Jacob D.; Silvestri, Guido; Lackner, Andrew A.; Paiardini, Mirko

    2014-01-01

    In rhesus macaques (RMs), experimental depletion of CD4+ T-cells prior to SIV infection results in higher viremia and emergence of CD4-independent SIV-envelopes. In this study we used the rhesus recombinant anti-CD4 antibody CD4R1 to deplete RM CD4+ T-cells prior to SIVmac251 infection and investigate the sources of the increased viral burden and the lifespan of productively infected cells. CD4-depleted animals showed (i) set-point viral load two-logs higher than controls; (ii) macrophages constituting 80% of all SIV vRNA+ cells in lymph node and mucosal tissues; (iii) substantial expansion of pro-inflammatory monocytes; (iv) aberrant activation and infection of microglial cells; and (v) lifespan of productively infected cells significantly longer in comparison to controls, but markedly shorter than previously estimated for macrophages. The net effect of CD4+ T-cell depletion is an inability to control SIV replication and a shift in the tropism of infected cells to macrophages, microglia, and, potentially, other CD4-low cells which all appear to have a shortened in vivo lifespan. We believe these findings have important implications for HIV eradication studies. PMID:25356757

  8. The macrophages in rheumatic diseases

    PubMed Central

    Laria, Antonella; Lurati, Alfredomaria; Marrazza, Mariagrazia; Mazzocchi, Daniela; Re, Katia Angela; Scarpellini, Magda

    2016-01-01

    Macrophages belong to the innate immune system giving us protection against pathogens. However it is known that they are also involved in rheumatic diseases. Activated macrophages have two different phenotypes related to different stimuli: M1 (classically activated) and M2 (alternatively activated). M1 macrophages release high levels of pro-inflammatory cytokines, reactive nitrogen and oxygen intermediates killing microorganisms and tumor cells; while M2 macrophages are involved in resolution of inflammation through phagocytosis of apoptotic neutrophils, reduced production of pro-inflammatory cytokines, and increased synthesis of mediators important in tissue remodeling, angiogenesis, and wound repair. The role of macrophages in the different rheumatic diseases is different according to their M1/M2 macrophages phenotype. PMID:26929657

  9. The (alpha2-->8)-linked polysialic acid capsule of group B Neisseria meningitidis modifies multiple steps during interaction with human macrophages.

    PubMed Central

    Read, R C; Zimmerli, S; Broaddus, C; Sanan, D A; Stephens, D S; Ernst, J D

    1996-01-01

    Group B Neisseria meningitidis causes systemic disease, including meningitis, after initial colonization and subsequent penetration of nasopharyngeal mucosa, a tissue which is richly populated by macrophages. In an initial effort to characterize the interaction of N. meningitidis and mature human macrophages, the influence of the alpha2-->8) -linked polysialic acid capsule on the interaction of N. meningitidis with human monocyte-derived macrophages was investigated with a capsulate case isolate and an isogenic Tn916-derived noncapsulate transformant. The capsulate strain was fourfold less adherent to the macrophage surface after cold incubation, although adherence of both strains was significantly increased after opsonization with nonimmune C5-depleted serum. When opsonized inocula were adjusted so that they adhered to macrophages in equal numbers, the two strains were internalized at equivalent rates and both entered membrane-bound compartments (phagosomes). Colocalization of bacteria with the late endosomal and lysosomal marker lysosome-associated membrane protein revealed that fusion of lysosomes with phagosomes containing the capsulate organism was significantly reduced 10 and 30 min after entry, but by 1 h, no difference between the strains was observed. Once internalized, meningococci were effectively killed, although more rapid killing of the capsulate strain was observed over the first 3 h. These results indicate that the (alpha2-->8)-linked polysialic acid capsule modifies the interaction of meningococci with human macrophages at multiple steps, including adherence to the macrophage surface and phagosome-lysosome fusion. Moreover, the discordance between the kinetics of phagosome- lysosome fusion and bacterial killing suggests that a nonlysosomal mechanism may be responsible for a significant fraction of macrophage killing of N. meningitidis. PMID:8757855

  10. Growth regulation by macrophages

    SciTech Connect

    Wharton, W.; Walker, E.; Stewart, C.C.

    1982-01-01

    The evidence reviewed here indicates that macrophages, either acting alone or in concert with other cells, influence the proliferation of multiple types of cells. Most of the data indicate that these effects are mediated by soluble macrophage-elaborated products (probably proteins) although the role of direct cell-to-cell contacts cannot be ruled out in all cases. A degree of success has been achieved on the biochemical characterization of these factors, due mainly to their low specific activity in conditioned medium and the lack of rapid, specific assays. Understanding the growth-regulating potential of macrophages is an important and needed area of research.

  11. Wormhole Travel for Macrophages.

    PubMed

    Okabe, Yasutaka; Medzhitov, Ruslan

    2016-04-21

    Leukocyte recruitment is generally achieved by rapid migration of inflammatory cells out of circulation, through modified blood vessels, and into affected tissues. Now, Wang and Kubes show that macrophages can be rapidly recruited from body cavities to the liver, via a non-vascular route, where they help to coordinate tissue repair.

  12. Haemophilus ducreyi Inhibits Phagocytosis by U-937 Cells, a Human Macrophage-Like Cell Line

    PubMed Central

    Wood, Gwendolyn E.; Dutro, Susan M.; Totten, Patricia A.

    2001-01-01

    Haemophilus ducreyi is a gram-negative obligate human pathogen that causes the genital ulcer disease chancroid. Chancroid lesions are deep necrotic ulcers with an immune cell infiltrate that includes macrophages. Despite the presence of these phagocytic cells, chancroid ulcers can persist for months and live H. ducreyi can be isolated from these lesions. To analyze the interaction of H. ducreyi with macrophages, we investigated the ability of H. ducreyi strain 35000 to adhere to, invade, and survive within U-937 cells, a human macrophage-like cell line. We found that although H. ducreyi strain 35000 adhered efficiently to U-937 cells, few bacteria were internalized, suggesting that H. ducreyi avoids phagocytosis by human macrophages. The few bacteria that were phagocytosed in these experiments were rapidly killed. We also found that H. ducreyi inhibits the phagocytosis of a secondary target (opsonized sheep red blood cells). Antiphagocytic activity was found in logarithmic, stationary-phase, and plate-grown cultures and was associated with whole, live bacteria but not with heat-killed cultures, sonicates, or culture supernatants. Phagocytosis was significantly inhibited after a 15-min exposure to H. ducreyi, and a multiplicity of infection of approximately 1 CFU per macrophage was sufficient to cause a significant reduction in phagocytosis by U-937 cells. Finally, all of nine H. ducreyi strains tested were antiphagocytic, suggesting that this is a common virulence mechanism for this organism. This finding suggests a mechanism by which H. ducreyi avoids killing and clearance by macrophages in chancroid lesions and inguinal lymph nodes. PMID:11447144

  13. Planning a dynamic kill

    SciTech Connect

    Abel, L.W.

    1996-05-01

    This article discusses the methodology, design philosophy, and guidelines for planning a dynamic-kill operation for a wild well. The topics covered are two methods of computer analysis for designing dynamic-kill requirements, the design process, determining the pumping spread, and the pitfalls that a designer faces in planning a dynamic kill.

  14. IFN-gamma enhances killing of methicillin-resistant Staphylococcus aureus by human monocytes more effectively than GM-CSF in the presence of daptomycin and other antibiotics.

    PubMed

    Smith, Raymond P; Baltch, Aldona L; Ritz, William J; Michelsen, Phyllis B; Bopp, Lawrence H

    2010-09-01

    Because cytokines have been utilized in treatment of sepsis in neonates, we studied the effects of interferon-gamma (IFN-gamma) and GM-CSF on killing of intracellular methicilin-resistant Staphylococcus aureus (MRSA) by human monocyte derived macrophages (MDM) in the presence of daptomycin (Dap), rifampin (Rif), gentamicin (Gen), and combinations of these drugs. MDM infected with MRSA were treated with Dap (1 x MIC), Gen (0.5 x MIC), or Rif (1 x MIC), singly or in combination, with or without cytokines. MDM were lysed and viable bacteria counted. With antibiotics, MDM activated by IFN-gamma had a more rapid and prolonged bacterial killing effect than MDM activated by GM-CSF. This effect was most obvious with the triple-drug combination. In contrast, GM-CSF reduced intracellular killing under most experimental conditions compared to the effect of antibiotics alone. Dap alone and two- and three-drug combinations demonstrated significant killing effect for the 48 h of the assay. IFN-gamma enhanced rapid intracellular killing of MRSA in the presence of triple-drug treatment or Dap alone. GM-CSF in combination with the antibiotics reduced killing under most conditions studied. Further studies to confirm these observations with IFN-gamma-activated MDM and other MRSA strains are needed to support clinical trials for difficult-to-treat MRSA infections.

  15. Augmentation of Human Macrophage Candidacidal Capacity by Recombinant Human Myeloperoxidase and GranulocyteMacrophage Colony-Stimulating Factor

    PubMed Central

    Maródi, László; Tournay, Christophe; Káposzta, Rita; Johnston, Richard B.; Moguilevsky, Nicole

    1998-01-01

    Phagocyte myeloperoxidase (MPO) is believed to be particularly important in defense against candida infection. We reported earlier that monocytes, rich in MPO, killed Candida albicans at a significantly higher rate and extent than did monocyte-derived macrophages, known to lack MPO, and that C. albicans is less resistant to MPO-dependent oxidants than less pathogenic Candida species. We hypothesized, therefore, that the capacity of macrophages to kill C. albicans might be improved in the presence of MPO. In this study, we evaluated the ability of recombinant human MPO (rhMPO) to augment the killing of C. albicans by resident macrophages and macrophages activated by recombinant human granulocyte-macrophage colony-stimulating factor. Addition of rhMPO (concentration range, 0.8 to 6.4 U/ml) to suspensions of resident and activated macrophages and opsonized C. albicans resulted in concentration-dependent and significant increases in candida killing. This enhancement was particularly pronounced with activated macrophages, whether C. albicans was opsonized or unopsonized and ingested through the macrophage mannose receptor. rhMPO did not affect the killing of C. albicans by monocytes, nor did it affect phagocytosis of opsonized or unopsonized C. albicans. These results indicate that exogenous rhMPO can augment the candidacidal capacity of both resident and activated macrophages, with a more profound effect on activated cells. We suggest that rhMPO may be effective in the treatment of invasive candidiasis. PMID:9596743

  16. In vivo bronchoalveolar macrophage defense against Rhizopus oryzae and Aspergillus fumigatus.

    PubMed

    Waldorf, A R; Levitz, S M; Diamond, R D

    1984-11-01

    The ability of bronchoalveolar macrophages from normal, diabetic, and cortisone-treated mice to inhibit spore germination and kill fungal spores in vivo was investigated. The data indicated that the normal host controls different fungal infections in the lungs by different mechanisms. Prevention of mucormycosis required inhibition of fungal spore germination by alveolar macrophages. In contrast, pulmonary defense against aspergillosis depended on early killing of conidia by alveolar macrophages and not on inhibition of germination by bronchoalveolar macrophages. Bronchoalveolar macrophages in diabetic and cortisone-treated animals allowed fungal spore germination, thereby permitting infection by Rhizopus oryzae. In the cortisone-treated mouse, bronchoalveolar macrophages did not kill fungal conidia and progressive infection by Aspergillus fumigatus occurred. Fungicidal activity of bronchoalveolar macrophages was measured with a new in vivo killing assay.

  17. DETC Induces Leishmania Parasite Killing in Human In Vitro and Murine In Vivo Models: A Promising Therapeutic Alternative in Leishmaniasis

    PubMed Central

    Khouri, Ricardo; Novais, Fernanda; Santana, Gisélia; de Oliveira, Camila Indiani; Vannier dos Santos, Marcos André; Barral, Aldina; Barral-Netto, Manoel; Van Weyenbergh, Johan

    2010-01-01

    Background Chemotherapy remains the primary tool for treatment and control of human leishmaniasis. However, currently available drugs present serious problems regarding side-effects, variable efficacy, and cost. Affordable and less toxic drugs are urgently needed for leishmaniasis. Methodology/Principal Findings We demonstrate, by microscopy and viability assays, that superoxide dismutase inhibitor diethyldithiocarbamate (DETC) dose-dependently induces parasite killing (p<0.001) and is able to “sterilize” Leishmania amazonensis infection at 2 mM in human macrophages in vitro. We also show that DETC-induced superoxide production (p<0.001) and parasite destruction (p<0.05) were reverted by the addition of the antioxidant N-acetylcysteine, indicating that DETC-induced killing occurs through oxidative damage. Furthermore, ultrastructural analysis by electron microscopy demonstrates a rapid and highly selective destruction of amastigotes in the phagosome upon DETC treatment, without any apparent damage to the host cell, including its mitochondria. In addition, DETC significantly induced parasite killing in Leishmania promastigotes in axenic culture. In murine macrophages infected with Leishmania braziliensis, DETC significantly induced in vitro superoxide production (p = 0.0049) and parasite killing (p = 0.0043). In vivo treatment with DETC in BALB/C mice infected with Leishmania braziliensis caused a significant decrease in lesion size (p<0.0001), paralleled by a 100-fold decrease (p = 0.0087) in parasite burden. Conclusions/Significance Due to its strong leishmanicidal effect in human macrophages in vitro, its in vivo effectiveness in a murine model, and its previously demonstrated in vivo safety profile in HIV treatment, DETC treatment might be considered as a valuable therapeutic option in human leishmaniasis, including HIV/Leishmania co-infection. PMID:21200432

  18. Acute cysticercosis favours rapid and more severe lesions caused by Leishmania major and Leishmania mexicana infection, a role for alternatively activated macrophages.

    PubMed

    Rodríguez-Sosa, Miriam; Rivera-Montoya, Irma; Espinoza, Arlett; Romero-Grijalva, Miriam; López-Flores, Roberto; González, Jorge; Terrazas, Luis I

    2006-08-01

    Parasitic helminths have developed complex mechanisms to modulate host immunity. In the present study we found that previous infection of mice with the cestode Taenia crassiceps favours parasitemia and induces larger cutaneous lesions during both Leishmania major and Leishmania mexicana co-infections. Analysis of cytokine responses into draining lymph nodes indicated that co-infection of T. crassiceps-Leishmania did not inhibit IFN-gamma production in response to Leishmania antigens, but significantly increased IL-4 production. Additionally, anti-Leishmania-specific IgG1 antibodies and total IgE increased in co-infected mice, whereas, IgG2a titers remained similar. Macrophages from Taenia-infected mice displayed increased mRNA transcripts of arginase-1, Ym1, and Mannose Receptor, as well as greater production of urea (all markers for an alternate activation state) compared to macrophages from Leishmania-infected mice. In contrast, lower mRNA transcripts for IL-12p35, IL-12p40, IL-23p19, and iNOS were detected in macrophages obtained from cestode-infected mice compared to uninfected and Leishmania-infected mice after LPS stimulation. The presence of cestode also generated impaired macrophage anti-leishmanicidal activity in vitro, as evidenced by the inability of these macrophages to prevent Leishmania growth compared to macrophages from uninfected mice. This was observed despite the fact that both groups of cells were exposed to IFN-gamma. Flow cytometry showed high IFN-gammaR expression on Taenia-induced macrophages. Thus, lack of response to IFN-gamma is not associated with the absence of its receptor. Our data suggest that cestode infection may favour Leishmania installation by inducing alternatively activated macrophages rather than inhibiting Th1-type responses.

  19. Ion-kill dosimetry

    NASA Technical Reports Server (NTRS)

    Katz, R.; Cucinotta, F. A.; Fromm, M.; Chambaudet, A.

    2001-01-01

    Unanticipated late effects in neutron and heavy ion therapy, not attributable to overdose, imply a qualitative difference between low and high LET therapy. We identify that difference as 'ion kill', associated with the spectrum of z/beta in the radiation field, whose measurement we label 'ion-kill dosimetry'.

  20. How neutrophils kill fungi.

    PubMed

    Gazendam, Roel P; van de Geer, Annemarie; Roos, Dirk; van den Berg, Timo K; Kuijpers, Taco W

    2016-09-01

    Neutrophils play a critical role in the prevention of invasive fungal infections. Whereas mouse studies have demonstrated the role of various neutrophil pathogen recognition receptors (PRRs), signal transduction pathways, and cytotoxicity in the murine antifungal immune response, much less is known about the killing of fungi by human neutrophils. Recently, novel primary immunodeficiencies have been identified in patients with a susceptibility to fungal infections. These human 'knock-out' neutrophils expand our knowledge to understand the role of PRRs and signaling in human fungal killing. From the studies with these patients it is becoming clear that neutrophils employ fundamentally distinct mechanisms to kill Candida albicans or Aspergillus fumigatus. PMID:27558342

  1. Passive transfer of leishmania lipopolysaccharide confers parasite survival in macrophages

    SciTech Connect

    Handman, E.; Schnur, L.F.; Spithill, T.W.; Mitchell, G.F.

    1986-12-01

    Infection of macrophages by the intracellular protozoan parasite Leishmania involves specific attachment to the host membrane, followed by phagocytosis and intracellular survival and growth. Two parasite molecules have been implicated in the attachment event: Leishmania lipopolysaccharide (L-LPS) and a glycoprotein (gp63). This study was designed to clarify the role of L-LPS in infection and the stage in the process of infection at which it operates. The authors have recently identified a Leishmania major strain (LRC-L119) which lacks the L-LPS molecule and is not infective for hamsters or mice. This parasite was isolated from a gerbil in Kenya and was identified phenotypically as L. major by isoenzyme and fatty acid analysis. In this study they have confirmed at the genotype level that LRC-L119 is L. major by analyzing and comparing the organization of cloned DNA sequences in the genome of different strains of L. major. Here they show that LRC-L119 promastigotes are phagocytosed rapidly by macrophages in vitro, but in contrast to virulent strains of L. major, they are then killed over a period of 18 hr. In addition, they show that transfer of purified L-LPS from a virulent clone of L. major (V121) into LRC-L119 promastigotes confers on them the ability to survive in macrophages in vitro.

  2. Expression of functions by normal sheep alveolar macrophages and their alteration by interaction with Mycoplasma ovipneumoniae.

    PubMed

    Niang, M; Rosenbusch, R F; Lopez-Virella, J; Kaeberle, M L

    1997-10-31

    Normal sheep alveolar macrophages collected by bronchial lavage were exposed to live or heat-killed Mycoplasma ovipneumoniae organisms, and their capability to ingest Staphylococcus aureus and to elicit antibody-dependent cellular cytotoxicity against sensitized chicken red blood cells was tested. Controls consisted of non-infected macrophages in M199 medium. In addition, the effect of M. ovipneumoniae on expression of surface molecules on these sheep alveolar macrophages was determined. The percentage of S. aureus ingested by nontreated sheep alveolar macrophages was significantly higher than that of infected macrophages. Live mycoplasmas were more effective in suppressing the ingestion of S. aureus by these macrophages than killed mycoplasmas. Both live and killed mycoplasmas suppressed the cytolytic effect of the sheep alveolar macrophages to a similar degree. About 78% and 45% of the normal sheep alveolar macrophages had IgG and complement receptors, respectively. Infection of these macrophages with M. ovipneumoniae decreased significantly the expression of IgG receptors but had no effects on complement receptors. There were substantial increases in the expression of both MHC class I and class II by the mycoplasma-induced macrophages as compared with unstimulated macrophages. Live mycoplasmas were more effective in inducing expression of both classes than killed mycoplasmas. The results, taken together, suggest that M. ovipneumoniae induced alterations in macrophage activities and this may be a contributing factor in the pathogenesis of respiratory disease induced by the organism.

  3. Phagocytosis of Staphylococcus aureus by bovine mammary gland macrophages and intracellular protection from antibiotic action in vitro and in vivo.

    PubMed

    Craven, N; Anderson, J C

    1984-11-01

    Macrophages isolated from the involuted bovine mammary gland were cultured in vitro. Phagocytosis of opsonized Staphylococcus aureus occurred rapidly, but intracellular killing of bacteria was slow. Many intracellular staphylococci survived for up to 4 d exposure to extracellular cloxacillin and emerged from within the macrophages to multiply extracellularly when the antibiotic was inactivated. Rifampicin was significantly more efficient than cloxacillin in killing intracellular S. aureus after 18 h incubation, but it too failed to sterilize the cultures within 3 d. Staphylococci, which had remained viable within macrophages during 20 h incubation with extracellular cloxacillin, showed an increased sensitivity to dilute lysostaphin on subsequent exposure. A 3 d course of intramammary therapy with cloxacillin, commencing simultaneously with an infecting inoculum of approximately 10(8) colony forming units (c.f.u.) S. aureus, apparently eliminated the infection from one quarter of the udders of each of three lactating cows, but bacteria were re-isolated from two cows after a delay of several days. However, when other quarters of the same cows were infected with approximately 10(8) c.f.u. S. aureus which had been phagocytosed by autologous mammary macrophages, similar simultaneous antibiotic therapy failed to affect these infections. The in vitro and in vivo findings indicate the significance of intracellular survival of S. aureus as a factor contributing to failure of antibiotic therapy.

  4. Mannheimia haemolytica and Its Leukotoxin Cause Macrophage Extracellular Trap Formation by Bovine Macrophages

    PubMed Central

    Aulik, Nicole A.; Hellenbrand, Katrina M.

    2012-01-01

    Human and bovine neutrophils release neutrophil extracellular traps (NETs), which are protein-studded DNA matrices capable of extracellular trapping and killing of pathogens. Recently, we reported that bovine neutrophils release NETs in response to the important respiratory pathogen Mannheimia haemolytica and its leukotoxin (LKT). Here, we demonstrate macrophage extracellular trap (MET) formation by bovine monocyte-derived macrophages exposed to M. haemolytica or its LKT. Both native fully active LKT and noncytolytic pro-LKT (produced by an lktC mutant of M. haemolytica) stimulated MET formation. Confocal and scanning electron microscopy revealed a network of DNA fibrils with colocalized histones in extracellular traps released from bovine macrophages. Formation of METs required NADPH oxidase activity, as previously demonstrated for NET formation. METs formed in response to LKT trapped and killed a portion of the M. haemolytica cells. Bovine alveolar macrophages, but not peripheral blood monocytes, also formed METs in response to M. haemolytica cells. MET formation was not restricted to bovine macrophages. We also observed MET formation by the mouse macrophage cell line RAW 264.7 and by human THP-1 cell-derived macrophages, in response to Escherichia coli hemolysin. The latter is a member of the repeats-in-toxin (RTX) toxin family related to the M. haemolytica leukotoxin. This study demonstrates that macrophages, like neutrophils, can form extracellular traps in response to bacterial pathogens and their exotoxins. PMID:22354029

  5. Mannheimia haemolytica and its leukotoxin cause macrophage extracellular trap formation by bovine macrophages.

    PubMed

    Aulik, Nicole A; Hellenbrand, Katrina M; Czuprynski, Charles J

    2012-05-01

    Human and bovine neutrophils release neutrophil extracellular traps (NETs), which are protein-studded DNA matrices capable of extracellular trapping and killing of pathogens. Recently, we reported that bovine neutrophils release NETs in response to the important respiratory pathogen Mannheimia haemolytica and its leukotoxin (LKT). Here, we demonstrate macrophage extracellular trap (MET) formation by bovine monocyte-derived macrophages exposed to M. haemolytica or its LKT. Both native fully active LKT and noncytolytic pro-LKT (produced by an lktC mutant of M. haemolytica) stimulated MET formation. Confocal and scanning electron microscopy revealed a network of DNA fibrils with colocalized histones in extracellular traps released from bovine macrophages. Formation of METs required NADPH oxidase activity, as previously demonstrated for NET formation. METs formed in response to LKT trapped and killed a portion of the M. haemolytica cells. Bovine alveolar macrophages, but not peripheral blood monocytes, also formed METs in response to M. haemolytica cells. MET formation was not restricted to bovine macrophages. We also observed MET formation by the mouse macrophage cell line RAW 264.7 and by human THP-1 cell-derived macrophages, in response to Escherichia coli hemolysin. The latter is a member of the repeats-in-toxin (RTX) toxin family related to the M. haemolytica leukotoxin. This study demonstrates that macrophages, like neutrophils, can form extracellular traps in response to bacterial pathogens and their exotoxins. PMID:22354029

  6. Why is particulate matter produced by wildfires toxic to lung macrophages?

    PubMed

    Franzi, Lisa M; Bratt, Jennifer M; Williams, Keisha M; Last, Jerold A

    2011-12-01

    The mechanistic basis of the high toxicity to lung macrophages of coarse PM from the California wildfires of 2008 was examined in cell culture experiments with mouse macrophages. Wildfire PM directly killed macrophages very rapidly in cell culture at relatively low doses. The wildfire coarse PM is about four times more toxic to macrophages on an equal weight basis than the same sized PM collected from normal ambient air (no wildfires) from the same region and season. There was a good correlation between the extent of cytotoxicity and the amount of oxidative stress observed at a given dose of wildfire PM in vitro. Our data implicate NF-κB signaling in the response of macrophages to wildfire PM, and suggest that most, if not all, of the cytotoxicity of wildfire PM to lung macrophages is the result of oxidative stress. The relative ratio of toxicity and of expression of biomarkers of oxidant stress between wildfire PM and "normal" PM collected from ambient air is consistent with our previous results in mice in vivo, also suggesting that most, if not all, of the cytotoxicity of wildfire PM to lung macrophages is the result of oxidative stress. Our findings from this and earlier studies suggest that the active components of coarse PM from the wildfire are heat-labile organic compounds. While we cannot rule out a minor role for endotoxin in coarse PM preparations from the collected wildfire PM in our observed results both in vitro and in vivo, based on experiments using the inhibitor Polymyxin B most of the oxidant stress and pro-inflammatory activity observed was not due to endotoxin.

  7. Killing vectors and anisotropy

    SciTech Connect

    Krisch, J. P.; Glass, E. N.

    2009-08-15

    We consider an action that can generate fluids with three unequal stresses for metrics with a spacelike Killing vector. The parameters in the action are directly related to the stress anisotropies. The field equations following from the action are applied to an anisotropic cosmological expansion and an extension of the Gott-Hiscock cosmic string.

  8. Children Who Kill.

    ERIC Educational Resources Information Center

    Natale, Jo Anna

    1999-01-01

    Two recent books, "When Good Kids Kill," by Michael D. Kelleher, and "Lost Boys," by James Garbarino, explore how children become killers and suggest ways to reduce our high-pressure society's epidemic levels of youth violence. Physically or psychologically distant parents and unaffirmative media messages are negative influences. (MLH)

  9. Redshifts and Killing vectors

    NASA Astrophysics Data System (ADS)

    Harvey, Alex; Schucking, Engelbert; Surowitz, Eugene J.

    2006-11-01

    Current approaches to physics stress the importance of conservation laws due to spacetime and internal symmetries. In special and general relativity the generators of these symmetries are known as Killing vectors. We use them for the rigorous determination of gravitational and cosmological redshifts.

  10. Direct imaging of macrophage activation during PDT treatment

    NASA Astrophysics Data System (ADS)

    Song, Sheng; Zhou, Feifan; Chen, Wei R.; Xing, Da

    2011-11-01

    Mounting evidence describes a more complex progress of macrophage activation during photodynamic therapy (PDT), which performing distinct immunological functions and different physiologies on surrounding cells and tissues. Macrophage-targeted PDT has been applied in the selective killing of cells involved in inflammation and tumor. We have previously shown that PDT-mediated tumor cells apoptosis can induce a higher level immune response than necrosis, and enhance the macrophage activation. However, the molecular mechanism of macrophage activation during PDT-induced apoptotic cells (AC) still unclear. Here, we use confocal microscopy to image the phagocytosis of tumor cells by macrophages. We also observed that PDT-treated AC can activate Toll-like receptors (TLRs) which are present on macrophages surface. Besides, the increase in nitric oxide (NO) formation in macrophages was detected in real time by a laser scanning microscopy. This study provided more details for understanding the molecular mechanism of the immune response induced by PDT-treated AC.

  11. Direct imaging of macrophage activation during PDT treatment

    NASA Astrophysics Data System (ADS)

    Song, Sheng; Zhou, Feifan; Chen, Wei R.; Xing, Da

    2012-03-01

    Mounting evidence describes a more complex progress of macrophage activation during photodynamic therapy (PDT), which performing distinct immunological functions and different physiologies on surrounding cells and tissues. Macrophage-targeted PDT has been applied in the selective killing of cells involved in inflammation and tumor. We have previously shown that PDT-mediated tumor cells apoptosis can induce a higher level immune response than necrosis, and enhance the macrophage activation. However, the molecular mechanism of macrophage activation during PDT-induced apoptotic cells (AC) still unclear. Here, we use confocal microscopy to image the phagocytosis of tumor cells by macrophages. We also observed that PDT-treated AC can activate Toll-like receptors (TLRs) which are present on macrophages surface. Besides, the increase in nitric oxide (NO) formation in macrophages was detected in real time by a laser scanning microscopy. This study provided more details for understanding the molecular mechanism of the immune response induced by PDT-treated AC.

  12. Effect of alveolar macrophages on Toxoplasma gondii.

    PubMed Central

    Ryning, F W; Remington, J S

    1977-01-01

    As pulmonary involvement can occur in disseminated toxoplasmosis in immunosuppressed patients, studies were initiated to define local mechanisms of resistance of the lung to Toxoplasma gondii. Alveolar macrophages were obtained from normal mice and mice chronically infected with T. gondii by bronchopulmonary lavage and cultured in vitro. Although normal alveolar macrophages were difficult to infect with Toxoplasma, they supported intracellular multiplication of this organism. When exposed to Toxoplasma that had been pretreated with heat-inactivated serum containing specific antibody, the number of intracellular organisms increased remarkably, and the macrophages destroyed the coated parasites. After development of chronic infections with Toxoplasma, there was a transient period during which a striking increase in numbers of alveolar macrophages was observed in lavage specimens. These macrophages differed from those of normal alveolar macrophages. There was a greater percentage of large cells, a greater tendency to spread on glass, and an increased number of intracellular Toxoplasma, and the cells were activated to kill or inhibit multiplication of the parasite. During the period when activated macrophages were demonstrable in bronchopulmonary washings, histological changes in the lungs revealed a marked mononuclear cell infiltrate. These studies support a role for the activated alveolar macrophage as an effector in resistance of the lung to infection with Toxoplasma. PMID:591065

  13. Homegrown Macrophages.

    PubMed

    Kim, Ki-Wook; Zhang, Nan; Choi, Kyunghee; Randolph, Gwendalyn J

    2016-09-20

    Macrophages residing in different organs have diverse gene-expression programs. Mass et al. (2016) propose that this diversity develops "at home"-within those organs-after the recruitment of a common precursor that had not made prior commitments to diversity. PMID:27653599

  14. Inability of tumour cells to elicit the respiratory burst in cytotoxic, activated macrophages.

    PubMed Central

    Bryant, S M; Hill, H R

    1982-01-01

    Activated macrophages from Corynebacterium parvum-treated mice are cytotoxic to non-antibody-coated tumour cells and have an augmented respiratory burst potential when compared to resident macrophages. We have investigated the possible involvement of the respiratory burst as an effector mechanism in this type of tumour killing. Scavengers of toxic metabolites of oxygen such as catalase, superoxide dismutase, 2,3-dihydroxybenzoate, ethanol, and cytochrome c did not inhibit macrophage cytotoxicity in this system. To investigate whether or not neoplastic cells stimulate the macrophage respiratory burst, we exposed activated macrophages to viable tumour cells and monitored macrophage superoxide anion production, chemiluminescence, and hexose monophosphate shunt activity. None of these indicators of the macrophage respiratory burst was stimulated by the tumour cells towards which the macrophages were cytotoxic. The data suggest that the macrophages burst is not utilized as an effector mechanism in the non-antibody-mediated macrophage tumour cytotoxicity reaction. PMID:6277777

  15. Role of Macrophage Scavenger Receptors in Response to Listeria monocytogenes Infection in Mice

    PubMed Central

    Ishiguro, Takuro; Naito, Makoto; Yamamoto, Takashi; Hasegawa, Go; Gejyo, Fumitake; Mitsuyama, Masao; Suzuki, Hiroshi; Kodama, Tatsuhiko

    2001-01-01

    Type I and type II macrophage scavenger receptors (SR-A I/II) recognize a variety of polyanions including bacterial cell-wall products such as lipopolysaccharide, suggesting a role for SR-A I/II in immunity against bacterial infection. SR-A I/II-deficient (MSR-A−/−) mice were more susceptible to infection with listeriolysin-O (LLO)-producing Listeria monocytogenes. After infection, Kupffer cells in wild-type (MSR-A+/+) mice phagocytized larger numbers of Listeria than those in MSR-A−/− mice. The number and the diameter of hepatic granulomas were larger in MSR-A−/− mice than MSR-A+/+ mice. L. monocytogenes replicated at higher levels in the liver of MSR-A−/− mice compared with MSR-A+/+ mice, and macrophages from MSR-A−/− mice showed impaired ability to kill Listeria in vitro. However, macrophages from MSR-A+/+ and MSR-A−/− mice showed similar levels of listericidal activity against isogenic mutant L. monocytogenes with an inactivated LLO gene. The listerial phagocytic activities of MSR-A+/+ macrophages treated with an anti-SR-A I/II antibody (2F8) and MSR-A−/− macrophages were significantly impaired compared with untreated MSR-A+/+ macrophages, indicating that SR-A I/II function as a receptor for L. monocytogenes. Electron microscopy revealed that most L. monocytogenes had been eliminated from the lysosomes of MSR-A+/+ macrophages in vivo and in vitro. In contrast, L. monocytogenes rapidly lysed the phagosomal membrane and escaped to the cytosol in MSR-A−/− macrophages and in MSR-A+/+ macrophages treated with 2F8 before phagosome-lysosome fusion. These findings imply that SR-A I/II plays a crucial role in host defense against listerial infection not only by functioning as a receptor but also by mediating listericidal mechanisms through the regulation of LLO-dependent listerial escape from the macrophages. PMID:11141491

  16. Cytolytic activity against tumor cells by macrophage cell lines and augmentation by macrophage stimulants.

    PubMed

    Taniyama, T; Holden, H T

    1980-07-15

    Previous studies have shown that macrophage cell lines retained the ability to phagocytize, to secrete lysosomal enzymes, and to function as effector cells in antibody-dependent cellular cytoxicity. In this paper, the cytolytic activity of murine macrophage cell lines against tumor target cells was assessed using an 18-h 51Cr release assay. Of the macrophage cell lines tested, RAW 264, PU5-1.8 and IC-21 had intermediate to high levels of spontaneous cytolytic activity, P388D, and J774 had low to intermediate levels, while /WEHI-3 showed little or no cytolytic activity against RBL-5, MBL-2 and TU-5 target cells. Tumor-cell killing by macrophage cell lines could be augmented by the addition of macrophage stimulants, such as bacterial lipopolysaccharide and poly I:C, indicating that the activation of macrophages by these stimulants does not require the participation of other cell types. Treatment with interferon also augmented the tumor-cell killing by macrophage cell lines. Although the mechanism by which these cell lines exert their spontaneous or boosted cytotoxic activity is not clear, it does not appear to be due to depletion of nutrients since cell lines with high metabolic and proliferative activities, such as WEHI-3 and RBL-5, showed little or no cytotoxicity and supernatants from the macrophage cell lines did not exert any cytotoxic effects in their essay. Thus, it appears that the different macrophage cell lines represent different levels of activation and/or differentiation and may be useful for studying the development of these processes as well as providing a useful tool for analyzing the mechanisms of macrophage-mediated cytolysis. PMID:6165690

  17. Generation and Characterization of Mouse Regulatory Macrophages.

    PubMed

    Carretero-Iglesia, Laura; Hill, Marcelo; Cuturi, Maria Cristina

    2016-01-01

    In the last years, cell therapy has become a promising approach to therapeutically manipulate immune responses in autoimmunity, cancer, and transplantation. Several types of lymphoid and myeloid cells origin have been generated in vitro and tested in animal models. Their efficacy to decrease pharmacological treatment has successfully been established. Macrophages play an important role in physiological and pathological processes. They represent an interesting cell population due to their high plasticity in vivo and in vitro. Here, we describe a protocol to differentiate murine regulatory macrophages in vitro from bone marrow precursors. We also describe several methods to assess macrophage classical functions, as their bacterial killing capacity and antigen endocytosis and degradation. Importantly, regulatory macrophages also display suppressive characteristics, which are addressed by the study of their hypostimulatory T lymphocyte capacity and polyclonal T lymphocyte activation suppression.

  18. Lipopolysaccharide rapidly modifies adenosine receptor transcripts in murine and human macrophages: role of NF-kappaB in A(2A) adenosine receptor induction.

    PubMed

    Murphree, Lauren J; Sullivan, Gail W; Marshall, Melissa A; Linden, Joel

    2005-11-01

    The A(2A) adenosine receptor (A(2A)AR) mediates anti-inflammatory actions of adenosine in a variety of cell types. LPS (lipopolysaccharide) was reported to induce a small (<2-fold) increase in the expression of A(2A)AR mRNA in human monocytes and monocytic cell lines. We investigated the effects of LPS on the expression of adenosine receptor mRNAs in primary mouse IPMPhi (intraperitoneal macrophages), human macrophages and Wehi-3 cells. Treatment with 10 ng/ml LPS for 4 h produced a >100-fold increase in A(2A)AR mRNA. LPS-induced increases in mRNA for A(2A)AR and TNFalpha (tumour necrosis factor alpha) are reduced by 90% in IPMPhi pretreated with the NF-kappaB (nuclear factor kappaB) inhibitor, BAY 11-7082 {(E)3-[(4-methylphenyl)sulphonyl]-2-propenenitrile; 10 microM}. In Wehi-3 cells exposed to LPS, A(2A)AR and A(2B)AR transcripts are elevated by 290- and 10-fold respectively, the A(1)AR transcript is unchanged and the A(3)AR transcript is decreased by 67%. The induction of A(2A)AR mRNA by LPS is detectable after 1 h, reaches a peak at 6 h at 600 times control and remains elevated beyond 24 h. The ED50 (effective dose) of LPS is 2.3 ng/ml. A(2A)AR receptor number, measured by 125I-ZM241385 binding to whole cells, is undetectable in naïve cells and increases linearly at a rate of 23 receptors x cell(-1) x min(-1) to a B(max) of 348 fmol/mg (28000 receptors/cell) in 20 h. The increase in receptor number is correlated with an increase in the potency of an A(2A) agonist (4-{3-[6-amino-9-(5-ethylcarbamoyl-3,4-dihydroxy-tetrahydro-furan-2-yl)-9H-purin-2-yl]-prop-2-ynyl}-cyclohexanecarboxylic acid methyl ester; referred to as ATL146e) to stimulate cAMP in these cells. After LPS pretreatment, the potency of the A(2A) agonist, ATL146e, to reduce TNFalpha release from IPMPhi was increased by 200-fold. The results support the hypothesis that regulation of adenosine receptor expression, especially up-regulation of the A(2A)AR, is part of a delayed feedback mechanism

  19. Antagonism of miR-328 Increases the Antimicrobial Function of Macrophages and Neutrophils and Rapid Clearance of Non-typeable Haemophilus Influenzae (NTHi) from Infected Lung

    PubMed Central

    Tay, Hock L.; Kaiko, Gerard E.; Plank, Maximilian; Li, JingJing; Maltby, Steven; Essilfie, Ama-Tawiah; Jarnicki, Andrew; Yang, Ming; Mattes, Joerg; Hansbro, Philip M.; Foster, Paul S.

    2015-01-01

    Pathogenic bacterial infections of the lung are life threatening and underpin chronic lung diseases. Current treatments are often ineffective potentially due to increasing antibiotic resistance and impairment of innate immunity by disease processes and steroid therapy. Manipulation miRNA directly regulating anti-microbial machinery of the innate immune system may boost host defence responses. Here we demonstrate that miR-328 is a key element of the host response to pulmonary infection with non-typeable haemophilus influenzae and pharmacological inhibition in mouse and human macrophages augments phagocytosis, the production of reactive oxygen species, and microbicidal activity. Moreover, inhibition of miR-328 in respiratory models of infection, steroid-induced immunosuppression, and smoke-induced emphysema enhances bacterial clearance. Thus, miRNA pathways can be targeted in the lung to enhance host defence against a clinically relevant microbial infection and offer a potential new anti-microbial approach for the treatment of respiratory diseases. PMID:25894560

  20. Surface structure influences contact killing of bacteria by copper

    PubMed Central

    Zeiger, Marco; Solioz, Marc; Edongué, Hervais; Arzt, Eduard; Schneider, Andreas S

    2014-01-01

    Copper kills bacteria rapidly by a mechanism that is not yet fully resolved. The antibacterial property of copper has raised interest in its use in hospitals, in place of plastic or stainless steel. On the latter surfaces, bacteria can survive for days or even weeks. Copper surfaces could thus provide a powerful accessory measure to curb nosocomial infections. We here investigated the effect of the copper surface structure on the efficiency of contact killing of Escherichia coli, an aspect which so far has received very little attention. It was shown that electroplated copper surfaces killed bacteria more rapidly than either polished copper or native rolled copper. The release of ionic copper was also more rapid from electroplated copper compared to the other materials. Scanning electron microscopy revealed that the bacteria nudged into the grooves between the copper grains of deposited copper. The findings suggest that, in terms of contact killing, more efficient copper surfaces can be engineered. PMID:24740976

  1. Charged conformal Killing spinors

    SciTech Connect

    Lischewski, Andree

    2015-01-15

    We study the twistor equation on pseudo-Riemannian Spin{sup c}-manifolds whose solutions we call charged conformal Killing spinors (CCKSs). We derive several integrability conditions for the existence of CCKS and study their relations to spinor bilinears. A construction principle for Lorentzian manifolds admitting CCKS with nontrivial charge starting from CR-geometry is presented. We obtain a partial classification result in the Lorentzian case under the additional assumption that the associated Dirac current is normal conformal and complete the classification of manifolds admitting CCKS in all dimensions and signatures ≤5 which has recently been initiated in the study of supersymmetric field theories on curved space.

  2. Elevated CO2 selectively inhibits interleukin-6 and tumor necrosis factor expression and decreases phagocytosis in the macrophage

    PubMed Central

    Wang, Naizhen; Gates, Khalilah L.; Trejo, Humberto; Favoreto, Silvio; Schleimer, Robert P.; Sznajder, Jacob I.; Beitel, Greg J.; Sporn, Peter H. S.

    2010-01-01

    Elevated blood and tissue CO2, or hypercapnia, is common in severe lung disease. Patients with hypercapnia often develop lung infections and have an increased risk of death following pneumonia. To explore whether hypercapnia interferes with host defense, we studied the effects of elevated PCO2 on macrophage innate immune responses. In differentiated human THP-1 macrophages and human and mouse alveolar macrophages stimulated with lipopolysaccharide (LPS) and other Toll-like receptor ligands, hypercapnia inhibited expression of tumor necrosis factor and interleukin (IL)-6, nuclear factor (NF)-κB-dependent cytokines critical for antimicrobial host defense. Inhibition of IL-6 expression by hypercapnia was concentration dependent, rapid, reversible, and independent of extracellular and intracellular acidosis. In contrast, hypercapnia did not down-regulate IL-10 or interferon-β, which do not require NF-κB. Notably, hypercapnia did not affect LPS-induced degradation of IκBα, nuclear translocation of RelA/p65, or activation of mitogen-activated protein kinases, but it did block IL-6 promoter-driven luciferase activity in mouse RAW 264.7 macrophages. Elevated PCO2 also decreased phagocytosis of opsonized polystyrene beads and heat-killed bacteria in THP-1 and human alveolar macrophages. By interfering with essential innate immune functions in the macrophage, hypercapnia may cause a previously unrecognized defect in resistance to pulmonary infection in patients with advanced lung disease.—Wang, N., Gates, K. L., Trejo, H., Favoreto, Jr., S., Schleimer, R. P., Sznajder, J. I., Beitel, G. J., Sporn, P. H. S. Elevated CO2 selectively inhibits interleukin-6 and tumor necrosis factor expression and decreases phagocytosis in the macrophage. PMID:20181940

  3. Aging Enhances the Production of Reactive Oxygen Species and Bactericidal Activity in Peritoneal Macrophages by Upregulating Classical Activation Pathways

    SciTech Connect

    Smallwood, Heather S.; López-Ferrer, Daniel; Squier, Thomas C.

    2011-10-07

    Maintenance of macrophages in their basal state and their rapid activation in response to pathogen detection are central to the innate immune system, acting to limit nonspecific oxidative damage and promote pathogen killing following infection. To identify possible age-related alterations in macrophage function, we have assayed the function of peritoneal macrophages from young (3–4 months) and aged (14–15 months) Balb/c mice. In agreement with prior suggestions, we observe age-dependent increases in the extent of recruitment of macrophages into the peritoneum, as well as ex vivo functional changes involving enhanced nitric oxide production under resting conditions that contribute to a reduction in the time needed for full activation of senescent macrophages following exposure to lipopolysaccharides (LPS). Further, we observe enhanced bactericidal activity following Salmonella uptake by macrophages isolated from aged Balb/c mice in comparison with those isolated from young animals. Pathways responsible for observed phenotypic changes were interrogated using tandem mass spectrometry, which identified age-dependent increases in levels of proteins linked to immune cell pathways under basal conditions and following LPS activation. Immune pathways upregulated in macrophages isolated from aged mice include proteins critical to the formation of the immunoproteasome. Detection of these latter proteins is dramatically enhanced following LPS exposure for macrophages isolated from aged animals; in comparison, the identification of immunoproteasome subunits is insensitive to LPS exposure for macrophages isolated from young animals. Consistent with observed global changes in the proteome, quantitative proteomic measurements indicate that there are age-dependent abundance changes involving specific proteins linked to immune cell function under basal conditions. LPS exposure selectively increases the levels of many proteins involved in immune cell function in aged Balb/c mice

  4. Macrophage depletion by free bisphosphonates and zoledronate-loaded red blood cells.

    PubMed

    Sabatino, Raffaella; Antonelli, Antonella; Battistelli, Serafina; Schwendener, Reto; Magnani, Mauro; Rossi, Luigia

    2014-01-01

    Bisphosphonates, besides being important drugs for the treatment of various bone diseases, could also be used to induce apoptosis in macrophage-like and cancer cells. However, their activity in vivo is limited by a short plasma half-life and rapid uptake within bone. Therefore, several delivery systems have been proposed to modify their pharmacokinetic profile and biodistribution. Among these, red blood cells (RBCs) represent one of the most promising biological carriers. The aim of this study was to select the best performing compound among Clodronate, Pamidronate, Ibandronate and Zoledronate in killing macrophages and to investigate RBCs as innovative carrier system to selectively target bisphosphonates to macrophages. To this end, the encapsulation of the selected bisphosphonates in autologous RBCs as well as the effect on macrophages, both in vitro and in vivo were studied. This work shows that, among the tested bisphosphonates, Zoledronate has proven to be the most active molecule. Human and murine RBCs have been successfully loaded with Zoledronate by a procedure of hypotonic dialysis and isotonic resealing, obtaining a dose-dependent drug entrapment with a maximal loading of 7.96±2.03, 6.95±3.9 and 7.0±1.89 µmoles of Zoledronate/ml of packed RBCs for human, Swiss and Balb/C murine RBCs, respectively. Engineered RBCs were able to detach human and murine macrophages in vitro, leading to a detachment of 66±8%, 67±8% and 60.5±3.5% for human, Swiss and Balb/C RBCs, respectively. The in vivo efficacy of loaded RBCs was tested in Balb/C mice administering 59 µg/mouse of RBC-encapsulated Zoledronate. By a single administration, depletion of 29.0±16.38% hepatic macrophages and of 67.84±5.48% spleen macrophages was obtained, confirming the ability of encapsulated Zoledronate to deplete macrophages in vivo. In conclusion, RBCs loaded with Zoledronate should be considered a suitable system for targeted delivery to macrophages, both in vitro and in vivo.

  5. Macrophage Depletion by Free Bisphosphonates and Zoledronate-Loaded Red Blood Cells

    PubMed Central

    Sabatino, Raffaella; Antonelli, Antonella; Battistelli, Serafina; Schwendener, Reto; Magnani, Mauro; Rossi, Luigia

    2014-01-01

    Bisphosphonates, besides being important drugs for the treatment of various bone diseases, could also be used to induce apoptosis in macrophage-like and cancer cells. However, their activity in vivo is limited by a short plasma half-life and rapid uptake within bone. Therefore, several delivery systems have been proposed to modify their pharmacokinetic profile and biodistribution. Among these, red blood cells (RBCs) represent one of the most promising biological carriers. The aim of this study was to select the best performing compound among Clodronate, Pamidronate, Ibandronate and Zoledronate in killing macrophages and to investigate RBCs as innovative carrier system to selectively target bisphosphonates to macrophages. To this end, the encapsulation of the selected bisphosphonates in autologous RBCs as well as the effect on macrophages, both in vitro and in vivo were studied. This work shows that, among the tested bisphosphonates, Zoledronate has proven to be the most active molecule. Human and murine RBCs have been successfully loaded with Zoledronate by a procedure of hypotonic dialysis and isotonic resealing, obtaining a dose-dependent drug entrapment with a maximal loading of 7.96±2.03, 6.95±3.9 and 7.0±1.89 µmoles of Zoledronate/ml of packed RBCs for human, Swiss and Balb/C murine RBCs, respectively. Engineered RBCs were able to detach human and murine macrophages in vitro, leading to a detachment of 66±8%, 67±8% and 60.5±3.5% for human, Swiss and Balb/C RBCs, respectively. The in vivo efficacy of loaded RBCs was tested in Balb/C mice administering 59 µg/mouse of RBC-encapsulated Zoledronate. By a single administration, depletion of 29.0±16.38% hepatic macrophages and of 67.84±5.48% spleen macrophages was obtained, confirming the ability of encapsulated Zoledronate to deplete macrophages in vivo. In conclusion, RBCs loaded with Zoledronate should be considered a suitable system for targeted delivery to macrophages, both in vitro and in vivo. PMID

  6. Probiotic Bacillus amyloliquefaciens mediate M1 macrophage polarization in mouse bone marrow-derived macrophages.

    PubMed

    Ji, Jian; Hu, Sheng-Lan; Cui, Zhi-Wen; Li, Wei-Fen

    2013-05-01

    Depending on the microenvironment, macrophages can acquire distinct functional phenotypes, referred to as classically activated M1 and M2. M1 macrophages are considered potent effector cells that kill intracellular pathogens, and M2 macrophages promote the resolution of wound healing. In this study, we are interested to know whether probiotic Bacillus amyloliquefaciens (Ba) can induce macrophages polarization. Real-time fluorescence PCR analysis demonstrated that the expression of IL-1β, iNOS, TNF-α and IL-6 genes for M1 macrophages was significantly increased at 1.5 h after probiotic Ba treatment compared to the probiotic Ba-free treatment (P < 0.01), whereas the expression of M2 macrophage marker genes (Arg1, Fizz1, MR, Ym1) was decreased (P < 0.05). Furthermore, the phagocytic activity was dramatically increased in the Ba-treated BMDMs using a FITC-dextran endocytosis assay. Together, these findings indicated that probiotic Ba facilitated polarization of M1 macrophages and enhanced its phagocytic capacity. The results expanded our knowledge about probiotic function-involved macrophage polarization.

  7. Effect of lipopolysaccharide on protein accumulation by murine peritoneal macrophages: the correlation to activation for macrophage tumoricidal function

    SciTech Connect

    Tannenbaum, C.S.

    1987-01-01

    The protein synthetic patterns of tumoricidal murine peritoneal macrophage populations have been compared to those of non-tumoricidal populations utilizing two dimensional polyacrylamide gel electrophoresis (2D PAGE) of (/sup 35/S)-methionine-labeled proteins. While the protein synthetic patterns exhibited by resident, inflammatory and activated macrophages had numerous common features which distinguished them from the other normal non-macrophage cell types examined, unique proteins also distinguished each macrophage population from the others. Peritoneal macrophages elicited by treatment with heat killed Propionibacterium acnes, the live, attenuated Mycobacterium bovis strain BCG, Listeria monocytogenes and the protozoan flagellate Trypanosoma rhodesiense, all exhibited tumoricidal activity in 16h or 72h functional assays, and shared a common protein synthetic profile which differentiated them from the synthetic patterns characteristic of the non-tumoricidal resident and inflammatory macrophages.

  8. Mycobacterium massiliense Induces Macrophage Extracellular Traps with Facilitating Bacterial Growth

    PubMed Central

    Yoon, Yina; Na, Yirang; Kim, Bum-Joon; Seok, Seung Hyeok

    2016-01-01

    Human neutrophils have been known to release neutrophil extracellular traps (NETs), antimicrobial DNA structures capable of capturing and killing microbes. Recently, a similar phenomenon has been reported in macrophages infected with various pathogens. However, a role for macrophages extracellular traps (METs) in host defense responses against Mycobacterium massiliense (M. mass) has yet to be described. In this study, we show that M. mass, a rapid growing mycobacterium (RGM), also induces the release of METs from PMA-differentiated THP-1 cells. Intriguingly, this process is not dependent on NADPH oxidase activity, which regulates NET formation. Instead, M. mass-induced MET formation partially depends on calcium influx and requires phagocytosis of high bacterial load. The METs consist of a DNA backbone embedded with microbicidal proteins such as histone, MPO and elastase. Released METs entrap M. mass and prevent their dissemination, but do not have bactericidal activity. Instead, they result in enhanced bacterial growth. In this regard, METs were considered to provide interaction of M. mass with cells and an environment for bacterial aggregation, which may facilitate mycobacterial survival and growth. In conclusion, our results demonstrate METs as an innate defense response against M. mass infection, and suggest that extracellular traps play a multifaceted role in the interplay between host and bacteria. PMID:27191593

  9. How to kill creativity.

    PubMed

    Amabile, T M

    1998-01-01

    In today's knowledge economy, creativity is more important than ever. But many companies unwittingly employ managerial practices that kill it. How? By crushing their employees' intrinsic motivation--the strong internal desire to do something based on interests and passions. Managers don't kill creativity on purpose. Yet in the pursuit of productivity, efficiency, and control--all worthy business imperatives--they undermine creativity. It doesn't have to be that way, says Teresa Amabile. Business imperatives can comfortably coexist with creativity. But managers will have to change their thinking first. Specifically, managers will need to understand that creativity has three parts: expertise, the ability to think flexibly and imaginatively, and motivation. Managers can influence the first two, but doing so is costly and slow. It would be far more effective to increase employees' intrinsic motivation. To that end, managers have five levers to pull: the amount of challenge they give employees, the degree of freedom they grant around process, the way they design work groups, the level of encouragement they give, and the nature of organizational support. Take challenge as an example. Intrinsic motivation is high when employees feel challenged but not overwhelmed by their work. The task for managers, therefore, becomes matching people to the right assignments. Consider also freedom. Intrinsic motivation--and thus creativity--soars when managers let people decide how to achieve goals, not what goals to achieve. Managers can make a difference when it comes to employee creativity. The result can be truly innovative companies in which creativity doesn't just survive but actually thrives.

  10. Kill operation requires thorough analysis

    SciTech Connect

    Abel, L.W.

    1995-05-15

    Full control of a blowout well requires a properly designed post-capping kill operation because failures in regaining well control usually occur during the kill operation, not during capping. Capping (the installation of pressure control or diverter equipment on the wellhead) is generally very reliable in gaining control of a blowout well. The following techniques are some of the viable means of killing blowout wells once the capping assemblies are in place: direct shut in of the flow; bullheading; momentum kill; volumetric control for migration of fluids or lubrication after migration ceases; and dynamic kills (friction-based dynamic kills or mass flow rate kills) The objective of most post-capping operations is to stop the flow and put the well under hydrostatic control. The means of killing a blowout once capping assemblies are in place should be chosen with care to avoid problems such as cratering, equipment failure, and underground blowouts. The particular circumstances and well integrity will dictate which kill method will be the most viable. Each of these five methods are explained.

  11. Effects of murine leukemia virus env gene proteins on macrophage-mediated cytotoxicity in vitro

    NASA Technical Reports Server (NTRS)

    Chapes, S. K.; Takemoto, L. J.; Spooner, B. S. (Principal Investigator)

    1991-01-01

    F5b Tumor cells were incubated with concentrated culture supernatants taken from cells resistant (F5m) or sensitive (F5b) to contact-dependent macrophage cytotoxicity. Macrophage cell line B6MP102 and murine peritoneal macrophages killed targets incubated with supernatants taken from sensitive cells but poorly killed cells incubated in supernatants isolated from resistant cells. Membranes from cells resistant to macrophage killing, F5m, were fused into F5b cells. The fused F5b cells were killed significantly less than F5b cells fused with F5b cell membranes or untreated F5b cells. The decreased killing of F5b cells corresponded to increased concentrations of gp70(a) molecules on F5b cells. Affinity purified gp70(a) was added to cytotoxicity assays but failed to inhibit macrophage cytotoxicity. P15E molecules were detectable on both F5b and F5m cells. In addition, a synthetic peptide found to exhibit the inhibitory properties of p15E was added to cytotoxicity assays. P15E synthetic peptide also did not inhibit macrophage cytotoxicity. Therefore, env gene proteins of murine leukemia virus do not appear responsible for inducing tumor cell resistance to activated macrophage contact-dependent cytotoxicity.

  12. Pseudomonas aeruginosa PAO1 Kills Caenorhabditis elegans by Cyanide Poisoning

    PubMed Central

    Gallagher, Larry A.; Manoil, Colin

    2001-01-01

    In this report we describe experiments to investigate a simple virulence model in which Pseudomonas aeruginosa PAO1 rapidly paralyzes and kills the nematode Caenorhabditis elegans. Our results imply that hydrogen cyanide is the sole or primary toxic factor produced by P. aeruginosa that is responsible for killing of the nematode. Four lines of evidence support this conclusion. First, a transposon insertion mutation in a gene encoding a subunit of hydrogen cyanide synthase (hcnC) eliminated nematode killing. Second, the 17 avirulent mutants examined all exhibited reduced cyanide synthesis, and the residual production levels correlated with killing efficiency. Third, exposure to exogenous cyanide alone at levels comparable to the level produced by PAO1 killed nematodes with kinetics similar to those observed with bacteria. The killing was not enhanced if hcnC mutant bacteria were present during cyanide exposure. And fourth, a nematode mutant (egl-9) resistant to P. aeruginosa was also resistant to killing by exogenous cyanide in the absence of bacteria. A model for nematode killing based on inhibition of mitochondrial cytochrome oxidase is presented. The action of cyanide helps account for the unusually broad host range of virulence of P. aeruginosa and may contribute to the pathogenesis in opportunistic human infections due to the bacterium. PMID:11591663

  13. THE PARTICULATE HYDROLASES OF MACROPHAGES

    PubMed Central

    Cohn, Zanvil A.; Wiener, Edith

    1963-01-01

    The influence of phagocytosis on the morphological and biochemical properties of macrophage hydrolase-containing granules has been studied in vitro. Following the uptake of large numbers of heat-killed bacteria, an intracellular rearrangement of hydrolytic enzymes occurred. This was associated with the solubilization of 50 to 60 per cent of the total cell content of acid phosphatase, cathepsin, lysozyme, beta glucuronidase, acid ribonuclease, and acid desoxyribonuclease and with a corresponding decrease in granule-bound enzyme. With more prolonged incubation the majority of the soluble intracellular pool of acid ribonuclease and lysozyme was lost to the extracellular medium. No change in the total content of any of the hydrolases was noted during 180 minutes of incubation in vitro. The morphological fate of the granules was studied by a histochemical method for acid phosphatase. After the phagocytosis of yeast cell walls there was a disappearance of acid phosphatase-positive granules and an accumulation of reaction product about the ingested particle. Experiments employing macrophages which were supravitally stained with neutral red also demonstrated the loss of neutral red-positive granules and the accumulation of the dye about the yeast cell walls. These results strongly suggest that lysis of macrophage granules occurs following phagocytosis and that a portion of the granule contents are then resegregated within the newly formed phagocytic vacuole. PMID:14112262

  14. Macrophages in resistance to candidiasis.

    PubMed Central

    Vázquez-Torres, A; Balish, E

    1997-01-01

    Candida albicans, an increasingly common opportunistic pathogenic fungus, frequently causes disease in immunodeficient but not immunocompetent hosts. Clarifying the role of the phagocytic cells that participate in resistance to candidiasis not only is basic to understanding how the host copes with this dimorphic pathogen but also will expedite the development of innovative prophylactic and therapeutic approaches for treating the multiple clinical presentations that candidiasis encompasses. In this review, we present evidence that a diverse population of mononuclear phagocytes, in different states of activation and differentiation and from a variety of host species, can phagocytize C. albicans blastoconidia via an array of opsonic and nonopsonic mechanisms and can kill C. albicans blastoconidia and hyphae by means of oxygen-dependent and -independent mechanisms. Reactive nitrogen intermediates should now be added to the well-established candidacidal reactive oxygen intermediates of macrophages. Furthermore, what were thought to be two independent pathways, i.e., nitric oxide and superoxide anion, have now been shown to combine to form a potent macrophage candidacidal molecule, peroxynitrite. In contrast to monocytes and neutrophils, which are important in resistance to early stages of C. albicans infections, more differentiated macrophages activated by cytokines such as gamma interferon participate in the acquired resistance of hosts with C. albicans-specific, cell-mediated immunity. Evidence presented in this review demonstrates that mononuclear phagocytes, in some instances in the absence of other professional phagocytes such as neutrophils, play an import role in resistance to systemic and mucosal candidiasis. PMID:9184009

  15. Modulating macrophage response to biomaterials

    NASA Astrophysics Data System (ADS)

    Zaveri, Toral

    Zn, parameters which are likely interdependent. Considering the toxicity of ZnO nanorod surface towards macrophages, their role as an antibacterial surface was explored. Antibacterial coating approaches are being investigated to modify implants to reduce bacterial adhesion and viability in order to reduce implant-associated infection. To assess the efficacy of ZnO nanorod surfaces as an anti-bacterial coating, we evaluated bacterial adhesion and viability, compared to sputtered ZnO and glass substrates. Common implant-associated pathogens, Pseudomonas aeruginosa and Staphylococcus epidermidis were investigated. ZnO nanorod surface and sputtered ZnO demonstrated a significant bactericidal effect, killing respectively 2.5x and 1.7x times the number of bacteria dead on glass. A similar bactericidal effect of ZnO substrates on S. epidermidis was also evident, with sputtered ZnO and ZnO nanorod substrates killing respectively 22x and 32x times bacteria dead on glass. These data support the further investigation of ZnO nanorod coatings for bacterial adhesion resistance and bactericidal properties.

  16. How electroshock weapons kill!

    NASA Astrophysics Data System (ADS)

    Lundquist, Marjorie

    2010-03-01

    Growing numbers of law enforcement officers now carry an electroshock weapon (ESW). Over 500 U.S. deaths have followed ESW use in the past 26 years; over 450 of these deaths followed use of an electromuscular disruptor in the past 9 years. Most training courses teach that ESWs are safe; that they can kill only by the direct effect of electric current on the heart; and that a death following use of an ESW always has some other cause. All these teachings are false! The last was disproved by Lundquist.^1 Williams^2 ruled out direct electrical effects as a cause of almost all the 213 deaths he studied, leaving disruption of normal physiological processes as the only alternative explanation. Careful study of all such deaths identifies 4 different ways that death has or could have been brought about by the ESW: kidney failure following rhabdomyolysis [rare]; cardiac arrest from hyperkalemia following rhabdomyolysis [undocumented]; lactic acid-induced ventricular fibrillation [conclusive proof impossible]; and [most common] anoxia from so much lactic acid in the circulating blood that it acts as an oxygen scavenger, continuously depleting the blood of oxygen until most of the lactate has been metabolized. ^1M. Lundquist, BAPS 54(1) K1.270(2009). ^2Howard E. Williams, Taser Electronic Control Devices and Sudden In-Custody Death, 2008.

  17. Induction of heme oxygenase-1 contributes to survival of Mycobacterium abscessus in human macrophages-like THP-1 cells

    PubMed Central

    Abdalla, Maher Y.; Ahmad, Iman M.; Switzer, Barbara; Britigan, Bradley E.

    2015-01-01

    Mycobacterium abscessus (M.abs) is a rapidly growing mycobacterial species that infects macrophages, and is an important pathogen in patients with cystic fibrosis. We studied the early stages of M.abs infection of macrophages, with emphasis on the role of heme-oxygenase-1 (HO-1) in this infection. THP-1 cells were activated using TPA into macrophage-like cells and infected with M.abs for different time points. M.abs infection robustly induced HO-1 expression in the THP-1 cells. Production of HO-1 was p38 MAPK-dependent, as p38 inhibitors suppressed HO-1 induction. Pretreatment with HO-1 inhibitors tin-protoporphyrin (SnPP) significantly inhibited M.abs growth inside macrophages. Furthermore, inhibiting HO-1 using HO-1 siRNA or the HO-1 upstream signaling molecule; Nrf2 using Nrf2 siRNA resulted in similar inhibition of M.abs. In contrast, inducing HO-1 did not increase M.abs intracellular growth above control. Products of HO-1 metabolism of heme are bilirubin, biliverdin, carbon monoxide (CO) and iron. The addition of either bilirubin or biliverdin, but not CO, completely restored the SnPP inhibitory effect and partially that with HO-1 siRNA. To understand the mechanisms, we used Syto-62 labeled M.abs to infect macrophages. Interestingly, HO-1 inhibition promoted M.abs-containing phagosome fusion with lysosomes, which should enhance M.abs killing. M.abs infection enhanced THP-1 ROS production as demonstrated by increased DHE, DCF fluorescence, and EPR signal. HO-1 inhibition further increased ROS production in infected macrophages. Our results indicate that HO-1 induction is important for M.abs growth during the early stages of infection, and that the HO-1 products bilirubin and biliverdin, perhaps through modulation of intracellular ROS levels, may be involved. PMID:25638774

  18. Peroxynitrite, a potent macrophage-derived oxidizing cytotoxin to combat invading pathogens

    PubMed Central

    Prolo, Carolina; Álvarez, María Noel; Radi, Rafael

    2013-01-01

    Macrophages are among the first cellular actors facing the invasion of microorganisms. These cells are able to internalize pathogens and destroy them by means of toxic mediators, many of which are produced enzymatically and have strong oxidizing capacity. Indeed, macrophages count on the NADPH oxidase complex activity, which is triggered during pathogen invasion and leads to the production of superoxide radical inside the phagosome. At the same time, the induction of nitric oxide synthase results in the production of nitric oxide in the cytosol which is able to readily diffuse to the phagocytic vacuole. Superoxide radical and nitric oxide react at diffusion controlled rates with each other inside the phagosome to yield peroxynitrite, a powerful oxidant capable to kill microorganisms. Peroxynitrite toxicity resides on oxidations and nitrations of biomolecules in the target cell. The central role of peroxynitrite as a key effector molecule in the control of infections has been proven in a wide number of models. However, some microorganisms and virulent strains adapt to survive inside the potentially hostile oxidizing microenvironment of the phagosome by either impeding peroxynitrite formation or rapidly detoxifying it once formed. In this context, the outcome of the infection process is a result of the interplay between the macrophage-derived oxidizing cytotoxins such as peroxynitrite and the antioxidant defense machinery of the invading pathogens. PMID:24281946

  19. Preadipocyte conversion to macrophage. Evidence of plasticity.

    PubMed

    Charrière, Guillaume; Cousin, Béatrice; Arnaud, Emmanuelle; André, Mireille; Bacou, Francis; Penicaud, Luc; Casteilla, Louis

    2003-03-14

    Preadipocytes are present throughout adult life in adipose tissues and can proliferate and differentiate into mature adipocytes according to the energy balance. An increasing number of reports demonstrate that cells from adipose lineages (preadipocytes and adipocytes) and macrophages share numerous functional or antigenic properties. No large scale comparison reflecting the phenotype complexity has been performed between these different cell types until now. We used profiling analysis to define the common features shared by preadipocyte, adipocyte, and macrophage populations. Our analysis showed that the preadipocyte profile is surprisingly closer to the macrophage than to the adipocyte profile. From these data, we hypothesized that in a macrophage environment preadipocytes could effectively be converted into macrophages. We injected labeled stroma-vascular cells isolated from mouse white adipose tissue or 3T3-L1 preadipocyte cell line into the peritoneal cavity of nude mice and investigated changes in their phenotype. Preadipocytes rapidly and massively acquired high phagocytic activity and index. 60-70% of preadipocytes also expressed five macrophage-specific antigens: F4/80, Mac-1, CD80, CD86, and CD45. These values were similar to those observed for peritoneal macrophages. In vitro experiments showed that cell-to-cell contact between preadipocytes and peritoneal macrophages partially induced this preadipocyte phenotype conversion. Overall, these results suggest that preadipocyte and macrophage phenotypes are very similar and that preadipocytes have the potential to be very efficiently and rapidly converted into macrophages. This work emphasizes the great cellular plasticity of adipose precursors and reinforces the link between adipose tissue and innate immunity processes. PMID:12519759

  20. How Mouse Macrophages Sense What Is Going On

    PubMed Central

    Ley, Klaus; Pramod, Akula Bala; Croft, Michael; Ravichandran, Kodi S.; Ting, Jenny P.

    2016-01-01

    Macrophages are central to both innate and adaptive immunity. With few exceptions, macrophages are the first cells that sense trouble and respond to disturbances in almost all tissues and organs. They sense their environment, inhibit or kill pathogens, take up apoptotic and necrotic cells, heal tissue damage, and present antigens to T cells. Although the origins (yolk sac versus monocyte-derived) and phenotypes (functions, gene expression profiles, surface markers) of macrophages vary between tissues, they have many receptors in common that are specific to one or a few molecular species. Here, we review the expression and function of almost 200 key macrophage receptors that help the macrophages sense what is going on, including pathogen-derived molecules, the state of the surrounding tissue cells, apoptotic and necrotic cell death, antibodies and immune complexes, altered self molecules, extracellular matrix components, and cytokines, including chemokines. PMID:27313577

  1. Decreased glycogen synthase kinase 3-beta levels and related physiological changes in Bacillus anthracis lethal toxin-treated macrophages.

    PubMed

    Tucker, Amy E; Salles, Isabelle I; Voth, Daniel E; Ortiz-Leduc, William; Wang, Han; Dozmorov, Igor; Centola, Michael; Ballard, Jimmy D

    2003-08-01

    The lethal factor (LF) component of Bacillus anthracis lethal toxin (LeTx) cleaves mitogen activated protein kinase kinases (MAPKKs) in a variety of different cell types, yet only macrophages are rapidly killed by this toxin. The reason for this selective killing is unclear, but suggests other factors may also be involved in LeTx intoxication. In the current study, DNA membrane arrays were used to identify broad changes in macrophage physiology after treatment with LeTx. Expression of genes regulated by MAPKK activity did not change significantly, yet a series of genes under glycogen synthase kinase-3-beta (GSK-3beta) regulation changed expression following LeTx treatment. Correlating with these transcriptional changes GSK-3beta was found to be below detectable levels in toxin-treated cells and an inhibitor of GSK-3beta, LiCl, sensitized resistant IC-21 macrophages to LeTx. In addition, zebrafish embryos treated with LeTx showed signs of delayed pigmentation and cardiac hypertrophy; both processes are subject to regulation by GSK-3beta. A putative compensatory response to loss of GSK-3beta was indicated by differential expression of three motor proteins following toxin treatment and Kif1C, a motor protein involved in sensitivity to LeTx, increased expression in toxin-sensitive cells yet decreased in resistant cells following toxin treatment. Differential expression of microtubule-associating proteins and a decrease in the level of cellular tubulin were detected in LeTx-treated cells, both of which can result from loss of GSK-3beta activity. These data provide new information on LeTx's overall influence on macrophage physiology and suggest loss of GSK-3beta contributes to cytotoxicity. PMID:12864812

  2. Notes on super Killing tensors

    NASA Astrophysics Data System (ADS)

    Howe, P. S.; Lindström, U.

    2016-03-01

    The notion of a Killing tensor is generalised to a superspace setting. Conserved quantities associated with these are defined for superparticles and Poisson brackets are used to define a supersymmetric version of the even Schouten-Nijenhuis bracket. Superconformal Killing tensors in flat superspaces are studied for spacetime dimensions 3,4,5,6 and 10. These tensors are also presented in analytic superspaces and super-twistor spaces for 3,4 and 6 dimensions. Algebraic structures associated with superconformal Killing tensors are also briefly discussed.

  3. Women who kill their children.

    PubMed

    d'Orbán, P T

    1979-06-01

    During a 6 year period (1970-75) 89 women charged with the killing or attempted murder of their children were examined in a female remand prison. Six types of maternal filicide were distinguished: battering mothers (36 cases), mentally ill mothers (24 cases), neonaticides (11 cases), retaliating mothers (9 cases), women who killed unwanted children (8 cases) and mercy killing (1 case). Types of filicide were compared on a number of social and psychiatric characteristics and on their offence patterns and court disposals. The operation of the Infanticide Act is discussed in the light of these findings.

  4. Mycobacterium avium subspecies paratuberculosis infection of cattle does not diminish peripheral blood-derived macrophage mycobactericidal activity.

    PubMed

    Hostetter, J; Zhang, W; Simutis, F

    2006-09-15

    Ruminants infected with Mycobacterium avium subspecies paratuberculosis consistently develop a multibacillary form of disease that is centered on the ileum. Mechanisms responsible for failure of macrophage function during multibacillary disease are incompletely characterized. Our data suggest that mycobactericidal functions are present, and potentially enhanced, in monocyte-derived macrophages from M. avium subsp. paratuberculosis infected cattle. Addition of CD4(+) T cells from infected animals to autologous in vitro infected macrophages did not increase bacterial killing. In contrast, CD4(+) T cells from non-infected animals did increase bacterial killing in autologous macrophages. In macrophages from both infected and non-infected cattle, bacterial killing appeared to be independent of interferon-gamma (IFN-gamma) and nitric oxide production.

  5. The effect of lipopolysaccharide on bovine mammary macrophage function.

    PubMed Central

    Politis, I; Zhao, X; McBride, B W; Burton, J H

    1991-01-01

    The effect of Escherichia coli lipopolysaccharide (LPS) on the expression of major histocompatibility complex (MHC) class II molecules by bovine mammary macrophages was examined. The ability of LPS-treated mammary macrophages to support antigen-specific T-cell proliferation, as a measure of their antigen presentation ability, was also evaluated. For this purpose, control and LPS-treated macrophages were pulsed with heat-killed Staphylococcus aureus and then cultured with S. aureus-sensitized T-cells. Our data show that LPS had no significant effect on the expression of MHC class II molecules on the surface of mammary macrophages. Furthermore, LPS-induced macrophages were no more active in supporting T-cell proliferation on a per cell basis than unstimulated macrophages. The lack of macrophage response to LPS with respect to expression of MHC class II molecules and the antigen presentation ability is another example of the hyporesponsive nature of macrophages isolated from the bovine mammary gland. PMID:1889031

  6. Hemophagocytic Macrophages Harbor Salmonella enterica during Persistent Infection

    PubMed Central

    Nix, Rebecca N; Altschuler, Sarah E; Henson, Peter M; Detweiler, Corrella S

    2007-01-01

    Salmonella enterica subspecies can establish persistent, systemic infections in mammals, including human typhoid fever. Persistent S. enterica disease is characterized by an initial acute infection that develops into an asymptomatic chronic infection. During both the acute and persistent stages, the bacteria generally reside within professional phagocytes, usually macrophages. It is unclear how salmonellae can survive within macrophages, cells that evolved, in part, to destroy pathogens. Evidence is presented that during the establishment of persistent murine infection, macrophages that contain S. enterica serotype Typhimurium are hemophagocytic. Hemophagocytic macrophages are characterized by the ingestion of non-apoptotic cells of the hematopoietic lineage and are a clinical marker of typhoid fever as well as certain other infectious and genetic diseases. Cell culture assays were developed to evaluate bacterial survival in hemophagocytic macrophages. S. Typhimurium preferentially replicated in macrophages that pre-phagocytosed viable cells, but the bacteria were killed in macrophages that pre-phagocytosed beads or dead cells. These data suggest that during persistent infection hemophagocytic macrophages may provide S. Typhimurium with a survival niche. PMID:18085823

  7. Enhanced resistance against Listeria monocytogenes at an early phase of primary infection in pregnant mice: activation of macrophages during pregnancy.

    PubMed Central

    Watanabe, Y; Mitsuyama, M; Sano, M; Nakano, H; Nomoto, K

    1986-01-01

    We investigated the pregnancy-induced changes in macrophage activity which are important in the expression of host defense against infections. Several macrophage functions were examined by using Listeria monocytogenes. In pregnant mice, prolonged survival and enhanced in vivo elimination of bacteria were observed in the early phase of primary infection. Functions of peritoneal macrophages, including in vitro phagocytosis intracellular killing, glucose consumption, generation of superoxide anion, and intracellular beta-glucuronidase activity were shown to be enhanced in pregnant mice. These findings indicate that pregnancy enhances macrophage functions qualitatively. Possible mechanisms for this enhancement and the significance of macrophage activation for pregnant hosts are discussed. PMID:3011673

  8. Killing, letting die and euthanasia.

    PubMed

    Husak, D N

    1979-12-01

    Medical ethicists debate whether or not the moral assessment of cases of euthanasia should depend on whether the patient is 'killed' or 'allowed to die'. The usual presupposition is that a clear distinction between killing and letting die can be drawn so that this substantive question is not begged. I contend that the categorisation of cases of instances of killing rather than as instances of letting die depends in part on a prior moral assessment of the case. Hence is it trivially rather than substantively true that the distinction has moral significance. But even if a morally neutral (ie non-question begging) distinction could be drawn, its application to the euthanasia controversy is problematic. I illustrate the difficulties of employing this distinction to reach moral conclusions by critically discussing Philippa Foot's recent treatment of euthanasia. I conclude that even if an act of euthanasia is an instance of killing, and there exists a prima facie moral duty not to kill, and no more stringent duty overrides this duty, one still cannot determine such an act to be morally impermissible.

  9. Regulation of macrophage-mediated larvicidal activity in Echinococcus granulosus and Mesocestoides corti (Cestoda) infection in mice.

    PubMed

    Jenkins, P; Dixon, J B; Rakha, N K; Carter, S D

    1990-04-01

    Killing of metacestodes by normal or post-infection macrophages and the regulation of this activity by cytokines were studied in vitro. The protoscolecidal activity of normal macrophages against Echinococcus granulosus was inhibited by a product of naive T-enriched lymphocytes co-cultured with protoscoleces (PSC). By contrast, supernates from co-cultures of Mesocestoides corti tetrathyridia (MCT) and T-enriched or B-enriched normal lymphocytes increased killing of MCT by normal macrophages. Larvicidal activity (against both PSC and MCT) was enhanced by high concentrations of macrophage-activating factors produced by Con A-stimulated rat lymphocytes (Con A-LK), but was reduced by low concentrations of these factors. Activation by synergism between Con A-LK and recombinant interferon-gamma(r. IFN-gamma) was demonstrated in macrophage-mediated killing of MCT at high effector to target ratio. Cytokine-activation of normal or post-MCT infection macrophages was compared. Macrophages from both 8 and 20 week post-infection mice were refractory to lymphokines from lymphocyte-MCT cultures and displayed greatly reduced killing of MCT. Macrophage activation by Con A-LK and r.IFN-gamma was also impaired, implying a general defect in the ability of these post-infection macrophages to respond to macrophage activating signals. The data indicate that two different mechanisms may exist by which metacestodes regulate potentially larvicidal effector mechanisms. E. granulosus can elicit the production of lymphokines suppressive for PSC killing, whereas M. corti appears directly to induce a refractory state in effector macrophages.

  10. Perivascular macrophages mediate neutrophil recruitment during bacterial skin infection

    PubMed Central

    Abtin, Arby; Jain, Rohit; Mitchell, Andrew J.; Roediger, Ben; Brzoska, Anthony J.; Tikoo, Shweta; Cheng, Qiang; Ng, Lai Guan; Cavanagh, Lois L.; von Andrian, Ulrich H.; Hickey, Michael J.; Firth, Neville; Weninger, Wolfgang

    2014-01-01

    Transendothelial migration of neutrophils in post-capillary venules is a key event in the inflammatory response against pathogens and tissue damage. The precise regulation of this process is incompletely understood. We report that perivascular macrophages are critical for neutrophil migration into skin infected with the pathogen Staphylococcus aureus. Using multiphoton intravital microscopy we show that neutrophils extravasate from inflamed dermal venules in close proximity to perivascular macrophages, which are a major source of neutrophil chemoattractants. The virulence factor alpha-hemolysin lyses perivascular macrophages leading to decreased neutrophil transmigration. Our data illustrate a previously unrecognized role for perivascular macrophages in neutrophil recruitment to inflamed skin, and indicate that Staphylococcus aureus uses hemolysin-dependent killing of these cells as an immune evasion strategy. PMID:24270515

  11. Macrophage and polymorphonuclear leukocyte function in patients with Alzheimer disease.

    PubMed

    Cameron, D J; Durst, G G; Majeski, J A

    1985-01-01

    Monocyte derived macrophages and polymorphonuclear leukocytes (PMNs) isolated from the peripheral blood of thirteen patients with Alzheimer disease were studied for their cytotoxic effects on a sensitive allogenic tumor target. PMN cells from 11 of the 13 patients with Alzheimer disease were able to kill the tumor cells. In addition, the macrophages from 12 of the 13 Alzheimer disease patients were cytotoxic towards the tumor targets. Four of these patients possessed a plasma inhibitory factor which was capable of suppressing macrophage mediated cytotoxicity. When the lymphocytes from these patients were studied for their ability to be stimulated with the specific antigen, streptokinase, to produce macrophage activating factor (MAF), only 5 of the 13 patients studied possessed lymphocytes which were capable of producing MAF. Thus, the only immunological defect in Alzheimer disease patients which was observed in this study was in the ability of the lymphocytes to synthesize MAF. PMID:4084662

  12. Perivascular macrophages mediate neutrophil recruitment during bacterial skin infection.

    PubMed

    Abtin, Arby; Jain, Rohit; Mitchell, Andrew J; Roediger, Ben; Brzoska, Anthony J; Tikoo, Shweta; Cheng, Qiang; Ng, Lai Guan; Cavanagh, Lois L; von Andrian, Ulrich H; Hickey, Michael J; Firth, Neville; Weninger, Wolfgang

    2014-01-01

    Transendothelial migration of neutrophils in postcapillary venules is a key event in the inflammatory response against pathogens and tissue damage. The precise regulation of this process is incompletely understood. We report that perivascular macrophages are critical for neutrophil migration into skin infected with the pathogen Staphylococcus aureus. Using multiphoton intravital microscopy we showed that neutrophils extravasate from inflamed dermal venules in close proximity to perivascular macrophages, which are a major source of neutrophil chemoattractants. The virulence factor α-hemolysin produced by S. aureus lyses perivascular macrophages, which leads to decreased neutrophil transmigration. Our data illustrate a previously unrecognized role for perivascular macrophages in neutrophil recruitment to inflamed skin and indicate that S. aureus uses hemolysin-dependent killing of these cells as an immune evasion strategy. PMID:24270515

  13. Does Assessment Kill Student Creativity?

    ERIC Educational Resources Information Center

    Beghetto, Ronald A.

    2005-01-01

    Does assessment kill creativity? In this article, creativity is defined and discussed and an overview of creativity and motivational research is provided to describe how assessment practices can influence students' creativity. Recommendations for protecting creativity when assessing students also are provided.

  14. Farm Education at Stony Kill.

    ERIC Educational Resources Information Center

    Parisio, Richard

    1986-01-01

    Describes typical winter farm lessons for students visiting Stony Kill Farm Environmental Education Center located 70 miles north of New York City: butter and corncake making, soil erosion experiments, dissecting and growing seeds. Emphasizes major theme of conservation of farmland from destructive farming practices and careless development. (NEC)

  15. O-Glycosylation in Cell Wall Proteins in Scedosporium prolificans Is Critical for Phagocytosis and Inflammatory Cytokines Production by Macrophages

    PubMed Central

    Xisto, Mariana I. D. S.; Bittencourt, Vera C. B.; Liporagi-Lopes, Livia Cristina; Haido, Rosa M. T.; Mendonça, Morena S. A.; Sassaki, Guilherme; Figueiredo, Rodrigo T.; Romanos, Maria Teresa V.; Barreto-Bergter, Eliana

    2015-01-01

    In this study, we analyze the importance of O-linked oligosaccharides present in peptidorhamnomannan (PRM) from the cell wall of the fungus Scedosporium prolificans for recognition and phagocytosis of conidia by macrophages. Adding PRM led to a dose-dependent inhibition of conidia phagocytosis, whereas de-O-glycosylated PRM did not show any effect. PRM induced the release of macrophage-derived antimicrobial compounds. However, O-linked oligosaccharides do not appear to be required for such induction. The effect of PRM on conidia-induced macrophage killing was examined using latex beads coated with PRM or de-O-glycosylated PRM. A decrease in macrophage viability similar to that caused by conidia was detected. However, macrophage killing was unaffected when beads coated with de-O-glycosylated PRM were used, indicating the toxic effect of O-linked oligosaccharides on macrophages. In addition, PRM triggered TNF-α release by macrophages. Chemical removal of O-linked oligosaccharides from PRM abolished cytokine induction, suggesting that the O-linked oligosaccharidic chains are important moieties involved in inflammatory responses through the induction of TNF-α secretion. In summary, we show that O-glycosylation plays a role in the recognition and uptake of S. prolificans by macrophages, killing of macrophages and production of pro- inflammatory cytokines. PMID:25875427

  16. Aging of mice is associated with p16(Ink4a)- and β-galactosidase-positive macrophage accumulation that can be induced in young mice by senescent cells.

    PubMed

    Hall, Brandon M; Balan, Vitaly; Gleiberman, Anatoli S; Strom, Evguenia; Krasnov, Peter; Virtuoso, Lauren P; Rydkina, Elena; Vujcic, Slavoljub; Balan, Karina; Gitlin, Ilya; Leonova, Katerina; Polinsky, Alexander; Chernova, Olga B; Gudkov, Andrei V

    2016-07-01

    Senescent cells (SCs) have been considered a source of age-related chronic sterile systemic inflammation and a target for anti-aging therapies. To understand mechanisms controlling the amount of SCs, we analyzed the phenomenon of rapid clearance of human senescent fibroblasts implanted into SCID mice, which can be overcome when SCs were embedded into alginate beads preventing them from immunocyte attack. To identify putative SC killers, we analyzed the content of cell populations in lavage and capsules formed around the SC-containing beads. One of the major cell types attracted by secretory factors of SCs was a subpopulation of macrophages characterized by p16(Ink4a) gene expression and β-galactosidase activity at pH6.0 (β-gal(pH6)), thus resembling SCs. Consistently, mice with p16(Ink4a) promoter-driven luciferase, developed bright luminescence of their peritoneal cavity within two weeks following implantation of SCs embedded in alginate beads. p16(Ink4a)/β-gal(pH6)-expressing cells had surface biomarkers of macrophages F4/80 and were sensitive to liposomal clodronate used for the selective killing of cells capable of phagocytosis. At the same time, clodronate failed to kill bona fide SCs generated in vitro by genotoxic stress. Old mice with elevated proportion of p16(Ink4a)/β-gal(pH6)-positive cells in their tissues demonstrated reduction of both following systemic clodronate treatment, indicating that a significant proportion of cells previously considered to be SCs are actually a subclass of macrophages. These observations point at a significant role of p16(Ink4a)/β-gal(pH6)-positive macrophages in aging, which previously was attributed solely to SCs. They require re-interpretation of the mechanisms underlying rejuvenating effects following eradication of p16(Ink4a)/β-gal(pH6)-positive cells and reconsideration of potential cellular target for anti-aging treatment.

  17. Aging of mice is associated with p16(Ink4a)- and β-galactosidase-positive macrophage accumulation that can be induced in young mice by senescent cells

    PubMed Central

    Hall, Brandon M.; Balan, Vitaly; Gleiberman, Anatoli S.; Strom, Evguenia; Krasnov, Peter; Virtuoso, Lauren P.; Rydkina, Elena; Vujcic, Slavoljub; Balan, Karina; Gitlin, Ilya; Leonova, Katerina; Polinsky, Alexander; Chernova, Olga B.; Gudkov, Andrei V.

    2016-01-01

    Senescent cells (SCs) have been considered a source of age-related chronic sterile systemic inflammation and a target for anti-aging therapies. To understand mechanisms controlling the amount of SCs, we analyzed the phenomenon of rapid clearance of human senescent fibroblasts implanted into SCID mice, which can be overcome when SCs were embedded into alginate beads preventing them from immunocyte attack. To identify putative SC killers, we analyzed the content of cell populations in lavage and capsules formed around the SC-containing beads. One of the major cell types attracted by secretory factors of SCs was a subpopulation of macrophages characterized by p16(Ink4a) gene expression and β-galactosidase activity at pH6.0 (β-galpH6), thus resembling SCs. Consistently, mice with p16(Ink4a) promoter-driven luciferase, developed bright luminescence of their peritoneal cavity within two weeks following implantation of SCs embedded in alginate beads. p16(Ink4a)/β-galpH6-expressing cells had surface biomarkers of macrophages F4/80 and were sensitive to liposomal clodronate used for the selective killing of cells capable of phagocytosis. At the same time, clodronate failed to kill bona fide SCs generated in vitro by genotoxic stress. Old mice with elevated proportion of p16(Ink4a)/β-galpH6-positive cells in their tissues demonstrated reduction of both following systemic clodronate treatment, indicating that a significant proportion of cells previously considered to be SCs are actually a subclass of macrophages. These observations point at a significant role of p16(Ink4a)/β-galpH6-positive macrophages in aging, which previously was attributed solely to SCs. They require re-interpretation of the mechanisms underlying rejuvenating effects following eradication of p16(Ink4a)/β-galpH6-positive cells and reconsideration of potential cellular target for anti-aging treatment. PMID:27391570

  18. Mycobacteria Clumping Increase Their Capacity to Damage Macrophages

    PubMed Central

    Brambilla, Cecilia; Llorens-Fons, Marta; Julián, Esther; Noguera-Ortega, Estela; Tomàs-Martínez, Cristina; Pérez-Trujillo, Miriam; Byrd, Thomas F.; Alcaide, Fernando; Luquin, Marina

    2016-01-01

    The rough morphotypes of non-tuberculous mycobacteria have been associated with the most severe illnesses in humans. This idea is consistent with the fact that Mycobacterium tuberculosis presents a stable rough morphotype. Unlike smooth morphotypes, the bacilli of rough morphotypes grow close together, leaving no spaces among them and forming large aggregates (clumps). Currently, the initial interaction of macrophages with clumps remains unclear. Thus, we infected J774 macrophages with bacterial suspensions of rough morphotypes of M. abscessus containing clumps and suspensions of smooth morphotypes, primarily containing isolated bacilli. Using confocal laser scanning microscopy and electron microscopy, we observed clumps of at least five rough-morphotype bacilli inside the phagocytic vesicles of macrophages at 3 h post-infection. These clumps grew within the phagocytic vesicles, killing 100% of the macrophages at 72 h post-infection, whereas the proliferation of macrophages infected with smooth morphotypes remained unaltered at 96 h post-infection. Thus, macrophages phagocytose large clumps, exceeding the bactericidal capacities of these cells. Furthermore, proinflammatory cytokines and granuloma-like structures were only produced by macrophages infected with rough morphotypes. Thus, the present study provides a foundation for further studies that consider mycobacterial clumps as virulence factors. PMID:27757105

  19. Antiorthostatic suspension stimulates profiles of macrophage activation in mice

    NASA Technical Reports Server (NTRS)

    Miller, E. S.; Bates, R. A.; Koebel, D. A.; Sonnenfeld, G.

    1999-01-01

    The antiorthostatic suspension model simulates certain physiological effects of spaceflight. We have previously reported BDF1 mice suspended by the tail in the antiorthostatic orientation for 4 days express high levels of resistance to virulent Listeria monocytogenesinfection. In the present study, we examined whether the increased resistance to this organism correlates with profiles of macrophage activation, given the role of the macrophage in killing this pathogen in vivo. We infected BDF1 mice with a lethal dose of virulent L. monocytogenes on day 4 of antiorthostatic suspension and 24 h later constructed profiles of macrophage activation. Viable listeria could not be detected in mice suspended in the antiorthostatic orientation 24 h after infection. Flow cytometric analysis revealed the numbers of granulocytes and mononuclear phagocytes in the spleen of infected mice were not significantly altered as a result of antiorthostatic suspension. Splenocytes from antiorthostatically suspended infected mice produced increased titers of IL-1. Serum levels of neopterin, a nucleotide metabolite secreted by activated macrophages, were enhanced in mice infected during antiorthostatic suspension, but not in antiorthostatically suspended naive mice. Splenic macrophages from mice infected on day 4 of suspension produced enhanced levels of lysozyme. In contrast to the results from antiorthostatically suspended infected mice, macrophages from antiorthostatically suspended uninfected mice did not express enhanced bactericidal activities. The collective results indicate that antiorthostatic suspension can stimulate profiles of macrophage activation which correlate with increased resistance to infection by certain classes of pathogenic bacteria.

  20. Macrophage mitochondrial and stress response to ingestion of Cryptococcus neoformans

    PubMed Central

    Coelho, Carolina; Souza, Ana Camila Oliveira; Derengowski, Lorena da Silveira; de Leon-Rodriguez, Carlos; Wang, Bo; Leon-Rivera, Rosiris; Bocca, Anamelia Lorenzetti; Gonçalves, Teresa; Casadevall, Arturo

    2015-01-01

    Human infection with Cryptococcus neoformans (Cn), a common fungal pathogen follows deposition of yeast spores in the lung alveoli. The subsequent host-pathogen interaction can result in either eradication, latency or extra-pulmonary dissemination. Successful control of Cn infection is dependent on host macrophages but macrophages display little ability to kill Cn in vitro. Recently, we reported that ingestion of Cn by mouse macrophages induces early cell cycle progression followed by mitotic arrest, an event that almost certainly reflects host cell damage. The goal of the present work was to understand macrophage pathways affected by Cn toxicity. Infection of macrophages by Cn was associated with alterations in protein translation rate and activation of several stress pathways such as Hypoxia Inducing Factor-1α (HIF-1α), Receptor-interacting Protein 1 (RIP1) and Apoptosis Inducing Factor (AIF). Concomitantly we observed mitochondrial depolarization in infected macrophages, an observation that was replicated in vivo. We also observed differences in the stress pathways activated depending on macrophage cell type, consistent with the non-specific nature of Cn virulence known to infect phylogenetically distant hosts. Our results indicate that Cn infection impairs multiple host cellular functions and undermines the health of these critical phagocytic cells, which can potentially interfere with their ability to clear this fungal pathogen. PMID:25646306

  1. Suppression of developmental anomalies by maternal macrophages in mice

    SciTech Connect

    Nomura, T.; Hata, S.; Kusafuka, T. )

    1990-11-01

    We tested whether nonspecific tumoricidal immune cells can suppress congenital malformations by killing precursor cells destined to cause such defects. Pretreatment of pregnant ICR mice with synthetic (Pyran copolymer) and biological (Bacillus Calmette-Guerin) agents significantly suppressed radiation- and chemical-induced congenital malformations (cleft palate, digit anomalies, tail anomalies, etc.). Such suppressive effects were associated with the activation of maternal macrophages by these agents, but were lost either after the disruption of activated macrophages by supersonic waves or by inhibition of their lysosomal enzyme activity with trypan blue. These results indicate that a live activated macrophage with active lysosomal enzymes can be an effector cell to suppress maldevelopment. A similar reduction by activated macrophages was observed in strain CL/Fr, which has a high spontaneous frequency of cleft lips and palates. Furthermore, Pyran-activated maternal macrophages could pass through the placenta, and enhanced urethane-induced cell killing (but not somatic mutation) in the embryo. It is likely that a maternal immunosurveillance system eliminating preteratogenic cells allows for the replacement with normal totipotent blast cells during the pregnancy to protect abnormal development.

  2. Anatomy of a Discovery: M1 and M2 Macrophages

    PubMed Central

    Mills, Charles Dudley

    2015-01-01

    M1 and M2 macrophage-type responses kill or repair in vivo. The unique ability of macrophages to make these polar opposite type of responses provides primary host protection and maintains tissue homeostasis throughout the animal kingdom. In humans and other higher animals, M1 and M2-type macrophage responses also initiate and direct T cells/adaptive immunity to provide additional protection such as Th1 (cytotoxic) or Th2 (antibody-mediated) type responses. Hence, macrophages were renamed M1 and M2 to indicate the central role of macrophages/innate immunity in immune systems. These findings indicate that the long held notion that adaptive immunity controls innate immunity was backward: a sea change in understanding how immune responses occur. The clinical impact of M1/kill and M2/repair responses is immense playing pivotal roles in curing (or causing) many diseases including infections, cancer, autoimmunity, and atherosclerosis. How M1/M2 came to be is an interesting story that, like life, involved Direction, Determination, Discouragement, and Discovery. PMID:25999950

  3. Purinergic Signaling to Terminate TLR Responses in Macrophages

    PubMed Central

    Hamidzadeh, Kajal; Mosser, David M.

    2016-01-01

    Macrophages undergo profound physiological alterations when they encounter pathogen-associated molecular patterns (PAMPs). These alterations can result in the elaboration of cytokines and mediators that promote immune responses and contribute to the clearance of pathogens. These innate immune responses by myeloid cells are transient. The termination of these secretory responses is not due to the dilution of stimuli, but rather to the active downregulation of innate responses induced by the very PAMPs that initiated them. Here, we describe a purinergic autoregulatory program whereby TLR-stimulated macrophages control their activation state. In this program, TLR-stimulated macrophages undergo metabolic alterations that result in the production of ATP and its release through membrane pannexin channels. This purine nucleotide is rapidly hydrolyzed to adenosine by ectoenzymes on the macrophage surface, CD39 and CD73. Adenosine then signals through the P1 class of seven transmembrane receptors to induce a regulatory state that is characterized by the downregulation of inflammatory cytokines and the production of anti-inflammatory cytokines and growth factors. This purinergic autoregulatory system mitigates the collateral damage that would be caused by the prolonged activation of macrophages and rather allows the macrophage to maintain homeostasis. The transient activation of macrophages can be prolonged by treating macrophages with IFN-γ. IFN-γ-treated macrophages become less sensitive to the regulatory effects of adenosine, allowing them to sustain macrophage activation for the duration of an adaptive immune response. PMID:26973651

  4. Purinergic Signaling to Terminate TLR Responses in Macrophages.

    PubMed

    Hamidzadeh, Kajal; Mosser, David M

    2016-01-01

    Macrophages undergo profound physiological alterations when they encounter pathogen-associated molecular patterns (PAMPs). These alterations can result in the elaboration of cytokines and mediators that promote immune responses and contribute to the clearance of pathogens. These innate immune responses by myeloid cells are transient. The termination of these secretory responses is not due to the dilution of stimuli, but rather to the active downregulation of innate responses induced by the very PAMPs that initiated them. Here, we describe a purinergic autoregulatory program whereby TLR-stimulated macrophages control their activation state. In this program, TLR-stimulated macrophages undergo metabolic alterations that result in the production of ATP and its release through membrane pannexin channels. This purine nucleotide is rapidly hydrolyzed to adenosine by ectoenzymes on the macrophage surface, CD39 and CD73. Adenosine then signals through the P1 class of seven transmembrane receptors to induce a regulatory state that is characterized by the downregulation of inflammatory cytokines and the production of anti-inflammatory cytokines and growth factors. This purinergic autoregulatory system mitigates the collateral damage that would be caused by the prolonged activation of macrophages and rather allows the macrophage to maintain homeostasis. The transient activation of macrophages can be prolonged by treating macrophages with IFN-γ. IFN-γ-treated macrophages become less sensitive to the regulatory effects of adenosine, allowing them to sustain macrophage activation for the duration of an adaptive immune response. PMID:26973651

  5. Host and Bacterial Factors Involved in the Innate Ability of Mouse Macrophages To Eliminate Internalized Unopsonized Escherichia coli

    PubMed Central

    Hamrick, Terri S.; Havell, Edward A.; Horton, John R.; Orndorff, Paul E.

    2000-01-01

    In an effort to better understand genetic and cellular factors that influence innate immunity, we examined host and bacterial factors involved in the nonopsonic phagocytosis and killing of Escherichia coli K-12 by mouse macrophages. Unelicited (resident) peritoneal macrophages from five different mouse strains, BALB/c, C57BL/6, CD-1, C3H/HeJ, and C3H/HeN, were employed. Additional macrophage populations were obtained from CD-1 mice (bone marrow-derived macrophages). Also, for BALB/c and C57BL/6 mice, peritoneal macrophages elicited with either thioglycolate or proteose peptone, bone marrow-derived macrophages, and macrophage-like cell lines derived from the two strains were employed. Two E. coli K-12 strains that differed specifically in their abilities to produce type 1 pili containing the adhesive protein FimH were examined. The parameters used to assess macrophage bacteriocidal activity were (i) the killing of internalized (gentamicin-protected) E. coli during the approximately 4-h assay and (ii) the initial rate at which internalized E. coli were eliminated. Data on these parameters allowed the following conclusions: (i) unelicited or proteose peptone-elicited peritoneal macrophages were significantly better at eliminating internalized bacteria than thioglycolate-elicited peritoneal macrophages, bone marrow-derived macrophages, or macrophage cell lines; (ii) the host genetic background had no significant effect upon the ability of unelicited peritoneal macrophages to kill E. coli (even though the mouse strains differ widely in their in vivo susceptibilities to bacterial infection); and (iii) the FimH phenotype had no significant effect upon E. coli survival once the bacterium was inside a macrophage. Additionally, there was no correlation between the bacteriocidal effectiveness of a macrophage population and the number of bacteria bound per macrophage. However, macrophage populations that were the least bacteriocidal tended to bind higher ratios of FimH+ to Fim

  6. The killing efficiency of soft iron shot

    USGS Publications Warehouse

    Andrews, R.; Longcore, J.R.

    1969-01-01

    A cooperative research effort between the ammunition industry and the Bureau of Sport Fisheries and Wildlife is aimed at finding a suitable non-toxic substitute for lead shot. A contract study by an independent research organization evaluated ways of coating or detoxifying lead shot or replacing it with another metal. As a result of that study, the only promising candidate is soft iron. Previous tests of hard iron shot had suggested that its killing effectiveness was poor at longer ranges due to the lower density. In addition, its hardness caused excessive damage to shotgun barrels. A unique, automated shooting facility was constructed at the Patuxent Wildlife Research Center to test the killing effectiveness of soft iron shot under controlled conditions. Tethered game-farm mallards were transported across a shooting point in a manner simulating free flight. A microswitch triggered a mounted shotgun so that each shot was 'perfect.' A soft iron shot, in Number 4 size, was produced by the ammunition industry and loaded in 12-gauge shells to give optimum ballistic performance. Commercial loads of lead shot in both Number 4 and Number 6 size were used for comparison. A total of 2,010 ducks were shot at ranges of 30 to 65 yards and at broadside and head-on angles in a statistically designed procedure. The following data were recorded for each duck: time until death, broken wing or leg bones, and number of embedded shot. Those ducks not killed outright were held for 10 days. From these data, ducks were categorized as 'probably bagged,' 'probably lost cripples,' or survivors. The test revealed that the killing effectiveness of this soft iron shot was superior to its anticipated performance and close to that obtained with commercial lead loads containing an equal number of pellets. Bagging a duck, in terms of rapid death or broken wing, was primarily dependent on the probability of a shot striking that vital area, and therefore a function of range. There was no indication

  7. Anion Exchanger 2 Regulates Dectin-1-Dependent Phagocytosis and Killing of Candida albicans.

    PubMed

    Urso, Katia; Charles, Julia F; Shull, Gary E; Aliprantis, Antonios O; Balestrieri, Barbara

    2016-01-01

    Anion exchanger 2 (Ae2; gene symbol, Slc4a2) is a plasma membrane Cl-/HCO3- exchanger expressed in the gastrointestinal tract, kidney and bone. We have previously shown that Ae2 is required for the function of osteoclasts, bone resorbing cells of the macrophage lineage, to maintain homeostatic cytoplasmic pH and electroneutrality during acid secretion. Macrophages require endosomal acidification for pathogen killing during the process known as phagocytosis. Chloride is thought to be the principal ion responsible for maintaining electroneutrality during organelle acidification, but whether Cl-/HCO3- exchangers such as Ae2 contribute to macrophage function is not known. In this study we investigated the role of Ae2 in primary macrophages during phagocytosis. We find that Ae2 is expressed in macrophages where it regulates intracellular pH and the binding of Zymosan, a fungal cell wall derivative. Surprisingly, the transcription and surface expression of Dectin-1, the major phagocytic receptor for Candida albicans (C. albicans) and Zymosan, is reduced in the absence of Ae2. As a consequence, Zymosan-induced Tnfα expression is also impaired in Ae2-deficient macrophages. Similar to Ae2 deficiency, pharmacological alkalinization of lysosomal pH with bafilomycin A decreases both Dectin-1 mRNA and cell surface expression. Finally, Ae2-deficient macrophages demonstrate defective phagocytosis and killing of the human pathogenic fungus C. albicans. Our results strongly suggest that Ae2 is a critical factor in the innate response to C. albicans. This study represents an important contribution to a better understanding of how Dectin-1 expression and fungal clearance is regulated. PMID:27391897

  8. Anion Exchanger 2 Regulates Dectin-1-Dependent Phagocytosis and Killing of Candida albicans

    PubMed Central

    Urso, Katia; Charles, Julia F.; Shull, Gary E.; Aliprantis, Antonios O.; Balestrieri, Barbara

    2016-01-01

    Anion exchanger 2 (Ae2; gene symbol, Slc4a2) is a plasma membrane Cl-/HCO3- exchanger expressed in the gastrointestinal tract, kidney and bone. We have previously shown that Ae2 is required for the function of osteoclasts, bone resorbing cells of the macrophage lineage, to maintain homeostatic cytoplasmic pH and electroneutrality during acid secretion. Macrophages require endosomal acidification for pathogen killing during the process known as phagocytosis. Chloride is thought to be the principal ion responsible for maintaining electroneutrality during organelle acidification, but whether Cl-/HCO3- exchangers such as Ae2 contribute to macrophage function is not known. In this study we investigated the role of Ae2 in primary macrophages during phagocytosis. We find that Ae2 is expressed in macrophages where it regulates intracellular pH and the binding of Zymosan, a fungal cell wall derivative. Surprisingly, the transcription and surface expression of Dectin-1, the major phagocytic receptor for Candida albicans (C. albicans) and Zymosan, is reduced in the absence of Ae2. As a consequence, Zymosan-induced Tnfα expression is also impaired in Ae2-deficient macrophages. Similar to Ae2 deficiency, pharmacological alkalinization of lysosomal pH with bafilomycin A decreases both Dectin-1 mRNA and cell surface expression. Finally, Ae2-deficient macrophages demonstrate defective phagocytosis and killing of the human pathogenic fungus C. albicans. Our results strongly suggest that Ae2 is a critical factor in the innate response to C. albicans. This study represents an important contribution to a better understanding of how Dectin-1 expression and fungal clearance is regulated. PMID:27391897

  9. The Many Alternative Faces of Macrophage Activation

    PubMed Central

    Hume, David A.

    2015-01-01

    Monocytes and macrophages provide the first line of defense against pathogens. They also initiate acquired immunity by processing and presenting antigens and provide the downstream effector functions. Analysis of large gene expression datasets from multiple cells and tissues reveals sets of genes that are co-regulated with the transcription factors that regulate them. In macrophages, the gene clusters include lineage-specific genes, interferon-responsive genes, early inflammatory genes, and genes required for endocytosis and lysosome function. Macrophages enter tissues and alter their function to deal with a wide range of challenges related to development and organogenesis, tissue injury, malignancy, sterile, or pathogenic inflammatory stimuli. These stimuli alter the gene expression to produce “activated macrophages” that are better equipped to eliminate the cause of their influx and to restore homeostasis. Activation or polarization states of macrophages have been classified as “classical” and “alternative” or M1 and M2. These proposed states of cells are not supported by large-scale transcriptomic data, including macrophage-associated signatures from large cancer tissue datasets, where the supposed markers do not correlate with other. Individual macrophage cells differ markedly from each other, and change their functions in response to doses and combinations of agonists and time. The most studied macrophage activation response is the transcriptional cascade initiated by the TLR4 agonist lipopolysaccharide. This response is reviewed herein. The network topology is conserved across species, but genes within the transcriptional network evolve rapidly and differ between mouse and human. There is also considerable divergence in the sets of target genes between mouse strains, between individuals, and in other species such as pigs. The deluge of complex information related to macrophage activation can be accessed with new analytical tools and new databases

  10. Post-transcriptional control of NLRP3 inflammasome activation in colonic macrophages

    PubMed Central

    Filardy, Alessandra A.; He, Jianping; Bennink, Jack; Yewdell, Jonathan; Kelsall, Brian L.

    2016-01-01

    Colonic macrophages (cMPs) are important for intestinal homeostasis as they kill microbes yet produce regulatory cytokines. Activity of the NLRP3 inflammasome, a major sensor of stress and microorganisms that results in pro-inflammatory cytokine production and cell death must be tightly controlled in the intestine. We demonstrate that resident cMPs are hyporesponsive to NLRP3 inflammasome activation due to a remarkable level of post-transcriptional control of NLRP3 and proIL-1β protein expression, which was also seen for TNF-α and IL-6, but lost during experimental colitis. Resident cMPs rapidly degraded NLRP3 and proIL-1β proteins by the ubiquitin/proteasome system. Finally, blocking IL-10R-signaling in vivo enhanced NLRP3 and proIL-1β protein, but not mRNA levels in resident cMPs implicating a role for IL-10 in environmental conditioning of cMPs. These data are the first to show dramatic post-transcriptional control of inflammatory cytokine production by a relevant tissue-derived macrophage population and proteasomal degradation of proIL-1β and NLRP3 as a mechanism to control inflammasome activation; findings which have broad implications for our understanding of intestinal and systemic inflammatory diseases. PMID:26627461

  11. Fully human MAP-fusion protein selectively targets and eliminates proliferating CD64(+) M1 macrophages.

    PubMed

    Hristodorov, Dmitrij; Mladenov, Radoslav; Fischer, Rainer; Barth, Stefan; Thepen, Theo

    2016-05-01

    Classical immunotoxins compromise a binding component (for example, a ligand, antibody or fragment thereof) and a cytotoxic component, usually derived from bacteria or plants (for example, Pseudomonas exotoxin A or ricin). Despite successful testing in vitro, the clinical development of immunotoxins has been hampered by immunogenicity and unsatisfactory safety profiles. Therefore, research has focused on fully human pro-apoptotic components suitable for the development of cytolytic fusion proteins (CFP). We recently reported that human microtubule-associated protein tau (MAP) can induce apoptosis when delivered to rapidly proliferating cancer cells. Here, we describe a new fully human CFP called H22(scFv)-MAP, which specifically targets CD64(+) cells. We show that H22(scFv)-MAP can efficiently kill proliferating HL-60 pro-monocytic cells in vitro. In addition, the human CFP specifically eliminates polarized M1 macrophages in a transgenic mouse model of cutaneous chronic inflammation. Because M1 macrophages promote the pathogenesis of many chronic inflammatory diseases, targeting this cell population with H22(scFv)-MAP could help to treat diseases such as atopic dermatitis, rheumatoid arthritis and inflammatory bowel disease.

  12. Beetle Kill Wall at NREL

    SciTech Connect

    2010-01-01

    When it comes to designing an interior decorative feature for one of the most energy efficient office buildings in the world, very few would consider bringing in a beetle to do the job. But thats what happened at the U.S. Department of Energy's (DOE) Research Support Facility (RSF) located on the National Renewable Energy Laboratory (NREL) campus.In June, the RSF will become home to more than 800 workers from DOE and NREL and building visitors will be greeted with a soaring, two-story high wall entirely covered with wood harvested from the bark beetle infestation that has killed millions of pine trees in the Western U.S. But, the use of beetle kill wood is just one example of the resources being leveraged to make the RSF a model for sustainability and one more step toward NRELs goal to be a net zero energy campus.

  13. Beetle Kill Wall at NREL

    ScienceCinema

    None

    2016-07-12

    When it comes to designing an interior decorative feature for one of the most energy efficient office buildings in the world, very few would consider bringing in a beetle to do the job. But thats what happened at the U.S. Department of Energy's (DOE) Research Support Facility (RSF) located on the National Renewable Energy Laboratory (NREL) campus.In June, the RSF will become home to more than 800 workers from DOE and NREL and building visitors will be greeted with a soaring, two-story high wall entirely covered with wood harvested from the bark beetle infestation that has killed millions of pine trees in the Western U.S. But, the use of beetle kill wood is just one example of the resources being leveraged to make the RSF a model for sustainability and one more step toward NRELs goal to be a net zero energy campus.

  14. Women who kill their mates.

    PubMed

    Bourget, Dominique; Gagné, Pierre

    2012-01-01

    Spousal homicide perpetrators are much more likely to be men than women. Accordingly, little research has focused on delineating characteristics of women who have committed spousal homicide. A retrospective clinical review of coroners' files containing all cases of spousal homicide occurring in Quebec over a 20-year period was carried out. A total of 276 spousal homicides occurred between 1991 and 2010, with 42 homicides by female spouses and 234 homicides by male spouses. Differences between homicides committed by female offenders and male offenders are discussed, and findings on spousal homicide committed by women are compared with those of previous studies. Findings regarding offenses perpetrated by females in the context of mental illness, domestic violence, and homicide-suicide are explored. The finding that only 28% of the female offenders in the Quebec sample had previously been subjected to violence by their victim is in contrast to the popular belief and reports that indicate that most female-perpetrated spousal homicide occurs in self-defense or in reaction to long-term abuse. In fact, women rarely gave a warning before killing their mates. Most did not suffer from a mental illness, although one-fifth were acutely intoxicated at the time of the killing. In the vast majority of cases of women who killed their mates, there were very few indicators that might have signaled the risk and helped predict the violent lethal behavior.

  15. Degradation of parathyroid hormone in macrophage endosomes

    SciTech Connect

    Diment, S.; Martin, K.J.; Stahl, P.D.

    1986-05-01

    Parathyroid hormone (PTH) is secreted as an 84 amino acid protein that is rapidly cleaved between amino acids 34 and 35 by Kupffer cells in liver. The resulting amino terminal peptide (1-34) is active at PTH target organs (kidney and bone). Cathepsin D can process PTH to 1-34 in vitro, and a cathepsin D-like protease, which may rapidly process proteins, is present in endosomes of alveolar macrophages. The authors set out to determine whether PTH is degraded to 1-34 in endosomes, and to elucidate the mechanism of hormone processing in vivo. Intracellular transport of /sup 125/I-PTH was assessed by binding to alveolar macrophages at 4/sup 0/C, followed by internalization at 37/sup 0/C. Distribution of PTH among plasma membranes, endosomes and lysosomes was determined by subcellular fractionation. Degradation of the ligand to TCA-soluble fragments in each compartment was assayed at neutral and acid pH. 1-34 in supernatants was separated from undergraded PTH by gel filtration and detected by bioassay on kidney membranes. The authors data suggest that: 1) macrophages rapidly degrade PTH to TCA-soluble fragments. 2) macrophages do not secrete proteases that degrade extracellular PTH. 3) PTH is internalized into endocytic vesicles after 5 mins, but not delivered to lysosomes within 30 mins. 4) A bioactive peptide is released into the extracellular medium after 20 mins. 5) PTH is degraded in endosomes at acid pH by a pepstatin-sensitive protease.

  16. 33 CFR 117.801 - Newtown Creek, Dutch Kills, English Kills and their tributaries.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 1 2014-07-01 2014-07-01 false Newtown Creek, Dutch Kills, English Kills and their tributaries. 117.801 Section 117.801 Navigation and Navigable Waters COAST GUARD....801 Newtown Creek, Dutch Kills, English Kills and their tributaries. (a) The following...

  17. 33 CFR 117.801 - Newtown Creek, Dutch Kills, English Kills and their tributaries.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Newtown Creek, Dutch Kills, English Kills and their tributaries. 117.801 Section 117.801 Navigation and Navigable Waters COAST GUARD....801 Newtown Creek, Dutch Kills, English Kills and their tributaries. (a) The following...

  18. 33 CFR 117.801 - Newtown Creek, Dutch Kills, English Kills and their tributaries.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Newtown Creek, Dutch Kills, English Kills and their tributaries. 117.801 Section 117.801 Navigation and Navigable Waters COAST GUARD....801 Newtown Creek, Dutch Kills, English Kills and their tributaries. (a) The following...

  19. 33 CFR 117.801 - Newtown Creek, Dutch Kills, English Kills and their tributaries.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Newtown Creek, Dutch Kills, English Kills and their tributaries. 117.801 Section 117.801 Navigation and Navigable Waters COAST GUARD....801 Newtown Creek, Dutch Kills, English Kills and their tributaries. (a) The following...

  20. 33 CFR 117.801 - Newtown Creek, Dutch Kills, English Kills and their tributaries.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 1 2013-07-01 2013-07-01 false Newtown Creek, Dutch Kills, English Kills and their tributaries. 117.801 Section 117.801 Navigation and Navigable Waters COAST GUARD....801 Newtown Creek, Dutch Kills, English Kills and their tributaries. (a) The following...

  1. LPS-inducible factor(s) from activated macrophages mediates cytolysis of Naegleria fowleri amoebae

    SciTech Connect

    Cleary, S.F.; Marciano-Cabral, F.

    1986-03-01

    Soluble cytolytic factors of macrophage origin have previously been described with respect to their tumoricidal activity. The purpose of this study was to investigate the mechanism and possible factor(s) responsible for cytolysis of the amoeba Naegleria fowleri by activated peritoneal macrophages from B6C3F1 mice. Macrophages or conditioned medium (CM) from macrophage cultures were incubated with /sup 3/H-Uridine labeled amoebae. Percent specific release of label served as an index of cytolysis. Bacille Calmette-Guerin (BCG) and Corynebacterium parvum macrophages demonstrated significant cytolysis of amoebae at 24 h with an effector to target ratio of 10:1. Treatment of macrophages with inhibitors of RNA or protein synthesis blocked amoebicidal activity. Interposition of a 1 ..mu..m pore membrane between macrophages and amoebae inhibited killing. Inhibition in the presence of the membrane was overcome by stimulating the macrophages with LPS. CM from SPS-stimulated, but not unstimulated, cultures of activated macrophages was cytotoxic for amoebae. The activity was heat sensitive and was recovered from ammonium sulfate precipitation of the CM. Results indicate that amoebicidal activity is mediated by a protein(s) of macrophage origin induced by target cell contact or stimulation with LPS.

  2. Glutamine Modulates Macrophage Lipotoxicity

    PubMed Central

    He, Li; Weber, Kassandra J.; Schilling, Joel D.

    2016-01-01

    Obesity and diabetes are associated with excessive inflammation and impaired wound healing. Increasing evidence suggests that macrophage dysfunction is responsible for these inflammatory defects. In the setting of excess nutrients, particularly dietary saturated fatty acids (SFAs), activated macrophages develop lysosome dysfunction, which triggers activation of the NLRP3 inflammasome and cell death. The molecular pathways that connect lipid stress to lysosome pathology are not well understood, but may represent a viable target for therapy. Glutamine uptake is increased in activated macrophages leading us to hypothesize that in the context of excess lipids glutamine metabolism could overwhelm the mitochondria and promote the accumulation of toxic metabolites. To investigate this question we assessed macrophage lipotoxicity in the absence of glutamine using LPS-activated peritoneal macrophages exposed to the SFA palmitate. We found that glutamine deficiency reduced lipid induced lysosome dysfunction, inflammasome activation, and cell death. Under glutamine deficient conditions mTOR activation was decreased and autophagy was enhanced; however, autophagy was dispensable for the rescue phenotype. Rather, glutamine deficiency prevented the suppressive effect of the SFA palmitate on mitochondrial respiration and this phenotype was associated with protection from macrophage cell death. Together, these findings reveal that crosstalk between activation-induced metabolic reprogramming and the nutrient microenvironment can dramatically alter macrophage responses to inflammatory stimuli. PMID:27077881

  3. Killing(-Yano) tensors in string theory

    NASA Astrophysics Data System (ADS)

    Chervonyi, Yuri; Lunin, Oleg

    2015-09-01

    We construct the Killing(-Yano) tensors for a large class of charged black holes in higher dimensions and study general properties of such tensors, in particular, their behavior under string dualities. Killing(-Yano) tensors encode the symmetries beyond isometries, which lead to insights into dynamics of particles and fields on a given geometry by providing a set of conserved quantities. By analyzing the eigenvalues of the Killing tensor, we provide a prescription for constructing several conserved quantities starting from a single object, and we demonstrate that Killing tensors in higher dimensions are always associated with ellipsoidal coordinates. We also determine the transformations of the Killing(-Yano) tensors under string dualities, and find the unique modification of the Killing-Yano equation consistent with these symmetries. These results are used to construct the explicit form of the Killing(-Yano) tensors for the Myers-Perry black hole in arbitrary number of dimensions and for its charged version.

  4. Biodegradation of carbon nanohorns in macrophage cells

    NASA Astrophysics Data System (ADS)

    Zhang, Minfang; Yang, Mei; Bussy, Cyrill; Iijima, Sumio; Kostarelos, Kostas; Yudasaka, Masako

    2015-02-01

    With the rapid developments in the medical applications of carbon nanomaterials such as carbon nanohorns (CNHs), carbon nanotubes, and graphene based nanomaterials, understanding the long-term fate, health impact, excretion, and degradation of these materials has become crucial. Herein, the in vitro biodegradation of CNHs was determined using a non-cellular enzymatic oxidation method and two types of macrophage cell lines. Approximately 60% of the CNHs was degraded within 24 h in a phosphate buffer solution containing myeloperoxidase. Furthermore, approximately 30% of the CNHs was degraded by both RAW 264.7 and THP-1 macrophage cells within 9 days. Inflammation markers such as pro-inflammatory cytokines interleukin 6 and tumor necrosis factor α were not induced by exposure to CNHs. However, reactive oxygen species were generated by the macrophage cells after uptake of CNHs, suggesting that these species were actively involved in the degradation of the nanomaterials rather than in an inflammatory pathway induction.With the rapid developments in the medical applications of carbon nanomaterials such as carbon nanohorns (CNHs), carbon nanotubes, and graphene based nanomaterials, understanding the long-term fate, health impact, excretion, and degradation of these materials has become crucial. Herein, the in vitro biodegradation of CNHs was determined using a non-cellular enzymatic oxidation method and two types of macrophage cell lines. Approximately 60% of the CNHs was degraded within 24 h in a phosphate buffer solution containing myeloperoxidase. Furthermore, approximately 30% of the CNHs was degraded by both RAW 264.7 and THP-1 macrophage cells within 9 days. Inflammation markers such as pro-inflammatory cytokines interleukin 6 and tumor necrosis factor α were not induced by exposure to CNHs. However, reactive oxygen species were generated by the macrophage cells after uptake of CNHs, suggesting that these species were actively involved in the degradation of the

  5. Fish kill from underwater explosions

    USGS Publications Warehouse

    Stuart, David J.

    1962-01-01

    The U.S. Geological Survey has used 23 different shotpoints during two seasons of field work in our seismic study of crustal structure in western United States. Without exception, it has been found that under-water shotpoints result in a more efficient conversion of explosive energy into seismic energy than do drilled-hole shotpoints. This experience, together with elimination of drilling costs, has led to the use of underwater shotpoints wherever possible. Three of the 23 shotpoints were in the Pacific Ocean, and for these we have no detailed information on the fish kill. Another six shotpoints were located in inland bodies of water. These are: * Soda Lake near Fallon, Nevada * Mono Lake near Lee Vining, California * Lake Mead near Boulder City, Nevada * Shasta Lake near Redding, California * C.J. Strike Reservoir near Bruneau, Idaho * Lucky Peak Reservoir near Boise, Idaho The 22 high-explosive charges, weighing a total of 95,100 pounds, that were fired in lakes containing fish life resulted in the known death of 2,413 game fish with a total weight of 759 pounds. The average mortality was 110 game fish or 34.5 pounds of game fish killed per average shot of 4,325 pounds of high-explosives.

  6. Receptor-Mediated Drug Delivery to Macrophages in Chemotherapy of Leishmaniasis

    NASA Astrophysics Data System (ADS)

    Mukhopadhyay, Amitabha; Chaudhuri, Gautam; Arora, Sunil K.; Sehgal, Shobha; Basu, Sandip K.

    1989-05-01

    Methotrexate coupled to maleylated bovine serum albumin was taken up efficiently through the ``scavenger'' receptors present on macrophages and led to selective killing of intracellular Leishmania mexicana amazonensis amastigotes in cultured hamster peritoneal macrophages. The drug conjugate was nearly 100 times as effective as free methotrexate in eliminating the intracellular parasites. Furthermore, in a model of experimental cutaneous leishmaniasis in hamsters, the drug conjugate brought about more than 90% reduction in the size of footpad lesions within 11 days. In contrast, the free drug at a similar concentration did not significantly affect lesion size. These studies demonstrate the potential of receptor-mediated drug delivery in the therapy of macrophage-associated diseases.

  7. Macrophages in gene therapy: cellular delivery vehicles and in vivo targets.

    PubMed

    Burke, B; Sumner, S; Maitland, N; Lewis, C E

    2002-09-01

    The appearance and activation of macrophages are thought to be rapid events in the development of many pathological lesions, including malignant tumors, atherosclerotic plaques, and arthritic joints. This has prompted recent attempts to use macrophages as novel cellular vehicles for gene therapy, in which macrophages are genetically modified ex vivo and then reintroduced into the body with the hope that a proportion will then home to the diseased site. Here, we critically review the efficacy of various gene transfer methods (viral, bacterial, protozoan, and various chemical and physical methods) in transfecting macrophages in vitro, and the results obtained when transfected macrophages are used as gene delivery vehicles. Finally, we discuss the use of various viral and nonviral methods to transfer genes to macrophages in vivo. As will be seen, definitive evidence for the use of macrophages as gene transfer vehicles has yet to be provided and awaits detailed trafficking studies in vivo. Moreover, although methods for transfecting macrophages have improved considerably in efficiency in recent years, targeting of gene transfer specifically to macrophages in vivo remains a problem. However, possible solutions to this include placing transgenes under the control of macrophage-specific promoters to limit expression to macrophages or stably transfecting CD34(+) precursors of monocytes/macrophages and then differentiating these cells into monocytes/macrophages ex vivo. The latter approach could conceivably lead to the bone marrow precursor cells of patients with inherited genetic disorders being permanently fortified or even replaced with genetically modified cells. PMID:12223508

  8. Macrophage tumoricidal mechanisms are selectively altered by prenatal chlordane exposure.

    PubMed

    Theus, S A; Tabor, D R; Soderberg, L S; Barnett, J B

    1992-09-01

    Macrophages (m phi) derived from mice treated in utero with chlordane show a significant delay of tumoricidal induction activity. In this study, m phi from chlordane-treated animals required a 48 h in vitro period of induction with interferon-gamma and lipopolysaccharide (IFN/LPS) before they could kill P815 targets. Similarly, m phi from chlordane-treated animals also failed to produce an immediate H2O2 burst upon perturbation. Conversely, their stimulated control m phi counterparts were tumoricidal by 2 h and exhibited a respiratory burst without any delay. Moreover, levels of the second messenger, inositol triphosphate (IP3), were significantly delayed in chlordane-treated animals following interaction with IFN/LPS. When nitrate/nitrite production was analyzed as an alternate mechanism for killing tumors, stimulated m phi from both normal and chlordane-treated animals responded equally. The data show that chlordane differentially introduces defects in m phi biochemical mechanisms associated with tumor killing.

  9. Tissue-resident macrophages

    PubMed Central

    Davies, Luke C.; Jenkins, Stephen J.; Allen, Judith E.; Taylor, Philip R.

    2014-01-01

    Tissue-resident macrophages are a heterogeneous population of immune cells that fulfill tissue-specific and niche-specific functions. These range from dedicated homeostatic functions, such as clearance of cellular debris and iron processing, to central roles in tissue immune-surveillance, response to infection and the resolution of inflammation. Recent studies highlight marked heterogeneity in the origins of tissue macrophages that arise from hematopoietic versus self-renewing embryo-derived populations. We discuss the tissue–niche-specific factors that dictate cell phenotype, the definition of which will allow novel strategies to promote the restoration of tissue homeostasis. Understanding the mechanisms that dictate tissue macrophage heterogeneity should explain why simplified paradigms of macrophage activation do not explain the extent of heterogeneity seen in vivo. PMID:24048120

  10. IL-33 Priming Enhances Peritoneal Macrophage Activity in Response to Candida albicans.

    PubMed

    Tran, Vuvi G; Cho, Hong R; Kwon, Byungsuk

    2014-08-01

    IL-33 is a member of the IL-1 cytokine family and plays a role in the host defense against bacteria, viruses, and fungi. In this study, we investigated the function of IL-33 and its receptor in in vitro macrophage responses to Candida albicans. Our results demonstrate that pre-sensitization of isolated peritoneal macrophages with IL-33 enhanced their pro-inflammatory cytokine production and phagocytic activity in response to C. albicans. These macrophage activities were entirely dependent on the ST2-MyD88 signaling pathway. In addition, pre-sensitization with IL-33 also increased ROS production and the subsequent killing ability of macrophages following C. albicans challenge. These results indicate that IL-33 may increase anti-fungal activity against Candida through macrophage-mediated resistance mechanisms. PMID:25177252

  11. Interferon-γ regulates cellular metabolism and mRNA translation to potentiate macrophage activation.

    PubMed

    Su, Xiaodi; Yu, Yingpu; Zhong, Yi; Giannopoulou, Eugenia G; Hu, Xiaoyu; Liu, Hui; Cross, Justin R; Rätsch, Gunnar; Rice, Charles M; Ivashkiv, Lionel B

    2015-08-01

    Interferon-γ (IFN-γ) primes macrophages for enhanced microbial killing and inflammatory activation by Toll-like receptors (TLRs), but little is known about the regulation of cell metabolism or mRNA translation during this priming. We found that IFN-γ regulated the metabolism and mRNA translation of human macrophages by targeting the kinases mTORC1 and MNK, both of which converge on the selective regulator of translation initiation eIF4E. Physiological downregulation of mTORC1 by IFN-γ was associated with autophagy and translational suppression of repressors of inflammation such as HES1. Genome-wide ribosome profiling in TLR2-stimulated macrophages showed that IFN-γ selectively modulated the macrophage translatome to promote inflammation, further reprogram metabolic pathways and modulate protein synthesis. These results show that IFN-γ-mediated metabolic reprogramming and translational regulation are key components of classical inflammatory macrophage activation.

  12. The Elusive Antifibrotic Macrophage

    PubMed Central

    Adhyatmika, Adhyatmika; Putri, Kurnia S. S.; Beljaars, Leonie; Melgert, Barbro N.

    2015-01-01

    Fibrotic diseases, especially of the liver, the cardiovascular system, the kidneys, and the lungs, account for approximately 45% of deaths in Western societies. Fibrosis is a serious complication associated with aging and/or chronic inflammation or injury and cannot be treated effectively yet. It is characterized by excessive deposition of extracellular matrix (ECM) proteins by myofibroblasts and impaired degradation by macrophages. This ultimately destroys the normal structure of an organ, which leads to loss of function. Most efforts to develop drugs have focused on inhibiting ECM production by myofibroblasts and have not yielded many effective drugs yet. Another option is to stimulate the cells that are responsible for degradation and uptake of excess ECM, i.e., antifibrotic macrophages. However, macrophages are plastic cells that have many faces in fibrosis, including profibrotic behavior-stimulating ECM production. This can be dependent on their origin, as the different organs have tissue-resident macrophages with different origins and a various influx of incoming monocytes in steady-state conditions and during fibrosis. To be able to pharmacologically stimulate the right kind of behavior in fibrosis, a thorough characterization of antifibrotic macrophages is necessary, as well as an understanding of the signals they need to degrade ECM. In this review, we will summarize the current state of the art regarding the antifibrotic macrophage phenotype and the signals that stimulate its behavior. PMID:26618160

  13. Supplementation of host response by targeting nitric oxide to the macrophage cytosol is efficacious in the hamster model of visceral leishmaniasis and adds to efficacy of amphotericin B.

    PubMed

    Pandya, Sanketkumar; Verma, Rahul Kumar; Khare, Prashant; Tiwari, Brajendra; Srinivasarao, Dadi A; Dube, Anuradha; Goyal, Neena; Misra, Amit

    2016-08-01

    We investigated efficacy of nitric oxide (NO) against Leishmania donovani. NO is a mediator of host response to infection, with direct parasiticidal activity in addition to its role in signalling to evoke innate macrophage responses. However, it is short-lived and volatile, and is therefore difficult to introduce into infected cells and maintain inracellular concentrations for meaningful periods of time. We incorporated diethylenetriamine NO adduct (DETA/NO), a prodrug, into poly(lactide-co-glycolide) particles of ∼200 nm, with or without amphotericin B (AMB). These particles sustained NO levels in mouse macrophage culture supernatants, generating an area under curve (AUC0.08-24h) of 591.2 ± 95.1 mM × h. Free DETA/NO resulted in NO peaking at 3 h and declining rapidly to yield an AUC of 462.5 ± 193.4. Particles containing AMB and DETA/NO were able to kill ∼98% of promastigotes and ∼76% of amastigotes in 12 h when tested in vitro. Promastigotes and amastigotes were killed less efficiently by particles containing a single drug- either DETA/NO (∼42%, 35%) or AMB (∼90%, 50%) alone, or by equivalent concentrations of drugs in solution. In a pre-clinical efficacy study of power >0.95 in the hamster model, DETA/NO particles were non-inferior to Fungizone® but not Ambisome®, resulting in significant (∼73%) reduction in spleen parasites in 7 days. Particles containing both DETA/NO and AMB were superior (∼93% reduction) to Ambisome®. We conclude that NO delivered to the cytosol of macrophages infected with Leishmania possesses intrinsic activity and adds significantly to the efficacy of AMB. PMID:27183429

  14. Macrophage polarization in inflammatory diseases.

    PubMed

    Liu, Yan-Cun; Zou, Xian-Biao; Chai, Yan-Fen; Yao, Yong-Ming

    2014-01-01

    Diversity and plasticity are two hallmarks of macrophages. M1 macrophages (classically activated macrophages) are pro-inflammatory and have a central role in host defense against infection, while M2 macrophages (alternatively activated macrophages) are associated with responses to anti-inflammatory reactions and tissue remodeling, and they represent two terminals of the full spectrum of macrophage activation. Transformation of different phenotypes of macrophages regulates the initiation, development, and cessation of inflammatory diseases. Here we reviewed the characters and functions of macrophage polarization in infection, atherosclerosis, obesity, tumor, asthma, and sepsis, and proposed that targeting macrophage polarization and skewing their phenotype to adapt to the microenvironment might hold great promise for the treatment of inflammatory diseases.

  15. Timelike Killing spinors in seven dimensions

    SciTech Connect

    Cariglia, Marco; Conamhna, Oisin A.P. Mac

    2004-12-15

    We employ the G-structure formalism to study supersymmetric solutions of minimal and SU(2) gauged supergravities in seven dimensions admitting Killing spinors with an associated timelike Killing vector. The most general such Killing spinor defines a SU(3) structure. We deduce necessary and sufficient conditions for the existence of a timelike Killing spinor on the bosonic fields of the theories, and find that such configurations generically preserve one out of 16 supersymmetries. Using our general supersymmetric ansatz we obtain numerous new solutions, including squashed or deformed anti-de Sitter solutions of the gauged theory, and a large class of Goedel-like solutions with closed timelike curves.

  16. A kill curve for Phanerozoic marine species

    NASA Technical Reports Server (NTRS)

    Raup, D. M.

    1991-01-01

    A kill curve for Phanerozoic species is developed from an analysis of the stratigraphic ranges of 17,621 genera, as compiled by Sepkoski. The kill curve shows that a typical species' risk of extinction varies greatly, with most time intervals being characterized by very low risk. The mean extinction rate of 0.25/m.y. is thus a mixture of long periods of negligible extinction and occasional pulses of much higher rate. Because the kill curve is merely a description of the fossil record, it does not speak directly to the causes of extinction. The kill curve may be useful, however, to li inverted question markmit choices of extinction mechanisms.

  17. Macrophage activation-induced thymosin beta 4 production: a tissue repair mechanism

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Macrophages play significant role in immunity which not only kill pathogens, produce cytokines but also clear dead tissues at the site of inflammation and stimulate wound healing. Much less is known how these cells contribute to tissue repair process. In course of our studies comparing the peptide...

  18. Tumour necrosis factor (TNF) as a mediator of macrophage helminthotoxic activity.

    PubMed

    James, S L; Glaven, J; Goldenberg, S; Meltzer, M S; Pearce, E

    1990-01-01

    Lymphokine-activated macrophages are cytotoxic for larvae of the helminth parasite Schistosoma mansoni. That soluble secreted factors may mediate this cytotoxicity was suggested by the observation that culture supernatant fluids from stimulated macrophages also exhibited larvacidal activity. These fluids contain the monokine tumour necrosis factor (TNF). Several observations indicated that TNF is directly toxic to schistosome larvae. Cytotoxic sera taken from BCG- or S. mansoni-immunized mice after endotoxin challenge killed schistosomula in vitro, and upon gel filtration the larvacidal factor(s) in the sera co-eluted with the tumoricidal activity defined as TNF. Recombinant-derived TNF exhibited direct toxicity to schistosomula at high concentrations, or at lower concentrations in the presence of IFN gamma. The larvacidal activity of macrophage supernatant fluids was abrogated by addition of either anti-TNF antisera or Zn+2, which has been shown to inhibit TNF-induced damage of tumour cells. Anti-TNF and Zn+2 likewise suppressed schistosomulum killing by lymphokine-activated peritoneal macrophages or the IC-21 macrophage line, indicating that TNF also plays a role in the effector mechanism of larval killing by whole cells. PMID:2314921

  19. Escherichia coli and Candida albicans Induced Macrophage Extracellular Trap-Like Structures with Limited Microbicidal Activity

    PubMed Central

    Liao, Chengshui; Liu, Xiaolei; Du, Jing; Shi, Haining; Wang, Xuelin; Bai, Xue; Peng, Peng; Yu, Lu; Wang, Feng; Zhao, Ying; Liu, Mingyuan

    2014-01-01

    The formation of extracellular traps (ETs) has recently been recognized as a novel defense mechanism in several types of innate immune cells. It has been suggested that these structures are toxic to microbes and contribute significantly to killing several pathogens. However, the role of ETs formed by macrophages (METs) in defense against microbes remains little known. In this study, we demonstrated that a subset of murine J774A.1 macrophage cell line (8% to 17%) and peritoneal macrophages (8.5% to 15%) form METs-like structures (METs-LS) in response to Escherichia coli and Candida albicans challenge. We found only a portion of murine METs-LS, which are released by dying macrophages, showed detectable killing effects on trapped E. coli but not C. albicans. Fluorescence and scanning electron microscopy analyses revealed that, in vitro, both microorganisms were entrapped in J774A.1 METs-LS composed of DNA and microbicidal proteins such as histone, myeloperoxidase and lysozyme. DNA components of both nucleus and mitochondrion origins were detectable in these structures. Additionally, METs-LS formation occurred independently of ROS produced by NADPH oxidase, and this process did not result in cell lysis. In summary, our results emphasized that microbes induced METs-LS in murine macrophage cells and that the microbicidal activity of these METs-LS differs greatly. We propose the function of METs-LS is to contain invading microbes at the infection site, thereby preventing the systemic diffusion of them, rather than significantly killing them. PMID:24587206

  20. Yersinia pestis Requires Host Rab1b for Survival in Macrophages

    PubMed Central

    Connor, Michael G.; Pulsifer, Amanda R.; Price, Christopher T.; Abu Kwaik, Yousef; Lawrenz, Matthew B.

    2015-01-01

    Yersinia pestis is a facultative intracellular pathogen that causes the disease known as plague. During infection of macrophages Y. pestis actively evades the normal phagosomal maturation pathway to establish a replicative niche within the cell. However, the mechanisms used by Y. pestis to subvert killing by the macrophage are unknown. Host Rab GTPases are central mediators of vesicular trafficking and are commonly targeted by bacterial pathogens to alter phagosome maturation and killing by macrophages. Here we demonstrate for the first time that host Rab1b is required for Y. pestis to effectively evade killing by macrophages. We also show that Rab1b is specifically recruited to the Yersinia containing vacuole (YCV) and that Y. pestis is unable to subvert YCV acidification when Rab1b expression is knocked down in macrophages. Furthermore, Rab1b knockdown also altered the frequency of association between the YCV with the lysosomal marker Lamp1, suggesting that Rab1b recruitment to the YCV directly inhibits phagosome maturation. Finally, we show that Rab1b knockdown also impacts the pH of the Legionella pneumophila containing vacuole, another pathogen that recruits Rab1b to its vacuole. Together these data identify a novel role for Rab1b in the subversion of phagosome maturation by intracellular pathogens and suggest that recruitment of Rab1b to the pathogen containing vacuole may be a conserved mechanism to control vacuole pH. PMID:26495854

  1. Human macrophage hemoglobin-iron metabolism in vitro

    SciTech Connect

    Custer, G.; Balcerzak, S.; Rinehart, J.

    1982-01-01

    An entirely in vitro technique was employed to characterize hemoglobin-iron metabolism by human macrophages obtained by culture of blood monocytes and pulmonary alveolar macrophages. Macrophages phagocytized about three times as many erythrocytes as monocytes and six times as many erythrocytes as pulmonary alveolar macrophages. The rate of subsequent release of /sup 59/Fe to the extracellular transferrin pool was two- to fourfold greater for macrophages as compared to the other two cell types. The kinetics of /sup 59/Fe-transferrin release were characterized by a relatively rapid early phase (hours 1-4) followed by a slow phase (hours 4-72) for all three cell types. Intracellular movement of iron was characterized by a rapid shift from hemoglobin to ferritin that was complete with the onset of the slow phase of extracellular release. A transient increase in /sup 59/Fe associated with an intracellular protein eluting with transferrin was also observed within 1 hour after phagocytosis. The process of hemoglobin-iron release to extracellular transferrin was inhibited at 4 degrees C but was unaffected by inhibitory of protein synthesis, glycolysis, microtubule function, and microfilament function. These data emphasize the rapidity of macrophage hemoglobin iron metabolism, provide a model for characterization of this process in vitro, and in general confirm data obtained utilizing in vivo animal models.

  2. Boromycin Kills Mycobacterial Persisters without Detectable Resistance

    PubMed Central

    Moreira, Wilfried; Aziz, Dinah B.; Dick, Thomas

    2016-01-01

    Boromycin is a boron-containing polyether macrolide antibiotic isolated from Streptomyces antibioticus. It was shown to be active against Gram positive bacteria and to act as an ionophore for potassium ions. The antibiotic is ineffective against Gram negative bacteria where the outer membrane appears to block access of the molecule to the cytoplasmic membrane. Here we asked whether boromycin is active against Mycobacterium tuberculosis which, similar to Gram negative bacteria, possesses an outer membrane. The results show that boromycin is a potent inhibitor of mycobacterial growth (MIC50 = 80 nM) with strong bactericidal activity against growing and non-growing drug tolerant persister bacilli. Exposure to boromycin resulted in a rapid loss of membrane potential, reduction of the intracellular ATP level and leakage of cytoplasmic protein. Consistent with boromycin acting as a potassium ionophore, addition of KCl to the medium blocked its antimycobacterial activity. In contrast to the potent antimycobacterial activities of the polyether macrolide, its cytotoxicity and haemolytic activity were low (CC50 = 30 μM, HC50 = 40 μM) with a selectivity index of more than 300. Spontaneous resistant mutants could not be isolated suggesting a mutation frequency of less than 10-9/CFU. Taken together, the results suggests that targeting mycobacterial transmembrane ion gradients may be an attractive chemotherapeutic intervention level to kill otherwise drug tolerant persister bacilli, and to slow down the development of genetic antibiotic resistance. PMID:26941723

  3. Boromycin Kills Mycobacterial Persisters without Detectable Resistance.

    PubMed

    Moreira, Wilfried; Aziz, Dinah B; Dick, Thomas

    2016-01-01

    Boromycin is a boron-containing polyether macrolide antibiotic isolated from Streptomyces antibioticus. It was shown to be active against Gram positive bacteria and to act as an ionophore for potassium ions. The antibiotic is ineffective against Gram negative bacteria where the outer membrane appears to block access of the molecule to the cytoplasmic membrane. Here we asked whether boromycin is active against Mycobacterium tuberculosis which, similar to Gram negative bacteria, possesses an outer membrane. The results show that boromycin is a potent inhibitor of mycobacterial growth (MIC50 = 80 nM) with strong bactericidal activity against growing and non-growing drug tolerant persister bacilli. Exposure to boromycin resulted in a rapid loss of membrane potential, reduction of the intracellular ATP level and leakage of cytoplasmic protein. Consistent with boromycin acting as a potassium ionophore, addition of KCl to the medium blocked its antimycobacterial activity. In contrast to the potent antimycobacterial activities of the polyether macrolide, its cytotoxicity and haemolytic activity were low (CC50 = 30 μM, HC50 = 40 μM) with a selectivity index of more than 300. Spontaneous resistant mutants could not be isolated suggesting a mutation frequency of less than 10(-9)/CFU. Taken together, the results suggests that targeting mycobacterial transmembrane ion gradients may be an attractive chemotherapeutic intervention level to kill otherwise drug tolerant persister bacilli, and to slow down the development of genetic antibiotic resistance. PMID:26941723

  4. Metalloproteinase-mediated Shedding of Integrin β2 Promotes Macrophage Efflux from Inflammatory Sites*

    PubMed Central

    Gomez, Ivan G.; Tang, Jingjing; Wilson, Carole L.; Yan, Wei; Heinecke, Jay W.; Harlan, John M.; Raines, Elaine W.

    2012-01-01

    Macrophage exiting from inflammatory sites is critical to limit the local innate immune response. With tissue insult, resident tissue macrophages rapidly efflux to lymph nodes where they modulate the adaptive immune response, and inflammatory macrophages attracted to the site of injury then exit during the resolution phase. However, the mechanisms that regulate macrophage efflux are poorly understood. This study has investigated soluble forms of integrin β2 whose levels are elevated in experimental peritonitis at times when macrophages are exiting the peritoneum, suggesting that its proteolytic shedding may be involved in macrophage efflux. Both constitutive and inducible metalloproteinase-dependent shedding of integrin β2 from mouse macrophages are demonstrated. Soluble integrin β2 is primarily released as a heterodimeric complex with αM that retains its ability to bind its ligands intracellular adhesion molecule-1, fibrin, and collagen and thus may serve as a soluble antagonist. In a model of accelerated exiting, administration of a metalloproteinase inhibitor prevents macrophage efflux by 50% and impedes loss of macrophage integrin β2 from the cell surface. Exiting of peritoneal macrophages in mice lacking integrin β2 is accelerated, and antibody disruption of integrin β2-substrate interactions can reverse 50% of the metalloprotease inhibitor blockade of macrophage exiting. Thus, our study demonstrates the ability of metalloproteinase-mediated shedding of integrin β2 to promote macrophage efflux from inflammatory sites, and the release of soluble integrin heterodimers may also limit local inflammation. PMID:22170060

  5. Macrophage Apoptosis Triggered by IpaD from Shigella flexneri.

    PubMed

    Arizmendi, Olivia; Picking, William D; Picking, Wendy L

    2016-06-01

    Shigellosis, a potentially severe bacillary dysentery, is an infectious gastrointestinal disease caused by Shigella spp. Shigella invades the human colonic epithelium and avoids clearance by promoting apoptosis of resident immune cells in the gut. This process is dependent on the Shigella type III secretion system (T3SS), which injects effector proteins into target cells to alter their normal cellular functions. Invasion plasmid antigen D (IpaD) is a structural component that forms a complex at the tip of the T3SS apparatus needle. Recently, IpaD has also been shown to indirectly induce apoptosis in B lymphocytes. In this study, we explored the cytotoxicity profile during macrophage infection by Shigella and discovered that the pathogen induces macrophage cell death independent of caspase-1. Our results demonstrate that IpaD triggers apoptosis in macrophages through activation of host caspases accompanied by mitochondrial disruption. Additionally, we found that the IpaD N-terminal domain is necessary for macrophage killing and SipD, a structural homologue from Salmonella, was found to promote similar cytotoxicity. Together, these findings indicate that IpaD is a contributing factor to macrophage cell death during Shigella infection. PMID:27068089

  6. Activation of murine macrophages and lymphocytes by Ureaplasma diversum.

    PubMed Central

    Chelmonska-Soyta, A; Miller, R B; Ruhnke, L; Rosendal, S

    1994-01-01

    Ureaplasma diversum is a pathogen in the bovine reproductive tract. The objective of the research was to study interactions with macrophages and lymphocytes which might elucidate aspects of pathogenetic mechanisms of this organism. We studied the activation of murine macrophages of C3H/HeN (LPS-responder) and C3H/HeJ (LPS-low-responder) genotype for TNF-alpha, IL-6, IL-1 and nitric oxide production and blastogenic response of C3H/HeJ splenocytes after Ureaplasma diversum stimulation. Live and heat-killed U. diversum induced TNF-alpha, IL-6 and IL-1 in peritoneal macrophage cultures of both C3H/HeN and C3H/HeJ mice in a dose dependent manner. Interferon-gamma modulated the cytokine production, by increasing the production of TNF-alpha, IL-6 and nitric oxide, but IL-1 secretion was only enhanced in C3H/HeJ macrophages stimulated by live ureaplasmas. Supernatant of U. diversum sonicate was mitogenic for murine spleen lymphocytes. The blastogenic response was dose dependent, and stimulation with both U. diversum and Concanavalin A seemed to have an additive effect. These results suggest that U. diversum, similar to other mycoplasmas, activates murine macrophages and lymphoid cells. The studies should be repeated with bovine cells in order to elucidate pathogenetic aspects of inflammation in cattle caused by U. diversum. PMID:7889459

  7. Macrophage Apoptosis Triggered by IpaD from Shigella flexneri.

    PubMed

    Arizmendi, Olivia; Picking, William D; Picking, Wendy L

    2016-06-01

    Shigellosis, a potentially severe bacillary dysentery, is an infectious gastrointestinal disease caused by Shigella spp. Shigella invades the human colonic epithelium and avoids clearance by promoting apoptosis of resident immune cells in the gut. This process is dependent on the Shigella type III secretion system (T3SS), which injects effector proteins into target cells to alter their normal cellular functions. Invasion plasmid antigen D (IpaD) is a structural component that forms a complex at the tip of the T3SS apparatus needle. Recently, IpaD has also been shown to indirectly induce apoptosis in B lymphocytes. In this study, we explored the cytotoxicity profile during macrophage infection by Shigella and discovered that the pathogen induces macrophage cell death independent of caspase-1. Our results demonstrate that IpaD triggers apoptosis in macrophages through activation of host caspases accompanied by mitochondrial disruption. Additionally, we found that the IpaD N-terminal domain is necessary for macrophage killing and SipD, a structural homologue from Salmonella, was found to promote similar cytotoxicity. Together, these findings indicate that IpaD is a contributing factor to macrophage cell death during Shigella infection.

  8. Candida albicans Is Phagocytosed, Killed, and Processed for Antigen Presentation by Human Dendritic Cells

    PubMed Central

    Newman, Simon L.; Holly, Angela

    2001-01-01

    Candida albicans is a component of the normal flora of the alimentary tract and also is found on the mucocutaneous membranes of the healthy host. Candida is the leading cause of invasive fungal disease in premature infants, diabetics, and surgical patients, and of oropharyngeal disease in AIDS patients. As the induction of cell-mediated immunity to Candida is of critical importance in host defense, we sought to determine whether human dendritic cells (DC) could phagocytose and degrade Candida and subsequently present Candida antigens to T cells. Immature DC obtained by culture of human monocytes in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4 phagocytosed unopsonized Candida in a time-dependent manner, and phagocytosis was not enhanced by opsonization of Candida in serum. Like macrophages (Mφ), DC recognized Candida by the mannose-fucose receptor. Upon ingestion, DC killed Candida as efficiently as human Mφ, and fungicidal activity was not enhanced by the presence of fresh serum. Although phagocytosis of Candida by DC stimulated the production of superoxide anion, inhibitors of the respiratory burst (or NO production) did not inhibit killing of Candida, even when phagocytosis was blocked by preincubation of DC with cytochalasin D. Further, although apparently only modest phagolysosomal fusion occurred upon DC phagocytosis of Candida, killing of Candida under anaerobic conditions was almost equivalent to killing under aerobic conditions. Finally, DC stimulated Candida-specific lymphocyte proliferation in a concentration-dependent manner after phagocytosis of both viable and heat-killed Candida cells. These data suggest that, in vivo, such interactions between DC and C. albicans may facilitate the induction of cell-mediated immunity. PMID:11598054

  9. Bull heading to kill live gas wells

    SciTech Connect

    Oudeman, P.; Avest, D. ter; Grodal, E.O.; Asheim, H.A.; Meissner, R.J.H.

    1994-12-31

    To kill a live closed-in gas well by bull heading down the tubing, the selected pump rate should be high enough to ensure efficient displacement of the gas into the formation (i.e., to avoid the kill fluid bypassing the gas). On the other hand, the pressures that develop during bull heading at high rate must not exceed wellhead pressure rating, tubing or casing burst pressures or the formation breakdown gradient, since this will lead, at best, to a very inefficient kill job. Given these constraints, the optimum kill rate, requited hydraulic horsepower, density and type of kill fluids have to be selected. For this purpose a numerical simulator has been developed, which predicts the sequence of events during bull heading. Pressures and flow rates in the well during the kill job are calculated, taking to account slip between the gas and kill fluid, hydrostatic and friction pressure drop, wellbore gas compression and leak-off to the formation. Comparison with the results of a dedicated field test demonstrates that these parameters can be estimated accurately. Example calculations will be presented to show how the simulator can be used to identify an optimum kill scenario.

  10. Macrophages and Iron Metabolism.

    PubMed

    Soares, Miguel P; Hamza, Iqbal

    2016-03-15

    Iron is a transition metal that due to its inherent ability to exchange electrons with a variety of molecules is essential to support life. In mammals, iron exists mostly in the form of heme, enclosed within an organic protoporphyrin ring and functioning primarily as a prosthetic group in proteins. Paradoxically, free iron also has the potential to become cytotoxic when electron exchange with oxygen is unrestricted and catalyzes the production of reactive oxygen species. These biological properties demand that iron metabolism is tightly regulated such that iron is available for core biological functions while preventing its cytotoxic effects. Macrophages play a central role in establishing this delicate balance. Here, we review the impact of macrophages on heme-iron metabolism and, reciprocally, how heme-iron modulates macrophage function.

  11. [Macrophages in asthma].

    PubMed

    Medina Avalos, M A; Orea Solano, M

    1997-01-01

    Every time they exist more demonstrations of the paper than performs the line monocytes-macrophage in the patogenesis of the bronchial asthma. The mononuclear phagocytes cells, as the alveolar macrophages, also they can be activated during allergic methods. The monocytes macrophages are possible efficient inductors of the inflammation; this due to the fact that they can secrete inflammatory mediators, between those which are counted the pre-forming granules of peptides, metabolites of oxidation activation, activator of platelets activator and metabolites of the arachidonic acid. The identification of IL-1 in the liquidate of the bronchial ablution of sick asthmatic, as well as the identification of IL-1 in the I bronchioalveolar washing of places of allergens cutaneous prick, supports the activation concept mononuclear of phagocytic cells in allergic sufferings. PMID:9432275

  12. Momentum kill procedure can quickly control blowouts

    SciTech Connect

    Watson, W.D. ); Moore, P. )

    1993-08-30

    The momentum kill method can help in quickly regaining control of a blowing well, providing the blowing well rate and fluid properties can be estimated reasonably. The momentum of the kill fluid counteracts and overcomes the flowing momentum of formation fluids. In other words, sufficient mud density pumped at a sufficient rate is directed into the flow stream to force the escaping fluid column back into the well bore. Sufficient kill fluid hydrostatic pressure must be stacked'' in the hole so that the well remains dead after the operation. The momentum kill is not a panacea for all blowouts. An assessment must be made of the potential problems unique to this method, and certain requirements must be met if the technique is to be successful. The paper discusses some of the considerations for evaluating the use of the momentum kill method.

  13. The efficacy of pneumococcal capsular polysaccharide-specific antibodies to serotype 3 Streptococcus pneumoniae requires macrophages.

    PubMed

    Fabrizio, Kevin; Manix, Catherine; Tian, Haijun; van Rooijen, Nico; Pirofski, Liise-anne

    2010-11-01

    The efficacy of antibody immunity against Streptococcus pneumoniae stems from the ability of opsonic, serotype (ST)-specific antibodies to pneumococcal capsular polysaccharide (PPS) to facilitate killing of the homologous ST by host phagocytes. However, PPS-specific antibodies have been identified that are protective in mice, but do not promote opsonic killing in vitro, raising the question of how they mediate protection in vivo. To probe this question, we investigated the dependence of antibody efficacy against lethal systemic (intraperitoneal, i.p.) infection with Streptococcus pneumoniae serotype 3 (ST3) on macrophages and neutrophils for the following PPS3-specific monoclonal antibodies (MAbs) in survival experiments in mice using a non-opsonic human IgM (A7), a non-opsonic mouse IgG1 (1E2) and an opsonic mouse IgG1 (5F6). The survival of A7- and PPS3-specific and isotype control MAb-treated neutrophil-depleted and neutrophil-sufficient and macrophage-depleted and macrophage-sufficient mice were determined after i.p. challenge with ST3 strains 6303 and WU2. Neutrophils were dispensable for A7 and the mouse MAbs to mediate protection in this model, but macrophages were required for the efficacy of A7 and optimal mouse MAb-mediated protection. For A7-treated mice, macrophage-depleted mice had higher blood CFU, cytokines and peripheral neutrophil levels than macrophage-sufficient mice, and macrophage-sufficient mice had lower tissue bacterial burdens than control MAb-treated mice. These findings demonstrate that macrophages contribute to opsonic and non-opsonic PPS3-specific MAb-mediated protection against ST3 infection by enhancing bacterial clearance and suggest that neutrophils do not compensate for the absence of macrophages in the model used in this study.

  14. Perforin-2 restricts growth of Chlamydia trachomatis in macrophages.

    PubMed

    Fields, K A; McCormack, R; de Armas, L R; Podack, E R

    2013-08-01

    Chlamydia trachomatis is a Gram-negative obligate intracellular bacterium that preferentially infects epithelial cells. Professional phagocytes provide C. trachomatis only a limited ability to survive and are proficient killers of chlamydiae. We present evidence herein that identifies a novel host defense protein, perforin-2, that plays a significant role in the eradication of C. trachomatis during the infection of macrophages. Knockdown of perforin-2 in macrophages did not alter the invasion of host cells but did result in chlamydial growth that closely mirrored that detected in HeLa cells. C trachomatis L2, serovar B, and serovar D and C. muridarum were all equally susceptible to perforin-2-mediated killing. Interestingly, induction of perforin-2 expression in epithelial cells is blocked during productive chlamydial growth, thereby protecting chlamydiae from bactericidal attack. Ectopic expression of perforin-2 in HeLa cells, however, does result in killing. Overall, our data implicate a new innate resistance protein in the control of chlamydial infection and may help explain why the macrophage environment is hostile to chlamydial growth. PMID:23753625

  15. Klebsiella pneumoniae survives within macrophages by avoiding delivery to lysosomes.

    PubMed

    Cano, Victoria; March, Catalina; Insua, Jose Luis; Aguiló, Nacho; Llobet, Enrique; Moranta, David; Regueiro, Verónica; Brennan, Gerard P; Millán-Lou, Maria Isabel; Martín, Carlos; Garmendia, Junkal; Bengoechea, José A

    2015-11-01

    Klebsiella pneumoniae is an important cause of community-acquired and nosocomial pneumonia. Evidence indicates that Klebsiella might be able to persist intracellularly within a vacuolar compartment. This study was designed to investigate the interaction between Klebsiella and macrophages. Engulfment of K. pneumoniae was dependent on host cytoskeleton, cell plasma membrane lipid rafts and the activation of phosphoinositide 3-kinase (PI3K). Microscopy studies revealed that K. pneumoniae resides within a vacuolar compartment, the Klebsiella-containing vacuole (KCV), which traffics within vacuoles associated with the endocytic pathway. In contrast to UV-killed bacteria, the majority of live bacteria did not co-localize with markers of the lysosomal compartment. Our data suggest that K. pneumoniae triggers a programmed cell death in macrophages displaying features of apoptosis. Our efforts to identify the mechanism(s) whereby K. pneumoniae prevents the fusion of the lysosomes to the KCV uncovered the central role of the PI3K-Akt-Rab14 axis to control the phagosome maturation. Our data revealed that the capsule is dispensable for Klebsiella intracellular survival if bacteria were not opsonized. Furthermore, the environment found by Klebsiella within the KCV triggered the down-regulation of the expression of cps. Altogether, this study proves evidence that K. pneumoniae survives killing by macrophages by manipulating phagosome maturation that may contribute to Klebsiella pathogenesis.

  16. The role of TREM-2 in internalization and intracellular survival of Brucella abortus in murine macrophages.

    PubMed

    Wei, Pan; Lu, Qiang; Cui, Guimei; Guan, Zhenhong; Yang, Li; Sun, Changjiang; Sun, Wanchun; Peng, Qisheng

    2015-02-15

    Triggering receptor expressed on myeloid cells-2 (TREM-2) is a cell surface receptor primarily expressed on macrophages and dendritic cells. TREM-2 functions as a phagocytic receptor for bacteria as well as an inhibitor of Toll like receptors (TLR) induced inflammatory cytokines. However, the role of TREM-2 in Brucella intracellular growth remains unknown. To investigate whether TREM-2 is involved in Brucella intracellular survival, we chose bone marrow derived macrophages (BMDMs), in which TREM-2 is stably expressed, as cell model. Colony formation Units (CFUs) assay suggests that TREM-2 is involved in the internalization of Brucella abortus (B. abortus) by macrophages, while silencing of TREM-2 decreases intracellular survival of B. abortus. To further study the underlying mechanisms of TREM-2-mediated bacterial intracellular survival, we examined the activation of B. abortus-infected macrophages through determining the kinetics of activation of the three MAPKs, including ERK, JNK and p38, and measuring TNFα production in response to lipopolysaccharide (LPS) of Brucella (BrLPS) or B. abortus stimulation. Our data show that TREM-2 deficiency promotes activation of Brucella-infected macrophages. Moreover, our data also demonstrate that macrophage activation promotes killing of Brucella by enhancing nitric oxygen (NO), but not reactive oxygen species (ROS) production, macrophage apoptosis or cellular death. Taken together, these findings provide a novel interpretation of Brucella intracellular growth through inhibition of NO production produced by TREM-2-mediated activated macrophages.

  17. Role of Cathepsins in Mycobacterium tuberculosis Survival in Human Macrophages.

    PubMed

    Pires, David; Marques, Joana; Pombo, João Palma; Carmo, Nuno; Bettencourt, Paulo; Neyrolles, Olivier; Lugo-Villarino, Geanncarlo; Anes, Elsa

    2016-01-01

    Cathepsins are proteolytic enzymes that function in the endocytic pathway, especially in lysosomes, where they contribute directly to pathogen killing or indirectly, by their involvement in the antigen presentation pathways. Mycobacterium tuberculosis (MTB) is a facultative intracellular pathogen that survives inside the macrophage phagosomes by inhibiting their maturation to phagolysosomes and thus avoiding a low pH and protease-rich environment. We previously showed that mycobacterial inhibition of the proinflammatory transcription factor NF-κB results in impaired delivery of lysosomal enzymes to phagosomes and reduced pathogen killing. Here, we elucidate how MTB also controls cathepsins and their inhibitors, cystatins, at the level of gene expression and proteolytic activity. MTB induced a general down-regulation of cathepsin expression in infected cells, and inhibited IFNγ-mediated increase of cathepsin mRNA. We further show that a decrease in cathepsins B, S and L favours bacterial survival within human primary macrophages. A siRNA knockdown screen of a large set of cathepsins revealed that almost half of these enzymes have a role in pathogen killing, while only cathepsin F coincided with MTB resilience. Overall, we show that cathepsins are important for the control of MTB infection, and as a response, it manipulates their expression and activity to favour its intracellular survival. PMID:27572605

  18. Role of Cathepsins in Mycobacterium tuberculosis Survival in Human Macrophages

    PubMed Central

    Pires, David; Marques, Joana; Pombo, João Palma; Carmo, Nuno; Bettencourt, Paulo; Neyrolles, Olivier; Lugo-Villarino, Geanncarlo; Anes, Elsa

    2016-01-01

    Cathepsins are proteolytic enzymes that function in the endocytic pathway, especially in lysosomes, where they contribute directly to pathogen killing or indirectly, by their involvement in the antigen presentation pathways. Mycobacterium tuberculosis (MTB) is a facultative intracellular pathogen that survives inside the macrophage phagosomes by inhibiting their maturation to phagolysosomes and thus avoiding a low pH and protease-rich environment. We previously showed that mycobacterial inhibition of the proinflammatory transcription factor NF-κB results in impaired delivery of lysosomal enzymes to phagosomes and reduced pathogen killing. Here, we elucidate how MTB also controls cathepsins and their inhibitors, cystatins, at the level of gene expression and proteolytic activity. MTB induced a general down-regulation of cathepsin expression in infected cells, and inhibited IFNγ-mediated increase of cathepsin mRNA. We further show that a decrease in cathepsins B, S and L favours bacterial survival within human primary macrophages. A siRNA knockdown screen of a large set of cathepsins revealed that almost half of these enzymes have a role in pathogen killing, while only cathepsin F coincided with MTB resilience. Overall, we show that cathepsins are important for the control of MTB infection, and as a response, it manipulates their expression and activity to favour its intracellular survival. PMID:27572605

  19. Household Air Pollution Causes Dose-Dependent Inflammation and Altered Phagocytosis in Human Macrophages

    PubMed Central

    Fullerton, Duncan G.; Scriven, James; Aljurayyan, Abdullah N.; Mzinza, David; Barrett, Steve; Wright, Adam K. A.; Wootton, Daniel G.; Glennie, Sarah J.; Baple, Katy; Knott, Amy; Mortimer, Kevin; Russell, David G.; Heyderman, Robert S.; Gordon, Stephen B.

    2015-01-01

    Three billion people are exposed to household air pollution from biomass fuel use. Exposure is associated with higher incidence of pneumonia, and possibly tuberculosis. Understanding mechanisms underlying these defects would improve preventive strategies. We used human alveolar macrophages obtained from healthy Malawian adults exposed naturally to household air pollution and compared them with human monocyte-derived macrophages exposed in vitro to respirable-sized particulates. Cellular inflammatory response was assessed by IL-6 and IL-8 production in response to particulate challenge; phagosomal function was tested by uptake and oxidation of fluorescence-labeled beads; ingestion and killing of Streptococcus pneumoniae and Mycobacterium tuberculosis were measured by microscopy and quantitative culture. Particulate ingestion was quantified by digital image analysis. We were able to reproduce the carbon loading of naturally exposed alveolar macrophages by in vitro exposure of monocyte-derived macrophages. Fine carbon black induced IL-8 release from monocyte-derived and alveolar macrophages (P < 0.05) with similar magnitude responses (log10 increases of 0.93 [SEM = 0.2] versus 0.74 [SEM = 0.19], respectively). Phagocytosis of pneumococci and mycobacteria was impaired with higher particulate loading. High particulate loading corresponded with a lower oxidative burst capacity (P = 0.0015). There was no overall effect on killing of M. tuberculosis. Alveolar macrophage function is altered by particulate loading. Our macrophage model is comparable morphologically to the in vivo uptake of particulates. Wood smoke–exposed cells demonstrate reduced phagocytosis, but unaffected mycobacterial killing, suggesting defects related to chronic wood smoke inhalation limited to specific innate immune functions. PMID:25254931

  20. Household air pollution causes dose-dependent inflammation and altered phagocytosis in human macrophages.

    PubMed

    Rylance, Jamie; Fullerton, Duncan G; Scriven, James; Aljurayyan, Abdullah N; Mzinza, David; Barrett, Steve; Wright, Adam K A; Wootton, Daniel G; Glennie, Sarah J; Baple, Katy; Knott, Amy; Mortimer, Kevin; Russell, David G; Heyderman, Robert S; Gordon, Stephen B

    2015-05-01

    Three billion people are exposed to household air pollution from biomass fuel use. Exposure is associated with higher incidence of pneumonia, and possibly tuberculosis. Understanding mechanisms underlying these defects would improve preventive strategies. We used human alveolar macrophages obtained from healthy Malawian adults exposed naturally to household air pollution and compared them with human monocyte-derived macrophages exposed in vitro to respirable-sized particulates. Cellular inflammatory response was assessed by IL-6 and IL-8 production in response to particulate challenge; phagosomal function was tested by uptake and oxidation of fluorescence-labeled beads; ingestion and killing of Streptococcus pneumoniae and Mycobacterium tuberculosis were measured by microscopy and quantitative culture. Particulate ingestion was quantified by digital image analysis. We were able to reproduce the carbon loading of naturally exposed alveolar macrophages by in vitro exposure of monocyte-derived macrophages. Fine carbon black induced IL-8 release from monocyte-derived and alveolar macrophages (P < 0.05) with similar magnitude responses (log10 increases of 0.93 [SEM = 0.2] versus 0.74 [SEM = 0.19], respectively). Phagocytosis of pneumococci and mycobacteria was impaired with higher particulate loading. High particulate loading corresponded with a lower oxidative burst capacity (P = 0.0015). There was no overall effect on killing of M. tuberculosis. Alveolar macrophage function is altered by particulate loading. Our macrophage model is comparable morphologically to the in vivo uptake of particulates. Wood smoke-exposed cells demonstrate reduced phagocytosis, but unaffected mycobacterial killing, suggesting defects related to chronic wood smoke inhalation limited to specific innate immune functions.

  1. Magnetometric evaluation for the effect of chrysotile on alveolar macrophages.

    PubMed

    Keira, T; Okada, M; Katagiri, H; Aizawa, Y; Okayasu, I; Kotani, M

    1998-10-01

    Alveolar macrophages are thought to play an important role in fibrogenesis due to asbestos exposure. In this experiment, we evaluated the effect mainly by unique magnetometry and also by conventional methods such as lactate dehydrogenase (LDH) activity measurement and morphological observations. Alveolar macrophages obtained from Syrian golden hamsters by bronchoalveolar lavages were exposed 18 hours in vitro to Fe3O4 as an indicator for magnetometry and chrysotile for experiments. A rapid decrease of the remanent magnetic field, so called "relaxation", was observed after the cessation of an external magnetic field in macrophages phagocytizing Fe3O4 alone, while relaxation was delayed in those concurrently exposed to chrysotile. Since relaxation is thought due to the cytoskeleton-driven random rotation of phagosomes containing iron oxide particles, chrysotile is considered to interfere with the cytoskeletal function of macrophages. Release of LDH from chrysotile-exposed macrophages into the medium was recognized, but it was not significantly higher than the controls. Apoptosis was negligible in macrophages exposed to chrysotile by the DNA ladder detection, the terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling method and morphological observations. Electron microscopical examinations revealed early necrotic changes in macrophages exposed to chrysotile. These findings indicate that cell magnetometry detects impaired cytoskeletal function due to in vitro exposure to chrysotile. PMID:10223613

  2. Candida albicans induces selective development of macrophages and monocyte derived dendritic cells by a TLR2 dependent signalling.

    PubMed

    Yáñez, Alberto; Megías, Javier; O'Connor, José-Enrique; Gozalbo, Daniel; Gil, M Luisa

    2011-01-01

    As TLRs are expressed by haematopoietic stem and progenitor cells (HSPCs), these receptors may play a role in haematopoiesis in response to pathogens during infection. We have previously demonstrated that in in vitro defined conditions inactivated yeasts and hyphae of Candida albicans induce HSPCs proliferation and differentiation towards the myeloid lineage by a TLR2/MyD88 dependent pathway. In this work, we showed that C. albicans invasive infection with a low virulence strain results in a rapid expansion of HSPCs (identified as LKS cells: Lin(-) c-Kit(+) Sca-1(+) IL-7Rα(-)), that reach the maximum at day 3 post-infection. This in vivo expansion of LKS cells in TLR2(-/-) mice was delayed until day 7 post- infection. Candidiasis was, as expected, accompanied by an increase in granulopoiesis and decreased lymphopoiesis in the bone marrow. These changes were more pronounced in TLR2(-/-) mice correlating with their higher fungal burden. Accordingly, emigration of Ly6C(high) monocytes and neutrophils to spleen was increased in TLR2(-/-) mice, although the increase in macrophages and inflammatory macrophages was completely dependent on TLR2. Similarly, we detected for the first time, in the spleen of C. albicans infected control mice, a newly generated population of dendritic cells that have the phenotype of monocyte derived dendritic cells (moDCs) that were not generated in TLR2(-/-) infected mice. In addition, C. albicans signalling through TLR2/MyD88 and Dectin-1 promotes in vitro the differentiation of Lin(-) cells towards moDCs that secrete TNF-α and are able to kill the microorganism. Therefore, our results indicate that during infection C. albicans can directly stimulate progenitor cells through TLR2 and Dectin-1 to generate newly formed inflammatory macrophages and moDCs that may fulfill an essential role in defense mechanisms against the pathogen.

  3. Candida albicans Induces Selective Development of Macrophages and Monocyte Derived Dendritic Cells by a TLR2 Dependent Signalling

    PubMed Central

    Yáñez, Alberto; Megías, Javier; O'Connor, José-Enrique; Gozalbo, Daniel; Gil, M. Luisa

    2011-01-01

    As TLRs are expressed by haematopoietic stem and progenitor cells (HSPCs), these receptors may play a role in haematopoiesis in response to pathogens during infection. We have previously demonstrated that in in vitro defined conditions inactivated yeasts and hyphae of Candida albicans induce HSPCs proliferation and differentiation towards the myeloid lineage by a TLR2/MyD88 dependent pathway. In this work, we showed that C. albicans invasive infection with a low virulence strain results in a rapid expansion of HSPCs (identified as LKS cells: Lin− c-Kit+ Sca-1+ IL-7Rα−), that reach the maximum at day 3 post-infection. This in vivo expansion of LKS cells in TLR2−/− mice was delayed until day 7 post- infection. Candidiasis was, as expected, accompanied by an increase in granulopoiesis and decreased lymphopoiesis in the bone marrow. These changes were more pronounced in TLR2−/− mice correlating with their higher fungal burden. Accordingly, emigration of Ly6Chigh monocytes and neutrophils to spleen was increased in TLR2−/− mice, although the increase in macrophages and inflammatory macrophages was completely dependent on TLR2. Similarly, we detected for the first time, in the spleen of C. albicans infected control mice, a newly generated population of dendritic cells that have the phenotype of monocyte derived dendritic cells (moDCs) that were not generated in TLR2−/− infected mice. In addition, C. albicans signalling through TLR2/MyD88 and Dectin-1 promotes in vitro the differentiation of Lin− cells towards moDCs that secrete TNF-α and are able to kill the microorganism. Therefore, our results indicate that during infection C. albicans can directly stimulate progenitor cells through TLR2 and Dectin-1 to generate newly formed inflammatory macrophages and moDCs that may fulfill an essential role in defense mechanisms against the pathogen. PMID:21935459

  4. Spacetime encodings. III. Second order Killing tensors

    SciTech Connect

    Brink, Jeandrew

    2010-01-15

    This paper explores the Petrov type D, stationary axisymmetric vacuum (SAV) spacetimes that were found by Carter to have separable Hamilton-Jacobi equations, and thus admit a second-order Killing tensor. The derivation of the spacetimes presented in this paper borrows from ideas about dynamical systems, and illustrates concepts that can be generalized to higher-order Killing tensors. The relationship between the components of the Killing equations and metric functions are given explicitly. The origin of the four separable coordinate systems found by Carter is explained and classified in terms of the analytic structure associated with the Killing equations. A geometric picture of what the orbital invariants may represent is built. Requiring that a SAV spacetime admits a second-order Killing tensor is very restrictive, selecting very few candidates from the group of all possible SAV spacetimes. This restriction arises due to the fact that the consistency conditions associated with the Killing equations require that the field variables obey a second-order differential equation, as opposed to a fourth-order differential equation that imposes the weaker condition that the spacetime be SAV. This paper introduces ideas that could lead to the explicit computation of more general orbital invariants in the form of higher-order Killing tensors.

  5. [Killing of cattle via electrical stunning].

    PubMed

    Maurer, B; Forster, S

    2007-04-01

    For disease control in the case of epidemics killing of cattle via electrical stunning is a method of choice. The official veterinarian is responsible for monitoring the adhesion to animal welfare principles during electrical stunning and killing. This requires specialised knowledge and experience as the symptoms of effective stunning are quite variable in cattle. Signs of effective and ineffective stunning are described below. In addition to suitable technical equipment, restraint of the animals and correct use of the equipment, neurophysiological processes have to be considered. Calm handling of the animals avoiding stress is a prerequisite for ensuring animal welfare and minimising pain especially when killing cattle using electrical methods.

  6. Oxidative stress, polarization of macrophages and tumour angiogenesis: Efficacy of caffeic acid.

    PubMed

    Oršolić, Nada; Kunštić, Martina; Kukolj, Marina; Gračan, Romana; Nemrava, Johann

    2016-08-25

    Macrophage polarization is a process when macrophage expresses different functional programs in response to microenvironmental signals and two extreme forms exist; M1 and M2 macrophages. M1 macrophages are highly microbicidal and anticancer with enhanced ability to kill and phagocytose pathogens, upregulate pro-inflammatory cytokines and reactive molecular species, and present antigens; M2 macrophages and the related tumour associated macrophages (TAMs) regulate tissue remodelling and promote tissue repair and angiogenesis and can amplification of metabolic pathways that can suppress adaptive immune responses. It is demonstrated that ROS production, critical for the activation and functions of M1 macrophages, is necessary for the differentiation of M2 macrophages and TAMs, and that antioxidant therapy blocks TAMs differentiation and tumorigenesis in mouse models of cancer. In order to study how caffeic acid (CA), a natural antioxidant, affects macrophage function, polarization, angiogenesis and tumour growth we injected mice with Ehrlich ascites tumour (EAT) cells and treated them for 10 days with CA in a dose of 40 and/or 80 mg kg(-1.) Macrophage polarization was further characterized by quantifying secreted pro- and anti-inflammatory cytokines, nitric oxide and arginase 1 activity. CA may increase the cytotoxic actions of M1 macrophages and inhibit tumour growth; inhibitory activity on TAMs may be mediated through its antioxidative activity. Taken together, we conclude that the antitumour activity of CA was the result of the synergistic activities of different mechanisms by which CA acts on proliferation, angiogenesis, immunomodulation and survival. The continuous administration of CA efficiently blocked the occurrence of TAMs and markedly suppressed tumorigenesis in mouse cancer models. Targeting TAMs by antioxidants can be a potentially effective method for cancer treatment.

  7. Polyanhydride Nanoparticle Delivery Platform Dramatically Enhances Killing of Filarial Worms

    PubMed Central

    Binnebose, Andrea M.; Haughney, Shannon L.; Martin, Richard; Imerman, Paula M.; Narasimhan, Balaji; Bellaire, Bryan H.

    2015-01-01

    Filarial diseases represent a significant social and economic burden to over 120 million people worldwide and are caused by endoparasites that require the presence of symbiotic bacteria of the genus Wolbachia for fertility and viability of the host parasite. Targeting Wolbachia for elimination is a therapeutic approach that shows promise in the treatment of onchocerciasis and lymphatic filariasis. Here we demonstrate the use of a biodegradable polyanhydride nanoparticle-based platform for the co-delivery of the antibiotic doxycycline with the antiparasitic drug, ivermectin, to reduce microfilarial burden and rapidly kill adult worms. When doxycycline and ivermectin were co-delivered within polyanhydride nanoparticles, effective killing of adult female Brugia malayi filarial worms was achieved with approximately 4,000-fold reduction in the amount of drug used. Additionally the time to death of the macrofilaria was also significantly reduced (five-fold) when the anti-filarial drug cocktail was delivered within polyanhydride nanoparticles. We hypothesize that the mechanism behind this dramatically enhanced killing of the macrofilaria is the ability of the polyanhydride nanoparticles to behave as a Trojan horse and penetrate the cuticle, bypassing excretory pumps of B. malayi, and effectively deliver drug directly to both the worm and Wolbachia at high enough microenvironmental concentrations to cause death. These provocative findings may have significant consequences for the reduction in the amount of drug and the length of treatment required for filarial infections in terms of patient compliance and reduced cost of treatment. PMID:26496201

  8. Transcriptional Regulation and Macrophage Differentiation.

    PubMed

    Hume, David A; Summers, Kim M; Rehli, Michael

    2016-06-01

    Monocytes and macrophages are professional phagocytes that occupy specific niches in every tissue of the body. Their survival, proliferation, and differentiation are controlled by signals from the macrophage colony-stimulating factor receptor (CSF-1R) and its two ligands, CSF-1 and interleukin-34. In this review, we address the developmental and transcriptional relationships between hematopoietic progenitor cells, blood monocytes, and tissue macrophages as well as the distinctions from dendritic cells. A huge repertoire of receptors allows monocytes, tissue-resident macrophages, or pathology-associated macrophages to adapt to specific microenvironments. These processes create a broad spectrum of macrophages with different functions and individual effector capacities. The production of large transcriptomic data sets in mouse, human, and other species provides new insights into the mechanisms that underlie macrophage functional plasticity. PMID:27337479

  9. Antimicrobial Mechanisms of Macrophages and the Immune Evasion Strategies of Staphylococcus aureus

    PubMed Central

    Flannagan, Ronald S.; Heit, Bryan; Heinrichs, David E.

    2015-01-01

    Habitually professional phagocytes, including macrophages, eradicate microbial invaders from the human body without overt signs of infection. Despite this, there exist select bacteria that are professional pathogens, causing significant morbidity and mortality across the globe and Staphylococcus aureus is no exception. S. aureus is a highly successful pathogen that can infect virtually every tissue that comprises the human body causing a broad spectrum of diseases. The profound pathogenic capacity of S. aureus can be attributed, in part, to its ability to elaborate a profusion of bacterial effectors that circumvent host immunity. Macrophages are important professional phagocytes that contribute to both the innate and adaptive immune response, however from in vitro and in vivo studies, it is evident that they fail to eradicate S. aureus. This review provides an overview of the antimicrobial mechanisms employed by macrophages to combat bacteria and describes the immune evasion strategies and some representative effectors that enable S. aureus to evade macrophage-mediated killing. PMID:26633519

  10. Pulmonary Responses to Stachybotrys chartarum and Its Toxins: Mouse Strain Affects Clearance and Macrophage Cytotoxicity

    PubMed Central

    Lichtenstein, Jamie H. Rosenblum; Molina, Ramon M.; Donaghey, Thomas C.; Amuzie, Chidozie J.; Pestka, James J.; Coull, Brent A.; Brain, Joseph D.

    2010-01-01

    We investigated differences in the pulmonary and systemic clearance of Stachybotrys chartarum spores in two strains of mice, BALB/c and C57BL/6J. To evaluate clearance, mice were intratracheally instilled with a suspension of radiolabeled S. chartarum spores or with unlabeled spores. The lungs of C57BL/6J mice showed more rapid spore clearance than the lungs of BALB/c mice, which correlated with increased levels of spore-associated radioactivity in the GI tracts of C57BL/6J as compared with BALB/c mice. To identify mechanisms responsible for mouse strain differences in spore clearance and previously described lung inflammatory responses, we exposed alveolar macrophages (AMs) lavaged from BALB/c and C57BL/6J mice to S. chartarum spores, S. chartarum spore toxin (SST), and satratoxin G (SG) in vitro. The S. chartarum spores were found to be highly toxic with most cells from either mouse strain being killed within 24 h when exposed to a spore:cell ratio of 1:75. The spores were more lethal to AMs from C57BL/6J than those from BALB/c mice. In mice, the SST elicited many of the same inflammatory responses as the spores in vivo, including AM recruitment, pulmonary hemorrhage, and cytokine production. Our data suggest that differences in pulmonary spore clearance may contribute to the differences in pulmonary responses to S. chartarum between BALB/c and C57BL/6J mice. Enhanced AM survival and subsequent macrophage-mediated inflammation may also contribute to the higher susceptibility of BALB/c mice to S. chartarum pulmonary effects. Analogous genetic differences among humans may contribute to reported variable sensitivity to S. chartarum. PMID:20385656

  11. Quantum integrability of quadratic Killing tensors

    SciTech Connect

    Duval, C.; Valent, G.

    2005-05-01

    Quantum integrability of classical integrable systems given by quadratic Killing tensors on curved configuration spaces is investigated. It is proven that, using a 'minimal' quantization scheme, quantum integrability is ensured for a large class of classic examples.

  12. Killing Initial Data on spacelike conformal boundaries

    NASA Astrophysics Data System (ADS)

    Paetz, Tim-Torben

    2016-08-01

    We analyze Killing Initial Data on Cauchy surfaces in conformally rescaled vacuum space-times satisfying Friedrich's conformal field equations. As an application, we derive the KID equations on a spacelike ℐ-.

  13. Catch me if you can: phagocytosis and killing avoidance by Cryptococcus neoformans.

    PubMed

    García-Rodas, Rocío; Zaragoza, Oscar

    2012-03-01

    After inhalation of infectious particles, Cryptococcus neoformans resides in the alveolar spaces, where it can survive and replicate in the extracellular environment. This yeast has developed different mechanisms to avoid internalization by phagocytic cells, the main one being a polysaccharide capsule around the cell body, which inhibits the uptake of the yeast by macrophages. In addition, capsule-independent mechanisms have also been described, such as the production of antiphagocytic proteins. Despite these mechanisms, phagocytosis can occur in the presence of opsonins, and once C. neoformans is internalized, multiple outcomes are possible, including pathogen killing or intracellular replication and escape from macrophages. For this reason, C. neoformans is considered a facultative intracellular pathogen. As alveolar macrophages are the first component of the host immune system to confront C. neoformans, the outcome of this interaction could determine the degree of infection, producing either a severe disseminated disease or a latency state. In this review, we will tackle the complexity of the interaction between C. neoformans and macrophages, including the phagocytic avoidance mechanisms and all the possible outcomes that have been described for this interaction. Finally, we will discuss the consequences of the different outcomes for the type of infection produced in the host.

  14. Mechanisms for macrophage-mediated HIV-1 induction.

    PubMed

    Devadas, Krishnakumar; Hardegen, Neil J; Wahl, Larry M; Hewlett, Indira K; Clouse, Kathleen A; Yamada, Kenneth M; Dhawan, Subhash

    2004-12-01

    Viral latency is a long-term pathogenic condition in patients infected with HIV-1. Low but sustained virus replication in chronically infected cells can be activated by stimulation with proinflammatory cytokines such as TNF-alpha, IL-1 beta, or other host factors. However, the precise mechanism by which cellular activation induces latently infected cells to produce virions has remained unclear. In the present report, we present evidence that activation of HIV-1 replication in latently infected U1 or ACH2 cells by human macrophages is mediated by a rapid nuclear localization of NF-kappaB p50/p65 dimer with concomitant increased expression of proinflammatory cytokines. Multiplexed RT-PCR amplification of mRNA isolated from cocultures of macrophages and U1 and ACH2 cells showed significant induction of IL-1beta, IL-6, IL-8, TNF-alpha, and TGF-beta expression within 3 h of coincubation. Fixation of macrophages, U-1, or ACH2 cells with paraformaldehyde before coculture completely abrogated the induction of NF-kappaB subunits and HIV-1 replication, suggesting that cooperative interaction between the two cell types is an essential process for cellular activation. Pretreatment of macrophage-U1 or macrophage-ACH2 cocultures with neutralizing anti-TNF-alpha Ab down-regulated the replication of HIV-1. In addition, pretreatment of macrophage-U1 or macrophage-ACH2 cocultures with the NF-kappaB inhibitor (E)3-[(4-methylphenyl)sulfonyl]-2-propenenitrile (BAY 11-7082) prevented the induction of cytokine expression, indicating a pivotal role of NF-kappaB-mediated signaling in the reactivation of HIV-1 in latently infected cells by macrophages. These results provide a mechanism by which macrophages induce HIV-1 replication in latently infected cells.

  15. Killing vector fields and harmonic superfield theories

    SciTech Connect

    Groeger, Josua

    2014-09-15

    The harmonic action functional allows a natural generalisation to semi-Riemannian supergeometry, also referred to as harmonic, which resembles the supersymmetric sigma models studied in high energy physics. We show that Killing vector fields are infinitesimal supersymmetries of this harmonic action and prove three different Noether theorems in this context. En passant, we provide a homogeneous treatment of five characterisations of Killing vector fields on semi-Riemannian supermanifolds, thus filling a gap in the literature.

  16. Effects of Macrophage Depletion on Sleep in Mice.

    PubMed

    Ames, Conner; Boland, Erin; Szentirmai, Éva

    2016-01-01

    The reciprocal interaction between the immune system and sleep regulation has been widely acknowledged but the cellular mechanisms that underpin this interaction are not completely understood. In the present study, we investigated the role of macrophages in sleep loss- and cold exposure-induced sleep and body temperature responses. Macrophage apoptosis was induced in mice by systemic injection of clodronate-containing liposomes (CCL). We report that CCL treatment induced an immediate and transient increase in non-rapid-eye movement sleep (NREMS) and fever accompanied by decrease in rapid-eye movement sleep, motor activity and NREMS delta power. Chronically macrophage-depleted mice had attenuated NREMS rebound after sleep deprivation compared to normal mice. Cold-induced increase in wakefulness and decrease in NREMS, rapid-eye movement sleep and body temperature were significantly enhanced in macrophage-depleted mice indicating increased cold sensitivity. These findings provide further evidence for the reciprocal interaction among the immune system, sleep and metabolism, and identify macrophages as one of the key cellular elements in this interplay. PMID:27442442

  17. Effects of Macrophage Depletion on Sleep in Mice

    PubMed Central

    Ames, Conner; Boland, Erin; Szentirmai, Éva

    2016-01-01

    The reciprocal interaction between the immune system and sleep regulation has been widely acknowledged but the cellular mechanisms that underpin this interaction are not completely understood. In the present study, we investigated the role of macrophages in sleep loss- and cold exposure-induced sleep and body temperature responses. Macrophage apoptosis was induced in mice by systemic injection of clodronate-containing liposomes (CCL). We report that CCL treatment induced an immediate and transient increase in non-rapid-eye movement sleep (NREMS) and fever accompanied by decrease in rapid-eye movement sleep, motor activity and NREMS delta power. Chronically macrophage-depleted mice had attenuated NREMS rebound after sleep deprivation compared to normal mice. Cold-induced increase in wakefulness and decrease in NREMS, rapid-eye movement sleep and body temperature were significantly enhanced in macrophage-depleted mice indicating increased cold sensitivity. These findings provide further evidence for the reciprocal interaction among the immune system, sleep and metabolism, and identify macrophages as one of the key cellular elements in this interplay. PMID:27442442

  18. Alteration of leukotriene release by macrophages ingesting Toxoplasma gondii.

    PubMed Central

    Locksley, R M; Fankhauser, J; Henderson, W R

    1985-01-01

    Mouse resident peritoneal macrophages incubated with ionophore A23187 or opsonized zymosan released leukotrienes (LT) B4 and C4 (LTB4 and LTC4) and LTC4 and LTD4, respectively. In contrast, incubation with Toxoplasma gondii, an obligate intracellular protozoan, led to the formation of 11-, 12-, and 15-hydroxyicosatetraenoic acids (HETEs), together with an unidentified compound, designated compound X. Each of these compounds incorporated [3H]arachidonic acid from the macrophage during phagocytosis of T. gondii. Compound X migrated immediately prior to 15-HETE by reverse-phase HPLC and was distinct from authentic monoHETE, monohydroperoxyicosatetraenoic acid (mono-HPETE), and dihydroxyicosatetraenoic acid (diHETE) standards. The generation of compound X by macrophages correlated with the extent of phagocytosis of T. gondii and with intracellular survival of the organisms. Prior antibody-coating of T. gondii or activation of macrophages, either of which inhibited survival and replication of ingested organisms, was associated with production of LTD4 but not compound X. Killed organisms also stimulated LTD4 release only. Although T. gondii concentrated arachidonic acid, they did not metabolize the compound to identifiable lipoxygenase products. Preincubation of macrophages with the relative lipoxygenase inhibitors nordihydroguaiaretic acid or 5,8,11,14-icosatetraynoic acid inhibited the formation of compound X. The absence of leukotriene production by macrophages ingesting T. gondii may explain the relative lack of a neutrophil inflammatory response in diseases due to obligate intracellular organisms. Alternatively, compound X may have functional activities that might mediate some of the host responses to cellular parasitism. PMID:2995993

  19. Killing of Actinobacillus actinomycetemcomitans by human lactoferrin.

    PubMed Central

    Kalmar, J R; Arnold, R R

    1988-01-01

    Actinobacillus actinomycetemcomitans is a fastidious, facultative gram-negative rod associated with endocarditis, certain forms of periodontal disease, and other focal infections. Human neutrophils have demonstrated bactericidal activity against A. actinomycetemcomitans, and much of the oxygen-dependent killing has been attributed to the myeloperoxidase-H2O2-halide system. However, the contribution of other neutrophil components to killing activity is obscure. Lactoferrin, an iron-binding glycoprotein, is a major constituent of neutrophil-specific granules and is also found in mucosal secretions. In this report, we show that human lactoferrin is bactericidal for A. actinomycetemcomitans. Killing activity required an unsaturated (iron- and anion-free) molecule that produced a 2-log decrease in viability within 120 min at 37 degrees C at a concentration of 1.9 microM. Besides exhibiting concentration dependence, killing kinetics were affected by minor variations in temperature and pH. Magnesium, a divalent cation thought to stabilize lipopolysaccharide interactions on the surface of gram-negative organisms, enhanced lactoferrin killing of A. actinomycetemcomitans, while other cations, such as potassium and calcium, had no effect. Our data suggest that lactoferrin contributes to killing of A. actinomycetemcomitans by human neutrophils and that it may also play a significant role in innate secretory defense against this potential periodontopathogen. PMID:3417349

  20. Hazardous materials in Fresh Kills landfill

    SciTech Connect

    Hirschhorn, J.S.

    1997-12-31

    No environmental monitoring and corrective action programs can pinpoint multiple locations of hazardous materials the total amount of them in a large landfill. Yet the consequences of hazardous materials in MSW landfills are considerable, in terms of public health concerns, environmental damage, and cleanup costs. In this paper a rough estimation is made of how much hazardous material may have been disposed in Fresh Kills landfill in Staten Island, New York. The logic and methods could be used for other MSW landfills. Fresh Kills has frequently been described as the world`s largest MSW landfill. While records of hazardous waste disposal at Fresh Kills over nearly 50 years of operation certainly do not exist, no reasonable person would argue with the conclusion that large quantities of hazardous waste surely have been disposed at Fresh Kills, both legally and illegally. This study found that at least 2 million tons of hazardous wastes and substances have been disposed at Fresh Kills since 1948. Major sources are: household hazardous waste, commercial RCRA hazardous waste, incinerator ash, and commercial non-RCRA hazardous waste, governmental RCRA hazardous waste. Illegal disposal of hazardous waste surely has contributed even more. This is a sufficient amount to cause serious environmental contamination and releases, especially from such a landfill without an engineered liner system, for example. This figure is roughly 1% of the total amount of waste disposed in Fresh Kills since 1948, probably at least 200 million tons.

  1. Glucocorticoid-induced impairment of macrophage antimicrobial activity: mechanisms and dependence on the state of activation.

    PubMed

    Schaffner, A; Schaffner, T

    1987-01-01

    Experimental observations indicate that tissue macrophages deployed in great numbers at critical anatomic sites such as the liver, spleen, and lung are major targets for glucocorticoids compromising natural resistance of the host. Therapeutic concentrations of glucocorticoids appear to prevent destruction of microorganisms ingested by macrophages without interfering with phagocytosis, phagolysosomal fusion, and/or secretion of reactive oxygen intermediates. These findings indicate that at the cellular level the glucocorticoid target should be sought for in the nonoxidative armature of the phagocyte and that nonoxidative killing systems of resident tissue macrophages play an important role in natural resistance to opportunistic pathogens. Glucocorticoids do not prevent lymphokine-induced activation of oxidative killing systems. Thus, lymphokines such as interferon-gamma can restore the microbicidal activity of macrophages functionally impaired by glucocorticoids. Counterbalance of the suppressive effect of glucocorticoids by lymphokines might only be possible, however, for pathogens susceptible to oxidative killing and not for microorganisms that are more resistant to reactive oxygen intermediates such as Aspergillus spores and Nocardia, opportunists that appear to be particularly associated with hypercortisolism.

  2. Fosfomycin enhances phagocyte-mediated killing of Staphylococcus aureus by extracellular traps and reactive oxygen species

    PubMed Central

    Shen, Fengge; Tang, Xudong; Cheng, Wei; Wang, Yang; Wang, Chao; Shi, Xiaochen; An, Yanan; Zhang, Qiaoli; Liu, Mingyuan; Liu, Bo; Yu, Lu

    2016-01-01

    The successful treatment of bacterial infections is the achievement of a synergy between the host’s immune defences and antibiotics. Here, we examined whether fosfomycin (FOM) could improve the bactericidal effect of phagocytes, and investigated the potential mechanisms. FOM enhanced the phagocytosis and extra- or intracellular killing of S. aureus by phagocytes. And FOM enhanced the extracellular killing of S. aureus in macrophage (MФ) and in neutrophils mediated by extracellular traps (ETs). ET production was related to NADPH oxidase-dependent reactive oxygen species (ROS). Additionally, FOM increased the intracellular killing of S. aureus in phagocytes, which was mediated by ROS through the oxidative burst process. Our results also showed that FOM alone induced S. aureus producing hydroxyl radicals in order to kill the bacterial cells in vitro. In a mouse peritonitis model, FOM treatment increased the bactericidal extra- and intracellular activity in vivo, and FOM strengthened ROS and ET production from peritoneal lavage fluid ex vivo. An IVIS imaging system assay further verified the observed in vivo bactericidal effect of the FOM treatment. This work may provide a deeper understanding of the role of the host’s immune defences and antibiotic interactions in microbial infections. PMID:26778774

  3. Legionella pneumophila Suppresses Interleukin-12 Production by Macrophages

    PubMed Central

    Matsunaga, Kazuto; Klein, Thomas W.; Newton, Catherine; Friedman, Herman; Yamamoto, Yoshimasa

    2001-01-01

    In vitro infection of macrophages with Legionella pneumophila induced interleukin-1α (IL-1α), IL-10, monocyte chemotactic protein 1 (MCP-1), and MCP-3 but not IL-12. The lipopolysaccharide (LPS)-induced production of IL-12 was down-regulated by infection with virulent L. pneumophila, but other cytokines were not affected. In contrast, avirulent L. pneumophila or UV-killed, virulent L. pneumophila did not induce any suppression of IL-12. The IL-12 suppression occurred at the level of mRNA accumulation for IL-12 genes in response to LPS stimulation, but the infection induced a marked accumulation of mRNA for both MCP-1 and MCP-3, which are known to suppress IL-12 production in LPS-stimulated macrophages. However, pretreatment of macrophages with MCP-1 did not suppress LPS-induced IL-12 production at the concentrations induced by L. pneumophila infection. These results suggest that L. pneumophila selectively suppresses IL-12 production induced by LPS from macrophages in vitro by an MCP-independent mechanism. PMID:11179377

  4. PDT-treated apoptotic cells induce macrophage synthesis NO

    NASA Astrophysics Data System (ADS)

    Song, S.; Xing, D.; Zhou, F. F.; Chen, W. R.

    2009-11-01

    Nitric oxide (NO) is a biologically active molecule which has multi-functional in different species. As a second messenger and neurotransmitter, NO is not only an important regulatory factor between cells' information transmission, but also an important messenger in cell-mediated immunity and cytotoxicity. On the other side, NO is involving in some diseases' pathological process. In pathological conditions, the macrophages are activated to produce a large quantity of nitric oxide synthase (iNOS), which can use L-arginine to produce an excessive amount of NO, thereby killing bacteria, viruses, parasites, fungi, tumor cells, as well as in other series of the immune process. In this paper, photofrin-based photodynamic therapy (PDT) was used to treat EMT6 mammary tumors in vitro to induce apoptotic cells, and then co-incubation both apoptotic cells and macrophages, which could activate macrophage to induce a series of cytotoxic factors, especially NO. This, in turn, utilizes macrophages to activate a cytotoxic response towards neighboring tumor cells. These results provided a new idea for us to further study the immunological mechanism involved in damaging effects of PDT, also revealed the important function of the immune effect of apoptotic cells in PDT.

  5. Macrophage polarization following chitosan implantation.

    PubMed

    Vasconcelos, Daniela P; Fonseca, Ana C; Costa, Madalena; Amaral, Isabel F; Barbosa, Mário A; Águas, Artur P; Barbosa, Judite N

    2013-12-01

    Macrophages are a key cell in the host response to implants and can be polarized into different phenotypes capable of inducing both detrimental and beneficial outcomes in tissue repair and remodeling, being important in tissue engineering and regenerative medicine. The objective of this study was to evaluate the macrophage response to 3D porous chitosan (Ch) scaffolds with different degrees of acetylation (DA, 5% and 15%). The M1/M2 phenotypic polarization profile of macrophages was investigated in vivo using a rodent air-pouch model. Our results show that the DA affects the macrophage response. Ch scaffolds with DA 5% induced the adhesion of lower numbers of inflammatory cells, being the M2 the predominant phenotypic profile among the adherent macrophages. In the inflammatory exudates F4/80(+)/CD206(+) cells (M2 macrophages) appeared in higher numbers then F4/80(+)/CCR7(+) cells (M1 macrophages), in addition, lower levels of pro-inflammatory cytokines together with higher levels of anti-inflammatory cytokines were found. Ch scaffolds with DA 15% showed opposite results, since M1 were the predominant macrophages both adherent to the scaffold and in the exudates, together with high levels of pro-inflammatory cytokines. In conclusion, Ch scaffolds with DA 5% induced a benign M2 anti-inflammatory macrophage response, whereas Ch scaffolds with DA 15% caused a macrophage M1 pro-inflammatory response.

  6. Imaging macrophages with nanoparticles

    NASA Astrophysics Data System (ADS)

    Weissleder, Ralph; Nahrendorf, Matthias; Pittet, Mikael J.

    2014-02-01

    Nanomaterials have much to offer, not only in deciphering innate immune cell biology and tracking cells, but also in advancing personalized clinical care by providing diagnostic and prognostic information, quantifying treatment efficacy and designing better therapeutics. This Review presents different types of nanomaterial, their biological properties and their applications for imaging macrophages in human diseases, including cancer, atherosclerosis, myocardial infarction, aortic aneurysm, diabetes and other conditions. We anticipate that future needs will include the development of nanomaterials that are specific for immune cell subsets and can be used as imaging surrogates for nanotherapeutics. New in vivo imaging clinical tools for noninvasive macrophage quantification are thus ultimately expected to become relevant to predicting patients' clinical outcome, defining treatment options and monitoring responses to therapy.

  7. Early hematopoiesis and macrophage development.

    PubMed

    McGrath, Kathleen E; Frame, Jenna M; Palis, James

    2015-12-01

    The paradigm that all blood cells are derived from hematopoietic stem cells (HSCs) has been challenged by two findings. First, there are tissue-resident hematopoietic cells, including subsets of macrophages that are not replenished by adult HSCs, but instead are maintained by self-renewal of fetal-derived cells. Second, during embryogenesis, there is a conserved program of HSC-independent hematopoiesis that precedes HSC function and is required for embryonic survival. The presence of waves of HSC-independent hematopoiesis as well as fetal HSCs raises questions about the origin of fetal-derived adult tissue-resident macrophages. In the murine embryo, historical examination of embryonic macrophage and monocyte populations combined with recent reports utilizing genetic lineage-tracing approaches has led to a model of macrophage ontogeny that can be integrated with existing models of hematopoietic ontogeny. The first wave of hematopoiesis contains primitive erythroid, megakaryocyte and macrophage progenitors that arise in the yolk sac, and these macrophage progenitors are the source of early macrophages throughout the embryo, including the liver. A second wave of multipotential erythro-myeloid progenitors (EMPs) also arises in the yolk sac. EMPs colonize the fetal liver, initiating myelopoiesis and forming macrophages. Lineage tracing indicates that this second wave of macrophages are distributed in most fetal tissues, although not appreciably in the brain. Thus, fetal-derived adult tissue-resident macrophages, other than microglia, appear to predominately derive from EMPs. While HSCs emerge at midgestation and colonize the fetal liver, the relative contribution of fetal HSCs to tissue macrophages at later stages of development is unclear. The inclusion of macrophage potential in multiple waves of hematopoiesis is consistent with reports of their functional roles throughout development in innate immunity, phagocytosis, and tissue morphogenesis and remodeling

  8. Emodin Bidirectionally Modulates Macrophage Polarization and Epigenetically Regulates Macrophage Memory.

    PubMed

    Iwanowycz, Stephen; Wang, Junfeng; Altomare, Diego; Hui, Yvonne; Fan, Daping

    2016-05-27

    Macrophages are pleiotropic cells capable of performing a broad spectrum of functions. Macrophage phenotypes are classified along a continuum between the extremes of proinflammatory M1 macrophages and anti-inflammatory M2 macrophages. The seemingly opposing functions of M1 and M2 macrophages must be tightly regulated for an effective and proper response to foreign molecules or damaged tissue. Excessive activation of either M1 or M2 macrophages contributes to the pathology of many diseases. Emodin is a Chinese herb-derived compound and has shown potential to inhibit inflammation in various settings. In this study, we tested the ability of emodin to modulate the macrophage response to both M1 and M2 stimuli. Primary mouse macrophages were stimulated with LPS/IFNγ or IL4 with or without emodin, and the effects of emodin on gene transcription, cell signaling pathways, and histone modifications were examined by a variety of approaches, including microarray, quantitative real-time PCR, Western blotting, chromatin immunoprecipitation, and functional assays. We found that emodin bidirectionally tunes the induction of LPS/IFNγ- and IL4-responsive genes through inhibiting NFκB/IRF5/STAT1 signaling and IRF4/STAT6 signaling, respectively. Thereby, emodin modulates macrophage phagocytosis, migration, and NO production. Furthermore, emodin inhibited the removal of H3K27 trimethylation (H3K27m3) marks and the addition of H3K27 acetylation (H3K27ac) marks on genes required for M1 or M2 polarization of macrophages. In conclusion, our data suggest that emodin is uniquely able to suppress the excessive response of macrophages to both M1 and M2 stimuli and therefore has the potential to restore macrophage homeostasis in various pathologies.

  9. Epigenomics of macrophages.

    PubMed

    Gosselin, David; Glass, Christopher K

    2014-11-01

    Macrophages play essential roles in tissue homeostasis, pathogen elimination, and tissue repair. A defining characteristic of these cells is their ability to efficiently adapt to a variety of abruptly changing and complex environments. This ability is intrinsically linked to a capacity to quickly alter their transcriptome, and this is tightly associated with the epigenomic organization of these cells and, in particular, their enhancer repertoire. Indeed, enhancers are genomic sites that serve as platforms for the integration of signaling pathways with the mechanisms that regulate mRNA transcription. Notably, transcription is pervasive at active enhancers and enhancer RNAs (eRNAs) are tightly coupled to regulated transcription of protein-coding genes. Furthermore, given that each cell type possesses a defining enhancer repertoire, studies on enhancers provide a powerful method to study how specialization of functions among the diverse macrophage subtypes may arise. Here, we review recent studies providing insights into the distinct mechanisms that contribute to the establishment of enhancers and their role in the regulation of transcription in macrophages.

  10. Epigenomics of macrophages

    PubMed Central

    Gosselin, David; Glass, Christopher K

    2014-01-01

    Summary Macrophages play essential roles in tissue homeostasis, pathogen elimination, and tissue repair. A defining characteristic of these cells is their ability to efficiently adapt to a variety of abruptly changing and complex environments. This ability is intrinsically linked to a capacity to quickly alter their transcriptome, and this is tightly associated with the epigenomic organization of these cells and, in particular, their enhancer repertoire. Indeed, enhancers are genomic sites that serve as platforms for the integration of signaling pathways with the mechanisms that regulate mRNA transcription. Notably, transcription is pervasive at active enhancers and enhancer RNAs (eRNAs) are tightly coupled to regulated transcription of protein-coding genes. Furthermore, given that each cell type possesses a defining enhancer repertoire, studies on enhancers provide a powerful method to study how specialization of functions among the diverse macrophage subtypes may arise. Here, we review recent studies providing insights into the distinct mechanisms that contribute to the establishment of enhancers and their role in the regulation of transcription in macrophages. PMID:25319330

  11. Antigen-Mediated Fusion of Specifically Sensitized Rabbit Alveolar Macrophages

    PubMed Central

    Galindo, B.

    1972-01-01

    Rabbits sensitized intravenously with heat-killed Mycobacterium tuberculosis (strain H37Ra) suspended in mineral oil developed a strong pulmonary granulomatous response which reached its peak about 3 to 4 weeks after injection. Alveolar cells (4 × 106 cells/ml of tissue culture medium 199) procured 6 weeks after sensitization showed extensive development of multinucleated giant cells after 12 hr of incubation in tissue culture flasks containing heat-killed H37Ra (5 μg/ml). Giant cells measured 80 μm to 2.5 mm in length and contained between 30 and 700 nuclei. In contrast, no giant cells were observed when similar samples of the same cell populations were incubated in flasks containing: (i) no mycobacteria; (ii) heat-killed Escherichia coli; (iii) heat-killed Bacillus subtilis; (iv) latex particles; (v) ovalbumin; or (vi) phytohemagglutinin. The addition of immune (anti-H37Ra) sera potentiated the phenomenon of giant cell formation. In addition, supernatant fluids obtained from sensitive alveolar cells incubated with H37Ra were capable of inducing giant cell formation when incubated with nonsensitized alveolar cells. The results suggest that fusion of alveolar macrophages is mediated by an immunological mechanism. Images PMID:4629127

  12. Imaging and measuring the biophysical properties of Fc gamma receptors on single macrophages using atomic force microscopy

    SciTech Connect

    Li, Mi; Liu, Lianqing; Xi, Ning; Wang, Yuechao; Xiao, Xiubin; Zhang, Weijing

    2013-09-06

    Highlights: •Nanoscale cellular ultra-structures of macrophages were observed. •The binding affinities of FcγRs were measured directly on macrophages. •The nanoscale distributions of FcγRs were mapped on macrophages. -- Abstract: Fc gamma receptors (FcγR), widely expressed on effector cells (e.g., NK cells, macrophages), play an important role in clinical cancer immunotherapy. The binding of FcγRs to the Fc portions of antibodies that are attached to the target cells can activate the antibody-dependent cell-mediated cytotoxicity (ADCC) killing mechanism which leads to the lysis of target cells. In this work, we used atomic force microscopy (AFM) to observe the cellular ultra-structures and measure the biophysical properties (affinity and distribution) of FcγRs on single macrophages in aqueous environments. AFM imaging was used to obtain the topographies of macrophages, revealing the nanoscale cellular fine structures. For molecular interaction recognition, antibody molecules were attached onto AFM tips via a heterobifunctional polyethylene glycol (PEG) crosslinker. With AFM single-molecule force spectroscopy, the binding affinities of FcγRs were quantitatively measured on single macrophages. Adhesion force mapping method was used to localize the FcγRs, revealing the nanoscale distribution of FcγRs on local areas of macrophages. The experimental results can improve our understanding of FcγRs on macrophages; the established approach will facilitate further research on physiological activities involved in antibody-based immunotherapy.

  13. Killing of Brucella abortus by bovine serum.

    PubMed

    Corbeil, L B; Blau, K; Inzana, T J; Nielsen, K H; Jacobson, R H; Corbeil, R R; Winter, A J

    1988-12-01

    Studies of the serum bactericidal system in bovine brucellosis were undertaken to investigate the role of the humoral immune response in protection of cattle against the facultative intracellular parasite Brucella abortus. Fresh sera from normal control cattle, infected cattle, and cattle immunized with B. abortus cell envelopes were collected before treatment and during the course of immunization or infection. Normal fresh bovine serum or fresh agammaglobulinemic serum from colostrum-deprived calves was effective in killing smooth virulent B. abortus 2308, but rough strains RB51 (a rough mutant of strain 2308) and 45/20 were much more sensitive to serum. The difference in susceptibility to serum was shown to be correlated with differences in lipopolysaccharide chemotype, with the more resistant strain 2308 having O polysaccharide and the more susceptible strains 45/20 and RB51 lacking O side chains. By treatment of fresh serum with MgCl2 and EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] killing was shown to occur via the classical pathway of complement activation. When antibody to B. abortus was present, killing of strain RB51 increased but killing of smooth strain 2308 decreased. The earliest antibody response in serum from infected animals did not interfere with killing. When affinity-purified bovine immunoglobulins specific for B. abortus smooth lipopolysaccharide were added to fresh normal bovine serum, immunoglobulin G1 (IgG1) and IgG2 isotypes blocked killing but IgM and IgA isotypes did not. Thus, it appears that serum from previously unexposed animals or animals early during infection can kill smooth B. abortus, an appropriate defense mechanism before the organism becomes intracellular. At later stages of infection, blocking antibodies predominate.

  14. Photodynamic quenched cathepsin activity based probes for cancer detection and macrophage targeted therapy.

    PubMed

    Ben-Nun, Yael; Merquiol, Emmanuelle; Brandis, Alexander; Turk, Boris; Scherz, Avigdor; Blum, Galia

    2015-01-01

    Elevated cathepsins levels and activities are found in several types of human cancer, making them valuable biomarkers for detection and targeting therapeutics. We designed small molecule quenched activity-based probes (qABPs) that fluoresce upon activity-dependent covalent modification, yielding cell killing by Photodynamic Therapy (PDT). These novel molecules are highly selective theranostic probes that enable both detection and treatment of cancer with minimal side effects. Our qABPs carry a photosensitizer (PS), which is activated by light, resulting in oxidative stress and subsequent cell ablation, and a quencher that when removed by active cathepsins allow the PS to fluoresce and demonstrate PD properties. Our most powerful and stable PS-qABP, YBN14, consists of a selective cathepsin recognition sequence, a QC-1 quencher and a new bacteriochlorin derivative as a PS. YBN14 allowed rapid and selective non-invasive in vivo imaging of subcutaneous tumors and induced specific tumor macrophage apoptosis by light treatment, resulting in a substantial tumor shrinkage in an aggressive breast cancer mouse model. These results demonstrate for the first time that the PS-qABPs technology offers a functional theranostic tool, which can be applied to numerous tumor types and other inflammation-associated diseases.

  15. Photodynamic Quenched Cathepsin Activity Based Probes for Cancer Detection and Macrophage Targeted Therapy

    PubMed Central

    Ben-Nun, Yael; Merquiol, Emmanuelle; Brandis, Alexander; Turk, Boris; Scherz, Avigdor; Blum, Galia

    2015-01-01

    Elevated cathepsins levels and activities are found in several types of human cancer, making them valuable biomarkers for detection and targeting therapeutics. We designed small molecule quenched activity-based probes (qABPs) that fluoresce upon activity-dependent covalent modification, yielding cell killing by Photodynamic Therapy (PDT). These novel molecules are highly selective theranostic probes that enable both detection and treatment of cancer with minimal side effects. Our qABPs carry a photosensitizer (PS), which is activated by light, resulting in oxidative stress and subsequent cell ablation, and a quencher that when removed by active cathepsins allow the PS to fluoresce and demonstrate PD properties. Our most powerful and stable PS-qABP, YBN14, consists of a selective cathepsin recognition sequence, a QC-1 quencher and a new bacteriochlorin derivative as a PS. YBN14 allowed rapid and selective non-invasive in vivo imaging of subcutaneous tumors and induced specific tumor macrophage apoptosis by light treatment, resulting in a substantial tumor shrinkage in an aggressive breast cancer mouse model. These results demonstrate for the first time that the PS-qABPs technology offers a functional theranostic tool, which can be applied to numerous tumor types and other inflammation-associated diseases. PMID:26000057

  16. MicroRNA-155 in exosomes secreted from helicobacter pylori infection macrophages immunomodulates inflammatory response

    PubMed Central

    Wang, Jianjun; Deng, Zhiyong; Wang, Zeyou; Wu, Jianhong; Gu, Tao; Jiang, Yibiao; Li, Guangxin

    2016-01-01

    Exosomes containing microRNA-155 act as molecule carriers during immune cell-cell communication and play an important role in the inflammatory response of H. pylori infection macrophages. Previous reports have found that miR-155 was over-expressed in H. pylori infection macrophages, but the significance of which is still unknown. In this study, we analyzed the impact of miR-155 loaded in exosomes derived from macrophages to the inflammatory response of H. pylori infection macrophages and possible mechanisms. We found that miR-155 promoted the expression of inflammatory cytokines including TNF-a, IL-6, IL-23, but also increased the expression of CD40, CD63, CD81, and MCH-I. Meanwhile, inflammatory signal pathways proteins, such as MyD88, NF-κB in H. pylori infection macrophages were down-regulated due to the over-expression of miR-155. Experiments in vitro or in vivo revealed that miR-155 promoted macrophages to inhibit or kill H. pylori by regulating the inflammatory response of cells to prevent the gastritis caused by H. pylori infection. These findings contribute to the understanding of miR-155 contained in exosomes in inflammatory responses of H. pylori infection macrophages. PMID:27725852

  17. Glycoconjugates as Mediators of Nitric Oxide Production upon Exposure to Bacterial Spores by Macrophages

    NASA Astrophysics Data System (ADS)

    Lahiani, Mohamed; Soderberg, Lee; Tarasenko, Olga

    2011-06-01

    Phagocytes generate nitric oxide (NO) in large quantities to combat bacteria. The spore-producing Gram-positive organisms of Bacillus cereus family are causative agents from mild to a life threatening infection in humans and domestic animals. Our group have shown that glycoconjugates (GCs) activate macrophages and enhance killing of Bacillus spores. In this investigation, we will explore the effect of different GCs structures on NO production. The objective of this study is to study effects of GCs 2, 4, 6, 8, 10 on NO release upon exposure to B. cereus and Bacillus anthracis spores by macrophages. Our results demonstrated that GCs activated macrophages and increased NO production using studied GCs ligands compared to macrophage only (p<0.001). GC2 and GC8 were able to further increase NO production in macrophages compared to the B. anthracis spores treated macrophages (p<0.001). Our finding suggests that GCs could be used as potential mediators of NO production in macrophages to fight B. anthracis and other pathogens.

  18. Epigenetic Control of Macrophage Polarisation and Soluble Mediator Gene Expression during Inflammation

    PubMed Central

    2016-01-01

    Macrophages function as sentinel cells, which constantly monitor the host environment for infection or injury. Macrophages have been shown to exhibit a spectrum of activated phenotypes, which can often be categorised under the M1/M2 paradigm. M1 macrophages secrete proinflammatory cytokines and chemokines, such as TNF-α, IL-6, IL-12, CCL4, and CXCL10, and induce phagocytosis and oxidative dependent killing mechanisms. In contrast, M2 macrophages support wound healing and resolution of inflammation. In the past decade, interest has grown in understanding the mechanisms involved in regulating macrophage activation. In particular, epigenetic control of M1 or M2 activation states has been shown to rely on posttranslational modifications of histone proteins adjacent to inflammatory-related genes. Changes in methylation and acetylation of histones by methyltransferases, demethylases, acetyltransferases, and deacetylases can all impact how macrophage phenotypes are generated. In this review, we summarise the latest advances in the field of epigenetic regulation of macrophage polarisation to M1 or M2 states, with particular focus on the cytokine and chemokine profiles associated with these phenotypes. PMID:27143818

  19. Cryptococcus neoformans-induced macrophage lysosome damage crucially contributes to fungal virulence1

    PubMed Central

    Davis, Michael J.; Eastman, Alison J.; Qiu, Yafeng; Gregorka, Brian; Kozel, Thomas R.; Osterholzer, John J.; Curtis, Jeffrey L.; Swanson, Joel A.; Olszewski, Michal A.

    2015-01-01

    Upon ingestion by macrophages, Cryptococcus neoformans (Cn) can survive and replicate intracellularly unless the macrophages become classically activated. The mechanism enabling intracellular replication is not fully understood; neither are the mechanisms which allow classical activation to counteract replication. Cn-induced lysosome damage was observed in infected murine bone marrow-derived macrophages, increased with time and required yeast viability. To demonstrate lysosome damage in the infected host, we developed a novel flow-cytometric method for measuring lysosome damage. Increased lysosome damage was found in Cn-containing lung cells compared to Cn–free cells. Among Cn-containing myeloid cells, recently recruited cells displayed lower damage than resident cells, consistent with the protective role of recruited macrophages. The magnitude of lysosome damage correlated with increased Cn replication. Experimental induction of lysosome damage increased Cn replication. Activation of macrophages with IFN-γ abolished macrophage lysosome damage and enabled increased killing of Cn. We conclude that induction of lysosome damage is an important Cn survival strategy and that classical activation of host macrophages counters replication by preventing damage. Thus, therapeutic strategies which decrease lysosomal damage, or increase resistance to such damage, could be valuable in treating cryptococcal infections. PMID:25637026

  20. Interleukin-32 induces the differentiation of monocytes into macrophage-like cells

    PubMed Central

    Netea, Mihai G.; Lewis, Eli C.; Azam, Tania; Joosten, Leo A. B.; Jaekal, Jun; Bae, Su-Young; Dinarello, Charles A.; Kim, Soo-Hyun

    2008-01-01

    After emigration from the bone marrow to the peripheral blood, monocytes enter tissues and differentiate into macrophages, the prototype scavenger of the immune system. By ingesting and killing microorganisms and removing cellular debris, macrophages also process antigens as a first step in mounting a specific immune response. IL-32 is a cytokine inducing proinflammatory cytokines and chemokines via p38-MAPK and NF-κB. In the present study, we demonstrate that IL-32 induces differentiation of human blood monocytes as well as THP-1 leukemic cells into macrophage-like cells with functional phagocytic activity for live bacteria. Muramyl dipepide (MDP), the ligand for the intracellular nuclear oligomerization domain (NOD) 2 receptor, has no effect on differentiation alone but augments the monocyte-to-macrophage differentiation by IL-32. Unexpectedly, IL-32 reversed GM-CSF/IL-4-induced dendritic cell differentiation to macrophage-like cells. Whereas the induction of TNFα, IL-1β, and IL-6 by IL-32 is mediated by p38-MAPK, IL-32-induced monocyte-to-macrophage differentiation is mediated through nonapoptotic, caspase-3-dependent mechanisms. Thus, IL-32 not only contributes to host responses through the induction of proinflammatory cytokines but also directly affects specific immunity by differentiating monocytes into macrophage-like cells. PMID:18296636

  1. System x(c)(-) regulates microglia and macrophage glutamate excitotoxicity in vivo.

    PubMed

    Kigerl, Kristina A; Ankeny, Daniel P; Garg, Sanjay K; Wei, Ping; Guan, Zhen; Lai, Wenmin; McTigue, Dana M; Banerjee, Ruma; Popovich, Phillip G

    2012-01-01

    It is widely believed that microglia and monocyte-derived macrophages (collectively referred to as central nervous system (CNS) macrophages) cause excitotoxicity in the diseased or injured CNS. This view has evolved mostly from in vitro studies showing that neurotoxic concentrations of glutamate are released from CNS macrophages stimulated with lipopolysaccharide (LPS), a potent inflammogen. We hypothesized that excitotoxic killing by CNS macrophages is more rigorously controlled in vivo, requiring both the activation of the glutamate/cystine antiporter (system x(c)(-)) and an increase in extracellular cystine, the substrate that drives glutamate release. Here, we show that non-traumatic microinjection of low-dose LPS into spinal cord gray matter activates CNS macrophages but without causing overt neuropathology. In contrast, neurotoxic inflammation occurs when LPS and cystine are co-injected. Simultaneous injection of NBQX, an antagonist of AMPA glutamate receptors, reduces the neurotoxic effects of LPS+cystine, implicating glutamate as a mediator of neuronal cell death in this model. Surprisingly, neither LPS nor LPS+cystine adversely affects survival of oligodendrocytes or oligodendrocyte progenitor cells. Ex vivo analyses show that redox balance in microglia and macrophages is controlled by induction of system x(c)(-) and that high GSH:GSSG ratios predict the neurotoxic potential of these cells. Together, these data indicate that modulation of redox balance in CNS macrophages, perhaps through regulating system x(c)(-), could be a novel approach for attenuating injurious neuroinflammatory cascades.

  2. Binding and Phagocytosis by Opsonized and Nonopsonized Channel Catfish Macrophages of Viable DsRed-fluorescent-labeled Edwardsiella ictaluri

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Phagocyte-mediated killing of bacterial pathogens is one of the major defensive mechanisms in fish. The binding, uptake and destruction of recombinant fluorescent protein DsRed transformed Edwardsiella ictaluri by opsonized and nonopsonized channel catfish (Ictalurus punctatus) macrophages was chara...

  3. Bactericidal activity of macrophages against Streptococcus uberis is different in mammary gland secretions of lactating and drying off cows.

    PubMed

    Denis, Michel; Parlane, Natalie A; Lacy-Hulbert, S Jane; Summers, Emma L; Buddle, Bryce M; Wedlock, D Neil

    2006-11-15

    The aim of this study was to compare the ability of milk macrophages and macrophages from the mammary gland secretions during the mid-dry period for their interaction with the mastitis-causing Streptococcus uberis. We also aimed to determine if S. uberis induced the release of the cytokine tumour necrosis alpha (TNF-alpha) and the bactericidal moiety nitric oxide (NO) from milk macrophages of lactating cows and macrophages from the mammary gland secretions at the mid-dry period. Macrophages were isolated from the mammary gland secretions of cows during the mid-lactation or mid-dry period, and compared with blood monocytes for their interaction with the important mastitis-causing pathogen S. uberis. When infected in vitro with S. uberis, milk macrophages from lactating cows with S. uberis released modest amounts of the cytokine tumour necrosis factor alpha (TNF-alpha) (139 pg/ml) and the bactericidal moiety nitric oxide (NO) (3-4 microM of nitrite). Blood monocytes from lactating cows released significantly higher amounts of TNF-alpha (345 +/- 143 pg/ml) and NO (7 +/- 2 microM of nitrite) after interaction with S. uberis, compared to milk macrophages (P < 0.01 for both TNF-alpha and NO). Stimulation of blood monocytes with the cytokine interferon-gamma (IFN-gamma) enhanced significantly the release of NO and TNF-alpha, but IFN-gamma did not significantly enhance the production of NO and TNF-alpha by milk macrophages from lactating cows. Milk macrophages from all lactating cows failed to kill S. uberis efficiently, and this lack of killing was unaffected by prior treatment with gamma interferon (IFN-gamma) (P > 0.05). Rather, S. uberis multiplied significantly inside infected milk macrophages from lactating cows, with a two-fold increase in bacterial numbers at 2 h post-infection. Milk macrophages from lactating cows were able however, to kill a significant proportion (50-60%, P < 0.01) of phagocytosed Staphylococcus aureus. Blood monocytes from all cows were found

  4. Hydroxyapatite and urate crystal induced cytokine release by macrophages.

    PubMed Central

    Alwan, W H; Dieppe, P A; Elson, C J; Bradfield, J W

    1989-01-01

    Destructive osteoarthritis is characterised by rapidly progressive joint destruction associated with intra-articular deposition of hydroxyapatite crystals. The possible role of such crystals in the pathogenesis of this condition was investigated by testing the ability of hydroxyapatite crystals to stimulate the production of bone resorbing activity from mouse peritoneal macrophages. Urate crystals were used for comparison. Culture supernatants were tested for bone resorbing activity using the mouse calvarial bone resorption assay, for interleukin 1 using a standard lymphocyte activation assay, and for prostaglandin E2 by radioimmunoassay. Culture supernatants from macrophages incubated with hydroxyapatite crystals contained dialysable bone resorbing activity, high concentrations of prostaglandin E2, but no interleukin 1 like activity. The production of the bone resorbing agent was prevented by culturing macrophages with hydroxyapatite crystals in the presence of indomethacin. By contrast, culture supernatants from macrophages incubated with urate crystals contained bone resorbing activity, which was only partly removed by dialysis, and interleukin 1 like activity. The latter was shown to be increased in culture supernatants from macrophages incubated with urate crystals in the presence of indomethacin, while production of bone resorbing activity was partially inhibited. It is considered that the bone resorbing activity liberated from macrophages stimulated by hydroxyapatite crystals can be explained by the presence of prostaglandin E2 alone, whereas the activity liberated by urate crystals is due to both prostaglandin E2 and interleukin 1. PMID:2545171

  5. Macrophage Roles Following Myocardial Infarction

    PubMed Central

    Lambert, Jessica M.; Lopez, Elizabeth F.; Lindsey, Merry L.

    2010-01-01

    Following myocardial infarction (MI), circulating blood monocytes respond to chemotactic factors, migrate into the infarcted myocardium, and differentiate into macrophages. At the injury site, macrophages remove necrotic cardiac myocytes and apoptotic neutrophils; secrete cytokines, chemokines, and growth factors; and modulate phases of the angiogenic response. As such, the macrophage is a primary responder cell type that is involved in the regulation of post-MI wound healing at multiple levels. This review summarizes what is currently known about macrophage functions post-MI and borrows literature from other injury and inflammatory models to speculate on additional roles. Basic science and clinical avenues that remain to be explored are also discussed. PMID:18656272

  6. Bioelectric modulation of macrophage polarization

    NASA Astrophysics Data System (ADS)

    Li, Chunmei; Levin, Michael; Kaplan, David L.

    2016-02-01

    Macrophages play a critical role in regulating wound healing and tissue regeneration by changing their polarization state in response to local microenvironmental stimuli. The native roles of polarized macrophages encompass biomaterials and tissue remodeling needs, yet harnessing or directing the polarization response has been largely absent as a potential strategy to exploit in regenerative medicine to date. Recent data have revealed that specific alteration of cells’ resting potential (Vmem) is a powerful tool to direct proliferation and differentiation in a number of complex tissues, such as limb regeneration, craniofacial patterning and tumorigenesis. In this study, we explored the bioelectric modulation of macrophage polarization by targeting ATP sensitive potassium channels (KATP). Glibenclamide (KATP blocker) and pinacidil (KATP opener) treatment not only affect macrophage polarization, but also influence the phenotype of prepolarized macrophages. Furthermore, modulation of cell membrane electrical properties can fine-tune macrophage plasticity. Glibenclamide decreased the secretion and gene expression of selected M1 markers, while pinacidil augmented M1 markers. More interestingly, glibencalmide promoted macrophage alternative activation by enhancing certain M2 markers during M2 polarization. These findings suggest that control of bioelectric properties of macrophages could offer a promising approach to regulate macrophage phenotype as a useful tool in regenerative medicine.

  7. Macrophages in homeostatic immune function.

    PubMed

    Jantsch, Jonathan; Binger, Katrina J; Müller, Dominik N; Titze, Jens

    2014-01-01

    Macrophages are not only involved in inflammatory and anti-infective processes, but also play an important role in maintaining tissue homeostasis. In this review, we summarize recent evidence investigating the role of macrophages in controlling angiogenesis, metabolism as well as salt and water balance. Particularly, we summarize the importance of macrophage tonicity enhancer binding protein (TonEBP, also termed nuclear factor of activated T-cells 5 [NFAT5]) expression in the regulation of salt and water homeostasis. Further understanding of homeostatic macrophage function may lead to new therapeutic approaches to treat ischemia, hypertension and metabolic disorders. PMID:24847274

  8. Macrophages in homeostatic immune function

    PubMed Central

    Jantsch, Jonathan; Binger, Katrina J.; Müller, Dominik N.; Titze, Jens

    2014-01-01

    Macrophages are not only involved in inflammatory and anti-infective processes, but also play an important role in maintaining tissue homeostasis. In this review, we summarize recent evidence investigating the role of macrophages in controlling angiogenesis, metabolism as well as salt and water balance. Particularly, we summarize the importance of macrophage tonicity enhancer binding protein (TonEBP, also termed nuclear factor of activated T-cells 5 [NFAT5]) expression in the regulation of salt and water homeostasis. Further understanding of homeostatic macrophage function may lead to new therapeutic approaches to treat ischemia, hypertension and metabolic disorders. PMID:24847274

  9. Kill fluid for oil field operations

    SciTech Connect

    Sydansk, R.D.

    1990-08-14

    This patent describes a process employing a kill fluid to substantially reduce the volumetric flow of formation fluid into a wellbore penetrating a formation containing the formation fluid below an earthen surface. It comprises: admixing components of a continuous flowing gel at the surface comprising of water-soluble carboxylate-containing polymer, a complex capable of crosslinking the polymer and formed of at least one electropositive chromium III species and at least one electronegative carboxylatespecies, and an aqueous solvent for the polymer and the complex; crosslinking the polymer and the complex to form the gel, wherein the kill fluid comprises the gel; placing a volume of the kill fluid in the wellbore sufficient to create a hydrostatic head which exerts a kill fluid pressure against the formation fluid substantially equal to or greater than the formation fluid pressure and thereby substantially reduces the volumetric flow of the formation fluid into the wellbore; performing an oil field operation after placing the volume of the kill fluid in the wellbore; and removing the gel from the wellbore to substantially restore the volumetric flow of the formation fluid into the wellbore.

  10. Killing superalgebras for Lorentzian four-manifolds

    NASA Astrophysics Data System (ADS)

    de Medeiros, Paul; Figueroa-O'Farrill, José; Santi, Andrea

    2016-06-01

    We determine the Killing superalgebras underpinning field theories with rigid unextended supersymmetry on Lorentzian four-manifolds by re-interpreting them as filtered deformations of mathbb{Z} -graded subalgebras with maximum odd dimension of the N = 1 Poincaré superalgebra in four dimensions. Part of this calculation involves computing a Spencer cohomology group which, by analogy with a similar result in eleven dimensions, prescribes a notion of Killing spinor, which we identify with the defining condition for bosonic supersymmetric backgrounds of minimal off-shell supergravity in four dimensions. We prove that such Killing spinors always generate a Lie superalgebra, and that this Lie superalgebra is a filtered deformation of a subalgebra of the N = 1 Poincaré superalgebra in four dimensions. Demanding the flatness of the connection defining the Killing spinors, we obtain equations satisfied by the maximally supersymmetric backgrounds. We solve these equations, arriving at the classification of maximally supersymmetric backgrounds whose associated Killing superalgebras are precisely the filtered deformations we classify in this paper.

  11. Non-standard symmetries and Killing tensors

    NASA Astrophysics Data System (ADS)

    Visinescu, Mihai

    2009-10-01

    Higher order symmetries corresponding to Killing tensors are investigated. The intimate relation between Killing-Yano tensors and non-standard supersymmetries is pointed out. The gravitational anomalies are absent if the hidden symmetry is associated with a Killing-Yano tensor. In the Dirac theory on curved spaces, Killing-Yano tensors generate Dirac type operators involved in interesting algebraic structures as dynamical algebras or even infinite dimensional algebras or superalgebras. The general results are applied to space-times which appear in modern studies. The 4-dimensional Euclidean Taub-NUT space and its generalizations introduced by Iwai and Katayama are analyzed from the point of view of hidden symmetries. One presents the infinite dimensional superalgebra of Dirac type operators on Taub-NUT space that can be seen as a twisted loop algebra. The axial anomaly, interpreted as the index of the Dirac operator, is computed for the generalized Taub-NUT metrics. The existence of the conformal Killing-Yano tensors is investigated for some spaces with mixed Sasakian structures.

  12. 75 FR 62469 - Drawbridge Operation Regulations; Newtown Creek, Dutch Kills, English Kills, and Their...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-12

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF HOMELAND SECURITY Coast Guard 33 CFR Part 117 Drawbridge Operation Regulations; Newtown Creek, Dutch Kills, English Kills, and Their Tributaries, NY, Maintenance AGENCY: Coast Guard, DHS. ACTION: Notice of...

  13. Trafficking of Estrella lausannensis in human macrophages.

    PubMed

    Rusconi, Brigida; Kebbi-Beghdadi, Carole; Greub, Gilbert

    2015-07-01

    Estrella lausannensis is a new member of the Chlamydiales order. Like other Chlamydia-related bacteria, it is able to replicate in amoebae and in fish cell lines. A preliminary study investigating the pathogenic potential of Chlamydia-related bacteria found a correlation between antibody response to E. lausannensis and pneumonia in children. To further investigate the pathogenic potential of E. lausannensis, we determined its ability to grow in human macrophages and its intracellular trafficking. The replication in macrophages resulted in viable E. lausannensis; however, it caused a significant cytopathic effect. The intracellular trafficking of E. lausannensis was analyzed by determining the interaction of the Estrella-containing inclusions with various endocytic markers as well as host organelles. The E. lausannensis inclusion escaped the endocytic pathway rapidly avoiding maturation into phagolysosomes by preventing both EEA-1 and LAMP-1 accumulation. Compared to Waddlia chondrophila, another Chlamydia-related bacteria, the recruitment of mitochondria and endoplasmic reticulum was minimal for E. lausannensis inclusions. Estrella lausannensis appears to use a distinct source of nutrients and energy compared to other members of the Chlamydiales order. In conclusion, we hypothesize that E. lausannensis has a restricted growth in human macrophages, due to its reduced capacity to control programmed cell death. PMID:25857735

  14. 9 CFR 113.211 - Feline Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Feline Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.211 Feline Rhinotracheitis Vaccine, Killed Virus. Feline Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  15. 9 CFR 113.211 - Feline Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Feline Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.211 Feline Rhinotracheitis Vaccine, Killed Virus. Feline Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  16. 9 CFR 113.203 - Feline Panleukopenia Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Feline Panleukopenia Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.203 Feline Panleukopenia Vaccine, Killed Virus. Feline Panleukopenia Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  17. 9 CFR 113.206 - Wart Vaccine, Killed Virus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Wart Vaccine, Killed Virus. 113.206... Killed Virus Vaccines § 113.206 Wart Vaccine, Killed Virus. Wart Vaccine, Killed Virus, shall be prepared... content as prescribed in § 113.200(f). (d) Potency and efficacy. The efficacy of wart vaccine has...

  18. 9 CFR 113.211 - Feline Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Feline Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.211 Feline Rhinotracheitis Vaccine, Killed Virus. Feline Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  19. 9 CFR 113.210 - Feline Calicivirus Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Feline Calicivirus Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.210 Feline Calicivirus Vaccine, Killed Virus. Feline Calicivirus Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  20. 9 CFR 113.203 - Feline Panleukopenia Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Feline Panleukopenia Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.203 Feline Panleukopenia Vaccine, Killed Virus. Feline Panleukopenia Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  1. 9 CFR 113.210 - Feline Calicivirus Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Feline Calicivirus Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.210 Feline Calicivirus Vaccine, Killed Virus. Feline Calicivirus Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  2. 9 CFR 113.210 - Feline Calicivirus Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Feline Calicivirus Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.210 Feline Calicivirus Vaccine, Killed Virus. Feline Calicivirus Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  3. 9 CFR 113.206 - Wart Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Wart Vaccine, Killed Virus. 113.206... Killed Virus Vaccines § 113.206 Wart Vaccine, Killed Virus. Wart Vaccine, Killed Virus, shall be prepared... content as prescribed in § 113.200(f). (d) Potency and efficacy. The efficacy of wart vaccine has...

  4. 9 CFR 113.206 - Wart Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Wart Vaccine, Killed Virus. 113.206... Killed Virus Vaccines § 113.206 Wart Vaccine, Killed Virus. Wart Vaccine, Killed Virus, shall be prepared... content as prescribed in § 113.200(f). (d) Potency and efficacy. The efficacy of wart vaccine has...

  5. 9 CFR 113.203 - Feline Panleukopenia Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Feline Panleukopenia Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.203 Feline Panleukopenia Vaccine, Killed Virus. Feline Panleukopenia Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  6. 9 CFR 113.206 - Wart Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Wart Vaccine, Killed Virus. 113.206... Killed Virus Vaccines § 113.206 Wart Vaccine, Killed Virus. Wart Vaccine, Killed Virus, shall be prepared... content as prescribed in § 113.200(f). (d) Potency and efficacy. The efficacy of wart vaccine has...

  7. 9 CFR 113.216 - Bovine Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Bovine Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.216 Bovine Rhinotracheitis Vaccine, Killed Virus. Infectious Bovine Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  8. 9 CFR 113.216 - Bovine Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Bovine Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.216 Bovine Rhinotracheitis Vaccine, Killed Virus. Infectious Bovine Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  9. 9 CFR 113.216 - Bovine Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Bovine Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.216 Bovine Rhinotracheitis Vaccine, Killed Virus. Infectious Bovine Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  10. 9 CFR 113.206 - Wart Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Wart Vaccine, Killed Virus. 113.206... Killed Virus Vaccines § 113.206 Wart Vaccine, Killed Virus. Wart Vaccine, Killed Virus, shall be prepared... content as prescribed in § 113.200(f). (d) Potency and efficacy. The efficacy of wart vaccine has...

  11. 9 CFR 113.203 - Feline Panleukopenia Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Feline Panleukopenia Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.203 Feline Panleukopenia Vaccine, Killed Virus. Feline Panleukopenia Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  12. 9 CFR 113.211 - Feline Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Feline Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.211 Feline Rhinotracheitis Vaccine, Killed Virus. Feline Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  13. 9 CFR 113.210 - Feline Calicivirus Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Feline Calicivirus Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.210 Feline Calicivirus Vaccine, Killed Virus. Feline Calicivirus Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  14. 9 CFR 113.216 - Bovine Rhinotracheitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Bovine Rhinotracheitis Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.216 Bovine Rhinotracheitis Vaccine, Killed Virus. Infectious Bovine Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master...

  15. Single-cell analysis reveals new subset markers of murine peritoneal macrophages and highlights macrophage dynamics upon Staphylococcus aureus peritonitis.

    PubMed

    Accarias, Solène; Genthon, Clémence; Rengel, David; Boullier, Séverine; Foucras, Gilles; Tabouret, Guillaume

    2016-07-01

    Resident macrophages play a central role in maintaining tissue homeostasis and immune surveillance. Here, we used single cell-based qPCR coupled with flow cytometry analysis to further define the phenotypes of large and small resident peritoneal macrophages (LPMs and SPMs, respectively) in mice. We demonstrated that the expression of Cxcl13, IfngR1, Fizz-1 and Mrc-1 clearly distinguished between LPMs and SPMs subsets. Using these markers, the dynamics of peritoneal macrophages in a Staphylococcus aureus-induced peritonitis model were analyzed. We found that S. aureus infection triggers a massive macrophage disappearance reaction in both subsets. Thereafter, inflammatory monocytes rapidly infiltrated the cavity and differentiated to replenish the SPMs. Although phenotypically indistinguishable from resident SPMs by flow cytometry, newly recruited SPMs had a different pattern of gene expression dominated by M2 markers combined with M1 associated features (inos expression). Interestingly, S. aureus elicited SPMs showed a robust expression of Cxcl13, suggesting that these cells may endorse the role of depleted LPMs and contribute to restoring peritoneal homeostasis. These data provide information on both resident and recruited macrophages dynamics upon S. aureus infection and demonstrate that single-cell phenotyping is a promising and highly valuable approach to unraveling macrophage diversity and plasticity. PMID:27220602

  16. Strain ŽP - the first bacterial conjugation-based "kill"-"anti-kill" antimicrobial system.

    PubMed

    Starčič Erjavec, Marjanca; Petkovšek, Živa; Kuznetsova, Marina V; Maslennikova, Irina L; Žgur-Bertok, Darja

    2015-11-01

    As multidrug resistant bacteria pose one of the greatest risks to human health new alternative antibacterial agents are urgently needed. One possible mechanism that can be used as an alternative to traditional antibiotic therapy is transfer of killing agents via conjugation. Our work was aimed at providing a proof of principle that conjugation-based antimicrobial systems are possible. We constructed a bacterial conjugation-based "kill"-"anti-kill" antimicrobial system employing the well known Escherichia coli probiotic strain Nissle 1917 genetically modified to harbor a conjugative plasmid carrying the "kill" gene (colicin ColE7 activity gene) and a chromosomally encoded "anti-kill" gene (ColE7 immunity gene). The constructed strain acts as a donor in conjugal transfer and its efficiency was tested in several types of conjugal assays. Our results clearly demonstrate that conjugation-based antimicrobial systems can be highly efficient. PMID:26436830

  17. Substance P does not alter interleukin-1 expression by splenic or granuloma macrophages in murine schistosomiasis.

    PubMed

    Cook, G A; Blum, A M; Ballas, Z; Weinstock, J V

    1991-09-01

    Substance P (SP) is an undecapeptide with neurotransmitter and immunoregulatory properties. In murine schistosomiasis, ova naturally induce liver and intestinal granulomas. These granulomas contain macrophages, and eosinophils that produce SP. A report showed that human blood monocytes isolated by adherence release interleukin-1 (IL-1) in response to SP (Lotz et al. (1989) Science 241, 1218). IL-1 is important for initiation of hypersensitivity granulomas. Therefore, it was determined whether SP modulates granuloma macrophage IL-1 production in murine schistosomiasis. Macrophages were obtained from lung and liver granulomas, and from spleens of infected mice. A thymocyte proliferation assay measured IL-1 activity in culture supernatants. Total RNA, extracted from macrophages, was assayed for IL-1 alpha and beta mRNA by Northern blotting using cDNA probes. In response to lipopolysaccharide (LPS), splenic macrophages and macrophages from young lung granulomas released appreciable IL-1. Macrophages from liver granulomas, that were lesions older than the lung granulomas, were unresponsive to LPS with regard to IL-1 secretion. Yet, granuloma macrophages spontaneously expressed IL-1 alpha and beta mRNA. LPS enhanced IL-1 mRNA expression in both splenic and granuloma macrophages. Exposure of macrophages from all sources to SP did not alter IL-1 secretion or gene expression. Similarly, the responsiveness of macrophages to LPS was not affected by concomitant exposure to SP. It is concluded that, in the murine system, SP does not directly influence splenic or granuloma macrophage IL-1 secretion or gene expression. Also, it appears that macrophage secretion of IL-1 is rapidly down-regulated following granuloma elicitation.

  18. Membrane Lipid Peroxidation in Copper Alloy-Mediated Contact Killing of Escherichia coli

    PubMed Central

    Hong, Robert; Kang, Tae Y.; Michels, Corinne A.

    2012-01-01

    Copper alloy surfaces are passive antimicrobial sanitizing agents that kill bacteria, fungi, and some viruses. Studies of the mechanism of contact killing in Escherichia coli implicate the membrane as the target, yet the specific component and underlying biochemistry remain unknown. This study explores the hypothesis that nonenzymatic peroxidation of membrane phospholipids is responsible for copper alloy-mediated surface killing. Lipid peroxidation was monitored with the thiobarbituric acid-reactive substances (TBARS) assay. Survival, TBARS levels, and DNA degradation were followed in cells exposed to copper alloy surfaces containing 60 to 99.90% copper or in medium containing CuSO4. In all cases, TBARS levels increased with copper exposure levels. Cells exposed to the highest copper content alloys, C11000 and C24000, exhibited novel characteristics. TBARS increased immediately at a very rapid rate but peaked at about 30 min. This peak was associated with the period of most rapid killing, loss in membrane integrity, and DNA degradation. DNA degradation is not the primary cause of copper-mediated surface killing. Cells exposed to the 60% copper alloy for 60 min had fully intact genomic DNA but no viable cells. In a fabR mutant strain with increased levels of unsaturated fatty acids, sensitivity to copper alloy surface-mediated killing increased, TBARS levels peaked earlier, and genomic DNA degradation occurred sooner than in the isogenic parental strain. Taken together, these results suggest that copper alloy surface-mediated killing of E. coli is triggered by nonenzymatic oxidative damage of membrane phospholipids that ultimately results in the loss of membrane integrity and cell death. PMID:22247141

  19. HIV transcription is induced with cell killing

    SciTech Connect

    Woloschak, G.E.; Schreck, S.; Chang-Liu, Chin-Mei; Panozzo, J.; Libertin, C.R.

    1993-11-01

    In this report, we demonstrate that this induction of HIV-LTR transcription occurs when stably transfected HeLa cells are exposed to agents which mediate cell killing, such as UV radiation, electroporation of sucrose buffer, prolonged heating, and low and high pH. Cells cultured following UV exposure demonstrated a peak in CAT expression that is evident in viable (but not necessarily cell division-competent) cells 24 h after exposure; this inductive response continued until at least 72 h after exposure. HIV-LTR induction was dose-dependent, and the amount of CAT transcription induced was correlated with the amount of cell killing that occurred in the culture.

  20. Cytotoxic Killing and Immune Evasion by Repair

    NASA Astrophysics Data System (ADS)

    Chan, Cliburn; George, Andrew J. T.; Stark, Jaroslav

    2007-07-01

    The interaction between the immune system and pathogens is a complex one, with pathogens constantly developing new ways of evading destruction by the immune system. The immune system's task is made even harder when the pathogen in question is an intra-cellular one (such as a virus or certain bacteria) and it is necessary to kill the infected host cell in order to eliminate the pathogen. This causes damage to the host, and such killing therefore needs to be carefully controlled, particularly in tissues with poor regenerative potential, or those involved in the immune response itself. Host cells therefore possess repair mechanisms which can counteract killing by immune cells. These in turn can be subverted by pathogens which up-regulate the resistance of infected cells to killing. In this paper, we explore the hypothesis that this repair process plays an important role in determining the efficacy of evasion and escape from immune control. We model a situation where cytotoxic T lymphocytes (CTL) and natural killer (NK) cells kill pathogen-infected and tumour cells by directed secretion of preformed granules containing perforin and granzymes. Resistance to such killing can be conferred by the expression of serine protease inhibitors (serpins). These are utilized by several virally infected and tumour cells, as well as playing a role in the protection of host bystander, immune and immuneprivileged cells. We build a simple stochastic model of cytotoxic killing, where serpins can neutralize granzymes stoichiometrically by forming an irreversible complex, and the survival of the cell is determined by the balance between serpin depletion and replenishment, which in its simplest form is equivalent to the well known shot noise process. We use existing analytical results for this process, and additional simulations to analyse the effects of repair on cytotoxic killing. We then extend the model to the case of a replicating target cell population, which gives a branching process

  1. Failed CTL/NK cell killing and cytokine hypersecretion are directly linked through prolonged synapse time.

    PubMed

    Jenkins, Misty R; Rudd-Schmidt, Jesse A; Lopez, Jamie A; Ramsbottom, Kelly M; Mannering, Stuart I; Andrews, Daniel M; Voskoboinik, Ilia; Trapani, Joseph A

    2015-03-01

    Failure of cytotoxic T lymphocytes (CTLs) or natural killer (NK) cells to kill target cells by perforin (Prf)/granzyme (Gzm)-induced apoptosis causes severe immune dysregulation. In familial hemophagocytic lymphohistiocytosis, Prf-deficient infants suffer a fatal "cytokine storm" resulting from macrophage overactivation, but the link to failed target cell death is not understood. We show that prolonged target cell survival greatly amplifies the quanta of inflammatory cytokines secreted by CTLs/NK cells and that interferon-γ (IFN-γ) directly invokes the activation and secondary overproduction of proinflammatory IL-6 from naive macrophages. Furthermore, using live cell microscopy to visualize hundreds of synapses formed between wild-type, Prf-null, or GzmA/B-null CTLs/NK cells and their targets in real time, we show that hypersecretion of IL-2, TNF, IFN-γ, and various chemokines is linked to failed disengagement of Prf- or Gzm-deficient lymphocytes from their targets, with mean synapse time increased fivefold, from ∼8 to >40 min. Surprisingly, the signal for detachment arose from the dying target cell and was caspase dependent, as delaying target cell death with various forms of caspase blockade also prevented their disengagement from fully competent CTLs/NK cells and caused cytokine hypersecretion. Our findings provide the cellular mechanism through which failed killing by lymphocytes causes systemic inflammation involving recruitment and activation of myeloid cells. PMID:25732304

  2. Macrophages migrate in an activation-dependent manner to chemokines involved in neuroinflammation

    PubMed Central

    2014-01-01

    Background In neuroinflammatory diseases, macrophages can play a dual role in the process of tissue damage, depending on their activation status (M1 / M2). M1 macrophages are considered to exert damaging effects to neurons, whereas M2 macrophages are reported to aid regeneration and repair of neurons. Their migration within the central nervous system may be of critical importance in the final outcome of neurodegeneration in neuroinflammatory diseases e.g. multiple sclerosis (MS). To provide insight into this process, we examined the migratory capacity of human monocyte-derived M1 and M2 polarised macrophages towards chemoattractants, relevant for neuroinflammatory diseases like MS. Methods Primary cultures of human monocyte-derived macrophages were exposed to interferon gamma and lipopolysaccharide (LPS) to evoke proinflammatory (M1) activation or IL-4 to evoke anti-inflammatory (M2) activation. In a TAXIScan assay, migration of M0, M1 and M2 towards chemoattractants was measured and quantified. Furthermore the adhesion capacity and the expression levels of integrins as well as chemokine receptors of M0, M1 and M2 were assessed. Alterations in cell morphology were analysed using fluorescent labelling of the cytoskeleton. Results Significant differences were observed between M1 and M2 macrophages in the migration towards chemoattractants. We show that M2 macrophages migrated over longer distances towards CCL2, CCL5, CXCL10, CXCL12 and C1q compared to non-activated (M0) and M1 macrophages. No differences were observed in the adhesion of M0, M1 and M2 macrophages to multiple matrix components, nor in the expression of integrins and chemokine receptors. Significant changes were observed in the cytoskeleton organization upon stimulation with CCL2, M0, M1 and M2 macrophages adopt a spherical morphology and the cytoskeleton is rapidly rearranged. M0 and M2 macrophages are able to form filopodia, whereas M1 macrophages only adapt a spherical morphology. Conclusions

  3. Mechanisms of Defense against Intracellular Pathogens Mediated by Human Macrophages.

    PubMed

    Bloom, Barry R; Modlin, Robert L

    2016-06-01

    The key question our work has sought to address has been, "What are the necessary and sufficient conditions that engender protection from intracellular pathogens in the human host?" The origins of this work derive from a long-standing interest in the mechanisms of protection against two such paradigmatic intracellular pathogens, Mycobacterium tuberculosis and Mycobacterium leprae, that have brilliantly adapted to the human host. It was obvious that these pathogens, which cause chronic diseases and persist in macrophages, must have acquired subtle strategies to resist host microbicidal mechanisms, yet since the vast majority of individuals infected with M. tuberculosis do not develop disease, there must be some potent human antimicrobial mechanisms. What follows is not a comprehensive review of the vast literature on the role of human macrophages in protection against infectious disease, but a summary of the research in our two laboratories with collaborators that we hope has contributed to some understanding of mechanisms of resistance and pathogenesis. While mouse models revealed some necessary conditions for protection, e.g., innate immunity, Th1 cells and their cytokines, and major histocompatibility complex class I-restricted T cells, here we emphasize multiple antimicrobial mechanisms that exist in human macrophages that differ from those of most experimental animals. Prominent here is the vitamin D-dependent antimicrobial pathway common to human macrophages activated by innate and acquired immune responses, mediated by antimicrobial peptides, e.g., cathelicidin, through an interleukin-15- and interleukin-32-dependent common pathway that is necessary for macrophage killing of M. tuberculosis in vitro. PMID:27337485

  4. Adipose tissue macrophages: amicus adipem?

    PubMed Central

    Odegaard, Justin I.; Ganeshan, Kirthana; Chawla, Ajay

    2014-01-01

    Chronic overnutrition drives complex adaptations within both professional metabolic and bystander tissues that, despite intense investigation, are still poorly understood. Xu et al. (2013) now describe the unexpected ability of adipose tissue macrophages to buffer lipids released from obese adipocytes in a manner independent of inflammatory macrophage activation. PMID:24315364

  5. Biology of Bony Fish Macrophages

    PubMed Central

    Hodgkinson, Jordan W.; Grayfer, Leon; Belosevic, Miodrag

    2015-01-01

    Macrophages are found across all vertebrate species, reside in virtually all animal tissues, and play critical roles in host protection and homeostasis. Various mechanisms determine and regulate the highly plastic functional phenotypes of macrophages, including antimicrobial host defenses (pro-inflammatory, M1-type), and resolution and repair functions (anti-inflammatory/regulatory, M2-type). The study of inflammatory macrophages in immune defense of teleosts has garnered much attention, and antimicrobial mechanisms of these cells have been extensively studied in various fish models. Intriguingly, both similarities and differences have been documented for the regulation of lower vertebrate macrophage antimicrobial defenses, as compared to what has been described in mammals. Advances in our understanding of the teleost macrophage M2 phenotypes likewise suggest functional conservation through similar and distinct regulatory strategies, compared to their mammalian counterparts. In this review, we discuss the current understanding of the molecular mechanisms governing teleost macrophage functional heterogeneity, including monopoetic development, classical macrophage inflammatory and antimicrobial responses as well as alternative macrophage polarization towards tissues repair and resolution of inflammation. PMID:26633534

  6. Rabies virus replication in primary murine bone marrow macrophages and in human and murine macrophage-like cell lines: implications for viral persistence.

    PubMed

    Ray, N B; Ewalt, L C; Lodmell, D L

    1995-02-01

    To determine whether rabies viruses replicate in macrophage or macrophage-like cells, several human and murine macrophage-like cell lines, as well as primary cultures of murine bone marrow macrophages, were incubated with the Evelyn-Rokitnicki-Abelseth (ERA) virus and several different street rabies viruses (SRV). ERA rabies virus replicated well in human monocytic U937 and THP-1 cells and murine macrophage IC-21 cells, as well as primary cultures of murine macrophages. Minimal replication was detected in murine monocytic WEHI-3BD- and PU5-1R cells, and ERA virus did not replicate in murine monocytic P388D1 or J774A.1 cells. A tissue culture-adapted SRV of bat origin also replicated in IC-21 and U937 cells. Non-tissue culture-adapted SRV isolated from different animal species, particularly bats, replicated minimally in U937, THP-1, IC-21 cells and primary murine bone marrow macrophages. To determine whether rabies virus replication is dependent upon the state of differentiation of the macrophage-like cell, human promyelocytic HL-60 cells were differentiated with 12-O-tetradecanoylphorbol-13-acetate (TPA). ERA rabies virus replicated in the differentiated HL-60 cells but not in undifferentiated HL-60 cells. Persistent infections were established in macrophage-like U937 cells with ERA rabies virus and SRV, and infectious SRV was isolated from adherent bone marrow cells of mice that had been infected 96 days previously. Virus harvested from persistently infected U937 cells and the adherent bone marrow cells had specifically adapted to each cell. This specificity was shown by the inability of the viruses to infect macrophages other than U937 cells and primary bone marrow macrophages, respectively. Virus titers of the persistently infected U937 cells fluctuated with extended cell passage. After 30 passages, virus released from the cells had lost virulence as shown by its inability to kill intracranially inoculated mice. However, the avirulent virus released from the

  7. Immunoactivating peptide p4 augments alveolar macrophage phagocytosis in two diverse human populations.

    PubMed

    Bangert, Mathieu; Wright, Adam K; Rylance, Jamie; Kelly, Matthew J; Wright, Angela D; Carlone, George M; Sampson, Jacquelyn S; Rajam, Gowrisankar; Ades, Edwin W; Kadioglu, Aras; Gordon, Stephen B

    2013-09-01

    New treatment strategies are urgently needed to overcome early mortality in acute bacterial infections. Previous studies have shown that administration of a novel immunoactivating peptide (P4) alongside passive immunotherapy prevents the onset of septicemia and rescues mice from lethal invasive disease models of pneumococcal pneumonia and sepsis. In this study, using two diverse populations of adult volunteers, we determined whether P4 treatment of human alveolar macrophages would upregulate phagocytic killing of Streptococcus pneumoniae ex vivo. We also measured macrophage intracellular oxidation, cytokine secretion, and surface marker expression following stimulation. Peptide treatment showed enhanced bacterial killing in the absence of nonspecific inflammation, consistent with therapeutic potential. This is the first demonstration of P4 efficacy on ex vivo-derived human lung cells.

  8. Interactions between calf alveolar macrophages and parainfluenza-3 virus.

    PubMed

    Probert, M; Stott, E J; Thomas, L H

    1977-02-01

    Cells washed from the lungs of freshly killed calves (lung wash cells; LWC) were cytotoxic for calf kidney (CK) target cells infected with parainfluenzavirus type 3 (Pi-3) when assayed by chromium release. LWC collected from 25 calves, including two gnotobiotic animals that had not previously been infected with Pi-3, were all cytotoxic, giving a specific chromium release between 11 and 50%. Cytotoxicity was detected at ratios of LWC to target cell as low as 5:1. The cytotoxic reaction required viable LWC, was inhibited by Pi-3 antiserum, and was not the result of virus-induced damage to the target cells. The cytotoxic cells in the LWC population were identified as alveolar macrophages from observations on glass adherence, phagocytic activity, killing by silica and fine-structural appearance. When LWC were added to CK cells or organ cultures of bovine trachea infected with Pi-3, the yield of virus was reduced for the first 2 to 3 days. However, subsequently, Pi-3 virus replicated in the LWC. Infection of LWC with Pi-3 virus reduced their cytotoxic activity. The significance of these interactions between alveolar macrophages and Pi-3 virus is discussed.

  9. Killed oral cholera vaccines: history, development and implementation challenges.

    PubMed

    Lopez, Anna Lena; Gonzales, Maria Liza Antoinette; Aldaba, Josephine G; Nair, G Balakrish

    2014-09-01

    Cholera is still a major global health problem, affecting mainly people living in unsanitary conditions and who are at risk for outbreaks of cholera. During the past decade, outbreaks are increasingly reported from more countries. From the early killed oral cholera vaccine, rapid improvements in vaccine development occurred as a result of a better understanding of the epidemiology of the disease, pathogenesis of cholera infection and immunity. The newer-generation oral killed cholera vaccines have been shown to be safe and effective in field trials conducted in cholera endemic areas. Likewise, they have been shown to be protective when used during outbreak settings. Aside from providing direct protection to vaccinated individuals, recent studies have demonstrated that these killed oral vaccines also confer indirect protection through herd immunity. Although new-generation oral cholera vaccines should not be considered in isolation from other preventive approaches in countries where they are most needed, especially improved water quality and sanitation, these vaccines serve as immediately available public health tools for preventing further morbidity and mortality from cholera. However, despite its availability for more than two decades, use of these vaccines has not been optimized. Although there are limitations of the currently available oral cholera vaccines, recent data show that the vaccines are safe, feasible to use even in difficult circumstances and able to provide protection in various settings. Clear identification of the areas and target population groups who will benefit from the use of the cholera vaccines will be required and strategies to facilitate accessibility and usage of these vaccines in these areas and population groups will need to be developed.

  10. Importance of antibody and complement for oxidative burst and killing of invasive nontyphoidal Salmonella by blood cells in Africans.

    PubMed

    Gondwe, Esther N; Molyneux, Malcolm E; Goodall, Margaret; Graham, Stephen M; Mastroeni, Pietro; Drayson, Mark T; MacLennan, Calman A

    2010-02-16

    Bacteremia caused by nontyphoidal strains of Salmonella is endemic among African children. Case-fatality rates are high and antibiotic resistance increasing, but no vaccine is currently available. T cells are important for clearance of Salmonella infection within macrophages, but in Africa, invasive Salmonella disease usually manifests in the blood and affects children between 4 months and 2 y of age, when anti-Salmonella antibody is absent. We have previously found a role for complement-fixing bactericidal antibody in protecting these children. Here we show that opsonic activity of antibody and complement is required for oxidative burst and killing of Salmonella by blood cells in Africans. Induction of neutrophil oxidative burst correlated with anti-Salmonella IgG and IgM titers and C3 deposition on bacteria and was significantly lower in African children younger than 2 y compared with older children. Preopsonizing Salmonella with immune serum overcame this deficit, indicating a requirement for antibody and/or complement. Using different opsonization procedures, both antibody and complement were found to be necessary for optimal oxidative burst, phagocytosis and killing of nontyphoidal Salmonella by peripheral blood cells in Africans. Although most strains of African nontyphoidal Salmonella can be killed with antibody and complement alone, phagocytes in the presence of specific antibody and complement can kill strains resistant to killing by immune serum. These findings increase the likelihood that an antibody-inducing vaccine will protect against invasive nontyphoidal Salmonella disease in African children.

  11. Mass killings and detection of impacts

    NASA Technical Reports Server (NTRS)

    Mclaren, Digby J.

    1988-01-01

    Highly energetic bolide impacts occur and their flux is known. For larger bodies the energy release is greater than for any other short-term global phenomenon. Such impacts produce or release a large variety of shock induced changes including major atmospheric, sedimentologic, seismic and volcanic events. These events must necessarily leave a variety of records in the stratigraphic column, including mass killings resulting in major changes in population density and reduction or extinction of many taxonomic groups, followed by characteristic patterns of faunal and flora replacement. Of these effects, mass killings, marked by large-scale loss of biomass, are the most easily detected evidence in the field but must be manifest on a near-global scale. Such mass killings that appear to be approximately synchronous and involve disappearance of biomass at a bedding plane in many sedimentologically independent sections globally suggest a common cause and probable synchroneity. Mass killings identify an horizon which may be examined for evidence of cause. Geochemical markers may be ephemeral and absence may not be significant. There appears to be no reason why ongoing phenomena such as climate and sea-level changes are primary causes of anomolous episodic events.

  12. Can Vet Schools Teach without Killing Animals?

    ERIC Educational Resources Information Center

    Mangan, Katherine S.

    2000-01-01

    Discusses a protest by students at the University of Illinois (Urbana) College of Veterinary Medicine over the killing of animals that led to temporary curtailing of lethal animal experiments. Examines the conflict between animal rights groups and some faculty who are openly skeptical about the effectiveness of alternatives to the hands-on…

  13. Methods of killing employed by psychotic parricides.

    PubMed

    Marleau, Jacques D

    2003-10-01

    Lewis, et al. in 1998 showed that psychotic women are more likely to use a weapon than nonpsychotic women to kill their children. This study presents data concerning psychotic parricide. Analysis indicated that a higher percentage used a weapon (81% versus 36%) than psychotic filicide. Reasons for this difference are discussed. PMID:14650686

  14. Peanut Roaster Temperatures Relative to Salmonella Kill

    Technology Transfer Automated Retrieval System (TEKTRAN)

    ARS, Market Quality and Handling Research Unit, Raleigh NC 27695 In response to the limited peanut butter contamination incident of 2006/7, studies were initiated to examine the effect of various time and temperature protocols on log kill levels for Salmonella on peanuts. The objective of the work ...

  15. School Shootings; Standards Kill Students and Society

    ERIC Educational Resources Information Center

    Angert, Betsy L.

    2008-01-01

    School shootings have been in the news of late. People ponder what occurs in classrooms today. Why would a young person wish to take a life? Within educational institutions, the killings are a concern. In our dire attempt to teach the children and ensure student success, it seems many of our offspring are lost. Some students feel separate from…

  16. Integrating Poetry and "To Kill a Mockingbird."

    ERIC Educational Resources Information Center

    Jolley, Susan Arpajian

    2002-01-01

    Outlines a method of teaching "To Kill a Mockingbird" along with the study of poetry. Notes that this method allows students to consider the themes of courage and developing compassion. Concludes that teaching such a multigenre unit allows students to look for connections among fact and fiction, the past and present, their own lives and…

  17. Killing Hitler: A Writer's Journey and Angst.

    ERIC Educational Resources Information Center

    Thaler, Paul

    2002-01-01

    Describes the author's experiences in preparing a talk that "evokes the specter" of Adolf Hitler and in writing an historical account of a British plot to kill Hitler. Address the question of why the British allowed him to live that final year of the war. Muses on why scholars write, and the impact of violence and terrorism. (SG)

  18. Nonlinear symmetries on spaces admitting Killing tensors

    NASA Astrophysics Data System (ADS)

    Visinescu, Mihai

    2010-04-01

    Nonlinear symmetries corresponding to Killing tensors are investigated. The intimate relation between Killing-Yano tensors and non-standard supersymmetries is pointed out. The gravitational anomalies are absent if the hidden symmetry is associated with a Killing-Yano tensor. In the case of the nonlinear symmetries the dynamical algebras of the Dirac-type operators is more involved and could be organized as infinite dimensional algebras or superalgebras. The general results are applied to some concrete spaces involved in theories of modern physics. As a first example it is considered the 4-dimensional Euclidean Taub-NUT space and its generalizations introduced by Iwai and Katayama. One presents the infinite dimensional superalgebra of Dirac type operators on Taub-NUT space that could be seen as a graded loop superalgebra of the Kac-Moody type. The axial anomaly, interpreted as the index of the Dirac operator, is computed for the generalized Taub-NUT metrics. Finally the existence of the conformal Killing-Yano tensors is investigated for some spaces with mixed Sasakian structures.

  19. Semiautomatic quantitation of macrophages in human renal biopsy specimens in proteinuric states.

    PubMed Central

    Furness, P N; Rogers-Wheatley, L; Harris, K P

    1997-01-01

    AIMS: To develop and validate a rapid and economical semiautomated approach to the measurement of immunostainable tissue components which is applicable to routine diagnostic practice. To apply this approach to the measurement of macrophages in renal biopsy specimens in nephrotic states, as protein in the renal tubules may induce macrophage infiltration, and the morphology of macrophages in tissue sections does not lend itself to cell counting. METHODS: Macrophages were identified by immunostaining with a pan-macrophage marker, followed by digital image capture and analysis using a macro procedure written for the freeware image analysis program NIH-Image. RESULTS: The method was rapid, robust and accurate to within the limits imposed by sampling error inherent in the use of small needle biopsy specimens. Very few macrophages are found in normal kidney (mean volume fraction (+/- 95% confidence limits) 0.04% (0.02%)) but infiltration of macrophages was detected in minimal change nephropathy (0.29% (0.12%)) and in membranous glomerulonephritis (0.42% (0.11%)). A statistically significant correlation was found between macrophage volume fraction and weight of proteinuria in minimal change nephropathy but not in membranous glomerulonephritis. Correlations were found in both diseases between macrophage volume fraction and serum creatinine at time of biopsy. CONCLUSIONS: The equipment is inexpensive and measurement takes less than one minute per biopsy specimen. The results indicate that macrophage infiltration is part of the pathological process in minimal change nephropathy and membranous glomerulonephritis. The correlation with creatinine at time of biopsy suggests that renal impairment in minimal change nephropathy may result from infiltration by immunologically active cells and not merely from haemodynamic changes in nephrons. However, the correlation is not close, indicating that the relation between macrophage infiltration and disease severity is not a simple one

  20. Contagion in Mass Killings and School Shootings

    PubMed Central

    Towers, Sherry; Gomez-Lievano, Andres; Khan, Maryam; Mubayi, Anuj; Castillo-Chavez, Carlos

    2015-01-01

    Background Several past studies have found that media reports of suicides and homicides appear to subsequently increase the incidence of similar events in the community, apparently due to the coverage planting the seeds of ideation in at-risk individuals to commit similar acts. Methods Here we explore whether or not contagion is evident in more high-profile incidents, such as school shootings and mass killings (incidents with four or more people killed). We fit a contagion model to recent data sets related to such incidents in the US, with terms that take into account the fact that a school shooting or mass murder may temporarily increase the probability of a similar event in the immediate future, by assuming an exponential decay in contagiousness after an event. Conclusions We find significant evidence that mass killings involving firearms are incented by similar events in the immediate past. On average, this temporary increase in probability lasts 13 days, and each incident incites at least 0.30 new incidents (p = 0.0015). We also find significant evidence of contagion in school shootings, for which an incident is contagious for an average of 13 days, and incites an average of at least 0.22 new incidents (p = 0.0001). All p-values are assessed based on a likelihood ratio test comparing the likelihood of a contagion model to that of a null model with no contagion. On average, mass killings involving firearms occur approximately every two weeks in the US, while school shootings occur on average monthly. We find that state prevalence of firearm ownership is significantly associated with the state incidence of mass killings with firearms, school shootings, and mass shootings. PMID:26135941

  1. Reciprocal interactions between endothelial cells and macrophages in angiogenic vascular niches

    SciTech Connect

    Baer, Caroline; Squadrito, Mario Leonardo; Iruela-Arispe, M. Luisa; De Palma, Michele

    2013-07-01

    The ability of macrophages to promote vascular growth has been associated with the secretion and local delivery of classic proangiogenic factors (e.g., VEGF-A and proteases). More recently, a series of studies have also revealed that physical contact of macrophages with growing blood vessels coordinates vascular fusion of emerging sprouts. Interestingly, the interactions between macrophages and vascular endothelial cells (ECs) appear to be bidirectional, such that activated ECs also support the expansion and differentiation of proangiogenic macrophages from myeloid progenitors. Here, we discuss recent findings suggesting that dynamic angiogenic vascular niches might also exist in vivo, e.g. in tumors, where sprouting blood vessels and immature myeloid cells like monocytes engage in heterotypic interactions that are required for angiogenesis. Finally, we provide an account of emerging mechanisms of cell-to-cell communication that rely on secreted microvesicles, such as exosomes, which can offer a vehicle for the rapid exchange of molecules and genetic information between macrophages and ECs engaged in angiogenesis. -- Highlights: • Macrophages promote angiogenesis by secreting proangiogenic factors. • Macrophages modulate angiogenesis via cell-to-cell contacts with endothelial cells. • Endothelial cells promote the differentiation of proangiogenic macrophages. • Macrophages and endothelial cells may cooperate to form angiogenic vascular niches.

  2. Antisense phosphorothioate oligonucleotides: selective killing of the intracellular parasite Leishmania amazonensis.

    PubMed Central

    Ramazeilles, C; Mishra, R K; Moreau, S; Pascolo, E; Toulmé, J J

    1994-01-01

    We targeted the mini-exon sequence, present at the 5' end of every mRNA of the protozoan parasite Leishmania amazonensis, by phosphorothioate oligonucleotides. A complementary 16-mer (16PS) was able to kill amastigotes--the intracellular stage of the parasite--in murine macrophages in culture. After 24 hr of incubation with 10 microM 16PS, about 30% infected macrophages were cured. The oligomer 16PS acted through antisense hybridization in a sequence-dependent way; no effect on parasites was observed with noncomplementary phosphorothioate oligonucleotides. The antisense oligonucleotide 16PS was a selective killer of the protozoans without any detrimental effect to the host macrophage. Using 16PS linked to a palmitate chain, which enabled it to complex with low density lipoproteins, improved the leishmanicidal efficiency on intracellular amastigotes, probably due to increased endocytosis. Phosphorothioate oligonucleotides complementary to the intron part of the mini-exon pre-RNA were also effective, suggesting that antisense oligomers could prevent trans-splicing in these parasites. Images PMID:8058724

  3. Inhibition of nuclear factor-kappa B activation decreases survival of Mycobacterium tuberculosis in human macrophages.

    PubMed

    Bai, Xiyuan; Feldman, Nicole E; Chmura, Kathryn; Ovrutsky, Alida R; Su, Wen-Lin; Griffin, Laura; Pyeon, Dohun; McGibney, Mischa T; Strand, Matthew J; Numata, Mari; Murakami, Seiji; Gaido, Loretta; Honda, Jennifer R; Kinney, William H; Oberley-Deegan, Rebecca E; Voelker, Dennis R; Ordway, Diane J; Chan, Edward D

    2013-01-01

    Nuclear factor-kappa B (NFκB) is a ubiquitous transcription factor that mediates pro-inflammatory responses required for host control of many microbial pathogens; on the other hand, NFκB has been implicated in the pathogenesis of other inflammatory and infectious diseases. Mice with genetic disruption of the p50 subunit of NFκB are more likely to succumb to Mycobacterium tuberculosis (MTB). However, the role of NFκB in host defense in humans is not fully understood. We sought to examine the role of NFκB activation in the immune response of human macrophages to MTB. Targeted pharmacologic inhibition of NFκB activation using BAY 11-7082 (BAY, an inhibitor of IκBα kinase) or an adenovirus construct with a dominant-negative IκBα significantly decreased the number of viable intracellular mycobacteria recovered from THP-1 macrophages four and eight days after infection. The results with BAY were confirmed in primary human monocyte-derived macrophages and alveolar macrophages. NFκB inhibition was associated with increased macrophage apoptosis and autophagy, which are well-established killing mechanisms of intracellular MTB. Inhibition of the executioner protease caspase-3 or of the autophagic pathway significantly abrogated the effects of BAY. We conclude that NFκB inhibition decreases viability of intracellular MTB in human macrophages via induction of apoptosis and autophagy.

  4. Inhibition of nuclear factor-kappa B activation decreases survival of Mycobacterium tuberculosis in human macrophages.

    PubMed

    Bai, Xiyuan; Feldman, Nicole E; Chmura, Kathryn; Ovrutsky, Alida R; Su, Wen-Lin; Griffin, Laura; Pyeon, Dohun; McGibney, Mischa T; Strand, Matthew J; Numata, Mari; Murakami, Seiji; Gaido, Loretta; Honda, Jennifer R; Kinney, William H; Oberley-Deegan, Rebecca E; Voelker, Dennis R; Ordway, Diane J; Chan, Edward D

    2013-01-01

    Nuclear factor-kappa B (NFκB) is a ubiquitous transcription factor that mediates pro-inflammatory responses required for host control of many microbial pathogens; on the other hand, NFκB has been implicated in the pathogenesis of other inflammatory and infectious diseases. Mice with genetic disruption of the p50 subunit of NFκB are more likely to succumb to Mycobacterium tuberculosis (MTB). However, the role of NFκB in host defense in humans is not fully understood. We sought to examine the role of NFκB activation in the immune response of human macrophages to MTB. Targeted pharmacologic inhibition of NFκB activation using BAY 11-7082 (BAY, an inhibitor of IκBα kinase) or an adenovirus construct with a dominant-negative IκBα significantly decreased the number of viable intracellular mycobacteria recovered from THP-1 macrophages four and eight days after infection. The results with BAY were confirmed in primary human monocyte-derived macrophages and alveolar macrophages. NFκB inhibition was associated with increased macrophage apoptosis and autophagy, which are well-established killing mechanisms of intracellular MTB. Inhibition of the executioner protease caspase-3 or of the autophagic pathway significantly abrogated the effects of BAY. We conclude that NFκB inhibition decreases viability of intracellular MTB in human macrophages via induction of apoptosis and autophagy. PMID:23634218

  5. Adoptive transfer of macrophages from adult mice reduces mortality in mice infected with human enterovirus 71.

    PubMed

    Liu, Jiangning; Li, Xiaoying; Fan, Xiaoxu; Ma, Chunmei; Qin, Chuan; Zhang, Lianfeng

    2013-02-01

    Human enterovirus 71 (EV71) causes hand, foot and mouth disease in children under 6 years of age, and the neurological complications of this virus can lead to death. Until now, no vaccines or drugs have been available for the clinical control of this epidemic. Macrophages can engulf pathogens and mediate a series of host immune responses that play a role in the defence against infectious diseases. Using immunohistochemistry, we observed the localizations of virus in muscle tissues of EV71-infected mice. The macrophages isolated from the adult mice could kill the virus gradually in vitro, as shown using quantitative real-time PCR (qRT-PCR) and virus titration. Co-localisation of lysosomes and virus within macrophages suggested that the lysosomes were possibly responsible for the phagocytosis of EV71. Activation of the macrophages in the peritoneal cavity of mice four days pre-infection reduced the mortality of mice upon lethal EV71 infection. The adoptive transfer of macrophages from adult mice inhibited virus replication in the muscle tissues of infected mice, and this was followed by a relief of symptoms and a significant reduction of mortality, which suggested that the adoptive transfer of macrophages from adult humans represents a potential strategy to treat EV71-infected patients.

  6. Enhancing effect of oxygen radical scavengers on murine macrophage anticryptococcal activity through production of nitric oxide

    PubMed Central

    TOHYAMA, M.; KAWAKAMI, K.; FUTENMA, M.; SAITO, A.

    1996-01-01

    We examined the roles of reactive nitrogen intermediates (RNI) and reactive oxygen intermediates (ROI) in interferon-gamma (IFN-γ)-induced cryptococcostatic activity of murine peritoneal macrophages using NG-monomethyl-L-arginine (L-NMMA), a competitive inhibitor of RNI synthesis, and superoxide dismutase (SOD) and catalase, oxygen radical scavengers. IFN-γ-activated macrophages produced nitric oxide (NO) in a dose-dependent manner, as measured by increased nitrite concentration in the culture supernatant. IFN-γ also enhanced the suppressive effect on cryptococcal growth in a similar dose-dependent manner. The induction of killing activity and NO production by an optimal dose of IFN-γ (100 U/ml) was virtually suppressed by 500 μM L-NMMA. These results confirmed the importance of the RNI-mediated effector mechanism in anticryptococcal activity of macrophages. SOD and catalase significantly enhanced the cryptococcostatic activity of macrophages induced by a suboptimal dose of IFN-γ (20 U/ml). The augmenting effect of these reagents was mediated by NO, since they potentiated the production of NO by macrophages and their effects were totally blocked by L-NMMA. Our results indicate that the IFN-γ-induced anticryptococcal activity of macrophages is dependent mostly on RNI, and suggest that the ROI system down-regulates the effector mechanism for cryptococcostasis by suppressing the RNI system. PMID:8608643

  7. Inhibition of Nuclear Factor-Kappa B Activation Decreases Survival of Mycobacterium tuberculosis in Human Macrophages

    PubMed Central

    Chmura, Kathryn; Ovrutsky, Alida R.; Su, Wen-Lin; Griffin, Laura; Pyeon, Dohun; McGibney, Mischa T.; Strand, Matthew J.; Numata, Mari; Murakami, Seiji; Gaido, Loretta; Honda, Jennifer R.; Kinney, William H.; Oberley-Deegan, Rebecca E.; Voelker, Dennis R.; Ordway, Diane J.; Chan, Edward D.

    2013-01-01

    Nuclear factor-kappa B (NFκB) is a ubiquitous transcription factor that mediates pro-inflammatory responses required for host control of many microbial pathogens; on the other hand, NFκB has been implicated in the pathogenesis of other inflammatory and infectious diseases. Mice with genetic disruption of the p50 subunit of NFκB are more likely to succumb to Mycobacterium tuberculosis (MTB). However, the role of NFκB in host defense in humans is not fully understood. We sought to examine the role of NFκB activation in the immune response of human macrophages to MTB. Targeted pharmacologic inhibition of NFκB activation using BAY 11-7082 (BAY, an inhibitor of IκBα kinase) or an adenovirus construct with a dominant-negative IκBα significantly decreased the number of viable intracellular mycobacteria recovered from THP-1 macrophages four and eight days after infection. The results with BAY were confirmed in primary human monocyte-derived macrophages and alveolar macrophages. NFκB inhibition was associated with increased macrophage apoptosis and autophagy, which are well-established killing mechanisms of intracellular MTB. Inhibition of the executioner protease caspase-3 or of the autophagic pathway significantly abrogated the effects of BAY. We conclude that NFκB inhibition decreases viability of intracellular MTB in human macrophages via induction of apoptosis and autophagy. PMID:23634218

  8. Lipocalin-2 ensures host defense against Salmonella Typhimurium by controlling macrophage iron homeostasis and immune response

    PubMed Central

    Nairz, Manfred; Schroll, Andrea; Haschka, David; Dichtl, Stefanie; Sonnweber, Thomas; Theurl, Igor; Theurl, Milan; Lindner, Ewald; Demetz, Egon; Aβhoff, Malte; Bellmann-Weiler, Rosa; Müller, Raphael; Gerner, Romana R.; Moschen, Alexander R.; Baumgartner, Nadja; Moser, Patrizia L.; Talasz, Heribert; Tilg, Herbert; Fang, Ferric C.; Weiss, Günter

    2015-01-01

    Lipocalin-2 (Lcn2) is an innate immune peptide with pleiotropic effects. Lcn2 binds iron-laden bacterial siderophores, chemo-attracts neutrophils and has immunomodulatory and apoptosis-regulating effects. In this study we show that upon infection with Salmonella enterica serovar Typhimurium, Lcn2 promotes iron export from Salmonella-infected macrophages, which reduces cellular iron content and enhances the generation of pro-inflammatory cytokines. Lcn2 represses IL-10 production while augmenting Nos2, TNF-α and IL-6 expression. Lcn2-/- macrophages have elevated IL-10 levels as a consequence of increased iron content. The crucial role of Lcn-2/IL-10 interactions was further demonstrated by the greater ability of Lcn2-/- IL-10-/- macrophages and mice to control intracellular Salmonella proliferation in comparison to Lcn2-/- counterparts. Over-expression of the iron exporter ferroportin-1 in Lcn2-/- macrophages represses IL-10 and restores TNF-α and IL-6 production to the levels found in wild-type macrophages, so that killing and clearance of intracellular Salmonella is promoted. Our observations suggest that Lcn2 promotes host resistance to Salmonella Typhimurium infection by binding bacterial siderophores and suppressing IL-10 production, and that both functions are linked to its ability to shuttle iron from macrophages. PMID:26332507

  9. Phorbal esters and calcium ionophore can prime murine peritoneal macrophages for tumor cell destruction

    SciTech Connect

    Somers, S.D.; Weiel, J.E.; Hamilton, T.A.; Adams, D.O.

    1986-06-01

    Murine macrophages from sites of inflammation develop toward tumoricidal competence by exposure to a macrophage-activating factor such as interferon-..gamma.. (IFN-..gamma..). To explore the biochemical transductional events initiated by IFN-..gamma.., peritoneal macrophages from C57BL/6J mice elicited by various sterile irritants were treated in vitro with two pharmacologic agents that mimic the action of certain second messengers. Phorbol myristate acetate (PMA) and the ionophore A23187 cooperatively reproduced the ability of IFN-..gamma.. to prime macrophages for tumoricidal function. Neither agent alone was able to prime macrophages. The two agents acted on the macrophages, and target susceptibility to kill was not altered by PMA and A23187. Only active phorbol esters, which are known to bind and stimulate protein kinase C, were able to cooperate with A23187 to induce priming. A cell-permeable synthetic diacylglycerol (sn-1,2-dioctanoyl glycerol) could also prime for cytolysis. In the presence of PMA, A23187, and EGTA, addition of Ca/sup + +/ was sufficient for priming, whereas the addition of Mg/sup + +/ was much less efficient. Priming by IFN-..gamma.., however, was not blocked by EGTA. Efflux of /sup 45/Ca/sup + +/ from preloaded cells was significantly increased by A23187 and by IFN-..gamma... Quin-2/AM, an intracellular chelator of Ca/sup + +/, blocked priming by IFN-..gamma...

  10. Macrophage Infiltration and Alternative Activation during Wound Healing Promote MEK1-Induced Skin Carcinogenesis.

    PubMed

    Weber, Christine; Telerman, Stephanie B; Reimer, Andreas S; Sequeira, Ines; Liakath-Ali, Kifayathullah; Arwert, Esther N; Watt, Fiona M

    2016-02-15

    Macrophages are essential for the progression and maintenance of many cancers, but their role during the earliest stages of tumor formation is unclear. To test this, we used a previously described transgenic mouse model of wound-induced skin tumorigenesis, in which expression of constitutively active MEK1 in differentiating epidermal cells results in chronic inflammation (InvEE mice). Upon wounding, the number of epidermal and dermal monocytes and macrophages increased in wild-type and InvEE skin, but the increase was greater, more rapid, and more sustained in InvEE skin. Macrophage ablation reduced tumor incidence. Furthermore, bioluminescent imaging in live mice to monitor macrophage flux at wound sites revealed that macrophage accumulation was predictive of tumor formation; wounds with the greatest number of macrophages at day 5 went on to develop tumors. Gene expression profiling of flow-sorted monocytes, macrophages, and T cells from InvEE and wild-type skin showed that as wound healing progressed, InvEE macrophages altered their phenotype. Throughout wound healing and after wound closure, InvEE macrophages demonstrated sustained upregulation of several markers implicated in alternative macrophage activation including arginase-1 (ARG1) and mannose receptor (CD206). Notably, inhibition of ARG1 activity significantly reduced tumor formation and epidermal proliferation in vivo, whereas addition of L-arginase to cultured keratinocytes stimulated proliferation. We conclude that macrophages play a key role in early, inflammation-mediated skin tumorigenesis, with mechanistic evidence suggesting that ARG1 secretion drives tumor development by stimulating epidermal cell proliferation. These findings highlight the importance of cancer immunotherapies aiming to polarize tumor-associated macrophages toward an antitumor phenotype. PMID:26754935

  11. Macrophage function in murine allogeneic bone marrow radiation chimeras in the early phase after transplantation

    SciTech Connect

    Roesler, J.; Baccarini, M.; Vogt, B.; Lohmann-Matthes, M.L. )

    1989-08-01

    We tested several of the functions of macrophages (M phi) in the early phase after allogeneic bone marrow transfer to get information about this important aspect of the nonspecific immune system in the T-cell-deficient recipient. On days 3-5 after transfer, the number of M phi was reduced in the spleen, liver, lungs, and peritoneal cavity (Pe). The phagocytosis of sheep red blood cells (SRBC) by these M phi was normal or even enhanced, as in the case of Pe-M phi. Already on days 8-12 after transfer, the number of M phi in spleen and liver exceeded that of controls, whereas the number was still reduced in lungs and Pe. We examined their ability to kill P815 tumor cells, to produce tumor necrosis factor-alpha (TNF alpha), to phagocytose SRBC, to produce reactive oxygen intermediates (ROI) in vitro and to kill Listeria monocytogenes in vivo. Most functions were normal and often even enhanced, depending on the organ origin, but the ability of Pe-M phi to produce ROI was reduced. Proliferative response to macrophage colony-stimulating factor (M-CSF) and killing of YAC-1 tumor cells revealed a high frequency of macrophage precursor cells in the spleen and liver and a high natural killer (NK) activity in the liver. Altogether, enhanced nonspecific immune function, especially preactivated M phi, may enable chimeras to survive attacks by opportunistic pathogens.

  12. Prostaglandin E2 Regulation of Macrophage Innate Immunity.

    PubMed

    Kimmel, Danielle W; Rogers, Lisa M; Aronoff, David M; Cliffel, David E

    2016-01-19

    Globally, maternal and fetal health is greatly impacted by extraplacental inflammation. Group B Streptococcus (GBS), a leading cause of chorioamnionitis, is thought to take advantage of the uterine environment during pregnancy in order to cause inflammation and infection. In this study, we demonstrate the metabolic changes of murine macrophages caused by GBS exposure. GBS alone prompted a delayed increase in lactate production, highlighting its ability to redirect macrophage metabolism from aerobic to anaerobic respiration. This production of lactate is thought to aid in the development and propagation of GBS throughout the surrounding tissue. Additionally, this study shows that PGE2 priming was able to exacerbate lactate production, shown by the rapid and substantial lactate increases seen upon GBS exposure. These data provide a novel model to study the role of GBS exposure to macrophages with and without PGE2 priming. PMID:26656203

  13. Monocytes, Macrophages and Other Inflammatory Mediators of Abdominal Aortic Aneurysm.

    PubMed

    Potteaux, Stephane; Tedgui, Alain

    2015-01-01

    Macrophages early invade the forming abdominal aortic aneurysm (AAA) and greatly contribute to its pathogenesis. Recent findings have shown that Ly-6C(high) and Ly-6C(low) monocytes are rapidly mobilized from the splenic reservoir in response to angiotensin II infusion and sequentially infiltrate the abdominal aorta. The first wave of Ly-6C(high) monocytes prevails in the aorta and promotes the accumulation of inflammatory macrophages, which most likely cause irreversible changes in the abdominal aorta. In this review, we discuss the current knowledge on the cellular mechanisms that initiate AAA in mice. We particularly focus on the role of monocyte and macrophage subsets during the early steps of the aneurysmal process. PMID:26306839

  14. Metabolic Reprograming in Macrophage Polarization

    PubMed Central

    Galván-Peña, Silvia; O’Neill, Luke A. J.

    2014-01-01

    Studying the metabolism of immune cells in recent years has emphasized the tight link existing between the metabolic state and the phenotype of these cells. Macrophages in particular are a good example of this phenomenon. Whether the macrophage obtains its energy through glycolysis or through oxidative metabolism can give rise to different phenotypes. Classically activated or M1 macrophages are key players of the first line of defense against bacterial infections and are known to obtain energy through glycolysis. Alternatively activated or M2 macrophages on the other hand are involved in tissue repair and wound healing and use oxidative metabolism to fuel their longer-term functions. Metabolic intermediates, however, are not just a source of energy but can be directly implicated in a particular macrophage phenotype. In M1 macrophages, the Krebs cycle intermediate succinate regulates HIF1α, which is responsible for driving the sustained production of the pro-inflammatory cytokine IL1β. In M2 macrophages, the sedoheptulose kinase carbohydrate kinase-like protein is critical for regulating the pentose phosphate pathway. The potential to target these events and impact on disease is an exciting prospect. PMID:25228902

  15. Sabretoothed carnivores and the killing of large prey.

    PubMed

    Andersson, Ki; Norman, David; Werdelin, Lars

    2011-01-01

    Sabre-like canines clearly have the potential to inflict grievous wounds leading to massive blood loss and rapid death. Hypotheses concerning sabretooth killing modes include attack to soft parts such as the belly or throat, where biting deep is essential to generate strikes reaching major blood vessels. Sabretoothed carnivorans are widely interpreted as hunters of larger and more powerful prey than that of their present-day nonsabretoothed relatives. However, the precise functional advantage of the sabretooth bite, particularly in relation to prey size, is unknown. Here, we present a new point-to-point bite model and show that, for sabretooths, depth of the killing bite decreases dramatically with increasing prey size. The extended gape of sabretooths only results in considerable increase in bite depth when biting into prey with a radius of less than ∼10 cm. For sabretooths, this size-reversed functional advantage suggests predation on species within a similar size range to those attacked by present-day carnivorans, rather than "megaherbivores" as previously believed. The development of the sabretooth condition appears to represent a shift in function and killing behaviour, rather than one in predator-prey relations. Furthermore, our results demonstrate how sabretoothed carnivorans are likely to have evolved along a functionally continuous trajectory: beginning as an extension of a jaw-powered killing bite, as adopted by present-day pantherine cats, followed by neck-powered biting and thereafter shifting to neck-powered shear-biting. We anticipate this new insight to be a starting point for detailed study of the evolution of pathways that encompass extreme specialisation, for example, understanding how neck-powered biting shifts into shear-biting and its significance for predator-prey interactions. We also expect that our model for point-to-point biting and bite depth estimations will yield new insights into the behaviours of a broad range of extinct predators

  16. Chlamydia muridarum Infection of Macrophages Elicits Bactericidal Nitric Oxide Production via Reactive Oxygen Species and Cathepsin B

    PubMed Central

    Rajaram, Krithika

    2015-01-01

    The ability of certain species of Chlamydia to inhibit the biogenesis of phagolysosomes permits their survival and replication within macrophages. The survival of macrophage-adapted chlamydiae correlates with the multiplicity of infection (MOI), and optimal chlamydial growth occurs in macrophages infected at an MOI of ≤1. In this study, we examined the replicative capacity of Chlamydia muridarum in the RAW 264.7 murine macrophage cell line at different MOIs. C. muridarum productively infected these macrophages at low MOIs but yielded few viable elementary bodies (EBs) when macrophages were infected at a moderate (10) or high (100) MOI. While high MOIs caused cytotoxicity and irreversible host cell death, macrophages infected at a moderate MOI did not show signs of cytotoxicity until late in the infectious cycle. Inhibition of host protein synthesis rescued C. muridarum in macrophages infected at a moderate MOI, implying that chlamydial growth was blocked by activated defense mechanisms. Conditioned medium from these macrophages was antichlamydial and contained elevated levels of interleukin 1β (IL-1β), IL-6, IL-10, and beta interferon (IFN-β). Macrophage activation depended on Toll-like receptor 2 (TLR2) signaling, and cytokine production required live, transcriptionally active chlamydiae. A hydroxyl radical scavenger and inhibitors of inducible nitric oxide synthase (iNOS) and cathepsin B also reversed chlamydial killing. High levels of reactive oxygen species (ROS) led to an increase in cathepsin B activity, and pharmacological inhibition of ROS and cathepsin B reduced iNOS expression. Our data demonstrate that MOI-dependent TLR2 activation of macrophages results in iNOS induction via a novel ROS- and cathepsin-dependent mechanism to facilitate C. muridarum clearance. PMID:26015483

  17. Phagocytic receptors on macrophages distinguish between different Sporothrix schenckii morphotypes.

    PubMed

    Guzman-Beltran, Silvia; Perez-Torres, Armando; Coronel-Cruz, Cristina; Torres-Guerrero, Haydee

    2012-10-01

    Sporothrix schenckii is a human pathogen that causes sporotrichosis, a cutaneous subacute or chronic mycosis. Little is known about the innate immune response and the receptors involved in host recognition and phagocytosis of S. schenckii. Here, we demonstrate that optimal phagocytosis of conidia and yeast is dependent on preimmune human serum opsonisation. THP-1 macrophages efficiently ingested opsonised conidia. Competition with D-mannose, methyl α-D-mannopyranoside, D-fucose, and N-acetyl glucosamine blocked this process, suggesting the involvement of the mannose receptor in binding and phagocytosis of opsonised conidia. Release of TNF-α was not stimulated by opsonised or non-opsonised conidia, although reactive oxygen species (ROS) were produced, resulting in the killing of conidia by THP-1 macrophages. Heat inactivation of the serum did not affect conidia internalization, which was markedly decreased for yeast cells, suggesting the role of complement components in yeast uptake. Conversely, release of TNF-α and production of ROS were induced by opsonised and non-opsonised yeast. These data demonstrate that THP-1 macrophages respond to opsonised conidia and yeast through different phagocytic receptors, inducing a differential cellular response. Conidia induces a poor pro-inflammatory response and lower rate of ROS-induced cell death, thereby enhancing the pathogen's survival.

  18. The Endoplasmic Reticulum Stress Sensor Inositol-Requiring Enzyme 1α Augments Bacterial Killing through Sustained Oxidant Production

    PubMed Central

    Abuaita, Basel H.; Burkholder, Kristin M.; Boles, Blaise R.

    2015-01-01

    ABSTRACT Bacterial infection can trigger cellular stress programs, such as the unfolded protein response (UPR), which occurs when misfolded proteins accumulate within the endoplasmic reticulum (ER). Here, we used the human pathogen methicillin-resistant Staphylococcus aureus (MRSA) as an infection model to probe how ER stress promotes antimicrobial function. MRSA infection activated the most highly conserved unfolded protein response sensor, inositol-requiring enzyme 1α (IRE1α), which was necessary for robust bacterial killing in vitro and in vivo. The macrophage IRE1-dependent bactericidal activity required reactive oxygen species (ROS). Viable MRSA cells excluded ROS from the nascent phagosome and strongly triggered IRE1 activation, leading to sustained generation of ROS that were largely Nox2 independent. In contrast, dead MRSA showed early colocalization with ROS but was a poor activator of IRE1 and did not trigger sustained ROS generation. The global ROS stimulated by IRE1 signaling was necessary, but not sufficient, for MRSA killing, which also required the ER resident SNARE Sec22B for accumulation of ROS in the phagosomal compartment. Taken together, these results suggest that IRE1-mediated persistent ROS generation might act as a fail-safe mechanism to kill bacterial pathogens that evade the initial macrophage oxidative burst. PMID:26173697

  19. Macrophages and the Viral Dissemination Super Highway

    PubMed Central

    Klepper, Arielle; Branch, Andrea D

    2016-01-01

    Monocytes and macrophages are key components of the innate immune system yet they are often the victims of attack by infectious agents. This review examines the significance of viral infection of macrophages. The central hypothesis is that macrophage tropism enhances viral dissemination and persistence, but these changes may come at the cost of reduced replication in cells other than macrophages. PMID:26949751

  20. Beyond killing: Can we find new ways to manage infection?

    PubMed

    Vale, Pedro F; McNally, Luke; Doeschl-Wilson, Andrea; King, Kayla C; Popat, Roman; Domingo-Sananes, Maria R; Allen, Judith E; Soares, Miguel P; Kümmerli, Rolf

    2016-01-01

    The antibiotic pipeline is running dry and infectious disease remains a major threat to public health. An efficient strategy to stay ahead of rapidly adapting pathogens should include approaches that replace, complement or enhance the effect of both current and novel antimicrobial compounds. In recent years, a number of innovative approaches to manage disease without the aid of traditional antibiotics and without eliminating the pathogens directly have emerged. These include disabling pathogen virulence-factors, increasing host tissue damage control or altering the microbiota to provide colonization resistance, immune resistance or disease tolerance against pathogens. We discuss the therapeutic potential of these approaches and examine their possible consequences for pathogen evolution. To guarantee a longer half-life of these alternatives to directly killing pathogens, and to gain a full understanding of their population-level consequences, we encourage future work to incorporate evolutionary perspectives into the development of these treatments. PMID:27016341

  1. Classical swine fever: morphological and morphometrical study of pulmonary intravascular macrophages.

    PubMed

    Carrasco, L; Ruiz-Villamor, E; Gómez-Villamandos, J C; Salguero, F J; Bautista, M J; Maciá, M; Quezada, M; Jover, A

    2001-07-01

    To gain further insight into the pathogenesis of classical swine fever (CSF), the changes induced by hog cholera (HC) virus in pulmonary intravascular macrophages (PIMs) were examined. Twelve pigs were inoculated by the intramuscular route with a virulent strain of HC virus (Quillota strain) and killed in groups of three at 4, 7, 10 and 14 days post-inoculation. Immunohistochemical and ultrastructural examination revealed HC virus infection in endothelial cells, PIMs, and interstitial and alveolar macrophages. In addition to viral replication, a predominant feature was the secretory activation of PIMs, characterized by expanded rough endoplasmic reticulum and hyperplastic Golgi complexes. The results obtained suggest that macrophage activation and the subsequent release of pro-inflammatory mediators play an important role in the pathogenesis of CSF.

  2. Mechanisms of glucocorticoid induced suppression of phagocytosis in murine peritoneal macrophage cultures

    SciTech Connect

    Becker, J.L.

    1986-01-01

    Glucocorticoids suppress phagocytosis of heat killed Saccharomyces cerevisiae in macrophage cultures. In order to determine the mechanisms by which this response occurs, this investigation was initiated to examine whether the suppression of phagocytosis is mediated by a steroid induced phagocytosis inhibitory protein (PIP). Furthermore, it is postulated that these suppressive effects may be associated with alterations in macrophage phospholipid metabolism. To assess the association between phospholipid metabolism and phagocytosis, control and 1 ..mu..M dexamethasone treated macrophages were exposed to the phospholipase inhibitor bromophenacylbromide. The enzyme inhibitor suppressed phagocytosis in a time and dose dependent manner. However, supplying dexamethasone treated cultures with arachidonate did not reverse the steroid induced suppression of phagocytosis, whether the arachidonate was supplied alone or together with indomethacin and nordihydroguaiaretic acid. Control cells, prelabeled with /sup 3/H-arachidonate, exhibited an increased percentage of the radiolabeled fatty acid in neutral lipids following phagocytosis, with a corresponding decrease in the percentage associated with phosphatidylcholine.

  3. Generation, culture and flow-cytometric characterization of primary mouse macrophages.

    PubMed

    Schleicher, Ulrike; Bogdan, Christian

    2009-01-01

    Macrophages are not only host cells for many pathogens, but also fulfill several key functions in the innate and adaptive immune response, including the release of pro- and anti-inflammatory cytokines, the generation of organic and inorganic autacoids, the phagocytosis and killing of intracellular microorganisms or tumor cells, and the degradation and presentation of antigens. Several of these functions are shared by other immune cells, including dendritic cells, granulocytes, NK cells, and/or T lymphocytes. Thus, the analysis of macrophage functions in vitro using primary mouse cell populations requires standardized methods for the generation and culture of macrophages that guarantee high cell purity as well as the absence of stimulatory microbial contaminants. This chapter presents methodology to achieve these aims.

  4. Phosphoethanolamine Modification of Neisseria gonorrhoeae Lipid A Reduces Autophagy Flux in Macrophages.

    PubMed

    Zughaier, Susu M; Kandler, Justin L; Balthazar, Jacqueline T; Shafer, William M

    2015-01-01

    Autophagy, an ancient homeostasis mechanism for macromolecule degradation, performs an important role in host defense by facilitating pathogen elimination. To counteract this host defense strategy, bacterial pathogens have evolved a variety of mechanisms to avoid or otherwise dysregulate autophagy by phagocytic cells so as to enhance their survival during infection. Neisseria gonorrhoeae is a strictly human pathogen that causes the sexually transmitted infection, gonorrhea. Phosphoethanolamine (PEA) addition to the 4' position of the lipid A (PEA-lipid A) moiety of the lipooligosaccharide (LOS) produced by gonococci performs a critical role in this pathogen's ability to evade innate defenses by conferring decreased susceptibility to cationic antimicrobial (or host-defense) peptides, complement-mediated killing by human serum and intraleukocytic killing by human neutrophils compared to strains lacking this PEA decoration. Heretofore, however, it was not known if gonococci can evade autophagy and if so, whether PEA-lipid A contributes to this ability. Accordingly, by using murine macrophages and human macrophage-like phagocytic cell lines we investigated if PEA decoration of gonococcal lipid A modulates autophagy formation. We report that infection with PEA-lipid A-producing gonococci significantly reduced autophagy flux in murine and human macrophages and enhanced gonococcal survival during their association with macrophages compared to a PEA-deficient lipid A mutant. Our results provide further evidence that PEA-lipid A produced by gonococci is a critical component in the ability of this human pathogen to evade host defenses. PMID:26641098

  5. Piscirickettsia salmonis induces apoptosis in macrophages and monocyte-like cells from rainbow trout.

    PubMed

    Rojas, Verónica; Galanti, Norbel; Bols, Niels C; Jiménez, Verónica; Paredes, Rodolfo; Marshall, Sergio H

    2010-05-15

    Piscirickettsia salmonis is the etiologic agent of the salmonid rickettsial septicemia (SRS) which causes significant losses in salmon production in Chile and other and in other regions in the southern hemisphere. As the killing of phagocytes is an important pathogenic mechanism for other bacteria to establish infections in vertebrates, we investigated whether P. salmonis kills trout macrophages by apoptosis. Apoptosis in infected macrophages was demonstrated by techniques based on morphological changes and host cell DNA fragmentation. Transmission electron microcopy showed classic apoptotic characteristics and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling showed fragmented DNA. Programmed cell death type I was further confirmed by increased binding of annexin V to externalized phosphatidylserine in infected macrophages. Moreover, significant increases of caspase 3 activation were detected in infected cells and treatment with caspase inhibitor caused a decrease in levels of apoptosis. This is the first evidence that P. salmonis induces cell death in trout macrophages. This could lead to bacterial survival and evasion of the host immune response and play an important role in the establishment of infection in the host.

  6. Salmonella enterica Replication in Hemophagocytic Macrophages Requires Two Type Three Secretion Systems▿

    PubMed Central

    Silva-Herzog, Eugenia; Detweiler, Corrella S.

    2010-01-01

    Salmonella enterica serotype Typhimurium is a natural pathogen of mice, which acquire the bacteria orally and develop systemic acute infections that can become subacute to chronic infections. S. Typhimurium can reside within hemophagocytic macrophages (HMs) in SV129S6 mice, an Slc11a1/Nramp1+/+ inbred strain. HMs are activated macrophages which have ingested viable hematopoietic cells and are a key characteristic of infectious and inflammatory diseases. Here we show that modest S. Typhimurium replication in HMs begins at 18 h postinfection, while activated macrophages kill the bacteria. For bacterial replication to occur, the phagocytosed viable cells must be grown to a low cell density and the multiplicity of infection must be low. HMs are able to kill phagocytosed Escherichia coli, produce reactive nitrogen species, and retain S. Typhimurium within membrane-bound vesicles. S. Typhimurium does not rescue E. coli upon coinfection of HMs. This indicates that S. Typhimurium does not cause HMs to become permissive for other microbes; rather, S. Typhimurium is especially equipped to survive within HMs. Two type three secretion systems (T3SS) encoded by S. Typhimurium are required for replication within HMs. While the T3SS within Salmonella pathogenicity island 2 (SPI-2) has been previously shown to be important for bacterial survival in cells, a role for SPI-1 in replication in macrophages has not been reported. The requirement for SPI-1 in HMs may help explain the role of SPI-1 during long-term colonization of mice. PMID:20515933

  7. Alternatively activated macrophages produce catecholamines to sustain adaptive thermogenesis

    PubMed Central

    Nguyen, Khoa D.; Qiu, Yifu; Cui, Xiaojin; Goh, Y.P. Sharon; Mwangi, Julia; David, Tovo; Mukundan, Lata; Brombacher, Frank; Locksley, Richard M.; Chawla, Ajay

    2011-01-01

    All homeotherms utilize thermogenesis to maintain core body temperature, ensuring that cellular functions and physiologic processes can ensue in cold environments1-3. In the prevailing model, when the hypothalamus senses cold temperatures, it triggers sympathetic discharge, resulting in the release of noradrenaline in brown adipose tissue (BAT) and white adipose tissue (WAT)4,5. Acting via the β3-adrenergic receptors, noradrenaline induces lipolysis in white adipocytes6, whereas it stimulates the expression of thermogenic genes, such as PPARγ coactivator 1a (Ppargc1a), uncoupling protein 1 (Ucp1), and acyl-CoA synthetase long-chain family member 1 (Acsl1), in brown adipocytes7-9. However, the precise nature of all the cell types involved in this efferent loop is not well established. Here we report an unexpected requirement for the interleukin 4 (IL4)-stimulated program of alternative macrophage activation in adaptive thermogenesis. Cold exposure rapidly promoted alternative activation of adipose tissue macrophages, which secrete catecholamines to induce thermogenic gene expression in BAT and lipolysis in WAT. Absence of alternatively activated macrophages impaired metabolic adaptations to cold, whereas administration of IL4 increased thermogenic gene expression, fatty acid mobilization, and energy expenditure, all in a macrophage-dependent manner. We have thus discovered a surprising role for alternatively activated macrophages in the orchestration of an important mammalian stress response, the response to cold. PMID:22101429

  8. Cellular metabolism and macrophage functional polarization.

    PubMed

    Zhu, Linnan; Zhao, Qingjie; Yang, Tao; Ding, Wenjun; Zhao, Yong

    2015-01-01

    Macrophages are a functionally heterogeneous cell population that is mainly shaped by a variety of microenvironmental stimuli. Interferon γ (IFN-γ), interleukin-1β (IL-1β), and lipopolysaccharide (LPS) induce a classical activation of macrophages (M1), whereas IL-4 and IL-13 induce an alternative activation program in macrophages (M2). Reprogramming of intracellular metabolisms is required for the proper polarization and functions of activated macrophages. Similar to the Warburg effect observed in tumor cells, M1 macrophages increase glucose consumption and lactate release and decreased oxygen consumption rate. In comparison, M2 macrophages mainly employ oxidative glucose metabolism pathways. In addition, fatty acids, vitamins, and iron metabolisms are also related to macrophage polarization. However, detailed metabolic pathways involved in macrophages have remained elusive. Understanding the bidirectional interactions between cellular metabolism and macrophage functions in physiological and pathological situations and the regulatory pathways involved may offer novel therapies for macrophage-associated diseases.

  9. Killing of an encapsulated strain of Escherichia coli by human serum.

    PubMed

    Taylor, P W; Kroll, H P

    1983-01-01

    Changes in cell viability and in factors affecting metabolic integrity were examined after exposure of Escherichia coli LP1092 to human serum. Antibody-dependent classical pathway activity accounted for the rapid killing of strain LP1092 by complement. Removal of serum lysozyme by bentonite absorption or by neutralization with anti-human lysozyme immunoglobulin G resulted in a reduction in the rate of killing; optimal activity could be restored by the addition of physiological amounts of egg-white lysozyme. The pattern of 86Rb+ and alkaline phosphatase release obtained after serum treatment did not support the view that complement simultaneously disrupts cytoplasmic and outer membrane integrity. Macromolecular synthesis was affected late in the reaction sequence; complete inhibition of precursor incorporation into RNA, DNA, and protein occurred only after almost total loss of bacterial colony-forming ability. Addition of chloramphenicol, an inhibitor of protein synthesis, to the bactericidal system resulted in a marked reduction in the rate of serum killing. Killing was completely inhibited by an inhibitor (KCN) and an uncoupler (2,4-dinitrophenol) of oxidative phosphorylation. Exposure of LP1092 cells to serum was followed by a rapid and large increase in intracellular ATP levels; ATP synthesis did not occur when bacteria were exposed to dialyzed serum, which killed LP1092 cells at a much reduced rate. Addition of glucose or serum ultrafiltrate to dialyzed serum restored optimal bactericidal activity. We suggest that optimal killing of gram-negative bacteria is an energy-dependent process requiring an input of bacterially generated ATP. PMID:6185430

  10. Deprivations, futures and the wrongness of killing.

    PubMed

    Marquis, D

    2001-12-01

    In my essay, Why abortion is immoral, I criticised discussions of the morality of abortion in which the crucial issue is whether fetuses are human beings or whether fetuses are persons. Both argument strategies are inadequate because they rely on indefensible assumptions. Why should being a human being or being a person make a moral difference? I argued that the correct account of the morality of abortion should be based upon a defensible account of why killing children and adults is wrong. I claimed that what makes killing us wrong is that our premature deaths deprive us of our futures of value, that is, the goods of life we would have experienced had we survived. This account of the wrongness of killing explains why killing is one of the worst of crimes and how killing greatly harms the victim. It coheres with the attitudes of those with cancer or HIV facing premature death. It explains why we believe it is wrong to kill infants (as personhood theories do not). It does not entail that it wrongs a human being to end her life if she is in persistent vegetative state or if her future must consist only of unbearable physical suffering and she wants to die (as sanctity of human life theories do not). This account of the wrongness of killing implies (with some defensible additional assumptions) that abortion is immoral because we were fetuses once and we know those fetuses had futures of value. Mark Brown claims that this potential future of value account is unsound because it implies that we have welfare rights to what we need to stay alive that most people would reject. I argue that Brown is incorrect in two ways: a welfare right to what we need to stay alive is not directly implied by my account and, in addition, most of us do believe that dependent human beings have substantial welfare rights to what they need to stay alive. Brown argues that depriving us of a future of value of which we have mental representations both is a better explanation of the wrongness of

  11. Conformal killing tensors and covariant Hamiltonian dynamics

    SciTech Connect

    Cariglia, M.; Gibbons, G. W.; Holten, J.-W. van; Horvathy, P. A.; Zhang, P.-M.

    2014-12-15

    A covariant algorithm for deriving the conserved quantities for natural Hamiltonian systems is combined with the non-relativistic framework of Eisenhart, and of Duval, in which the classical trajectories arise as geodesics in a higher dimensional space-time, realized by Brinkmann manifolds. Conserved quantities which are polynomial in the momenta can be built using time-dependent conformal Killing tensors with flux. The latter are associated with terms proportional to the Hamiltonian in the lower dimensional theory and with spectrum generating algebras for higher dimensional quantities of order 1 and 2 in the momenta. Illustrations of the general theory include the Runge-Lenz vector for planetary motion with a time-dependent gravitational constant G(t), motion in a time-dependent electromagnetic field of a certain form, quantum dots, the Hénon-Heiles and Holt systems, respectively, providing us with Killing tensors of rank that ranges from one to six.

  12. Killing activity of microwaves in milk.

    PubMed

    Kindle, G; Busse, A; Kampa, D; Meyer-König, U; Daschner, F D

    1996-08-01

    The killing activity of microwaves of 2450 MHz frequency and 600 W power on Pseudomonas aeruginosa, Escherichia coli, Enterobacter sakazakii, Klebsiella pneumoniae, Staphylococcus aureus, Candida albicans, Mycobacterium terrae and poliomyelitis vaccine-virus suspended in five infant formula preparations was investigated. The samples were brought to the boil (85-100 s depending on milk type). They had reached average temperatures of 82-93 degrees C at this point. Most of the vegetative organisms were killed. In those samples where growth was still detectable after microwave treatment, a significant reduction in viable micro-organisms (at least 5000-fold) was noted. We conclude that microwave beating to the boil is a convenient and fast method to reduce microbial contamination of infant feeds. However, care should be taken to ensure that milk is adequately cooled to the required temperature before it is fed to an infant. PMID:8864939

  13. Killing and replacing queen-laid eggs: low cost of worker policing in the honeybee.

    PubMed

    Kärcher, Martin H; Ratnieks, Francis L W

    2014-07-01

    Worker honeybees, Apis mellifera, police each other's reproduction by killing worker-laid eggs. Previous experiments demonstrated that worker policing is effective, killing most (∼98%) worker-laid eggs. However, many queen-laid eggs were also killed (∼50%) suggesting that effective policing may have high costs. In these previous experiments, eggs were transferred using forceps into test cells, mostly into unrelated discriminator colonies. We measured both the survival of unmanipulated queen-laid eggs and the proportion of removal errors that were rectified by the queen laying a new egg. Across 2 days of the 3-day egg stage, only 9.6% of the queen-laid eggs in drone cells and 4.1% in worker cells were removed in error. When queen-laid eggs were removed from cells, 85% from drone cells and 61% from worker cells were replaced within 3 days. Worker policing in the honeybee has a high benefit to policing workers because workers are more related to the queen's sons (brothers, r = 0.25) than sister workers' sons (0.15). This study shows that worker policing also has a low cost in terms of the killing of queen-laid eggs, as only a small proportion of queen-laid eggs are killed, most of which are rapidly replaced. PMID:24921604

  14. Killing and replacing queen-laid eggs: low cost of worker policing in the honeybee.

    PubMed

    Kärcher, Martin H; Ratnieks, Francis L W

    2014-07-01

    Worker honeybees, Apis mellifera, police each other's reproduction by killing worker-laid eggs. Previous experiments demonstrated that worker policing is effective, killing most (∼98%) worker-laid eggs. However, many queen-laid eggs were also killed (∼50%) suggesting that effective policing may have high costs. In these previous experiments, eggs were transferred using forceps into test cells, mostly into unrelated discriminator colonies. We measured both the survival of unmanipulated queen-laid eggs and the proportion of removal errors that were rectified by the queen laying a new egg. Across 2 days of the 3-day egg stage, only 9.6% of the queen-laid eggs in drone cells and 4.1% in worker cells were removed in error. When queen-laid eggs were removed from cells, 85% from drone cells and 61% from worker cells were replaced within 3 days. Worker policing in the honeybee has a high benefit to policing workers because workers are more related to the queen's sons (brothers, r = 0.25) than sister workers' sons (0.15). This study shows that worker policing also has a low cost in terms of the killing of queen-laid eggs, as only a small proportion of queen-laid eggs are killed, most of which are rapidly replaced.

  15. New Insights Into Tissue Macrophages: From Their Origin to the Development of Memory.

    PubMed

    Italiani, Paola; Boraschi, Diana

    2015-08-01

    Macrophages are the main effector cells of innate immunity and are involved in inflammatory and anti-infective processes. They also have an essential role in maintaining tissue homeostasis, supporting tissue development, and repairing tissue damage. Until few years ago, it was believed that tissue macrophages derived from circulating blood monocytes, which terminally differentiated in the tissue and unable to proliferate. Recent evidence in the biology of tissue macrophages has uncovered a series of immune and ontogenic features that had been neglected for long, despite old observations. These include origin, heterogeneity, proliferative potential (or self-renewal), polarization, and memory. In recent years, the number of publications on tissue resident macrophages has grown rapidly, highlighting the renewed interest of the immunologists for these key players of innate immunity. This mini-review aims to summarizing the new current knowledge in macrophage immunobiology, in order to offer a clear and immediate overview of the field.

  16. Neutrophils prime a long-lived effector macrophage phenotype that mediates accelerated helminth expulsion

    PubMed Central

    Chen, Fei; Wu, Wenhui; Millman, Ariel; Craft, Joshua F.; Chen, Eunice; Patel, Nirav; Boucher, Jean L.; Urban, Joseph F.; Kim, Charles C.; Gause, William C.

    2014-01-01

    We examined the role of innate cells in acquired resistance to the natural murine parasitic nematode, Nippostrongylus brasiliensis. Macrophages obtained as late as 45 days after N. brasiliensis inoculation were able to transfer accelerated parasite clearance to naive recipients. Primed macrophages adhered to larvae in vitro and triggered increased mortality of parasites. Neutrophil depletion in primed mice abrogated the protective effects of transferred macrophages and inhibited their in vitro binding to larvae. Neutrophils in parasite-infected mice showed a distinct transcriptional profile and promoted alternatively activated M2 macrophage polarization through secretory factors including IL-13. Differentially activated neutrophils in the context of a type 2 immune response therefore prime a long-lived effector macrophage phenotype that directly mediates rapid nematode damage and clearance. PMID:25173346

  17. Role for the class A macrophage scavenger receptor in the phagocytosis of apoptotic thymocytes in vitro.

    PubMed Central

    Platt, N; Suzuki, H; Kurihara, Y; Kodama, T; Gordon, S

    1996-01-01

    Numerous immature thymocytes undergo apoptosis and are rapidly engulfed by phagocytic thymic macrophages. The macrophage surface receptors involved in apoptotic thymocyte recognition are unknown. We have examined the role of the class A macrophage scavenger receptor (SR-A) in the engulfment of apoptotic thymocytes. Uptake of steroid-treated apoptotic thymocytes by thymic and inflammatory-elicited SR-A positive macrophages is partially inhibited by an anti-SR-A mAb and more completely by a range of scavenger receptor ligands. Thymic macrophages from mice with targeted disruption of the SR-A gene show a 50% reduction in phagocytosis of apoptotic thymocytes in vitro. These data suggest that SR-A may play a role in the clearance of dying cells in the thymus. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8901603

  18. Mycobacterium avium MAV_2941 mimics Phosphoinositol-3-Kinase to interfere with macrophage phagosome maturation

    PubMed Central

    Danelishvili, Lia; Bermudez, Luiz E.

    2015-01-01

    Mycobacterium avium subsp hominissuis (M. avium) is a pathogen that infects and survives in macrophages. Previously, we have identified the M. avium MAV_2941 gene encoding a 73 amino acid protein exported by the oligopeptide transporter OppA to the macrophage cytoplasm. Mutations in MAV_2941 were associated with significant impairment of M. avium growth in THP-1 macrophages. In this study, we investigated the molecular mechanism of MAV_2941 action and demonstrated that MAV_2941 interacts with the vesicle trafficking proteins syntaxin-8 (STX8), adaptor-related protein complex 3 (AP-3) complex subunit beta-1 (AP3B1) and Archain 1 (ARCN1) in mononuclear phagocytic cells. Sequencing analysis revealed that the binding site of MAV_2941 is structurally homologous to the human phosphatidylinositol 3-kinase (PI3K) chiefly in the region recognized by vesicle trafficking proteins. The β3A subunit of AP-3, encoded by AP3B1, is essential for trafficking cargo proteins, including lysosomal-associated membrane protein 1 (LAMP-1), to the phagosome and lysosome-related organelles. Here, we show that while the heat-killed M. avium when ingested by macrophages co-localizes with LAMP-1 protein, transfection of MAV_2941 in macrophages results in significant decrease of LAMP-1 co-localization with the heat-killed M. avium phagosomes. Mutated MAV_2941, where the amino acids homologous to the binding region of PI3K were changed, failed to interact with trafficking proteins. Inactivation of the AP3B1 gene led to alteration in the trafficking of LAMP-1. These results suggest that M. avium MAV_2941 interferes with the protein trafficking within macrophages altering the maturation of phagosome. PMID:26043821

  19. HIV transcription is induced with cell killing

    SciTech Connect

    Woloschak, G.E.; Schreck, S.; Chang-Liu, Chin Mei; Panozzo, J.; Libertin, C.R.

    1994-01-01

    Previous work has shown that HeLa cells stably transfected with an HIV-LTR-CAT construct are induced to express chloramphenicol acetyl transferase (CAT) following exposure to DNA-damaging agents such as ultraviolet radiation, {gamma} rays, neutrons, and others. In this report, the authors demonstrate that this induction of HIV-LTR transcription occurs when stably transfected HeLa cells are exposed to agents which mediate cell killing, such as UV radiation, electroporation of sucrose buffer, prolonged heating, and low and high pH. Cells cultured following UV exposure demonstrated a peak in CAT expression that is evidence in viable (but not necessarily cell division-competent) cells 24 h after exposure; this inductive response continued until at least 72 h after exposure. HIV-LTR induction was dose-dependent, and the amount of CAT transcription induced was correlated with the amount of cell killing that occurred in the culture. Other agents which caused no cell killing (such as heat-shock for up to 2 h, treatment with metronidazole, exposure to sunlight, vitamin C treatment, and others) had no effect on HIV-LTR induction. These results suggest that HIV transcription is induced as a consequence of the turn on of a cellular death or apoptotic pathway.

  20. Designing surfaces that kill bacteria on contact

    NASA Astrophysics Data System (ADS)

    Tiller, Joerg C.; Liao, Chun-Jen; Lewis, Kim; Klibanov, Alexander M.

    2001-05-01

    Poly(4-vinyl-N-alkylpyridinium bromide) was covalently attached to glass slides to create a surface that kills airborne bacteria on contact. The antibacterial properties were assessed by spraying aqueous suspensions of bacterial cells on the surface, followed by air drying and counting the number of cells remaining viable (i.e., capable of growing colonies). Amino glass slides were acylated with acryloyl chloride, copolymerized with 4-vinylpyridine, and N-alkylated with different alkyl bromides (from propyl to hexadecyl). The resultant surfaces, depending on the alkyl group, were able to kill up to 94 ± 4% of Staphylococcus aureus cells sprayed on them. A surface alternatively created by attaching poly(4-vinylpyridine) to a glass slide and alkylating it with hexyl bromide killed 94 ± 3% of the deposited S. aureus cells. On surfaces modified with N-hexylated poly(4-vinylpyridine), the numbers of viable cells of another Gram-positive bacterium, Staphylococcus epidermidis, as well as of the Gram-negative bacteria Pseudomonas aeruginosa and Escherichia coli, dropped more than 100-fold compared with the original amino glass. In contrast, the number of viable bacterial cells did not decline significantly after spraying on such common materials as ceramics, plastics, metals, and wood.

  1. GOETHALS BRIDGE FROM NORTH SIDE OVER ARTHUR KILL. RAILROAD BRIDGE ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    GOETHALS BRIDGE FROM NORTH SIDE OVER ARTHUR KILL. RAILROAD BRIDGE IN FOREGROUND - Goethals Bridge, Spanning Arthur Kill from New Jersey to Staten Island, Staten Island (subdivision), Richmond County, NY

  2. 11. GENERAL INTERIOR VIEW OF KILLING FLOOR ON LEVEL 4; ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    11. GENERAL INTERIOR VIEW OF KILLING FLOOR ON LEVEL 4; LOOKING SOUTHWEST TOWARD SPLITTERS' PLATFORMS - Rath Packing Company, Beef Killing Building, Sycamore Street between Elm & Eighteenth Streets, Waterloo, Black Hawk County, IA

  3. Macrophage Bactericidal Activities against Staphylococcus aureus Are Enhanced In Vivo by Selenium Supplementation in a Dose-Dependent Manner

    PubMed Central

    Aribi, Mourad; Meziane, Warda; Habi, Salim; Boulatika, Yasser

    2015-01-01

    Background Dietary selenium is of fundamental importance to maintain optimal immune function and enhance immunity during infection. To this end, we examined the effect of selenium on macrophage bactericidal activities against Staphylococcus aureus. Methods Assays were performed in golden Syrian hamsters and peritoneal macrophages cultured with S. aureus and different concentrations of selenium. Results Infected and selenium-supplemented animals have significantly decreased levels of serum nitric oxide (NO) production when compared with infected but non-selenium-supplemented animals at day 7 post-infection (p < 0.05). A low dose of 5 ng/mL selenium induced a significant decrease in macrophage NO production, but significant increase in hydrogen peroxide (H2O2) levels (respectively, p = 0.009, p < 0.001). The NO production and H2O2 levels were significantly increased with increasing concentrations of selenium; the optimal macrophage activity levels were reached at 20 ng/mL. The concentration of 5 ng/mL of selenium induced a significant decrease in the bacterial arginase activity but a significant increase in the macrophage arginase activity. The dose of 20 ng/mL selenium induced a significant decrease of bacterial growth (p < 0.0001) and a significant increase in macrophage phagocytic activity, NO production/arginase balance and S. aureus killing (for all comparisons, p < 0.001). Conclusions Selenium acts in a dose-dependent manner on macrophage activation, phagocytosis and bacterial killing suggesting that inadequate doses may cause a loss of macrophage bactericidal activities and that selenium supplementation could enhance the in vivo control of immune response to S. aureus. PMID:26340099

  4. In vivo manipulation (depletion versus activation) of testicular macrophages: central and local effects.

    PubMed

    Gaytan, F; Bellido, C; Morales, C; García, M; van Rooijen, N; Aguilar, E

    1996-07-01

    Testicular macrophages are a relevant cell type for the regulation of Leydig cell steroidogenesis. The availability of liposome technology allows in vivo manipulation of macrophages in order to analyze their role in the regulation of the hypothalamic-pituitary-testicular axis. In this study, adult (70 days of age) and prepubertal (22 days of age) rats were injected intratesticularly with liposomes containing either dichloromethylene diphosphonate (C12MDP) to deplete testicular macrophages or muramyl tripeptide (MTP-PE) to activate them. Control rats were injected with the corresponding volumes of 0.9% NaCl. Animals were killed 10 days after treatment. Adult rats injected bilaterally or unilaterally with C12MDP liposomes showed increased serum LH and testosterone concentrations, as well as increased testosterone concentrations in the testicular interstitial fluid. In unilaterally injected rats, testosterone concentrations in the interstitial fluid were higher in the macrophage-containing testes than in the contralateral, macrophage-depleted testes. Adult rats treated bilaterally with MTP-PE liposomes showed increased numbers of testicular macrophages, whereas the number of Leydig cells was unchanged. Serum LH concentrations were decreased, but no changes were found in testosterone concentrations. Prepubertal rats treated bilaterally with C12MDP liposomes showed decreased numbers of Leydig cells. However, serum LH and testosterone concentrations were increased. Otherwise, prepubertal rats treated bilaterally with MTP-PE liposomes showed increased numbers of macrophages and Leydig cells, as well as increased serum testosterone concentrations. These data suggest that testicular macrophage-derived factors act at two different levels in the pituitary-testicular axis: first, at a central level by inhibiting LH secretion, and secondly, at a local level by stimulating Leydig cell steroidogenesis.

  5. 78 FR 43063 - Drawbridge Operation Regulations; Arthur Kill, NY

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-19

    ... SECURITY Coast Guard 33 CFR Part 117 Drawbridge Operation Regulations; Arthur Kill, NY AGENCY: Coast Guard... District, has issued a temporary deviation from the regulations governing the operation of the Arthur Kill AK Railroad Bridge across Arthur Kill, mile 11.6, between Staten Island, New York and Elizabeth,...

  6. Road-Killed Animals as Resources for Ecological Studies.

    ERIC Educational Resources Information Center

    Adams, Clark E.

    1983-01-01

    Summarizes 19 literature sources identifying road-killed vertebrates and frequency of kill by numbers. Examples of how these animals can be incorporated into curricula (integrating biology, society, people, and values) are given, followed by an illustrated example of how a road-killed raccoon's skull demonstrated a human/wildlife interaction prior…

  7. 7 CFR 29.1018 - Fire-killed.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 2 2011-01-01 2011-01-01 false Fire-killed. 29.1018 Section 29.1018 Agriculture... Type 92) § 29.1018 Fire-killed. Any leaf of which 5 percent or more of its surface has a set green... tobacco may be described as fire-killed. (See Rule 23.)...

  8. 7 CFR 29.1018 - Fire-killed.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Fire-killed. 29.1018 Section 29.1018 Agriculture... Type 92) § 29.1018 Fire-killed. Any leaf of which 5 percent or more of its surface has a set green... tobacco may be described as fire-killed. (See Rule 23.)...

  9. 7 CFR 29.1018 - Fire-killed.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Fire-killed. 29.1018 Section 29.1018 Agriculture... Type 92) § 29.1018 Fire-killed. Any leaf of which 5 percent or more of its surface has a set green... tobacco may be described as fire-killed. (See Rule 23.)...

  10. 7 CFR 29.1018 - Fire-killed.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 2 2012-01-01 2012-01-01 false Fire-killed. 29.1018 Section 29.1018 Agriculture... Type 92) § 29.1018 Fire-killed. Any leaf of which 5 percent or more of its surface has a set green... tobacco may be described as fire-killed. (See Rule 23.)...

  11. 7 CFR 29.1018 - Fire-killed.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 2 2013-01-01 2013-01-01 false Fire-killed. 29.1018 Section 29.1018 Agriculture... Type 92) § 29.1018 Fire-killed. Any leaf of which 5 percent or more of its surface has a set green... tobacco may be described as fire-killed. (See Rule 23.)...

  12. 9 CFR 113.214 - Parvovirus Vaccine, Killed Virus (Canine).

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Parvovirus Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.214 Parvovirus Vaccine, Killed Virus (Canine). Parvovirus Vaccine... established as follows: (1) Twenty-five parvovirus susceptible dogs (20 vaccinates and 5 controls) shall...

  13. 9 CFR 113.212 - Bursal Disease Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Bursal Disease Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.212 Bursal Disease Vaccine, Killed Virus. Bursal Disease Vaccine... postvaccination, challenge 20 vaccinates and 10 controls by eyedrop with a virulent infectious bursal...

  14. 9 CFR 113.212 - Bursal Disease Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Bursal Disease Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.212 Bursal Disease Vaccine, Killed Virus. Bursal Disease Vaccine... postvaccination, challenge 20 vaccinates and 10 controls by eyedrop with a virulent infectious bursal...

  15. 9 CFR 113.212 - Bursal Disease Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Bursal Disease Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.212 Bursal Disease Vaccine, Killed Virus. Bursal Disease Vaccine... postvaccination, challenge 20 vaccinates and 10 controls by eyedrop with a virulent infectious bursal...

  16. 9 CFR 113.212 - Bursal Disease Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Bursal Disease Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.212 Bursal Disease Vaccine, Killed Virus. Bursal Disease Vaccine... postvaccination, challenge 20 vaccinates and 10 controls by eyedrop with a virulent infectious bursal...

  17. 9 CFR 113.208 - Avian Encephalomyelitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Avian Encephalomyelitis Vaccine... STANDARD REQUIREMENTS Killed Virus Vaccines § 113.208 Avian Encephalomyelitis Vaccine, Killed Virus. Avian Encephalomyelitis Vaccine (Killed Virus) shall be prepared from virus-bearing tissues or fluids obtained...

  18. 9 CFR 113.208 - Avian Encephalomyelitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Avian Encephalomyelitis Vaccine... STANDARD REQUIREMENTS Killed Virus Vaccines § 113.208 Avian Encephalomyelitis Vaccine, Killed Virus. Avian Encephalomyelitis Vaccine (Killed Virus) shall be prepared from virus-bearing tissues or fluids obtained...

  19. 9 CFR 113.204 - Mink Enteritis Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Mink Enteritis Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.204 Mink Enteritis Vaccine, Killed Virus. Mink Enteritis Vaccine... prior to challenge. If unfavorable reactions attributable to the vaccine occur, the serial...

  20. 9 CFR 113.214 - Parvovirus Vaccine, Killed Virus (Canine).

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Parvovirus Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.214 Parvovirus Vaccine, Killed Virus (Canine). Parvovirus Vaccine... Master Seed which has been established as pure, safe, and immunogenic shall be used for...

  1. 9 CFR 113.208 - Avian Encephalomyelitis Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Avian Encephalomyelitis Vaccine... STANDARD REQUIREMENTS Killed Virus Vaccines § 113.208 Avian Encephalomyelitis Vaccine, Killed Virus. Avian Encephalomyelitis Vaccine (Killed Virus) shall be prepared from virus-bearing tissues or fluids obtained...

  2. 9 CFR 113.214 - Parvovirus Vaccine, Killed Virus (Canine).

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Parvovirus Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.214 Parvovirus Vaccine, Killed Virus (Canine). Parvovirus Vaccine... Master Seed which has been established as pure, safe, and immunogenic shall be used for...

  3. 9 CFR 113.214 - Parvovirus Vaccine, Killed Virus (Canine).

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Parvovirus Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.214 Parvovirus Vaccine, Killed Virus (Canine). Parvovirus Vaccine... Master Seed which has been established as pure, safe, and immunogenic shall be used for...

  4. 9 CFR 113.214 - Parvovirus Vaccine, Killed Virus (Canine).

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Parvovirus Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.214 Parvovirus Vaccine, Killed Virus (Canine). Parvovirus Vaccine... Master Seed which has been established as pure, safe, and immunogenic shall be used for...

  5. 9 CFR 113.204 - Mink Enteritis Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Mink Enteritis Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.204 Mink Enteritis Vaccine, Killed Virus. Mink Enteritis Vaccine... prior to challenge. If unfavorable reactions attributable to the vaccine occur, the serial...

  6. 9 CFR 113.204 - Mink Enteritis Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Mink Enteritis Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.204 Mink Enteritis Vaccine, Killed Virus. Mink Enteritis Vaccine... prior to challenge. If unfavorable reactions attributable to the vaccine occur, the serial...

  7. 9 CFR 113.204 - Mink Enteritis Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Mink Enteritis Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.204 Mink Enteritis Vaccine, Killed Virus. Mink Enteritis Vaccine... prior to challenge. If unfavorable reactions attributable to the vaccine occur, the serial...

  8. 9 CFR 113.204 - Mink Enteritis Vaccine, Killed Virus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Mink Enteritis Vaccine, Killed Virus... REQUIREMENTS Killed Virus Vaccines § 113.204 Mink Enteritis Vaccine, Killed Virus. Mink Enteritis Vaccine... prior to challenge. If unfavorable reactions attributable to the vaccine occur, the serial...

  9. Macrophage defense mechanisms against intracellular bacteria

    PubMed Central

    Weiss, Günter; Schaible, Ulrich E

    2015-01-01

    Macrophages and neutrophils play a decisive role in host responses to intracellular bacteria including the agent of tuberculosis (TB), Mycobacterium tuberculosis as they represent the forefront of innate immune defense against bacterial invaders. At the same time, these phagocytes are also primary targets of intracellular bacteria to be abused as host cells. Their efficacy to contain and eliminate intracellular M. tuberculosis decides whether a patient initially becomes infected or not. However, when the infection becomes chronic or even latent (as in the case of TB) despite development of specific immune activation, phagocytes have also important effector functions. Macrophages have evolved a myriad of defense strategies to combat infection with intracellular bacteria such as M. tuberculosis. These include induction of toxic anti-microbial effectors such as nitric oxide and reactive oxygen intermediates, the stimulation of microbe intoxication mechanisms via acidification or metal accumulation in the phagolysosome, the restriction of the microbe's access to essential nutrients such as iron, fatty acids, or amino acids, the production of anti-microbial peptides and cytokines, along with induction of autophagy and efferocytosis to eliminate the pathogen. On the other hand, M. tuberculosis, as a prime example of a well-adapted facultative intracellular bacterium, has learned during evolution to counter-balance the host's immune defense strategies to secure survival or multiplication within this otherwise hostile environment. This review provides an overview of innate immune defense of macrophages directed against intracellular bacteria with a focus on M. tuberculosis. Gaining more insights and knowledge into this complex network of host-pathogen interaction will identify novel target sites of intervention to successfully clear infection at a time of rapidly emerging multi-resistance of M. tuberculosis against conventional antibiotics. PMID:25703560

  10. Macrophage defense mechanisms against intracellular bacteria.

    PubMed

    Weiss, Günter; Schaible, Ulrich E

    2015-03-01

    Macrophages and neutrophils play a decisive role in host responses to intracellular bacteria including the agent of tuberculosis (TB), Mycobacterium tuberculosis as they represent the forefront of innate immune defense against bacterial invaders. At the same time, these phagocytes are also primary targets of intracellular bacteria to be abused as host cells. Their efficacy to contain and eliminate intracellular M. tuberculosis decides whether a patient initially becomes infected or not. However, when the infection becomes chronic or even latent (as in the case of TB) despite development of specific immune activation, phagocytes have also important effector functions. Macrophages have evolved a myriad of defense strategies to combat infection with intracellular bacteria such as M. tuberculosis. These include induction of toxic anti-microbial effectors such as nitric oxide and reactive oxygen intermediates, the stimulation of microbe intoxication mechanisms via acidification or metal accumulation in the phagolysosome, the restriction of the microbe's access to essential nutrients such as iron, fatty acids, or amino acids, the production of anti-microbial peptides and cytokines, along with induction of autophagy and efferocytosis to eliminate the pathogen. On the other hand, M. tuberculosis, as a prime example of a well-adapted facultative intracellular bacterium, has learned during evolution to counter-balance the host's immune defense strategies to secure survival or multiplication within this otherwise hostile environment. This review provides an overview of innate immune defense of macrophages directed against intracellular bacteria with a focus on M. tuberculosis. Gaining more insights and knowledge into this complex network of host-pathogen interaction will identify novel target sites of intervention to successfully clear infection at a time of rapidly emerging multi-resistance of M. tuberculosis against conventional antibiotics. PMID:25703560

  11. Gold nanorods as a theranostic platform for in vitro and in vivo imaging and photothermal therapy of inflammatory macrophages

    NASA Astrophysics Data System (ADS)

    Qin, Jinbao; Peng, Zhiyou; Li, Bo; Ye, Kaichuang; Zhang, Yuxin; Yuan, Fukang; Yang, Xinrui; Huang, Lijia; Hu, Junqing; Lu, Xinwu

    2015-08-01

    Inflammatory macrophages play pivotal roles in the development of atherosclerosis. Theranostics, a promising approach for local imaging and photothermal therapy of inflammatory macrophages, has drawn increasing attention in biomedical research. In this study, gold nanorods (Au NRs) were synthesized, and their in vitro photothermal effects on the macrophage cell line (Ana-1 cells) under 808 nm near infrared reflection (NIR) were investigated by the CCK8 assay, calcein AM/PI staining, flow cytometry, transmission electron microscopy (TEM), silver staining and in vitro micro-computed tomography (CT) imaging. These Au NRs were then applied to an apolipoprotein E knockout (Apo E) mouse model to evaluate their effects on in vivo CT imaging and their effectiveness as for the subsequent photothermal therapy of macrophages in femoral artery restenosis under 808 nm laser irradiation. In vitro photothermal ablation treatment using Au NRs exhibited a significant cell-killing efficacy of macrophages, even at relatively low concentrations of Au NRs and low NIR powers. In addition, the in vivo results demonstrated that the Au NRs are effective for in vivo imaging and photothermal therapy of inflammatory macrophages in femoral artery restenosis. This study shows that Au nanorods are a promising theranostic platform for the diagnosis and photothermal therapy of inflammation-associated diseases.Inflammatory macrophages play pivotal roles in the development of atherosclerosis. Theranostics, a promising approach for local imaging and photothermal therapy of inflammatory macrophages, has drawn increasing attention in biomedical research. In this study, gold nanorods (Au NRs) were synthesized, and their in vitro photothermal effects on the macrophage cell line (Ana-1 cells) under 808 nm near infrared reflection (NIR) were investigated by the CCK8 assay, calcein AM/PI staining, flow cytometry, transmission electron microscopy (TEM), silver staining and in vitro micro-computed tomography

  12. Yersinia pestis can bypass protective antibodies to LcrV and activation with gamma interferon to survive and induce apoptosis in murine macrophages.

    PubMed

    Noel, Betty L; Lilo, Sarit; Capurso, Daniel; Hill, Jim; Bliska, James B

    2009-10-01

    Yersinia pestis, the agent of plague, uses a type III secretion injectisome to deliver Yop proteins into macrophages to counteract phagocytosis and induce apoptosis. Additionally, internalized Y. pestis can survive in the phagosomes of naïve or gamma interferon (IFN-gamma)-activated macrophages by blocking vacuole acidification. The Y. pestis LcrV protein is a target of protective antibodies. The binding of antibodies to LcrV at the injectisome tip results in neutralization of the apoptosis of Y. pestis-infected macrophages and is used as an in vitro correlate of protective immunity. The cytokines IFN-gamma and tumor necrosis factor alpha can cooperate with anti-LcrV to promote protection against lethal Y. pestis infection in mice. It is not known if these phagocyte-activating cytokines cooperate with anti-LcrV to increase the killing of the pathogen and decrease apoptosis in macrophages. We investigated how anti-LcrV and IFN-gamma impact bacterial survival and apoptosis in cultured murine macrophages infected with Y. pestis KIM5. Y. pestis KIM5 opsonized with polyclonal or monoclonal anti-LcrV was used to infect macrophages treated with or without IFN-gamma. The phagocytosis and survival of KIM5 and the apoptosis of macrophages were measured at different time points postinfection. The results show that anti-LcrV reduced apoptosis at an early time point (5 h) but not at a later time point (24 h). Polyclonal anti-LcrV was unable to inhibit apoptosis at either time point in IFN-gamma-activated macrophages. Additionally, anti-LcrV was ineffective at promoting the killing of KIM5 in naïve or activated macrophages. We conclude that Y. pestis can bypass protective antibodies to LcrV and activation with IFN-gamma to survive and induce apoptosis in murine macrophages.

  13. Yersinia pestis Can Bypass Protective Antibodies to LcrV and Activation with Gamma Interferon To Survive and Induce Apoptosis in Murine Macrophages

    PubMed Central

    Noel, Betty L.; Lilo, Sarit; Capurso, Daniel; Hill, Jim; Bliska, James B.

    2009-01-01

    Yersinia pestis, the agent of plague, uses a type III secretion injectisome to deliver Yop proteins into macrophages to counteract phagocytosis and induce apoptosis. Additionally, internalized Y. pestis can survive in the phagosomes of naïve or gamma interferon (IFN-γ)-activated macrophages by blocking vacuole acidification. The Y. pestis LcrV protein is a target of protective antibodies. The binding of antibodies to LcrV at the injectisome tip results in neutralization of the apoptosis of Y. pestis-infected macrophages and is used as an in vitro correlate of protective immunity. The cytokines IFN-γ and tumor necrosis factor alpha can cooperate with anti-LcrV to promote protection against lethal Y. pestis infection in mice. It is not known if these phagocyte-activating cytokines cooperate with anti-LcrV to increase the killing of the pathogen and decrease apoptosis in macrophages. We investigated how anti-LcrV and IFN-γ impact bacterial survival and apoptosis in cultured murine macrophages infected with Y. pestis KIM5. Y. pestis KIM5 opsonized with polyclonal or monoclonal anti-LcrV was used to infect macrophages treated with or without IFN-γ. The phagocytosis and survival of KIM5 and the apoptosis of macrophages were measured at different time points postinfection. The results show that anti-LcrV reduced apoptosis at an early time point (5 h) but not at a later time point (24 h). Polyclonal anti-LcrV was unable to inhibit apoptosis at either time point in IFN-γ-activated macrophages. Additionally, anti-LcrV was ineffective at promoting the killing of KIM5 in naïve or activated macrophages. We conclude that Y. pestis can bypass protective antibodies to LcrV and activation with IFN-γ to survive and induce apoptosis in murine macrophages. PMID:19710295

  14. A whole-genome RNAi screen uncovers a novel role for human potassium channels in cell killing by the parasite Entamoeba histolytica

    PubMed Central

    Marie, Chelsea; Verkerke, Hans P.; Theodorescu, Dan; Petri, William A.

    2015-01-01

    The parasite Entamoeba histolytica kills human cells resulting in ulceration, inflammation and invasion of the colonic epithelium. We used the cytotoxic properties of ameba to select a genome-wide RNAi library to reveal novel host factors that control susceptibility to amebic killing. We identified 281 candidate susceptibility genes and bioinformatics analyses revealed that ion transporters were significantly enriched among susceptibility genes. Potassium (K+) channels were the most common transporter identified. Their importance was further supported by colon biopsy of humans with amebiasis that demonstrated suppressed K+ channel expression. Inhibition of human K+ channels by genetic silencing, pharmacologic inhibitors and with excess K+ protected diverse cell types from E. histolytica-induced death. Contact with E. histolytica parasites triggered K+ channel activation and K+ efflux by intestinal epithelial cells, which preceded cell killing. Specific inhibition of Ca2+-dependent K+ channels was highly effective in preventing amebic cytotoxicity in intestinal epithelial cells and macrophages. Blockade of K+ efflux also inhibited caspase-1 activation, IL-1β secretion and pyroptotic death in THP-1 macrophages. We concluded that K+ channels are host mediators of amebic cytotoxicity in multiple cells types and of inflammasome activation in macrophages. PMID:26346926

  15. A whole-genome RNAi screen uncovers a novel role for human potassium channels in cell killing by the parasite Entamoeba histolytica.

    PubMed

    Marie, Chelsea; Verkerke, Hans P; Theodorescu, Dan; Petri, William A

    2015-01-01

    The parasite Entamoeba histolytica kills human cells resulting in ulceration, inflammation and invasion of the colonic epithelium. We used the cytotoxic properties of ameba to select a genome-wide RNAi library to reveal novel host factors that control susceptibility to amebic killing. We identified 281 candidate susceptibility genes and bioinformatics analyses revealed that ion transporters were significantly enriched among susceptibility genes. Potassium (K(+)) channels were the most common transporter identified. Their importance was further supported by colon biopsy of humans with amebiasis that demonstrated suppressed K(+) channel expression. Inhibition of human K(+) channels by genetic silencing, pharmacologic inhibitors and with excess K(+) protected diverse cell types from E. histolytica-induced death. Contact with E. histolytica parasites triggered K(+) channel activation and K(+) efflux by intestinal epithelial cells, which preceded cell killing. Specific inhibition of Ca(2+)-dependent K(+) channels was highly effective in preventing amebic cytotoxicity in intestinal epithelial cells and macrophages. Blockade of K(+) efflux also inhibited caspase-1 activation, IL-1β secretion and pyroptotic death in THP-1 macrophages. We concluded that K(+) channels are host mediators of amebic cytotoxicity in multiple cells types and of inflammasome activation in macrophages. PMID:26346926

  16. A whole-genome RNAi screen uncovers a novel role for human potassium channels in cell killing by the parasite Entamoeba histolytica.

    PubMed

    Marie, Chelsea; Verkerke, Hans P; Theodorescu, Dan; Petri, William A

    2015-09-08

    The parasite Entamoeba histolytica kills human cells resulting in ulceration, inflammation and invasion of the colonic epithelium. We used the cytotoxic properties of ameba to select a genome-wide RNAi library to reveal novel host factors that control susceptibility to amebic killing. We identified 281 candidate susceptibility genes and bioinformatics analyses revealed that ion transporters were significantly enriched among susceptibility genes. Potassium (K(+)) channels were the most common transporter identified. Their importance was further supported by colon biopsy of humans with amebiasis that demonstrated suppressed K(+) channel expression. Inhibition of human K(+) channels by genetic silencing, pharmacologic inhibitors and with excess K(+) protected diverse cell types from E. histolytica-induced death. Contact with E. histolytica parasites triggered K(+) channel activation and K(+) efflux by intestinal epithelial cells, which preceded cell killing. Specific inhibition of Ca(2+)-dependent K(+) channels was highly effective in preventing amebic cytotoxicity in intestinal epithelial cells and macrophages. Blockade of K(+) efflux also inhibited caspase-1 activation, IL-1β secretion and pyroptotic death in THP-1 macrophages. We concluded that K(+) channels are host mediators of amebic cytotoxicity in multiple cells types and of inflammasome activation in macrophages.

  17. Phagocytosis of aggregated lipoprotein by macrophages: Low density lipoprotein receptor-dependent foam-cell formation

    SciTech Connect

    Suits, A.G.; Chait, A.; Aviram, M.; Heinecke, J.W. )

    1989-04-01

    Low density lipoprotein (LDL) modified by incubation with phospholipase C (PLC-LDL) aggregates in solution and is rapidly taken up and degraded by human and mouse macrophages, producing foam cells in vitro. Human, mouse, and rabbit macrophages degraded {sup 125}I-labeled PLC-LDL ({sup 125}I-PLC-LDL) more rapidly than native {sup 125}I-labeled LDL ({sup 125}I-LDL), while nonphagocytic cells such as human fibroblasts and bovine aortic endothelial cells degraded {sup 125}I-PLC-LDL more slowly than {sup 125}I-LDL. This suggested the mechanism for internalization of PLC-LDL was phagocytosis. When examined by electron microscopy, mouse peritoneal macrophages appeared to be phagocytosing PLC-LDL. The uptake and degradation of {sup 125}I-PLC-LDL by human macrophages was inhibited >80% by the monoclonal antibody C7 (IgG2b) produced by hybridoma C7, which blocks the ligand binding domain of the LDL receptor. Similarly, methylation of {sup 125}I-LDL ({sup 125}I-MeLDL) prior to treatment with phospholipase C decreased its subsequent uptake and degradation by human macrophages by >90%. The uptake and degradation of phospholipase C-modified {sup 125}I-MeLDL by macrophages could be restored by incubation of the methylated lipoprotein with apoprotein E, a ligand recognized by the LDL receptor. These results indicate that macrophages internalize PLC-LDL by LDL receptor-dependent phagocytosis.

  18. Cathepsin Activity-Based Probes and Inhibitor for Preclinical Atherosclerosis Imaging and Macrophage Depletion

    PubMed Central

    Abd-Elrahman, Ihab; Kosuge, Hisanori; Wises Sadan, Tommy; Ben-Nun, Yael; Meir, Karen; Rubinstein, Chen; Bogyo, Matthew; McConnell, Michael V.

    2016-01-01

    Background and Purpose Cardiovascular disease is the leading cause of death worldwide, mainly due to an increasing prevalence of atherosclerosis characterized by inflammatory plaques. Plaques with high levels of macrophage infiltration are considered “vulnerable” while those that do not have significant inflammation are considered stable; cathepsin protease activity is highly elevated in macrophages of vulnerable plaques and contributes to plaque instability. Establishing novel tools for non-invasive molecular imaging of macrophages in plaques could aid in preclinical studies and evaluation of therapeutics. Furthermore, compounds that reduce the macrophage content within plaques should ultimately impact care for this disease. Methods We have applied quenched fluorescent cathepsin activity-based probes (ABPs) to a murine atherosclerosis model and evaluated their use for in vivo imaging using fluorescent molecular tomography (FMT), as well as ex vivo fluorescence imaging and fluorescent microscopy. Additionally, freshly dissected human carotid plaques were treated with our potent cathepsin inhibitor and macrophage apoptosis was evaluated by fluorescent microscopy. Results We demonstrate that our ABPs accurately detect murine atherosclerotic plaques non-invasively, identifying cathepsin activity within plaque macrophages. In addition, our cathepsin inhibitor selectively induced cell apoptosis of 55%±10% of the macrophage within excised human atherosclerotic plaques. Conclusions Cathepsin ABPs present a rapid diagnostic tool for macrophage detection in atherosclerotic plaque. Our inhibitor confirms cathepsin-targeting as a promising approach to treat atherosclerotic plaque inflammation. PMID:27532109

  19. Anoctamin 6 mediates effects essential for innate immunity downstream of P2X7 receptors in macrophages

    NASA Astrophysics Data System (ADS)

    Ousingsawat, Jiraporn; Wanitchakool, Podchanart; Kmit, Arthur; Romao, Ana M.; Jantarajit, Walailak; Schreiber, Rainer; Kunzelmann, Karl

    2015-02-01

    Purinergic P2X7 receptors (P2X7R) are fundamental to innate immune response. In macrophages, transient stimulation of P2X7R activates several transport mechanisms and induces the scrambling of phospholipids with subsequent membrane blebbing and apoptosis. These processes support phagocytosis and subsequent killing of phagocytosed bacteria. Here we demonstrate that the stimulation of P2X7 receptors activates anoctamin 6 (ANO6, TMEM16F), a protein that functions as Ca2+ dependent phospholipid scramblase and Ca2+-activated Cl- channel. Inhibition or knockdown of ANO6 attenuates ATP-induced cell shrinkage, cell migration and phospholipid scrambling. In mouse macrophages, Ano6 produces large ion currents by stimulation of P2X7 receptors and contributes to ATP-induced membrane blebbing and apoptosis, which is largely reduced in macrophages from Ano6-/- mice. ANO6 supports bacterial phagocytosis and killing by mouse and human THP-1 macrophages. Our data demonstrate that anoctamin 6 is an essential component of the immune defense by macrophages.

  20. Cross-resistance of Leishmania infantum isolates to nitric oxide from patients refractory to antimony treatment, and greater tolerance to antileishmanial responses by macrophages.

    PubMed

    de Moura, Tatiana R; Santos, Micheli Luize Barbosa; Braz, Juciene M; Santos, Luis Felipe V C; Aragão, Matheus T; de Oliveira, Fabricia A; Santos, Priscila L; da Silva, Ângela Maria; de Jesus, Amélia Ribeiro; de Almeida, Roque P

    2016-02-01

    Visceral leishmaniasis is a life-threatening disease characterized by intense parasitism of the spleen, liver, and bone marrow. Antimonials have served as front-line antileishmanial therapeutics for decades, but the increasing failure rates under antimonial treatment have challenged the continued use of these drugs. Pentavalent antimonials are known to reinforce the killing mechanisms of macrophages, although the associated mechanism remains unclear. Here, for the first time, we determined whether Leishmania infantum strains isolated from patients refractory to antimony treatment (relapse cases) were cross-resistant to antimonials, liposomal amphotericin B, and/or nitric oxide, and also whether these strains modulate macrophage infection. We selected four clinical isolates from relapse cases and two clinical isolates from antimony-responsive patients (control group) for the present study. The L. infantum promastigotes from all four relapse cases were resistant to trivalent antimonial treatment and nitric oxide, while only one isolate was resistant to liposomal amphotericin B. We evaluated whether the resistant strains from relapse cases showed enhanced infectivity and amastigote survival in macrophages, or macrophage-killing mechanisms in macrophages activated by lipopolysaccharide plus interferon gamma. Infection indexes calculated using macrophages infected with isolates from relapse were higher than those observed with control strains that were stimulated independently. Macrophage infection was higher with L. infantum isolates from relapse cases and correlated with enhanced interleukin 1-β production but showed similar nitrite production. Our results demonstrate that L. infantum field isolates from relapse cases were resistant to antimonials and nitric oxide and that these parasites stimulated inflammatory cytokines and were resistant to macrophage-killing mechanisms, factors that may contribute to disease severity.

  1. Mutations of Francisella novicida that Alter the Mechanism of Its Phagocytosis by Murine Macrophages

    PubMed Central

    Lai, Xin-He; Shirley, Renee L.; Crosa, Lidia; Kanistanon, Duangjit; Tempel, Rebecca; Ernst, Robert K.; Gallagher, Larry A.; Manoil, Colin; Heffron, Fred

    2010-01-01

    Infection with the bacterial pathogen Francisella tularensis tularensis (F. tularensis) causes tularemia, a serious and debilitating disease. Francisella tularensis novicida strain U112 (abbreviated F. novicida), which is closely related to F. tularensis, is pathogenic for mice but not for man, making it an ideal model system for tularemia. Intracellular pathogens like Francisella inhibit the innate immune response, thereby avoiding immune recognition and death of the infected cell. Because activation of inflammatory pathways may lead to cell death, we reasoned that we could identify bacterial genes involved in inhibiting inflammation by isolating mutants that killed infected cells faster than the wild-type parent. We screened a comprehensive transposon library of F. novicida for mutant strains that increased the rate of cell death following infection in J774 macrophage-like cells, as compared to wild-type F. novicida. Mutations in 28 genes were identified as being hypercytotoxic to both J774 and primary macrophages of which 12 were less virulent in a mouse infection model. Surprisingly, we found that F. novicida with mutations in four genes (lpcC, manB, manC and kdtA) were taken up by and killed macrophages at a much higher rate than the parent strain, even upon treatment with cytochalasin D (cytD), a classic inhibitor of macrophage phagocytosis. At least 10-fold more mutant bacteria were internalized by macrophages as compared to the parent strain if the bacteria were first fixed with formaldehyde, suggesting a surface structure is required for the high phagocytosis rate. However, bacteria were required to be viable for macrophage toxicity. The four mutant strains do not make a complete LPS but instead have an exposed lipid A. Interestingly, other mutations that result in an exposed LPS core were not taken up at increased frequency nor did they kill host cells more than the parent. These results suggest an alternative, more efficient macrophage uptake mechanism

  2. Growth rate paradox of Salmonella typhimurium within host macrophages.

    PubMed Central

    Abshire, K Z; Neidhardt, F C

    1993-01-01

    The growth rate of Salmonella typhimurium U937 within host macrophages was estimated by two independent methods. The extent to which ribosomal protein L12 is acetylated to produce ribosomal protein L7 changes markedly with the growth rate. By this measure, the intracellular bacteria appeared to be growing rapidly. Measurements of viable bacteria, however, indicated that the bacteria were growing slowly. A solution of this apparent growth rate paradox was sought by treating U937 cells infected with S. typhimurium X3306 with ampicillin or chloramphenicol to help determine the number of bacteria that were actively growing and dividing in the intracellular condition. Use of these antibiotics showed that by 2 h after invasion, the intracellular bacteria consisted of at least two populations, one static and the other rapidly dividing. This finding implies that previously described changes in the gene expression of S. typhimurium are important for the survival and/or multiplication of the bacteria within the macrophage. Images PMID:8509329

  3. Dynamic kill: controlling wild wells a new way

    SciTech Connect

    Blount, E.M.; Soeiinah, E.

    1981-10-01

    Dynamic kill describes a technique for terminating a blowout utilizing flowing frictional pressure to supplement the hydrostatic pressure of the kill fluid being injected through the relief well and up the blowing well. Therefore, a lighter kill fluid such as water can be implemented. The objective is to allow a blowout to be killed without breaking down the formation so the maximum amount of fluid can be circulated through the relief well by not losing fluid to a fractured formation. This allows optimum control during the kill operation and stable communication between the two wells. By allowing more fluid to be applied to the kill through one relief well, dynamic kill also increases the probability that one relief well will be sufficient. When the well is dynamically dead, the initial kill fluid, which will usually be too light to hold the well dead in a static condition, is replaced with a heavier kill mud. In fact, three weights of mud may be required to allow control during the transition from low density initial dynamic kill mud to heavy final kill mud. 5 refs.

  4. [The question of killing in animal protection ethics].

    PubMed

    Luy, J

    2000-03-01

    About twenty years ago the traditional question whether humans are allowed to kill animals was replaced by two new questions: the question of killing and the question of suffering. The question of killing ist the abstract question if--in isolation from the question of suffering--finishing an animal's life is moral. So the question of killing can be raised for every single case of an animal killed by man. The moral evaluation of a killing without suffering (of all directly and indirectly involved beings) concerns just the question of killing; for example, an experiment in which the animal is made unconscious by an anesthetic prior to the experiment being performed and is then killed before it regains consciousness.--The comparative analysis of the popular arguments demonstrates fallacies in some of them and shows a new view of the problem. Because not the killing itself (without fear or pain) but the innate fear of death is the real motive to reject ones own killing, it is necessary to demand that every killing of an animal by man is done without any fear (or pain) for the animal. To claim an animal's right to life however is not yet justified.

  5. Oral Inflammatory Diseases and Systemic Inflammation: Role of the Macrophage

    PubMed Central

    Hasturk, Hatice; Kantarci, Alpdogan; Van Dyke, Thomas E.

    2012-01-01

    Inflammation is a complex reaction to injurious agents and includes vascular responses, migration, and activation of leukocytes. Inflammation starts with an acute reaction, which evolves into a chronic phase if allowed to persist unresolved. Acute inflammation is a rapid process characterized by fluid exudation and emigration of leukocytes, primarily neutrophils, whereas chronic inflammation extends over a longer time and is associated with lymphocyte and macrophage infiltration, blood vessel proliferation, and fibrosis. Inflammation is terminated when the invader is eliminated, and the secreted mediators are removed; however, many factors modify the course and morphologic appearance as well as the termination pattern and duration of inflammation. Chronic inflammatory illnesses such as diabetes, arthritis, and heart disease are now seen as problems that might have an impact on the periodontium. Reciprocal effects of periodontal diseases are potential factors modifying severity in the progression of systemic inflammatory diseases. Macrophages are key cells for the inflammatory processes as regulators directing inflammation to chronic pathological changes or resolution with no damage or scar tissue formation. As such, macrophages are involved in a remarkably diverse array of homeostatic processes of vital importance to the host. In addition to their critical role in immunity, macrophages are also widely recognized as ubiquitous mediators of cellular turnover and maintenance of extracellular matrix homeostasis. In this review, our objective is to identify macrophage-mediated events central to the inflammatory basis of chronic diseases, with an emphasis on how control of macrophage function can be used to prevent or treat harmful outcomes linked to uncontrolled inflammation. PMID:22623923

  6. Role of ceruloplasmin in macrophage iron efflux during hypoxia.

    PubMed

    Sarkar, Joydeep; Seshadri, Vasudevan; Tripoulas, Nicholas A; Ketterer, Michael E; Fox, Paul L

    2003-11-01

    The reticuloendothelial system has a central role in erythropoiesis and iron homeostasis. An important function of reticuloendothelial macrophages is phagocytosis of senescent red blood cells. The iron liberated from heme is recycled for delivery to erythrocyte precursors for a new round of hemoglobin synthesis. The molecular mechanism by which recycled iron is released from macrophages remains unresolved. We have investigated the mechanism of macrophage iron efflux, focusing on the role of ceruloplasmin (Cp), a copper protein with a potent ferroxidase activity that converts Fe2+ to Fe3+ in the presence of molecular oxygen. As shown by others, Cp markedly increased iron binding to apotransferrin at acidic pH; however, the physiological significance of this finding is uncertain because little stimulation was observed at neutral pH. Introduction of a hypoxic atmosphere resulted in marked Cp-stimulated binding of iron to apotransferrin at physiological pH. The role of Cp in cellular iron release was examined in U937 monocytic cells induced to differentiate to the macrophage lineage. Cp added at its normal plasma concentration increased the rate of 55Fe release from U937 cells by about 250%. The stimulation was absolutely dependent on the presence of apotransferrin and hypoxia. Cp-stimulated iron release was confirmed in mouse peritoneal macrophages. Stimulation of iron release required an intracellular "labile iron pool" that was rapidly depleted in the presence of Cp and apotransferrin. Ferroxidase-mediated loading of iron into apotransferrin was critical for iron release because ferroxidase-deficient Cp was inactive and because holotransferrin could not substitute for apotransferrin. The extracellular iron concentration was critical as shown by inhibition of iron release by exogenous free iron, and marked enhancement of release by an iron chelator. Together these data show that Cp stimulates iron release from macrophages under hypoxic conditions by a ferroxidase

  7. Micro-sociology of mass rampage killings.

    PubMed

    Collins, Randall

    2014-01-01

    Spectacular but very rare violent events such as mass killings by habitual non-criminals cannot be explained by factors which are very widespread, such as possession of firearms, being a victim of bullying, an introvert, or a career failure. A stronger clue is clandestine preparation of attack by one or two individuals, against randomly chosen representatives of a hated collective identity. Mass killers develop a deep back-stage, obsessed with planning their attack, overcoming social inferiority and isolation by an emotion of clandestine excitement. PMID:25179819

  8. Targeting the Checkpoint to Kill Cancer Cells.

    PubMed

    Benada, Jan; Macurek, Libor

    2015-01-01

    Cancer treatments such as radiotherapy and most of the chemotherapies act by damaging DNA of cancer cells. Upon DNA damage, cells stop proliferation at cell cycle checkpoints, which provides them time for DNA repair. Inhibiting the checkpoint allows entry to mitosis despite the presence of DNA damage and can lead to cell death. Importantly, as cancer cells exhibit increased levels of endogenous DNA damage due to an excessive replication stress, inhibiting the checkpoint kinases alone could act as a directed anti-cancer therapy. Here, we review the current status of inhibitors targeted towards the checkpoint effectors and discuss mechanisms of their actions in killing of cancer cells. PMID:26295265

  9. Targeting the Checkpoint to Kill Cancer Cells

    PubMed Central

    Benada, Jan; Macurek, Libor

    2015-01-01

    Cancer treatments such as radiotherapy and most of the chemotherapies act by damaging DNA of cancer cells. Upon DNA damage, cells stop proliferation at cell cycle checkpoints, which provides them time for DNA repair. Inhibiting the checkpoint allows entry to mitosis despite the presence of DNA damage and can lead to cell death. Importantly, as cancer cells exhibit increased levels of endogenous DNA damage due to an excessive replication stress, inhibiting the checkpoint kinases alone could act as a directed anti-cancer therapy. Here, we review the current status of inhibitors targeted towards the checkpoint effectors and discuss mechanisms of their actions in killing of cancer cells. PMID:26295265

  10. Progress at Fresh Kills improving water quality

    SciTech Connect

    Londres, E.J.

    1991-06-01

    This paper reports that in December 1987, the federal district court in Nevada issued a consent order forcing New York City (NYC) to improve its handling of solid waste and reduce the discharge of solid waste into the surrounding waterway. Implementation of the consent order by NYC resulted in many improvements in the transport of solid waste from the Marine Transfer Station (MTS) to Fresh Kills Landfill. The end result was a marked reduction in solid waste discharge and an improvement in water quality along the New Jersey shore areas.

  11. Killing-Yano forms and Killing tensors on a warped space

    NASA Astrophysics Data System (ADS)

    Krtouš, Pavel; KubizÅák, David; Kolář, Ivan

    2016-01-01

    We formulate several criteria under which the symmetries associated with the Killing and Killing-Yano tensors on the base space can be lifted to the symmetries of the full warped geometry. The procedure is explicitly illustrated on several examples, providing new prototypes of spacetimes admitting such tensors. In particular, we study a warped product of two Kerr-NUT-(A)dS spacetimes and show that it gives rise to a new class of highly symmetric vacuum (with a cosmological constant) black hole solutions that inherit many of the properties of the Kerr-NUT-(A)dS geometry.

  12. Caprine arthritis encephalitis virus dysregulates the expression of cytokines in macrophages.

    PubMed Central

    Lechner, F; Machado, J; Bertoni, G; Seow, H F; Dobbelaere, D A; Peterhans, E

    1997-01-01

    Caprine arthritis encephalitis virus (CAEV) is a lentivirus of goats that leads to chronic mononuclear infiltration of various tissues, in particular, the radiocarpal joints. Cells of the monocyte/macrophage lineage are the major host cells of CAEV in vivo. We have shown that infection of cultured goat macrophages with CAEV results in an alteration of cytokine expression in vitro. Constitutive expression of interleukin 8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) was increased in infected macrophages, whereas transforming growth factor beta1 (TGF-beta1) mRNA was down-regulated. When macrophages were infected with a CAEV clone lacking the trans-acting nuclear regulatory gene tat, IL-8 and MCP-1 were also increased. No significant differences from cells infected with the wild-type clone were observed, suggesting that Tat is not required for the increased expression of IL-8 and MCP-1 in infected macrophages. Furthermore, infection with CAEV led to an altered pattern of cytokine expression in response to lipopolysaccharide (LPS), heat-killed Listeria monocytogenes plus gamma interferon, or fixed cells of Staphylococcus aureus Cowan I. In infected macrophages, tumor necrosis factor alpha, IL-1beta, IL-6, and IL-12 p40 mRNA expression was reduced in response to all stimuli tested whereas changes in expression of granulocyte-macrophage colony-stimulating factor depended on the stimulating agent. Electrophoretic mobility shift assays demonstrated that, in contrast to effects of human immunodeficiency virus infection of macrophages, CAEV infection had no effect on the level of constitutive nuclear factor-kappaB (NF-kappaB) activity or on the level of LPS-stimulated NF-kappaB activity, suggesting that NF-kappaB is not involved in altered regulation of cytokine expression in CAEV-infected cells. In contrast, activator protein 1 (AP-1) binding activity was decreased in infected macrophages. These data show that CAEV infection may result in a dysregulation of

  13. Macrophage cell lines derived from major histocompatibility complex II-negative mice

    NASA Technical Reports Server (NTRS)

    Beharka, A. A.; Armstrong, J. W.; Chapes, S. K.; Spooner, B. S. (Principal Investigator)

    1998-01-01

    Two bone-marrow-derived macrophage cell lines, C2D and C2Dt, were isolated from major histocompatibility class II negative knock-out mice. The C2D cell line was stabilized by continuous culture in colony-stimulating factor-1 and the C2Dt cell line was transformed with SV40 virus large T antigen. These cells exhibited phenotypic properties of macrophages including morphology and expression of Mac 1 and Mac 2 cell surface molecules. These cells also had comparable growth to the bone-marrow-derived macrophage cell line B6MP102. These new cell lines were not spontaneously cytotoxic and were only capable of modest killing of F5b tumor cells when stimulated with LPS and interferon-gamma, but not when stimulated with LPS alone or with staphylococcal exotoxin. C2D and C2Dt cells phagocytosed labeled Staphylococcus aureus similarly to B6MP102 cells but less well than C2D peritoneal macrophages. These cell lines secreted interleukin-6, but not tumor necrosis factor or nitric oxide in response to LPS or staphlococcal enterotoxins A or B C2D(t) cells were tumorigenic in C2D and C57BL/6J mice but C2D cells were not. These data suggest that macrophage cell lines can be established from bone marrow cells of major histocompatibility complex II-negative mice.

  14. Interactions between macrophage/Kupffer cells and hepatocytes in surgical sepsis

    SciTech Connect

    West, M.A.

    1988-01-01

    Experiments were performed to investigate the role of Kupffer cell/macrophage interactions with hepatocytes in modulating liver function during infections using direct in vitro cocultivation of rat macrophages or Kupffer cells with rat hepatocytes. Protein synthesis was assayed as a sensitive indicator of integrated hepatocellular function by measuring {sup 3}H-leucine incorporation into hepatocyte protein. Septic stimuli such as lipoploysaccharide and killed bacteria were added to cocultures of hepatocytes and macrophages or Kupffer cells and the responses compared to hepatocytes alone. Information about the types of proteins synthesized by hepatocytes under various culture conditions was determined using polyacrylamide gel electrophoresis and autoradiography. These experiments showed that septic stimuli alter the amount and type of protein synthesized by hepatocytes and had no direct effect on hepatocytes in the absence of macrophages or Kupffer cells. The mediator(s) appears to be a heat labile, soluble monokine(s) which is distinct from interleukin-1 or tumor necrosis factor. The important role of Kupffer cells/macrophages in mediating alterations in hepatocellular function in sepsis may ultimately improve patient care.

  15. Interferon-Gamma Improves Macrophages Function against M. tuberculosis in Multidrug-Resistant Tuberculosis Patients.

    PubMed

    Khan, Taj Ali; Mazhar, Humaira; Saleha, Shamim; Tipu, Hamid Nawaz; Muhammad, Niaz; Abbas, Muhammad Nasser

    2016-01-01

    Background. Mycobacterium tuberculosis (M. tuberculosis) that causes tuberculosis (TB) kills millions of infected people annually especially multidrug-resistant tuberculosis (MDR-TB). On infection, macrophages recognize the mycobacteria by toll-like receptor (TLR) followed by phagocytosis and control of mycobacteria. In addition, macrophages also secrete IL-12 to induce IFN-γ production by T, which, in turn, increases the phagocytosis and oxidative burst. Individuals with defects in innate or adaptive immunity exhibit increased susceptibility to M. tuberculosis. Understanding these immunologic mechanisms will help in TB control. We aimed to investigate the immunopathologic mechanisms in MDR-TB and role of recombinant human interferon-gamma (rhIFN-γ). Study Design and Methods. Monocyte-derived macrophages (MDMs) were generated from peripheral blood mononuclear cells of MDR-TB patients and healthy subjects and were investigated for immunologic response by ELISA and flow cytometry. Results. Different functional and molecular anomalies were observed in macrophages. In addition, a defective immune response to M. tuberculosis from the patient's MDMs was characterized, which in turn improved by pretreatment with rhIFN-γ. Conclusion. This work highlights the fact that rhIFN-γ improves macrophages function against M. tuberculosis and treatment of patients with poor responsiveness to TB therapy may be needed in future to include IFN-γ as adjuvant therapy after the full characterization of pathological and molecular mechanisms in these and in other more multidrug-resistant TB patients. PMID:27478636

  16. Interferon-Gamma Improves Macrophages Function against M. tuberculosis in Multidrug-Resistant Tuberculosis Patients

    PubMed Central

    Mazhar, Humaira; Muhammad, Niaz; Abbas, Muhammad Nasser

    2016-01-01

    Background. Mycobacterium tuberculosis (M. tuberculosis) that causes tuberculosis (TB) kills millions of infected people annually especially multidrug-resistant tuberculosis (MDR-TB). On infection, macrophages recognize the mycobacteria by toll-like receptor (TLR) followed by phagocytosis and control of mycobacteria. In addition, macrophages also secrete IL-12 to induce IFN-γ production by T, which, in turn, increases the phagocytosis and oxidative burst. Individuals with defects in innate or adaptive immunity exhibit increased susceptibility to M. tuberculosis. Understanding these immunologic mechanisms will help in TB control. We aimed to investigate the immunopathologic mechanisms in MDR-TB and role of recombinant human interferon-gamma (rhIFN-γ). Study Design and Methods. Monocyte-derived macrophages (MDMs) were generated from peripheral blood mononuclear cells of MDR-TB patients and healthy subjects and were investigated for immunologic response by ELISA and flow cytometry. Results. Different functional and molecular anomalies were observed in macrophages. In addition, a defective immune response to M. tuberculosis from the patient's MDMs was characterized, which in turn improved by pretreatment with rhIFN-γ. Conclusion. This work highlights the fact that rhIFN-γ improves macrophages function against M. tuberculosis and treatment of patients with poor responsiveness to TB therapy may be needed in future to include IFN-γ as adjuvant therapy after the full characterization of pathological and molecular mechanisms in these and in other more multidrug-resistant TB patients. PMID:27478636

  17. Lymphocyte-conditioned medium protects human monocyte-macrophages from cholesteryl ester accumulation.

    PubMed Central

    Fogelman, A M; Seager, J; Haberland, M E; Hokom, M; Tanaka, R; Edwards, P A

    1982-01-01

    Exposure of human monocyte-macrophages to as little as 50 microliters of cultured medium from lymphocytes stimulated by concanavalin A (Con A) resulted in a dramatic decrease in the activities of the low density lipoprotein (LDL) receptor pathway, the LDL-dextran sulfate pathway, and the scavenger receptor pathway. This effect was not seen when the monocyte-macrophages were exposed to culture medium from lymphocytes cultured without Con A or with Con A together with alpha-methyl mannoside or control medium without lymphocytes. The activity of 3-hydroxy-3-methyglutaryl-coenzyme A reductase also decreased in monocyte-macrophages exposed to culture medium from stimulated lymphocytes. Acyl-CoA:cholesterol O-acyltransferase activity, protein synthesis, protein content, phagocytosis of heat-killed yeast, and non-receptor-mediated endocytosis were not inhibited. Monocyte-macrophages exposed to malondialdehyde altered-LDL in the presence of stimulated lymphocyte culture medium accumulated substantially less cholesteryl esters than did cells in control medium. We propose that substances produced by stimulated lymphocytes may be useful in protecting macrophages from cholesteryl ester accumulation. Images PMID:6278500

  18. Using RNA-interference to Investigate the Innate Immune Response in Mouse Macrophages

    PubMed Central

    De Arras, Lesly; Guthrie, Brandon S.; Alper, Scott

    2014-01-01

    Macrophages are key phagocytic innate immune cells. When macrophages encounter a pathogen, they produce antimicrobial proteins and compounds to kill the pathogen, produce various cytokines and chemokines to recruit and stimulate other immune cells, and present antigens to stimulate the adaptive immune response. Thus, being able to efficiently manipulate macrophages with techniques such as RNA-interference (RNAi) is critical to our ability to investigate this important innate immune cell. However, macrophages can be technically challenging to transfect and can exhibit inefficient RNAi-induced gene knockdown. In this protocol, we describe methods to efficiently transfect two mouse macrophage cell lines (RAW264.7 and J774A.1) with siRNA using the Amaxa Nucleofector 96-well Shuttle System and describe procedures to maximize the effect of siRNA on gene knockdown. Moreover, the described methods are adapted to work in 96-well format, allowing for medium and high-throughput studies. To demonstrate the utility of this approach, we describe experiments that utilize RNAi to inhibit genes that regulate lipopolysaccharide (LPS)-induced cytokine production. PMID:25407484

  19. Using RNA-interference to investigate the innate immune response in mouse macrophages.

    PubMed

    De Arras, Lesly; Guthrie, Brandon S; Alper, Scott

    2014-11-03

    Macrophages are key phagocytic innate immune cells. When macrophages encounter a pathogen, they produce antimicrobial proteins and compounds to kill the pathogen, produce various cytokines and chemokines to recruit and stimulate other immune cells, and present antigens to stimulate the adaptive immune response. Thus, being able to efficiently manipulate macrophages with techniques such as RNA-interference (RNAi) is critical to our ability to investigate this important innate immune cell. However, macrophages can be technically challenging to transfect and can exhibit inefficient RNAi-induced gene knockdown. In this protocol, we describe methods to efficiently transfect two mouse macrophage cell lines (RAW264.7 and J774A.1) with siRNA using the Amaxa Nucleofector 96-well Shuttle System and describe procedures to maximize the effect of siRNA on gene knockdown. Moreover, the described methods are adapted to work in 96-well format, allowing for medium and high-throughput studies. To demonstrate the utility of this approach, we describe experiments that utilize RNAi to inhibit genes that regulate lipopolysaccharide (LPS)-induced cytokine production.

  20. Role of high-avidity binding of human neutrophil myeloperoxidase in the killing of Actinobacillus actinomycetemcomitans.

    PubMed Central

    Miyasaki, K T; Zambon, J J; Jones, C A; Wilson, M E

    1987-01-01

    The binding of the neutrophil enzyme myeloperoxidase (MPO) to microbial surfaces is believed to be the first step in its microbicidal activity. The MPO-H2O2-Cl- system is responsible for most oxidative killing of Actinobacillus actinomycetemcomitans by human neutrophils. There appear to be three forms of MPO (MPO I, II, and III), all of which can kill this organism in the presence of H2O2 and chloride. In this study, we characterized the binding of native human neutrophil MPO to A. actinomycetemcomitans by an elution procedure dependent on the cationic detergent cetyltrimethylammonium bromide. Binding of native MPO was rapid and reached apparent equilibrium within 1 min. A proportion of binding under equilibrium conditions was saturable and highly avid, with a capacity of 4,500 sites per cell and a dissociation constant of 7.9 X 10(-10) M. At equal protein concentrations, more MPO III bound than MPO II, and more MPO II bound than MPO I. The high-avidity interaction was inhibitable with yeast mannan and with the serotype-defining mannan of A. actinomycetemcomitans. Binding was also partially reversible with yeast mannan. MPO bound to the high-avidity sites did not oxidize guaiacol but oxidized chloride, as detected by the chlorination of taurine. MPO bound to the high-avidity sites was incapable of killing A. actinomycetemcomitans alone in the presence of H2O2 and Cl-, but potentiated killing when sufficient additional MPO was provided. The killing of A. actinomycetemcomitans by the MPO-H2O2-Cl- system was inhibited by yeast mannan and a serotype-defining mannan of A. actinomycetemcomitans. We conclude that high-avidity binding of MPO to the surface of A. actinomycetemcomitans is a mannan-specific interaction and that MPO bound to the high-avidity sites is essential but not alone sufficient to kill A. actinomycetemcomitans. PMID:3032796

  1. Macrophage functions in Biozzi mice.

    PubMed Central

    Dockrell, H M; Taverne, J; Lelchuk, R; Depledge, P; Brown, I N; Playfair, J H

    1985-01-01

    The faster degradation of antigen by macrophages in Biozzi low (L) responder mice, compared to Biozzi high (H) responder mice, is thought to be responsible for their lower antibody response. We have measured four functions associated with macrophages to see whether macrophages from L mice were generally more active than those from H mice. Peritoneal macrophages obtained from normal mice were compared with those from groups of mice given Mycobacterium bovis BCG or Propionibacterium acnes. Cells from normal H mice gave a stronger oxidative burst when triggered with phorbol myristate acetate, and were more cytotoxic for tumour cells than cells from L mice. Cells from all mice injected with BCG or P. acnes gave a stronger oxidative burst, and were more cytotoxic for tumour cells; again, both responses were higher in H mice than in L mice. By contrast, when groups of mice that had received P. acnes were given endotoxin and bled, higher titres of tumour necrosis factor were found in the sera of L mice. Spleen cells from both lines of mice released similar levels of interleukin-1, both spontaneously and in response to lipopolysaccharide. Our results suggest that these various macrophage responses are expressed independently in H and L mice. PMID:3894222

  2. [Differential growth inhibition of mycobacteria by interferon-gamma-or tumor necrosis factor-alpha-treated murine peritoneal macrophages].

    PubMed

    Sato, K; Tomioka, H; Saito, H

    1996-11-01

    Growth inhibition of the intracellular mycobacteria such as Mycobacterium tuberculosis, M. bovis, M. kansasii, M. avium, M. intracellulare, M. fortuitum, and M. chelonae subsp. abscessus by interferon-gamma (IFN-gamma)- or tumor necrosis factor-alpha (TNF-alpha)-treated murine peritoneal macrophages elicited by proteose peptone was studied in vitro. Macrophages were infected with slowly growing mycobacteria and the extracellular mycobacteria were washed out. Then, macrophages were treated with IFN-gamma or TNF-alpha at a concentration of 10 to 1000 U/ml for 2 days. In another experiment, macrophages were pretreated with these cytokines for 1 day then infected with rapidly growing mycobacteria as before. Macrophages were cultured with or without IFN-gamma or TNF-alpha for additional day. Mycobacterial growth was assessed by determination of colony-forming units on 7H11 agar plates after destruction of the macrophages. Stimulation of macrophages with IFN-gamma reduced the growth of mycobacteria. However, except for M. tuberculosis and M. bovis, growth was not inhibited by macrophages treated with TNF-alpha. IFN-gamma seems to be an important cytokine for the activation of mycobactericidal mechanisms in murine macrophages. Stimulation with IFN-gamma or TNF-alpha and subsequent phagocytosis of M. tuberculosis or M. intracellulare increased O2- production, which was assayed by the method of cytochrome C reduction by murine peritoneal macrophages. Phorbol myristate acetate-triggered-O2- production was also elevated by the cytokine pretreatment of the macrophages, suggesting that mycobacterial growth inhibition did not parallel the production of reactive oxygen intermediates in TNF alpha-activated murine peritoneal macrophages. These data suggest that bactericidal mechanisms of murine macrophages against nontuberculous mycobacteria may not depend on reactive oxygen intermediates. PMID:8958673

  3. Why are potential women being killed?

    PubMed

    Thomson, A

    1993-12-01

    The persistence of traditional practices that provide disincentives to having daughters is giving rise to widespread infanticide in India. In a survey conducted in Madras in 1993, over half of the mothers interviewed acknowledged having killed an infant girl. The infanticide rate is believed to be even higher in India's rural areas. Families who can afford ultrasound to determine the fetal sex are reportedly using selective abortion to avert the birth of a daughter. Of 8000 abortions induced in a Bombay clinic, 7999 involved a female fetus. Families cite the financial burden inherent in providing a dowry as the primary reason for female infanticide. Also cited is the need for a son to both provide financial support to parents in old age and to light their funeral pyre. There are reports of mothers who refuse to kill female infants being abandoned or physically battered by their husbands. At present, there are 116 males to every 100 females in India--an imbalance that is likely to increase in the future and make it impossible for many men to form families. Just as television has been implicated in creating a demand for large dowries that would enable husbands' families to purchase Western luxury items, the mass media should use its influence to alter the attitudes that perpetuate the low status of women in India.

  4. The Pathogenic Role of Macrophage Migration Inhibitory Factor in Immunologically Induced Kidney Disease in the Rat

    PubMed Central

    Lan, Hui Y.; Bacher, Michael; Yang, Niansheng; Mu, Wei; Nikolic-Paterson, David J.; Metz, Christine; Meinhardt, Andreas; Bucala, Richard; Atkins, Robert C.

    1997-01-01

    Macrophage migration inhibitory factor (MIF) plays a pivotal role in the inflammatory response in endotoxemia and in the delayed-type hypersensitivity response, but its potential as a regulator of immunologically induced disease is unknown. We have addressed this issue by administering a neutralizing anti-MIF antibody in a rat model of immunologically induced crescentic anti-glomerular basement membrane (GBM) glomerulonephritis. Six individual experiments using paired inbred littermates were performed. Rats were primed with rabbit immunoglobulin on day −5 and then injection with rabbit anti–rat GBM serum on day 0. Pairs of animals were treated with anti-MIF or a control monoclonal antibody from the time of anti-GBM serum administration until being killed 14 d later. Control antibody-treated animals developed severe proteinuria and renal function impairment with severe histological damage due to marked leukocytic infiltration and activation within the kidney. In contrast, anti-MIF treatment substantially reduced proteinuria, prevented the loss of renal function, significantly reduced histological damage including glomerular crescent formation, and substantially inhibited renal leukocytic infiltration and activation (all P <0.001 compared with control treatment). Inhibition of renal disease by anti-MIF treatment was attributed to preventing the marked upregulation of interleukin-1β, leukocyte adhesion molecules including intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, and inducible nitric oxide synthase expression seen in the control antibody-treated animals. This inhibition of progressive renal injury was mirrored by the complete suppression of the skin delayed-type hypersensitivity response to the challenge antigen (rabbit IgG). Interestingly, anti-MIF treatment did not effect the secondary antibody response or immune deposition within the kidney, indicating that MIF participates in cellular-based immunity in this primed macrophage

  5. Effect of aqueous extract of Tinospora cordifolia on functions of peritoneal macrophages isolated from CCl4 intoxicated male albino mice

    PubMed Central

    2011-01-01

    Background The current practice of ingesting phytochemicals for supporting the immune system or fighting infections is based on centuries-old tradition. Macrophages are involved at all the stages of an immune response. The present study focuses on the immunostimulant properties of Tinospora cordifolia extract that are exerted on circulating macrophages isolated from CCl4 (0.5 ml/kg body weight) intoxicated male albino mice. Methods Apart from damaging the liver system, carbon tetrachloride also inhibits macrophage functions thus, creating an immunocompromised state, as is evident from the present study. Such cell functions include cell morphology, adhesion property, phagocytosis, enzyme release (myeloperoxidase or MPO), nitric oxide (NO) release, intracellular survival of ingested bacteria and DNA fragmentation in peritoneal macrophages isolated from these immunocompromised mice. T. cordifolia extract was tested for acute toxicity at the given dose (150 mg/kg body weight) by lactate dehydrogenase (LDH) assay. Results The number of morphologically altered macrophages was increased in mice exposed to CCl4. Administration of CCl4 (i.p.) also reduced the phagocytosis, cell adhesion, MPO release, NO release properties of circulating macrophages of mice. The DNA fragmentation of peritoneal macrophages was observed to be higher in CCl4 intoxicated mice. The bacterial killing capacity of peritoneal macrophages was also adversely affected by CCl4. However oral administration of aqueous fraction of Tinospora cordifolia stem parts at a dose of 40 mg/kg body weight (in vivo) in CCl4 exposed mice ameliorated the effect of CCl4, as the percentage of morphologically altered macrophages, phagocytosis activity, cell adhesion, MPO release, NO release, DNA fragmentation and intracellular killing capacity of CCl4 intoxicated peritoneal macrophages came closer to those of the control group. No acute toxicity was identified in oral administration of the aqueous extract of Tinospora

  6. Localization and activity of various lysosomal proteases in Leishmania amazonensis-infected macrophages.

    PubMed Central

    Prina, E; Antoine, J C; Wiederanders, B; Kirschke, H

    1990-01-01

    In mammalian hosts, Leishmania amastigotes are obligatory intracellular parasites of macrophages and multiply within parasitophorous vacuoles of phagolysosomal origin. To understand how they escape the harmful strategies developed by macrophages to kill ingested microorganisms, it is important to obtain information on the functional state of parasitophorous vacuole. For this purpose, we studied the intracellular distribution and activity of host lysosomal proteases in rat bone marrow-derived macrophages infected with Leishmania amazonensis amastigotes. Localization of cathepsins B, H, L, and D was investigated by using specific immunoglobulins. In uninfected macrophages, these enzymes were located in perinuclear granules (most of them were probably secondary lysosomes) which, after infection, disappeared progressively. In infected macrophages, cathepsins were detected mainly in the parasitophorous vacuoles, suggesting that the missing secondary lysosomes had fused with these organelles. Biochemical assays of various proteases (cathepsins B, H, and D and dipeptidyl peptidases I and II) showed that infection was accompanied by a progressive increase of all activities tested, except that of dipeptidyl peptidase II, which remained constant. No more than 1 to 10% of these activities could be attributed to amastigotes. These data indicate that (i) Leishmania infection is followed by an increased synthesis and/or a reduced catabolism of host lysosomal proteases, and (ii) amastigotes grow in a compartment rich in apparently fully active proteases. Unexpectedly, it was found that infected and uninfected macrophages degraded endocytosed proteins similarly. The lack of correlation in infected macrophages between increase of protease activities and catabolism of exogenous proteins could be linked to the huge increase in volume of the lysosomal compartment. Images PMID:2187806

  7. Interferon-γ promotes phagocytosis of Cryptococcus neoformans but not Cryptococcus gattii by murine macrophages.

    PubMed

    Ikeda-Dantsuji, Yurika; Ohno, Hideaki; Tanabe, Koichi; Umeyama, Takashi; Ueno, Keigo; Nagi, Minoru; Yamagoe, Satoshi; Kinjo, Yuki; Miyazaki, Yoshitsugu

    2015-12-01

    Among invasive fungal infections, cryptococcosis caused by inhalation of Cryptococcus neoformans or Cryptococcus gattii is particularly dangerous because it can disseminate to the central nervous system and cause life-threatening meningitis or meningoencephalitis. Previous reports described significant differences in the histopathological features of C. neoformans and C. gattii infection, such as greater pathogen proliferation and a limited macrophage response in mouse lung infected by C. gattii. To elucidate the difference in pathogenicity of these two Cryptococcus species, we investigated the interaction of C. neoformans and C. gattii with murine macrophages, the first line of host defense, by confocal laser microscopy. Only thin-capsulated, and not thick-capsulated C. neoformans and C. gattii were phagocytosed by macrophages. Preactivation with interferon-γ increased the phagocytic rate of thin-capsulated C. neoformans up to two-fold, but did not promote phagocytosis of thin-capsulated C. gattii. Lipopolysaccharide preactivation or Aspergillus fumigatus conidia co-incubation had no effect on internalization of thin-capsulated C. neoformans or C. gattii by macrophages. Phagocytosis of live thin-capsulated C. neoformans, but not that of live thin-capsulated C. gattii, induced interleukin-12 release from macrophages. However, phagocytosis of heat-killed or paraformaldehyde-fixed thin-capsulated C. neoformans did not increase IL-12 release, showing that the internalization of live yeast is important for initiating the immune response during C. neoformans-macrophage interactions. Our data suggest that macrophage response to C. gattii is limited compared with that to C. neoformans and that these results may partially explain the limited immune response and the greater pathogenicity of C. gattii.

  8. Cisplatin stimulates protein tyrosine phosphorylation in macrophages.

    PubMed

    Kumar, R; Shrivastava, A; Sodhi, A

    1995-03-01

    Cisplatin [cis-dichlorodiamine platinum (II)], a potent anti-tumor compound, stimulates immune responses by activating monocyte-macrophages and other cells of the immune system. The mechanism by which cisplatin activates these cells is poorly characterized. Since protein tyrosine phosphorylation appears to be a major intracellular signalling event that mediates cellular responses, we examined whether cisplatin alters tyrosine phosphorylation in macrophages. We found that cisplatin increased tyrosine phosphorylation of several proteins in peritoneal macrophages and in P388D1 and IC-21 macrophage cell lines. Treatment of macrophages with tyrosine kinase inhibitors, genestein and lavendustin A, inhibited cisplatin-stimulated protein tyrosine phosphorylation in macrophages. Macrophages treated with cisplatin also exhibit increased fluorescence with anti-phosphotyrosine-FITC antibody. These data indicate that protein tyrosine phosphorylation plays a role in cisplatin-induced activation of macrophages. PMID:7539662

  9. Phenotypic Transitions of Macrophages Orchestrate Tissue Repair

    PubMed Central

    Novak, Margaret L.; Koh, Timothy J.

    2014-01-01

    Macrophages are essential for the efficient healing of numerous tissues, and they contribute to impaired healing and fibrosis. Tissue repair proceeds through overlapping phases of inflammation, proliferation, and remodeling, and macrophages are present throughout this progression. Macrophages exhibit transitions in phenotype and function as tissue repair progresses, although the precise factors regulating these transitions remain poorly defined. In efficiently healing injuries, macrophages present during a given stage of repair appear to orchestrate transition into the next phase and, in turn, can promote debridement of the injury site, cell proliferation and angiogenesis, collagen deposition, and matrix remodeling. However, dysregulated macrophage function can contribute to failure to heal or fibrosis in several pathological situations. This review will address current knowledge of the origins and functions of macrophages during the progression of tissue repair, with emphasis on skin and skeletal muscle. Dysregulation of macrophages in disease states and therapies targeting macrophage activation to promote tissue repair are also discussed. PMID:24091222

  10. Macrophage-induced thymic lymphocyte maturation.

    PubMed Central

    Van den Tweel, J G; Walker, W S

    1977-01-01

    Guinea-pig peritoneal macrophages were found to influence the functional maturation of thymic lymphocytes. Autologous thymic lymphocytes obtained from macrophage co-cultures responded to three different mitogens and were reduced in their ability to reassociate spontaneously with macrophages. Neither of these properties were found in thymic lymphocytes that had not been cultured with macrophages. These functional changes appeared to be specific for macrophages since thymic lymphocytes incubated with skin fibroblasts failed to respond to the test mitogens. Furthermore, they were not the result of either the inactivation, by macrophages, of a putative suppressor thymocyte or a soluble macrophage product. In addition to influencing the functional maturation of thymic lymphocytes, macrophages also appeared to play a direct role in inducing the mitogen response of functionally mature cells. PMID:304037

  11. Isolation and culture of murine macrophages.

    PubMed

    Davies, John Q; Gordon, Siamon

    2005-01-01

    The two most convenient sources of primary murine macrophages are the bone marrow and the peritoneal cavity. Resident peritoneal macrophages can readily be harvested from mice and purified by adherence to tissue culture plastic. The injection of Bio-Gel polyacrylamide beads or thioglycollate broth into the peritoneal cavity produces an inflammatory response allowing the purification of large numbers of elicited macrophages. The production of an activated macrophage population can be achieved by using Bacillus-Calmette-Guerin as the inflammatory stimulus. Resident bone marrow macrophages can be isolated following enzymatic separation of cells from bone marrow plugs and enrichment on 30% fetal calf serum containing medium or Ficoll-Hypaque gradients. Bone marrow-derived macrophages can be produced by differentiating nonadherent macrophage precursors with medium containing macrophage colony-stimulating factor.

  12. Macrophages and HIV-1: An Unhealthy Constellation.

    PubMed

    Sattentau, Quentin J; Stevenson, Mario

    2016-03-01

    Lentiviruses have a long-documented association with macrophages. Abundant evidence exists for in vitro and, in a tissue-specific manner, in vivo infection of macrophages by the primate lentiviruses HIV-1 and SIV. However, macrophage contribution to aspects of HIV-1 and SIV pathogenesis, and their role in viral persistence in individuals on suppressive antiretroviral therapy, remains unclear. Here we discuss recent evidence implicating macrophages in HIV-1-mediated disease and highlight directions for further investigation.

  13. Macrophage Heterogeneity and Plasticity: Impact of Macrophage Biomarkers on Atherosclerosis

    PubMed Central

    Rojas, Joselyn; Salazar, Juan; Martínez, María Sofía; Palmar, Jim; Bautista, Jordan; Chávez-Castillo, Mervin; Gómez, Alexis; Bermúdez, Valmore

    2015-01-01

    Cardiovascular disease (CVD) is a global epidemic, currently representing the worldwide leading cause of morbidity and mortality. Atherosclerosis is the fundamental pathophysiologic component of CVD, where the immune system plays an essential role. Monocytes and macrophages are key mediators in this aspect: due to their heterogeneity and plasticity, these cells may act as either pro- or anti-inflammatory mediators. Indeed, monocytes may develop heterogeneous functional phenotypes depending on the predominating pro- or anti-inflammatory microenvironment within the lesion, resulting in classic, intermediate, and non-classic monocytes, each with strikingly differing features. Similarly, macrophages may also adopt heterogeneous profiles being mainly M1 and M2, the former showing a proinflammatory profile while the latter demonstrates anti-inflammatory traits; they are further subdivided in several subtypes with more specialized functions. Furthermore, macrophages may display plasticity by dynamically shifting between phenotypes in response to specific signals. Each of these distinct cell profiles is associated with diverse biomarkers which may be exploited for therapeutic intervention, including IL-10, IL-13, PPAR-γ, LXR, NLRP3 inflammasomes, and microRNAs. Direct modulation of the molecular pathways concerning these potential macrophage-related targets represents a promising field for new therapeutic alternatives in atherosclerosis and CVD. PMID:26491604

  14. Investigation of macrophage polarization using bone marrow derived macrophages.

    PubMed

    Ying, Wei; Cheruku, Patali S; Bazer, Fuller W; Safe, Stephen H; Zhou, Beiyan

    2013-06-23

    The article describes a readily easy adaptive in vitro model to investigate macrophage polarization. In the presence of GM-CSF/M-CSF, hematopoietic stem/progenitor cells from the bone marrow are directed into monocytic differentiation, followed by M1 or M2 stimulation. The activation status can be tracked by changes in cell surface antigens, gene expression and cell signaling pathways.

  15. Caspase-1 activation in macrophages infected with Yersinia pestis KIM requires the type III secretion system effector YopJ.

    PubMed

    Lilo, Sarit; Zheng, Ying; Bliska, James B

    2008-09-01

    Pathogenic Yersinia species utilize a type III secretion system (T3SS) to translocate effectors called Yersinia outer proteins (Yops) into infected host cells. Previous studies demonstrated a role for effector Yops in the inhibition of caspase-1-mediated cell death and secretion of interleukin-1beta (IL-1beta) in naïve macrophages infected with Yersinia enterocolitica. Naïve murine macrophages were infected with a panel of different Yersinia pestis and Yersinia pseudotuberculosis strains to determine whether Yops of these species inhibit caspase-1 activation. Cell death was measured by release of lactate dehydrogenase (LDH), and enzyme-linked immunosorbent assay for secreted IL-1beta was used to measure caspase-1 activation. Surprisingly, isolates derived from the Y. pestis KIM strain (e.g., KIM5) displayed an unusual ability to activate caspase-1 and kill infected macrophages compared to other Y. pestis and Y. pseudotuberculosis strains tested. Secretion of IL-1beta following KIM5 infection was reduced in caspase-1-deficient macrophages compared to wild-type macrophages. However, release of LDH was not reduced in caspase-1-deficient macrophages, indicating that cell death occurred independently of caspase-1. Analysis of KIM-derived strains defective for production of functional effector or translocator Yops indicated that translocation of catalytically active YopJ into macrophages was required for caspase-1 activation and cell death. Release of LDH and secretion of IL-1beta were not reduced when actin polymerization was inhibited in KIM5-infected macrophages, indicating that extracellular bacteria translocating YopJ could trigger cell death and caspase-1 activation. This study uncovered a novel role for YopJ in the activation of caspase-1 in macrophages.

  16. Caspase-1 Activation in Macrophages Infected with Yersinia pestis KIM Requires the Type III Secretion System Effector YopJ▿

    PubMed Central

    Lilo, Sarit; Zheng, Ying; Bliska, James B.

    2008-01-01

    Pathogenic Yersinia species utilize a type III secretion system (T3SS) to translocate effectors called Yersinia outer proteins (Yops) into infected host cells. Previous studies demonstrated a role for effector Yops in the inhibition of caspase-1-mediated cell death and secretion of interleukin-1β (IL-1β) in naïve macrophages infected with Yersinia enterocolitica. Naïve murine macrophages were infected with a panel of different Yersinia pestis and Yersinia pseudotuberculosis strains to determine whether Yops of these species inhibit caspase-1 activation. Cell death was measured by release of lactate dehydrogenase (LDH), and enzyme-linked immunosorbent assay for secreted IL-1β was used to measure caspase-1 activation. Surprisingly, isolates derived from the Y. pestis KIM strain (e.g., KIM5) displayed an unusual ability to activate caspase-1 and kill infected macrophages compared to other Y. pestis and Y. pseudotuberculosis strains tested. Secretion of IL-1β following KIM5 infection was reduced in caspase-1-deficient macrophages compared to wild-type macrophages. However, release of LDH was not reduced in caspase-1-deficient macrophages, indicating that cell death occurred independently of caspase-1. Analysis of KIM-derived strains defective for production of functional effector or translocator Yops indicated that translocation of catalytically active YopJ into macrophages was required for caspase-1 activation and cell death. Release of LDH and secretion of IL-1β were not reduced when actin polymerization was inhibited in KIM5-infected macrophages, indicating that extracellular bacteria translocating YopJ could trigger cell death and caspase-1 activation. This study uncovered a novel role for YopJ in the activation of caspase-1 in macrophages. PMID:18559430

  17. Heterosigma bloom and associated fish kill

    USGS Publications Warehouse

    Hershberger, P.K.; Rensel, J.E.; Postel, J.R.; Taub, F.B.

    1997-01-01

    A bloom of the harmful marine phytoplankton, Heterosigma carterae occurred in upper Case Inlet, south Puget Sound, Washington in late September, 1994, correlating with the presence of at least 35 dead salmon. This marks the first time that this alga has been closely correlated with a wild fish kill; in the past it was thought to be associated with kills of penned fish at fish farms only. We were informed of the presence of a possible harmful algal bloom and dead salinois Ilear the town of Allyn on 27 September and a team was formed to investigate. We arrived at the Allyn waterfront at 17:30 hours the same day. Prior to our arrival, state agency personnel walked approximatcly two miles of shoreline from the powerlines north of the dock, to the mouth of Sherwood Creek and conducted the only official count of dead fish present along the shore consisting of 12 coho salmon (Oncorhynchus kisutch), 11 chum salmon (Oncorhynchus keta), 12 chinook salmon (Oncorhynchus tschawytscha), one flat fish, and one sculpin on the morning of 9/27. Since previous harmful blooms of Heterosigma have resultedin the majority of net penreared salmon sinking to the bottom of pens, and only approximately two miles of shoreline were sampled, it is suspected that many more exposed fish may have succumbed than were counted. Witnesses who explored the east side of the bay reported seeing many dead salmon there as well, but no counts were made. State agency personnel who observed the fish kill reported seeing “dying fish coming to the beach, gulping at the surface, trying to get out of the water” Scavengers were seen consuming the salmon carcasses; these included two harbor seals, a house cat, and Hymenopteran insects. None suffered any noticeable acute ill effects. Although precise cause of death has not been ascertained, visual inspection of the reproductive organs from a deceased male chum salmon found on the shore at Allyn confirmed that the fish was not yet reproductively mature and

  18. Tissue macrophage identity and self-renewal.

    PubMed

    Gentek, Rebecca; Molawi, Kaaweh; Sieweke, Michael H

    2014-11-01

    Macrophages are cellular components of the innate immune system that reside in virtually all tissues and contribute to immunity, repair, and homeostasis. The traditional view that all tissue-resident macrophages derive from the bone marrow through circulating monocyte intermediates has dramatically shifted recently with the observation that macrophages from embryonic progenitors can persist into adulthood and self-maintain by local proliferation. In several tissues, however, monocytes also contribute to the resident macrophage population, on which the local environment can impose tissue-specific macrophage functions. These observations have raised important questions: What determines resident macrophage identity and function, ontogeny or environment? How is macrophage proliferation regulated? In this review, we summarize the current knowledge about the identity, proliferation, and turnover of tissue-resident macrophages and how they differ from freshly recruited short-lived monocyte-derived cells. We examine whether macrophage proliferation can be qualified as self-renewal of mature differentiated cells and whether the concepts and molecular pathways are comparable to self-renewal mechanisms in stem cells. Finally, we discuss how improved understanding of macrophage identity and self-renewal could be exploited for therapeutic intervention of macrophage-mediated pathologies by selectively targeting freshly recruited or resident macrophages.

  19. Stepwise cytoskeletal polarization as a series of checkpoints in innate but not adaptive cytolytic killing

    NASA Astrophysics Data System (ADS)

    Wülfing, Christoph; Purtic, Bozidar; Klem, Jennifer; Schatzle, John D.

    2003-06-01

    Cytolytic killing is a major effector mechanism in the elimination of virally infected and tumor cells. The innate cytolytic effectors, natural killer (NK) cells, and the adaptive effectors, cytotoxic T cells (CTL), despite differential immune recognition, both use the same lytic mechanism, cytolytic granule release. Using live cell video fluorescence microscopy in various primary cell models of NK cell and CTL killing, we show here that on tight target cell contact, a majority of the NK cells established cytoskeletal polarity required for effective lytic function slowly or incompletely. In contrast, CTLs established cytoskeletal polarity rapidly. In addition, NK cell killing was uniquely sensitive to minor interference with cytoskeletal dynamics. We propose that the stepwise NK cell cytoskeletal polarization constitutes a series of checkpoints in NK cell killing. In addition, the use of more deliberate progression to effector function to compensate for inferior immune recognition specificity provides a mechanistic explanation for how the same effector function can be used in the different functional contexts of the innate and adaptive immune response.

  20. Protection of Candida parapsilosis from neutrophil killing through internalization by human endothelial cells

    PubMed Central

    Glass, Kyle A; Longley, Sarah J; Bliss, Joseph M; Shaw, Sunil K

    2015-01-01

    Candida parapsilosis is a fungal pathogen that is associated with hematogenously disseminated disease in premature neonates, acutely ill or immunocompromised patients. In cell culture, C. parapsilosis cells are actively and avidly endocytosed by endothelial cells via actin polymerization mediated by N-WASP. Here we present evidence that C. parapsilosis that were internalized by endothelial cells remained alive, and avoided being acidified or otherwise damaged via the host cell. Internalized fungal cells reproduced intracellularly and eventually burst out of the host endothelial cell. When neutrophils were added to endothelium and C. parapsilosis, they patrolled the endothelial surface and efficiently killed most adherent fungal cells prior to endocytosis. But after endocytosis by endothelial cells, internalized fungal cells evaded neutrophil killing. Silencing endothelial N-WASP blocked endocytosis of C. parapsilosis and left fungal cells stranded on the cell surface, where they were susceptible to neutrophil killing. These observations suggest that for C. parapsilosis to escape from the bloodstream, fungi may adhere to and be internalized by endothelial cells before being confronted and phagocytosed by a patrolling leukocyte. Once internalized by endothelial cells, C. parapsilosis may safely replicate to cause further rounds of infection. Immunosurveillance of the intravascular lumen by leukocytes crawling on the endothelial surface and rapid killing of adherent yeast may play a major role in controlling C. parapsilosis dissemination and infected endothelial cells may be a significant reservoir for fungal persistence. PMID:26039751

  1. In Vivo Killing Capacity of Cytotoxic T Cells Is Limited and Involves Dynamic Interactions and T Cell Cooperativity.

    PubMed

    Halle, Stephan; Keyser, Kirsten Anja; Stahl, Felix Rolf; Busche, Andreas; Marquardt, Anja; Zheng, Xiang; Galla, Melanie; Heissmeyer, Vigo; Heller, Katrin; Boelter, Jasmin; Wagner, Karen; Bischoff, Yvonne; Martens, Rieke; Braun, Asolina; Werth, Kathrin; Uvarovskii, Alexey; Kempf, Harald; Meyer-Hermann, Michael; Arens, Ramon; Kremer, Melanie; Sutter, Gerd; Messerle, Martin; Förster, Reinhold

    2016-02-16

    According to in vitro assays, T cells are thought to kill rapidly and efficiently, but the efficacy and dynamics of cytotoxic T lymphocyte (CTL)-mediated killing of virus-infected cells in vivo remains elusive. We used two-photon microscopy to quantify CTL-mediated killing in mice infected with herpesviruses or poxviruses. On average, one CTL killed 2-16 virus-infected cells per day as determined by real-time imaging and by mathematical modeling. In contrast, upon virus-induced MHC class I downmodulation, CTLs failed to destroy their targets. During killing, CTLs remained migratory and formed motile kinapses rather than static synapses with targets. Viruses encoding the calcium sensor GCaMP6s revealed strong heterogeneity in individual CTL functional capacity. Furthermore, the probability of death of infected cells increased for those contacted by more than two CTLs, indicative of CTL cooperation. Thus, direct visualization of CTLs during killing of virus-infected cells reveals crucial parameters of CD8(+) T cell immunity. PMID:26872694

  2. In Vivo Killing Capacity of Cytotoxic T Cells Is Limited and Involves Dynamic Interactions and T Cell Cooperativity

    PubMed Central

    Halle, Stephan; Keyser, Kirsten Anja; Stahl, Felix Rolf; Busche, Andreas; Marquardt, Anja; Zheng, Xiang; Galla, Melanie; Heissmeyer, Vigo; Heller, Katrin; Boelter, Jasmin; Wagner, Karen; Bischoff, Yvonne; Martens, Rieke; Braun, Asolina; Werth, Kathrin; Uvarovskii, Alexey; Kempf, Harald; Meyer-Hermann, Michael; Arens, Ramon; Kremer, Melanie; Sutter, Gerd; Messerle, Martin; Förster, Reinhold

    2016-01-01

    Summary According to in vitro assays, T cells are thought to kill rapidly and efficiently, but the efficacy and dynamics of cytotoxic T lymphocyte (CTL)-mediated killing of virus-infected cells in vivo remains elusive. We used two-photon microscopy to quantify CTL-mediated killing in mice infected with herpesviruses or poxviruses. On average, one CTL killed 2–16 virus-infected cells per day as determined by real-time imaging and by mathematical modeling. In contrast, upon virus-induced MHC class I downmodulation, CTLs failed to destroy their targets. During killing, CTLs remained migratory and formed motile kinapses rather than static synapses with targets. Viruses encoding the calcium sensor GCaMP6s revealed strong heterogeneity in individual CTL functional capacity. Furthermore, the probability of death of infected cells increased for those contacted by more than two CTLs, indicative of CTL cooperation. Thus, direct visualization of CTLs during killing of virus-infected cells reveals crucial parameters of CD8+ T cell immunity. PMID:26872694

  3. Female resistance to pneumonia identifies lung macrophage nitric oxide synthase-3 as a therapeutic target

    PubMed Central

    Yang, Zhiping; Huang, Yuh-Chin T; Koziel, Henry; de Crom, Rini; Ruetten, Hartmut; Wohlfart, Paulus; Thomsen, Reimar W; Kahlert, Johnny A; Sørensen, Henrik Toft; Jozefowski, Szczepan; Colby, Amy; Kobzik, Lester

    2014-01-01

    To identify new approaches to enhance innate immunity to bacterial pneumonia, we investigated the natural experiment of gender differences in resistance to infections. Female and estrogen-treated male mice show greater resistance to pneumococcal pneumonia, seen as greater bacterial clearance, diminished lung inflammation, and better survival. In vitro, lung macrophages from female mice and humans show better killing of ingested bacteria. Inhibitors and genetically altered mice identify a critical role for estrogen-mediated activation of lung macrophage nitric oxide synthase-3 (NOS3). Epidemiologic data show decreased hospitalization for pneumonia in women receiving estrogen or statins (known to activate NOS3). Pharmacologic targeting of NOS3 with statins or another small-molecule compound (AVE3085) enhanced macrophage bacterial killing, improved bacterial clearance, and increased host survival in both primary and secondary (post-influenza) pneumonia. The data identify a novel mechanism for host defense via NOS3 and suggest a potential therapeutic strategy to reduce secondary bacterial pneumonia after influenza. DOI: http://dx.doi.org/10.7554/eLife.03711.001 PMID:25317947

  4. Induction of Macrophage Function in Human THP-1 Cells Is Associated with Rewiring of MAPK Signaling and Activation of MAP3K7 (TAK1) Protein Kinase

    PubMed Central

    Richter, Erik; Ventz, Katharina; Harms, Manuela; Mostertz, Jörg; Hochgräfe, Falko

    2016-01-01

    Macrophages represent the primary human host response to pathogen infection and link the immediate defense to the adaptive immune system. Mature tissue macrophages convert from circulating monocyte precursor cells by terminal differentiation in a process that is not fully understood. Here, we analyzed the protein kinases of the human monocytic cell line THP-1 before and after induction of macrophage differentiation by using kinomics and phosphoproteomics. When comparing the macrophage-like state with the monocytic precursor, 50% of the kinome was altered in expression and even 71% of covered kinase phosphorylation sites were affected. Kinome rearrangements are for example characterized by a shift of overrepresented cyclin-dependent kinases associated with cell cycle control in monocytes to calmodulin-dependent kinases and kinases involved in proinflammatory signaling. Eventually, we show that monocyte-to-macrophage differentiation is associated with major rewiring of mitogen-activated protein kinase signaling networks and demonstrate that protein kinase MAP3K7 (TAK1) acts as the key signaling hub in bacterial killing, chemokine production and differentiation. Our study proves the fundamental role of protein kinases and cellular signaling as major drivers of macrophage differentiation and function. The finding that MAP3K7 is central to macrophage function suggests MAP3K7 and its networking partners as promising targets in host-directed therapy for macrophage-associated disease. PMID:27066479

  5. The eyeball killer: serial killings with postmortem globe enucleation.

    PubMed

    Coyle, Julie; Ross, Karen F; Barnard, Jeffrey J; Peacock, Elizabeth; Linch, Charles A; Prahlow, Joseph A

    2015-05-01

    Although serial killings are relatively rare, they can be the cause of a great deal of anxiety while the killer remains at-large. Despite the fact that the motivations for serial killings are typically quite complex, the psychological analysis of a serial killer can provide valuable insight into how and why certain individuals become serial killers. Such knowledge may be instrumental in preventing future serial killings or in solving ongoing cases. In certain serial killings, the various incidents have a variety of similar features. Identification of similarities between separate homicidal incidents is necessary to recognize that a serial killer may be actively killing. In this report, the authors present a group of serial killings involving three prostitutes who were shot to death over a 3-month period. Scene and autopsy findings, including the unusual finding of postmortem enucleation of the eyes, led investigators to recognize the serial nature of the homicides.

  6. The eyeball killer: serial killings with postmortem globe enucleation.

    PubMed

    Coyle, Julie; Ross, Karen F; Barnard, Jeffrey J; Peacock, Elizabeth; Linch, Charles A; Prahlow, Joseph A

    2015-05-01

    Although serial killings are relatively rare, they can be the cause of a great deal of anxiety while the killer remains at-large. Despite the fact that the motivations for serial killings are typically quite complex, the psychological analysis of a serial killer can provide valuable insight into how and why certain individuals become serial killers. Such knowledge may be instrumental in preventing future serial killings or in solving ongoing cases. In certain serial killings, the various incidents have a variety of similar features. Identification of similarities between separate homicidal incidents is necessary to recognize that a serial killer may be actively killing. In this report, the authors present a group of serial killings involving three prostitutes who were shot to death over a 3-month period. Scene and autopsy findings, including the unusual finding of postmortem enucleation of the eyes, led investigators to recognize the serial nature of the homicides. PMID:25682709

  7. Killing, letting die and moral perception.

    PubMed

    Gillett, Grant

    1994-10-01

    There are a number of arguments that purport to show, in general terms, that there is no difference between killing and letting die. These are used to justify active euthanasia on the basis of the reasons given for allowing patients to die. I argue that the general and abstract arguments fail to take account of the complex and particular situations which are found in the care of those with terminal illness. When in such situations, there are perceptions and intuitions available that do not easily find propositional form but lead most of those whose practice is in the care of the dying to resist active euthanasia. I make a plea for their intuitions to be heeded above the sterile voice of abstract premises and arguments by examining the completeness of the outline form of the pro-euthanasia argument. In doing so, I make use of Nussbaum's discussion of moral perception and general claims to be found in the literature of moral particularism.

  8. f(R)-gravity from Killing tensors

    NASA Astrophysics Data System (ADS)

    Paliathanasis, Andronikos

    2016-04-01

    We consider f(R)-gravity in a Friedmann-Lemaître-Robertson-Walker spacetime with zero spatial curvature. We apply the Killing tensors of the minisuperspace in order to specify the functional form of f(R) and for the field equations to be invariant under Lie-Bäcklund transformations, which are linear in momentum (contact symmetries). Consequently, the field equations to admit quadratic conservation laws given by Noether’s theorem. We find three new integrable f(R)-models, for which, with the application of the conservation laws, we reduce the field equations to a system of two first-order ordinary differential equations. For each model we study the evolution of the cosmological fluid. We find that for each integrable model the cosmological fluid has an equation of state parameter, in which there is linear behavior in terms of the scale factor which describes the Chevallier, Polarski and Linder parametric dark energy model.

  9. Killed poliovirus antigen titration in humans.

    PubMed

    Salk, J; Cohen, H; Fillastre, C; Stoeckel, P; Rey, J L; Schlumberger, M; Nicolas, A; van Steenis, G; van Wezel, A L; Triau, R; Saliou, P; Barry, L F; Moreau, J P; Mérieux, C

    1978-01-01

    To establish the antigen content of a killed poliovirus vaccine sufficiently potent to induce immunity with one or two doses and to establish a reference standard vaccine which has been tested under field conditions, a titration was carried out in infants to determine the amount of each of the three antigenic types of poliovirus vaccine required to induce seroconversion with a single dose. It has been observed that over a critical range of antigen concentration there is an essentially linear relationship between antibody response and quantity of antigen administered. More than 90 percent of the groups studied had detectable antibody after receiving single injections of 80, 8 and 64 D-antigen units of Types I, II and III, respectively. Four-fold less antigen for each of the three types was less effective. The implications of these findings for an efficient immunization procedure are discussed.

  10. Advanced gel propulsion controls for kill vehicles

    NASA Astrophysics Data System (ADS)

    Yasuhara, W. K.; Olson, A.; Finato, S.

    1993-06-01

    A gel propulsion control concept for tactical applications is reviewed, and the status of the individual component technologies currently under development at the Aerojet Propulsion Division is discussed. It is concluded that a gel propellant Divert and Attitude Control Subsystem (DACS) provides a safe, insensitive munitions compliant alternative to current liquid Theater Missile Defense (TMD) DACS approaches. The gel kill vehicle (KV) control system packages a total impulse typical of a tactical weapon interceptor for the ground- or sea-based TMD systems. High density packaging makes it possible to increase firepower and to eliminate long-term high pressure gas storage associated with bipropellant systems. The integrated control subsystem technologies encompass solid propellant gas generators, insulated composite overwrapped propellant tanks, lightweight endoatmospheric thrusters, and insensitive munition gel propellants, which meet the requirements of a deployable, operationally safe KV.

  11. Self-forces from generalized Killing fields

    NASA Astrophysics Data System (ADS)

    Harte, Abraham I.

    2008-12-01

    A non-perturbative formalism is developed that simplifies the understanding of self-forces and self-torques acting on extended scalar charges in curved spacetimes. Laws of motion are locally derived using momenta generated by a set of generalized Killing fields. Self-interactions that may be interpreted as arising from the details of a body's internal structure are shown to have very simple geometric and physical interpretations. Certain modifications to the usual definition for a center-of-mass are identified that significantly simplify the motions of charges with strong self-fields. A derivation is also provided for a generalized form of the Detweiler Whiting axiom that pointlike charges should react only to the so-called regular component of their self-field. Standard results are shown to be recovered for sufficiently small charge distributions.

  12. Transverse conformal Killing forms on Kähler foliations

    NASA Astrophysics Data System (ADS)

    Jung, Seoung Dal

    2015-04-01

    On a closed, connected Riemannian manifold with a Kähler foliation of codimension q = 2 m, any transverse Killing r(≥ 2) -form is parallel (Jung and Jung, 2012). In this paper, we study transverse conformal Killing forms on Kähler foliations. In fact, if the foliation is minimal, then for any transverse conformal Killing r-form ϕ(2 ≤ r ≤ q - 2), Jϕ is parallel. Here J is defined in Section 4.

  13. Killing Sections and Sigma Models with Lie Algebroid Targets

    NASA Astrophysics Data System (ADS)

    Bruce, Andrew James

    2016-08-01

    We define and examine the notion of a Killing section of a Riemannian Lie algebroid as a natural generalisation of a Killing vector field. We show that the various expression for a vector field to be Killing naturally generalise to the setting of Lie algebroids. As an application we examine the internal symmetries of a class of sigma models for which the target space is a Riemannian Lie algebroid. Critical points of these sigma models are interpreted as generalised harmonic maps.

  14. Toxin-induced resistance in Bacillus anthracis lethal toxin-treated macrophages

    PubMed Central

    Salles, Isabelle I.; Tucker, Amy E.; Voth, Daniel E.; Ballard, Jimmy D.

    2003-01-01

    In the current study, we show that macrophages adaptively resist anthrax lethal toxin (LT) through a toxin-activated process termed toxin-induced resistance (TIR). TIR was triggered by pretreatment of RAW 264.7 or J774A.1 macrophages with a low dose of LT for at least 6 h, which resulted in resistance to high doses of LT for 96 h. Activation of TIR required functional toxin, because LT subunits, mutants, and heat-inactivated toxin were unable to trigger resistance. TIR macrophages were not altered in toxin receptor levels or cell cycle profiles. Treatment of TIR macrophages with high doses of LT resulted in a sustained decline in full-length mitogen-activated protein kinase kinase 2, a known target of lethal factor, and a marked reduction in diphosphorylated extracellular response kinases 1,2 for 24 h. However, despite the sustained loss of full-length mitogen-activated protein kinase kinase 2, by 48 h, TIR macrophages regained diphosphorylated extracellular response kinases 1,2, suggesting an adaptation led to recovery of this signaling pathway. TIR macrophages were also able to maintain normal levels of ubiquitinylated proteins, whereas sensitive cells show a rapid reduction in ubiquitin-modified proteins before cell death, indicating a possible alteration in proteasome activity contributed to resistance. These results provide a paradigm for toxin-cell interactions and suggest macrophages are capable of adapting to and tolerating toxic doses of LT. PMID:14519843

  15. TLR Stimulation Dynamically Regulates Heme and Iron Export Gene Expression in Macrophages

    PubMed Central

    Philip, Mary; Chiu, Edison Y.; Hajjar, Adeline M.; Abkowitz, Janis L.

    2016-01-01

    Pathogenic bacteria have evolved multiple mechanisms to capture iron or iron-containing heme from host tissues or blood. In response, organisms have developed defense mechanisms to keep iron from pathogens. Very little of the body's iron store is available as free heme; rather nearly all body iron is complexed with heme or other proteins. The feline leukemia virus, subgroup C (FeLV-C) receptor, FLVCR, exports heme from cells. It was unknown whether FLVCR regulates heme-iron availability after infection, but given that other heme regulatory proteins are upregulated in macrophages in response to bacterial infection, we hypothesized that macrophages dynamically regulate FLVCR. We stimulated murine primary macrophages or macrophage cell lines with LPS and found that Flvcr is rapidly downregulated in a TLR4/MD2-dependent manner; TLR1/2 and TLR3 stimulation also decreased Flvcr expression. We identified several candidate TLR-activated transcription factors that can bind to the Flvcr promoter. Macrophages must balance the need to sequester iron from systemic circulating or intracellular pathogens with the macrophage requirement for heme and iron to produce reactive oxygen species. Our findings underscore the complexity of this regulation and point to a new role for FLVCR and heme export in macrophages responses to infection and inflammation. PMID:27006955

  16. Macrophage depletion reduces postsurgical tumor recurrence and metastatic growth in a spontaneous murine model of melanoma

    PubMed Central

    Tham, Muly; Khoo, Karen; Yeo, Kim Pin; Kato, Masashi; Prevost-Blondel, Amelle; Angeli, Veronique; Abastado, Jean-Pierre

    2015-01-01

    Surgical resection of tumors is often followed by regrowth at the primary site and metastases may emerge rapidly following removal of the primary tumor. Macrophages are important drivers of tumor growth, and here we investigated their involvement in postoperative relapse as well as explore macrophage depletion as an adjuvant to surgical resection. RETAAD mice develop spontaneous metastatic melanoma that begins in the eye. Removal of the eyes as early as 1 week of age did not prevent the development of metastases; rather, surgery led to increased proliferation of tumor cells locally and in distant metastases. Surgery-induced increase in tumor cell proliferation correlated with increased macrophage density within the tumor. Moreover, macrophages stimulate tumor sphere formation from tumor cells of post-surgical but not control mice. Macrophage depletion with a diet containing the CSF-1R specific kinase inhibitor Ki20227 following surgery significantly reduced postoperative tumor recurrence and abrogated enhanced metastatic outgrowth. Our results confirm that tumor cells disseminate early, and show that macrophages contribute both to post-surgical tumor relapse and growth of metastases, likely through stimulating a population of tumor-initiating cells. Thus macrophage depletion warrants exploration as an adjuvant to surgical resection. PMID:25762633

  17. Interaction of Mycoplasma dispar with bovine alveolar macrophages.

    PubMed Central

    Almeida, R A; Wannemuehler, M J; Rosenbusch, R F

    1992-01-01

    The capacity to avoid phagocytosis and the activation of bovine alveolar macrophages (BAM) by encapsulated Mycoplasma dispar or purified M. dispar capsule was investigated. Encapsulated and unencapsulated M. dispar were cocultured with BAM in the presence or absence of antisera prepared against unencapsulated M. dispar or purified capsule antiserum. Unopsonized mycoplasmas resisted phagocytosis, while only anti-capsule antibodies enhanced the phagocytosis of encapsulated mycoplasmas. BAM were cultured in the presence of purified M. dispar capsule or either live or heat-killed encapsulated or unencapsulated M. dispar. These BAM were then activated with Escherichia coli endotoxin or left without further activation. The supernatants of these cultures were assayed for tumor necrosis factor, interleukin 1, and glucose consumption as indicators of macrophage activation. Tumor necrosis factor and interleukin 1 were produced by BAM stimulated with unencapsulated M. dispar but not when encapsulated M. dispar or its purified capsule was used. Similarly, glucose consumption was increased in the presence of unencapsulated M. dispar, but not when BAM were cocultured with encapsulated M. dispar or purified capsule. When BAM were treated with purified capsule or encapsulated mycoplasmas, they could not be subsequently activated by endotoxin. These results indicate that encapsulated M. dispar or purified capsule exerts an inhibitory effect on the activity of BAM and prevents the activation of these cells. PMID:1612758

  18. Isoniazid (INH) treatment of INH-resistant Mycobacterium tuberculosis inhibits infected macrophage to produce TNF-alpha.

    PubMed

    Wibawa, Tri; Pangemanan, Lisa; Rachmawaty, Farida J; Rintiswati, Ning; Mustofa; Soesatyo, Marsetyawan H N E

    2014-09-01

    Macrophages undergo apoptosis after infected by Mycobacterium tuber- culosis (M.tb), which is regulated by tumor necrosis factor alpha (TNF-alpha) and has a direct correlation with killing of intracellular bacilli. In order to clarify the role of isoniazid (INH) in the modulation of macrophages apoptosis and intracellular bacilli replication, we performed the following studies using an INH-resistant clinical M.tb isolate (INHres). Macrophages derived from peripheral blood were infected with INHres and treated or not treated with INH. Apoptosis was measured using an Ag-capture ELISA for histone and fragmented DNA. Production of TNF-alpha by INHres infected was assayed using ELISA and viability of intracellular M.tb was determined using bacterial culture of macrophage lysates on Lowenstein-Jensen (LJ) medium. INH pre-treatment to INHres reduced macrophages apoptosis, production of TNF-alpha and intracellular INHres viability. This study indicated that INH affected TNF-alpha release resulting in reduction of the extent macrophages apoptosis and of intracellular INH-resistant M.tb viability.

  19. Gold nanorods as a theranostic platform for in vitro and in vivo imaging and photothermal therapy of inflammatory macrophages.

    PubMed

    Qin, Jinbao; Peng, Zhiyou; Li, Bo; Ye, Kaichuang; Zhang, Yuxin; Yuan, Fukang; Yang, Xinrui; Huang, Lijia; Hu, Junqing; Lu, Xinwu

    2015-09-01

    Inflammatory macrophages play pivotal roles in the development of atherosclerosis. Theranostics, a promising approach for local imaging and photothermal therapy of inflammatory macrophages, has drawn increasing attention in biomedical research. In this study, gold nanorods (Au NRs) were synthesized, and their in vitro photothermal effects on the macrophage cell line (Ana-1 cells) under 808 nm near infrared reflection (NIR) were investigated by the CCK8 assay, calcein AM/PI staining, flow cytometry, transmission electron microscopy (TEM), silver staining and in vitro micro-computed tomography (CT) imaging. These Au NRs were then applied to an apolipoprotein E knockout (Apo E) mouse model to evaluate their effects on in vivo CT imaging and their effectiveness as for the subsequent photothermal therapy of macrophages in femoral artery restenosis under 808 nm laser irradiation. In vitro photothermal ablation treatment using Au NRs exhibited a significant cell-killing efficacy of macrophages, even at relatively low concentrations of Au NRs and low NIR powers. In addition, the in vivo results demonstrated that the Au NRs are effective for in vivo imaging and photothermal therapy of inflammatory macrophages in femoral artery restenosis. This study shows that Au nanorods are a promising theranostic platform for the diagnosis and photothermal therapy of inflammation-associated diseases. PMID:26228112

  20. Advancements in dynamic kill calculations for blowout wells

    SciTech Connect

    Kouba, G.E. . Production Fluids Div.); MacDougall, G.R. ); Schumacher, B.W. . Information Technology Dept.)

    1993-09-01

    This paper addresses the development, interpretation, and use of dynamic kill equations. To this end, three simple calculation techniques are developed for determining the minimum dynamic kill rate. Two techniques contain only single-phase calculations and are independent of reservoir inflow performance. Despite these limitations, these two methods are useful for bracketing the minimum flow rates necessary to kill a blowing well. For the third technique, a simplified mechanistic multiphase-flow model is used to determine a most-probable minimum kill rate.

  1. Antibacterial activity of silver-killed bacteria: the "zombies" effect

    NASA Astrophysics Data System (ADS)

    Wakshlak, Racheli Ben-Knaz; Pedahzur, Rami; Avnir, David

    2015-04-01

    We report a previously unrecognized mechanism for the prolonged action of biocidal agents, which we denote as the zombies effect: biocidally-killed bacteria are capable of killing living bacteria. The concept is demonstrated by first killing Pseudomonas aeruginosa PAO1 with silver nitrate and then challenging, with the dead bacteria, a viable culture of the same bacterium: Efficient antibacterial activity of the killed bacteria is observed. A mechanism is suggested in terms of the action of the dead bacteria as a reservoir of silver, which, due to Le-Chatelier's principle, is re-targeted to the living bacteria. Langmuirian behavior, as well as deviations from it, support the proposed mechanism.

  2. Antibacterial activity of silver-killed bacteria: the "zombies" effect.

    PubMed

    Wakshlak, Racheli Ben-Knaz; Pedahzur, Rami; Avnir, David

    2015-04-23

    We report a previously unrecognized mechanism for the prolonged action of biocidal agents, which we denote as the zombies effect: biocidally-killed bacteria are capable of killing living bacteria. The concept is demonstrated by first killing Pseudomonas aeruginosa PAO1 with silver nitrate and then challenging, with the dead bacteria, a viable culture of the same bacterium: Efficient antibacterial activity of the killed bacteria is observed. A mechanism is suggested in terms of the action of the dead bacteria as a reservoir of silver, which, due to Le-Chatelier's principle, is re-targeted to the living bacteria. Langmuirian behavior, as well as deviations from it, support the proposed mechanism.

  3. Killing spinors as a characterisation of rotating black hole spacetimes

    NASA Astrophysics Data System (ADS)

    Cole, Michael J.; Valiente Kroon, Juan A.

    2016-06-01

    We investigate the implications of the existence of Killing spinors in a spacetime. In particular, we show that in vacuum and electrovacuum a Killing spinor, along with some assumptions on the associated Killing vector in an asymptotic region, guarantees that the spacetime is locally isometric to the Kerr or Kerr–Newman solutions. We show that the characterisation of these spacetimes in terms of Killing spinors is an alternative expression of characterisation results of Mars (Kerr) and Wong (Kerr–Newman) involving restrictions on the Weyl curvature and matter content.

  4. [Killing and dignity of animals: a problem for veterinarians?].

    PubMed

    Fahrion; Dürr, S; Doherr, M G; Hartnack, S; Kunzmann, P

    2011-05-01

    Killing of animals is an important task to be performed by veterinarians. Killing decisions and their implementation often raise ethical questions. As a result of an interdisciplinary workshop targeting the subject "killing of animals" with veterinarians and ethicists, a three-dimensional dimension scheme was developed. Whereas the first two dimensions are focused on the animal's past and future life and are discussed with regard to life quality and life accomplishment (the "telos"), the third dimension incorporates the reason to kill and may integrate the concept of dignity. This form of dignity and the weighing of interests are applied to example scenarios and the resulting responsibilities of veterinarians and society are discussed.

  5. [Progresses on macrophages of male reproductive tract].

    PubMed

    Li, Jing-Jing; Wang, Tao; Wang, Geng-Xin

    2002-12-01

    The review summarized the recent progress on macrophages of male reproductive tract and the action of macrophages on male reproductive physiology and pathology. The close correlation and effect between testicular macrophages and Leydig cells, Sertoli cells, germ cells, hypothalamic-pituitary-gonadal axis were introduced, respectively. At the same time, it pointed out the changes of macrophages' morphology and function in immune orchitis, and their regulation on the development of orchitis. So the complex immune regulation network in testes and testicular macrophages playing an important role on spermatogenesis and the stableness of spermatogenetic microenvironment in testes were further illuminated, which can provide theoretical basis for clinic therapy.

  6. Contribution of Macrophage Polarization to Metabolic Diseases.

    PubMed

    Komohara, Yoshihiro; Fujiwara, Yukio; Ohnishi, Koji; Shiraishi, Daisuke; Takeya, Motohiro

    2016-01-01

    Macrophage activation is one of the major immunological events in the pathogenesis of various diseases. Recent studies have disclosed that complicated mechanisms are involved in macrophage activation and polarization, and many published research articles have been based on the M1/M2 polarization concept. It is considered that M1- and M2-like macrophages are associated with T helper (Th)1-type and Th2-type immune responses, respectively, via several immune mediators. In this article, we summarize the correlations between macrophage polarization and metabolic disorders in both humans and mice and discuss the contribution of macrophage polarization to the pathogenic process of metabolic diseases.

  7. Macrophage Polarization during Murine Lyme Borreliosis.

    PubMed

    Lasky, Carrie E; Olson, Rachel M; Brown, Charles R

    2015-07-01

    Infection of C3H mice with Borrelia burgdorferi, the causative agent of Lyme disease, reliably produces an infectious arthritis and carditis that peak around 3 weeks postinfection and then spontaneously resolve. Macrophage polarization has been suggested to drive inflammation, the clearance of bacteria, and tissue repair and resolution in a variety of infectious disease models. During Lyme disease it is clear that macrophages are capable of clearing Borrelia spirochetes and exhausted neutrophils; however, the role of macrophage phenotype in disease development or resolution has not been studied. Using classical (NOS2) and alternative (CD206) macrophage subset-specific markers, we determined the phenotype of F4/80(+) macrophages within the joints and heart throughout the infection time course. Within the joint, CD206(+) macrophages dominated throughout the course of infection, and NOS2(+) macrophage numbers became elevated only during the peak of inflammation. We also found dual NOS2(+) CD206(+) macrophages which increased during resolution. In contrast to findings for the ankle joints, numbers of NOS2(+) and CD206(+) macrophages in the heart were similar at the peak of inflammation. 5-Lipoxygenase-deficient (5-LOX(-/-)) mice, which display a failure of Lyme arthritis resolution, recruited fewer F4/80(+) cells to the infected joints and heart, but macrophage subset populations were unchanged. These results highlight differences in the inflammatory infiltrates during Lyme arthritis and carditis and demonstrate the coexistence of multiple macrophage subsets within a single inflammatory site.

  8. Identification of polarized macrophage subsets in zebrafish

    PubMed Central

    Nguyen-Chi, Mai; Laplace-Builhe, Béryl; Travnickova, Jana; Luz-Crawford, Patricia; Tejedor, Gautier; Phan, Quang Tien; Duroux-Richard, Isabelle; Levraud, Jean-Pierre; Kissa, Karima; Lutfalla, Georges

    2015-01-01

    While the mammalian macrophage phenotypes have been intensively studied in vitro, the dynamic of their phenotypic polarization has never been investigated in live vertebrates. We used the zebrafish as a live model to identify and trail macrophage subtypes. We generated a transgenic line whose macrophages expressing tumour necrosis factor alpha (tnfa), a key feature of classically activated (M1) macrophages, express fluorescent proteins Tg(mpeg1:mCherryF/tnfa:eGFP-F). Using 4D-confocal microscopy, we showed that both aseptic wounding and Escherichia coli inoculation triggered macrophage recruitment, some of which started to express tnfa. RT-qPCR on Fluorescence Activated Cell Sorting (FACS)-sorted tnfa+ and tnfa− macrophages showed that they, respectively, expressed M1 and alternatively activated (M2) mammalian markers. Fate tracing of tnfa+ macrophages during the time-course of inflammation demonstrated that pro-inflammatory macrophages converted into M2-like phenotype during the resolution step. Our results reveal the diversity and plasticity of zebrafish macrophage subsets and underline the similarities with mammalian macrophages proposing a new system to study macrophage functional dynamic. DOI: http://dx.doi.org/10.7554/eLife.07288.001 PMID:26154973

  9. Macrophages and cellular immunity in Drosophila melanogaster.

    PubMed

    Gold, Katrina S; Brückner, Katja

    2015-12-01

    The invertebrate Drosophila melanogaster has been a powerful model for understanding blood cell development and immunity. Drosophila is a holometabolous insect, which transitions through a series of life stages from embryo, larva and pupa to adulthood. In spite of this, remarkable parallels exist between Drosophila and vertebrate macrophages, both in terms of development and function. More than 90% of Drosophila blood cells (hemocytes) are macrophages (plasmatocytes), making this highly tractable genetic system attractive for studying a variety of questions in macrophage biology. In vertebrates, recent findings revealed that macrophages have two independent origins: self-renewing macrophages, which reside and proliferate in local microenvironments in a variety of tissues, and macrophages of the monocyte lineage, which derive from hematopoietic stem or progenitor cells. Like vertebrates, Drosophila possesses two macrophage lineages with a conserved dual ontogeny. These parallels allow us to take advantage of the Drosophila model when investigating macrophage lineage specification, maintenance and amplification, and the induction of macrophages and their progenitors by local microenvironments and systemic cues. Beyond macrophage development, Drosophila further serves as a paradigm for understanding the mechanisms underlying macrophage function and cellular immunity in infection, tissue homeostasis and cancer, throughout development and adult life. PMID:27117654

  10. On the Lie subalgebra of Killing-Milne and Killing-Cartan vector fields in Newtonian spacetime

    NASA Astrophysics Data System (ADS)

    Chamel, Nicolas

    2015-12-01

    The Galilean (and more generally Milne) invariance of Newtonian theory allows for Killing vector fields of a general kind, whereby the Lie derivative of a field is not required to vanish but only to be cancellable by some infinitesimal Galilean (respectively Milne) gauge transformation. In this paper, it is shown that both the Killing-Milne vector fields, which preserve the background Newtonian spacetime structure and the Killing-Cartan vector fields, which in addition preserve the gravitational field, form a Lie subalgebra.

  11. Tissue-Resident Macrophage Ontogeny and Homeostasis.

    PubMed

    Ginhoux, Florent; Guilliams, Martin

    2016-03-15

    Defining the origins and developmental pathways of tissue-resident macrophages should help refine our understanding of the role of these cells in various disease settings and enable the design of novel macrophage-targeted therapies. In recent years the long-held belief that macrophage populations in the adult are continuously replenished by monocytes from the bone marrow (BM) has been overturned with the advent of new techniques to dissect cellular ontogeny. The new paradigm suggests that several tissue-resident macrophage populations are seeded during waves of embryonic hematopoiesis and self-maintain independently of BM contribution during adulthood. However, the exact nature of the embryonic progenitors that give rise to adult tissue-resident macrophages is still debated, and the mechanisms enabling macrophage population maintenance in the adult are undefined. Here, we review the emergence of these concepts and discuss current controversies and future directions in macrophage biology. PMID:26982352

  12. Macrophages in Tissue Repair, Regeneration, and Fibrosis.

    PubMed

    Wynn, Thomas A; Vannella, Kevin M

    2016-03-15

    Inflammatory monocytes and tissue-resident macrophages are key regulators of tissue repair, regeneration, and fibrosis. After tissue injury, monocytes and macrophages undergo marked phenotypic and functional changes to play critical roles during the initiation, maintenance, and resolution phases of tissue repair. Disturbances in macrophage function can lead to aberrant repair, such that uncontrolled production of inflammatory mediators and growth factors, deficient generation of anti-inflammatory macrophages, or failed communication between macrophages and epithelial cells, endothelial cells, fibroblasts, and stem or tissue progenitor cells all contribute to a state of persistent injury, and this could lead to the development of pathological fibrosis. In this review, we discuss the mechanisms that instruct macrophages to adopt pro-inflammatory, pro-wound-healing, pro-fibrotic, anti-inflammatory, anti-fibrotic, pro-resolving, and tissue-regenerating phenotypes after injury, and we highlight how some of these mechanisms and macrophage activation states could be exploited therapeutically.

  13. Control of the Inflammatory Macrophage Transcriptional Signature by miR-155

    PubMed Central

    Jablonski, Kyle A.; Gaudet, Andrew D.; Amici, Stephanie A.; Popovich, Phillip G.

    2016-01-01

    Inflammatory M1 spectrum macrophages protect from infection but can cause inflammatory disease and tissue damage, whereas alternatively activated/M2 spectrum macrophages reduce inflammation and promote tissue repair. Modulation of macrophage phenotype may be therapeutically beneficial and requires further understanding of the molecular programs that control macrophage differentiation. A potential mechanism by which macrophages differentiate may be through microRNA (miRNA), which bind to messenger RNA and post-transcriptionally modify gene expression, cell phenotype and function. We hypothesized that the inflammation-associated miRNA, miR-155, would be required for typical development of macrophage inflammatory state. miR-155 was rapidly up-regulated over 100-fold in inflammatory M1(LPS + IFN-γ), but not M2(IL-4), macrophages. Inflammatory genes Inos, Il1b and Tnfa and their corresponding protein or enzymatic products were reduced up to 72% in miR-155 knockout mouse M1(LPS + IFN-γ) macrophages, but miR-155 deficiency did not affect expression of the M2-associated gene Arg1 in M2(IL-4) macrophages. Additionally, a miR-155 oligonucleotide inhibitor efficiently suppressed Inos and Tnfa gene expression in wild-type M1(LPS + IFN-γ) macrophages. Comparative transcriptional profiling of unstimulated and M1(LPS + IFN-γ) macrophages derived from wild-type (WT) and miR-155 knockout (KO) mice revealed that half (approximately 650 genes) of the signature we previously identified in WT M1(LPS + IFN-γ) macrophages was dependent on miR-155. Real-Time PCR of independent datasets confirmed that miR-155 contributed to suppression of its validated mRNA targets Inpp5d, Tspan14, Ptprj and Mafb and induction of Inos, Il1b, Tnfa, Il6 and Il12. Overall, these data indicate that miR-155 plays an essential role in driving the inflammatory phenotype of M1(LPS+ IFN-γ) macrophages. PMID:27447824

  14. Synthesis of antimony complexes of yeast mannan and mannan derivatives and their effect on Leishmania-infected macrophages.

    PubMed

    Cantos, G; Barbieri, C L; Iacomini, M; Gorin, P A; Travassos, L R

    1993-01-01

    Antimony(Sb)-yeast mannan complexes were synthesized as a strategy to introduce Sb into macrophages infected with Leishmania amastigotes. The complexes were taken up by endocytosis after specific recognition by alpha-D-mannosyl receptors on the macrophage membrane. About 90% of the intracellular parasites were destroyed by Sb-mannan in vitro, whereas the corresponding Sb concentration used as the pentavalent antimonial drug glucantime destroyed about 60% of the amastigotes. None of the Sb complexes prepared with mannan acid or basic derivatives was as effective as the simple Sb-mannan complex in clearing macrophage infection by Leishmania (L) amazonensis. The leishmanicidal effect of Sb-mannan was also demonstrated in vivo with infected hamsters. The alternative use of Sb-mannan complex in the treatment of human leishmaniasis is envisaged on the basis of parasite-killing efficiency and the use of a low antimony dose.

  15. Epirubicin-Adsorbed Nanodiamonds Kill Chemoresistant Hepatic Cancer Stem Cells

    PubMed Central

    2015-01-01

    Chemoresistance is a primary cause of treatment failure in cancer and a common property of tumor-initiating cancer stem cells. Overcoming mechanisms of chemoresistance, particularly in cancer stem cells, can markedly enhance cancer therapy and prevent recurrence and metastasis. This study demonstrates that the delivery of Epirubicin by nanodiamonds is a highly effective nanomedicine-based approach to overcoming chemoresistance in hepatic cancer stem cells. The potent physical adsorption of Epirubicin to nanodiamonds creates a rapidly synthesized and stable nanodiamond–drug complex that promotes endocytic uptake and enhanced tumor cell retention. These attributes mediate the effective killing of both cancer stem cells and noncancer stem cells in vitro and in vivo. Enhanced treatment of both tumor cell populations results in an improved impairment of secondary tumor formation in vivo compared with treatment by unmodified chemotherapeutics. On the basis of these results, nanodiamond-mediated drug delivery may serve as a powerful method for overcoming chemoresistance in cancer stem cells and markedly improving overall treatment against hepatic cancers. PMID:25437772

  16. Nanotechnology for the detection and kill of circulating tumor cells

    PubMed Central

    2014-01-01

    Circulating tumor cells (CTCs) represent a surrogate biomarker of hematogenous metastases and thus could be considered as a ‘liquid biopsy’ which reveals metastasis in action. But it is absolutely a challenge to detect CTCs due to their extreme rarity. At present, the most common principle is to take advantage of the epithelial surface markers of CTCs which attach to a specific antibody. Antibody-magnetic nanobeads combine with the epithelial surface markers, and then the compound is processed by washing, separation, and detection. However, a proportion of CTC antigen expressions are down-regulated or lost in the process of epithelial-mesenchymal transition (EMT), and thus, this part of CTCs cannot be detected by classical detection methods such as CellSearch. To resolve this problem, some multiple-marker CTC detections have been developed rapidly. Additionally, nanotechnology is a promising approach to kill CTCs with high efficiency. Implantable nanotubes coated with apoptosis-promoting molecules improve the disease-free survival and overall survival. The review introduces some novel CTC detection techniques and therapeutic methods by virtue of nanotechnology to provide a better knowledge of the progress about CTC study. PMID:25258614

  17. Nanotechnology for the detection and kill of circulating tumor cells

    NASA Astrophysics Data System (ADS)

    Gao, Yang; Yuan, Zhou

    2014-09-01

    Circulating tumor cells (CTCs) represent a surrogate biomarker of hematogenous metastases and thus could be considered as a `liquid biopsy' which reveals metastasis in action. But it is absolutely a challenge to detect CTCs due to their extreme rarity. At present, the most common principle is to take advantage of the epithelial surface markers of CTCs which attach to a specific antibody. Antibody-magnetic nanobeads combine with the epithelial surface markers, and then the compound is processed by washing, separation, and detection. However, a proportion of CTC antigen expressions are down-regulated or lost in the process of epithelial-mesenchymal transition (EMT), and thus, this part of CTCs cannot be detected by classical detection methods such as CellSearch. To resolve this problem, some multiple-marker CTC detections have been developed rapidly. Additionally, nanotechnology is a promising approach to kill CTCs with high efficiency. Implantable nanotubes coated with apoptosis-promoting molecules improve the disease-free survival and overall survival. The review introduces some novel CTC detection techniques and therapeutic methods by virtue of nanotechnology to provide a better knowledge of the progress about CTC study.

  18. Time-kill behavior against eight bacterial species and cytotoxicity of antibacterial monomers

    PubMed Central

    Li, Fang; Weir, Michael D.; Fouad, Ashraf F.; Xu, Hockin H. K.

    2013-01-01

    Objectives The objectives of this study were to investigate: (1) the antibacterial activity of two antibacterial monomers, dimethylaminododecyl methacrylate (DMADDM) and dimethylammoniumethyl dimethacrylate (DMAEDM), against eight different species of oral pathogens for the first time; (2) the cytotoxicity of DMAEDM and DMADDM. Methods DMAEDM and DMADDM were synthesized by reacting a tertiary amine group with an organo-halide. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against eight species of bacteria were tested. Time-kill determinations were performed to examine the bactericidal kinetics. Cytotoxicity of monomers on human gingival fibroblasts (HGF) was assessed using a methyl thiazolyltetrazolium assay and live/dead viability assay. Results DMADDM showed strong bactericidal activity against all bacteria, with MIC of 1.2 to 9.8μg/mL. DMAEDM had MIC of 20 to 80mg/mL. Time-kill determinations indicated that DMADDM and DMAEDM had rapid killing effects against eight species of bacteria, and eliminated all bacteria in 30min at the concentration of 4-fold MBC. Median lethal concentration for DMADDM and DMAEDM was between 20 to 40μg/mL, which was 20-fold higher than 1 to 2μg/mL for BisGMA control. Conclusions DMAEDM and DMADDM were tested in time-kill assay against eight species of oral bacteria for the first time. Both were effective in bacteria-inhibition, but DMADDM had a higher potency than DMAEDM. Different killing efficacy was found against different bacteria species. DMAEDM and DMADDM had much lower cytotoxicity than BisGMA. Therefore, DMADDM and DMAEDM are promising for use in bonding agents and other restorative/preventive materials to combat a variety of oral pathogens. PMID:23876930

  19. Inhibition of the Plasma-Membrane-Associated Serine Protease Cathepsin G by Mycobacterium tuberculosis Rv3364c Suppresses Caspase-1 and Pyroptosis in Macrophages

    PubMed Central

    Danelishvili, Lia; Everman, Jamie L.; McNamara, Michael J.; Bermudez, Luiz E.

    2012-01-01

    Tuberculosis is a disease associated with the infection of a great part of the world’s population and is responsible for the death of two to three million people annually. Mycobacterium tuberculosis infects macrophages and subverts its mechanisms of killing. The pathogen suppresses macrophage apoptosis by many different mechanisms. We describe that, upon uptake by macrophages, M. tuberculosis overexpresses an operon Rv3361c-Rv3365c and secretes Rv3364c. The Rv3365c knockout strain is deficient in apoptosis inhibition. The Rv3364c protein binds to the serine protease cathepsin G on the membrane, inhibiting its enzymatic activity and the downstream activation of caspase-1-dependent apoptosis. In summary, M. tuberculosis prevents macrophage pyroptosis by a novel mechanism involving cytoplasmic surveillance proteins. PMID:22275911

  20. Extracellular ATP is cytotoxic to mononuclear phagocytes but does not induce killing of intracellular Mycobacterium avium subsp. paratuberculosis.

    PubMed

    Woo, Seng-Ryong; Barletta, Raúl G; Czuprynski, Charles J

    2007-09-01

    Mycobacterium avium subsp. paratuberculosis is the etiologic agent of Johne's disease, a chronic granulomatous enteritis in ruminants. ATP has been reported to induce cell death of macrophages and killing of Mycobacterium species in human and murine macrophages. In this study we investigated the short-term effect of ATP on the viability of M. avium subsp. paratuberculosis-infected bovine mononuclear phagocytes and the bacilli within them. Addition of 5 mM ATP to M. avium subsp. paratuberculosis-infected bovine monocytes resulted in 50% cytotoxicity of bovine monocytes at 24 h. Addition of 2'(3')-O-(4-benzoylbenzoyl) ATP triethylammonium salt (Bz-ATP), which is a longer-lived ATP homologue and purinergic receptor agonist, significantly increased the uptake of YO-PRO, which is a marker for membrane pore activation by P2X receptors. Addition of Bz-ATP also stimulated lactate dehydrogenase release and caspase-3 activity in infected bovine monocytes. Neither ATP nor Bz-ATP reduced the survival of M. avium subsp. paratuberculosis in bovine mononuclear phagocytes. Likewise, addition of ATP or Bz-ATP was cytotoxic to murine macrophage cell lines (RAW 264.7 and J774A.1 cells) but did not affect the intracellular survival of M. avium subsp. paratuberculosis, nor were the numbers of viable Mycobacterium avium subsp. avium or Mycobacterium bovis BCG cells altered in bovine mononuclear phagocytes or J774A.1 cells following ATP or Bz-ATP treatment. These data suggest that extracellular ATP does not induce the killing of intracellular M. avium subsp. paratuberculosis in bovine mononuclear phagocytes.