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Sample records for lps-activated macrophage cultures

  1. Soybean-derived Bowman-Birk inhibitor inhibits neurotoxicity of LPS-activated macrophages

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    Persidsky Yuri

    2011-02-01

    Full Text Available Abstract Background Lipopolysaccharide (LPS, the major component of the outer membrane of gram-negative bacteria, can activate immune cells including macrophages. Activation of macrophages in the central nervous system (CNS contributes to neuronal injury. Bowman-Birk inhibitor (BBI, a soybean-derived protease inhibitor, has anti-inflammatory properties. In this study, we examined whether BBI has the ability to inhibit LPS-mediated macrophage activation, reducing the release of pro-inflammatory cytokines and subsequent neurotoxicity in primary cortical neural cultures. Methods Mixed cortical neural cultures from rat were used as target cells for testing neurotoxicity induced by LPS-treated macrophage supernatant. Neuronal survival was measured using a cell-based ELISA method for expression of the neuronal marker MAP-2. Intracellular reactive oxygen species (ROS production in macrophages was measured via 2', 7'-dichlorofluorescin diacetate (DCFH2DA oxidation. Cytokine expression was determined by quantitative real-time PCR. Results LPS treatment of macrophages induced expression of proinflammatory cytokines (IL-1β, IL-6 and TNF-α and of ROS. In contrast, BBI pretreatment (1-100 μg/ml of macrophages significantly inhibited LPS-mediated induction of these cytokines and ROS. Further, supernatant from BBI-pretreated and LPS-activated macrophage cultures was found to be less cytotoxic to neurons than that from non-BBI-pretreated and LPS-activated macrophage cultures. BBI, when directly added to the neuronal cultures (1-100 μg/ml, had no protective effect on neurons with or without LPS-activated macrophage supernatant treatment. In addition, BBI (100 μg/ml had no effect on N-methyl-D-aspartic acid (NMDA-mediated neurotoxicity. Conclusions These findings demonstrate that BBI, through its anti-inflammatory properties, protects neurons from neurotoxicity mediated by activated macrophages.

  2. Full Spectrum of LPS Activation in Alveolar Macrophages of Healthy Volunteers by Whole Transcriptomic Profiling.

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    Miguel Pinilla-Vera

    Full Text Available Despite recent advances in understanding macrophage activation, little is known regarding how human alveolar macrophages in health calibrate its transcriptional response to canonical TLR4 activation. In this study, we examined the full spectrum of LPS activation and determined whether the transcriptomic profile of human alveolar macrophages is distinguished by a TIR-domain-containing adapter-inducing interferon-β (TRIF-dominant type I interferon signature. Bronchoalveolar lavage macrophages were obtained from healthy volunteers, stimulated in the presence or absence of ultrapure LPS in vitro, and whole transcriptomic profiling was performed by RNA sequencing (RNA-Seq. LPS induced a robust type I interferon transcriptional response and Ingenuity Pathway Analysis predicted interferon regulatory factor (IRF7 as the top upstream regulator of 89 known gene targets. Ubiquitin-specific peptidase (USP-18, a negative regulator of interferon α/β responses, was among the top up-regulated genes in addition to IL10 and USP41, a novel gene with no known biological function but with high sequence homology to USP18. We determined whether IRF-7 and USP-18 can influence downstream macrophage effector cytokine production such as IL-10. We show that IRF-7 siRNA knockdown enhanced LPS-induced IL-10 production in human monocyte-derived macrophages, and USP-18 overexpression attenuated LPS-induced production of IL-10 in RAW264.7 cells. Quantitative PCR confirmed upregulation of USP18, USP41, IL10, and IRF7. An independent cohort confirmed LPS induction of USP41 and IL10 genes. These results suggest that IRF-7 and predicted downstream target USP18, both elements of a type I interferon gene signature identified by RNA-Seq, may serve to fine-tune early cytokine response by calibrating IL-10 production in human alveolar macrophages.

  3. Simvastatin induces caspase-independent apoptosis in LPS-activated RAW264.7 macrophage cells

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    Kim, Yong Chan; Song, Seok Bean; Lee, Mi Hee; Kang, Kwang Il; Lee, Hayyoung; Paik, Sang-Gi; Kim, Kyoon Eon; Kim, Young Sang

    2006-01-01

    Macrophages participate in several inflammatory pathologies such as sepsis and arthritis. We examined the effect of simvastatin on the LPS-induced proinflammatory macrophage RAW264.7 cells. Co-treatment of LPS and a non-toxic dose of simvastatin induced cell death in RAW264.7 cells. The cell death was accompanied by disruption of mitochondrial membrane potential (MMP), genomic DNA fragmentation, and caspase-3 activation. Surprisingly, despite caspase-dependent apoptotic cascade being completely blocked by Z-VAD-fmk, a pan-caspase inhibitor, the cell death was only partially repressed. In the presence of Z-VAD-fmk, DNA fragmentation was blocked, but DNA condensation, disruption of MMP, and nuclear translocation of apoptosis inducing factor were obvious. The cell death by simvastatin and LPS was effectively decreased by both the FPP and GGPP treatments as well as mevalonate. Our findings indicate that simvastatin triggers the cell death of LPS-treated RAW264.7 cells through both caspase-dependent and -independent apoptotic pathways, suggesting a novel mechanism of statins for the severe inflammatory disease therapy

  4. Aqueous Extract of Tomato (Solanum lycopersicum L. and Ferulic Acid Reduce the Expression of TNF-α and IL-1β in LPS-Activated Macrophages

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    Simón Navarrete

    2015-08-01

    Full Text Available Acute inflammation is essential for defending the body against pathogens; however, when inflammation becomes chronic, it is harmful to the body and is part of the pathophysiology of various diseases such as Diabetes Mellitus type 2 (DM2 and Cardiovascular Disease (CVD among others. In chronic inflammation macrophages play an important role, mainly through the secretion of proinflammatory cytokines such as Tumor necrosis factor (TNF-α and Interleukin (IL-1β, explained in part by activation of the Toll-like receptor 4 (TLR4, a signaling pathway which culminates in the activation of Nuclear factor (NF-κB, an important transcription factor in the expression of these proinflammatory genes. On the other hand, the benefits on health of a diet rich in fruit and vegetables are well described. In this work, the effects of aqueous extract of tomato and ferulic acid on the expression of proinflammatory cytokines in LPS activated monocyte-derived THP-1 macrophages were investigated. In addition, using Western blot, we investigated whether the inhibition was due to the interference on activation of NF-κB. We found that both the tomato extract and ferulic acid presented inhibitory activity on the expression of TNF-α and IL-1β cytokine by inhibiting the activation of NF-κB. The current results suggest that tomatoes and ferulic acid may contribute to prevention of chronic inflammatory diseases.

  5. Extra virgin olive oil polyphenolic extracts downregulate inflammatory responses in LPS-activated murine peritoneal macrophages suppressing NFκB and MAPK signalling pathways.

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    Cárdeno, A; Sánchez-Hidalgo, M; Aparicio-Soto, M; Sánchez-Fidalgo, S; Alarcón-de-la-Lastra, C

    2014-06-01

    Extra virgin olive oil (EVOO) is obtained from the fruit of the olive tree Olea europaea L. Phenolic compounds present in EVOO have recognized anti-oxidant and anti-inflammatory properties. However, the activity of the total phenolic fraction extracted from EVOO and the action mechanisms involved are not well defined. The present study was designed to evaluate the potential anti-inflammatory mechanisms of the polyphenolic extract (PE) from EVOO on LPS-stimulated peritoneal murine macrophages. Nitric oxide (NO) production was analyzed by the Griess method and intracellular reactive oxygen species (ROS) by fluorescence analysis. Moreover, changes in the protein expression of the pro-inflammatory enzymes, inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2 and microsomal prostaglandin E synthase-1 (mPGES-1), as well as the role of nuclear transcription factor kappa B (NFκB) and mitogen-activated protein kinase (MAPK) signalling pathways, were analyzed by Western blot. PE from EVOO reduced LPS-induced oxidative stress and inflammatory responses through decreasing NO and ROS generation. In addition, PE induced a significant down-regulation of iNOS, COX-2 and mPGES-1 protein expressions, reduced MAPK phosphorylation and prevented the nuclear NFκB translocation. This study establishes that PE from EVOO possesses anti-inflammatory activities on LPS-stimulated murine macrophages.

  6. Anti-inflammatory effect of a selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor via the stimulation of heme oxygenase-1 in LPS-activated mice and J774.1 murine macrophages

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    Sung Bum Park

    2016-08-01

    Full Text Available 11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1 converts inactive cortisone to the active cortisol. 11β-HSD1 may be involved in the resolution of inflammation. In the present study, we investigate the anti-inflammatory effects of 2-(3-benzoyl-4-hydroxy-1,1-dioxo-2H-1,2-benzothiazine-2-yl-1-phenylethanone (KR-66344, a selective 11β-HSD1 inhibitor, in lipopolysaccharide (LPS-activated C57BL/6J mice and macrophages. LPS increased 11β-HSD1 activity and expression in macrophages, which was inhibited by KR-66344. In addition, KR-66344 increased survival rate in LPS treated C57BL/6J mice. HO-1 mRNA expression level was increased by KR-66344, and this effect was reversed by the HO competitive inhibitor, ZnPP, in macrophages. Moreover, ZnPP reversed the suppression of ROS formation and cell death induced by KR-66344. ZnPP also suppressed animal survival rate in LPS plus KR-66344 treated C57BL/6J mice. In the spleen of LPS-treated mice, KR-66344 prevented cell death via suppression of inflammation, followed by inhibition of ROS, iNOS and COX-2 expression. Furthermore, LPS increased NFκB-p65 and MAPK phosphorylation, and these effects were abolished by pretreatment with KR-66344. Taken together, KR-66344 protects against LPS-induced animal death and spleen injury by inhibition of inflammation via induction of HO-1 and inhibition of 11β-HSD1 activity. Thus, we concluded that the selective 11β-HSD1 inhibitor may provide a novel strategy in the prevention/treatment of inflammatory disorders in patients.

  7. Excretory/secretory products from two Fasciola hepatica isolates induce different transcriptional changes and IL-10 release in LPS-activated bovine "BOMA" macrophages.

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    Bąska, Piotr; Norbury, Luke James; Zawistowska-Deniziak, Anna; Wiśniewski, Marcin; Januszkiewicz, Kamil

    2017-10-01

    Fasciola hepatica are trematodes that reside in the bile ducts of mammals. Infection causes US$3 billion in losses annually in animal production and is considered a zoonosis of growing importance. An under-represented area in F. hepatica research has been the examination of the different immunomodulatory abilities of various parasite isolates on the host immune system. In this paper, this issue was explored, with the bovine macrophage cell line "BOMA". The cells were matured by LPS treatment and stimulated with excretory/secretory antigens (ES) from two Fasciola hepatica isolates: a laboratory isolate "Weybridge" (Fh-WeyES) and a wild isolate (Fh-WildES). As expected, stimulation with antigen mixtures with highly similar compositions resulted in mild transcriptomic differences. However, there were significant differences in cytokine levels. Compared to Fh-WeyES, exposure to Fh-WildES upregulated 27 and downregulated 30 genes. Fh-ES from both isolates diminished the release of TNF-α, whereas only Fh-WildES decreased IL-10 secretion. Neither Fh-WeyES nor Fh-WildES had an impact on IL-12 release. Our results indicate that various isolates can have different immunomodulatory abilities and impacts on the bovine immune system.

  8. Recombinant CC16 protein inhibits the production of pro-inflammatory cytokines via NF-κB and p38 MAPK pathways in LPS-activated RAW264.7 macrophages

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    Min Pang; Guoping Zheng; Baofeng Yu; Hailong Wang; Yangyang Yuan; Dong Wang; Ting Li; Dan Wang; Xiaohong Shi; Min Guo; Chunfang Wang; Xinri Zhang

    2017-01-01

    Accumulating evidence indicates that Clara cell protein-16 (CC16) has anti-inflammatory functions,although the involved molecular pathways have not been completely elucidated.Here,we evaluated the effect of recombinant rat CC16 (rCC16) on the expression of tumor necrosis factor alpha (TNF-α),interleukin-6 (IL-6),and IL-8 in lipopolysaccharide (LPS)-stimulated mouse macrophages (RAW264.7 cells) and explored the underlying molecular mechanisms.It was found that rCC16 inhibited LPS-induced TNF-α,IL-6,and IL-8 expression at both the messenger ribonucleicacid (mRNA) level and protein level in a concentration-dependent manner,as demonstrated by realtime reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay.Such suppressive effects were accompanied by the inhibition of transcriptional activity and the deoxyribonucleic acid binding activity of nuclear factor (NF)-κB but not activator protein (AP)-1.Western blot analysis further revealed that rCC16 inhibited the increase of nuclear NF-κB and the reduction of cytosolic NF-κB,the phosphorylation and reduction of NF-κB inhibitory protein IκBα,and the p38 mitogen-activated protein kinase (MAPK)-dependent NF-κB activation by phosphoryl-ation at Ser276 of its p65 subunit.Furthermore,rCC16 was found to have no effect on the phosphorylation of c-Jun N-terminal kinase,c-Jun,or the nuclear translocation of c-Jun.In addition,reduction of TNF-α,IL-6,and IL-8 were reversed when the level of endogenous uteroglobin-binding protein was reduced by RNA interference in rCC16-and LPS-treated RAW264.7 cells.Our data suggest that rCC16 suppresses LPS-mediated inflammatory mediator TNF-α,IL-6,and IL-8 production by inactivating NF-κB and p38 MAPK but not AP-1 in RAW264.7 cells.

  9. Leucine supplementation attenuates macrophage foam-cell formation: Studies in humans, mice, and cultured macrophages.

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    Grajeda-Iglesias, Claudia; Rom, Oren; Hamoud, Shadi; Volkova, Nina; Hayek, Tony; Abu-Saleh, Niroz; Aviram, Michael

    2018-02-05

    Whereas atherogenicity of dietary lipids has been largely studied, relatively little is known about the possible contribution of dietary amino acids to macrophage foam-cell formation, a hallmark of early atherogenesis. Recently, we showed that leucine has antiatherogenic properties in the macrophage model system. In this study, an in-depth investigation of the role of leucine in macrophage lipid metabolism was conducted by supplementing humans, mice, or cultured macrophages with leucine. Macrophage incubation with serum obtained from healthy adults supplemented with leucine (5 g/d, 3 weeks) significantly decreased cellular cholesterol mass by inhibiting the rate of cholesterol biosynthesis and increasing cholesterol efflux from macrophages. Similarly, leucine supplementation to C57BL/6 mice (8 weeks) resulted in decreased cholesterol content in their harvested peritoneal macrophages (MPM) in relation with reduced cholesterol biosynthesis rate. Studies in J774A.1 murine macrophages revealed that leucine dose-dependently decreased cellular cholesterol and triglyceride mass. Macrophages treated with leucine (0.2 mM) showed attenuated uptake of very low-density lipoproteins and triglyceride biosynthesis rate, with a concurrent down-regulation of diacylglycerol acyltransferase-1, a key enzyme catalyzing triglyceride biosynthesis in macrophages. Similar effects were observed when macrophages were treated with α-ketoisocaproate, a key leucine metabolite. Finally, both in vivo and in vitro leucine supplementation significantly improved macrophage mitochondrial respiration and ATP production. The above studies, conducted in human, mice, and cultured macrophages, highlight a protective role for leucine attenuating macrophage foam-cell formation by mechanisms related to the metabolism of cholesterol, triglycerides, and energy production. © 2018 BioFactors, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

  10. Carapanolides J–L from the Seeds of Carapa guianensis (Andiroba and Their Effects on LPS-Activated NO Production

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    Yuuki Matsui

    2014-10-01

    Full Text Available A novel gedunin and two novel phragmalin-type limonoids, named carapanolides J–L (compounds 1–3 as well as a known gedunin-type limonoid 4 were isolated from the seeds of Carapa guianensis (andiroba. Their structures were determined on the basis of 1D and 2D NMR spectroscopy and HRFABMS. Compounds 1–4 were evaluated for their effects on the production of NO in LPS-activated mouse peritoneal macrophages.

  11. Alternatively Activated (M2) Macrophage Phenotype Is Inducible by Endothelin-1 in Cultured Human Macrophages.

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    Soldano, Stefano; Pizzorni, Carmen; Paolino, Sabrina; Trombetta, Amelia Chiara; Montagna, Paola; Brizzolara, Renata; Ruaro, Barbara; Sulli, Alberto; Cutolo, Maurizio

    2016-01-01

    Alternatively activated (M2) macrophages are phenotypically characterized by the expression of specific markers, mainly macrophage scavenger receptors (CD204 and CD163) and mannose receptor-1 (CD206), and participate in the fibrotic process by over-producing pro-fibrotic molecules, such as transforming growth factor-beta1 (TGFbeta1) and metalloproteinase (MMP)-9. Endothelin-1 (ET-1) is implicated in the fibrotic process, exerting its pro-fibrotic effects through the interaction with its receptors (ETA and ETB). The study investigated the possible role of ET-1 in inducing the transition from cultured human macrophages into M2 cells. Cultured human monocytes (THP-1 cell line) were activated into macrophages (M0 macrophages) with phorbol myristate acetate and subsequently maintained in growth medium (M0-controls) or treated with either ET-1 (100nM) or interleukin-4 (IL-4, 10ng/mL, M2 inducer) for 72 hours. Similarly, primary cultures of human peripheral blood monocyte (PBM)-derived macrophages obtained from healthy subjects, were maintained in growth medium (untreated cells) or treated with ET-1 or IL-4 for 6 days. Both M0 and PBM-derived macrophages were pre-treated with ET receptor antagonist (ETA/BRA, bosentan 10-5M) for 1 hour before ET-1 stimulation. Protein and gene expression of CD204, CD206, CD163, TGFbeta1 were analysed by immunocytochemistry, Western blotting and quantitative real time polymerase chain reaction (qRT-PCR). Gene expression of interleukin(IL)-10 and macrophage derived chemokine (CCL-22) was evaluated by qRT-PCR. MMP-9 production was investigated by gel zymography. ET-1 significantly increased the expression of M2 phenotype markers CD204, CD206, CD163, IL-10 and CCL-22, and the production of MMP-9 in both cultures of M0 and PBM-derived macrophages compared to M0-controls and untreated cells. In cultured PBM-derived macrophages, ET-1 increased TGFbeta1 protein and gene expression compared to untreated cells. The ET-1-mediated effects were

  12. Effect of irradiation on lysosomal enzyme activation in cultured macrophages

    International Nuclear Information System (INIS)

    Clarke, C.; Wills, E.D.

    1980-01-01

    The effect of γrays on lysosomal enzyme activity of normal and immune macrophages of DBA/2 mice cultured in vitro has been studied. A dose of 500 rad did not significantly affect lysosomal enzyme activity 3 hours after irradiation but caused the activity to increase to 1.4 times the control value 22.5 hours after irradiation. 22.5 hours after a dose of 3000 rad the enzyme activity increased to 2.5 times the control. Lysosomal enzyme activity of the macrophages was also markedly increased by immunization of the mice with D lymphoma cells, before culture in vitro, but irradiation of these cells with a dose of 500 rad caused a further increase in lysosomal enzyme activity. The results indicate that immunization and irradiation both cause stimulation of lysosomal enzyme activity in macrophages but that the mechanisms of activation are unlikely to be identical. (author)

  13. The effects of three types of macrophages culture supernatant on CFU-GM in irradiated mice

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    Quan Hongxun; Fu Li; Zhao Fengchen; Han Fen

    2008-01-01

    Objective: To study the effects of peritional macrophyge(PM), alveolar macrophage (AM), and Kupffer cell (KC) on colony forming unite granulacyte/macrophage (CFU -GM) in irradiated mice. Methods: Using techniques of hemopoietic progenitors in vitro, the authors studied the effects of three types of macrophages culture supernatant on CFU - GM. Results: It is shown that three types of macrophages culture supernatant may stimulate proliferation and differentiation of CFU-GM in irradiated mice, and KC is the best one in comparison to others. Conclusion: three types of macrophages culture supernatant may protect CFU-GM irradiated mice with KC being the best method. (authors)

  14. Thermo-responsive cell culture carrier: Effects on macrophage functionality and detachment efficiency.

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    Rennert, Knut; Nitschke, Mirko; Wallert, Maria; Keune, Natalie; Raasch, Martin; Lorkowski, Stefan; Mosig, Alexander S

    2017-01-01

    Harvesting cultivated macrophages for tissue engineering purposes by enzymatic digestion of cell adhesion molecules can potentially result in unintended activation, altered function, or behavior of these cells. Thermo-responsive polymer is a promising tool that allows for gentle macrophage detachment without artificial activation prior to subculture within engineered tissue constructs. We therefore characterized different species of thermo-responsive polymers for their suitability as cell substrate and to mediate gentle macrophage detachment by temperature shift. Primary human monocyte- and THP-1-derived macrophages were cultured on thermo-responsive polymers and characterized for phagocytosis and cytokine secretion in response to lipopolysaccharide stimulation. We found that both cell types differentially respond in dependence of culture and stimulation on thermo-responsive polymers. In contrast to THP-1 macrophages, primary monocyte-derived macrophages showed no signs of impaired viability, artificial activation, or altered functionality due to culture on thermo-responsive polymers compared to conventional cell culture. Our study demonstrates that along with commercially available UpCell carriers, two other thermo-responsive polymers based on poly(vinyl methyl ether) blends are attractive candidates for differentiation and gentle detachment of primary monocyte-derived macrophages. In summary, we observed similar functionality and viability of primary monocyte-derived macrophages cultured on thermo-responsive polymers compared to standard cell culture surfaces. While this first generation of custom-made thermo-responsive polymers does not yet outperform standard culture approaches, our results are very promising and provide the basis for exploiting the unique advantages offered by custom-made thermo-responsive polymers to further improve macrophage culture and recovery in the future, including the covalent binding of signaling molecules and the reduction of

  15. Dissolution of short and long rockwool and glasswool fibers by macrophages in flowthrough cell culture.

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    Luoto, K; Holopainen, M; Kangas, J; Kalliokoski, P; Savolainen, K

    1998-07-01

    Dissolution of MMVF (man-made vitreous fibers) by macrophages has previously been studied utilizing cell cultures in wells. A new, more dynamic method has been developed to explore the effects of macrophages on MMVF dissolution. In this method, the culture medium flows through a membrane on which the macrophages and fibers are placed. The dissolution of short and long rockwool and glasswool fibers was investigated in the present study by macrophages by assessing the dissolution of Si (silicon), Fe (iron), and Al (aluminium) from the fibers. Dissolution of these elements usually increased as a function of time. Generally, the dissolution of elements from the fibers in the flowthrough culture exceeded that observed with the culture in wells system. The dissolution of glasswool fibers was greater in medium than in cell culture, whereas the opposite was true for rockwool fibers. Dissolution of Si was greater from glasswool than from rockwool fibers, while the opposite was true for Fe and Al. Macrophages that had phagocytized fibers in flowthrough culture contained Si, and there were also precipitations with Si in the samples. The fibers in the flowthrough culture also exhibited surface changes such as breakings, pittings, etching, and peeling. The short rockwool fibers tended to fracture more than short glasswool fibers, while long glasswool fibers were more extensively broken than short glasswool fibers. The results with this new, dynamic, flowthrough culture method with macrophages demonstrate that this method provides valuable information on the abilities of macrophages to dissolve MMVF leading to subsequent morphological changes of fibers.

  16. Induction of different activated phenotypes of mouse peritoneal macrophages grown in different tissue culture media.

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    Kawakami, Tomoya; Koike, Atsushi; Amano, Fumio

    2017-08-01

    The role of activated macrophages in the host defense against pathogens or tumor cells has been investigated extensively. Many researchers have been using various culture media in in vitro experiments using macrophages. We previously reported that J774.1/JA-4 macrophage-like cells showed great differences in their activated macrophage phenotypes, such as production of reactive oxygen, nitric oxide (NO) or cytokines depending on the culture medium used, either F-12 (Ham's F-12 nutrient mixture) or Dulbecco modified Eagle's medium (DMEM). To examine whether a difference in the culture medium would influence the functions of primary macrophages, we used BALB/c mouse peritoneal macrophages in this study. Among the activated macrophage phenotypes, the expression of inducible NO synthase in LPS- and/or IFN-γ-treated peritoneal macrophages showed the most remarkable differences between F-12 and DMEM; i.e., NO production by LPS- and/or IFN-γ-treated cells was far lower in DMEM than in F-12. Similar results were obtained with C57BL mouse peritoneal macrophages. Besides, dilution of F-12 medium with saline resulted in a slight decrease in NO production, whereas that of DMEM with saline resulted in a significant increase, suggesting the possibility that DMEM contained some inhibitory factor(s) for NO production. However, such a difference in NO production was not observed when macrophage-like cell lines were examined. These results suggest that phenotypes of primary macrophages could be changed significantly with respect to host inflammatory responses by the surrounding environment including nutritional factors and that these altered macrophage phenotypes might influence the biological host defense.

  17. Enhancement of Anti-Inflammatory Activity of Curcumin Using Phosphatidylserine-Containing Nanoparticles in Cultured Macrophages

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    Ji Wang

    2016-06-01

    Full Text Available Macrophages are one kind of innate immune cells, and produce a variety of inflammatory cytokines in response to various stimuli, such as oxidized low density lipoprotein found in the pathogenesis of atherosclerosis. In this study, the effect of phosphatidylserine on anti-inflammatory activity of curcumin-loaded nanostructured lipid carriers was investigated using macrophage cultures. Different amounts of phosphatidylserine were used in the preparation of curcumin nanoparticles, their physicochemical properties and biocompatibilities were then compared. Cellular uptake of the nanoparticles was investigated using a confocal laser scanning microscope and flow cytometry analysis in order to determine the optimal phosphatidylserine concentration. In vitro anti-inflammatory activities were evaluated in macrophages to test whether curcumin and phosphatidylserine have interactive effects on macrophage lipid uptake behavior and anti-inflammatory responses. Here, we showed that macrophage uptake of phosphatidylserine-containing nanostructured lipid carriers increased with increasing amount of phosphatidylserine in the range of 0%–8%, and decreased when the phosphatidylserine molar ratio reached over 12%. curcumin-loaded nanostructured lipid carriers significantly inhibited lipid accumulation and pro-inflammatory factor production in cultured macrophages, and evidently promoted release of anti-inflammatory cytokines, when compared with curcumin or phosphatidylserine alone. These results suggest that the delivery system using PS-based nanoparticles has great potential for efficient delivery of drugs such as curcumin, specifically targeting macrophages and modulation of their anti-inflammatory functions.

  18. Tick-borne encephalitis virus infection of cultured mouse macrophages

    Czech Academy of Sciences Publication Activity Database

    Ahantarig, A.; Růžek, Daniel; Vancová, Marie; Janowitz, A.; Šťastná, Hana; Tesařová, Martina; Grubhoffer, Libor

    2009-01-01

    Roč. 52, č. 5 (2009), s. 283-290 ISSN 0300-5526 R&D Projects: GA ČR GA524/06/1479; GA MŠk(CZ) LC06009 Institutional research plan: CEZ:AV0Z60220518 Keywords : tick-borne encephalitis * macrophage s * electron microscopy Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 1.106, year: 2009

  19. A new and efficient culture method for porcine bone marrow-derived M1- and M2-polarized macrophages.

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    Gao, Jiye; Scheenstra, Maaike R; van Dijk, Albert; Veldhuizen, Edwin J A; Haagsman, Henk P

    2018-06-01

    Macrophages play an important role in the innate immune system as part of the mononuclear phagocyte system (MPS). They have a pro-inflammatory signature (M1-polarized macrophages) or anti-inflammatory signature (M2-polarized macrophages) based on expression of surface receptors and secretion of cytokines. However, very little is known about the culture of macrophages from pigs and more specific about the M1 and M2 polarization in vitro. Porcine monocytes or mononuclear bone marrow cells were used to culture M1- and M2-polarized macrophages in the presence of GM-CSF and M-CSF, respectively. Surface receptor expression was measured with flow cytometry and ELISA was used to quantify cytokine secretion in response to LPS and PAM 3 CSK 4 stimulation. Human monocyte-derived macrophages were used as control. Porcine M1- and M2-polarized macrophages were cultured best using porcine GM-CSF and murine M-CSF, respectively. Cultures from bone marrow cells resulted in a higher yield M1- and M2-polarized macrophages which were better comparable to human monocyte-derived macrophages than cultures from porcine monocytes. Porcine M1-polarized macrophages displayed the characteristic fried egg shape morphology, lower CD163 expression and low IL-10 production. Porcine M2-polarized macrophages contained the spindle-like morphology, higher CD163 expression and high IL-10 production. Porcine M1- and M2-polarized macrophages can be most efficiently cultured from mononuclear bone marrow cells using porcine GM-CSF and murine M-CSF. The new culture method facilitates more refined studies of porcine macrophages in vitro, important for both porcine and human health since pigs are increasingly used as model for translational research. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  20. Labelling of cultured macrophages with novel magnetic nanoparticles

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    Hsiao, J.-K. [Department of Medical Imaging, National Taiwan University Hospital and College of Medicine, Taipei 100, Taiwan (China); Institute of Biomedical Engineering, National Taiwan University, Taipei 100, Taiwan (China); Tai, M.-F. [Department of Electronic Engineering and Graduate School of Opto-Mechatronics and Materials, WuFeng Institute of Technology, 117, Chian-Kuo Rd., Sec. 2, Ming-Hsiung, Chia-yi 621, Taiwan (China)]. E-mail: mftai@mail.wfc.edu.tw; Lee, Y.-C. [Department of Physics, National Chung Cheng University, Ming-Hsiung, Chia-yi 621, Taiwan (China); Yang, C.-Y. [Department of Medical Imaging, National Taiwan University Hospital and College of Medicine, Taipei 100, Taiwan (China); Wang, H.-Y. [Department of Physics, National Chung Cheng University, Ming-Hsiung, Chia-yi 621, Taiwan (China); Liu, H.-M. [Department of Medical Imaging, National Taiwan University Hospital and College of Medicine, Taipei 100, Taiwan (China); Fang, J.-S. [Department of Material Science and Engineering, National Formosa University, Huwei, Yunlin, Taiwan (China); Chen, S.-T. [Musculoskeletal Disease Center, J.L. Pettis VA Medical Center, Department of Biochemistry Loma Linda University, Loma Linda, CA 92357 (United States)

    2006-09-15

    Magnetic resonance (MR) imaging is capable of demonstrating human anatomy and pathological conditions. Iron oxide magnetic nanoparticles (MNPs) have been used in MR imaging as liver-specific contrast medium, cellular and molecular imaging probes. Because few studies focused on the MNPs other than iron oxides, we developed FeNi alloy MNPs coated with polyethylenimine (PEI). In this study, we demonstrated PEI-coated FeNi MNPs are able to label the cells, which could be detected in MR imaging. For labelling purpose, MNPs were incubated with mouse macrophage cell line (Raw 264.7) for 24 h and these PEI-labelled FeNi alloy MNPs can be uptaken by macrophages efficiently compared with Ferucarbotran, a commercialized superparamagnetic iron oxide (SPIO) under flow cytometry measurement. Besides, these cells labelled with MNPs could be imaged in MR with the identical potency as Ferucarbotran. Further investigation of the cells using Prussian blue staining revealed that FeNi alloy MNPs inside the cells is not oxidized. This phenomenon alleviated the consideration of potential risk of nickel toxicity. We conclude that PEI-coated FeNi MNPs could be candidate for MR contrast medium.

  1. Labelling of cultured macrophages with novel magnetic nanoparticles

    International Nuclear Information System (INIS)

    Hsiao, J.-K.; Tai, M.-F.; Lee, Y.-C.; Yang, C.-Y.; Wang, H.-Y.; Liu, H.-M.; Fang, J.-S.; Chen, S.-T.

    2006-01-01

    Magnetic resonance (MR) imaging is capable of demonstrating human anatomy and pathological conditions. Iron oxide magnetic nanoparticles (MNPs) have been used in MR imaging as liver-specific contrast medium, cellular and molecular imaging probes. Because few studies focused on the MNPs other than iron oxides, we developed FeNi alloy MNPs coated with polyethylenimine (PEI). In this study, we demonstrated PEI-coated FeNi MNPs are able to label the cells, which could be detected in MR imaging. For labelling purpose, MNPs were incubated with mouse macrophage cell line (Raw 264.7) for 24 h and these PEI-labelled FeNi alloy MNPs can be uptaken by macrophages efficiently compared with Ferucarbotran, a commercialized superparamagnetic iron oxide (SPIO) under flow cytometry measurement. Besides, these cells labelled with MNPs could be imaged in MR with the identical potency as Ferucarbotran. Further investigation of the cells using Prussian blue staining revealed that FeNi alloy MNPs inside the cells is not oxidized. This phenomenon alleviated the consideration of potential risk of nickel toxicity. We conclude that PEI-coated FeNi MNPs could be candidate for MR contrast medium

  2. Macrophage secretory products selectively stimulate dermatan sulfate proteoglycan production in cultured arterial smooth muscle cells

    International Nuclear Information System (INIS)

    Edwards, I.J.; Wagner, W.D.; Owens, R.T.

    1990-01-01

    Arterial dermatan sulfate proteoglycan has been shown to increase with atherosclerosis progression, but factors responsible for this increase are unknown. To test the hypothesis that smooth muscle cell proteoglycan synthesis may be modified by macrophage products, pigeon arterial smooth muscle cells were exposed to the media of either cholesteryl ester-loaded pigeon peritoneal macrophages or a macrophage cell line P388D1. Proteoglycans radiolabeled with [35S]sulfate and [3H]serine were isolated from culture media and smooth muscle cells and purified following precipitation with 1-hexadecylpyridinium chloride and chromatography. Increasing concentrations of macrophage-conditioned media were associated with a dose-response increase in [35S]sulfate incorporation into secreted proteoglycans, but there was no change in cell-associated proteoglycans. Incorporation of [3H]serine into total proteoglycan core proteins was not significantly different (5.2 X 10(5) dpm and 5.5 X 10(5) disintegrations per minute (dpm) in control and conditioned media-treated cultures, respectively), but selective effects were observed on individual proteoglycan types. Twofold increases in dermatan sulfate proteoglycan and limited degradation of chondroitin sulfate proteoglycan were apparent based on core proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Immunoinhibition studies indicated that interleukin-1 was involved in the modulation of proteoglycan synthesis by macrophage-conditioned media. These data provide support for the role of macrophages in alteration of the matrix proteoglycans synthesized by smooth muscle cells and provide a mechanism to account for the reported increased dermatan sulfate/chondroitin sulfate ratios in the developing atherosclerotic lesion

  3. Development of ostrich thrombocytes and monocyte-derived macrophages in culture and the control of Toxoplasma gondii reproduction after macrophage activation.

    Science.gov (United States)

    Miranda, Farlen J B; Damasceno-Sá, João Cláudio; DaMatta, Renato A

    2016-01-01

    Raising ostriches became an important economic activity after their products became commodities. The health of farm animals is of paramount importance, so assessing basic immunological responses is necessary to better understand health problems. We developed a method to obtain ostrich thrombocytes and macrophages. The thrombocytes died by apoptosis after 48 h in culture, and the macrophages expanded in size and increased the number of acidic compartments. Macrophages were activated by chicken interferon-γ, producing high levels of nitric oxide. Toxoplasma gondii was able to infect these macrophages, and activation controlled parasitic reproduction. T. gondii, however, persisted in these cells, and infection reduced the production of nitric oxide. These results are important for the future assessment of the basic cellular and immunobiology of ostriches and demonstrate T. gondii suppression of nitric oxide production. © 2016 Poultry Science Association Inc.

  4. In vitro methods of assessing ocular biocompatibility using THP-1-derived macrophages.

    Science.gov (United States)

    McCanna, David Joseph; Barthod-Malat, Aurore V; Gorbet, Maud B

    2015-01-01

    Macrophages play an important role in the elimination of infections, the removal of debris and in tissue repair after infection and trauma. In vitro models that assess ocular biomaterials for toxicity typically focus on the effects of these materials on epithelial or fibroblast cells. This investigation evaluated known ocular toxins deposited on model materials for their effects on the viability and activation of macrophages. THP-1-derived macrophages were cultured onto silicone films (used as a base biomaterial) deposited with chemical toxins (benzalkonium chloride (BAK), zinc diethyldithiocarbamate (ZDEC) and lipopolysaccharide (LPS)). Utilizing three fluorescent dyes calcein, ethidium homodimer-1 (EthD-1) and annexin V, the viability of macrophages attached to the biomaterial was determined using confocal microscopy. Propidium iodide (PI) staining and alamarBlue® (resazurin) reduction were used to assess cell death and metabolic activity. CD14, CD16, CD33, CD45, and CD54 expression of adherent macrophages, were also evaluated to detect LPS activation of macrophages using flow cytometry. The sensitivity of this test battery was demonstrated as significant toxicity from treated surfaces with ZDEC (0.001-0.01%), and BAK (0.001%-0.1%) was detected. Also, macrophage activation could be detected by measuring CD54 expression after exposure to adsorbed LPS. These in vitro methods will be helpful in determining the toxicity potential of new ocular biomaterials.

  5. Functional crosstalk in culture between macrophages and trigeminal sensory neurons of a mouse genetic model of migraine

    Directory of Open Access Journals (Sweden)

    Franceschini Alessia

    2012-11-01

    Full Text Available Abstract Background Enhanced activity of trigeminal ganglion neurons is thought to underlie neuronal sensitization facilitating the onset of chronic pain attacks, including migraine. Recurrent headache attacks might establish a chronic neuroinflammatory ganglion profile contributing to the hypersensitive phenotype. Since it is difficult to study this process in vivo, we investigated functional crosstalk between macrophages and sensory neurons in primary cultures from trigeminal sensory ganglia of wild-type (WT or knock-in (KI mice expressing the Cacna1a gene mutation (R192Q found in familial hemiplegic migraine-type 1. After studying the number and morphology of resident macrophages in culture, the consequences of adding host macrophages on macrophage phagocytosis and membrane currents mediated by pain-transducing P2X3 receptors on sensory neurons were examined. Results KI ganglion cultures constitutively contained a larger number of active macrophages, although no difference in P2X3 receptor expression was found. Co-culturing WT or KI ganglia with host macrophages (active as much as resident cells strongly stimulated single cell phagocytosis. The same protocol had no effect on P2X3 receptor expression in WT or KI co-cultures, but it largely enhanced WT neuron currents that grew to the high amplitude constitutively seen for KI neurons. No further potentiation of KI neuronal currents was observed. Conclusions Trigeminal ganglion cultures from a genetic mouse model of migraine showed basal macrophage activation together with enhanced neuronal currents mediated by P2X3 receptors. This phenotype could be replicated in WT cultures by adding host macrophages, indicating an important functional crosstalk between macrophages and sensory neurons.

  6. Functional crosstalk in culture between macrophages and trigeminal sensory neurons of a mouse genetic model of migraine.

    Science.gov (United States)

    Franceschini, Alessia; Nair, Asha; Bele, Tanja; van den Maagdenberg, Arn Mjm; Nistri, Andrea; Fabbretti, Elsa

    2012-11-21

    Enhanced activity of trigeminal ganglion neurons is thought to underlie neuronal sensitization facilitating the onset of chronic pain attacks, including migraine. Recurrent headache attacks might establish a chronic neuroinflammatory ganglion profile contributing to the hypersensitive phenotype. Since it is difficult to study this process in vivo, we investigated functional crosstalk between macrophages and sensory neurons in primary cultures from trigeminal sensory ganglia of wild-type (WT) or knock-in (KI) mice expressing the Cacna1a gene mutation (R192Q) found in familial hemiplegic migraine-type 1. After studying the number and morphology of resident macrophages in culture, the consequences of adding host macrophages on macrophage phagocytosis and membrane currents mediated by pain-transducing P2X3 receptors on sensory neurons were examined. KI ganglion cultures constitutively contained a larger number of active macrophages, although no difference in P2X3 receptor expression was found. Co-culturing WT or KI ganglia with host macrophages (active as much as resident cells) strongly stimulated single cell phagocytosis. The same protocol had no effect on P2X3 receptor expression in WT or KI co-cultures, but it largely enhanced WT neuron currents that grew to the high amplitude constitutively seen for KI neurons. No further potentiation of KI neuronal currents was observed. Trigeminal ganglion cultures from a genetic mouse model of migraine showed basal macrophage activation together with enhanced neuronal currents mediated by P2X3 receptors. This phenotype could be replicated in WT cultures by adding host macrophages, indicating an important functional crosstalk between macrophages and sensory neurons.

  7. Solubilization of 241AmO2 in alveolar macrophage cultures

    International Nuclear Information System (INIS)

    Robinson, A.V.; Schneider, R.P.

    1980-01-01

    Cultured rabbit alveolar macrophages were used to study the effect of phagocytosis on the solubilization of 241 AmO 2 . A comparison was made of the solubility of phagocytized AmO 2 and AmO 2 in cell-free media, in the presence and absence of 0.1 mM DTPA. A time-dependent increase of 26% in the soluble (0.1-μm filtrate) intracellular americium fraction was seen in macrophages cultured for 3 days. The addition of 0.1mM DTPA to culture medium resulted in an increase of 36% over the same time period. In contrast, cell-free media without DTPA resulted in less than a 2% increase in solubility after 4 days of incubation, while addition of 0.1mM DTPA resulted in a 5% increase over the same time period. These results indicate cell-mediated solbuilization of phagocytized AmO 2 by macrophages

  8. High level of expression of recombinant human granulocyte-macrophage colony stimulating factor in transgenic rice cell suspension culture

    DEFF Research Database (Denmark)

    Shin, Yun-Ji; Hong, Shin-Young; Kwon, Tae-Ho

    2003-01-01

    Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco cell suspension cultures. However, the amount of hGM-CSF accumulated in the culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome...... of recombinant hGM-CSF in transgenic rice cell suspension culture and protease activity of this culture medium was low compared to that of tobacco culture system....

  9. Radiometric macrophage culture assay for rapid evaluation of antileprosy activity of rifampin

    Energy Technology Data Exchange (ETDEWEB)

    Mittal, A.; Seshadri, P.S.; Prasad, H.K.; Sathish, M.; Nath, I.

    1983-10-01

    The antileprosy effect of rifampin was evaluated by a newly developed rapid in vitro assay wherein 31 human-derived strains and 1 armadillo-derived strain of Mycobacterium leprae were maintained for 2 and 3 weeks, respectively, in murine and human macrophages in the presence of (3H)thymidine. Of these strains, 27 showed significant incorporation of the radiolabel in cultures of live bacilli as compared with control cultures of heat-killed bacilli of the same strain. Consistent and significant inhibition of (3H)thymidine uptake was observed in M. leprae resident cultures with 3 to 200 ng of rifampin per ml as compared with similar cultures without the drug. In general, an increase in percent inhibition was seen from 3 to 20 ng/ml, with marginal increases at 40, 50, and 100 ng/ml. M. leprae strains appear to be remarkably susceptible to this drug in the in vitro assay.

  10. Radiometric macrophage culture assay for rapid evaluation of antileprosy activity of rifampin

    International Nuclear Information System (INIS)

    Mittal, A.; Seshadri, P.S.; Prasad, H.K.; Sathish, M.; Nath, I.

    1983-01-01

    The antileprosy effect of rifampin was evaluated by a newly developed rapid in vitro assay wherein 31 human-derived strains and 1 armadillo-derived strain of Mycobacterium leprae were maintained for 2 and 3 weeks, respectively, in murine and human macrophages in the presence of [3H]thymidine. Of these strains, 27 showed significant incorporation of the radiolabel in cultures of live bacilli as compared with control cultures of heat-killed bacilli of the same strain. Consistent and significant inhibition of [3H]thymidine uptake was observed in M. leprae resident cultures with 3 to 200 ng of rifampin per ml as compared with similar cultures without the drug. In general, an increase in percent inhibition was seen from 3 to 20 ng/ml, with marginal increases at 40, 50, and 100 ng/ml. M. leprae strains appear to be remarkably susceptible to this drug in the in vitro assay

  11. Isolation of human monocytes by double gradient centrifugation and their differentiation to macrophages in teflon-coated cell culture bags.

    Science.gov (United States)

    Menck, Kerstin; Behme, Daniel; Pantke, Mathias; Reiling, Norbert; Binder, Claudia; Pukrop, Tobias; Klemm, Florian

    2014-09-09

    Human macrophages are involved in a plethora of pathologic processes ranging from infectious diseases to cancer. Thus they pose a valuable tool to understand the underlying mechanisms of these diseases. We therefore present a straightforward protocol for the isolation of human monocytes from buffy coats, followed by a differentiation procedure which results in high macrophage yields. The technique relies mostly on commonly available lab equipment and thus provides a cost and time effective way to obtain large quantities of human macrophages. Briefly, buffy coats from healthy blood donors are subjected to a double density gradient centrifugation to harvest monocytes from the peripheral blood. These monocytes are then cultured in fluorinated ethylene propylene (FEP) Teflon-coated cell culture bags in the presence of macrophage colony-stimulating factor (M-CSF). The differentiated macrophages can be easily harvested and used for subsequent studies and functional assays. Important methods for quality control and validation of the isolation and differentiation steps will be highlighted within the protocol. In summary, the protocol described here enables scientists to routinely and reproducibly isolate human macrophages without the need for cost intensive tools. Furthermore, disease models can be studied in a syngeneic human system circumventing the use of murine macrophages.

  12. Effects of extracellular matrix proteins on macrophage differentiation, growth, and function: comparison of liquid and agar culture systems

    Science.gov (United States)

    Armstrong, J. W.; Chapes, S. K.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Both spaceflight and skeletal unloading suppress the haematopoietic differentiation of macrophages (Sonnenfeld et al., Aviat. Space Environ. Med., 61:648-653, 1990; Armstrong et al., J. Appl. Physiol., 75:2734-2739, 1993). The mechanism behind this reduction in haematopoiesis has yet to be elucidated. However, changes in bone marrow extracellular matrix (ECM) may be involved. To further understand the role of ECM products in macrophage differentiation, we have performed experiments evaluating the effects of fibronectin, laminin, collagen type I, and collagen type IV on macrophage development and function. Bone marrow-derived macrophages cultured on four different ECM substrates in liquid culture medium showed less growth than those cultured on plastic. Significant morphological differences were seen on each of the substrates used. Phenotypically and functionally, as measured by class II major histocompatibility molecule (MHCII) expression, MAC-2 expression, and the secretion of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha), these macrophages were similar. In contrast, bone marrow-derived macrophages cultured in suspension, using agar, showed no difference in growth when exposed to ECM proteins. However, IL-6 and TNF-alpha secretion was affected by fibronectin, laminin, collagen type I, and collagen type IV in a concentration-dependent manner. We conclude that the ECM products fibronectin, laminin, collagen type I, and collagen type IV have profound effects on macrophage development and function. Additionally, we suggest that an ECM-supplemented agar culture system provides an environment more analogous to in vivo bone marrow than does a traditional liquid culture system.

  13. Structural characterization of blackberry wine polysaccharides and immunomodulatory effects on LPS-activated RAW 264.7 macrophages.

    Science.gov (United States)

    Cordeiro Caillot, Adriana Rute; de Lacerda Bezerra, Iglesias; Palhares, Laís Cristina Gusmão Ferreira; Santana-Filho, Arquimedes Paixão; Chavante, Suely Ferreira; Sassaki, Guilherme Lanzi

    2018-08-15

    Three polysaccharide fractions were isolated from blackberry wine. The crude extract BWPs was obtained with ethanol precipitation and freeze-thawing process, it was then submitted to Fehling treatment, giving soluble BWPFs and insoluble BWPFp fractions. These fractions were characterized by Gas Chromatography-Mass Spectrometry (GC-MS) and Nuclear Magnetic Resonance (NMR). Major polysaccharides were identified for each fraction: mannan, type II arabinogalactan and type I rhamnogalacturonan for BWPs, a mannan formed by a major chain of α-Manp(1 → 6)-linked units, O-2 substituted with α-d-Manp(1 → 2)-linked side chains for BWPFp and a AG II formed by a major chain of β-d-Galp(1 → 3)-linked, substituted at O-6 by side chains of the β-d-Galp(1 → 6)-linked, which then are substituted at O-3 by non-reducing units of α-l-Araf and a RG I, formed by [→4)-α-d-GalpA-(1 → 2)-α-l-Rhap-(1→] n for BWPFs. Anti-inflammatory effects of polysaccharide fractions were evaluated in RAW 264.7 cells. Fractions markedly reduced nitric oxide (NO) and pro-inflammatory cytokine production (TNF-α and IL-1β) in LPS-treated cells. Copyright © 2018 Elsevier Ltd. All rights reserved.

  14. Stimulating effects of vitamin A on PHA- and LPS-activated lymphocytes

    International Nuclear Information System (INIS)

    Liu Fengju; Su Liaoyuan

    1993-03-01

    A method of 3 H-TdR incorporation in lymphocytes activated by PHA was applied. The incubation with vitamin A at a concentration of 5 μg/ml for 16 hours was followed by the incorporation, the counts per minute (cpm) of radioactivity in lymphocytes increased significantly. When the concentration was greater than 25 μg/ml or the incubation at concentration of 10 μg/ml for 72 hours, the incorporation value of 3 H-TdR was remarkably decreased. The lymphocytes irradiated by 60 Co gamma rays at different doses were incubated in vitro with vitamin A of 5 μg/ml concentration for 16 hours, the cpm from 3 H-TdR incorporation in DNA was higher than that in control group with no vitamin A. Tests on 10 cancer patients showed that after one course of treatment with radiotherapy (75 ∼ 184 Gy), the incorporation value of 3 H-TdR was decreased significantly. After adding more 5 μg/ml concentration of vitamin A, the incorporation value of 3 H-TdR was remarkably greater than in the control group. These results indicated that vitamin A at adequate level is able to enhance the effect of transformation of PHA-activated lymphocytes, but the incorporation in LPS-activated B lymphocytes enhanced by vitamin A was not observed

  15. A randomized clinical trial to evaluate the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) in embryo culture medium for in vitro fertilization

    DEFF Research Database (Denmark)

    Ziebe, Søren; Loft, Anne; Povlsen, Betina B

    2013-01-01

    To evaluate the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) in embryo culture medium on ongoing implantation rate (OIR).......To evaluate the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) in embryo culture medium on ongoing implantation rate (OIR)....

  16. Evaluation of gold nanoparticles biocompatibility: a multiparametric study on cultured endothelial cells and macrophages

    International Nuclear Information System (INIS)

    Orlando, Antonina; Colombo, Miriam; Prosperi, Davide; Corsi, Fabio; Panariti, Alice; Rivolta, Ilaria; Masserini, Massimo; Cazzaniga, Emanuela

    2016-01-01

    Colloidal gold nanoparticles (AuNPs) have been considered an established advanced tool in biomedicine thanks to their physicochemical properties combined with nanoscale size ideal for the interrogation of biological systems. However, such properties are believed to be a possible major cause of “unsafety” of these materials. For this reason, increasing attention has been due to assess how AuNPs affect cell behaviour in cultures. In the present work, we investigate the effects of PMA polymer-coated Au@PMA PEGylated (8.9 ± 0.2 nm) or not (6.6 ± 0.6 nm) on HUVECs and macrophages, which are model cell types likely to interact with Au@PMA after systemic administration in vivo, using a multiparametric approach. Testing different NPs concentrations and incubation times, we analysed the effect of such NPs on cell viability, oxidative stress, inflammatory processes, and cell uptake. Our data suggested that Au@PMA reduced the cell viability mostly through oxidative stress and TNF-α production after the uptake by HUVECs and macrophages, respectively. PEGylation conferred improved biocompatibility to Au@PMA in particular, no significant effects on any parameter tested could be observed at a concentration of 20 µg mL"−"1. This approach allowed us to explore different aspects of cell-NPs interaction and to suggest that these NPs could be potentially used for the in vivo studies.

  17. Evaluation of gold nanoparticles biocompatibility: a multiparametric study on cultured endothelial cells and macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Orlando, Antonina [University of Milano-Bicocca, Department of Health Sciences (Italy); Colombo, Miriam; Prosperi, Davide [University of Milano-Bicocca, Department of Biotechnology and Biosciences (Italy); Corsi, Fabio [University of Milano, Department of Biomedical and Clinical Sciences (Italy); Panariti, Alice; Rivolta, Ilaria; Masserini, Massimo; Cazzaniga, Emanuela, E-mail: emanuela.cazzaniga@unimib.it [University of Milano-Bicocca, Department of Health Sciences (Italy)

    2016-03-15

    Colloidal gold nanoparticles (AuNPs) have been considered an established advanced tool in biomedicine thanks to their physicochemical properties combined with nanoscale size ideal for the interrogation of biological systems. However, such properties are believed to be a possible major cause of “unsafety” of these materials. For this reason, increasing attention has been due to assess how AuNPs affect cell behaviour in cultures. In the present work, we investigate the effects of PMA polymer-coated Au@PMA PEGylated (8.9 ± 0.2 nm) or not (6.6 ± 0.6 nm) on HUVECs and macrophages, which are model cell types likely to interact with Au@PMA after systemic administration in vivo, using a multiparametric approach. Testing different NPs concentrations and incubation times, we analysed the effect of such NPs on cell viability, oxidative stress, inflammatory processes, and cell uptake. Our data suggested that Au@PMA reduced the cell viability mostly through oxidative stress and TNF-α production after the uptake by HUVECs and macrophages, respectively. PEGylation conferred improved biocompatibility to Au@PMA in particular, no significant effects on any parameter tested could be observed at a concentration of 20 µg mL{sup −1}. This approach allowed us to explore different aspects of cell-NPs interaction and to suggest that these NPs could be potentially used for the in vivo studies.

  18. Evaluation of gold nanoparticles biocompatibility: a multiparametric study on cultured endothelial cells and macrophages

    Science.gov (United States)

    Orlando, Antonina; Colombo, Miriam; Prosperi, Davide; Corsi, Fabio; Panariti, Alice; Rivolta, Ilaria; Masserini, Massimo; Cazzaniga, Emanuela

    2016-03-01

    Colloidal gold nanoparticles (AuNPs) have been considered an established advanced tool in biomedicine thanks to their physicochemical properties combined with nanoscale size ideal for the interrogation of biological systems. However, such properties are believed to be a possible major cause of "unsafety" of these materials. For this reason, increasing attention has been due to assess how AuNPs affect cell behaviour in cultures. In the present work, we investigate the effects of PMA polymer-coated Au@PMA PEGylated (8.9 ± 0.2 nm) or not (6.6 ± 0.6 nm) on HUVECs and macrophages, which are model cell types likely to interact with Au@PMA after systemic administration in vivo, using a multiparametric approach. Testing different NPs concentrations and incubation times, we analysed the effect of such NPs on cell viability, oxidative stress, inflammatory processes, and cell uptake. Our data suggested that Au@PMA reduced the cell viability mostly through oxidative stress and TNF-α production after the uptake by HUVECs and macrophages, respectively. PEGylation conferred improved biocompatibility to Au@PMA in particular, no significant effects on any parameter tested could be observed at a concentration of 20 µg mL-1. This approach allowed us to explore different aspects of cell-NPs interaction and to suggest that these NPs could be potentially used for the in vivo studies.

  19. The effects of beryllium metal particles on the viability and function of cultured rat alveolar macrophages

    International Nuclear Information System (INIS)

    Finch, G.L.; Lowther, W.T.; Hoover, M.D.; Brooks, A.L.

    1988-01-01

    Rat pulmonary alveolar macrophages (PAM) were exposed in vitro to beryllium metal particles. The particles used were relatively large (Be-II) and small (Be-V) size fractions of beryllium metal obtained from an aerosol cyclone, and a beryllium metal aerosol generated by laser vaporization of beryllium metal in an argon atmosphere (Be-L). Glass beads (GB) were used as a negative control particle. The endpoints examined included cell killing (trypan blue dye exclusion) and phagocytic ability (sheep red blood cell uptake). Phagocytic ability was inhibited by beryllium particles at concentrations that did not cause appreciable cell killing. Results based on the mass concentration of particles in culture medium were transformed by the amount of specific surface area of the particles to permit expression of toxicity on the basis of amount of surface area of particles per unit volume of culture medium. On a mass concentration basis, the order of cytotoxicity was Be-L > Be-V ∼ Be-II > GB; for inhibition of phagacytosis, the cytotoxicity order was Be-L ∼ Be-V > Be-II > GB. On a surface area concentration basis, the order of toxicity for viability was altered to Be-II > Be-L ∼ Be-V (with GB indeterminant) and to Be-V > Be-II ∼ Be-L > GB for inhibition of phagocytosis. We conclude that there are factors in addition to specific surface area that influence the expression of toxic effects in cultured PAM. (author)

  20. Cultivation of murine bone marrow macrophages in sponges: a method that permits recovery of viable cultured cells

    Energy Technology Data Exchange (ETDEWEB)

    Akporiaye, E T; Stewart, S; Stewart, C C

    1984-01-01

    Various investigators have cultured murine bone marrow or peritoneal cells in vitro on glass or plastic surfaces with the ultimate aim of retrieving adherent macrophages for morphologic and functional evaluation. The removal of these adherent macrophages by conventional techniques has been consistently accompanied by low yield and significant cell damage. The authors report here a simple technique for culturing murine bone marrow cells in gelatin sponges (Spongostan and Gelfoam) in growth medium containing 10% fetal bovine serum and 10% L-cell conditioned medium. Viable cells were retrieved from the sponges in 10 min by digestion with collagenase. The in situ growth kinetics were similar to those found for cells cultured on plastic dishes. The recovered cells were adherent, phagocytic, positive for Fc ..gamma.. receptors, and had esterase activity. 23 references, 1 figure, 1 table.

  1. Macrophages Under Low Oxygen Culture Conditions Respond to Ion Parametric Resonance Magnetic Fields

    Science.gov (United States)

    Macrophages, when entering inflamed tissue, encounter low oxygen tension due to the impairment of blood supply and/or the massive infiltration of cells that consume oxygen. Previously, we showed that such macrophages release more bacteriotoxic hydrogen peroxide (H202) when expose...

  2. Isoferritins in rat Kupffer cells, hepatocytes, and extrahepatic macrophages. Biosynthesis in cell suspensions and cultures in response to iron

    International Nuclear Information System (INIS)

    Doolittle, R.L.; Richter, G.W.

    1981-01-01

    Cultures of Kupffer cells and of hepatocytes, prepared from single rat livers, synthesized ferritin protein equally efficiently. In culture but not in suspension, both sorts of cells responded significantly to stimulation with iron by increased ferritin synthesis. As determined by isoelectric focusing, the isoferritin profiles of newly synthesized 14 -labeled Kupffer cell and hepatocyte ferritin were identical, each having three bands. However, unlabeled ferritin, extracted from nonparenchymal liver cells (mainly Kupffer and endothelial cells) of iron-loaded rats, contained an acidic isoferritin that was not present in hepatocyte ferritin. Investigation of ferritin synthesis in cultured peritoneal and alveolar macrophages yielded similar results. The isofocusing profile of newly synthesized peritoneal macrophage ferritin was indistinguishable from the profile of fresh Kupffer cell or hepatocyte ferritin. Thus, the three isoferritins common to Kupffer cells, hepatocytes, and extrahepatic macrophages are neither cell- nor tissue-specific. However, modifications on intracellular storage may affect the isofocusing properties. The findings, although consistent with the LnH24-n subunit model of ferritin protein, indicate identical restrictive genomic control of the H:L ratios in these sorts of cells. Further, they make it probable that Kupffer cell ferritin iron, originating by endogenous synthesis, is the principal source of Kupffer cell hemosiderin iron

  3. [EVALUATION OF THE HUMAN SENSITIVITY TO SMALLPOX VIRUS BY THE PRIMARY CULTURES OF THE MONOCYTE-MACROPHAGES].

    Science.gov (United States)

    Zamedyanskaya, A S; Titova, K A; Sergeev, Al A; Kabanov, A S; Bulychev, L E; Sergeev, Ar A; Galakhova, D O; Nesterov, A E; Nosareva, O V; Shishkina, L N; Taranov, O S; Omigov, V V; Agafonov, A P; Sergeev, A N

    2016-01-01

    Studies of the primary cultures of granulocytes, mononuclear, and monocyte-macrophage cells derived from human blood were performed using variola virus (VARV) in the doses of 0.001-0.021 PFU/cell (plaques-forming units per cell). Positive dynamics of the virus accumulation was observed only in the monocyte-macrophages with maximum values of virus concentration (5.0-5.5 Ig PFU/ml) mainly within six days after the infection. The fact of VARV replication in the monocyte-macrophages was confirmed by the data of electron microscopy. At the same time, virus vaccines when tested in doses 3.3 and 4.2 Ig PFU/ml did not show the ability to reproduce in these human cells. The people sensitivity to VARV as assessed from the data obtained on human monocyte-macrophages corresponded to -1 PFU (taking into account the smooth interaction of the virus in the body to the cells of this type), which is consistent to previously found theoretical data on the virus sensitivity. The human susceptibility to VARV assessed experimentally can be used to predict the adequacy of developed smallpox models (in vivo) based on susceptible animals. This is necessary for reliable assessment of the efficiency of development of drugs for treatment and prophylaxis of the smallpox.

  4. Degradation of connective tissue matrices by macrophages. III. Morphological and biochemical studies on extracellular, pericellular, and intracellular events in matrix proteolysis by macrophages in culture

    International Nuclear Information System (INIS)

    Werb, Z.; Bainton, D.F.; Jones, P.A.

    1980-01-01

    The aim of the present study was to determine the localization of macrophage-mediated degradation of matrix proteins. The sites of matrix degradation were examined ultrastructurally, and the effects of modulation of macrophage secretion, endocytosis, and activity of macrophage hydrolases on matrix degradation were monitored biochemically

  5. Identification of blue staining vaccine-derived material in inflammatory lesions using cultured canine macrophages.

    Science.gov (United States)

    Scruggs, Jennifer L; LeBlanc, Casey J

    2015-03-01

    Vaccine reactions are described in cytology textbooks as having eosinophilic to magenta colored globules within and admixed with inflammatory cells. Recently, we have seen increased numbers of inflammatory lesions containing blue to blue-gray globular material, with historical information suggesting an association with rabies vaccination. The purpose of the study was to confirm the blue-gray and the eosinophilic material observed microscopically in some inflammatory lesions as being vaccine-derived. Three different vaccines were cytocentrifuged and Wright stained. Vaccine aliquots were also added to the culture media of canine-derived macrophages for 24 hours and the cells subsequently harvested, cytocentrifuged, and Wright stained. The globular material present in both preparations was compared to that observed in vaccine-induced inflammatory lesions. Morin staining was used to identify metal within vaccine material in both in vitro- and in vivo-derived cytology samples. Vaccine-derived material has a characteristic color and appearance. Appearance of the material was consistent in cytologic samples, in cells incubated with the vaccine, and in cytocentrifuged preparations of the vaccine vial contents. The blue-gray globules stained positively for Morin stain, while the eosinophilic material did not stain. Vaccine-induced inflammatory lesions may contain blue to blue-gray or magenta stained globular material. Blue-gray material was associated with administration of rabies vaccine Imrab 3 TF and the observed material may be metal-containing adjuvant. Magenta material was associated with other vaccines and negative for Morin stain, suggesting a metal-free adjuvant. © 2014 American Society for Veterinary Clinical Pathology.

  6. Oxidative damage to DNA by diesel exhaust particle exposure in co-cultures of human lung epithelial cells and macrophages

    DEFF Research Database (Denmark)

    Jantzen, Kim; Roursgaard, Martin; Madsen, Claus Desler

    2012-01-01

    Studies in mono-culture of cells have shown that diesel exhaust particles (DEPs) increase the production of reactive oxygen species (ROS) and oxidative stress-related damage to DNA. However, the level of particle-generated genotoxicity may depend on interplay between different cell types, e.g. lung...... treatment with standard reference DEPs, SRM2975 and SRM1650b. The exposure to DEPs did not affect the colony-forming ability of A549 cells in co-culture with THP-1a cells. The DEPs generated DNA strand breaks and oxidatively damaged DNA, measured using the alkaline comet assay as formamidopyrimidine...... relationship between levels of respiration and ROS production. In conclusion, exposure of mono-cultured cells to DEPs generated oxidative stress to DNA, whereas co-cultures with macrophages had lower levels of oxidatively damaged DNA than A549 epithelial cells....

  7. Three-Dimensional Organotypic Co-Culture Model of Intestinal Epithelial Cells and Macrophages to Study "Salmonella Enterica" Colonization Patterns

    Science.gov (United States)

    Ott, Mark; Yang, J; Barilla, J.; Crabbe, A.; Sarker, S. F.; Liu, Y.

    2017-01-01

    Three-dimensional/3-D organotypic models of human intestinal epithelium mimic the differentiated form and function of parental tissues often not exhibited by 2-D monolayers and respond to Salmonella in ways that reflect in vivo infections. To further enhance the physiological relevance of 3-D models to more closely approximate in vivo intestinal microenvironments during infection, we developed and validated a novel 3-D intestinal co-culture model containing multiple epithelial cell types and phagocytic macrophages, and applied to study enteric infection by different Salmonella pathovars.

  8. Inhibitors of nuclear factor kappa B cause apoptosis in cultured macrophages

    Directory of Open Access Journals (Sweden)

    E. E. Mannick

    1997-01-01

    Full Text Available The precise role of the transcription factor nuclear factor kappa B (NF- κB in the regulation of cell survival and cell death is still unresolved and may depend on cell type and position in the cell cycle. The aim of this study was to determine if three pharmacologic inhibitors of NF-κB, pyrrolidine dithiocarbamate, N-tosyl-L-lysl chloromethyl ketone and calpain I inhibitor, induce apoptosis in a murine macrophage cell line (RAW 264.7 at doses similar to those required for NF-κB inhibition. We found that each of the three inhibitors resulted in a dose- and time-dependent increase in morphologic indices of apoptosis in unstimulated, LPS-stimulated and TNF-stimulated cells. Lethal doses were consistent with those required for NF- κB inhibition. We conclude that nuclear NF-κB activation may represent an important survival mechanism in macrophages.

  9. Metformin affects the features of a human hepatocellular cell line (HepG2) by regulating macrophage polarization in a co-culture microenviroment.

    Science.gov (United States)

    Chen, Miaojiao; Zhang, Jingjing; Hu, Fang; Liu, Shiping; Zhou, Zhiguang

    2015-11-01

    Accumulating evidence suggests an association between diabetes and cancer. Inflammation is a key event that underlies the pathological processes of the two diseases. Metformin displays anti-cancer effects, but the mechanism is not completely clear. This study investigated whether metformin regulated the microenvironment of macrophage polarization to affect the characteristics of HepG2 cells and the possible role of the Notch-signalling pathway. RAW264.7 macrophages were cultured alone or co-cultured with HepG2 cells and treated with metformin. We analysed classical (M1) and alternative (M2) gene expression in RAW264.7 cells using quantitative real-time polymerase chain reaction. Changes in mRNA and protein expressions of Notch signalling in both cell types were also detected using quantitative real-time polymerase chain reaction and Western-blotting analyses. The proliferation, apoptosis and migration of HepG2 cells were detected using Cell Titer 96 AQueous One Solution Cell Proliferation Assay (MTS) (Promega Corporation, Fitchburg, WI, USA), Annexin V-FITC/PI (7SeaPharmTech, Shanghai, China) and the cell scratch assay, respectively. Metformin induced single-cultured RAW264.7 macrophages with an M2 phenotype but attenuated the M2 macrophage differentiation and inhibited monocyte chemoattractant protein-1 (MCP-1) secretion in a co-culture system. The co-cultured group of metformin pretreatment activated Notch signalling in macrophages but repressed it inHepG2 cells. Co-culture also promoted the proliferation and migration of HepG2 cells. However, along with the enhanced apoptosis, the proliferation and the migration of HepG2 cells were remarkably inhibited in another co-culture system with metformin pretreatment. Metformin can skew RAW264.7 macrophages toward different phenotypes according to changes in the microenvironment, which may affect the inflammatory conditions mediated by macrophages, induce apoptosis and inhibit the proliferation and migration of HepG2

  10. A fibroblast/macrophage co-culture model to evaluate the biocompatibility of an electrospun Dextran/PLGA scaffold and its potential to induce inflammatory responses

    International Nuclear Information System (INIS)

    Pan Hui; Kantharia, Sarah; Jiang Hongliang; Chen Weiliam

    2011-01-01

    Fibroblasts and macrophages are the two major types of cells responding to implanted biomaterials. They play crucial roles in inflammatory responses, host-material interactions and tissue remodeling. However, the synergistic interactions of these two cell types with biomaterials are not fully understood. In this investigation, an in vitro fibroblast/macrophage co-culture system was utilized to examine the biocompatibility and the potential to induce inflammatory responses of an electrospun Dextran/PLGA scaffold. The scaffold did not affect the morphologies, attachments, proliferations and viabilities of both the fibroblasts and macrophages, cultured separately or together. Moreover, it only activated a small subset of the macrophages implicating a low potential to induce either severe acute or chronic inflammatory response. Additionally, fibroblasts played a role in prolonging macrophage activation in the presence of the scaffolds. Using antibody arrays, IL-10, SDF-1, MIP-1 gamma and RANTES were found to be up-regulated when the cells were incubated with the scaffolds. The results of subdermal implantation of the Dextran/PLGA scaffolds confirmed its biocompatibility and low inflammatory potential.

  11. A fibroblast/macrophage co-culture model to evaluate the biocompatibility of an electrospun Dextran/PLGA scaffold and its potential to induce inflammatory responses

    Energy Technology Data Exchange (ETDEWEB)

    Pan Hui; Kantharia, Sarah [Department of Biomedical Engineering, State University of New York-Stony Brook, Stony Brook, NY 11794-2580 (United States); Jiang Hongliang [Department of Polymer Science and Engineering, Zhejiang University, Hangzhou 310027 (China); Chen Weiliam, E-mail: weiliam.chen@nyumc.org [Division of Wound Healing and Regenerative Medicine, Department of Surgery, New York University School of Medicine, New York, NY 10016 (United States)

    2011-12-15

    Fibroblasts and macrophages are the two major types of cells responding to implanted biomaterials. They play crucial roles in inflammatory responses, host-material interactions and tissue remodeling. However, the synergistic interactions of these two cell types with biomaterials are not fully understood. In this investigation, an in vitro fibroblast/macrophage co-culture system was utilized to examine the biocompatibility and the potential to induce inflammatory responses of an electrospun Dextran/PLGA scaffold. The scaffold did not affect the morphologies, attachments, proliferations and viabilities of both the fibroblasts and macrophages, cultured separately or together. Moreover, it only activated a small subset of the macrophages implicating a low potential to induce either severe acute or chronic inflammatory response. Additionally, fibroblasts played a role in prolonging macrophage activation in the presence of the scaffolds. Using antibody arrays, IL-10, SDF-1, MIP-1 gamma and RANTES were found to be up-regulated when the cells were incubated with the scaffolds. The results of subdermal implantation of the Dextran/PLGA scaffolds confirmed its biocompatibility and low inflammatory potential.

  12. Different particle determinants induce apoptosis and cytokine release in primary alveolar macrophage cultures

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    Schwarze Per E

    2006-06-01

    Full Text Available Abstract Background Particles are known to induce both cytokine release (MIP-2, TNF-α, a reduction in cell viability and an increased apoptosis in alveolar macrophages. To examine whether these responses are triggered by the same particle determinants, alveolar macrophages were exposed in vitro to mineral particles of different physical-chemical properties. Results The crystalline particles of the different stone types mylonite, gabbro, basalt, feldspar, quartz, hornfels and fine grain syenite porphyr (porphyr, with a relatively equal size distribution (≤ 10 μm, but different chemical/mineral composition, all induced low and relatively similar levels of apoptosis. In contrast, mylonite and gabbro induced a marked MIP-2 response compared to the other particles. For particles of smaller size, quartz (≤ 2 μm seemed to induce a somewhat stronger apoptotic response than even smaller quartz (≤ 0.5 μm and larger quartz (≤ 10 μm in relation to surface area, and was more potent than hornfels and porphyr (≤ 2 μm. The reduction in cell viability induced by quartz of the different sizes was roughly similar when adjusted to surface area. With respect to cytokines, the release was more marked after exposure to quartz ≤ 0.5 μm than to quartz ≤ 2 μm and ≤ 10 μm. Furthermore, hornfels (≤ 2 μm was more potent than the corresponding hornfels (≤ 10 μm and quartz (≤ 2 μm to induce cytokine responses. Pre-treatment of hornfels and quartz particles ≤ 2 μm with aluminium lactate, to diminish the surface reactivity, did significantly reduce the MIP-2 response to hornfels. In contrast, the apoptotic responses to the particles were not affected. Conclusion These results indicate that different determinants of mineral/stone particles are critical for inducing cytokine responses, reduction in cell viability and apoptosis in alveolar macrophages. The data suggest that the particle surface reactivity was critical for cytokine responses

  13. Effect of nanoparticles binding ß-amyloid peptide on nitric oxide production by cultured endothelial cells and macrophages

    Directory of Open Access Journals (Sweden)

    Orlando A

    2013-04-01

    Full Text Available Antonina Orlando,1 Francesca Re,1 Silvia Sesana,1 Ilaria Rivolta,1 Alice Panariti,1 Davide Brambilla,2 Julien Nicolas,2 Patrick Couvreur,2 Karine Andrieux,2 Massimo Masserini,1 Emanuela Cazzaniga1 1Department of Health Sciences, University of Milano-Bicocca, Monza, Italy; 2Institut Galien Paris Sud, University Paris-Sud, Châtenay-Malabry, France Background: As part of a project designing nanoparticles for the treatment of Alzheimer’s disease, we have synthesized and characterized a small library of nanoparticles binding with high affinity to the β-amyloid peptide and showing features of biocompatibility in vitro, which are important properties for administration in vivo. In this study, we focused on biocompatibility issues, evaluating production of nitric oxide by cultured human umbilical vein endothelial cells and macrophages, used as models of cells which would be exposed to nanoparticles after systemic administration. Methods: The nanoparticles tested were liposomes and solid lipid nanoparticles carrying phosphatidic acid or cardiolipin, and PEGylated poly(alkyl cyanoacrylate nanoparticles (PEG-PACA. We measured nitric oxide production using the Griess method as well as phosphorylation of endothelial nitric oxide synthase and intracellular free calcium, which are biochemically related to nitric oxide production. MTT viability tests and caspase-3 detection were also undertaken. Results: Exposure to liposomes did not affect the viability of endothelial cells at any concentration tested. Increased production of nitric oxide was detected only with liposomes carrying phosphatidic acid or cardiolipin at the highest concentration (120 µg/mL, together with increased synthase phosphorylation and intracellular calcium levels. Macrophages exposed to liposomes showed a slightly dose-dependent decrease in viability, with no increase in production of nitric oxide. Exposure to solid lipid nanoparticles carrying phosphatidic acid decreased viability in

  14. Tumor-cytolytic human macrophages cultured as nonadherent cells: potential for the adoptive immunotherapy of cancer.

    Science.gov (United States)

    Helinski, E H; Hurley, E L; Streck, R J; Bielat, K L; Pauly, J L

    1990-01-01

    Tumor-cytolytic lymphokine (e.g., interleukin-2; IL-2)-activated killer cells are currently being evaluated in IL-2/LAK cell adoptive immunotherapy regimens for the treatment of cancer. Monocyte-derived macrophages (M phi) are also known to be efficient tumor killer cells; accordingly, M phi that have been activated in vitro may also be of therapeutic merit. However, attempts to cultivate M phi for morphological and functional studies have often been compromised because M phi adhere rapidly and tenaciously to cultureware. Studies that we have conducted to address this problem have proven successful in developing procedures for the long-term cultivation of non-adherent immunocompetent M phi in serum-free medium using petri dishes containing a thin Teflon liner. The utility of this technology is documented by the results of studies presented herein in which light and scanning electron microscopy was used to analyze tumor-cytolytic human M phi. In these experiments, we demonstrated that nonadherent immunocompetent human M phi can be prepared for detailed examinations of their pleomorphic membrane architecture. Moreover, nonadherent human M phi could readily be collected for preparing conjugates of M phi and tumor cells. It is anticipated that this technology should prove useful for future structure-function studies defining the topographical location and spatial distribution of antigens and receptors on M phi membrane ultrastructures, particularly the microvilli-like projections that bridge together an immunocompetent effector M phi and target cell (e.g., tumor cells and microbial pathogens) and which provide the physical interaction required for the initial phases of a cellular immune response that includes antigen recognition and cell-to-cell adhesion.

  15. Macrophage involvement affects matrix stiffness-related influences on cell osteogenesis under three-dimensional culture conditions.

    Science.gov (United States)

    He, Xiao-Tao; Wu, Rui-Xin; Xu, Xin-Yue; Wang, Jia; Yin, Yuan; Chen, Fa-Ming

    2018-04-15

    Accumulating evidence indicates that the physicochemical properties of biomaterials exert profound influences on stem cell fate decisions. However, matrix-based regulation selected through in vitro analyses based on a given cell population do not genuinely reflect the in vivo conditions, in which multiple cell types are involved and interact dynamically. This study constitutes the first investigation of how macrophages (Mφs) in stiffness-tunable transglutaminase cross-linked gelatin (TG-gel) affect the osteogenesis of bone marrow-derived mesenchymal stem cells (BMMSCs). When a single cell type was cultured, low-stiffness TG-gels promoted BMMSC proliferation, whereas high-stiffness TG-gels supported cell osteogenic differentiation. However, Mφs in high-stiffness TG-gels were more likely to polarize toward the pro-inflammatory M1 phenotype. Using either conditioned medium (CM)-based incubation or Transwell-based co-culture, we found that Mφs encapsulated in the low-stiffness matrix exerted a positive effect on the osteogenesis of co-cultured BMMSCs. Conversely, Mφs in high-stiffness TG-gels negatively affected cell osteogenic differentiation. When both cell types were cultured in the same TG-gel type and placed into the Transwell system, the stiffness-related influences of Mφs on BMMSCs were significantly altered; both the low- and high-stiffness matrix induced similar levels of BMMSC osteogenesis. Although the best material parameter for synergistically affecting Mφs and BMMSCs remains unknown, our data suggest that Mφ involvement in the co-culture system alters previously identified material-related influences on BMMSCs, such as matrix stiffness-related effects, which were identified based on a culture system involving a single cell type. Such Mφ-stem cell interactions should be considered when establishing proper matrix parameter-associated cell regulation in the development of biomimetic biomaterials for regenerative applications. The substrate stiffness

  16. Radiation effects on cultured human monocytes and on monocyte-derived macrophages

    International Nuclear Information System (INIS)

    Buescher, E.S.; Gallin, J.I.

    1984-01-01

    Prior to administration, leukocyte transfusions are commonly irradiated with up to 5,000 R to eliminate lymphocytes and thereby prevent graft-versus-host disease in the recipient. It has been widely believed that phagocytes are resistant to this irradiation. In a recent report, it was noted that phagocyte oxidative metabolism was compromised during preparation of white cells for transfusion. As part of the effort to examine the basis for this inhibition of phagocyte function during white cell preparation, an assessment was made of the effects of irradiation on the long-lived monocytes that have been shown to persist at inflammatory foci posttransfusion. Human monocytes were irradiated for up to 3 min, receiving 2,500-5,000 R. This irradiation damaged human monocytes, significantly decreasing their in vitro survival for the first 3 wk of culture, and growth as assessed by two-dimensional cell size measurements during the first 2 wk of culture. Despite smaller cell size, total cell protein was significantly increased over time in irradiated cultures. Extracellular release of lysozyme and beta-glucuronidase per cell was not affected by irradiation, but extracellular lactate dehydrogenase (LDH) release was significantly increased after irradiation. Irradiated monocytes killed Listeria monocytogenes at a slower rate than the nonirradiated controls. Thus, the data indicate that irradiation in doses used to prevent graft-versus-host disease in leukocyte transfusion recipients has a deleterious effect on in vitro human monocyte survival and function

  17. Granulocyte Macrophage Colony Stimulating Factor Supplementation in Culture Media for Subfertile Women Undergoing Assisted Reproduction Technologies: A Systematic Review

    Science.gov (United States)

    Siristatidis, Charalampos; Vogiatzi, Paraskevi; Salamalekis, George; Creatsa, Maria; Vrachnis, Nikos; Glujovsky, Demián; Iliodromiti, Zoe; Chrelias, Charalampos

    2013-01-01

    Granulocyte macrophage colony stimulating factor (GM-CSF) is a cytokine/growth factor produced by epithelial cells that exerts embryotrophic effects during the early stages of embryo development. We performed a systematic review, and six studies that were performed in humans undergoing assisted reproduction technologies (ART) were located. We wanted to evaluate if embryo culture media supplementation with GM-CSF could improve success rates. As the type of studies and the outcome parameters investigated were heterogeneous, we decided not to perform a meta-analysis. Most of them had a trend favoring the supplementation with GM-CSF, when outcomes were measured in terms of increased percentage of good-quality embryos reaching the blastocyst stage, improved hatching initiation and number of cells in the blastocyst, and reduction of cell death. However, no statistically significant differences were found in implantation and pregnancy rates in all apart from one large multicenter trial, which reported favorable outcomes, in terms of implantation and live birth rates. We propose properly conducted and adequately powered randomized controlled trials (RCTs) to further validate and extrapolate the current findings with the live birth rate to be the primary outcome measure. PMID:23509457

  18. Functional and Structural Analysis of a Highly-Expressed Yersinia pestis Small RNA following Infection of Cultured Macrophages.

    Directory of Open Access Journals (Sweden)

    Nan Li

    Full Text Available Non-coding small RNAs (sRNAs are found in practically all bacterial genomes and play important roles in regulating gene expression to impact bacterial metabolism, growth, and virulence. We performed transcriptomics analysis to identify sRNAs that are differentially expressed in Yersinia pestis that invaded the human macrophage cell line THP-1, compared to pathogens that remained extracellular in the presence of host. Using ultra high-throughput sequencing, we identified 37 novel and 143 previously known sRNAs in Y. pestis. In particular, the sRNA Ysr170 was highly expressed in intracellular Yersinia and exhibited a log2 fold change ~3.6 higher levels compared to extracellular bacteria. We found that knock-down of Ysr170 expression attenuated infection efficiency in cell culture and growth rate in response to different stressors. In addition, we applied selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE analysis to determine the secondary structure of Ysr170 and observed structural changes resulting from interactions with the aminoglycoside antibiotic gentamycin and the RNA chaperone Hfq. Interestingly, gentamicin stabilized helix 4 of Ysr170, which structurally resembles the native gentamicin 16S ribosomal binding site. Finally, we modeled the tertiary structure of Ysr170 binding to gentamycin using RNA motif modeling. Integration of these experimental and structural methods can provide further insight into the design of small molecules that can inhibit function of sRNAs required for pathogen virulence.

  19. Thermo-radiosensitivity of the granulocyte and macrophage precursor cells of mice. I.-Development of the in vivo culture and effects induced by the hyperthermia

    International Nuclear Information System (INIS)

    Bueren, J. A.; Nieto, M.

    1983-01-01

    The present report shows the agar diffusion chamber technique for culturing granulocyte- macrophage precursor cells, obtained from mice bone marrow. Diffusion chambers containing the bone marrow suspension are implanted intraperitoneally Into mice and constitute a compartment which avoids the migration of cells, but allows the transit of the mouse biological fluxes, necessary for the cellular proliferation. By means of this technique, we studied the lethal effects of the hyperthermia on the precursors and their capacity to repair sublethal damage. (Author) 129 refs

  20. X-ray-induced production of granulocyte-macrophage colony-stimulating factor (GM-CSF) by mouse spleen cells in culture

    International Nuclear Information System (INIS)

    Onoda, M.; Shinoda, M.; Tsuneoka, K.; Shikita, M.

    1980-01-01

    Spleen cells were collected from normal mice and cultured in a medium containing 20% calf serum. Addition of lipopolysaccharide (LPS) in the culture significantly increased the production of granulocyte-macrophage colony-stimulating factor (GM-CSF), and a maximum induction was attained in 5 days. Irradiation of the spleen cells with 300 to 3000 R x rays also enhanced the production of GM-CSF, but there was a latent period of about 5 days before the factor appeared in the culture medium. The observed difference between LPS and x rays in the timing of inducing GM-CSF production in the spleen cell culture was consistent with the difference observed in animals. These results suggest that different mechanisms of GM-CSF production operate in the spleen in response to either LPS or x rays

  1. “Stealth dissemination” of macrophage-tumor cell fusions cultured from blood of patients with pancreatic ductal adenocarcinoma

    Science.gov (United States)

    Circulating tumor cells (CTCs) appear to be involved in early dissemination of many cancers, although which characteristics are important in metastatic spread are not clear. Here we describe isolation and characterization of macrophage-tumor cell fusions (MTFs) from the blood of pancreatic ductal a...

  2. A transgenic rat hepatocyte - Kupffer cell co-culture model for evaluation of direct and macrophage-related effect of poly(amidoamine) dendrimers.

    Science.gov (United States)

    Jemnitz, Katalin; Bátai-Konczos, Attila; Szabó, Mónika; Ioja, Enikő; Kolacsek, Orsolya; Orbán, Tamás I; Török, György; Homolya, László; Kovács, Eszter; Jablonkai, István; Veres, Zsuzsa

    2017-02-01

    Increasing number of papers demonstrate that Kupffer cells (KCs) play a role in the development of drug induced liver injury (DILI). Furthermore, elevated intracellular Ca 2+ level of hepatocytes is considered as a common marker of DILI. Here we applied an in vitro model based on hepatocyte mono- and hepatocyte/KC co-cultures (H/KC) isolated from transgenic rats stably expressing the GCaMP2 fluorescent Ca 2+ sensor protein to investigate the effects of polycationic (G5), polyanionic (G4.5) and polyethylene-glycol coated neutral (G5 Peg) dendrimers known to accumulate in the liver, primarily in KCs. Following dendrimer exposure, hepatocyte homeostasis was measured by MTT cytotoxicity assay and by Ca 2+ imaging, while hepatocyte functions were studied by CYP2B1/2 inducibility, and bilirubin and taurocholate transport. G5 was significantly more cytotoxic than G4.5 for hepatocytes and induced Ca 2+ oscillation and sustained Ca 2+ signals at 1μM and10 μM, respectively both in hepatocytes and KCs. Dendrimer-induced Ca 2+ signals in hepatocytes were attenuated by macrophages. Activation of KCs by lipopolysaccharide and G5 decreased the inducibility of CYP2B1/2, which was restored by depleting the KCs with gadolinium-chloride and pentoxyphylline, suggesting a role of macrophages in the hindrance of CYP2B1/2 induction by G5 and lipopolysaccharide. In the H/KC, but not in the hepatocyte mono-culture, G5 reduced the canalicular efflux of bilirubin and stimulated the uptake and canalicular efflux of taurocholate. In conclusion, H/KC provides a good model for the prediction of hepatotoxic potential of drugs, especially of nanomaterials known to be trapped by macrophages, activation of which presumably contributes to DILI. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Control of microorganisms of oral health interest with Arctium lappa L. (burdock) extract non-cytotoxic to cell culture of macrophages (RAW 264.7).

    Science.gov (United States)

    de Oliveira, Jonatas Rafael; de Aguiar Almeida, Rosilene Batista; das Graças Figueiredo Vilela, Polyana; de Oliveira, Felipe Eduardo; da Rocha, Rosilene Fernandes; Jorge, Antonio Olavo Cardoso; de Oliveira, Luciane Dias

    2014-08-01

    To evaluate the antimicrobial activity of Arctium lappa L. extract on Staphylococcus aureus, S. epidermidis, Streptococcus mutans, Candida albicans, C. tropicalis and C. glabrata. In addition, the cytotoxicity of this extract was analyzed on macrophages (RAW 264.7). By broth microdilution method, different concentrations of the extract (250-0.4 mg/mL) were used in order to determine the minimum microbicidal concentration (MMC) in planktonic cultures and the most effective concentration was used on biofilms on discs made of acrylic resin. The cytotoxicity A. lappa L. extract MMC was evaluated on RAW 264.7 by MTT assay and the quantification of IL-1β and TNF-α by ELISA. The most effective concentration was 250 mg/mL and also promoted significant reduction (log₁₀) in the biofilms of S. aureus (0.438 ± 0.269), S. epidermidis (0.377 ± 0.298), S. mutans (0.244 ± 0.161) and C. albicans (0.746 ± 0.209). Cell viability was similar to 100%. The production of IL-1β was similar to the control group (p>0.05) and there was inhibition of TNF-α (plappa L. extract was microbicidal for all the evaluated strains in planktonic cultures, microbiostatic for biofilms and not cytotoxic to the macrophages. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. Heme oxygenase-1 induction alters chemokine regulation and ameliorates human immunodeficiency virus-type-1 infection in lipopolysaccharide-stimulated macrophages

    International Nuclear Information System (INIS)

    Zhou, Zhao-Hua; Kumari, Namita; Nekhai, Sergei; Clouse, Kathleen A.; Wahl, Larry M.; Yamada, Kenneth M.; Dhawan, Subhash

    2013-01-01

    Highlights: •Lipopolysaccharide stimulation of heme oxygenase-1 (HO-1) ameliorated HIV-1 infection of primary human macrophages. •The partial protection by HO-1 against HIV infection was associated with induction of chemokines such as MIP1α and MIP1β. •This mechanism explains lipopolysaccharide-stimulated HO-1-mediated inhibition of HIV-1 infection of macrophages. -- Abstract: We have elucidated a putative mechanism for the host resistance against HIV-1 infection of primary human monocyte-derived macrophages (MDM) stimulated with lipopolysaccharide (LPS). We show that LPS-activated MDM both inhibited HIV-1 entry into the cells and were refractory to post-entry productive viral replication. LPS-treated cells were virtually negative for mature virions as revealed by transmission electron microscopy. LPS activation of MDM markedly enhanced the expression of heme oxygenase-1 (HO-1), a potent inducible cytoprotective enzyme. Increased HO-1 expression was accompanied by elevated production of macrophage inflammatory chemokines (MIP1α and MIP1β) by LPS-activated MDM, significantly decreased surface chemokine receptor-5 (CCR-5) expression, and substantially reduced virus replication. Treatment of cells with HO-1 inhibitor SnPP IX (tin protoporphyrin IX) attenuated the LPS-mediated responses, HIV-1 replication and secretion of MIP1α, MIP1β, and LD78β chemokines with little change in surface CCR-5 expression. These results identify a novel role for HO-1 in the modulation of host immune response against HIV infection of MDM

  5. Anti-Inflammatory Activity of the Methanol Extract of Moutan Cortex in LPS-Activated Raw264.7 Cells

    Directory of Open Access Journals (Sweden)

    Seung-Chul Chun

    2007-01-01

    Full Text Available Moutan Cortex (MCE has been used in traditional medicine to remove heat from the blood, promote blood circulation and alleviate blood stasis. This study was conducted to evaluate the effects of MCE on regulatory mechanisms of cytokines and nitric oxide (NO involved in immunological activity of Raw264.7 cells. Cells were pretreated with methanolic extracts of MCE, and further cultured for an appropriate time after lipopolyssacharide (LPS addition. During the entire experimental period, 0.1 and 0.3 mg ml−1 of MCE had no cytotoxicity. In these concentrations, MCE inhibited the production of NO and prostaglandin E2 (PGE2, the expression of inducible NO synthase (iNOS, cyclooxygenase-2 (COX-2 and phosphorylated inhibitor of κBα (p-IκBα, and the activation of nuclear factor κB (NF-κB. MCE also reduced the concentration of tumor necrosis factor-α (TNF-α, interleukin-1β (IL-1β and interleukin-6 (IL-6 in the Raw264.7 cells that were activated by LPS. These results demonstrate that MCE has anti-inflammatory effects through the inhibition of iNOS and COX-2 expression by suppressing the phosphorylation of I-κBα and the activation of NF-κB.

  6. Inhibition of TNF-α production in LPS-activated THP-1 monocytic cells by the crude extracts of seven Bhutanese medicinal plants.

    Science.gov (United States)

    Wangchuk, Phurpa; Keller, Paul A; Pyne, Stephen G; Taweechotipatr, Malai

    2013-07-30

    Seven studied medicinal plants; Aconitum laciniatum, Ajania nubigena, Codonopsis bhutanica, Corydalis crispa, Corydalis dubia, Meconopsis simplicifolia and Pleurospermum amabile, are currently used in the Bhutanese Traditional Medicine (BTM) for the management of different types of disorders including the diseases that bore relevance to various inflammatory conditions. This study aimed to evaluate the inhibition of TNF-α production in LPS-activated THP-1 monocytic cells by the crude extracts of seven selected Bhutanese medicinal plants. It is expected to; (a) generate a scientific basis for their use in the BTM and (b) form a basis for prioritization of the seven plants for further phytochemical and anti-inflammatory studies. Seven plants were selected using an ethno-directed bio-rational approach and their crude extracts were prepared using four different solvents (methanol, hexane, dichloromethane and chloroform). The TNF-α inhibitory activity of these extracts was determined by cytokine-specific sandwich quantitative enzyme-linked immunosorbent assays (ELISAs). The results were quantified statistically and the statistical significance were evaluated by GraphPad Prism version 5.01 using Student's t-test with one-tailed distribution. A p-value ≤0.05 was considered statistically significant. Of the seven plants studied, the crude extracts of six of them inhibited the production of pro-inflammatory cytokine, TNF-α in LPS-activated THP-1 monocytic cells. Amongst the six plants, Corydalis crispa gave the best inhibitory activity followed by Pleurospermum amabile, Ajania nubigena, Corydalis dubia, Meconopsis simplicifolia and Codonopsis bhutanica. Of the 13 extracts that exhibited statistically significant TNF-α inhibitory activity (p<0.05; p<0.01), five of them showed very strong inhibition when compared to the DMSO control and RPMI media. Six medicinal plants studied here showed promising TNF-α inhibitory activity. These findings rationalize the traditional

  7. Short-term exposure of umbilical cord blood CD34+ cells to granulocyte-macrophage colony-stimulating factor early in culture improves ex vivo expansion of neutrophils.

    Science.gov (United States)

    Marturana, Flavia; Timmins, Nicholas E; Nielsen, Lars K

    2011-03-01

    Despite the availability of modern antibiotics/antimycotics and cytokine support, neutropenic infection accounts for the majority of chemotherapy-associated deaths. While transfusion support with donor neutrophils is possible, cost and complicated logistics make such an option unrealistic on a routine basis. A manufactured neutrophil product could enable routine prophylactic administration of neutrophils, preventing the onset of neutropenia and substantially reducing the risk of infection. We examined the use of pre-culture strategies and various cytokine/modulator combinations to improve neutrophil expansion from umbilical cord blood (UCB) hematopoietic stem and progenitor cells (HPC). Enriched UCB HPC were cultured using either two-phase pre-culture strategies or a single phase using various cytokine/modulator combinations. Outcome was assessed with respect to numerical expansion, cell morphology, granulation and respiratory burst activity. Pre-culture in the absence of strong differentiation signals (e.g. granulocyte colony-stimulating factor; G-CSF) failed to provide any improvement to final neutrophil yields. Similarly, removal of differentiating cells during pre-culture failed to improve neutrophil yields to an appreciable extent. Of the cytokine/modulator combinations, the addition of granulocyte-macrophage (GM)-colony-stimulating factor (CSF) alone gave the greatest increase. In order to avoid production of monocytes, it was necessary to remove GM-CSF on day 5. Using this strategy, neutrophil expansion improved 2.7-fold. Although all cytokines and culture strategies employed have been reported previously to enhance HPC expansion, we found that the addition of GM-CSF alone was sufficient to improve total cell yields maximally. The need to remove GM-CSF on day 5 to avoid monocyte differentiation highlights the context and time-dependent complexity of exogenous signaling in hematopoietic cell differentiation and growth.

  8. Carbon monoxide-releasing molecule-3 suppresses Prevotella intermedia lipopolysaccharide-induced production of nitric oxide and interleukin-1β in murine macrophages.

    Science.gov (United States)

    Choi, Eun-Young; Choe, So-Hui; Hyeon, Jin-Yi; Choi, Jeom-Il; Choi, In Soon; Kim, Sung-Jo

    2015-10-05

    This study was performed to analyze the effect of carbon monoxide (CO)-releasing molecule-3 (CORM-3) in alleviating the production of proinflammatory mediators in macrophages treated with lipopolysaccharide (LPS) from Prevotella intermedia, a pathogen associated with periodontal disease, and its possible mechanisms of action. LPS was isolated using the hot phenol-water method. Culture supernatants were assayed for nitric oxide (NO) and interleukin-1β (IL-1β). Gene expression was quantified by real-time PCR, and protein expression by immunoblotting. DNA-binding activities of NF-κB subunits were determined using an ELISA-based kit. CORM-3 suppressed the production of inducible NO synthase (iNOS)-derived NO and IL-1β at both gene transcription and translation levels in P. intermedia LPS-activated RAW264.7 cells. CORM-3 enhanced heme oxygenase-1 (HO-1) expression in cells stimulated with P. intermedia LPS, and inhibition of HO-1 activity by SnPP notably reversed the suppressive effect of CORM-3 on LPS-induced production of NO. LPS-induced phosphorylation of p38 and JNK was not affected by CORM-3. CORM-3 did not influence P. intermedia LPS-induced degradation of IκB-α. Instead, nuclear translocation of NF-κB p65 and p50 subunits was blocked by CORM-3 in LPS-treated cells. In addition, CORM-3 reduced LPS-induced p65 and p50 binding to DNA. Besides, CORM-3 significantly suppressed P. intermedia LPS-induced phosphorylation of STAT1. Overall, this study indicates that CORM-3 suppresses the production of NO and IL-1β in P. intermedia LPS-activated murine macrophages via HO-1 induction and inhibition of NF-κB and STAT1 pathways. The modulation of host inflammatory response by CORM-3 would be an attractive therapeutic approach to attenuate the progression of periodontal disease. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Administration of Protein kinase D1 induce an immunomodulatory effect on lipopolysaccharide-induced intestinal inflammation in a co-culture model of intestinal epithelial Caco-2 cells and RAW 264.7 macrophage cells

    DEFF Research Database (Denmark)

    Nielsen, Ditte Søvsø Gundelund; Fredborg, Marlene; Andersen, Vibeke

    2017-01-01

    the effects of human PKD1 in relation to intestinal inflammation, using a co-culture model of intestinal epithelial Caco-2 cells and RAW264.7 macrophages. An inflammatory response was induced in the macrophages by lipopolysaccharide (LPS), upregulating the expression of tumour necrosis factor alpha (TNF......-α), interleukin- (IL-) 1β, and IL-6 besides increasing the secretion of TNF-α protein. The effect of administering PKD1 to Caco-2 was evaluated in relation to both amelioration of inflammation and the ability to suppress inflammation initiation. Administration of PKD1 (10–100 ng/ml) following induction...

  10. Bulky PAH-DNA induced by exposure of a co-culture model of human alveolar macrophages and embryonic epithelial cells to atmospheric particulate pollution

    International Nuclear Information System (INIS)

    Abbas, Imane; Garcon, Guillaume; Billet, Sylvain; Shirali, Pirouz; Andre, Veronique; Le Goff, Jeremie; Sichel, Francois; Roy Saint-Georges, Francoise; Mulliez, Philippe

    2012-01-01

    Because of their deep penetration in human lungs, fine airborne particulate matter were described as mainly responsible for the deleterious effects of exposure to air pollution on health. Organic constituents are adsorbed on particles surface and, after inhalation, some (polycyclic aromatic hydrocarbons, PAHs) can be activated into reactive metabolites and can bind to DNA. The formation of bulky DNA adducts has been researched after exposure of mono-and co-cultures of alveolar macrophages (AM) and human embryonic human lung epithelial (L132), to fine air pollution particulate matter Air samples have been collected with cascade impactor and characterized: size distribution (92.15% 2 /g), inorganic (Fe, AI, Ca, Na, K, Mg, Pb, etc.) and organic compounds (PAHs, etc.). 32 P post-labeling method was applied to detect bulky DNA adducts in AM and L132, in mono-and co-cultures, 72 h after their exposure to atmospheric particles at their Lethals and Effects concentrations or (LC or CE) to 50% (i.e. MA: EC 50 = 74.63 μg/mL and L132: LC-5-0 = 75.36 μg/mL). Exposure to desorbed particles (MA: C1= 61.11 μg/mL and L132 : C2 = 61.71 μg/mL) and B[a]P (1 μM) were included. Bulky PAH-DNA adducts were detected in AM in mono-culture after exposure to total particles (Pt), to B[a]P and desorbed particles (Pd). Whatever the exposure, no DNA adduct was detected in L132 in mono-culture. These results are coherent with the enzymatic activities of cytochrome P450 l Al in AM and L132. Exposure of co-culture to Pt, or Pd induced bulky adducts to DNA in AM but not in L132. Exposure to B[a]P alone has altered the DNA of AM and L132, in co-culture. Exposure to Pt is closer to the environmental conditions, but conferred an exposure to amounts of genotoxic agents compared to studies using organic extracts. The formation of bulky DNA adducts was nevertheless observed in AM exposed to Pt, in mono- or co-culture, indicating that they were competent in terms of metabolic activation of PAHs. The

  11. Bone marrow-derived mesenchymal stem cells propagate immunosuppressive/anti-inflammatory macrophages in cell-to-cell contact-independent and -dependent manners under hypoxic culture.

    Science.gov (United States)

    Takizawa, Naoki; Okubo, Naoto; Kamo, Masaharu; Chosa, Naoyuki; Mikami, Toshinari; Suzuki, Keita; Yokota, Seiji; Ibi, Miho; Ohtsuka, Masato; Taira, Masayuki; Yaegashi, Takashi; Ishisaki, Akira; Kyakumoto, Seiko

    2017-09-15

    Immunosuppressive/anti-inflammatory macrophage (Mφ), M2-Mφ that expressed the typical M2-Mφs marker, CD206, and anti-inflammatory cytokine, interleukin (IL)-10, is beneficial and expected tool for the cytotherapy against inflammatory diseases. Here, we demonstrated that bone marrow-derived lineage-positive (Lin+) blood cells proliferated and differentiated into M2-Mφs by cooperation with the bone marrow-derived mesenchymal stem cells (MSCs) under hypoxic condition: MSCs not only promoted proliferation of undifferentiated M2-Mφs, pre-M2-Mφs, in the Lin+ fraction via a proliferative effect of the MSCs-secreted macrophage colony-stimulating factor, but also promoted M2-Mφ polarization of the pre-M2-Mφs through cell-to-cell contact with the pre-M2-Mφs. Intriguingly, an inhibitor for intercellular adhesion molecule (ICAM)-1 receptor/lymphocyte function-associated antigen (LFA)-1, Rwj50271, partially suppressed expression of CD206 in the Lin+ blood cells but an inhibitor for VCAM-1 receptor/VLA-4, BIO5192, did not, suggesting that the cell-to-cell adhesion through LFA-1 on pre-M2-Mφs and ICAM-1 on MSCs was supposed to promoted the M2-Mφ polarization. Thus, the co-culture system consisting of bone marrow-derived Lin+ blood cells and MSCs under hypoxic condition was a beneficial supplier of a number of M2-Mφs, which could be clinically applicable to inflammatory diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Culture media-based selection of endothelial cells, pericytes, and perivascular-resident macrophage-like melanocytes from the young mouse vestibular system.

    Science.gov (United States)

    Zhang, Jinhui; Chen, Songlin; Cai, Jing; Hou, Zhiqiang; Wang, Xiaohan; Kachelmeier, Allan; Shi, Xiaorui

    2017-03-01

    The vestibular blood-labyrinth barrier (BLB) is comprised of perivascular-resident macrophage-like melanocytes (PVM/Ms) and pericytes (PCs), in addition to endothelial cells (ECs) and basement membrane (BM), and bears strong resemblance to the cochlear BLB in the stria vascularis. Over the past few decades, in vitro cell-based models have been widely used in blood-brain barrier (BBB) and blood-retina barrier (BRB) research, and have proved to be powerful tools for studying cell-cell interactions in their respective organs. Study of both the vestibular and strial BLB has been limited by the unavailability of primary culture cells from these barriers. To better understand how barrier component cells interact in the vestibular system to control BLB function, we developed a novel culture medium-based method for obtaining EC, PC, and PVM/M primary cells from tiny explants of the semicircular canal, sacculus, utriculus, and ampullae tissue of young mouse ears at post-natal age 8-12 d. Each phenotype is grown in a specific culture medium which selectively supports the phenotype in a mixed population of vestibular cell types. The unwanted phenotypes do not survive passaging. The protocol does not require additional equipment or special enzyme treatment. The harvesting process takes less than 2 h. Primary cell types are generated within 7-10 d. The primary culture ECs, PCs, and PVM/M shave consistent phenotypes more than 90% pure after two passages (∼ 3 weeks). The highly purified primary cell lines can be used for studying cell-cell interactions, barrier permeability, and angiogenesis. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Differential in Gel Electrophoresis (DIGE Comparative Proteomic Analysis of Macrophages Cell Cultures in Response to Perthamide C Treatment

    Directory of Open Access Journals (Sweden)

    Raffaele Riccio

    2013-04-01

    Full Text Available Secondary metabolites contained in marine organisms disclose diverse pharmacological activities, due to their intrinsic ability to recognize bio-macromolecules, which alter their expression and modulate their function. Thus, the identification of the cellular pathways affected by marine natural products is crucial to provide important functional information concerning their mechanism of action at the molecular level. Perthamide C, a marine sponge metabolite isolated from the polar extracts of Theonella swinhoei and endowed with a broad and interesting anti-inflammatory profile, was found in a previous study to specifically interact with heat shock protein-90 and glucose regulated protein-94, also disclosing the ability to reduce cisplatin-mediated apoptosis. In this paper, we evaluated the effect of this compound on the whole proteome of murine macrophages cells by two-dimensional DIGE proteomics. Thirty-three spots were found to be altered in expression by at least 1.6-fold and 29 proteins were identified by LC ESI-Q/TOF-MS. These proteins are involved in different processes, such as metabolism, structural stability, protein folding assistance and gene expression. Among them, perthamide C modulates the expression of several chaperones implicated in the folding of proteins correlated to apoptosis, such as Hsp90 and T-complexes, and in this context our data shed more light on the cellular effects and pathways altered by this marine cyclo-peptide.

  14. Influence of conversion of penicillin G into a basic derivative on its accumulation and subcellular localization in cultured macrophages

    International Nuclear Information System (INIS)

    Renard, C.; Vanderhaeghe, H.J.; Claes, P.J.; Zenebergh, A.; Tulkens, P.M.

    1987-01-01

    beta-Lactam antibiotics do not accumulate in phagocytes, probably because of their acidic character. We therefore synthesized a basic derivative of penicillin G, namely, 14 C-labeled N-(3-dimethylamino-propyl)benzylpenicillinamide (ABP), and studied its uptake and subcellular localization in J774 macrophages compared with that of 14 C-labeled penicillin G. Whereas the intracellular concentration (Ci) of penicillin G remained lower than its extracellular concentration (Ce), ABP reached a Ci/Ce ratio of 4 to 5. Moreover, approximately 50% of intracellular ABP was found associated with lysosomes after isopycnic centrifugation of cell homogenates in isoosmotic Percoll or hyperosmotic sucrose gradients. The behavior of ABP was thus partly consistent with the model of de Duve et al., in which they described the intralysosomal accumulation of weak organic bases in lysosomes. Although ABP is microbiologically inactive, our results show that beta-lactam antibiotics can be driven into cells by appropriate modification. Further efforts therefore may be warranted in the design of active compounds or prodrugs that may prove useful in the chemotherapy of intracellular infections

  15. Culture of human oocytes with granulocyte-macrophage colony-stimulating factor has no effect on embryonic chromosomal constitution

    DEFF Research Database (Denmark)

    Agerholm, Inge; Loft, Anne; Hald, Finn

    2010-01-01

    -vitro culture of human embryos in the presence of 2 ng/ml GM-CSF resulted in 34.8% (8/23) uniformly normal embryos. Culture without 2 ng/ml GM-CSF resulted in 33.3% (9/27) uniformly normal embryos. A trend towards a higher number of TQE in the test group was observed; however, due to lack of TQE in the control...... women donating 86 oocytes. The primary endpoint was to investigate the chromosomal constitution of human embryos (fluorescence in-situ hybridization analysis for chromosomes 13, 16, 18, 21, 22, X and Y) cultured with or without GM-CSF. The secondary endpoints were number of top-quality embryos (TQE......) and number of normally developed embryos evaluated morphologically on day 3. The cytogenetic analyses demonstrated non-inferiority and therefore the chromosomal constitution of human embryos cultured in vitro in the presence of 2 ng/ml GM-CSF was no worse than the control group cultured without GM-CSF. In...

  16. Culture of human oocytes with granulocyte-macrophage colony-stimulating factor has no effect on embryonic chromosomal constitution

    DEFF Research Database (Denmark)

    Agerholm, Inge; Loft, Anne; Hald, Finn

    2010-01-01

    women donating 86 oocytes. The primary endpoint was to investigate the chromosomal constitution of human embryos (fluorescence in-situ hybridization analysis for chromosomes 13, 16, 18, 21, 22, X and Y) cultured with or without GM-CSF. The secondary endpoints were number of top-quality embryos (TQE......) and number of normally developed embryos evaluated morphologically on day 3. The cytogenetic analyses demonstrated non-inferiority and therefore the chromosomal constitution of human embryos cultured in vitro in the presence of 2 ng/ml GM-CSF was no worse than the control group cultured without GM-CSF. In......-vitro culture of human embryos in the presence of 2 ng/ml GM-CSF resulted in 34.8% (8/23) uniformly normal embryos. Culture without 2 ng/ml GM-CSF resulted in 33.3% (9/27) uniformly normal embryos. A trend towards a higher number of TQE in the test group was observed; however, due to lack of TQE in the control...

  17. Riboflavin Reduces Pro-Inflammatory Activation of Adipocyte-Macrophage Co-culture. Potential Application of Vitamin B2 Enrichment for Attenuation of Insulin Resistance and Metabolic Syndrome Development.

    Science.gov (United States)

    Mazur-Bialy, Agnieszka Irena; Pocheć, Ewa

    2016-12-15

    Due to the progressive increase in the incidence of obese and overweight individuals, cardiometabolic syndrome has become a worldwide pandemic in recent years. Given the immunomodulatory properties of riboflavin, the current study was performed to investigate the potency of riboflavin in reducing obesity-related inflammation, which is the main cause of insulin resistance, diabetes mellitus 2 or arteriosclerosis. We determined whether pretreatment with a low dose of riboflavin (10.4-1000 nM) affected the pro-inflammatory activity of adipocyte-macrophage co-culture (3T3 L1-RAW 264.7) following lipopolysaccharide stimulation (LPS; 100 ng/mL) which mimics obesity-related inflammation. The apoptosis of adipocytes and macrophages as well as tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), interleukin 1beta (IL-1β), monocyte chemotactic protein 1 (MCP-1), high-mobility group box 1 (HMGB1), transforming growth factor-beta 1 (TGFβ), interleukin 10 (IL-10), inducible nitric oxide synthase (iNOS), nitric oxide (NO), matrix metalloproteinase 9 (MMP-9), tissue inhibitor of metalloproteinases-1 (TIMP-1) expression and release, macrophage migration and adipokines (adiponectin and leptin) were determined. Our results indicated an efficient reduction in pro-inflammatory factors (TNFα, IL-6, MCP-1, HMGB1) upon culture with riboflavin supplementation (500-1000 nM), accompanied by elevation in anti-inflammatory adiponectin and IL-10. Moreover, macrophage migration was reduced by the attenuation of chemotactic MCP-1 release and degradation of the extracellular matrix by MMP-9. In conclusion, riboflavin effectively inhibits the pro-inflammatory activity of adipocyte and macrophage co-cultures, and therefore we can assume that its supplementation may reduce the likelihood of conditions associated with the mild inflammation linked to obesity.

  18. Riboflavin Reduces Pro-Inflammatory Activation of Adipocyte-Macrophage Co-culture. Potential Application of Vitamin B2 Enrichment for Attenuation of Insulin Resistance and Metabolic Syndrome Development

    Directory of Open Access Journals (Sweden)

    Agnieszka Irena Mazur-Bialy

    2016-12-01

    Full Text Available Due to the progressive increase in the incidence of obese and overweight individuals, cardiometabolic syndrome has become a worldwide pandemic in recent years. Given the immunomodulatory properties of riboflavin, the current study was performed to investigate the potency of riboflavin in reducing obesity-related inflammation, which is the main cause of insulin resistance, diabetes mellitus 2 or arteriosclerosis. We determined whether pretreatment with a low dose of riboflavin (10.4–1000 nM affected the pro-inflammatory activity of adipocyte-macrophage co-culture (3T3 L1-RAW 264.7 following lipopolysaccharide stimulation (LPS; 100 ng/mL which mimics obesity-related inflammation. The apoptosis of adipocytes and macrophages as well as tumor necrosis factor-alpha (TNF-α, interleukin 6 (IL-6, interleukin 1beta (IL-1β, monocyte chemotactic protein 1 (MCP-1, high-mobility group box 1 (HMGB1, transforming growth factor–beta 1 (TGFβ, interleukin 10 (IL-10, inducible nitric oxide synthase (iNOS, nitric oxide (NO, matrix metalloproteinase 9 (MMP-9, tissue inhibitor of metalloproteinases-1 (TIMP-1 expression and release, macrophage migration and adipokines (adiponectin and leptin were determined. Our results indicated an efficient reduction in pro-inflammatory factors (TNFα, IL-6, MCP-1, HMGB1 upon culture with riboflavin supplementation (500–1000 nM, accompanied by elevation in anti-inflammatory adiponectin and IL-10. Moreover, macrophage migration was reduced by the attenuation of chemotactic MCP-1 release and degradation of the extracellular matrix by MMP-9. In conclusion, riboflavin effectively inhibits the pro-inflammatory activity of adipocyte and macrophage co-cultures, and therefore we can assume that its supplementation may reduce the likelihood of conditions associated with the mild inflammation linked to obesity.

  19. DHA suppresses Prevotella intermedia lipopolysaccharide-induced production of proinflammatory mediators in murine macrophages.

    Science.gov (United States)

    Choi, Eun-Young; Jin, Ji-Young; Choi, Jeom-Il; Choi, In Soon; Kim, Sung-Jo

    2014-04-14

    Several reports have indicated that dietary intake of DHA is associated with lower prevalence of periodontitis. In the present study, we investigated the effect of DHA on the production of proinflammatory mediators in murine macrophage-like RAW264.7 cells stimulated with lipopolysaccharide (LPS) isolated from Prevotella intermedia, a pathogen implicated in inflammatory periodontal disease, and its mechanisms of action. LPS was isolated from lyophilised P. intermedia ATCC 25,611 cells using the standard hot-phenol-water protocol. Culture supernatants were collected and assayed for NO, IL-1β and IL-6. Real-time PCR analysis was carried out to detect the expression of inducible NO synthase (iNOS), IL-1β, IL-6 and haeme oxygenase-1 (HO-1) mRNA. Immunoblot analysis was carried out to quantify the expression of iNOS and HO-1 protein and concentrations of signalling proteins. DNA-binding activities of NF-κB subunits were determined using an ELISA-based assay kit. DHA significantly attenuated the production of NO, IL-1β and IL-6 at both gene transcription and translation levels in P. intermedia LPS-activated RAW264.7 cells. DHA induced the expression of HO-1 in cells treated with P. intermedia LPS. Selective inhibition of HO-1 activity by tin protoporphyrin IX significantly mitigated the inhibitory effects of DHA on LPS-induced NO production. DHA significantly attenuated the phosphorylation of c-Jun N-terminal kinase induced by LPS. In addition, DHA suppressed the transcriptional activity of NF-κB by regulating the nuclear translocation and DNA-binding activity of NF-κB p50 subunit and inhibited the phosphorylation of signal transducer and activator of transcription 1. Further in vivo studies are needed to better evaluate the potential of DHA in humans as a therapeutic agent to treat periodontal disease.

  20. Effects of granulocyte-macrophage colony-stimulating factor and interleukin 6 on the growth of leukemic blasts in suspension culture.

    Science.gov (United States)

    Tsao, C J; Cheng, T Y; Chang, S L; Su, W J; Tseng, J Y

    1992-05-01

    We examined the stimulatory effects of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 6 (IL)-6 on the in vitro proliferation of leukemic blast cells from patients with acute leukemia. Bone marrow or peripheral blood leukemic blast cells were obtained from 21 patients, including 14 cases of acute myeloblastic leukemia (AML), four cases of acute lymphoblastic leukemia (ALL), two cases of acute undifferentiated leukemia, and one case of acute mixed-lineage leukemia. The proliferation of leukemic blast cells was evaluated by measuring the incorporation of 3H-thymidine into cells incubated with various concentrations of cytokines for 3 days. GM-CSF stimulated the DNA synthesis (with greater than 2.0 stimulation index) of blast cells in 9 of 14 (64%) AML cases, two cases of acute undifferentiated leukemia and one case of acute mixed-lineage leukemia. Only two cases of AML blasts responded to IL-6 to grow in the short-term suspension cultures. GM-CSF and IL-6 did not display a synergistic effect on the growth of leukemic cells. Moreover, GM-CSF and IL-6 did not stimulate the proliferation of ALL blast cells. Binding study also revealed the specific binding of GM-CSF on the blast cells of acute undifferentiated leukemia and acute mixed-lineage leukemia. Our results indicated that leukemic blast cells of acute undifferentiated leukemia and acute mixed-lineage leukemia possessed functional GM-CSF receptors.

  1. Biochemical and ultrastructural analysis of β-VLDL and AC-LDL metabolism by pigeon monocyte-derived macrophages in culture

    International Nuclear Information System (INIS)

    Henson, D.A.

    1987-01-01

    It is proposed that monocyte-derived foam cells in atherosclerotic lesions of White Carneau pigeons become lipid-filled through the uptake of lipoproteins including β-migrating very low density lipoproteins (β-VLDL) and acetylated low density lipoproteins (Ac-LDL). Using iodinated forms of the above lipoproteins, specific and saturable receptors for both β-VLDL and Ac-LDL were detected on the surface of White Carneau pigeon monocyte-derived macrophages in culture. Competition studies demonstrated the high degree of binding specificity for 125 I-Ac-LDL. Likewise, binding of 125 I-β-VLDL to its receptor was significantly inhibited by excess β-VLDL, however LDL from both hyper- and normocholesterolemic pigeons were also recognized by the receptor. Upon binding of β-VLDL and Ac-LDL to their respective receptors, the lipoproteins were rapidly internalized and delivered to intracellular sites of degradation. As measured by the amount of 14 C-oleate incorporated into cholesteryl 14 C-oleate, the cholesterole liberated from the degradation of both β-VLDL and Ac-LDL stimulated cholesteryl ester synthesis in the pigeon cells. Using lipoproteins conjugated to colloidal gold of visualization with transmission electron microscopy, a major difference in the binding and uptake properties of β-VLDL-Gold and Ac-LDL-Gold was documented

  2. Genomic variation in macrophage-cultured European porcine reproductive and respiratory syndrome virus Olot/91 revealed using ultra-deep next generation sequencing.

    Science.gov (United States)

    Lu, Zen H; Brown, Alexander; Wilson, Alison D; Calvert, Jay G; Balasch, Monica; Fuentes-Utrilla, Pablo; Loecherbach, Julia; Turner, Frances; Talbot, Richard; Archibald, Alan L; Ait-Ali, Tahar

    2014-03-04

    Porcine Reproductive and Respiratory Syndrome (PRRS) is a disease of major economic impact worldwide. The etiologic agent of this disease is the PRRS virus (PRRSV). Increasing evidence suggest that microevolution within a coexisting quasispecies population can give rise to high sequence heterogeneity in PRRSV. We developed a pipeline based on the ultra-deep next generation sequencing approach to first construct the complete genome of a European PRRSV, strain Olot/9, cultured on macrophages and then capture the rare variants representative of the mixed quasispecies population. Olot/91 differs from the reference Lelystad strain by about 5% and a total of 88 variants, with frequencies as low as 1%, were detected in the mixed population. These variants included 16 non-synonymous variants concentrated in the genes encoding structural and nonstructural proteins; including Glycoprotein 2a and 5. Using an ultra-deep sequencing methodology, the complete genome of Olot/91 was constructed without any prior knowledge of the sequence. Rare variants that constitute minor fractions of the heterogeneous PRRSV population could successfully be detected to allow further exploration of microevolutionary events.

  3. Dexamethasone targeted directly to macrophages induces macrophage niches that promote erythroid expansion

    DEFF Research Database (Denmark)

    Falchi, Mario; Varricchio, Lilian; Martelli, Fabrizio

    2015-01-01

    Cultures of human CD34(pos) cells stimulated with erythroid growth factors plus dexamethasone, a model for stress erythropoiesis, generate numerous erythroid cells plus a few macrophages (approx. 3%; 3:1 positive and negative for CD169). Interactions occurring between erythroblasts and macrophages...... in these cultures and the biological effects associated with these interactions were documented by live phase-contrast videomicroscopy. Macrophages expressed high motility interacting with hundreds/thousands of erythroblasts per hour. CD169(pos) macrophages established multiple rapid 'loose' interactions...... with proerythroblasts leading to formation of transient erythroblastic island-like structures. By contrast, CD169(neg) macrophages established 'tight' interactions with mature erythroblasts and phagocytosed these cells. 'Loose' interactions of CD169(pos) macrophages were associated with proerythroblast cytokinesis (the...

  4. Fish oil-derived long-chain n-3 polyunsaturated fatty acids reduce expression of M1-associated macrophage markers in an ex vivo adipose tissue culture model, in part through adiponectin

    Directory of Open Access Journals (Sweden)

    Anna A. De Boer

    2015-10-01

    Full Text Available Adipose tissue (AT macrophages (ATM play a key role in obesity-associated pathologies, and their phenotype can be influenced by the local tissue microenvironment. Interestingly, long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFA and the LC n-3 PUFA-upregulated adipokine, adiponectin (Ad, may mitigate excessive ATM inflammatory M1-polarization responses. However, to what extent LC n-3 PUFA and Ad work in concert to affect macrophage phenotype has not been examined. Thus, we used an established ex vivo AT organ culture model using visceral AT from mice fed a control (CON; 10% w/w safflower oil n-6 PUFA-rich diet or an isocaloric fish-oil (FO; 3% w/w menhaden oil + 7% w/w safflower oil-derived LC n-3 PUFA-rich diet to generate AT conditioned media (ACM. We then evaluated if CON or FO ACM affected macrophage polarization markers in a model designed to mimic acute (18 h ACM plus LPS for the last 6 h or chronic (macrophages treated with LPS-challenged CON or FO ACM for 24 h inflammation ± Ad-neutralizing antibody and the LPS-neutralizing agent, polymyxin B. In the acute inflammation model, macrophages treated with FO ACM had decreased lipid uptake and mRNA expression of M1 markers (Nos2, Nfκb, Il6, Il18, Ccl2 and Ccl5 compared with CON ACM (p≤0.05; however, these effects were largely attenuated when Ad was neutralized (p>0.05. Further, in the chronic inflammation model, macrophages treated with FO ACM had decreased mRNA expression of M1 markers (Nos2, Tnfα, Ccl2 and Il1β and IL-6 and CCL2 secretion (p≤0.05; however, some of these effects were lost when Ad was neutralized, and were further exacerbated when both Ad and LPS were neutralized. Taken together, this work shows that LC n-3 PUFA and Ad work in concert to suppress certain M1 macrophage responses. Thus, future strategies to modulate the ATM phenotype should consider the role of both LC n-3 PUFA and Ad in mitigating obese AT inflammation.

  5. Fish Oil-Derived Long-Chain n-3 Polyunsaturated Fatty Acids Reduce Expression of M1-Associated Macrophage Markers in an ex vivo Adipose Tissue Culture Model, in Part through Adiponectin.

    Science.gov (United States)

    De Boer, Anna A; Monk, Jennifer M; Liddle, Danyelle M; Power, Krista A; Ma, David W L; Robinson, Lindsay E

    2015-01-01

    Adipose tissue (AT) macrophages (ATM) play a key role in obesity-associated pathologies, and their phenotype can be influenced by the local tissue microenvironment. Interestingly, long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFA) and the LC n-3 PUFA-upregulated adipokine, adiponectin (Ad), may mitigate excessive ATM inflammatory M1-polarization responses. However, to what extent LC n-3 PUFA and Ad work in concert to affect macrophage phenotype has not been examined. Thus, we used an established ex vivo AT organ culture model using visceral AT from mice fed a control (CON; 10% w/w safflower oil) n-6 PUFA-rich diet or an isocaloric fish oil (FO; 3% w/w menhaden oil + 7% w/w safflower oil)-derived LC n-3 PUFA-rich diet to generate AT conditioned media (ACM). We then evaluated if CON or FO ACM affected macrophage polarization markers in a model designed to mimic acute [18 h ACM plus lipopolysaccharide (LPS) for the last 6 h] or chronic (macrophages treated with LPS-challenged CON or FO ACM for 24 h) inflammation ± Ad-neutralizing antibody and the LPS-neutralizing agent, polymyxin B. In the acute inflammation model, macrophages treated with FO ACM had decreased lipid uptake and mRNA expression of M1 markers (Nos2, Nfκb, Il6, Il18, Ccl2, and Ccl5) compared with CON ACM (p ≤ 0.05); however, these effects were largely attenuated when Ad was neutralized (p > 0.05). Furthermore, in the chronic inflammation model, macrophages treated with FO ACM had decreased mRNA expression of M1 markers (Nos2, Tnfα, Ccl2, and Il1β) and IL-6 and CCL2 secretion (p ≤ 0.05); however, some of these effects were lost when Ad was neutralized, and were further exacerbated when both Ad and LPS were neutralized. Taken together, this work shows that LC n-3 PUFA and Ad work in concert to suppress certain M1 macrophage responses. Thus, future strategies to modulate the ATM phenotype should consider the role of both LC n-3 PUFA and Ad in mitigating obese AT

  6. The Nuclear Orphan Receptor NR4A1 is Involved in the Apoptotic Pathway Induced by LPS and Simvastatin in RAW 264.7 Macrophages.

    Science.gov (United States)

    Kim, Yong Chan; Song, Seok Bean; Lee, Sang Kyu; Park, Sang Min; Kim, Young Sang

    2014-04-01

    Macrophage death plays a role in several physiological and inflammatory pathologies such as sepsis and arthritis. In our previous work, we showed that simvastatin triggers cell death in LPS-activated RAW 264.7 mouse macrophage cells through both caspase-dependent and independent apoptotic pathways. Here, we show that the nuclear orphan receptor NR4A1 is involved in a caspase-independent apoptotic process induced by LPS and simvastatin. Simvastatin-induced NR4A1 expression in RAW 264.7 macrophages and ectopic expression of a dominant-negative mutant form of NR4A1 effectively suppressed both DNA fragmentation and the disruption of mitochondrial membrane potential (MMP) during LPS- and simvastatin-induced apoptosis. Furthermore, apoptosis was accompanied by Bcl-2-associated X protein (Bax) translocation to the mitochondria. Our findings suggest that NR4A1 expression and mitochondrial translocation of Bax are related to simvastatin-induced apoptosis in LPS-activated RAW 264.7 macrophages.

  7. Concurrent suppression of NF-κB, p38 MAPK and reactive oxygen species formation underlies the effect of a novel compound isolated from Curcuma comosa Roxb. in LPS-activated microglia.

    Science.gov (United States)

    Jiamvoraphong, Nittaya; Jantaratnotai, Nattinee; Sanvarinda, Pantip; Tuchinda, Patoomratana; Piyachaturawat, Pawinee; Thampithak, Anusorn; Sanvarinda, Pimtip

    2017-07-01

    We investigated the molecular mechanisms underlying the effect of (3S)-1-(3,4-dihydroxyphenyl)-7-phenyl-(6E)-6-hepten-3-ol, also known as compound 092, isolated from Curcuma comosa Roxb on the production of pro-inflammatory mediators and oxidative stress in lipopolysaccharide (LPS)-activated highly aggressive proliferating immortalized (HAPI) microglial cell lines. Nitric oxide (NO) production was determined using the Griess reaction, and reverse transcription polymerase chain reaction was used to measure the expression of inducible nitric oxide synthase (iNOS) mRNA. Western blotting was used to determine the levels of pro-inflammatory mediators and their related upstream proteins. Compound 092 suppressed NO production and iNOS expression in LPS-stimulated HAPI cells. These effects originated from the ability of compound 092 to attenuate the activation of nuclear factor (NF)-κB as determined by the reduction in p-NF-κB and p-IκB kinase (IKK) protein levels. Compound 092 also significantly lowered LPS-activated intracellular reactive oxygen species production and p38 mitogen-activated protein kinase (MAPK) activation. Compound 092 suppresses microglial activation through attenuation of p38 MAPK and NF-κB activation. Compound 092 thus holds the potential to treat neurodegenerative disorders associated with neuroinflammation and oxidative stress. © 2017 Royal Pharmaceutical Society.

  8. Nicotine Impairs Macrophage Control of Mycobacterium tuberculosis.

    Science.gov (United States)

    Bai, Xiyuan; Stitzel, Jerry A; Bai, An; Zambrano, Cristian A; Phillips, Matthew; Marrack, Philippa; Chan, Edward D

    2017-09-01

    Pure nicotine impairs macrophage killing of Mycobacterium tuberculosis (MTB), but it is not known whether the nicotine component in cigarette smoke (CS) plays a role. Moreover, the mechanisms by which nicotine impairs macrophage immunity against MTB have not been explored. To neutralize the effects of nicotine in CS extract, we used a competitive inhibitor to the nicotinic acetylcholine receptor (nAChR)-mecamylamine-as well as macrophages derived from mice with genetic disruption of specific subunits of nAChR. We also determined whether nicotine impaired macrophage autophagy and whether nicotine-exposed T regulatory cells (Tregs) could subvert macrophage anti-MTB immunity. Mecamylamine reduced the CS extract increase in MTB burden by 43%. CS extract increase in MTB was also significantly attenuated in macrophages from mice with genetic disruption of either the α7, β2, or β4 subunit of nAChR. Nicotine inhibited autophagosome formation in MTB-infected THP-1 cells and primary murine alveolar macrophages, as well as increased the intracellular MTB burden. Nicotine increased migration of THP-1 cells, consistent with the increased number of macrophages found in the lungs of smokers. Nicotine induced Tregs to produce transforming growth factor-β. Naive mouse macrophages co-cultured with nicotine-exposed Tregs had significantly greater numbers of viable MTB recovered with increased IL-10 production and urea production, but no difference in secreted nitric oxide as compared with macrophages cocultured with unexposed Tregs. We conclude that nicotine in CS plays an important role in subverting macrophage control of MTB infection.

  9. A co-culture system with three different primary human cell populations reveals that biomaterials and MSC modulate macrophage-driven fibroblast recruitment.

    Science.gov (United States)

    Caires, Hugo R; Barros da Silva, Patrícia; Barbosa, Mário A; Almeida, Catarina R

    2018-03-01

    The biological response to implanted biomaterials is a complex and highly coordinated phenomenon involving many different cell types that interact within 3D microenvironments. Here, we increased the complexity of a 3D platform to include at least 3 cell types that play a role in the host response upon scaffold implantation. With this system, it was possible to address how immune responses triggered by 3D biomaterials mediate recruitment of stromal cells that promote tissue regeneration, mesenchymal stromal/stem cells (MSC), or a foreign body response, fibroblasts. Primary human macrophages yielded the highest fibroblast recruitment when interacting with chitosan scaffolds but not polylactic acid. Interestingly, when there were MSC and fibroblasts in the same environment, macrophages in chitosan scaffolds again promoted a significant increase on fibroblast recruitment, but not of MSC. However, macrophages that were firstly allowed to interact with MSC within the scaffolds were no longer able to recruit fibroblasts. This study illustrates the potential to use different scaffolds to regulate the dynamics of recruitment of proregenerative or fibrotic cell types through immunomodulation. Overall, this work strengths the idea that ex vivo predictive systems need to consider the different players involved in the biological response to biomaterials and that timing of arrival of specific cell types will affect the outcome. Copyright © 2017 John Wiley & Sons, Ltd.

  10. Lactoferricin mediates Anti-Inflammatory and Anti-Catabolic Effects via Inhibition of IL-1 and LPS Activity in the Intervertebral Disc†

    Science.gov (United States)

    Kim, Jae-Sung; Ellman, Michael B.; Yan, Dongyao; An, Howard S.; Kc, Ranjan; Li, Xin; Chen, Di; Xiao, Guozhi; Cs-Zabo, Gabriella; Hoskin, David W.; Buechter, D.D.; Van Wijnen, Andre J.; Im, Hee-Jeong

    2013-01-01

    The catabolic cytokine interleukin-1 (IL-1) and endotoxin lipopolysaccharide (LPS) are well-known inflammatory mediators involved in degenerative disc disease, and inhibitors of IL-1 and LPS may potentially be used to slow or prevent disc degeneration in vivo. Here, we elucidate the striking anti-catabolic and anti-inflammatory effects of bovine lactoferricin (LfcinB) in the intervertebral disc (IVD) via antagonism of both IL-1 and LPS-mediated catabolic activity using in vitro and ex vivo analyses. Specifically, we demonstrate the biological counteraction of LfcinB against IL-1 and LPS-mediated proteoglycan (PG) depletion, matrix-degrading enzyme production and enzyme activity in long-term (alginate beads) and short-term (monolayer) culture models using bovine and human nucleus pulposus (NP) cells. LfcinB significantly attenuates the IL-1 and LPS-mediated suppression of PG production and synthesis, and thus restores PG accumulation and pericellular matrix formation. Simultaneously, LfcinB antagonizes catabolic factor mediated induction of multiple cartilage-degrading enzymes, including MMP-1, MMP-3, MMP-13, ADAMTS-4, and ADAMTS-5, in bovine NP cells at both mRNA and protein levels. LfcinB also suppresses the catabolic factor-induced stimulation of oxidative and inflammatory factors such as iNOS, IL-6, and toll-like receptor-2 (TLR-2) and TLR-4. Finally, the ability of LfcinB to antagonize IL-1 and LPS-mediated suppression of PG is upheld in an en bloc intradiscal microinjection model followed by ex vivo organ culture using both mouse and rabbit IVD tissue, suggesting a potential therapeutic benefit of LfcinB on degenerative disc disease in the future. PMID:23460134

  11. Lactoferricin mediates anti-inflammatory and anti-catabolic effects via inhibition of IL-1 and LPS activity in the intervertebral disc.

    Science.gov (United States)

    Kim, Jae-Sung; Ellman, Michael B; Yan, Dongyao; An, Howard S; Kc, Ranjan; Li, Xin; Chen, Di; Xiao, Guozhi; Cs-Szabo, Gabriella; Hoskin, David W; Buechter, Doug D; Van Wijnen, Andre J; Im, Hee-Jeong

    2013-09-01

    The catabolic cytokine interleukin-1 (IL-1) and endotoxin lipopolysaccharide (LPS) are well-known inflammatory mediators involved in degenerative disc disease, and inhibitors of IL-1 and LPS may potentially be used to slow or prevent disc degeneration in vivo. Here, we elucidate the striking anti-catabolic and anti-inflammatory effects of bovine lactoferricin (LfcinB) in the intervertebral disc (IVD) via antagonism of both IL-1 and LPS-mediated catabolic activity using in vitro and ex vivo analyses. Specifically, we demonstrate the biological counteraction of LfcinB against IL-1 and LPS-mediated proteoglycan (PG) depletion, matrix-degrading enzyme production, and enzyme activity in long-term (alginate beads) and short-term (monolayer) culture models using bovine and human nucleus pulposus (NP) cells. LfcinB significantly attenuates the IL-1 and LPS-mediated suppression of PG production and synthesis, and thus restores PG accumulation and pericellular matrix formation. Simultaneously, LfcinB antagonizes catabolic factor mediated induction of multiple cartilage-degrading enzymes, including MMP-1, MMP-3, MMP-13, ADAMTS-4, and ADAMTS-5, in bovine NP cells at both mRNA and protein levels. LfcinB also suppresses the catabolic factor-induced stimulation of oxidative and inflammatory factors such as iNOS, IL-6, and toll-like receptor-2 (TLR-2) and TLR-4. Finally, the ability of LfcinB to antagonize IL-1 and LPS-mediated suppression of PG is upheld in an en bloc intradiscal microinjection model followed by ex vivo organ culture using both mouse and rabbit IVD tissue, suggesting a potential therapeutic benefit of LfcinB on degenerative disc disease in the future. Copyright © 2013 Wiley Periodicals, Inc.

  12. LL-37 directs macrophage differentiation toward macrophages with a proinflammatory signature.

    Science.gov (United States)

    van der Does, Anne M; Beekhuizen, Henry; Ravensbergen, Bep; Vos, Tim; Ottenhoff, Tom H M; van Dissel, Jaap T; Drijfhout, Jan W; Hiemstra, Pieter S; Nibbering, Peter H

    2010-08-01

    The human cathelicidin LL-37 has broad-spectrum antimicrobial activity. It also participates at the interface of innate and adaptive immunity by chemoattracting immune effector cells, modulating the production of a variety of inflammatory mediators by different cell types, and regulating the differentiation of monocytes into dendritic cells. In this study, we investigated the effects of LL-37 on the differentiation of human monocytes into anti-inflammatory macrophages (MPhi-2; driven by M-CSF) versus proinflammatory macrophages (MPhi-1; driven by GM-CSF) as well as on fully differentiated MPhi-1 and MPhi-2. Results revealed that monocytes cultured with M-CSF in the presence of LL-37 resulted in macrophages displaying a proinflammatory signature, namely, low expression of CD163 and little IL-10 and profound IL-12p40 production on LPS stimulation. The effects of LL-37 on M-CSF-driven macrophage differentiation were dose- and time-dependent with maximal effects observed at 10 microg/ml when the peptide was present from the start of the cultures. The peptide enhanced the GM-CSF-driven macrophage differentiation. Exposure of fully differentiated MPhi-2 to LL-37 for 6 d resulted in macrophages that produced less IL-10 and more IL-12p40 on LPS stimulation than control MPhi-2. In contrast, LL-37 had no effect on fully differentiated MPhi-1. Peptide mapping using a set of 16 overlapping 22-mer peptides covering the complete LL-37 sequence revealed that the C-terminal portion of LL-37 is responsible for directing macrophage differentiation. Our results furthermore indicate that the effects of LL-37 on macrophage differentiation required internalization of the peptide. Together, we conclude that LL-37 directs macrophage differentiation toward macrophages with a proinflammatory signature.

  13. Helicobacter pylori protein HP0986 (TieA) interacts with mouse TNFR1 and triggers proinflammatory and proapoptotic signaling pathways in cultured macrophage cells (RAW 264.7).

    Science.gov (United States)

    Ansari, Suhail A; Devi, Savita; Tenguria, Shivendra; Kumar, Ashutosh; Ahmed, Niyaz

    2014-08-01

    HP0986 protein of Helicobacter pylori has been shown to trigger induction of proinflammatory cytokines (IL-8 and TNF-α) through the activation of NF-κB and also to induce Fas mediated apoptosis of human macrophage cells (THP-1). In this study, we unravel mechanistic details of the biological effects of this protein in a murine macrophage environment. Up regulation of MCP-1 and TNF-α in HP0986-induced RAW 264.7 cells occurred subsequent to the activation and translocation of NF-κB to the cell nucleus. Further, HP0986 induced apoptosis of RAW 264.7 cells through Fas activation and this was in agreement with previous observations made with THP-1 cells. Our studies indicated activation of TNFR1 through interaction with HP0986 and this elicited the aforementioned responses independent of TLR2, TLR4 or TNFR2. We found that mouse TNFR1 activation by HP0986 facilitates formation of a complex comprising of TNFR1, TRADD and TRAF2, and this occurs upstream of NF-κB activation. Furthermore, FADD also forms a second complex, at a later stage, together with TNFR1 and TRADD, resulting in caspase-8 activation and thereby the apoptosis of RAW 264.7 cells. In summary, our observations reveal finer details of the functional activity of HP0986 protein in relation to its behavior in a murine macrophage cell environment. These findings reconfirm the proinflammatory and apoptotic role of HP0986 signifying it to be an important trigger of innate responses. These observations form much needed baseline data entailing future in vivo studies of the functions of HP0986 in a murine model. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Comparative microscopic and biochemical study of the uptake of fluorescent and 125I-labeled lipoproteins by skin fibroblasts, smooth muscle cells, and peritoneal macrophages in culture

    International Nuclear Information System (INIS)

    Reynolds, G.D.; St Clair, R.W.

    1985-01-01

    Uptake of low density lipoprotein (LDL) and of acetyl LDL was compared in skin fibroblasts, smooth muscle cells, and peritoneal macrophages with the use of lipoproteins labeled with either 125 I or the fluorescent probe 3,3'-dioctadecylindocarbocyanine (DiI). The uptake of DiI-labeled lipoproteins was assessed by quantitative spectrofluorometry and by fluorescence microscopy. The DiI was quantitatively retained by the cells, while the 125 I-LDL was degraded and 125 I-labeled degradation products were excreted from the cells. In smooth muscle cells and fibroblasts the uptake of LDL was virtually the same whether measured with the use of the DiI or 125 I-label. The labeling of acetyl LDL with DiI enhanced its uptake in peritoneal macrophages by an average of 18%. With the DiI label, lipoprotein uptake could be determined after as little as 10 minutes of incubation at 37 C. The pattern of uptake of the DiI-labeled lipoproteins was consistent with binding to specific receptors, because no DiI could be detected in mutant cells without LDL receptors, and uptake was competitively inhibited by addition of excess unlabeled lipoprotein. When the DiI-labeled lipoproteins were removed from the medium, there was a 5-15% loss of DiI from all cell types studied over the first 24 hours

  15. BMP pathway regulation of and by macrophages.

    Directory of Open Access Journals (Sweden)

    Megha Talati

    Full Text Available Pulmonary arterial hypertension (PAH is a disease of progressively increasing pulmonary vascular resistance, associated with mutations of the type 2 receptor for the BMP pathway, BMPR2. The canonical signaling pathway for BMPR2 is through the SMAD family of transcription factors. BMPR2 is expressed in every cell type, but the impact of BMPR2 mutations affecting SMAD signaling, such as Bmpr2delx4+, had only previously been investigated in smooth muscle and endothelium. In the present study, we created a mouse with universal doxycycline-inducible expression of Bmpr2delx4+ in order to determine if broader expression had an impact relevant to the development of PAH. We found that the most obvious phenotype was a dramatic, but patchy, increase in pulmonary inflammation. We crossed these double transgenic mice onto an NF-κB reporter strain, and by luciferase assays on live mice, individual organs and isolated macrophages, we narrowed down the origin of the inflammatory phenotype to constitutive activation of tissue macrophages. Study of bone marrow-derived macrophages from mutant and wild-type mice suggested a baseline difference in differentiation state in Bmpr2 mutants. When activated with LPS, both mutant and wild-type macrophages secrete BMP pathway inhibitors sufficient to suppress BMP pathway activity in smooth muscle cells (SMC treated with conditioned media. Functionally, co-culture with macrophages results in a BMP signaling-dependent increase in scratch closure in cultured SMC. We conclude that SMAD signaling through BMP is responsible, in part, for preventing macrophage activation in both live animals and in cells in culture, and that activated macrophages secrete BMP inhibitors in sufficient quantity to cause paracrine effect on vascular smooth muscle.

  16. In vitro studies of interaction of rickettsia and macrophages: effect of ultraviolet light on Coxiella burnetti inactivation and macrophage enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Little, J.S.; Kishimoto, R.A.; Canonico, P.G.

    1980-03-01

    The inactivation of Coxiella burnetii in suspension or in cultures of guinea pig peritoneal macrophages by ultraviolet (uv) light was studied. The effect of uv treatment on the activity of macrophage organelle marker enzymes and their subsequent equilibration in linear sucrose gradients was also determined. It was shown that uv treatment for 15 s at a distance of 10 cm inactivated C. burnetti, either in suspension or within guinea pig peritoneal macrophages. Similar uv treatment had little effect on the activity or equilibration of macrophage organelle marker enzymes in linear sucrose gradients.

  17. In vitro studies of interaction of rickettsia and macrophages: effect of ultraviolet light on Coxiella burnetti inactivation and macrophage enzymes

    International Nuclear Information System (INIS)

    Little, J.S.; Kishimoto, R.A.; Canonico, P.G.

    1980-01-01

    The inactivation of Coxiella burnetii in suspension or in cultures of guinea pig peritoneal macrophages by ultraviolet (uv) light was studied. The effect of uv treatment on the activity of macrophage organelle marker enzymes and their subsequent equilibration in linear sucrose gradients was also determined. It was shown that uv treatment for 15 s at a distance of 10 cm inactivated C. burnetti, either in suspension or within guinea pig peritoneal macrophages. Similar uv treatment had little effect on the activity or equilibration of macrophage organelle marker enzymes in linear sucrose gradients

  18. Macrophages in synovial inflammation

    Directory of Open Access Journals (Sweden)

    Aisling eKennedy

    2011-10-01

    Full Text Available AbstractSynovial macrophages are one of the resident cell types in synovial tissue and while they remain relatively quiescent in the healthy joint, they become activated in the inflamed joint and, along with infiltrating monocytes/macrophages, regulate secretion of pro-inflammatory cytokines and enzymes involved in driving the inflammatory response and joint destruction. Synovial macrophages are positioned throughout the sub-lining layer and lining layer at the cartilage-pannus junction and mediate articular destruction. Sub-lining macrophages are now also considered as the most reliable biomarker for disease severity and response to therapy in rheumatoid arthritis (RA. There is a growing understanding of the molecular drivers of inflammation and an appreciation that the resolution of inflammation is an active process rather than a passive return to homeostasis, and this has implications for our understanding of the role of macrophages in inflammation. Macrophage phenotype determines the cytokine secretion profile and tissue destruction capabilities of these cells. Whereas inflammatory synovial macrophages have not yet been classified into one phenotype or another it is widely known that TNFα and IL-l, characteristically released by M1 macrophages, are abundant in RA while IL-10 activity, characteristic of M2 macrophages, is somewhat diminished.Here we will briefly review our current understanding of macrophages and macrophage polarisation in RA as well as the elements implicated in controlling polarisation, such as cytokines and transcription factors like NFκB, IRFs and NR4A, and pro-resolving factors, such as LXA4 and other lipid mediators which may promote a non-inflammatory, pro-resolving phenotype and may represent a novel therapeutic paradigm.

  19. [Macrophages in human semen].

    Science.gov (United States)

    Bouvet, Beatriz Reina; Brufman, Adriana Silvia; Paparella, Cecilia Vicenta; Feldman, Rodolfo Nestor; Gatti, Vanda Nora; Solis, Edita Amalia

    2003-11-01

    To investigate the presence of macrophages in human semen samples and the function they carry out in the seminal fluid. Their presence was studied in relation to spermatic morphology, percentage of spermatozoids with native DNA, and presence of antispermatic antibodies. The work was performed with semen samples from 31 unfertile males from 63 couples in which the "female factor" was ruled out as the cause of infertility. Sperm study according to WHO (1992) was carried out in all samples, in addition to: DNA study with acridine orange as fluorocrom, macrophage concentration by neutral red in a Neubauer camera, and detection of antispermatic antibodies with a mixed agglutination test (TAC II) (validated with Mar Screen-Fertility technologies). Sperm morphology was evaluated by Papanicolaou test. 19/31 selected sperm samples (61.3%) showed increased concentration of macrophages, 13 of them (41.9%) with denaturalized DNA, and 8 (25.8%) abnormal morphology. Six samples showed increased macrophage concentration and predominance of native DNA, whereas 11 samples showed increased macrophages and abnormal morphology. Among 18 (58.1%) samples showing antispermatic antibodies 14 (77.7%) had an increased concentration of macrophages. Statistical analysis resulted in a high correlation between macrophage concentration and increased percentage of spermatozoids with denaturalized DNA (p < 0.05). An increased concentration of macrophages is associated with the presence of antispermatic antibodies (p < 0.05). There was not evidence of significant association between concentration of macrophages and percentage of morphologically normal spermatozoids (p < 0.05). We can conclude that macrophages are present in human semen and participate in immunovigilance contributing to improve the seminal quality.

  20. Lipopolysaccharide modulation of a CD14-like molecule on porcine alveolar macrophages

    Science.gov (United States)

    Kielian, T. L.; Ross, C. R.; McVey, D. S.; Chapes, S. K.; Blecha, F.; Spooner, B. S. (Principal Investigator)

    1995-01-01

    Cluster of differentiation antigen 14 (CD14) functions as a receptor for lipopolysaccharide (LPS) LPS-binding protein (LBP) complexes. Because LPS has varying effects on CD14 expression in vitro, we evaluated CD14 expression in response to LPS with a fully differentiated macrophage phenotype, the alveolar macrophage. By using flow microfluorometric analysis and a radioimmunoassay with an anti-human CD14 monoclonal antibody (My4) that cross-reacts with porcine CD14, we found that macrophages stimulated with LPS for 24 h exhibited a two- to fivefold increase in CD14-like antigen compared with unstimulated cells. At low concentrations of LPS, up-regulation of the CD14-like antigen was dependent on serum; at higher concentrations of LPS, serum was not required. In the absence of serum a 10-fold higher dose of LPS (10 ng/ml) was required to increase CD14-like expression. In addition, LPS-induced CD14-like up-regulation correlated with secretion of tumor necrosis factor-alpha, regardless of serum concentration. Blockade with My4 antibody significantly inhibited LPS-induced tumor necrosis factor-alpha secretion at 1 ng/ml of LPS. However, inhibition decreased as we increased the LPS concentration, suggesting the existence of CD14-independent pathways of macrophage activation in response to LPS. Alternatively, My4 may have a lower affinity for the porcine CD14 antigen than LPS, which may have only partially blocked the LPS-LBP binding site at high concentrations of LPS. Therefore, these data suggest that LPS activation of porcine alveolar macrophages for 24 h increased CD14-like receptor expression. The degree of CD14-like up-regulation was related to LPS concentration, however, activation did not require the presence of serum at high concentrations of LPS.

  1. L-Plastin promotes podosome longevity and supports macrophage motility

    Science.gov (United States)

    Zhou, Julie Y.; Szasz, Taylor P.; Stewart-Hutchinson, Phillip J.; Sivapalan, Janardan; Todd, Elizabeth M.; Deady, Lauren E.; Cooper, John A.; Onken, Michael D.; Morley, S. Celeste

    2016-01-01

    Elucidating the molecular regulation of macrophage migration is essential for understanding the patho-physiology of multiple human diseases, including host responses to infection and autoimmune disorders. Macrophage migration is supported by dynamic rearrangements of the actin cytoskeleton, with formation of actin-based structures such as podosomes and lamellipodia. Here we provide novel insights into the function of the actin-bundling protein l-plastin (LPL) in primary macrophages. We found that podosome stability is disrupted in primary resident peritoneal macrophages from LPL−/− mice. Live-cell imaging of F-actin using resident peritoneal macrophages from LifeACT-RFP+ mice demonstrated that loss of LPL led to decreased longevity of podosomes, without reducing the number of podosomes initiated. Additionally, macrophages from LPL−/− mice failed to elongate in response to chemotactic stimulation. These deficiencies in podosome stabilization and in macrophage elongation correlated with impaired macrophage transmigration in culture and decreased monocyte migration into murine peritoneum. Thus, we have identified a role for LPL in stabilizing long-lived podosomes and in enabling macrophage motility. PMID:27614263

  2. Agomelatine, a MT1/MT2 melatonergic receptor agonist with serotonin 5-HT2C receptor antagonistic properties, suppresses Prevotella intermedia lipopolysaccharide-induced production of proinflammatory mediators in murine macrophages.

    Science.gov (United States)

    Hyeon, Jin-Yi; Choi, Eun-Young; Choe, So-Hui; Park, Hae Ryoun; Choi, Jeom-Il; Choi, In Soon; Kim, Sung-Jo

    2017-10-01

    This study was performed in an attempt to examine the influence of agomelatine in mitigating the generation of proinflammatory mediators in RAW264.7 murine macrophages exposed to lipopolysaccharide (LPS) obtained from Prevotella intermedia, a gram-negative anaerobic bacterium that is related with various types of periodontal diseases, and the molecular mechanisms behind its effects. LPS from P. intermedia strain ATCC 25611 was prepared employing the conventional phenol-water procedure. Conditioned culture media were analyzed for the levels of nitric oxide (NO), interleukin-1β (IL-1β) and IL-6. Real-time PCR analysis was carried out to determine the mRNA levels of inducible NO synthase (iNOS), IL-1β, IL-6 and SOCS1. Protein expression levels were evaluated by immunoblot test. NF-κB-dependent SEAP reporter assay was performed using a reporter cell line. DNA-binding activities of NF-κB subunits were analyzed utilizing the ELISA-based kits. Agomelatine was found to down-regulate significantly the generation of iNOS-derived NO, IL-1β and IL-6 as well as the expression of their mRNAs in cells activated with P. intermedia LPS. Agomelatine decreased NF-κB-dependent SEAP release caused by P. intermedia LPS. Agomelatine did not inhibit NF-κB transcription induced by LPS at the level of IκB-α degradation. Instead, LPS-induced nuclear translocation and DNA binding of NF-κB p50 subunit was blocked by agomelatine. P. intermedia LPS-elicited activation of STAT1 and STAT3 was reduced notably by co-treatment with agomelatine. Agomelatine showed a tendency to enhance mRNA level of SOCS1 in LPS-activated cells as well. Agomelatine merits further evaluation to reveal its usefulness on the host modulation of periodontal disease. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Differential S1P Receptor Profiles on M1- and M2-Polarized Macrophages Affect Macrophage Cytokine Production and Migration.

    Science.gov (United States)

    Müller, Jan; von Bernstorff, Wolfram; Heidecke, Claus-Dieter; Schulze, Tobias

    2017-01-01

    Introduction . Macrophages are key players in complex biological processes. In response to environmental signals, macrophages undergo polarization towards a proinflammatory (M1) or anti-inflammatory (M2) phenotype. Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid that acts via 5 G-protein coupled receptors (S1P 1-5 ) in order to influence a broad spectrum of biological processes. This study assesses S1P receptor expression on macrophages before and after M1 and M2 polarization and performs a comparative analysis of S1P signalling in the two activational states of macrophages. Methods . Bone marrow derived macrophages (BMDM) from C57 BL/6 mice were cultured under either M1- or M2-polarizing conditions. S1P-receptor expression was determined by quantitative RT-PCR. Influence of S1P on macrophage activation, migration, phagocytosis, and cytokine secretion was assessed in vitro. Results . All 5 S1P receptor subclasses were expressed in macrophages. Culture under both M1- and M2-polarizing conditions led to significant downregulation of S1P 1 . In contrast, M1-polarized macrophages significantly downregulated S1P 4 . The expression of the remaining three S1P receptors did not change. S1P increased expression of iNOS under M2-polarizing conditions. Furthermore, S1P induced chemotaxis in M1 macrophages and changed cytokine production in M2 macrophages. Phagocytosis was not affected by S1P-signalling. Discussion . The expression of different specific S1P receptor profiles may provide a possibility to selectively influence M1- or M2-polarized macrophages.

  4. TNF-alpha, produced by feline infectious peritonitis virus (FIPV)-infected macrophages, upregulates expression of type II FIPV receptor feline aminopeptidase N in feline macrophages.

    Science.gov (United States)

    Takano, Tomomi; Hohdatsu, Tsutomu; Toda, Ayako; Tanabe, Maki; Koyama, Hiroyuki

    2007-07-20

    The pathogenicity of feline infectious peritonitis virus (FIPV) is known to depend on macrophage tropism, and this macrophage infection is enhanced by mediation via anti-S antibody (antibody-dependent enhancement, ADE). In this study, we found that TNF-alpha production was increased with viral replication in macrophages inoculated with a mixture of FIPV and anti-S antibody, and demonstrated that this culture supernatant had feline PBMC apoptosis-inducing activity. We also demonstrated that the expression level of the FIPV virus receptor, feline aminopeptidase N (fAPN), was increased in macrophages of FIP cats. For upregulation of TNF-alpha and fAPN in macrophages, viral replication in macrophages is necessary, and their expressions were increased by ADE of FIPV infection. It was demonstrated that a heat-resistant fAPN-inducing factor was present in the culture supernatant of FIPV-infected macrophages, and this factor was TNF-alpha: fAPN expression was upregulated in recombinant feline TNF-alpha-treated macrophages, and FIPV infectivity was increased in these macrophages. These findings suggested that FIPV replication in macrophages increases TNF-alpha production in macrophages, and the produced TNF-alpha acts and upregulates fAPN expression, increasing FIPV sensitivity.

  5. The elusive antifibrotic macrophage

    Directory of Open Access Journals (Sweden)

    Adhyatmika eAdhyatmika

    2015-11-01

    Full Text Available 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 behaviour 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 behaviour 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 behaviour.

  6. Therapeutic potential of regulatory macrophages generated from peritoneal dialysate in adriamycin nephropathy.

    Science.gov (United States)

    Cao, Qi; Wang, Yiping; Wang, Changqi; Wang, Xin M; Lee, Vincent W S; Zheng, Guoping; Zhao, Ye; Alexander, Stephen I; Harris, David C H

    2018-04-01

    Cell therapy using macrophages requires large amounts of cells, which are difficult to collect from patients. Patients undergoing peritoneal dialysis (PD) discard huge numbers of peritoneal macrophages in dialysate daily. Macrophages can be modulated to become regulatory macrophages, which have shown great promise as a therapeutic strategy in experimental kidney disease and human kidney transplantation. This study aimed to examine the potential of using peritoneal macrophages (PMs) from peritoneal dialysate to treat kidney disease. Monocytes/macrophages accounted for >40% of total peritoneal leukocytes in both patients and mice undergoing PD. PMs from patients and mice undergoing PD were more mature than peripheral monocytes/macrophages, as shown by low expression of C-C motif chemokine receptor 2 (CCR2) and morphological changes during in vitro culture. PMs from patients and mice undergoing PD displayed normal macrophage function and could be modulated into a regulatory (M2) phenotype. In vivo, adoptive transfer of peritoneal M2 macrophages derived from PD mice effectively protected against kidney injury in mice with adriamycin nephropathy (AN). Importantly, the transfused peritoneal M2 macrophages maintained their M2 phenotype in kidney of AN mice. In conclusion, PMs derived from patients and mice undergoing PD exhibited conventional macrophage features. Peritoneal M2 macrophages derived from PD mice are able to reduce kidney injury in AN, suggesting that peritoneal macrophages from patients undergoing PD may have the potential for clinical therapeutic application.

  7. Phenotypic, functional, and quantitative characterization of canine peripheral blood monocyte-derived macrophages

    Directory of Open Access Journals (Sweden)

    R Bueno

    2005-08-01

    Full Text Available The yield as well as phenotypic and functional parameters of canine peripheral blood monocyte-derived macrophages were analyzed. The cells that remained adherent to Teflon after 10 days of culture had high phagocytic activity when inoculated with Leishmania chagasi. Flow cytometric analysis demonstrated that more than 80% of cultured cells were positive for the monocyte/macrophage marker CD14.

  8. Thermo-radiosensitivity of the granulocyte and macrophage precursor cells of mice. I.-Development of the in vivo culture and effects induced by the hyperthermia; Termo-radiosensibilidad del precursor hematopoyetico que origina las series granulocitica y macrofaga de raton. I.- Desarrollo del cultivo in vivo y efectos producidos por la hipertermia

    Energy Technology Data Exchange (ETDEWEB)

    Bueren, J A; Nieto, M

    1983-07-01

    The present report shows the agar diffusion chamber technique for culturing granulocyte- macrophage precursor cells, obtained from mice bone marrow. Diffusion chambers containing the bone marrow suspension are implanted intraperitoneally Into mice and constitute a compartment which avoids the migration of cells, but allows the transit of the mouse biological fluxes, necessary for the cellular proliferation. By means of this technique, we studied the lethal effects of the hyperthermia on the precursors and their capacity to repair sublethal damage. (Author) 129 refs.

  9. Evaluation of cytotoxicity of two endodontic cements in a macrophage culture Avaliação da citotoxicidade de dois cimentos endodônticos em cultura de macrófagos

    Directory of Open Access Journals (Sweden)

    Celso Emanoel de Souza Queiroz

    2005-09-01

    Full Text Available Compared to gutta-percha, the endodontic cements are used in small quantity to seal root canals, but are indispensable to achieve hermetically sealed margins, where its biocompatibility depends on the sum of responses of each cell present in the periapical region. The object of this study was to evaluate the cytotoxicity of two endodontic cements, one based on epoxy resin (Sealer 26 and the other containing zinc oxide eugenol (Endofill by using cultured peritoneal macrophages from Swiss mice to measure the induced production of nitric oxide. After solidification and pulverization, aliquots of 100mul of suspension containing 18mg/mL of the respective cements were added to 96-well tissue culture plates containing the tissue culture of macrophages at a concentration of 5.0X10(6 cells/ml. In the positive control group the cell culture was treated with 10mg/mL of lipopolyssaccharide from Escherichia coli 026:B6 and the cell culture alone represented the negative control. After 48 hours of incubation, at 37ºC, in 5% CO2, the cultures were placed in an ELISA automatic reader to evaluate the release of nitric oxide. The production of nitric oxide for cement Sealer 26 was between 36.1 and 313.0 mumols, with a mean of 143.82±111.03mumols, while for the Endofill these values were significantly less (p=0.01, varying from 50.8 to 125.7mumols, with a mean of 80.33±28.42 mumols. In the positive and negative control groups the mean release of nitric oxide was of 162.75mumols and 4.42mumols, respectively. There was no significant difference between the positive control group and cement Sealer 26 (p>0.05. Therefore, the cement Sealer 26 caused significantly greater toxicity to the macrophages, possibly due to the components from the epoxy resin and formaldehyde release during polymerization.Comparativamente à guta-percha, os cimentos endodônticos são utilizados em pequena quantidade nas obturações dos canais radiculares, mas são imprescindíveis para

  10. The influence of macrophage growth factors on Theiler's Murine Encephalomyelitis Virus (TMEV) infection and activation of macrophages.

    Science.gov (United States)

    Schneider, Karin M; Watson, Neva B; Minchenberg, Scott B; Massa, Paul T

    2018-02-01

    Macrophages are common targets for infection and innate immune activation by many pathogenic viruses including the neurotropic Theiler's Murine Encephalomyelitis Virus (TMEV). As both infection and innate activation of macrophages are key determinants of viral pathogenesis especially in the central nervous system (CNS), an analysis of macrophage growth factors on these events was performed. C3H mouse bone-marrow cells were differentiated in culture using either recombinant macrophage colony stimulating factor (M-CSF) or granulocyte-macrophage colony-stimulating factor (GM-CSF), inoculated with TMEV (BeAn) and analyzed at various times thereafter. Cytokine RNA and protein analysis, virus titers, and flow cytometry were performed to characterize virological parameters under these culture conditions. GM-CSF-differentiated macrophages showed higher levels of TMEV viral RNA and proinflammatory molecules compared to infected M-CSF-differentiated cells. Thus, GM-CSF increases both TMEV infection and TMEV-induced activation of macrophages compared to that seen with M-CSF. Moreover, while infectious viral particles decreased from a peak at 12h to undetectable levels at 48h post infection, TMEV viral RNA remained higher in GM-CSF- compared to M-CSF-differentiated macrophages in concert with increased proinflammatory gene expression. Analysis of a possible basis for these differences determined that glycolytic rates contributed to heightened virus replication and proinflammatory cytokine secretion in GM-CSF compared to M-CSF-differentiated macrophages. In conclusion, we provide evidence implicating a role for GM-CSF in promoting virus replication and proinflammatory cytokine expression in macrophages, indicating that GM-CSF may be a key factor for TMEV infection and the induction of chronic TMEV-induced immunopathogenesis in the CNS. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Consistent inhibition of cyclooxygenase drives macrophages towards the inflammatory phenotype.

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    Yi Rang Na

    Full Text Available Macrophages play important roles in defense against infection, as well as in homeostasis maintenance. Thus alterations of macrophage function can have unexpected pathological results. Cyclooxygenase (COX inhibitors are widely used to relieve pain, but the effects of long-term usage on macrophage function remain to be elucidated. Using bone marrow-derived macrophage culture and long-term COX inhibitor treatments in BALB/c mice and zebrafish, we showed that chronic COX inhibition drives macrophages into an inflammatory state. Macrophages differentiated in the presence of SC-560 (COX-1 inhibitor, NS-398 (COX-2 inhibitor or indomethacin (COX-1/2 inhibitor for 7 days produced more TNFα or IL-12p70 with enhanced p65/IκB phosphoylation. YmI and IRF4 expression was reduced significantly, indicative of a more inflammatory phenotype. We further observed that indomethacin or NS-398 delivery accelerated zebrafish death rates during LPS induced sepsis. When COX inhibitors were released over 30 days from an osmotic pump implant in mice, macrophages from peritoneal cavities and adipose tissue produced more TNFα in both the basal state and under LPS stimulation. Consequently, indomethacin-exposed mice showed accelerated systemic inflammation after LPS injection. Our findings suggest that macrophages exhibit a more inflammatory phenotype when COX activities are chronically inhibited.

  12. Culture.

    Science.gov (United States)

    Smith, Timothy B; Rodríguez, Melanie Domenech; Bernal, Guillermo

    2011-02-01

    This article summarizes the definitions, means, and research of adapting psychotherapy to clients' cultural backgrounds. We begin by reviewing the prevailing definitions of cultural adaptation and providing a clinical example. We present an original meta-analysis of 65 experimental and quasi-experimental studies involving 8,620 participants. The omnibus effect size of d = .46 indicates that treatments specifically adapted for clients of color were moderately more effective with that clientele than traditional treatments. The most effective treatments tended to be those with greater numbers of cultural adaptations. Mental health services targeted to a specific cultural group were several times more effective than those provided to clients from a variety of cultural backgrounds. We recommend a series of research-supported therapeutic practices that account for clients' culture, with culture-specific treatments being more effective than generally culture-sensitive treatments. © 2010 Wiley Periodicals, Inc.

  13. Essential oil from leaves of Liquidambar formosana ameliorates inflammatory response in lipopolysaccharide-activated mouse macrophages.

    Science.gov (United States)

    Hua, Kuo-Feng; Yang, Tzu-Jung; Chiu, Huan-Wen; Ho, Chen-Lung

    2014-06-01

    The essential oil from Liquidambar formosana leaves (EOLF) was demonstrated to exhibit anti-inflammatory activity in mouse macrophages. EOLF reduced nitrite oxide generation, secretion levels of tumor necrosis factor-alpha and interleukin-6, and expression levels of prointerleukin-beta, inducible nitric oxide synthase, and cyclooxygenase-2 in lipopolysaccharide (LPS)-activated mouse macrophages. EOLF also reduced NLRP3 inflammasome-derived interleukin-1beta secretion. The underlying mechanisms for the EOLF-mediated anti-inflammatory activity were (1) reduction of LPS-induced reactive oxygen species generation; (2) reduction of LPS-induced activation of c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38 MAP kinase; (3) reduction of LPS-induced nuclear factor-kappaBeta activation. Furthermore, 25 compounds were identified in the EOLF using GC-FID and GC-MS and the major compounds were terpinen-4-ol (32.0%), beta-pinene (18.0%), gamma-terpinene (13.8%), and alpha-terpinene (9.7%). We found that LPS-induced nitrite oxide generation was inhibited significantly by terpinen-4-ol. Our results indicated that EOLF has anti-inflammatory activity and may provide a molecular rationale for future therapeutic interventions in immune modulation.

  14. Macrophage heterogeneity in tissues: phenotypic diversity and functions

    Science.gov (United States)

    Gordon, Siamon; Plüddemann, Annette; Martinez Estrada, Fernando

    2014-01-01

    During development and throughout adult life, macrophages derived from hematopoietic progenitors are seeded throughout the body, initially in the absence of inflammatory and infectious stimuli as tissue-resident cells, with enhanced recruitment, activation, and local proliferation following injury and pathologic insults. We have learned a great deal about macrophage properties ex vivo and in cell culture, but their phenotypic heterogeneity within different tissue microenvironments remains poorly characterized, although it contributes significantly to maintaining local and systemic homeostasis, pathogenesis, and possible treatment. In this review, we summarize the nature, functions, and interactions of tissue macrophage populations within their microenvironment and suggest questions for further investigation. PMID:25319326

  15. Macrophage inflammatory protein-3α influences growth of K562 leukemia cells in co-culture with anticancer drug-pretreated HS-5 stromal cells

    International Nuclear Information System (INIS)

    Lee, Y.C.; Chiou, T.-J.; Tzeng, W.-F.; Chu, S.T.

    2008-01-01

    Stromal cell monolayers have been an important means of studying the regulation of hematopoiesis, because they produce cytokines. Cytosine arabinoside, vincristine, daunorubicin, and doxorubicin are common drugs for hematological cancer therapy, and they may have some effects on bone marrow stroma during chemotherapy. The aim of this study was to elucidate interactions between the bone marrow stromal microenvironment and leukemic cells after drug treatment. We tested the hypothesis that human HS-5 stromal cells, pretreated with anticancer drugs, affected the growth of leukemic K562 cells by changing the cytokines in the culture microenvironment. Thereafter, proliferation of K562 cells increased nearly 2.5-fold compared the co-cultivation with drugs-pretreated HS-5 stromal cells and drugs-untreated HS-5 stromal cells. The results indicated that co-cultivation with HS-5 stromal cells pretreated with drugs caused significant K562 cell proliferation. Cytokines in the microenvironment were detected via the RayBio Human Cytokine Antibody Array Membrane. The levels of the cytokines CKβ, IL-12, IL-13, IGFBP-2, MCP-1, MCP-3, MCP-4, MDC, MIP-1β and MIP-1δ were decreased, with a particularly marked decrease in MIP-3α. In co-culture medium, there was a 20-fold decrease in MIP-3α in daunorubicin-pretreated HS-5 cells and at least a 3-fold decrease in Ara-C-pretreated cells. This indicated a significant effect of anticancer drugs on the stromal cell line. Using phosphorylated Erk and pRb proteins as cell proliferation markers, we found that phosphorylation of these markers in K562 cells was inhibited during co-cultivation with drug-pretreated stromal cells in MIP-3α-supplemented medium and restored by MIP-3α antibody supplement. In conclusion, anticancer drug pretreatment suppresses the negative control exerted by HS-5 cells on leukemic cell proliferation, via modulation of cytokines in the microenvironment, especially at the level of MIP-3α

  16. Metabolic activation of volatile organic compounds and polycyclic aromatic hydrocarbons coated onto airborne PM{sub 2.5} in isolated human alveolar macrophages; Etude de l'activation metabolique des composes organiques volatils et des hydrocarbures aromatiques polycycliques d'un aerosol anthropogenique par des macrophages alveolaires humains en culture primaire

    Energy Technology Data Exchange (ETDEWEB)

    Saint-Georges, F.; Mulliez, P. [Hopital Saint Philibert - GHICL-FLM, Service de Pneumologie, 59 - Lomme (France); Saint-Georges, F.; Abbas, I.; Garcon, G.; Billet, S.; Verdin, A.; Shirali, P. [LCE-EA2598, Lab. de Recherche en toxicologie Industrielle et Environnementale - ULCO-MREI, 59 - Dunkerque (France); Gosset, P. [Hopital Saint Vincent, Laboratoire d' Anatomie et de Cytologie Pathologiques - GHICL-FLM, 59 - Lille (France); Courcot, D. [LCE-EA2598, Lab. de Catalyse et Environnement - ULCO-MREI, 59 - Dunkerque (France)

    2009-01-15

    To contribute to improve the knowledge of the underlying mechanisms of action involved in air pollution Particulate Matter (PM)-induced cytotoxicity, we were interested in the metabolic activation of Volatile Organic Compounds (VOC) and/or Polycyclic Aromatic Hydrocarbons (PAH)-coated onto Dunkerque City's PM{sub 2.5} in human Alveolar Macrophages (AM) isolated from Bronchoalveolar Lavage Fluid (BALF). This in vitro cell lung model is very close to the normal in vivo situation, notably in the characteristics that AM display in terms of gene expression of phase I and phase II-metabolizing enzymes. The bronchoscopic examinations and BAL procedures were carried out without any complications. The exposure of AM, during 24, 48 or 72 h, to increasing concentrations of the collected aerosol induced significant variations of the activities of the extracellular lactate dehydrogenase and the mitochondrial dehydrogenase. The lethal concentrations at 10% and 50% were 14.93 and 74.63 {mu}g/mL for AM, respectively, and indicated the relatively higher sensibility of such target lung cells. VOC and/or PAH-coated at low levels onto the surface of the particulate fraction significantly induced gene expression of cytochrome P450 (GYP) 1A1, CYP2E1, NADPH Quinone oxido-reductase (NQO)-1) and Glutathione S-Transferase (GST)P1 and M3, versus controls, suggesting thereby the formation of biologically reactive metabolites. Moreover, these results suggested the role of physical vector of carbonaceous core of PM, which can, therefore, increase both the penetration and the retention of attached-VOC into the cells, thereby enabling them to exert a more durable induction. Hence, we concluded that the metabolic activation of the very low doses of VOC and/or PAH-coated onto Dunkerque City's PM{sub 2.5} is one of the underlying mechanisms of action closely involved in its cytotoxicity in isolated human AM in culture. (author)

  17. Live Imaging of HIV-1 Transfer across T Cell Virological Synapse to Epithelial Cells that Promotes Stromal Macrophage Infection.

    Science.gov (United States)

    Real, Fernando; Sennepin, Alexis; Ganor, Yonatan; Schmitt, Alain; Bomsel, Morgane

    2018-05-08

    During sexual intercourse, HIV-1 crosses epithelial barriers composing the genital mucosa, a poorly understood feature that requires an HIV-1-infected cell vectoring efficient mucosal HIV-1 entry. Therefore, urethral mucosa comprising a polarized epithelium and a stroma composed of fibroblasts and macrophages were reconstructed in vitro. Using this system, we demonstrate by live imaging that efficient HIV-1 transmission to stromal macrophages depends on cell-mediated transfer of the virus through virological synapses formed between HIV-1-infected CD4 + T cells and the epithelial cell mucosal surface. We visualized HIV-1 translocation through mucosal epithelial cells via transcytosis in regions where virological synapses occurred. In turn, interleukin-13 is secreted and HIV-1 targets macrophages, which develop a latent state of infection reversed by lipopolysaccharide (LPS) activation. The live observation of virological synapse formation reported herein is key in the design of vaccines and antiretroviral therapies aimed at blocking HIV-1 access to cellular reservoirs in genital mucosa. Copyright © 2018. Published by Elsevier Inc.

  18. Cell Elasticity Determines Macrophage Function

    Science.gov (United States)

    Patel, Naimish R.; Bole, Medhavi; Chen, Cheng; Hardin, Charles C.; Kho, Alvin T.; Mih, Justin; Deng, Linhong; Butler, James; Tschumperlin, Daniel; Fredberg, Jeffrey J.; Krishnan, Ramaswamy; Koziel, Henry

    2012-01-01

    Macrophages serve to maintain organ homeostasis in response to challenges from injury, inflammation, malignancy, particulate exposure, or infection. Until now, receptor ligation has been understood as being the central mechanism that regulates macrophage function. Using macrophages of different origins and species, we report that macrophage elasticity is a major determinant of innate macrophage function. Macrophage elasticity is modulated not only by classical biologic activators such as LPS and IFN-γ, but to an equal extent by substrate rigidity and substrate stretch. Macrophage elasticity is dependent upon actin polymerization and small rhoGTPase activation, but functional effects of elasticity are not predicted by examination of gene expression profiles alone. Taken together, these data demonstrate an unanticipated role for cell elasticity as a common pathway by which mechanical and biologic factors determine macrophage function. PMID:23028423

  19. Cell elasticity determines macrophage function.

    Directory of Open Access Journals (Sweden)

    Naimish R Patel

    Full Text Available Macrophages serve to maintain organ homeostasis in response to challenges from injury, inflammation, malignancy, particulate exposure, or infection. Until now, receptor ligation has been understood as being the central mechanism that regulates macrophage function. Using macrophages of different origins and species, we report that macrophage elasticity is a major determinant of innate macrophage function. Macrophage elasticity is modulated not only by classical biologic activators such as LPS and IFN-γ, but to an equal extent by substrate rigidity and substrate stretch. Macrophage elasticity is dependent upon actin polymerization and small rhoGTPase activation, but functional effects of elasticity are not predicted by examination of gene expression profiles alone. Taken together, these data demonstrate an unanticipated role for cell elasticity as a common pathway by which mechanical and biologic factors determine macrophage function.

  20. Proliferating macrophages prevail in atherosclerosis.

    Science.gov (United States)

    Randolph, Gwendalyn J

    2013-09-01

    Macrophages accumulate in atherosclerotic lesions during the inflammation that is part of atherosclerosis development and progression. A new study in mice indicates that the accumulation of macrophages in atherosclerotic plaques depends on local macrophage proliferation rather than the recruitment of circulating monocytes.

  1. Macrophage mitochondrial oxidative stress promotes atherosclerosis and nuclear factor-κB-mediated inflammation in macrophages.

    Science.gov (United States)

    Wang, Ying; Wang, Gary Z; Rabinovitch, Peter S; Tabas, Ira

    2014-01-31

    Mitochondrial oxidative stress (mitoOS) has been shown to correlate with the progression of human atherosclerosis. However, definitive cell type-specific causation studies in vivo are lacking, and the molecular mechanisms of potential proatherogenic effects remain to be determined. Our aims were to assess the importance of macrophage mitoOS in atherogenesis and to explore the underlying molecular mechanisms. We first validated Western diet-fed Ldlr(-/-) mice as a model of human mitoOS-atherosclerosis association by showing that non-nuclear oxidative DNA damage, a marker of mitoOS in lesional macrophages, correlates with aortic root lesion development. To investigate the importance of macrophage mitoOS, we used a genetic engineering strategy in which the OS suppressor catalase was ectopically expressed in mitochondria (mCAT) in macrophages. MitoOS in lesional macrophages was successfully suppressed in these mice, and this led to a significant reduction in aortic root lesional area. The mCAT lesions had less monocyte-derived cells, less Ly6c(hi) monocyte infiltration into lesions, and lower levels of monocyte chemotactic protein-1. The decrease in lesional monocyte chemotactic protein-1 was associated with the suppression of other markers of inflammation and with decreased phosphorylation of RelA (NF-κB p65), indicating decreased activation of the proinflammatory NF-κB pathway. Using models of mitoOS in cultured macrophages, we showed that mCAT suppressed monocyte chemotactic protein-1 expression by decreasing the activation of the IκB-kinase β-RelA NF-κB pathway. MitoOS in lesional macrophages amplifies atherosclerotic lesion development by promoting NF-κB-mediated entry of monocytes and other inflammatory processes. In view of the mitoOS-atherosclerosis link in human atheromata, these findings reveal a potentially new therapeutic target to prevent the progression of atherosclerosis.

  2. Differential effects of Th1, monocyte/macrophage and Th2 cytokine mixtures on early gene expression for molecules associated with metabolism, signaling and regulation in central nervous system mixed glial cell cultures

    Directory of Open Access Journals (Sweden)

    Studzinski Diane

    2009-01-01

    Full Text Available Abstract Background Cytokines secreted by immune cells and activated glia play central roles in both the pathogenesis of and protection from damage to the central nervous system (CNS in multiple sclerosis (MS. Methods We have used gene array analysis to identify the initial direct effects of cytokines on CNS glia by comparing changes in early gene expression in CNS glial cultures treated for 6 hours with cytokines typical of those secreted by Th1 and Th2 lymphocytes and monocyte/macrophages (M/M. Results In two previous papers, we summarized effects of these cytokines on immune-related molecules, and on neural and glial related proteins, including neurotrophins, growth factors and structural proteins. In this paper, we present the effects of the cytokines on molecules involved in metabolism, signaling and regulatory mechanisms in CNS glia. Many of the changes in gene expression were similar to those seen in ischemic preconditioning and in early inflammatory lesions in experimental autoimmune encephalomyelitis (EAE, related to ion homeostasis, mitochondrial function, neurotransmission, vitamin D metabolism and a variety of transcription factors and signaling pathways. Among the most prominent changes, all three cytokine mixtures markedly downregulated the dopamine D3 receptor, while Th1 and Th2 cytokines downregulated neuropeptide Y receptor 5. An unexpected finding was the large number of changes related to lipid metabolism, including several suggesting a switch from diacylglycerol to phosphatidyl inositol mediated signaling pathways. Using QRT-PCR we validated the results for regulation of genes for iNOS, arginase and P glycoprotein/multi-drug resistance protein 1 (MDR1 seen at 6 hours with microarray. Conclusion Each of the three cytokine mixtures differentially regulated gene expression related to metabolism and signaling that may play roles in the pathogenesis of MS, most notably with regard to mitochondrial function and neurotransmitter

  3. Nanopatterned bulk metallic glass-based biomaterials modulate macrophage polarization.

    Science.gov (United States)

    Shayan, Mahdis; Padmanabhan, Jagannath; Morris, Aaron H; Cheung, Bettina; Smith, Ryan; Schroers, Jan; Kyriakides, Themis R

    2018-06-01

    Polarization of macrophages by chemical, topographical and mechanical cues presents a robust strategy for designing immunomodulatory biomaterials. Here, we studied the ability of nanopatterned bulk metallic glasses (BMGs), a new class of metallic biomaterials, to modulate murine macrophage polarization. Cytokine/chemokine analysis of IL-4 or IFNγ/LPS-stimulated macrophages showed that the secretion of TNF-α, IL-1α, IL-12, CCL-2 and CXCL1 was significantly reduced after 24-hour culture on BMGs with 55 nm nanorod arrays (BMG-55). Additionally, under these conditions, macrophages increased phagocytic potential and exhibited decreased cell area with multiple actin protrusions. These in vitro findings suggest that nanopatterning can modulate biochemical cues such as IFNγ/LPS. In vivo evaluation of the subcutaneous host response at 2 weeks demonstrated that the ratio of Arg-1 to iNOS increased in macrophages adjacent to BMG-55 implants, suggesting modulation of polarization. In addition, macrophage fusion and fibrous capsule thickness decreased and the number and size of blood vessels increased, which is consistent with changes in macrophage responses. Our study demonstrates that nanopatterning of BMG implants is a promising technique to selectively polarize macrophages to modulate the immune response, and also presents an effective tool to study mechanisms of macrophage polarization and function. Implanted biomaterials elicit a complex series of tissue and cellular responses, termed the foreign body response (FBR), that can be influenced by the polarization state of macrophages. Surface topography can influence polarization, which is broadly characterized as either inflammatory or repair-like. The latter has been linked to improved outcomes of the FBR. However, the impact of topography on macrophage polarization is not fully understood, in part, due to a lack of high moduli biomaterials that can be reproducibly processed at the nanoscale. Here, we studied

  4. [Macrophage colony stimulating factor enhances non-small cell lung cancer invasion and metastasis by promoting macrophage M2 polarization].

    Science.gov (United States)

    Li, Y J; Yang, L; Wang, L P; Zhang, Y

    2017-06-23

    Objective: To investigate the key cytokine which polarizes M2 macrophages and promotes invasion and metastasis in non-small cell lung cancer (NSCLC). Methods: After co-culture with A549 cells in vitro, the proportion of CD14(+) CD163(+) M2 macrophages in monocytes and macrophage colony stimulating factor (M-CSF) levels in culture supernatant were detected by flow cytometry, ELISA assay and real-time qPCR, respectively. The effects of CD14(+) CD163(+) M2 macrophages on invasion of A549 cells and angiogenesis of HUVEC cells were measured by transwell assay and tubule formation assay, respectively. The clinical and prognostic significance of M-CSF expression in NSCLC was further analyzed. Results: The percentage of CD14(+) CD163(+) M2 macrophages in monocytes and the concentration of M-CSF in the supernatant followed by co-culture was (12.03±0.46)% and (299.80±73.76)pg/ml, respectively, which were significantly higher than those in control group [(2.80±1.04)% and (43.07±11.22)pg/ml, respectively, P macrophages in vitro . M2 macrophages enhanced the invasion of A549 cells (66 cells/field vs. 26 cells/field) and the angiogenesis of HUVEC cells (22 tubes/field vs. 8 tubes/field). The mRNA expression of M-CSF in stage Ⅰ-Ⅱ patients (16.23±4.83) was significantly lower than that in stage Ⅲ-Ⅳ (53.84±16.08; P macrophages, which can further promote the metastasis and angiogenesis of NSCLC. M-CSF could be used as a potential therapeutic target of NSCLC.

  5. Mitogen-activated protein kinase phosphatase-1 expression in macrophages is controlled by lymphocytes during macrophage activation.

    Science.gov (United States)

    Luo, Chong; Yang, Xiqiang; Yao, Lan; Jiang, Liping; Liu, Wei; Li, Xin; Wang, Lijia

    2012-01-01

    The viewpoints on the control of innate immune cells by the adaptive immune system during sepsis remain controversial. Mitogen-activated protein kinase phosphatase-1 (MKP-1) is essential to the negative control of innate immunity and suppresses the activation of macrophages by inhibiting activated mitogen-activated protein kinase (MAPK). The purpose of the current study was to observe inflammatory response and macrophage activation in mice with severe combined immunodeficiency (SCID) with endotoxemia and to determine the role of MKP-1 in the control of macrophage activation by the adaptive immune system. Endotoxemia was induced in wild-type and SCID mice by an intraperitoneal injection of lipopolysaccharide (LPS), and all of the SCID mice died. SCID mice produced more inflammatory cytokines than BALB/c mice systemically and locally. TNF-α mRNA expression was higher and MKP-1 mRNA expression was lower in peritoneal macrophages (PMa) from SCID mice compared to PMa from wild-type mice after and even before LPS injection. Thioglycollate-stimulated PMa from wild-type mice were stimulated with LPS in vitro in the presence or absence of pan-T cells. The levels of TNF-α and IL-6 were higher in the supernatants from PMa cultured alone compared to PMa co-cultured with pan-T cells, and PMa MKP-1 mRNA and protein expression were higher when PMa were co-cultured with pan-T cells. Therefore, pan-T cells can up-regulate MKP-1 expression in macrophages and inhibit the secretion of inflammatory cytokines secretion by macrophages. In SCID mice, lymphocyte deficiency, especially T cell deficiency, causes insufficient MKP-1 expression in macrophages, which can be responsible for the severe inflammation and bad prognosis of septic SCID mice. MKP-1 plays an important role in the control of macrophage activation by the adaptive immune system.

  6. Lineage-specific enhancers activate self-renewal genes in macrophages and embryonic stem cells

    OpenAIRE

    Soucie, E.L.; Weng, Z.; Geirsdottir, L.; Molawi, K.; Maurizio, J.; Fenouil, R.; Mossadegh-Keller, N.; Gimenez, G.; VanHille, L.; Beniazza, M.; Favret, J.; Berruyer, C.; Perrin, P.; Hacohen, N.; Andrau, J.C.

    2016-01-01

    Differentiated macrophages can self-renew in tissues and expand long-term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network controlling self-renewal. Single cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down...

  7. Mechanisms of macrophage accumulation in the lungs of asbestos-exposed subjects

    International Nuclear Information System (INIS)

    Spurzem, J.R.; Saltini, C.; Rom, W.; Winchester, R.J.; Crystal, R.G.

    1987-01-01

    Chronic asbestos exposure is associated with the accumulation of mononuclear phagocytes in the lower respiratory tract. This process can be both protective and injurious, since macrophages can aid in asbestos clearance yet also modulate structural derangements of the alveolar walls. To understand why macrophages accumulate in the lungs of asbestos-exposed persons, 2 possible mechanisms were evaluated using alveolar macrophages from subjects with histories of chronic high exposure to airborne asbestos: enhanced recruitment of blood monocytes to the lung, and an increased rate of replication of macrophages in situ. Monoclonal antibody analysis with antibodies that detect surface antigens on the majority of circulating blood monocytes but only on a minority of mature alveolar macrophages demonstrated that an increased proportion of alveolar macrophages of asbestos workers expressed monocyte lineage antigens, suggesting the presence of young newly recruited macrophages and thus enhanced recruitment. Culture of the alveolar macrophages from these subjects with [ 3 H]thymidine followed by autoradiography demonstrated an increased proportion of alveolar macrophages synthesizing DNA, suggesting the macrophages are replicating at an increased rate in situ. These observations are consistent with the concept that both enhanced recruitment of blood monocytes and increased local proliferation of alveolar macrophages contribute to the accumulation mononuclear phagocytes in the lung of persons with chronic asbestos exposure

  8. Induced-Pluripotent-Stem-Cell-Derived Primitive Macrophages Provide a Platform for Modeling Tissue-Resident Macrophage Differentiation and Function.

    Science.gov (United States)

    Takata, Kazuyuki; Kozaki, Tatsuya; Lee, Christopher Zhe Wei; Thion, Morgane Sonia; Otsuka, Masayuki; Lim, Shawn; Utami, Kagistia Hana; Fidan, Kerem; Park, Dong Shin; Malleret, Benoit; Chakarov, Svetoslav; See, Peter; Low, Donovan; Low, Gillian; Garcia-Miralles, Marta; Zeng, Ruizhu; Zhang, Jinqiu; Goh, Chi Ching; Gul, Ahmet; Hubert, Sandra; Lee, Bernett; Chen, Jinmiao; Low, Ivy; Shadan, Nurhidaya Binte; Lum, Josephine; Wei, Tay Seok; Mok, Esther; Kawanishi, Shohei; Kitamura, Yoshihisa; Larbi, Anis; Poidinger, Michael; Renia, Laurent; Ng, Lai Guan; Wolf, Yochai; Jung, Steffen; Önder, Tamer; Newell, Evan; Huber, Tara; Ashihara, Eishi; Garel, Sonia; Pouladi, Mahmoud A; Ginhoux, Florent

    2017-07-18

    Tissue macrophages arise during embryogenesis from yolk-sac (YS) progenitors that give rise to primitive YS macrophages. Until recently, it has been impossible to isolate or derive sufficient numbers of YS-derived macrophages for further study, but data now suggest that induced pluripotent stem cells (iPSCs) can be driven to undergo a process reminiscent of YS-hematopoiesis in vitro. We asked whether iPSC-derived primitive macrophages (iMacs) can terminally differentiate into specialized macrophages with the help of growth factors and organ-specific cues. Co-culturing human or murine iMacs with iPSC-derived neurons promoted differentiation into microglia-like cells in vitro. Furthermore, murine iMacs differentiated in vivo into microglia after injection into the brain and into functional alveolar macrophages after engraftment in the lung. Finally, iPSCs from a patient with familial Mediterranean fever differentiated into iMacs with pro-inflammatory characteristics, mimicking the disease phenotype. Altogether, iMacs constitute a source of tissue-resident macrophage precursors that can be used for biological, pathophysiological, and therapeutic studies. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Rapid, radiolabeled-microculture method that uses macrophages for in vitro evaluation of Mycobacterium leprae viability and drug susceptibility.

    Science.gov (United States)

    Mittal, A; Sathish, M; Seshadri, P S; Nath, I

    1983-04-01

    This paper describes a microculture rapid assay using radiolabeling and mouse macrophages to determine the viability and the drug susceptibility or resistance of Mycobacterium leprae. Comparison of M. leprae resident macrophage cultures maintained in 96-well flat-bottomed plates showed results for viability and susceptibility or resistance to dapsone that were similar to results for concurrent cultures in Leighton tubes with greater numbers of bacilli and macrophages.

  10. Rapid, Radiolabeled-Microculture Method That Uses Macrophages for In Vitro Evaluation of Mycobacterium leprae Viability and Drug Susceptibility

    OpenAIRE

    Mittal, A.; Sathish, M.; Seshadri, P. S.; Nath, Indira

    1983-01-01

    This paper describes a microculture rapid assay using radiolabeling and mouse macrophages to determine the viability and the drug susceptibility or resistance of Mycobacterium leprae. Comparison of M. leprae resident macrophage cultures maintained in 96-well flat-bottomed plates showed results for viability and susceptibility or resistance to dapsone that were similar to results for concurrent cultures in Leighton tubes with greater numbers of bacilli and macrophages.

  11. The Effect of Biomaterials Used for Tissue Regeneration Purposes on Polarization of Macrophages

    NARCIS (Netherlands)

    G.S.A. ter Hoeve-Boersema (Simone); N. Grotenhuis (Nienke); Y. Bayon (Yves); J.F. Lange (Johan); Y.M. Bastiaansen-Jenniskens (Yvonne)

    2016-01-01

    textabstractActivation of macrophages is critical in the acute phase of wound healing after implantation of surgical biomaterials. To understand the response of macrophages, they are often cultured in vitro on biomaterials. Since a wide range of biomaterials is currently used in the clinics, we

  12. Granulocyte-macrophage colony-stimulating factor primes interleukin-13 production by macrophages via protease-activated receptor-2.

    Science.gov (United States)

    Aoki, Manabu; Yamaguchi, Rui; Yamamoto, Takatoshi; Ishimaru, Yasuji; Ono, Tomomichi; Sakamoto, Arisa; Narahara, Shinji; Sugiuchi, Hiroyuki; Hirose, Eiji; Yamaguchi, Yasuo

    2015-04-01

    Chronic inflammation is often linked to the presence of type 2-polarized macrophages, which are induced by the T helper type 2 cytokines interleukin-4 and interleukin-13 (IL-13). IL-13 is a key mediator of tissue fibrosis caused by T helper type 2-based inflammation. Human neutrophil elastase (HNE) plays a pivotal role in the pathogenesis of pulmonary fibrosis. This study investigated the priming effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on IL-13 expression by macrophages stimulated with HNE. Adherent macrophages were obtained from primary cultures of human mononuclear cells. Expression of IL-13 mRNA and protein by GM-CSF-dependent macrophages was investigated after stimulation with HNE, using the polymerase chain reaction and enzyme-linked immunosorbent assay. GM-CSF had a priming effect on IL-13 mRNA and protein expression by macrophages stimulated with HNE, while this effect was not observed for various other cytokines. GM-CSF-dependent macrophages showed a significant increase in the expression of protease activated receptor-2 (PAR-2) mRNA and protein. The response of IL-13 mRNA to HNE was significantly decreased by pretreatment with alpha1-antitrypsin, a PAR-2 antibody (SAM11), or a PAR-2 antagonist (ENMD-1068). These findings suggest that stimulation with HNE can induce IL-13 production by macrophages, especially GM-CSF-dependent macrophages. Accordingly, neutrophil elastase may have a key role in fibrosis associated with chronic inflammation. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Fish oil-derived long-chain n-3 polyunsaturated fatty acids reduce expression of M1-associated macrophage markers in an ex vivo adipose tissue culture model, in part through adiponectin

    OpenAIRE

    Anna A. De Boer; Jennifer M. Monk; Jennifer M. Monk; Danyelle M. Liddle; Krista A. Power; David W.L. Ma; Lindsay E. Robinson

    2015-01-01

    Adipose tissue (AT) macrophages (ATM) play a key role in obesity-associated pathologies, and their phenotype can be influenced by the local tissue microenvironment. Interestingly, long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFA) and the LC n-3 PUFA-upregulated adipokine, adiponectin (Ad), may mitigate excessive ATM inflammatory M1-polarization responses. However, to what extent LC n-3 PUFA and Ad work in concert to affect macrophage phenotype has not been examined. Thus, we used an es...

  14. Macrophage immunoregulatory pathways in tuberculosis.

    Science.gov (United States)

    Rajaram, Murugesan V S; Ni, Bin; Dodd, Claire E; Schlesinger, Larry S

    2014-12-01

    Macrophages, the major host cells harboring Mycobacterium tuberculosis (M.tb), are a heterogeneous cell type depending on their tissue of origin and host they are derived from. Significant discord in macrophage responses to M.tb exists due to differences in M.tb strains and the various types of macrophages used to study tuberculosis (TB). This review will summarize current concepts regarding macrophage responses to M.tb infection, while pointing out relevant differences in experimental outcomes due to the use of divergent model systems. A brief description of the lung environment is included since there is increasing evidence that the alveolar macrophage (AM) has immunoregulatory properties that can delay optimal protective host immune responses. In this context, this review focuses on selected macrophage immunoregulatory pattern recognition receptors (PRRs), cytokines, negative regulators of inflammation, lipid mediators and microRNAs (miRNAs). Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Conditioned medium from alternatively activated macrophages induce mesangial cell apoptosis via the effect of Fas

    International Nuclear Information System (INIS)

    Huang, Yuan; Luo, Fangjun; Li, Hui; Jiang, Tao; Zhang, Nong

    2013-01-01

    During inflammation in the glomerulus, the proliferation of myofiroblast-like mesangial cells is commonly associated with the pathological process. Macrophages play an important role in regulating the growth of resident mesangial cells in the glomeruli. Alternatively activated macrophage (M2 macrophage) is a subset of macrophages induced by IL-13/IL-4, which is shown to play a repair role in glomerulonephritis. Prompted by studies of development, we performed bone marrow derived macrophage and rat mesangial cell co-culture study. Conditioned medium from IL-4 primed M2 macrophages induced rat mesangial cell apoptosis. The pro-apoptotic effect of M2 macrophages was demonstrated by condensed nuclei stained with Hoechst 33258, increased apoptosis rates by flow cytometry analysis and enhanced caspase-3 activation by western blot. Fas protein was up-regulated in rat mesangial cells, and its neutralizing antibody ZB4 partly inhibited M2 macrophage-induced apoptosis. The up-regulated arginase-1 expression in M2 macrophage also contributed to this apoptotic effect. These results indicated that the process of apoptosis triggered by conditioned medium from M2 macrophages, at least is partly conducted through Fas in rat mesangial cells. Our findings provide compelling evidence that M2 macrophages control the growth of mesangial cells in renal inflammatory conditions. - Highlights: • Conditioned-medium from M2 macrophages induces rat mesangial cell (MsC) apoptosis. • M2 macrophage conditioned medium exerts its pro-apoptotic effects via Fas ligand. • Arginase-1 activity in M2 macrophages plays a role in inducing apoptosis in rat MsC

  16. Conditioned medium from alternatively activated macrophages induce mesangial cell apoptosis via the effect of Fas

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Yuan; Luo, Fangjun; Li, Hui; Jiang, Tao; Zhang, Nong, E-mail: nzhang@fudan.edu.cn

    2013-11-15

    During inflammation in the glomerulus, the proliferation of myofiroblast-like mesangial cells is commonly associated with the pathological process. Macrophages play an important role in regulating the growth of resident mesangial cells in the glomeruli. Alternatively activated macrophage (M2 macrophage) is a subset of macrophages induced by IL-13/IL-4, which is shown to play a repair role in glomerulonephritis. Prompted by studies of development, we performed bone marrow derived macrophage and rat mesangial cell co-culture study. Conditioned medium from IL-4 primed M2 macrophages induced rat mesangial cell apoptosis. The pro-apoptotic effect of M2 macrophages was demonstrated by condensed nuclei stained with Hoechst 33258, increased apoptosis rates by flow cytometry analysis and enhanced caspase-3 activation by western blot. Fas protein was up-regulated in rat mesangial cells, and its neutralizing antibody ZB4 partly inhibited M2 macrophage-induced apoptosis. The up-regulated arginase-1 expression in M2 macrophage also contributed to this apoptotic effect. These results indicated that the process of apoptosis triggered by conditioned medium from M2 macrophages, at least is partly conducted through Fas in rat mesangial cells. Our findings provide compelling evidence that M2 macrophages control the growth of mesangial cells in renal inflammatory conditions. - Highlights: • Conditioned-medium from M2 macrophages induces rat mesangial cell (MsC) apoptosis. • M2 macrophage conditioned medium exerts its pro-apoptotic effects via Fas ligand. • Arginase-1 activity in M2 macrophages plays a role in inducing apoptosis in rat MsC.

  17. Biology of Bony Fish Macrophages

    OpenAIRE

    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 ...

  18. Targeting androgen receptor to suppress macrophage-induced EMT and benign prostatic hyperplasia (BPH) development.

    Science.gov (United States)

    Lu, Tianjing; Lin, Wen-Jye; Izumi, Kouji; Wang, Xiaohai; Xu, Defeng; Fang, Lei-Ya; Li, Lei; Jiang, Qi; Jin, Jie; Chang, Chawnshang

    2012-10-01

    Early studies suggested macrophages might play roles in inflammation-associated benign prostatic hyperplasia (BPH) development, yet the underlying mechanisms remain unclear. Here we first showed that CD68(+) macrophages were identified in both epithelium and the stromal area of human BPH tissues. We then established an in vitro co-culture model with prostate epithelial and macrophage cell lines to study the potential impacts of infiltrating macrophages in the BPH development and found that co-culturing prostate epithelial cells with macrophages promoted migration of macrophages. In a three-dimensional culture system, the sphere diameter of BPH-1 prostate cells was significantly increased during coculture with THP-1 macrophage cells. Mechanism dissection suggested that expression levels of epithelial-mesenchymal transition (EMT) markers, such as N-cadherin, Snail, and TGF-β2, were increased, and administration of anti-TGF-β2 neutralizing antibody during co-culture suppressed the EMT and THP-1-mediated growth of BPH-1 cells, suggesting THP-1 might go through EMT to influence the BPH development and progression. Importantly, we found that modulation of androgen receptor (AR) in BPH-1 and mPrE cells significantly increased THP-1 and RAW264.7 cell migration, respectively, and enhanced expression levels of EMT markers, suggesting that AR in prostate epithelial cells might play a role in promoting macrophage-mediated EMT in prostate epithelial cells. Silencing AR function via an AR degradation enhancer, ASC-J9, decreased the macrophage migration to BPH-1 cells and suppressed EMT marker expression. Together, these results provide the first evidence to demonstrate that prostate epithelial AR function is important for macrophage-mediated EMT and proliferation of prostate epithelial cells, which represents a previously unrecognized role of AR in the cross-talk between macrophages and prostate epithelial cells. These results may provide new insights for a new therapeutic

  19. Bioelectric modulation of macrophage polarization

    Science.gov (United States)

    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.

  20. Anti-inflammatory effect of garlic 14-kDa protein on LPS-stimulated-J774A.1 macrophages.

    Science.gov (United States)

    Rabe, Shahrzad Zamani Taghizadeh; Ghazanfari, Tooba; Siadat, Zahra; Rastin, Maryam; Rabe, Shahin Zamani Taghizadeh; Mahmoudi, Mahmoud

    2015-04-01

    Garlic 14-kDa protein is purified from garlic (Allium sativum L.) which is used in traditional medicine and exerts various immunomodulatory activities. The present study investigated the suppressive effect of garlic 14-kDa protein on LPS-induced expression of pro-inflammatory mediators and underlying mechanism in inflammatory macrophages. J774A.1 macrophages were treated with 14-kDa protein (5-30 μg/ml) with/without LPS (1 μg/ml) and the production of inflammatory mediators such as prostaglandin E2 (PGE2), TNF-α, and IL-1β released were measured using ELISA. Nitric oxide (NO) production was determined using the Griess method. The anti-inflammatory activity of 14-kDa protein was examined by measuring inducible nitric oxide synthase and cyclooxygenase-2 proteins using western blot. The expression of nuclear NF-κB p65 subunit was assessed by western blot. Garlic 14-kDa protein significantly inhibited the excessive production of NO, PGE, TNF-α, and IL-1β in lipopolysaccharide (LPS)-activated J774A.1 macrophages in a concentration-related manner without cytotoxic effect. Western blot analysis demonstrated that garlic 14-kDa protein suppressed corresponding inducible NO synthase expression and activated cyclooxygenase-2 protein expression. The inhibitory effect was mediated partly by a reduction in the activity and expression of transcription factor NF-κB protein. Our results suggested, for the first time, garlic 14-kDa protein exhibits anti-inflammatory properties in macrophages possibly by suppressing the inflammatory mediators via the inhibition of transcription factor NF-κB signaling pathway. The traditional use of garlic as anti-inflammatory remedy could be ascribed partly to 14-kDa protein content. This protein might be a useful candidate for controlling inflammatory diseases and further investigations in vivo.

  1. Inhibition of herpes simplex virus multiplication by activated macrophages: a role for arginase?

    Science.gov (United States)

    Wildy, P; Gell, P G; Rhodes, J; Newton, A

    1982-07-01

    Proteose-peptone-activated mouse macrophages can prevent productive infection by herpes simplex virus in neighboring cells in vitro whether or not those cells belong to the same animal species. The effect does not require contact between the macrophages and the infected cells, may be prevented by adding extra arginine to the medium, and may be reversed when extra arginine is added 24 h after the macrophages. Arginase activity was found both intracellularly and released from the macrophages. The extracellular enzyme is quite stable; 64% activity was found after 48 h of incubation at 37 degrees C in tissue culture medium. No evidence was found that the inefficiency of virus replication in macrophages was due to self-starvation by arginase. As might be predicted macrophages can, by the same mechanism, limit productive infection by vaccinia virus.

  2. Human macrophage hemoglobin-iron metabolism in vitro

    International Nuclear Information System (INIS)

    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 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 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 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

  3. cultural

    Directory of Open Access Journals (Sweden)

    Irene Kreutz

    2006-01-01

    Full Text Available Es un estudio cualitativo que adoptó como referencial teorico-motodológico la antropología y la etnografía. Presenta las experiencias vivenciadas por mujeres de una comunidad en el proceso salud-enfermedad, con el objetivo de comprender los determinantes sócio-culturales e históricos de las prácticas de prevención y tratamiento adoptados por el grupo cultural por medio de la entrevista semi-estructurada. Los temas que emergieron fueron: la relación entre la alimentación y lo proceso salud-enfermedad, las relaciones con el sistema de salud oficial y el proceso salud-enfermedad y lo sobrenatural. Los dados revelaron que los moradores de la comunidad investigada tienen un modo particular de explicar sus procedimientos terapéuticos. Consideramos que es papel de los profesionales de la salud en sus prácticas, la adopción de abordajes o enfoques que consideren al individuo en su dimensión sócio-cultural e histórica, considerando la enorme diversidad cultural en nuestro país.

  4. Infectious bronchitis corona virus establishes productive infection in avian macrophages interfering with selected antimicrobial functions.

    Directory of Open Access Journals (Sweden)

    Aruna Amarasinghe

    Full Text Available Infectious bronchitis virus (IBV causes respiratory disease leading to loss of egg and meat production in chickens. Although it is known that macrophage numbers are elevated in the respiratory tract of IBV infected chickens, the role played by macrophages in IBV infection, particularly as a target cell for viral replication, is unknown. In this study, first, we investigated the ability of IBV to establish productive replication in macrophages in lungs and trachea in vivo and in macrophage cell cultures in vitro using two pathogenic IBV strains. Using a double immunofluorescent technique, we observed that both IBV Massachusetts-type 41 (M41 and Connecticut A5968 (Conn A5968 strains replicate in avian macrophages at a low level in vivo. This in vivo observation was substantiated by demonstrating IBV antigens in macrophages following in vitro IBV infection. Further, IBV productive infection in macrophages was confirmed by demonstrating corona viral particles in macrophages and IBV ribonucleic acid (RNA in culture supernatants. Evaluation of the functions of macrophages following infection of macrophages with IBV M41 and Conn A5968 strains revealed that the production of antimicrobial molecule, nitric oxide (NO is inhibited. It was also noted that replication of IBV M41 and Conn A5968 strains in macrophages does not interfere with the induction of type 1 IFN activity by macrophages. In conclusion, both M41 and Con A5968 IBV strains infect macrophages in vivo and in vitro resulting productive replications. During the replication of IBV in macrophages, their ability to produce NO can be affected without affecting the ability to induce type 1 IFN activity. Further studies are warranted to uncover the significance of macrophage infection of IBV in the pathogenesis of IBV infection in chickens.

  5. M1 and M2 macrophages derived from THP-1 cells differentially modulate the response of cancer cells to etoposide

    International Nuclear Information System (INIS)

    Genin, Marie; Clement, Francois; Fattaccioli, Antoine; Raes, Martine; Michiels, Carine

    2015-01-01

    Tumor associated macrophages (TAMs) are present in high density in solid tumors. TAMs share many characteristics with alternatively activated macrophages, also called M2. They have been shown to favor tumor development and a role in chemoresistance has also been suggested. Here, we investigated the effects of M2 in comparison to M1 macrophages on cancer cell sensitivity to etoposide. We set up a model of macrophage polarization, starting from THP-1 monocytes differentiated into macrophages using PMA (Phorbol 12-myristate 13-acetate). Once differentiated (M0 macrophages), they were incubated with IL-4 and IL-13 in order to obtain M2 polarized macrophages or with IFN-gamma and LPS for classical macrophage activation (M1). To mimic the communication between cancer cells and TAMs, M0, M1 or M2 macrophages and HepG2 or A549 cancer cells were co-cultured during respectively 16 (HepG2) or 24 (A549) hours, before etoposide exposure for 24 (HepG2) or 16 (A549) hours. After the incubation, the impact of etoposide on macrophage polarization was studied and cancer cell apoptosis was assessed by western-blot for cleaved caspase-3 and cleaved PARP-1 protein, caspase activity assay and FACS analysis of Annexin V and PI staining. mRNA and protein expression of M1 and M2 markers confirmed the polarization of THP-1-derived macrophages, which provide a new, easy and well-characterized model of polarized human macrophages. Etoposide-induced cancer cell apoptosis was markedly reduced in the presence of THP-1 M2 macrophages, while apoptosis was increased in cells co-cultured with M1 macrophages. On the other hand, etoposide did not influence M1 or M2 polarization. These results evidence for the first time a clear protective effect of M2 on the contrary to M1 macrophages on etoposide-induced cancer cell apoptosis

  6. Yersinia pestis intracellular parasitism of macrophages from hosts exhibiting high and low severity of plague.

    Directory of Open Access Journals (Sweden)

    Duraisamy Ponnusamy

    Full Text Available BACKGROUND: Yersinia pestis causes severe disease in natural rodent hosts, but mild to inapparent disease in certain rodent predators such as dogs. Y. pestis initiates infection in susceptible hosts by parasitizing and multiplying intracellularly in local macrophages prior to systemic dissemination. Thus, we hypothesize that Y. pestis disease severity may depend on the degree to which intracellular Y. pestis overcomes the initial host macrophage imposed stress. METHODOLOGY/PRINCIPAL FINDINGS: To test this hypothesis, the progression of in vitro infection by Y. pestis KIM62053.1+ of mouse splenic and RAW264.7 tissue culture macrophages and dog peripheral blood-derived and DH82 tissue culture macrophages was studied using microscopy and various parameters of infection. The study showed that during the early stage of infection, intracellular Y. pestis assumed filamentous cellular morphology with multiple copies of the genome per bacterium in both mouse and dog macrophages. Later, in mouse macrophages, the infection elicited spacious vacuolar extension of Yersinia containing vacuoles (YCV, and the filamentous Y. pestis reverted to coccobacillary morphology with genomic equivalents approximately equaling colony forming units. In contrast, Y. pestis infected dog macrophages did not show noticeable extension of YCV, and intracellular Y. pestis retained the filamentous cellular morphology for the entire experiment in DH82 cells or were killed by blood-derived macrophages. In addition, during the later stage of infection, Y. pestis infected mouse macrophages exhibited cell lysis whereas dog macrophages did not. CONCLUSION/SIGNIFICANCE: Overall, these results support our hypothesis that Y. pestis in mouse macrophages can overcome the initial intracellular stress necessary for subsequent systemic infection. However, in dogs, failure of Y. pestis to overcome macrophage imposed stress may result in mild or in apparent disease in dogs.

  7. Role of macrophages in the immune response to hepatocytes

    International Nuclear Information System (INIS)

    Bumgardner, G.L.; Chen, S.; Almond, S.P.; Ascher, N.L.; Payne, W.D.; Matas, A.J.

    1990-01-01

    The purpose of this study was to determine the role of host macrophages in the development of allospecific cytolytic T cells (allo-CTLs) in response to purified allogeneic MHC Class I+, Class II- hepatocytes in vivo in hepatocyte sponge matrix allografts (HC-SMA). Depletion of antigen-presenting cells (APCs) from responder splenocytes in mixed lymphocyte hepatocyte culture (MLHC) inhibits the development of allo-CTLs in response to purified hepatocytes. First the ability of sponge macrophages to function as accessory cells in indirect presentation of hepatocyte Class I antigen was tested in MLHC. We found that addition of irradiated sponge cells (a source of sponge macrophages) restored the development of allo-CTLs in MLHC depleted of responder APCs. Therefore, radioresistant sponge macrophages can function as accessory cells in MLHC. We next employed silica as an immunotherapy targeted against host macrophages and assessed the effect on development of allo-CTLs in HC-SMA. We found that local (intrasponge) silica treatment completely inhibited the development of allo-CTLs in HC-SMA. Combined local and systemic silica treatment resulted in inhibition of allocytotoxicity comparable to local silica treatment alone in the doses tested. We conclude that host macrophages which infiltrate HC-SMA can function as accessory cells in vitro in MLHC and that both infiltrating host macrophages and lymphocytes participate in the development of an alloimmune response to purified hepatocytes in vivo. This interaction may involve indirect antigen presentation of hepatocyte Class I antigen by macrophages to host lymphocytes which accumulate in HC-SMA

  8. Decreased inducibility of TNF expression in lipid-loaded macrophages

    Directory of Open Access Journals (Sweden)

    Kallin Bengt

    2002-10-01

    Full Text Available Abstract Background Inflammation and immune responses are considered to be very important in the pathogenesis of atherosclerosis. Lipid accumulation in macrophages of the arterial intima is a characteristic feature of atherosclerosis which can influence the inflammatory potential of macrophages. We studied the effects of lipid loading on the regulation of TNF expression in human monocyte-derived macrophages. Results In macrophages incubated with acetylated low density lipoprotein (ac-LDL for 2 days, mRNA expression of TNF in cells stimulated with TNF decreased by 75%. In cell cultures stimulated over night with IL-1β, lipid loading decreased secretion of TNF into culture medium by 48%. These results suggest that lipid accumulation in macrophages makes them less responsive to inflammatory stimuli. Decreased basal activity and inducibility of transcription factor AP-1 was observed in lipid-loaded cells, suggesting a mechanism for the suppression of cytokine expression. NF-κB binding activity and inducibility were only marginally affected by ac-LDL. LDL and ac-LDL did not activate PPARγ. In contrast, oxidized LDL stimulated AP-1 and PPARγ but inhibited NF-κB, indicating that the effects of lipid loading with ac-LDL were not due to oxidation of lipids. Conclusions Accumulation of lipid, mainly cholesterol, results in down-regulation of TNF expression in macrophages. Since monocytes are known to be activated by cell adhesion, these results suggest that foam cells in atherosclerotic plaques may contribute less potently to an inflammatory reaction than newly arrived monocytes/macrophages.

  9. Effect of Triptolide on Functions of Monocytes/ Macrophages in ...

    African Journals Online (AJOL)

    The number of monocytes/macrophages under the varying conditions was subsequently determined by methyl thiazolyl tetrazolium (MTT) assay. The supernatants were collected after 24-h culture, and the content of VEGF and VEGF-C in each supernatant measured by enzyme-linked immunosorbent assay (ELISA).

  10. Siderocalin inhibits the intracellular replication of Mycobacterium tuberculosis in macrophages

    DEFF Research Database (Denmark)

    Johnson, Erin E; Srikanth, Chittur V; Sandgren, Andreas

    2010-01-01

    Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show that sideroc...

  11. Epigenetic regulation of macrophage function

    NARCIS (Netherlands)

    Hoeksema, M.A.

    2016-01-01

    Atherosclerosis is a lipid-driven chronic inflammatory disorder with a key role for macrophages in all disease stages. Macrophages are involved as scavengers of lipids, regulate inflammation, attract other immune cells and contribute to the resolution of inflammation, fibrosis and plaque stability.

  12. Biology of Bony Fish Macrophages

    Directory of Open Access Journals (Sweden)

    Jordan W. Hodgkinson

    2015-11-01

    Full Text Available 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.

  13. Biology of Bony Fish Macrophages.

    Science.gov (United States)

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

    2015-11-30

    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.

  14. Networked T cell death following macrophage infection by Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Stephen H-F Macdonald

    Full Text Available BACKGROUND: Depletion of T cells following infection by Mycobacterium tuberculosis (Mtb impairs disease resolution, and interferes with clinical test performance that relies on cell-mediated immunity. A number of mechanisms contribute to this T cell suppression, such as activation-induced death and trafficking of T cells out of the peripheral circulation and into the diseased lungs. The extent to which Mtb infection of human macrophages affects T cell viability however, is not well characterised. METHODOLOGY/PRINCIPAL FINDINGS: We found that lymphopenia (<1.5 × 10(9 cells/l was prevalent among culture-positive tuberculosis patients, and lymphocyte counts significantly improved post-therapy. We previously reported that Mtb-infected human macrophages resulted in death of infected and uninfected bystander macrophages. In the current study, we sought to examine the influence of infected human alveolar macrophages on T cells. We infected primary human alveolar macrophages (the primary host cell for Mtb or PMA-differentiated THP-1 cells with Mtb H37Ra, then prepared cell-free supernatants. The supernatants of Mtb-infected macrophages caused dose-dependent, caspase-dependent, T cell apoptosis. This toxic effect of infected macrophage secreted factors did not require TNF-α or Fas. The supernatant cytotoxic signal(s were heat-labile and greater than 50 kDa in molecular size. Although ESAT-6 was toxic to T cells, other Mtb-secreted factors tested did not influence T cell viability; nor did macrophage-free Mtb bacilli or broth from Mtb cultures. Furthermore, supernatants from Mycobacterium bovis Bacille de Calmette et Guerin (BCG- infected macrophages also elicited T cell death suggesting that ESAT-6 itself, although cytotoxic, was not the principal mediator of T cell death in our system. CONCLUSIONS: Mtb-Infected macrophages secrete heat-labile factors that are toxic to T cells, and may contribute to the immunosuppression seen in tuberculosis as well as

  15. Macrophages under pressure: the role of macrophage polarization in hypertension.

    Science.gov (United States)

    Harwani, Sailesh C

    2018-01-01

    Hypertension is a multifactorial disease involving the nervous, renal, and cardiovascular systems. Macrophages are the most abundant and ubiquitous immune cells, placing them in a unique position to serve as key mediators between these components. The polarization of macrophages confers vast phenotypic and functional plasticity, allowing them to act as proinflammatory, homeostatic, and anti-inflammatory agents. Key differences between the M1 and M2 phenotypes, the 2 subsets at the extremes of this polarization spectrum, place macrophages at a juncture to mediate many mechanisms involved in the pathogenesis of hypertension. Neuronal and non-neuronal regulation of the immune system, that is, the "neuroimmuno" axis, plays an integral role in the polarization of macrophages. In hypertension, the neuroimmuno axis results in synchronization of macrophage mobilization from immune cell reservoirs and their chemotaxis, via increased expression of chemoattractants, to end organs critical in the development of hypertension. This complicated system is largely coordinated by the dichotomous actions of the autonomic neuronal and non-neuronal activation of cholinergic, adrenergic, and neurohormonal receptors on macrophages, leading to their ability to "switch" between phenotypes at sites of active inflammation. Data from experimental models and human studies are in concordance with each other and support a central role for macrophage polarization in the pathogenesis of hypertension. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Microbial stasis of Leishmania enriettii in activated guinea pig macrophages

    International Nuclear Information System (INIS)

    Groocock, C.M.; Soulsby, E.J.L.

    1980-01-01

    Peritoneal exudate cells (PEC) from Leishmania-sensitized guinea pigs were cultured in vitro in the presence (activated) or absence (non-activated) of leishmanial antigen for 24 or 48 hours. These were then labelled with 51 Cr and challenged with 125 I-labelled promastigotes. The changing relationship between the macrophage and the parasite was monitored by observing changes in the ratio of the cell-associated isotopes. Highly significant differences in the ratio change developed during culture. These differences were a result of the activated cultures showing a higher release of 51 Cr and a lower release of 125 I when compared with the non-activated cells, at 12 hours the percentage release of 125 I from the parasite within the activated macrophage was fourfold less than that released by parasites within non-activated cells (9.2% versus 38.3%) and tenfold less than that released from glutaraldehyde-killed organisms phagocytosed by activated macrophages (91.6%). These studies indicate that stasis rather than killing of leishmaniae occurs in the activated macrophage in vitro. Parallel experiments evaluated by the visual counting of leishmaniae within the macrophages support these data. PEC from tuberculin-sensitized guinea pigs activated in vitro by purified protein derivative showed little or no activity against leishmaniae, indicating a specific requirement for this microbial stasis by activated macrophages. As a corollary of this, peritoneal exudate lymphocytes separated from the same preparations of PEC were shown to be specifically reactive to leishmanial antigen by transformation and incorporation of 3 H-thymidine. (author)

  17. Cyclophilin B attenuates the expression of TNF-α in lipopolysaccharide-stimulated macrophages through the induction of B cell lymphoma-3.

    Science.gov (United States)

    Marcant, Adeline; Denys, Agnès; Melchior, Aurélie; Martinez, Pierre; Deligny, Audrey; Carpentier, Mathieu; Allain, Fabrice

    2012-08-15

    Extracellular cyclophilin A (CyPA) and CyPB have been well described as chemotactic factors for various leukocyte subsets, suggesting their contribution to inflammatory responses. Unlike CyPA, CyPB accumulates in extracellular matrixes, from which it is released by inflammatory proteases. Hence, we hypothesized that it could participate in tissue inflammation by regulating the activity of macrophages. In the current study, we confirmed that CyPB initiated in vitro migration of macrophages, but it did not induce production of proinflammatory cytokines. In contrast, pretreatment of macrophages with CyPB attenuated the expression of inflammatory mediators induced by LPS stimulation. The expression of TNF-α mRNA was strongly reduced after exposure to CyPB, but it was not accompanied by significant modification in LPS-induced activation of MAPK and NF-κB pathways. LPS activation of a reporter gene under the control of TNF-α gene promoter was also markedly decreased in cells treated with CyPB, suggesting a transcriptional mechanism of inhibition. Consistent with this hypothesis, we found that CyPB induced the expression of B cell lymphoma-3 (Bcl-3), which was accompanied by a decrease in the binding of NF-κB p65 to the TNF-α promoter. As expected, interfering with the expression of Bcl-3 restored cell responsiveness to LPS, thus confirming that CyPB acted by inhibiting initiation of TNF-α gene transcription. Finally, we found that CyPA was not efficient in attenuating the production of TNF-α from LPS-stimulated macrophages, which seemed to be due to a modest induction of Bcl-3 expression. Collectively, these findings suggest an unexpected role for CyPB in attenuation of the responses of proinflammatory macrophages.

  18. Ketamine inhibits tumor necrosis factor-α and interleukin-6 gene expressions in lipopolysaccharide-stimulated macrophages through suppression of toll-like receptor 4-mediated c-Jun N-terminal kinase phosphorylation and activator protein-1 activation

    International Nuclear Information System (INIS)

    Wu, G.-J.; Chen, T.-L.; Ueng, Y.-F.; Chen, R.-M.

    2008-01-01

    Our previous study showed that ketamine, an intravenous anesthetic agent, has anti-inflammatory effects. In this study, we further evaluated the effects of ketamine on the regulation of tumor necrosis factor-α (TNF-α) and interlukin-6 (IL-6) gene expressions and its possible signal-transducing mechanisms in lipopolysaccharide (LPS)-activated macrophages. Exposure of macrophages to 1, 10, and 100 μM ketamine, 100 ng/ml LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. A concentration of 1000 μM of ketamine alone or in combined treatment with LPS caused significant cell death. Administration of LPS increased cellular TNF-α and IL-6 protein levels in concentration- and time-dependent manners. Meanwhile, treatment with ketamine concentration- and time-dependently alleviated the enhanced effects. LPS induced TNF-α and IL-6 mRNA syntheses. Administration of ketamine at a therapeutic concentration (100 μM) significantly inhibited LPS-induced TNF-α and IL-6 mRNA expressions. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA into macrophages decreased cellular TLR4 levels. Co-treatment of macrophages with ketamine and TLR4 siRNA decreased the LPS-induced TNF-α and IL-6 productions more than alone administration of TLR4 siRNA. LPS stimulated phosphorylation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos from the cytoplasm to nuclei. However, administration of ketamine significantly decreased LPS-induced activation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos. LPS increased the binding of nuclear extracts to activator protein-1 consensus DNA oligonucleotides. Administration of ketamine significantly ameliorated LPS-induced DNA binding activity of activator protein-1. Therefore, a clinically relevant concentration of ketamine can inhibit TNF-α and IL-6 gene expressions in LPS-activated macrophages. The suppressive mechanisms occur through suppression of TLR4-mediated

  19. Mesenchymal stem cells ameliorate rhabdomyolysis-induced acute kidney injury via the activation of M2 macrophages

    Science.gov (United States)

    2014-01-01

    Introduction The mortality of rhabdomyolysis-induced acute kidney injury (AKI) is still high, as there is no effective therapy. It has been shown that bone marrow-derived mesenchymal stem cells (MSCs) can induce M2 macrophages, which mediate MSC protection in other experimental inflammation-related organ injury. This study was designed to investigate the protective effects of macrophage activation in MSC therapy of rhabdomyolysis-induced AKI. Methods MSCs were injected into glycerol-induced rhabdomyolysis mice. Renal injury was evaluated using the serum creatinine, urea nitrogen, renal pathology and acute tubular necrosis score. The distribution of MSCs was detected using two-photon fluorescence confocal imaging. Immunofluorescence of anti-F4/80 and anti-CD206 was performed to determine macrophages and M2 macrophages in the tissues of the kidney, and M2 macrophage infiltration was also evaluated using western blotting analyses. After depletion of macrophages using clodronate liposomes at the phase of kidney repair, renal injury was re-evaluated. RAW 264.7 macrophages were incubated with lipopolysaccharide and co-cultured with MSCs and subsequently visualised using immunofluorescence staining and flow cytometry analysis. Finally, disparate phenotype macrophages, including normal macrophages (M0), lipopolysaccharide-stimulated macrophages (M1), and MSC-co-cultured macrophages (M2), were infused into mice with AKI, which were pre-treated with liposomal clodronate. Results In vivo infusion of MSCs protected AKI mice from renal function impairment and severe tubular injury, which was accompanied by a time-dependent increase in CD206-positive M2 macrophage infiltration. In addition, depleting macrophages with clodronate delayed restoration of AKI. In vitro, macrophages co-cultured with MSCs acquired an anti-inflammatory M2 phenotype, which was characterised by an increased expression of CD206 and the secretory cytokine interleukin (IL)-10. The concentrations of IL-10, IL

  20. DMPD: Macrophage-stimulating protein and RON receptor tyrosine kinase: potentialregulators of macrophage inflammatory activities. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 12472665 Macrophage-stimulating protein and RON receptor tyrosine kinase: potential...:545-53. (.png) (.svg) (.html) (.csml) Show Macrophage-stimulating protein and RON receptor tyrosine kinase:...le Macrophage-stimulating protein and RON receptor tyrosine kinase: potentialregulators of macrophage inflam

  1. Effect of azithromycin on Prevotella intermedia lipopolysaccharide-induced production of interleukin-6 in murine macrophages.

    Science.gov (United States)

    Choi, Eun-Young; Jin, Ji-Young; Choi, Jeom-Il; Choi, In Soon; Kim, Sung-Jo

    2014-04-15

    Interleukin-6 (IL-6) is a key proinflammatory cytokine which plays a central role in the pathogenesis of periodontal disease. Host modulatory agents targeting at inhibiting IL-6, therefore, appear to be beneficial in slowing the progression of periodontal disease and potentially reducing destructive aspects of the host response. The present study was designed to investigate the effect of the macrolide antibiotic azithromycin on IL-6 generation in murine macrophages treated with lipopolysaccharide (LPS) from Prevotella intermedia, a pathogen implicated in inflammatory periodontal disease, and its mechanisms of action. Azithromycin significantly suppressed IL-6 production as well as its mRNA expression in P. intermedia LPS-activated RAW264.7 cells. LPS-induced activation of JNK and p38 was not affected by azithromycin treatment. Azithromycin failed to prevent P. intermedia LPS from degrading IκB-α. Instead, azithromycin significantly diminished nuclear translocation and DNA binding activity of NF-κB p50 subunit induced with LPS. Azithromycin inhibited P. intermedia LPS-induced STAT1 and STAT3 phosphorylation. In addition, azithromycin up-regulated the mRNA level of SOCS1 in cells treated with LPS. In conclusion, azithromycin significantly attenuated P. intermedia LPS-induced production of IL-6 in murine macrophages via inhibition of NF-κB, STAT1 and STAT3 activation, which is possibly related to the activation of SOCS1 signaling. Further in vivo studies are required to better evaluate the potential of azithromycin in the treatment of periodontal disease. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Synthesis of pro-inflammatory cytokines and adhesion molecules expression by the irradiated human monocyte/macrophage

    International Nuclear Information System (INIS)

    Pons, I.

    1997-09-01

    As lesions induced by ionizing radiations are essentially noticed in organs the functional and structural organisation of which depend on the highly proliferative stem cell pool, the author reports an in-vivo investigation of the effect of a gamma irradiation on the expression and secretion of pro-inflammatory cytokines par human monocytes/macrophages. In order to study the role of the cell environment in the radiation-induced inflammation, the author studied whether a co-stimulation of monocytes/macrophages by gamma irradiation, or the exposure of co-cultures of monocytes/macrophages and lymphocytes, could modulate the regulation of inflammatory cytokines. The author also studied the modulation of the expression of adhesion molecules mainly expressed by the monocyte/macrophage, and the membrane density of the CD14 receptor after irradiation of monocytes/macrophages during 24 hours, and of totally differentiated macrophages after seven days of culture

  3. Exocytosis of macrophage lysosomes leads to digestion of apoptotic adipocytes and foam cell formation[S

    Science.gov (United States)

    Haka, Abigail S.; Barbosa-Lorenzi, Valéria C.; Lee, Hyuek Jong; Falcone, Domenick J.; Hudis, Clifford A.; Dannenberg, Andrew J.

    2016-01-01

    Many types of apoptotic cells are phagocytosed and digested by macrophages. Adipocytes can be hundreds of times larger than macrophages, so they are too large to be digested by conventional phagocytic processes. The nature of the interaction between macrophages and apoptotic adipocytes has not been studied in detail. We describe a cellular process, termed exophagy, that is important for macrophage clearance of dead adipocytes and adipose tissue homeostasis. Using mouse models of obesity, human tissue, and a cell culture model, we show that macrophages form hydrolytic extracellular compartments at points of contact with dead adipocytes using local actin polymerization. These compartments are acidic and contain lysosomal enzymes delivered by exocytosis. Uptake and complete degradation of adipocyte fragments, which are released by extracellular hydrolysis, leads to macrophage foam cell formation. Exophagy-mediated foam cell formation is a highly efficient means by which macrophages internalize large amounts of lipid, which may ultimately overwhelm the metabolic capacity of the macrophage. This process provides a mechanism for degradation of objects, such as dead adipocytes, that are too large to be phagocytosed by macrophages. PMID:27044658

  4. Inhibition of HIV Expression and Integration in Macrophages by Methylglyoxal-Bis-Guanylhydrazone.

    Science.gov (United States)

    Jin, Xia; McGrath, Michael S; Xu, Hua

    2015-11-01

    Macrophages are a target for infection with HIV and represent one of the viral reservoirs that are relatively resistant to current antiretroviral drugs. Here we demonstrate that methylglyoxal-bis-guanylhydrazone (MGBG), a polyamine analog and potent S-adenosylmethionine decarboxylase inhibitor, decreases HIV expression in monocytes and macrophages. MGBG is selectively concentrated by these cells through a mechanism consistent with active transport by the polyamine transporter. Using a macrophage-tropic reporter virus tagged with the enhanced green fluorescent protein, we demonstrate that MGBG decreases the frequency of HIV-infected cells. The effect is dose dependent and correlates with the production of HIV p24 in culture supernatants. This anti-HIV effect was further confirmed using three macrophage-tropic primary HIV isolates. Viral life cycle mapping studies show that MGBG inhibits HIV DNA integration into the cellular DNA in both monocytes and macrophages. Our work demonstrates for the first time the selective concentration of MGBG by monocytes/macrophages, leading to the inhibition of HIV-1 expression and a reduction in proviral load within macrophage cultures. These results suggest that MGBG may be useful in adjunctive macrophage-targeted therapy for HIV infection. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  5. Neutrophil Microvesicles from Healthy Control and Rheumatoid Arthritis Patients Prevent the Inflammatory Activation of Macrophages

    Directory of Open Access Journals (Sweden)

    Hefin I. Rhys

    2018-03-01

    Full Text Available Microvesicles (MVs are emerging as a novel means to enact cell-to-cell communication in inflammation. Here, we aimed to ascertain the ability of neutrophil-derived MVs to modulate target cell behaviour, the focus being the macrophage.MVs were generated in response to tumour necrosis factor-α, from healthy control neutrophils or those from rheumatoid arthritis patients. MVs were used to stimulate human monocyte-derived macrophages in vitro, or administered intra-articularly in the K/BxN mouse model of arthritis. A macrophage/fibroblast-like synoviocyte co-culture system was used to study the effects of vesicles on the crosstalk between these cells.We demonstrate a direct role for phosphatidylserine and annexin-A1 exposed by the MVs to counteract classical activation of the macrophages, and promote the release of transforming growth factor-β, respectively. Classically-activated macrophages exposed to neutrophil MVs no longer activated fibroblast-like synoviocytes in subsequent co-culture settings. Finally, intra-articular administration of neutrophil MVs from rheumatoid arthritis patients in arthritic mice affected the phenotype of joint macrophages.Altogether these data, with the identification of specific MV determinants, open new opportunities to modulate on-going inflammation in the synovia – mainly by affecting macrophage polarization and potentially also fibroblast-like synoviocytes - through the delivery of autologous or heterologous MVs produced from neutrophils. Keywords: Neutrophils, Macrophages, Vesicles, Rheumatoid arthritis

  6. Identification and comparison of macrophage-induced proteins and proteins induced under various stress conditions in Brucella abortus.

    OpenAIRE

    Rafie-Kolpin, M; Essenberg, R C; Wyckoff, J H

    1996-01-01

    Brucella abortus is a facultative intracellular pathogen of cattle and humans that is capable of survival inside macrophages. In order to understand how B. abortus copes with the conditions during intracellular growth in macrophages, the protein synthesis pattern of the bacteria grown inside bovine macrophages has been compared with that of bacteria grown in the cell culture medium by two-dimensional polyacrylamide gel electrophoresis. Approximately 24 new proteins that are not detected in th...

  7. Progastrin represses the alternative activation of human macrophages and modulates their influence on colon cancer epithelial cells.

    Directory of Open Access Journals (Sweden)

    Carlos Hernández

    Full Text Available Macrophage infiltration is a negative prognostic factor for most cancers but gastrointestinal tumors seem to be an exception. The effect of macrophages on cancer progression depends on their phenotype, which may vary between M1 (pro-inflammatory, defensive to M2 (tolerogenic, pro-tumoral. Gastrointestinal cancers often become an ectopic source of gastrins and macrophages present receptors for these peptides. The aim of the present study is to analyze whether gastrins can affect the pattern of macrophage infiltration in colorectal tumors. We have evaluated the relationship between gastrin expression and the pattern of macrophage infiltration in samples from colorectal cancer and the influence of these peptides on the phenotype of macrophages differentiated from human peripheral monocytes in vitro. The total number of macrophages (CD68+ cells was similar in tumoral and normal surrounding tissue, but the number of M2 macrophages (CD206+ cells was significantly higher in the tumor. However, the number of these tumor-associated M2 macrophages correlated negatively with the immunoreactivity for gastrin peptides in tumor epithelial cells. Macrophages differentiated from human peripheral monocytes in the presence of progastrin showed lower levels of M2-markers (CD206, IL10 with normal amounts of M1-markers (CD86, IL12. Progastrin induced similar effects in mature macrophages treated with IL4 to obtain a M2-phenotype or with LPS plus IFNγ to generate M1-macrophages. Macrophages differentiated in the presence of progastrin presented a reduced expression of Wnt ligands and decreased the number and increased cell death of co-cultured colorectal cancer epithelial cells. Our results suggest that progastrin inhibits the acquisition of a M2-phenotype in human macrophages. This effect exerted on tumor associated macrophages may modulate cancer progression and should be taken into account when analyzing the therapeutic value of gastrin immunoneutralization.

  8. Preparation of guinea pig macrophage for electrophoretic experiments in space

    Science.gov (United States)

    1979-01-01

    Methods of storage and cultivation of macrophage cells in preparation for space experiments were investigated. Results show that freezing and thawing immediately after extraction did not cause any change in viability or electrophoretic mobility of the cells. A prolonged storage at -80 C did cause cell damage as indicated by a 95% reduction in variable cells. Cell damage was decreased when Glycerol or Dimethyl Sulfoxide (DMSO) was added as a cryogenic protective agent. A 100% viability was observed in cultivation experiments after two weeks due to the additional serum. Results from gamma-glutamyl transpeptidase study showed a zero activity rate. It is suggested that a flat stationary field be used for the collection and use of macrophage. It was found that a 24-hour delay in obtaining macrophage cells helps to maintain a pure culture.

  9. Periodontitis-activated monocytes/macrophages cause aortic inflammation

    Science.gov (United States)

    Miyajima, Shin-ichi; Naruse, Keiko; Kobayashi, Yasuko; Nakamura, Nobuhisa; Nishikawa, Toru; Adachi, Kei; Suzuki, Yuki; Kikuchi, Takeshi; Mitani, Akio; Mizutani, Makoto; Ohno, Norikazu; Noguchi, Toshihide; Matsubara, Tatsuaki

    2014-01-01

    A relationship between periodontal disease and atherosclerosis has been suggested by epidemiological studies. Ligature-induced experimental periodontitis is an adequate model for clinical periodontitis, which starts from plaque accumulation, followed by inflammation in the periodontal tissue. Here we have demonstrated using a ligature-induced periodontitis model that periodontitis activates monocytes/macrophages, which subsequently circulate in the blood and adhere to vascular endothelial cells without altering the serum TNF-α concentration. Adherent monocytes/macrophages induced NF-κB activation and VCAM-1 expression in the endothelium and increased the expression of the TNF-α signaling cascade in the aorta. Peripheral blood-derived mononuclear cells from rats with experimental periodontitis showed enhanced adhesion and increased NF-κB/VCAM-1 in cultured vascular endothelial cells. Our results suggest that periodontitis triggers the initial pathogenesis of atherosclerosis, inflammation of the vasculature, through activating monocytes/macrophages. PMID:24893991

  10. The macrophage-histiocytic system

    Energy Technology Data Exchange (ETDEWEB)

    Cross, A

    1971-04-01

    The macrophage-histiocytic system is primarily concerned with the phagocytosis and degradation either of foreign material that enters the organism or of senile and damaged cells belonging to the organism itself. The system includes various kinds of cells with the common ability to process and eventually degrade and digest the ingested material. Two morphological characteristics of these cells are linked to their phagocytic functions: intra-cytoplasmic vacuoles and lysosomes. Although endothelial and fibroblastic cells can ingest particles, it seems that most cells of the macrophage-histiocytic system belong to the monocyte series. The stem cell of the system is still a matter for discussion and the mature cells have attracted a large and confusing array of names. Most of the experimental work with irradiation has involved macrophages of the peritoneal cavity and lymph nodes. It is likely that the other cells of the macrophage-histiocytic system are affected in the same way by irradiation, but this is not certain.

  11. DMPD: The actions of bacterial DNA on murine macrophages. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 10534106 The actions of bacterial DNA on murine macrophages. Sester DP, Stacey KJ, ... Show The actions of bacterial DNA on murine macrophages. PubmedID 10534106 Title The actions of bacterial DNA on murine macrophage

  12. BIGH3 protein and macrophages in retinal endothelial cell apoptosis.

    Science.gov (United States)

    Mondragon, Albert A; Betts-Obregon, Brandi S; Moritz, Robert J; Parvathaneni, Kalpana; Navarro, Mary M; Kim, Hong Seok; Lee, Chi Fung; LeBaron, Richard G; Asmis, Reto; Tsin, Andrew T

    2015-01-01

    Diabetes is a pandemic disease with a higher occurrence in minority populations. The molecular mechanism to initiate diabetes-associated retinal angiogenesis remains largely unknown. We propose an inflammatory pathway of diabetic retinopathy in which macrophages in the diabetic eye provide TGFβ to retinal endothelial cells (REC) in the retinal microvasculature. In response to TGFβ, REC synthesize and secrete a pro-apoptotic BIGH3 (TGFβ-Induced Gene Human Clone 3) protein, which acts in an autocrine loop to induce REC apoptosis. Rhesus monkey retinal endothelial cells (RhREC) were treated with dMCM (cell media of macrophages treated with high glucose and LDL) and assayed for apoptosis (TUNEL), BIGH3 mRNA (qPCR), and protein (Western blots) expressions. Cells were also treated with ΤGFβ1 and 2 for BIGH3 mRNA and protein expression. Inhibition assays were carried out using antibodies for TGFβ1 and for BIGH3 to block apoptosis and mRNA expression. BIGH3 in cultured RhREC cells were identified by immunohistochemistry (IHC). Distribution of BIGH3 and macrophages in the diabetic mouse retina was examined with IHC. RhRECs treated with dMCM or TGFβ showed a significant increase in apoptosis and BIGH3 protein expression. Recombinant BIGH3 added to RhREC culture medium led to a dose-dependent increase in apoptosis. Antibodies (Ab) directed against BIGH3 and TGFβ, as well as TGFβ receptor blocker resulted in a significant reduction in apoptosis induced by either dMCM, TGFβ or BIGH3. IHC showed that cultured RhREC constitutively expressed BIGH3. Macrophage and BIGH3 protein were co-localized to the inner retina of the diabetic mouse eye. Our results support a novel inflammatory pathway for diabetic retinopathy. This pathway is initiated by TGFβ released from macrophages, which promotes synthesis and release of BIGH3 protein by REC and REC apoptosis.

  13. Monocyte chemoattractant protein-1 (MCP-1 regulates macrophage cytotoxicity in abdominal aortic aneurysm.

    Directory of Open Access Journals (Sweden)

    Qiwei Wang

    Full Text Available AIMS: In abdominal aortic aneurysm (AAA, macrophages are detected in the proximity of aortic smooth muscle cells (SMCs. We have previously demonstrated in a murine model of AAA that apoptotic SMCs attract monocytes and other leukocytes by producing MCP-1. Here we tested whether infiltrating macrophages also directly contribute to SMC apoptosis. METHODS AND RESULTS: Using a SMC/RAW264.7 macrophage co-culture system, we demonstrated that MCP-1-primed RAWs caused a significantly higher level of apoptosis in SMCs as compared to control macrophages. Next, we detected an enhanced Fas ligand (FasL mRNA level and membrane FasL protein expression in MCP-1-primed RAWs. Neutralizing FasL blocked SMC apoptosis in the co-culture. In situ proximity ligation assay showed that SMCs exposed to primed macrophages contained higher levels of receptor interacting protein-1 (RIP1/Caspase 8 containing cell death complexes. Silencing RIP1 conferred apoptosis resistance to SMCs. In the mouse elastase injury model of aneurysm, aneurysm induction increased the level of RIP1/Caspase 8 containing complexes in medial SMCs. Moreover, TUNEL-positive SMCs in aneurysmal tissues were frequently surrounded by CD68(+/FasL(+ macrophages. Conversely, elastase-treated arteries from MCP-1 knockout mice display a reduction of both macrophage infiltration and FasL expression, which was accompanied by diminished apoptosis of SMCs. CONCLUSION: Our data suggest that MCP-1-primed macrophages are more cytotoxic. MCP-1 appears to modulate macrophage cytotoxicity by increasing the level of membrane bound FasL. Thus, we showed that MCP-1-primed macrophages kill SMCs through a FasL/Fas-Caspase8-RIP1 mediated mechanism.

  14. Substance P enhances tissue factor release from granulocyte-macrophage colony-stimulating factor-dependent macrophages via the p22phox/β-arrestin 2/Rho A signaling pathway.

    Science.gov (United States)

    Yamaguchi, Rui; Yamamoto, Takatoshi; Sakamoto, Arisa; Ishimaru, Yasuji; Narahara, Shinji; Sugiuchi, Hiroyuki; Yamaguchi, Yasuo

    2016-03-01

    Granulocyte-macrophage colony stimulating factor (GM-CSF) induces procoagulant activity of macrophages. Tissue factor (TF) is a membrane-bound glycoprotein and substance P (SP) is a pro-inflammatory neuropeptide involved in the formation of membrane blebs. This study investigated the role of SP in TF release by GM-CSF-dependent macrophages. SP significantly decreased TF levels in whole-cell lysates of GM-CSF-dependent macrophages. TF was detected in the culture supernatant by enzyme-linked immunosorbent assay after stimulation of macrophages by SP. Aprepitant (an SP/neurokinin 1 receptor antagonist) reduced TF release from macrophages stimulated with SP. Pretreatment of macrophages with a radical scavenger(pyrrolidinedithiocarbamate) also limited the decrease of TF in whole-cell lysates after stimulation with SP. A protein kinase C inhibitor (rottlerin) partially blocked this macrophage response to SP, while it was significantly inhibited by a ROCK inhibitor (Y-27632) or a dynamin inhibitor (dinasore). An Akt inhibitor (perifosine) also partially blocked this response. Furthermore, siRNA targeting p22phox, β-arrestin 2, or Rho A, blunted the release of TF from macrophages stimulated with SP. In other experiments, visceral adipocytes derived from cryopreserved preadipocytes were found to produce SP. In conclusion, SP enhances the release of TF from macrophages via the p22phox/β-arrestin 2/Rho A signaling pathway. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Imaging of macrophage-related lung diseases

    International Nuclear Information System (INIS)

    Marten, Katharina; Hansell, David M.

    2005-01-01

    Macrophage-related pulmonary diseases are a heterogeneous group of disorders characterized by macrophage accumulation, activation or dysfunction. These conditions include smoking-related interstitial lung diseases, metabolic disorders such as Niemann-Pick or Gaucher disease, and rare primary lung tumors. High-resolution computed tomography abnormalities include pulmonary ground-glass opacification secondary to infiltration by macrophages, centrilobular nodules or interlobular septal thickening reflecting peribronchiolar or septal macrophage accumulation, respectively, emphysema caused by macrophage dysfunction, and honeycombing following macrophage-related lung matrix remodeling. (orig.)

  16. Effect of Shark Liver Oil on Peritoneal Murine Macrophages in Responses to Killed-Candida albicans

    Directory of Open Access Journals (Sweden)

    Monire Hajimoradi

    2009-09-01

    Full Text Available Objective(sShark Liver Oil (SLO is an immunomodulator. Macrophages play a key role in host defense against pathogens like fungi. Candida albicans have mechanisms to escape immune system. We determined the effect of killed-Candida on the in vitro viability of macrophages and the effect of SLO on augmentation of this potency.Materials and MethodsPeritoneal macrophages were separated and cultured (3×105/well. At first, the effect of killed-Candida (200 cells/well on macrophage viability was evaluated, using MTT test. Then, MTT was performed on macrophages stimulated with killed-Candida in the presence of SLO. ResultsKilled-Candida suppressed the ability of MTT reduction and hence macrophages viability (P=0.026, but addition of SLO (100 mg/ml significantly enhanced cell viability (P=0.00. So, SLO could neutralize the inhibitory effect of Candida.ConclusionSimultaneous with cytotoxic effect of killed-Candida cells on macrophages viability, SLO augment macrophages viability. So, it can be applied in candidiasis as a complement.

  17. Ability of certain plant extracts traditionally used to treat ciguatera fish poisoning to inhibit nitric oxide production in RAW 264.7 macrophages.

    Science.gov (United States)

    Kumar-Roiné, Shilpa; Matsui, Mariko; Reybier, Karine; Darius, Hélène Taiana; Chinain, Mireille; Pauillac, Serge; Laurent, Dominique

    2009-06-25

    Ciguatera fish poisoning (CFP) is an intertropical ichthyosarcotoxism that manifests in complex assortment of symptoms in humans. Ciguatoxins (CTXs), issued from Gambierdicus spp., are causative agents of this intoxication. We have recently demonstrated that a Pacific CTX (P-CTX-1B) strongly modulated iNOS expression, leading to overproduction of nitric oxide (NO) in RAW 264.7 murine macrophage cells. NO produced in large amounts is involved in a wide range of pathophysiological processes. Many traditional remedies are commonly used in the Pacific against CFP. In this context, bioassay-guided screening was carried out to study NO inhibiting capacity of 28 selected plant extracts. We prepared aqueous extracts of plants used in New Caledonia in the treatment of CFP and screened their NO inhibitory activity in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Among 28 plants tested, Euphorbia hirta (Euphorbiaceae), Syzygium malaccense (Myrtaceae), Schinus terebenthifolius (Anacardiaceae), Punica granatum (Punicaceae), Cerbera manghas (Apocynaceae), Vitex trifolia (Labiateae) and Ximenia americana (Olacaceae) showed inhibitory activity, validating their use as traditional remedies in CFP, and the potential for use in the treatment of conditions accompanied by NO overproduction. These plants are promising candidates for further screening of their active compounds through activity-guided fractionation.

  18. Activated macrophages create lineage-specific microenvironments for pancreatic acinar- and β-cell regeneration in mice.

    Science.gov (United States)

    Criscimanna, Angela; Coudriet, Gina M; Gittes, George K; Piganelli, Jon D; Esni, Farzad

    2014-11-01

    Although the cells that contribute to pancreatic regeneration have been widely studied, little is known about the mediators of this process. During tissue regeneration, infiltrating macrophages debride the site of injury and coordinate the repair response. We investigated the role of macrophages in pancreatic regeneration in mice. We used a saporin-conjugated antibody against CD11b to reduce the number of macrophages in mice following diphtheria toxin receptor-mediated cell ablation of pancreatic cells, and evaluated the effects on pancreatic regeneration. We analyzed expression patterns of infiltrating macrophages after cell-specific injury or from the pancreas of nonobese diabetic mice. We developed an in vitro culture system to study the ability of macrophages to induce cell-specific regeneration. Depletion of macrophages impaired pancreatic regeneration. Macrophage polarization, as assessed by expression of tumor necrosis factor-α, interleukin 6, interleukin 10, and CD206, depended on the type of injury. The signals provided by polarized macrophages promoted lineage-specific generation of acinar or endocrine cells. Macrophage from nonobese diabetic mice failed to provide signals necessary for β-cell generation. Macrophages produce cell type-specific signals required for pancreatic regeneration in mice. Additional study of these processes and signals might lead to new approaches for treating type 1 diabetes or pancreatitis. Copyright © 2014 AGA Institute. Published by Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

    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.

  20. Siderocalin inhibits the intracellular replication of Mycobacterium tuberculosis in macrophages

    DEFF Research Database (Denmark)

    Johnson, Erin E; Srikanth, Chittur V; Sandgren, Andreas

    2010-01-01

    Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show that sideroc......Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show...... findings are consistent with an important role for siderocalin in protection against M.tb infection and suggest that exogenously administered siderocalin may have therapeutic applications in tuberculosis....

  1. Macrophage functions measured by magnetic microparticles in vivo and in vitro

    International Nuclear Information System (INIS)

    Moeller, Winfried; Kreyling, Wolfgang G.; Kohlhaeufl, Martin; Haeussinger, Karl; Heyder, Joachim

    2001-01-01

    Monodisperse ferrimagnetic iron-oxide particles of 1.4 μm geometric diameter were used to study alveolar macrophage functions (phagocytosis, phagosome transport) and cytoskeletal integrity in healthy subjects and in patients with idiopathic pulmonary fibrosis as well as in cultured macrophages. Dysfunctions in phagocytosis, in phagosome transport and cytoskeletal integrity correlated with an impaired alveolar clearance and could be induced in vitro by cytoskeletal drugs

  2. Effect of porcine reproductive and respiratory syndrome virus (PRRSV) on alveolar lung macrophage survival and function

    DEFF Research Database (Denmark)

    Oleksiewicz, Martin B.; Nielsen, Jens

    1999-01-01

    Porcine reproductive and respiratory syndrome virus (PRRSV) recently emerged as an important cause of reproductive disorders and pneumonia in domestic pigs throughout the world. Acute cytocidal replication of PRRSV in alveolar lung macrophages causes the acute pneumonia; however, it remains largely...... infection in this system. In short, in our minimal system containing only a single cell type, phagocytosis-suppressive effects of PRRSV infection were detected, that acted at the culture level by reducing the total number of alveolar lung macrophages....

  3. Macrophage functions measured by magnetic microparticles in vivo and in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Moeller, Winfried E-mail: moeller@gsf.de; Kreyling, Wolfgang G.; Kohlhaeufl, Martin; Haeussinger, Karl; Heyder, Joachim

    2001-07-01

    Monodisperse ferrimagnetic iron-oxide particles of 1.4 {mu}m geometric diameter were used to study alveolar macrophage functions (phagocytosis, phagosome transport) and cytoskeletal integrity in healthy subjects and in patients with idiopathic pulmonary fibrosis as well as in cultured macrophages. Dysfunctions in phagocytosis, in phagosome transport and cytoskeletal integrity correlated with an impaired alveolar clearance and could be induced in vitro by cytoskeletal drugs.

  4. The role of macrophage derived growth factors in pulmonary fibrosis

    International Nuclear Information System (INIS)

    Pickrell, J.A.; Jarpe, M.; Benson, J.M.; Henderson, R.F.

    1988-01-01

    Factors released from rat alveolar macrophages exposed to high (95 μg/mL) concentrations of the fibrogenic agent, nickel subsulfide, were found to inhibit the proliferation of cultured lung epithelial cells and stimulate the growth of fibroblasts. Such factors, if present in the alveoli of rats exposed by inhalation to nickel subsulfide in vivo, may play a role in inhibiting re-epithelization of nickel-damaged lungs and in stimulating fibroblast proliferation, leading to pulmonary fibrosis. (author)

  5. IAP survivin regulates atherosclerotic macrophage survival

    NARCIS (Netherlands)

    Blanc-Brude, Olivier P.; Teissier, Elisabeth; Castier, Yves; Lesèche, Guy; Bijnens, Ann-Pascal; Daemen, Mat; Staels, Bart; Mallat, Ziad; Tedgui, Alain

    2007-01-01

    Inflammatory macrophage apoptosis is critical to atherosclerotic plaque formation, but its mechanisms remain enigmatic. We hypothesized that inhibitor of apoptosis protein (IAP) survivin regulates macrophage death in atherosclerosis. Western blot analysis revealed discrete survivin expression in

  6. Adipose tissue macrophages impair preadipocyte differentiation in humans.

    Directory of Open Access Journals (Sweden)

    Li Fen Liu

    Full Text Available The physiologic mechanisms underlying the relationship between obesity and insulin resistance are not fully understood. Impaired adipocyte differentiation and localized inflammation characterize adipose tissue from obese, insulin-resistant humans. The directionality of this relationship is not known, however. The aim of the current study was to investigate whether adipose tissue inflammation is causally-related to impaired adipocyte differentiation.Abdominal subcutaneous(SAT and visceral(VAT adipose tissue was obtained from 20 human participants undergoing bariatric surgery. Preadipocytes were isolated, and cultured in the presence or absence of CD14+ macrophages obtained from the same adipose tissue sample. Adipocyte differentiation was quantified after 14 days via immunofluorescence, Oil-Red O, and adipogenic gene expression. Cytokine secretion by mature adipocytes cultured with or without CD14+macrophages was quantified.Adipocyte differentiation was significantly lower in VAT than SAT by all measures (p<0.001. With macrophage removal, SAT preadipocyte differentiation increased significantly as measured by immunofluorescence and gene expression, whereas VAT preadipocyte differentiation was unchanged. Adipocyte-secreted proinflammatory cytokines were higher and adiponectin lower in media from VAT vs SAT: macrophage removal reduced inflammatory cytokine and increased adiponectin secretion from both SAT and VAT adipocytes. Differentiation of preadipocytes from SAT but not VAT correlated inversely with systemic insulin resistance.The current results reveal that proinflammatory immune cells in human SAT are causally-related to impaired preadipocyte differentiation, which in turn is associated with systemic insulin resistance. In VAT, preadipocyte differentiation is poor even in the absence of tissue macrophages, pointing to inherent differences in fat storage potential between the two depots.

  7. Role of Osteal Macrophages in Bone Metabolism

    Directory of Open Access Journals (Sweden)

    Sun Wook Cho

    2015-03-01

    Full Text Available Macrophages have been shown to have pleiotropic functions in various pathophysiologies, especially in terms of anti-inflammatory and regenerative activity. Recently, the novel functions of bone marrow resident macrophages (called osteal macrophages were intensively studied in bone development, remodeling and tissue repair processes. This review discusses the current evidence for a role of osteal macrophages in bone modeling, remodeling, and fracture healing processes.

  8. Bulky PAH-DNA induced by exposure of a co-culture model of human alveolar macrophages and embryonic epithelial cells to atmospheric particulate pollution; Adduits encombrants a l'ADN dans des cocultures de cellules pulmonaires humaines exposees a une pollution atmospherique particulaire

    Energy Technology Data Exchange (ETDEWEB)

    Abbas, Imane; Garcon, Guillaume; Billet, Sylvain; Shirali, Pirouz [Universite Lille Nord de France - Lille (France); Unite de Chimie Environnementale et Interactions sur le Vivant, MREI, Universite du Littoral Cote d' Opale, Dunkerque (France); Andre, Veronique; Le Goff, Jeremie; Sichel, Francois [GRECAN, Universite de Caen Basse-Normandie et centre Francois Baclesse, Caen (France); Roy Saint-Georges, Francoise; Mulliez, Philippe [Service de Pneumologie, Hopital Saint-Philibert, GHICL, Lille (France)

    2012-01-15

    Because of their deep penetration in human lungs, fine airborne particulate matter were described as mainly responsible for the deleterious effects of exposure to air pollution on health. Organic constituents are adsorbed on particles surface and, after inhalation, some (polycyclic aromatic hydrocarbons, PAHs) can be activated into reactive metabolites and can bind to DNA. The formation of bulky DNA adducts has been researched after exposure of mono-and co-cultures of alveolar macrophages (AM) and human embryonic human lung epithelial (L132), to fine air pollution particulate matter Air samples have been collected with cascade impactor and characterized: size distribution (92.15% < 2.5{mu}.m), specific surface area (1 m{sup 2}/g), inorganic (Fe, AI, Ca, Na, K, Mg, Pb, etc.) and organic compounds (PAHs, etc.). {sup 32}P post-labeling method was applied to detect bulky DNA adducts in AM and L132, in mono-and co-cultures, 72 h after their exposure to atmospheric particles at their Lethals and Effects concentrations or (LC or CE) to 50% (i.e. MA: EC{sub 50} = 74.63 {mu}g/mL and L132: LC-5-0 = 75.36 {mu}g/mL). Exposure to desorbed particles (MA: C1= 61.11 {mu}g/mL and L132 : C2 = 61.71 {mu}g/mL) and B[a]P (1 {mu}M) were included. Bulky PAH-DNA adducts were detected in AM in mono-culture after exposure to total particles (Pt), to B[a]P and desorbed particles (Pd). Whatever the exposure, no DNA adduct was detected in L132 in mono-culture. These results are coherent with the enzymatic activities of cytochrome P450 l Al in AM and L132. Exposure of co-culture to Pt, or Pd induced bulky adducts to DNA in AM but not in L132. Exposure to B[a]P alone has altered the DNA of AM and L132, in co-culture. Exposure to Pt is closer to the environmental conditions, but conferred an exposure to amounts of genotoxic agents compared to studies using organic extracts. The formation of bulky DNA adducts was nevertheless observed in AM exposed to Pt, in mono- or co-culture, indicating that

  9. Pseudomonas aeruginosa infection alters the macrophage phenotype switching process during wound healing in diabetic mice.

    Science.gov (United States)

    Chen, Sinuo; Li, Renren; Cheng, Chun; Xu, Jing-Ying; Jin, Caixia; Gao, Furong; Wang, Juan; Zhang, Jieping; Zhang, Jingfa; Wang, Hong; Lu, Lixia; Xu, Guo-Tong; Tian, Haibin

    2018-03-07

    Macrophages play critical roles in wound healing process. They switch from "classically activated" (M1) phenotype in the early inflammatory phase to "alternatively activated" (M2) phenotype in the later healing phase. However, the dynamic process of macrophage phenotype switching in diabetic wounds burdened with bacteria is unclear. In this report, Pseudomonas aeruginosa, frequently detected in diabetic foot ulcers, was inoculated into cutaneous wounds of db/db diabetic mice to mimic bacterium-infected diabetic wound healing. We observed that P. aeruginosa infection impaired diabetic wound healing and quickly promoted the expression of pro-inflammatory genes (M1 macrophage markers) tumor necrosis factor-α (tnf-α), interleukin-1β (il-1β) and il-6 in wounds. The expression of markers of M2 macrophages, including il-10, arginase-1, and ym1 were also upregulated. In addition, similar gene expression patterns were observed in macrophages isolated directly from wounds. Immunostaining showed that P. aeruginosa infection increased both the ratios of M1 and M2 macrophages in wounds compared with that in control groups, which was further confirmed by in vitro culturing macrophages with P. aeruginosa and skin fibroblast conditioned medium. However, the ratios of the expression levels of pro-inflammatory genes to anti-inflammatory gene il-10 was increased markedly in P. aeruginosa infected wounds and macrophages compared with that in control groups, and P. aeruginosa prolonged the presence of M1 macrophages in the wounds. These data demonstrated that P. aeruginosa in diabetic wounds activates a mixed M1/M2 macrophage phenotype with an excessive activation of M1 phenotype or relatively inadequate activation of M2 phenotype. © 2018 International Federation for Cell Biology.

  10. Micro RNA in Exosomes from HIV-Infected Macrophages

    Directory of Open Access Journals (Sweden)

    William W. Roth

    2015-12-01

    Full Text Available Exosomes are small membrane-bound vesicles secreted by cells that function to shuttle RNA and proteins between cells. To examine the role of exosomal micro RNA (miRNA during the early stage of HIV-1 infection we characterized miRNA in exosomes from HIV-infected macrophages, compared with exosomes from non-infected macrophages. Primary human monocytes from uninfected donors were differentiated to macrophages (MDM which were either mock-infected or infected with the macrophage-tropic HIV-1 BaL strain. Exosomes were recovered from culture media and separated from virus particles by centrifugation on iodixanol density gradients. The low molecular weight RNA fraction was prepared from purified exosomes. After pre-amplification, RNA was hybridized to microarrays containing probes for 1200 miRNA species of known and unknown function. We observed 48 miRNA species in both infected and uninfected MDM exosomes. Additionally, 38 miRNAs were present in infected-cell exosomes but not uninfected-cell exosomes. Of these, 13 miRNAs were upregulated in exosomes from HIV-infected cells, including 4 miRNA species that were increased by more than 10-fold. Though numerous miRNA species have been identified in HIV-infected cells, relatively little is known about miRNA content in exosomes from these cells. In the future, we plan to investigate whether the upregulated miRNA species we identified are increased in exosomes from HIV-1-positive patients.

  11. HIV-1 and the macrophage

    NARCIS (Netherlands)

    Bol, Sebastiaan M.; Cobos-Jimenez, Viviana; Kootstra, Neeltje A.; van 't Wout, Angelique B.

    2011-01-01

    Macrophages and CD4(+) T cells are natural target cells for HIV-1, and both cell types contribute to the establishment of the viral reservoir that is responsible for continuous residual virus replication during antiretroviral therapy and viral load rebound upon treatment interruption. Scientific

  12. Aloe vera downregulates LPS-induced inflammatory cytokine production and expression of NLRP3 inflammasome in human macrophages.

    Science.gov (United States)

    Budai, Marietta M; Varga, Aliz; Milesz, Sándor; Tőzsér, József; Benkő, Szilvia

    2013-12-01

    Aloe vera has been used in traditional herbal medicine as an immunomodulatory agent inducing anti-inflammatory effects. However, its role on the IL-1β inflammatory cytokine production has not been studied. IL-1β production is strictly regulated both at transcriptional and posttranslational levels through the activity of Nlrp3 inflammasome. In this study we aimed to determine the effect of Aloe vera on the molecular mechanisms of Nlrp3 inflammasome-mediated IL-1β production in LPS-activated human THP-1 cells and monocyte-derived macrophages. Our results show that Aloe vera significantly reduced IL-8, TNFα, IL-6 and IL-1β cytokine production in a dose dependent manner. The inhibitory effect was substantially more pronounced in the primary cells. We found that Aloe vera inhibited the expression of pro-IL-1β, Nlrp3, caspase-1 as well as that of the P2X7 receptor in the LPS-induced primary macrophages. Furthermore, LPS-induced activation of signaling pathways like NF-κB, p38, JNK and ERK were inhibited by Aloe vera in these cells. Altogether, we show for the first time that Aloe vera-mediated strong reduction of IL-1β appears to be the consequence of the reduced expression of both pro-IL-1β as well as Nlrp3 inflammasome components via suppressing specific signal transduction pathways. Furthermore, we show that the expression of the ATP sensor P2X7 receptor is also downregulated by Aloe vera that could also contribute to the attenuated IL-1β cytokine secretion. These results may provide a new therapeutic approach to regulate inflammasome-mediated responses. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Effect of 14-kDa and 47-kDa protein molecules of age garlic extract on peritoneal macrophages.

    Science.gov (United States)

    Daneshmandi, Saeed; Hajimoradi, Monire; Ahmadabad, Hasan Namdar; Hassan, Zuhair Mohammad; Roudbary, Maryam; Ghazanfari, Tooba

    2011-03-01

    Garlic (Allium sativum), traditionally being used as a spice worldwide, has different applications and is claimed to possess beneficial effects in several health ailments such as tumor and atherosclerosis. Garlic is also an immunomodulator and its different components are responsible for different properties. The present work aimed to assess the effect of protein fractions of garlic on peritoneal macrophages. 14-kDa and 47-kDa protein fractions of garlic were purified. Mice peritoneal macrophages were lavaged and cultured in a microtiter plate and exposed to different concentrations of garlic proteins. MTT assay was performed to evaluate the viability of macrophage. The amount of nitric oxide (NO) was detected in culture supernatants of macrophages by Griess reagent and furthermore, the cytotoxicity study of culture supernatants was carried out on WEHI-164 fibrosarcoma cell line as tumor necrosis factor-α bioassay. MTT assay results for both 14-kDa and 47-kDa protein fractions of stimulated macrophages were not significant (P > 0.05). Both 14-kDa and 47-kDa fractions significantly suppressed production of NO from macrophages (P = 0.007 and P = 0.003, respectively). Cytotoxicity of macrophages' supernatant on WEHI-164 fibrosarcoma cells was not affected by garlic protein fractions (P = 0.066 for 14-kDa and P = 0.085 for 47-kDa fractions). according to our finding, 14-kDa and 47-kDa fractions of aged garlic extract are able to suppress NO production from macrophages, which can be used as a biological advantage. These molecules had no cytotoxic effect on macrophages and do not increase tumoricidal property of macrophages.

  14. Macrophage phenotypic subtypes diametrically regulate epithelial-mesenchymal plasticity in breast cancer cells

    International Nuclear Information System (INIS)

    Yang, Min; Ma, Bo; Shao, Hanshuang; Clark, Amanda M.; Wells, Alan

    2016-01-01

    Metastatic progression of breast cancer involves phenotypic plasticity of the carcinoma cells moving between epithelial and mesenchymal behaviors. During metastatic seeding and dormancy, even highly aggressive carcinoma cells take on an E-cadherin-positive epithelial phenotype that is absent from the emergent, lethal metastatic outgrowths. These phenotypes are linked to the metastatic microenvironment, though the specific cells and induction signals are still to be deciphered. Recent evidence suggests that macrophages impact tumor progression, and may alter the balance between cancer cell EMT and MErT in the metastatic microenvironment. Here we explore the role of M1/M2 macrophages in epithelial-mesenchymal plasticity of breast cancer cells by coculturing epithelial and mesenchymal cells lines with macrophages. We found that after polarizing the THP-1 human monocyte cell line, the M1 and M2-types were stable and maintained when co-cultured with breast cancer cells. Surprisingly, M2 macrophages may conferred a growth advantage to the epithelial MCF-7 cells, with these cells being driven to a partial mesenchymal phenotypic as indicated by spindle morphology. Notably, E-cadherin protein expression is significantly decreased in MCF-7 cells co-cultured with M2 macrophages. M0 and M1 macrophages had no effect on the MCF-7 epithelial phenotype. However, the M1 macrophages impacted the highly aggressive mesenchymal-like MDA-MB-231 breast cancer cells to take on a quiescent, epithelial phenotype with re-expression of E-cadherin. The M2 macrophages if anything exacerbated the mesenchymal phenotype of the MDA-MB-231 cells. Our findings demonstrate M2 macrophages might impart outgrowth and M1 macrophages may contribute to dormancy behaviors in metastatic breast cancer cells. Thus EMT and MErT are regulated by selected macrophage phenotype in the liver metastatic microenvironment. These results indicate macrophage could be a potential therapeutic target for limiting death due

  15. Mechanism of interleukin-13 production by granulocyte-macrophage colony-stimulating factor-dependent macrophages via protease-activated receptor-2.

    Science.gov (United States)

    Yamaguchi, Rui; Yamamoto, Takatoshi; Sakamoto, Arisa; Ishimaru, Yasuji; Narahara, Shinji; Sugiuchi, Hiroyuki; Hirose, Eiji; Yamaguchi, Yasuo

    2015-06-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) promotes classically activated M1 macrophages. GM-CSF upregulates protease-activated receptor-2 (PAR-2) protein expression and activation of PAR-2 by human neutrophil elastase (HNE) regulates cytokine production. This study investigated the mechanism of PAR-2-mediated interleukin (IL)-13 production by GM-CSF-dependent macrophages stimulated with HNE. Adherent macrophages were obtained from primary cultures of human mononuclear cells. After stimulation with HNE to activate the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway, IL-13 mRNA and protein levels were assessed by the reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. PAR-2 protein was detected in GM-CSF-dependent macrophages by Western blotting. Unexpectedly, PD98059 (an ERK1 inhibitor) increased IL-13 production, even at higher concentrations. Interestingly, U0126 (an ERK1/2 inhibitor) reduced IL-13 production in a concentration-dependent manner. Neither SB203580 (a p38alpha/p38beta inhibitor) nor BIRB796 (a p38gamma/p38delta inhibitor) affected IL-13 production, while TMB-8 (a calcium chelator) diminished IL-13 production. Stimulation with HNE promoted the production of IL-13 (a Th2 cytokine) by GM-CSF-dependent M1 macrophages. PAR-2-mediated IL-13 production may be dependent on the Ca(2+)/ERK2 signaling pathway. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Propofol pretreatment attenuates LPS-induced granulocyte-macrophage colony-stimulating factor production in cultured hepatocytes by suppressing MAPK/ERK activity and NF-κB translocation

    International Nuclear Information System (INIS)

    Jawan, Bruno; Kao, Y.-H.; Goto, Shigeru; Pan, M.-C.; Lin, Y.-C.; Hsu, L.-W.; Nakano, Toshiaki; Lai, C.-Y.; Sun, C.-K.; Cheng, Y.-F.; Tai, M.-H.

    2008-01-01

    Propofol (PPF), a widely used intravenous anesthetic for induction and maintenance of anesthesia during surgeries, was found to possess suppressive effect on host immunity. This study aimed at investigating whether PPF plays a modulatory role in the lipopolysaccharide (LPS)-induced inflammatory cytokine expression in a cell line of rat hepatocytes. Morphological observation and viability assay showed that PPF exhibits no cytotoxicity at concentrations up to 300 μM after 48 h incubation. Pretreatment with 100 μM PPF for 24 h prior to LPS stimulation was performed to investigate the modulatory effect on LPS-induced inflammatory gene production. The results of semi-quantitative RT-PCR demonstrated that PPF pretreatment significantly suppressed the LPS-induced toll-like receptor (TLR)-4, CD14, tumor necrosis factor (TNF)-α, and granulocyte-macrophage colony-stimulating factor (GM-CSF) gene expression. Western blotting analysis showed that PPF pretreatment potentiated the LPS-induced TLR-4 downregulation. Flow cytometrical analysis revealed that PPF pretreatment showed no modulatory effect on the LPS-upregulated CD14 expression on hepatocytes. In addition, PPF pretreatment attenuated the phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) and IκBα, as well as the nuclear translocation of NF-κB primed by LPS. Moreover, addition of PD98059, a MAPK kinase inhibitor, significantly suppressed the LPS-induced NF-κB nuclear translocation and GM-CSF production, suggesting that the PPF-attenuated GM-CSF production in hepatocytes may be attributed to its suppressive effect on MAPK/ERK signaling pathway. In conclusion, PPF as an anesthetic may clinically benefit those patients who are vulnerable to sepsis by alleviating sepsis-related inflammatory response in livers

  17. Effect of Vibrio cholerae neuraminidase on the mitogen response of T lymphocytes. I. Enhancement of macrophage T-lymphocyte cooperation in concanavalin-A-induced lymphocyte activation.

    Science.gov (United States)

    Knop, J

    1980-12-01

    Vibrio cholerae neuraminidase (VCN) enhances the immune response of lymphocytes in various systems, such as antigen- and mitogen-induced blastogenesis, mixed lymphocyte culture (MLC) and tumor-cell response. We used macrophage-depleted and reconstituted murine lymph-node T-cells to investigate the effect of VCN on macrophage-T-lymphocyte co-operation in Con-A-induced lymphocyte activation. In unfractionated lymph-node cells VCN enhanced the Con-A-induced lymphocyte activation as measured by 3H-thymidine (3H-dThd) incorporation. Removing macrophages from the cells resulted in a significantly diminished response. In addition the enhancing effect of VCN was greatly reduced. Reconstitution of the lymphocyte cultures with macrophages in increasing numbers and from various sources rstored the lymphocyte response and the enhancing effect of VCN. VCN proved to be most efficient in cultures reconstituted with normal peritoneal macrophages. Some effect was also observed using bone-marrow-derived (BM) macrophages. However, higher numbers of normal PE macrophages in the presence of VCN inhibited lymphocyte activation, and inhibition by thioglycollate-broth-induced macrophages was considerably increased by VCN. These results suggest that VCN acts by increasing the efficiency of macrophage-T lymphocyte interaction.

  18. [Macrophage activation in atherosclerosis. Message 1: Activation of macrophages normally and in atherosclerotic lesions].

    Science.gov (United States)

    Nikiforov, N G; Kornienko, V Y; Karagodin, V P; Orekhov, A N

    2015-01-01

    Macrophages play important role in initiation and progression of inflammation in atherosclerosis. Plaque macrophages were shown to exhibit a phenotypic range that is intermediate between two extremes, M1 (proinflammatory) and M2 (anti-inflammatory). Indeed, in atherosclerosis, macrophages demonstrate phenotypic plasticity to rapidly adjust to changing microenvironmental conditions. In plaque macrophages demonstrate different phenotypes, and besides macrophage phenotypes could be changed. Phenotypes M1, M2, M4, Mhem, HA-mac, M(Hb) u Mox are described in the article. Ability of macrophages change their phenotype also considered.

  19. Alternate radiolabeled markers for detecting metabolic activity of Mycobacterium leprae residing in murine macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Prasad, H.K.; Hastings, R.C.

    1985-05-01

    This study demonstrated the utility of using 4% NaOH as a murine macrophage cell-solubilizing agent to discriminate between host macrophage metabolism and that of intracellular Mycobacterium leprae. A 4% concentration of NaOH had no deleterious effect on labeled mycobacteria. Thereby, alternate radiolabeled indicators of the metabolic activity of intracellular M. leprae could be experimented with. Significant incorporation of /sup 14/C-amino acid mixture, (/sup 14/C)leucine, (/sup 14/C)uridine, and carrier-free /sup 32/P was observed in cultures containing freshly extracted (''live'') strains of M. leprae as compared with control cultures containing autoclaved bacilli.

  20. Alternate radiolabeled markers for detecting metabolic activity of Mycobacterium leprae residing in murine macrophages

    International Nuclear Information System (INIS)

    Prasad, H.K.; Hastings, R.C.

    1985-01-01

    This study demonstrated the utility of using 4% NaOH as a murine macrophage cell-solubilizing agent to discriminate between host macrophage metabolism and that of intracellular Mycobacterium leprae. A 4% concentration of NaOH had no deleterious effect on labeled mycobacteria. Thereby, alternate radiolabeled indicators of the metabolic activity of intracellular M. leprae could be experimented with. Significant incorporation of 14 C-amino acid mixture, [ 14 C]leucine, [ 14 C]uridine, and carrier-free 32 P was observed in cultures containing freshly extracted (''live'') strains of M. leprae as compared with control cultures containing autoclaved bacilli

  1. Immunomodulatory role for membrane vesicles released by THP-1 macrophages and respiratory pathogens during macrophage infection.

    Science.gov (United States)

    Volgers, Charlotte; Benedikter, Birke J; Grauls, Gert E; Savelkoul, Paul H M; Stassen, Frank R M

    2017-11-13

    During infection, inflammation is partially driven by the release of mediators which facilitate intercellular communication. Amongst these mediators are small membrane vesicles (MVs) that can be released by both host cells and Gram-negative and -positive bacteria. Bacterial membrane vesicles are known to exert immuno-modulatory and -stimulatory actions. Moreover, it has been proposed that host cell-derived vesicles, released during infection, also have immunostimulatory properties. In this study, we assessed the release and activity of host cell-derived and bacterial MVs during the first hours following infection of THP-1 macrophages with the common respiratory pathogens non-typeable Haemophilus influenzae, Moraxella catarrhalis, Streptococcus pneumoniae, and Pseudomonas aeruginosa. Using a combination of flow cytometry, tunable resistive pulse sensing (TRPS)-based analysis and electron microscopy, we demonstrated that the release of MVs occurs by both host cells and bacteria during infection. MVs released during infection and bacterial culture were found to induce a strong pro-inflammatory response by naive THP-1 macrophages. Yet, these MVs were also found to induce tolerance of host cells to secondary immunogenic stimuli and to enhance bacterial adherence and the number of intracellular bacteria. Bacterial MVs may play a dual role during infection, as they can both trigger and dampen immune responses thereby contributing to immune defence and bacterial survival.

  2. Toll-like receptor 4-positive macrophages protect mice from Pasteurella pneumotropica-induced pneumonia

    Science.gov (United States)

    Hart, Marcia L.; Mosier, Derek A.; Chapes, Stephen K.

    2003-01-01

    This study investigates Toll-like receptor 4 (TLR4)-positive macrophages in early recognition and clearance of pulmonary bacteria. TLR4 is a trans-membrane receptor that is the primary recognition molecule for lipopolysaccharide of gram-negative bacteria. The TLR4(Lps-del) mouse strains C57BL10/ScN (B10) and STOCK Abb(tm1) TLR4(Lps-del) Slc11a1(s)(B10 x C2D) are susceptible to pulmonary infections and develop pneumonia when naturally or experimentally infected by the opportunistic bacterium Pasteurella pneumotropica. Since these mice have the TLR4(Lps-del) genotype, we hypothesized that reconstitution of mice with TLR4-positive macrophages would provide resistance to this bacterium. A cultured macrophage cell line (C2D macrophages) and bone marrow cells from C2D mice were adoptively transferred to B10 and B10 x C2D mice by intraperitoneal injection. C2D macrophages increased B10 and B10 x C2D mouse resistance to P. pneumotropica. In C2D-recipient mice there was earlier transcription of tumor necrosis factor alpha and chemokines JE and macrophage inflammatory protein 2 (MIP-2) in the lungs of B10 and B10 x C2D mice, and there was earlier transcription of KC and MIP-1alpha in B10 x C2D mice. In addition, the course of inflammation following experimental Pasteurella challenge was altered in C2D recipients. C2D macrophages also protected B10 x C2D mice, which lack CD4(+) T cells. These data indicate that macrophages are critical for pulmonary immunity and can provide host resistance to P. pneumotropica. This study indicates that TLR4-positive macrophages are important for early recognition and clearance of pulmonary bacterial infections.

  3. Brucella Dissociation Is Essential for Macrophage Egress and Bacterial Dissemination

    Directory of Open Access Journals (Sweden)

    Thomas A Ficht

    2014-03-01

    Full Text Available It has long been observed that smooth Brucella can dissociate into rough mutants that are cytotoxic to macrophages. However, the in vivo biological significance and/or mechanistic de-tails of Brucella dissociation and cytotoxicity remain incomplete. In the current report, a plaque assay was developed using Brucella strains exhibiting varying degrees of cytotoxicity. Infected monolayers were observed daily using phase contrast microscopy for plaque formation while Brucella uptake and replication were monitored using an immunofluorescence assay (IFA. Vis-ible plaques were detected at 4-5 days post infection (p.i. with cytotoxic Brucella 16M∆manBA at an MOI of 0.1. IFA staining demonstrated that the plaques consisted of macrophages with replicating Brucella. Visible plaques were not detected in monolayers infected with non-cytotoxic 16M∆manBA∆virB2 at an MOI of 0.1. However, IFA staining did reveal small groups of macrophages (foci with replicating Brucella in the monolayers infected with 16M∆manBA∆virB2. The size of the foci observed in macrophage monolayers infected with rough Brucella correlated directly with cytotoxicity measured in liquid culture, suggesting that cytotoxicity was essential for Brucella egress and dissemination. In monolayers infected with 16M, small and large foci were observed. Double antibody staining revealed spontaneous rough mutants within the large, but not the small foci in 16M infected monolayers. Furthermore, plaque formation was observed in the large foci derived from 16M infections. Finally, the addi-tion of gentamicin to the culture medium inhibited plaque formation, suggesting that the cell-to-cell spreading occurred only following release of the organisms from the cells. Taken together, these results demonstrate that Brucella induced cytotoxicity is critical for Brucella egress and dissemination.

  4. Brucella dissociation is essential for macrophage egress and bacterial dissemination.

    Science.gov (United States)

    Pei, Jianwu; Kahl-McDonagh, Melissa; Ficht, Thomas A

    2014-01-01

    It has long been observed that smooth Brucella can dissociate into rough mutants that are cytotoxic to macrophages. However, the in vivo biological significance and/or mechanistic details of Brucella dissociation and cytotoxicity remain incomplete. In the current report, a plaque assay was developed using Brucella strains exhibiting varying degrees of cytotoxicity. Infected monolayers were observed daily using phase contrast microscopy for plaque formation while Brucella uptake and replication were monitored using an immunofluorescence assay (IFA). Visible plaques were detected at 4-5 days post infection (p.i.) with cytotoxic Brucella 16MΔmanBA at an MOI of 0.1. IFA staining demonstrated that the plaques consisted of macrophages with replicating Brucella. Visible plaques were not detected in monolayers infected with non-cytotoxic 16MΔmanBAΔvirB2 at an MOI of 0.1. However, IFA staining did reveal small groups of macrophages (foci) with replicating Brucella in the monolayers infected with 16MΔmanBAΔvirB2. The size of the foci observed in macrophage monolayers infected with rough Brucella correlated directly with cytotoxicity measured in liquid culture, suggesting that cytotoxicity was essential for Brucella egress and dissemination. In monolayers infected with 16M, small and large foci were observed. Double antibody staining revealed spontaneous rough mutants within the large, but not the small foci in 16M infected monolayers. Furthermore, plaque formation was observed in the large foci derived from 16M infections. Finally, the addition of gentamicin to the culture medium inhibited plaque formation, suggesting that cell-to-cell spread occurred only following release of the organisms from the cells. Taken together, these results demonstrate that Brucella-induced cytotoxicity is critical for Brucella egress and dissemination.

  5. Dexamethasone palmitate ameliorates macrophages-rich graft-versus-host disease by inhibiting macrophage functions.

    Science.gov (United States)

    Nishiwaki, Satoshi; Nakayama, Takayuki; Murata, Makoto; Nishida, Tetsuya; Terakura, Seitaro; Saito, Shigeki; Kato, Tomonori; Mizuno, Hiroki; Imahashi, Nobuhiko; Seto, Aika; Ozawa, Yukiyasu; Miyamura, Koichi; Ito, Masafumi; Takeshita, Kyosuke; Kato, Hidefumi; Toyokuni, Shinya; Nagao, Keisuke; Ueda, Ryuzo; Naoe, Tomoki

    2014-01-01

    Macrophage infiltration of skin GVHD lesions correlates directly with disease severity, but the mechanisms underlying this relationship remain unclear and GVHD with many macrophages is a therapeutic challenge. Here, we characterize the macrophages involved in GVHD and report that dexamethasone palmitate (DP), a liposteroid, can ameliorate such GVHD by inhibiting macrophage functions. We found that host-derived macrophages could exacerbate GVHD in a mouse model through expression of higher levels of pro-inflammatory TNF-α and IFN-γ, and lower levels of anti-inflammatory IL-10 than resident macrophages in mice without GVHD. DP significantly decreased the viability and migration capacity of primary mouse macrophages compared to conventional dexamethasone in vitro. DP treatment on day 7 and day 14 decreased macrophage number, and attenuated GVHD score and subsequent mortality in a murine model. This is the first study to provide evidence that therapy for GVHD should be changed on the basis of infiltrating cell type.

  6. DMPD: Macrophage differentiation and function in health and disease. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available in health and disease. PubmedID 18251777 Title Macrophage differentiation and function in health and disease...thol Int. 2008 Mar;58(3):143-55. (.png) (.svg) (.html) (.csml) Show Macrophage differentiation and function

  7. Increased metabolite levels of glycolysis and pentose phosphate pathway in rabbit atherosclerotic arteries and hypoxic macrophage.

    Directory of Open Access Journals (Sweden)

    Atsushi Yamashita

    Full Text Available AIMS: Inflammation and possibly hypoxia largely affect glucose utilization in atherosclerotic arteries, which could alter many metabolic systems. However, metabolic changes in atherosclerotic plaques remain unknown. The present study aims to identify changes in metabolic systems relative to glucose uptake and hypoxia in rabbit atherosclerotic arteries and cultured macrophages. METHODS: Macrophage-rich or smooth muscle cell (SMC-rich neointima was created by balloon injury in the iliac-femoral arteries of rabbits fed with a 0.5% cholesterol diet or a conventional diet. THP-1 macrophages stimulated with lipopolysaccharides (LPS and interferon-γ (INFγ were cultured under normoxic and hypoxic conditions. We evaluated comprehensive arterial and macrophage metabolism by performing metabolomic analyses using capillary electrophoresis-time of flight mass spectrometry. We evaluated glucose uptake and its relationship to vascular hypoxia using (18F-fluorodeoxyglucose ((18F-FDG and pimonidazole, a marker of hypoxia. RESULTS: The levels of many metabolites increased in the iliac-femoral arteries with macrophage-rich neointima, compared with those that were not injured and those with SMC-rich neointima (glycolysis, 4 of 9; pentose phosphate pathway, 4 of 6; tricarboxylic acid cycle, 4 of 6; nucleotides, 10 of 20. The uptake of (18F-FDG in arterial walls measured by autoradiography positively correlated with macrophage- and pimonidazole-immunopositive areas (r = 0.76, and r = 0.59 respectively; n = 69 for both; p<0.0001. Pimonidazole immunoreactivity was closely localized with the nuclear translocation of hypoxia inducible factor-1α and hexokinase II expression in macrophage-rich neointima. The levels of glycolytic (8 of 8 and pentose phosphate pathway (4 of 6 metabolites increased in LPS and INFγ stimulated macrophages under hypoxic but not normoxic condition. Plasminogen activator inhibitor-1 protein levels in the supernatant were closely

  8. Mesenchymal stem cell-educated macrophages

    OpenAIRE

    Eggenhofer Elke; Hoogduijn Martin J

    2012-01-01

    Abstract Mesenchymal stem cells (MSC) mediate their immunosuppressive effects via a variety of mechanisms. One of these mechanisms involves the induction of macrophages with immunomodulatory capacities. This effect of MSC may be exploited when MSC are used as a cell therapeutic product. Furthermore, MSC are resident in tissues where they may locally target infiltrating macrophages to adapt more regulatory properties. The present review discusses the interaction between MSC and macrophages, th...

  9. Quantitative GPCR and ion channel transcriptomics in primary alveolar macrophages and macrophage surrogates

    Directory of Open Access Journals (Sweden)

    Groot-Kormelink Paul J

    2012-10-01

    Full Text Available Abstract Background Alveolar macrophages are one of the first lines of defence against invading pathogens and play a central role in modulating both the innate and acquired immune systems. By responding to endogenous stimuli within the lung, alveolar macrophages contribute towards the regulation of the local inflammatory microenvironment, the initiation of wound healing and the pathogenesis of viral and bacterial infections. Despite the availability of protocols for isolating primary alveolar macrophages from the lung these cells remain recalcitrant to expansion in-vitro and therefore surrogate cell types, such as monocyte derived macrophages and phorbol ester-differentiated cell lines (e.g. U937, THP-1, HL60 are frequently used to model macrophage function. Methods The availability of high throughput gene expression technologies for accurate quantification of transcript levels enables the re-evaluation of these surrogate cell types for use as cellular models of the alveolar macrophage. Utilising high-throughput TaqMan arrays and focussing on dynamically regulated families of integral membrane proteins, we explore the similarities and differences in G-protein coupled receptor (GPCR and ion channel expression in alveolar macrophages and their widely used surrogates. Results The complete non-sensory GPCR and ion channel transcriptome is described for primary alveolar macrophages and macrophage surrogates. The expression of numerous GPCRs and ion channels whose expression were hitherto not described in human alveolar macrophages are compared across primary macrophages and commonly used macrophage cell models. Several membrane proteins known to have critical roles in regulating macrophage function, including CXCR6, CCR8 and TRPV4, were found to be highly expressed in macrophages but not expressed in PMA-differentiated surrogates. Conclusions The data described in this report provides insight into the appropriate choice of cell models for

  10. M2 macrophages activate WNT signaling pathway in epithelial cells: relevance in ulcerative colitis.

    Directory of Open Access Journals (Sweden)

    Jesús Cosín-Roger

    Full Text Available Macrophages, which exhibit great plasticity, are important components of the inflamed tissue and constitute an essential element of regenerative responses. Epithelial Wnt signalling is involved in mechanisms of proliferation and differentiation and expression of Wnt ligands by macrophages has been reported. We aim to determine whether the macrophage phenotype determines the expression of Wnt ligands, the influence of the macrophage phenotype in epithelial activation of Wnt signalling and the relevance of this pathway in ulcerative colitis. Human monocyte-derived macrophages and U937-derived macrophages were polarized towards M1 or M2 phenotypes and the expression of Wnt1 and Wnt3a was analyzed by qPCR. The effects of macrophages and the role of Wnt1 were analyzed on the expression of β-catenin, Tcf-4, c-Myc and markers of cell differentiation in a co-culture system with Caco-2 cells. Immunohistochemical staining of CD68, CD206, CD86, Wnt1, β-catenin and c-Myc were evaluated in the damaged and non-damaged mucosa of patients with UC. We also determined the mRNA expression of Lgr5 and c-Myc by qPCR and protein levels of β-catenin by western blot. Results show that M2, and no M1, activated the Wnt signaling pathway in co-culture epithelial cells through Wnt1 which impaired enterocyte differentiation. A significant increase in the number of CD206+ macrophages was observed in the damaged mucosa of chronic vs newly diagnosed patients. CD206 immunostaining co-localized with Wnt1 in the mucosa and these cells were associated with activation of canonical Wnt signalling pathway in epithelial cells and diminution of alkaline phosphatase activity. Our results show that M2 macrophages, and not M1, activate Wnt signalling pathways and decrease enterocyte differentiation in co-cultured epithelial cells. In the mucosa of UC patients, M2 macrophages increase with chronicity and are associated with activation of epithelial Wnt signalling and diminution in

  11. Macrophage antioxidant protection within atherosclerotic plaques.

    Science.gov (United States)

    Gieseg, Steven P; Leake, David S; Flavall, Elizabeth M; Amit, Zunika; Reid, Linzi; Yang, Ya-Ting

    2009-01-01

    Macrophage cells within inflammatory lesions are exposed to a wide range of degrading and cytotoxic molecules including reactive oxygen species. Unlike neutrophils, macrophages do not normally die in this environment but continue to generate oxidants, phagocytose cellular remains, and release a range of cyto-active agents which modulate the immune response. It is this potential of the macrophage cell to survive in an oxidative environment that allows the growth and complexity of advanced atherosclerotic plaques. This review will examine the oxidants encountered by macrophages within an atherosclerotic plaque and describe some of the potential antioxidant mechanisms which enable macrophages to function within inflammatory lesions. Ascorbate, a-tocopherol, and glutathione appear to be central to the protection of macrophages yet additional antioxidant mechanisms appear to be involved. Gamma-Interferon causes macrophages to generate 7,8-dihydroneopterin, neopterin and 3-hydroxyanthranilic acid both of which have antioxidant properties. Manganese superoxide dismutase is also upregulated in macrophages. The evidence that these antioxidants provide further protection, so allowing the macrophage cells to survive within sites of chronic inflammation such as atherosclerotic plaques, will be described.

  12. MONOCYTES AND MACROPHAGES IN PREGNANCY AND PREECLAMPSIA

    Directory of Open Access Journals (Sweden)

    Marijke M Faas

    2014-06-01

    Full Text Available Preeclampsia is an important complication in pregnancy, characterized byhypertension and proteinuria in the second half of pregnancy. Generalizedactivation of the inflammatory response is thought to play a role in thepathogenesis of preeclampsia. Monocytes may play a central role in thisinflammatory response. Monocytes are short lived cells, that mature in thecirculation and invade into tissues upon an inflammatory stimulus anddevelop into macrophages. Macrophages are abundantly present in theendometrium and play a role in implantation and placentation in normalpregnancy. In preeclampsia, these macrophages appear to be present in largernumbers and are also activated. In the present review we focused on the roleof monocytes and macrophages in the pathophysiology of preeclampsia.

  13. Macrophage Plasticity in Skeletal Muscle Repair

    Directory of Open Access Journals (Sweden)

    Elena Rigamonti

    2014-01-01

    Full Text Available Macrophages are one of the first barriers of host defence against pathogens. Beyond their role in innate immunity, macrophages play increasingly defined roles in orchestrating the healing of various injured tissues. Perturbations of macrophage function and/or activation may result in impaired regeneration and fibrosis deposition as described in several chronic pathological diseases. Heterogeneity and plasticity have been demonstrated to be hallmarks of macrophages. In response to environmental cues they display a proinflammatory (M1 or an alternative anti-inflammatory (M2 phenotype. A lot of evidence demonstrated that after acute injury M1 macrophages infiltrate early to promote the clearance of necrotic debris, whereas M2 macrophages appear later to sustain tissue healing. Whether the sequential presence of two different macrophage populations results from a dynamic shift in macrophage polarization or from the recruitment of new circulating monocytes is a subject of ongoing debate. In this paper, we discuss the current available information about the role that different phenotypes of macrophages plays after injury and during the remodelling phase in different tissue types, with particular attention to the skeletal muscle.

  14. Diarctigenin, a lignan constituent from Arctium lappa, down-regulated zymosan-induced transcription of inflammatory genes through suppression of DNA binding ability of nuclear factor-kappaB in macrophages.

    Science.gov (United States)

    Kim, Byung Hak; Hong, Seong Su; Kwon, Soon Woo; Lee, Hwa Young; Sung, Hyeran; Lee, In-Jeong; Hwang, Bang Yeon; Song, Sukgil; Lee, Chong-Kil; Chung, Daehyun; Ahn, Byeongwoo; Nam, Sang-Yoon; Han, Sang-Bae; Kim, Youngsoo

    2008-11-01

    Diarctigenin was previously isolated as an inhibitor of nitric oxide (NO) production in macrophages from the seeds of Arctium lappa used as an alternative medicine for the treatment of inflammatory disorders. However, little is known about the molecular basis of these effects. Here, we demonstrated that diarctigenin inhibited the production of NO, prostaglandin E(2), tumor necrosis factor-alpha, and interleukin (IL)-1beta and IL-6 with IC(50) values of 6 to 12 miciroM in zymosan- or lipopolysaccharide-(LPS) activated macrophages. Diarctigenin attenuated zymosan-induced mRNA synthesis of inducible NO synthase (iNOS) and also inhibited promoter activities of iNOS and cytokine genes in the cells. Because nuclear factor (NF)-kappaB plays a pivotal role in inflammatory gene transcription, we next investigated the effect of diarctigenin on NF-kappaB activation. Diarctigenin inhibited the transcriptional activity and DNA binding ability of NF-kappaB in zymosan-activated macrophages but did not affect the degradation and phosphorylation of inhibitory kappaB (IkappaB) proteins. Moreover, diarctigenin suppressed expression vector NF-kappaB p65-elicited NF-kappaB activation and also iNOS promoter activity, indicating that the compound could directly target an NF-kappa-activating signal cascade downstream of IkappaB degradation and inhibit NF-kappaB-regulated iNOS expression. Diarctigenin also inhibited the in vitro DNA binding ability of NF-kappaB but did not affect the nuclear import of NF-kappaB p65 in the cells. Taken together, diarctigenin down-regulated zymosan- or LPS-induced inflammatory gene transcription in macrophages, which was due to direct inhibition of the DNA binding ability of NF-kappaB. Finally, this study provides a pharmacological potential of diarctigenin in the NF-kappaB-associated inflammatory disorders.

  15. 2',4-Dihydroxy-3',4',6'-trimethoxychalcone from Chromolaena odorata possesses anti-inflammatory effects via inhibition of NF-κB and p38 MAPK in lipopolysaccharide-activated RAW 264.7 macrophages.

    Science.gov (United States)

    Dhar, Rana; Kimseng, Rungruedee; Chokchaisiri, Ratchanaporn; Hiransai, Poonsit; Utaipan, Tanyarath; Suksamrarn, Apichart; Chunglok, Warangkana

    2018-02-01

    Immune dysregulation has been implicated in the pathogenesis of many diseases. Macrophages play a crucial role contributing to the onset, progression, and resolution of inflammation. Macrophage inflammatory mediators are of considerable interest as potential targets to treat inflammatory diseases. The present study was conducted to elucidate the anti-inflammatory mechanism of 2',4-dihydroxy-3',4',6'-trimethoxychalcone (1), the major chalcone isolated from Chromolaena odorata (L.) R.M.King & H.Rob, against lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 macrophages. Cell viability, nitric oxide (NO), and proinflammatory cytokines of LPS-activated RAW 264.7 cells were measured by MTT, Griess, and ELISA assays, respectively. Cell lysates were subjected to Western blotting for investigation of protein expression. Treatment with the major chalcone 1 significantly attenuated the production of NO and proinflammatory cytokines, tumor necrosis factor-α, interleukin-1β, and interleukin-6 in a dose-dependent manner. The chalcone suppressed nuclear factor-κB (NF-κB) stimulation by preventing activation of inhibitor κB kinase (IKK) α/β, degradation of inhibitor κB (IκB) α, and translocation of p65 NF-κB into the nucleus. Additionally, the chalcone markedly repressed the phosphorylation of p38 mitogen-activated protein kinase (MAPK), but no further inhibition was detected for c-Jun N-terminal activated kinases or extracellular regulated kinases. Thus, suppression of NF-κB and p38 MAPK activation may be the core mechanism underlying the anti-inflammatory activity of 2',4-dihydroxy-3',4',6'-trimethoxychalcone (1). These findings provide evidence that 2',4-dihydroxy-3',4',6'-trimethoxychalcone (1) possesses anti-inflammatory activity via targeting proinflammatory macrophages. This anti-inflammatory chalcone is a promising compound for reducing inflammation.

  16. DMPD: Shaping of monocyte and macrophage function by adenosine receptors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17056121 Shaping of monocyte and macrophage function by adenosine receptors. Hasko ...tml) (.csml) Show Shaping of monocyte and macrophage function by adenosine receptors. PubmedID 17056121 Titl...e Shaping of monocyte and macrophage function by adenosine receptors. Authors Has

  17. DMPD: Macrophage activation by endogenous danger signals. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18161744 Macrophage activation by endogenous danger signals. Zhang X, Mosser DM. J ...Pathol. 2008 Jan;214(2):161-78. (.png) (.svg) (.html) (.csml) Show Macrophage activation by endogenous dange...r signals. PubmedID 18161744 Title Macrophage activation by endogenous danger signals. Authors Zhang X, Moss

  18. DMPD: Regulation of endogenous apolipoprotein E secretion by macrophages. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18388328 Regulation of endogenous apolipoprotein E secretion by macrophages. Kockx ...svg) (.html) (.csml) Show Regulation of endogenous apolipoprotein E secretion by macrophages. PubmedID 18388...328 Title Regulation of endogenous apolipoprotein E secretion by macrophages. Aut

  19. DMPD: Macrophage migration inhibitory factor and host innate immune responses tomicrobes. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 14620137 Macrophage migration inhibitory factor and host innate immune responses to...microbes. Calandra T. Scand J Infect Dis. 2003;35(9):573-6. (.png) (.svg) (.html) (.csml) Show Macrophage migration... inhibitory factor and host innate immune responses tomicrobes. PubmedID 14620137 Title Macrophage migration

  20. DMPD: Cellular signaling in macrophage migration and chemotaxis. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 11073096 Cellular signaling in macrophage migration and chemotaxis. Jones GE. J Leu...koc Biol. 2000 Nov;68(5):593-602. (.png) (.svg) (.html) (.csml) Show Cellular signaling in macrophage migration... and chemotaxis. PubmedID 11073096 Title Cellular signaling in macrophage migration and chemotaxis. Autho

  1. DMPD: Monocyte/macrophage traffic in HIV and SIV encephalitis. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 12960230 Monocyte/macrophage traffic in HIV and SIV encephalitis. Kim WK, Corey S, ...Alvarez X, Williams K. J Leukoc Biol. 2003 Nov;74(5):650-6. Epub 2003 Aug 11. (.png) (.svg) (.html) (.csml) Show Monocyte/macrophage... traffic in HIV and SIV encephalitis. PubmedID 12960230 Title Monocyte/macrophage tr

  2. DMPD: CSF-1 and cell cycle control in macrophages. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 8981359 CSF-1 and cell cycle control in macrophages. Hamilton JA. Mol Reprod Dev. 1...997 Jan;46(1):19-23. (.png) (.svg) (.html) (.csml) Show CSF-1 and cell cycle control in macrophages. PubmedI...D 8981359 Title CSF-1 and cell cycle control in macrophages. Authors Hamilton JA. Publication Mol Reprod Dev

  3. DMPD: Silica binding and toxicity in alveolar macrophages. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18226603 Silica binding and toxicity in alveolar macrophages. Hamilton RF Jr, Thaku...l) Show Silica binding and toxicity in alveolar macrophages. PubmedID 18226603 Title Silica binding and toxicity in alveolar macropha...ges. Authors Hamilton RF Jr, Thakur SA, Holian A. Public

  4. DMPD: Iron regulation of hepatic macrophage TNFalpha expression. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 11841920 Iron regulation of hepatic macrophage TNFalpha expression. Tsukamoto H. Fr...ee Radic Biol Med. 2002 Feb 15;32(4):309-13. (.png) (.svg) (.html) (.csml) Show Iron regulation of hepatic macrophage... TNFalpha expression. PubmedID 11841920 Title Iron regulation of hepatic macrophage TNFalpha expres

  5. Modulation of allogeneic stimulation in man. I. Characterization of an in vitro induced suppressor macrophage population

    International Nuclear Information System (INIS)

    Stux, S.V.; Dubey, D.P.; Yunis, E.J.

    1981-01-01

    Cultured human peripheral blood mononuclear cells suppressed the allogeneic response of fresh autologous lymphocytes. This suppressor activity developed gradually over a period of one week. The cells primarily responsible for this effect were enriched by Ficoll density gradient centrifugation. It was found that the suppressor cell is a large, low density nylon wool adherent, radioresistant, phagocytic, and nonspecific esterase positive mononuclear cell. Moreover, these cells did not form E rosettes and were Fc positive. Electron microscopy confirmed that suppressor cells were macrophage like. Suppressor activity was not due to cytotoxicity, crowding, or steric hinderance by the cultured cells. The suppressor macrophage population did not appear to inhibit the allogeneic response via prostaglandin or arginase release, or interfere with the tritiated thymidine uptake by release of endogenous thymidine. The above system is viewed as an in vitro model of immune regulation by suppressor macrophages, in the context of allogeneic response

  6. Human macrophage foam cells degrade atherosclerotic plaques through cathepsin K mediated processes

    DEFF Research Database (Denmark)

    Barascuk, Natasha; Skjøt-Arkil, Helene; Register, Thomas C

    2010-01-01

    BACKGROUND: Proteolytic degradation of Type I Collagen by proteases may play an important role in remodeling of atherosclerotic plaques, contributing to increased risk of plaque rupture.The aim of the current study was to investigate whether human macrophage foam cells degrade the extracellular...... matrix (ECM) of atherosclerotic plaques by cathepsin K mediated processes. METHODS: We 1) cultured human macrophages on ECM and measured cathepsin K generated fragments of type I collagen (C-terminal fragments of Type I collagen (CTX-I) 2) investigated the presence of CTX-I in human coronary arteries......-I in areas of intimal hyperplasia and in shoulder regions of advanced plaques. Treatment of human monocytes with M-CSF or M-CSF+LDL generated macrophages and foam cells producing CTX-I when cultured on type I collagen enriched matrix. Circulating levels of CTX-I were not significantly different in women...

  7. β-Glucan from Lentinus edodes inhibits nitric oxide and tumor necrosis factor-α production and phosphorylation of mitogen-activated protein kinases in lipopolysaccharide-stimulated murine RAW 264.7 macrophages.

    Science.gov (United States)

    Xu, Xiaojuan; Yasuda, Michiko; Nakamura-Tsuruta, Sachiko; Mizuno, Masashi; Ashida, Hitoshi

    2012-01-06

    Lentinan (LNT), a β-glucan from the fruiting bodies of Lentinus edodes, is well known to have immunomodulatory activity. NO and TNF-α are associated with many inflammatory diseases. In this study, we investigated the effects of LNT extracted by sonication (LNT-S) on the NO and TNF-α production in LPS-stimulated murine RAW 264.7 macrophages. The results suggested that treatment with LNT-S not only resulted in the striking inhibition of TNF-α and NO production in LPS-activated macrophage RAW 264.7 cells, but also the protein expression of inducible NOS (iNOS) and the gene expression of iNOS mRNA and TNF-α mRNA. It is surprising that LNT-S enhanced LPS-induced NF-κB p65 nuclear translocation and NF-κB luciferase activity, but severely inhibited the phosphorylation of JNK1/2 and ERK1/2. The neutralizing antibodies of anti-Dectin-1 and anti-TLR2 hardly affected the inhibition of NO production. All of these results suggested that the suppression of LPS-induced NO and TNF-α production was at least partially attributable to the inhibition of JNK1/2 and ERK1/2 activation. This work discovered a promising molecule to control the diseases associated with overproduction of NO and TNF-α.

  8. Cardiosphere-Derived Cells Facilitate Heart Repair by Modulating M1/M2 Macrophage Polarization and Neutrophil Recruitment.

    Directory of Open Access Journals (Sweden)

    Al Shaimaa Hasan

    Full Text Available Cardiosphere-derived cells (CDCs, one of the promising stem cell sources for myocardial repair, have been tested in clinical trials and resulted in beneficial effects; however, the relevant mechanisms are not fully understood. In this study, we examined the hypothesis that CDCs favor heart repair by switching the macrophages from a pro-inflammatory phenotype (M1 into a regulatory anti-inflammatory phenotype (M2. Macrophages from mice were cultured with CDCs-conditioned medium or with fibroblasts-conditioned medium as a control. Immunostaining showed that CDCs-conditioned medium significantly enhanced the expression of CD206 (a marker for M2 macrophages, but decreased the expression of CD86 (a marker for M1 macrophages 3 days after culture. For animal studies, we used an acute myocardial infarction model of mice. We injected CDCs, fibroblasts, or saline only into the border zone of infarction. Then we collected the heart tissues for histological analysis 5 and 14 days after treatment. Compared with control animals, CDCs treatment significantly decreased M1 macrophages and neutrophils but increased M2 macrophages in the infarcted heart. Furthermore, CDCs-treated mice had reduced infarct size and fewer apoptotic cells compared to the controls. Our data suggest that CDCs facilitate heart repair by modulating M1/M2 macrophage polarization and neutrophil recruitment, which may provide a new insight into the mechanisms of stem cell-based myocardial repair.

  9. Cardiosphere-Derived Cells Facilitate Heart Repair by Modulating M1/M2 Macrophage Polarization and Neutrophil Recruitment

    Science.gov (United States)

    Hasan, Al Shaimaa; Luo, Lan; Yan, Chen; Zhang, Tian-Xia; Urata, Yoshishige; Goto, Shinji; Mangoura, Safwat A.; Abdel-Raheem, Mahmoud H.; Zhang, Shouhua; Li, Tao-Sheng

    2016-01-01

    Cardiosphere-derived cells (CDCs), one of the promising stem cell sources for myocardial repair, have been tested in clinical trials and resulted in beneficial effects; however, the relevant mechanisms are not fully understood. In this study, we examined the hypothesis that CDCs favor heart repair by switching the macrophages from a pro-inflammatory phenotype (M1) into a regulatory anti-inflammatory phenotype (M2). Macrophages from mice were cultured with CDCs-conditioned medium or with fibroblasts-conditioned medium as a control. Immunostaining showed that CDCs-conditioned medium significantly enhanced the expression of CD206 (a marker for M2 macrophages), but decreased the expression of CD86 (a marker for M1 macrophages) 3 days after culture. For animal studies, we used an acute myocardial infarction model of mice. We injected CDCs, fibroblasts, or saline only into the border zone of infarction. Then we collected the heart tissues for histological analysis 5 and 14 days after treatment. Compared with control animals, CDCs treatment significantly decreased M1 macrophages and neutrophils but increased M2 macrophages in the infarcted heart. Furthermore, CDCs-treated mice had reduced infarct size and fewer apoptotic cells compared to the controls. Our data suggest that CDCs facilitate heart repair by modulating M1/M2 macrophage polarization and neutrophil recruitment, which may provide a new insight into the mechanisms of stem cell-based myocardial repair. PMID:27764217

  10. Adrenomedullin Regulates IL-1β Gene Expression in F4/80+ Macrophages during Synovial Inflammation

    Science.gov (United States)

    Takano, Shotaro; Miyagi, Masayuki; Inoue, Gen; Aikawa, Jun; Iwabuchi, Kazuya; Takaso, Masashi

    2017-01-01

    Adrenomedullin (AM) plays an important role in the regulation of inflammatory processes; however, the role and expression of AM in synovial inflammation have not been determined. To investigate the expression and role of AM in inflamed synovial tissue (ST), the gene expression profiles of AM in the ST, including synovial macrophages and fibroblasts, of a murine patellar surgical dislocation model were characterized. In addition, the effects of interleukin- (IL-) 1β and AM in cultured synovial cells were also examined. CD11c+ macrophages were found to be elevated in ST of the surgically dislocated patella. Higher gene expression of CD11c, IL-1β, AM, receptor activity-modifying proteins 2 (RAMP2), and 3 (RAMP3) was also observed in ST obtained from the dislocated side. AM expression was also significantly increased in synovial fibroblasts and macrophages in response to IL-1β treatment. Synovial macrophages also highly expressed RAMP3 compared to fibroblasts and this expression was further stimulated by exogenously added IL-1β. Further, the treatment of the F4/80-positive cell fraction obtained from ST with AM inhibited IL-1β expression. Taken together, these findings demonstrated that AM was produced by synovial fibroblasts and macrophages in inflamed ST and that increased levels of AM may exert anti-inflammatory effects on synovial macrophages. PMID:28299347

  11. Transferrin-derived synthetic peptide induces highly conserved pro-inflammatory responses of macrophages.

    Science.gov (United States)

    Haddad, George; Belosevic, Miodrag

    2009-02-01

    We examined the induction of macrophage pro-inflammatory responses by transferrin-derived synthetic peptide originally identified following digestion of transferrin from different species (murine, bovine, human N-lobe and goldfish) using elastase. The mass spectrometry analysis of elastase-digested murine transferrin identified a 31 amino acid peptide located in the N2 sub-domain of the transferrin N-lobe, that we named TMAP. TMAP was synthetically produced and shown to induce a number of pro-inflammatory genes by quantitative PCR. TMAP induced chemotaxis, a potent nitric oxide response, and TNF-alpha secretion in different macrophage populations; P338D1 macrophage-like cells, mouse peritoneal macrophages, mouse bone marrow-derived macrophages (BMDM) and goldfish macrophages. The treatment of BMDM cultures with TMAP stimulated the production of nine cytokines and chemokines (IL-6, MCP-5, MIP-1 alpha, MIP-1 gamma, MIP-2, GCSF, KC, VEGF, and RANTES) that was measured using cytokine antibody array and confirmed by Western blot. Our results indicate that transferrin-derived peptide, TMAP, is an immunomodulating molecule capable of inducing pro-inflammatory responses in lower and higher vertebrates.

  12. HIV-1-infected macrophages induce astrogliosis by SDF-1α and matrix metalloproteinases

    International Nuclear Information System (INIS)

    Okamoto, Mika; Wang, Xin; Baba, Masanori

    2005-01-01

    Brain macrophages/microglia and astrocytes are known to be involved in the pathogenesis of HIV-1-associated dementia (HAD). To clarify their interaction and contribution to the pathogenesis, HIV-1-infected or uninfected macrophages were used as a model of brain macrophages/microglia, and their effects on human astrocytes in vitro were examined. The culture supernatants of HIV-1-infected or uninfected macrophages induced significant astrocyte proliferation, which was annihilated with a neutralizing antibody to stromal cell-derived factor (SDF)-1α or a matrix metalloproteinase (MMP) inhibitor. In these astrocytes, CXCR4, MMP, and tissue inhibitors of matrix metalloproteinase mRNA expression and SDF-1α production were significantly up-regulated. The supernatants of infected macrophages were always more effective than those of uninfected cells. Moreover, the enhanced production of SDF-1α was suppressed by the MMP inhibitor. These results indicate that the activated and HIV-1-infected macrophages can indirectly induce astrocyte proliferation through up-regulating SDF-1α and MMP production, which implies a mechanism of astrogliosis in HAD

  13. Lactoferricin Peptides Increase Macrophages' Capacity To Kill Mycobacterium avium.

    Science.gov (United States)

    Silva, Tânia; Moreira, Ana C; Nazmi, Kamran; Moniz, Tânia; Vale, Nuno; Rangel, Maria; Gomes, Paula; Bolscher, Jan G M; Rodrigues, Pedro N; Bastos, Margarida; Gomes, Maria Salomé

    2017-01-01

    Mycobacterial infections cause a significant burden of disease and death worldwide. Their treatment is long, toxic, costly, and increasingly prone to failure due to bacterial resistance to currently available antibiotics. New therapeutic options are thus clearly needed. Antimicrobial peptides represent an important source of new antimicrobial molecules, both for their direct activity and for their immunomodulatory potential. We have previously reported that a short version of the bovine antimicrobial peptide lactoferricin with amino acids 17 to 30 (LFcin17-30), along with its variants obtained by specific amino acid substitutions, killed Mycobacterium avium in broth culture. In the present work, those peptides were tested against M. avium living inside its natural host cell, the macrophage. We found that the peptides increased the antimicrobial action of the conventional antibiotic ethambutol inside macrophages. Moreover, the d-enantiomer of the lactoferricin peptide (d-LFcin17-30) was more stable and induced significant killing of intracellular mycobacteria by itself. Interestingly, d-LFcin17-30 did not localize to M. avium -harboring phagosomes but induced the production of proinflammatory cytokines and increased the formation of lysosomes and autophagosome-like vesicles. These results lead us to conclude that d-LFcin17-30 primes macrophages for intracellular microbial digestion through phagosomal maturation and/or autophagy, culminating in mycobacterial killing. IMPORTANCE The genus Mycobacterium comprises several pathogenic species, including M. tuberculosis , M. leprae , M. avium , etc. Infections caused by these bacteria are particularly difficult to treat due to their intrinsic impermeability, low growth rate, and intracellular localization. Antimicrobial peptides are increasingly acknowledged as potential treatment tools, as they have a high spectrum of activity, low tendency to induce bacterial resistance, and immunomodulatory properties. In this study, we

  14. Proprotein convertase 1/3 inhibited macrophages: A novel therapeutic based on drone macrophages.

    Science.gov (United States)

    Duhamel, Marie; Rodet, Franck; Murgoci, Adriana; Wisztorski, Maxence; Day, Robert; Fournier, Isabelle; Salzet, Michel

    2016-06-01

    We demonstrated here thanks to proteomic, that proprotein convertase 1/3 knockdown macrophages present all the characteristic of activated pro-inflammatory macrophages. TLR4 and TLR9 signaling pathways can be enhanced leading to the secretion of pro-inflammatory factors and antitumor factors. We can control their activation by controlling one enzyme, PC1/3. In a tumor context, PC1/3 inhibition in macrophages may reactivate them and lead to a cytokine storm after stimulation "at distance" with a TLR ligand. Therefore, we name these proprotein convertase inhibited macrophages the "drone macrophages". They constitute an innovative cell therapy to treat efficiently tumors.

  15. Role of Tellurite Resistance Operon in Filamentous Growth of Yersinia pestis in Macrophages.

    Science.gov (United States)

    Ponnusamy, Duraisamy; Clinkenbeard, Kenneth D

    2015-01-01

    Yersinia pestis initiates infection by parasitism of host macrophages. In response to macrophage infections, intracellular Y. pestis can assume a filamentous cellular morphology which may mediate resistance to host cell innate immune responses. We previously observed the expression of Y. pestis tellurite resistance proteins TerD and TerE from the terZABCDE operon during macrophage infections. Others have observed a filamentous response associated with expression of tellurite resistance operon in Escherichia coli exposed to tellurite. Therefore, in this study we examine the potential role of Y. pestis tellurite resistance operon in filamentous cellular morphology during macrophage infections. In vitro treatment of Y. pestis culture with sodium tellurite (Na2TeO3) caused the bacterial cells to assume a filamentous phenotype similar to the filamentous phenotype observed during macrophage infections. A deletion mutant for genes terZAB abolished the filamentous morphologic response to tellurite exposure or intracellular parasitism, but without affecting tellurite resistance. However, a terZABCDE deletion mutant abolished both filamentous morphologic response and tellurite resistance. Complementation of the terZABCDE deletion mutant with terCDE, but not terZAB, partially restored tellurite resistance. When the terZABCDE deletion mutant was complemented with terZAB or terCDE, Y. pestis exhibited filamentous morphology during macrophage infections as well as while these complemented genes were being expressed under an in vitro condition. Further in E. coli, expression of Y. pestis terZAB, but not terCDE, conferred a filamentous phenotype. These findings support the role of Y. pestis terZAB mediation of the filamentous response phenotype; whereas, terCDE confers tellurite resistance. Although the beneficial role of filamentous morphological responses by Y. pestis during macrophage infections is yet to be fully defined, it may be a bacterial adaptive strategy to macrophage

  16. Chitosan drives anti-inflammatory macrophage polarisation and pro-inflammatory dendritic cell stimulation

    Directory of Open Access Journals (Sweden)

    MI Oliveira

    2012-07-01

    Full Text Available Macrophages and dendritic cells (DC share the same precursor and play key roles in immunity. Modulation of their behaviour to achieve an optimal host response towards an implanted device is still a challenge. Here we compare the differentiation process and polarisation of these related cell populations and show that they exhibit different responses to chitosan (Ch, with human monocyte-derived macrophages polarising towards an anti-inflammatory phenotype while their DC counterparts display pro-inflammatory features. Macrophages and DC, whose interactions with biomaterials are frequently analysed using fully differentiated cells, were cultured directly on Ch films, rather than exposed to the polymer after complete differentiation. Ch was the sole stimulating factor and activated both macrophages and DC, without leading to significant T cell proliferation. After 10 d on Ch, macrophages significantly down-regulated expression of pro-inflammatory markers, CD86 and MHCII. Production of pro-inflammatory cytokines, particularly TNF-α, decreased with time for cells cultured on Ch, while anti-inflammatory IL-10 and TGF-β1, significantly increased. Altogether, these results suggest an M2c polarisation. Also, macrophage matrix metalloproteinase activity was augmented and cell motility was stimulated by Ch. Conversely, DC significantly enhanced CD86 expression, reduced IL-10 secretion and increased TNF-α and IL-1β levels. Our findings indicate that cells with a common precursor may display different responses, when challenged by the same biomaterial. Moreover, they help to further comprehend macrophage/DC interactions with Ch and the balance between pro- and anti-inflammatory signals associated with implant biomaterials. We propose that an overall pro-inflammatory reaction may hide the expression of anti-inflammatory cytokines, likely relevant for tissue repair/regeneration.

  17. Carp macrophages and neutrophilic granulocytes secrete an interleukin-1-like factor.

    NARCIS (Netherlands)

    Verburg-van Kemenade, B.M.L.; Weyts, F.A.A.; Debets, R.; Flik, G.

    1995-01-01

    Carp, Cyprinus carpio L, macrophages and neutrophilic granulocytes obtained from pronephros were cultured. Supernatant was harvested after 48 h and tested for interleukin-1 (IL-1) bioactivity. A concentration-dependent stimulation of proliferation was found of carp Ig− lymphocytes as well as of the

  18. Ameloginins promote an alternatively activated macrophage phenotype in vitro

    DEFF Research Database (Denmark)

    Almqvist, S; Werthen, M; Lyngstadas, SP

    2011-01-01

    aggregates were visualised by transmission electron microscopy. The amelogenin treatment of macrophages increased several pro- and anti-inflammatory cytokines, including alternative macrophage activation marker AMAC-1 (p

  19. Macrophage diversity in renal injury and repair

    NARCIS (Netherlands)

    Ricardo, Sharon D.; van Goor, Harry; Eddy, Allison A.

    Monocyte-derived macrophages can determine the outcome of the immune response and whether this response contributes to tissue repair or mediates tissue destruction. In addition to their important role in immune-mediated renal disease and host defense, macrophages play a fundamental role in tissue

  20. Macrophage polarization: the epigenetic point of view

    NARCIS (Netherlands)

    van den Bossche, Jan; Neele, Annette E.; Hoeksema, Marten A.; de Winther, Menno P. J.

    2014-01-01

    The first functions of macrophages to be identified by Metchnikoff were phagocytosis and microbial killing. Although these are important features, macrophages are functionally very complex and involved in virtually all aspects of life, from immunity and host defense, to homeostasis, tissue repair

  1. Macrophages Promote Axon Regeneration with Concurrent Neurotoxicity

    NARCIS (Netherlands)

    Gensel, J.C.; Nakamura, S.; Guan, Z.; Rooijen, van N.; Ankeny, D.P.; Popovich, P.G.

    2009-01-01

    Activated macrophages can promote regeneration of CNS axons. However, macrophages also release factors that kill neurons. These opposing functions are likely induced simultaneously but are rarely considered together in the same experimental preparation. A goal of this study was to unequivocally

  2. Genesis and kinetics of peritoneal macrophages

    International Nuclear Information System (INIS)

    Wacker, H.H.

    1982-01-01

    The author intended to develop an experimental model for investigations of the proliferation kinetics of tissue macrophages, using the example of peritoneal macrophages. To get a suitable cell population, a blood cell population was labelled with 3 H-thymidine and transferred in a parabiotic test. (orig./MG) [de

  3. Human Cord Blood and Bone Marrow CD34+ Cells Generate Macrophages That Support Erythroid Islands.

    Directory of Open Access Journals (Sweden)

    Eyayu Belay

    Full Text Available Recently, we developed a small molecule responsive hyperactive Mpl-based Cell Growth Switch (CGS that drives erythropoiesis associated with macrophages in the absence of exogenous cytokines. Here, we compare the physical, cellular and molecular interaction between the macrophages and erythroid cells in CGS expanded CD34+ cells harvested from cord blood, marrow or G-CSF-mobilized peripheral blood. Results indicated that macrophage based erythroid islands could be generated from cord blood and marrow CD34+ cells but not from G-CSF-mobilized CD34+ cells. Additional studies suggest that the deficiency resides with the G-CSF-mobilized CD34+ derived monocytes. Gene expression and proteomics studies of the in vitro generated erythroid islands detected the expression of erythroblast macrophage protein (EMP, intercellular adhesion molecule 4 (ICAM-4, CD163 and DNASE2. 78% of the erythroblasts in contact with macrophages reached the pre reticulocyte orthochromatic stage of differentiation within 14 days of culture. The addition of conditioned medium from cultures of CD146+ marrow fibroblasts resulted in a 700-fold increase in total cell number and a 90-fold increase in erythroid cell number. This novel CD34+ cell derived erythroid island may serve as a platform to explore the molecular basis of red cell maturation and production under normal, stress and pathological conditions.

  4. Macrophage Activation Mechanisms in Human Monocytic Cell Line-derived Macrophages.

    Science.gov (United States)

    Sumiya, Yu; Ishikawa, Mami; Inoue, Takahiro; Inui, Toshio; Kuchiike, Daisuke; Kubo, Kentaro; Uto, Yoshihiro; Nishikata, Takahito

    2015-08-01

    Although the mechanisms of macrophage activation are important for cancer immunotherapy, they are poorly understood. Recently, easy and robust assay systems for assessing the macrophage-activating factor (MAF) using monocytic cell line-derived macrophages were established. Gene-expression profiles of U937- and THP-1-derived macrophages were compared using gene expression microarray analysis and their responses against several MAFs were examined by in vitro experiments. Activated states of these macrophages could not be assigned to a specific sub-type but showed, however, different unique characteristics. The unique of monocytic cell line-derived macrophages could provide clues to understand the activation mechanism of macrophages and, therefore, help to develop effective cancer immunotherapy with MAFs. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  5. Purinergic signaling during macrophage differentiation results in M2 alternative activated macrophages.

    Science.gov (United States)

    Barberà-Cremades, Maria; Baroja-Mazo, Alberto; Pelegrín, Pablo

    2016-02-01

    Macrophages represent a highly heterogenic cell population of the innate immune system, with important roles in the initiation and resolution of the inflammatory response. Purinergic signaling regulates both M1 and M2 macrophage function at different levels by controlling the secretion of cytokines, phagocytosis, and the production of reactive oxygen species. We found that extracellular nucleotides arrest macrophage differentiation from bone marrow precursors via adenosine and P2 receptors. This results in a mature macrophage with increased expression of M2, but not M1, genes. Similar to adenosine and ATP, macrophage growth arrested with LPS treatment resulted in an increase of the M2-related marker Ym1. Recombinant Ym1 was able to affect macrophage proliferation and could, potentially, be involved in the arrest of macrophage growth during hematopoiesis. © Society for Leukocyte Biology.

  6. Alternatively activated macrophages (M2 macrophages) in the skin of patient with localized scleroderma.

    Science.gov (United States)

    Higashi-Kuwata, Nobuyo; Makino, Takamitsu; Inoue, Yuji; Takeya, Motohiro; Ihn, Hironobu

    2009-08-01

    Localized scleroderma is a connective tissue disorder that is limited to the skin and subcutaneous tissue. Macrophages have been reported to be particularly activated in patients with skin disease including systemic sclerosis and are potentially important sources for fibrosis-inducing cytokines, such as transforming growth factor beta. To clarify the features of immunohistochemical characterization of the immune cell infiltrates in localized scleroderma focusing on macrophages, skin biopsy specimens were analysed by immunohistochemistry. The number of cells stained with monoclonal antibodies, CD68, CD163 and CD204, was calculated. An evident macrophage infiltrate and increased number of alternatively activated macrophages (M2 macrophages) in their fibrotic areas were observed along with their severity of inflammation. This study revealed that alternatively activated macrophages (M2 macrophages) may be a potential source of fibrosis-inducing cytokines in localized scleroderma, and may play a crucial role in the pathogenesis of localized scleroderma.

  7. Mycobacteria, Metals, and the Macrophage

    Science.gov (United States)

    Niederweis, Michael; Wolschendorf, Frank; Mitra, Avishek; Neyrolles, Olivier

    2015-01-01

    Summary Mycobacterium tuberculosis is a facultative intracellular pathogen that thrives inside host macrophages. A key trait of M. tuberculosis is to exploit and manipulate metal cation trafficking inside infected macrophages to ensure survival and replication inside the phagosome. Here we describe the recent fascinating discoveries that the mammalian immune system responds to infections with M. tuberculosis by overloading the phagosome with copper and zinc, two metals which are essential nutrients in small quantities but are toxic in excess. M. tuberculosis has developed multi-faceted resistance mechanisms to protect itself from metal toxicity including control of uptake, sequestration inside the cell, oxidation, and efflux. The host response to infections combines this metal poisoning strategy with nutritional immunity mechanisms that deprive M. tuberculosis from metals such as iron and manganese to prevent bacterial replication. Both immune mechanisms rely on the translocation of metal transporter proteins to the phagosomal membrane during the maturation process of the phagosome. This review summarizes these recent findings and discusses how metal-targeted approaches might complement existing TB chemotherapeutic regimens with novel anti-infective therapies. PMID:25703564

  8. Effects of Opsonization and Gamma Interferon on Growth of Brucella Melitensis 16M in Mouse Peritoneal Macrophages In Vitro

    Science.gov (United States)

    2000-01-01

    bacterial CFU (Fig. 3). Macrophages cultured without brucel - lae or cultured with brucellae but without IFN-7 made ɘ.3 nmol of nitrite/200 JJLI well...receptors used for the uptake of nonopsonized brucel - lae are unknown. Synergy between the receptor for the Fc domain of immunoglobulin G (FcR) and

  9. Moracin C, A Phenolic Compound Isolated from Artocarpus heterophyllus, Suppresses Lipopolysaccharide-Activated Inflammatory Responses in Murine Raw264.7 Macrophages.

    Science.gov (United States)

    Yao, Xue; Wu, Dang; Dong, Ningning; Ouyang, Ping; Pu, Jiaqian; Hu, Qian; Wang, Jingyuan; Lu, Weiqiang; Huang, Jin

    2016-07-25

    Artocarpus heterophyllus, a popular tropical fruit commonly known as the jackfruit tree, is normally planted in subtropical or tropical areas. Since a variety of phytochemicals isolated from A. heterophyllus have been found to possess potently anti-inflammatory, antiviral and antimalarial activities, researchers have devoted much interest to its potential pharmaceutical value. However, the exact mechanism underlying its anti-inflammatory activity is not well characterized. In this study, seven natural products isolated from A. heterophyllus, including 25-Hydroxycycloart-23-en-3-one (HY), Artocarpin (AR), Dadahol A (DA), Morachalcone A (MA), Artoheterophyllin B (AB), Cycloheterophyllin (CY) and Moracin C (MC) were collected. Lipopolysaccharide (LPS)-stimulated inflammatory response in RAW264.7 macrophages were used in this study. Among these compounds, MC significantly inhibited LPS-activated reactive oxygen species (ROS) and nitric oxide (NO) release without marked cytotoxicity. Furthermore, MC effectively reduced LPS stimulated up-regulation of mRNA and protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and serval pro-inflammatory cytokines (interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor α (TNF-α)). Mechanistic studies revealed that the anti-inflammatory effect of MC was associated with the activation of the mitogen activated protein kinases (MAPKs) (including p38, ERK and JNK) and nuclear factor-κB (NF-κB) pathways, especially reducing the nuclear translocation of NF-κB p65 subunit as revealed by nuclear separation experiment and confocal microscopy.

  10. Unexpected macrophage-independent dyserythropoiesis in Gaucher disease.

    Science.gov (United States)

    Reihani, Nelly; Arlet, Jean-Benoit; Dussiot, Michael; de Villemeur, Thierry Billette; Belmatoug, Nadia; Rose, Christian; Colin-Aronovicz, Yves; Hermine, Olivier; Le Van Kim, Caroline; Franco, Melanie

    2016-12-01

    Gaucher disease is a rare inherited disease caused by a deficiency in glucocerebrosidase leading to lipid accumulation in cells of mononuclear-macrophage lineage known as Gaucher cells. Visceral enlargement, bone involvement, mild anemia and thrombocytopenia are the major manifestations of Gaucher disease. We have previously demonstrated that the red blood cells from patients exhibit abnormal properties, which indicates a new role in Gaucher disease pathophysiology. To investigate whether erythroid progenitors are affected, we examined the in vitro erythropoiesis from the peripheral CD34 + cells of patients and controls. CD34- cells were differentiated into macrophages and co-cultivated with erythroblasts. We showed an accelerated differentiation of erythroid progenitors without maturation arrest from patients compared to controls. This abnormal differentiation persisted in the patients when the same experiments were performed without macrophages, which strongly suggested that dyserythropoiesis in Gaucher disease is secondary to an inherent defect in the erythroid progenitors. The accelerated differentiation was associated with reduced cell proliferation. As a result, less mature erythroid cells were generated in vitro in the Gaucher disease cultures compared to the control. We then compared the biological characteristics of untreated patients according to their anemic status. Compared to the non-anemic group, the anemic patients exhibit higher plasma levels of growth differentiation factor-15, a marker of ineffective erythropoiesis, but they had no indicators of hemolysis and similar reticulocyte counts. Taken together, these results demonstrated an unsuspected dyserythropoiesis that was independent of the macrophages and could participate, at least in part, to the basis of anemia in Gaucher disease. Copyright© Ferrata Storti Foundation.

  11. B-1 cells modulate the murine macrophage response to Leishmania major infection.

    Science.gov (United States)

    Arcanjo, Angelica F; Nunes, Marise P; Silva-Junior, Elias B; Leandro, Monique; da Rocha, Juliana Dutra Barbosa; Morrot, Alexandre; Decote-Ricardo, Debora; Freire-de-Lima, Celio Geraldo

    2017-05-26

    To investigate the modulatory effect of B-1 cells on murine peritoneal macrophages infected with Leishmania major ( L. major ) in vitro . Peritoneal macrophages obtained from BALB/c and BALB/c XID mice were infected with L. major and cultured in the presence or absence of B-1 cells obtained from wild-type BALB/c mice. Intracellular amastigotes were counted, and interleukin-10 (IL-10) production was quantified in the cellular supernatants using an enzyme-linked immunosorbent assay. The levels of the lipid mediator prostaglandin E2 (PGE 2 ) were determined using a PGE 2 enzyme immunoassay kit (Cayman Chemical, Ann Arbor, MI), and the number of lipid bodies was quantified in the cytoplasm of infected macrophages in the presence and absence of B-1 cells. Culturing the cells with selective PGE 2 -neutralizing drugs inhibited PGE 2 production and confirmed the role of this lipid mediator in IL-10 production. In contrast, we demonstrated that B-1 cells derived from IL-10 KO mice did not favor the intracellular growth of L. major . We report that B-1 cells promote the growth of L. major amastigotes inside peritoneal murine macrophages. We demonstrated that the modulatory effect was independent of physical contact between the cells, suggesting that soluble factor(s) were released into the cultures. We demonstrated in our co-culture system that B-1 cells trigger IL-10 production by L. major -infected macrophages. Furthermore, the increased secretion of IL-10 was attributed to the presence of the lipid mediator PGE 2 in supernatants of L. major -infected macrophages. The presence of B-1 cells also favors the production of lipid bodies by infected macrophages. In contrast, we failed to obtain the same effect on parasite replication inside L. major -infected macrophages when the B-1 cells were isolated from IL-10 knockout mice. Our results show that elevated levels of PGE 2 and IL-10 produced by B-1 cells increase L. major growth, as indicated by the number of parasites in cell

  12. A recombinant fusion toxin based on enzymatic inactive C3bot1 selectively targets macrophages.

    Directory of Open Access Journals (Sweden)

    Lydia Dmochewitz

    Full Text Available BACKGROUND: The C3bot1 protein (~23 kDa from Clostridium botulinum ADP-ribosylates and thereby inactivates Rho. C3bot1 is selectively taken up into the cytosol of monocytes/macrophages but not of other cell types such as epithelial cells or fibroblasts. Most likely, the internalization occurs by a specific endocytotic pathway via acidified endosomes. METHODOLOGY/PRINCIPAL FINDINGS: Here, we tested whether enzymatic inactive C3bot1E174Q serves as a macrophage-selective transport system for delivery of enzymatic active proteins into the cytosol of such cells. Having confirmed that C3bot1E174Q does not induce macrophage activation, we used the actin ADP-ribosylating C2I (∼50 kDa from Clostridium botulinum as a reporter enzyme for C3bot1E174Q-mediated delivery into macrophages. The recombinant C3bot1E174Q-C2I fusion toxin was cloned and expressed as GST-protein in Escherichia coli. Purified C3bot1E174Q-C2I was recognized by antibodies against C2I and C3bot and showed C2I-specific enzyme activity in vitro. When applied to cultured cells C3bot1E174Q-C2I ADP-ribosylated actin in the cytosol of macrophages including J774A.1 and RAW264.7 cell lines as well as primary cultured human macrophages but not of epithelial cells. Together with confocal fluorescence microscopy experiments, the biochemical data indicate the selective uptake of a recombinant C3-fusion toxin into the cytosol of macrophages. CONCLUSIONS/SIGNIFICANCE: In summary, we demonstrated that C3bot1E174Q can be used as a delivery system for fast, selective and specific transport of enzymes into the cytosol of living macrophages. Therefore, C3-based fusion toxins can represent valuable molecular tools in experimental macrophage pharmacology and cell biology as well as attractive candidates to develop new therapeutic approaches against macrophage-associated diseases.

  13. Unraveling Macrophage Heterogeneity in Erythroblastic Islands

    Directory of Open Access Journals (Sweden)

    Katie Giger Seu

    2017-09-01

    Full Text Available Mammalian erythropoiesis occurs within erythroblastic islands (EBIs, niches where maturing erythroblasts interact closely with a central macrophage. While it is generally accepted that EBI macrophages play an important role in erythropoiesis, thorough investigation of the mechanisms by which they support erythropoiesis is limited largely by inability to identify and isolate the specific macrophage sub-population that constitute the EBI. Early studies utilized immunohistochemistry or immunofluorescence to study EBI morphology and structure, while more recent efforts have used flow cytometry for high-throughput quantitative characterization of EBIs and their central macrophages. However, these approaches based on the expectation that EBI macrophages are a homogeneous population (F4/80+/CD169+/VCAM-1+ for example provide an incomplete picture and potentially overlook critical information about the nature and biology of the islands and their central macrophages. Here, we present a novel method for analysis of EBI macrophages from hematopoietic tissues of mice and rats using multispectral imaging flow cytometry (IFC, which combines the high-throughput advantage of flow cytometry with the morphological and fluorescence features derived from microscopy. This method provides both quantitative analysis of EBIs, as well as structural and morphological details of the central macrophages and associated cells. Importantly, the images, combined with quantitative software features, can be used to evaluate co-expression of phenotypic markers which is crucial since some antigens used to identify macrophages (e.g., F4/80 and CD11b can be expressed on non-erythroid cells associated with the islands instead of, or in addition to the central macrophage itself. We have used this method to analyze native EBIs from different hematopoietic tissues and evaluated the expression of several markers that have been previously reported to be expressed on EBI macrophages. We

  14. Caprine arthritis encephalitis virus dysregulates the expression of cytokines in macrophages.

    Science.gov (United States)

    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

  15. Adoptive transfer of M2 macrophages reduces neuropathic pain via opioid peptides.

    Science.gov (United States)

    Pannell, Maria; Labuz, Dominika; Celik, Melih Ö; Keye, Jacqueline; Batra, Arvind; Siegmund, Britta; Machelska, Halina

    2016-10-07

    During the inflammation which occurs following nerve damage, macrophages are recruited to the site of injury. Phenotypic diversity is a hallmark of the macrophage lineage and includes pro-inflammatory M1 and anti-inflammatory M2 populations. Our aim in this study was to investigate the ability of polarized M0, M1, and M2 macrophages to secrete opioid peptides and to examine their relative contribution to the modulation of neuropathic pain. Mouse bone marrow-derived cells were cultured as unstimulated M0 macrophages or were stimulated into an M1 phenotype using lipopolysaccharide and interferon-γ or into an M2 phenotype using interleukin-4. The macrophage phenotypes were verified using flow cytometry for surface marker analysis and cytokine bead array for cytokine profile assessment. Opioid peptide levels were measured by radioimmunoassay and enzyme immunoassay. As a model of neuropathic pain, a chronic constriction injury (CCI) of the sciatic nerve was employed. Polarized M0, M1, and M2 macrophages (5 × 10 5 cells) were injected perineurally twice, on days 14 and 15 following CCI or sham surgery. Mechanical and heat sensitivity were measured using the von Frey and Hargreaves tests, respectively. To track the injected macrophages, we also transferred fluorescently stained polarized cells and analyzed the surface marker profile of endogenous and injected cells in the nerves ex vivo. Compared to M0 and M1 cells, M2 macrophages contained and released higher amounts of opioid peptides, including Met-enkephalin, dynorphin A (1-17), and β-endorphin. M2 cells transferred perineurally at the nerve injury site reduced mechanical, but not heat hypersensitivity following the second injection. The analgesic effect was reversed by the perineurally applied opioid receptor antagonist naloxone methiodide. M2 cells did not affect sensitivity following sham surgery. Neither M0 nor M1 cells altered mechanical and heat sensitivity in CCI or sham-operated animals. Tracing the

  16. Triamcinolone acetonide activates an anti-inflammatory and folate receptor-positive macrophage that prevents osteophytosis in vivo.

    Science.gov (United States)

    Siebelt, Michiel; Korthagen, Nicoline; Wei, Wu; Groen, Harald; Bastiaansen-Jenniskens, Yvonne; Müller, Christina; Waarsing, Jan Hendrik; de Jong, Marion; Weinans, Harrie

    2015-12-05

    Triamcinolone acetonide (TA) is used for osteoarthritis management to reduce pain, and pre-clinical studies have shown that TA limits osteophyte formation. Osteophyte formation is known to be facilitated by synovial macrophage activation. TA injections might influence macrophage activation and subsequently reduce osteophytosis. Although widely applied in clinical care, the mechanism through which TA exerts this effect remains unknown. In this animal study, we investigated the in vivo effects of TA injections on macrophage activation, osteophyte development and joint degeneration. Furthermore, in vitro macrophage differentiation experiments were conducted to further explain working mechanisms of TA effects found in vivo. Osteoarthritis was induced in rat knees using papain injections and a running protocol. Untreated and TA-treated animals were longitudinally monitored for 12 weeks with in vivo micro-computed tomography (μCT) to measure subchondral bone changes. Synovial macrophage activation was measured in vivo using folate receptor β (FRβ)-targeted single-photon emission computed tomography/computed tomography. Articular cartilage was analyzed at 6 and 12 weeks with ex vivo contrast-enhanced μCT and histology. To further explain the outcomes of our in vivo study, TA on macrophages was also studied in vitro. These cultured macrophages were either M1- or M2-activated, and they were analyzed using fluorescence-activated cell sorting for CD163 and FRβ expression as well as for messenger RNA (mRNA) expression of interleukin (IL)-10. Our in vivo study showed that intra-articular injections with TA strongly enhanced FRβ(+) macrophage activation. Despite stimulated macrophage activation, osteophyte formation was fully prevented. There was no beneficial effect of TA against cartilage degradation or subchondral bone sclerosis. In vitro macrophage cultures showed that TA strongly induced monocyte differentiation towards CD163(+) and FRβ(+) macrophages. Furthermore

  17. Suppressive effects of ketamine on macrophage functions

    International Nuclear Information System (INIS)

    Chang Yi; Chen, T.-L.; Sheu, J.-R.; Chen, R.-M.

    2005-01-01

    Ketamine is an intravenous anesthetic agent. Clinically, induction of anesthesia with ketamine can cause immunosuppression. Macrophages play important roles in host defense. In this study, we attempted to evaluate the effects of ketamine on macrophage functions and its possible mechanism using mouse macrophage-like Raw 264.7 cells as the experimental model. Exposure of macrophages to 10 and 100 μM ketamine, which correspond to 0.1 and 1 times the clinically relevant concentration, for 1, 6, and 24 h had no effect on cell viability or lactate dehydrogenase release. When the administered concentration reached 1000 μM, ketamine caused a release of lactate dehydrogenase and cell death. Ketamine, at 10 and 100 μM, did not affect the chemotactic activity of macrophages. Administration of 1000 μM ketamine in macrophages resulted in a decrease in cell migration. Treatment of macrophages with ketamine reduced phagocytic activities. The oxidative ability of macrophages was suppressed by ketamine. Treatment with lipopolysaccharide induced TNF-α, IL-1β, and IL-6 mRNA in macrophages. Administration of ketamine alone did not influence TNF-α, IL-1β, or IL-6 mRNA production. Meanwhile, cotreatment with ketamine and lipopolysaccharide significantly inhibited lipopolysaccharide-induced TNF-α, IL-1β, and IL-6 mRNA levels. Exposure to ketamine led to a decrease in the mitochondrial membrane potential. However, the activity of mitochondrial complex I NADH dehydrogenase was not affected by ketamine. This study shows that a clinically relevant concentration of ketamine (100 μM) can suppress macrophage function of phagocytosis, its oxidative ability, and inflammatory cytokine production possibly via reduction of the mitochondrial membrane potential instead of direct cellular toxicity

  18. Inhibition of P-glycoprotein by HIV protease inhibitors increases intracellular accumulation of berberine in murine and human macrophages.

    Directory of Open Access Journals (Sweden)

    Weibin Zha

    Full Text Available HIV protease inhibitor (PI-induced inflammatory response in macrophages is a major risk factor for cardiovascular diseases. We have previously reported that berberine (BBR, a traditional herbal medicine, prevents HIV PI-induced inflammatory response through inhibiting endoplasmic reticulum (ER stress in macrophages. We also found that HIV PIs significantly increased the intracellular concentrations of BBR in macrophages. However, the underlying mechanisms of HIV PI-induced BBR accumulation are unknown. This study examined the role of P-glycoprotein (P-gp in HIV PI-mediated accumulation of BBR in macrophages.Cultured mouse RAW264.7 macrophages, human THP-1-derived macrophages, Wild type MDCK (MDCK/WT and human P-gp transfected (MDCK/P-gp cells were used in this study. The intracellular concentration of BBR was determined by HPLC. The activity of P-gp was assessed by measuring digoxin and rhodamine 123 (Rh123 efflux. The interaction between P-gp and BBR or HIV PIs was predicated by Glide docking using Schrodinger program. The results indicate that P-gp contributed to the efflux of BBR in macrophages. HIV PIs significantly increased BBR concentrations in macrophages; however, BBR did not alter cellular HIV PI concentrations. Although HIV PIs did not affect P-gp expression, P-gp transport activities were significantly inhibited in HIV PI-treated macrophages. Furthermore, the molecular docking study suggests that both HIV PIs and BBR fit the binding pocket of P-gp, and HIV PIs may compete with BBR to bind P-gp.HIV PIs increase the concentration of BBR by modulating the transport activity of P-gp in macrophages. Understanding the cellular mechanisms of potential drug-drug interactions is critical prior to applying successful combinational therapy in the clinic.

  19. Improved survival of mesenchymal stem cells by macrophage migration inhibitory factor

    OpenAIRE

    Xia, Wenzheng; Xie, Congying; Jiang, Miaomiao; Hou, Meng

    2015-01-01

    Macrophage migration inhibitory factor (MIF) is a critical inflammatory cytokine that was recently associated with progenitor cell survival and potently inhibits apoptosis. We examined the protective effect of MIF on hypoxia/serum deprivation (SD)-induced apoptosis of mesenchymal stem cells (MSCs), as well as the possible mechanisms. MSCs were obtained from rat bone marrow and cultured in vitro. Apoptosis was induced by culturing MSCs under hypoxia/SD conditions for up to 24?h and assessed by...

  20. Suppression of annexin A2 by prostaglandin E₂ impairs phagocytic ability of peritoneal macrophages in women with endometriosis.

    Science.gov (United States)

    Wu, Meng-Hsing; Chuang, Pei-Chin; Lin, Yiu-Juian; Tsai, Shaw-Jenq

    2013-04-01

    Is annexin A2 involved in the reduced phagocytic ability of macrophages in endometriosis? Data from women with endometriosis and a murine model of the disease show that expression of annexin A2 in peritoneal macrophages is inhibited by prostaglandin E2 (PGE2) and this impairs the phagocytic ability of macrophages. Endometriosis is a chronic inflammatory disease that recruits many immune cells, especially macrophages, to the peritoneal cavity. The phagocytic ability of peritoneal macrophages isolated from women with endometriosis is reduced. A laboratory study. Thirty-five patients (20 with and 15 without endometriosis) of reproductive age with normal menstrual cycles were recruited. Peritoneal macrophages isolated from women with or without endometriosis were cultured and treated with vehicle, PGE2 and different EP receptor agonists, and the expression of annexin A2 was quantified by RT-PCR and western blotting. Annexin A2 was knocked down (by small interfering RNA) in normal macrophages or overexpressed (by treatment with recombinant protein) in endometriotic macrophages and their phagocytic ability was measured by flow cytometry. Peritoneal macrophages were isolated from a mouse model of endometriosis and treated with PGE2 or cyclo-oxygenase (COX) inhibitors, and annexin A2 mRNA was quantified. Levels of annexin A2 were markedly reduced in peritoneal macrophages from women with endometriosis versus controls (mRNA: P endometriosis versus control) via the EP2/EP4 receptor-dependent signaling pathway. Treatment with PGE2 or knockdown of annexin A2 inhibited the phagocytic ability of macrophages (P peritoneal macrophages were markedly reduced in mice treated with PGE2 (P peritoneal macrophages (P peritoneal cells from patients with endometriosis or that their endometriotic fluid contains increased amounts of PGE2 when compared with control subjects. Inhibiting PGE2 signaling, in order to restore or enhance the phagocytic capability of macrophages, may represent a new

  1. Susceptibility of bone marrow-derived macrophages to influenza virus infection is dependent on macrophage phenotype.

    Science.gov (United States)

    Campbell, Gillian M; Nicol, Marlynne Q; Dransfield, Ian; Shaw, Darren J; Nash, Anthony A; Dutia, Bernadette M

    2015-10-01

    The role of the macrophage in influenza virus infection is complex. Macrophages are critical for resolution of influenza virus infections but implicated in morbidity and mortality in severe infections. They can be infected with influenza virus and consequently macrophage infection is likely to have an impact on the host immune response. Macrophages display a range of functional phenotypes, from the prototypical pro-inflammatory classically activated cell to alternatively activated anti-inflammatory macrophages involved in immune regulation and wound healing. We were interested in how macrophages of different phenotype respond to influenza virus infection and therefore studied the infection of bone marrow-derived macrophages (BMDMs) of classical and alternative phenotype in vitro. Our results show that alternatively activated macrophages are more readily infected and killed by the virus than classically activated. Classically activated BMDMs express the pro-inflammatory markers inducible nitric oxide synthase (iNOS) and TNF-α, and TNF-α expression was further upregulated following infection. Alternatively activated macrophages express Arginase-1 and CD206; however, following infection, expression of these markers was downregulated whilst expression of iNOS and TNF-α was upregulated. Thus, infection can override the anti-inflammatory state of alternatively activated macrophages. Importantly, however, this results in lower levels of pro-inflammatory markers than those produced by classically activated cells. Our results showed that macrophage phenotype affects the inflammatory macrophage response following infection, and indicated that modulating the macrophage phenotype may provide a route to develop novel strategies to prevent and treat influenza virus infection.

  2. HIV-1 Latency in Monocytes/Macrophages

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    Amit Kumar

    2014-04-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 targets CD4+ T cells and cells of the monocyte/macrophage lineage. HIV pathogenesis is characterized by the depletion of T lymphocytes and by the presence of a population of cells in which latency has been established called the HIV-1 reservoir. Highly active antiretroviral therapy (HAART has significantly improved the life of HIV-1 infected patients. However, complete eradication of HIV-1 from infected individuals is not possible without targeting latent sources of infection. HIV-1 establishes latent infection in resting CD4+ T cells and findings indicate that latency can also be established in the cells of monocyte/macrophage lineage. Monocyte/macrophage lineage includes among others, monocytes, macrophages and brain resident macrophages. These cells are relatively more resistant to apoptosis induced by HIV-1, thus are important stable hideouts of the virus. Much effort has been made in the direction of eliminating HIV-1 resting CD4+ T-cell reservoirs. However, it is impossible to achieve a cure for HIV-1 without considering these neglected latent reservoirs, the cells of monocyte/macrophage lineage. In this review we will describe our current understanding of the mechanism of latency in monocyte/macrophage lineage and how such cells can be specifically eliminated from the infected host.

  3. Inflammatory Macrophages Promotes Development of Diabetic Encephalopathy.

    Science.gov (United States)

    Wang, Beiyun; Miao, Ya; Zhao, Zhe; Zhong, Yuan

    2015-01-01

    Diabetes and Alzheimer's disease are often associated with each other, whereas the relationship between two diseases is ill-defined. Although hyperglycemia during diabetes is a major cause of encephalopathy, diabetes may also cause chronic inflammatory complications including peripheral neuropathy. Hence the role and the characteristics of inflammatory macrophages in the development of diabetic encephalopathy need to be clarified. Diabetes were induced in mice by i.p. injection of streptozotocin (STZ). Two weeks after STZ injection and confirmation of development of diabetes, inflammatory macrophages were eliminated by i.p. injection of 20µg saporin-conjugated antibody against a macrophage surface marker CD11b (saporin-CD11b) twice per week, while a STZ-treated group received injection of rat IgG of same frequency as a control. The effects of macrophage depletion on brain degradation markers, brain malondialdehyde (MDA), catalase, superoxidase anion-positive cells and nitric oxide (NO) were measured. Saporin-CD11b significantly reduced inflammatory macrophages in brain, without affecting mouse blood glucose, serum insulin, glucose responses and beta cell mass. However, reduced brain macrophages significantly inhibited the STZ-induced decreases in brain MDA, catalase and superoxidase anion-positive cells, and the STZ-induced decreases in brain NO. Inflammatory macrophages may promote development of diabetic encephalopathy. © 2015 S. Karger AG, Basel.

  4. Chondroitin-6-sulfate attenuates inflammatory responses in murine macrophages via suppression of NF-κB nuclear translocation.

    Science.gov (United States)

    Tan, Guak-Kim; Tabata, Yasuhiko

    2014-06-01

    Inflammation is a host protective response to noxious stimuli, and excessive production of pro-inflammatory mediators by macrophages (mφ) can lead to numerous pathological conditions. In this study, immunomodulatory effects of immobilized and soluble glycosaminoglycans (GAGs) on mouse-bone-marrow-derived mφ were compared by measuring nitric oxide (NO). We demonstrate here that all GAGs studied except for heparin were able to modulate interferon-γ/lipopolysaccharide (IFN-γ/LPS)-induced NO release by mφ to varying extents after 24h of incubation. In particular, the modulatory activities of soluble chondroitin-6-sulfate (C6S), hyaluronic acid and heparan sulfate altered markedly after covalent immobilization. Of these, soluble C6S exhibited the strongest NO inhibitory activity, and the inhibition was dose- and time-dependent. Moreover, C6S significantly reduced pro-inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α production by IFN-γ/LPS- or LPS-activated mφ. Specifically, the C6S-mediated suppression of mφ pro-inflammatory phenotype was accompanied by an increase in the IL-10 level, suggesting a possible switch towards anti-inflammatory/wound healing M2 state. In addition, the highest magnitude of inhibitory effects was obtained when cells were pre-treated with C6S prior to IFN-γ/LPS or LPS challenge, suggesting an additional role for C6S in protection against microbial infection. Further investigations reveal that the anti-inflammatory effects of C6S on activated mφ may be ascribed at least in part to suppression of NF-κB nuclear translocation. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  5. Signal regulatory protein α associated with the progression of oral leukoplakia and oral squamous cell carcinoma regulates phenotype switch of macrophages.

    Science.gov (United States)

    Ye, Xiaojing; Zhang, Jing; Lu, Rui; Zhou, Gang

    2016-12-06

    Signal regulatory protein α (SIRPα) is a cell-surface protein expressed on macrophages that are regarded as an important component of the tumor microenvironment. The expression of SIRPα in oral leukoplakia (OLK) and oral squamous cell carcinoma (OSCC), and further explored the role of SIRPα on the phenotype, phagocytosis ability, migration, and invasion of macrophages in OSCC were investigated. The expression of SIRPα in OLK was higher than in OSCC, correlating with the expression of CD68 and CD163 on macrophages. After cultured with the conditioned media of oral cancer cells, the expression of SIRPα on THP-1 cells was decreased gradually. In co-culture system, macrophages were induced into M2 phenotype by oral cancer cells. Blockade of SIRPα inhibited phagocytosis ability and IL-6, TNF-α productions of macrophages. In addition, the proliferation, migration, and IL-10, TGF-β productions of macrophages were upregulated after blockade of SIRPα. Macrophages upregulated the expression of SIRPα and phagocytosis ability, and inhibited the migration and invasion when the activation of NF-κB was inhibited by pyrrolidine dithiocarbamate ammonium (PDTC). Hence, SIRPα might play an important role in the progression of OLK and oral cancer, and could be a pivotal therapeutic target in OSCC by regulating the phenotype of macrophages via targeting NF-κB.

  6. Stimulation of mast cells leads to cholesterol accumulation in macrophages in vitro by a mast cell granule-mediated uptake of low density lipoprotein

    International Nuclear Information System (INIS)

    Kokkonen, J.O.; Kovanen, P.T.

    1987-01-01

    The uptake of low density lipoprotein (LDL) by cultured mouse macrophages was markedly promoted by isolated rat mast cell granules present in the culture medium. The granule-mediated uptake of 125 I-LDL enhanced the rate of cholesteryl ester synthesis in the macrophages, the result being accumulation of cholesteryl esters in these cells. Binding of LDL to the granules was essential for the granule-mediated uptake of LDL by macrophages, for the uptake process was prevented by treating the granules with avidin or protamine chloride or by treating LDL with 1,2-cyclohexanedione, all of which inhibit the binding of LDL to the granules. Inhibition of granule phagocytosis by the macrophages with cytochalasin B also abolished the granule-mediated uptake of LDL. Finally, mouse macrophage monolayers and LDL were incubated in the presence of isolated rat serosal mast cells. Stimulation of the mast cells with compound 48/80, a degranulating agent, resulted in dose-dependent release of secretory granules from the mast cells and a parallel increase in 14 C cholesteryl ester synthesis in the macrophages. The results show that, in this in vitro model, the sequence of events leading to accumulation of cholesteryl esters in macrophages involves initial stimulation of mast cells, subsequent release of their secretory granules, binding of LDL to the exocytosed granules, and, finally, phagocytosis of the LDL-containing granules by macrophages

  7. Mycobacterium avium MAV2052 protein induces apoptosis in murine macrophage cells through Toll-like receptor 4.

    Science.gov (United States)

    Lee, Kang-In; Choi, Han-Gyu; Son, Yeo-Jin; Whang, Jake; Kim, Kwangwook; Jeon, Heat Sal; Park, Hye-Soo; Back, Yong Woo; Choi, Seunga; Kim, Seong-Woo; Choi, Chul Hee; Kim, Hwa-Jung

    2016-04-01

    Mycobacterium avium and its sonic extracts induce apoptosis in macrophages. However, little is known about the M. avium components regulating macrophage apoptosis. In this study, using multidimensional fractionation, we identified MAV2052 protein, which induced macrophage apoptosis in M. avium culture filtrates. The recombinant MAV2052 induced macrophage apoptosis in a caspase-dependent manner. The loss of mitochondrial transmembrane potential (ΔΨm), mitochondrial translocation of Bax, and release of cytochrome c from mitochondria were observed in macrophages treated with MAV2052. Further, reactive oxygen species (ROS) production was required for the apoptosis induced by MAV2052. In addition, ROS and mitogen-activated protein kinases were involved in MAV2052-mediated TNF-α and IL-6 production. ROS-mediated activation of apoptosis signal-regulating kinase 1 (ASK1)-JNK pathway was a major signaling pathway for MAV2052-induced apoptosis. Moreover, MAV2052 bound to Toll-like receptor (TLR) 4 molecule and MAV2052-induced ROS production, ΔΨm loss, and apoptosis were all significantly reduced in TLR4(-/-) macrophages. Altogether, our results suggest that MAV2052 induces apoptotic cell death through TLR4 dependent ROS production and JNK pathway in murine macrophages.

  8. In vitro and in vivo models of cerebral ischemia show discrepancy in therapeutic effects of M2 macrophages.

    Directory of Open Access Journals (Sweden)

    Virginie Desestret

    Full Text Available THE INFLAMMATORY RESPONSE FOLLOWING ISCHEMIC STROKE IS DOMINATED BY INNATE IMMUNE CELLS: resident microglia and blood-derived macrophages. The ambivalent role of these cells in stroke outcome might be explained in part by the acquisition of distinct functional phenotypes: classically (M1 and alternatively activated (M2 macrophages. To shed light on the crosstalk between hypoxic neurons and macrophages, an in vitro model was set up in which bone marrow-derived macrophages were co-cultured with hippocampal slices subjected to oxygen and glucose deprivation. The results showed that macrophages provided potent protection against neuron cell loss through a paracrine mechanism, and that they expressed M2-type alternative polarization. These findings raised the possibility of using bone marrow-derived M2 macrophages in cellular therapy for stroke. Therefore, 2 million M2 macrophages (or vehicle were intravenously administered during the subacute stage of ischemia (D4 in a model of transient middle cerebral artery occlusion. Functional neuroscores and magnetic resonance imaging endpoints (infarct volumes, blood-brain barrier integrity, phagocytic activity assessed by iron oxide uptake were longitudinally monitored for 2 weeks. This cell-based treatment did not significantly improve any outcome measure compared with vehicle, suggesting that this strategy is not relevant to stroke therapy.

  9. Macrophages and Uveitis in Experimental Animal Models

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    Salvador Mérida

    2015-01-01

    Full Text Available Resident and infiltrated macrophages play relevant roles in uveitis as effectors of innate immunity and inductors of acquired immunity. They are major effectors of tissue damage in uveitis and are also considered to be potent antigen-presenting cells. In the last few years, experimental animal models of uveitis have enabled us to enhance our understanding of the leading role of macrophages in eye inflammation processes, including macrophage polarization in experimental autoimmune uveoretinitis and the major role of Toll-like receptor 4 in endotoxin-induced uveitis. This improved knowledge should guide advantageous iterative research to establish mechanisms and possible therapeutic targets for human uveitis resolution.

  10. Induction of heat-shock proteins and phagocytic function of chicken macrophage following in vitro heat exposure

    International Nuclear Information System (INIS)

    Miller, L.; Qureshi, M.A.

    1992-01-01

    The protein profiles and phagocytic ability of Sephadex-elicited chicken peritoneal macrophages were examined following heat-shock exposure. Macrophage cultures were exposed to various temperatures, time exposures and recovery periods. Densitometric analysis of SDS-PAGE autoradiographs revealed that heat-induced macrophages synthesized three major (23, 70 and 90 kD) heat-shock proteins (HSPs). The optimal temperature and time for induction of these HSPs was 45-46 degrees C for 1 h, with a variable recovery period for each HSP. Macrophages exposed to 45 degrees C for 30 and 60 min were significantly depressed in phagocytosis of uncoated sheep erythrocytes (SE) under 45 degrees C incubation conditions. However, phagocytosis of antibody-coated SE was not affected when compared to 41 degrees C control cultures. Macrophages allowed to recover at 41 degrees C following heat-shock exhibited no alterations in their phagocytic ability for either antibody-coated or uncoated SE. This study suggests that heat shock induces three major HSPs in chicken peritoneal macrophages in addition to maintaining their Fc-mediated phagocytic function while significantly depressing their nonspecific phagocytosis

  11. Intratumoral macrophages contribute to epithelial-mesenchymal transition in solid tumors

    International Nuclear Information System (INIS)

    Bonde, Anne-Katrine; Tischler, Verena; Kumar, Sushil; Soltermann, Alex; Schwendener, Reto A

    2012-01-01

    Several stromal cell subtypes including macrophages contribute to tumor progression by inducing epithelial-mesenchymal transition (EMT) at the invasive front, a mechanism also linked to metastasis. Tumor associated macrophages (TAM) reside mainly at the invasive front but they also infiltrate tumors and in this process they mainly assume a tumor promoting phenotype. In this study, we asked if TAMs also regulate EMT intratumorally. We found that TAMs through TGF-β signaling and activation of the β-catenin pathway can induce EMT in intratumoral cancer cells. We depleted macrophages in F9-teratocarcinoma bearing mice using clodronate-liposomes and analyzed the tumors for correlations between gene and protein expression of EMT-associated and macrophage markers. The functional relationship between TAMs and EMT was characterized in vitro in the murine F9 and mammary gland NMuMG cells, using a conditioned medium culture approach. The clinical relevance of our findings was evaluated on a tissue microarray cohort representing 491 patients with non-small cell lung cancer (NSCLC). Gene expression analysis of F9-teratocarcinomas revealed a positive correlation between TAM-densities and mesenchymal marker expression. Moreover, immunohistochemistry showed that TAMs cluster with EMT phenotype cells in the tumors. In vitro, long term exposure of F9-and NMuMG-cells to macrophage-conditioned medium led to decreased expression of the epithelial adhesion protein E-cadherin, activation of the EMT-mediating β-catenin pathway, increased expression of mesenchymal markers and an invasive phenotype. In a candidate based screen, macrophage-derived TGF-β was identified as the main inducer of this EMT-associated phenotype. Lastly, immunohistochemical analysis of NSCLC patient samples identified a positive correlation between intratumoral macrophage densities, EMT markers, intraepithelial TGF-β levels and tumor grade. Data presented here identify a novel role for macrophages in EMT

  12. The neuropathological basis to the functional role of microglia/macrophages in gliomas.

    Science.gov (United States)

    Schiffer, Davide; Mellai, Marta; Bovio, Enrica; Annovazzi, Laura

    2017-09-01

    The paper wants to be a tracking shot of the main recent acquisitions on the function and significance of microglia/macrophages in gliomas. The observations have been principally carried out on in vitro cultures and on tumor transplants in animals. Contrary to what is deduced from microglia in non-neoplastic pathologic conditions of central nervous system (CNS), most conclusions indicate that microglia acts favoring tumor proliferation through an immunosuppression induced by glioma cells. By immunohistochemistry, different microglia phenotypes are recognized in gliomas, from ramified microglia to frank macrophagic aspect. One wonders whether the functional conclusions drawn from many microglia studies, but not in conditions of human pathology, apply to all the phenotypes recognizable in them. It is difficult to verify in human pathology a prognostic significance of microglia. Only CD163-positive microglia/macrophages inversely correlate with glioma patients' survival, whereas the total number of microglia does not change with the malignancy grade.

  13. Lineage-specific enhancers activate self-renewal genes in macrophages and embryonic stem cells.

    Science.gov (United States)

    Soucie, Erinn L; Weng, Ziming; Geirsdóttir, Laufey; Molawi, Kaaweh; Maurizio, Julien; Fenouil, Romain; Mossadegh-Keller, Noushine; Gimenez, Gregory; VanHille, Laurent; Beniazza, Meryam; Favret, Jeremy; Berruyer, Carole; Perrin, Pierre; Hacohen, Nir; Andrau, J-C; Ferrier, Pierre; Dubreuil, Patrice; Sidow, Arend; Sieweke, Michael H

    2016-02-12

    Differentiated macrophages can self-renew in tissues and expand long term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network that controls self-renewal. Single-cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells. Copyright © 2016, American Association for the Advancement of Science.

  14. Activation of Alveolar Macrophages after Plutonium Oxide Inhalation in Rats: Involvement in the Early Inflammatory Response

    Energy Technology Data Exchange (ETDEWEB)

    Van der Meeren, A.; Tourdes, F.; Gremy, O.; Grillon, G.; Abram, M.C.; Poncy, J.L.; Griffiths, N. [CEA, DSV, DRR, SRCA, Centre DAM Ile de France, F-91297 Bruyeres Le Chatel, Arpajon (France)

    2008-07-01

    Alveolar macrophages play an important role in the distribution, clearance and inflammatory reactions after particle inhalation, which may influence long-term events such as fibrosis and tumorigenesis. The objectives of the present study were to investigate the early inflammatory events after plutonium oxide inhalation in rats and involvement of alveolar macrophages. Lung changes were studied from 3 days to 3 months after inhalation of PuO{sub 2} or different isotopic compositions (70% or 97% {sup 239}Pu) and initial lung deposits (range 2.1 to 43.4 kBq/rat). Analyses of bronchoalveolar lavages showed early increases in the numbers of granulocytes, lymphocytes and multi-nucleated macrophages. The activation of macrophages was evaluated ex vivo by measurement of inflammatory mediator levels in culture supernatants. TNF-alpha and chemokine MCP-1, MIP-2 and CINC-1 production was elevated from 7 days after inhalation and remained so up to 3 months. In contrast, IL-1 beta, IL-6 and IL-10 production was unchanged. At 6 weeks, pulmonary macrophage numbers and activation state were increased as observed from an immunohistochemistry study of lung sections with anti-ED1. Similarly, histological analyses of lung sections also showed evidence of inflammatory responses. In conclusion, our results indicate early inflammatory changes in the lungs of PuO{sub 2}-contaminated animals and the involvement of macrophages in this process. A dose-effect relationship was observed between the amount of radionuclide inhaled or retained at the time of analysis and inflammatory mediator production by alveolar macrophages 14 days after exposure. For similar initial lung deposits, the inflammatory manifestation appears higher for 97% {sup 239}Pu than for 70% {sup 239}Pu. (authors)

  15. Effect of Tityus serrulatus venom on cytokine production and the activity of murine macrophages

    Directory of Open Access Journals (Sweden)

    Vera L. Petricevich

    2002-01-01

    Full Text Available The purpose of this study was to investigate the effects of Tityus serrulatus venom (TSV on murine peritoneal macrophages evaluated in terms of activation. The effects of crude TSV were analysed by detection of cytokines, oxygen intermediate metabolites (H2O2 and nitric oxide (NO in supernatants of peritoneal macrophages. Several functional bioassays were employed including an in vitro model for envenomating: cytotoxicity of TSV was assessed using the lyses percentage. Tumor necrosis factor (TNF activity was assayed by measuring its cytotoxic activity on L-929 cells, and interleukin-6 (IL-6 and interferon-γ (IFN-γ were assayed by enzyme-linked immunosorbent assay, whereas NO levels were detected by Griess colorimetric reactions in culture supernatant of macrophages incubated with TSV and subsequently exposed to either lipopolysaccharide or IFN-γ. Incubation of macrophages with TSV increased production of IL-6 and IFN-γ in a dose-dependent manner. TNF production was not detected in supernatants treated with TSV at any concentration. The increase in IL-6 secretion was not associated with concentration-dependent cytoxicity of TSV on these cells. These data suggest that the cytotoxicity does not appear to be the main cause of an increased cytokine production by these cells. Although NO is an important effector molecule in macrophage microbicidal activity, the inducing potential of the test compounds for its release was found to be very moderate, ranging from 125 to 800 mM. Interestingly, NO levels of peritoneal macrophages were increased after IFN-γ. Moreover, NO production had an apparent effect on macrophage activity. The results obtained here also shown that the TSV induces an important elevation in H2O2 release. These results combined with NO production suggest that TSV possesses significant immunomodulatory activities capable of stimulating immune functions in vitro.

  16. Macrophage proinflammatory response to the titanium alloy equipment in dental implantation.

    Science.gov (United States)

    Chen, X; Li, H S; Yin, Y; Feng, Y; Tan, X W

    2015-08-07

    Titanium alloy and stainless steel (SS) had been widely used as dental implant materials because of their affinity with epithelial tissue and connective tissue, and good physical, chemical, biological, mechanical properties and processability. We compared the effects of titanium alloy and SS on macrophage cytokine expression as well as their biocompatibility. Mouse macrophage RAW264.7 cells were cultured on titanium alloy and SS surfaces. Cells were counted by scanning electron microscopy. A nitride oxide kit was used to detect released nitric oxide by macrophages on the different materials. An enzyme linked immunosorbent assay was used to detect monocyte chemoattractant protein-1 levels. Scanning electron microscopy revealed fewer macrophages on the surface of titanium alloy (48.2 ± 6.4 x 10(3) cells/cm(2)) than on SS (135 ± 7.3 x 10(3) cells/cm(2)). The nitric oxide content stimulated by titanium alloy was 22.5 mM, which was lower than that stimulated by SS (26.8 mM), but the difference was not statistically significant (P = 0.07). The level of monocyte chemoattractant protein-1 released was significantly higher in the SS group (OD value = 0.128) than in the titanium alloy group (OD value = 0.081) (P = 0.024). The transforming growth factor-b1 mRNA expression levels in macrophages after stimulation by titanium alloy for 12 and 36 h were significantly higher than that after stimulation by SS (P = 0.31 and 0.25, respectively). Macrophages participate in the inflammatory response by regulating cytokines such as nitric oxide, monocyte chemoattractant protein-1, and transforming growth factor-b1. There were fewer macrophages and lower inflammation on the titanium alloy surface than on the SS surface. Titanium alloy materials exhibited better biological compatibility than did SS.

  17. The response of macrophages to titanium particles is determined by macrophage polarization.

    Science.gov (United States)

    Pajarinen, Jukka; Kouri, Vesa-Petteri; Jämsen, Eemeli; Li, Tian-Fang; Mandelin, Jami; Konttinen, Yrjö T

    2013-11-01

    Aseptic loosening of total joint replacements is driven by the reaction of macrophages to foreign body particles released from the implant. It was hypothesized that the macrophages' response to these particles is dependent, in addition to particle characteristics and contaminating biomolecules, on the state of macrophage polarization as determined by the local cytokine microenvironment. To test this hypothesis we differentiated M1 and M2 macrophages from human peripheral blood monocytes and compared their responses to titanium particles using genome-wide microarray analysis and a multiplex cytokine assay. In comparison to non-activated M0 macrophages, the overall chemotactic and inflammatory responses to titanium particles were greatly enhanced in M1 macrophages and effectively suppressed in M2 macrophages. In addition, the genome-wide approach revealed several novel, potentially osteolytic, particle-induced mediators, and signaling pathway analysis suggested the involvement of toll-like and nod-like receptor signaling in particle recognition. It is concluded that the magnitude of foreign body reaction caused by titanium particles is dependent on the state of macrophage polarization. Thus, by limiting the action of M1 polarizing factors, e.g. bacterial biofilm formation, in peri-implant tissues and promoting M2 macrophage polarization by biomaterial solutions or pharmacologically, it might be possible to restrict wear-particle-induced inflammation and osteolysis. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  18. The anti-inflammatory effects of the tellurium redox modulating compound, AS101, are associated with regulation of NFκB signaling pathway and nitric oxide induction in macrophages

    Directory of Open Access Journals (Sweden)

    Sredni Benjamin

    2010-01-01

    Full Text Available Abstract Background LPS-activated macrophages produce mediators which are involved in inflammation and tissue injury, and especially those associated with endotoxic shock. The non toxic tellurium compound ammonium tri-chloro(dioxoethylene-O,O'-tellurate, AS101, has been recently shown to exert profound anti-inflammatory properties in animal models, associated with its Te(IV redox chemistry. This study explores the anti-inflammatory properties of AS101 with respect to modulation of inflammatory cytokines production and regulation of iNOS transcription and expression in activated macrophages via targeting the NFkB complex. Results AS101 decreased production of IL-6 and in parallel down-regulated LPS-induced iNOS expression and NO secretion by macrophages. AS101 reduced IkB phosphorylation and degradation, and reduced NFkB nuclear translocalization, albeit these effects were exerted at different kinetics. Chromatin immunoprecipitation assays showed that AS101 treatment attenuated p50-subunit ability to bind DNA at the NFkB consensus site in the iNOS promotor following LPS induction. Conclusions Besides AS101, the investigation of therapeutic activities of other tellurium(IV compounds is scarce in the literature, although tellurium is the fourth most abundant trace element in the human body. Since IKK and NFkB may be regulated by thiol modifications, we may thus envisage, inview of our integrated results, that Te(IV compounds, may have important roles in thiol redox biological activity in the human body and represent a new class of anti-inflammatory compounds.

  19. Botanical polysaccharides: macrophage immunomodulation and therapeutic potential.

    Science.gov (United States)

    Schepetkin, Igor A; Quinn, Mark T

    2006-03-01

    Botanical polysaccharides exhibit a number of beneficial therapeutic properties, and it is thought that the mechanisms involved in these effects are due to the modulation of innate immunity and, more specifically, macrophage function. In this review, we summarize our current state of understanding of the macrophage modulatory effects of botanical polysaccharides isolated from a wide array of different species of flora, including higher plants, mushrooms, lichens and algae. Overall, the primary effect of botanical polysaccharides is to enhance and/or activate macrophage immune responses, leading to immunomodulation, anti-tumor activity, wound-healing and other therapeutic effects. Furthermore, botanical and microbial polysaccharides bind to common surface receptors and induce similar immunomodulatory responses in macrophages, suggesting that evolutionarily conserved polysaccharide structural features are shared between these organisms. Thus, the evaluation of botanical polysaccharides provides a unique opportunity for the discovery of novel therapeutic agents and adjuvants that exhibit beneficial immunomodulatory properties.

  20. Epigenetic Regulation of Monocyte and Macrophage Function

    NARCIS (Netherlands)

    Hoeksema, Marten A.; de Winther, Menno P. J.

    2016-01-01

    Monocytes and macrophages are key players in tissue homeostasis and immune responses. Epigenetic processes tightly regulate cellular functioning in health and disease. Recent Advances: Recent technical developments have allowed detailed characterizations of the transcriptional circuitry underlying

  1. Neutrophils reduce the parasite burden in Leishmania (Leishmania amazonensis-infected macrophages.

    Directory of Open Access Journals (Sweden)

    Erico Vinícius de Souza Carmo

    2010-11-01

    Full Text Available Studies on the role of neutrophils in Leishmania infection were mainly performed with L. (L major, whereas less information is available for L. (L amazonensis. Previous results from our laboratory showed a large infiltrate of neutrophils in the site of infection in a mouse strain resistant to L. (L. amazonensis (C3H/HePas. In contrast, the susceptible strain (BALB/c displayed a predominance of macrophages harboring a high number of amastigotes and very few neutrophils. These findings led us to investigate the interaction of inflammatory neutrophils with L. (L. amazonensis-infected macrophages in vitro.Mouse peritoneal macrophages infected with L. (L. amazonensis were co-cultured with inflammatory neutrophils, and after four days, the infection was quantified microscopically. Data are representative of three experiments with similar results. The main findings were 1 intracellular parasites were efficiently destroyed in the co-cultures; 2 the leishmanicidal effect was similar when cells were obtained from mouse strains resistant (C3H/HePas or susceptible (BALB/c to L. (L. amazonensis; 3 parasite destruction did not require contact between infected macrophages and neutrophils; 4 tumor necrosis factor alpha (TNF-α, neutrophil elastase and platelet activating factor (PAF were involved with the leishmanicidal activity, and 5 destruction of the parasites did not depend on generation of oxygen or nitrogen radicals, indicating that parasite clearance did not involve the classical pathway of macrophage activation by TNF-α, as reported for other Leishmania species.The present results provide evidence that neutrophils in concert with macrophages play a previously unrecognized leishmanicidal effect on L. (L. amazonensis. We believe these findings may help to understand the mechanisms involved in innate immunity in cutaneous infection by this Leishmania species.

  2. A method for multiple sequential analyses of macrophage functions using a small single cell sample

    Directory of Open Access Journals (Sweden)

    F.R.F. Nascimento

    2003-09-01

    Full Text Available Microbial pathogens such as bacillus Calmette-Guérin (BCG induce the activation of macrophages. Activated macrophages can be characterized by the increased production of reactive oxygen and nitrogen metabolites, generated via NADPH oxidase and inducible nitric oxide synthase, respectively, and by the increased expression of major histocompatibility complex class II molecules (MHC II. Multiple microassays have been developed to measure these parameters. Usually each assay requires 2-5 x 10(5 cells per well. In some experimental conditions the number of cells is the limiting factor for the phenotypic characterization of macrophages. Here we describe a method whereby this limitation can be circumvented. Using a single 96-well microassay and a very small number of peritoneal cells obtained from C3H/HePas mice, containing as little as <=2 x 10(5 macrophages per well, we determined sequentially the oxidative burst (H2O2, nitric oxide production and MHC II (IAk expression of BCG-activated macrophages. More specifically, with 100 µl of cell suspension it was possible to quantify H2O2 release and nitric oxide production after 1 and 48 h, respectively, and IAk expression after 48 h of cell culture. In addition, this microassay is easy to perform, highly reproducible and more economical.

  3. Phytosterols Differentially Influence ABC transporter Expression, Cholesterol Efflux and Inflammatory Cytokine Secretion in Macrophage Foam Cells

    Science.gov (United States)

    Sabeva, Nadezhda S; McPhaul, Christopher M; Li, Xiangan; Cory, Theodore J.; Feola, David J.; Graf, Gregory A

    2010-01-01

    Phytosterol supplements lower low density lipoprotein (LDL) cholesterol, but accumulate in vascular lesions of patients and limit the anti-atherosclerotic effects of LDL lowering in apolipoprotein E deficient mice, suggesting that the cholesterol lowering benefit of phytosterol supplementation may not be fully realized. Individual phytosterols have cell-type specific effects that may either be beneficial or deleterious with respect to atherosclerosis, but little is known concerning their effects on macrophage function. The effects of phytosterols on ABCA1 and ABCG1 abundance, cholesterol efflux, and inflammatory cytokine secretion were determined in cultured macrophage foam cells. Among the commonly consumed phytosterols, stigmasterol increased expression of ABCA1 and ABCG1 and increased efflux of cholesterol to apolipoprotein (Apo) AI and high density lipoprotein (HDL). Campesterol and sitosterol had no effect on ABCA1 or ABCG1 levels. Sitosterol had no effect of cholesterol efflux to Apo AI or HDL, whereas campesterol had a modest, but significant reduction in cholesterol efflux to HDL in THP-1 macrophages. Whereas stigmasterol blunted aggregated LDL-induced increases in tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1β secretion, sitosterol exacerbated these effects. The presence of campesterol had no effect on agLDL-induced inflammatory cytokine secretion from THP-1 macrophages. In conclusion, the presence of stigmasterol in modified lipoproteins promoted cholesterol efflux and suppressed inflammatory cytokine secretion in response to lipid loading in macrophage foam cells. While campesterol was largely inert, the presence of sitosterol increased the proinflammatory cytokine secretion. PMID:21111593

  4. Nanoparticles Containing Curcumin Useful for Suppressing Macrophages In Vivo in Mice.

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    Chie Amano

    Full Text Available To explore a novel method using liposomes to suppress macrophages, we screened food constituents through cell culture assays. Curcumin was one of the strongest compounds exhibiting suppressive effects on macrophages. We subsequently tried various methods to prepare liposomal curcumin, and eventually succeeded in preparing liposomes with sufficient amounts of curcumin to suppress macrophages by incorporating a complex of curcumin and bovine serum albumin. The diameter of the resultant nanoparticles, the liposomes containing curcumin, ranged from 60 to 100 nm. Flow cytometric analyses revealed that after intraperitoneal administration of the liposomes containing curcumin into mice, these were incorporated mainly by macrophages positive for F4/80, CD36, and CD11b antigens. Peritoneal cells prepared from mice injected in vivo with the liposomes containing curcumin apparently decreased interleukin-6-producing activities. Major changes in body weight and survival rates in the mice were not observed after administrating the liposomes containing curcumin. These results indicate that the liposomes containing curcumin are safe and useful for the selective suppression of macrophages in vivo in mice.

  5. Microsystem to evaluate the incorporation of 3H-uridine in macrophage RNA

    International Nuclear Information System (INIS)

    Varesio, L.; Naglich, J.; Brunda, M.J.; Taramelli, D.; Eva, A.

    1981-01-01

    A method is described for the evaluation of the total 3H-uridine incorporated by macrophages in vitro into trichloroacetic acid (TCA)-precipitable material. The technique is based upon solubilization of the macrophage monolayers by guanidine-HC1, followed by TCA precipitation. The recovery of RNA into the precipitate and the reproducibility of the results were strictly dependent on the use of filtered reagents and on incubation of the TCA precipitate for 2 or more hours at 4 degree C before harvesting. Treatment with quanidine-HC1 did not affect the recovery of labeled RNA. Moreover, we observed that radioactive precipitate had the characteristics of RNA, since its recovery was sensitive to the addition of unlabeled uridine in the culture medium and to the treatment of the macrophages with inhibitors of RNA synthesis, but not of protein synthesis. Tritiated uridine incorporation in microcultures of macrophages can be assessed with this technique, by processing the cells directly in the wells. The main advantages of this procedure are: 1) the radioactivity can be measured by semiautomatic cell harvesters, 2) a small number of macrophages are required, and 3) many samples can be processed simultaneously. Overall, the technique is simple, rapid, and could be successfully adapted to study other metabolic pathways

  6. Metabolic Reprogramming of Macrophages Exposed to Silk, Poly(lactic-co-glycolic acid), and Silica Nanoparticles.

    Science.gov (United States)

    Saborano, Raquel; Wongpinyochit, Thidarat; Totten, John D; Johnston, Blair F; Seib, F Philipp; Duarte, Iola F

    2017-07-01

    Monitoring macrophage metabolism in response to nanoparticle exposure provides new insights into biological outcomes, such as inflammation or toxicity, and supports the design of tailored nanomedicines. This paper describes the metabolic signature of macrophages exposed to nanoparticles ranging in diameter from 100 to 125 nm and made from silk, poly(lactic-co-glycolic acid) or silica. Nanoparticles of this size and type are currently at various stages of preclinical and clinical development for drug delivery applications. 1 H NMR analysis of cell extracts and culture media is used to quantify the changes in the intracellular and extracellular metabolomes of macrophages in response to nanoparticle exposure. Increased glycolytic activity, an altered tricarboxylic acid cycle, and reduced ATP generation are consistent with a proinflammatory phenotype. Furthermore, amino acids possibly arising from autophagy, the creatine kinase/phosphocreatine system, and a few osmolytes and antioxidants emerge as important players in the metabolic reprogramming of macrophages exposed to nanoparticles. This metabolic signature is a common response to all nanoparticles tested; however, the direction and magnitude of some variations are clearly nanoparticle specific, indicating material-induced biological specificity. Overall, metabolic reprogramming of macrophages can be achieved with nanoparticle treatments, modulated through the choice of the material, and monitored using 1 H NMR metabolomics. © 2017 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Dose-dependent cytotoxicity of clinically relevant cobalt nanoparticles and ions on macrophages in vitro

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    Kwon, Young-Min; Xia Zhidao; Glyn-Jones, Sion; Beard, David; Gill, Harinderjit S; Murray, David W, E-mail: young-min.kwon@ndos.ox.ac.u [Nuffield Department of Orthopaedic Surgery, University of Oxford, Oxford OX3 7LD (United Kingdom)

    2009-04-15

    Despite the satisfactory short-term implant survivorship of metal-on-metal hip resurfacing arthroplasty, periprosthetic soft-tissue masses such as pseudotumours are being increasingly reported. Cytotoxic effects of cobalt or chromium have been suggested to play a role in its aetiology. The aim of this study was to investigate the effects of clinically relevant metal nanoparticles and ions on the viability of macrophages in vitro. A RAW 264.7 murine macrophage cell line was cultured in the presence of either: (1) cobalt, chromium and titanium nanoparticles sized 30-35 nm; or (2) cobalt sulphate and chromium chloride. Two methods were used to quantify cell viability: Alamar Blue assay and Live/Dead assay. The cytotoxicity was observed only with cobalt. Cobalt nanoparticles and ions demonstrated dose-dependent cytotoxic effects on macrophages in vitro: the cytotoxic concentrations of nanoparticles and ions were 1 x 10{sup 12} particles ml{sup -1} and 1000 {mu}M, respectively. The high concentration of cobalt nanoparticles required for cytotoxicity of macrophages in vitro suggests that increased production of cobalt nanoparticles in vivo, due to excessive MoM implant wear, may lead to local adverse biological effects. Therefore, cytotoxicity of high concentrations of metal nanoparticles phagocytosed by macrophages located in the periprosthetic tissues may be an important factor in pathogenesis of pseudotumours.

  8. Lack of RNase L attenuates macrophage functions.

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    Xin Yi

    Full Text Available Macrophages are one of the major cell types in innate immunity against microbial infection. It is believed that the expression of proinflammatory genes such as tumor necrosis factor-α (TNF-α, interleukin (IL-1β, IL-6, and cyclooxygenase-2 (Cox-2 by macrophages is also crucial for activation of both innate and adaptive immunities. RNase L is an interferon (IFN inducible enzyme which is highly expressed in macrophages. It has been demonstrated that RNase L regulates the expression of certain inflammatory genes. However, its role in macrophage function is largely unknown.Bone marrow-derived macrophages (BMMs were generated from RNase L(+/+and (-/- mice. The migration of BMMs was analyzed by using Transwell migration assays. Endocytosis and phagocytosis of macrophages were assessed by using fluorescein isothiocyanate (FITC-Dextran 40,000 and FITC-E. coli bacteria, respectively. The expression of inflammatory genes was determined by Western Blot and ELISA. The promoter activity of Cox-2 was measured by luciferase reporter assays.Lack of RNase L significantly decreased the migration of BMMs induced by M-CSF, but at a less extent by GM-CSF and chemokine C-C motif ligand-2 (CCL2. Interestingly, RNase L deficient BMMs showed a significant reduction of endocytic activity to FITC-Dextran 40,000, but no any obvious effect on their phagocytic activity to FITC-bacteria under the same condition. RNase L impacts the expression of certain genes related to cell migration and inflammation such as transforming growth factor (TGF-β, IL-1β, IL-10, CCL2 and Cox-2. Furthermore, the functional analysis of the Cox-2 promoter revealed that RNase L regulated the expression of Cox-2 in macrophages at its transcriptional level. Taken together, our findings provide direct evidence showing that RNase L contributes to innate immunity through regulating macrophage functions.

  9. The Probiotic Mixture VSL#3 Alters the Morphology and Secretion Profile of Both Polarized and Unpolarized Human Macrophages in a Polarization-Dependent Manner

    Science.gov (United States)

    Isidro, Raymond A.; Bonilla, Fernando J.; Pagan, Hendrick; Cruz, Myrella L.; Lopez, Pablo; Godoy, Lenin; Hernandez, Siomara; Loucil-Alicea, Raisa Y.; Rivera-Amill, Vanessa; Yamamura, Yasuhiro; Isidro, Angel A.; Appleyard, Caroline B.

    2014-01-01

    Background Patients with Inflammatory Bowel Disease (IBD), most commonly Crohn’s disease (CD) or ulcerative colitis (UC), suffer from chronic intestinal inflammation of unknown etiology. Increased proinflammatory macrophages (M1) have been documented in tissue from patients with CD. Anti-inflammatory macrophages (M2) may play a role in UC given the preponderance of Th2 cytokines in this variant of IBD. Animal and clinical studies have shown that the probiotic VSL#3 can ameliorate signs and symptoms of IBD. Although animal data suggests a modulatory effect on macrophage phenotype, the effect of VSL#3 on human macrophages remains unknown. Objective To determine the effect of the probiotic VSL#3 on the phenotype of polarized (M1/M2) and unpolarized (MΦ) human macrophages. Methods Human monocyte-derived macrophages, generated by culturing monocytes with M-CSF, were left unpolarized or were polarized towards an M1 or an M2 phenotype by culture with LPS and IFN-γ or IL-4, respectively, and were then cultured in the presence or absence of VSL#3 for 3 days. Changes in macrophage morphology were assessed. Cytokine and chemokine levels in supernatants were determined by multiplex assay. Results VSL#3 decreased the granuloma-like aggregates of M1 macrophages, increased fibroblast-like M2 macrophages, and decreased fibroblast-like MΦ macrophages. VSL#3 increased the secretion of IL-1β, IL-6, IL-10, and G-CSF by M1, M2, and MΦ macrophages. VSL#3 exposure maintained the proinflammatory phenotype of M1 macrophages, sustaining IL-12 secretion, increasing IL-23 secretion, and decreasing MDC secretion. Both VSL#3-treated M2 and MΦ macrophages secreted higher levels of anti-inflammatory and pro-healing factors such as IL-1Ra, IL-13, EGF, FGF-2, TGF-α, and VEGF, as well as proinflammatory cytokines, including IL-12 and TNF-α. Conclusion Under our experimental conditions VSL#3 induced a mixed proinflammatory and anti-inflammatory phenotype in polarized and unpolarized

  10. 'In vitro' studies on the interaction of rickettsia and macrophages. I. Effect of ultraviolet light on 'Coxiella burnetii' inactivation and macrophage enzymes: uv-inactivated 'C. burnetii'/macrophage enzymes. Interim report

    Energy Technology Data Exchange (ETDEWEB)

    Little, J.S.; Kishimoto, R.A.; Canonico, P.G.

    1979-09-04

    The inactivation of Coxiella burnetii in suspension or in cultures of guinea pig peritoneal macrophages by ultraviolet (UV) light was studied. The effect of UV treatment on the activity of macrophage organelle marker enzymes and their subsequent equilibration in linear sucrose gradients was also determined. It was shown that UV treatment of 600 microwatts/sq cm for 15 sec at a distance of 10 cm inactivated C. burnetii, either in suspension (10 to the 8th power organisms/ML) or within guinea pig peritoneal macrophages. Similar UV treatment had little effect on the activity or equilibration of macrophage organelle marker enzymes in linear sucrose gradients. However, longer exposure caused considerable inactivatioin of these enzymes.

  11. Macrophages in intestinal homeostasis and inflammation

    Science.gov (United States)

    Bain, Calum C; Mowat, Allan McI

    2014-01-01

    The intestine contains the largest pool of macrophages in the body which are essential for maintaining mucosal homeostasis in the face of the microbiota and the constant need for epithelial renewal but are also important components of protective immunity and are involved in the pathology of inflammatory bowel disease (IBD). However, defining the biological roles of intestinal macrophages has been impeded by problems in defining the phenotype and origins of different populations of myeloid cells in the mucosa. Here, we discuss how multiple parameters can be used in combination to discriminate between functionally distinct myeloid cells and discuss the roles of macrophages during homeostasis and how these may change when inflammation ensues. We also discuss the evidence that intestinal macrophages do not fit the current paradigm that tissue-resident macrophages are derived from embryonic precursors that self-renew in situ, but require constant replenishment by blood monocytes. We describe our recent work demonstrating that classical monocytes constantly enter the intestinal mucosa and how the environment dictates their subsequent fate. We believe that understanding the factors that drive intestinal macrophage development in the steady state and how these may change in response to pathogens or inflammation could provide important insights into the treatment of IBD. PMID:24942685

  12. Endometriosis, a disease of the macrophage

    Directory of Open Access Journals (Sweden)

    Annalisa eCapobianco

    2013-01-01

    Full Text Available Endometriosis, a common cause of pelvic pain and female infertility, depends on the growth of vascularised endometrial tissue at ectopic sites. Endometrial fragments reach the peritoneal cavity during the fertile years: local cues decide whether they yield endometriotic lesions. Macrophages are recruited at sites of hypoxia and tissue stress, where they clear cell debris and heme-iron and generate pro-life and pro-angiogenesis signals. Macrophages are abundant in endometriotic lesions, where are recruited and undergo alternative activation. In rodents macrophages are required for lesions to establish and to grow; bone-marrow derived Tie-2 expressing macrophages specifically contribute to lesions neovasculature, possibly because they concur to the recruitment of circulating endothelial progenitors, and sustain their survival and the integrity of the vessel wall. Macrophages sense cues (hypoxia, cell death, iron overload in the lesions and react delivering signals to restore the local homeostasis: their action represents a necessary, non-redundant step in the natural history of the disease. Endometriosis may be due to a misperception of macrophages about ectopic endometrial tissue. They perceive it as a wound, they activate programs leading to ectopic cell survival and tissue vascularization. Clearing this misperception is a critical area for the development of novel medical treatments of endometriosis, an urgent and unmet medical need.

  13. Macrophages and nerve fibres in peritoneal endometriosis.

    Science.gov (United States)

    Tran, Lu Vinh Phuc; Tokushige, Natsuko; Berbic, Marina; Markham, Robert; Fraser, Ian S

    2009-04-01

    Endometriosis is considered to be an inflammatory disease, and macrophages are the most numerous immune cells in endometriotic lesions. However, the mechanisms underlying the elevation of macrophages and their role in the pathogenesis and manifestations of endometriosis still remain unclear. The number of macrophages stained for CD68 in endometriotic lesions (n = 24) and in peritoneum distant from the lesions (n = 14) from women with endometriosis was compared with the number of macrophages in normal peritoneum from women without endometriosis (n = 18). Peritoneal lesions were also double-stained for CD68 and protein gene product 9.5 to study the relationship between macrophages and nerve fibres. The densities of macrophages in peritoneal endometriotic lesions and unaffected peritoneum from women with endometriosis were both significantly higher than that in normal peritoneum from women without endometriosis (P peritoneal lesions from women with endometriosis compared with normal peritoneum from women without endometriosis. These cells may well play roles in the growth and development of endometriotic lesions and in the generation of pain through interaction with nerve fibres.

  14. The in vitro effects of macrophages on the osteogenic capabilities of MC3T3-E1 cells encapsulated in a biomimetic poly(ethylene glycol) hydrogel.

    Science.gov (United States)

    Saleh, Leila S; Carles-Carner, Maria; Bryant, Stephanie J

    2018-04-15

    Poly(ethylene glycol) PEG-based hydrogels are promising for cell encapsulation and tissue engineering, but are known to elicit a foreign body response (FBR) in vivo. The goal of this study was to investigate the impact of the FBR, and specifically the presence of inflammatory macrophages, on encapsulated cells and their ability to synthesize new extracellular matrix. This study employed an in vitro co-culture system with murine macrophages and MC3T3-E1 pre-osteoblasts encapsulated in a bone-mimetic hydrogel, which were cultured in transwell inserts, and exposed to an inflammatory stimulant, lipopolysaccharide (LPS). The co-culture was compared to mono-cultures of the cell-laden hydrogels alone and with LPS over 28 days. Two macrophage cell sources, RAW 264.7 and primary derived, were investigated. The presence of LPS-stimulated primary macrophages led to significant changes in the cell-laden hydrogel by a 5.3-fold increase in percent apoptotic osteoblasts at day 28, 4.2-fold decrease in alkaline phosphatase activity at day 10, and 7-fold decrease in collagen deposition. The presence of LPS-stimulated RAW macrophages led to significant changes in the cell-laden hydrogel by 5-fold decrease in alkaline phosphatase activity at day 10 and 4-fold decrease in collagen deposition. Mineralization, as measured by von Kossa stain or quantified by calcium content, was not sensitive to macrophages or LPS. Elevated interleukin-6 and tumor necrosis factor-α secretion were detected in mono-cultures with LPS and co-cultures. Overall, primary macrophages had a more severe inhibitory effect on osteoblast differentiation than the macrophage cell line, with greater apoptosis and collagen I reduction. In summary, this study highlights the detrimental effects of macrophages on encapsulated cells for bone tissue engineering. Poly(ethylene glycol) (PEG)-based hydrogels are promising for cell encapsulation and tissue engineering, but are known to elicit a foreign body response (FBR) in

  15. The role of substrate morphology for the cytokine release profile of immature human primary macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Bartneck, Matthias [Department of Medicine III, Medical Faculty, RWTH Aachen, Pauwelsstr. 30, 52074 Aachen (Germany); Heffels, Karl-Heinz [Department and Chair of Functional Materials in Medicine and Dentistry, University of Würzburg, Pleicherwall 2, 97070 Würzburg (Germany); Bovi, Manfred [Electron Microscopic Facility, Medical Faculty, RWTH Aachen (Germany); Groll, Jürgen [Department and Chair of Functional Materials in Medicine and Dentistry, University of Würzburg, Pleicherwall 2, 97070 Würzburg (Germany); Zwadlo-Klarwasser, Gabriele [Interdisciplinary Center for Clinical Research and Dept. of Dermatology, Medical Faculty, RWTH Aachen, Pauwelsstr. 30, 52056 Aachen (Germany)

    2013-12-01

    There is increasing evidence that the physicochemical nature of any given material is a dominant factor for the release of cytokines by innate immune cells, specifically of macrophages, and thus majorly influences their interaction with other cell types. Recently, we could show that the 3D structure of star shaped polytheylene oxide–polypropylene oxide co-polymers (sP(EO-stat-PO))-hydrogel coated substrates has a stronger influence on the release pattern of cytokines after 7 days of culture than surface chemistry. Here, we focused on the analysis of cytokine release over time and a more detailed analysis of cell morphology by scanning electron microscopy (SEM). Therefore, we compared different strategies for SEM sample preparation and found that using osmium tetroxide combined with aqua bidest led to best preparation results. For cytokine release we show significant changes from day 3 to day 7 of cell culture. After 3 days, the sP(EO-stat-PO)-coated substrates led to an induction of pro-angiogenic CCL3 and CCL4, and of low amounts of the anti-inflammatory IL10, which declined at day 7. In contrast, pleiotropic IL6 and the pro-inflammatory TNFα and IL1β were expressed stronger at day 7 than at day 3. - Highlights: • Strategies for the preparation of macrophages on hydrogel materials (Fig. 1) • Cytokine release of immature macrophages on the substrates (Fig. 2 and Table 1) • Changes in cytokine release during macrophage maturation (Table 2)

  16. The role of substrate morphology for the cytokine release profile of immature human primary macrophages

    International Nuclear Information System (INIS)

    Bartneck, Matthias; Heffels, Karl-Heinz; Bovi, Manfred; Groll, Jürgen; Zwadlo-Klarwasser, Gabriele

    2013-01-01

    There is increasing evidence that the physicochemical nature of any given material is a dominant factor for the release of cytokines by innate immune cells, specifically of macrophages, and thus majorly influences their interaction with other cell types. Recently, we could show that the 3D structure of star shaped polytheylene oxide–polypropylene oxide co-polymers (sP(EO-stat-PO))-hydrogel coated substrates has a stronger influence on the release pattern of cytokines after 7 days of culture than surface chemistry. Here, we focused on the analysis of cytokine release over time and a more detailed analysis of cell morphology by scanning electron microscopy (SEM). Therefore, we compared different strategies for SEM sample preparation and found that using osmium tetroxide combined with aqua bidest led to best preparation results. For cytokine release we show significant changes from day 3 to day 7 of cell culture. After 3 days, the sP(EO-stat-PO)-coated substrates led to an induction of pro-angiogenic CCL3 and CCL4, and of low amounts of the anti-inflammatory IL10, which declined at day 7. In contrast, pleiotropic IL6 and the pro-inflammatory TNFα and IL1β were expressed stronger at day 7 than at day 3. - Highlights: • Strategies for the preparation of macrophages on hydrogel materials (Fig. 1) • Cytokine release of immature macrophages on the substrates (Fig. 2 and Table 1) • Changes in cytokine release during macrophage maturation (Table 2)

  17. Tubular lysosome morphology and distribution within macrophages depend on the integrity of cytoplasmic microtubules

    International Nuclear Information System (INIS)

    Swanson, J.; Bushnell, A.; Silverstein, S.C.

    1987-01-01

    Pinocytosis of the fluorescent dye lucifer yellow labels elongated, membrane-bound tubular organelles in several cell types, including cultured human monocytes, thioglycolate-elicited mouse peritoneal macrophages, and the macrophage-like cell line J774.2. These tubular structures can be identified as lysosomes by acid phosphatase histochemistry and immunofluorescence localization of cathepsin L. The abundance of tubular lysosomes is markedly increased by treatment with phorbol 12-myristate 13-acetate. When labeled by pinocytosis of microperoxidase and examined by electron microscopic histochemistry, the tubular lysosomes have an outside diameter of ≅ 75 nm and a length of several micrometers; they radiate from the cell's centrosphere in alignment with cytoplasmic microtubules and intermediate filaments. Incubation of phorbol myristate acetate-treated macrophages at 4 0 C or in medium containing 5 μM colchicine or nocodazole at 37 0 C leads to disassembly of microtubules and fragmentation of the tubular lysosomes. Return of the cultures to 37 0 C or removal of nocodazole from the medium leads to reassembly of microtubules and the reappearance of tubular lysosomes within 10-20 min. The authors conclude that microtubules are essential for the maintenance of tubular lysosome morphology and that, in macrophages, a significant proportion of the lysosomal compartment is contained within these tubular structures

  18. Legionella pneumophila transcriptome during intracellular multiplication in human macrophages

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    Sebastien P Faucher

    2011-04-01

    Full Text Available Legionella pneumophila is the causative agent of Legionnaires’ disease, an acute pulmonary infection. L. pneumophila is able to infect and multiply in both phagocytic protozoa, such as Acanthamoeba castellanii, and mammalian professional phagocytes. The best-known L. pneumophila virulence determinant is the Icm/Dot Type IVB secretion system (TFBSS, which is used to translocate more than 150 effector proteins to host cells. While the transcriptional response of Legionella to the intracellular environment of A. castellanii has been investigated, much less is known about the Legionella transcriptional response inside human macrophages. In this study, the transcriptome of L. pneumophila was monitored during exponential and post-exponential phase in rich AYE broth as well as during infection of human cultured macrophages. This was accomplished with microarrays and an RNA amplification procedure called SCOTS to detect small amounts of mRNA from low numbers of intracellular bacteria. Among the genes induced intracellularly are those involved in amino acid biosynthetic pathways leading to L-arginine, L-histidine and L-proline as well as many transport systems involved in amino acid and iron uptake. Gene involved in catabolism of glycerol is also induced during intracellular growth and could be used as a carbon source. The genes encoding the Icm/Dot system are not differentially expressed inside cells compared to control bacteria grown in rich broth, but the genes encoding several translocated effectors are strongly induced. Moreover, we used the transcriptome data to predict previously unrecognized Icm/Dot effector genes based on their expression pattern and confirmed translocation for three candidates. This study provides a comprehensive view of how L. pneumophila responds to the human macrophage intracellular environment.

  19. The uptake of tocopherols by RAW 264.7 macrophages

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    Papas Andreas M

    2002-10-01

    Full Text Available Abstract Background Alpha-Tocopherol and gamma-tocopherol are the two major forms of vitamin E in human plasma and the primary lipid soluble antioxidants. The dietary intake of gamma-tocopherol is generally higher than that of alpha-tocopherol. However, alpha-tocopherol plasma levels are about four fold higher than those of gamma-tocopherol. Among other factors, a preferential cellular uptake of gamma-tocopherol over alpha-tocopherol could contribute to the observed higher plasma alpha-tocopherol levels. In this investigation, we studied the uptake and depletion of both alpha-tocopherol and gamma-tocopherol (separately and together in cultured RAW 264.7 macrophages. Similar studies were performed with alpha-tocopheryl quinone and gamma-tocopheryl quinone, which are oxidation products of tocopherols. Results RAW 264.7 macrophages showed a greater uptake of gamma-tocopherol compared to alpha-tocopherol (with uptake being defined as the net difference between tocopherol transported into the cells and loss due to catabolism and/or in vitro oxidation. Surprisingly, we also found that the presence of gamma-tocopherol promoted the cellular uptake of alpha-tocopherol. Mass balance considerations suggest that products other than quinone were formed during the incubation of tocopherols with macrophages. Conclusion Our data suggests that gamma-tocopherol could play a significant role in modulating intracellular antioxidant defence mechanisms. Moreover, we found the presence of gamma-tocopherol dramatically influenced the cellular accumulation of alpha-tocopherol, i.e., gamma-tocopherol promoted the accumulation of alpha-tocopherol. If these results could be extrapolated to in vivo conditions they suggest that gamma-tocopherol is selectively taken up by cells and removed from plasma more rapidly than alpha-tocopherol. This could, in part, contribute to the selective maintenance of alpha-tocopherol in plasma compared to gamma-tocopherol.

  20. Lemongrass and citral effect on cytokines production by murine macrophages.

    Science.gov (United States)

    Bachiega, Tatiana Fernanda; Sforcin, José Maurício

    2011-09-01

    Cymbopogon citratus (DC) Stapf (Poaceae-Gramineae), an herb commonly known as lemongrass (LG), is an important source of ethnomedicines as well as citral, the major constituent of Cymbopogon citratus, used in perfumery, cosmetic and pharmaceutical industries for controlling pathogens. Thus, the goal of this work was to analyze the effect of LG and citral on cytokines production (IL-1β, IL-6 and IL-10) in vitro, as well as before or after LPS incubation. Peritoneal macrophages from BALB/c mice were treated with LG or citral in different concentrations for 24h. The concentrations that inhibited cytokines production were tested before or after macrophages challenge with LPS, in order to evaluate a possible anti-inflammatory action. Supernatants of cell cultures were used for cytokines determination by ELISA. As to IL-1β, only citral inhibited its release, exerting an efficient action before LPS challenge. LG and citral inhibited IL-6 release. Cymbopogon citratus showed inhibitory effects only after LPS challenge, whereas citral prevented efficiently LPS effects before and after LPS addition. Citral inhibited IL-10 production and although LG did not inhibit its production, the concentration of 100 μg/well was tested in the LPS-challenge protocol, because it inhibited IL-6 production. LG inhibited LPS action after macrophages incubation with LPS, while citral counteracted LPS action when added before or after LPS incubation. LG exerted an anti-inflammatory action and citral may be involved in its inhibitory effects on cytokines production. We suggest that a possible mechanism involved in such results could be the inhibition of the transcription factor NF-κB. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  1. M2 macrophages induce ovarian cancer cell proliferation via a heparin binding epidermal growth factor/matrix metalloproteinase 9 intercellular feedback loop.

    Science.gov (United States)

    Carroll, Molly J; Kapur, Arvinder; Felder, Mildred; Patankar, Manish S; Kreeger, Pamela K

    2016-12-27

    In ovarian cancer, a high ratio of anti-inflammatory M2 to pro-inflammatory M1 macrophages correlates with poor patient prognosis. The mechanisms driving poor tumor outcome as a result of the presence of M2 macrophages in the tumor microenvironment remain unclear and are challenging to study with current techniques. Therefore, in this study we utilized a micro-culture device previously developed by our lab to model concentrated paracrine signaling in order to address our hypothesis that interactions between M2 macrophages and ovarian cancer cells induce tumor cell proliferation. Using the micro-culture device, we determined that co-culture with M2-differentiated primary macrophages or THP-1 increased OVCA433 proliferation by 10-12%. This effect was eliminated with epidermal growth factor receptor (EGFR) or heparin-bound epidermal growth factor (HB-EGF) neutralizing antibodies and HBEGF expression in peripheral blood mononuclear cells from ovarian cancer patients was 9-fold higher than healthy individuals, suggesting a role for HB-EGF in tumor progression. However, addition of HB-EGF at levels secreted by macrophages or macrophage-conditioned media did not induce proliferation to the same extent, indicating a role for other factors in this process. Matrix metalloproteinase-9, MMP-9, which cleaves membrane-bound HB-EGF, was elevated in co-culture and its inhibition decreased proliferation. Utilizing inhibitors and siRNA against MMP9 in each population, we determined that macrophage-secreted MMP-9 released HB-EGF from macrophages, which increased MMP9 in OVCA433, resulting in a positive feedback loop to drive HB-EGF release and increase proliferation in co-culture. Identification of multi-cellular interactions such as this may provide insight into how to most effectively control ovarian cancer progression.

  2. DMPD: Nuclear receptors in macrophages: a link between metabolism and inflammation. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18022390 Nuclear receptors in macrophages: a link between metabolism and inflammati...on. Szanto A, Roszer T. FEBS Lett. 2008 Jan 9;582(1):106-16. Epub 2007 Nov 20. (.png) (.svg) (.html) (.csml) Show Nuclear... receptors in macrophages: a link between metabolism and inflammation. PubmedID 18022390 Title Nuclear

  3. DMPD: Receptor tyrosine kinases and the regulation of macrophage activation. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 14726496 Receptor tyrosine kinases and the regulation of macrophage activation. Cor...osine kinases and the regulation of macrophage activation. PubmedID 14726496 Title Receptor tyrosine...rell PH, Morrison AC, Lutz MA. J Leukoc Biol. 2004 May;75(5):731-7. Epub 2004 Jan 14. (.png) (.svg) (.html) (.csml) Show Receptor tyr

  4. Colonic macrophage polarization in homeostasis, inflammation, and cancer

    Science.gov (United States)

    Appleyard, Caroline B.

    2016-01-01

    Our review focuses on the colonic macrophage, a monocyte-derived, tissue-resident macrophage, and the role it plays in health and disease, specifically in inflammatory conditions such as inflammatory bowel disease and cancer of the colon and rectum. We give special emphasis to macrophage polarization, or phenotype, in these different states. We focus on macrophages because they are one of the most numerous leukocytes in the colon, and because they normally contribute to homeostasis through an anti-inflammatory phenotype. However, in conditions such as inflammatory bowel disease, proinflammatory macrophages are increased in the colon and have been linked to disease severity and progression. In colorectal cancer, tumor cells may employ anti-inflammatory macrophages to promote tumor growth and dissemination, whereas proinflammatory macrophages may antagonize tumor growth. Given the key roles that this cell type plays in homeostasis, inflammation, and cancer, the colonic macrophage is an intriguing therapeutic target. As such, potential macrophage-targeting strategies are discussed. PMID:27229123

  5. Macrophage colony stimulating factor (M-CSF) induces Fc receptor expression on macrophages

    International Nuclear Information System (INIS)

    Magee, D.M.; Wing, E.J.; Waheed, A.; Shadduck, R.K.

    1986-01-01

    M-CSF is a glycoprotein that stimulates bone marrow progenitor cells to proliferate and differentiate into macrophages (M theta). In addition, M-CSF can modulate the function of mature M theta. In this study, the authors determined the effect of M-CSF on expression of receptors for IgG (Fc receptors). Murine resident peritoneal M theta monolayers were incubated with either M-CSF, recombinant gamma interferon (IFN), or left untreated for 48 hrs. Expression of Fc receptors was assessed by microscopy using an antibody coated sheet erythrocytes (EA) rosette assay. The results indicated that M-CSF treated M theta had significantly higher numbers of bound EA (7.1 erythrocytes/M theta), than IFN M theta (4.4), or untreated M theta (2.5) (p 51 Cr labelled EA assay, CSF M theta (16,411 cpm), IFN M theta (10,887), untreated M theta (6897) (p < 0.001). Additionally, the maximal response was noted between 10 and 500 units M-CSF. Purified anti-M-CSF IgG, when included in the cultures, ablated the enhancement of EA binding, whereas normal rabbit IgG did not. These findings indicate that M-CSF is a potent inducer of Fc receptor expression on M theta and supports other data concerning the role of M-CSF as a biological response modifier

  6. Macrophage polarisation: an immunohistochemical approach for identifying M1 and M2 macrophages.

    Directory of Open Access Journals (Sweden)

    Mário Henrique M Barros

    Full Text Available Macrophage polarization is increasingly recognised as an important pathogenetic factor in inflammatory and neoplastic diseases. Proinflammatory M1 macrophages promote T helper (Th 1 responses and show tumoricidal activity. M2 macrophages contribute to tissue repair and promote Th2 responses. CD68 and CD163 are used to identify macrophages in tissue sections. However, characterisation of polarised macrophages in situ has remained difficult. Macrophage polarisation is regulated by transcription factors, pSTAT1 and RBP-J for M1, and CMAF for M2. We reasoned that double-labelling immunohistochemistry for the detection of macrophage markers together with transcription factors may be suitable to characterise macrophage polarisation in situ. To test this hypothesis, we have studied conditions associated with Th1- and Th2-predominant immune responses: infectious mononucleosis and Crohn's disease for Th1 and allergic nasal polyps, oxyuriasis, wound healing and foreign body granulomas for predominant Th2 response. In all situations, CD163+ cells usually outnumbered CD68+ cells. Moreover, CD163+ cells, usually considered as M2 macrophages, co-expressing pSTAT1 and RBP-J were found in all conditions examined. The numbers of putative M1 macrophages were higher in Th1- than in Th2-associated diseases, while more M2 macrophages were seen in Th2- than in Th1 related disorders. In most Th1-related diseases, the balance of M1 over M2 cells was shifted towards M1 cells, while the reverse was observed for Th2-related conditions. Hierarchical cluster analysis revealed two distinct clusters: cluster I included Th1 diseases together with cases with high numbers of CD163+pSTAT1+, CD68+pSTAT1+, CD163+RBP-J+ and CD68+RBP-J+ macrophages; cluster II comprised Th2 conditions together with cases displaying high numbers of CD163+CMAF+ and CD68+CMAF+ macrophages. These results suggest that the detection of pSTAT1, RBP-J, and CMAF in the context of CD68 or CD163 expression is a

  7. Macrophage polarisation: an immunohistochemical approach for identifying M1 and M2 macrophages.

    Science.gov (United States)

    Barros, Mário Henrique M; Hauck, Franziska; Dreyer, Johannes H; Kempkes, Bettina; Niedobitek, Gerald

    2013-01-01

    Macrophage polarization is increasingly recognised as an important pathogenetic factor in inflammatory and neoplastic diseases. Proinflammatory M1 macrophages promote T helper (Th) 1 responses and show tumoricidal activity. M2 macrophages contribute to tissue repair and promote Th2 responses. CD68 and CD163 are used to identify macrophages in tissue sections. However, characterisation of polarised macrophages in situ has remained difficult. Macrophage polarisation is regulated by transcription factors, pSTAT1 and RBP-J for M1, and CMAF for M2. We reasoned that double-labelling immunohistochemistry for the detection of macrophage markers together with transcription factors may be suitable to characterise macrophage polarisation in situ. To test this hypothesis, we have studied conditions associated with Th1- and Th2-predominant immune responses: infectious mononucleosis and Crohn's disease for Th1 and allergic nasal polyps, oxyuriasis, wound healing and foreign body granulomas for predominant Th2 response. In all situations, CD163+ cells usually outnumbered CD68+ cells. Moreover, CD163+ cells, usually considered as M2 macrophages, co-expressing pSTAT1 and RBP-J were found in all conditions examined. The numbers of putative M1 macrophages were higher in Th1- than in Th2-associated diseases, while more M2 macrophages were seen in Th2- than in Th1 related disorders. In most Th1-related diseases, the balance of M1 over M2 cells was shifted towards M1 cells, while the reverse was observed for Th2-related conditions. Hierarchical cluster analysis revealed two distinct clusters: cluster I included Th1 diseases together with cases with high numbers of CD163+pSTAT1+, CD68+pSTAT1+, CD163+RBP-J+ and CD68+RBP-J+ macrophages; cluster II comprised Th2 conditions together with cases displaying high numbers of CD163+CMAF+ and CD68+CMAF+ macrophages. These results suggest that the detection of pSTAT1, RBP-J, and CMAF in the context of CD68 or CD163 expression is a suitable tool for

  8. A macrophage activation switch (MAcS)-index for assessment of monocyte/macrophage activation

    DEFF Research Database (Denmark)

    Maniecki, Maciej Bogdan; Lauridsen, Mette; Knudsen, Troels Bygum

    2008-01-01

    , simplified by the M1-M2 dichotomy of classically activated (M1), pro-inflammatory cells and alternatively activated (M2), anti-inflammatory cells. Macrophages, however, display a large degree of flexibility and are able to switch between activation states (1). The hemoglobin scavenger receptor CD163...... is expressed exclusively on monocytes and macrophages, and its expression is strongly induced by anti-inflammatory stimuli like IL10 and glucocorticoid, making CD163 an ideal M2 macrophage marker (2). Furthermore a soluble variant of CD163 (sCD163) is shed from the cell surface to plasma by protease mediated.......058-5139) (panti-inflammatory state.   CONCLUSION: We present a CD163-derived macrophage activation switch (MAcS)-index, which seems able to differentiate between (predominantly) pro-inflammatory and anti-inflammatory macrophage activation. The index needs...

  9. Variation in HIV-1 R5 macrophage-tropism correlates with sensitivity to reagents that block envelope: CD4 interactions but not with sensitivity to other entry inhibitors

    Directory of Open Access Journals (Sweden)

    Simmonds Peter

    2008-01-01

    Full Text Available Abstract Background HIV-1 R5 viruses cause most of the AIDS cases worldwide and are preferentially transmitted compared to CXCR4-using viruses. Furthermore, R5 viruses vary extensively in capacity to infect macrophages and highly macrophage-tropic variants are frequently identified in the brains of patients with dementia. Here, we investigated the sensitivity of R5 envelopes to a range of inhibitors and antibodies that block HIV entry. We studied a large panel of R5 envelopes, derived by PCR amplification without culture from brain, lymph node, blood and semen. These R5 envelopes conferred a wide range of macrophage tropism and included highly macrophage-tropic variants from brain and non-macrophage-tropic variants from lymph node. Results R5 macrophage-tropism correlated with sensitivity to inhibition by reagents that inhibited gp120:CD4 interactions. Thus, increasing macrophage-tropism was associated with increased sensitivity to soluble CD4 and to IgG-CD4 (PRO 542, but with increased resistance to the anti-CD4 monoclonal antibody (mab, Q4120. These observations were highly significant and are consistent with an increased affinity of envelope for CD4 for macrophage-tropic envelopes. No overall correlations were noted between R5 macrophage-tropism and sensitivity to CCR5 antagonists or to gp41 specific reagents. Intriguingly, there was a relationship between increasing macrophage-tropism and increased sensitivity to the CD4 binding site mab, b12, but decreased sensitivity to 2G12, a mab that binds a glycan complex on gp120. Conclusion Variation in R5 macrophage-tropism is caused by envelope variation that predominantly influences sensitivity to reagents that block gp120:CD4 interactions. Such variation has important implications for therapy using viral entry inhibitors and for the design of envelope antigens for vaccines.

  10. Macrophage-induced cytostasis: kinetic analysis of bromodeoxyuridine-pulsed cells

    International Nuclear Information System (INIS)

    Stevenson, A.P.; Crissman, H.A.; Stewart, C.C.

    1985-01-01

    The effect of tumoricidal macrophages on the cell cycle progression of six different cell lines was studied using an anti-bromodeoxyuridine (BrdUrd) monoclonal antibody to follow the traverse of BrdUrd-labeled cells. Exponentially growing cultured mammalian cells, from six different cell lines, were prepulsed with BrdUrd before exposure to tumoricidal macrophages. The cultured cells were then analyzed as a function of time for DNA content (by propidium iodide staining) and for BrdUrd incorporation (using a fluoresceini-sothiocyanate [FITC]-conjugated anti-BrdUrd monoclonal antibody). The position of the cells in cycle and the progression of the BrdUrd-labeled cohort was followed using flow cytometry. The cell lines examined were: Colon 26; BALB/c-3T3, ST3T3 (a spontaneously transformed, tumorigenic clone of 3T3), WCHE5 (a clone of whole Chinese hamster embryo cells), RIF (a radiation-induced fibrosarcoma), and A101D (a human melanoma). The bivariate distributions showed that for all six cell lines the BrdUrd-labeled cohort in the control cultures progressed around the cell cycle during the first 12 h of culture, as the cells exponentially increased. In contrast, when each cell line was incubated with tumoricidal macrophages, the BrdUrd-labeled cohort did not progress through cell cycle but remained in S phase throughout the 12-h culture period. There was also no evidence for progression of cells out of G 1 . The data show that cells were arrested in every phase of cell cycle. This study suggests that cytostasis, as manifested by the termination of progression in all phases of the cell cycle, is a universal phenomenon induced by tumoricidal macrophages. 20 references, 4 figures

  11. Release of lysosomal enzymes in Candida albicans phagocytosis by rat peritoneal macrophages.

    Science.gov (United States)

    Fontenla de Petrino, S E; Sirena, A

    1984-02-15

    The present paper reports the in vitro release of lysosomal enzymes in the supernatant of cultures of rat peritoneal macrophages, with the addition of Candida albicans cells. Macrophages were taken from the rat peritoneal cavity 72 hr after non-specific activation with Brain-Heart-Infusion (B.H.I.) broth containing 10% proteose-peptone No. 3. They were then cultured in Parker medium No. 199 (TC 199). After 24 hr a suspension of Candida albicans cells, in a determined concentration, was added to the peritoneal macrophage cultures. At that time, and during pre-determined periods, the following enzymes in the culture supernatants were studied using colorimetric methods: beta-glucuronidase, beta-galactosidase and acid phosphatase. It is concluded that, under identical conditions, the release of beta-galactosidase and acid phosphatase is higher than for beta-glucuronidase. The release rate of all three enzymes is the highest at a 6 hr incubation period, after which, a gradual decrease leads to the rate down to 50% at 24 hr.

  12. The Poly-γ-D-Glutamic Acid Capsule of Bacillus licheniformis, a Surrogate of Bacillus anthracis Capsule Induces Interferon-Gamma Production in NK Cells through Interactions with Macrophages.

    Science.gov (United States)

    Lee, Hae-Ri; Jeon, Jun Ho; Rhie, Gi-Eun

    2017-05-28

    The poly-γ- D -glutamic acid (PGA) capsule, a major virulence factor of Bacillus anthracis , provides protection of the bacterium from phagocytosis and allows its unimpeded growth in the host. We investigated crosstalk between murine natural killer (NK) cells and macrophages stimulated with the PGA capsule of Bacillus licheniformis , a surrogate of the B. anthracis capsule. PGA induced interferon-gamma production from NK cells cultured with macrophages. This effect was dependent on macrophage-derived IL-12 and cell-cell contact interaction with macrophages through NK cell receptor NKG2D and its ligand RAE-1. The results showed that PGA could enhance NK cell activation by inducing IL-12 production in macrophages and a contact-dependent crosstalk with macrophages.

  13. Macrophages in Mildly Acid Microenvironment

    African Journals Online (AJOL)

    1College of Stomatology, Guiyang Medical University, 2Guiyang Stomatological Hospital, Guiyang, Guizhou 550004, PR China ... supernatants were collected after 24-h culture, and the content of VEGF and VEGF-C in each ... concentration at the same pH value, and reached a minimum at 50 ug/mL and pH 6.6. Conclusion ...

  14. Macrophages inhibit human osteosarcoma cell growth after activation with the bacterial cell wall derivative liposomal muramyl tripeptide in combination with interferon-γ.

    Science.gov (United States)

    Pahl, Jens H W; Kwappenberg, Kitty M C; Varypataki, Eleni M; Santos, Susy J; Kuijjer, Marieke L; Mohamed, Susan; Wijnen, Juul T; van Tol, Maarten J D; Cleton-Jansen, Anne-Marie; Egeler, R Maarten; Jiskoot, Wim; Lankester, Arjan C; Schilham, Marco W

    2014-03-10

    In osteosarcoma, the presence of tumor-infiltrating macrophages positively correlates with patient survival in contrast to the negative effect of tumor-associated macrophages in patients with other tumors. Liposome-encapsulated muramyl tripeptide (L-MTP-PE) has been introduced in the treatment of osteosarcoma patients, which may enhance the potential anti-tumor activity of macrophages. Direct anti-tumor activity of human macrophages against human osteosarcoma cells has not been described so far. Hence, we assessed osteosarcoma cell growth after co-culture with human macrophages. Monocyte-derived M1-like and M2-like macrophages were polarized with LPS + IFN-γ, L-MTP-PE +/- IFN-γ or IL-10 and incubated with osteosarcoma cells. Two days later, viable tumor cell numbers were analyzed. Antibody-dependent effects were investigated using the therapeutic anti-EGFR antibody cetuximab. M1-like macrophages inhibited osteosarcoma cell growth when activated with LPS + IFN-γ. Likewise, stimulation of M1-like macrophages with liposomal muramyl tripeptide (L-MTP-PE) inhibited tumor growth, but only when combined with IFN-γ. Addition of the tumor-reactive anti-EGFR antibody cetuximab did not further improve the anti-tumor activity of activated M1-like macrophages. The inhibition was mediated by supernatants of activated M1-like macrophages, containing TNF-α and IL-1β. However, specific blockage of these cytokines, nitric oxide or reactive oxygen species did not inhibit the anti-tumor effect, suggesting the involvement of other soluble factors released upon macrophage activation. While LPS + IFN-γ-activated M2-like macrophages had low anti-tumor activity, IL-10-polarized M2-like macrophages were able to reduce osteosarcoma cell growth in the presence of the anti-EGFR cetuximab involving antibody-dependent tumor cell phagocytosis. This study demonstrates that human macrophages can be induced to exert direct anti-tumor activity against osteosarcoma cells. Our

  15. Adipocyte-Macrophage Cross-Talk in Obesity.

    Science.gov (United States)

    Engin, Ayse Basak

    2017-01-01

    Obesity is characterized by the chronic low-grade activation of the innate immune system. In this respect, macrophage-elicited metabolic inflammation and adipocyte-macrophage interaction has a primary importance in obesity. Large amounts of macrophages are accumulated by different mechanisms in obese adipose tissue. Hypertrophic adipocyte-derived chemotactic monocyte chemoattractant protein-1 (MCP-1)/C-C chemokine receptor 2 (CCR2) pathway also promotes more macrophage accumulation into the obese adipose tissue. However, increased local extracellular lipid concentrations is a final mechanism for adipose tissue macrophage accumulation. A paracrine loop involving free fatty acids and tumor necrosis factor-alpha (TNF-alpha) between adipocytes and macrophages establishes a vicious cycle that aggravates inflammatory changes in the adipose tissue. Adipocyte-specific caspase-1 and production of interleukin-1beta (IL-1beta) by macrophages; both adipocyte and macrophage induction by toll like receptor-4 (TLR4) through nuclear factor-kappaB (NF-kappaB) activation; free fatty acid-induced and TLR-mediated activation of c-Jun N-terminal kinase (JNK)-related pro-inflammatory pathways in CD11c+ immune cells; are effective in macrophage accumulation and in the development of adipose tissue inflammation. Old adipocytes are removed by macrophages through trogocytosis or sending an "eat me" signal. The obesity-induced changes in adipose tissue macrophage numbers are mainly due to increases in the triple-positive CD11b+ F4/80+ CD11c+ adipose tissue macrophage subpopulation. The ratio of M1-to-M2 macrophages is increased in obesity. Furthermore, hypoxia along with higher concentrations of free fatty acids exacerbates macrophage-mediated inflammation in obesity. The metabolic status of adipocytes is a major determinant of macrophage inflammatory output. Macrophage/adipocyte fatty-acid-binding proteins act at the interface of metabolic and inflammatory pathways. Both macrophages and

  16. Excess 25-hydroxyvitamin D3 exacerbates tubulointerstitial injury in mice by modulating macrophage phenotype.

    Science.gov (United States)

    Kusunoki, Yasuo; Matsui, Isao; Hamano, Takayuki; Shimomura, Akihiro; Mori, Daisuke; Yonemoto, Sayoko; Takabatake, Yoshitsugu; Tsubakihara, Yoshiharu; St-Arnaud, René; Isaka, Yoshitaka; Rakugi, Hiromi

    2015-11-01

    Vitamin D hydroxylated at carbon 25 (25(OH)D) is generally recognized as a precursor of active vitamin D. Despite its low affinity for the vitamin D receptor (VDR), both deficient and excessive 25(OH)D levels are associated with poor clinical outcomes. Here we studied direct effects of 25(OH)D3 on the kidney using 25(OH)D-1α-hydroxylase (CYP27B1) knockout mice. The effects of 25(OH)D3 on unilateral ureteral obstruction were analyzed as proximal tubular cells and macrophages are two major cell types that take up 25(OH)D and contribute to the pathogenesis of kidney injury. Excess 25(OH)D3 in obstructed mice worsened oxidative stress and tubulointerstitial fibrosis, whereas moderate levels of 25(OH)D3 had no effects. The exacerbating effects of excess 25(OH)D3 were abolished in CYP27B1/VDR double-knockout mice and in macrophage-depleted CYP27B1 knockout mice. Excess 25(OH)D3 upregulated both M1 marker (TNF-α) and M2 marker (TGF-β1) levels of kidney-infiltrating macrophages. In vitro analyses verified that excess 25(OH)D3 directly upregulated TNF-α and TGF-β1 in cultured macrophages but not in tubular cells. TNF-α and 25(OH)D3 cooperatively induced oxidative stress by upregulating iNOS in tubular cells. Aggravated tubulointerstitial fibrosis in mice with excess 25(OH)D3 indicated that macrophage-derived TGF-β1 also had a key role in the pathogenesis of surplus 25(OH)D3. Thus, excess 25(OH)D3 worsens tubulointerstitial injury by modulating macrophage phenotype.

  17. Entrance and Survival of Brucella pinnipedialis Hooded Seal Strain in Human Macrophages and Epithelial Cells

    Science.gov (United States)

    Briquemont, Benjamin; Sørensen, Karen K.; Godfroid, Jacques

    2013-01-01

    Marine mammal Brucella spp. have been isolated from pinnipeds (B. pinnipedialis) and cetaceans (B. ceti) from around the world. Although the zoonotic potential of marine mammal brucellae is largely unknown, reports of human disease exist. There are few studies of the mechanisms of bacterial intracellular invasion and multiplication involving the marine mammal Brucella spp. We examined the infective capacity of two genetically different B. pinnipedialis strains (reference strain; NTCT 12890 and a hooded seal isolate; B17) by measuring the ability of the bacteria to enter and replicate in cultured phagocytes and epithelial cells. Human macrophage-like cells (THP-1), two murine macrophage cell lines (RAW264.7 and J774A.1), and a human malignant epithelial cell line (HeLa S3) were challenged with bacteria in a gentamicin protection assay. Our results show that B. pinnipedialis is internalized, but is then gradually eliminated during the next 72 – 96 hours. Confocal microscopy revealed that intracellular B. pinnipedialis hooded seal strain colocalized with lysosomal compartments at 1.5 and 24 hours after infection. Intracellular presence of B. pinnipedialis hooded seal strain was verified by transmission electron microscopy. By using a cholesterol-scavenging lipid inhibitor, entrance of B. pinnipedialis hooded seal strain in human macrophages was significantly reduced by 65.8 % (± 17.3), suggesting involvement of lipid-rafts in intracellular entry. Murine macrophages invaded by B. pinnipedialis do not release nitric oxide (NO) and intracellular bacterial presence does not induce cell death. In summary, B. pinnipedialis hooded seal strain can enter human and murine macrophages, as well as human epithelial cells. Intracellular entry of B. pinnipedialis hooded seal strain involves, but seems not to be limited to, lipid-rafts in human macrophages. Brucella pinnipedialis does not multiply or survive for prolonged periods intracellulary. PMID:24376851

  18. IQGAP1 is involved in post-ischemic neovascularization by regulating angiogenesis and macrophage infiltration.

    Directory of Open Access Journals (Sweden)

    Norifumi Urao

    2010-10-01

    Full Text Available Neovascularization is an important repair mechanism in response to ischemic injury and is dependent on inflammation, angiogenesis and reactive oxygen species (ROS. IQGAP1, an actin-binding scaffold protein, is a key regulator for actin cytoskeleton and motility. We previously demonstrated that IQGAP1 mediates vascular endothelial growth factor (VEGF-induced ROS production and migration of cultured endothelial cells (ECs; however, its role in post-ischemic neovascularization is unknown.Ischemia was induced by left femoral artery ligation, which resulted in increased IQGAP1 expression in Mac3(+ macrophages and CD31(+ capillary-like ECs in ischemic legs. Mice lacking IQGAP1 exhibited a significant reduction in the post-ischemic neovascularization as evaluated by laser Doppler blood flow, capillary density and α-actin positive arterioles. Furthermore, IQGAP1(-/- mice showed a decrease in macrophage infiltration and ROS production in ischemic muscles, leading to impaired muscle regeneration and increased necrosis and fibrosis. The numbers of bone marrow (BM-derived cells in the peripheral blood were not affected in these knockout mice. BM transplantation revealed that IQGAP1 expressed in both BM-derived cells and tissue resident cells, such as ECs, is required for post-ischemic neovascularization. Moreover, thioglycollate-induced peritoneal macrophage recruitment and ROS production were inhibited in IQGAP1(-/- mice. In vitro, IQGAP1(-/- BM-derived macrophages showed inhibition of migration and adhesion capacity, which may explain the defective macrophage recruitment into the ischemic tissue in IQGAP1(-/- mice.IQGAP1 plays a key role in post-ischemic neovascularization by regulating, not only, ECs-mediated angiogenesis but also macrophage infiltration as well as ROS production. Thus, IQGAP1 is a potential therapeutic target for inflammation- and angiogenesis-dependent ischemic cardiovascular diseases.

  19. Efferocytosis is impaired in Gaucher macrophages.

    Science.gov (United States)

    Aflaki, Elma; Borger, Daniel K; Grey, Richard J; Kirby, Martha; Anderson, Stacie; Lopez, Grisel; Sidransky, Ellen

    2017-04-01

    Gaucher disease, the inherited deficiency of lysosomal glucocerebrosidase, is characterized by the presence of glucosylceramide-laden macrophages resulting from impaired digestion of aged erythrocytes or apoptotic leukocytes. Studies of macrophages from patients with type 1 Gaucher disease with genotypes N370S/N370S, N370S/L444P or N370S/c.84dupG revealed that Gaucher macrophages have impaired efferocytosis resulting from reduced levels of p67 phox and Rab7. The decreased Rab7 expression leads to impaired fusion of phagosomes with lysosomes. Moreover, there is defective translocation of p67 phox to phagosomes, resulting in reduced intracellular production of reactive oxygen species. These factors contribute to defective deposition and clearance of apoptotic cells in phagolysosomes, which may have an impact on the inflammatory response and contribute to the organomegaly and inflammation seen in patients with Gaucher disease. Copyright© Ferrata Storti Foundation.

  20. Lysine Deacetylases and Regulated Glycolysis in Macrophages.

    Science.gov (United States)

    Shakespear, Melanie R; Iyer, Abishek; Cheng, Catherine Youting; Das Gupta, Kaustav; Singhal, Amit; Fairlie, David P; Sweet, Matthew J

    2018-06-01

    Regulated cellular metabolism has emerged as a fundamental process controlling macrophage functions, but there is still much to uncover about the precise signaling mechanisms involved. Lysine acetylation regulates the activity, stability, and/or localization of metabolic enzymes, as well as inflammatory responses, in macrophages. Two protein families, the classical zinc-dependent histone deacetylases (HDACs) and the NAD-dependent HDACs (sirtuins, SIRTs), mediate lysine deacetylation. We describe here mechanisms by which classical HDACs and SIRTs directly regulate specific glycolytic enzymes, as well as evidence that links these protein deacetylases to the regulation of glycolysis-related genes. In these contexts, we discuss HDACs and SIRTs as key control points for regulating immunometabolism and inflammatory outputs from macrophages. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Adipocyte fetuin-A contributes to macrophage migration into adipose tissue and polarization of macrophages.

    Science.gov (United States)

    Chatterjee, Priyajit; Seal, Soma; Mukherjee, Sandip; Kundu, Rakesh; Mukherjee, Sutapa; Ray, Sukanta; Mukhopadhyay, Satinath; Majumdar, Subeer S; Bhattacharya, Samir

    2013-09-27

    Macrophage infiltration into adipose tissue during obesity and their phenotypic conversion from anti-inflammatory M2 to proinflammatory M1 subtype significantly contributes to develop a link between inflammation and insulin resistance; signaling molecule(s) for these events, however, remains poorly understood. We demonstrate here that excess lipid in the adipose tissue environment may trigger one such signal. Adipose tissue from obese diabetic db/db mice, high fat diet-fed mice, and obese diabetic patients showed significantly elevated fetuin-A (FetA) levels in respect to their controls; partially hepatectomized high fat diet mice did not show noticeable alteration, indicating adipose tissue to be the source of this alteration. In adipocytes, fatty acid induces FetA gene and protein expressions, resulting in its copious release. We found that FetA could act as a chemoattractant for macrophages. To simulate lipid-induced inflammatory conditions when proinflammatory adipose tissue and macrophages create a niche of an altered microenvironment, we set up a transculture system of macrophages and adipocytes; the addition of fatty acid to adipocytes released FetA into the medium, which polarized M2 macrophages to M1. This was further confirmed by direct FetA addition to macrophages. Taken together, lipid-induced FetA from adipocytes is an efficient chemokine for macrophage migration and polarization. These findings open a new dimension for understanding obesity-induced inflammation.

  2. Signal-transducing mechanisms of ketamine-caused inhibition of interleukin-1β gene expression in lipopolysaccharide-stimulated murine macrophage-like Raw 264.7 cells

    International Nuclear Information System (INIS)

    Chen, T.-L.; Chang, C.-C.; Lin, Y.-L.; Ueng, Y.-F.; Chen, R.-M.

    2009-01-01

    Ketamine may affect the host immunity. Interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α) are pivotal cytokines produced by macrophages. This study aimed to evaluate the effects of ketamine on the regulation of inflammatory cytokine gene expression, especially IL-1β, in lipopolysaccharide (LPS)-activated murine macrophage-like Raw 264.7 cells and its possible signal-transducing mechanisms. Administration of Raw 264.7 cells with a therapeutic concentration of ketamine (100 μM), LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. Exposure to 100 μM ketamine decreased the binding affinity of LPS and LPS-binding protein but did not affect LPS-induced RNA and protein synthesis of TLR4. Treatment with LPS significantly increased IL-1β, IL-6, and TNF-α gene expressions in Raw 264.7 cells. Ketamine at a clinically relevant concentration did not affect the synthesis of these inflammatory cytokines, but significantly decreased LPS-caused increases in these cytokines. Immunoblot analyses, an electrophoretic mobility shift assay, and a reporter luciferase activity assay revealed that ketamine significantly decreased LPS-induced translocation and DNA binding activity of nuclear factor-kappa B (NFκB). Administration of LPS sequentially increased the phosphorylations of Ras, Raf, MEK1/2, ERK1/2, and IKK. However, a therapeutic concentration of ketamine alleviated such augmentations. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA reduced cellular TLR4 amounts and ameliorated LPS-induced RAS activation and IL-1β synthesis. Co-treatment with ketamine and TLR4 siRNA synergistically ameliorated LPS-caused enhancement of IL-1β production. Results of this study show that a therapeutic concentration of ketamine can inhibit gene expression of IL-1β possibly through suppressing TLR4-mediated signal-transducing phosphorylations of Ras, Raf, MEK1/2, ERK1/2, and IKK and subsequent translocation and

  3. The influence of protein malnutrition on the production of GM-CSF and M-CSF by macrophages

    Directory of Open Access Journals (Sweden)

    Dalila Cunha de Oliveira

    Full Text Available ABSTRACT It is well established that protein malnutrition (PM impairs immune defenses and increases susceptibility to infection. Macrophages are cells that play a central role in innate immunity, constituting one of the first barriers against infections. Macrophages produce several soluble factors, including cytokines and growth factors, important to the immune response. Among those growth factors, granulocyte-macrophage colony-stimulating factor (GM-CSF and macrophage colony-stimulating factor (M-CSF. GM-CSF and M-CSF are important to monocyte and macrophage development and stimulation of the immune response process. Knowing the importance of GM-CSF and M-CSF, we sought to investigate the influence of PM on macrophage production of these growth factors. Two-month-old male BALB/c mice were subjected to PM with a low-protein diet (2% and compared to a control diet (12% mouse group. Nutritional status, hemogram and the number of peritoneal cells were evaluated. Additionally, peritoneal macrophages were cultured and the production of GM-CSF and M-CSF and mRNA expression were evaluated. To determine if PM altered macrophage production of GM-CSF and M-CSF, they were stimulated with TNF-α. The PM animals had anemia, leukopenia and a reduced number of peritoneal cells. The production of M-CSF was not different between groups; however, cells from PM animals, stimulated with or without TNF-α, presented reduced capability to produce GM-CSF. These data imply that PM interferes with the production of GM-CSF, and consequently would affect the production and maturation of hematopoietic cells and the immune response.

  4. Long-term culture of organotypic multicellular glioma spheroids: a good culture model for studying gliomas

    NARCIS (Netherlands)

    Kaaijk, P.; Troost, D.; Das, P. K.; Leenstra, S.; Bosch, D. A.

    1995-01-01

    Gliomas, as well as other solid tumours, contain tumour stroma composed of connective tissue, macrophages, capillaries and other non-cellular constituents. Therefore, a homogeneous culture of tumour cells alone, as is often used as a culture model for gliomas, is not ideal to study all aspects of

  5. Drug Trafficking into Macrophages via the Endocytotic Receptor CD163

    DEFF Research Database (Denmark)

    Graversen, Jonas Heilskov; Moestrup, Søren Kragh

    2015-01-01

    for cytotoxic or phenotype-modulating drugs in the treatment of inflammatory and cancerous diseases. Such targeting of macrophages has been tried using the natural propensity of macrophages to non-specifically phagocytose circulating foreign particulate material. In addition, the specific targeting...... of macrophage-expressed receptors has been used in order to obtain a selective uptake in macrophages and reduce adverse effects of off-target delivery of drugs. CD163 is a highly expressed macrophage-specific endocytic receptor that has been studied for intracellular delivery of small molecule drugs...... to macrophages using targeted liposomes or antibody drug conjugates. This review will focus on the biology of CD163 and its potential role as a target for selective macrophage targeting compared with other macrophage targeting approaches....

  6. Wip1-dependent modulation of macrophage migration and phagocytosis

    DEFF Research Database (Denmark)

    Tang, Yiting; Pan, Bing; Zhou, Xin

    2017-01-01

    Macrophage accumulation within the vascular wall is a hallmark of atherosclerosis. Controlling macrophage conversion into foam cells remains a major challenge for treatment of atherosclerotic diseases. Here, we show that Wip1, a member of the PP2C family of Ser/Thr protein phosphatases, modulates...... macrophage migration and phagocytosis associated with atherosclerotic plaque formation. Wip1 deficiency increases migratory and phagocytic activities of the macrophage under stress conditions. Enhanced migration of Wip1-/- macrophages is mediated by Rac1-GTPase and PI3K/AKT signalling pathways. Elevated...... phagocytic ability of Wip1-/- macrophages is linked to CD36 plasma membrane recruitment that is regulated by AMPK activity. Our study identifies Wip1 as an intrinsic negative regulator of macrophage chemotaxis. We propose that Wip1-dependent control of macrophage function may provide avenues for preventing...

  7. Macrophages and Their Role in Atherosclerosis: Pathophysiology and Transcriptome Analysis

    Directory of Open Access Journals (Sweden)

    Yuri V. Bobryshev

    2016-01-01

    Full Text Available Atherosclerosis can be regarded as a chronic inflammatory state, in which macrophages play different and important roles. Phagocytic proinflammatory cells populate growing atherosclerotic lesions, where they actively participate in cholesterol accumulation. Moreover, macrophages promote formation of complicated and unstable plaques by maintaining proinflammatory microenvironment. At the same time, anti-inflammatory macrophages contribute to tissue repair and remodelling and plaque stabilization. Macrophages therefore represent attractive targets for development of antiatherosclerotic therapy, which can aim to reduce monocyte recruitment to the lesion site, inhibit proinflammatory macrophages, or stimulate anti-inflammatory responses and cholesterol efflux. More studies are needed, however, to create a comprehensive classification of different macrophage phenotypes and to define their roles in the pathogenesis of atherosclerosis. In this review, we provide an overview of the current knowledge on macrophage diversity, activation, and plasticity in atherosclerosis and describe macrophage-based cellular tests for evaluation of potential antiatherosclerotic substances.

  8. The macrophage scavenger receptor CD163

    DEFF Research Database (Denmark)

    Nielsen, Marianne Jensby; Madsen, Mette; Møller, Holger J

    2006-01-01

    CD163 is the monocyte/macrophage-specific receptor for haptoglobin-hemoglobin (Hp-Hb) complexes. The cytoplasmic tail of human CD163 exists as a short tail variant and two long tail variants. Reverse transcriptase-polymerase chain reaction analysis indicated that all three CD163 variants are subs......CD163 is the monocyte/macrophage-specific receptor for haptoglobin-hemoglobin (Hp-Hb) complexes. The cytoplasmic tail of human CD163 exists as a short tail variant and two long tail variants. Reverse transcriptase-polymerase chain reaction analysis indicated that all three CD163 variants...

  9. Anti-inflammation effect of methyl salicylate 2-O-β-D-lactoside on adjuvant induced-arthritis rats and lipopolysaccharide (LPS)-treated murine macrophages RAW264.7 cells.

    Science.gov (United States)

    Zhang, Xue; Sun, Jialin; Xin, Wenyu; Li, Yongjie; Ni, Lin; Ma, Xiaowei; Zhang, Dan; Zhang, Dongming; Zhang, Tiantai; Du, Guanhua

    2015-03-01

    Methyl salicylate 2-O-β-D-lactoside (MSL) is a derivative of natural salicylate isolated from Gaultheria yunnanensis (Franch.) Rehder, which is widely used for treating rheumatoid arthritis (RA), swelling and pain. The aim of the present study was to investigate the effect of MSL on the progression of adjuvant-induced arthritis (AIA) in rat in vivo and explore the anti-inflammatory effects and mechanism of MSL in lipopolysaccharide (LPS)-treated murine macrophages RAW264.7 cells in vitro. Our results showed that MSL significantly inhibited the arthritis progression in AIA rats, decreasing the right hind paw swelling and ankle diameter, attenuating histopathological changes and suppressing the plasma levels of TNF-α and IL-1β in AIA rats. Besides, MSL had potent anti-inflammatory effects on the LPS-activated RAW264.7. MSL dose-dependently inhibited the activity of COX-1, and COX-2. Moreover, MSL prominently inhibited LPS-induced activation of MAPK in RAW264.7 cells by blocking phosphorylation of p38 and ERK. Our study suggests that MSL may be effective in the treatment of inflammatory diseases by inhibiting the pro-inflammatory cytokine production and regulating the MAPK signal pathway. Copyright © 2015. Published by Elsevier B.V.

  10. Immunity to Schistosoma mansoni in guinea-pigs vaccinated with radiation-attenuated cercariae. T-cell activation of macrophages for larval killing

    International Nuclear Information System (INIS)

    Gordon, J.R.; McLaren, D.J.

    1988-01-01

    This study addresses macrophage activation in guinea-pigs vaccinated with radiation-attenuated cercariae of Schistosom mansoni. Peritoneal exudate macrophages elicited in vaccinated animals by mineral oil injection were activated to kill larval schistosomes in vitro. Killing efficiency is dependent upon the cell:target ratio employed and is enhanced by, but is not strictly dependent on, the presence of specific antibodies. Macrophages co-cultured with parasites release superoxide radicals and hydrogen peroxide, but the use of inhibitors has shown that neither of these reactive oxygen intermediates are the causal agents of cellular cytotoxicity in this system. Oil-elicited macrophages from naive guinea-pigs do not show comparable activation; they can, however, be activated in vitro by incubation with culture supernatant fluids from schistosome antigen-stimulated spleen, or lymph node cells harvested from vaccinated guinea-pigs. Naive macrophages activated in this way kill schistosomula in vitro and release the activation markers IL-l and superoxide anion. The macrophage-activating factor (MAF) present in spleen cell culture supernatant fluids has a MW of 35,000-55,000, but does not have the chemical characteristics of gamma-interferon. (author)

  11. Enhanced rifampicin delivery to alveolar macrophages by solid lipid nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Chuan Junlan [West China School of Pharmacy, Sichuan University, Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education (China); Li Yanzhen [Tianjin Institute of Pharmaceutical Research, State Key Laboratory of Drug Delivery Technology and Pharmacokinetics (China); Yang Likai; Sun Xun [West China School of Pharmacy, Sichuan University, Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education (China); Zhang Qiang [Peking University, State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences (China); Gong Tao, E-mail: gongtaoy@126.com; Zhang Zhirong, E-mail: zrzzl@vip.sina.com [West China School of Pharmacy, Sichuan University, Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education (China)

    2013-05-15

    The present study aimed at developing a drug delivery system targeting the densest site of tuberculosis infection, the alveolar macrophages (AMs). Rifampicin (RFP)-loaded solid lipid nanoparticles (RFP-SLNs) with an average size of 829.6 {+-} 16.1 nm were prepared by a modified lipid film hydration method. The cytotoxicity of RFP-SLNs to AMs and alveolar epithelial type II cells (AECs) was examined using MTT assays. The viability of AMs and AECs was above 80 % after treatment with RFP-SLNs, which showed low toxicity to both AMs and AECs. Confocal Laser Scanning Microscopy was employed to observe the interaction between RFP-SLNs and both AMs and AECs. After incubating the cells with RFP-SLNs for 2 h, the fluorescent intensity in AMs was more and remained longer (from 0.5 to 12 h) when compared with that in AECs (from 0.5 to 8 h). In vitro uptake characteristics of RFP-SLNs in AMs and AECs were also investigated by detection of intracellular RFP by High performance liquid chromatography. Results showed that RFP-SLNs delivered markedly higher RFP into AMs (691.7 ng/mg in cultured AMs, 662.6 ng/mg in primary AMs) than that into AECs (319.2 ng/mg in cultured AECs, 287.2 ng/mg in primary AECs). Subsequently, in vivo delivery efficiency and the selectivity of RFP-SLNs were further verified in Sprague-Dawley rats. Under pulmonary administration of RFP-SLNs, the amount of RFP in AMs was significantly higher than that in AECs at each time point. Our results demonstrated that solid lipid nanoparticles are a promising strategy for the delivery of rifampicin to alveolar macrophages selectively.

  12. In vitro cytotoxicity of Manville Code 100 glass fibers: Effect of fiber length on human alveolar macrophages

    Directory of Open Access Journals (Sweden)

    Jones William

    2006-03-01

    observed in fiber-exposed human macrophage cultures. In contrast, rat macrophages exhibited both incomplete phagocytosis of long fibers and length-dependent toxicity. The results of the human and rat cell studies suggest that incomplete engulfment may enhance cytotoxicity of fiber glass. However, the possibility should not be ruled out that differences between human versus rat macrophages other than cell diameter could account for differences in fiber effects.

  13. Autoradiographic examinations on the synthesis of the virus herpes simplex in macrophages, rabbit kidney- and BHK-cells by labelling with 3H-thymidine, 3H-uridine and UDP-3H-galactose

    International Nuclear Information System (INIS)

    Lindermann, J.

    1979-01-01

    After injection, an autoradiographically measurable reduplication of virus-specific DNA takes place in NMRI-macrophages infected with HSV (herpes simplex virus). For this process, a sufficient duration of the 3 H-thymidine pulses is of essential importance. An infection with HSV of the Lennette strain induces in considerably more cells of a macrophage population the production of virus-specific DNA than the infection with HSV of the D-316 type. After injection a reduplication of virus-specific DNA occurs in more cells of a HSV-infected population of old macrophages than in that one consisting of young macrophages. In the 7th hour after injection a reduction of RNA synthesis was observed in the HSV-infected macrophage cultures. After HSV-infection, a synthesis of virus-specific DNA could not be detected autoradiographically in macrophages of C 57/bl mice. After HSV-infection the virus-specific DNA synthesis was also observed in activated macrophages of NMRI mice, which had been treated with thioglucollate 72 hours before cell extraction. In general, the previous thioglucollate treatment increased the susceptibility of the macrophages of infection with subsequent reduplication of herpes-specific DNA. Compared to other not infected but activated macrophages, the RNA synthesis after injection was found to be not (HSV Lennette) or only minutely (HSV D-316) reduced in HSV-infected cultures of activated macrophages. In the investigations about the utilisation of exogenic UDP- 3 H-galactose by BHK and rabbit cells it was observed that only few cells of the cultures provide an externally accessible galactosyl transferase activity. Due to the influence of cell-damaging mechanisms a cell proliferation is induced, which is accompanied by externally accessible galactosyl transferase activities in the cell cultures. (orig./MG) [de

  14. Inhibition of inducible Nitric Oxide Synthase by a mustard gas analog in murine macrophages

    Directory of Open Access Journals (Sweden)

    Smith Milton

    2006-11-01

    Full Text Available Abstract Background 2-Chloroethyl ethyl sulphide (CEES is a sulphur vesicating agent and an analogue of the chemical warfare agent 2,2'-dichlorodiethyl sulphide, or sulphur mustard gas (HD. Both CEES and HD are alkylating agents that influence cellular thiols and are highly toxic. In a previous publication, we reported that lipopolysaccharide (LPS enhances the cytotoxicity of CEES in murine RAW264.7 macrophages. In the present investigation, we studied the influence of CEES on nitric oxide (NO production in LPS stimulated RAW264.7 cells since NO signalling affects inflammation, cell death, and wound healing. Murine macrophages stimulated with LPS produce NO almost exclusively via inducible nitric oxide synthase (iNOS activity. We suggest that the influence of CEES or HD on the cellular production of NO could play an important role in the pathophysiological responses of tissues to these toxicants. In particular, it is known that macrophage generated NO synthesised by iNOS plays a critical role in wound healing. Results We initially confirmed that in LPS stimulated RAW264.7 macrophages NO is exclusively generated by the iNOS form of nitric oxide synthase. CEES treatment inhibited the synthesis of NO (after 24 hours in viable LPS-stimulated RAW264.7 macrophages as measured by either nitrite secretion into the culture medium or the intracellular conversion of 4,5-diaminofluorescein diacetate (DAF-2DA or dichlorofluorescin diacetate (DCFH-DA. Western blots showed that CEES transiently decreased the expression of iNOS protein; however, treatment of active iNOS with CEES in vitro did not inhibit its enzymatic activity Conclusion CEES inhibits NO production in LPS stimulated macrophages by decreasing iNOS protein expression. Decreased iNOS expression is likely the result of CEES induced alteration in the nuclear factor kappa B (NF-κB signalling pathway. Since NO can act as an antioxidant, the CEES induced down-regulation of iNOS in LPS

  15. Atherogenicity of amino acids in the lipid-laden macrophage model system in vitro and in atherosclerotic mice: a key role for triglyceride metabolism.

    Science.gov (United States)

    Rom, Oren; Grajeda-Iglesias, Claudia; Najjar, Mahmoud; Abu-Saleh, Niroz; Volkova, Nina; Dar, Dalit Esther; Hayek, Tony; Aviram, Michael

    2017-07-01

    Atherosclerosis-related research has focused mainly on the effects of lipids on macrophage foam cell formation and atherogenesis, whereas the role of amino acids (AAs) was understudied. The current study aimed to identify anti- or pro-atherogenic AA in the macrophage model system and to elucidate the underlying metabolic and molecular mechanisms. J774A.1 cultured macrophages were treated with increasing concentrations of each 1 of the 20 AAs. Macrophage atherogenicity was assessed in terms of cellular toxicity, generation of reactive oxygen species (ROS) and cellular cholesterol or triglyceride content. At nontoxic concentrations (up to 1 mM), modest effects on ROS generation or cholesterol content were noted, but six specific AAs significantly affected macrophage triglyceride content. Glycine, cysteine, alanine and leucine significantly decreased macrophage triglyceride content (by 24%-38%), through attenuated uptake of triglyceride-rich very low-density lipoprotein (VLDL) by macrophages. In contrast, glutamate and glutamine caused a marked triglyceride accumulation in macrophages (by 107% and 129%, respectively), via a diacylglycerol acyltransferase-1 (DGAT1)-dependent increase in triglyceride biosynthesis rate with a concurrent maturation of the sterol regulatory element-binding protein-1 (SREBP1). Supplementation of apolipoprotein E-deficient (apoE -/- ) mice with glycine for 40 days significantly decreased the triglyceride levels in serum and in peritoneal macrophages (MPMs) isolated from the mice (by 19%). In contrast, glutamine supplementation significantly increased MPM ROS generation and the accumulation of cholesterol and that of triglycerides (by 48%), via enhanced uptake of LDL and VLDL. Altogether, the present findings reveal some novel roles for specific AA in macrophage atherogenicity, mainly through modulation of cellular triglyceride metabolism. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Proprotein convertase 1/3 inhibited macrophages: A novel therapeutic based on drone macrophages

    Directory of Open Access Journals (Sweden)

    Marie Duhamel

    2016-06-01

    Full Text Available We demonstrated here thanks to proteomic, that proprotein convertase 1/3 knockdown macrophages present all the characteristic of activated pro-inflammatory macrophages. TLR4 and TLR9 signaling pathways can be enhanced leading to the secretion of pro-inflammatory factors and antitumor factors. We can control their activation by controlling one enzyme, PC1/3. In a tumor context, PC1/3 inhibition in macrophages may reactivate them and lead to a cytokine storm after stimulation “at distance” with a TLR ligand. Therefore, we name these proprotein convertase inhibited macrophages the “drone macrophages”. They constitute an innovative cell therapy to treat efficiently tumors.

  17. Prolonged triglyceride storage in macrophages: pHo trumps pO2 and TLR4.

    Science.gov (United States)

    Lu, Mingfang; Kho, Terry; Munford, Robert S

    2014-08-01

    Lipid-laden macrophages contribute to pathologies as diverse as atherosclerosis and tuberculosis. Three common stimuli are known to promote macrophage lipid storage: low tissue oxygen tension (pO2), low extracellular pH (pHo), and exposure to agonists such as bacterial LPS. Noting that cells responding to low pO2 or agonistic bacterial molecules often decrease pHo by secreting lactic and other carboxylic acids, we studied how pHo influences the stimulation of triacylglycerol (TAG) storage by low pO2 and LPS. We found that TAG retention after incubation for 48-72 h was inversely related to pHo when primary macrophages were cultured in 21% oxygen, 4% oxygen, or with LPS at either oxygen concentration. Maintaining pHo at ~7.4 was sufficient to prevent the increase in prolonged TAG storage induced by either low pO2 or LPS. The strong influence of pHo on TAG retention may explain why lipid-laden macrophages are found in some tissue environments and not in others. It is also possible that other long-term cellular changes currently attributed to low pO2 or bacterial agonists may be promoted, at least in part, by the decrease in pHo that these stimuli induce.

  18. Macrophage serum markers in pneumococcal bacteremia: Prediction of survival by soluble CD163

    DEFF Research Database (Denmark)

    Møller, Holger Jon; K. Moestrup, Søren; Weis, Nina Margrethe

    2006-01-01

    OBJECTIVE: Soluble CD163 (sCD163) is a new macrophage-specific serum marker. This study investigated sCD163 and other markers of macrophage activation (neopterin, ferritin, transcobalamin, and soluble urokinase plasminogen activator receptor [suPAR]) as prognostic factors in patients...... analyses at the time of first positive blood culture. MEASUREMENTS AND MAIN RESULTS: sCD163 was highly correlated with other macrophage markers and was significantly elevated (median [25-75 percentiles], 4.6 mg/L [2.8-8.9]) compared with healthy controls (2.7 mg/L [2.1-3.3], p ..., all macrophage markers were increased in patients who died from their infection compared with survivors, whereas no change was observed in any of the markers in the very old age. At cutoff levels of 9.5 mg/L (sCD163) and 1650 nmol/L (C-reactive protein), the relative risk for fatal outcome in patients...

  19. Misbehaving macrophages in the pathogenesis of psoriasis.

    Science.gov (United States)

    Clark, Rachael A; Kupper, Thomas S

    2006-08-01

    Psoriasis is a chronic inflammatory skin disease unique to humans. In this issue of the JCI, 2 studies of very different mouse models of psoriasis both report that macrophages play a key role in inducing psoriasis-like skin disease. Psoriasis is clearly a polygenic, inherited disease of uncontrolled cutaneous inflammation. The debate that currently rages in the field is whether psoriasis is a disease of autoreactive T cells or whether it reflects an intrinsic defect within the skin--or both. However, these questions have proven difficult to dissect using molecular genetic tools. In the current studies, the authors have used 2 different animal models to address the role of macrophages in disease pathogenesis: Wang et al. use a mouse model in which inflammation is T cell dependent, whereas the model used by Stratis et al. is T cell independent (see the related articles beginning on pages 2105 and 2094, respectively). Strikingly, both groups report an important contribution by macrophages, implying that macrophages can contribute to both epithelial-based and T cell-mediated pathways of inflammation.

  20. Metabolic-epigenetic crosstalk in macrophage activation

    NARCIS (Netherlands)

    Baardman, Jeroen; Licht, Iris; de Winther, Menno P. J.; van den Bossche, Jan

    2015-01-01

    Epigenetic enzymes are emerging as crucial controllers of macrophages, innate immune cells that determine the outcome of many inflammatory diseases. Recent studies demonstrate that the activity of particular chromatin-modifying enzymes is regulated by the availability of specific metabolites like

  1. The macrophage scavenger receptor CD163

    NARCIS (Netherlands)

    Fabriek, Babs O.; Dijkstra, Christine D.; van den Berg, Timo K.

    2005-01-01

    Mature tissue macrophages form a first line of defense to recognize and eliminate potential pathogens; these specialized cells are capable of phagocytosis, degradation of self and foreign materials, establishment of cell-cell interactions, and the production of inflammatory mediators. Mature tissue

  2. Mouse adenovirus type 1 infection of macrophages

    NARCIS (Netherlands)

    Ashley, S.L.; Welton, A.R.; Harwood, K.M.; Rooijen, van N.; Spindler, K.R.

    2009-01-01

    Mouse adenovirus type 1 (MAV-1) causes acute and persistent infections in mice, with high levels of virus found in the brain, spinal cord and spleen in acute infections. MAV-1 infects endothelial cells throughout the mouse, and monocytes/macrophages have also been implicated as targets of the virus.

  3. NFAT5-Regulated Macrophage Polarization Supports the Proinflammatory Function of Macrophages and T Lymphocytes.

    Science.gov (United States)

    Tellechea, Mónica; Buxadé, Maria; Tejedor, Sonia; Aramburu, Jose; López-Rodríguez, Cristina

    2018-01-01

    Macrophages are exquisite sensors of tissue homeostasis that can rapidly switch between pro- and anti-inflammatory or regulatory modes to respond to perturbations in their microenvironment. This functional plasticity involves a precise orchestration of gene expression patterns whose transcriptional regulators have not been fully characterized. We had previously identified the transcription factor NFAT5 as an activator of TLR-induced responses, and in this study we explore its contribution to macrophage functions in different polarization settings. We found that both in classically and alternatively polarized macrophages, NFAT5 enhanced functions associated with a proinflammatory profile such as bactericidal capacity and the ability to promote Th1 polarization over Th2 responses. In this regard, NFAT5 upregulated the Th1-stimulatory cytokine IL-12 in classically activated macrophages, whereas in alternatively polarized ones it enhanced the expression of the pro-Th1 mediators Fizz-1 and arginase 1, indicating that it could promote proinflammatory readiness by regulating independent genes in differently polarized macrophages. Finally, adoptive transfer assays in vivo revealed a reduced antitumor capacity in NFAT5-deficient macrophages against syngeneic Lewis lung carcinoma and ID8 ovarian carcinoma cells, a defect that in the ID8 model was associated with a reduced accumulation of effector CD8 T cells at the tumor site. Altogether, detailed analysis of the effect of NFAT5 in pro- and anti-inflammatory macrophages uncovered its ability to regulate distinct genes under both polarization modes and revealed its predominant role in promoting proinflammatory macrophage functions. Copyright © 2017 by The American Association of Immunologists, Inc.

  4. DMPD: The oxidation of lipoproteins by monocytes-macrophages. Biochemical andbiological mechanisms. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 10473535 The oxidation of lipoproteins by monocytes-macrophages. Biochemical andbio.... (.png) (.svg) (.html) (.csml) Show The oxidation of lipoproteins by monocytes-macrophages. Biochemical and...onocytes-macrophages. Biochemical andbiological mechanisms. Authors Chisolm GM 3rd, Hazen SL, Fox PL, Cathca

  5. DMPD: Pathogen-induced apoptosis of macrophages: a common end for different pathogenicstrategies. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 11207583 Pathogen-induced apoptosis of macrophages: a common end for different path...ml) Show Pathogen-induced apoptosis of macrophages: a common end for different pathogenicstrategies. PubmedI...D 11207583 Title Pathogen-induced apoptosis of macrophages: a common end for diff

  6. DMPD: Mechanism of age-associated up-regulation in macrophage PGE2 synthesis. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15331118 Mechanism of age-associated up-regulation in macrophage PGE2 synthesis. Wu...e-associated up-regulation in macrophage PGE2 synthesis. PubmedID 15331118 Title Mechanism of age-associated... up-regulation in macrophage PGE2 synthesis. Authors Wu D, Meydani SN. Publicatio

  7. MiR-146a modulates macrophage polarization by inhibiting Notch1 pathway in RAW264.7 macrophages.

    Science.gov (United States)

    Huang, Cheng; Liu, Xue-Jiao; QunZhou; Xie, Juan; Ma, Tao-Tao; Meng, Xiao-Ming; Li, Jun

    2016-03-01

    Macrophages are heterogeneous and plastic cells which are able to undergo dynamic transition between M1 and M2 polarized phenotypes in response to the microenvironment signals. However, the underlying molecular mechanisms of macrophage polarization are still obscure. In the current study, it was revealed that miR-146a might play a pivotal role in macrophage polarization. As our results indicated, miR-146a was highly expressed in M2 macrophages rather than M1 macrophages. Over-expression of miR-146a resulted in significantly decreased production of pro-inflammatory cytokines including iNOS and TNF-α in M1 macrophages, while increased production of M2 marker genes such as Arg1 and CD206 in M2 macrophages. In contrast, knockdown of miR-146a promoted M1 macrophage polarization but diminished M2 macrophage polarization. Mechanistically, it was revealed that miR-146a modulated macrophage polarization by targeting Notch1. Of note, PPARγ was responsible as another target for miR-146a-mediated macrophage polarization. Taken together, it was suggested that miR-146a might serve as a molecular regulator in macrophage polarization and is a potential therapeutic target for inflammatory diseases. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. DMPD: Molecular mechanisms of macrophage activation and deactivation bylipopolysaccharide: roles of the receptor complex. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 14609719 Molecular mechanisms of macrophage activation and deactivation bylipopolys...acol Ther. 2003 Nov;100(2):171-94. (.png) (.svg) (.html) (.csml) Show Molecular mechanisms of macrophage act...medID 14609719 Title Molecular mechanisms of macrophage activation and deactivation bylipopolysaccharide: ro

  9. DMPD: Genetic regulation of macrophage priming/activation: the Lsh gene story. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 1757110 Genetic regulation of macrophage priming/activation: the Lsh gene story. Bl... (.svg) (.html) (.csml) Show Genetic regulation of macrophage priming/activation: the Lsh gene story. Pubmed...ID 1757110 Title Genetic regulation of macrophage priming/activation: the Lsh gen

  10. Yersinia pestis and host macrophages: immunodeficiency of mouse macrophages induced by YscW.

    Science.gov (United States)

    Bi, Yujing; Du, Zongmin; Han, Yanping; Guo, Zhaobiao; Tan, Yafang; Zhu, Ziwen; Yang, Ruifu

    2009-09-01

    The virulence of the pathogenic Yersinia species depends on a plasmid-encoded type III secretion system (T3SS) that transfers six Yersinia outer protein (Yop) effector proteins into the cytoplasm of eukaryotic cells, leading to disruption of host defence mechanisms. It is shown in this study that Yersinia pestis YscW, a protein of the T3SS injectisome, contributes to the induction of a deficiency in phagocytosis in host macrophages and a reduction in their antigen-presenting capacity. A Y. pestis strain lacking yscW had no effect on uptake by host macrophages. In mice infected with wild-type Y. pestis, the yscW mutant or a complement strain, immunodeficiency was observed in host macrophages compared with those from uninfected mice. However, the phagocytosis and antigen presenting capacities of macrophages infected by yscW mutant strain both in vivo and in vitro were significantly higher than those by wild type strain. Consistent with this finding, when YscW was expressed in the RAW264.7 macrophage cell line, phagocytosis and antigen-presenting capacities were significantly lower than those of the control groups. These results indicate that Y. pestis YscW may directly induce immunodeficiency in murine macrophages by crippling their phagocytosis and antigen-presenting capacities. These data provide evidences to Y. pestis pathogenesis that some proteins in T3SS injectisome, such as YscW protein, might play independent roles in disrupting host defense apart from their known functions.

  11. Macrophage-Mediated Lymphangiogenesis: The Emerging Role of Macrophages as Lymphatic Endothelial Progenitors

    International Nuclear Information System (INIS)

    Ran, Sophia; Montgomery, Kyle E.

    2012-01-01

    It is widely accepted that macrophages and other inflammatory cells support tumor progression and metastasis. During early stages of neoplastic development, tumor-infiltrating macrophages (TAMs) mount an immune response against transformed cells. Frequently, however, cancer cells escape the immune surveillance, an event that is accompanied by macrophage transition from an anti-tumor to a pro-tumorigenic type. The latter is characterized by high expression of factors that activate endothelial cells, suppress immune response, degrade extracellular matrix, and promote tumor growth. Cumulatively, these products of TAMs promote tumor expansion and growth of both blood and lymphatic vessels that facilitate metastatic spread. Breast cancers and other epithelial malignancies induce the formation of new lymphatic vessels (i.e., lymphangiogenesis) that leads to lymphatic and subsequently, to distant metastasis. Both experimental and clinical studies have shown that TAMs significantly promote tumor lymphangiogenesis through paracrine and cell autonomous modes. The paracrine effect consists of the expression of a variety of pro-lymphangiogenic factors that activate the preexisting lymphatic vessels. The evidence for cell-autonomous contribution is based on the observed tumor mobilization of macrophage-derived lymphatic endothelial cell progenitors (M-LECP) that integrate into lymphatic vessels prior to sprouting. This review will summarize the current knowledge of macrophage-dependent growth of new lymphatic vessels with specific emphasis on an emerging role of macrophages as lymphatic endothelial cell progenitors (M-LECP)

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

    OpenAIRE

    Jorj?o, Adeline Lacerda; de Oliveira, Felipe Eduardo; Le?o, Mariella Vieira Pereira; Carvalho, Cl?udio 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-?, IL-1?, 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 cul...

  13. Moracin C, A Phenolic Compound Isolated from Artocarpus heterophyllus, Suppresses Lipopolysaccharide-Activated Inflammatory Responses in Murine Raw264.7 Macrophages

    Directory of Open Access Journals (Sweden)

    Xue Yao

    2016-07-01

    Full Text Available Artocarpus heterophyllus, a popular tropical fruit commonly known as the jackfruit tree, is normally planted in subtropical or tropical areas. Since a variety of phytochemicals isolated from A. heterophyllus have been found to possess potently anti-inflammatory, antiviral and antimalarial activities, researchers have devoted much interest to its potential pharmaceutical value. However, the exact mechanism underlying its anti-inflammatory activity is not well characterized. In this study, seven natural products isolated from A. heterophyllus, including 25-Hydroxycycloart-23-en-3-one (HY, Artocarpin (AR, Dadahol A (DA, Morachalcone A (MA, Artoheterophyllin B (AB, Cycloheterophyllin (CY and Moracin C (MC were collected. Lipopolysaccharide (LPS-stimulated inflammatory response in RAW264.7 macrophages were used in this study. Among these compounds, MC significantly inhibited LPS-activated reactive oxygen species (ROS and nitric oxide (NO release without marked cytotoxicity. Furthermore, MC effectively reduced LPS stimulated up-regulation of mRNA and protein expression of inducible nitric oxide synthase (iNOS, cyclooxygenase-2 (COX-2, and serval pro-inflammatory cytokines (interleukin-1β (IL-1β, interleukin-6 (IL-6 and tumor necrosis factor α (TNF-α. Mechanistic studies revealed that the anti-inflammatory effect of MC was associated with the activation of the mitogen activated protein kinases (MAPKs (including p38, ERK and JNK and nuclear factor-κB (NF-κB pathways, especially reducing the nuclear translocation of NF-κB p65 subunit as revealed by nuclear separation experiment and confocal microscopy.

  14. Mechanism of inhibitory effect of atorvastatin on resistin expression induced by tumor necrosis factor-α in macrophages

    Directory of Open Access Journals (Sweden)

    Chua Su-Kiat

    2009-05-01

    Full Text Available Abstract Atorvastatin has been shown to reduce resistin expression in macrophages after pro-inflammatory stimulation. However, the mechanism of reducing resistin expression by atorvastatin is not known. Therefore, we sought to investigate the molecular mechanisms of atorvastatin for reducing resistin expression after proinflammatory cytokine, tumor necrosis factor-α (TNF-α stimulation in cultured macrophages. Cultured macrophages were obtained from human peripheral blood mononuclear cells. TNF-α stimulation increased resistin protein and mRNA expression and atorvastatin inhibited the induction of resistin by TNF-α. Addition of mevalonate induced resistin protein expression similar to TNF-α stimulation. However, atorvastatin did not have effect on resistin protein expression induced by mevalonate. SP600125 and JNK small interfering RNA (siRNA completely attenuated the resistin protein expression induced by TNF-α and mevalonate. TNF-α induced phosphorylation of Rac, while atorvastatin and Rac-1 inhibitor inhibited the phosphorylation of Rac induced by TNF-α. The gel shift and promoter activity assay showed that TNF-α increased AP-1-binding activity and resistin promoter activity, while SP600125 and atorvastatin inhibited the AP-1-binding activity and resistin promoter activity induced by TNF-α. Recombinant resistin and TNF-α significantly reduced glucose uptake in cultured macrophages, while atorvastatin reversed the reduced glucose uptake by TNF-α. In conclusion, JNK and Rac pathway mediates the inhibitory effect of atorvastatin on resistin expression induced by TNF-α.

  15. Molecular Mechanisms Modulating the Phenotype of Macrophages and Microglia

    Directory of Open Access Journals (Sweden)

    Stephanie A. Amici

    2017-11-01

    Full Text Available Macrophages and microglia play crucial roles during central nervous system development, homeostasis and acute events such as infection or injury. The diverse functions of tissue macrophages and microglia are mirrored by equally diverse phenotypes. A model of inflammatory/M1 versus a resolution phase/M2 macrophages has been widely used. However, the complexity of macrophage function can only be achieved by the existence of varied, plastic and tridimensional macrophage phenotypes. Understanding how tissue macrophages integrate environmental signals via molecular programs to define pathogen/injury inflammatory responses provides an opportunity to better understand the multilayered nature of macrophages, as well as target and modulate cellular programs to control excessive inflammation. This is particularly important in MS and other neuroinflammatory diseases, where chronic inflammatory macrophage and microglial responses may contribute to pathology. Here, we perform a comprehensive review of our current understanding of how molecular pathways modulate tissue macrophage phenotype, covering both classic pathways and the emerging role of microRNAs, receptor-tyrosine kinases and metabolism in macrophage phenotype. In addition, we discuss pathway parallels in microglia, novel markers helpful in the identification of peripheral macrophages versus microglia and markers linked to their phenotype.

  16. Macrophage Polarization in Chronic Inflammatory Diseases: Killers or Builders?

    Science.gov (United States)

    Baci, Denisa; Tremolati, Marco; Fanuli, Matteo; Farronato, Giampietro; Mortara, Lorenzo

    2018-01-01

    Macrophages are key cellular components of the innate immunity, acting as the main player in the first-line defence against the pathogens and modulating homeostatic and inflammatory responses. Plasticity is a major feature of macrophages resulting in extreme heterogeneity both in normal and in pathological conditions. Macrophages are not homogenous, and they are generally categorized into two broad but distinct subsets as either classically activated (M1) or alternatively activated (M2). However, macrophages represent a continuum of highly plastic effector cells, resembling a spectrum of diverse phenotype states. Induction of specific macrophage functions is closely related to the surrounding environment that acts as a relevant orchestrator of macrophage functions. This phenomenon, termed polarization, results from cell/cell, cell/molecule interaction, governing macrophage functionality within the hosting tissues. Here, we summarized relevant cellular and molecular mechanisms driving macrophage polarization in “distant” pathological conditions, such as cancer, type 2 diabetes, atherosclerosis, and periodontitis that share macrophage-driven inflammation as a key feature, playing their dual role as killers (M1-like) and/or builders (M2-like). We also dissect the physio/pathological consequences related to macrophage polarization within selected chronic inflammatory diseases, placing polarized macrophages as a relevant hallmark, putative biomarkers, and possible target for prevention/therapy. PMID:29507865

  17. Macrophage Polarization in Chronic Inflammatory Diseases: Killers or Builders?

    Directory of Open Access Journals (Sweden)

    Luca Parisi

    2018-01-01

    Full Text Available Macrophages are key cellular components of the innate immunity, acting as the main player in the first-line defence against the pathogens and modulating homeostatic and inflammatory responses. Plasticity is a major feature of macrophages resulting in extreme heterogeneity both in normal and in pathological conditions. Macrophages are not homogenous, and they are generally categorized into two broad but distinct subsets as either classically activated (M1 or alternatively activated (M2. However, macrophages represent a continuum of highly plastic effector cells, resembling a spectrum of diverse phenotype states. Induction of specific macrophage functions is closely related to the surrounding environment that acts as a relevant orchestrator of macrophage functions. This phenomenon, termed polarization, results from cell/cell, cell/molecule interaction, governing macrophage functionality within the hosting tissues. Here, we summarized relevant cellular and molecular mechanisms driving macrophage polarization in “distant” pathological conditions, such as cancer, type 2 diabetes, atherosclerosis, and periodontitis that share macrophage-driven inflammation as a key feature, playing their dual role as killers (M1-like and/or builders (M2-like. We also dissect the physio/pathological consequences related to macrophage polarization within selected chronic inflammatory diseases, placing polarized macrophages as a relevant hallmark, putative biomarkers, and possible target for prevention/therapy.

  18. HIV Infection of Macrophages: Implications for Pathogenesis and Cure

    Directory of Open Access Journals (Sweden)

    Kiera Leigh Clayton

    2017-05-01

    Full Text Available Although CD4+ T cells represent the major reservoir of persistent HIV and SIV infection, accumulating evidence suggests that macrophages also contribute. However, investigations of the role of macrophages are often underrepresented at HIV pathogenesis and cure meetings. This was the impetus for a scientific workshop dedicated to this area of study, held in Cambridge, MA in January 2017. The workshop brought together experts in the fields of HIV/SIV immunology/virology, macrophage biology and immunology, and animal models of HIV/SIV infection to facilitate discussions regarding the role of macrophages as a physiologically relevant viral reservoir, and the implications of macrophage infection for HIV pathogenesis and cure strategies. An emerging consensus that infected macrophages likely persist in the setting of combination antiretroviral therapy, driving persistent inflammation and contributing to the viral reservoir, indicate the importance of addressing macrophages as well as CD4+ T cells with future therapeutic strategies.

  19. Lysosomal Disorders Drive Susceptibility to Tuberculosis by Compromising Macrophage Migration

    Science.gov (United States)

    Berg, Russell D.; Levitte, Steven; O’Sullivan, Mary P.; O’Leary, Seónadh M.; Cambier, C.J.; Cameron, James; Takaki, Kevin K.; Moens, Cecilia B.; Tobin, David M.; Keane, Joseph; Ramakrishnan, Lalita

    2016-01-01

    Summary A zebrafish genetic screen for determinants of susceptibility to Mycobacterium marinum identified a hypersusceptible mutant deficient in lysosomal cysteine cathepsins that manifests hallmarks of human lysosomal storage diseases. Under homeostatic conditions, mutant macrophages accumulate undigested lysosomal material, which disrupts endocytic recycling and impairs their migration to, and thus engulfment of, dying cells. This causes a buildup of unengulfed cell debris. During mycobacterial infection, macrophages with lysosomal storage cannot migrate toward infected macrophages undergoing apoptosis in the tuberculous granuloma. The unengulfed apoptotic macrophages undergo secondary necrosis, causing granuloma breakdown and increased mycobacterial growth. Macrophage lysosomal storage similarly impairs migration to newly infecting mycobacteria. This phenotype is recapitulated in human smokers, who are at increased risk for tuberculosis. A majority of their alveolar macrophages exhibit lysosomal accumulations of tobacco smoke particulates and do not migrate to Mycobacterium tuberculosis. The incapacitation of highly microbicidal first-responding macrophages may contribute to smokers’ susceptibility to tuberculosis. PMID:27015311

  20. CD163-Macrophages Are Involved in Rhabdomyolysis-Induced Kidney Injury and May Be Detected by MRI with Targeted Gold-Coated Iron Oxide Nanoparticles.

    Science.gov (United States)

    Rubio-Navarro, Alfonso; Carril, Mónica; Padro, Daniel; Guerrero-Hue, Melanie; Tarín, Carlos; Samaniego, Rafael; Cannata, Pablo; Cano, Ainhoa; Villalobos, Juan Manuel Amaro; Sevillano, Ángel Manuel; Yuste, Claudia; Gutiérrez, Eduardo; Praga, Manuel; Egido, Jesús; Moreno, Juan Antonio

    2016-01-01

    Macrophages play an important role in rhabdomyolysis-acute kidney injury (AKI), although the molecular mechanisms involved in macrophage differentiation are poorly understood. We analyzed the expression and regulation of CD163, a membrane receptor mainly expressed by anti-inflammatory M2 macrophages, in rhabdomyolysis-AKI and developed targeted probes for its specific detection in vivo by MRI. Intramuscular injection of glycerol in mice promoted an early inflammatory response, with elevated proportion of M1 macrophages, and partial differentiation towards a M2 phenotype in later stages, where increased CD163 expression was observed. Immunohistological studies confirmed the presence of CD163-macrophages in human rhabdomyolysis-AKI. In cultured macrophages, myoglobin upregulated CD163 expression via HO-1/IL-10 axis. Moreover, we developed gold-coated iron oxide nanoparticles vectorized with an anti-CD163 antibody that specifically targeted CD163 in kidneys from glycerol-injected mice, as determined by MRI studies, and confirmed by electron microscopy and immunological analysis. Our findings are the first to demonstrate that CD163 is present in both human and experimental rhabdomyolysis-induced AKI, suggesting an important role of this molecule in this pathological condition. Therefore, the use of probes targeting CD163-macrophages by MRI may provide important information about the cellular composition of renal lesion in rhabdomyolysis.

  1. Macrophage Immune Response Suppression by Recombinant Mycobacterium tuberculosis Antigens, the ESAT-6, CFP-10, and ESAT-6/CFP-10 Fusion Proteins

    Science.gov (United States)

    Seghatoleslam, Atefeh; Hemmati, Mina; Ebadat, Saeedeh; Movahedi, Bahram; Mostafavi-Pour, Zohreh

    2016-01-01

    Background: Macrophage immune responses are affected by the secretory proteins of Mycobacterium tuberculosis (Mtb). This study aimed to examine the immune responses of macrophages to Mtb secretory antigens, namely ESAT-6, CFP-10, and ESAT-6/CFP-10. Methods: THP-1 cells (a human monocytic cell line) were cultured and differentiated to macrophages by phorbol 12-myristate 13-acetate. The cytotoxicity of the recombinant Mtb proteins was assessed using the MTT assay. Two important immune responses of macrophages, namely NO and ROS production, were measured in response to the ESAT-6, CFP-10, and ESAT-6/CFP-10 antigens. The data were analyzed using one-way ANOVA with SPSS, version 16, and considered significant at Pproteins markedly reduced macrophage immune response. The treatment of the THP-1-differentiated cells with ESAT-6, CFP-10, and ESAT-6/CFP-10 reduced NO and ROS production. The treated THP-1-differentiated cells exhibited less inducible NO synthase activity than did the untreated cells. No toxic effect on macrophage viability was observed for the applied proteins at the different concentrations. Conclusion: It seems that the decline in macrophage immune response is due to the suppression of NO and ROS production pathways without any effect on cell viability. PMID:27365551

  2. Studies on the binding and transport processes of americium-241 hydroxide polymers in rat lung and bovine alveolar macrophages

    International Nuclear Information System (INIS)

    Taya, A.

    1986-03-01

    The binding of Am-241 hydroxide polymers to the cell components of rat lung was investigated using differential centrifugation, density gradient centrifugation with different media, gel chromatography, free flow electrophoresis and electron microscopic autoradiography with Pu-241. The bovine alveolar macrophage cultures were introduced as an in vitro test system for Am-241 uptake. Form the biochemical and electron microscopic studies it can be concluded that Am-241 is taken up by pulmonary macrophages, where its first storage site is probably the lysosome. Then the Am-241 seems to be solubilized in the lysosomes and to be bound to the cytosolic ferritin of macrophages. Am-241 might be released from the cells and crosses the alveolar membranes as bound to transferrin or as low molecular weight form. (orig.) [de

  3. Ex vivo expansion of alveolar macrophages with Mycobacterium tuberculosis from the resected lungs of patients with pulmonary tuberculosis

    Science.gov (United States)

    Petrunina, Ekaterina; Umpeleva, Tatiana; Karskanova, Svetlana; Bayborodin, Sergey; Vakhrusheva, Diana; Kravchenko, Marionella; Skornyakov, Sergey

    2018-01-01

    Tuberculosis (TB), with the Mycobacterium tuberculosis (Mtb) as the causative agent, remains to be a serious world health problem. Traditional methods used for the study of Mtb in the lungs of TB patients do not provide information about the number and functional status of Mtb, especially if Mtb are located in alveolar macrophages. We have developed a technique to produce ex vivo cultures of cells from different parts of lung tissues surgically removed from patients with pulmonary TB and compared data on the number of cells with Mtb inferred by the proposed technique to the results of bacteriological and histological analyses used for examination of the resected lungs. The ex vivo cultures of cells obtained from the resected lungs of all patients were largely composed of CD14-positive alveolar macrophages, foamy or not, with or without Mtb. Lymphocytes, fibroblasts, neutrophils, and multinucleate Langhans giant cells were also observed. We found alveolar macrophages with Mtb in the ex vivo cultures of cells from the resected lungs of even those TB patients, whose sputum smears and lung tissues did not contain acid-fast Mtb or reveal growing Mtb colonies on dense medium. The detection of alveolar macrophages with Mtb in ex vivo culture as soon as 16–18 h after isolation of cells from the resected lungs of all TB patients suggests that the technique proposed for assessing the level of infection in alveolar macrophages of TB patients has higher sensitivity than do prolonged bacteriological or pathomorphological methods. The proposed technique allowed us to rapidly (in two days after surgery) determine the level of infection with Mtb in the cells of the resected lungs of TB patients and, by the presence or absence of Mtb colonies, including those with cording morphology, the functional status of the TB agent at the time of surgery. PMID:29401466

  4. circRNA Mediates Silica-Induced Macrophage Activation Via HECTD1/ZC3H12A-Dependent Ubiquitination

    Science.gov (United States)

    Zhou, Zewei; Jiang, Rong; Yang, Xiyue; Guo, Huifang; Fang, Shencun; Zhang, Yingming; Cheng, Yusi; Wang, Jing; Yao, Honghong; Chao, Jie

    2018-01-01

    Rationale: Phagocytosis of silicon dioxide (SiO2) into lung cells causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Circular RNAs (circRNAs) are a subclass of non-coding RNAs detected within mammalian cells; however, researchers have not determined whether circRNAs are involved in the pathophysiological process of silicosis. The upstream molecular mechanisms and functional effects on cell apoptosis, proliferation and migration were investigated to elucidate the role of circRNAs in SiO2-induced inflammation in pulmonary macrophages. Methods: Primary cultures of alveolar macrophages from healthy donors and patients as well as the RAW264.7 macrophage cell line were used to explore the functions of circHECTD1 (HECT domain E3 ubiquitin protein ligase 1) in macrophage activation. Results: The results of the experiments indicated that 1) SiO2 concomitantly decreased circHECTD1 levels and increased HECTD1 protein expression; 2) circHECTD1 and HECTD1 were involved in SiO2-induced macrophage activation via ubiquitination; and 3) SiO2-activated macrophages promoted fibroblast proliferation and migration via the circHECTD1/HECTD1 pathway. Tissue samples from silicosis patients confirmed the upregulation of HECTD1. Conclusions: Our study elucidated a link between SiO2-induced macrophage activation and the circHECTD1/HECTD1 pathway, thereby providing new insight into the potential use of HECTD1 in the development of novel therapeutic strategies for treating silicosis. PMID:29290828

  5. circRNA Mediates Silica-Induced Macrophage Activation Via HECTD1/ZC3H12A-Dependent Ubiquitination.

    Science.gov (United States)

    Zhou, Zewei; Jiang, Rong; Yang, Xiyue; Guo, Huifang; Fang, Shencun; Zhang, Yingming; Cheng, Yusi; Wang, Jing; Yao, Honghong; Chao, Jie

    2018-01-01

    Rationale: Phagocytosis of silicon dioxide (SiO 2 ) into lung cells causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Circular RNAs (circRNAs) are a subclass of non-coding RNAs detected within mammalian cells; however, researchers have not determined whether circRNAs are involved in the pathophysiological process of silicosis. The upstream molecular mechanisms and functional effects on cell apoptosis, proliferation and migration were investigated to elucidate the role of circRNAs in SiO 2 -induced inflammation in pulmonary macrophages. Methods: Primary cultures of alveolar macrophages from healthy donors and patients as well as the RAW264.7 macrophage cell line were used to explore the functions of circHECTD1 (HECT domain E3 ubiquitin protein ligase 1) in macrophage activation. Results: The results of the experiments indicated that 1) SiO 2 concomitantly decreased circHECTD1 levels and increased HECTD1 protein expression; 2) circHECTD1 and HECTD1 were involved in SiO 2 -induced macrophage activation via ubiquitination; and 3) SiO 2 -activated macrophages promoted fibroblast proliferation and migration via the circHECTD1/HECTD1 pathway. Tissue samples from silicosis patients confirmed the upregulation of HECTD1. Conclusions: Our study elucidated a link between SiO 2 -induced macrophage activation and the circHECTD1/HECTD1 pathway, thereby providing new insight into the potential use of HECTD1 in the development of novel therapeutic strategies for treating silicosis.

  6. Tumor cell-macrophage interactions increase angiogenesis through secretion of EMMPRIN

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    Bat-Chen eAmit-Cohen

    2013-07-01

    Full Text Available Tumor macrophages are generally considered to be alternatively/M2 activated to induce secretion of pro-angiogenic factors such as VEGF and MMPs. EMMPRIN (CD147, basigin is overexpressed in many tumor types, and has been shown to induce fibroblasts and endothelial cell expression of MMPs and VEGF. We first show that tumor cell interactions with macrophages resulted in increased expression of EMMPRIN and induction of MMP-9 and VEGF. Human A498 renal carcinoma or MCF-7 breast carcinoma cell lines were co-cultured with the U937 monocytic-like cell line in the presence of TNFalpha (1 ng/ml. Membranal EMMPRIN expression was increased in the co-cultures (by 3-4 folds, p<0.01, as was the secretion of MMP-9 and VEGF (by 2-5 folds for both MMP-9 and VEGF, p<0.01, relative to the single cultures with TNFalpha. Investigating the regulatory mechanisms, we show that EMMPRIN was post-translationally regulated by miR-146a, as no change was observed in the tumoral expression of EMMPRIN mRNA during co-culture, expression of miR-146a was increased and its neutralization by its antagomir inhibited EMMPRIN expression. The secretion of EMMPRIN was also enhanced (by 2-3 folds, p<0.05, only in the A498 co-culture via shedding off of the membranal protein by a serine protease that is yet to be identified, as demonstrated by the use of wide range protease inhibitors. Finally, soluble EMMPRIN enhanced monocytic secretion of MMP-9 and VEGF, as inhibition of its expression levels by neutralizing anti-EMMPRIN or siRNA in the tumor cells lead to subsequent decreased induction of these two pro-angiogenic proteins. These results reveal a mechanism whereby tumor cell-macrophage interactions promote angiogenesis via an EMMPRIN-mediated pathway.

  7. Evaluation of the alveolar macrophage role in the pulmonary distribution of actinide oxides

    International Nuclear Information System (INIS)

    Guezingar-Liebard, Florence

    1999-01-01

    Actinide oxide inhalation is potentially a risk during the fuel fabrication process in the electronuclear industry. These particles can induce pulmonary lesions. The alveolar macrophage play an important role in the particle sequestration and transport but the actinide toxicity towards these cells is not well known. The aim of this work was to characterize the evolution of particle localisation in lungs after inhalation and to evaluate the role of macrophages in the lesion histo-genesis. We have used of a solid track detector to visualise alpha dose distribution within lung tissue. After 237 NpO 2 , MOX or PuO 2 inhalation by rats, different kinetics of clearance were observed for the sub-pleural and peri-bronchial areas compared to the others alveolar areas. For initial lung burdens that alter the lung clearance, particle aggregates were observed. Their kinetic and localisation vary depending on the aerosol, for a same global dose delivered to the lungs. This could be due to the different specific alpha activities of the particles and to the particle number deposited in the lung to obtain a similar burden but it could be also due to a chemical toxicity of neptunium higher than that of the others actinides. The flow cytometry methods developed allow us to measure apoptosis, phagocytosis and free radicals generation. After addition of soluble uranium to the culture medium, similar results were obtained using either alveolar macrophages extracted from rats or a macrophage cell line. This work confirms that alveolar macrophages are involved in the aggregate formation which induces heterogeneous dose distribution within the different lung tissues. (author) [fr

  8. Voluntary exercise attenuates obesity-associated inflammation through ghrelin expressed in macrophages.

    Science.gov (United States)

    Kizaki, Takako; Maegawa, Taketeru; Sakurai, Takuya; Ogasawara, Jun-etsu; Ookawara, Tomomi; Oh-ishi, Shuji; Izawa, Tetsuya; Haga, Shukoh; Ohno, Hideki

    2011-09-30

    Chronic low-level inflammation is associated with obesity and a sedentary lifestyle, causing metabolic disturbances such as insulin resistance. Exercise training has been shown to decrease chronic low-level systemic inflammation in high-fat diet (HFD)-induced obesity. However, the molecular mechanisms mediating its beneficial effects are not fully understood. Ghrelin is a peptide hormone predominantly produced in the stomach that stimulates appetite and induces growth hormone release. In addition to these well-known functions, recent studies suggest that ghrelin localizes to immune cells and exerts an anti-inflammatory effect. The purpose of the current study was to investigate the role of ghrelin expressed in macrophages in the anti-inflammatory effects of voluntary exercise training. Expression of tumor necrosis factor-α (TNF-α), monocyte chemotactic protein (MCP)-1 and F4/80 was increased in adipose tissue from mice fed a HFD (HFD mice) compared with mice fed a standard diet (SD mice), whereas the expression of these inflammatory cytokines was markedly decreased in mice performing voluntary wheel running during the feeding of a HFD (HFEx mice). The expression of TNF-α was also increased in peritoneal macrophages by a HFD and exercise training inhibited the increase of TNF-α expression. Interestingly, expression of ghrelin in peritoneal macrophages was decreased by a HFD and recovered by exercise training. Suppression of ghrelin expression by siRNA increased TNF-α expression and LPS-stimulated NF-κB activation in RAW264 cells, which is a macrophage cell line. TNF-α expression by stimulation with LPS was significantly suppressed in RAW264 cells cultured in the presence of ghrelin. These results suggest that ghrelin exerts potent anti-inflammatory effects in macrophages and functions as a mediator of the beneficial effects of exercise training. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. Do chondroitin sulfates with different structures have different activities on chondrocytes and macrophages?

    Science.gov (United States)

    da Cunha, André L; Aguiar, Jair A K; Correa da Silva, Flavio S; Michelacci, Yara M

    2017-10-01

    The aim of the present study was to investigate the activities of natural chondroitin sulfates (CS) with different structures on cultured chondrocytes and macrophages. CS were isolated from cartilages of bovine trachea (BT), porcine trachea (PT), chicken sternum (Ch) and skate (Sk). The preparations were 90-98% pure, with ∼1% proteins, nucleic acids and keratan sulfate contaminants. Structural analysis of these CS and of commercial chondroitin 4- and 6-sulfate (C4S, C6S) have shown that most of their disaccharides are monosulfated, with varying proportions of 4- and 6-sulfation, and 2-7% non-sulfated disaccharides. Sk-CS and C6S contained detectable amounts of disulfated disaccharides. All the CS were polydisperse, with modal molecular weights of 26-135kDa. These CS had anti-inflammatory activities on both chondrocytes and macrophages, but with different efficiencies. On horse and human chondrocytes, they reduced the IL-1β-induced liberation of NO and PGE 2 , and on RAW 264.7 immortalized macrophage-like cell line, C4S, C6S, Ch and Sk-CS decreased the LPS-induced liberation of TNF-α, but did not affect IL-6. In contrast, on bone marrow derived macrophages, C4S, C6S, BT and PT-CS reduced the LPS-induced liberation of TNF-α, IL-6, IL-1β and NO, indicating that the RAW response to CS was different from that of primary macrophages. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. In vitro biocorrosion of Co-Cr-Mo implant alloy by macrophage cells.

    Science.gov (United States)

    Lin, Hsin-Yi; Bumgardner, Joel D

    2004-11-01

    We hypothesized that macrophage cells and their released reactive chemical species (RCS) affect Co-Cr-Mo alloy's corrosion properties and that alloy corrosion products change macrophage cell behavior. A custom cell culture corrosion cell was used to evaluate how culture medium, cells, and RCS altered alloy corrosion in 3-day tests. Corrosion was evaluated by measuring total charge transfer at a constant potential using a potentiostat and metal ion release by atomic emission spectroscopy. Viability, proliferation, and NO (nitric oxide) and IL-1beta (interlukin-1beta) release were used to assess cellular response to alloy corrosion products. In the presence of activated cells, total charge transfers and Co ion release were the lowest (p < 0.05). This was attributed to an enhancement of the surface oxide by RCS. Cr and Mo release were not different between cells and activated cells. Low levels of metal ions did not affect cell viability, proliferation, or NO release, though IL-1beta released from the activated cells was higher on the alloy compared to the controls. These data support the hypothesis that macrophage cells and their RCS affect alloy corrosion. Changes in alloy corrosion by cells may be important to the development of host responses to the alloy and its corrosion products.

  11. Up-regulation of TLR2 and TLR4 in high mobility group Box1-stimulated macrophages in pulpitis patients

    Science.gov (United States)

    Mahmoudi, Javad; Sabermarouf, Babak; Baradaran, Behzad; Sadat-Hatamnezhad, Leila; Shotorbani, Siamak Sandoghchian

    2017-01-01

    Objective(s): High Mobility Group Box1 (HMGB1) is a nonhistone, DNA-binding protein that serves a crucial role in regulating gene transcription and is involved in a variety of proinflammatory, extracellular activities. The aim of this study was to explore whether HMGB1 stimulation can up-regulate the expression of Toll-like Receptor 2 (TLR2) and Toll-like Receptor 4 (TLR4) on macrophages from pulpitis and to clarify the subsequent events involving Th17 cells and Th17 cell-associated cytokine changes. Materials and Methods: Having prepared dental pulp tissues of pulpitis and healthy controls, macrophage were isolated and cultured. Macrophages were thereafter stimulated by HMGB1 time course. RT-QPCR, flowcytometer, immunofluorescence, Western blotting, and ELISA techniques were used in the present research. Results: Our results showed that the expression of TLR2 and TLR4 on macrophages stimulated with HMGB1 increased in pulpitis compared with controls (macrophages without HMGB1 stimulation) with a statistical significance (Ppulpitis increased, and NF-kB, the downstream target of TLR2 and TLR4, also showed a marked elevation after macrophages’ stimulation by HMGB1. Conclusion: The evidence from the present study suggests that the enhanced TLR2 and TLR4 pathways and Th17 cell polarization may be due to HMGB1 stimulation in pulpitis. PMID:28293399

  12. Experimental Evolution of Mycobacterium tuberculosis in Human Macrophages Results in Low-Frequency Mutations Not Associated with Selective Advantage.

    Directory of Open Access Journals (Sweden)

    Valentina Guerrini

    Full Text Available Isolates of the human pathogen Mycobacterium tuberculosis recovered from clinical samples exhibit genetic heterogeneity. Such variation may result from the stressful environment encountered by the pathogen inside the macrophage, which is the host cell tubercle bacilli parasitize. To study the evolution of the M. tuberculosis genome during growth inside macrophages, we developed a model of intracellular culture in which bacteria were serially passaged in macrophage-like THP-1 cells for about 80 bacterial generations. Genome sequencing of single bacterial colonies isolated before and after the infection cycles revealed that M. tuberculosis developed mutations at a rate of about 5.7 × 10-9 / bp/ generation, consistent with mutation rates calculated during in vivo infection. Analysis of mutant growth in macrophages and in mice showed that the mutations identified after the cyclic infection conferred no advantage to the mutants relative to wild-type. Furthermore, activity testing of the recombinant protein harboring one of these mutations showed that the presence of the mutation did not affect the enzymatic activity. The serial infection protocol developed in this work to study M. tuberculosis genome microevolution can be applied to exposure to stressors to determine their effect on genome remodeling during intra-macrophage growth.

  13. Leishmania hijacking of the macrophage intracellular compartments.

    Science.gov (United States)

    Liévin-Le Moal, Vanessa; Loiseau, Philippe M

    2016-02-01

    Leishmania spp., transmitted to humans by the bite of the sandfly vector, are responsible for the three major forms of leishmaniasis, cutaneous, diffuse mucocutaneous and visceral. Leishmania spp. interact with membrane receptors of neutrophils and macrophages. In macrophages, the parasite is internalized within a parasitophorous vacuole and engages in a particular intracellular lifestyle in which the flagellated, motile Leishmania promastigote metacyclic form differentiates into non-motile, metacyclic amastigote form. This phenomenon is induced by Leishmania-triggered events leading to the fusion of the parasitophorous vacuole with vesicular members of the host cell endocytic pathway including recycling endosomes, late endosomes and the endoplasmic reticulum. Maturation of the parasitophorous vacuole leads to the intracellular proliferation of the Leishmania amastigote forms by acquisition of host cell nutrients while escaping host defense responses. © 2015 FEBS.

  14. Molecular Characterization of Macrophage-Biomaterial Interactions.

    Science.gov (United States)

    Moore, Laura Beth; Kyriakides, Themis R

    2015-01-01

    Implantation of biomaterials in vascularized tissues elicits the sequential engagement of molecular and cellular elements that constitute the foreign body response. Initial events include the non-specific adsorption of proteins to the biomaterial surface that render it adhesive for cells such as neutrophils and macrophages. The latter undergo unique activation and in some cases undergo cell-cell fusion to form foreign body giant cells that contribute to implant damage and fibrotic encapsulation. In this review, we discuss the molecular events that contribute to macrophage activation and fusion with a focus on the role of the inflammasome, signaling pathways such as JAK/STAT and NF-κB, and the putative involvement of micro RNAs in the regulation of these processes.

  15. Purinergic signaling to terminate TLR responses in macrophages

    Directory of Open Access Journals (Sweden)

    Kajal eHamidzadeh

    2016-03-01

    Full Text Available 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 down-regulation 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 down-regulation 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.

  16. Macrophage Phenotype and Function in Different Stages of Atherosclerosis

    Science.gov (United States)

    Tabas, Ira; Bornfeldt, Karin E.

    2016-01-01

    The remarkable plasticity and plethora of biological functions performed by macrophages have enticed scientists to study these cells in relation to atherosclerosis for more than 50 years, and major discoveries continue to be made today. It is now understood that macrophages play important roles in all stages of atherosclerosis, from initiation of lesions and lesion expansion, to necrosis leading to rupture and the clinical manifestations of atherosclerosis, to resolution and regression of atherosclerotic lesions. Lesional macrophages are derived primarily from blood monocytes, although recent research has shown that lesional macrophage-like cells can also be derived from smooth muscle cells. Lesional macrophages take on different phenotypes depending on their environment and which intracellular signaling pathways are activated. Rather than a few distinct populations of macrophages, the phenotype of the lesional macrophage is more complex and likely changes during the different phases of atherosclerosis and with the extent of lipid and cholesterol loading, activation by a plethora of receptors, and metabolic state of the cells. These different phenotypes allow the macrophage to engulf lipids, dead cells, and other substances perceived as danger signals; efflux cholesterol to HDL; proliferate and migrate; undergo apoptosis and death; and secrete a large number of inflammatory and pro-resolving molecules. This review article, part of the Compendium on Atherosclerosis, discusses recent advances in our understanding of lesional macrophage phenotype and function in different stages of atherosclerosis. With the increasing understanding of the roles of lesional macrophages, new research areas and treatment strategies are beginning to emerge. PMID:26892964

  17. Human Induced Pluripotent Stem Cell-Derived Macrophages for Unraveling Human Macrophage Biology.

    Science.gov (United States)

    Zhang, Hanrui; Reilly, Muredach P

    2017-11-01

    Despite a substantial appreciation for the critical role of macrophages in cardiometabolic diseases, understanding of human macrophage biology has been hampered by the lack of reliable and scalable models for cellular and genetic studies. Human induced pluripotent stem cell (iPSC)-derived macrophages (IPSDM), as an unlimited source of subject genotype-specific cells, will undoubtedly play an important role in advancing our understanding of the role of macrophages in human diseases. In this review, we summarize current literature in the differentiation and characterization of IPSDM at phenotypic, functional, and transcriptomic levels. We emphasize the progress in differentiating iPSC to tissue resident macrophages, and in understanding the ontogeny of in vitro differentiated IPSDM that resembles primitive hematopoiesis, rather than adult definitive hematopoiesis. We review the application of IPSDM in modeling both Mendelian genetic disorders and host-pathogen interactions. Finally, we highlighted the potential areas of research using IPSDM in functional validation of coronary artery disease loci in genome-wide association studies, functional genomic analyses, drug testing, and cell therapeutics in cardiovascular diseases. © 2017 American Heart Association, Inc.

  18. Lewis Lung Cancer Cells Promote SIGNR1(CD209b)-Mediated Macrophages Polarization Induced by IL-4 to Facilitate Immune Evasion.

    Science.gov (United States)

    Yan, Xiaolong; Li, Wenhai; Pan, Lei; Fu, Enqing; Xie, Yonghong; Chen, Min; Mu, Deguang

    2016-05-01

    Tumor-associated macrophages are a prominent component of lung cancer and contribute to tumor progression by facilitating the immune evasion of cancer cells. DC-SIGN (CD209) assists in the immune evasion of a broad spectrum of pathogens and neoplasms by inhibiting the maturation of DCs and subsequent cytokines production. However, the expression of DC-SIGN in macrophages and its role in mediating immune evasion in lung cancer and the underlying mechanism remain unclear. Our study aimed to identify the immunosuppressive role of SIGNR1 in murine macrophage differentiation and lung cancer progression. We found that SIGNR1-positive RAW264.7 macrophages were enriched in mixed cultures with Lewis lung cancer cells (LLC) (ratio of RAW 264.7 to LLC being 1:1) after stimulation with IL-4. Moreover, LLC-educated macrophages exhibited significantly higher levels of IL-10 but lower IL-12 in response to IL-4 treatment as determined by RT-PCR and ELISA. However, inhibition of SIGNR1 markedly hampered the production of IL-10, indicating that SIGNR1 was indispensable for IL-4+LLC induced macrophage polarization towards the M2 subtype. Furthermore, polarized M2 cells immersed in a tumor microenvironment promoted the migration of LLCs, as measured by transwell assays, but migration was suppressed after blockade of SIGNR1 using CD209b antibody. In addition, IL-4+LLC-educated macrophages reduced the proliferation of the activated T cells and reduced IFN-γ-mediated Th1 response in T cells, while SIGNR1 inhibition rescued Th1 cell functions. In conclusion, murine SIGNR1 expressed in LLC-educated macrophages appears to mediate IL-4-induced RAW264.7 macrophage polarization and thus facilitate lung cancer evasion. © 2015 Wiley Periodicals, Inc.

  19. Efficient, long term production of monocyte-derived macrophages from human pluripotent stem cells under partly-defined and fully-defined conditions.

    Directory of Open Access Journals (Sweden)

    Bonnie van Wilgenburg

    Full Text Available Human macrophages are specialised hosts for HIV-1, dengue virus, Leishmania and Mycobacterium tuberculosis. Yet macrophage research is hampered by lack of appropriate cell models for modelling infection by these human pathogens, because available myeloid cell lines are, by definition, not terminally differentiated like tissue macrophages. We describe here a method for deriving monocytes and macrophages from human Pluripotent Stem Cells which improves on previously published protocols in that it uses entirely defined, feeder- and serum-free culture conditions and produces very consistent, pure, high yields across both human Embryonic Stem Cell (hESC and multiple human induced Pluripotent Stem Cell (hiPSC lines over time periods of up to one year. Cumulatively, up to ∼3×10(7 monocytes can be harvested per 6-well plate. The monocytes produced are most closely similar to the major blood monocyte (CD14(+, CD16(low, CD163(+. Differentiation with M-CSF produces macrophages that are highly phagocytic, HIV-1-infectable, and upon activation produce a pro-inflammatory cytokine profile similar to blood monocyte-derived macrophages. Macrophages are notoriously hard to genetically manipulate, as they recognise foreign nucleic acids; the lentivector system described here overcomes this, as pluripotent stem cells can be relatively simply genetically manipulated for efficient transgene expression in the differentiated cells, surmounting issues of transgene silencing. Overall, the method we describe here is an efficient, effective, scalable system for the reproducible production and genetic modification of human macrophages, facilitating the interrogation of human macrophage biology.

  20. Identification of BCAP-{sub L} as a negative regulator of the TLR signaling-induced production of IL-6 and IL-10 in macrophages by tyrosine phosphoproteomics

    Energy Technology Data Exchange (ETDEWEB)

    Matsumura, Takayuki [Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku-ku, Tokyo 162-0041 (Japan); Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480 (Japan); Oyama, Masaaki; Kozuka-Hata, Hiroko [Medical Proteomics Laboratory, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Ishikawa, Kosuke; Inoue, Takafumi [Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku-ku, Tokyo 162-0041 (Japan); Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480 (Japan); Muta, Tatsushi [Laboratory of Cell Recognition and Response, Graduate School of Life Sciences, Tohoku University, Sendai 980-8578 (Japan); Semba, Kentaro, E-mail: ksemba@waseda.jp [Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku-ku, Tokyo 162-0041 (Japan); Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480 (Japan); Inoue, Jun-ichiro, E-mail: jun-i@ims.u-tokyo.ac.jp [Medical Proteomics Laboratory, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Division of Cellular and Molecular Biology, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan)

    2010-09-17

    Research highlights: {yields} Twenty five tyrosine-phosphorylated proteins in LPS-stimulated macrophages were determined. {yields} BCAP is a novel tyrosine-phosphorylated protein in LPS-stimulated macrophages. {yields} BCAP-{sub L} inhibits IL-6 and IL-10 production in LPS-stimulated macrophages. -- Abstract: Toll-like receptor (TLR) signaling in macrophages is essential for anti-pathogen responses such as cytokine production and antigen presentation. Although numerous reports suggest that protein tyrosine kinases (PTKs) are involved in cytokine induction in response to lipopolysaccharides (LPS; TLR4 ligand) in macrophages, the PTK-mediated signal transduction pathway has yet to be analyzed in detail. Here, we carried out a comprehensive and quantitative dynamic tyrosine phosphoproteomic analysis on the TLR4-mediated host defense system in RAW264.7 macrophages using stable isotope labeling by amino acids in cell culture (SILAC). We determined the temporal profiles of 25 proteins based on SILAC-encoded peptide(s). Of these, we focused on the tyrosine phosphorylation of B-cell adaptor for phosphatidylinositol 3-kinase (BCAP) because the function of BCAP remains unknown in TLR signaling in macrophages. Furthermore, Bcap has two distinct transcripts, a full-length (Bcap-{sub L}) and an alternatively initiated or spliced (Bcap-{sub S}) mRNA, and little is known about the differential functions of the BCAP-{sub L} and BCAP-{sub S} proteins. Our study showed, for the first time, that RNAi-mediated selective depletion of BCAP-{sub L} enhanced IL-6 and IL-10 production but not TNF-{alpha} production in TLR ligand-stimulated macrophages. We propose that BCAP-{sub L} (but not BCAP-{sub S}) is a negative regulator of the TLR-mediated host defense system in macrophages.

  1. Misbehaving macrophages in the pathogenesis of psoriasis

    OpenAIRE

    Clark, Rachael A.; Kupper, Thomas S.

    2006-01-01

    Psoriasis is a chronic inflammatory skin disease unique to humans. In this issue of the JCI, 2 studies of very different mouse models of psoriasis both report that macrophages play a key role in inducing psoriasis-like skin disease. Psoriasis is clearly a polygenic, inherited disease of uncontrolled cutaneous inflammation. The debate that currently rages in the field is whether psoriasis is a disease of autoreactive T cells or whether it reflects an intrinsic defect within the skin — or both....

  2. Molecular Characterization of Macrophage-Biomaterial Interactions

    OpenAIRE

    Moore, Laura Beth; Kyriakides, Themis R.

    2015-01-01

    Implantation of biomaterials in vascularized tissues elicits the sequential engagement of molecular and cellular elements that constitute the foreign body response. Initial events include the non-specific adsorption of proteins to the biomaterial surface that render it adhesive for cells such as neutrophils and macrophages. The latter undergo unique activation and in some cases undergo cell-cell fusion to form foreign body giant cells that contribute to implant damage and fibrotic encapsulati...

  3. Macrophage specific drug delivery in experimental leishmaniasis.

    Science.gov (United States)

    Basu, Mukul Kumar; Lala, Sanchaita

    2004-09-01

    Macrophage-specific delivery systems are the subject of much interest nowadays, because of the fact that macrophages act as host cells for many parasites and bacteria, which give rise to outbreak of so many deadly diseases(eg. leishmaniasis, tuberculosis etc.) in humans. To combat these deadly diseases initially macrophage specific liposomal delivery system were thought of and tested in vivo against experimental leishmaniasis in hamsters using a series of indigenous or synthetic antileishmanial compounds and the results were critically discussed. In vitro testing was also done against macrophages infected with Leishmania donovani, the causative agent for visceral leishmaniasis. The common problem of liposome therapy being their larger size, stability and storage, non-ionic surfactant vesicles, niosomes were prepared, for their different drug distribution and release characteristics compared to liposomes. When tested in vivo, the retention capacity of niosomes was found to be higher than that of liposomes due to the absence of lipid molecules and their smaller size. Thus the therapeutic efficacy of certain antileishmanial compounds was found to be better than that in the liposomal form. The niosomes, being cheaper, less toxic, biodegradable and non-immunogenic, were considered for sometime as suitable alternatives to liposomes as drug carriers. Besides the advent of other classical drugs carriers(e.g. neoglycoproteins), the biggest challenge came from polymeric delivery vehicles, specially the polymeric nanoparticles which were made of cost effective biodegradable polymers and different natural polymers. Because of very small size and highly stable nature, use of nanoparticles as effective drug carriers has been explored in experimental leishmaniasis using a series of antileishmanial compounds, both of indigenous and synthetic origin. The feasibility of application in vivo, when tested for biological as well as for other physicochemical parameters, the polymeric

  4. M2 polarization enhances silica nanoparticle uptake by macrophages

    Directory of Open Access Journals (Sweden)

    Jessica eHoppstädter

    2015-03-01

    Full Text Available While silica nanoparticles have enabled numerous industrial and medical applications, their toxicological safety requires further evaluation. Macrophages are the major cell population responsible for nanoparticle clearance in vivo. The prevailing macrophage phenotype largely depends on the local immune status of the host. Whereas M1-polarized macrophages are considered as pro-inflammatory macrophages involved in host defense, M2 macrophages exhibit anti-inflammatory and wound-healing properties, but also promote tumor growth.We employed different models of M1 and M2 polarization: GM-CSF/LPS/IFN-gamma was used to generate primary human M1 cells and M-CSF/IL-10 to differentiate M2 monocyte-derived macrophages. PMA-differentiated THP-1 cells were polarized towards an M1 type by LPS/IFN-gamma and towards M2 by IL-10. Uptake of fluorescent silica nanoparticles (Ø 26 and 41 nm and microparticles (Ø 1.75 µm was quantified. At the concentration used (50 µg/ml, silica nanoparticles did not influence cell viability as assessed by MTT assay. Nanoparticle uptake was enhanced in M2-polarized primary human monocyte-derived macrophages compared with M1 cells, as shown by flow cytometric and microscopic approaches. In contrast, the uptake of microparticles did not differ between M1 and M2 phenotypes. M2 polarization was also associated with increased nanoparticle uptake in the macrophage-like THP-1 cell line. In accordance, in vivo polarized M2-like primary human tumor-associated macrophages (TAM obtained from lung tumors took up more nanoparticles than M1-like alveolar macrophages isolated from the surrounding lung tissue.In summary, our data indicate that the M2 polarization of macrophages promotes nanoparticle internalization. Therefore, the phenotypical differences between macrophage subsets should be taken into consideration in future investigations on nanosafety, but might also open up therapeutic perspectives allowing to specifically target M2

  5. Legumain is activated in macrophages during pancreatitis

    Science.gov (United States)

    Wartmann, Thomas; Fleming, Alicia K.; Gocheva, Vasilena; van der Linden, Wouter A.; Withana, Nimali P.; Verdoes, Martijn; Aurelio, Luigi; Edgington-Mitchell, Daniel; Lieu, TinaMarie; Parker, Belinda S.; Graham, Bim; Reinheckel, Thomas; Furness, John B.; Joyce, Johanna A.; Storz, Peter; Halangk, Walter; Bogyo, Matthew; Bunnett, Nigel W.

    2016-01-01

    Pancreatitis is an inflammatory disease of the pancreas characterized by dysregulated activity of digestive enzymes, necrosis, immune infiltration, and pain. Repeated incidence of pancreatitis is an important risk factor for pancreatic cancer. Legumain, a lysosomal cysteine protease, has been linked to inflammatory diseases such as atherosclerosis, stroke, and cancer. Until now, legumain activation has not been studied during pancreatitis. We used a fluorescently quenched activity-based probe to assess legumain activation during caerulein-induced pancreatitis in mice. We detected activated legumain by ex vivo imaging, confocal microscopy, and gel electrophoresis. Compared with healthy controls, legumain activity in the pancreas of caerulein-treated mice was increased in a time-dependent manner. Legumain was localized to CD68+ macrophages and was not active in pancreatic acinar cells. Using a small-molecule inhibitor of legumain, we found that this protease is not essential for the initiation of pancreatitis. However, it may serve as a biomarker of disease, since patients with chronic pancreatitis show strongly increased legumain expression in macrophages. Moreover, the occurrence of legumain-expressing macrophages in regions of acinar-to-ductal metaplasia suggests that this protease may influence reprogramming events that lead to inflammation-induced pancreatic cancer. PMID:27514475

  6. Pegylated silica nanoparticles: cytotoxicity and macrophage uptake

    Science.gov (United States)

    Glorani, Giulia; Marin, Riccardo; Canton, Patrizia; Pinto, Marcella; Conti, Giamaica; Fracasso, Giulio; Riello, Pietro

    2017-08-01

    Here, we present a thorough study of pegylated silica nanoparticle (SNP) interaction with different biological environments. The SNPs have a mean diameter of about 40 nm and are coated with polyethylene glycol (PEG) of different molecular weights. The physicochemical characterization of SNPs allowed the confirmation of the binding of PEG chains to the silica surface, the reproducibility of the synthesis and the narrow size-dispersion. In view of clarifying the SNP interaction with biological environments, we first assessed the SNP reactivity after the incubation with two cell lines (macrophages RAW 264.7 and primary human fibroblasts), observing a reduced toxicity of pegylated SNPs compared to the bare ones. Then, we investigated the effect of the protein adsorption on the SNP surface using the model serum protein, bovine serum albumin (BSA). We found that the protein adsorption takes place more heavily on poorly pegylated SNPs, promoting the uptake of the latter by macrophages and leading to an increased mortality of these cells. To better understand this mechanism by means of flow cytometry, the dye Ru(bpy)3Cl2 was incorporated in the SNPs. The overall results highlight the SNP potentialities as a drug delivery system, thanks to the low interactions with the macrophages.

  7. Burkholderia pseudomallei transcriptional adaptation in macrophages

    Directory of Open Access Journals (Sweden)

    Chieng Sylvia

    2012-07-01

    Full Text Available Abstract Background Burkholderia pseudomallei is a facultative intracellular pathogen of phagocytic and non-phagocytic cells. How the bacterium interacts with host macrophage cells is still not well understood and is critical to appreciate the strategies used by this bacterium to survive and how intracellular survival leads to disease manifestation. Results Here we report the expression profile of intracellular B. pseudomallei following infection of human macrophage-like U937 cells. During intracellular growth over the 6 h infection period, approximately 22 % of the B. pseudomallei genome showed significant transcriptional adaptation. B. pseudomallei adapted rapidly to the intracellular environment by down-regulating numerous genes involved in metabolism, cell envelope, motility, replication, amino acid and ion transport system and regulatory function pathways. Reduced expression in catabolic and housekeeping genes suggested lower energy requirement and growth arrest during macrophage infection, while expression of genes encoding anaerobic metabolism functions were up regulated. However, whilst the type VI secretion system was up regulated, expression of many known virulence factors was not significantly modulated over the 6hours of infection. Conclusions The transcriptome profile described here provides the first comprehensive view of how B. pseudomallei survives within host cells and will help identify potential virulence factors and proteins that are important for the survival and growth of B. pseudomallei within human cells.

  8. Stochastic differentiation into an osteoclast lineage from cloned macrophage-like cells

    Energy Technology Data Exchange (ETDEWEB)

    Hayashi, Shin-Ichi, E-mail: shayashi@med.tottori-u.ac.jp [Division of Immunology, Department of Molecular and Cellular Biology, School of Life Science, Faculty of Medicine, Tottori University, 86 Nishi-Cho, Yonago, Tottori 683-8503 (Japan); Murata, Akihiko; Okuyama, Kazuki; Shimoda, Yuhki; Hikosaka, Mari [Division of Immunology, Department of Molecular and Cellular Biology, School of Life Science, Faculty of Medicine, Tottori University, 86 Nishi-Cho, Yonago, Tottori 683-8503 (Japan); Yasuda, Hisataka [Planning and Development, Bioindustry Division, Oriental Yeast Co., Ltd, Itabashi-Ku, Tokyo 174-8505 (Japan); Yoshino, Miya [Division of Immunology, Department of Molecular and Cellular Biology, School of Life Science, Faculty of Medicine, Tottori University, 86 Nishi-Cho, Yonago, Tottori 683-8503 (Japan)

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer The frequency of C7 differentiation into osteoclast was low and constant. Black-Right-Pointing-Pointer Only extended C7 cell cultures exponentially increased osteoclast+ cultures. Black-Right-Pointing-Pointer C7 cell differentiation into committed osteoclast precursors is on 'autopilot'. Black-Right-Pointing-Pointer The system may maintain the stem cell self-renewal and differentiation. -- Abstract: Differentiation into osteoclasts is induced by a macrophage colony-stimulating factor and receptor activator of nuclear-factor {kappa}B ligand. The macrophage-like cell line, C7 has the potential to differentiate into osteoclasts when it is cultured with both factors for 6 days. Although C7 is an established cell line, the frequency of differentiation into this lineage was less than 10%, and the ratio was maintained at a constant level, even after repeated cloning. In this study, to increase the differentiation of C7 cells to osteoclasts, C7 derivative treatments with several activators and/or inhibitors were performed for 3 days prior to setting osteoclast induction analysis; however, a reagent to significantly up-regulate the frequency of differentiation was not found. Only extended cultures for osteoclastogenesis exponentially increased the frequency of osteoclast precursors. It is likely that C7 cell differentiation into committed osteoclast precursors is on 'autopilot' rather than requiring specific signals to drive this process.

  9. Stochastic differentiation into an osteoclast lineage from cloned macrophage-like cells

    International Nuclear Information System (INIS)

    Hayashi, Shin-Ichi; Murata, Akihiko; Okuyama, Kazuki; Shimoda, Yuhki; Hikosaka, Mari; Yasuda, Hisataka; Yoshino, Miya

    2012-01-01

    Highlights: ► The frequency of C7 differentiation into osteoclast was low and constant. ► Only extended C7 cell cultures exponentially increased osteoclast+ cultures. ► C7 cell differentiation into committed osteoclast precursors is on ‘autopilot’. ► The system may maintain the stem cell self-renewal and differentiation. -- Abstract: Differentiation into osteoclasts is induced by a macrophage colony-stimulating factor and receptor activator of nuclear-factor κB ligand. The macrophage-like cell line, C7 has the potential to differentiate into osteoclasts when it is cultured with both factors for 6 days. Although C7 is an established cell line, the frequency of differentiation into this lineage was less than 10%, and the ratio was maintained at a constant level, even after repeated cloning. In this study, to increase the differentiation of C7 cells to osteoclasts, C7 derivative treatments with several activators and/or inhibitors were performed for 3 days prior to setting osteoclast induction analysis; however, a reagent to significantly up-regulate the frequency of differentiation was not found. Only extended cultures for osteoclastogenesis exponentially increased the frequency of osteoclast precursors. It is likely that C7 cell differentiation into committed osteoclast precursors is on ‘autopilot’ rather than requiring specific signals to drive this process.

  10. Macrophages are required for dendritic cell uptake of respiratory syncytial virus from an infected epithelium.

    Science.gov (United States)

    Ugonna, Kelechi; Bingle, Colin D; Plant, Karen; Wilson, Kirsty; Everard, Mark L

    2014-01-01

    We have previously shown that the respiratory syncytial virus [RSV] can productively infect monocyte derived dendritic cells [MoDC] and remain dormant within the same cells for prolonged periods. It is therefore possible that infected dendritic cells act as a reservoir within the airways of individuals between annual epidemics. In the present study we explored the possibility that sub-epithelial DCs can be infected with RSV from differentiated bronchial epithelium and that in turn RSV from DCs can infect the epithelium. A dual co-culture model was established in which a differentiated primary airway epithelium on an Air Liquid Interface (ALI) was cultured on a transwell insert and MoDCs were subsequently added to the basolateral membrane of the insert. Further experiments were undertaken using a triple co-culture model in which in which macrophages were added to the apical surface of the differentiated epithelium. A modified RSV [rr-RSV] expressing a red fluorescent protein marker of replication was used to infect either the MoDCs or the differentiated epithelium and infection of the reciprocal cell type was assessed using confocal microscopy. Our data shows that primary epithelium became infected when rr-RSV infected MoDCs were introduced onto the basal surface of the transwell insert. MoDCs located beneath the epithelium did not become infected with virus from infected epithelial cells in the dual co-culture model. However when macrophages were present on the apical surface of the primary epithelium infection of the basal MoDCs occurred. Our data suggests that RSV infected dendritic cells readily transmit infection to epithelial cells even when they are located beneath the basal layer. However macrophages appear to be necessary for the transmission of infection from epithelial cells to basal dendritic cells.

  11. Origins and Hallmarks of Macrophages: Development, Homeostasis, and Disease

    Science.gov (United States)

    Wynn, Thomas A.; Chawla, Ajay; Pollard, Jeffrey W.

    2013-01-01

    Preface Macrophages the most plastic cells of the hematopoietic system are found in all tissues and exhibit great functional diversity. They have roles in development, homeostasis, tissue repair, and immunity. While anatomically distinct, resident tissue macrophages exhibit different transcriptional profiles, and functional capabilities, they are all required for the maintenance of homeostasis. However, these reparative and homeostatic functions can be subverted by chronic insults, resulting in a causal association of macrophages with disease states. In this review, we discuss how macrophages regulate normal physiology and development and provide several examples of their pathophysiologic roles in disease. We define the “hallmarks” of macrophages performing particular functions, taking into account novel insights into the diversity of their lineages, identity, and regulation. This diversity is essential to understand because macrophages have emerged as important therapeutic targets in many important human diseases. PMID:23619691

  12. Regulation of macrophage development and function in peripheral tissues

    Science.gov (United States)

    Lavin, Yonit; Mortha, Arthur; Rahman, Adeeb; Merad, Miriam

    2015-01-01

    Macrophages are immune cells of haematopoietic origin that provide crucial innate immune defence and have tissue-specific functions in the regulation and maintenance of organ homeostasis. Recent studies of macrophage ontogeny, as well as transcriptional and epigenetic identity, have started to reveal the decisive role of the tissue stroma in the regulation of macrophage function. These findings suggest that most macrophages seed the tissues during embryonic development and functionally specialize in response to cytokines and metabolites that are released by the stroma and drive the expression of unique transcription factors. In this Review, we discuss how recent insights into macrophage ontogeny and macrophage–stroma interactions contribute to our understanding of the crosstalk that shapes macrophage function and the maintenance of organ integrity. PMID:26603899

  13. Tumor-Associated Macrophages in Oncolytic Virotherapy: Friend or Foe?

    Directory of Open Access Journals (Sweden)

    Nicholas L. Denton

    2016-07-01

    Full Text Available Cancer therapy remains a challenge due to toxicity limitations of chemotherapy and radiation therapy. Oncolytic viruses that selectively replicate and destroy cancer cells are of increasing interest. In addition to direct cell lysis, these vectors stimulate an anti-tumor immune response. A key regulator of tumor immunity is the tumor-associated macrophage population. Macrophages can either support oncolytic virus therapy through pro-inflammatory stimulation of the anti-tumor response at the cost of hindering direct oncolysis or through immunosuppressive protection of virus replication at the cost of hindering the anti-tumor immune response. Despite similarities in macrophage interaction between adult and pediatric tumors and the abundance of research supporting macrophage modulation in adult tumors, there are few studies investigating macrophage modulation in pediatric cancers or modulation of immunotherapy. We review the current state of knowledge regarding macrophages in cancers and their influence on oncolytic virotherapy.

  14. The differential role of human macrophage in triggering secondary bystander effects after either gamma-ray or carbon beam irradiation.

    Science.gov (United States)

    Dong, Chen; He, Mingyuan; Tu, Wenzhi; Konishi, Teruaki; Liu, Weili; Xie, Yuexia; Dang, Bingrong; Li, Wenjian; Uchihori, Yukio; Hei, Tom K; Shao, Chunlin

    2015-07-10

    The abscopal effect could be an underlying factor in evaluating prognosis of radiotherapy. This study established an in vitro system to examine whether tumor-generated bystander signals could be transmitted by macrophages to further trigger secondary cellular responses after different irradiations, where human lung cancer NCI-H446 cells were irradiated with either γ-rays or carbon ions and co-cultured with human macrophage U937 cells, then these U937 cells were used as a bystander signal transmitter and co-cultured with human bronchial epithelial cells BEAS-2B. Results showed that U937 cells were only activated by γ-irradiated NCI-H446 cells so that the secondary injuries in BEAS-2B cells under carbon ion irradiation were weaker than γ-rays. Both TNF-α and IL-1α were involved in the γ-irradiation induced secondary bystander effect but only TNF-α contributed to the carbon ion induced response. Further assay disclosed that IL-1α but not TNF-α was largely responsible for the activation of macrophages and the formation of micronucleus in BEAS-2B cells. These data suggest that macrophages could transfer secondary bystander signals and play a key role in the secondary bystander effect of photon irradiation, while carbon ion irradiation has conspicuous advantage due to its reduced secondary injury. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  15. Polyethylene and methyl methacrylate particle-stimulated inflammatory tissue and macrophages up-regulate bone resorption in a murine neonatal calvaria in vitro organ system.

    Science.gov (United States)

    Ren, Weiping; Wu, Bin; Mayton, Lois; Wooley, Paul H

    2002-09-01

    There is considerable evidence that orthopaedic wear debris plays a crucial role in the pathology of aseptic loosening of joint prostheses. This study examined the effect of inflammatory membranes stimulated with methyl methacrylate and polyethylene on bone resorption, using the murine air pouch model. The capacity of RAW 264.7 mouse macrophages exposed to polymer particles to produce factors affecting bone metabolism was also studied. Neonatal calvaria bones were co-cultured with either pouch membranes or conditioned media from activated macrophages. Bone resorption was measured by the release of calcium from cultured bones, and the activity of tartrate-resistant acid phosphatase in both bone sections and culture medium was also assayed. Results showed that inflammatory pouch membrane activated by methyl methacrylate and polyethylene enhanced osteoclastic bone resorption. Conditioned media from particles stimulated mouse macrophages also stimulated bone resorption, although this effect was weaker than resorption induced by inflammatory pouch membranes. The addition of the particles directly into the medium of cultured calvaria bones had little effect on bone resorption. Our observations indicate that both inflammatory tissue and macrophages provoked by particles can stimulate bone resorption in cultured mouse neonatal calvaria bones. This simple in vitro bone resorption system allows us to investigate the fundamental cellular and molecular mechanism of wear debris induced bone resorption and to screen potential therapeutic approaches for aseptic loosening.

  16. Role of Macrophage-Induced Inflammation in Mesothelioma

    Science.gov (United States)

    2011-07-01

    macrophages). Normal pleura becomes available intermittently , serving to slow the completion of this task. All is set in place for us to complete the...GFP) regulated by a Csf1r-promoter (Sasmono et al. 2003) show that macrophages travel up and down these fibers at a fast rate and also “jump” between...2010). Macrophages have also recently been shown to be important in adipogenesis at least during obesity , through their secretion of adipocyte growth

  17. Cord blood-derived macrophage-lineage cells rapidly stimulate osteoblastic maturation in mesenchymal stem cells in a glycoprotein-130 dependent manner.

    Directory of Open Access Journals (Sweden)

    Tania J Fernandes

    Full Text Available In bone, depletion of osteoclasts reduces bone formation in vivo, as does osteal macrophage depletion. How osteoclasts and macrophages promote the action of bone forming osteoblasts is, however, unclear. Since recruitment and differentiation of multi-potential stromal cells/mesenchymal stem cells (MSC generates new active osteoblasts, we investigated whether human osteoclasts and macrophages (generated from cord blood-derived hematopoietic progenitors induce osteoblastic maturation in adipose tissue-derived MSC. When treated with an osteogenic stimulus (ascorbate, dexamethasone and β-glycerophosphate these MSC form matrix-mineralising, alkaline phosphatase-expressing osteoblastic cells. Cord blood-derived progenitors were treated with macrophage colony stimulating factor (M-CSF to form immature proliferating macrophages, or with M-CSF plus receptor activator of NFκB ligand (RANKL to form osteoclasts; culture medium was conditioned for 3 days by these cells to study their production of osteoblastic factors. Both osteoclast- and macrophage-conditioned medium (CM greatly enhanced MSC osteoblastic differentiation in both the presence and absence of osteogenic medium, evident by increased alkaline phosphatase levels within 4 days and increased mineralisation within 14 days. These CM effects were completely ablated by antibodies blocking gp130 or oncostatin M (OSM, and OSM was detectable in both CM. Recombinant OSM very potently stimulated osteoblastic maturation of these MSC and enhanced bone morphogenetic protein-2 (BMP-2 actions on MSC. To determine the influence of macrophage activation on this OSM-dependent activity, CM was collected from macrophage populations treated with M-CSF plus IL-4 (to induce alternative activation or with GM-CSF, IFNγ and LPS to cause classical activation. CM from IL-4 treated macrophages stimulated osteoblastic maturation in MSC, while CM from classically-activated macrophages did not. Thus, macrophage-lineage cells

  18. Tumor associated macrophages protect colon cancer cells from TRAIL-induced apoptosis through IL-1beta-dependent stabilization of Snail in tumor cells.

    Directory of Open Access Journals (Sweden)

    Pawan Kaler

    2010-07-01

    Full Text Available We recently reported that colon tumor cells stimulate macrophages to release IL-1beta, which in turn inactivates GSK3beta and enhances Wnt signaling in colon cancer cells, generating a self-amplifying loop that promotes the growth of tumor cells.Here we describe that macrophages protect HCT116 and Hke-3 colon cancer cells from TRAIL-induced apoptosis. Inactivation of IL-1beta by neutralizing IL-1beta antibody, or silencing of IL-1beta in macrophages inhibited their ability to counter TRAIL-induced apoptosis. Accordingly, IL-1beta was sufficient to inhibit TRAIL-induced apoptosis. TRAIL-induced collapse of the mitochondrial membrane potential (Delta psi and activation of caspases were prevented by macrophages or by recombinant IL-1beta. Pharmacological inhibition of IL-1beta release from macrophages by vitamin D(3, a potent chemopreventive agent for colorectal cancer, restored the ability of TRAIL to induce apoptosis of tumor cells cultured with macrophages. Macrophages and IL-1beta failed to inhibit TRAIL-induced apoptosis in HCT116 cells expressing dnIkappaB, dnAKT or dnTCF4, confirming that they oppose TRAIL-induced cell death through induction of Wnt signaling in tumor cells. We showed that macrophages and IL-1beta stabilized Snail in tumor cells in an NF-kappaB/Wnt dependent manner and that Snail deficient tumor cells were not protected from TRAIL-induced apoptosis by macrophages or by IL-1beta, demonstrating a crucial role of Snail in the resistance of tumor cells to TRAIL.We have identified a positive feedback loop between tumor cells and macrophages that propagates the growth and promotes the survival of colon cancer cells: tumor cells stimulate macrophages to secrete IL-1beta, which in turn, promotes Wnt signaling and stabilizes Snail in tumor cells, conferring resistance to TRAIL. Vitamin D(3 halts this amplifying loop by interfering with the release of IL-1beta from macrophages. Accordingly, vitamin D(3 sensitizes tumor cells to TRAIL

  19. Macrophages: contributors to allograft dysfunction, repair, or innocent bystanders?

    Science.gov (United States)

    Mannon, Roslyn B

    2012-02-01

    Macrophages are members of the innate immune response. However, their role in the adaptive immune response is not known. The purpose of this review is to highlight our current understanding of macrophage structure and function and how they may participate in allograft injury. Studies in acute kidney injury models identify macrophages as key mediators of inflammatory injury, while more recent studies indicate that they may play a reparative role, depending on phenotype - M1 or M2 type macrophages. Mregs, generated in vitro, appear to have immune suppressive abilities and a unique phenotype. In solid-organ transplant, the emphasis of studies has been on acute or chronic injury. These data are derived from animal models using depletion of macrophages or antagonizing their activation and inflammatory responses. The relative contribution of macrophage phenotype in transplantation has not been explored. These studies suggest that macrophages play an injurious role in acute cellular allograft rejection, as well as in chronic injury. Infiltration of an allograft with macrophages is also associated with worse graft function and poor prognosis. Further studies are needed to understand the mechanisms of macrophage-mediated injury, explore their potential reparative role, and determine if they or their functional products are biomarkers of poor graft outcomes.

  20. Functional modifications of macrophage activity after sublethal irradiation

    International Nuclear Information System (INIS)

    Swartz, R.P.

    1982-01-01

    The modifications of macrophage activity following sublethal irradiation, both in vivo and in vitro, were studied using spreading and C3b-receptor-mediated ingestion assays. Nonelicited peritoneal washout cells were examined for changes in activity and selected population characteristics. The cells from irradiated mice were from a resident peritoneal population and not immigrating cells. The macrophage population showed enhanced activity early with a refractory period (24-48) when the macrophages were unresponsive to stimulation by irradiated lymphocytes. The enhanced activity was inversely dose dependent on macrophage. The lymphocytes showed a regulatory function(s) on the time post irradiation at which they were examined. Early lymphocytes exhibited the ability to enhance the activity of normal macrophages while lymphocytes removed 24 hours post irradiation could suppress the activity of already activated macrophages. The effect(s) of the various lymphocyte populations were reproduced with cell-free supernatants which was indicative of the production of lymphokines. Separation on nylon wool columns indicated that the activity resided primarily in the T-cell population of lymphocytes. In vitro irradiation indicated that stimulation of the lymphocytes is macrophage dependent. Additional work indicated that sublethally irradiated macrophages did not inhibit replication of the coccidian protozoon Toxoplasma gondii although they did show increased phagocytosis. Examination of the serum from whole body irradiated mice showed the presence of a postirradiation substance which enhanced the phagocytosis of normal macrophages. It was not present in the serum of normal mice and was not endotoxin

  1. Macrophage migration inhibitory factor is associated with aneurysmal expansion

    DEFF Research Database (Denmark)

    Pan, Jie-Hong; Lindholt, Jes Sanddal; Sukhova, Galina K

    2003-01-01

    Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine released mainly from macrophages and activated lymphocytes. Both atherosclerosis and abdominal aortic aneurysm (AAA) are inflammatory diseases tightly linked to the function of these cells. The correlation and contribution o...... of MIF to these human diseases remain unknown, although a recent rabbit study showed expression of this cytokine in atherosclerotic lesions.......Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine released mainly from macrophages and activated lymphocytes. Both atherosclerosis and abdominal aortic aneurysm (AAA) are inflammatory diseases tightly linked to the function of these cells. The correlation and contribution...

  2. Delayed growth of EL4 lymphoma in SR-A-deficient mice is due to upregulation of nitric oxide and interferon-gamma production by tumor-associated macrophages.

    Science.gov (United States)

    Komohara, Yoshihiro; Takemura, Kenichi; Lei, Xiao Feng; Sakashita, Naomi; Harada, Mamoru; Suzuki, Hiroshi; Kodama, Tatsuhiko; Takeya, Motohiro

    2009-11-01

    Class A scavenger receptors (SR-A, CD204) are highly expressed in tumor-associated macrophages (TAM). To investigate the function of SR-A in TAM, wild-type and SR-A-deficient (SR-A(-/-)) mice were injected with EL4 cells. Although these groups of mice did not differ in the numbers of infiltrating macrophages and lymphocytes and in neovascularization, SR-A(-/-) mice had delayed growth of EL4 tumors. Expression of inducible nitric oxide (NO) synthase and interferon (IFN)-gamma mRNA increased significantly in tumor tissues from SR-A(-/-) mice. Engulfment of necrotic EL4 cells induced upregulation of NO and IFN-gamma production by cultured macrophages, and production of NO and IFN-gamma increased in SR-A(-/-) macrophages in vitro. IFN-beta production by cultured macrophages was also elevated in SR-A(-/-) macrophages in vitro. These results suggested that the antitumor activity of macrophages increased in SR-A(-/-) mice because of upregulation of NO and IFN-gamma production. These data indicate an important role of SR-A in regulating TAM function by inhibiting toll-like receptor (TLR)4-IFN-beta signaling.

  3. Moderate restriction of macrophage-tropic human immunodeficiency virus type 1 by SAMHD1 in monocyte-derived macrophages.

    Science.gov (United States)

    Taya, Kahoru; Nakayama, Emi E; Shioda, Tatsuo

    2014-01-01

    Macrophage-tropic human immunodeficiency virus type 1 (HIV-1) strains are able to grow to high titers in human monocyte-derived macrophages. However, it was recently reported that cellular protein SAMHD1 restricts HIV-1 replication in human cells of the myeloid lineage, including monocyte-derived macrophages. Here we show that degradation of SAMHD1 in monocyte-derived macrophages was associated with moderately enhanced growth of the macrophage-tropic HIV-1 strain. SAMHD1 degradation was induced by treating target macrophages with vesicular stomatitis virus glycoprotein-pseudotyped human immunodeficiency virus type 2 (HIV-2) particles containing viral protein X. For undifferentiated monocytes, HIV-2 particle treatment allowed undifferentiated monocytes to be fully permissive for productive infection by the macrophage-tropic HIV-1 strain. In contrast, untreated monocytes were totally resistant to HIV-1 replication. These results indicated that SAMHD1 moderately restricts even a macrophage-tropic HIV-1 strain in monocyte-derived macrophages, whereas the protein potently restricts HIV-1 replication in undifferentiated monocytes.

  4. Macrophage heterogeneity and cholesterol homeostasis: classically-activated macrophages are associated with reduced cholesterol accumulation following treatment with oxidized LDL.

    Science.gov (United States)

    Chu, Eugene M; Tai, Daven C; Beer, Jennifer L; Hill, John S

    2013-02-01

    Macrophages are centrally involved during atherosclerosis development and are the predominant cell type that accumulates cholesterol in the plaque. Macrophages however, are heterogeneous in nature reflecting a variety of microenvironments and different phenotypes may be more prone to contribute towards atherosclerosis progression. Using primary human monocyte-derived macrophages, we sought to evaluate one aspect of atherogenic potential of different macrophage phenotypes by determining their propensity to associate with and accumulate oxidized low density lipoprotein (oxLDL). Classically-activated macrophages treated simultaneously with interferon γ (IFNγ) and tumor necrosis factor α (TNFα) associated with less oxLDL and accumulated less cholesterol compared to untreated controls. The combined treatment of IFNγ and TNFα reduced the mRNA expression of CD36 and the expression of both cell surface CD36 and macrophage scavenger receptor 1 (MSR1) protein. Under oxLDL loaded conditions, IFNγ and TNFα did not reduce macrophage protein expression of the transcription factor peroxisome proliferator-actived receptor γ (PPARγ) which is known to positively regulate CD36 expression. However, macrophages treated with IFNγ attenuated the ability of the PPARγ-specific agonist rosiglitazone from upregulating cell surface CD36 protein expression. Our results demonstrate that the observed reduction of cholesterol accumulation in macrophages treated with IFNγ and TNFα following oxLDL treatment was due at least in part to reduced cell surface CD36 and MSR1 protein expression. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. DMPD: Signal integration between IFNgamma and TLR signalling pathways in macrophages. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 16920490 Signal integration between IFNgamma and TLR signalling pathways in macroph...tml) (.csml) Show Signal integration between IFNgamma and TLR signalling pathways in macrophages. PubmedID 16920490 Title Signal inte...gration between IFNgamma and TLR signalling pathways in

  6. Modulation of human macrophage activity by Ascaris antigens is dependent on macrophage polarization state

    DEFF Research Database (Denmark)

    Almeida, Sara; Nejsum, Peter; Williams, Andrew R.

    2018-01-01

    Parasitic worms (helminths) are known to actively modulate host immune responses and inflammation. The aim of this study was to investigate if adult body fluid (ABF) from the helminth Ascaris suum has immunomodulatory effects on different subtypes of human monocyte-derived macrophages (Mɸ) in vitro...

  7. Nicotinamide: a vitamin able to shift macrophage differentiation toward macrophages with restricted inflammatory features.

    Science.gov (United States)

    Weiss, Ronald; Schilling, Erik; Grahnert, Anja; Kölling, Valeen; Dorow, Juliane; Ceglarek, Uta; Sack, Ulrich; Hauschildt, Sunna

    2015-11-01

    The differentiation of human monocytes into macrophages is influenced by environmental signals. Here we asked in how far nicotinamide (NAM), a vitamin B3 derivative known to play a major role in nicotinamide adenine dinucleotide (NAD)-mediated signaling events, is able to modulate monocyte differentiation into macrophages developed in the presence of granulocyte macrophage colony-stimulating factor (GM-MØ) or macrophage colony-stimulating factor (M-MØ). We found that GM-MØ undergo biochemical, morphological and functional modifications in response to NAM, whereas M-MØ were hardly affected. GM-MØ exposed to NAM acquired an M-MØ-like structure while the LPS-induced production of pro-inflammatory cytokines and COX-derived eicosanoids were down-regulated. In contrast, NAM had no effect on the production of IL-10 or the cytochrome P450-derived eicosanoids. Administration of NAM enhanced intracellular NAD concentrations; however, it did not prevent the LPS-mediated drain on NAD pools. In search of intracellular molecular targets of NAM known to be involved in LPS-induced cytokine and eicosanoid synthesis, we found NF-κB activity to be diminished. In conclusion, our data show that vitamin B3, when present during the differentiation of monocytes into GM-MØ, interferes with biochemical pathways resulting in strongly reduced pro-inflammatory features. © The Author(s) 2015.

  8. Immunomodulatory Molecule IRAK-M Balances Macrophage Polarization and Determines Macrophage Responses during Renal Fibrosis.

    Science.gov (United States)

    Steiger, Stefanie; Kumar, Santhosh V; Honarpisheh, Mohsen; Lorenz, Georg; Günthner, Roman; Romoli, Simone; Gröbmayr, Regina; Susanti, Heni-Eka; Potempa, Jan; Koziel, Joanna; Lech, Maciej

    2017-08-15

    Activation of various innate immune receptors results in IL-1 receptor-associated kinase (IRAK)-1/IRAK-4-mediated signaling and secretion of proinflammatory cytokines such as IL-12, IL-6, or TNF-α, all of which are implicated in tissue injury and elevated during tissue remodeling processes. IRAK-M, also known as IRAK-3, is an inhibitor of proinflammatory cytokine and chemokine expression in intrarenal macrophages. Innate immune activation contributes to both acute kidney injury and tissue remodeling that is associated with chronic kidney disease (CKD). Our study assessed the contribution of macrophages in CKD and the role of IRAK-M in modulating disease progression. To evaluate the effect of IRAK-M in chronic renal injury in vivo, a mouse model of unilateral ureteral obstruction (UUO) was employed. The expression of IRAK-M increased within 2 d after UUO in obstructed compared with unobstructed kidneys. Mice deficient in IRAK-M were protected from fibrosis and displayed a diminished number of alternatively activated macrophages. Compared to wild-type mice, IRAK-M-deficient mice showed reduced tubular injury, leukocyte infiltration, and inflammation following renal injury as determined by light microscopy, immunohistochemistry, and intrarenal mRNA expression of proinflammatory and profibrotic mediators. Taken together, these results strongly support a role for IRAK-M in renal injury and identify IRAK-M as a possible modulator in driving an alternatively activated profibrotic macrophage phenotype in UUO-induced CKD. Copyright © 2017 by The American Association of Immunologists, Inc.

  9. Polyglucose nanoparticles with renal elimination and macrophage avidity facilitate PET imaging in ischaemic heart disease

    NARCIS (Netherlands)

    Keliher, Edmund J.; Ye, Yu-Xiang; Wojtkiewicz, Gregory R.; Aguirre, Aaron D.; Tricot, Benoit; Senders, Max L.; Groenen, Hannah; Fay, Francois; Perez-Medina, Carlos; Calcagno, Claudia; Carlucci, Giuseppe; Reiner, Thomas; Sun, Yuan; Courties, Gabriel; Iwamoto, Yoshiko; Kim, Hye-Yeong; Wang, Cuihua; Chen, John W.; Swirski, Filip K.; Wey, Hsiao-Ying; Hooker, Jacob; Fayad, Zahi A.; Mulder, Willem J. M.; Weissleder, Ralph; Nahrendorf, Matthias

    2017-01-01

    Tissue macrophage numbers vary during health versus disease. Abundant inflammatory macrophages destruct tissues, leading to atherosclerosis, myocardial infarction and heart failure. Emerging therapeutic options create interest in monitoring macrophages in patients. Here we describe positron emission

  10. Paracrine Interactions between Adipocytes and Tumor Cells Recruit and Modify Macrophages to the Mammary Tumor Microenvironment: The Role of Obesity and Inflammation in Breast Adipose Tissue

    Energy Technology Data Exchange (ETDEWEB)

    Santander, Ana M.; Lopez-Ocejo, Omar; Casas, Olivia; Agostini, Thais; Sanchez, Lidia; Lamas-Basulto, Eduardo; Carrio, Roberto [Department of Microbiology and Immunology, University of Miami Miller School of Medicine, 1600 NW 10th Ave, Miami, FL 33136 (United States); Cleary, Margot P. [Hormel Institute, University of Minnesota, Austin, MN 55912 (United States); Gonzalez-Perez, Ruben R. [Department of Microbiology, Biochemistry and Immunology, Morehouse School of Medicine, Atlanta, GA 30314 (United States); Torroella-Kouri, Marta, E-mail: mtorroella@med.miami.edu [Department of Microbiology and Immunology, University of Miami Miller School of Medicine, 1600 NW 10th Ave, Miami, FL 33136 (United States); Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, 1475 NW 12th Ave, Miami, FL 33136 (United States)

    2015-01-15

    The relationship between obesity and breast cancer (BC) has focused on serum factors. However, the mammary gland contains adipose tissue (AT) which may enable the crosstalk between adipocytes and tumor cells contributing to tumor macrophage recruitment. We hypothesize that the breast AT (bAT) is inflamed in obese females and plays a major role in breast cancer development. The effects of this interplay on macrophage chemotaxis were examined in vitro, using co-cultures of mouse macrophages, mammary tumor cells and adipocytes. Macrophages were exposed to the adipocyte and tumor paracrine factors leptin, CCL2 and lauric acid (alone or in combinations). In cell supernatants Luminex identified additional molecules with chemotactic and other pro-tumor functions. Focus on the adipokine leptin, which has been shown to have a central role in breast cancer pathogenesis, indicated it modulates macrophage phenotypes and functions. In vivo experiments demonstrate that mammary tumors from obese mice are larger and that bAT from obese tumor-bearers contains higher numbers of macrophages/CLS and hypertrophic adipocytes than bAT from lean tumor-bearers, thus confirming it is more inflamed. Also, bAT distal from the tumor is more inflamed in obese than in lean mice. Our results reveal that bAT plays a role in breast cancer development in obesity.

  11. Paracrine Interactions between Adipocytes and Tumor Cells Recruit and Modify Macrophages to the Mammary Tumor Microenvironment: The Role of Obesity and Inflammation in Breast Adipose Tissue

    International Nuclear Information System (INIS)

    Santander, Ana M.; Lopez-Ocejo, Omar; Casas, Olivia; Agostini, Thais; Sanchez, Lidia; Lamas-Basulto, Eduardo; Carrio, Roberto; Cleary, Margot P.; Gonzalez-Perez, Ruben R.; Torroella-Kouri, Marta

    2015-01-01

    The relationship between obesity and breast cancer (BC) has focused on serum factors. However, the mammary gland contains adipose tissue (AT) which may enable the crosstalk between adipocytes and tumor cells contributing to tumor macrophage recruitment. We hypothesize that the breast AT (bAT) is inflamed in obese females and plays a major role in breast cancer development. The effects of this interplay on macrophage chemotaxis were examined in vitro, using co-cultures of mouse macrophages, mammary tumor cells and adipocytes. Macrophages were exposed to the adipocyte and tumor paracrine factors leptin, CCL2 and lauric acid (alone or in combinations). In cell supernatants Luminex identified additional molecules with chemotactic and other pro-tumor functions. Focus on the adipokine leptin, which has been shown to have a central role in breast cancer pathogenesis, indicated it modulates macrophage phenotypes and functions. In vivo experiments demonstrate that mammary tumors from obese mice are larger and that bAT from obese tumor-bearers contains higher numbers of macrophages/CLS and hypertrophic adipocytes than bAT from lean tumor-bearers, thus confirming it is more inflamed. Also, bAT distal from the tumor is more inflamed in obese than in lean mice. Our results reveal that bAT plays a role in breast cancer development in obesity

  12. Expression of bacterial virulence factors and cytokines during in vitro macrophage infection by enteroinvasive Escherichia coli and Shigella flexneri: a comparative study

    Directory of Open Access Journals (Sweden)

    Silvia Y Bando

    2010-09-01

    Full Text Available Enteroinvasive Escherichia coli (EIEC and Shigellaspp cause bacillary dysentery in humans by invading and multiplying within epithelial cells of the colonic mucosa. Although EIEC and Shigellashare many genetic and biochemical similarities, the illness caused by Shigellais more severe. Thus, genomic and structure-function molecular studies on the biological interactions of these invasive enterobacteria with eukaryotic cells have focused on Shigella rather than EIEC. Here we comparatively studied the interactions of EIEC and of Shigella flexneriwith cultured J774 macrophage-like cells. We evaluated several phenotypes: (i bacterial escape from macrophages after phagocytosis, (ii macrophage death induced by EIEC and S. flexneri, (iii macrophage cytokine expression in response to infection and (iv expression of plasmidial (pINV virulence genes. The results showed thatS. flexneri caused macrophage killing earlier and more intensely than EIEC. Both pathogens induced significant macrophage production of TNF, IL-1 and IL-10 after 7 h of infection. Transcription levels of the gene invasion plasmid antigen-C were lower in EIEC than in S. flexneri throughout the course of the infection; this could explain the diminished virulence of EIEC compared to S. flexneri.

  13. Macrophage Efferocytosis and Prostate Cancer Bone Metastasis

    Science.gov (United States)

    2018-03-01

    E) ELISA for total CXCL1 and CXCL5 levels in supernatants of MΦs alone or cocultured with RM1(HA) or PC3(HA). (F) Transcriptional activ- ity cell...marrow macrophages (Fig- ure 1D). ELISA evaluation for CXCL1 and CXCL5 proteins in the coculture media for apoptotic cancer cells (Figure 1E) confirmed... ELISA analysis of total pro- tein lysates from VEH- (n = 10) and AP-treated (n = 11) tumor vossicles. (G) Graphs depicting the correlation between

  14. Lactoferrin Efficiently Counteracts the Inflammation-Induced Changes of the Iron Homeostasis System in Macrophages.

    Science.gov (United States)

    Cutone, Antimo; Rosa, Luigi; Lepanto, Maria Stefania; Scotti, Mellani Jinnett; Berlutti, Francesca; Bonaccorsi di Patti, Maria Carmela; Musci, Giovanni; Valenti, Piera

    2017-01-01

    Human lactoferrin (hLf), an 80-kDa multifunctional iron-binding cationic glycoprotein, is constitutively secreted by exocrine glands and by neutrophils during inflammation. hLf is recognized as a key element in the host immune defense system. The in vitro and in vivo experiments are carried out with bovine Lf (bLf), which shares high sequence homology and identical functions with hLf, including anti-inflammatory activity. Here, in "pure" M1 human macrophages, obtained by stimulation with a mixture of 10 pg/ml LPS and 20 ng/ml IFN-γ, as well as in a more heterogeneous macrophage population, challenged with high-dose of LPS (1 µg/ml), the effect of bLf on the expression of the main proteins involved in iron and inflammatory homeostasis, namely ferroportin (Fpn), membrane-bound ceruloplasmin (Cp), cytosolic ferritin (Ftn), transferrin receptor 1, and cytokines has been investigated. The increase of IL-6 and IL-1β cytokines, following the inflammatory treatments, is associated with both upregulation of cytosolic Ftn and downregulation of Fpn, membrane-bound Cp, and transferrin receptor 1. All these changes take part into intracellular iron overload, a very unsafe condition leading in vivo to higher host susceptibility to infections as well as iron deficiency in the blood and anemia of inflammation. It is, therefore, of utmost importance to counteract the persistence of the inflammatory status to rebalance iron levels between tissues/secretions and blood. Moreover, levels of the antiinflammatory cytokine IL-10 were increased in cells treated with high doses of LPS. Conversely, IL-10 decreased when the LPS/IFN-γ mix was used, suggesting that only the inflammation triggered by LPS high doses can switch on an anti-inflammatory response in our macrophagic model. Here, we demonstrate that bLf, when included in the culture medium, significantly reduced IL-6 and IL-1β production and efficiently prevented the changes of Fpn, membrane-bound Cp, cytosolic Ftn, and

  15. Lactoferrin Efficiently Counteracts the Inflammation-Induced Changes of the Iron Homeostasis System in Macrophages

    Directory of Open Access Journals (Sweden)

    Antimo Cutone

    2017-06-01

    Full Text Available Human lactoferrin (hLf, an 80-kDa multifunctional iron-binding cationic glycoprotein, is constitutively secreted by exocrine glands and by neutrophils during inflammation. hLf is recognized as a key element in the host immune defense system. The in vitro and in vivo experiments are carried out with bovine Lf (bLf, which shares high sequence homology and identical functions with hLf, including anti-inflammatory activity. Here, in “pure” M1 human macrophages, obtained by stimulation with a mixture of 10 pg/ml LPS and 20 ng/ml IFN-γ, as well as in a more heterogeneous macrophage population, challenged with high-dose of LPS (1 µg/ml, the effect of bLf on the expression of the main proteins involved in iron and inflammatory homeostasis, namely ferroportin (Fpn, membrane-bound ceruloplasmin (Cp, cytosolic ferritin (Ftn, transferrin receptor 1, and cytokines has been investigated. The increase of IL-6 and IL-1β cytokines, following the inflammatory treatments, is associated with both upregulation of cytosolic Ftn and downregulation of Fpn, membrane-bound Cp, and transferrin receptor 1. All these changes take part into intracellular iron overload, a very unsafe condition leading in vivo to higher host susceptibility to infections as well as iron deficiency in the blood and anemia of inflammation. It is, therefore, of utmost importance to counteract the persistence of the inflammatory status to rebalance iron levels between tissues/secretions and blood. Moreover, levels of the antiinflammatory cytokine IL-10 were increased in cells treated with high doses of LPS. Conversely, IL-10 decreased when the LPS/IFN-γ mix was used, suggesting that only the inflammation triggered by LPS high doses can switch on an anti-inflammatory response in our macrophagic model. Here, we demonstrate that bLf, when included in the culture medium, significantly reduced IL-6 and IL-1β production and efficiently prevented the changes of Fpn, membrane-bound Cp

  16. Nitric oxide production from macrophages is regulated by arachidonic acid metabolites.

    Science.gov (United States)

    Imai, Y; Kolb, H; Burkart, V

    1993-11-30

    In activated macrophages the inducible form of the enzyme nitric oxide (NO) synthase generates high amounts of the toxic mediator NO. After 20 h of treatment with LPS rat peritoneal macrophages release 12-16 nmol NO2-/10(5) cells which is detectable in the culture supernatant by the Griess reaction as a measure of NO formation. The addition of aminoguanidine (1 mM), a preferential inhibitor of the inducible NO-synthase, completely abolished NO2-accumulation. Incubation with indomethacin or acetyl-salicylic acid, preferential inhibitors of the cyclooxygenase pathway of the arachidonic acid metabolism, did not influence NO2- levels. Nordihydro-guaiaretic acid (50 microM), a preferential inhibitor of the lipoxygenase pathway, caused strong reduction of NO2- accumulation to 1.9 +/- 0.3 nmol/200 microliter. Simultaneous inhibition of cyclo- and lipoxygenase by BW755c resulted in an intermediate effect (7.3 +/- 1.1 nmol/200 microliter NO2-). These results show that the induction of NO production in activated macrophages is regulated by products of the lipoxygenase-pathway of the arachidonic acid metabolism.

  17. Human mesenchymal stromal cell-secreted lactate induces M2-macrophage differentiation by metabolic reprogramming

    Science.gov (United States)

    Civini, Sara; Pacelli, Consiglia; Dieng, Mame Massar; Lemieux, William; Jin, Ping; Bazin, Renée; Patey, Natacha; Marincola, Francesco M.; Moldovan, Florina; Zaouter, Charlotte; Trudeau, Louis-Eric; Benabdhalla, Basma; Louis, Isabelle; Beauséjour, Christian; Stroncek, David; Le Deist, Françoise; Haddad, Elie

    2016-01-01

    Human mesenchymal stromal cells (MSC) have been shown to dampen immune response and promote tissue repair, but the underlying mechanisms are still under investigation. Herein, we demonstrate that umbilical cord-derived MSC (UC-MSC) alter the phenotype and function of monocyte-derived dendritic cells (DC) through lactate-mediated metabolic reprogramming. UC-MSC can secrete large quantities of lactate and, when present during monocyte-to-DC differentiation, induce instead the acquisition of M2-macrophage features in terms of morphology, surface markers, migratory properties and antigen presentation capacity. Microarray expression profiling indicates that UC-MSC modify the expression of metabolic-related genes and induce a M2-macrophage expression signature. Importantly, monocyte-derived DC obtained in presence of UC-MSC, polarize naïve allogeneic CD4+ T-cells into Th2 cells. Treatment of UC-MSC with an inhibitor of lactate dehydrogenase strongly decreases lactate concentration in culture supernatant and abrogates the effect on monocyte-to-DC differentiation. Metabolic analysis further revealed that UC-MSC decrease oxidative phosphorylation in differentiating monocytes while strongly increasing the spare respiratory capacity proportional to the amount of secreted lactate. Because both MSC and monocytes are recruited in vivo at the site of tissue damage and inflammation, we propose the local increase of lactate concentration induced by UC-MSC and the consequent enrichment in M2-macrophage generation as a mechanism to achieve immunomodulation. PMID:27070086

  18. Mertk deficiency affects macrophage directional migration via disruption of cytoskeletal organization.

    Directory of Open Access Journals (Sweden)

    Yong Tang

    Full Text Available Mertk belongs to the Tyro3, Axl and Mertk (TAM family of receptor tyrosine kinases, and plays a pivotal role in regulation of cytoskeletal rearrangement during phagocytosis. Phagocytosis by either professional or non-professional phagocytes is impaired in the Mertk deficient individual. In the present study, we further investigated the effects of Mertk mutation on peritoneal macrophage morphology, attachment, spreading and movement. Mertk-mutated macrophages exhibited decreased attachment, weak spreading, loss of spindle-like body shape and lack of clear leading and trailing edges within the first few hours of culture, as observed by environmental scanning electron microscopy. Time-lapse video photography recording showed that macrophage without Mertk conducted mainly random movement with oscillating swing around the cell body, and lost the directional migration action seen on the WT cells. Western blotting showed a decreased phosphorylation of focal adhesion kinase (FAK. Immunocytochemistry revealed that actin filaments and dynamic protein myosin II failed to concentrate in the leading edge of migrating cells. Microtubules were localized mainly in one side of mutant cell body, with no clear MTOC and associated radially-distributed microtubule bundles, which were clearly evident in the WT cells. Our results suggest that Mertk deficiency affects not only phagocytosis but also cell shape and migration, likely through a common regulatory mechanism on cytoskeletons.

  19. Expression of Nocardia brasiliensis superoxide dismutase during the early infection of murine peritoneal macrophages.

    Science.gov (United States)

    Revol, Agnès; Espinoza-Ruiz, Marisol; Medina-Villanueva, Igor; Salinas-Carmona, Mario Cesar

    2006-12-01

    Nocardia brasiliensis is the main agent of actinomycetoma in Mexico, but little is known about its virulence and molecular pathogenic pathways. These facultative intracellular bacteria are able to survive and divide within the host phagocytic cells, in part by neutralizing the reactive oxygen intermediates. Superoxide dismutase (SOD) participates in the intracellular survival of several bacterial species and, in particular, constitutes one of Nocardia asteroides virulence factors. To clarify SOD participation in the N. brasiliensis early infective process, we report its isolation and the consequent comparison of its transcript level. A 630 bp polymerase chain reaction fragment that included most of the coding sequence of N. brasiliensis sodA was cloned. A competitive assay was developed, allowing comparison of bacterial sod expression in exponential culture and 1 h after infecting peritoneal macrophages from BALB/c mice. At that time, there were viable bacteria in the macrophages. The intracellular bacteria presented a clear decrease in their sod transcript amount, although their 16S rRNA (used as an internal control) and hsp levels were maintained or slightly increased, respectively. These results indicate that sodA transcription is not maintained within the SOS bacterial response induced by phagosomal conditions. Further kinetics will be necessary to precisely define sod transcriptional regulation during N. brasiliensis intra-macrophage growth.

  20. Lysosomal cross-correction by hematopoietic stem cell-derived macrophages via tunneling nanotubes

    Science.gov (United States)

    Naphade, Swati; Sharma, Jay; Chevronnay, Héloïse P. Gaide; Shook, Michael A.; Yeagy, Brian A.; Rocca, Celine J.; Ur, Sarah N.; Lau, Athena J.; Courtoy, Pierre J.; Cherqui, Stephanie

    2014-01-01

    Despite controversies on the potential of hematopoietic stem cells (HSCs) to promote tissue repair, we previously showed that HSC transplantation could correct cystinosis, a multi-systemic lysosomal storage disease, caused by a defective lysosomal membrane cystine transporter, cystinosin (CTNS). Addressing the cellular mechanisms, we here report vesicular cross-correction after HSC differentiation into macrophages. Upon co-culture with cystinotic fibroblasts, macrophages produced tunneling nanotubes (TNTs) allowing transfer of cystinosin-bearing lysosomes into Ctns-deficient cells, which exploited the same route to retrogradely transfer cystine-loaded lysosomes to macrophages, providing a bidirectional correction mechanism. TNT formation was enhanced by contact with diseased cells. In vivo, HSCs grafted to cystinotic kidneys also generated nanotubular extensions resembling invadopodia that crossed the dense basement membranes and delivered cystinosin into diseased proximal tubular cells. This is the first report of correction of a genetic lysosomal defect by bidirectional vesicular exchange via TNTs and suggests broader potential for HSC transplantation for other disorders due to defective vesicular proteins. PMID:25186209

  1. Vitamin D Is Required for IFN-γ–Mediated Antimicrobial Activity of Human Macrophages

    Science.gov (United States)

    Fabri, Mario; Stenger, Steffen; Shin, Dong-Min; Yuk, Jae-Min; Liu, Philip T.; Realegeno, Susan; Lee, Hye-Mi; Krutzik, Stephan R.; Schenk, Mirjam; Sieling, Peter A.; Teles, Rosane; Montoya, Dennis; Iyer, Shankar S.; Bruns, Heiko; Lewinsohn, David M.; Hollis, Bruce W.; Hewison, Martin; Adams, John S.; Steinmeyer, Andreas; Zügel, Ulrich; Cheng, Genhong; Jo, Eun-Kyeong; Bloom, Barry R.; Modlin, Robert L.

    2012-01-01

    Control of tuberculosis worldwide depends on our understanding of human immune mechanisms, which combat the infection. Acquired T cell responses are critical for host defense against microbial pathogens, yet the mechanisms by which they act in humans remain unclear. We report that T cells, by the release of interferon-γ (IFN-γ), induce autophagy, phagosomal maturation, the production of antimicrobial peptides such as cathelicidin, and antimicrobial activity against Mycobacterium tuberculosis in human macrophages via a vitamin D–dependent pathway. IFN-γ induced the antimicrobial pathway in human macrophages cultured in vitamin D–sufficient sera, but not in sera from African-Americans that have lower amounts of vitamin D and who are more susceptible to tuberculosis. In vitro supplementation of vitamin D–deficient serum with 25-hydroxyvitamin D3 restored IFN-γ–induced antimicrobial peptide expression, autophagy, phagosome-lysosome fusion, and antimicrobial activity. These results suggest a mechanism in which vitamin D is required for acquired immunity to overcome the ability of intracellular pathogens to evade macrophage-mediated antimicrobial responses. The present findings underscore the importance of adequate amounts of vitamin D in all human populations for sustaining both innate and acquired immunity against infection. PMID:21998409

  2. Localization of CORO1A in the Macrophages Containing Mycobacterium leprae

    International Nuclear Information System (INIS)

    Suzuki, Koichi; Takeshita, Fumihiko; Nakata, Noboru; Ishii, Norihisa; Makino, Masahiko

    2006-01-01

    Mycobacteria have acquired an intracellular lifestyle within the macrophage, which is best exemplified by the enlarged infected histiocytes seen in lepromatous leprosy. To survive within the cell, mycobacteria must escape intracellular bactericidal mechanisms. In a study of Mycobacterium bovis Bacille Calmette-Guérin (M. bovis BCG) infection, it was shown that the host protein, CORO1A, also known as tryptophan aspartate-containing coat protein (TACO), accumulates on the phagosomal membrane, resulting in inhibition of phagosome-lysosome fusion, and thus augmenting intracellular survival. In this study, we show that CORO1A strongly localizes on the membrane of phagosomes that contain Mycobacterium leprae (M. leprae), where Toll-like receptor 2 was also visualized by immunostaining. When cultured macrophages were infected with M. leprae, CORO1A recruitment from the plasma membrane to the phagosomal membrane was observed. Moderate to strong CORO1A retention was observed in late lesions that contained foamy histiocytes, in which M. leprae were difficult to detect by acid-fast staining. These results suggest that components accumulating within the phagosome rather than viable bacilli are responsible for the retention of CORO1A, and that there is also a bactericidal mechanism in the macrophage that might counter the effects of CORO1A

  3. Enhanced Replication of Virulent Newcastle Disease Virus in Chicken Macrophages Is due to Polarized Activation of Cells by Inhibition of TLR7.

    Science.gov (United States)

    Zhang, Pingze; Ding, Zhuang; Liu, Xinxin; Chen, Yanyu; Li, Junjiao; Tao, Zhi; Fei, Yidong; Xue, Cong; Qian, Jing; Wang, Xueli; Li, Qingmei; Stoeger, Tobias; Chen, Jianjun; Bi, Yuhai; Yin, Renfu

    2018-01-01

    Newcastle disease (ND), caused by infections with virulent strains of Newcastle disease virus (NDV), is one of the most important infectious disease affecting wild, peridomestic, and domestic birds worldwide. Vaccines constructed from live, low-virulence (lentogenic) viruses are the most accepted prevention and control strategies for combating ND in poultry across the globe. Avian macrophages are one of the first cell lines of defense against microbial infection, responding to signals in the microenvironment. Although macrophages are considered to be one of the main target cells for NDV infection in vivo , very little is known about the ability of NDV to infect chicken macrophages, and virulence mechanisms of NDV as well as the polarized activation patterns of macrophages and correlation with viral infection and replication. In the present study, a cell culture model (chicken bone marrow macrophage cell line HD11) and three different virulence and genotypes of NDV (including class II virulent NA-1, class II lentogenic LaSota, and class I lentogenic F55) were used to solve the above underlying questions. Our data indicated that all three NDV strains had similar replication rates during the early stages of infection. Virulent NDV titers were shown to increase compared to the other lentogenic strains, and this growth was associated with a strong upregulation of both pro-inflammatory M1-like markers/cytokines and anti-inflammatory M2-like markers/cytokines in chicken macrophages. Virulent NDV was found to block toll-like receptor (TLR) 7 expression, inducing higher expression of type I interferons in chicken macrophages at the late stage of viral infection. Only virulent NDV replication can be inhibited by pretreatment with TLR7 ligand. Overall, this study demonstrated that virulent NDV activates a M1-/M2-like mixed polarized activation of chicken macrophages by inhibition of TLR7, resulting in enhanced replication compared to lentogenic viruses.

  4. Silica-induced initiation of circular ZC3H4 RNA/ZC3H4 pathway promotes the pulmonary macrophage activation.

    Science.gov (United States)

    Yang, Xiyue; Wang, Jing; Zhou, Zewei; Jiang, Rong; Huang, Jie; Chen, Lulu; Cao, Zhouli; Chu, Han; Han, Bing; Cheng, Yusi; Chao, Jie

    2018-01-22

    Phagocytosis of silicon dioxide (SiO 2 ) into lung cells causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Circular RNAs (circRNAs) are a subclass of noncoding RNAs that are present within mammalian cells; however, researchers have not determined whether circRNAs are involved in the pathophysiologic process of silicosis. To elucidate the role of these RNAs in SiO 2 -induced inflammation in pulmonary macrophages, we investigated the upstream molecular mechanisms and functional effects of circRNAs on cell apoptosis, proliferation, and migration. Primary cultures of alveolar macrophages from healthy donors and from patients and the RAW264.7 macrophage cell line were used to explore the functions of circZC3H4 RNA in macrophage activation. The experimental results indicated the following: 1) SiO 2 concomitantly increased circZC3H4 RNA expression and increased ZC3H4 protein levels; 2) circular ZC3H4 (circZC3H4) RNA and ZC3H4 protein participated in SiO 2 -induced macrophage activation; and 3) SiO 2 -activated macrophages promoted fibroblast proliferation and migration via the circZC3H4 RNA/ZC3H4 pathway. The up-regulation of the ZC3H4 protein was confirmed in tissue samples from patients with silicosis. Our study elucidates a link between SiO 2 -induced macrophage activation and the circZC3H4 RNA/ZC3H4 pathway, thereby providing novel insight into the potential use of ZC3H4 to develop novel therapeutic strategies for silicosis.-Yang, X., Wang, J., Zhou, Z., Jiang, R., Huang, J., Chen, L., Cao, Z., Chu, H., Han, B., Cheng, Y., Chao, J. Silica-induced initiation of circular ZC3H4 RNA/ZC3H4 pathway promotes the pulmonary macrophage activation.

  5. Probiotic Bacteria Alter Pattern-Recognition Receptor Expression and Cytokine Profile in a Human Macrophage Model Challenged with Candida albicans and Lipopolysaccharide

    Directory of Open Access Journals (Sweden)

    Victor H. Matsubara

    2017-11-01

    Full Text Available Probiotics are live microorganisms that confer benefits to the host health. The infection rate of potentially pathogenic organisms such as Candida albicans, the most common agent associated with mucosal candidiasis, can be reduced by probiotics. However, the mechanisms by which the probiotics interfere with the immune system are largely unknown. We evaluated the effect of probiotic bacteria on C. albicans challenged human macrophages. Macrophages were pretreated with lactobacilli alone (Lactobacillus rhamnosus LR32, Lactobacillus casei L324m, or Lactobacillus acidophilus NCFM or associated with Escherichia coli lipopolysaccharide (LPS, followed by the challenge with C. albicans or LPS in a co-culture assay. The expression of pattern-recognition receptors genes (CLE7A, TLR2, and TLR4 was determined by RT-qPCR, and dectin-1 reduced levels were confirmed by flow cytometry. The cytokine profile was determined by ELISA using the macrophage cell supernatant. Overall probiotic lactobacilli down-regulated the transcription of CLEC7A (p < 0.05, resulting in the decreased expression of dectin-1 on probiotic pretreated macrophages. The tested Lactobacillus species down-regulated TLR4, and increased TLR2 mRNA levels in macrophages challenged with C. albicans. The cytokines profile of macrophages challenged with C. albicans or LPS were altered by the probiotics, which generally led to increased levels of IL-10 and IL-1β, and reduction of IL-12 production by macrophages (p < 0.05. Our data suggest that probiotic lactobacilli impair the recognition of PAMPs by macrophages, and alter the production of pro/anti-inflammatory cytokines, thus modulating inflammation.

  6. Macropinocytosis is decreased in diabetic mouse macrophages and is regulated by AMPK

    Directory of Open Access Journals (Sweden)

    Chakour Kenneth S

    2008-07-01

    Full Text Available Abstract Background Macrophages (MΦs utilize macropinocytosis to integrate immune and metabolic signals in order to initiate an effective immune response. Diabetes is characterized by metabolic abnormalities and altered immune function. Here we examine the influence of diabetes on macropinocytosis in primary mouse macrophages and in an in vitro diabetes model. Results The data demonstrate that peritoneal MΦs from diabetic (db/db mice had reduced macropinocytosis when compared to MΦs from non-diabetic (db/+ mice. Additionally, MΦs cultured in hyperglycemic conditions were less adept at macropinocytosis than those cultured in low glucose. Notably, AMP-activated protein kinase (AMPK activity was decreased in MΦs cultured in hyperglycemic conditions. Activation of AMPK with leptin or 5-aminoimidazole-4-carboxamide-1-β-riboside (AICAR increased macropinocytosis and inhibition of AMPK with compound C decreased macropinocytosis. Conclusion Taken together, these findings indicate that MΦs from diabetic mice have decreased macropinocytosis. This decrease appears dependent on reduced AMPK activity. These results demonstrate a previously unrealized role for AMPK in MΦs and suggest that increasing AMPK activity in diabetic MΦs could improve innate immunity and decrease susceptibility to infection.

  7. Activation of macrophages for microbicidal and tumoricidal effector functions by soluble factors from EL-4, a continuous T cell line.

    OpenAIRE

    Nacy, C A; James, S L; Benjamin, W R; Farrar, J J; Hockmeyer, W T; Meltzer, M S

    1983-01-01

    Macrophages treated with culture fluids from EL-4 cells, a continuous T cell line, were activated to kill mKSA-TU-5 fibrosarcoma cells, amastigotes of Leishmania tropica, and schistosomula of Schistosoma mansoni. Active EL-4 factors eluted from Sephadex G-100 in two distinct regions: molecular weight 45,000 (activities induced killing of unrelated intracellular and extracellular targets) and molecular weight 23,000 (activities induced killing of extracellular targets only). These results conf...

  8. Host-Pathogen Coupled Networks: Model for Bacillus Anthracis Interaction with Host Macrophages

    Science.gov (United States)

    2015-09-01

    circles). Data points represent absorbance values at 570 nm following the MTT assay. Fitted values are KX2 = 10-6 nM-1s-1 and R = 0.21 nM s-1. (B...represent absorbance values at 570 nm following MTT assay. Here cell “death”, as indicated by the diminution of the signal (corrected for residual...cleaves the N-terminus of MAPKKs and induces tyrosine /threonine phosphorylation of MAPKs in cultured macrophages. Biochem Biophys Res Commun. 248(3):706

  9. Accumulation of M1-like macrophages in type 2 diabetic islets is followed by a systemic shift in macrophage polarization.

    Science.gov (United States)

    Cucak, Helena; Grunnet, Lars Groth; Rosendahl, Alexander

    2014-01-01

    Human T2D is characterized by a low-grade systemic inflammation, loss of β-cells, and diminished insulin production. Local islet immunity is still poorly understood, and hence, we evaluated macrophage subpopulations in pancreatic islets in the well-established murine model of T2D, the db/db mouse. Already at 8 weeks of disease, on average, 12 macrophages were observed in the diabetic islets, whereas only two were recorded in the nondiabetic littermates. On a detailed level, the islet resident macrophages increased fourfold compared with nondiabetic littermates, whereas a pronounced recruitment (eightfold) of a novel subset of macrophages (CD68+F4/80-) was observed. The majority of the CD68+F4/80+ but only 40% of the CD68+F4/80- islet macrophages expressed CD11b. Both islet-derived macrophage subsets expressed moderate MHC-II, high galectin-3, and low CD80/CD86 levels, suggesting the cells to be macrophages rather than DCs. On a functional level, the vast majority of the macrophages in the diabetic islets was of the proinflammatory, M1-like phenotype. The systemic immunity in diabetic animals was characterized by a low-grade inflammation with elevated cytokine levels and increase of splenic cytokine, producing CD68+F4/80- macrophages. In late-stage diabetes, the cytokine signature changed toward a TGF-β-dominated profile, coinciding with a significant increase of galectin-3-positive macrophages in the spleen. In summary, our results show that proinflammatory M1-like galectin-3+ CD80/CD86(low) macrophages invade diabetic islets. Moreover, the innate immunity matures in a diabetes-dependent manner from an initial proinflammatory toward a profibrotic phenotype, supporting the concept that T2D is an inflammatory disease.

  10. Transcriptional landscape of Mycobacterium tuberculosis infection in macrophages

    KAUST Repository

    Roy, Sugata; Schmeier, Sebastian; Kaczkowski, Bogumil; Arner, Erik; Alam, Tanvir; Ozturk, Mumin; Tamgue, Ousman; Parihar, Suraj P.; Kawaji, Hideya; Itoh, Masayoshi; Lassmann, Timo; Carninci, Piero; Hayashizaki, Yoshihide; Forrest, Alistair R. R.; Guler, Reto; Bajic, Vladimir B.; Brombacher, Frank; Suzuki, Harukazu

    2018-01-01

    landscape of IFNγ (M1) or IL-4/IL-13 (M2) stimulated macrophages during Mtb infection in a time-kinetic manner. Mtb infection widely and drastically altered macrophage-specific gene expression, which is far larger than that of M1 or M2 activations. Gene

  11. DNA Damage Signaling Instructs Polyploid Macrophage Fate in Granulomas

    DEFF Research Database (Denmark)

    Herrtwich, Laura; Nanda, Indrajit; Evangelou, Konstantinos

    2016-01-01

    to a chronic stimulus, though critical for disease outcome, have not been defined. Here, we delineate a macrophage differentiation pathway by which a persistent Toll-like receptor (TLR) 2 signal instructs polyploid macrophage fate by inducing replication stress and activating the DNA damage response. Polyploid...

  12. Niacin and its metabolites as master regulators of macrophage activation.

    Science.gov (United States)

    Montserrat-de la Paz, Sergio; Naranjo, M Carmen; Lopez, Sergio; Abia, Rocio; Muriana, Francisco J Garcia; Bermudez, Beatriz

    2017-01-01

    Niacin is a broad-spectrum lipid-regulating drug used for clinical therapy of chronic high-grade inflammatory diseases. However, the mechanisms by which either niacin or the byproducts of its catabolism ameliorate these inflammatory diseases are not clear yet. Human circulating monocytes and mature macrophages were used to analyze the effects of niacin and its metabolites (NAM, NUA and 2-Pyr) on oxidative stress, plasticity and inflammatory response by using biochemical, flow cytometry, quantitative real-time PCR and Western blot technologies. Niacin, NAM and 2-Pyr significantly decreased ROS, NO and NOS2 expression in LPS-treated human mature macrophages. Niacin and NAM skewed macrophage polarization toward antiinflammatory M2 macrophage whereas a trend toward proinflammatory M1 macrophage was noted following treatment with NUA. Niacin and NAM also reduced the inflammatory competence of LPS-treated human mature macrophages and promoted bias toward antiinflammatory CD14 + CD16 ++ nonclassical human primary monocytes. This study reveals for the first time that niacin and its metabolites possess antioxidant, reprogramming and antiinflammatory properties on human primary monocytes and monocyte-derived macrophages. Our findings imply a new understanding of the mechanisms by which niacin and its metabolites favor a continuous and gradual plasticity process in the human monocyte/macrophage system. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Epigenetic pathways in macrophages emerge as novel targets in atherosclerosis

    NARCIS (Netherlands)

    Neele, Annette E.; van den Bossche, Jan; Hoeksema, Marten A.; de Winther, Menno P. J.

    2015-01-01

    Atherosclerosis is a lipid-driven chronic inflammatory disorder. Monocytes and macrophages are key immune cells in the development of disease and clinical outcome. It is becoming increasingly clear that epigenetic pathways govern many aspects of monocyte and macrophage differentiation and

  14. Nanomedicine Strategies to Target Tumor-Associated Macrophages

    NARCIS (Netherlands)

    Binnemars-Postma, Karin A.; Storm, G; Prakash, Jai

    2017-01-01

    In recent years, the influence of the tumor microenvironment (TME) on cancer progression has been better understood. Macrophages, one of the most important cell types in the TME, exist in different subtypes, each of which has a different function. While classically activated M1 macrophages are

  15. Nanomedicine strategies to target tumor-associated macrophages

    NARCIS (Netherlands)

    Binnemars-Postma, Karin; Storm, Gert; Prakash, Jai

    2017-01-01

    In recent years, the influence of the tumor microenvironment (TME) on cancer progression has been better understood. Macrophages, one of the most important cell types in the TME, exist in different subtypes, each of which has a different function. While classically activated M1 macrophages are

  16. Biomaterials Influence Macrophage-Mesenchymal Stem Cell Interaction In Vitro

    NARCIS (Netherlands)

    N. Grotenhuis (Nienke); S.F. De Witte (Samantha Fh); G.J.V.M. van Osch (Gerjo); Y. Bayon (Yves); J.F. Lange (Johan); Y.M. Bastiaansen-Jenniskens (Yvonne)

    2016-01-01

    textabstractBackground: Macrophages and mesenchymal stem cells (MSCs) are important cells in wound healing. We hypothesized that the cross-talk between macrophages and adipose tissue-derived MSCs (ASCs) is biomaterial dependent, thereby influencing processes involved in wound healing. Materials and

  17. Macrophages are critical effectors of antibody therapies for cancer.

    Science.gov (United States)

    Weiskopf, Kipp; Weissman, Irving L

    2015-01-01

    Macrophages are innate immune cells that derive from circulating monocytes, reside in all tissues, and participate in many states of pathology. Macrophages play a dichotomous role in cancer, where they promote tumor growth but also serve as critical immune effectors of therapeutic antibodies. Macrophages express all classes of Fcγ receptors, and they have immense potential to destroy tumors via the process of antibody-dependent phagocytosis. A number of studies have demonstrated that macrophage phagocytosis is a major mechanism of action of many antibodies approved to treat cancer. Consequently, a number of approaches to augment macrophage responses to therapeutic antibodies are under investigation, including the exploration of new targets and development of antibodies with enhanced functions. For example, the interaction of CD47 with signal-regulatory protein α (SIRPα) serves as a myeloid-specific immune checkpoint that limits the response of macrophages to antibody therapies, and CD47-blocking agents overcome this barrier to augment phagocytosis. The response of macrophages to antibody therapies can also be enhanced with engineered Fc variants, bispecific antibodies, or antibody-drug conjugates. Macrophages have demonstrated success as effectors of cancer immunotherapy, and further investigation will unlock their full potential for the benefit of patients.

  18. Of macrophages and red blood cells; a complex love story

    NARCIS (Netherlands)

    de Back, Djuna Z.; Kostova, Elena B.; van Kraaij, Marian; van den Berg, Timo K.; van Bruggen, Robin

    2014-01-01

    Macrophages tightly control the production and clearance of red blood cells (RBC). During steady state hematopoiesis, approximately 10(10) RBC are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with

  19. Of macrophages and red blood cells; a complex love story.

    Directory of Open Access Journals (Sweden)

    Djuna Zoe de Back

    2014-01-01

    Full Text Available Macrophages tightly control the production and clearance of red blood cells (RBC. During steady state haematopoiesis, approximately 1010 red blood cells are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

  20. Of macrophages and red blood cells; a complex love story.

    Science.gov (United States)

    de Back, Djuna Z; Kostova, Elena B; van Kraaij, Marian; van den Berg, Timo K; van Bruggen, Robin

    2014-01-01

    Macrophages tightly control the production and clearance of red blood cells (RBC). During steady state hematopoiesis, approximately 10(10) RBC are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

  1. Cytokine expression of macrophages in HIV-1-associated vacuolar myelopathy.

    Science.gov (United States)

    Tyor, W R; Glass, J D; Baumrind, N; McArthur, J C; Griffin, J W; Becker, P S; Griffin, D E

    1993-05-01

    Macrophages are frequently present within the periaxonal and intramyelinic vacuoles that are located primarily in the posterior and lateral funiculi of the thoracic spinal cord in HIV-associated vacuolar myelopathy. But the role of these macrophages in the formation of the vacuoles is unclear. One hypothesis is that cytokines, such as interleukin-1 (IL-1) and tumor necrosis factor (TNF)-alpha, are produced locally by macrophages and have toxic effects on myelin or oligodendrocytes. The resulting myelin damage eventually culminates in the removal of myelin by macrophages and vacuole formation. We studied thoracic spinal cord specimens taken at autopsy from HIV-positive (+) and HIV-negative individuals. The predominant mononuclear cells present in HIV+ spinal cords are macrophages. They are located primarily in the posterior and lateral funiculi regardless of the presence or absence of vacuolar myelopathy. Macrophages and microglia are more frequent in HIV+ than HIV-negative individuals and these cells frequently stain for class I and class II antigens, IL-1, and TNF-alpha. Activated macrophages positive for IL-1 and TNF-alpha are great increased in the posterior and lateral funiculi of HIV+ individuals with and without vacuolar myelopathy, suggesting they are present prior to the development of vacuoles. Cytokines, such as TNF-alpha, may be toxic for myelin or oligodendrocytes, leading to myelin damage and removal by macrophages and vacuole formation.

  2. Vascular endothelial growth factor (VEGF), produced by feline infectious peritonitis (FIP) virus-infected monocytes and macrophages, induces vascular permeability and effusion in cats with FIP.

    Science.gov (United States)

    Takano, Tomomi; Ohyama, Taku; Kokumoto, Aiko; Satoh, Ryoichi; Hohdatsu, Tsutomu

    2011-06-01

    Feline infectious peritonitis virus (FIPV) causes a fatal disease called FIP in Felidae. The effusion in body cavity is commonly associated with FIP. However, the exact mechanism of accumulation of effusion remains unclear. We investigated vascular endothelial growth factor (VEGF) to examine the relationship between VEGF levels and the amounts of effusion in cats with FIP. Furthermore, we examined VEGF production in FIPV-infected monocytes/macrophages, and we used feline vascular endothelial cells to examine vascular permeability induced by the culture supernatant of FIPV-infected macrophages. In cats with FIP, the production of effusion was related with increasing plasma VEGF levels. In FIPV-infected monocytes/macrophages, the production of VEGF was associated with proliferation of virus. Furthermore, the culture supernatant of FIPV-infected macrophages induced hyperpermeability of feline vascular endothelial cells. It was suggested that vascular permeability factors, including VEGF, produced by FIPV-infected monocytes/macrophages might increase the vascular permeability and the amounts of effusion in cats with FIP. Copyright © 2011 Elsevier B.V. All rights reserved.

  3. Toxicity of penicillic acid for rat alveolar macrophages in vitro

    International Nuclear Information System (INIS)

    Sorenson, W.G.; Simpson, J.

    1985-01-01

    Penicillic acid (PA) is a polyketide mycotoxin produced by several species of Aspergillus and Penicillium. This mycotoxin is toxic in experimental animals and has also been reported to be carcinogenic. The cytotoxicity of penicillic acid was studied in rat albeolar macrophages (AM) in vitro. The effects of penicillic acid on membrane integrity were studied by measuring cell volume changes and 51 Cr release. There was a significant decrease in adenosine triphosphate (ATP) in cell cultures exposed to 1.0 mM penicillic acid for 4 hr. Inhibition of the incorporation of [ 3 H]leucine into protein was both dose- and time-dependent and protein synthesis was inhibited significantly after 2 hr exposure to ≥0.1 mM penicillic acid. RNA synthesis was inhibited to a lesser extent than protein synthesis. There was significant inhibition of phagocytosis after 2 hr exposure at ≥0.3 mM penicillic acid and the ED 50 for phagocytosis was 0.09 mM. Thus phagocytosis was more sensitive to the toxic effects of penicillic acid than any other cellular process studied. The data suggest the possibility of a respiratory hazard to agricultural workers exposed to contaminated grain

  4. Arachidonic acid metabolism in silica-stimulated bovine alveolar macrophages

    International Nuclear Information System (INIS)

    Englen, M.D.

    1989-01-01

    The in vitro production of arachidonic acid (AA) metabolites in adherent bovine alveolar macrophages (BAM) incubated with silica was investigated. BAM were pre-labelled with 3 H-AA, and lipid metabolites released into the culture medium were analyzed by high performance liquid chromatography (HPLC). Lactate dehydrogenase (LDH) release was simultaneously assayed to provide an indication of cell injury. Increasing doses of silica selectively stimulated the 5-lipoxygenase pathway of AA metabolism, while cyclooxygenase metabolite output was suppressed. LDH release increased in a linear, dose-dependent fashion over the range of silica doses used. Moreover, within 15 min following addition of a high silica dose, a shift to the production of 5-lipoxygenase metabolites occurred, accompanied by a reduction in cyclooxygenase products. This rapid alteration in AA metabolism preceded cell injury. To examine the relationship between cytotoxicity and AA metabolite release by BAM exposed to silicas with different cytotoxic and fibrogenic activities, BAM were exposed to different doses of DQ-12, Minusil-5, and Sigma silicas, and carbonyl iron beads. The median effective dose (ED 50 ) of each particulate to stimulate the release of AA metabolites and LDH was calculated. The ED 50 values for DQ-12, Minusil-5, and Sigma silica showed that the relative cytotoxicities of the different silicas for BAM corresponded to the relative potencies of the silicas to elicit 5-lipoxygenase metabolites from BAM. These results indicate that the cytotoxic, and presumed fibrogenic potential, of a silica is correlated with the potency to stimulate the release of leukotrienes from AM

  5. Modulation of macrophage Ia expression by lipopolysaccharide: Stem cell requirements, accessory lymphocyte involvement, and IA-inducing factor production

    International Nuclear Information System (INIS)

    Wentworth, P.A.; Ziegler, H.K.

    1989-01-01

    The mechanism of induction of murine macrophage Ia expression by lipopolysaccharide (LPS) was studied. Intraperitoneal injection of 1 microgram of LPS resulted in a 3- to 10-fold increase in the number of IA-positive peritoneal macrophages (flow cytometry and immunofluorescence) and a 6-to 16-fold increase by radioimmunoassay. The isolated lipid A moiety of LPS was a potent inducer of macrophage Ia expression. Ia induction required a functional myelopoietic system as indicated by the finding that the response to LPS was eliminated in irradiated (900 rads) mice and reinstated by reconstitution with bone marrow cells. Comparison of LPS-induced Ia expression in normal and LPS-primed mice revealed a faster secondary response to LPS. The memory response could be adoptively transferred to normal mice with nonadherent spleen cells prepared 60 days after LPS injection. Spleen cells prepared 5 days after LPS injection caused Ia induction in LPS-nonresponder mice; such induction was not observed in irradiated (900 rads) recipients. The cell responsible for this phenomenon was identified as a Thy-1+, immunoglobulin-negative nonadherent cell. The biosynthesis and expression of Ia were not increased by direct exposure of macrophages to LPS in vitro. Small amounts of LPS inhibited Ia induction by gamma interferon. LPS showed positive regulatory effects on Ia expression by delaying the loss of Ia expression on cultured macrophages and by stimulating the production of Ia-inducing factors. Supernatants from cultured spleen cells stimulated with LPS in vitro contained antiviral and Ia-inducing activity that was acid labile, indicating that the active factor is gamma interferon. We conclude that induction of Ia expression by LPS in vivo is a bone-marrow-dependent, radiation-sensitive process which involves the stimulation of a gamma interferon-producing accessory lymphocyte and a delay in Ia turnover

  6. Selective phosphorylation during early macrophage differentiation

    KAUST Repository

    Zhang, Huoming

    2015-08-26

    The differentiation of macrophages from monocytes is a tightly controlled and complex biological process. Although numerous studies have been conducted using biochemical approaches or global gene/gene profiling, the mechanisms of the early stages of differentiation remain unclear. Here we used SILAC-based quantitative proteomics approach to perform temporal phosphoproteome profiling of early macrophage differentiation. We identified a large set of phosphoproteins and grouped them as PMA-regulated and non-regulated phosphoproteins in the early stages of differentiation. Further analysis of the PMA-regulated phosphoproteins revealed that transcriptional suppression, cytoskeletal reorganization and cell adhesion were among the most significantly activated pathways. Some key involved regulators of these pathways are mTOR, MYB, STAT1 and CTNNB. Moreover, we were able to classify the roles and activities of several transcriptional factors during different differentiation stages and found that E2F is likely to be an important regulator during the relatively late stages of differentiation. This study provides the first comprehensive picture of the dynamic phosphoproteome during myeloid cells differentiation, and identifies potential molecular targets in leukemic cells.

  7. Hemocyanins Stimulate Innate Immunity by Inducing Different Temporal Patterns of Proinflammatory Cytokine Expression in Macrophages.

    Science.gov (United States)

    Zhong, Ta-Ying; Arancibia, Sergio; Born, Raimundo; Tampe, Ricardo; Villar, Javiera; Del Campo, Miguel; Manubens, Augusto; Becker, María Inés

    2016-06-01

    Hemocyanins induce a potent Th1-dominant immune response with beneficial clinical outcomes when used as a carrier/adjuvant in vaccines and nonspecific immunostimulant in cancer. However, the mechanisms by which hemocyanins trigger innate immune responses, leading to beneficial adaptive immune responses, are unknown. This response is triggered by a proinflammatory signal from various components, of which macrophages are an essential part. To understand how these proteins influence macrophage response, we investigated the effects of mollusks hemocyanins with varying structural and immunological properties, including hemocyanins from Concholepas concholepas, Fissurella latimarginata, and Megathura crenulata (keyhole limpet hemocyanin), on cultures of peritoneal macrophages. Hemocyanins were phagocytosed and slowly processed. Analysis of this process showed differential gene expression along with protein levels of proinflammatory markers, including IL-1β, IL-6, IL-12p40, and TNF-α. An extended expression analysis of 84 cytokines during a 24-h period showed a robust proinflammatory response for F. latimarginata hemocyanin in comparison with keyhole limpet hemocyanin and C. concholepas hemocyanin, which was characterized by an increase in the transcript levels of M1 cytokines involved in leukocyte recruitment. These cytokine genes included chemokines (Cxcl1, Cxcl3, Cxcl5, Ccl2, and Ccl3), ILs (Il1b and Ifng), growth factors (Csf2 and Csf3), and TNF family members (Cd40lg). The protein levels of certain cytokines were increased. However, every hemocyanin maintains downregulated key M2 cytokine genes, including Il4 and Il5 Collectively, our data demonstrate that hemocyanins are able to trigger the release of proinflammatory factors with different patterns of cytokine expression, suggesting differential signaling pathways and transcriptional network mechanisms that lead to the activation of M1-polarized macrophages. Copyright © 2016 by The American Association of

  8. Viral Inhibition of Bacterial Phagocytosis by Human Macrophages: Redundant Role of CD36.

    Directory of Open Access Journals (Sweden)

    Grace E Cooper

    Full Text Available Macrophages are essential to maintaining lung homoeostasis and recent work has demonstrated that influenza-infected lung macrophages downregulate their expression of the scavenger receptor CD36. This receptor has also been shown to be involved in phagocytosis of Streptococcus pneumoniae, a primary agent associated with pneumonia secondary to viral infection. The aim of this study was to investigate the role of CD36 in the effects of viral infection on macrophage phagocytic function. Human monocyte-derived macrophages (MDM were exposed to H3N2 X31 influenza virus, M37 respiratory syncytial virus (RSV or UV-irradiated virus. No infection of MDM was seen upon exposure to UV-irradiated virus but incubation with live X31 or M37 resulted in significant levels of viral detection by flow cytometry or RT-PCR respectively. Infection resulted in significantly diminished uptake of S. pneumoniae by MDM and significantly decreased expression of CD36 at both the cell surface and mRNA level. Concurrently, there was a significant increase in IFNβ gene expression in response to infection and we observed a significant decrease in bacterial phagocytosis (p = 0.031 and CD36 gene expression (p = 0.031 by MDM cultured for 24 h in 50IU/ml IFNβ. Knockdown of CD36 by siRNA resulted in decreased phagocytosis, but this was mimicked by transfection reagent alone. When MDM were incubated with CD36 blocking antibodies no effect on phagocytic ability was observed. These data indicate that autologous IFNβ production by virally-infected cells can inhibit bacterial phagocytosis, but that decreased CD36 expression by these cells does not play a major role in this functional deficiency.

  9. Extracellular polysaccharides produced by Ganoderma formosanum stimulate macrophage activation via multiple pattern-recognition receptors

    Directory of Open Access Journals (Sweden)

    Wang Cheng-Li

    2012-08-01

    Full Text Available Abstract Background The fungus of Ganoderma is a traditional medicine in Asia with a variety of pharmacological functions including anti-cancer activities. We have purified an extracellular heteropolysaccharide fraction, PS-F2, from the submerged mycelia culture of G. formosanum and shown that PS-F2 exhibits immunostimulatory activities. In this study, we investigated the molecular mechanisms of immunostimulation by PS-F2. Results PS-F2-stimulated TNF-α production in macrophages was significantly reduced in the presence of blocking antibodies for Dectin-1 and complement receptor 3 (CR3, laminarin, or piceatannol (a spleen tyrosine kinase inhibitor, suggesting that PS-F2 recognition by macrophages is mediated by Dectin-1 and CR3 receptors. In addition, the stimulatory effect of PS-F2 was attenuated in the bone marrow-derived macrophages from C3H/HeJ mice which lack functional Toll-like receptor 4 (TLR4. PS-F2 stimulation triggered the phosphorylation of mitogen-activated protein kinases JNK, p38, and ERK, as well as the nuclear translocation of NF-κB, which all played essential roles in activating TNF-α expression. Conclusions Our results indicate that the extracellular polysaccharides produced by G. formosanum stimulate macrophages via the engagement of multiple pattern-recognition receptors including Dectin-1, CR3 and TLR4, resulting in the activation of Syk, JNK, p38, ERK, and NK-κB and the production of TNF-α.

  10. [DNA hydroxymethylase 10-11 translocation 2 (TET2) inhibits mouse macrophage activation and polarization].

    Science.gov (United States)

    Li, Bingyi; Huo, Yi; Lin, Zhifeng; Wang, Tao

    2017-09-01

    Objective To study the role of DNA hydroxymethylase 10-11 translocation 2 (TET2) in macrophage activation and polarization. Methods RAW264.7 macrophages were cultured in vitro and stimulated with 100 ng/mL LPS for 0, 1, 2, 4, 6 hours. Real-time quantitative PCR was used to detect TET2 mRNA expression. TET2 expression was knocked down with siRNA and the knock-down efficiency was evaluated by real-time quantitative PCR and Western blotting. Following siRNA transfection for 48 hours, RAW264.7 cells were stimulated by LPS for 4 hours, and then real-time quantitative PCR and ELISA were performed to detect the expressions of interleukin 6 (IL-6), tumor necrosis factor α (TNF-α) and IL-12. The M1 polarizing markers TNF-α, inducible nitric oxide synthase (iNOS) and IL-12, and M2 polarizing markers mannose receptor (MR), arginase 1 (Arg-1) and chitinase 3-like molecule 1 (Ym1) were tested after M1 or M2 induction by LPS/IFN-γ or IL-4. Results TET2 expression increased after LPS treatment in RAW264.7 cells and reached the peak at 2 hours later. The siRNA effectively reduced the expression of TET2. The expressions of IL-6, TNF-α and IL-12 mRNAs increased after TET2 knock-down and LPS stimulation. The expressions of M1 polarization markers and M2 markers were up-regulated by the corresponding stimulations after TET2 knock-down. Conclusion TET2 has the effect of inhibiting LPS-induced macrophage activation and plays an inhibitory role in macrophage M1 and M2 polarization.

  11. Therapeutic potential of carbohydrates as regulators of macrophage activation.

    Science.gov (United States)

    Lundahl, Mimmi L E; Scanlan, Eoin M; Lavelle, Ed C

    2017-12-15

    It is well established for a broad range of disease states, including cancer and Mycobacterium tuberculosis infection, that pathogenesis is bolstered by polarisation of macrophages towards an anti-inflammatory phenotype, known as M2. As these innate immune cells are relatively long-lived, their re-polarisation to pro-inflammatory, phagocytic and bactericidal "classically activated" M1 macrophages is an attractive therapeutic approach. On the other hand, there are scenarios where the resolving inflammation, wound healing and tissue remodelling properties of M2 macrophages are beneficial - for example the successful introduction of biomedical implants. Although there are numerous endogenous and exogenous factors that have an impact on the macrophage polarisation spectrum, this review will focus specifically on prominent macrophage-modulating carbohydrate motifs with a view towards highlighting structure-function relationships and therapeutic potential. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Macrophages recognize size and shape of their targets.

    Directory of Open Access Journals (Sweden)

    Nishit Doshi

    2010-04-01

    Full Text Available Recognition by macrophages is a key process in generating immune response against invading pathogens. Previous studies have focused on recognition of pathogens through surface receptors present on the macrophage's surface. Here, using polymeric particles of different geometries that represent the size and shape range of a variety of bacteria, the importance of target geometry in recognition was investigated. The studies reported here reveal that attachment of particles of different geometries to macrophages exhibits a strong dependence on size and shape. For all sizes and shapes studied, particles possessing the longest dimension in the range of 2-3 microm exhibited highest attachment. This also happens to be the size range of most commonly found bacteria in nature. The surface features of macrophages, in particular the membrane ruffles, might play an important role in this geometry-based target recognition by macrophages. These findings have significant implications in understanding the pathogenicity of bacteria and in designing drug delivery carriers.

  13. Soluble ICAM-1 activates lung macrophages and enhances lung injury

    DEFF Research Database (Denmark)

    Schmal, H; Czermak, B J; Lentsch, A B

    1998-01-01

    production of TNF-alpha and the CXC chemokine, macrophage inflammatory protein-2 (MIP-2). Alveolar macrophages exhibited cytokine responses to both sICAM-1 and immobilized sICAM-1, while rat PBMCs failed to demonstrate similar responses. Exposure of alveolar macrophages to sICAM-1 resulted in NFkappa......B activation (which was blocked by the presence of the aldehyde peptide inhibitor of 28S proteosome and by genistein, a tyrosine kinase inhibitor). As expected, cross-linking of CD18 on macrophages with Ab resulted in generation of TNF-alpha and MIP-2. This response was also inhibited in the presence...... of TNF-alpha and MIP-2 and increased neutrophil recruitment. Therefore, through engagement of beta2 integrins, sICAM-1 enhances alveolar macrophage production of MIP-2 and TNF-alpha, the result of which is intensified lung injury after intrapulmonary disposition of immune complexes....

  14. Engineering mechanical microenvironment of macrophage and its biomedical applications.

    Science.gov (United States)

    Li, Jing; Li, Yuhui; Gao, Bin; Qin, Chuanguang; He, Yining; Xu, Feng; Yang, Hui; Lin, Min

    2018-03-01

    Macrophages are the most plastic cells in the hematopoietic system and can be widely found in almost all tissues. Recently studies have shown that mechanical cues (e.g., matrix stiffness and stress/strain) can significantly affect macrophage behaviors. Although existing reviews on the physical and mechanical cues that regulate the macrophage's phenotype are available, engineering mechanical microenvironment of macrophages in vitro as well as a comprehensive overview and prospects for their biomedical applications (e.g., tissue engineering and immunotherapy) has yet to be summarized. Thus, this review provides an overview on the existing methods for engineering mechanical microenvironment of macrophages in vitro and then a section on their biomedical applications and further perspectives are presented.

  15. Effect of Surface Modification and Macrophage Phenotype on Particle Internalization

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Daniel [Iowa State University; Phan, Ngoc [Iowa State University; Isely, Christopher [Iowa State University; Bruene, Lucas [Iowa State University; Bratlie, Kaitlin M [Ames Laboratory

    2014-11-10

    Material properties play a key role in the cellular internalization of polymeric particles. In the present study, we have investigated the effects of material characteristics such as water contact angle, zeta potential, melting temperature, and alternative activation of complement on particle internalization for pro-inflammatory, pro-angiogenic, and naïve macrophages by using biopolymers (~600 nm), functionalized with 13 different molecules. Understanding how material parameters influence particle internalization for different macrophage phenotypes is important for targeted delivery to specific cell populations. Here, we demonstrate that material parameters affect the alternative pathway of complement activation as well as particle internalization for different macrophage phenotypes. Here, we show that the quantitative structure–activity relationship method (QSAR) previously used to predict physiochemical properties of materials can be applied to targeting different macrophage phenotypes. These findings demonstrated that targeted drug delivery to macrophages could be achieved by exploiting material parameters.

  16. Macrophages Contribute to the Spermatogonial Niche in the Adult Testis

    Directory of Open Access Journals (Sweden)

    Tony DeFalco

    2015-08-01

    Full Text Available The testis produces sperm throughout the male reproductive lifespan by balancing self-renewal and differentiation of spermatogonial stem cells (SSCs. Part of the SSC niche is thought to lie outside the seminiferous tubules of the testis; however, specific interstitial components of the niche that regulate spermatogonial divisions and differentiation remain undefined. We identified distinct populations of testicular macrophages, one of which lies on the surface of seminiferous tubules, in close apposition to areas of tubules enriched for undifferentiated spermatogonia. These macrophages express spermatogonial proliferation- and differentiation-inducing factors, such as colony-stimulating factor 1 (CSF1 and enzymes involved in retinoic acid (RA biosynthesis. We show that transient depletion of macrophages leads to a disruption in spermatogonial differentiation. These findings reveal an unexpected role for macrophages in the spermatogonial niche in the testis and raise the possibility that macrophages play previously unappreciated roles in stem/progenitor cell regulation in other tissues.

  17. Airway cellularity, lipid laden macrophages and microbiology of gastric juice and airways in children with reflux oesophagitis

    Directory of Open Access Journals (Sweden)

    Lewindon PJ

    2005-07-01

    Full Text Available Abstract Background Gastroesophageal reflux disease (GORD can cause respiratory disease in children from recurrent aspiration of gastric contents. GORD can be defined in several ways and one of the most common method is presence of reflux oesophagitis. In children with GORD and respiratory disease, airway neutrophilia has been described. However, there are no prospective studies that have examined airway cellularity in children with GORD but without respiratory disease. The aims of the study were to compare (1 BAL cellularity and lipid laden macrophage index (LLMI and, (2 microbiology of BAL and gastric juices of children with GORD (G+ to those without (G-. Methods In 150 children aged Results BAL neutrophil% in G- group (n = 63 was marginally but significantly higher than that in the G+ group (n = 77, (median of 7.5 and 5 respectively, p = 0.002. Lipid laden macrophage index (LLMI, BAL percentages of lymphocyte, eosinophil and macrophage were similar between groups. Viral studies were negative in all, bacterial cultures positive in 20.7% of BALs and in 5.3% of gastric aspirates. BAL cultures did not reflect gastric aspirate cultures in all but one child. Conclusion In children without respiratory disease, GORD defined by presence of reflux oesophagitis, is not associated with BAL cellular profile or LLMI abnormality. Abnormal microbiology of the airways, when present, is not related to reflux oesophagitis and does not reflect that of gastric juices.

  18. Extracts of Crinum latifolium inhibit the cell viability of mouse lymphoma cell line EL4 and induce activation of anti-tumour activity of macrophages in vitro.

    Science.gov (United States)

    Nguyen, Hoang-Yen T; Vo, Bach-Hue T; Nguyen, Lac-Thuy H; Bernad, Jose; Alaeddine, Mohamad; Coste, Agnes; Reybier, Karine; Pipy, Bernard; Nepveu, Françoise

    2013-08-26

    Crinum latifolium L. (CL) leaf extracts have been traditionally used in Vietnam and are now used all over the world for the treatment of prostate cancer. However, the precise cellular mechanisms of the action of CL extracts remain unclear. To examine the effects of CL samples on the anti-tumour activity of peritoneal murine macrophages. The properties of three extracts (aqueous, flavonoid, alkaloid), one fraction (alkaloid), and one pure compound (6-hydroxycrinamidine) obtained from CL, were studied (i) for redox capacities (DPPH and bleaching beta-carotene assays), (ii) on murine peritoneal macrophages (MTT assay) and on lymphoma EL4-luc2 cells (luciferine assay) for cytotoxicity, (iii) on macrophage polarization (production of ROS and gene expression by PCR), and (iv) on the tumoricidal functions of murine peritoneal macrophages (lymphoma cytotoxicity by co-culture with syngeneic macrophages). The total flavonoid extract with a high antioxidant activity (IC50=107.36 mg/L, DPPH assay) showed an inhibitory action on cancer cells. Alkaloid extracts inhibited the proliferation of lymphoma cells either by directly acting on tumour cells or by activating of the tumoricidal functions of syngeneic macrophages. The aqueous extract induced mRNA expression of tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin 6 (IL-6) indicating differentiation of macrophages into pro-inflammatory M1 polarized macrophages. The total flavonoid, alkaloid extracts and an alkaloid fraction induced the expression of the formyl peptide receptor (FPR) on the surface of the polarized macrophages that could lead to the activation of macrophages towards the M1 phenotype. Aqueous and flavonoid extracts enhanced NADPH quinine oxido-reductase 1 (NQO1) mRNA expression in polarized macrophages which could play an important role in cancer chemoprevention. All the samples studied were non-toxic to normal living cells and the pure alkaloid tested, 6-hydroxycrinamidine, was not

  19. DMPD: Differential responses of human monocytes and macrophages to IL-4 and IL-13. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 10534111 Differential responses of human monocytes and macrophages to IL-4 and IL-1...):575-8. (.png) (.svg) (.html) (.csml) Show Differential responses of human monocytes and macrophages to IL-...4 and IL-13. PubmedID 10534111 Title Differential responses of human monocytes an

  20. DMPD: Macrophage activation through CCR5- and CXCR4-mediated gp120-elicited signalingpathways. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 12960231 Macrophage activation through CCR5- and CXCR4-mediated gp120-elicited sign...82. Epub 2003 Jul 22. (.png) (.svg) (.html) (.csml) Show Macrophage activation through CCR5- and CXCR4-media...on through CCR5- and CXCR4-mediated gp120-elicited signalingpathways. Authors Lee C, Liu QH, Tomkowicz B, Yi