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Sample records for monocytes monocyte-derived macrophages

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

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    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.

  2. The TLR Expression Pattern on Monocyte-Derived Macrophages for Lipopolysaccharid Stimulation of Calves

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    GUO Yi-jie; ZHAO Guo-Qi; HUO Yong-jiu; Sachi Tana-ka; Hisashi Aso; Takahiro Yamaguchi

    2009-01-01

    In this paper, toll-like receptor expression pattern in monocytes-derived macrophages by lipopolysaccharid (LPS) stimulation was examined. Jugular venous blood samples from 4 Japanese calves were obtained and the peripheral blood mononuclear cells (PBMC) were isolated. The PBMC were cultured for 7 d so as to collect monocytes-derived macrophages in Repcell. The PBMC were stimulated by LPS for 24 h and the mRNA expression pattern of TLR and cytokines in monocytes-derived macrophages (Mod-Mφ) was analyzed. Results showed that LPS stimulation of Mod-Mφ could increase the mRNA levels of the genes of TNF-α, IL-6, and IL-8. In addition, the mRNA levels of the genes of TNF-α and IL-6 in the group of LPS stimulation were most significantly (P<0.01) higher than those in control group and the mRNA levels of TLR1, 3, 5, 8, and 10 were significantly (P<0.05) decreased after LPS stimulation. There was no difference in the mRNA expressions of TLR2, 4, 6, and 7 between the groups of the control and LPS stimulation. Besides, expression of TLR9 was not found. It suggested that monocytes-derived macrophages could respond to LPS and they might take an important role in the innate immunity. The important function of the cells might contribute to better disease treatment.

  3. HIV-1-infected monocytes and monocyte-derived macrophages are impaired in their ability to produce superoxide radicals.

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    Howell, A L; Groveman, D S; Wallace, P K; Fanger, M W

    1997-01-01

    Monocytes and monocyte-derived macrophages play a key role in immune defense against pathogenic organisms. Superoxide anion production is a key mechanism by which phagocytes kill pathogens. We sought to determine whether human immunodeficiency virus-infected monocytes and monocyte-derived macrophages are compromised in their ability to produce the superoxide anion following stimulation with phorbol myristate acetate (PMA) or after cross-linking the type I Fc receptor for IgG (Fc gamma RI). Fc gamma RI was cross-linked by the binding of monoclonal antibody 197, which reacts with an epitope of Fc gamma RI via its Fc region. Monocytes and monocyte-derived macrophages obtained from seronegative donors were infected in vitro with human immunodeficiency virus-1JR-FL and used in effector assays that measured superoxide anion production by the reduction of nitroblue tetrazolium. Reduced nitroblue tetrazolium was measured spectrophotometrically and by microscopy in which the percentage of cells containing intracellular deposits of the dye was assessed. By spectrophotometric measurement, we found that human immunodeficiency virus-infected monocytes and monocyte-derived macrophages produced less superoxide anion following either phorbol myristate acetate stimulation or Fc gamma RI cross-linking than uninfected cells from the same donor. Using microscopy we saw no difference in the percentage of infected and uninfected macrophages containing intracellular deposits of nitroblue tetrazolium suggesting that human immunodeficiency virus-infected macrophages produce less superoxide anion on a per cell basis than uninfected macrophages. Activation of human immunodeficiency virus-infected monocytes with interferon-gamma for 72 h prior to stimulation with phorbol myristate acetate or monoclonal antibody 197 increased their ability to reduce nitroblue tetrazolium. These findings suggest that impairment in the production of reactive oxygen intermediates may, in some cases, contribute to

  4. Nogo-B is associated with cytoskeletal structures in human monocyte-derived macrophages

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    Gredler Viktoria

    2011-01-01

    Full Text Available Abstract Background The reticulon Nogo-B participates in cellular and immunological processes in murine macrophages. Since leukocytes are an essential part of the immune system in health and disease, we decided to investigate the expression of Nogo-A, Nogo-B and Nogo-C in different human immune cell subpopulations. Furthermore, we analyzed the localization of Nogo-B in human monocyte-derived macrophages by indirect immunofluorescence stainings to gain further insight into its possible function. Findings We describe an association of Nogo-B with cytoskeletal structures and the base of filopodia, but not with focal or podosomal adhesion sites of monocyte-derived macrophages. Nogo-B positive structures are partially co-localized with RhoA staining and Rac1 positive membrane ruffles. Furthermore, Nogo-B is associated with the tubulin network, but not accumulated in the Golgi region. Although Nogo-B is present in the endoplasmic reticulum, it can also be translocated to large cell protrusions or the trailing end of migratory cells, where it is homogenously distributed. Conclusions Two different Nogo-B staining patterns can be distinguished in macrophages: firstly we observed ER-independent Nogo-B localization in cell protrusions and at the trailing end of migrating cells. Secondly, the localization of Nogo-B in actin/RhoA/Rac1 positive regions supports an influence on cytoskeletal organization. To our knowledge this is the first report on Nogo-B expression at the base of filopodia, thus providing further insight into the distribution of this protein.

  5. TNF and PGE2 in human monocyte-derived macrophages infected with Chlamydia trachomatis

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    E. Manor

    1993-01-01

    Full Text Available In this study levels of prostaglandin E2 (PGE2, tumour necrosis factor (TNF and interleukin-1 (IL-1 alpha in medium from monocyte derived macrophages (MdM infected with Chlamydia trachomatis (L2/434/Bu or K biovars. TNF and PGE2 were found in both cases while IL-1 alpha was not detected. Both TNF and PGE2 levels were higher in the medium of the MdM infected with K biovars. TNF reached maximum levels 24 h postinfection, and then declined, while PGE2 levels increased continuously during the infection time up to 96 h post-infection. Addition of dexamethasone inhibited production of TNF and PGE2. Inhibition of PGE2 production by indomethacin resulted in increased production of TNF, while addition of PGE2 caused partial inhibition of TNF production from infected MdM.

  6. HIV-1 Vpr induces interferon-stimulated genes in human monocyte-derived macrophages.

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    Muhammad Atif Zahoor

    Full Text Available Macrophages act as reservoirs of human immunodeficiency virus type 1 (HIV-1 and play an important role in its transmission to other cells. HIV-1 Vpr is a multi-functional protein involved in HIV-1 replication and pathogenesis; however, its exact role in HIV-1-infected human macrophages remains poorly understood. In this study, we used a microarray approach to explore the effects of HIV-1 Vpr on the transcriptional profile of human monocyte-derived macrophages (MDMs. More than 500 genes, mainly those involved in the innate immune response, the type I interferon pathway, cytokine production, and signal transduction, were differentially regulated (fold change >2.0 after infection with a recombinant adenovirus expressing HIV-1 Vpr protein. The differential expression profiles of select interferon-stimulated genes (ISGs and genes involved in the innate immune response, including STAT1, IRF7, MX1, MX2, ISG15, ISG20, IFIT1, IFIT2, IFIT3, IFI27, IFI44L, APOBEC3A, DDX58 (RIG-I, TNFSF10 (TRAIL, and RSAD2 (viperin were confirmed by real-time quantitative PCR and were consistent with the microarray data. In addition, at the post-translational level, HIV-1 Vpr induced the phosphorylation of STAT1 at tyrosine 701 in human MDMs. These results demonstrate that HIV-1 Vpr leads to the induction of ISGs and expand the current understanding of the function of Vpr and its role in HIV-1 immune pathogenesis.

  7. The transcriptome of Legionella pneumophila-infected human monocyte-derived macrophages.

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    Christopher T D Price

    Full Text Available Legionella pneumophila is an intracellular bacterial pathogen that invades and replicates within alveolar macrophages through injection of ∼ 300 effector proteins by its Dot/Icm type IV translocation apparatus. The bona fide F-box protein, AnkB, is a nutritional virulence effector that triggers macrophages to generate a surplus of amino acids, which is essential for intravacuolar proliferation. Therefore, the ankB mutant represents a novel genetic tool to determine the transcriptional response of human monocyte-derived macrophages (hMDMs to actively replicating L. pneumophila.Here, we utilized total human gene microarrays to determine the global transcriptional response of hMDMs to infection by wild type or the ankB mutant of L. pneumophila. The transcriptomes of hMDMs infected with either actively proliferating wild type or non-replicative ankB mutant bacteria were remarkably similar. The transcriptome of infected hMDMs was predominated by up-regulation of inflammatory pathways (IL-10 anti-inflammatory, interferon signaling and amphoterin signaling, anti-apoptosis, and down-regulation of protein synthesis pathways. In addition, L. pneumophila modulated diverse metabolic pathways, particularly those associated with bio-active lipid metabolism, and SLC amino acid transporters expression.Taken together, the hMDM transcriptional response to L. pneumophila is independent of intra-vacuolar replication of the bacteria and primarily involves modulation of the immune response and metabolic as well as nutritional pathways.

  8. Transcriptional analysis of diverse strains Mycobacterium avium subspecies paratuberculosis in primary bovine monocyte derived macrophages.

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    Zhu, Xiaochun; Tu, Zheng J; Coussens, Paul M; Kapur, Vivek; Janagama, Harish; Naser, Saleh; Sreevatsan, Srinand

    2008-10-01

    In this study we analyzed the macrophage-induced gene expression of three diverse genotypes of Mycobacterium avium subsp. paratuberculosis (MAP). Using selective capture of transcribed sequences (SCOTS) on three genotypically diverse MAP isolates from cattle, human, and sheep exposed to primary bovine monocyte derived macrophages for 48 h and 120 h we created and sequenced six cDNA libraries. Sequence annotations revealed that the cattle isolate up-regulated 27 and 241 genes; the human isolate up-regulated 22 and 53 genes, and the sheep isolate up-regulated 35 and 358 genes, at the two time points respectively. Thirteen to thirty-three percent of the genes identified did not have any annotated function. Despite variations in the genes identified, the patterns of expression fell into overlapping cellular functions as inferred by pathway analysis. For example, 10-12% of the genes expressed by all three strains at each time point were associated with cell-wall biosynthesis. All three strains of MAP studied up-regulated genes in pathways that combat oxidative stress, metabolic and nutritional starvation, and cell survival. Taken together, this comparative transcriptional analysis suggests that diverse MAP genotypes respond with similar modus operandi for survival in the host.

  9. Effect of size of man-made and natural mineral fibers on chemiluminescent response in human monocyte-derived macrophages.

    OpenAIRE

    2001-01-01

    Fiber size is an important factor in the tumorigenicity of various mineral fibers and asbestos fibers in animal experiments. We examined the time course of the ability to induce lucigenin-dependent chemiluminescence (CL) from human monocyte-derived macrophages exposed to Japan Fibrous Material standard reference samples (glass wool, rock wool, micro glass fiber, two types of refractory ceramic fiber, refractory mullite fiber, potassium titanium whisker, silicon carbide whisker, titanium oxide...

  10. Fate mapping reveals that microglia and recruited monocyte-derived macrophages are definitively distinguishable by phenotype in the retina.

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    O'Koren, E G; Mathew, R; Saban, D R

    2016-02-09

    The recent paradigm shift that microglia are yolk sac-derived, not hematopoietic-derived, is reshaping our knowledge about the isolated role of microglia in CNS diseases, including degenerative conditions of the retina. However, unraveling microglial-specific functions has been hindered by phenotypic overlap of microglia with monocyte-derived macrophages. The latter are differentiated from recruited monocytes in neuroinflammation, including retina. Here we demonstrate the use of fate mapping wherein microglia and monocyte-derived cells are endogenously labeled with different fluorescent reporters. Combining this method with 12-color flow cytometry, we show that these two populations are definitively distinguishable by phenotype in retina. We prove that retinal microglia have a unique CD45(lo) CD11c(lo) F4/80(lo) I-A/I-E(-) signature, conserved in the steady state and during retinal injury. The latter was observed in the widely used light-induced retinal degeneration model and corroborated in other models, including whole-body irradiation/bone-marrow transplantation. The literature contains conflicting observations about whether microglia, including in the retina, increase expression of these markers in neuroinflammation. We show that monocyte-derived macrophages have elevated expression of these surface markers, not microglia. Our resolution of such phenotypic differences may serve as a robust way to help characterize isolated roles of these cells in retinal neuroinflammation and possibly elsewhere in CNS.

  11. Establishing porcine monocyte-derived macrophage and dendritic cell systems for studying the interaction with PRRSV-1

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    Helen eSingleton

    2016-06-01

    Full Text Available Monocyte-derived macrophages (MoMØ and monocyte-derived dendritic cells (MoDC are two model systems well established in human and rodent systems that can be used to study the interaction of pathogens with host cells. Porcine reproductive and respiratory syndrome virus (PRRSV is known to infect myeloid cells, such as macrophages (MØ and dendritic cells (DC. Therefore, this study aimed to establish systems for the differentiation and characterization of MoMØ and MoDC for subsequent infection with PRRSV-1. M-CSF differentiated monocyte-derived macrophages (MoMØ were stimulated with activators for classical (M1 or alternative (M2 activation. GM-CSF and IL-4 generated monocyte-derived dendritic cells (MoDC were activated with the well established maturation cocktail containing PAMPs and cytokines. In addition, MoMØ and MoDC were treated with dexamethasone and IL-10, which are known immuno-suppressive reagents. Cells were characterized by morphology, phenotype and function and porcine MØ subsets highlighted some divergence from described human counterparts, while MoDC, appeared more similar to mouse and human DCs. The infection with PRRSV-1 strain Lena demonstrated different replication kinetics between MoMØ and MoDC and within subsets of each cell type. While MoMØ susceptibility was significantly increased by dexamethasone and IL-10 with an accompanying increase in CD163/CD169 expression, MoDC supported only a minimal replication of PRRSV These findings underline the high variability in the susceptibility of porcine myeloid cells towards PRRSV-1 infection.

  12. The impact of telmisartan on angiotensin converting enzyme 2 mRNA expression in monocyte-derived macrophages of diabetic hypertensive patients

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    李永勤

    2013-01-01

    Objective To investigate the effects of telmisartan on the expression of angiotensin converting enzyme 2(ACE2) mRNA in monocyte-derived macrophages of hypertensive patients accompanied with diabetes. Methods 62 essential hypertensive patients accompanied with

  13. The identification of markers of macrophage differentiation in PMA-stimulated THP-1 cells and monocyte-derived macrophages.

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    Marc Daigneault

    Full Text Available Differentiated macrophages are the resident tissue phagocytes and sentinel cells of the innate immune response. The phenotype of mature tissue macrophages represents the composite of environmental and differentiation-dependent imprinting. Phorbol-12-myristate-13-acetate (PMA and 1,25-dihydroxyvitamin D3 (VD(3 are stimuli commonly used to induce macrophage differentiation in monocytic cell lines but the extent of differentiation in comparison to primary tissue macrophages is unclear. We have compared the phenotype of the promonocytic THP-1 cell line after various protocols of differentiation utilising VD(3 and PMA in comparison to primary human monocytes or monocyte-derived macrophages (MDM. Both stimuli induced changes in cell morphology indicative of differentiation but neither showed differentiation comparable to MDM. In contrast, PMA treatment followed by 5 days resting in culture without PMA (PMAr increased cytoplasmic to nuclear ratio, increased mitochondrial and lysosomal numbers and altered differentiation-dependent cell surface markers in a pattern similar to MDM. Moreover, PMAr cells showed relative resistance to apoptotic stimuli and maintained levels of the differentiation-dependent anti-apoptotic protein Mcl-1 similar to MDM. PMAr cells retained a high phagocytic capacity for latex beads, and expressed a cytokine profile that resembled MDM in response to TLR ligands, in particular with marked TLR2 responses. Moreover, both MDM and PMAr retained marked plasticity to stimulus-directed polarization. These findings suggest a modified PMA differentiation protocol can enhance macrophage differentiation of THP-1 cells and identify increased numbers of mitochondria and lysosomes, resistance to apoptosis and the potency of TLR2 responses as important discriminators of the level of macrophage differentiation for transformed cells.

  14. Proteomic alteration of equine monocyte-derived macrophages infected with equine infectious anemia virus.

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    Du, Cheng; Liu, Hai-Fang; Lin, Yue-Zhi; Wang, Xue-Feng; Ma, Jian; Li, Yi-Jing; Wang, Xiaojun; Zhou, Jian-Hua

    2015-06-01

    Similar to the well-studied viruses human immunodeficiency virus (HIV)-1 and simian immunodeficiency virus (SIV), equine infectious anemia virus (EIAV) is another member of the Lentivirus genus in the family Retroviridae. Previous studies revealed that interactions between EIAV and the host resulted in viral evolution in pathogenicity and immunogenicity, as well as adaptation to the host. Proteomic analysis has been performed to examine changes in protein expression and/or modification in host cells infected with viruses and has revealed useful information for virus-host interactions. In this study, altered protein expression in equine monocyte-derived macrophages (eMDMs, the principle target cell of EIAV in vivo) infected with the EIAV pathogenic strain EIAV(DLV34) (DLV34) was examined using 2D-LC-MS/MS coupled with the iTRAQ labeling technique. The expression levels of 210 cellular proteins were identified to be significantly upregulated or downregulated by infection with DLV34. Alterations in protein expression were confirmed by examining the mRNA levels of eight selected proteins using quantitative real-time reverse-transcription PCR, and by verifying the levels of ten selected proteins using parallel reaction monitoring (PRM). Further analysis of GO and Kyoto Encyclopedia of Genes and Genomes (KEGG)-Pathway enrichment demonstrated that these differentially expressed proteins are primarily related to the biological processes of oxidative phosphorylation, protein folding, RNA splicing, and ubiquitylation. Our results can facilitate a better understanding of the host response to EIAV infection and the cellular processes required for EIAV replication and pathogenesis. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Effect of cytokines on Siglec-1 and HIV-1 entry in monocyte-derived macrophages: the importance of HIV-1 envelope V1V2 region.

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    Jobe, Ousman; Trinh, Hung V; Kim, Jiae; Alsalmi, Wadad; Tovanabutra, Sodsai; Ehrenberg, Philip K; Peachman, Kristina K; Gao, Guofen; Thomas, Rasmi; Kim, Jerome H; Michael, Nelson L; Alving, Carl R; Rao, Venigalla B; Rao, Mangala

    2016-06-01

    Monocytes and monocyte-derived macrophages express relatively low levels of CD4. Despite this, macrophages can be effectively infected with human immunodeficiency virus type 1. Macrophages have a critical role in human immunodeficiency virus type 1 transmission; however, the mechanism or mechanisms of virus infection are poorly understood. We report that growth factors, such as granulocyte macrophage colony-stimulating factor and macrophage colony-stimulating factor affect the phenotypic profile and permissiveness of macrophages to human immunodeficiency virus type 1. Human immunodeficiency virus type 1 infection of monocyte-derived macrophages derived from granulocyte macrophage and macrophage colony-stimulating factors was predominantly facilitated by the sialic acid-binding immunoglobulin-like lectin-1. The number of sialic acid-binding immunoglobulin-like lectin receptors on macrophage colony-stimulating factor-derived monocyte-derived macrophages was significantly greater than on granulocyte macrophage colony-stimulating factor-derived monocyte-derived macrophages, and correspondingly, human immunodeficiency virus type 1 infection was greater in the macrophage colony-stimulating factor-derived monocyte-derived macrophages. Single-genome analysis and quantitative reverse transcriptase-polymerase chain reaction revealed that the differences in infectivity was not due to differences in viral fitness or in viral variants with differential infectivity but was due to reduced viral entry into the granulocyte macrophage colony-stimulating factor-derived monocyte-derived macrophages. Anti-sialic acid-binding immunoglobulin-like lectin, trimeric glycoprotein 145, and scaffolded V1V2 proteins were bound to sialic acid-binding immunoglobulin-like lectin and significantly reduced human immunodeficiency virus type 1 entry and infection. Furthermore, sialic acid residues present in the V1V2 region of the envelope protein mediated human immunodeficiency virus type 1

  16. The effect of low oxygen with and without steady-state hydrogen peroxide on cytokine gene and protein expression of monocyte-derived macrophages - biomed 2011

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    Owegi, H.; Bouwens, M.; Egot-Lemaire, S.; Mueller, S.; Geib, R.W.; Waite, G.N.

    2011-01-01

    An early event during inflammation and infection is the migration of monocytes into tissues where they differentiate into macrophages. Such monocyte-derived macrophages face an unfavorable environment characterized by extremely low oxygen tension and accumulation of reactive oxygen species such as h

  17. Activated human mast cells induce LOX-1-specific scavenger receptor expression in human monocyte-derived macrophages.

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    Mervi Alanne-Kinnunen

    Full Text Available Activated mast cells in atherosclerotic lesions degranulate and release bioactive compounds capable of regulating atherogenesis. Here we examined the ability of activated human primary mast cells to regulate the expression of the major scavenger receptors in cultured human primary monocyte-derived macrophages (HMDMs.Components released by immunologically activated human primary mast cells induced a transient expression of lectin-like oxidized LDL receptor (LOX-1 mRNA in HMDMs, while the expression of two other scavenger receptors, MSR1 and CD36, remained unaffected. The LOX-1-inducing secretory components were identified as histamine, tumor necrosis factor alpha (TNF-α, and transforming growth factor beta (TGF-β1, which exhibited a synergistic effect on LOX-1 mRNA expression. Histamine induced a transient expression of LOX-1 protein. Mast cell -induced increase in LOX-1 expression was not associated with increased uptake of oxidized LDL by the macrophages.Mast cell-derived histamine, TNF-α, and TGF-β1 act in concert to induce a transient increase in LOX-1 expression in human primary monocyte-derived macrophages. The LOX-1-inducing activity potentially endows mast cells a hitherto unrecognized role in the regulation of innate immune reactions in atherogenesis.

  18. High density lipoprotein suppresses lipoprotein associated phospholipase A2 in human monocytes-derived macrophages through peroxisome proliferator-activated receptor-γ pathway

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    HAN Guan-ping; REN Jing-yi; QIN Li; SONG Jun-xian; WANG Lan; CHEN Hong

    2012-01-01

    Background Lipoprotein-associated phospholipase A2 (Lp-PLA2) is mainly secreted by macrophages,serving as a specific marker of atherosclerotic plaque and exerting pro-atherogenic effects.It is known that high-density lipoprotein (HDL) plays an important role against atherosclerosis by inhibiting pro-inflammatory factors,however,the relationship between HDL and Lp-PLA2 remains elusive.Methods In this study,reverse transcription-polymerase chain reaction (RT-PCR),Western blotting,and a platelet-activating factor (PAF) acetylhydrolase assay were performed to determine the Lp-PLA2 mRNA level,protein expression and activity in human monocyte-derived macrophages upon HDL treatment of different concentrations and durations.To investigate the underlying mechanism of HDL-induced Lp-PLA2 action,pioglitazone,a peroxisome proliferator-activated receptor-y (PPARy) ligand,was introduced to human monocyte-derived macrophages and mRNA and protein levels of Lp-PLA2,as well as its activity,were determined.Results Lp-PLA2 mRNA levels,protein expression and activity were significantly inhibited in response to HDL treatment in a dose and time dependent manner in human monocyte-derived macrophages.Pioglitazone treatment (1-10 ng/ml) upregulated the Lp-PLA2 mRNA level,protein expression and activity in human monocyte-derived macrophages,while the effects were markedly reversed by HDL.In addition,pioglitazone resulted in a significant increase in PPARY phosphorylation in human monocyte-derived macrophages,which could be inhibited by HDL.Conclusion These findings indicate that HDL suppresses the expression and activity of Lp-PLA2 in human monocyte-derived macrophages,and the underlying mechanisms may be mediated through the PPARY pathway.

  19. In vitro generation of monocyte-derived macrophages under serum-free conditions improves their tumor promoting functions.

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    Flora Rey-Giraud

    Full Text Available The tumor promoting role of M2 macrophages has been described in in vivo models and the presence of macrophages in certain tumor types has been linked to a poor clinical outcome. In light of burgeoning activities to clinically develop new therapies targeting tumor-associated macrophages (TAMs, reliable in vitro models faithfully mimicking the tumor promoting functions of TAMs are required. Generation and activation of human monocyte-derived macrophages (MDM in vitro, described as M1 or M2 macrophages attributed with tumoricidal or tumor-promoting functions, respectively, has been widely reported using mainly serum containing culture methods. In this study, we compared the properties of macrophages originating from monocytes cultured either in media containing serum together with M-CSF for M2 and GM-CSF for M1 macrophages or in serum-free media supplemented with M-CSF or GM-CSF and cytokines such as IL-4, IL-10 to induce activated M2 or LPS together with IFN-γ to generate activated M1 phenotype. We observed differences in cell morphology as well as increased surface receptor expression levels in serum-containing culture whereas similar or higher cytokine production levels were detected under serum-free culture conditions. More importantly, MDM differentiated under serum-free conditions displayed enhanced tumoricidal activity for M1 and tumor promoting property for M2 macrophages in contrast to MDM differentiated in the presence of serum. Moreover, evaluation of MDM phagocytic activity in serum free condition resulted in greater phagocytic properties of M2 compared to M1. Our data therefore confirm the tumor promoting properties of M2 macrophages in vitro and encourage the targeting of TAMs for cancer therapy.

  20. Comparative nitric oxide production by LPS-stimulated monocyte-derived macrophages from Ovis canadensis and Ovis aries.

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    Sacco, R E; Waters, W R; Rudolph, K M; Drew, M L

    2006-01-01

    Bighorn sheep are more susceptible to respiratory infection by Mannheimia haemolytica than are domestic sheep. In response to bacterial challenge, macrophages produce a number of molecules that play key roles in the inflammatory response, including highly reactive nitrogen intermediates such as nitric oxide (NO). Supernatants from monocyte-derived macrophages cultured with M. haemolytica LPS were assayed for nitric oxide activity via measurement of the NO metabolite, nitrite. In response to LPS stimulation, bighorn sheep macrophages secreted significantly higher levels of NO compared to levels for non-stimulated macrophages. In contrast, levels of NO produced by domestic sheep macrophages in response to M. haemolytica LPS did not differ from levels detected in non-stimulated cell cultures. Nitrite levels detected in supernatants of LPS-stimulated bighorn macrophage cultures treated with an inducible nitric oxide synthase (INOS) inhibitor, N(G)-monomethyl-L-arginine, were similar to that observed in non-stimulated cultures indicating a role for the iNOS pathway.

  1. HIV-1 inhibits phagocytosis and inflammatory cytokine responses of human monocyte-derived macrophages to P. falciparum infected erythrocytes.

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    Louise E Ludlow

    Full Text Available HIV-1 infection increases the risk and severity of malaria by poorly defined mechanisms. We investigated the effect of HIV-1(Ba-L infection of monocyte-derived macrophages (MDM on phagocytosis of opsonised P. falciparum infected erythrocytes (IE and subsequent proinflammatory cytokine secretion. Compared to mock-infected MDM, HIV-1 infection significantly inhibited phagocytosis of IE (median (IQR (10 (0-28 versus (34 (27-108; IE internalised/100 MDM; p = 0.001 and decreased secretion of IL-6 (1,116 (352-3,387 versus 1,552 (889-6,331; pg/mL; p = 0.0078 and IL-1β (16 (7-21 versus 33 (27-65; pg/mL; p = 0.0078. Thus inadequate phagocytosis and cytokine production may contribute to impaired control of malaria in HIV-1 infected individuals.

  2. Mycobacterium tuberculosis ESAT6 and CPF10 Induce Adenosine Deaminase 2 mRNA Expression in Monocyte-Derived Macrophages

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    Bae, Mi Jung; Ryu, Suyeon; Kim, Ha-Jeong; Cha, Seung Ick

    2017-01-01

    Background Delayed hypersensitivity plays a large role in the pathogenesis of tuberculous pleural effusion (TPE). Macrophages infected with live Mycobacterium tuberculosis (MTB) increase the levels of adenosine deaminase2 (ADA2) in the pleural fluid of TPE patients. However, it is as yet unclear whether ADA2 can be produced by macrophages when challenged with MTB antigens alone. This study therefore evaluated the levels of ADA2 mRNA expression, using monocyte-derived macrophages (MDMs) stimulated with MTB antigens. Methods Purified monocytes from the peripheral blood mononuclear cells of healthy volunteers were differentiated into macrophages using granulocyte-macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF). The MDMs were stimulated with early secretory antigenic target protein 6 (ESAT6) and culture filtrate protein 10 (CFP10). The mRNA expression levels for the cat eye syndrome chromosome region, candidate 1 (CECR1) gene encoding ADA2 were then measured. Results CECR1 mRNA expression levels were significantly higher in MDMs stimulated with ESAT6 and CFP10, than in the unstimulated MDMs. When stimulated with ESAT6, M-CSF-treated MDMs showed more pronounced CECR1 mRNA expression than GM-CSF-treated MDMs. Interferon-γ decreased the ESAT6- and CFP10-induced CECR1 mRNA expression in MDMs. CECR1 mRNA expression levels were positively correlated with mRNA expression of tumor necrosis factor α and interleukin 10, respectively. Conclusion ADA2 mRNA expression increased when MDMs were stimulated with MTB antigens alone. This partly indicates that pleural fluid ADA levels could increase in patients with culture-negative TPE. Our results may be helpful in improving the understanding of TPE pathogenesis.

  3. Characterization of HIV-1 Infection and Innate Sensing in Different Types of Primary Human Monocyte-Derived Macrophages

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    Elisabeth A. Diget

    2013-01-01

    Full Text Available Macrophages play an important role in human immunodeficiency virus (HIV pathogenesis and contribute to establishment of a viral reservoir responsible for continuous virus production and virus transmission to T cells. In this study, we investigated the differences between various monocyte-derived macrophages (MDMs generated through different differentiation protocols and evaluated different cellular, immunological, and virological properties. We found that elevated and persistent HIV-1 pWT/BaL replication could be obtained only in MDMs grown in RPMI containing macrophage colony-stimulating factor (M-CSF. Interestingly, this MDM type was also most responsive to toll-like receptor stimulation. By contrast, all MDM types were activated to a comparable extent by intracellular DNA, and the macrophage serum-free medium-(Mac-SFM-differentiated MDMs responded strongly to membrane fusion through expression of CXCL10. Finally, we found that HIV infection of RPMI/M-CSF-differentiated MDMs induced low-grade expression of two interferon-stimulated genes in some donors. In conclusion, our study demonstrates that the differentiation protocol used greatly influences the ability of MDMs to activate innate immune reactions and support HIV-1 replication. Paradoxically, the data show that the MDMs with the strongest innate immune response were also the most permissive for HIV-1 replication.

  4. Helminth-induced Ly6Chi monocyte-derived alternatively activated macrophages suppress experimental autoimmune encephalomyelitis

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    Terrazas, Cesar; de Dios Ruiz-Rosado, Juan; Amici, Stephanie A.; Jablonski, Kyle A.; Martinez-Saucedo, Diana; Webb, Lindsay M.; Cortado, Hanna; Robledo-Avila, Frank; Oghumu, Steve; Satoskar, Abhay R.; Rodriguez-Sosa, Miriam; Terrazas, Luis I.; Guerau-de-Arellano, Mireia; Partida-Sánchez, Santiago

    2017-01-01

    Helminths cause chronic infections and affect the immune response to unrelated inflammatory diseases. Although helminths have been used therapeutically to ameliorate inflammatory conditions, their anti-inflammatory properties are poorly understood. Alternatively activated macrophages (AAMϕs) have been suggested as the anti-inflammatory effector cells during helminth infections. Here, we define the origin of AAMϕs during infection with Taenia crassiceps, and their disease-modulating activity on the Experimental Autoimmune Encephalomyelitis (EAE). Our data show two distinct populations of AAMϕs, based on the expression of PD-L1 and PD-L2 molecules, resulting upon T. crassiceps infection. Adoptive transfer of Ly6C+ monocytes gave rise to PD-L1+/PD-L2+, but not PD-L1+/PD-L2− cells in T. crassiceps-infected mice, demonstrating that the PD-L1+/PD-L2+ subpopulation of AAMϕs originates from blood monocytes. Furthermore, adoptive transfer of PD-L1+/PD-L2+ AAMϕs into EAE induced mice reduced disease incidence, delayed disease onset, and diminished the clinical disability, indicating the critical role of these cells in the regulation of autoimmune disorders. PMID:28094319

  5. Inhibition of HIV-1 replication in human monocyte-derived macrophages by parasite Trypanosoma cruzi.

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    Guadalupe Andreani

    Full Text Available BACKGROUND: Cells of monocyte/macrophage lineage are one of the major targets of HIV-1 infection and serve as reservoirs for viral persistence in vivo. These cells are also the target of the protozoa Trypanosoma cruzi, the causative agent of Chagas disease, being one of the most important endemic protozoonoses in Latin America. It has been demonstrated in vitro that co-infection with other pathogens can modulate HIV replication. However, no studies at cellular level have suggested an interaction between T. cruzi and HIV-1 to date. METHODOLOGY/PRINCIPAL FINDINGS: By using a fully replicative wild-type virus, our study showed that T. cruzi inhibits HIV-1 antigen production by nearly 100% (p99% being stronger than HIV-T. cruzi (approximately 90% for BaL and approximately 85% for VSV-G infection. In MDM with established HIV-1 infection, T. cruzi significantly inhibited luciferate activity (p<0.01. By quantifying R-U5 and U5-gag transcripts by real time PCR, our study showed the expression of both transcripts significantly diminished in the presence of trypomastigotes (p<0.05. Thus, T. cruzi inhibits viral post-integration steps, early post-entry steps and entry into MDM. Trypomastigotes also caused a approximately 60-70% decrease of surface CCR5 expression on MDM. Multiplication of T. cruzi inside the MDM does not seem to be required for inhibiting HIV-1 replication since soluble factors secreted by trypomastigotes have shown similar effects. Moreover, the major parasite antigen cruzipain, which is secreted by the trypomastigote form, was able to inhibit viral production in MDM over 90% (p<0.01. CONCLUSIONS/SIGNIFICANCE: Our study showed that T. cruzi inhibits HIV-1 replication at several replication stages in macrophages, a major cell target for both pathogens.

  6. Analysis of the bovine monocyte-derived macrophage response to Mycobacterium avium subspecies paratuberculosis infection using RNA-seq

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    Maura E Casey

    2015-02-01

    Full Text Available Johne’s disease, caused by infection with Mycobacterium avium subsp. paratuberculosis, (MAP, is a chronic intestinal disease of ruminants with serious economic consequences for cattle production in the United States and elsewhere. During infection, MAP bacilli are phagocytosed and subvert host macrophage processes, resulting in subclinical infections that can lead to immunopathology and dissemination of disease. Analysis of the host macrophage transcriptome during infection can therefore shed light on the molecular mechanisms and host-pathogen interplay associated with Johne’s disease. Here we describe results of an in vitro study of the bovine monocyte-derived macrophage (MDM transcriptome response during MAP infection using RNA-seq. MDM were obtained from seven age- and sex-matched Holstein-Friesian cattle and were infected with MAP across a six-hour infection time course with non-infected controls. We observed 245 and 574 differentially expressed genes in MAP-infected versus non-infected control samples (adjusted P value ≤ 0.05 at 2 and 6 hours post-infection, respectively. Functional analyses of these differentially expressed genes, including biological pathway enrichment, highlighted potential functional roles for genes that have not been previously described in the host response to infection with MAP bacilli. In addition, differential expression of pro- and anti-inflammatory cytokine genes, such as those associated with the IL-10 signaling pathway, and other immune-related genes that encode proteins involved in the bovine macrophage response to MAP infection emphasize the balance between protective host immunity and bacilli survival and proliferation. Systematic comparisons of RNA-seq gene expression results with Affymetrix® microarray data generated from the same experimental samples also demonstrated that RNA-seq represents a superior technology for studying host transcriptional responses to intracellular infection.

  7. Monocyte-Derived Suppressor Cells in Transplantation.

    Science.gov (United States)

    Ochando, Jordi; Conde, Patricia; Bronte, Vincenzo

    Myeloid-derived suppressor cells (MDSC) are cells of myeloid origin with enhanced suppressive function. They are negative regulators of the immune responses and comprise a heterogeneous mixture of immunosuppressive cells of monocytic (M-MDSC) and granulocytic (G-MDSC) origin. A more recent nomenclature proposes the term "suppressive monocyte derived cells" (suppressive MCs) to define CSF1/CSF2-dependent mouse suppressor cells that develop from common monocyte progenitors (cMoPs) after birth. Here, we review the literature about monocytic-derived cells with demonstrated suppressor function in vitro and in vivo within the context of solid organ transplantation.

  8. Screening of Mycobacterium avium subsp. paratuberculosis Mutants for Attenuation in a Bovine Monocyte-Derived Macrophage Model

    Directory of Open Access Journals (Sweden)

    Elise A Lamont

    2014-06-01

    Full Text Available Vaccination remains a major tool for prevention and progression of Johne’s disease, a chronic enteritis of ruminants worldwide. Currently there is only one licensed vaccine within the United States and two vaccines licensed internationally against Johne’s disease. All licensed vaccines reduce fecal shedding of Mycobacterium avium subsp. paratuberculosis (MAP and delay disease progression. However, there are no available vaccines that prevent disease onset. A joint effort by the Johne’s Disease Integrated Program (JDIP, a USDA-funded consortium, and USDA- APHIS/VS sought to identify transposon insertion mutant strains as vaccine candidates in part of a three phase study. The focus of the Phase I study was to evaluate MAP mutant attenuation in a well-defined in vitro bovine monocyte-derived macrophage (MDM model. Attenuation was determined by colony forming unit (CFUs counts and slope estimates. Based on CFU counts alone, the MDM model did not identify any mutant that significantly differed from the wild-type control, MAP K-10. Slope estimates using mixed models approach identified six mutants as being attenuated. These were enrolled in protection studies involving murine and baby goat vaccination-challenge models. MDM based approach identified trends in attenuation but this did not correlate with protection in a natural host model. These results suggest the need for alternative strategies for Johne’s disease vaccine candidate screening and evaluation.

  9. ALV-J strain SCAU-HN06 induces innate immune responses in chicken primary monocyte-derived macrophages.

    Science.gov (United States)

    Feng, Min; Dai, Manman; Cao, Weisheng; Tan, Yan; Li, Zhenhui; Shi, Meiqing; Zhang, Xiquan

    2017-01-01

    Avian leucosis virus subgroup J (ALV-J) can cause lifelong infection and can escape from the host immune defenses in chickens. Since macrophages act as the important defense line against invading pathogens in host innate immunity, we investigated the function and innate immune responses of chicken primary monocyte-derived macrophages (MDM) after ALV-J infection in this study. Our results indicated that ALV-J was stably maintained in MDM cells but that the viral growth rate was significantly lower than that in DF-1 cells. We also found that ALV-J infection significantly increased nitric oxide (NO) production, but had no effect on MDM phagocytic capacity. Interestingly, infection with ALV-J rapidly promoted the expression levels of Myxovirus resistance 1 (Mx) (3 h, 6 h), ISG12 (6 h), and interleukin-1β (IL-1β) (3 h, 12 h) at an early infection stage, whereas it sharply decreased the expression of Mx (24 h, 36 h), ISG12 (36 h), and made little change on IL-1β (24 h, 36 h) production at a late infection stage in MDM cells. Moreover, the protein levels of interferon-β (IFN-β) and interleukin-6 (IL-6) had sharply increased in infected MDM cells from 3 to 36 h post infection (hpi) of ALV-J. And, the protein level of interleukin-10 (IL-10) was dramatically decreased at 36 hpi in MDM cells infected with ALV-J. These results demonstrate that ALV-J can induce host innate immune responses and we hypothesize that macrophages play an important role in host innate immune attack and ALV-J immune escape.

  10. Evaluation of the cytotoxicity of organic dust components on THP1 monocytes-derived macrophages using high content analysis.

    Science.gov (United States)

    Ramery, Eve; O'Brien, Peter J

    2014-03-01

    Organic dust contains pathogen-associated molecular patterns (PAMPs) which can induce significant airway diseases following chronic exposure. Mononuclear phagocytes are key protecting cells of the respiratory tract. Several studies have investigated the effects of PAMPs and mainly endotoxins, on cytokine production. However the sublethal cytotoxicity of organic dust components on macrophages has not been tested yet. The novel technology of high content analysis (HCA) is already used to assess subclinical drug-induced toxicity. It combines the capabilities of flow cytometry, intracellular fluorescence probes, and image analysis and enables rapid multiple analyses in large numbers of samples. In this study, HCA was used to investigate the cytotoxicity of the three major PAMPs contained in organic dust, i.e., endotoxin (LPS), peptidoglycan (PGN) and β-glucans (zymosan) on THP-1 monocyte-derived macrophages. LPS was used at concentrations of 0.005, 0.01, 0.02, 0.05, 0.1, and 1 μg/mL; PGN and zymosan were used at concentrations of 1, 5, 10, 50, 100, and 500 μg/mL. Cells were exposed to PAMPs for 24 h. In addition, the oxidative burst and the phagocytic capabilities of the cells were tested. An overlap between PGN intrinsic fluorescence and red/far-red fluorescent dyes occurred, rendering the evaluation of some parameters impossible for PGN. LPS induced sublethal cytotoxicity at the lowest dose (from 50 ng/mL). However, the greatest cytotoxic changes occurred with zymosan. In addition, zymosan, but not LPS, induced phagosome maturation and oxidative burst. Given the fact that β-glucans can be up to 100-fold more concentrated in organic dust than LPS, these results suggest that β-glucans could play a major role in macrophage impairment following heavy dust exposure and will merit further investigation in the near future.

  11. Integrated microRNA-mRNA-analysis of human monocyte derived macrophages upon Mycobacterium avium subsp. hominissuis infection.

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    Jutta Sharbati

    Full Text Available BACKGROUND: Many efforts have been made to understand basal mechanisms of mycobacterial infections. Macrophages are the first line of host immune defence to encounter and eradicate mycobacteria. Pathogenic species have evolved different mechanisms to evade host response, e.g. by influencing macrophage apoptotic pathways. However, the underlying molecular regulation is not fully understood. A new layer of eukaryotic regulation of gene expression is constituted by microRNAs. Therefore, we present a comprehensive study for identification of these key regulators and their targets in the context of host macrophage response to mycobacterial infections. METHODOLOGY/PRINCIPAL FINDINGS: We performed microRNA as well as mRNA expression analysis of human monocyte derived macrophages infected with several Mycobacterium avium hominissuis strains by means of microarrays as well as quantitative reverse transcription PCR (qRT-PCR. The data revealed the ability of all strains to inhibit apoptosis by transcriptional regulation of BCL2 family members. Accordingly, at 48 h after infection macrophages infected with all M. avium strains showed significantly decreased caspase 3 and 7 activities compared to the controls. Expression of let-7e, miR-29a and miR-886-5p were increased in response to mycobacterial infection at 48 h. The integrated analysis of microRNA and mRNA expression as well as target prediction pointed out regulative networks identifying caspase 3 and 7 as potential targets of let-7e and miR-29a, respectively. Consecutive reporter assays verified the regulation of caspase 3 and 7 by these microRNAs. CONCLUSIONS/SIGNIFICANCE: We show for the first time that mycobacterial infection of human macrophages causes a specific microRNA response. We furthermore outlined a regulatory network of potential interactions between microRNAs and mRNAs. This study provides a theoretical concept for unveiling how distinct mycobacteria could manipulate host cell response

  12. Acute stress reduces wound-induced activation of microbicidal potential of ex vivo isolated human monocyte-derived macrophages.

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    Ulrike Kuebler

    Full Text Available BACKGROUND: Psychological stress delays wound healing but the precise underlying mechanisms are unclear. Macrophages play an important role in wound healing, in particular by killing microbes. We hypothesized that (a acute psychological stress reduces wound-induced activation of microbicidal potential of human monocyte-derived macrophages (HMDM, and (b that these reductions are modulated by stress hormone release. METHODS: Fourty-one healthy men (mean age 35 ± 13 years were randomly assigned to either a stress or stress-control group. While the stress group underwent a standardized short-term psychological stress task after catheter-induced wound infliction, stress-controls did not. Catheter insertion was controlled. Assessing the microbicidal potential, we investigated PMA-activated superoxide anion production by HMDM immediately before and 1, 10 and 60 min after stress/rest. Moreover, plasma norepinephrine and epinephrine and salivary cortisol were repeatedly measured. In subsequent in vitro studies, whole blood was incubated with norepinephrine in the presence or absence of phentolamine (norepinephrine blocker before assessing HMDM microbicidal potential. RESULTS: Compared with stress-controls, HMDM of the stressed subjects displayed decreased superoxide anion-responses after stress (p's <.05. Higher plasma norepinephrine levels statistically mediated lower amounts of superoxide anion-responses (indirect effect 95% CI: 4.14-44.72. Norepinephrine-treated HMDM showed reduced superoxide anion-production (p<.001. This effect was blocked by prior incubation with phentolamine. CONCLUSIONS: Our results suggest that acute psychological stress reduces wound-induced activation of microbicidal potential of HMDM and that this reduction is mediated by norepinephrine. This might have implications for stress-induced impairment in wound healing.

  13. High resolution preparation of monocyte-derived macrophages (MDM protein fractions for clinical proteomics

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    Olivieri Oliviero

    2009-02-01

    Full Text Available Abstract Background Macrophages are involved in a number of key physiological processes and complex responses such as inflammatory, immunological, infectious diseases and iron homeostasis. These cells are specialised for iron storage and recycling from senescent erythrocytes so they play a central role in the fine tuning of iron balancing and distribution. The comprehension of the many physiological responses of macrophages implies the study of the related molecular events. To this regard, proteomic analysis, is one of the most powerful tools for the elucidation of the molecular mechanisms, in terms of changes in protein expression levels. Results Our aim was to optimize a protocol for protein fractionation and high resolution mapping using human macrophages for clinical studies. We exploited a fractionation protocol based on the neutral detergent Triton X-114. The 2D maps of the fractions obtained showed high resolution and a good level of purity. Western immunoblotting and mass spectrometry (MS/MS analysis indicated no fraction cross contamination. On 2D-PAGE mini gels (7 × 8 cm we could count more than five hundred protein spots, substantially increasing the resolution and the number of detectable proteins for the macrophage proteome. The fractions were also evaluated, with preliminary experiments, using Surface Enhanced Laser Desorption Ionization Time of Flight Mass Spectrometry (SELDI-TOF-MS. Conclusion This relatively simple method allows deep investigation into macrophages proteomics producing discrete and accurate protein fractions, especially membrane-associated and integral proteins. The adapted protocol seems highly suitable for further studies of clinical proteomics, especially for the elucidation of the molecular mechanisms controlling iron homeostasis in normal and disease conditions.

  14. Helminth-induced Ly6Chi monocyte-derived alternatively activated macrophages suppress experimental autoimmune encephalomyelitis

    OpenAIRE

    Terrazas, Cesar; de Dios Ruiz-Rosado, Juan; Stephanie A. Amici; Jablonski, Kyle A.; Martinez-Saucedo, Diana; Lindsay M Webb; Cortado, Hanna; Robledo-Avila, Frank; Oghumu, Steve; Satoskar, Abhay R.; Rodriguez-Sosa, Miriam; Terrazas, Luis I.; Guerau-de-Arellano, Mireia; Partida-Sánchez, Santiago

    2017-01-01

    Helminths cause chronic infections and affect the immune response to unrelated inflammatory diseases. Although helminths have been used therapeutically to ameliorate inflammatory conditions, their anti-inflammatory properties are poorly understood. Alternatively activated macrophages (AAMϕs) have been suggested as the anti-inflammatory effector cells during helminth infections. Here, we define the origin of AAMϕs during infection with Taenia crassiceps, and their disease-modulating activity o...

  15. Inhibition of nitric oxide enhances ovine lentivirus replication in monocyte-derived macrophages.

    Science.gov (United States)

    Keane, Kevin A; Mason, Gary L; DeMartini, James C

    2002-12-01

    Ovine lentivirus (OvLV) also known as maedi-visna virus, infects and replicates primarily in macrophages. This investigation examined the role of nitric oxide in the replication of OvLV in cultured macrophages. Peripheral blood mononuclear cells were collected from OvLV-free sheep and cultured in Teflon coated flasks at a high concentration of lamb serum. The cells were subsequently infected with OvLV strain 85/34. OvLV replication was assessed under different experimental treatments by comparison of reverse transcriptase (RT) activity in culture supernatant. Cultures that were treated with exogenous nitric oxide via S-nitroso-acetylpenicillamine did not have altered levels of RT activity compared to cultures treated with the inactive control compound, acetylpenicillamine. However, blockage of nitric oxide production by treatment with aminoguanidine, a competitive inhibitor of inducible nitric oxide synthase (iNOS), led to a significant rise in RT activity. This rise in RT activity was partially reversed in aminoguanidine treated cultures by L-arginine, the normal substrate for iNOS. Finally, the number of viral antigen producing cells was also quantified after aminoguanidine treatment and found to be significantly higher than untreated cultures. Collectively, these results indicate that nitric oxide is a negative regulator of OvLV replication in macrophages.

  16. Intestinal Monocyte-Derived Macrophages Control Commensal-Specific Th17 Responses

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    Casandra Panea

    2015-08-01

    Full Text Available Generation of different CD4 T cell responses to commensal and pathogenic bacteria is crucial for maintaining a healthy gut environment, but the associated cellular mechanisms are poorly understood. Dendritic cells (DCs and macrophages (Mfs integrate microbial signals and direct adaptive immunity. Although the role of DCs in initiating T cell responses is well appreciated, how Mfs contribute to the generation of CD4 T cell responses to intestinal microbes is unclear. Th17 cells are critical for mucosal immune protection and at steady state are induced by commensal bacteria, such as segmented filamentous bacteria (SFB. Here, we examined the roles of mucosal DCs and Mfs in Th17 induction by SFB in vivo. We show that Mfs, and not conventional CD103+ DCs, are essential for the generation of SFB-specific Th17 responses. Thus, Mfs drive mucosal T cell responses to certain commensal bacteria.

  17. Genome-wide association study identifies single nucleotide polymorphism in DYRK1A associated with replication of HIV-1 in monocyte-derived macrophages.

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    Sebastiaan M Bol

    Full Text Available BACKGROUND: HIV-1 infected macrophages play an important role in rendering resting T cells permissive for infection, in spreading HIV-1 to T cells, and in the pathogenesis of AIDS dementia. During highly active anti-retroviral treatment (HAART, macrophages keep producing virus because tissue penetration of antiretrovirals is suboptimal and the efficacy of some is reduced. Thus, to cure HIV-1 infection with antiretrovirals we will also need to efficiently inhibit viral replication in macrophages. The majority of the current drugs block the action of viral enzymes, whereas there is an abundance of yet unidentified host factors that could be targeted. We here present results from a genome-wide association study identifying novel genetic polymorphisms that affect in vitro HIV-1 replication in macrophages. METHODOLOGY/PRINCIPAL FINDINGS: Monocyte-derived macrophages from 393 blood donors were infected with HIV-1 and viral replication was determined using Gag p24 antigen levels. Genomic DNA from individuals with macrophages that had relatively low (n = 96 or high (n = 96 p24 production was used for SNP genotyping with the Illumina 610 Quad beadchip. A total of 494,656 SNPs that passed quality control were tested for association with HIV-1 replication in macrophages, using linear regression. We found a strong association between in vitro HIV-1 replication in monocyte-derived macrophages and SNP rs12483205 in DYRK1A (p = 2.16 × 10(-5. While the association was not genome-wide significant (p<1 × 10(-7, we could replicate this association using monocyte-derived macrophages from an independent group of 31 individuals (p = 0.0034. Combined analysis of the initial and replication cohort increased the strength of the association (p = 4.84 × 10(-6. In addition, we found this SNP to be associated with HIV-1 disease progression in vivo in two independent cohort studies (p = 0.035 and p = 0.0048. CONCLUSIONS/SIGNIFICANCE: These findings suggest that the kinase

  18. Efficient, long term production of monocyte-derived macrophages from human pluripotent stem cells under partly-defined and fully-defined conditions.

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    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.

  19. Monocytes-derived macrophages mediated stable expression of human brain-derived neurotrophic factor, a novel therapeutic strategy for neuroAIDS.

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    Jing Tong

    Full Text Available HIV-1 associated dementia remains a significant public health burden. Clinical and experimental research has shown that reduced levels of brain-derived neurotrophic factor (BDNF may be a risk factor for neurological complications associated with HIV-1 infection. We are actively testing genetically modified macrophages for their possible use as the cell-based gene delivery vehicle for the central nervous system (CNS. It can be an advantage to use the natural homing/migratory properties of monocyte-derived macrophages to deliver potentially neuroprotective BDNF into the CNS, as a non-invasive manner. Lentiviral-mediated gene transfer of human (hBDNF plasmid was constructed and characterized. Defective lentiviral stocks were generated by transient transfection of 293T cells with lentiviral transfer plasmid together with packaging and envelope plasmids. High titer lentiviral vector stocks were harvested and used to transduce human neuronal cell lines, primary cultures of human peripheral mononocyte-derived macrophages (hMDM and murine myeloid monocyte-derived macrophages (mMDM. These transduced cells were tested for hBDNF expression, stability, and neuroprotective activity. The GenomeLab GeXP Genetic Analysis System was used to evaluate transduced cells for any adverse effects by assessing gene profiles of 24 reference genes. High titer vectors were prepared for efficient transduction of neuronal cell lines, hMDM, and mMDM. Stable secretion of high levels of hBDNF was detected in supernatants of transduced cells using western blot and ELISA. The conditioned media containing hBDNF were shown to be protective to neuronal and monocytic cell lines from TNF-α and HIV-1 Tat mediated cytotoxicity. Lentiviral vector-mediated gene transduction of hMDM and mMDM resulted in high-level, stable expression of the neuroprotective factorBDNF in vitro. These findings form the basis for future research on the potential use of BDNF as a novel therapy for neuroAIDS.

  20. Monocytes-derived macrophages mediated stable expression of human brain-derived neurotrophic factor, a novel therapeutic strategy for neuroAIDS.

    Science.gov (United States)

    Tong, Jing; Buch, Shilpa; Yao, Honghong; Wu, Chengxiang; Tong, Hsin-I; Wang, Youwei; Lu, Yuanan

    2014-01-01

    HIV-1 associated dementia remains a significant public health burden. Clinical and experimental research has shown that reduced levels of brain-derived neurotrophic factor (BDNF) may be a risk factor for neurological complications associated with HIV-1 infection. We are actively testing genetically modified macrophages for their possible use as the cell-based gene delivery vehicle for the central nervous system (CNS). It can be an advantage to use the natural homing/migratory properties of monocyte-derived macrophages to deliver potentially neuroprotective BDNF into the CNS, as a non-invasive manner. Lentiviral-mediated gene transfer of human (h)BDNF plasmid was constructed and characterized. Defective lentiviral stocks were generated by transient transfection of 293T cells with lentiviral transfer plasmid together with packaging and envelope plasmids. High titer lentiviral vector stocks were harvested and used to transduce human neuronal cell lines, primary cultures of human peripheral mononocyte-derived macrophages (hMDM) and murine myeloid monocyte-derived macrophages (mMDM). These transduced cells were tested for hBDNF expression, stability, and neuroprotective activity. The GenomeLab GeXP Genetic Analysis System was used to evaluate transduced cells for any adverse effects by assessing gene profiles of 24 reference genes. High titer vectors were prepared for efficient transduction of neuronal cell lines, hMDM, and mMDM. Stable secretion of high levels of hBDNF was detected in supernatants of transduced cells using western blot and ELISA. The conditioned media containing hBDNF were shown to be protective to neuronal and monocytic cell lines from TNF-α and HIV-1 Tat mediated cytotoxicity. Lentiviral vector-mediated gene transduction of hMDM and mMDM resulted in high-level, stable expression of the neuroprotective factorBDNF in vitro. These findings form the basis for future research on the potential use of BDNF as a novel therapy for neuroAIDS.

  1. In vitro evidence for the protective role of Sida rhomboidea. Roxb extract against LDL oxidation and oxidized LDL-induced apoptosis in human monocyte-derived macrophages.

    Science.gov (United States)

    Thounaojam, Menaka C; Jadeja, Ravirajsinh N; Devkar, Ranjisinh V; Ramachandran, A V

    2011-06-01

    The present study was undertaken to evaluate protective role of S. rhomboidea. Roxb (SR) leaf extract against in vitro low-density lipoprotein (LDL) oxidation and oxidized LDL (Ox-LDL) induced macrophage apoptosis. Copper and cell-mediated LDL oxidation, Ox-LDL-induced peroxyl radical generation, mitochondrial activity, and apoptosis in human monocyte-derived macrophages (HMDMs) were assessed in presence of SR extract. Results clearly indicated that SR was capable of reducing LDL oxidation and formation of intermediary oxidation products. Also, SR successfully attenuated peroxyl radical formation, mitochondrial dysfunction, nuclear condensation, and apoptosis in Ox-LDL-exposed HMDMs. This scientific report is the first detailed investigation that establishes anti-atherosclerotic potential of SR extract.

  2. Surface modification of biomaterials based on high-molecular polylactic acid and their effect on inflammatory reactions of primary human monocyte-derived macrophages: perspective for personalized therapy.

    Science.gov (United States)

    Stankevich, Ksenia S; Gudima, Alexandru; Filimonov, Victor D; Klüter, Harald; Mamontova, Evgeniya M; Tverdokhlebov, Sergei I; Kzhyshkowska, Julia

    2015-06-01

    Polylactic acid (PLA) based implants can cause inflammatory complications. Macrophages are key innate immune cells that control inflammation. To provide higher biocompatibility of PLA-based implants with local innate immune cells their surface properties have to be improved. In our study surface modification technique for high-molecular PLA (MW=1,646,600g/mol) based biomaterials was originally developed and successfully applied. Optimal modification conditions were determined. Treatment of PLA films with toluene/ethanol=3/7 mixture for 10min with subsequent exposure in 0.001M brilliant green dye (BGD) solution allows to entrap approximately 10(-9)mol/cm(2) model biomolecules. The modified PLA film surface was characterized by optical microscopy, SERS, FT-IR, UV and TG/DTA/DSC analysis. Tensile strain of modified films was determined as well. The effect of PLA films modified with BGD on the inflammatory reactions of primary human monocyte-derived macrophages was investigated. We developed in vitro test-system by differentiating primary monocyte-derived macrophages on a coating material. Type 1 and type 2 inflammatory cytokines (TNFα, CCL18) secretion and histological biomarkers (CD206, stabilin-1) expression were analyzed by ELISA and confocal microscopy respectively. BGD-modified materials have improved thermal stability and good mechanical properties. However, BGD modifications induced additional donor-specific inflammatory reactions and suppressed tolerogenic phenotype of macrophages. Therefore, our test-system successfully demonstrated specific immunomodulatory effects of original and modified PLA-based biomaterials, and can be further applied for the examination of improved coatings for implants and identification of patient-specific reactions to implants.

  3. HSV-1-induced chemokine expression via IFI16-dependent and IFI16-independent pathways in human monocyte-derived macrophages

    DEFF Research Database (Denmark)

    Søby, Stine; Laursen, Rune R; Østergaard, Lars Jørgen;

    2012-01-01

    ABSTRACT: BACKGROUND: Innate recognition is essential in the antiviral response against infection by herpes simplex virus (HSV). Chemokines are important for control of HSV via recruitment of natural killer cells, T lymphocytes, and antigen-presenting cells. We previously found that early HSV-1......-mediated chemokine responses are not dependent on TLR2 and TLR9 in human macrophages. Here, we investigated the role of the recently identified innate IFN-inducible DNA receptor IFI16 during HSV-1 infection in human macrophages. METHODS: Peripheral blood mononuclear cells were purified from buffy coats...... and monocytes were differentiated to macrophages. Macrophages infected with HSV-1 were analyzed using siRNA-mediated knock-down of IFI16 by real-time PCR, ELISA, and Western blotting. RESULTS: We determined that both CXCL10 and CCL3 are induced independent of HSV-1 replication. IFI16 mediates CCL3 m...

  4. Analysis of the human monocyte-derived macrophage transcriptome and response to lipopolysaccharide provides new insights into genetic aetiology of inflammatory bowel disease.

    Science.gov (United States)

    Baillie, J Kenneth; Arner, Erik; Daub, Carsten; De Hoon, Michiel; Itoh, Masayoshi; Kawaji, Hideya; Lassmann, Timo; Carninci, Piero; Forrest, Alistair R R; Hayashizaki, Yoshihide; Faulkner, Geoffrey J; Wells, Christine A; Rehli, Michael; Pavli, Paul; Summers, Kim M; Hume, David A

    2017-03-01

    The FANTOM5 consortium utilised cap analysis of gene expression (CAGE) to provide an unprecedented insight into transcriptional regulation in human cells and tissues. In the current study, we have used CAGE-based transcriptional profiling on an extended dense time course of the response of human monocyte-derived macrophages grown in macrophage colony-stimulating factor (CSF1) to bacterial lipopolysaccharide (LPS). We propose that this system provides a model for the differentiation and adaptation of monocytes entering the intestinal lamina propria. The response to LPS is shown to be a cascade of successive waves of transient gene expression extending over at least 48 hours, with hundreds of positive and negative regulatory loops. Promoter analysis using motif activity response analysis (MARA) identified some of the transcription factors likely to be responsible for the temporal profile of transcriptional activation. Each LPS-inducible locus was associated with multiple inducible enhancers, and in each case, transient eRNA transcription at multiple sites detected by CAGE preceded the appearance of promoter-associated transcripts. LPS-inducible long non-coding RNAs were commonly associated with clusters of inducible enhancers. We used these data to re-examine the hundreds of loci associated with susceptibility to inflammatory bowel disease (IBD) in genome-wide association studies. Loci associated with IBD were strongly and specifically (relative to rheumatoid arthritis and unrelated traits) enriched for promoters that were regulated in monocyte differentiation or activation. Amongst previously-identified IBD susceptibility loci, the vast majority contained at least one promoter that was regulated in CSF1-dependent monocyte-macrophage transitions and/or in response to LPS. On this basis, we concluded that IBD loci are strongly-enriched for monocyte-specific genes, and identified at least 134 additional candidate genes associated with IBD susceptibility from reanalysis

  5. Wear particles from studded tires and granite pavement induce pro-inflammatory alterations in human monocyte-derived macrophages: a proteomic study.

    Science.gov (United States)

    Karlsson, Helen; Lindbom, John; Ghafouri, Bijar; Lindahl, Mats; Tagesson, Christer; Gustafsson, Mats; Ljungman, Anders G

    2011-01-14

    Airborne particulate matter is considered to be one of the environmental contributors to the mortality in cancer, respiratory, and cardiovascular diseases. For future preventive actions, it is of major concern to investigate the toxicity of defined groups of airborne particles and to clarify their pathways in biological tissues. To expand the knowledge beyond general inflammatory markers, this study examined the toxicoproteomic effects on human monocyte derived macrophages after exposure to wear particles generated from the interface of studded tires and a granite-containing pavement. As comparison, the effect of endotoxin was also investigated. The macrophage proteome was separated using two-dimensional gel electrophoresis. Detected proteins were quantified, and selected proteins were identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry. Among analyzed proteins, seven were significantly decreased and three were increased by exposure to wear particles as compared to unexposed control cells. Endotoxin exposure resulted in significant changes in the expression of six proteins: four decreased and two increased. For example, macrophage capping protein was significantly increased after wear particle exposure only, whereas calgizzarin and galectin-3 were increased by both wear particle and endotoxin exposure. Overall, proteins associated with inflammatory response were increased and proteins involved in cellular functions such as redox balance, anti-inflammatory response, and glycolysis were decreased. Investigating the effects of characterized wear particles on human macrophages with a toxicoproteomic approach has shown to be useful in the search for more detailed information about specific pathways and possible biological markers.

  6. Analysis of the human monocyte-derived macrophage transcriptome and response to lipopolysaccharide provides new insights into genetic aetiology of inflammatory bowel disease.

    Directory of Open Access Journals (Sweden)

    J Kenneth Baillie

    2017-03-01

    Full Text Available The FANTOM5 consortium utilised cap analysis of gene expression (CAGE to provide an unprecedented insight into transcriptional regulation in human cells and tissues. In the current study, we have used CAGE-based transcriptional profiling on an extended dense time course of the response of human monocyte-derived macrophages grown in macrophage colony-stimulating factor (CSF1 to bacterial lipopolysaccharide (LPS. We propose that this system provides a model for the differentiation and adaptation of monocytes entering the intestinal lamina propria. The response to LPS is shown to be a cascade of successive waves of transient gene expression extending over at least 48 hours, with hundreds of positive and negative regulatory loops. Promoter analysis using motif activity response analysis (MARA identified some of the transcription factors likely to be responsible for the temporal profile of transcriptional activation. Each LPS-inducible locus was associated with multiple inducible enhancers, and in each case, transient eRNA transcription at multiple sites detected by CAGE preceded the appearance of promoter-associated transcripts. LPS-inducible long non-coding RNAs were commonly associated with clusters of inducible enhancers. We used these data to re-examine the hundreds of loci associated with susceptibility to inflammatory bowel disease (IBD in genome-wide association studies. Loci associated with IBD were strongly and specifically (relative to rheumatoid arthritis and unrelated traits enriched for promoters that were regulated in monocyte differentiation or activation. Amongst previously-identified IBD susceptibility loci, the vast majority contained at least one promoter that was regulated in CSF1-dependent monocyte-macrophage transitions and/or in response to LPS. On this basis, we concluded that IBD loci are strongly-enriched for monocyte-specific genes, and identified at least 134 additional candidate genes associated with IBD susceptibility

  7. Analysis of the human monocyte-derived macrophage transcriptome and response to lipopolysaccharide provides new insights into genetic aetiology of inflammatory bowel disease

    Science.gov (United States)

    Arner, Erik; De Hoon, Michiel; Carninci, Piero; Hayashizaki, Yoshihide; Pavli, Paul; Summers, Kim M.; Hume, David A.

    2017-01-01

    The FANTOM5 consortium utilised cap analysis of gene expression (CAGE) to provide an unprecedented insight into transcriptional regulation in human cells and tissues. In the current study, we have used CAGE-based transcriptional profiling on an extended dense time course of the response of human monocyte-derived macrophages grown in macrophage colony-stimulating factor (CSF1) to bacterial lipopolysaccharide (LPS). We propose that this system provides a model for the differentiation and adaptation of monocytes entering the intestinal lamina propria. The response to LPS is shown to be a cascade of successive waves of transient gene expression extending over at least 48 hours, with hundreds of positive and negative regulatory loops. Promoter analysis using motif activity response analysis (MARA) identified some of the transcription factors likely to be responsible for the temporal profile of transcriptional activation. Each LPS-inducible locus was associated with multiple inducible enhancers, and in each case, transient eRNA transcription at multiple sites detected by CAGE preceded the appearance of promoter-associated transcripts. LPS-inducible long non-coding RNAs were commonly associated with clusters of inducible enhancers. We used these data to re-examine the hundreds of loci associated with susceptibility to inflammatory bowel disease (IBD) in genome-wide association studies. Loci associated with IBD were strongly and specifically (relative to rheumatoid arthritis and unrelated traits) enriched for promoters that were regulated in monocyte differentiation or activation. Amongst previously-identified IBD susceptibility loci, the vast majority contained at least one promoter that was regulated in CSF1-dependent monocyte-macrophage transitions and/or in response to LPS. On this basis, we concluded that IBD loci are strongly-enriched for monocyte-specific genes, and identified at least 134 additional candidate genes associated with IBD susceptibility from reanalysis

  8. Elevated ARG1 expression in primary monocytes-derived macrophages as a predictor of radiation-induced acute skin toxicities in early breast cancer patients.

    Science.gov (United States)

    Jung, Karen; Sabri, Siham; Hanson, John; Xu, Yaoxian; Wang, Ying Wayne; Lai, Raymond; Abdulkarim, Bassam S

    2015-01-01

    Radiation therapy (RT) the front-line treatment after surgery for early breast cancer patients is associated with acute skin toxicities in at least 40% of treated patients. Monocyte-derived macrophages are polarized into functionally distinct (M1 or M2) activated phenotypes at injury sites by specific systemic cytokines known to play a key role in the transition between damage and repair in irradiated tissues. The role of M1 and M2 macrophages in RT-induced acute skin toxicities remains to be defined. We investigated the potential value of M1 and M2 macrophages as predictive factors of RT-induced skin toxicities in early breast cancer patients treated with adjuvant RT after lumpectomy. Blood samples collected from patients enrolled in a prospective clinical study (n = 49) were analyzed at baseline and after the first delivered 2Gy RT dose. We designed an ex vivo culture system to differentiate patient blood monocytes into macrophages and treated them with M1 or M2-inducing cytokines before quantitative analysis of their "M1/M2" activation markers, iNOS, Arg1, and TGFß1. Statistical analysis was performed to correlate experimental data to clinical assessment of acute skin toxicity using Common Toxicity Criteria (CTC) grade for objective evaluation of skin reactions. Increased ARG1 mRNA significantly correlated with higher grades of erythema, moist desquamation, and CTC grade. Multivariate analysis revealed that increased ARG1 expression in macrophages after a single RT dose was an independent prognostic factor of erythema (p = 0 .032), moist desquamation (p = 0 .027), and CTC grade (p = 0 .056). Interestingly, multivariate analysis of ARG1 mRNA expression in macrophages stimulated with IL-4 also revealed independent prognostic value for predicting acute RT-induced toxicity factors, erythema (p = 0 .069), moist desquamation (p = 0 .037), and CTC grade (p = 0 .046). To conclude, our findings underline for the first time the biological significance of increased ARG1 m

  9. High Intracellular Concentrations of Posaconazole Do Not Impact on Functional Capacities of Human Polymorphonuclear Neutrophils and Monocyte-Derived Macrophages In Vitro.

    Science.gov (United States)

    Farowski, Fedja; Cornely, Oliver A; Hartmann, Pia

    2016-06-01

    Posaconazole is a commonly used antifungal for the prophylaxis and treatment of invasive fungal infections. We previously demonstrated that the intracellular concentration of posaconazole in peripheral blood mononuclear cells (PBMCs) and polymorphonuclear neutrophils (PMNs) was greatly increased compared to the plasma concentration. As these professional phagocytes are crucial to combat fungal infections, we set out to investigate if and how, beneficial or deleterious, this high loading of intracellular posaconazole impacts the functional capacities of these cells. Here, we show that high intracellular concentrations of posaconazole do not significantly impact PMN and monocyte-derived macrophage function in vitro In particular, killing capacity and cytoskeletal features of PMN, such as migration, are not affected, indicating that these cells serve as vehicles for posaconazole to the site of infection. Moreover, since posaconazole as such slowed the germination of Aspergillus fumigatus conidia, infected neutrophils released less reactive oxygen species (ROS). Based on these findings, we propose that the delivery of posaconazole by neutrophils to the site of Aspergillus species infection warrants control of the pathogen and preservation of tissue integrity at the same time.

  10. Pathogenic prion protein fragment (PrP106–126) promotes human immunodeficiency virus type-1 infection in peripheral blood monocyte-derived macrophages

    Science.gov (United States)

    Bacot, Silvia M.; Feldman, Gerald M.; Yamada, Kenneth M.; Dhawan, Subhash

    2017-01-01

    Transfusion of blood and blood products contaminated with the pathogenic form of prion protein Prpsc, thought to be the causative agent of variant a Creutzfeldt–Jakob disease (vCJD), may result in serious consequences in recipients with a compromised immune system, for example, as seen in HIV-1 infection. In the present study, we demonstrate that treatment of peripheral blood monocyte-derived macrophages (MDM) with PrP106–126, a synthetic domain of PrPsc that has intrinsic functional activities related to the full-length protein, markedly increased their susceptibility to HIV-1 infection, induced cytokine secretion, and enhanced their migratory behavior in response to N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP). Live-cell imaging of MDM cultured in the presence of PrP106–126 showed large cell clusters indicative of cellular activation. Tyrosine kinase inhibitor STI-571, protein kinase C inhibitor K252B, and cyclin-dependent kinase inhibitor olomoucine attenuated PrP106–126-induced altered MDM functions. These findings delineate a previously undefined functional role of PrP106–126-mediated host cell response in promoting HIV-1 pathogenesis. PMID:25589240

  11. Induction of cyclooxygenase-2 expression during HIV-1-infected monocyte-derived macrophage and human brain microvascular endothelial cell interactions

    NARCIS (Netherlands)

    Pereira, CF; Boven, LA; Middel, J; Verhoef, J; Nottet, HSLM

    2000-01-01

    Human immunodeficiency virus type-1 (HIV-1)-associated dementia (HAD) is a neurodegenerative disease characterized by HIV infection and replication in brain tissue. HIV-1-infected monocytes overexpress inflammatory molecules that facilitate their entry into the brain. Prostanoids are lipid mediators

  12. Killing of Escherichia coli by Crohn's Disease Monocyte-derived Macrophages and Its Enhancement by Hydroxychloroquine and Vitamin D

    OpenAIRE

    Flanagan, Paul K.; Chiewchengchol, Direkrit; Helen L Wright; Edwards, Steven W.; Alswied, Abdullah; Satsangi, Jack; Subramanian, Sreedhar; Rhodes, Jonathan M.; Campbell, Barry J.

    2015-01-01

    BACKGROUND: Crohn's disease (CD) is associated with defective innate immunity, including impaired neutrophil chemotaxis, and mucosal invasion by bacteria, particularly adherent and invasive Escherichia coli that replicate inside macrophage phagolysosomes. We compared CD and healthy control (HC) macrophages for their abilities to kill E. coli and generate neutrophil chemoattractants and also assessed the effects of hydroxychloroquine (HCQ) and vitamin D on killing of phagocytosed E. coli.METHO...

  13. Infection of equine monocyte-derived macrophages with an attenuated equine infectious anemia virus (EIAV) strain induces a strong resistance to the infection by a virulent EIAV strain.

    Science.gov (United States)

    Ma, Jian; Wang, Shan-Shan; Lin, Yue-Zhi; Liu, Hai-Fang; Liu, Qiang; Wei, Hua-Mian; Wang, Xue-Feng; Wang, Yu-Hong; Du, Cheng; Kong, Xian-Gang; Zhou, Jian-Hua; Wang, Xiaojun

    2014-08-09

    The Chinese attenuated equine infectious anemia virus (EIAV) vaccine has successfully protected millions of equine animals from EIA disease in China. Given that the induction of immune protection results from the interactions between viruses and hosts, a better understanding of the characteristics of vaccine strain infection and host responses would be useful for elucidating the mechanism of the induction of immune protection by the Chinese attenuated EIAV strain. In this study, we demonstrate in equine monocyte-derived macrophages (eMDM) that EIAVFDDV13, a Chinese attenuated EIAV strain, induced a strong resistance to subsequent infection by a pathogenic strain, EIAVUK3. Further experiments indicate that the expression of the soluble EIAV receptor sELR1, Toll-like receptor 3 (TLR3) and interferon β (IFNβ) was up-regulated in eMDM infected with EIAVFDDV13 compared with eMDM infected with EIAVUK3. Stimulating eMDM with poly I:C resulted in similar resistance to EIAV infection as induced by EIAVFDDV13 and was correlated with enhanced TLR3, sELR1 and IFNβ expression. The knock down of TLR3 mRNA significantly impaired poly I:C-stimulated resistance to EIAV, greatly reducing the expression of sELR1 and IFNβ and lowered the level of infection resistance induced by EIAVFDDV13. These results indicate that the induction of restraining infection by EIAVFDDV13 in macrophages is partially mediated through the up-regulated expression of the soluble viral receptor and IFNβ, and that the TLR3 pathway activation plays an important role in the development of an EIAV-resistant intracellular environment.

  14. Global gene expression and systems biology analysis of bovine monocyte-derived macrophages in response to in vitro challenge with Mycobacterium bovis.

    Directory of Open Access Journals (Sweden)

    David A Magee

    Full Text Available BACKGROUND: Mycobacterium bovis, the causative agent of bovine tuberculosis, is a major cause of mortality in global cattle populations. Macrophages are among the first cell types to encounter M. bovis following exposure and the response elicited by these cells is pivotal in determining the outcome of infection. Here, a functional genomics approach was undertaken to investigate global gene expression profiles in bovine monocyte-derived macrophages (MDM purified from seven age-matched non-related females, in response to in vitro challenge with M. bovis (multiplicity of infection 2:1. Total cellular RNA was extracted from non-challenged control and M. bovis-challenged MDM for all animals at intervals of 2 hours, 6 hours and 24 hours post-challenge and prepared for global gene expression analysis using the Affymetrix® GeneChip® Bovine Genome Array. RESULTS: Comparison of M. bovis-challenged MDM gene expression profiles with those from the non-challenged MDM controls at each time point identified 3,064 differentially expressed genes 2 hours post-challenge, with 4,451 and 5,267 differentially expressed genes detected at the 6 hour and 24 hour time points, respectively (adjusted P-value threshold ≤ 0.05. Notably, the number of downregulated genes exceeded the number of upregulated genes in the M. bovis-challenged MDM across all time points; however, the fold-change in expression for the upregulated genes was markedly higher than that for the downregulated genes. Systems analysis revealed enrichment for genes involved in: (1 the inflammatory response; (2 cell signalling pathways, including Toll-like receptors and intracellular pathogen recognition receptors; and (3 apoptosis. CONCLUSIONS: The increased number of downregulated genes is consistent with previous studies showing that M. bovis infection is associated with the repression of host gene expression. The results also support roles for MyD88-independent signalling and intracellular PRRs in

  15. Differential expression of HIV-1 interfering factors in monocyte-derived macrophages stimulated with polarizing cytokines or interferons

    Science.gov (United States)

    Jiménez, Viviana Cobos; Booiman, Thijs; de Taeye, Steven W.; van Dort, Karel A.; Rits, Maarten A. N.; Hamann, Jörg; Kootstra, Neeltje A.

    2012-10-01

    HIV-1 replication in macrophages can be regulated by cytokines and infection is restricted in macrophages activated by type I interferons and polarizing cytokines. Here, we observed that the expression levels of the cellular factors Trim5α, CypA, APOBEC3G, SAMHD-1, Trim22, tetherin and TREX-1, and the anti-HIV miRNAs miR-28, miR-150, miR-223 and miR-382 was upregulated by IFN-α and IFN-β in macrophages, which may account for the inhibiting effect on viral replication and the antiviral state of these cells. Expression of these factors was also increased by IFN-γ +/- TNF-α, albeit to a lesser extent; yet, HIV-1 replication in these cells was not restricted at the level of proviral synthesis, indicating that these cellular factors only partially contribute to the observed restriction. IL-4, IL-10 or IL-32 polarization did not affect the expression of cellular factors and miRNAs, suggesting only a limited role for these cellular factors in restricting HIV-1 replication in macrophages.

  16. Transcriptional Response of Bovine Monocyte-Derived Macrophages after the Infection with Different Argentinean Mycobacterium bovis Isolates

    Directory of Open Access Journals (Sweden)

    Karina Caimi

    2013-01-01

    Full Text Available Infection of bovines with Mycobacterium bovis causes important financial hardship in many countries presenting also a risk for humans. M. bovis is known to be adapted to survive and thrive within the intramacrophage environment. In spite of its relevance, at present the information about macrophage expression patterns is scarce, particularly regarding the bovine host. In this study, transcriptomic analysis was used to detect genes differentially expressed in macrophages derived from peripheral blood mononuclear cells at early stages of infection with two Argentinean strains of M. bovis, a virulent and an attenuated strains. The results showed that the number of differentially expressed genes in the cells infected with the virulent strain (5 was significantly lower than those in the cells infected with the attenuated strain (172. Several genes were more strongly expressed in infected macrophages. Among them, we detected encoding transcription factors, anthrax toxin receptor, cell division and apoptosis regulator, ankyrin proteins, cytoskeleton proteins, protein of cell differentiation, and regulators of endocytic traffic of membrane. Quantitative real-time PCR of a selected group of differentially expressed genes confirmed the microarrays results. Altogether, the present results contribute to understanding the mechanisms involved in the early interaction of M. bovis with the bovine macrophage.

  17. Enhanced Inhibitory Effect of Ultra-Fine Granules of Red Ginseng on LPS-induced Cytokine Expression in the Monocyte-Derived Macrophage THP-1 Cells

    Directory of Open Access Journals (Sweden)

    Hong-Yeoul Kim

    2008-08-01

    Full Text Available Red ginseng is one of the most popular traditional medicines in Korea because its soluble hot-water extract is known to be very effective on enhancing immunity as well as inhibiting inflammation. Recently, we developed a new technique, called the HACgearshift system, which can pulverize red ginseng into the ultra-fine granules ranging from 0.2 to 7.0 μm in size. In this study, the soluble hot-water extract of those ultra-fine granules of red ginseng (URG was investigated and compared to that of the normal-sized granules of red ginseng (RG. The high pressure liquid chromatographic analyses of the soluble hot-water extracts of both URG and RG revealed that URG had about 2-fold higher amounts of the ginsenosides, the biologically active components in red ginseng, than RG did. Using quantitative RT-PCR, cytokine profiling against the Escherichia coli lipopolysaccharide (LPS in the monocyte-derived macrophage THP-1 cells demonstrated that the URG-treated cells showed a significant reduction in cytokine expression than the RG-treated ones. Transcription expression of the LPS-induced cytokines such as TNF-α, IL-1β, IL-6, IL-8, IL-10, and TGF-β was significantly inhibited by URG compared to RG. These results suggest that some biologically active and soluble components in red ginseng can be more effectively extracted from URG than RG by standard hot-water extraction.

  18. Differential intracellular fate of Burkholderia pseudomallei 844 and Burkholderia thailandensis UE5 in human monocyte-derived dendritic cells and macrophages

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    Engering Anneke

    2009-04-01

    Full Text Available Abstract Background Burkholderia pseudomallei (Bp is a category B biothreat organism that causes a potentially fatal disease in humans and animals, namely melioidosis. Burkholderia thailandensis (Bt is another naturally occurring species that is very closely related to Bp. However, despite this closely related genotype, Bt is considered avirulent as it does not cause the disease. In the present study, we compared the growth kinetics of B. pseudomallei strain 844 (Bp-844 in human monocyte-derived dendritic cells (MoDCs and macrophages (Mφs, as well as its ability to stimulate host cell responses with those of B. thailandensis strain UE5 (Bt-UE5. Results Primary human MoDCs and Mφs were infected with Bp-844 and its intracellular growth kinetics and ability to induce host cell responses were evaluated. The results were compared with those obtained using the Bt-UE5. In human MoDCs, both bacteria were similar in respect to their ability to survive and replicate intracellularly, induce upregulation of costimulatory molecules and cytokines and bias T helper cell differentiation toward a Th1 phenotype. By contrast, the two bacteria exhibited different growth kinetics in human Mφs, where the intracellular growth of Bt-UE5, but not Bp-844, was significantly suppressed. Moreover, the ability of Mφs to kill Bp-844 was markedly enhanced following stimulation with IFN-γ. Conclusion The data presented showed that while both strains were similar in their ability to survive and replicate in human MoDCs, only Bp-844 could readily replicate in human Mφs. Both bacteria induced similar host cellular responses, particularly with regard to their ability to bias T cell differentiation toward a Th1 phenotype.

  19. Monocyte-derived dendritic cells in bipolar disorder

    NARCIS (Netherlands)

    Knijff, EM; Ruwhof, C; de Wit, HJ; Kupka, RW; Vonk, R; Akkerhuis, GW; Nolen, WA; Drexhage, HA

    2006-01-01

    Background: Dendritic cells (DC) are key regulators of the immune system, which is compromised in patients with bipolar disorder. We sought to study monocyte-derived DC in bipolar disorder. Methods: Monocytes purified from blood collected from DSM-IV bipolar disorder outpatients (n = 53, 12 without

  20. Sphingosylphosphorylcholine stimulates human monocyte-derived dendritic cell chemotaxis

    Institute of Scientific and Technical Information of China (English)

    Ha-young LEE; Eun-ha SHIN; Yoe-sik BAE

    2006-01-01

    Aim: To investigate the effects of Sphingosylphosphorylcholine (SPC) on human monocyte-derived dendritic cell (DC) chemotaxis. Methods: Human DC were generated from peripheral blood monocytes by culturing them with granulocyte macrophage-colony stimulating factor and interleukin-4. The effect of SPC on the DC chemotactic migration was measured by chemotaxis assay. Intracellular signaling event involved in the SPC-induced DC chemotaxis was investigated with several inhibitors for specific kinase. The expression of the SPC receptors was examined by reverse transcription polymerase chain reaction. Results: We found that SPC induced chemotactic migration in immature DC (iDC) and mature DC (mDC). In terms of SPC-induced signaling events, mitogen activated protein kinase activation and Akt activation in iDC and mDC were stimulated. SPC-induced chemotaxis was mediated by extracellular signal-regulated protein kinase and phosphoino-sitide-3-kinase, but not by calcium in both iDC and mDC. Although mDC express ovarian cancer G protein-coupled receptor 1, but not G protein-coupled receptor 4, iDC do not express any of these receptors. To examine the involvement of sphin-gosine-1-phosphate (SIP) receptors, we checked the effect of an SIP receptor antagonist (VPC23019) on SPC-induced DC chemotaxis. VPC23019 did not affect SPC-induced DC chemotaxis. Conclusion: The results suggest that SPC may play a role in regulating DC trafficking during phagocytosis and the T cell-stimulating phase, and the unique SPC receptor, which is different from SIP receptors, is involved in SPC-induced chemotaxis.

  1. Acanthamoeba castellanii Genotype T4 Stimulates the Production of Interleukin-10 as Well as Proinflammatory Cytokines in THP-1 Cells, Human Peripheral Blood Mononuclear Cells, and Human Monocyte-Derived Macrophages.

    Science.gov (United States)

    Mattana, Antonella; Sanna, Manuela; Cano, Antonella; Delogu, Giuseppe; Erre, Giuseppe; Roberts, Craig W; Henriquez, Fiona L; Fiori, Pier Luigi; Cappuccinelli, Piero

    2016-10-01

    Free-living amoebae of the genus Acanthamoeba can cause severe and chronic infections in humans, mainly localized in immune privileged sites, such as the brain and the eye. Monocytes/macrophages are thought to be involved in Acanthamoeba infections, but little is known about how these facultative parasites influence their functions. The aim of this work was to investigate the effects of Acanthamoeba on human monocytes/macrophages during the early phase of infection. Here, THP-1 cells, primary human monocytes isolated from peripheral blood, and human monocyte-derived macrophages were either coincubated with trophozoites of a clinical isolate of Acanthamoeba (genotype T4) or stimulated with amoeba-derived cell-free conditioned medium. Production of proinflammatory cytokines (tumor necrosis factor alpha [TNF-α], interleukin-6 [IL-6], and IL-12), anti-inflammatory cytokine (IL-10), and chemokine (IL-8) was evaluated at specific hours poststimulation (ranging from 1.5 h to 23 h). We showed that both Acanthamoeba trophozoites and soluble amoebic products induce an early anti-inflammatory monocyte-macrophage phenotype, characterized by significant production of IL-10; furthermore, challenge with either trophozoites or their soluble metabolites stimulate both proinflammatory cytokines and chemokine production, suggesting that this protozoan infection results from the early induction of coexisting, opposed immune responses. Results reported in this paper confirm that the production of proinflammatory cytokines and chemokines by monocytes and macrophages can play a role in the development of the inflammatory response during Acanthamoeba infections. Furthermore, we demonstrate for the first time that Acanthamoeba stimulates IL-10 production in human innate immune cells, which might both promote the immune evasion of Acanthamoeba and limit the induced inflammatory response.

  2. Acanthamoeba castellanii Genotype T4 Stimulates the Production of Interleukin-10 as Well as Proinflammatory Cytokines in THP-1 Cells, Human Peripheral Blood Mononuclear Cells, and Human Monocyte-Derived Macrophages

    Science.gov (United States)

    Sanna, Manuela; Cano, Antonella; Delogu, Giuseppe; Erre, Giuseppe; Roberts, Craig W.; Henriquez, Fiona L.; Fiori, Pier Luigi; Cappuccinelli, Piero

    2016-01-01

    Free-living amoebae of the genus Acanthamoeba can cause severe and chronic infections in humans, mainly localized in immune privileged sites, such as the brain and the eye. Monocytes/macrophages are thought to be involved in Acanthamoeba infections, but little is known about how these facultative parasites influence their functions. The aim of this work was to investigate the effects of Acanthamoeba on human monocytes/macrophages during the early phase of infection. Here, THP-1 cells, primary human monocytes isolated from peripheral blood, and human monocyte-derived macrophages were either coincubated with trophozoites of a clinical isolate of Acanthamoeba (genotype T4) or stimulated with amoeba-derived cell-free conditioned medium. Production of proinflammatory cytokines (tumor necrosis factor alpha [TNF-α], interleukin-6 [IL-6], and IL-12), anti-inflammatory cytokine (IL-10), and chemokine (IL-8) was evaluated at specific hours poststimulation (ranging from 1.5 h to 23 h). We showed that both Acanthamoeba trophozoites and soluble amoebic products induce an early anti-inflammatory monocyte-macrophage phenotype, characterized by significant production of IL-10; furthermore, challenge with either trophozoites or their soluble metabolites stimulate both proinflammatory cytokines and chemokine production, suggesting that this protozoan infection results from the early induction of coexisting, opposed immune responses. Results reported in this paper confirm that the production of proinflammatory cytokines and chemokines by monocytes and macrophages can play a role in the development of the inflammatory response during Acanthamoeba infections. Furthermore, we demonstrate for the first time that Acanthamoeba stimulates IL-10 production in human innate immune cells, which might both promote the immune evasion of Acanthamoeba and limit the induced inflammatory response. PMID:27481240

  3. Tumour-cytolytic human monocyte-derived macrophages: a simple and efficient method for the generation and long-term cultivation as non-adherent cells in a serum-free medium.

    Science.gov (United States)

    Streck, R J; Hurley, E L; Epstein, D A; Pauly, J L

    1992-01-01

    We report a simple and efficient culture procedure for the generation of tumour-cytolytic human monocyte-derived macrophages (MAC). In this method, normal human peripheral blood mononuclear cells, isolated using a conventional Ficoll-Hypaque density gradient procedure, are cultured as a heterogenous leukocyte population in Teflon or other hydrophobic cultureware, in a commercially available serum-free culture medium (M-SFM) that has been formulated specifically for the cultivation and ex vivo stimulation of human monocytes and MAC, and in the absence of exogenous mitogens, antigens, cytokines or other stimulants. This procedure features a negative-selection technique that takes advantage of the differential survival of blood leukocytes. Using the prescribed in vitro conditions, lymphocytes survived relatively poorly, whereas monocytes differentiated in the absence of exogenous stimulants into mature tumour-cytolytic MAC. The MAC were present as non-adherent, single cells that expressed good viability (greater than 95%) for a prolonged period (greater than 60 days). When compared to conventional procedures for generating MAC, the prescribed technique is thought to offer several important advantages in that it: (a) eliminates the tedious and cumbersome monocyte isolation procedures, thus providing a significant savings not only in time and money but also in eliminating repetitive cell manipulations that have often been associated with damage to monocyte morphology and/or function; (b) reduces the loss of monocyte subsets that are not recovered during specific isolation procedures; (c) facilitates harvesting a single cell, non-adherent suspension of immunocompetent MAC suitable for various examinations including analyses defining MAC morphology, cytochemistry, phenotype and function; and (d) eliminates variability and artifacts associated with different sera that are utilised frequently as medium supplements. The utility of the prescribed method is illustrated by the

  4. The abcEDCBA-Encoded ABC Transporter and the virB Operon-Encoded Type IV Secretion System of Brucella ovis Are Critical for Intracellular Trafficking and Survival in Ovine Monocyte-Derived Macrophages.

    Directory of Open Access Journals (Sweden)

    Auricelio A Macedo

    Full Text Available Brucella ovis infection is associated with epididymitis, orchitis and infertility in rams. Most of the information available on B. ovis and host cell interaction has been generated using murine macrophages or epithelial cell lines, but the interaction between B. ovis and primary ovine macrophages has not been studied. The aim of this study was to evaluate the role of the B. ovis abcEDCBA-encoded ABC transporter and the virB operon-encoded Type IV Secretion System (T4SS during intracellular survival of B. ovis in ovine peripheral blood monocyte-derived macrophages. ΔabcBA and ΔvirB2 mutant strains were unable to survive in the intracellular environment when compared to the WT B. ovis at 48 hours post infection (hpi. In addition, these mutant strains cannot exclude the lysosomal marker LAMP1 from its vacuolar membrane, and their vacuoles do not acquire the endoplasmic reticulum marker calreticulin, which takes place in the WT B. ovis containing vacuole. Higher levels of nitric oxide production were observed in macrophages infected with WT B. ovis at 48 hpi when compared to macrophages infected with the ΔabcBA or ΔvirB2 mutant strains. Conversely, higher levels of reactive oxygen species were detected in macrophages infected with the ΔabcBA or ΔvirB2 mutant strains at 48 hpi when compared to macrophages infected with the WT strain. Our results demonstrate that B. ovis is able to persist and multiply in ovine macrophages, while ΔabcBA and ΔvirB2 mutations prevent intracellular multiplication, favor phagolysosome fusion, and impair maturation of the B. ovis vacuole towards an endoplasmic reticulum-derived compartment.

  5. Role of neoplastic monocyte-derived fibrocytes in primary myelofibrosis

    Science.gov (United States)

    Bueso-Ramos, Carlos E.; Newberry, Kate J.; Knez, Liza; Post, Sean M.; Ahn, Jihae; Levine, Ross L.; Kantarjian, Hagop M.

    2016-01-01

    Primary myelofibrosis (PMF) is a fatal neoplastic disease characterized by clonal myeloproliferation and progressive bone marrow (BM) fibrosis thought to be induced by mesenchymal stromal cells stimulated by overproduced growth factors. However, tissue fibrosis in other diseases is associated with monocyte-derived fibrocytes. Therefore, we sought to determine whether fibrocytes play a role in the induction of BM fibrosis in PMF. In this study, we show that BM from patients with PMF harbors an abundance of clonal, neoplastic collagen- and fibronectin-producing fibrocytes. Immunodeficient mice transplanted with myelofibrosis patients’ BM cells developed a lethal myelofibrosis-like phenotype. Treatment of the xenograft mice with the fibrocyte inhibitor serum amyloid P (SAP; pentraxin-2) significantly prolonged survival and slowed the development of BM fibrosis. Collectively, our data suggest that neoplastic fibrocytes contribute to the induction of BM fibrosis in PMF, and inhibiting fibrocyte differentiation with SAP may interfere with this process. PMID:27481130

  6. Divergent JAM-C Expression Accelerates Monocyte-Derived Cell Exit from Atherosclerotic Plaques.

    Directory of Open Access Journals (Sweden)

    Paul F Bradfield

    Full Text Available Atherosclerosis, caused in part by monocytes in plaques, continues to be a disease that afflicts the modern world. Whilst significant steps have been made in treating this chronic inflammatory disease, questions remain on how to prevent monocyte and macrophage accumulation in atherosclerotic plaques. Junctional Adhesion Molecule C (JAM-C expressed by vascular endothelium directs monocyte transendothelial migration in a unidirectional manner leading to increased inflammation. Here we show that interfering with JAM-C allows reverse-transendothelial migration of monocyte-derived cells, opening the way back out of the inflamed environment. To study the role of JAM-C in plaque regression we used a mouse model of atherosclerosis, and tested the impact of vascular JAM-C expression levels on monocyte reverse transendothelial migration using human cells. Studies in-vitro under inflammatory conditions revealed that overexpression or gene silencing of JAM-C in human endothelium exposed to flow resulted in higher rates of monocyte reverse-transendothelial migration, similar to antibody blockade. We then transplanted atherosclerotic, plaque-containing aortic arches from hyperlipidemic ApoE-/- mice into wild-type normolipidemic recipient mice. JAM-C blockade in the recipients induced greater emigration of monocyte-derived cells and further diminished the size of atherosclerotic plaques. Our findings have shown that JAM-C forms a one-way vascular barrier for leukocyte transendothelial migration only when present at homeostatic copy numbers. We have also shown that blocking JAM-C can reduce the number of atherogenic monocytes/macrophages in plaques by emigration, providing a novel therapeutic strategy for chronic inflammatory pathologies.

  7. Divergent JAM-C Expression Accelerates Monocyte-Derived Cell Exit from Atherosclerotic Plaques.

    Science.gov (United States)

    Bradfield, Paul F; Menon, Arjun; Miljkovic-Licina, Marijana; Lee, Boris P; Fischer, Nicolas; Fish, Richard J; Kwak, Brenda; Fisher, Edward A; Imhof, Beat A

    2016-01-01

    Atherosclerosis, caused in part by monocytes in plaques, continues to be a disease that afflicts the modern world. Whilst significant steps have been made in treating this chronic inflammatory disease, questions remain on how to prevent monocyte and macrophage accumulation in atherosclerotic plaques. Junctional Adhesion Molecule C (JAM-C) expressed by vascular endothelium directs monocyte transendothelial migration in a unidirectional manner leading to increased inflammation. Here we show that interfering with JAM-C allows reverse-transendothelial migration of monocyte-derived cells, opening the way back out of the inflamed environment. To study the role of JAM-C in plaque regression we used a mouse model of atherosclerosis, and tested the impact of vascular JAM-C expression levels on monocyte reverse transendothelial migration using human cells. Studies in-vitro under inflammatory conditions revealed that overexpression or gene silencing of JAM-C in human endothelium exposed to flow resulted in higher rates of monocyte reverse-transendothelial migration, similar to antibody blockade. We then transplanted atherosclerotic, plaque-containing aortic arches from hyperlipidemic ApoE-/- mice into wild-type normolipidemic recipient mice. JAM-C blockade in the recipients induced greater emigration of monocyte-derived cells and further diminished the size of atherosclerotic plaques. Our findings have shown that JAM-C forms a one-way vascular barrier for leukocyte transendothelial migration only when present at homeostatic copy numbers. We have also shown that blocking JAM-C can reduce the number of atherogenic monocytes/macrophages in plaques by emigration, providing a novel therapeutic strategy for chronic inflammatory pathologies.

  8. Differential regulatory activities of viral protein X for anti-viral efficacy of nucleos(t)ide reverse transcriptase inhibitors in monocyte-derived macrophages and activated CD4(+) T cells.

    Science.gov (United States)

    Hollenbaugh, Joseph A; Schader, Susan M; Schinazi, Raymond F; Kim, Baek

    2015-11-01

    Vpx encoded by HIV-2 and SIVsm enhances retroviral reverse transcription in macrophages in vitro by mediating the degradation of the host SAMHD1 protein that hydrolyzes dNTPs and by elevating cellular dNTP levels. Here we employed RT-SHIV constructs (SIV encoding HIV-1 RT) to investigate the contribution of Vpx to the potency of NRTIs, which compete against dNTPs, in monocyte-derived macrophages (MDMs) and activated CD4(+) T cells. Relative to HIV-1, both SIV and RT-SHIV exhibited reduced sensitivities to AZT, 3TC and TDF in MDMs but not in activated CD4(+) T cells. However, when SIV and RT-SHIV constructs not coding for Vpx were utilized, we observed greater sensitivities to all NRTIs tested using activated CD4(+) T cells relative to the Vpx-coding counterparts. This latter phenomenon was observed for AZT only when using MDMs. Our data suggest that Vpx in RT-SHIVs may underestimate the antiviral efficacy of NRTIs in a cell type dependent manner.

  9. HCMV Displays a Unique Transcriptome of Immunomodulatory Genes in Primary Monocyte-Derived Cell Types

    Science.gov (United States)

    Van Damme, Ellen; Thys, Kim; Tuefferd, Marianne; Van Hove, Carl; Aerssens, Jeroen; Van Loock, Marnix

    2016-01-01

    Human cytomegalovirus (HCMV) is a betaherpesvirus which rarely presents problems in healthy individuals, yet may result in severe morbidity in immunocompromised patients and in immune-naïve neonates. HCMV has a large 235 kb genome with a coding capacity of at least 165 open reading frames (ORFs). This large genome allows complex gene regulation resulting in different sets of transcripts during lytic and latent infection. While latent virus mainly resides within monocytes and CD34+ progenitor cells, reactivation to lytic infection is driven by differentiation towards terminally differentiated myeloid dendritic cells and macrophages. Consequently, it has been suggested that macrophages and dendritic cells contribute to viral spread in vivo. Thus far only limited knowledge is available on the expression of HCMV genes in terminally differentiated myeloid primary cells and whether or not the virus exhibits a different set of lytic genes in primary cells compared with lytic infection in NHDF fibroblasts. To address these questions, we used Illumina next generation sequencing to determine the HCMV transcriptome in macrophages and dendritic cells during lytic infection and compared it to the transcriptome in NHDF fibroblasts. Here, we demonstrate unique expression profiles in macrophages and dendritic cells which significantly differ from the transcriptome in fibroblasts mainly by modulating the expression of viral transcripts involved in immune modulation, cell tropism and viral spread. In a head to head comparison between macrophages and dendritic cells, we observed that factors involved in viral spread and virion composition are differentially regulated suggesting that the plasticity of the virion facilitates the infection of surrounding cells. Taken together, this study provides the full transcript expression analysis of lytic HCMV genes in monocyte-derived type 1 and type 2 macrophages as well as in monocyte-derived dendritic cells. Thereby underlining the potential

  10. Human Monocyte-Derived Osteoclasts Are Targeted by Staphylococcal Pore-Forming Toxins and Superantigens.

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    Sacha Flammier

    Full Text Available Staphylococcus aureus is the leading cause of bone and joint infections (BJIs. Staphylococcal pathogenesis involves numerous virulence factors including secreted toxins such as pore-forming toxins (PFTs and superantigens. The role of these toxins on BJI outcome is largely unknown. In particular, few studies have examined how osteoclasts, the bone-resorbing cells, respond to exposure to staphylococcal PFTs and superantigens. We investigated the direct impact of recombinant staphylococcal toxins on human primary mature monocyte-derived osteoclasts, in terms of cytotoxicity and cell activation with cell death and bone resorption assays, using macrophages of the corresponding donors as a reference. Monocyte-derived osteoclasts displayed similar toxin susceptibility profiles compared to macrophages. Specifically, we demonstrated that the Panton-Valentine leukocidin, known as one of the most powerful PFT which lyses myeloid cells after binding to the C5a receptor, was able to induce the death of osteoclasts. The archetypal superantigen TSST-1 was not cytotoxic but enhanced the bone resorption activity of osteoclasts, suggesting a novel mechanism by which superantigen-producing S. aureus can accelerate the destruction of bone tissue during BJI. Altogether, our data indicate that the diverse clinical presentations of BJIs could be related, at least partly, to the toxin profiles of S. aureus isolates involved in these severe infections.

  11. Genome-Wide Association Study Identifies Single Nucleotide Polymorphism in DYRK1A Associated with Replication of HIV-1 in Monocyte-Derived Macrophages

    NARCIS (Netherlands)

    Bol, S.M.; Moerland, P.D.; Limou, S.; van Remmerden, Y.; Coulonges, C.; Manen, D.; Herbeck, J.T.; Fellay, J.; Sieberer, M.; Sietzema, J.G.; van 't Slot, R.; Martinson, J.; Zagury, J.F.; Schuitemaker, H.; van 't Wout, A.B.

    2011-01-01

    Background: HIV-1 infected macrophages play an important role in rendering resting T cells permissive for infection, in spreading HIV-1 to T cells, and in the pathogenesis of AIDS dementia. During highly active anti-retroviral treatment (HAART), macrophages keep producing virus because tissue penetr

  12. Characterization of canine monocyte-derived dendritic cells with phenotypic and functional differentiation.

    Science.gov (United States)

    Wang, Yu-Shan; Chi, Kwan-Hwa; Liao, Kuang-Wen; Liu, Cheng-Chi; Cheng, Chiao-Lei; Lin, Yi-Chun; Cheng, Chiung-Hsiang; Chu, Rea-Min

    2007-07-01

    For therapeutic purposes, large numbers of dendritic cells (DCs) are essential. In this study, we used 2% autologous canine plasma, granulocyte/macrophage colony-stimulating factor (GM-CSF), fms-like tyrosine kinase 3 ligand (Flt3L), and interleukin 4 (IL-4) in generating monocyte-derived DCs from peripheral blood mononuclear cells of dogs. The plasma enriched the population of CD14-positive monocytes by greatly enhancing the efficiency of monocyte adherence, the proportion of adherent cells increasing from 6.6% with 10% fetal bovine serum to 15.3% with 2% autologous canine plasma. Culturing the adherent monocytes for 6 d with human GM-CSF, canine IL-4, and human Flt3L significantly increased the yield of DCs, more than 90% of which were CD14-negative. Because, in the presence of lipopolysaccharide (LPS), monocytes that were CD14-positive expressed tumor necrosis factor ac much more than DCs with low levels of CD14, it is important to decrease the numbers of CD14-positive cells in generating monocyte-derived DCs. With flow cytometry and real-time reverse-transcriptase-mediated polymerase chain reaction assays, we found that in canine immature DCs (iDCs) the expression of DLA class II molecules, CD1a, CD11c, CD40, and CD86 was high and the expression of CD80, CD83, and CD14 either low or negative. During maturation (stimulated by LPS), the expression of CDla, CD40, CD83, and CD80 was upregulated. However, the expression of DLA class II molecules, CD11c, and CD86 was not increased in mature DCs. Incubating the iDCs with LPS decreased antigen uptake and increased the cells' immunostimulatory capacity (assessed by the allogeneic mixed-lymphocyte reaction), indicating that LPS accelerates the functional maturation of DCs. This protocol may facilitate the use of DCs in cellular immunotherapy.

  13. Role of HIV-1 subtype C envelope V3 to V5 regions in viral entry, coreceptor utilization and replication efficiency in primary T-lymphocytes and monocyte-derived macrophages

    Directory of Open Access Journals (Sweden)

    Gopalan Sarla

    2007-11-01

    Full Text Available Abstract Background Several subtypes of HIV-1 circulate in infected people worldwide, including subtype B in the United States and subtype C in Africa and India. To understand the biological properties of HIV-1 subtype C, including cellular tropism, virus entry, replication efficiency and cytopathic effects, we reciprocally inserted our previously characterized envelope V3–V5 regions derived from 9 subtype C infected patients from India into a subtype B molecular clone, pNL4-3. Equal amounts of the chimeric viruses were used to infect T-lymphocyte cell lines (A3.01 and MT-2, coreceptor cell lines (U373-MAGI-CCR5/CXCR4, primary blood T-lymphocytes (PBL and monocyte-derived macrophages (MDM. Results We found that subtype C envelope V3–V5 region chimeras failed to replicate in T-lymphocyte cell lines but replicated in PBL and MDM. In addition, these chimeras were able to infect U373MAGI-CD4+-CCR5+ but not U373MAGI-CD4+-CXCR4+ cell line, suggesting CCR5 coreceptor utilization and R5 phenotypes. These subtype C chimeras were unable to induce syncytia in MT-2 cells, indicative of non-syncytium inducing (NSI phenotypes. More importantly, the subtype C envelope chimeras replicated at higher levels in PBL and MDM compared with subtype B chimeras and isolates. Furthermore, the higher levels subtype C chimeras replication in PBL and MDM correlated with increased virus entry in U373MAGI-CD4+-CCR5+. Conclusion Taken together, these results suggest that the envelope V3 to V5 regions of subtype C contributed to higher levels of HIV-1 replication compared with subtype B chimeras, which may contribute to higher viral loads and faster disease progression in subtype C infected individuals than other subtypes as well as rapid HIV-1 subtype C spread in India.

  14. cGAS Senses Human Cytomegalovirus and Induces Type I Interferon Responses in Human Monocyte-Derived Cells

    Science.gov (United States)

    Paijo, Jennifer; Döring, Marius; Spanier, Julia; Grabski, Elena; Nooruzzaman, Mohammed; Schmidt, Tobias; Witte, Gregor; Messerle, Martin; Hornung, Veit; Kaever, Volkhard; Kalinke, Ulrich

    2016-01-01

    Human cytomegalovirus (HCMV) infections of healthy individuals are mostly unnoticed and result in viral latency. However, HCMV can also cause devastating disease, e.g., upon reactivation in immunocompromised patients. Yet, little is known about human immune cell sensing of DNA-encoded HCMV. Recent studies indicated that during viral infection the cyclic GMP/AMP synthase (cGAS) senses cytosolic DNA and catalyzes formation of the cyclic di-nucleotide cGAMP, which triggers stimulator of interferon genes (STING) and thus induces antiviral type I interferon (IFN-I) responses. We found that plasmacytoid dendritic cells (pDC) as well as monocyte-derived DC and macrophages constitutively expressed cGAS and STING. HCMV infection further induced cGAS, whereas STING expression was only moderately affected. Although pDC expressed particularly high levels of cGAS, and the cGAS/STING axis was functional down-stream of STING, as indicated by IFN-I induction upon synthetic cGAMP treatment, pDC were not susceptible to HCMV infection and mounted IFN-I responses in a TLR9-dependent manner. Conversely, HCMV infected monocyte-derived cells synthesized abundant cGAMP levels that preceded IFN-I production and that correlated with the extent of infection. CRISPR/Cas9- or siRNA-mediated cGAS ablation in monocytic THP-1 cells and primary monocyte-derived cells, respectively, impeded induction of IFN-I responses following HCMV infection. Thus, cGAS is a key sensor of HCMV for IFN-I induction in primary human monocyte-derived DC and macrophages. PMID:27058035

  15. Galectin-3 Binding Protein Secreted by Breast Cancer Cells Inhibits Monocyte-Derived Fibrocyte Differentiation.

    Science.gov (United States)

    White, Michael J V; Roife, David; Gomer, Richard H

    2015-08-15

    To metastasize, tumor cells often need to migrate through a layer of collagen-containing scar tissue which encapsulates the tumor. A key component of scar tissue and fibrosing diseases is the monocyte-derived fibrocyte, a collagen-secreting profibrotic cell. To test the hypothesis that invasive tumor cells may block the formation of the fibrous sheath, we determined whether tumor cells secrete factors that inhibit monocyte-derived fibrocyte differentiation. We found that the human metastatic breast cancer cell line MDA-MB-231 secretes activity that inhibits human monocyte-derived fibrocyte differentiation, whereas less aggressive breast cancer cell lines secrete less of this activity. Purification indicated that Galectin-3 binding protein (LGALS3BP) is the active factor. Recombinant LGALS3BP inhibits monocyte-derived fibrocyte differentiation, and immunodepletion of LGALS3BP from MDA-MB 231 conditioned media removes the monocyte-derived fibrocyte differentiation-inhibiting activity. LGALS3BP inhibits the differentiation of monocyte-derived fibrocytes from wild-type mouse spleen cells, but not from SIGN-R1(-/-) mouse spleen cells, suggesting that CD209/SIGN-R1 is required for the LGALS3BP effect. Galectin-3 and galectin-1, binding partners of LGALS3BP, potentiate monocyte-derived fibrocyte differentiation. In breast cancer biopsies, increased levels of tumor cell-associated LGALS3BP were observed in regions of the tumor that were invading the surrounding stroma. These findings suggest LGALS3BP and galectin-3 as new targets to treat metastatic cancer and fibrosing diseases.

  16. Inhibition of the differentiation of monocyte-derived dendritic cells by human gingival fibroblasts.

    Directory of Open Access Journals (Sweden)

    Sylvie Séguier

    Full Text Available We investigated whether gingival fibroblasts (GFs can modulate the differentiation and/or maturation of monocyte-derived dendritic cells (DCs and analyzed soluble factors that may be involved in this immune modulation. Experiments were performed using human monocytes in co-culture with human GFs in Transwell® chambers or using monocyte cultures treated with conditioned media (CM from GFs of four donors. The four CM and supernatants from cell culture were assayed by ELISA for cytokines involved in the differentiation of dendritic cells, such as IL-6, VEGF, TGFβ1, IL-13 and IL-10. The maturation of monocyte-derived DCs induced by LPS in presence of CM was also studied. Cell surface phenotype markers were analyzed by flow cytometry. In co-cultures, GFs inhibited the differentiation of monocyte-derived DCs and the strength of this blockade correlated with the GF/monocyte ratio. Conditioned media from GFs showed similar effects, suggesting the involvement of soluble factors produced by GFs. This inhibition was associated with a lower stimulatory activity in MLR of DCs generated with GFs or its CM. Neutralizing antibodies against IL-6 and VEGF significantly (P<0.05 inhibited the inhibitory effect of CM on the differentiation of monocytes-derived DCs and in a dose dependent manner. Our data suggest that IL-6 is the main factor responsible for the inhibition of DCs differentiation mediated by GFs but that VEGF is also involved and constitutes an additional mechanism.

  17. CHI3L1 nuclear localization in monocyte derived dendritic cells.

    Science.gov (United States)

    Di Rosa, Michelino; Tibullo, Daniele; Saccone, Salvatore; Distefano, Gisella; Basile, Maria Sofia; Di Raimondo, Francesco; Malaguarnera, Lucia

    2016-02-01

    Chitinase-3-like-1 protein (CHI3L1) is a glycosyl hydrolase (GH) highly expressed in a variety of inflammatory diseases at infectious and non-infectious etiology. CHI3L1 is produced by a wide variety of cells including monocyte-derived macrophages cell lines such as polarized M1 and M2 type macrophages, osteoclasts and Kupffer cells. In this study we have examined the expression of CHI3L1 during the differentiation and maturation of dendritic cells. Magnetically-isolated peripheral blood monocytes were differentiated toward immature DCs (iDC) and mature DCs (mDCs) through a combination of factors and cytokines. Our result showed, for the first time, that CHI3L1 is expressed during the process of differentiation and maturation of dendritic cells in time dependent manner. Furthermore, the CHI3L1 is evenly distributed in cytoplasm and in the nucleus of both the iDCs and mDCs. These results suggest that CHI3L1 may play crucial role in the DCs immunoresponse.

  18. The effect of short-chain fatty acids on human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Nastasi, Claudia; Candela, Marco; Bonefeld, Charlotte Menné

    2015-01-01

    negligible effects, while both butyrate and propionate strongly modulated gene expression in both immature and mature human monocyte-derived DC. An Ingenuity pathway analysis based on the differentially expressed genes suggested that propionate and butyrate modulate leukocyte trafficking, as SCFA strongly......The gut microbiota is essential for human health and plays an important role in the pathogenesis of several diseases. Short-chain fatty acids (SCFA), such as acetate, butyrate and propionate, are end-products of microbial fermentation of macronutrients that distribute systemically via the blood....... The aim of this study was to investigate the transcriptional response of immature and LPS-matured human monocyte-derived DC to SCFA. Our data revealed distinct effects exerted by each individual SCFA on gene expression in human monocyte-derived DC, especially in the mature ones. Acetate only exerted...

  19. Intracranial transplantation of monocyte-derived multipotential cells enhances recovery after ischemic stroke in rats.

    Science.gov (United States)

    Hattori, Hidenori; Suzuki, Shigeaki; Okazaki, Yuka; Suzuki, Norihiro; Kuwana, Masataka

    2012-02-01

    Cell transplantation has emerged as a potential therapy to reduce the neurological deficits caused by ischemic stroke. We previously reported a primitive cell population, monocyte-derived multipotential cells (MOMCs), which can differentiate into mesenchymal, neuronal, and endothelial lineages. In this study, MOMCs and macrophages were prepared from rat peripheral blood and transplanted intracranially into the ischemic core of syngeneic rats that had undergone a left middle cerebral artery occlusion procedure. Neurological deficits, as evaluated by the corner test, were less severe in the MOMC-transplanted rats than in macrophage-transplanted or mock-treated rats. Histological evaluations revealed that the number of microvessels that had formed in the ischemic boundary area by 4 weeks after transplantation was significantly greater in the MOMC-transplanted rats than in the control groups. The blood vessel formation was preceded by the appearance of round CD31(+) cells, which we confirmed were derived from the transplanted MOMCs. Small numbers of bloodvessels incorporating MOMC-derived endothelial cells expressing a mature endothelial marker RECA-1 were detected at 4 weeks after transplantation. In addition, MOMCs expressed a series of angiogenic factors, including vascular endothelial growth factor, angiopoetin-1, and placenta growth factor (PlGF). These findings provide evidence that the intracranial delivery of MOMCs enhances functional recovery by promoting neovascularization in a rat model for ischemic stroke.

  20. Inducing Maturation of Monocyte-Derived Dendritic Cells on Human Epithelial Cell Feeder Layer

    Directory of Open Access Journals (Sweden)

    Delirezh N

    2012-02-01

    Full Text Available Background: Nowadays, dendritic cells (DCs have a special place in cancer treatment strategies and they have been used for tumor immunotherapy as they can induce immune response against tumor cells. Researchers have been trying to generate efficient dendritic cells in vitro; therefore, this research was done to generate them for use in research and tumor immunotherapy. Methods: This study took place at Urmia University in 2010-2011 years. In this study plastic adherent monocytes were incubated with granulocyte-macrophage colony stimulating factor (GM-CSF and interleukin-4 (IL-4 for five days. Finally, fully matured and stable DCs were generated by 48 hours of incubation in a monocyte conditioned medium (MCM containing tumor necrosis factor-α (TNF-α and epithelial cells. Phenotypic and functional analysis were carried out by using anti-CD14, anti-CD80, anti-CD86, and anti-CD83 monoclonal antibodies, and by determining their phagocytic activity, mixed lymphocyte reaction (MLR and cytokine production, respectively. Results: Dendritic cells were produced with high levels of surface molecule, i.e. of CD80, CD83, CD86, HLA-DR, expression and low levels of CD14 expression. Dendritic cells showed efficient phagocytosis and ability to stimulate T-lymphocytes. Moreover, dendritic cells could secrete high levels of interleukin-12 (IL-12 cytokine which was depictive of their full maturation. Measurement of the produced cytokines showed the generation of type-1 dendritic cells (DC1. Conclusion: Our study showed that skin epithelial cells could induce maturation of monocyte-derived dendritic cells (DCs. This feeder layer led to the production of efficient dendritic cells with the ability to be used for tumor immunotherapy.

  1. A novel in vitro human microglia model: characterization of human monocyte-derived microglia.

    Science.gov (United States)

    Etemad, Samar; Zamin, Rasheeda Mohd; Ruitenberg, Marc J; Filgueira, Luis

    2012-07-30

    Microglia are the innate immune cells of the central nervous system. They help maintaining physiological homeostasis and contribute significantly to inflammatory responses in the course of infection, injury and degenerative processes. To date, there is no standardized simple model available to investigate the biology of human microglia. The aim of this study was to establish a new human microglia model. For that purpose, human peripheral blood monocytes were cultured in serum free medium in the presence of M-CSF, GM-CSF, NGF and CCL2 to generate monocyte-derived microglia (M-MG). M-MG were clearly different in morphology, phenotype and function from freshly isolated monocytes, cultured monocytes in the absence of the cytokines and monocyte-derived dendritic cells (M-DC) cultured in the presence of GM-CSF and IL-4. M-MG acquired a ramified morphology with primary and secondary processes. M-MG displayed a comparable phenotype to the human microglia cell line HMC3, expressing very low levels of CD45, CD14 and HLA-DR, CD11b and CD11c; and undetectable levels of CD40, CD80 and CD83, and a distinct pattern of chemokine receptors (positive for CCR1, CCR2, CCR4, CCR5, CXCR1, CXCR3, CX3CR1; negative for CCR6 and CCR7). In comparison with M-DC, M-MG displayed lower T-lymphocyte stimulatory capacity, as well as lower phagocytosis activity. The described protocol for the generation of human monocyte-derived microglia is feasible, well standardized and reliable, as it uses well defined culture medium and recombinant cytokines, but no serum or conditioned medium. This protocol will certainly be very helpful for future studies investigating the biology and pathology of human microglia.

  2. Monocyte-derived inflammatory Langerhans cells and dermal dendritic cells mediate psoriasis-like inflammation.

    Science.gov (United States)

    Singh, Tej Pratap; Zhang, Howard H; Borek, Izabela; Wolf, Peter; Hedrick, Michael N; Singh, Satya P; Kelsall, Brian L; Clausen, Bjorn E; Farber, Joshua M

    2016-12-16

    Dendritic cells (DCs) have been implicated in the pathogenesis of psoriasis but the roles for specific DC subsets are not well defined. Here we show that DCs are required for psoriasis-like changes in mouse skin induced by the local injection of IL-23. However, Flt3L-dependent DCs and resident Langerhans cells are dispensable for the inflammation. In epidermis and dermis, the critical DCs are TNF-producing and IL-1β-producing monocyte-derived DCs, including a population of inflammatory Langerhans cells. Depleting Ly6C(hi) blood monocytes reduces DC accumulation and the skin changes induced either by injecting IL-23 or by application of the TLR7 agonist imiquimod. Moreover, we find that IL-23-induced inflammation requires expression of CCR6 by DCs or their precursors, and that CCR6 mediates monocyte trafficking into inflamed skin. Collectively, our results imply that monocyte-derived cells are critical contributors to psoriasis through production of inflammatory cytokines that augment the activation of skin T cells.

  3. Monocyte-derived inflammatory Langerhans cells and dermal dendritic cells mediate psoriasis-like inflammation

    Science.gov (United States)

    Singh, Tej Pratap; Zhang, Howard H.; Borek, Izabela; Wolf, Peter; Hedrick, Michael N.; Singh, Satya P.; Kelsall, Brian L.; Clausen, Bjorn E.; Farber, Joshua M.

    2016-01-01

    Dendritic cells (DCs) have been implicated in the pathogenesis of psoriasis but the roles for specific DC subsets are not well defined. Here we show that DCs are required for psoriasis-like changes in mouse skin induced by the local injection of IL-23. However, Flt3L-dependent DCs and resident Langerhans cells are dispensable for the inflammation. In epidermis and dermis, the critical DCs are TNF-producing and IL-1β-producing monocyte-derived DCs, including a population of inflammatory Langerhans cells. Depleting Ly6Chi blood monocytes reduces DC accumulation and the skin changes induced either by injecting IL-23 or by application of the TLR7 agonist imiquimod. Moreover, we find that IL-23-induced inflammation requires expression of CCR6 by DCs or their precursors, and that CCR6 mediates monocyte trafficking into inflamed skin. Collectively, our results imply that monocyte-derived cells are critical contributors to psoriasis through production of inflammatory cytokines that augment the activation of skin T cells. PMID:27982014

  4. Dysfunctional HDL from HIV+ individuals promotes monocyte-derived foam cell formation in vitro.

    Science.gov (United States)

    Angelovich, Thomas A; Hearps, Anna C; Oda, Michael N; Borja, Mark S; Huynh, Diana; Homann, Stefanie; Jaworowski, Anthony; Kelesidis, Theodoros

    2017-09-18

    The role of HDL function in HIV-related atherosclerotic cardiovascular disease (CVD) is unclear. HDLs isolated from HIV+ [HIV(+)HDL] and HIV-uninfected individuals (HDL) were assessed for HDL function and ability to promote monocyte-derived foam cell formation (MDFCF) (a key event in HIV-related CVD) ex vivo. Using an established in vitro model of atherogenesis and plasma samples from an established cross-sectional study of virologically-suppressed HIV+ males on stable effective antiretroviral therapy (ART) and with low CVD risk (median age: 42 years; n = 10), we explored the impact of native HDL [HIV(+)HDL] on MDFCF. In this exploratory study we selected HIV-HDL known to be dysfunctional based on two independent measures of impaired HDL function: a) antioxidant (high HDLox) b) ability of HDL to release apoA-I [low HDL-apoA-I exchange (HAE %)]. Five healthy males matched by age and race to the HIV+ group were included. Given that oxidation of HDL leads to abnormal HDL function, we also compared proatherogenic effects of HIV-HDL versus chemically-derived HDLox. The ex vivo atherogenesis assay was performed using lipoproteins (purchased or isolated from plasma using ultracentrifugation) and monocytes purified via negative selection from healthy donors. HIV(+)HDL known to have reduced antioxidant function and rate of HDL/ApoAI exchange promoted MDFCF to a greater extent than HDL (33.0% vs 26.2% foam cells; p = 0.015). HDL oxidized in vitro also enhanced foam cell formation as compared to non-oxidized HDL (p HDL in virologically suppressed HIV+ individuals may potentiate atherosclerosis in HIV infection by promoting monocyte-derived foam cell formation.The role of HDL function in HIV-related atherosclerotic cardiovascular disease is unclear. HDL isolated from HIV+ [HIV(+)HDL] and HIV-uninfected individuals [HIV(-)HDL] were assessed for HDL function and ability to promote foam cell formation ex vivo. HIV(+)HDL known to have reduced antioxidant function and

  5. [Induction of monocyte-derived dendritic cell differentiation by asthmatic serum in a transendothelial trafficking model].

    Science.gov (United States)

    Zhou, Lin-fu; Wang, Wen-lu; Li, Hong-yan; Zhang, Ming-shun; Ji, Xiao-hui; He, Shao-heng; Huang, Mao; Yin, Kai-sheng

    2011-03-01

    To explore the effect of asthmatic and healthy serum on differentiation and function of monocyte-derived dendritic cells (MDDC) in a transendothelial trafficking model. The sera and peripheral blood mononuclear cells (PBMC) were separated from 12 asthmatic patients and 12 healthy volunteers, and monocytes were selected from PBMC using magnetic beads. The trypsin-digested human umbilical vein endothelial cells (HUVEC) at passage 2 from 5 healthy lying-in women were used to construct the transendothelial trafficking model under asthmatic or healthy serum, wherein MDDC were identified by silver nitrate staining and scanning electron microscopy. Nuclear factor κB (NF-κB) activity was determined by electrophoretic mobility shift assay. Flow cytometry, ELISA and mixed leukocyte reaction were relevantly utilized to detect the phenotype, cytokine and T cell proliferation. (1) Monocytes traversed through HUVEC monolayer after 2 h, and reverse-transmigrated to develop into DC 48 h later. (2) The healthy serum stimulated monocytes into immature MDDC with lower CD(14) [(20 ± 5)%] (F = 49.01, P 0.05), higher CD(80) and CD(83) [(49.7 ± 10.2)% and (30.2 ± 6.8)%] (F = 4.01 and 20.68, all P trafficking model, which provides a promising experimental platform for both investigation of immunological mechanisms in asthma and screening of novel anti-asthma drugs in vitro.

  6. Isolation of IL-12p70-competent human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Søndergaard, Jonas Nørskov; Pedersen, Susanne Brix

    2012-01-01

    that moDCs generated under standard conditions develop into two subsets based on CD1a-expression with the CD1a+ moDCs being the main IL-12p70 producers. This has however not been generally accepted, which we show here because the subset described as CD1a-negative does express CD1a, but at a lower level......Diverse methodologies ranging from experimental immunological studies to immunotherapy involve the application of human monocyte-derived dendritic cells (moDCs). Considerable donor-dependent variations in the moDC production of IL-12p70 affect the outcome of these methodologies. It has been shown...

  7. In vitro interaction of Stenotrophomonas maltophilia with human monocyte-derived dendritic cells

    Directory of Open Access Journals (Sweden)

    Emanuela eRoscetto

    2015-07-01

    Full Text Available Stenotrophomonas maltophilia is increasingly identified as an opportunistic pathogen in immunocompromised, cancer and cystic fibrosis (CF patients. Knowledge on innate immune responses to S. maltophilia and its potential modulation is poor. The present work investigated the ability of 12 clinical S. maltophilia strains (5 from CF patients, 7 from non-CF patients and one environmental strain to survive inside human monocyte-derived dendritic cells (DCs. The effects of the bacteria on maturation of and cytokine secretion by DCs were also measured. S. maltophilia strains presented a high degree of heterogeneity in internalization and intracellular replication efficiencies as well as in the ability of S. maltophilia to interfere with normal DCs maturation. By contrast, all S. maltophilia strains were able to activate DCs, as measured by increase in the expression of surface maturation markers and proinflammatory cytokines secretion.

  8. Impaired migration capacity in monocytes derived from patients with Gaucher disease.

    Science.gov (United States)

    Bettman, Noam; Avivi, Irit; Rosenbaum, Hanna; Bisharat, Lina; Katz, Tamar

    2015-08-01

    Gaucher disease (GD) is characterized by glucocerebroside (GC) accumulation due to defective activity of the glucocerebrosidase (GlcCerase) enzyme. Monocytes and macrophages exhibit the highest GlcCerase activity and are most prominently affected by GC engorgement. As GD patients tend to exert various immune system-related changes, this study was designed to investigate potential effects of monocyte dysfunction on these alterations. Monocytes were isolated from peripheral blood mononuclear cells (PBMCs) of untreated GD patients and healthy volunteers. Monocyte migration capacity towards SDF1α was assessed. The GD patients exhibited reduced numbers of monocytes and decreased capability of SDF1α-dependent monocyte migration. Evaluation of CXCR4, the SDF1α receptor, revealed reduced expression of surface CXCR4 on GD-derived monocytes, despite similar CXCR4 mRNA transcript levels in the monocytes of healthy volunteers and GD patients. Reduction of surface CXCR4 was accompanied by increased intracellular CXCR4 levels in patient monocytes. This elevated intracellular CXCR4 might reflect significantly increased SDF1α concentrations characterizing patients' serum and the lysosomal impairment of GD, resulting in decreased degradation of CXCR4. Different distributions of CXCR4 expression observed in the two groups explain impaired SDF1α-dependent monocyte migration. Reduced numbers and impaired migration capacity of GD-derived monocytes could contribute to abnormal inflammation and GD-associated immune alterations seen in these patients.

  9. Differentiation and function of human monocyte-derived dendritic cells under the influence of leflunomide

    Directory of Open Access Journals (Sweden)

    Stojić-Vukanić Z.

    2011-01-01

    Full Text Available Leflunomide is an immunosuppressive drug effective in experimental models of transplantation and autoimmune diseases and in the treatment of active rheumatoid arthritis (RA. Having in mind that it has been shown that some other immunosuppressive drugs (glucocorticoids, mycophenolate mofetil, sirolimus etc. impair dendritic cell (DC phenotype and function, we investigated the effect of A77 1726, an active metabolite of leflunomide, on the differentiation and function of human monocyte-derived dendritic cells (MDDC in vitro. Immature MDDC were generated by cultivating monocytes in medium supplemented with GM-CSF and IL-4. To induce maturation, immature MDDC were cultured for 2 additional days with LPS. A77 1726 (100 μM was added at the beginning of cultivation. Flow cytometric analysis showed that MDDC differentiated in the presence of A77 1726 exhibited an altered phenotype, with a down-regulated surface expression of CD80, CD86, CD54 and CD40 molecules. Furthermore, the continuous presence of A77 1726 during differentiation and maturation prevented successful maturation, judging by the decreased expression of maturation marker CD83, costimulatory and adhesive molecules on A77 1726-treated mature MDDC. In addition, A77 1726-pretreated MDDC exhibited a poor stimulatory capacity of the allogeneic T cells and a low production of IL-10 and IL-18. These data suggest that leflunomide impairs the differentiation, maturation and function of human MDDC in vitro, which is an additional mechanism of its immunosuppressive effect.

  10. Phenotype and Function of Monocyte-Derived Dendritic Cells from Chinese Rhesus Macaques

    Institute of Scientific and Technical Information of China (English)

    Houjun Xia; Hongliang Liu; Gaohong Zhang; Yongtang Zheng

    2009-01-01

    Dendritic cells (DCs) play a pivotal role in linking the innate immunity and acquired immunity in responses to pathogen. Non-human primates such as Chinese Rhesus Macaque (CRM) are the favorable models for preclinical study of potential therapeutic drugs, vaccines and mechanisms of human diseases. However, the phenotypicai characterization of monocyte-derived dendritic cells (MDDCs) from CRM has not been elucidated. Monocytes from CRM were cultured with GM-CSF and IL-4 in RPMI-1640. Six days later, these cells were differentiated with typical dendritical morphology. CDllc and DC-SIGN were highly expressed. The immature MDDCs expressed the low levels of CD25, CD80, CD83, moderate CD40, CD86, and high MHC. After stimulation, the mature MDDCs increased expression of mature molecules CD25 and CD83, co-stimulatory molecules such as CD80, CD86 and CD40, and kept a high level of MHC. The capacity of endocytosis decreased with maturation. The mature MDDCs have strong ability of inducing allogeneic T cell proliferation and producing IL-12. In conclusion, we have characterized the phenotype and ultimate function of MDDCs from CRM for the first time. Cellular & Molecular Immunology. 2009;6(3):159-165.

  11. Novel characterization of monocyte-derived cell populations in the meninges and choroid plexus and their rates of replenishment in bone marrow chimeric mice.

    Science.gov (United States)

    Chinnery, Holly R; Ruitenberg, Marc J; McMenamin, Paul G

    2010-09-01

    The mouse dura mater, pia mater, and choroid plexus contain resident macrophages and dendritic cells (DCs). These cells participate in immune surveillance, phagocytosis of cellular debris, uptake of antigens from the surrounding cerebrospinal fluid and immune regulation in many pathologic processes. We used Cx3cr1 knock-in, CD11c-eYFP transgenic and bone marrow chimeric mice to characterize the phenotype, density and replenishment rate of monocyte-derived cells in the meninges and choroid plexus and to assess the role of the chemokine receptor CX3CR1 on their number and tissue distribution. Iba-1 major histocompatibility complex (MHC) Class II CD169 CD68 macrophages and CD11c putative DCs were identified in meningeal and choroid plexus whole mounts. Comparison of homozygous and heterozygous Cx3cr1 mice did not reveal CX3CR1-dependancy on density, distribution or phenotype of monocyte-derived cells. In turnover studies, wild type lethally irradiated mice were reconstituted with Cx3cr1/-positive bone marrow and were analyzed at 3 days, 1, 2, 4 and 8 weeks after transplantation. There was a rapid replenishment of CX3CR1-positive cells in the dura mater (at 4 weeks) and the choroid plexus was fully reconstituted by 8 weeks. These data provide the foundation for future studies on the role of resident macrophages and DCs in conditions such as meningitis, autoimmune inflammatory disease and in therapies involving irradiation and hematopoietic or stem cell transplantation.

  12. A protocol for generation of clinical grade mRNA-transfected monocyte-derived dendritic cells for cancer vaccines.

    Science.gov (United States)

    Mu, L J; Gaudernack, G; Saebøe-Larssen, S; Hammerstad, H; Tierens, A; Kvalheim, G

    2003-11-01

    With the aim of producing large quantities of mRNA-transfected monocyte-derived dendritic cells (DCs) to be used as cancer vaccines, a new clinical grade procedure has been developed. Peripheral blood mononuclear cells (PBMCs) obtained by leukapheresis were enriched for monocytes by immunomagnetic depletion of CD19+ B cells and CD2+ T cells employing the ISOLEX 300i device. After 5 days of culture of enriched monocytes in gas permeable Teflon bags, using serum-free medium supplemented with granulocyte/macrophage-colony stimulating factor and interleukin-4 (IL-4), immature DCs were generated. Following transfection with mRNA from three human prostate cancer cell lines (DU145, LNCaP and PC-3), employing a newly developed square wave electroporation procedure, the immature DCs were immediately transferred to Teflon bags and matured for 48 h, using serum-free medium supplemented with IL-1alpha, IL-6, tumour necrosis factor-alpha and PGE2. The electroporation procedure efficiently transferred mRNA into the DCs with minor effect on the viability of the cells. The generated matured transfected DCs show high expression of the antigens CD83, CD80, CD86 and human leucocyte antigen-DR. Freezing and thawing of the transfected matured DCs had minor effect on cell viability and the phenotype. From 4 x 109 PBMCs, about 1 x 108 transfected matured DCs are produced. The thawed transfected DCs were able to elicit primary T-cell responses in vitro against antigens encoded by the prostate cancer mRNA as shown by enzyme-linked immunospot assay using mock-transfected DCs as control. Based on these results, clinical trials in cancer patients have been initiated.

  13. Δ(9)-Tetrahydrocannabinol (THC) enhances lipopolysaccharide-stimulated tissue factor in human monocytes and monocyte-derived microvesicles.

    Science.gov (United States)

    Williams, Julie C; Klein, Thomas W; Goldberger, Bruce A; Sleasman, John W; Mackman, Nigel; Goodenow, Maureen M

    2015-01-01

    Immunomodulatory effects in humans of Δ(9-)Tetrahydrocannabinol (THC), the psychoactive component of marijuana are controversial. Tissue factor (TF), the activator of the extrinsic coagulation cascade, is increased on circulating activated monocytes and is expressed on microvesicles released from activated monocytes during inflammatory conditions, which perpetuate coagulopathies in a number of diseases. In view of the increased medicinal use of marijuana, effects of THC on human monocytes and monocyte-derived microvesicles activated by lipopolysaccharide (LPS) were investigated. Peak levels of TF procoagulant activity developed in monocytes or microvesicles 6 h following LPS treatment and were unaltered by THC. After 24 h of LPS stimulation, TF activity declined in control-treated or untreated cells and microvesicles, but persisted with THC treatment. Peak TF protein occurred within 6 h of LPS treatment independent of THC; by 24 h, TF protein declined to almost undetectable levels without THC, but was about 4-fold greater with THC. Steady-state TF mRNA levels were similar up to 2 h in the presence of LPS with or without THC, while 10-fold greater TF mRNA levels persisted over 3-24 h with THC treatment. Activation of MAPK or NF-κB pathways was unaltered by THC treatment and inflammatory cytokine IL-6 levels were unchanged. In contrast, TNF and IL-8 levels were enhanced by 20-50 %. THC enhances TF expression in activated monocytes resulting in elevated procoagulant activity. Marijuana use could potentiate coagulopathies in individuals with chronic immune activation such as HIV-1 infection or inflammatory bowel disease.

  14. Cooperation between Monocyte-Derived Cells and Lymphoid Cells in the Acute Response to a Bacterial Lung Pathogen.

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    Andrew S Brown

    2016-06-01

    Full Text Available Legionella pneumophila is the causative agent of Legionnaires' disease, a potentially fatal lung infection. Alveolar macrophages support intracellular replication of L. pneumophila, however the contributions of other immune cell types to bacterial killing during infection are unclear. Here, we used recently described methods to characterise the major inflammatory cells in lung after acute respiratory infection of mice with L. pneumophila. We observed that the numbers of alveolar macrophages rapidly decreased after infection coincident with a rapid infiltration of the lung by monocyte-derived cells (MC, which, together with neutrophils, became the dominant inflammatory cells associated with the bacteria. Using mice in which the ability of MC to infiltrate tissues is impaired it was found that MC were required for bacterial clearance and were the major source of IL12. IL12 was needed to induce IFNγ production by lymphoid cells including NK cells, memory T cells, NKT cells and γδ T cells. Memory T cells that produced IFNγ appeared to be circulating effector/memory T cells that infiltrated the lung after infection. IFNγ production by memory T cells was stimulated in an antigen-independent fashion and could effectively clear bacteria from the lung indicating that memory T cells are an important contributor to innate bacterial defence. We also determined that a major function of IFNγ was to stimulate bactericidal activity of MC. On the other hand, neutrophils did not require IFNγ to kill bacteria and alveolar macrophages remained poorly bactericidal even in the presence of IFNγ. This work has revealed a cooperative innate immune circuit between lymphoid cells and MC that combats acute L. pneumophila infection and defines a specific role for IFNγ in anti-bacterial immunity.

  15. Cooperation between Monocyte-Derived Cells and Lymphoid Cells in the Acute Response to a Bacterial Lung Pathogen.

    Science.gov (United States)

    Brown, Andrew S; Yang, Chao; Fung, Ka Yee; Bachem, Annabell; Bourges, Dorothée; Bedoui, Sammy; Hartland, Elizabeth L; van Driel, Ian R

    2016-06-01

    Legionella pneumophila is the causative agent of Legionnaires' disease, a potentially fatal lung infection. Alveolar macrophages support intracellular replication of L. pneumophila, however the contributions of other immune cell types to bacterial killing during infection are unclear. Here, we used recently described methods to characterise the major inflammatory cells in lung after acute respiratory infection of mice with L. pneumophila. We observed that the numbers of alveolar macrophages rapidly decreased after infection coincident with a rapid infiltration of the lung by monocyte-derived cells (MC), which, together with neutrophils, became the dominant inflammatory cells associated with the bacteria. Using mice in which the ability of MC to infiltrate tissues is impaired it was found that MC were required for bacterial clearance and were the major source of IL12. IL12 was needed to induce IFNγ production by lymphoid cells including NK cells, memory T cells, NKT cells and γδ T cells. Memory T cells that produced IFNγ appeared to be circulating effector/memory T cells that infiltrated the lung after infection. IFNγ production by memory T cells was stimulated in an antigen-independent fashion and could effectively clear bacteria from the lung indicating that memory T cells are an important contributor to innate bacterial defence. We also determined that a major function of IFNγ was to stimulate bactericidal activity of MC. On the other hand, neutrophils did not require IFNγ to kill bacteria and alveolar macrophages remained poorly bactericidal even in the presence of IFNγ. This work has revealed a cooperative innate immune circuit between lymphoid cells and MC that combats acute L. pneumophila infection and defines a specific role for IFNγ in anti-bacterial immunity.

  16. Immature monocyte derived dendritic cells gene expression profile in response to Virus-Like Particles stimulation

    Directory of Open Access Journals (Sweden)

    Marincola Francesco M

    2005-12-01

    Full Text Available Abstract We have recently developed a candidate HIV-1 vaccine model based on HIV-1 Pr55gag Virus-Like Particles (HIV-VLPs, produced in a baculovirus expression system and presenting a gp120 molecule from an Ugandan HIV-1 isolate of the clade A (HIV-VLPAs. The HIV-VLPAs induce in Balb/c mice systemic and mucosal neutralizing Antibodies as well as cytotoxic T lymphocytes, by intra-peritoneal as well as intra-nasal administration. Moreover, we have recently shown that the baculovirus-expressed HIV-VLPs induce maturation and activation of monocyte-derived dendritic cells (MDDCs which, in turn, produce Th1- and Th2-specific cytokines and stimulate in vitro a primary and secondary response in autologous CD4+ T cells. In the present manuscript, the effects of the baculovirus-expressed HIV-VLPAs on the genomic transcriptional profile of MDDCs obtained from normal healthy donors have been evaluated. The HIV-VLPA stimulation, compared to both PBS and LPS treatment, modulate the expression of genes involved in the morphological and functional changes characterizing the MDDCs activation and maturation. The results of gene profiling analysis here presented are highly informative on the global pattern of gene expression alteration underlying the activation of MDDCs by HIV-VLPAs at the early stages of the immune response and may be extremely helpful for the identification of exclusive activation markers.

  17. PGE2 confers survivin-dependent apoptosis resistance in human monocyte-derived dendritic cells.

    Science.gov (United States)

    Baratelli, Felicita; Krysan, Kostyantyn; Heuzé-Vourc'h, Nathalie; Zhu, Li; Escuadro, Brian; Sharma, Sherven; Reckamp, Karen; Dohadwala, Mariam; Dubinett, Steven M

    2005-08-01

    Control of apoptosis is fundamental for dendritic cell (DC) homeostasis. Numerous factors maintain DC viability throughout their lifespan, including inhibitor of apoptosis proteins. Among them, survivin is overexpressed in many human malignancies, but its physiological function in normal cells has not been fully delineated. Prostaglandin E2 (PGE2), also overproduced in several malignancies, has shown to induce proapoptotic and antiapoptotic effects in different cell types, including immune cells. In DC, PGE2 predominantly affects maturation and modulates immune functions. Here, we show that exposure of monocyte-derived DC to PGE2 (10(-5) M) for 72 h significantly increased DC survivin mRNA and protein expression. In contrast, DC, matured with lipopolysaccharide or tumor necrosis factor alpha, did not reveal survivin induction in response to PGE2. Following exposure to apoptotic stimuli, DC treated with PGE2 exhibited an overall increased viability compared with control DC, and this effect was correlated inversely with caspase-3 activation. Moreover, PGE2-treated, survivin-deficient DC demonstrated reduced viability in response to apoptotic stimuli. Further analysis indicated that PGE2 induced DC survivin expression in an E prostanoid (EP)2/EP4 receptor and phosphatidylinositol-3 kinase-dependent manner. These findings suggest that PGE2-dependent regulation of survivin is important in modulating apoptosis resistance in human DC.

  18. Environmentally relevant dose of arsenic interferes in functions of human monocytes derived dendritic cells.

    Science.gov (United States)

    Bahari, Abbas; Salmani, Vahid

    2017-06-05

    Arsenic is a major environmental pollutant and highly hazardous toxin to human health, which well established as carcinogen and immune deregulatory properties. Dendritic cells (DCs) have a pivotal role in cell-mediated immunity for T-cell activation and antigen presentation. In this study, T cell activation, some key functional genes expression, cell stability and phagocytosis capacity of human monocytes derived DCs (MDDCs) were analyzed after in vitro exposure to very low dose of arsenic for 12 and 24h. Arsenic decreased continually phagocytosis capacity of MDDCs. Furthermore, down-regulation of the cell-surface expression of the co-stimulatory molecule CD40 after 24h post treatment with arsenic, confirmed arsenic interferers in the phagocytosis process. Pro inflammatory cytokines, IL1β and TNFα were more expressed in arsenic-treated MDDCs while IL6 transiently was down regulated. In general, our novel findings here strongly suggest that low level of arsenic dysregulates four fundamental immune processes of DCs. Mechanistically; this could explain the observed immunodeficiency activity of Arsenic, and give direction for comprehension the pathogenesis of Arsenic-induced diseases. Copyright © 2017. Published by Elsevier B.V.

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

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    Grob, P.M.; David, E.; Warren, T.C.; DeLeon, R.P.; Farina, P.R.; Homon, C.A. (Boehringer Ingelheim Pharmaceuticals, Inc., Ridgefield, CT (USA))

    1990-05-15

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

  20. Salvianolic acid B suppresses maturation of human monocyte-derived dendritic cells by activating PPARγ

    Science.gov (United States)

    Sun, Aijun; Liu, Hongying; Wang, Shijun; Shi, Dazhuo; Xu, Lei; Cheng, Yong; Wang, Keqiang; Chen, Keji; Zou, Yunzeng; Ge, Junbo

    2011-01-01

    BACKGROUND AND PURPOSE Salvianolic acid B (Sal B), a water-soluble antioxidant derived from a Chinese medicinal herb, is known to be effective in the prevention of atherosclerosis. Here, we tested the hypothesis that the anti-atherosclerotic effect of Sal B might be mediated by suppressing maturation of human monocyte-derived dendritic cells (h-monDC). EXPERIMENTAL APPROACH h-monDC were derived by incubating purified human monocytes with GM-CSF and IL-4. h-monDC were pre-incubated with or without Sal B and stimulated by oxidized low-density lipoprotein (ox-LDL) in the presence or absence of PPARγ siRNA. Expression of h-monDC membrane molecules (CD40, CD86, CD1a, HLA-DR) were analysed by FACS, cytokines were measured by elisa and the TLR4-associated signalling pathway was determined by Western blotting. KEY RESULTS Ox-LDL promoted h-monDC maturation, stimulated CD40, CD86, CD1a, HLA-DR expression and IL-12, IL-10, TNF-α production; and up-regulated TLR4 signalling. These effects were inhibited by Sal B. Sal B also triggered PPARγ activation and promoted PPARγ nuclear translocation, attenuated ox-LDL-induced up-regulation of TLR4 and myeloid differentiation primary-response protein 88 and inhibited the downstream p38-MAPK signalling cascade. Knocking down PPARγ with the corresponding siRNA blocked these effects of Sal B. CONCLUSIONS AND IMPLICATIONS Our data suggested that Sal B effectively suppressed maturation of h-monDC induced by ox-LDL through PPARγ activation. PMID:21649636

  1. Leukoreduction system chambers are an efficient, valid, and economic source of functional monocyte-derived dendritic cells and lymphocytes.

    Science.gov (United States)

    Pfeiffer, Isabell A; Zinser, Elisabeth; Strasser, Erwin; Stein, Marcello F; Dörrie, Jan; Schaft, Niels; Steinkasserer, Alexander; Knippertz, Ilka

    2013-11-01

    The demand for human monocyte-derived dendritic cells (moDCs), as well as for primary human B and T lymphocytes for immunological research purposes has been increased in recent years. Classically, these monocytes are isolated from blood, leukapheresis products or buffy coats of healthy donors by plastic adherence of peripheral blood mononuclear cells (PBMCs), followed by stimulation with granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4, while lymphocytes are usually isolated from the non-adherent fraction (NAF) by magnetic cell sorting. However, donor-blood is a limited resource and not every blood bank offers leukapheresis products or buffy coats for laboratory use. Additionally, a leukapheresis is very expensive and also the generation/isolation of cells is time- and cost-intensive. To overcome some of these obstacles, we evaluated if low-cost leukoreduction system chambers (LRSCs), which arise after routine donor plateletpheresis procedures, and are usually discarded, would be an alternative and appropriate source of PBMCs to generate moDCs and to isolate lymphocytes. By analyzing the number and phenotype of immature and mature dendritic cells (DCs), as well as of B and T lymphocytes derived from LRSCs, we found all cells to be of high quantity and quality. Further investigations on DCs comprising transwell migration assays, allogeneic mixed lymphocyte reactions (MLR), cytokine secretion assays, and cytotoxic T cell induction assays revealed high migratory, as well as stimulatory capacity of these cells. In addition, DCs and T cells were efficiently electroporated with mRNA and showed characteristic cytokine production after co-culture, demonstrating LRSCs as an efficient, valid, and economic source for generation of moDCs and lymphocytes for research purposes.

  2. Alcohol and cannabinoids differentially affect HIV infection and function of human monocyte-derived dendritic cells (MDDC

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    Marisela eAgudelo

    2015-12-01

    Full Text Available During human immunodeficiency virus (HIV infection, alcohol has been known to induce inflammation while cannabinoids have been shown to have an anti-inflammatory role. For instance cannabinoids have been shown to reduce susceptibility to HIV-1 infection and attenuate HIV replication in macrophages. Recently, we demonstrated that alcohol induces cannabinoid receptors and regulates cytokine production by monocyte-derived dendritic cells (MDDC. However, the ability of alcohol and cannabinoids to alter MDDC function during HIV infection has not been clearly elucidated yet. In order to study the potential impact of alcohol and cannabinoids on differentiated MDDC infected with HIV, monocytes were cultured for 7 days with GM-CSF and IL-4, differentiated MDDC were infected with HIV-1Ba-L and treated with EtOH (0.1 and 0.2%, THC (5 and 10 uM, or JWH-015 (5 and 10 uM for 4-7 days. HIV infection of MDDC was confirmed by p24 and Long Terminal Repeats (LTR estimation. MDDC endocytosis assay and cytokine array profiles were measured to investigate the effects of HIV and substances of abuse on MDDC function. Our results show the HIV+EtOH treated MDDC had the highest levels of p24 production and expression when compared with the HIV positive controls and the cannabinoid treated cells. Although both cannabinoids, THC and JWH-015 had lower levels of p24 production and expression, the HIV+JWH-015 treated MDDC had the lowest levels of p24 when compared to the HIV+THC treated cells. In addition, MDDC endocytic function and cytokine production were also differentially altered after alcohol and cannabinoid treatments. Our results show a differential effect of alcohol and cannabinoids, which may provide insights into the divergent inflammatory role of alcohol and cannabinoids to modulate MDDC function in the context of HIV infection.

  3. Evaluating the Effects of Cytomegalovirus Glycoprotein B on the Maturation and Function of Monocyte-derived dendritic cells

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    Afsson shariat

    2015-11-01

    Full Text Available Background & Objectives: Interaction of cytomegalovirus glycoprotein B with toll-like receptors of dendritic cells leads to early signaling and innate immune responses. The aim of this study is to evaluate the effects of cytomegalovirus glycoprotein B on the maturation and function of monocyte-derived dendritic cells in treated groups in comparison with control groups. Materials & Methods: Blood samples were taken from 5 healthy volunteers. Following the generation of monocyte-derived dendritic cells on the fifth day of cell culture, half of the immature dendritic cells were treated with cytomegalovirus glycoprotein B, and the rest of them were induced to mature dendritic untreated cells and were used as the control group. The maturation and function of dendritic cells were evaluated in these two groups. Results: The gene expression level of toll-like receptor-4 significantly increased in the group treated with glycoprotein B (p < 0.05, whereas there were no significant differences in the expression rates of CD83, CD86, CD1a, and HLA-DR and the secretion of IL-23 from monocyte-derived dendritic cells between the treated groups and the controls. Conclusion: The increase in the gene expression of toll-like receptor-4 in monocyte-derived dendritic cells treated with cytomegalovirus glycoprotein B showed that cell contact is required to elicit cellular antiviral response and toll-like receptor activation. Thus, it is critical to recognize the viral and cellular determinants of the immune system in order to develop new therapeutic strategies against cytomegalovirus.

  4. Expression and regulation of Schlafen (SLFN family members in primary human monocytes, monocyte-derived dendritic cells and T cells

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    Alexander Puck

    2015-01-01

    Full Text Available Schlafen (SLFN/Slfn family members have been investigated for their involvement in fundamental cellular processes including growth regulation, differentiation and control of viral replication. However, most research has been focused on the characterization of Slfns within the murine system or in human cell lines. Since little is known about SLFNs in primary human immune cells, we set out to analyze the expression and regulation of the six human SLFN genes in monocytes, monocyte-derived dendritic cells (moDCs and T cells. Comparison of SLFN gene expression across these three cell types showed high mRNA expression of SLFN11 in monocytes and moDCs and high SLFN5 expression in T cells, indicating functional importance within these cell types. Differentiation of monocytes to moDCs leads to the gradual upregulation of SLFN12L and SLFN13 while SLFN12 levels were decreased by differentiation stimuli. Stimulation of moDCs via human rhinovirus, lipopolysaccharide, or IFN-α lead to strong upregulation of SLFN gene expression, while peptidoglycan poorly stimulated regulation of both SLFNs and the classical interferon-stimulated gene MxA. T cell activation was found to downregulate the expression of SLFN5, SLFN12 and SLFN12L, which was reversible upon addition of exogenous IFN-α. In conclusion, we demonstrate, that SLFN gene upregulation is mainly dependent on autocrine type I interferon signaling in primary human immune cells. Rapid decrease of SLFN expression levels following T cell receptor stimulation indicates a role of SLFNs in the regulation of human T cell quiescence.

  5. Leukotrienes inhibit early stages of HIV-1 infection in monocyte-derived microglia-like cells

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    Bertin Jonathan

    2012-03-01

    Full Text Available Abstract Background Microglia are one of the main cell types to be productively infected by HIV-1 in the central nervous system (CNS. Leukotriene B4 (LTB4 and cysteinyl-leukotrienes such as LTC4 are some of the proinflammatory molecules produced in infected individuals that contribute to neuroinflammation. We therefore sought to investigate the role of leukotrienes (LTs in HIV-1 infection of microglial cells. Methods To evaluate the role of LTs on HIV-1 infection in the CNS, monocyte-derived microglial-like cells (MDMis were utilized in this study. Leukotriene-treated MDMis were infected with either fully replicative brain-derived HIV-1 isolates (YU2 or R5-tropic luciferase-encoding particles in order to assess viral production and expression. The efficacy of various steps of the replication cycle was evaluated by means of p24 quantification by ELISA, luciferase activity determination and quantitative real-time polymerase chain reaction (RT-PCR. Results We report in this study that virus replication is reduced upon treatment of MDMis with LTB4 and LTC4. Additional experiments indicate that these proinflammatory molecules alter the pH-independent entry and early post-fusion events of the viral life cycle. Indeed, LT treatment induced a diminution in integrated proviral DNA while reverse-transcribed viral products remained unaffected. Furthermore, decreased C-C chemokine receptor type 5 (CCR5 surface expression was observed in LT-treated MDMis. Finally, the effect of LTs on HIV-1 infection in MDMis appears to be mediated partly via a signal transduction pathway involving protein kinase C. Conclusions These data show for the first time that LTs influence microglial cell infection by HIV-1, and may be a factor in the control of viral load in the CNS.

  6. Gene expression profiling of human monocyte-derived dendritic cells - Searching for molecular regulators of tolerogenicity

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    Katina eSchinnerling

    2015-10-01

    Full Text Available The ability of dendritic cells (DCs to initiate and modulate antigen-specific immune responses has made them attractive targets for immunotherapy. Since DC research in humans is limited by the scarcity of DC populations in the blood circulation, most of our knowledge about DC biology and function has been obtained in vitro from monocyte-derived DCs (moDCs, which can be readily generated in sufficient numbers and are able to differentiate into distinct functional subsets depending on the nature of stimulus. In particular, moDCs with tolerogenic properties (tolDCs possess great therapeutic potential for the treatment of autoimmune diseases. Several protocols have been developed to generate tolDCs in vitro, able to reinstruct auto-reactive T cells and to promote regulatory cells. While ligands and soluble mediators, by which DCs shape immune responses, have been vastly studied, the intracellular pathways and transcriptional regulators that govern tolDC differentiation and function are poorly understood. Whole-genome microarrays and proteomics provide useful strategies to dissect the complex molecular processes that promote tolerogenicity. Only few attempts have been made to understand tolDC biology through a global view on ‘omics’ profiles. So far, the identification of a common regulator of tolerogenicity has been hampered by the fact that each protocol, used for tolDC generation, targets distinct signaling pathways. Here we review the progress in understanding the transcriptional regulation of moDC differentiation, with a special focus on tolDCs, and highlight candidate molecules that might be associated with DC tolerogenicity.

  7. Leukotrienes inhibit early stages of HIV-1 infection in monocyte-derived microglia-like cells.

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    Bertin, Jonathan; Barat, Corinne; Bélanger, Dave; Tremblay, Michel J

    2012-03-16

    Microglia are one of the main cell types to be productively infected by HIV-1 in the central nervous system (CNS). Leukotriene B4 (LTB4) and cysteinyl-leukotrienes such as LTC4 are some of the proinflammatory molecules produced in infected individuals that contribute to neuroinflammation. We therefore sought to investigate the role of leukotrienes (LTs) in HIV-1 infection of microglial cells. To evaluate the role of LTs on HIV-1 infection in the CNS, monocyte-derived microglial-like cells (MDMis) were utilized in this study. Leukotriene-treated MDMis were infected with either fully replicative brain-derived HIV-1 isolates (YU2) or R5-tropic luciferase-encoding particles in order to assess viral production and expression. The efficacy of various steps of the replication cycle was evaluated by means of p24 quantification by ELISA, luciferase activity determination and quantitative real-time polymerase chain reaction (RT-PCR). We report in this study that virus replication is reduced upon treatment of MDMis with LTB4 and LTC4. Additional experiments indicate that these proinflammatory molecules alter the pH-independent entry and early post-fusion events of the viral life cycle. Indeed, LT treatment induced a diminution in integrated proviral DNA while reverse-transcribed viral products remained unaffected. Furthermore, decreased C-C chemokine receptor type 5 (CCR5) surface expression was observed in LT-treated MDMis. Finally, the effect of LTs on HIV-1 infection in MDMis appears to be mediated partly via a signal transduction pathway involving protein kinase C. These data show for the first time that LTs influence microglial cell infection by HIV-1, and may be a factor in the control of viral load in the CNS.

  8. Staphylococcus aureus from atopic dermatitis skin alters cytokine production triggered by monocyte-derived Langerhans cell.

    Science.gov (United States)

    Iwamoto, Kazumasa; Moriwaki, Masaya; Niitsu, Yoshie; Saino, Masachika; Takahagi, Shunsuke; Hisatsune, Junzo; Sugai, Motoyuki; Hide, Michihiro

    2017-08-05

    Atopic dermatitis (AD) is one of the most common chronic inflammatory skin diseases. The skin of patients with AD presents as a disbalance of the microbiome with a strong colonization by Staphylococcus aureus, which positively correlates with the severity of the disease. However, the effect of colonized S. aureus on the skin immune system has not been fully elucidated. The aim of this study is to explore whether S. aureus isolated from AD skin is able to skew T cell responses via Langerhans cells (LC) as compared to a standard strain of S. aureus and S. epidermidis. We prepared monocyte-derived LC (MoLC) from healthy controls and patients with AD, and stimulated MoLC with a standard strain of S. aureus NCTC8325, S. aureus TF3378 isolated from AD skin, or S. epidermidis. Stimulated MoLC were co-cultured with autologous CD4(pos) T cells and then T cell responses were analyzed by T cell polarization assays, cytokine analysis and real-time PCR. MoLC stimulated by S. aureus TF3378 induced significantly high and rapid proliferation of T cells as compared to those by S. aureus NCTC8325 and S. epidermidis. Cytokine productions from T cells cultured with S. aureus TF3378-stimulated MoLC showed significantly high amounts of IL-2 and less IFN-γ production with imbalanced Th1/Th2 (decreased TBX21/GATA3 ratio) mRNA expression. The T cell proliferation with increased IL-2 production via S. aureus TF3378-stimulated MoLC was diminished by treatment of proteinase K. S. aureus TF3378 on AD skin can skew T cell responses via LC toward imbalanced Th1/Th2 skin immunity. Copyright © 2017 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.

  9. Neutrophil extracellular traps downregulate lipopolysaccharide-induced activation of monocyte-derived dendritic cells.

    Science.gov (United States)

    Barrientos, Lorena; Bignon, Alexandre; Gueguen, Claire; de Chaisemartin, Luc; Gorges, Roseline; Sandré, Catherine; Mascarell, Laurent; Balabanian, Karl; Kerdine-Römer, Saadia; Pallardy, Marc; Marin-Esteban, Viviana; Chollet-Martin, Sylvie

    2014-12-01

    Polymorphonuclear neutrophils (PMN) play a central role in inflammation and participate in its control, notably by modulating dendritic cell (DC) functions via soluble mediators or cell-cell contacts. Neutrophil extracellular traps (NETs) released by PMN could play a role in this context. To evaluate NET effects on DC maturation, we developed a model based on monocyte-derived DC (moDC) and calibrated NETs isolated from fresh human PMN. We found that isolated NETs alone had no discernable effect on moDC. In contrast, they downregulated LPS-induced moDC maturation, as shown by decreased surface expression of HLA-DR, CD80, CD83, and CD86, and by downregulated cytokine production (TNF-α, IL-6, IL-12, IL-23), with no increase in the expression of tolerogenic DC genes. Moreover, the presence of NETs during moDC maturation diminished the capacity of these moDC to induce T lymphocyte proliferation in both autologous and allogeneic conditions, and modulated CD4(+) T lymphocyte polarization by promoting the production of Th2 cytokines (IL-5 and IL-13) and reducing that of Th1 and Th17 cytokines (IFN-γ and IL-17). Interestingly, the expression and activities of the lymphoid chemokine receptors CCR7 and CXCR4 on moDC were not altered when moDC matured in the presence of NETs. Together, these findings reveal a new role for NETs in adaptive immune responses, modulating some moDC functions and thereby participating in the control of inflammation.

  10. HIV infection of monocytes-derived dendritic cells inhibits Vγ9Vδ2 T cells functions.

    Directory of Open Access Journals (Sweden)

    Alessandra Sacchi

    Full Text Available DCs act as sentinel cells against incoming pathogens and represent the most potent antigen presenting cells, having the unique capability to prime naïve T cells. In addition to their role in induction of adaptive immune responses, DC are also able to activate innate cells as γδ T cells; in particular, a reciprocal crosstalk between DC and γδ T cells was demonstrated. However, whether HIV infection may alter DC-Vγ9Vδ2 T cells cross-talk was not yet described. To clarify this issue, we cultured activated Vγ9Vδ2 T cells with HIV infected monocyte derived DC (MoDC. After 5 days we evaluated MoDC phenotype, and Vγ9Vδ2 T cells activation and proliferation. In our model, Vγ9Vδ2 T cells were not able to proliferate in response to HIV-infected MoDC, although an up-regulation of CD69 was observed. Upon phosphoantigens stimulation, Vγ9Vδ2 T cells proliferation and cytokine production were inhibited when cultured with HIV-infected MoDC in a cell-contact dependent way. Moreover, HIV-infected MoDC are not able to up-regulate CD86 molecules when cultured with activated Vγ9Vδ2 T cells, compared with uninfected MoDC. Further, activated Vγ9Vδ2 T cells are not able to induce HLA DR up-regulation and CCR5 down-regulation on HIV-infected MoDC. These data indicate that HIV-infected DC alter the capacity of Vγ9Vδ2 T cells to respond to their antigens, pointing out a new mechanisms of induction of Vγ9Vδ2 T cells anergy carried out by HIV, that could contribute to immune evasion.

  11. HIV infection of monocytes-derived dendritic cells inhibits Vγ9Vδ2 T cells functions.

    Science.gov (United States)

    Sacchi, Alessandra; Rinaldi, Alessandra; Tumino, Nicola; Casetti, Rita; Agrati, Chiara; Turchi, Federica; Bordoni, Veronica; Cimini, Eleonora; Martini, Federico

    2014-01-01

    DCs act as sentinel cells against incoming pathogens and represent the most potent antigen presenting cells, having the unique capability to prime naïve T cells. In addition to their role in induction of adaptive immune responses, DC are also able to activate innate cells as γδ T cells; in particular, a reciprocal crosstalk between DC and γδ T cells was demonstrated. However, whether HIV infection may alter DC-Vγ9Vδ2 T cells cross-talk was not yet described. To clarify this issue, we cultured activated Vγ9Vδ2 T cells with HIV infected monocyte derived DC (MoDC). After 5 days we evaluated MoDC phenotype, and Vγ9Vδ2 T cells activation and proliferation. In our model, Vγ9Vδ2 T cells were not able to proliferate in response to HIV-infected MoDC, although an up-regulation of CD69 was observed. Upon phosphoantigens stimulation, Vγ9Vδ2 T cells proliferation and cytokine production were inhibited when cultured with HIV-infected MoDC in a cell-contact dependent way. Moreover, HIV-infected MoDC are not able to up-regulate CD86 molecules when cultured with activated Vγ9Vδ2 T cells, compared with uninfected MoDC. Further, activated Vγ9Vδ2 T cells are not able to induce HLA DR up-regulation and CCR5 down-regulation on HIV-infected MoDC. These data indicate that HIV-infected DC alter the capacity of Vγ9Vδ2 T cells to respond to their antigens, pointing out a new mechanisms of induction of Vγ9Vδ2 T cells anergy carried out by HIV, that could contribute to immune evasion.

  12. SLAM/SLAM interactions inhibit CD40-induced production of inflammatory cytokines in monocyte-derived dendritic cells.

    Science.gov (United States)

    Réthi, Bence; Gogolák, Péter; Szatmari, Istvan; Veres, Agota; Erdôs, Erika; Nagy, Laszlo; Rajnavölgyi, Eva; Terhorst, Cox; Lányi, Arpád

    2006-04-01

    Signaling lymphocyte activation molecule (SLAM, CD150, or SLAMF1) is a self-ligand receptor on the surface of activated T- and B-lymphocytes, macrophages, and dendritic cells (DCs). Here we examine the effect of SLAM/SLAM interactions on CD40L-induced CD40 signaling pathways in human DCs. CD40L-expressing L929 cells induced DCs to produce interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and IL-12, which was strongly inhibited by coexpression of SLAM on the surface of the L929 cells. Similarly, transfection of DCs with SLAM strongly reduced CD40L-induced IL-12 production. Furthermore, the negative effect of SLAM/SLAM interactions on CD40L-induced DC activation was also detected in the presence of lipopolysaccharide (LPS). LPS-induced IL-12 secretion, however, was not inhibited by SLAM engagement. CD40L-activated DCs affected by exposure to SLAM/SLAM engagement were impaired in their ability to induce differentiation of naive T lymphocytes into interferon-gamma (IFN-gamma)-producing T-helper 1 (Th1) effector cells. These inhibitory effects were not the result of a general unresponsiveness of DCs to CD40L, as SLAM/SLAM interactions did not prevent CD40L-induced up-regulation of CD83, CD86, or human leukocyte antigen (HLA)-DQ on the surface of DCs. Taken together, the results indicate that SLAM/SLAM interactions inhibit CD40-induced signal transduction in monocyte-derived dendritic cells, an effect that was not detectable in earlier studies using anti-SLAM monoclonal antibodies.

  13. Comparative analysis of signature genes in PRRSV-infected porcine monocyte-derived dendritic cells at differential activation statuses

    Science.gov (United States)

    Activation statuses of monocytic cells including monocytes, macrophages and dendritic cells (DCs) are critically important for antiviral immunity. In particular, some devastating viruses, including porcine reproductive and respiratory syndrome virus (PRRSV), are capable of directly infecting these c...

  14. Characterization of monocyte-derived dendritic cells maturated with IFN-alpha

    DEFF Research Database (Denmark)

    Svane, I M; Nikolajsen, K; Walter, M R;

    2006-01-01

    Dendritic cells (DC) are promising candidates for cancer immunotherapy. These cells can be generated from peripheral blood monocytes cultured with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). In order to obtain full functional capacity, maturation is required......, maturation with IFN-alpha has only a small effect on induction of autologous T-cell stimulatory capacity of the DC. However, an increase in DC allogeneic T-cell stimulatory capacity was observed. These data suggest that IFN-alpha has a potential as a maturation agent used in DC-based cancer vaccine trials...

  15. Monocyte and Macrophage Plasticity in Tissue Repair and Regeneration

    Science.gov (United States)

    Das, Amitava; Sinha, Mithun; Datta, Soma; Abas, Motaz; Chaffee, Scott; Sen, Chandan K.; Roy, Sashwati

    2016-01-01

    Heterogeneity and high versatility are the characteristic features of the cells of monocyte-macrophage lineage. The mononuclear phagocyte system, derived from the bone marrow progenitor cells, is primarily composed of monocytes, macrophages, and dendritic cells. In regenerative tissues, a central role of monocyte-derived macrophages and paracrine factors secreted by these cells is indisputable. Macrophages are highly plastic cells. On the basis of environmental cues and molecular mediators, these cells differentiate to proinflammatory type I macrophage (M1) or anti-inflammatory or proreparative type II macrophage (M2) phenotypes and transdifferentiate into other cell types. Given a central role in tissue repair and regeneration, the review focuses on the heterogeneity of monocytes and macrophages with current known mechanisms of differentiation and plasticity, including microenvironmental cues and molecular mediators, such as noncoding RNAs. PMID:26118749

  16. Host hindrance to HIV-1 replication in monocytes and macrophages

    Directory of Open Access Journals (Sweden)

    Pancino Gianfranco

    2010-04-01

    Full Text Available Abstract Monocytes and macrophages are targets of HIV-1 infection and play critical roles in multiple aspects of viral pathogenesis. HIV-1 can replicate in blood monocytes, although only a minor proportion of circulating monocytes harbor viral DNA. Resident macrophages in tissues can be infected and function as viral reservoirs. However, their susceptibility to infection, and their capacity to actively replicate the virus, varies greatly depending on the tissue localization and cytokine environment. The susceptibility of monocytes to HIV-1 infection in vitro depends on their differentiation status. Monocytes are refractory to infection and become permissive upon differentiation into macrophages. In addition, the capacity of monocyte-derived macrophages to sustain viral replication varies between individuals. Host determinants regulate HIV-1 replication in monocytes and macrophages, limiting several steps of the viral life-cycle, from viral entry to virus release. Some host factors responsible for HIV-1 restriction are shared with T lymphocytes, but several anti-viral mechanisms are specific to either monocytes or macrophages. Whilst a number of these mechanisms have been identified in monocytes or in monocyte-derived macrophages in vitro, some of them have also been implicated in the regulation of HIV-1 infection in vivo, in particular in the brain and the lung where macrophages are the main cell type infected by HIV-1. This review focuses on cellular factors that have been reported to interfere with HIV-1 infection in monocytes and macrophages, and examines the evidences supporting their role in vivo, highlighting unique aspects of HIV-1 restriction in these two cell types.

  17. iNKT Cell Emigration out of the Lung Vasculature Requires Neutrophils and Monocyte-Derived Dendritic Cells in Inflammation

    Science.gov (United States)

    Thanabalasuriar, A; Neupane, A.S; Wang, J; Krummel, M.F; Kubes, P

    2017-01-01

    iNKT cells are a subset of innate T cells that recognize glycolipids presented on CD1d molecules and protect against a variety of bacterial infections including S. pneumoniae. Using lung intravital imaging, we examined the behavior and mechanism of pulmonary iNKT cell activation in response to the potent iNKT cell ligand α-galactosylceramide or during S. pneumoniae infection. In untreated mice the major fraction of iNKT cells resided in the vasculature, but a small critical population resided in the extravascular space in proximity to monocyte-derived DCs. Administration of either α-GalCer or S. pneumoniae, induced CD1d dependent rapid recruitment of neutrophils out of the vasculature. This neutrophil exodus paved the way for extravasation of iNKT cells from the lung vasculature via CCL17. Depletion of monocyte-derived DCs abrogated both the neutrophil and subsequent iNKT cell extravasation. Moreover, impairing iNKT cell migration out of the lung vasculature by blocking CCL17 greatly increased susceptibility to S. pneumoniae infection, suggesting a critical role for the secondary wave of iNKT cells in host defense. PMID:27653688

  18. Evidence for a dual function of monocyte-derived mononuclear phagocytes during chronic intestinal inflammation

    DEFF Research Database (Denmark)

    Rivollier, Aymeric Marie Christian; Pool, Lieneke; Frising, Ulrika

    Mononuclear phagocytes derived from tissue-infiltrating monocytes play diverse roles in immunity, ranging from pathogen killing to immune regulation. We and others showed that, upon recruitment to the intestinal mucosa, the differentiation of Ly6Chi monocytes into phagocytes with anti- versus pro...... suggest a dual and time-restricted contribution of MDP during the development and healing phases of the disease....

  19. Chemoresistance of human monocyte-derived dendritic cells is regulated by IL-17A.

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    Selma Olsson Åkefeldt

    Full Text Available Dendritic cells initiate adaptive immune responses, leading either to control cancer by effector T cells or to exacerbate cancer by regulatory T cells that inhibit IFN-γ-mediated Th1-type response. Dendritic cells can also induce Th17-type immunity, mediated by IL-17A. However, the controversial role of this cytokine in cancer requires further investigations. We generated dendritic cells from peripheral blood monocytes to investigate lifespan, phenotype and chemoresistance of dendritic cells, treated with IL-17A with or without IFN-γ. Studying the expression of Bcl-2 family members, we demonstrated that dendritic cells constitutively express one pro-survival Bcl-2 member: MCL1. Immature dendritic cells were CD40(lowHLADR(low CD1a(+ MCL1(+, did not express CD14, CD68 or BCL2A1, and displayed a short 2-day lifespan. IL-17A-treated DC exhibited a semi-mature (CD40(high HLADR(low pre-M2 (CCL22(+ CD206(+ CD163(+ IL1RN(+ IL-10(- CXCL10(- IL-12(- mixed (CD1a(+ CD14+ CD68(+ macrophage-dendritic cell phenotype. They efficiently exerted mannose receptor-mediated endocytosis and did not produce superoxide anions, in the absence of TLR engagement. Interestingly, IL-17A promoted a long-term survival of dendritic cells, beyond 12 days, that correlated to BCL2A1 induction, a pro-survival Bcl-2 family member. BCL2A1 transcription was activated by NF-κB, downstream of IL-17A transduction. Thus, immature dendritic cells only express MCL1, whereas IL-17A-treated dendritic cells concomitantly expressed two pro-survival Bcl-2 family members: MCL1 and BCL2A1. These latter developed chemoresistance to 11 of the 17 chemotherapy agents tested. However, high doses of either vinblastine or cytarabine decreased MCL1 expression and induced dendritic cell death. When IL-17A is produced in vivo, administration of anti-IL-17A biotherapy may impair dendritic cell survival by targeting BCL2A1 expression. Consequently, depending on the effector or regulatory role of dendritic

  20. The effect of cytosolic extract of Alternaria aternata fungus on Monocyte-derived dendritic cell maturation and T-lymphocyte polarization in the presence of myelin basic protein

    Directory of Open Access Journals (Sweden)

    Loghmanni A

    2013-03-01

    Full Text Available Background: Multiple Sclerosis (MS is an autoimmune disease with impairment in function of central nervous system. Macrophages and dendritic cells play important roles in alleviating or progression of the disease. These cells can cause inflammation and damage to the myelin of nerve cells by realizing of harmful substances when these cells get matured. We studied the effect of Alternaria alternata extract on maturation of monocyte- derived dendritic cell (modc and T-cell responses in the presence of Myelin Basic Protein (MBP as a laboratory model of multiple sclerosis (MS. The purpose of this study is suitable dendritic cells production for usage in MS immunotherapy.Methods: For this study plastic adherent monocytes were cultured with granulocyte/ macrophage- colony stimulating factor (GM-CSF and interleukin -4 for converting these cells to modc and pulsed with MBP and matured in the presence of monocyte-conditioned medium (MCM in control group and MCM + Alternaria alternata extract in treatment groups. Anti-CD14, anti-CD83, anti-human leukocyte antigen-DR (anti HLA-DR monoclonal antibody were carried out for phenotyping. Autologos T cell responses and cytokine production were evaluated.Results: The results showed that the expression of CD14 decreased and CD83, HLA-DR increased in treatment groups in comparison with control groups. The production amount of IL-10 overcame IL-12 and in T cell the production of cytokines, IL-17 and Interferon-γ (IFN-γ decreased and IL-4 was increased (P<0.05. These effects escalated with increasing of dosage from 50 to 100 (mg/ml (P<0.001.Conclusion: Alternaria alternata extract can cause maturation of MBP-pulsed modc and skewing of T- lymphocyte toward Th2 and thereby can evolve into a new strategy in immunotherapy of MS.

  1. Prophylactic vaccines are potent activators of monocyte-derived dendritic cells and drive effective anti-tumor responses in melanoma patients at the cost of toxicity

    NARCIS (Netherlands)

    Bol, K.F.; Aarntzen, E.H.J.G.; Pots, J.M.; Olde Nordkamp, M.A.M.; Rakt, M.W.M.M. van de; Scharenborg, N.M.; Boer, A.J. de; Oorschot, T.G.M. van; Croockewit, S.; Blokx, W.A.M.; Oyen, W.J.G.; Boerman, O.C.; Mus, R.D.M.; Rossum, M.M. van; Graaf, C.A.A. van der; Punt, C.J.; Adema, G.J.; Figdor, C.G.; Vries, I.J. de; Schreibelt, G.

    2016-01-01

    Dendritic cell (DC)-based immunotherapy is explored worldwide in cancer patients, predominantly with DC matured with pro-inflammatory cytokines and prostaglandin E2. We studied the safety and efficacy of vaccination with monocyte-derived DC matured with a cocktail of prophylactic vaccines that

  2. SMAC Mimetic BV6 Induces Cell Death in Monocytes and Maturation of Monocyte-Derived Dendritic Cells

    Science.gov (United States)

    Holtz, Philipp; Kapp, Markus; Grigoleit, Götz Ulrich; Schmuck, Carsten; Wajant, Harald; Siegmund, Daniela

    2011-01-01

    Background Compounds mimicking the inhibitory effect of SMAC / DIABLO on X-linked inhibitor of apoptosis (XIAP) have been developed with the aim to achieve sensitization for apoptosis of tumor cells resistant due to deregulated XIAP expression. It turned out that SMAC mimetics also have complex effects on the NFκB system and TNF signaling. In view of the overwhelming importance of the NFκB transcription factors in the immune system, we analyzed here the effects of the SMAC mimetic BV6 on immune cells. Principal Findings BV6 induced apoptotic and necrotic cell death in monocytes while T-cells, dendritic cells and macrophages were largely protected against BV6-induced cell death. In immature dendritic cells BV6 treatment resulted in moderate activation of the classical NFκB pathway, but it also diminished the stronger NFκB-inducing effect of TNF and CD40L. Despite its inhibitory effect on TNF- and CD40L signaling, BV6 was able to trigger maturation of immature DCs as indicated by upregulation of CD83, CD86 and IL12. Significance The demonstrated effects of SMAC mimetics on immune cells may complicate the development of tumor therapeutic concepts based on these compounds but also arise the possibility to exploit them for the development of immune stimulatory therapies. PMID:21738708

  3. SMAC mimetic BV6 induces cell death in monocytes and maturation of monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Nicole Müller-Sienerth

    Full Text Available BACKGROUND: Compounds mimicking the inhibitory effect of SMAC/DIABLO on X-linked inhibitor of apoptosis (XIAP have been developed with the aim to achieve sensitization for apoptosis of tumor cells resistant due to deregulated XIAP expression. It turned out that SMAC mimetics also have complex effects on the NFκB system and TNF signaling. In view of the overwhelming importance of the NFκB transcription factors in the immune system, we analyzed here the effects of the SMAC mimetic BV6 on immune cells. PRINCIPAL FINDINGS: BV6 induced apoptotic and necrotic cell death in monocytes while T-cells, dendritic cells and macrophages were largely protected against BV6-induced cell death. In immature dendritic cells BV6 treatment resulted in moderate activation of the classical NFκB pathway, but it also diminished the stronger NFκB-inducing effect of TNF and CD40L. Despite its inhibitory effect on TNF- and CD40L signaling, BV6 was able to trigger maturation of immature DCs as indicated by upregulation of CD83, CD86 and IL12. SIGNIFICANCE: The demonstrated effects of SMAC mimetics on immune cells may complicate the development of tumor therapeutic concepts based on these compounds but also arise the possibility to exploit them for the development of immune stimulatory therapies.

  4. Investigating the Role of Surface Materials and Three Dimensional Architecture on In Vitro Differentiation of Porcine Monocyte-Derived Dendritic Cells.

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    Sofie Bruun Hartmann

    Full Text Available In vitro generation of dendritic-like cells through differentiation of peripheral blood monocytes is typically done using two-dimensional polystyrene culture plates. In the process of optimising cell culture techniques, engineers have developed fluidic micro-devises usually manufactured in materials other than polystyrene and applying three-dimensional structures more similar to the in vivo environment. Polydimethylsiloxane (PDMS is an often used polymer for lab-on-a-chip devices but not much is known about the effect of changing the culture surface material from polystyrene to PDMS. In the present study the differentiation of porcine monocytes to monocyte-derived dendritic cells (moDCs was investigated using CD172apos pig blood monocytes stimulated with GM-CSF and IL-4. Monocytes were cultured on surfaces made of two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS and carbonised three-dimensional PDMS. Cells cultured conventionally (on two-dimensional polystyrene differentiated into moDCs as expected. Interestingly, gene expression of a wide range of cytokines, chemokines, and pattern recognition receptors was influenced by culture surface material and architecture. Distinct clustering of cells, based on similar expression patterns of 46 genes of interest, was seen for cells isolated from two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS. Changing the material from polystyrene to PDMS resulted in cells with expression patterns usually associated with macrophage expression (upregulation of CD163 and downregulation of CD1a, FLT3, LAMP3 and BATF3. However, this was purely based on gene expression level, and no functional assays were included in this study which would be necessary in order to classify the cells as being macrophages. When changing to three-dimensional culture the cells became increasingly activated in terms of IL6, IL8, IL10 and CCR5 gene expression. Further stimulation

  5. Investigating the Role of Surface Materials and Three Dimensional Architecture on In Vitro Differentiation of Porcine Monocyte-Derived Dendritic Cells

    Science.gov (United States)

    Hartmann, Sofie Bruun; Mohanty, Soumyaranjan; Skovgaard, Kerstin; Brogaard, Louise; Flagstad, Frederikke Bjergvang; Emnéus, Jenny; Wolff, Anders; Summerfield, Artur; Jungersen, Gregers

    2016-01-01

    In vitro generation of dendritic-like cells through differentiation of peripheral blood monocytes is typically done using two-dimensional polystyrene culture plates. In the process of optimising cell culture techniques, engineers have developed fluidic micro-devises usually manufactured in materials other than polystyrene and applying three-dimensional structures more similar to the in vivo environment. Polydimethylsiloxane (PDMS) is an often used polymer for lab-on-a-chip devices but not much is known about the effect of changing the culture surface material from polystyrene to PDMS. In the present study the differentiation of porcine monocytes to monocyte-derived dendritic cells (moDCs) was investigated using CD172apos pig blood monocytes stimulated with GM-CSF and IL-4. Monocytes were cultured on surfaces made of two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS and carbonised three-dimensional PDMS. Cells cultured conventionally (on two-dimensional polystyrene) differentiated into moDCs as expected. Interestingly, gene expression of a wide range of cytokines, chemokines, and pattern recognition receptors was influenced by culture surface material and architecture. Distinct clustering of cells, based on similar expression patterns of 46 genes of interest, was seen for cells isolated from two- and three-dimensional polystyrene as well as two- and three-dimensional PDMS. Changing the material from polystyrene to PDMS resulted in cells with expression patterns usually associated with macrophage expression (upregulation of CD163 and downregulation of CD1a, FLT3, LAMP3 and BATF3). However, this was purely based on gene expression level, and no functional assays were included in this study which would be necessary in order to classify the cells as being macrophages. When changing to three-dimensional culture the cells became increasingly activated in terms of IL6, IL8, IL10 and CCR5 gene expression. Further stimulation with LPS resulted

  6. Electroporated Antigen-Encoding mRNA Is Not a Danger Signal to Human Mature Monocyte-Derived Dendritic Cells

    Directory of Open Access Journals (Sweden)

    Stefanie Hoyer

    2015-01-01

    Full Text Available For therapeutic cancer vaccination, the adoptive transfer of mRNA-electroporated dendritic cells (DCs is frequently performed, usually with monocyte-derived, cytokine-matured DCs (moDCs. However, DCs are rich in danger-sensing receptors which could recognize the exogenously delivered mRNA and induce DC activation, hence influencing the DCs’ immunogenicity. Therefore, we examined whether electroporation of mRNA with a proper cap and a poly-A tail of at least 64 adenosines had any influence on cocktail-matured moDCs. We used 16 different RNAs, encoding tumor antigens (MelanA, NRAS, BRAF, GNAQ, GNA11, and WT1, and variants thereof. None of those RNAs induced changes in the expression of CD25, CD40, CD83, CD86, and CD70 or the secretion of the cytokines IL-8, IL-6, and TNFα of more than 1.5-fold compared to the control condition, while an mRNA encoding an NF-κB-activation protein as positive control induced massive secretion of the cytokines. To determine whether mRNA electroporation had any effect on the whole transcriptome of the DCs, we performed microarray analyses of DCs of 6 different donors. None of 60,000 probes was significantly different between mock-electroporated DCs and MelanA-transfected DCs. Hence, we conclude that no transcriptional programs were induced within cocktail-matured DCs by electroporation of single tumor-antigen-encoding mRNAs.

  7. In vitro detection of contact allergens: development of an optimized protocol using human peripheral blood monocyte-derived dendritic cells.

    Science.gov (United States)

    Reuter, Hendrik; Spieker, Jochem; Gerlach, Silke; Engels, Ursula; Pape, Wolfgang; Kolbe, Ludger; Schmucker, Robert; Wenck, Horst; Diembeck, Walter; Wittern, Klaus-Peter; Reisinger, Kerstin; Schepky, Andreas G

    2011-02-01

    Allergic contact dermatitis is a delayed T-cell mediated allergic response associated with relevant social and economic impacts. Animal experiments (e.g. the local lymph node assay) are still supplying most of the data used to assess the sensitization potential of new chemicals. However, the 7th amendment to the EU Cosmetic Directive will introduce a testing ban for cosmetic ingredients after 2013. In vitro alternative methods are thus being actively developed. Although promising results have been obtained with cell lines, their reduced functionality and inherent genomic instability led us to reinvestigate the use of peripheral blood monocyte-derived dendritic cells (PBMDCs) for the establishment of a reliable in vitro sensitization test. To solve the issues associated with the use of primary cells, the culture and exposure conditions (cytokine concentrations, incubation time, readout, pooled vs. single donors and cytotoxicity) were re-assessed and optimized. Here we propose a stable and reproducible protocol based on PBMDCs. This should allow a wider acceptance of PBMDCs as a reliable test system for the detection of human skin sensitizers and the inclusion of this protocol in an integrated testing strategy.

  8. Levamisole enhances immune response by affecting the activation and maturation of human monocyte-derived dendritic cells

    Science.gov (United States)

    Chen, L-Y; Lin, Y-L; Chiang, B-L

    2008-01-01

    Levamisole is a synthetic phenylimidazolthiazole that was first introduced in 1966 as an anti-helmintic agent. Current studies have been focused upon its effect on immune response and on cancer treatment. We examined the molecular mechanisms of levamisole in the activation and maturation of human monocyte-derived dendritic cells (DC) and human T cells. Treatment of DC with levamisole increased the presentation of CD80, CD86, CD83 and human leucocyte antigen D-related (HLA-DR) molecules on the cell membrane, as well as the production of interleukin (IL)-12 p40 and IL-10. Levamisole-treated human DC also enhanced T cell activation towards type 1 T helper immune response by inducing interferon-γ secretion. Neutralization with antibodies against Toll-like receptor (TLR)-2 inhibited levamisole-induced production of IL-12 p40 and IL-10, suggesting a vital role for TLR-2 in signalling DC upon incubation with levamisole. The inhibition of nuclear factor-κB, extracellular signal-regulated kinases 1/2 or c-Jun N-terminal kinases pathways also prevented the effects of levamisole on DC in producing IL-12 p40 or IL-10. Taken together, levamisole could enhance immune response towards T helper 1 development through the activation of dendritic cells or T cell aspects. PMID:18005262

  9. Immunomodulatory effects of adult Haemonchus contortus excretory/secretory products on human monocyte-derived dendritic cells.

    Science.gov (United States)

    Rehman, Z U; Knight, J S; Koolaard, J; Simpson, H V; Pernthaner, A

    2015-12-01

    The levels of expression of surface molecules and release of cytokines and chemokines of human monocyte-derived dendritic cells were determined after their exposure to adult H. contortus excretory/secretory (ES) products or a combination of ES products and bacterial lipopolysaccharide (LPS). Worm products provoked a weak response and only partial maturation of the dendritic cells, consistent with the hyporesponsiveness and more tolerogenic immune environment present in parasitized animals and humans. Co-stimulation with LPS demonstrated that H. contortus secretions, like those of other helminths, contain immunomodulators capable of reducing some aspects of the strong T(H)1/T(H)2 response evoked by bacterial LPS. There were significant reductions in the release of some cytokine/chemokines by LPS-stimulated mdDCs and a trend (although not significant at P < 0.05) for reduced expression levels of CD40, CD80 and HLA-DR. A prominent feature was the variability in responses of dendritic cells from the four donors, even on different days in repeat experiments, suggesting that generalized conclusions may be difficult to make, except in genetically related animals. Such observations may therefore be applicable only to restricted populations. In addition, previous exposure to parasites in a target population for immunomodulatory therapy may be an important factor in assessing the likelihood of adverse reactions or failures in the treatment to worm therapy.

  10. The effect of caffeic acid phenethyl ester on the functions of human monocyte-derived dendritic cells

    Directory of Open Access Journals (Sweden)

    Wu Wen-Mein

    2009-07-01

    Full Text Available Abstract Background Propolis, an ancient herbal medicine, has been reported the beneficial effect both in asthma patients and murine model of asthma, but the mechanism was not clearly understood. In this study, the effect of caffeic acid phenethyl ester (CAPE, the most extensively studied components in propolis, on the functions of human monocyte-derived dendritic cells (MoDCs was investigated. Results CAPE significantly inhibited IL-12 p40, IL-12 p70, IL-10 protein expression in mature healthy human MoDCs stimulated by lipopolysaccharides (LPS and IL-12 p40, IL-10, IP-10 stimulated by crude mite extract. CAPE significantly inhibited IL-10 and IP-10 but not IL-12 expression in allergic patients' MoDCs stimulated by crude mite extract. In contrast, the upregulation of costimulatory molecules in mature MoDCs was not suppressed by CAPE. Further, the antigen presenting ability of DCs was not inhibited by CAPE. CAPE inhibited IκBα phosphorylation and NF-κB activation but not mitogen-activated protein kinase (MAPK family phosphorylation in human MoDCs. Conclusion These results indicated that CAPE inhibited cytokine and chemokine production by MoDCs which might be related to the NF-κB signaling pathway. This study provided a new insight into the mechanism of CAPE in immune response and the rationale for propolis in the treatment of asthma and other allergic disorders.

  11. Indoor pollutant hexabromocyclododecane enhances house dust mite-induced activation of human monocyte-derived dendritic cells.

    Science.gov (United States)

    Canbaz, Derya; Lebre, M Cristina; Logiantara, Adrian; van Ree, Ronald; van Rijt, Leonie S

    2016-11-01

    The indoor pollutant hexabromocyclododecane (HBCD) has been added as flame retardant to many consumer products but detaches and accumulates in house dust. Inhalation of house dust leads to exposure to house dust mite (HDM) allergens in the presence of HBCD. Activation of dendritic cells is crucial in the sensitization to HDM allergens. The current study examined whether exposure to HBCD affected activation/maturation of HDM-exposed human dendritic cells (DC). Human monocyte-derived DC (moDC) were exposed simultaneously to HDM and a concentration range of HBCD (0.1-20 μM) in vitro. HDM exposure of moDC induced expression of co-stimulatory molecule CD80 and production of pro-inflammatory cytokines interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF)-α. However, simultaneous exposure of moDC to HBCD and HDM enhanced the expression of antigen presenting molecule HLA-DR, co-stimulatory molecule CD86 and pro-inflammatory cytokine IL-8 depending on the dose of HBCD. Our results indicate that simultaneous exposure of HDM and HBCD can enhance the antigen presentation and maturation/activation of DC.

  12. Monocyte-derived dendritic cells from HLA-B27+ axial spondyloarthritis (SpA) patients display altered functional capacity and deregulated gene expression.

    Science.gov (United States)

    Talpin, Alice; Costantino, Félicie; Bonilla, Nelly; Leboime, Ariane; Letourneur, Franck; Jacques, Sébastien; Dumont, Florent; Amraoui, Sonia; Dutertre, Charles-Antoine; Garchon, Henri-Jean; Breban, Maxime; Chiocchia, Gilles

    2014-08-21

    This study aimed to compare the functional capacity and gene expression profile of monocyte-derived dendritic cells (MD-DCs) in HLA-B27+ axial spondyloarthritis (SpA) patients and healthy controls. MD-DCs were differentiated with interleukin 4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) for seven days, starting from purified CD14+ monocytes and stimulated with lipopolysaccharide (LPS) for six and twenty four hours. Their capacity to stimulate allogeneic CD4+ T cells from unrelated healthy donor was tested. Transcriptomic study was performed with Affymetrix HuGene 1.0 ST microarrays. Gene expression levels were compared between patients and controls using a multivariate design under a linear model (LIMMA). Real-time quantitative PCR (qRT-PCR) was performed for validation of the most striking gene expression differences. The stimulatory capacity of allogeneic CD4+ T cells by MD-DCs from SpA patients was decreased. Transcriptomic analysis revealed 81 genes differentially expressed in MD-DCs between SpA patients and controls (P 1.5). Four selected genes were validated by q ADAMTS15, CITED2, F13A1 and SELL. Expression levels of ADAMTS15 and CITED2, encoding a metallopeptidase and a transcription factor, respectively, were inversely correlated with each other (R = 0.75, P = 0.0003). Furthermore, in silico analysis identified several genes of the Wnt signaling pathway having expression co-regulated with CITED2. This study revealed altered function and gene expression pattern in MD-DCs from HLA-B27+ axial SpA. Co-expression study showed an inverse correlation between ADAMTS15 and CITED2. Moreover, the Wnt signaling pathway appeared as deregulated in SpA MD-DCs, a finding which may be connected to Th17-driven inflammatory responses.

  13. Enhanced lentiviral transduction of monocyte-derived dendritic cells in the presence of conditioned medium from dying monocytes.

    Science.gov (United States)

    Masurier, C; Boutin, S; Veron, P; Bernard, J; Danos, O; Davoust, J

    2007-02-01

    Lentiviral vectors (LVs) are attractive vehicles for the transduction of human dendritic cells (DCs) in order to mobilize their endogenous antigen presentation pathways. We analyzed here how to improve the efficiency of LV transduction, which we performed at the initial stages of the differentiation of purified monocytes into dendritic cells (Mo-DCs). Using LVs pseudotyped with the vesicular stomatitis virus envelope G glycoprotein (VSV-G), we found that a conditioned medium derived from dying monocytes (MCM) improved by 2- to 10- fold the proportion of transduced Mo-DCs. This enhanced transduction efficiency requires the presence of MCM during the initial stage of LV transduction and does not affect the phenotype and antigen presentation function of terminally differentiated Mo-DCs. Importantly, we found that MCM derived from a human acute monocytic leukemia cell line, THP-1, was equally effective. The MCM activity was heat stable (56 degrees C) and was present in the soluble fraction after high-speed centrifugation. Altogether our results show that a soluble factor present in dying monocyte cultures can replace advantageously facilitating agents such as Polybrene, to achieve high LV transductions levels. This protocol can be performed with autologous monocytes and is therefore applicable in clinical settings.

  14. Prophylactic vaccines are potent activators of monocyte-derived dendritic cells and drive effective anti-tumor responses in melanoma patients at the cost of toxicity

    OpenAIRE

    Bol, Kalijn F.; Aarntzen, Erik H. J. G.; Pots, Jeanette M.; Olde Nordkamp, Michel A. M.; van de Rakt, Mandy W. M. M.; Scharenborg, Nicole M.; de Boer, Annemiek J.; van Oorschot, Tom G. M.; Croockewit, Sandra A. J.; Blokx, Willeke A. M.; Oyen, Wim J. G.; Boerman, Otto C.; Mus, Roel D. M.; van Rossum, Michelle M.; van der Graaf, Chantal A. A.

    2016-01-01

    Dendritic cell (DC)-based immunotherapy is explored worldwide in cancer patients, predominantly with DC matured with pro-inflammatory cytokines and prostaglandin E2. We studied the safety and efficacy of vaccination with monocyte-derived DC matured with a cocktail of prophylactic vaccines that contain clinical-grade Toll-like receptor ligands (BCG, Typhim, Act-HIB) and prostaglandin E2 (VAC-DC). Stage III and IV melanoma patients were vaccinated via intranodal injection (12 patients) or combi...

  15. Activation of the aryl hydrocarbon receptor affects activation and function of human monocyte-derived dendritic cells.

    Science.gov (United States)

    Wang, C; Ye, Z; Kijlstra, A; Zhou, Y; Yang, P

    2014-08-01

    Aryl hydrocarbon receptor (AhR) is well known for mediating the toxic effects of dioxin-containing pollutants, but has also been shown to be involved in the natural regulation of the immune response. In this study, we investigated the effect of AhR activation by its endogenous ligands 6-formylindolo[3,2-b]carbazole (FICZ) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) on the differentiation, maturation and function of monocyte-derived DCs in Behçet's disease (BD) patients. In this study, we showed that AhR activation by FICZ and ITE down-regulated the expression of co-stimulatory molecules including human leucocyte antigen D-related (HLA-DR), CD80 and CD86, while it had no effect on the expression of CD83 and CD40 on DCs derived from BD patients and normal controls. Lipopolysaccharide (LPS)-treated dendritic cells (DCs) from active BD patients showed a higher level of interleukin (IL)-1β, IL-6, IL-23 and tumour necrosis factor (TNF)-α production. FICZ or ITE significantly inhibited the production of IL-1β, IL-6, IL-23 and TNF-α, but induced IL-10 production by DCs derived from active BD patients and normal controls. FICZ or ITE-treated DCs significantly inhibited the T helper type 17 (Th17) and Th1 cell response. Activation of AhR either by FICZ or ITE inhibits DC differentiation, maturation and function. Further studies are needed to investigate whether manipulation of the AhR pathway may be used to treat BD or other autoimmune diseases.

  16. Differential Activation of Human Monocyte-Derived and Plasmacytoid Dendritic Cells by West Nile Virus Generated in Different Host Cells▿

    Science.gov (United States)

    Silva, Maria Carlan; Guerrero-Plata, Antonieta; Gilfoy, Felicia D.; Garofalo, Roberto P.; Mason, Peter W.

    2007-01-01

    Dendritic cells (DCs) play a central role in innate immunity and antiviral responses. In this study, we investigated the production of alpha interferon (IFN-α) and inducible chemokines by human monocyte-derived dendritic cells (mDCs) and plasmacytoid dendritic cells (pDCs) infected with West Nile virus (WNV), an emergent pathogen whose infection can lead to severe cases of encephalitis in the elderly, children, and immunocompromised individuals. Our experiments demonstrated that WNV grown in mammalian cells (WNVVero) was a potent inducer of IFN-α secretion in pDCs and, to a lesser degree, in mDCs. The ability of WNVVero to induce IFN-α in pDCs did not require viral replication and was prevented by the treatment of cells with bafilomycin A1 and chloroquine, suggesting that it was dependent on endosomal Toll-like receptor recognition. On the other hand, IFN-α production in mDCs required viral replication and was associated with the nuclear translocation of IRF3 and viral antigen expression. Strikingly, pDCs failed to produce IFN-α when stimulated with WNV grown in mosquito cells (WNVC7/10), while mDCs responded similarly to WNVVero or WNVC7/10. Moreover, the IFN-dependent chemokine IP-10 was produced in substantial amounts by pDCs in response to WNVVero but not WNVC7/10, while interleukin-8 was produced in greater amounts by mDCs infected with WNVC7/10 than in those infected with WNVVero. These findings suggest that cell-specific mechanisms of WNV recognition leading to the production of type I IFN and inflammatory chemokines by DCs may contribute to both the innate immune response and disease pathogenesis in human infections. PMID:17913823

  17. In vitro interactions of Candida parapsilosis wild type and lipase deficient mutants with human monocyte derived dendritic cells

    Directory of Open Access Journals (Sweden)

    Vágvölgyi Csaba

    2011-05-01

    Full Text Available Abstract Background Candida parapsilosis typically is a commensal of human skin. However, when host immune defense is compromised or the normal microflora balance is disrupted, C. parapsilosis transforms itself into an opportunistic pathogen. Candida-derived lipase has been identified as potential virulence factor. Even though cellular components of the innate immune response, such as dendritic cells, represent the first line of defense against invading pathogens, little is known about the interaction of these cells with invading C. parapsilosis. Thus, the aim of our study was to assess the function of dendritic cells in fighting C. parapsilosis and to determine the role that C. parapsilosis-derived lipase plays in the interaction with dendritic cells. Results Monocyte-derived immature and mature dendritic cells (iDCs and mDCs, respectively co-cultured with live wild type or lipase deficient C. parapsilosis strains were studied to determine the phagocytic capacity and killing efficiency of host cells. We determined that both iDCs and mDCs efficiently phagocytosed and killed C. parapsilosis, furthermore our results show that the phagocytic and fungicidal activities of both iDCs and mDCs are more potent for lipase deficient compared to wild type yeast cells. In addition, the lipase deficient C. parapsilosis cells induce higher gene expression and protein secretion of proinflammatory cytokines and chemokines in both DC types relative to the effect of co-culture with wild type yeast cells. Conclusions Our results show that DCs are activated by exposure to C. parapsilosis, as shown by increased phagocytosis, killing and proinflammatory protein secretion. Moreover, these data strongly suggest that C. parapsilosis derived lipase has a protective role during yeast:DC interactions, since lipase production in wt yeast cells decreased the phagocytic capacity and killing efficiency of host cells and downregulated the expression of host effector molecules.

  18. Comparative analysis of signature genes in porcine reproductive and respiratory syndrome virus (PRRSV)-infected porcine monocyte-derived dendritic cells at differential activation statuses

    Science.gov (United States)

    Activation statuses of monocytic cells, e.g. monocytes, macrophages and dendritic cells (DCs), are critically important for antiviral immunity. In particular, some devastating viruses, including porcine reproductive and respiratory syndrome virus (PRRSV), are capable of directly infecting these cell...

  19. HIV-1 gp120 activates the STAT3/interleukin-6 axis in primary human monocyte-derived dendritic cells.

    Science.gov (United States)

    Del Cornò, Manuela; Donninelli, Gloria; Varano, Barbara; Da Sacco, Letizia; Masotti, Andrea; Gessani, Sandra

    2014-10-01

    Dendritic cells (DCs) are fundamental for the initiation of immune responses and are important players in AIDS immunopathogenesis. The modulation of DC functional activities represents a strategic mechanism for HIV-1 to evade immune surveillance. Impairment of DC function may result from bystander effects of HIV-1 envelope proteins independently of direct HIV-1 infection. In this study, we report that exposure of immature monocyte-derived DCs (MDDCs) to HIV-1 R5 gp120 resulted in the CCR5-dependent production of interleukin-6 (IL-6) via mitogen-activated protein kinase (MAPK)/NF-κB pathways. IL-6 in turn activated STAT3 by an autocrine loop. Concomitantly, gp120 promoted an early activation of STAT3 that further contributed to IL-6 induction. This activation paralleled a concomitant upregulation of the STAT3 inhibitor PIAS3. Notably, STAT3/IL-6 pathway activation was not affected by the CCR5-specific ligand CCL4. These results identify STAT3 as a key signaling intermediate activated by gp120 in MDDCs and highlight the existence of a virus-induced dysregulation of the IL-6/STAT3 axis. HIV-1 gp120 signaling through STAT3 may provide an explanation for the impairment of DC function observed upon HIV exposure. This study provides new evidence for the molecular mechanisms and signaling pathways triggered by HIV-1 gp120 in human DCs in the absence of productive infection, emphasizing a role of aberrant signaling in early virus-host interaction, contributing to viral pathogenesis. We identified STAT3 as a key component in the gp120-mediated signaling cascade involving MAPK and NF-κB components and ultimately leading to IL-6 secretion. STAT3 now is recognized as a key regulator of DC functions. Thus, the identification of this transcription factor as a signaling molecule mediating some of gp120's biological effects unveils a new mechanism by which HIV-1 may deregulate DC functions and contribute to AIDS pathogenesis. Copyright © 2014, American Society for Microbiology

  20. Mesenchymal stem cells derived from human exfoliated deciduous teeth (SHEDs induce immune modulatory profile in monocyte-derived dendritic cells.

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    Fernando de Sá Silva

    Full Text Available BACKGROUND: Mesenchymal stem cells have prominent immune modulatory properties, which may have clinical applications; however their major source, bone marrow, is of limited availability. On the other hand, mesenchymal stem cells derived from human exfoliated deciduous teeth (SHEDs are readily accessible, but their immune regulatory properties have not been completely investigated. This study was designed, therefore, to evaluate the SHEDs influence on DCs differentiation, maturation, ability to activate T cells and to expand CD4(+Foxp3(+ T cells. METHODOLOGY/PRINCIPAL FINDINGS: The experiments were based in cellular co-culture during differentiation and maturation of monocyte derived-DCs (moDCs, with, or not, presence of SHEDs. After co-culture with SHEDs, (moDCs presented lower expression of BDCA-1 and CD11c, in comparison to DC cultivated without SHEDs. CD40, CD80, CD83 and CD86 levels were also decreased in mature DCs (mDCs after co-cultivation with SHEDs. To assess the ability of SHEDs-exposed moDCs to modulate T cell responses, the former were separated from SHEDs, and co-cultured with peripheral blood lymphocytes. After 5 days, the proliferation of CD4(+ and CD8(+ T cells was evaluated and found to be lower than that induced by moDCs cultivated without SHEDs. In addition, an increase in the proportion of CD4(+Foxp3(+IL-10(+ T cells was observed among cells stimulated by mature moDCs that were previously cultivated with SHEDs. Soluble factors released during co-cultures also showed a reduction in the pro-inflammatory cytokines (IL-2, TNF-α and IFN-γ, and an increase in the anti-inflammatory molecule IL-10. CONCLUSION/SIGNIFICANCE: This study shows that SHEDs induce an immune regulatory phenotype in moDCs cells, evidenced by changes in maturation and differentiation rates, inhibition of lymphocyte stimulation and ability to expand CD4(+Foxp3(+ T cells. Further characterization and validation of this phenomenon could support the use of SHEDs

  1. PGE2 differentially regulates monocyte-derived dendritic cell cytokine responses depending on receptor usage (EP2/EP4).

    Science.gov (United States)

    Poloso, Neil J; Urquhart, Paula; Nicolaou, Anna; Wang, Jenny; Woodward, David F

    2013-07-01

    Dendritic cells (DCs) are central players in coordinating immune responses, both innate and adaptive. While the role of lipid mediators in the immune response has been the subject of many investigations, the precise role of prostaglandins has often been plagued by contradictory studies. In this study, we examined the role of PGE(2) on human DC function. Although studies have suggested that PGE(2) specifically plays a role in DC motility and cytokine release profile, the precise receptor usage and signaling pathways involved remain unclear. In this report we found that irrespective of the human donor, monocyte-derived dendritic cells (MoDCs) express three of the four PGE(2) receptor subtypes (EP(2-4)), although only EP(2) and EP(4) were active with respect to cytokine production. Using selective EP receptor antagonists and agonists, we demonstrate that PGE(2) coordinates control of IL-23 release (a promoter of Th17, an autoimmune associated T cell subset) in a dose-dependent manner by differential use of EP(2) and EP(4) receptors in LPS-activated MoDCs. This is in contrast to IL-12, which is dose dependently inhibited by PGE(2) through both receptor subtypes. Low concentrations (∼1-10nM) of PGE(2) promoted IL-23 production via EP(4) receptors, while at higher (>50 nM), but still physiologically relevant concentrations, IL-23 is suppressed by an EP(2) dependent mechanism. These results can be explained by differential regulation of the common subunit, IL-12p40, and IL-23p19, by EP(2) and EP(4). By these means, PGE(2) can act as a regulatory switch of immune responses depending on its concentration in the microenvironment. In addition, we believe these results may also explain why seemingly conflicting biological functions assigned to PGE(2) have been reported in the literature, as the concentration of ligand (PGE(2)) fundamentally alters the nature of the response. This finding also highlights the potential of designing therapeutics which differentially target

  2. Pharmacological effects of mitraphylline from Uncaria tomentosa in primary human monocytes: Skew toward M2 macrophages.

    Science.gov (United States)

    Montserrat-de la Paz, S; de la Puerta, R; Fernandez-Arche, A; Quilez, A M; Muriana, F J G; Garcia-Gimenez, M D; Bermudez, B

    2015-07-21

    Uncaria tomentosa (Willdenow ex Roemer & Schultes) DC. (Rubiaceae) is a Peruvian thorny liana, commonly known as "cat׳s claw", and traditionally used in folk medicine to deal with several inflammatory diseases. Mitraphylline (MTP) is the most abundant pentacyclic oxindolic alkaloid (POA) from U. Tomentosa and has been reported to modify the inflammatory response. Herein, we have sought to identify the mechanisms underlying this modulatory effect of MTP on primary human monocytes and its ability to regulate differentiation processes on human primary monocyte and monocyte-derived macrophages. In vitro studies with human primary monocytes and monocyte-derived macrophages were performed. Monocytes and M0 macrophages were exposed to MTP (25μM) and LPS (100ng/mL). M0 macrophages were polarized to M1 and M2 phenotypes in the absence or presence of MTP. The activation state of monocytes/macrophages was assessed by flow cytometry, gene expression and protein analysis of different specific markers. In human primary monocytes, the incubation of MTP for 24h reduced the number of classical (CD14(++)CD16(-)) and intermediate (CD14(++)CD16(+)) subsets when compared to untreated or LPS-treated cells. MTP also reduced the chemotactic capacity of human primary monocytes. In addition, MTP promoted the polarization of M0 macrophages toward an anti-inflammatory M2 phenotype, the abrogation of the release of pro-inflammatory cytokines such as TNFα, IL-6 or IL-1β, as well as the restoration of markers for M2 macrophages in LPS-treated M1 macrophages. Our results suggest that MTP may be a key modulator for regulating the plasticity of monocytes/macrophages and the attenuation of the inflammatory response. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  3. Endogenous epoxygenases are modulators of monocyte/macrophage activity.

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    Jonas Bystrom

    Full Text Available BACKGROUND: Arachidonic acid is metabolized through three major metabolic pathways, the cyclooxygenase, lipoxygenase and CYP450 enzyme systems. Unlike cyclooxygenase and lipoxygenases, the role of CYP450 epoxygenases in monocyte/macrophage-mediated responses is not known. METHODOLOGY/PRINCIPAL FINDINGS: When transfected in vitro, CYP2J2 is an efficient activator of anti-inflammatory pathways through the nuclear receptor peroxisome proliferator-activated receptor (PPAR α. Human monocytes and macrophages contain PPARα and here we show they express the epoxygenases CYP2J2 and CYP2C8. Inhibition of constitutive monocyte epoxygenases using the epoxygenase inhibitor SKF525A induces cyclooxygenase (COX-2 expression and activity, and the release of TNFα, and can be reversed by either add back of the endogenous epoxygenase products and PPARα ligand 11,12- epoxyeicosatrienoic acid (EET or the addition of the selective synthetic PPARα ligand GW7647. In alternatively activated (IL-4-treated monocytes, in contrast to classically activated cells, epoxygenase inhibition decreased TNFα release. Epoxygenases can be pro-inflammatory via superoxide anion production. The suppression of TNFα by SKF525A in the presence of IL-4 was associated with a reduction in superoxide anion generation and reproduced by the superoxide dismutase MnCl(2. Similar to these acute activation studies, in monocyte derived macrophages, epoxygenase inhibition elevates M1 macrophage TNFα mRNA and further decreases M2 macrophage TNFα. CONCLUSIONS/SIGNIFICANCE: In conclusion, epoxygenase activity represents an important endogenous pathway which limits monocyte activation. Moreover endogenous epoxygenases are immuno-modulators regulating monocyte/macrophage activation depending on the underlying activation state.

  4. Monocyte derived dendritic cells generated by IFN-α acquire mature dendritic and natural killer cell properties as shown by gene expression analysis

    OpenAIRE

    Czibere Akos; Winter Meike; Diaz Blanco Elena; Papewalis Claudia; Schott Matthias; Maihöfer Dagmar; Kronenwett Ralf; Safaian Nancy; Korthals Mark; Haas Rainer; Kobbe Guido; Fenk Roland

    2007-01-01

    Abstract Background Dendritic cell (DC) vaccines can induce antitumor immune responses in patients with malignant diseases, while the most suitable DC culture conditions have not been established yet. In this study we compared monocyte derived human DC from conventional cultures containing GM-CSF and IL-4/TNF-α (IL-4/TNF-DC) with DC generated by the novel protocol using GM-CSF and IFN-α (IFN-DC). Methods To characterise the molecular differences of both DC preparations, gene expression profil...

  5. Human XCR1+ Dendritic Cells Derived In Vitro from CD34+ Progenitors Closely Resemble Blood Dendritic Cells, Including Their Adjuvant Responsiveness, Contrary to Monocyte-Derived Dendritic Cells

    OpenAIRE

    S. Balan; Ollion, V.; Colletti, N.; Chelbi, R.; Montanana-Sanchis, F.; LIU, H.; Vu Manh, T.-P.; Sanchez, C.; Savoret, J.; Perrot, I.; Doffin, A.-C.; Fossum, E.; Bechlian, D.; Chabannon, C.; Bogen, B

    2014-01-01

    Human monocyte-derived dendritic cell (MoDC) have been used in the clinic with moderately encouraging results. Mouse XCR1+ DC excel at cross-presentation, can be targeted in vivo to induce protective immunity, and share characteristics with XCR1+ human DC. Assessment of the immunoactivation potential of XCR1+ human DC is hindered by their paucity in vivo and by their lack of a well-defined in vitro counterpart. We report in this study a protocol generating both XCR1+ and XCR1− human DC in CD3...

  6. Comparative Analysis of the Interaction of Helicobacter pylori with Human Dendritic Cells, Macrophages, and Monocytes

    Science.gov (United States)

    Fehlings, Michael; Drobbe, Lea; Moos, Verena; Renner Viveros, Pablo; Hagen, Jana; Beigier-Bompadre, Macarena; Pang, Ervinna; Belogolova, Elena; Churin, Yuri; Schneider, Thomas; Meyer, Thomas F.; Aebischer, Toni

    2012-01-01

    Helicobacter pylori may cause chronic gastritis, gastric cancer, or lymphoma. Myeloid antigen-presenting cells (APCs) are most likely involved in the induction and expression of the underlying inflammatory responses. To study the interaction of human APC subsets with H. pylori, we infected monocytes, monocyte-derived dendritic cells (DCs), and monocyte-derived (classically activated; M1) macrophages with H. pylori and analyzed phenotypic alterations, cytokine secretion, phagocytosis, and immunostimulation. Since we detected CD163+ (alternatively activated; M2) macrophages in gastric biopsy specimens from H. pylori-positive patients, we also included monocyte-derived M2 macrophages in the study. Upon H. pylori infection, monocytes secreted interleukin-1β (IL-1β), IL-6, IL-10, and IL-12p40 (partially secreted as IL-23) but not IL-12p70. Infected DCs became activated, as shown by the enhanced expression of CD25, CD80, CD83, PDL-1, and CCR7, and secreted IL-1β, IL-6, IL-10, IL-12p40, IL-12p70, and IL-23. However, infection led to significantly downregulated CD209 and suppressed the constitutive secretion of macrophage migration inhibitory factor (MIF). H. pylori-infected M1 macrophages upregulated CD14 and CD32, downregulated CD11b and HLA-DR, and secreted mainly IL-1β, IL-6, IL-10, IL-12p40, and IL-23. Activation of DCs and M1 macrophages correlated with increased capacity to induce T-cell proliferation and decreased phagocytosis of dextran. M2 macrophages upregulated CD14 and CD206 and secreted IL-10 but produced less of the proinflammatory cytokines than M1 macrophages. Thus, H. pylori affects the functions of human APC subsets differently, which may influence the course and the outcome of H. pylori infection. The suppression of MIF in DCs constitutes a novel immune evasion mechanism exploited by H. pylori. PMID:22615251

  7. Comparison of alpha-Type-1 polarizing and standard dendritic cell cytokine cocktail for maturation of therapeutic monocyte-derived dendritic cell preparations from cancer patients

    DEFF Research Database (Denmark)

    Trepiakas, Redas; Pedersen, Anders Elm; Met, Ozcan

    2008-01-01

    polarized dendritic cells (alphaDC1) in serum-free medium was published based on maturation of monocyte-derived DCs with TNF-alpha/IL-1-beta/polyinosinic:polycytidylic acid (poly-I:C)/interferon (IFN)-alpha and IFN-gamma. This DC maturation cocktail was described to fulfill the criteria for optimal DC......The current "gold standard" for generation of dendritic cell (DC) used in DC-based cancer vaccine studies is maturation of monocyte-derived DCs with tumor necrosis factor-alpha (TNF-alpha)/IL-1beta/IL-6 and prostaglandin E(2) (PGE(2)). Recently, a protocol for producing so-called alpha-Type-1...... of alphaDC1 maturation cocktail to a protocol for clinical grade DC generation from cancer patients performed in X-VIVO 15 medium. We showed that alphaDC1 in this protocol induce lower up-regulation of CD83 and several other maturation markers, co-stimulatory molecules and CCR7 together with higher up...

  8. Lentiviral-mediated gene delivery in human monocyte-derived dendritic cells: optimized design and procedures for highly efficient transduction compatible with clinical constraints.

    Science.gov (United States)

    Rouas, Redouane; Uch, Rathviro; Cleuter, Yvette; Jordier, François; Bagnis, Claude; Mannoni, Patrice; Lewalle, Philippe; Martiat, Philippe; Van den Broeke, Anne

    2002-09-01

    Gene delivery to dendritic cells (DCs) could represent a powerful method of inducing potent, long-lasting immunity. Although recent studies underline the intense interest in lentiviral vector-mediated monocyte-derived DC transduction, efficient gene transfer methods currently require high multiplicities of infection and are not compatible with clinical constraints. We have designed a strategy to optimize the efficiency and clinical relevance of this approach. Initially, using a third generation lentiviral vector expressing green fluorescent protein, we found that modifying the vector design, the DC precursor cell type, and the DC differentiation stage for transduction results in sustained transgene expression in 75-85% of immature DCs (transduction at a multiplicity of infection of 8). This high efficiency was reproducible among different donors irrespective of whether DCs were expanded from fresh or cryopreserved CD14(+) precursors. We then developed procedures that bypass the need for highly concentrated lentiviral preparations and the addition of polybrene to achieve efficient transduction. DCs transduced under these conditions retain their immature phenotype and immunostimulatory potential in both autologous and allogeneic settings. Furthermore, genetically modified DCs maintain their ability to respond to maturation signals and secrete bioactive IL-12, indicating that they are fully functional. Finally, the level of transgene expression is preserved in the therapeutically relevant mature DCs, demonstrating that there is neither promoter-silencing nor loss of transduced cells during maturation. The novel approach described should advance lentiviral-mediated monocyte-derived DC transduction towards a clinical reality.

  9. Treatment with dexamethasone and monophosphoryl lipid A removes disease-associated transcriptional signatures in monocyte-derived dendritic cells from rheumatoid arthritis patients and confers tolerogenic features

    Directory of Open Access Journals (Sweden)

    Paulina Andrea García-González

    2016-10-01

    Full Text Available Tolerogenic dendritic cells (TolDCs are promising tools for therapy of autoimmune diseases such as rheumatoid arthritis (RA. Here we characterise monocyte-derived TolDCs from RA patients modulated with dexamethasone and activated with monophosphoryl lipid A (MPLA, referred to as MPLA-tDCs, in terms of gene expression, phenotype, cytokine profile, migratory properties and T cell-stimulatory capacity, in order to explore their suitability for cellular therapy. MPLA-tDCs derived from RA patients displayed an anti-inflammatory profile with reduced expression of costimulatory molecules and high IL-10/IL-12 ratio, but were capable of migrating towards the lymphoid chemokines CXCL12 and CCL19. These MPLA-tDCs induced hyporesponsiveness of autologous CD4+ T cells specific for synovial antigens in vitro. Global transcriptome analysis confirmed a unique transcriptional profile of MPLA-tDCs and revealed that RA-associated genes, which were upregulated in untreated DCs from RA patients, returned to expression levels of healthy donor-derived DCs after treatment with dexamethasone and MPLA. Thus, monocyte-derived DCs from RA patients have the capacity to develop tolerogenic features at transcriptional as well as at translational level, when modulated with dexamethasone and MPLA, overcoming disease-related effects. Furthermore, the ability of MPLA-tDCs to impair T cell responses to synovial antigens validates their potential as cellular treatment for RA.

  10. Human monocyte-derived insulin-like growth factor-2 enhances the infection of human arterial endothelial cells by Chlamydia pneumoniae.

    Science.gov (United States)

    Lin, T M; Campbell, L A; Rosenfeld, M E; Kuo, C C

    2001-05-01

    It has been shown that infection of human endothelial cells by Chlamydia pneumoniae is enhanced by co-culturing endothelial cells with human monocytes and is mediated by monocyte-derived soluble factors. This study was conducted to identify the infectivity-enhancing factor. Serum-free conditioned medium of human monocytic cells was fractionated by ultrafiltration. The enhancing activity was found in the fraction in the molecular mass range between 5000 and 10,000 kDa. Recombinant human insulin-like growth factor (IGF)-1 or -2, with a molecular mass of 7500 kDa, was added to the culture medium of human endothelial cells for growing C. pneumoniae. Only IGF-2 enhanced C. pneumoniae growth. Pretreatment of the conditioned medium with a monoclonal antibody against IGF-2 blocked the enhancing activity. This suggests that the infectivity-enhancing factor is IGF-2 and that paracrine interactions between monocytes and endothelial cells in vivo can induce secretory products and sustain infection with C. pneumoniae within atherosclerotic lesions.

  11. Th1 disabled function in response to TLR4 stimulation of monocyte-derived DC from patients chronically-infected by hepatitis C virus.

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    Laure Perrin-Cocon

    Full Text Available BACKGROUND: Lack of protective antibodies and inefficient cytotoxic responses are characteristics of chronic hepatitis C infection. A defect in dendritic cell (DC function has thus been suspected, but this remains a controversial issue. METHODS AND FINDINGS: Here we show that monocyte-derived DC (MoDC from chronically-infected patients can mature in response to TLR1/2, TLR2/6 or TLR3 ligands. In contrast, when stimulated with the TLR4 ligand LPS, MoDC from patients show a profound defect in inducing IFNgamma secretion by allogeneic T cells. This defect is not due to defective phenotypic maturation or to the presence of HCV-RNA in DC or monocytes but is correlated to reduced IL-12 secretion by DC. Restoration of DC ability to stimulate IFNgamma secretion can be obtained by blocking MEK activation in DC, indicating that MEK/ERK pathway is involved in the Th1 defect of MoDC. Monocytes from HCV patients present increased spontaneous secretion of cytokines and chemokines, especially MIP-1beta. Addition of MIP-1beta on healthy monocytes during differentiation results in DC that have Th1 defect characteristic of MoDC from HCV patients, suggesting that MIP-1beta secretion by HCV monocytes participates in the Th1 defect of DC. CONCLUSIONS: Our data indicate that monocytes from HCV patients are activated in vivo. This interferes with their differentiation into DC, leading to deficient TLR4 signaling in these cells that are enable to induce a Th1 response. This specific defect is linked to the activation of the MEK/ERK pathway.

  12. Acidosis differently modulates the inflammatory program in monocytes and macrophages.

    Science.gov (United States)

    Riemann, Anne; Wußling, Hanna; Loppnow, Harald; Fu, Hang; Reime, Sarah; Thews, Oliver

    2016-01-01

    Inflammation, ischemia or the microenvironment of solid tumors is often accompanied by a reduction of extracellular pH (acidosis) that stresses the cells and acts on cellular signaling and transcription. The effect of acidosis on the expression of various inflammatory markers, on functional parameters (migration, phagocytic activity) and on signaling pathways involved was studied in monocytic cells and macrophages. In monocytic cell lines acidosis led to a reduction in expression of most of the inflammatory mediators, namely IL-1ß, IL-6, TNF-α, MCP-1, COX-2 and osteopontin. In primary human monocytes MCP-1 and TNF-α were reduced but COX-2 and IL-6 were increased. In RAW264.7 macrophage cell line IL-1ß, COX-2 and iNOS expression was increased, whereas MCP-1 was reduced similar to the effect in monocytic cells. For primary human monocyte-derived macrophages the regulation of inflammatory markers by acidosis depended on activation state, except for the acidosis-induced downregulation of MCP-1 and TNF-α. Acidosis affected functional immune cell behavior when looking at phagocytic activity which was increased in a time-dependent manner, but cellular motility was not changed. Neither ERK1/2 nor CREB signaling was stimulated by the reduction of extracellular pH. However, p38 was activated by acidosis in RAW264.7 cells and this activation was critical for the induction of IL-1ß, COX-2 and iNOS expression. In conclusion, acidosis may impede the recruitment of immune cells, but fosters inflammation when macrophages are present by increasing the level of COX-2 and iNOS and by functionally forcing up the phagocytic activity.

  13. Human monocyte-derived dendritic cells expressing both chemotactic cytokines IL-8, MCP-1, RANTES and their receptors,and their selective migration to these chemokines

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To characterize the mRNA expression of CXC chemokine IL-8, CC chemokine monocyte chemothractant protein-1 (MCP-1) and regulated on activation,normal T cell expressed and secreted (RANTES), and a newly defined DC chemokine DC- CK1 as well as the expression of IL-8 receptor, MCP-1 receptor and RANTES receptor in human monocyte derived dendritic cells (MoDCs).The migratory responsiveness of MoDC to IL-8, MCP-1 and RANTES was alsso studied. Methods In vitro generated MoDCs were obtained by differentiating monocytes in the presence of GM-CSF and IL-4 for 5 days. The time course of RNA expression was analyzed by RT-PCR and migratoly ability was assessed by a micromultiwell chemotaxis chamber assay. Results IL-8, MCP-1, RANTES and their corres ponding receptors were consistently expressed in MoDCs. DC-CK-1 expression was detectable efter 48 hours of differentiation. MoDC selectively migrated in response to MCP-1 and RANTES but not to IL-8 though transcripts of IL-8 receptor were present. Conclusion Because the capacity of dendritic cells to initiate immune responses depends on their specialized migratory and tissue homing properties, the expression of chemokines and their receptors along with the migratory responsiveness to chemokines of MoDC in our study suggests a potential role of chemokines in the interaction between dendritic cells and T cells and the induction of immune responses.

  14. PU.1 is essential for CD11c expression in CD8(+/CD8(- lymphoid and monocyte-derived dendritic cells during GM-CSF or FLT3L-induced differentiation.

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    Xue-Jun Zhu

    Full Text Available Dendritic cells (DCs regulate innate and acquired immunity through their roles as antigen-presenting cells. Specific subsets of mature DCs, including monocyte-derived and lymphoid-derived DCs, can be distinguished based on distinct immunophenotypes and functional properties. The leukocyte integrin, CD11c, is considered a specific marker for DCs and it is expressed by all DC subsets. We created a strain of mice in which DCs and their progenitors could be lineage traced based on activity of the CD11c proximal promoter. Surprisingly, we observed levels of CD11c promoter activity that were similar in DCs and in other mature leukocytes, including monocytes, granulocytes, and lymphocytes. We sought to identify DNA elements and transcription factors that regulate DC-associated expression of CD11c. The ets transcription factor, PU.1, is a key regulator of DC development, and expression of PU.1 varies in different DC subsets. GM-CSF increased monocyte-derived DCs in mice and from mouse bone marrow cultured in vitro, but it did not increase CD8(+ lymphoid-derived DCs or B220(+ plasmacytoid DCs. FLT3L increased both monocyte-derived DCs and lymphoid-derived DCs from mouse bone marrow cultured in vitro. GM-CSF increased the 5.3 Kb CD11c proximal promoter activity in monocyte-derived DCs and CD8(+ lymphoid-derived DCs, but not in B220(+ plasmacytoid DCs. In contrast, FLT3L increased the CD11c proximal promoter activity in both monocyte-derived DCs and B220(+ plasmacytoid DCs. We used shRNA gene knockdown and chromatin immunoprecipitation to demonstrate that PU.1 is required for the effects of GM-CSF or FLT3L on monocyte-derived DCs. We conclude that both GM-CSF and FLT3L act through PU.1 to activate the 5.3 Kb CD11c proximal promoter in DCs and to induce differentiation of monocyte-derived DCs. We also confirm that the CD11c proximal promoter is not sufficient to direct lineage specificity of CD11c expression, and that additional DNA elements are required

  15. 萝卜硫素对慢性阻塞性肺疾病患者巨噬细胞Toll样受体4/髓样分化因子88信号通路的影响%Effects of sulforaphane on Toll-like receptor 4/myeloid differentiation factor 88 pathway of monocyte-derived macrophages from patients with chronic obstructive pulmonary disease

    Institute of Scientific and Technical Information of China (English)

    曾晓丽; 刘晓菊; 包海荣; 张艺; 王小虎; 施凯; 庞琪

    2014-01-01

    4 (TLR4)/ myeloid differentiation factor 88 (MyD88) pathway and its downstream inflammatory cytokines in patients with chronic obstructive pulmonary disease (COPD).Methods From Jan.2012 to Mar.2013,thirty-two stable COPD patients and thirty healthy donors (non-COPD group) from the First Hospital of Lanzhou University were recruited.The peripheral blood monocytes were isolated and induced to macrophages (monocyte-derived macrophages,MDMs).The MDMs of COPD patients were divided into a blank control group,a LPS group,a sulforaphane group,a sulforaphane and LPS group (combined group),while the MDMs from the non-COPD group received no drug intervention.The number of cells in each group was 3 ×106.The mRNA and protein expression of TLR4 and MyD88 were measured with real-time PCR and Western blot.The TNF-α and IL-6 levels in the culture supernatant were measured with ELISA.Oneway ANOVA and LSD-t test were used for statistical analysis.Results The levels of mRNA and protein of TLR4 and MyD88 and the contents of TNF-α and IL-6 in the culture supernatant were higher in the blank control group [3.7 ±0.5,1.9±0.4,0.45 ±0.18,1.11 ±0.65,(31 ±4) and (43 ±5) μg/L] than those in the nonCOPDgroup [1.00,1.00,0.26±0.14,0.58±0.40,(19±2) and (29±4) μg/L] (t=2.19-12.11,P <0.05 or P <0.01).After LPS treatment (LPS group),the above parameters [5.5 ± 1.1,3.4 ± 1.6,0.65 ± 0.20,1.66 ± 0.64,(47 ± 4) and (54 ± 5) μg/L] were increased as compared to those in the blank control group (t =2.39-11.9,P < 0.05 or P < 0.01),but after sulforaphane treatment (Sulforaphane group),these parameters [2.2 ± 0.4,1.0 ± 0.6,0.25 ± 0.09,0.62 ± 0.34,(20 ± 3) and (27 ±4) μg/L] were decreased as compared to those in the blank control group (t =2.13-8.46,P < 0.05 or P < 0.01).Similarly,these parameters in the combined group [3.2 ± 0.5,1.5 ± 0.8,0.33 ± 0.11,0.77 ±0.25,(31 ±3) and (33 ±4) μg/L] were also remarkably decreased as compared to those in the LPS group (t =3

  16. Prostaglandin E 2 Does Not Modulate CCR7 Expression and Functionality after Differentiation of Blood Monocytes into Macrophages

    OpenAIRE

    Marc-André Allaire; Bérengère Tanné; Côté, Sandra C.; Nancy Dumais

    2013-01-01

    Previously, we demonstrated that prostaglandin E2 (PGE2) induces C-C chemokine receptor type 7 (CCR7) expression on human monocytes, which stimulates their subsequent migration in response to the CCR7 natural ligands CCL19 and CCL21. In this study, we determined whether PGE2 affects CCR7 expression on macrophages. Flow cytometric analysis and chemotaxis assays were performed on Mono Mac-1-derived macrophage (MDMM-1) as well as unpolarized monocyte-derived macrophages (MDMs) to determine the C...

  17. Gallic Acid Is the Major Active Component of Cortex Moutan in Inhibiting Immune Maturation of Human Monocyte-Derived Dendritic Cells.

    Science.gov (United States)

    Chan, Ben Chung Lap; Li, Long Fei; Hu, Shui Qing; Wat, Elaine; Wong, Eric Chun Wai; Zhang, Vanilla Xin; Lau, Clara Bik San; Wong, Chun Kwok; Hon, Kam Lun Ellis; Hui, Patrick Chi Leung; Leung, Ping Chung

    2015-09-10

    Atopic dermatitis (AD) is a widely prevalent and chronically relapsing inflammatory skin disease. Penta Herbs Formula (PHF) is efficacious in improving the quality of life and reducing topical corticosteroid used in children with AD and one of the active herbs it contains is Cortex Moutan. Recent studies showed that altered functions of dendritic cells (DC) were observed in atopic individuals, suggesting that DC might play a major role in the generation and maintenance of inflammation by their production of pro-inflammatory cytokines. Hence, the aims of the present study were to identify the major active component(s) of Cortex Moutan, which might inhibit DC functions and to investigate their possible interactions with conventional corticosteroid on inhibiting the development of DC from monocytes. Monocyte-derived dendritic cells (moDC) culture model coupled with the high-speed counter-current chromatography (HSCCC), high pressure liquid chromatography (HPLC) and Liquid Chromatography-Mass Spectrometry (LCMS) analyses were used. Gallic acid was the major active component from Cortex Moutan which could dose dependently inhibit interleukin (IL)-12 p40 and the functional cluster of differentiation (CD) surface markers CD40, CD80, CD83 and CD86 expression from cytokine cocktail-activated moDC. Gallic acid could also lower the concentration of hydrocortisone required to inhibit the activation of DC.

  18. Gallic Acid Is the Major Active Component of Cortex Moutan in Inhibiting Immune Maturation of Human Monocyte-Derived Dendritic Cells

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    Ben Chung Lap Chan

    2015-09-01

    Full Text Available Atopic dermatitis (AD is a widely prevalent and chronically relapsing inflammatory skin disease. Penta Herbs Formula (PHF is efficacious in improving the quality of life and reducing topical corticosteroid used in children with AD and one of the active herbs it contains is Cortex Moutan. Recent studies showed that altered functions of dendritic cells (DC were observed in atopic individuals, suggesting that DC might play a major role in the generation and maintenance of inflammation by their production of pro-inflammatory cytokines. Hence, the aims of the present study were to identify the major active component(s of Cortex Moutan, which might inhibit DC functions and to investigate their possible interactions with conventional corticosteroid on inhibiting the development of DC from monocytes. Monocyte-derived dendritic cells (moDC culture model coupled with the high-speed counter-current chromatography (HSCCC, high pressure liquid chromatography (HPLC and Liquid Chromatography-Mass Spectrometry (LCMS analyses were used. Gallic acid was the major active component from Cortex Moutan which could dose dependently inhibit interleukin (IL-12 p40 and the functional cluster of differentiation (CD surface markers CD40, CD80, CD83 and CD86 expression from cytokine cocktail-activated moDC. Gallic acid could also lower the concentration of hydrocortisone required to inhibit the activation of DC.

  19. Effects of PPAR-γ agonist rosiglitazone on MMP-9 and TIMP-1 expression of monocyte-derived macrophages isolated from patients with acute coronary syndrome%过氧化物酶增殖物激活受体-γ调节体外诱导培养的人巨噬细胞表达基质金属蛋白酶-9及其抑制物-1的作用及机制

    Institute of Scientific and Technical Information of China (English)

    罗玉梅; 万新红; 姜德谦; 旷文勇; 郭洪波; 陈朝霞; 王合金; 谢丽华; 段雯

    2009-01-01

    Objective Coronary arterial plaque rupture and secondary thrombosis are the major pathogenesis of acute coronary syndrome ( ACS) . Metalloprotease ( MMPs) secreted by monocyte/ macrophage was the main predisposing factor of the plaque rupture and peroxisotne proliferator-activated receptor-γ (PPAR-γ) is involved in a variety of inflammatory cytokine gene transcriptional regulations. We explored the possible role of PPAR-γ in the regulation of MMP-9 and TIMP-1 expressed by peripheral monocyte-derived macrophages (MDMs) from patients with ACS. Methods Peripheral blood mononuclear cells were isolated from 48 patients with ACS and 28 healthy controls and stimulated by macrophage colony-stimulating factor (0. 1 μg/ml for 24 hours) to form MDMs. MDMs were then incubated under various concentrations of rosiglitazone (0, 1, 10, 20 μmol/L) for 48 hours. The concentrations of MMP-9 and TIMP-1 in the supernatant were measured by enzyme linked immunosorbent assay, and the mRNA expression of PPAR-γ, MMP-9 by RT-PCR and nuclear factor-KB P65 ( NF-kB P65 ) expression by immunohistochemistry. Results PPAR-γ mRNA expression was significantly lower while NF-kB P65 and MMP-9 expression as well as MMP-9 and TIMP-1 concentrations in supernatant were significantly higher in ACS group than those in control group (all P <0. 05). After rosiglitazone intervention, PPAR-γ mRNA expression was significantly upregulated in both ACS and control groups in a dose-dependent manner. Both the MMP-9 concentration in the supernatant and MMP-9 mRNA expression were reduced post intervention with rosiglitazone in both groups. The TIMP-1 mRNA expression and concentration in supernatant were not affected by rosiglitazone in both groups. Rosiglitazone induced significant downregulation of NF-kB P65 expression in both groups. Conclusion Rosiglitazone intervention may downregulate MMP-9 expression by upregulating PPAR-γ expression, and by downregulaiting NF-kB expression in MDMs isolated from patients

  20. Development of a standardized protocol for reproducible generation of matured monocyte-derived dendritic cells suitable for clinical application

    OpenAIRE

    Bohnenkamp, H.R.; Noll, T.

    2003-01-01

    There is increasing interest in the generation of dendritic cells (DC) for cancer immunotherapy. In order to utilize DC in clinical trials it is necessary to have standardized, reproducible and easy to use protocols. We describe here the process development for the generation of DC as the result of investigation of culture conditions as well as consumption rates of medium and cytokines. Our studies demonstrate that highly viable DC (93 ± 2%) can be produced from CD14+ enriched monocytes via i...

  1. A novel method to generate monocyte-derived dendritic cells during coculture with HaCaT facilitates detection of weak contact allergens in cosmetics.

    Science.gov (United States)

    Frombach, Janna; Sonnenburg, Anna; Krapohl, Björn-Dirk; Zuberbier, Torsten; Stahlmann, Ralf; Schreiner, Maximilian

    2017-01-01

    The in vitro sensitization assay LCSA (Loose-fit Coculture-based Sensitization Assay) has proved reliable for the detection of contact sensitizers in the past. However, the coculture of human monocyte-derived dendritic cells (DCs) with primary human keratinocytes (KCs) in serum-free medium is relatively complex compared to other sensitization assays which use continuous cell lines. To facilitate high-throughput screening of chemicals, we replaced KCs with the HaCaT cell line under various culture conditions. Coculture of HaCaT with peripheral blood mononuclear cells in serum-supplemented medium leads to generation of CD1a(+)/CD1c(+) DCs after addition of GM-CSF, IL-4, and TGF-β1 (as opposed to CD1a(-)/CD1c(-) DCs which arise in the "classic" LCSA coculture). These cells resemble monocyte-derived DCs generated in monoculture, but, unlike those, they show a marked upregulation CD86 after treatment with contact allergens. All of the nine sensitizers in this study were correctly identified by CD1a(+)/CD1c(+) DCs in coculture with HaCaT. Among the substances were weak contact allergens such as propylparaben (which is false negative in the local lymph node assay in mice) and resorcinol (which was not detected by CD1a(-)/CD1c(-) DCs in the "classic" LCSA). The level of CD86 upregulation on CD1a(+)/CD1c(+) DCs was higher for most allergens compared to CD1a(-)/CD1c(-) DCs, thus improving the assay's discriminatory power. Three out of four non-sensitizers were also correctly assessed by the coculture assay. A false-positive reaction to caprylic (octanoic) acid confirms earlier results that some fatty acids are able to induce CD86 on DC in vitro. In conclusion, change of the LCSA protocol led to reduction of time and cost while even increasing the assay's sensitivity and discriminatory power.

  2. Different Transcriptional Profiles of Human Monocyte-Derived Dendritic Cells Infected with Distinct Strains of Mycobacterium tuberculosis and Mycobacterium bovis Bacillus Calmette-Guérin

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    Nunzia Sanarico

    2011-01-01

    Full Text Available In order to analyze dendritic cells (DCs activation following infection with different mycobacterial strains, we studied the expression profiles of 165 genes of human monocyte-derived DCs infected with H37Rv, a virulent Mycobacterium tuberculosis (MTB laboratory strain, CMT97, a clinical MTB isolate, Mycobacterium bovis bacillus Calmette-Guérin (BCG, Aventis Pasteur, and BCG Japan, both employed as vaccine against tuberculosis. The analysis of the gene expression reveals that, despite a set of genes similarly modulated, DCs response resulted strain dependent. In particular, H37Rv significantly upregulated EBI3 expression compared with BCG Japan, while it was the only strain that failed to release a significant IL-10 amount. Of note, BCG Japan showed a marked increase in CCR7 and TNF-α expression regarding both MTB strains and it resulted the only strain failing in exponential intracellular growth. Our results suggest that DCs display the ability to elicit a tailored strain-specific immune response.

  3. Prophylactic vaccines are potent activators of monocyte-derived dendritic cells and drive effective anti-tumor responses in melanoma patients at the cost of toxicity.

    Science.gov (United States)

    Bol, Kalijn F; Aarntzen, Erik H J G; Pots, Jeanette M; Olde Nordkamp, Michel A M; van de Rakt, Mandy W M M; Scharenborg, Nicole M; de Boer, Annemiek J; van Oorschot, Tom G M; Croockewit, Sandra A J; Blokx, Willeke A M; Oyen, Wim J G; Boerman, Otto C; Mus, Roel D M; van Rossum, Michelle M; van der Graaf, Chantal A A; Punt, Cornelis J A; Adema, Gosse J; Figdor, Carl G; de Vries, I Jolanda M; Schreibelt, Gerty

    2016-03-01

    Dendritic cell (DC)-based immunotherapy is explored worldwide in cancer patients, predominantly with DC matured with pro-inflammatory cytokines and prostaglandin E2. We studied the safety and efficacy of vaccination with monocyte-derived DC matured with a cocktail of prophylactic vaccines that contain clinical-grade Toll-like receptor ligands (BCG, Typhim, Act-HIB) and prostaglandin E2 (VAC-DC). Stage III and IV melanoma patients were vaccinated via intranodal injection (12 patients) or combined intradermal/intravenous injection (16 patients) with VAC-DC loaded with keyhole limpet hemocyanin (KLH) and mRNA encoding tumor antigens gp100 and tyrosinase. Tumor antigen-specific T cell responses were monitored in blood and skin-test infiltrating-lymphocyte cultures. Almost all patients mounted prophylactic vaccine- or KLH-specific immune responses. Both after intranodal injection and after intradermal/intravenous injection, tumor antigen-specific immune responses were detected, which coincide with longer overall survival in stage IV melanoma patients. VAC-DC induce local and systemic CTC grade 2 and 3 toxicity, which is most likely caused by BCG in the maturation cocktail. The side effects were self-limiting or resolved upon a short period of systemic steroid therapy. We conclude that VAC-DC can induce functional tumor-specific responses. Unfortunately, toxicity observed after vaccination precludes the general application of VAC-DC, since in DC maturated with prophylactic vaccines BCG appears to be essential in the maturation cocktail.

  4. Monocyte-derived dendritic cells induce a house dust mite-specific Th2 allergic inflammation in the lung of humanized SCID mice: involvement of CCR7.

    Science.gov (United States)

    Hammad, Hamida; Lambrecht, Bart N; Pochard, Pierre; Gosset, Philippe; Marquillies, Philippe; Tonnel, André-Bernard; Pestel, Joël

    2002-08-01

    In rodents, airway dendritic cells (DCs) capture inhaled Ag, undergo maturation, and migrate to the draining mediastinal lymph nodes (MLN) to initiate the Ag-specific T cell response. However, the role of human DCs in the pathogenesis of the Th2 cell-mediated disease asthma remains to be clarified. Here, by using SCID mice engrafted with T cells from either house dust mite (HDM)-allergic patients or healthy donors, we show that DCs pulsed with Der p 1, one of the major allergens of HDM, and injected intratracheally into naive animals migrated into the MLN. In the MLN, Der p 1-pulsed DCs from allergic patients induced the proliferation of IL-4-producing CD4(+) T cells, whereas those from healthy donors induced IFN-gamma-secreting cells. In reconstituted human PBMC-reconstituted SCID mice primed with pulsed DCs from allergic patients, repeated exposure to aerosols of HDM induced 1) a strong pulmonary inflammatory reaction rich in T cells and eosinophils, 2) an increase in IL-4 and IL-5 production in the lung lavage fluid, and 3) increased IgE production compared with that in mice primed with unpulsed DCs. All these effects were reduced following in vivo neutralization of the CCR7 ligand secondary lymphoid tissue chemokine. These data in human PBMC-reconstituted SCID mice show that monocyte-derived DCs might play a key role in the pathogenesis of the pulmonary allergic response by inducing Th2 effector function following migration to the MLN.

  5. Leishmania mexicana promastigotes down regulate JNK and p-38 MAPK activation: Role in the inhibition of camptothecin-induced apoptosis of monocyte-derived dendritic cells.

    Science.gov (United States)

    Rodríguez-González, Jorge; Wilkins-Rodríguez, Arturo; Argueta-Donohué, Jesús; Aguirre-García, Magdalena; Gutiérrez-Kobeh, Laila

    2016-04-01

    Dendritic cells (DC) are one of the principal host cells of the obligate intracellular parasite Leishmania. Inhibition of host cell apoptosis is a strategy employed by multiple pathogens to ensure their survival in the infected cell. We have previously shown that the infection of monocyte-derived dendritic cells (moDC) with Leishmania mexicana inhibits campthotecin-induced apoptosis. Nevertheless, the mechanisms involved in the inhibition of apoptosis of dendritic cells by Leishmania have not been established. Mitogen-activated protein kinases (MAPK) are key participants in the process of apoptosis and different species of Leishmania have been shown to regulate these kinases. In the present study, we analyzed the effect of L. mexicana promastigotes in the activation of JNK and p38 MAP kinase and their participation in the inhibition of apoptosis. The infection of moDC with L. mexicana promastigotes diminished significantly the phosphorylation of the MAP kinases JNK and p38. The inhibition of both kinases diminished DNA fragmentation, but in a major extent was the reduction of DNA fragmentation when JNK was inhibited. The capacity of L. mexicana promastigotes to diminish MAP kinases activation is probably one of the strategies employed to delay apoptosis induction in the infected moDC and may have implications for Leishmania pathogenesis by favoring the invasion of its host and the persistence of the parasite in the infected cells.

  6. Induction of Th17 Lymphocytes and Treg Cells by Monocyte-Derived Dendritic Cells in Patients with Rheumatoid Arthritis and Systemic Lupus Erythematosus

    Science.gov (United States)

    Estrada-Capetillo, Lizbeth; Hernández-Castro, Berenice; Monsiváis-Urenda, Adriana; Alvarez-Quiroga, Crisol; Layseca-Espinosa, Esther; Abud-Mendoza, Carlos; Baranda, Lourdes; Urzainqui, Ana; Sánchez-Madrid, Francisco; González-Amaro, Roberto

    2013-01-01

    Dendritic cells (DCs) have a key role in the regulation of immune response. We herein explored, in patients with inflammatory diseases, the role of monocyte derived DC's (mo-DCs) on the generation of Th17 and T regulatory (Treg) lymphocytes. Peripheral blood was obtained from thirty-five patients with rheumatoid arthritis (RA), twelve with systemic lupus erythematosus (SLE), and twenty healthy subjects. Mo-DCs were generated under standard (IL-4/GM-CSF) or tolerogenic (IL-4/GM-CSF plus recombinant P-selectin or PD-1 or IL-10) conditions, and their ability to induce Th17 and Treg lymphocytes was tested. We detected that mo-DCs from patients with RA showed an enhanced release of IL-6 and IL-23 as well as an increased capability to induce Th17 cells. Although mo-DCs from SLE patients also released high levels of IL-6/IL-23, it did not show an increased ability to induce Th17 lymphocytes. In addition, mo-DCs, from patients with RA and SLE generated under the engagement of PSGL-1, showed a defective capability to induce Foxp3+ Treg cells. A similar phenomenon was observed in SLE, when DC's cells were generated under PDL-1 engagement. Our data indicate that DCs from patients with rheumatic inflammatory disease show an aberrant function that may have an important role in the pathogenesis of these conditions. PMID:24288552

  7. Induction of Th17 Lymphocytes and Treg Cells by Monocyte-Derived Dendritic Cells in Patients with Rheumatoid Arthritis and Systemic Lupus Erythematosus

    Directory of Open Access Journals (Sweden)

    Lizbeth Estrada-Capetillo

    2013-01-01

    Full Text Available Dendritic cells (DCs have a key role in the regulation of immune response. We herein explored, in patients with inflammatory diseases, the role of monocyte derived DC’s (mo-DCs on the generation of Th17 and T regulatory (Treg lymphocytes. Peripheral blood was obtained from thirty-five patients with rheumatoid arthritis (RA, twelve with systemic lupus erythematosus (SLE, and twenty healthy subjects. Mo-DCs were generated under standard (IL-4/GM-CSF or tolerogenic (IL-4/GM-CSF plus recombinant P-selectin or PD-1 or IL-10 conditions, and their ability to induce Th17 and Treg lymphocytes was tested. We detected that mo-DCs from patients with RA showed an enhanced release of IL-6 and IL-23 as well as an increased capability to induce Th17 cells. Although mo-DCs from SLE patients also released high levels of IL-6/IL-23, it did not show an increased ability to induce Th17 lymphocytes. In addition, mo-DCs, from patients with RA and SLE generated under the engagement of PSGL-1, showed a defective capability to induce Foxp3+ Treg cells. A similar phenomenon was observed in SLE, when DC’s cells were generated under PDL-1 engagement. Our data indicate that DCs from patients with rheumatic inflammatory disease show an aberrant function that may have an important role in the pathogenesis of these conditions.

  8. Development of a standardized protocol for reproducible generation of matured monocyte-derived dendritic cells suitable for clinical application.

    Science.gov (United States)

    Bohnenkamp, H R; Noll, T

    2003-09-01

    There is increasing interest in the generation of dendritic cells (DC) for cancer immunotherapy. In order to utilize DC in clinical trials it is necessary to have standardized, reproducible and easy to use protocols. We describe here the process development for the generation of DC as the result of investigation of culture conditions as well as consumption rates of medium and cytokines. Our studies demonstrate that highly viable DC (93 +/- 2%) can be produced from CD14(+) enriched monocytes via immunomagnetic beads in a high yield (31 +/- 6%) with X-VIVO 15, 400 U ml(-1) GM-CSF and 2000 U ml(-1) IL-4 without serum and feeding. For the maturation of DC different cocktails (TNF-alpha, IL-1beta, IL-6, PGE(2) and TNF-alpha, PGE(2)) were compared. In both cases cells expressed typical surface molecules of mature DC and induced high proliferative responses in mixed lymphocyte reactions which led to IFN-gamma producing T-lymphocytes. The data suggest that the use of this optimized, easy to use protocol results in highly mature DC.

  9. Generation of functional monocyte-derived fast dendritic cells suitable for clinical application in the absence of interleukin-6.

    Science.gov (United States)

    Ramadan, Gamal

    2011-10-01

    To develop dendritic cells (DCs)-based immunotherapy for cancer patients, it is necessary to have a standardized, reproducible, fast, and easy to use protocol for in vitro generation of fully functional DCs. Recently, a new strategy was described for differentiation and maturation of human monocyte (Mo)-derived fast-DCs with full T cell stimulatory capacity within only 48-72 h of in vitro culture. Interleukin (IL)-6 plus tumour necrosis factor (TNF)-α, IL-1β, and prostaglandin (PG)-E(2) were used in this strategy to induce maturation of the generated DCs. The present study further modifies this strategy by excluding IL-6 from the cytokines cocktail used for DCs maturation. The results showed that maturation of fast-DCs without IL-6 did not significantly alter the morphology, phenotype and the yield of mature DCs (P > 0.05, compared with those generated with IL-6). Moreover, fast-DCs generated without IL-6 are functional antigen presenting cells, have the ability to induce tetanus toxoid-specific autologous T cell proliferation, and are suitable for gene delivery through adenoviral vector transduction as those generated with IL-6 (P > 0.05). In conclusion, the present study proves that fully mature and functional Mo-derived fast-DCs can be generated in vitro without adding IL-6, which not only reduces the number of required recombinant cytokines, but may also resemble DCs development in vivo more closely.

  10. Differential Constitutive and Cytokine-Modulated Expression of Human Toll-like Receptors in Primary Neutrophils, Monocytes, and Macrophages

    Directory of Open Access Journals (Sweden)

    D. Shane O'Mahony, Uyenvy Pham, Ramesh Iyer, Thomas R. Hawn, W. Conrad Liles

    2008-01-01

    Full Text Available Human Toll-like receptors (TLRs comprise a family of proteins that recognizes pathogen-associated molecular patterns (PAMPs and initiates host innate immune responses. Neutrophils, monocytes, and macrophages are critical cellular components of the human innate immune system. Proinflammatory cytokines, such as granulocyte colony-stimulating factor (G-CSF, granulocyte-macrophage colony-stimulating factor (GM-CSF, macrophage colony-stimulating factor (M-CSF, and interferon-γ (IFN-γ, have been shown to up-regulate microbicidal activity in these effector cells of innate immunity. Currently, the cellular and molecular mechanisms responsible for these effects are not completely understood. We hypothesized that these cytokines may up-regulate TLR expression as a mechanism to facilitate microbial recognition and augment the innate immune response. Using quantitative realtime rt-PCR technology, we examined constitutive expression of TLR2, TLR4, TLR5, and TLR9 mRNA and the effects of G-CSF, GM-CSF, M-CSF, and IFN-γ on TLR mRNA expression in purified populations of normal human neutrophils, monocytes, and monocyte-derived macrophages. Relative constitutive expression of TLR2, TLR4, and TLR9 was similar in neutrophils and monocytes. Constitutive expression of TLR5 was less in neutrophils compared to monocytes. Constitutive expression of TLR4 was greater and that of TLR9 lower in monocyte-derived macrophages compared to monocytes. Of the cytokines examined, IFN-γ and GM-CSF caused the greatest effects on TLR expression. IFN- γ up-regulated TLR2 and TLR4 in neutrophils and monocytes. GM-CSF up-regulated expression of TLR2 and TLR4 in neutrophils and TLR2 in monocytes. TLR5 was down-regulated by inflammatory cytokines in monocytes. These results suggest a potential role for IFN- γ and/or GM-CSF as therapeutic immunomodulators of the host defense to infection.

  11. 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.

  12. 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, ...Show Monocyte/macrophage traffic in HIV and SIV encephalitis. PubmedID 12960230 Title Monocyte/macrophage traffic

  13. Ly6Chi monocyte recruitment is responsible for Th2 associated host-protective macrophage accumulation in liver inflammation due to schistosomiasis.

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    Marcia Nascimento

    2014-08-01

    Full Text Available Accumulation of M2 macrophages in the liver, within the context of a strong Th2 response, is a hallmark of infection with the parasitic helminth, Schistosoma mansoni, but the origin of these cells is unclear. To explore this, we examined the relatedness of macrophages to monocytes in this setting. Our data show that both monocyte-derived and resident macrophages are engaged in the response to infection. Infection caused CCR2-dependent increases in numbers of Ly6Chi monocytes in blood and liver and of CX3CR1+ macrophages in diseased liver. Ly6Chi monocytes recovered from liver had the potential to differentiate into macrophages when cultured with M-CSF. Using pulse chase BrdU labeling, we found that most hepatic macrophages in infected mice arose from monocytes. Consistent with this, deletion of monocytes led to the loss of a subpopulation of hepatic CD11chi macrophages that was present in infected but not naïve mice. This was accompanied by a reduction in the size of egg-associated granulomas and significantly exacerbated disease. In addition to the involvement of monocytes and monocyte-derived macrophages in hepatic inflammation due to infection, we observed increased incorporation of BrdU and expression of Ki67 and MHC II in resident macrophages, indicating that these cells are participating in the response. Expression of both M2 and M1 marker genes was increased in liver from infected vs. naive mice. The M2 fingerprint in the liver was not accounted for by a single cell type, but rather reflected expression of M2 genes by various cells including macrophages, neutrophils, eosinophils and monocytes. Our data point to monocyte recruitment as the dominant process for increasing macrophage cell numbers in the liver during schistosomiasis.

  14. Advanced glycation end products inhibit both infection and transmission in trans of HIV-1 from monocyte-derived dendritic cells to autologous T cells.

    Science.gov (United States)

    Nasreddine, Nadine; Borde, Chloé; Gozlan, Joël; Bélec, Laurent; Maréchal, Vincent; Hocini, Hakim

    2011-05-15

    Highly active antiretroviral therapy is associated with carbohydrate metabolic alterations that may lead to diabetes. One consequence of hyperglycemia is the formation of advanced glycation end products (AGEs) that are involved in diabetes complications. We investigated the impact of AGEs on the infection of monocyte-derived dendritic cells (MDDCs) by HIV-1 and the ability of MDDCs to transmit the virus to T cells. We showed that AGEs could inhibit infection of MDDCs with primary R5-tropic HIV-1(Ba-L) by up to 85 ± 9.2% and with primary X4-tropic HIV-1(VN44) by up to 60 ± 8.5%. This inhibitory effect of AGEs was not prevented by a neutralizing anti-receptor for advanced glycation end products (anti-RAGE) Ab, demonstrating a RAGE-independent mechanism. Moreover, AGEs inhibited by 70-80% the transmission in trans of the virus to CD4 T cells. Despite the inhibitory effect of AGEs on both MDDC infection and virus transmission in trans, no inhibition of virus attachment to cell membrane was observed, confirming that attachment and transmission of the virus involve independent mechanisms. The inhibitory effect of AGEs on infection was associated with a RAGE-independent downregulation of CD4 at the cell membrane and by a RAGE-dependent repression of the CXCR4 and CCR5 HIV-1 receptors. AGEs induce the secretion of proinflammatory cytokines IL-6, TNF-α, and IL-12, but not RANTES or MIP-1α, and did not lead to MDDC maturation as demonstrated by the lack of expression of the CD83 molecule. Taken together, our results suggest that AGEs can play an inhibiting role in HIV-1 infection in patients who accumulate circulating AGEs, including patients treated with protease inhibitors that developed diabetes.

  15. Human XCR1+ dendritic cells derived in vitro from CD34+ progenitors closely resemble blood dendritic cells, including their adjuvant responsiveness, contrary to monocyte-derived dendritic cells.

    Science.gov (United States)

    Balan, Sreekumar; Ollion, Vincent; Colletti, Nicholas; Chelbi, Rabie; Montanana-Sanchis, Frédéric; Liu, Hong; Vu Manh, Thien-Phong; Sanchez, Cindy; Savoret, Juliette; Perrot, Ivan; Doffin, Anne-Claire; Fossum, Even; Bechlian, Didier; Chabannon, Christian; Bogen, Bjarne; Asselin-Paturel, Carine; Shaw, Michael; Soos, Timothy; Caux, Christophe; Valladeau-Guilemond, Jenny; Dalod, Marc

    2014-08-15

    Human monocyte-derived dendritic cell (MoDC) have been used in the clinic with moderately encouraging results. Mouse XCR1(+) DC excel at cross-presentation, can be targeted in vivo to induce protective immunity, and share characteristics with XCR1(+) human DC. Assessment of the immunoactivation potential of XCR1(+) human DC is hindered by their paucity in vivo and by their lack of a well-defined in vitro counterpart. We report in this study a protocol generating both XCR1(+) and XCR1(-) human DC in CD34(+) progenitor cultures (CD34-DC). Gene expression profiling, phenotypic characterization, and functional studies demonstrated that XCR1(-) CD34-DC are similar to canonical MoDC, whereas XCR1(+) CD34-DC resemble XCR1(+) blood DC (bDC). XCR1(+) DC were strongly activated by polyinosinic-polycytidylic acid but not LPS, and conversely for MoDC. XCR1(+) DC and MoDC expressed strikingly different patterns of molecules involved in inflammation and in cross-talk with NK or T cells. XCR1(+) CD34-DC but not MoDC efficiently cross-presented a cell-associated Ag upon stimulation by polyinosinic-polycytidylic acid or R848, likewise to what was reported for XCR1(+) bDC. Hence, it is feasible to generate high numbers of bona fide XCR1(+) human DC in vitro as a model to decipher the functions of XCR1(+) bDC and as a potential source of XCR1(+) DC for clinical use.

  16. Histamine Regulates Actin Cytoskeleton in Human Toll-like Receptor 4-activated Monocyte-derived Dendritic Cells Tuning CD4+ T Lymphocyte Response.

    Science.gov (United States)

    Aldinucci, Alessandra; Bonechi, Elena; Manuelli, Cinzia; Nosi, Daniele; Masini, Emanuela; Passani, Maria Beatrice; Ballerini, Clara

    2016-07-08

    Histamine, a major mediator in allergic diseases, differentially regulates the polarizing ability of dendritic cells after Toll-like receptor (TLR) stimulation, by not completely explained mechanisms. In this study we investigated the effects of histamine on innate immune reaction during the response of human monocyte-derived DCs (mDCs) to different TLR stimuli: LPS, specific for TLR4, and Pam3Cys, specific for heterodimer molecule TLR1/TLR2. We investigated actin remodeling induced by histamine together with mDCs phenotype, cytokine production, and the stimulatory and polarizing ability of Th0. By confocal microscopy and RT-PCR expression of Rac1/CdC42 Rho GTPases, responsible for actin remodeling, we show that histamine selectively modifies actin cytoskeleton organization induced by TLR4, but not TLR2 and this correlates with increased IL4 production and decreased IFNγ by primed T cells. We also demonstrate that histamine-induced cytoskeleton organization is at least in part mediated by down-regulation of small Rho GTPase CdC42 and the protein target PAK1, but not by down-regulation of Rac1. The presence and relative expression of histamine receptors HR1-4 and TLRs were determined as well. Independently of actin remodeling, histamine down-regulates IL12p70 and CXCL10 production in mDCs after TLR2 and TLR4 stimulation. We also observed a trend of IL10 up-regulation that, despite previous reports, did not reach statistical significance.

  17. Evaluation of the sensitizing potential of antibiotics in vitro using the human cell lines THP-1 and MUTZ-LC and primary monocyte-derived dendritic cells.

    Science.gov (United States)

    Sebastian, Katrin; Ott, Hagen; Zwadlo-Klarwasser, Gabriele; Skazik-Voogt, Claudia; Marquardt, Yvonne; Czaja, Katharina; Merk, Hans F; Baron, Jens Malte

    2012-08-01

    Since the 7th amendment to the EU cosmetics directive foresees a complete ban on animal testing, alternative in vitro methods have been established to evaluate the sensitizing potential of small molecular weight compounds. To find out whether these novel in vitro assays are also capable to predict the sensitizing potential of small molecular weight drugs, model compounds such as beta-lactams and sulfonamides - which are the most frequent cause of adverse drug reactions - were co-incubated with THP-1, MUTZ-LC, or primary monocyte-derived dendritic cells for 48 h and subsequent expression of selected marker genes (IL-8, IL-1β, CES1, NQO1, GCLM, PIR and TRIM16) was studied by real time PCR. Benzylpenicillin and phenoxymethylpenicillin were recognized as sensitizing compounds because they are capable to induce the mRNA expression of these genes in moDCs and, except for IL-8, in THP-1 cells but not in MUTZ-LC. Ampicillin stimulated the expression of some marker genes in moDCs and THP-1 cells. SMX did not affect the expression of these genes in THP-1, however, in moDCs, at least PIR was enhanced and there was an increase of the release of IL-8. These data reveal that novel in vitro DC based assays might play a role in the evaluation of the allergenic potential of novel drug compounds, but these systems seem to lack the ability to detect the sensitizing potential of prohaptens that require metabolic activation prior to sensitization and moDCs seem to be superior with regard to the sensitivity compared with THP-1 and MUTZ-3 cell lines.

  18. Engineering monocyte-derived dendritic cells to secrete interferon-α enhances their ability to promote adaptive and innate anti-tumor immune effector functions.

    Science.gov (United States)

    Willemen, Yannick; Van den Bergh, Johan M J; Lion, Eva; Anguille, Sébastien; Roelandts, Vicky A E; Van Acker, Heleen H; Heynderickx, Steven D I; Stein, Barbara M H; Peeters, Marc; Figdor, Carl G; Van Tendeloo, Viggo F I; de Vries, I Jolanda; Adema, Gosse J; Berneman, Zwi N; Smits, Evelien L J

    2015-07-01

    Dendritic cell (DC) vaccination has demonstrated potential in clinical trials as a new effective cancer treatment, but objective and durable clinical responses are confined to a minority of patients. Interferon (IFN)-α, a type-I IFN, can bolster anti-tumor immunity by restoring or increasing the function of DCs, T cells and natural killer (NK) cells. Moreover, type-I IFN signaling on DCs was found to be essential in mice for tumor rejection by the innate and adaptive immune system. Targeted delivery of IFN-α by DCs to immune cells could boost the generation of anti-tumor immunity, while avoiding the side effects frequently associated with systemic administration. Naturally circulating plasmacytoid DCs, major producers of type-I IFN, were already shown capable of inducing tumor antigen-specific T cell responses in cancer patients without severe toxicity, but their limited number complicates their use in cancer vaccination. In the present work, we hypothesized that engineering easily generated human monocyte-derived mature DCs to secrete IFN-α using mRNA electroporation enhances their ability to promote adaptive and innate anti-tumor immunity. Our results show that IFN-α mRNA electroporation of DCs significantly increases the stimulation of tumor antigen-specific cytotoxic T cell as well as anti-tumor NK cell effector functions in vitro through high levels of IFN-α secretion. Altogether, our findings mark IFN-α mRNA-electroporated DCs as potent inducers of both adaptive and innate anti-tumor immunity and pave the way for clinical trial evaluation in cancer patients.

  19. Low CCR7-mediated migration of human monocyte derived dendritic cells in response to human respiratory syncytial virus and human metapneumovirus.

    Directory of Open Access Journals (Sweden)

    Cyril Le Nouën

    2011-06-01

    Full Text Available Human respiratory syncytial virus (HRSV and, to a lesser extent, human metapneumovirus (HMPV and human parainfluenza virus type 3 (HPIV3, can re-infect symptomatically throughout life without significant antigenic change, suggestive of incomplete or short-lived immunity. In contrast, re-infection by influenza A virus (IAV largely depends on antigenic change, suggestive of more complete immunity. Antigen presentation by dendritic cells (DC is critical in initiating the adaptive immune response. Antigen uptake by DC induces maturational changes that include decreased expression of the chemokine receptors CCR1, CCR2, and CCR5 that maintain DC residence in peripheral tissues, and increased expression of CCR7 that mediates the migration of antigen-bearing DC to lymphatic tissue. We stimulated human monocyte-derived DC (MDDC with virus and found that, in contrast to HPIV3 and IAV, HMPV and HRSV did not efficiently decrease CCR1, 2, and 5 expression, and did not efficiently increase CCR7 expression. Consistent with the differences in CCR7 mRNA and protein expression, MDDC stimulated with HRSV or HMPV migrated less efficiently to the CCR7 ligand CCL19 than did IAV-stimulated MDDC. Using GFP-expressing recombinant virus, we showed that the subpopulation of MDDC that was robustly infected with HRSV was particularly inefficient in chemokine receptor modulation. HMPV- or HRSV-stimulated MDDC responded to secondary stimulation with bacterial lipopolysaccharide or with a cocktail of proinflammatory cytokines by increasing CCR7 and decreasing CCR1, 2 and 5 expression, and by more efficient migration to CCL19, suggesting that HMPV and HRSV suboptimally stimulate rather than irreversibly inhibit MDDC migration. This also suggests that the low concentration of proinflammatory cytokines released from HRSV- and HMPV-stimulated MDDC is partly responsible for the low CCR7-mediated migration. We propose that inefficient migration of HRSV- and HMPV-stimulated DC to

  20. Broad antiviral activity of carbohydrate-binding agents against the four serotypes of dengue virus in monocyte-derived dendritic cells.

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    Marijke M F Alen

    Full Text Available BACKGROUND: Dendritic cells (DC, present in the skin, are the first target cells of dengue virus (DENV. Dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (DC-SIGN is present on DC and recognizes N-glycosylation sites on the E-glycoprotein of DENV. Thus, the DC-SIGN/E-glycoprotein interaction can be considered as an important target for inhibitors of viral replication. We evaluated various carbohydrate-binding agents (CBAs against all four described serotypes of DENV replication in Raji/DC-SIGN(+ cells and in monocyte-derived DC (MDDC. METHODOLOGY/PRINCIPAL FINDINGS: A dose-dependent anti-DENV activity of the CBAs Hippeastrum hybrid (HHA, Galanthus nivalis (GNA and Urtica dioica (UDA, but not actinohivin (AH was observed against all four DENV serotypes as analyzed by flow cytometry making use of anti-DENV antibodies. Remarkably, the potency of the CBAs against DENV in MDDC cultures was significantly higher (up to 100-fold than in Raji/DC-SIGN(+ cells. Pradimicin-S (PRM-S, a small-size non-peptidic CBA, exerted antiviral activity in MDDC but not in Raji/DC-SIGN(+ cells. The CBAs act at an early step of DENV infection as they bind to the viral envelope of DENV and subsequently prevent virus attachment. Only weak antiviral activity of the CBAs was detected when administered after the virus attachment step. The CBAs were also able to completely prevent the cellular activation and differentiation process of MDDC induced upon DENV infection. CONCLUSIONS/SIGNIFICANCE: The CBAs exerted broad spectrum antiviral activity against the four DENV serotypes, laboratory-adapted viruses and low passage clinical isolates, evaluated in Raji/DC-SIGN(+ cells and in primary MDDC.

  1. Th2 polarization by Der p 1--pulsed monocyte-derived dendritic cells is due to the allergic status of the donors.

    Science.gov (United States)

    Hammad, H; Charbonnier, A S; Duez, C; Jacquet, A; Stewart, G A; Tonnel, A B; Pestel, J

    2001-08-15

    The polarization of the immune response toward a Th2 or a Th1 profile can be mediated by dendritic cells (DCs) following antigen presentation and interaction with T cells. Costimulatory molecules such as CD80 and CD86 expressed by DCs, the polarizing cytokine environment during DC--T-cell interaction, and also the nature of the antigen are critical in the orientation of the immune response. In this study, the effect of the cysteine protease Der p 1, one of the major allergens of the house dust mite Dermatophagoides pteronyssinus, on these different parameters was evaluated comparatively on monocyte-derived DCs obtained from healthy donors, from pollen-sensitive patients, or from patients sensitive to Dermatophagoides pteronyssinus. Results showed that Der p 1 induced an increase in CD86 expression only on DCs from house dust mite--sensitive patients. This was also associated with a higher capacity to induce T-cell proliferation, a rapid increase in the production of proinflammatory cytokines, tumor necrosis factor--alpha and interleukin (IL)-1 beta, and the type 2 cytokine IL-10. No changes in the release of IL-12 p70 were induced by Der p 1. Finally, purified T cells from house dust mite-sensitive patients stimulated by autologous Der p 1--pulsed DCs preferentially produced IL-4 rather than interferon-gamma. These effects were abolished in the presence of the inactive precursor of Der p 1 (ProDer p 1). Taken together, these data suggest that DCs from house dust mite--sensitive patients, in contrast to DCs from healthy donors and from pollen-sensitive patients, exposed to Der p 1 play a pivotal role in the enhancement of the Th2 response associated with the allergic reaction developed in response to house dust mite exposure. (Blood. 2001;98:1135-1141)

  2. Human monocyte-derived dendritic cells from leukoreduction system chambers after plateletpheresis are functional in an in vitro co-culture assay with intestinal epithelial cells.

    Science.gov (United States)

    Tiscornia, Inés; Sánchez-Martins, Viviana; Hernández, Ana; Bollati-Fogolín, Mariela

    2012-10-31

    PP as an alternative source of PBMC, to be used in co-culture systems with IEC. The novelty of this protocol is the combination of the blood monocyte source with a simple and fast differentiation method to obtain DC, and their use in a combined culture with IEC and LAB to model microbial-host interaction. Since the initial PP volume is ten times lower than that of BC, the use of PP minimizes biological residue generation and reagent consumption. In addition, monocyte-derived DC from PP were suitable for use in co-culture assays as a first screening step to study the immunomodulatory properties of LAB.

  3. Mechanism involved in interleukin-21-induced phagocytosis in human monocytes and macrophages.

    Science.gov (United States)

    Vallières, F; Girard, D

    2017-02-01

    The interleukin (IL)-21/IL-21 receptor (R) is a promising system to be exploited for the development of therapeutic strategies. Although the biological activities of IL-21 and its cell signalling events have been largely studied in immunocytes, its interaction with human monocytes and macrophages have been neglected. Previously, we reported that IL-21 enhances Fc gamma receptor (FcRγ)-mediated phagocytosis in human monocytes and in human monocyte-derived macrophages (HMDM) and identified Syk as a novel molecular target of IL-21. Here, we elucidate further how IL-21 promotes phagocytosis in these cells. Unlike its ability to enhance phagocytosis of opsonized sheep red blood cells (SRBCs), IL-21 did not promote phagocytosis of Escherichia coli and zymosan by monocytes and did not alter the cell surface expression of CD16, CD32 and CD64. In HMDM, IL-21 was found to enhance phagocytosis of zymosan. In addition, we found that IL-21 activates p38, protein kinase B (Akt), signal transducer and activator of transcription (STAT)-1 and STAT-3 in monocytes and HMDM. Using a pharmacological approach, we demonstrate that IL-21 enhances phagocytosis by activating some mitogen-activated protein kinases (MAPKs) and phosphoinositide 3-kinase (PI3K)-Akt and Janus kinase (JAK)-STAT pathways. These results obtained in human monocytes and macrophages have to be considered for a better exploitation of the IL-21/IL-21R system for therapeutic purposes. © 2016 British Society for Immunology.

  4. Macrophage dynamics are regulated by local macrophage proliferation and monocyte recruitment in injured pancreas.

    Science.gov (United States)

    Van Gassen, Naomi; Van Overmeire, Eva; Leuckx, Gunter; Heremans, Yves; De Groef, Sofie; Cai, Ying; Elkrim, Yvon; Gysemans, Conny; Stijlemans, Benoît; Van de Casteele, Mark; De Baetselier, Patrick; De Leu, Nico; Heimberg, Harry; Van Ginderachter, Jo A

    2015-05-01

    Pancreas injury by partial duct ligation (PDL) activates a healing response, encompassing β-cell neogenesis and proliferation. Macrophages (MΦs) were recently shown to promote β-cell proliferation after PDL, but they remain poorly characterized. We assessed myeloid cell diversity and the factors driving myeloid cell dynamics following acute pancreas injury by PDL. In naive and sham-operated pancreas, the myeloid cell compartment consisted mainly of two distinct tissue-resident MΦ types, designated MHC-II(lo) and MHC-II(hi) MΦs, the latter being predominant. MHC-II(lo) and MHC-II(hi) pancreas MΦs differed at the molecular level, with MHC-II(lo) MΦs being more M2-activated. After PDL, there was an early surge of Ly6C(hi) monocyte infiltration in the pancreas, followed by a transient MHC-II(lo) MΦ peak and ultimately a restoration of the MHC-II(hi) MΦ-dominated steady-state equilibrium. These intricate MΦ dynamics in PDL pancreas depended on monocyte recruitment by C-C chemokine receptor 2 and macrophage-colony stimulating factor receptor as well as on macrophage-colony stimulating factor receptor-dependent local MΦ proliferation. Functionally, MHC-II(lo) MΦs were more angiogenic. We further demonstrated that, at least in C-C chemokine receptor 2-KO mice, tissue MΦs, rather than Ly6C(hi) monocyte-derived MΦs, contributed to β-cell proliferation. Together, our study fully characterizes the MΦ subsets in the pancreas and clarifies the complex dynamics of MΦs after PDL injury. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Monocyte derived dendritic cells generated by IFN-α acquire mature dendritic and natural killer cell properties as shown by gene expression analysis

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    Czibere Akos

    2007-09-01

    Full Text Available Abstract Background Dendritic cell (DC vaccines can induce antitumor immune responses in patients with malignant diseases, while the most suitable DC culture conditions have not been established yet. In this study we compared monocyte derived human DC from conventional cultures containing GM-CSF and IL-4/TNF-α (IL-4/TNF-DC with DC generated by the novel protocol using GM-CSF and IFN-α (IFN-DC. Methods To characterise the molecular differences of both DC preparations, gene expression profiling was performed using Affymetrix microarrays. The data were conformed on a protein level by immunophenotyping, and functional tests for T cell stimulation, migration and cytolytic activity were performed. Results Both methods resulted in CD11c+ CD86+ HLA-DR+ cells with a typical DC morphology that could efficiently stimulate T cells. But gene expression profiling revealed two distinct DC populations. Whereas IL-4/TNF-DC showed a higher expression of genes envolved in phagocytosis IFN-DC had higher RNA levels for markers of DC maturity and migration to the lymph nodes like DCLAMP, CCR7 and CD49d. This different orientation of both DC populations was confined by a 2.3 fold greater migration in transwell experiments (p = 0.01. Most interestingly, IFN-DC also showed higher RNA levels for markers of NK cells such as TRAIL, granzymes, KLRs and other NK cell receptors. On a protein level, intracytoplasmatic TRAIL and granzyme B were observed in 90% of IFN-DC. This translated into a cytolytic activity against K562 cells with a median specific lysis of 26% at high effector cell numbers as determined by propidium iodide uptake, whereas IL-4/TNF-DC did not induce any tumor cell lysis (p = 0.006. Thus, IFN-DC combined characteristics of mature DC and natural killer cells. Conclusion Our results suggest that IFN-DC not only stimulate adaptive but also mediate innate antitumor immune responses. Therefore, IFN-DC should be evaluated in clinical vaccination trials. In

  6. Monocyte to macrophage differentiation goes along with modulation of the plasmalogen pattern through transcriptional regulation.

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    Stefan Wallner

    Full Text Available BACKGROUND: Dysregulation of monocyte-macrophage differentiation is a hallmark of vascular and metabolic diseases and associated with persistent low grade inflammation. Plasmalogens represent ether lipids that play a role in diabesity and previous data show diminished plasmalogen levels in obese subjects. We therefore analyzed transcriptomic and lipidomic changes during monocyte-macrophage differentiation in vitro using a bioinformatic approach. METHODS: Elutriated monocytes from 13 healthy donors were differentiated in vitro to macrophages using rhM-CSF under serum-free conditions. Samples were taken on days 0, 1, 4 and 5 and analyzed for their lipidomic and transcriptomic profiles. RESULTS: Gene expression analysis showed strong regulation of lipidome-related transcripts. Enzymes involved in fatty acid desaturation and elongation were increasingly expressed, peroxisomal and ER stress related genes were induced. Total plasmalogen levels remained unchanged, while the PE plasmalogen species pattern became more similar to circulating granulocytes, showing decreases in PUFA and increases in MUFA. A partial least squares discriminant analysis (PLS/DA revealed that PE plasmalogens discriminate the stage of monocyte-derived macrophage differentiation. Partial correlation analysis could predict novel potential key nodes including DOCK1, PDK4, GNPTAB and FAM126A that might be involved in regulating lipid and especially plasmalogen homeostasis during differentiation. An in silico transcription analysis of lipid related regulation revealed known motifs such as PPAR-gamma and KLF4 as well as novel candidates such as NFY, RNF96 and Zinc-finger proteins. CONCLUSION: Monocyte to macrophage differentiation goes along with profound changes in the lipid-related transcriptome. This leads to an induction of fatty-acid desaturation and elongation. In their PE-plasmalogen profile macrophages become more similar to granulocytes than monocytes, indicating terminal

  7. Interleukin-27 is a potent inhibitor of cis HIV-1 replication in monocyte-derived dendritic cells via a type I interferon-independent pathway.

    Directory of Open Access Journals (Sweden)

    Qian Chen

    Full Text Available IL-27, a member of the IL-12 family of cytokines, plays an important and diverse role in the function of the immune system. Whilst generally recognized as an anti-inflammatory cytokine, in addition IL-27 has been found to have broad anti-viral effects. Recently, IL-27 has been shown to be a potent inhibitor of HIV-1 infection in CD4+ T cells and macrophages. The main objective of this study was to see whether IL-27 has a similar inhibitory effect on HIV-1 replication in dendritic cells (DCs. Monocytes were differentiated into immature DCs (iDCs and mature DCs (mDCs with standard techniques using a combination of GM-CSF, IL-4 and LPS. Following differentiation, iDCs were infected with HIV-1 and co-cultured in the presence or absence of IL-27. IL-27 treated DCs were shown to be highly potent inhibitors of cis HIV-1, particularly of CCR5 tropic strains. Of note, other IL-12 family members (IL-12, IL-23 and IL-35 had no effect on HIV-1 replication. Microarray studies of IL-27 treated DCs showed no up-regulation of Type I (IFN gene expression. Neutralization of the Type-I IFN receptor had no impact on the HIV inhibition. Lastly, IL-27 mediated inhibition was shown to act post-viral entry and prior to completion of reverse transcription. These results show for the first time that IL-27 is a potent inhibitor of cis HIV-1 infection in DCs by a Type I IFN independent mechanism. IL-27 has previously been reported to inhibit HIV-1 replication in CD4+ T cells and macrophages, thus taken together, this cytokine is a potent anti-HIV agent against all major cell types targeted by the HIV-1 virus and may have a therapeutic role in the future.

  8. 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

  9. Effect of in vitro digested cod liver oil of different quality on oxidative, proteomic and inflammatory responses in the yeast Saccharomyces cerevisiae and human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Larsson, Karin; Istenič, Katja; Wulff, Tune

    2015-01-01

    digested fresh and oxidised cod liver oils in vitro, monitored the levels of lipid peroxidation products and evaluated oxidative, proteomic and inflammatory responses to the two types of digests in the yeast Saccharomyces cerevisiae and human monocyte-derived dendritic cells. RESULTS: Digests of cod liver......BACKGROUND: Upon oxidation of the polyunsaturated fatty acids in fish oil, either before ingestion or, as recently shown, during the gastro-intestinal passage, a cascade of potentially cytotoxic peroxidation products, such as malondialdehyde and 4-hydroxy-2-hexenal, can form. In this study, we...... oil with 22–53 µmol L−1 malondialdehyde and 0.26–3.7 µmol L−1 4-hydroxy-2-hexenal increased intracellular oxidation and cell energy metabolic activity compared to a digested blank in yeast cells and the influence of digests on mitochondrial protein expression was more pronounced for oxidised cod liver...

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

    Institute of Scientific and Technical Information of China (English)

    詹康; 赵倩明; 隋雁南; 封飞飞; 占今舜; 赵国琦

    2016-01-01

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

  11. Accessory cells with a veiled morphology and movement pattern generated from monocytes after avoidance of plastic adherence and of NADPH oxidase activation. A comparison with GM-CSF/IL-4-induced monocyte-derived dendritic cells.

    Science.gov (United States)

    Ruwhof, Cindy; Canning, Martha O; Grotenhuis, Kristel; de Wit, Harm J; Florencia, Zenovia Z; de Haan-Meulman, Meeny; Drexhage, Hemmo A

    2002-07-01

    Veiled cells (VC) present in afferent lymph transport antigen from the periphery to the draining lymph nodes. Although VC in lymph form a heterogeneous population, some of the cells clearly belong on morphological grounds to the Langerhans cell (LC)/ dendritic cell (DC) series. Here we show that culturing monocytes for 24 hrs while avoiding plastic adherence (polypropylene tubes) and avoiding the activation of NADPH oxidase (blocking agents) results in the generation of a population of veiled accessory cells. The generated VC were actively moving cells like lymph-borne VC in vivo. The monocyte (mo)-derived VC population existed of CD14(dim/-) and CD14(brighT) cells. Of these the CD14(dim/-) VC were as good in stimulating allogeneic T cell proliferation as immature DC (iDC) obtained after one week of adherent culture of monocytes in granulocyte-macrophage-colony stimulating factor (GM-CSF)/interleukin (IL)-4. This underscores the accessory cell function of the mo-derived CD14(dim/-) VC. Although the CD14(dim/-)VC had a modest expression of the DC-specific marker CD83 and were positive for S100, expression of the DC-specific markers CD1a, Langerin, DC-SIGN, and DC-LAMP were absent. This indicates that the here generated CD14(dim/-) VC can not be considered as classical LC/DC. It was also impossible to turn the CD14(dim/-) mo-derived VC population into typical DC by culture for one week in GM-CSF/IL-4 or LPS. In fact the cells died tinder such circumstances, gaining some macrophage characteristics before dying. The IL-12 production from mo-derived CD14(dim/-) VC was lower, whereas the production of IL-10 was higher as compared to iDC. Consequently the T cells that were stimulated by these mo-derived VC produced less IFN-gamma as compared with T cells stimulated by iDC. Our data indicate that it is possible to rapidly generate a population of CD14(dim/-) veiled accessory cells from monocytes. The marker pattern and cytokine production of these VC indicate that this

  12. The Bacillus anthracis cholesterol-dependent cytolysin, Anthrolysin O, kills human neutrophils, monocytes and macrophages

    Directory of Open Access Journals (Sweden)

    Rest Richard F

    2006-06-01

    Full Text Available Abstract Background Bacillus anthracis is an animal and human pathogen whose virulence is characterized by lethal and edema toxin, as well as a poly-glutamic acid capsule. In addition to these well characterized toxins, B. anthracis secretes several proteases and phospholipases, and a newly described toxin of the cholesterol-dependent cytolysin (CDC family, Anthrolysin O (ALO. Results In the present studies we show that recombinant ALO (rALO or native ALO, secreted by viable B. anthracis, is lethal to human primary polymorphonuclear leukocytes (PMNs, monocytes, monocyte-derived macrophages (MDMs, lymphocytes, THP-1 monocytic human cell line and ME-180, Detroit 562, and A549 epithelial cells by trypan blue exclusion or lactate dehydrogenase (LDH release viability assays. ALO cytotoxicity is dose and time dependent and susceptibility to ALO-mediated lysis differs between cell types. In addition, the viability of monocytes and hMDMs was assayed in the presence of vegetative Sterne strains 7702 (ALO+, UT231 (ALO-, and a complemented strain expressing ALO, UT231 (pUTE544, and was dependent upon the expression of ALO. Cytotoxicity of rALO is seen as low as 0.070 nM in the absence of serum. All direct cytotoxic activity is inhibited by the addition of cholesterol or serum concentration as low as 10%. Conclusion The lethality of rALO and native ALO on human monocytes, neutrophils, macrophages and lymphocytes supports the idea that ALO may represent a previously unidentified virulence factor of B. anthracis. The study of other factors produced by B. anthracis, along with the major anthrax toxins, will lead to a better understanding of this bacterium's pathogenesis, as well as provide information for the development of antitoxin vaccines for treating and preventing anthrax.

  13. Ly6Chi Monocytes and Their Macrophage Descendants Regulate Neutrophil Function and Clearance in Acetaminophen-Induced Liver Injury

    Science.gov (United States)

    Graubardt, Nadine; Vugman, Milena; Mouhadeb, Odelia; Caliari, Gabriele; Pasmanik-Chor, Metsada; Reuveni, Debby; Zigmond, Ehud; Brazowski, Eli; David, Eyal; Chappell-Maor, Lousie; Jung, Steffen; Varol, Chen

    2017-01-01

    Monocyte-derived macrophages (MoMF) play a pivotal role in the resolution of acetaminophen-induced liver injury (AILI). Timely termination of neutrophil activity and their clearance are essential for liver regeneration following injury. Here, we show that infiltrating Ly6Chi monocytes, their macrophage descendants, and neutrophils spatially and temporally overlap in the centrilobular necrotic areas during the necroinflammatory and resolution phases of AILI. At the necroinflammatory phase, inducible ablation of circulating Ly6Chi monocytes resulted in reduced numbers and fractions of reactive oxygen species (ROS)-producing neutrophils. In alignment with this, neutrophils sorted from monocyte-deficient livers exhibited reduced expression of NADPH oxidase 2. Moreover, human CD14+ monocytes stimulated with lipopolysaccharide or hepatocyte apoptotic bodies directly induced ROS production by cocultured neutrophils. RNA-seq-based transcriptome profiling of neutrophils from Ly6Chi monocyte-deficient versus normal livers revealed 449 genes that were differentially expressed with at least twofold change (p ≤ 0.05). In the absence of Ly6Chi monocytes, neutrophils displayed gene expression alterations associated with decreased innate immune activity and increased cell survival. At the early resolution phase, Ly6Chi monocytes differentiated into ephemeral Ly6Clo MoMF and their absence resulted in significant accumulation of late apoptotic neutrophils. Further gene expression analysis revealed the induced expression of a specific repertoire of bridging molecules and receptors involved with apoptotic cell clearance during the transition from Ly6Chi monocytes to MoMF. Collectively, our findings establish a phase-dependent task division between liver-infiltrating Ly6Chi monocytes and their MoMF descendants with the former regulating innate immune functions and cell survival of neutrophils and the later neutrophil clearance. PMID:28620383

  14. Impairment of in vitro generation of monocyte-derived human dendritic cells by inactivated human immunodeficiency virus-1: Involvement of type I interferon produced from plasmacytoid dendritc cells.

    Science.gov (United States)

    Kodama, Akira; Tanaka, Reiko; Zhang, Li Feng; Adachi, Tetsuya; Saito, Mineki; Ansari, Aftab A; Tanaka, Yuetsu

    2010-06-01

    In an attempt to simplify the protocol of DC generation in vitro, studies conducted herein show that functional DCs could be generated from bulk peripheral blood mononuclear cells (PBMCs) in media containing GM-CSF and IL-4. Interestingly, when PBMCs, but not purified monocytes, were exposed to either CCR5- or CXCR4-tropic inactivated HIV-1 isolates (iHIV-1) at the initiation of the culture, DC yields were significantly reduced in a dose-dependent manner because of monocyte apoptosis. Similar impairment of DC generation was noted using type I IFNs and poly IC not only in cultures of PBMCs but also using highly enriched monocytes. This effect was reversed by antihuman type I IFN receptor, but not by anti-FasL, anti-TRAIL, anti-TNF, or a mixture of these antibodies. iHIV-1-exposed PBMCs, but not monocytes, produced high levels of IFN-alpha but not IFN-beta. PBMCs depleted of CD123(+) plasmacytoid DCs produced low levels of IFN-alpha and were resistant to iHIV-1-mediated DC impairment. Interestingly, exogenously added TNF reversed the impairment by iHIV-1 in the PBMC cultures. In conclusion, the present results indicate that iHIV-1 impairs the in vitro generation of functional DCs from PBMCs through the induction of IFN-alpha from plasmacytoid DCs in a CD4-dependent fashion in the absence of TNF.

  15. Acute exposure to apolipoprotein A1 inhibits macrophage chemotaxis in vitro and monocyte recruitment in vivo

    Science.gov (United States)

    Iqbal, Asif J; Barrett, Tessa J; Taylor, Lewis; McNeill, Eileen; Manmadhan, Arun; Recio, Carlota; Carmineri, Alfredo; Brodermann, Maximillian H; White, Gemma E; Cooper, Dianne; DiDonato, Joseph A; Zamanian-Daryoush, Maryam; Hazen, Stanley L; Channon, Keith M

    2016-01-01

    Apolipoprotein A1 (apoA1) is the major protein component of high-density lipoprotein (HDL) and has well documented anti-inflammatory properties. To better understand the cellular and molecular basis of the anti-inflammatory actions of apoA1, we explored the effect of acute human apoA1 exposure on the migratory capacity of monocyte-derived cells in vitro and in vivo. Acute (20–60 min) apoA1 treatment induced a substantial (50–90%) reduction in macrophage chemotaxis to a range of chemoattractants. This acute treatment was anti-inflammatory in vivo as shown by pre-treatment of monocytes prior to adoptive transfer into an on-going murine peritonitis model. We find that apoA1 rapidly disrupts membrane lipid rafts, and as a consequence, dampens the PI3K/Akt signalling pathway that coordinates reorganization of the actin cytoskeleton and cell migration. Our data strengthen the evidence base for therapeutic apoA1 infusions in situations where reduced monocyte recruitment to sites of inflammation could have beneficial outcomes. DOI: http://dx.doi.org/10.7554/eLife.15190.001 PMID:27572261

  16. Prostaglandin E2 Does Not Modulate CCR7 Expression and Functionality after Differentiation of Blood Monocytes into Macrophages

    Directory of Open Access Journals (Sweden)

    Marc-André Allaire

    2013-01-01

    Full Text Available Previously, we demonstrated that prostaglandin E2 (PGE2 induces C-C chemokine receptor type 7 (CCR7 expression on human monocytes, which stimulates their subsequent migration in response to the CCR7 natural ligands CCL19 and CCL21. In this study, we determined whether PGE2 affects CCR7 expression on macrophages. Flow cytometric analysis and chemotaxis assays were performed on Mono Mac-1-derived macrophage (MDMM-1 as well as unpolarized monocyte-derived macrophages (MDMs to determine the CCR7 expression and functionality in the presence of PGE2. Data revealed that a MDMM-1 exhibited markedly downregulated CCR7 expression and functionality that were partially restored by treatment with PGE2. In MDMs, we observed a drastic downregulation of CCR7 expression and functionality that were unaffected following PGE2 treatment. Our data indicate that monocyte differentiation induces the loss of CCR7 expression and that PGE2 is unable to modulate CCR7 expression and functionality as shown previously in monocytes.

  17. Human monocytes undergo excessive apoptosis following temozolomide activating the ATM/ATR pathway while dendritic cells and macrophages are resistant.

    Directory of Open Access Journals (Sweden)

    Martina Bauer

    Full Text Available Immunodeficiency is a severe therapy-limiting side effect of anticancer chemotherapy resulting from sensitivity of immunocompetent cells to DNA damaging agents. A central role in the immune system is played by monocytes that differentiate into macrophages and dendritic cells (DCs. In this study we compared human monocytes isolated from peripheral blood and cytokine matured macrophages and DCs derived from them and assessed the mechanism of toxicity of the DNA methylating anticancer drug temozolomide (TMZ in these cell populations. We observed that monocytes, but not DCs and macrophages, were highly sensitive to the killing effect of TMZ. Studies on DNA damage and repair revealed that the initial DNA incision was efficient in monocytes while the re-ligation step of base excision repair (BER can not be accomplished, resulting in an accumulation of DNA single-strand breaks (SSBs. Furthermore, monocytes accumulated DNA double-strand breaks (DSBs following TMZ treatment, while DCs and macrophages were able to repair DSBs. Monocytes lack the DNA repair proteins XRCC1, ligase IIIα and PARP-1 whose expression is restored during differentiation into macrophages and DCs following treatment with GM-CSF and GM-CSF plus IL-4, respectively. These proteins play a key role both in BER and DSB repair by B-NHEJ, which explains the accumulation of DNA breaks in monocytes following TMZ treatment. Although TMZ provoked an upregulation of XRCC1 and ligase IIIα, BER was not enhanced likely because PARP-1 was not upregulated. Accordingly, inhibition of PARP-1 did not sensitize monocytes, but monocyte-derived DCs in which strong PARP activation was observed. TMZ induced in monocytes the DNA damage response pathways ATM-Chk2 and ATR-Chk1 resulting in p53 activation. Finally, upon activation of the Fas-receptor and the mitochondrial pathway apoptosis was executed in a caspase-dependent manner. The downregulation of DNA repair in monocytes, resulting in their selective

  18. PAF-degrading acetylhydrolase is preferentially associated with dense LDL and VHDL-1 in human plasma. Catalytic characteristics and relation to the monocyte-derived enzyme.

    Science.gov (United States)

    Tselepis, A D; Dentan, C; Karabina, S A; Chapman, M J; Ninio, E

    1995-10-01

    In human plasma, platelet activating factor (PAF)-degrading acetylhydrolase (acetylhydrolase) is principally transported in association with LDLs and HDLs; this enzyme hydrolyzes PAF and short-chain forms of oxidized phosphatidylcholine, transforming them into lyso-PAF and lysophosphatidylcholine, respectively. We have examined the distribution, catalytic characteristics, and transfer of acetylhydrolase activity among plasma lipoprotein subspecies separated by isopycnic density gradient ultracentrifugation; the possibility that the plasma enzyme may be partially derived from adherent monocytes has also been evaluated. In normolipidemic subjects with Lp(a) levels VHDL-1; d = 1.156 to 1.179 g/mL), representing 23.9 +/- 1.7% and 20.6 +/- 3.2%, respectively, of total plasma activity. The apparent Km values for PAF of the enzyme associated with such lipoproteins were 89.7 +/- 23.4 and 34.8 +/- 4.5 mumol/L for LDL-5 and VHDL-1, respectively: indeed, the Km value for LDL-5 was some 10-fold higher than that of the light LDL-1, LDL-2, and LDL-3 subspecies, whereas the Km of VHDL-1 was some twofold greater than those of the HDL-2 and HDL-3 subspecies. Furthermore, when expressed on the basis of unit plasma volume, the Vmax of the acetylhydrolase associated with LDL-5 was some 150-fold greater than that in LDL-1 (d = 1.019 to 1.023 g/mL). No significant differences in the pH dependence of enzyme activity or in sensitivity to protease inactivation, sulfydryl reagents, the serine protease inhibitor Pefabloc, or the PAF antagonist CV 3988 could be detected between apo B-containing and apo A-I-containing lipoprotein particle subspecies. Incubation of LDL-1 (Km = 8.4 +/- 2.6 mumol/L) and LDL-2 (d = 1.023 to 1.029 g/mL; Km = 8.4 +/- 3.3 mumol/L) subspecies with LDL-5, in which acetylhydrolase had been inactivated by pretreatment with Pefabloc, demonstrated preferential transfer of acetylhydrolase to LDL-5. Acetylhydrolase transferred to LDL-5 from the light LDL subspecies exhibited

  19. Activation and cytokine profile of monocyte derived dendritic cells in leprosy: in vitro stimulation by sonicated Mycobacterium leprae induces decreased level of IL-12p70 in lepromatous leprosy

    Directory of Open Access Journals (Sweden)

    André Flores Braga

    2015-08-01

    Full Text Available Dendritic cells (DCs play a pivotal role in the connection of innate and adaptive immunity of hosts to mycobacterial infection. Studies on the interaction of monocyte-derived DCs (MO-DCs using Mycobacterium leprae in leprosy patients are rare. The present study demonstrated that the differentiation of MOs to DCs was similar in all forms of leprosy compared to normal healthy individuals. In vitro stimulation of immature MO-DCs with sonicated M. leprae induced variable degrees of DC maturation as determined by the increased expression of HLA-DR, CD40, CD80 and CD86, but not CD83, in all studied groups. The production of different cytokines by the MO-DCs appeared similar in all of the studied groups under similar conditions. However, the production of interleukin (IL-12p70 by MO-DCs from lepromatous (LL leprosy patients after in vitro stimulation with M. lepraewas lower than tuberculoid leprosy patients and healthy individuals, even after CD40 ligation with CD40 ligand-transfected cells. The present cumulative findings suggest that the MO-DCs of LL patients are generally a weak producer of IL-12p70 despite the moderate activating properties ofM. leprae. These results may explain the poor M. leprae-specific cell-mediated immunity in the LL type of leprosy.

  20. Activation and cytokine profile of monocyte derived dendritic cells in leprosy: in vitro stimulation by sonicated Mycobacterium leprae induces decreased level of IL-12p70 in lepromatous leprosy

    Science.gov (United States)

    Braga, André Flores; Moretto, Daniela Ferraz; Gigliotti, Patrícia; Peruchi, Mariela; Vilani-Moreno, Fátima Regina; Campanelli, Ana Paula; Latini, Ana Carla Pereira; Iyer, Anand; Das, Pranab Kumar; de Souza, Vânia Nieto Brito

    2015-01-01

    Dendritic cells (DCs) play a pivotal role in the connection of innate and adaptive immunity of hosts to mycobacterial infection. Studies on the interaction of monocyte-derived DCs (MO-DCs) using Mycobacterium leprae in leprosy patients are rare. The present study demonstrated that the differentiation of MOs to DCs was similar in all forms of leprosy compared to normal healthy individuals. In vitro stimulation of immature MO-DCs with sonicated M. leprae induced variable degrees of DC maturation as determined by the increased expression of HLA-DR, CD40, CD80 and CD86, but not CD83, in all studied groups. The production of different cytokines by the MO-DCs appeared similar in all of the studied groups under similar conditions. However, the production of interleukin (IL)-12p70 by MO-DCs from lepromatous (LL) leprosy patients after in vitro stimulation with M. leprae was lower than tuberculoid leprosy patients and healthy individuals, even after CD40 ligation with CD40 ligand-transfected cells. The present cumulative findings suggest that the MO-DCs of LL patients are generally a weak producer of IL-12p70 despite the moderate activating properties ofM. leprae. These results may explain the poor M. leprae-specific cell-mediated immunity in the LL type of leprosy. PMID:26222022

  1. Comparison of monocyte-derived dendritic cells from colorectal cancer patients, non-small-cell-lung-cancer patients and healthy donors

    DEFF Research Database (Denmark)

    Kvistborg, P; Bechmann, C M; Pedersen, A W;

    2009-01-01

    7, and the optimal expression pattern is considered as CD14(low), CD1a, CD83(high), CD86(high), MHC class II(high) and CCR7(high). In accordance with data from other studies including other types of cancer patients, especially breast cancer patients, we found that the maturation status of the DC...... blood monocytes loaded with allogeneic tumor cell lysate rich in cancer/testis antigens. This vaccine has at present been tested for activity in three phase II clinical trials including two cohorts of patients with advanced colorectal cancer (CRC) and one cohort of patients with advanced non...... were analyzed by flow cytometry and the obtained data were used as a basis to set guideline values for our quality control of GMP produced DC vaccines. Each vaccine batch was analyzed for the expression of the surface maturation and differentiation molecules CD14, CD1a, CD83, CD86, MHC class II and CCR...

  2. Monocyte-derived dendritic cells from late gestation cows have an impaired ability to mature in response to E. coli stimulation in a receptor and cytokine-mediated fashion.

    Science.gov (United States)

    Pomeroy, Brianna; Sipka, Anja; Klaessig, Suzanne; Schukken, Ynte

    2015-09-15

    During late gestation the bovine immune system is less capable of eliciting inflammatory responses and eliminating invading pathogens. The maternal immune system is directed toward tolerance in order to prevent fetal rejection due to recognition of paternal antigens. In humans and mice, dendritic cell (DC) populations maintain a tolerogenic phenotype essential in the generation and preservation of maternal immune tolerance throughout pregnancy. However, the primary mechanisms which facilitate maternal immune tolerance involved in bovine gestation remain poorly understood. In order to determine if DC phenotype and function were regulated toward tolerance during bovine gestation, we compared in vitro generated monocyte-derived DC (mo-DC) from monocytes isolated from cows in late gestation (LG) to those from non-pregnant (NP) cows in their ability to mature following stimulation with UV irradiated Escherichia coli. Our results show mo-DC from LG cows have an impaired ability to mature in response to E. coli stimulation in a receptor and cytokine-mediated fashion in comparison to those from NP cows. Specifically, mo-DC from LG cows were unable to upregulate MHC II and maintained high expression of CD14, both indicative of an immature phenotype following E. coli-stimulation. Only mo-DC from LG showed significant increase in IL-10 production and had a significantly lower ratio of production of the Th1-polarizing cytokine IL-12 to regulatory cytokine IL-10 following E. coli stimulation compared to mo-DC from NP cows. Our findings demonstrate mo-DC from LG cows have a stifled capacity to develop a mature phenotype and drive pro-inflammatory Th1-type responses to E. coli stimulation. Results from this study provide insight into DC immune modulation in bovine pregnancy and elucidate host factors which may contribute to the heightened susceptibility to infection in late gestation. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Facilitated monocyte-macrophage uptake and tissue distribution of superparmagnetic iron-oxide nanoparticles.

    Directory of Open Access Journals (Sweden)

    Arnaud Beduneau

    Full Text Available BACKGROUND: We posit that the same mononuclear phagocytes (MP that serve as target cells and vehicles for a host of microbial infections can be used to improve diagnostics and drug delivery. We also theorize that physical and biological processes such as particle shape, size, coating and opsonization that affect MP clearance of debris and microbes can be harnessed to facilitate uptake of nanoparticles (NP and tissue delivery. METHODS: Monocytes and monocyte-derived macrophages (MDM were used as vehicles of superparamagnetic iron oxide (SPIO NP and immunoglobulin (IgG or albumin coated SPIO for studies of uptake and distribution. IgG coated SPIO was synthesized by covalent linkage and uptake into monocytes and MDM investigated related to size, time, temperature, concentration, and coatings. SPIO and IgG SPIO were infused intravenously into naïve mice. T(2 measures using magnetic resonance imaging (MRI were used to monitor tissue distribution in animals. RESULTS: Oxidation of dextran on the SPIO surface generated reactive aldehyde groups and permitted covalent linkage to amino groups of murine and human IgG and F(ab'(2 fragments and for Alexa Fluor(R 488 hydroxylamine to form a Schiff base. This labile intermediate was immediately reduced with sodium cyanoborohydride in order to stabilize the NP conjugate. Optical density measurements of the oxidized IgG, F(ab'(2, and/or Alexa Fluor(R 488 SPIO demonstrated approximately 50% coupling yield. IgG-SPIO was found stable at 4 degrees C for a period of 1 month during which size and polydispersity index varied little from 175 nm and 200 nm, respectively. In vitro, NP accumulated readily within monocyte and MDM cytoplasm after IgG-SPIO exposure; whereas, the uptake of native SPIO in monocytes and MDM was 10-fold less. No changes in cell viability were noted for the SPIO-containing monocytes and MDM. Cell morphology was not changed as observed by transmission electron microscopy. Compared to unconjugated

  4. Monocyte-derived dendritic cells enhance cell proliferation and porcine circovirus type 2 replication in concanavalin A-stimulated swine peripheral blood lymphocytes in vitro.

    Science.gov (United States)

    Lin, Chun-Ming; Jeng, Chian-Ren; Hsiao, Shih-Hsuan; Lee, Yao; Tsai, Yi-Chieh; Chia, Mi-Yuan; Pang, Victor Fei

    2012-01-15

    Dendritic cells (DCs) are professional antigen presenting cells cooperating with other immune cells for the activation of innate and adaptive immune responses. The objective of the present study was to investigate the replication activity of porcine circovirus type 2 (PCV2) in DCs and/or lymphocytes during their cross talk and its possible mechanism. Two models were set, herein. Swine blood monocyte (Mo)-derived DCs (MoDCs) or peripheral blood lymphocytes (PBLs) were inoculated with PCV2 prior to their co-cultivation. Bacterial lipopolysaccharide (LPS) and concanavalin A (Con A) were used to stimulate MoDCs and PBLs, respectively. During 6 days of cultivation, a high PCV2 antigen-containing rate without detectable intranuclear signals and a slight but significant increase in the copy number of PCV2 genome were detected in PCV2-inoculated MoDCs. The presence of LPS alone or PCV2-free PBLs, however, had no effect on the location of PCV2 antigens or copy number of PCV2 genome in PCV2-inoculated MoDCs. On the contrary, active PCV2 replication occurred in Con A-stimulated PCV2-inoculated PBLs. When compared with blood Mos, MoDCs induced significantly higher cell proliferation and intensified PCV2 replication in Con A-stimulated PCV2-inoculated PBLs, for which direct contact between MoDCs and lymphocytes was required. Among the cytokines secreted by Con A-activated PBLs, interleukin (IL)-2, but not IL-4 or interferon-γ, could induce cell proliferation and PCV2 replication in PCV2-inoculated PBLs. The findings suggest that although MoDCs support only limited PCV2 replication in themselves, their accessory cell function is required for cell proliferation and PCV2 replication in PCV2-infected lymphocytes.

  5. Chemical and physical effects on the adhesion, maturation, and survival of monocytes, macrophages, and foreign body giant cells

    Science.gov (United States)

    Collier, Terry Odell, III

    Injury caused by biomedical device implantation initiates inflammatory and wound healing responses. Cells migrate to the site of injury to degrade bacteria and toxins, create new vasculature, and form new and repair injured tissue. Blood-proteins rapidly adsorb onto the implanted material surface and express adhesive ligands which mediate cell adhesion on the material surface. Monocyte-derived macrophages and multi-nucleated foreign body giant cells adhere to the surface and degrade the surface of the material. Due to the role of macrophage and foreign body giant cell on material biocompatibility and biostability, the effects of surface chemistry, surface topography and specific proteins on the maturation and survival of monocytes, macrophages and foreign body giant cells has been investigated. Novel molecularly designed materials were used to elucidate the dynamic interactions which occur between inflammatory cells, proteins and surfaces. The effect of protein and protein adhesion was investigated using adhesive protein depleted serum conditions on RGD-modified and silane modified surfaces. The effects of surface chemistry were investigated using temperature responsive surfaces of poly (N-isopropylacrylamide) and micropatterned surfaces of N-(2 aminoethyl)-3-aminopropyltrimethoxysilane regions on an interpenetrating polymer network of polyacrylamide and poly(ethylene glycol). The physical effects were investigated using polyimide scaffold materials and polyurethane materials with surface modifying end groups. The depletion of immunoglobulin G caused decreased levels of macrophage adhesion, foreign body giant cell formation and increased levels of apoptosis. The temporal nature of macrophage adhesion was observed with changing effectiveness of adherent cell detachment with time, which correlated to increased expression of beta1 integrin receptors on detached macrophages with time. The limited ability of the micropatterned surface, polyimide scaffold and surface

  6. Plasma lipoproteins and monocyte-macrophages in a peroxisome-deficient system: study of a patient with infantile refsum disease.

    Science.gov (United States)

    Mandel, H; Berant, M; Meiron, D; Aizin, A; Oiknine, J; Brook, J G; Aviram, M

    1992-01-01

    Hypocholesterolaemia in infantile Refsum disease (IRD) may link peroxisomes and lipoprotein metabolism. In our patient, plasma cholesterol levels were reduced to 26% and 29% of control in LDL and HDL fractions, respectively. Plasma apolipoproteins B-100 and A-I levels were 52% and 66% of controls, respectively. In the kindred, plasma cholesterol concentration was 61-73% of controls. The HDL-cholesterol/apo A-I ratios were: patient 0.12; kindred 0.17; controls 0.28. Analysis of the IRD patient's lipoprotein revealed compositional abnormalities in all fractions. The patient's LDL demonstrated a substantial reduction in its lipid-to-protein ratio. Alterations in plasma lipoproteins affect their interaction with macrophages. Upon incubation of the patient's LDL with J-774 macrophages, its cellular uptake, measured as cholesterol esterification rate, was only 66% of a control rate. The abnormal LDL of the IRD patient showed also only 25% of control susceptibility to in vitro oxidation. Studies of cellular cholesterol metabolism in the patient's monocyte-derived macrophages (MDM) showed 57% increased cholesterol esterification rate in comparison to normal MDM. The possible link between lipoprotein abnormalities and monocyte-macrophage cholesterol metabolism is discussed.

  7. Monocyte chemiluminescence and macrophage precursors in the aged.

    Directory of Open Access Journals (Sweden)

    Takahashi,Isao

    1985-12-01

    Full Text Available Age-related alterations in the host defense system have been vigorously investigated because of increased susceptibility to infection and neoplasms in the aged. Although monocyte-macrophages form a major part of the cellular defense against microorganisms, the majority of investigations has been limited to neutrophils and lymphocytes. The present study, designed to determine the influence of age on mononuclear phagocytes, revealed no significant decrease in the absolute number of blood monocytes, but did reveal a tendency for the chemiluminescence of blood monocytes to decrease (p less than 0.10 and a significant decrease in the numbers of macrophage precursors (p less than 0.05 in the aged (over 70 year old, in comparison with controls (under 40 years old. On the basis of these findings, functional alterations of monocyte-macrophages seem to participate in the increased susceptibility to infection in the aged.

  8. Psychedelic N,N-dimethyltryptamine and 5-methoxy-N,N-dimethyltryptamine modulate innate and adaptive inflammatory responses through the sigma-1 receptor of human monocyte-derived dendritic cells.

    Science.gov (United States)

    Szabo, Attila; Kovacs, Attila; Frecska, Ede; Rajnavolgyi, Eva

    2014-01-01

    The orphan receptor sigma-1 (sigmar-1) is a transmembrane chaperone protein expressed in both the central nervous system and in immune cells. It has been shown to regulate neuronal differentiation and cell survival, and mediates anti-inflammatory responses and immunosuppression in murine in vivo models. Since the details of these findings have not been elucidated so far, we studied the effects of the endogenous sigmar-1 ligands N,N-dimethyltryptamine (NN-DMT), its derivative 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) and the synthetic high affinity sigmar-1 agonist PRE-084 hydrochloride on human primary monocyte-derived dendritic cell (moDCs) activation provoked by LPS, polyI:C or pathogen-derived stimuli to induce inflammatory responses. Co-treatment of moDC with these activators and sigma-1 receptor ligands inhibited the production of pro-inflammatory cytokines IL-1β, IL-6, TNFα and the chemokine IL-8, while increased the secretion of the anti-inflammatory cytokine IL-10. The T-cell activating capacity of moDCs was also inhibited, and dimethyltryptamines used in combination with E. coli or influenza virus as stimulators decreased the differentiation of moDC-induced Th1 and Th17 inflammatory effector T-cells in a sigmar-1 specific manner as confirmed by gene silencing. Here we demonstrate for the first time the immunomodulatory potential of NN-DMT and 5-MeO-DMT on human moDC functions via sigmar-1 that could be harnessed for the pharmacological treatment of autoimmune diseases and chronic inflammatory conditions of the CNS or peripheral tissues. Our findings also point out a new biological role for dimethyltryptamines, which may act as systemic endogenous regulators of inflammation and immune homeostasis through the sigma-1 receptor.

  9. PRRSV-infected monocyte-derived dendritic cells express high levels of SLA-DR and CD80/86 but do not stimulate PRRSV-naïve regulatory T cells to proliferate.

    Science.gov (United States)

    Rodríguez-Gómez, Irene M; Käser, Tobias; Gómez-Laguna, Jaime; Lamp, Benjamin; Sinn, Leonie; Rümenapf, Till; Carrasco, Librado; Saalmüller, Armin; Gerner, Wilhelm

    2015-05-20

    In vitro generated monocyte-derived dendritic cells (moDCs) have frequently been used to study the influence of porcine reproductive and respiratory syndrome virus (PRRSV) infection on antigen presenting cells. However, obtained results have often been conflicting in regard to expression of co-stimulatory molecules and interaction with T cells. In this study we performed a detailed phenotypic characterisation of PRRSV-infected moDCs and non-infected moDCs. For CD163 and CD169, which are involved in PRRSV-entry into host cells, our results show that prior to infection porcine moDCs express high levels of CD163 but only very low levels for CD169. Following infection with either PRRSV-1 or PRRSV-2 strains after 24 h, PRRSV-nucleoprotein (N-protein)(+) and N-protein(-) moDCs derived from the same microculture were analyzed for expression of swine leukocyte antigen-DR (SLA-DR) and CD80/86. N-protein(+) moDCs consistently expressed higher levels of SLA-DR and CD80/86 compared to N-protein(-) moDCs. We also investigated the influence of PRRSV-infected moDCs on proliferation and frequency of Foxp3(+) regulatory T cells present within CD4(+) T cells in in vitro co-cultures. Neither CD3-stimulated nor unstimulated CD4(+) T cells showed differences in regard to proliferation and frequency of Foxp3(+) T cells following co-cultivation with either PRRSV-1 or PRRSV-2 infected moDCs. Our results suggest that a more detailed characterisation of PRRSV-infected moDCs will lead to more consistent results across different laboratories and PRRSV strains as indicated by the major differences in SLA-DR and CD80/86 expression between PRRSV-infected and non-infected moDCs present in the same microculture.

  10. Psychedelic N,N-dimethyltryptamine and 5-methoxy-N,N-dimethyltryptamine modulate innate and adaptive inflammatory responses through the sigma-1 receptor of human monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Attila Szabo

    Full Text Available The orphan receptor sigma-1 (sigmar-1 is a transmembrane chaperone protein expressed in both the central nervous system and in immune cells. It has been shown to regulate neuronal differentiation and cell survival, and mediates anti-inflammatory responses and immunosuppression in murine in vivo models. Since the details of these findings have not been elucidated so far, we studied the effects of the endogenous sigmar-1 ligands N,N-dimethyltryptamine (NN-DMT, its derivative 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT and the synthetic high affinity sigmar-1 agonist PRE-084 hydrochloride on human primary monocyte-derived dendritic cell (moDCs activation provoked by LPS, polyI:C or pathogen-derived stimuli to induce inflammatory responses. Co-treatment of moDC with these activators and sigma-1 receptor ligands inhibited the production of pro-inflammatory cytokines IL-1β, IL-6, TNFα and the chemokine IL-8, while increased the secretion of the anti-inflammatory cytokine IL-10. The T-cell activating capacity of moDCs was also inhibited, and dimethyltryptamines used in combination with E. coli or influenza virus as stimulators decreased the differentiation of moDC-induced Th1 and Th17 inflammatory effector T-cells in a sigmar-1 specific manner as confirmed by gene silencing. Here we demonstrate for the first time the immunomodulatory potential of NN-DMT and 5-MeO-DMT on human moDC functions via sigmar-1 that could be harnessed for the pharmacological treatment of autoimmune diseases and chronic inflammatory conditions of the CNS or peripheral tissues. Our findings also point out a new biological role for dimethyltryptamines, which may act as systemic endogenous regulators of inflammation and immune homeostasis through the sigma-1 receptor.

  11. Monocyte-derived dendritic cells exposed to Der p 1 allergen enhance the recruitment of Th2 cells: major involvement of the chemokines TARC/CCL17 and MDC/CCL22.

    Science.gov (United States)

    Hammad, Hamida; Smits, Hermelijn H; Ratajczak, Céline; Nithiananthan, Asokananthan; Wierenga, Eddy A; Stewart, Geoffrey A; Jacquet, Alain; Tonnel, Andre-Bernard; Pestel, Joël

    2003-01-01

    Dendritic cells (DC) are potent antigen - presenting cells that can orientate the immune response towards a Th1 or a Th2 type. DC produce chemokines that are involved in the recruitment of either Th1 cells, such as IP10 (CXCL10), Th2 cells such as TARC (CCL17) and MDC (CCL22), or non-polarized T cells such as RANTES (CCL5) and MIP-lalpha (CCL3). We investigated whether monocyte-derived DC (MD-DC) generated from healthy donors or from patients sensitive to Dermatophagoides pteronyssinus (Dpt) and exposed to the cysteine-protease Der p 1(allergen of Dpt), could upregulate the expression of chemokines involved in type 1 or type 2 T cell recruitment. MD-DC were pulsed with either Der p 1 or with LPS as the control and the chemokines produced were evaluated using ELISA and chemotaxis assays. Der p 1-pulsed DC from allergic patients showed increased TARC (CCL17) and MDC (CCL22) production without modifying IP-10 (CXCL10) release. Der p 1-pulsed DC from healthy donors showed only increased IP-10 (CXCL10) secretion. RANTES (CCL5) and MIP-lalpha (CCL3) production were similarly increased when DC were from healthy or allergic donors. The selective Th2 clone recruitment activity of supernatants from Der p 1-pulsed DC of allergic patients was inhibited by anti-TARC (CCL17) and anti-MDC (CCL22) neutralizing Abs. By using anti-IP10 (CXCL10) blocking Abs, supernatants of Der p 1-pulsed DC from healthy donors were shown to be involved in the recruitment of Th1 cells. These results suggest that in allergic patients exposed to house dust mites, DC may favour the exacerbation of the Th2 response via the increase in type 2 chemokine production. Copyright John Libbey Eurotext 2003.

  12. Single point mutations in the helicase domain of the NS3 protein enhance dengue virus replicative capacity in human monocyte-derived dendritic cells and circumvent the type I interferon response.

    Science.gov (United States)

    Silveira, G F; Strottmann, D M; de Borba, L; Mansur, D S; Zanchin, N I T; Bordignon, J; dos Santos, C N Duarte

    2016-01-01

    Dengue is the most prevalent arboviral disease worldwide. The outcome of the infection is determined by the interplay of viral and host factors. In the present study, we evaluated the cellular response of human monocyte-derived DCs (mdDCs) infected with recombinant dengue virus type 1 (DV1) strains carrying a single point mutation in the NS3hel protein (L435S or L480S). Both mutated viruses infect and replicate more efficiently and produce more viral progeny in infected mdDCs compared with the parental, non-mutated virus (vBACDV1). Additionally, global gene expression analysis using cDNA microarrays revealed that the mutated DVs induce the up-regulation of the interferon (IFN) signalling and pattern recognition receptor (PRR) canonical pathways in mdDCs. Pronounced production of type I IFN were detected specifically in mdDCs infected with DV1-NS3hel-mutated virus compared with mdDCs infected with the parental virus. In addition, we showed that the type I IFN produced by mdDCs is able to reduce DV1 infection rates, suggesting that cytokine function is effective but not sufficient to mediate viral clearance of DV1-NS3hel-mutated strains. Our results demonstrate that single point mutations in subdomain 2 have important implications for adenosine triphosphatase (ATPase) activity of DV1-NS3hel. Although a direct functional connection between the increased ATPase activity and viral replication still requires further studies, these mutations speed up viral RNA replication and are sufficient to enhance viral replicative capacity in human primary cell infection and circumvent type I IFN activity. This information may have particular relevance for attenuated vaccine protocols designed for DV.

  13. Macrophage diversity in renal injury and repair

    NARCIS (Netherlands)

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

    2008-01-01

    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 re

  14. 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

  15. Cholesterol enrichment of human monocyte/macrophages induces surface exposure of phosphatidylserine and the release of biologically-active tissue factor-positive microvesicles.

    Science.gov (United States)

    Liu, Ming-Lin; Reilly, Michael P; Casasanto, Peter; McKenzie, Steven E; Williams, Kevin Jon

    2007-02-01

    Biologically significant amounts of two procoagulant molecules, phosphatidylserine (PS) and tissue factor (TF), are transported by monocyte/macrophage-derived microvesicles (MVs). Because cellular cholesterol accumulation is an important feature of atherosclerotic vascular disease, we now examined effects of cholesterol enrichment on MV release from human monocytes and macrophages. Cholesterol enrichment of human THP-1 monocytes, alone or in combination with lipopolysaccharide (LPS), tripled their total MV generation, as quantified by flow cytometry based on particle size and PS exposure. The subset of these MVs that were also TF-positive was likewise increased by cellular cholesterol enrichment, and these TF-positive MVs exhibited a striking 10-fold increase in procoagulant activity. Moreover, cholesterol enrichment of primary human monocyte-derived macrophages also increased their total as well as TF-positive MV release, and these TF-positive MVs exhibited a similar 10-fold increase in procoagulant activity. To explore the mechanisms of enhanced MV release, we found that cholesterol enrichment of monocytes caused PS exposure on the cell surface by as early as 2 hours and genomic DNA fragmentation in a minority of cells by 20 hours. Addition of a caspase inhibitor at the beginning of these incubations blunted both cholesterol-induced apoptosis and MV release. Cholesterol enrichment of human monocyte/macrophages induces the generation of highly biologically active, PS-positive MVs, at least in part through induction of apoptosis. Cholesterol-induced monocyte/macrophage MVs, both TF-positive and TF-negative, may be novel contributors to atherothrombosis.

  16. Influence of phthalates on cytokine production in monocytes and macrophages

    DEFF Research Database (Denmark)

    Frohnert, Juliana; Bendtzen, Klaus; Boas, Malene;

    2015-01-01

    BACKGROUND: Phthalates are a group of endocrine disrupting chemicals suspected to influence the immune system. The aim of this systematic review is to summarise the present knowledge on the influence of phthalates on monocyte and macrophage production and secretion of cytokines, an influence which......-α secretion/production from monocytes or macrophages. A summary of cytokine measurements was not possible since few studies were comparable in study design and due to insufficient reporting of raw data for most of the included studies. CONCLUSION: Results from this review have suggested that at least one...

  17. 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

  18. Purification of Human Monocytes and Lymphocyte Populations by Counter Current Elutriation– A Short Protocol

    OpenAIRE

    Clarke, Elizabeth V.; Benoit, Marie E.; Tenner, Andrea J.

    2013-01-01

    Investigations of the activation processes involved in human monocytes and monocyte-derived macrophages and dendritic cells often required large numbers of cells that have not been possibly altered or activated by adherence to surfaces, by binding of antibodies to surface antigens during positive selection, or by release of activators by platelets or other non myeloid cells during isolation or co-culture. Human peripheral blood monocytes as well as lymphocytes from the same blood donor can be...

  19. Ameloginins promote an alternatively activated macrophage phenotype in vitro

    DEFF Research Database (Denmark)

    Almqvist, S; Werthen, M; Lyngstadas, SP

    2011-01-01

    Amelogenins are extracellular matrix proteins used for the topical treatment of chronically inflamed tissues. The influence of amelogenins on human monocyte-derived macrophages was studied by measuring the concentrations of cytokines in culture supernatants. The interactions of cells and protein...

  20. Effects of Two Fullerene Derivatives on Monocytes and Macrophages

    Directory of Open Access Journals (Sweden)

    Sabrina Pacor

    2015-01-01

    Full Text Available Two fullerene derivatives (fullerenes 1 and 2, bearing a hydrophilic chain on the pyrrolidinic nitrogen, were developed with the aim to deliver anticancer agents to solid tumors. These two compounds showed a significantly different behaviour on human neoplastic cell lines in vitro in respect to healthy leukocytes. In particular, the pyrrolidinium ring on the fullerene carbon cage brings to a more active compound. In the present work, we describe the effects of these fullerenes on primary cultures of human monocytes and macrophages, two kinds of immune cells representing the first line of defence in the immune response to foreign materials. These compounds are not recognized by circulating monocytes while they get into macrophages. The evaluation of the pronecrotic or proapoptotic effects, analysed by means of analysis of the purinergic receptor P2X7 activation and of ROS scavenging activity, has allowed us to show that fullerene 2, but not its analogue fullerene 1, displays toxicity, even though at concentrations higher than those shown to be active on neoplastic cells.

  1. Effects of Two Fullerene Derivatives on Monocytes and Macrophages.

    Science.gov (United States)

    Pacor, Sabrina; Grillo, Alberto; Đorđević, Luka; Zorzet, Sonia; Lucafò, Marianna; Da Ros, Tatiana; Prato, Maurizio; Sava, Gianni

    2015-01-01

    Two fullerene derivatives (fullerenes 1 and 2), bearing a hydrophilic chain on the pyrrolidinic nitrogen, were developed with the aim to deliver anticancer agents to solid tumors. These two compounds showed a significantly different behaviour on human neoplastic cell lines in vitro in respect to healthy leukocytes. In particular, the pyrrolidinium ring on the fullerene carbon cage brings to a more active compound. In the present work, we describe the effects of these fullerenes on primary cultures of human monocytes and macrophages, two kinds of immune cells representing the first line of defence in the immune response to foreign materials. These compounds are not recognized by circulating monocytes while they get into macrophages. The evaluation of the pronecrotic or proapoptotic effects, analysed by means of analysis of the purinergic receptor P2X7 activation and of ROS scavenging activity, has allowed us to show that fullerene 2, but not its analogue fullerene 1, displays toxicity, even though at concentrations higher than those shown to be active on neoplastic cells.

  2. Macrophage subsets and microglia in multiple sclerosis

    OpenAIRE

    2014-01-01

    Along with microglia and monocyte-derived macrophages, macrophages in the perivascular space, choroid plexus, and meninges are the principal effector cells in neuroinflammatory and neurodegenerative disorders. These phagocytes are highly heterogeneous cells displaying spatial- and temporal-dependent identities in the healthy, injured, and inflamed CNS. In the last decade, researchers have debated on whether phagocytes subtypes and phenotypes are pathogenic or protective in CNS pathologies. In...

  3. Ontogeny and Polarization of Macrophages in Inflammation: Blood monocytes versus tissue macrophages.

    Directory of Open Access Journals (Sweden)

    Adwitia eDey

    2015-01-01

    Full Text Available The explosion of new information in recent years on the origin of macrophages in the steady-state and in the context of inflammation has opened up numerous new avenues of investigation and possibilities for therapeutic intervention. In contrast to the classical model of macrophage development, it is clear that tissue-resident macrophages can develop from yolk sac-derived erythromyeloid progenitors, fetal liver progenitors and bone marrow-derived monocytes. Under both homeostatic conditions and in response to pathophysiological insult, the contribution of these distinct sources of macrophages varies significantly between tissues. Furthermore, while all of these populations of macrophages appear to be capable of adopting the polarized M1/M2 phenotypes, their respective contribution to inflammation, resolution of inflammation and tissue repair remains poorly understood and is likely to be tissue- and disease-dependent. A better understanding of the ontology and polarization capacity of macrophages in homeostasis and disease will be essential for the development of novel therapies that target the inherent plasticity of macrophages in the treatment of acute and chronic inflammatory disease.

  4. Aortic endothelial cells regulate proliferation of human monocytes in vitro via a mechanism synergistic with macrophage colony-stimulating factor. Convergence at the cyclin E/p27(Kip1) regulatory checkpoint.

    Science.gov (United States)

    Antonov, A S; Munn, D H; Kolodgie, F D; Virmani, R; Gerrity, R G

    1997-06-15

    Monocyte-derived macrophages (Mphis) are pivotal participants in the pathogenesis of atherosclerosis. Evidence from both animal and human plaques indicates that local proliferation may contribute to accumulation of lesion Mphis, and the major Mphi growth factor, macrophage colony stimulating factor (MCSF), is present in atherosclerotic plaques. However, most in vitro studies have failed to demonstrate that human monocytes/Mphis possess significant proliferative capacity. We now report that, although human monocytes cultured in isolation showed only limited MCSF-induced proliferation, monocytes cocultured with aortic endothelial cells at identical MCSF concentrations underwent enhanced (up to 40-fold) and prolonged (21 d) proliferation. In contrast with monocytes in isolation, this was optimal at low seeding densities, required endothelial cell contact, and could not be reproduced by coculture with smooth muscle cells. Intimal Mphi isolated from human aortas likewise showed endothelial cell contact-dependent, MCSF-induced proliferation. Consistent with a two-signal mechanism governing Mphi proliferation, the cell cycle regulatory protein, cyclin E, was rapidly upregulated by endothelial cell contact in an MCSFindependent fashion, but MCSF was required for successful downregulation of the cell cycle inhibitory protein p27(Kip1) before cell cycling. Thus endothelial cells and MCSF differentially and synergistically regulate two Mphi genes critical for progression through the cell cycle.

  5. Transduced monocyte/macrophages targeted to murine skin by UV light.

    Science.gov (United States)

    Zhang, Alexandra Y; Wu, Caiyun; Zhou, Lixin; Ismail, Sahar A; Tao, Jianming; McCormick, Laura L; Cooper, Kevin D; Gilliam, Anita C

    2006-01-01

    We have selectively targeted monocyte/macrophages overexpressing an immunomodulatory molecule, latency-associated peptide (LAP), a naturally occurring antagonist for transforming growth factor-beta1, to murine skin utilizing UV light to produce a cutaneous influx of transduced monocyte/macrophages. Bone marrow (BM) cells from BALB/c mice were transduced in vitro with a retroviral construct containing green fluorescent protein (GFP) for detection and human LAP (hLAP) as a test molecule. The transduced BM cells were then cultured in vitro with granulocyte-macrophage colony-stimulating factor (GM-CSF) to produce differentiation to monocyte/macrophages. More than 80% of transduced BM cells were CD11b-positive and MOMA-positive by fluorescence-activated cell-sorter analysis and secreted LAP by ELISA after 10 days of culture in granulocyte-macrophage colony-stimulating factor (GM-CSF). Transduced monocyte/macrophages containing either GFP or hLAP-GFP were then injected intravenously into BALB/c mice. One-half of recipients in each group were exposed to UVB (72 mJ) to induce monocyte/macrophage infiltration into skin. Recipients were sacrificed 60 h after UV irradiation. We found transduced cutaneous macrophages expressing GFP by examining with fluorescence microscopy frozen skin sections of recipient mice immunostained with antibodies to GFP and to macrophage marker F4/80. We identified hLAP sequences by polymerase chain reaction (PCR) of total DNA in recipient blood and UV-irradiated skin but not in unirradiated skin. LAP sequences were also detected at much lower levels in other organs (lung, spleen, and liver) by PCR. Therefore, we have shown that genetically altered monocytic cells can be injected intravenously and targeted to mouse skin by UV exposure. This macrophage-based gene-transfer method may be a potentially useful immunotherapeutic approach for delivering monocyte/macrophage-derived products to skin.

  6. A novel hybrid aspirin-NO-releasing compound inhibits TNFalpha release from LPS-activated human monocytes and macrophages

    Directory of Open Access Journals (Sweden)

    Fox Sarah

    2008-07-01

    Full Text Available Abstract Background The cytoprotective nature of nitric oxide (NO led to development of NO-aspirins in the hope of overcoming the gastric side-effects of aspirin. However, the NO moiety gives these hybrids potential for actions further to their aspirin-mediated anti-platelet and anti-inflammatory effects. Having previously shown that novel NO-aspirin hybrids containing a furoxan NO-releasing group have potent anti-platelet effects, here we investigate their anti-inflammatory properties. Here we examine their effects upon TNFα release from lipopolysaccharide (LPS-stimulated human monocytes and monocyte-derived macrophages and investigate a potential mechanism of action through effects on LPS-stimulated nuclear factor-kappa B (NF-κB activation. Methods Peripheral venous blood was drawn from the antecubital fossa of human volunteers. Mononuclear cells were isolated and cultured. The resultant differentiated macrophages were treated with pharmacologically relevant concentrations of either a furoxan-aspirin (B8, B7; 10 μM, their respective furazan NO-free counterparts (B16, B15; 10 μM, aspirin (10 μM, existing nitroaspirin (NCX4016; 10 μM, an NO donor (DEA/NO; 10 μM or dexamethasone (1 μM, in the presence and absence of LPS (10 ng/ml; 4 h. Parallel experiments were conducted on undifferentiated fresh monocytes. Supernatants were assessed by specific ELISA for TNFα release and by lactate dehydrogenase (LDH assay for cell necrosis. To assess NF-κB activation, the effects of the compounds on the loss of cytoplasmic inhibitor of NF-κB, IκBα (assessed by western blotting and nuclear localisation (assessed by immunofluorescence of the p65 subunit of NF-κB were determined. Results B8 significantly reduced TNFα release from LPS-treated macrophages to 36 ± 10% of the LPS control. B8 and B16 significantly inhibited monocyte TNFα release to 28 ± 5, and 49 ± 9% of control, respectively. The B8 effect was equivalent in magnitude to that of

  7. Immature monocytes recruited to the ischemic mouse brain differentiate into macrophages with features of alternative activation.

    Science.gov (United States)

    Miró-Mur, Francesc; Pérez-de-Puig, Isabel; Ferrer-Ferrer, Maura; Urra, Xabier; Justicia, Carles; Chamorro, Angel; Planas, Anna M

    2016-03-01

    Acute stroke induces a local inflammatory reaction causing leukocyte infiltration. Circulating monocytes are recruited to the ischemic brain and become tissue macrophages morphologically indistinguishable from reactive microglia. However, monocytes are a heterogeneous population of cells with different functions. Herein, we investigated the infiltration and fate of the monocyte subsets in a mouse model of focal brain ischemia by permanent occlusion of the distal portion of the middle cerebral artery. We separated two main subtypes of CD11b(hi) monocytes according to their expression of the surface markers Ly6C and CD43. Using adoptive transfer of reporter monocytes and monocyte depletion, we identified the pro-inflammatory Ly6C(hi)CD43(lo)CCR2(+) subset as the predominant monocytes recruited to the ischemic tissue. Monocytes were seen in the leptomeninges from where they entered the cortex along the penetrating arterioles. Four days post-ischemia, they had invaded the infarcted core, where they were often located adjacent to blood vessels. At this time, Iba-1(-) and Iba-1(+) cells in the ischemic tissue incorporated BrdU, but BrdU incorporation was rare in the reporter monocytes. The monocyte phenotype progressively changed by down-regulating Ly6C, up-regulating F4/80, expressing low or intermediate levels of Iba-1, and developing macrophage morphology. Moreover, monocytes progressively acquired the expression of typical markers of alternatively activated macrophages, like arginase-1 and YM-1. Collectively, the results show that stroke mobilized immature pro-inflammatory Ly6C(hi)CD43(lo) monocytes that acutely infiltrated the ischemic tissue reaching the core of the lesion. Monocytes differentiated to macrophages with features of alternative activation suggesting possible roles in tissue repair during the sub-acute phase of stroke.

  8. Release of salusin-beta from human monocytes/macrophages.

    Science.gov (United States)

    Sato, Kengo; Fujimoto, Kazumi; Koyama, Takatoshi; Shichiri, Masayoshi

    2010-06-01

    the release of salusin-beta. These data demonstrate that salusin-beta, which induces macrophage foam cell formation, is secreted in its authentic form from human monocytes/macrophages.

  9. Macrophage Responses to Epithelial Dysfunction Promote Lung Fibrosis in Aging

    Science.gov (United States)

    2016-10-01

    niche by monocyte- derived alveolar macrophages. Thus, we were able to overcome the technical issues and use regular shielded bone marrow chimeras for... protocols established and optimized. Major Task 3: Can the adoptive transfer of tissue-resident alveolar macrophages improve chronic stress in the lung...subtask. Since approval of the protocol by IRB and HRPO a total of 45 lung samples were processed , including donor lungs and lungs from patients with

  10. Macrophages, Dendritic Cells, and Regression of Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Jonathan E. Feig

    2012-07-01

    Full Text Available Atherosclerosis is the number one cause of death in the Western world. It results from the interaction between modified lipoproteins and monocyte-derived cells such as macrophages, dendritic cells, T cells, and other cellular elements of the arterial wall. This inflammatory process can ultimately lead to the development of complex lesions, or plaques, that protrude into the arterial lumen. Ultimately, plaque rupture and thrombosis can occur leading to the clinical complications of myocardial infarction or stroke. Although each of the cell types plays roles in the pathogenesis of atherosclerosis, in this review, the focus will be primarily on the monocyte derived cells- macrophages and dendritic cells. The roles of these cell types in atherogenesis will be highlighted. Finally, the mechanisms of atherosclerosis regression as it relates to these cells will be discussed.

  11. Infiltrating blood-derived macrophages are vital cells playing an anti-inflammatory role in recovery from spinal cord injury in mice.

    Directory of Open Access Journals (Sweden)

    Ravid Shechter

    2009-07-01

    Full Text Available BACKGROUND: Although macrophages (MPhi are known as essential players in wound healing, their contribution to recovery from spinal cord injury (SCI is a subject of debate. The difficulties in distinguishing between different MPhi subpopulations at the lesion site have further contributed to the controversy and led to the common view of MPhi as functionally homogenous. Given the massive accumulation in the injured spinal cord of activated resident microglia, which are the native immune occupants of the central nervous system (CNS, the recruitment of additional infiltrating monocytes from the peripheral blood seems puzzling. A key question that remains is whether the infiltrating monocyte-derived MPhi contribute to repair, or represent an unavoidable detrimental response. The hypothesis of the current study is that a specific population of infiltrating monocyte-derived MPhi is functionally distinct from the inflammatory resident microglia and is essential for recovery from SCI. METHODS AND FINDINGS: We inflicted SCI in adult mice, and tested the effect of infiltrating monocyte-derived MPhi on the recovery process. Adoptive transfer experiments and bone marrow chimeras were used to functionally distinguish between the resident microglia and the infiltrating monocyte-derived MPhi. We followed the infiltration of the monocyte-derived MPhi to the injured site and characterized their spatial distribution and phenotype. Increasing the naïve monocyte pool by either adoptive transfer or CNS-specific vaccination resulted in a higher number of spontaneously recruited cells and improved recovery. Selective ablation of infiltrating monocyte-derived MPhi following SCI while sparing the resident microglia, using either antibody-mediated depletion or conditional ablation by diphtheria toxin, impaired recovery. Reconstitution of the peripheral blood with monocytes resistant to ablation restored the lost motor functions. Importantly, the infiltrating monocyte-derived

  12. Different Regulation of Interleukin-1 Production and Activity in Monocytes and Macrophages: Innate Memory as an Endogenous Mechanism of IL-1 Inhibition

    Directory of Open Access Journals (Sweden)

    Mariusz P. Madej

    2017-06-01

    Full Text Available Production and activity of interleukin (IL-1β are kept under strict control in our body, because of its powerful inflammation-promoting capacity. Control of IL-1β production and activity allows IL-1 to exert its defensive activities without causing extensive tissue damage. Monocytes are the major producers of IL-1β during inflammation, but they are also able to produce significant amounts of IL-1 inhibitors such as IL-1Ra and the soluble form of the decoy receptor IL-1R2, in an auto-regulatory feedback loop. Here, we investigated how innate immune memory could modulate production and activity of IL-1β by human primary monocytes and monocyte-derived tissue-like/deactivated macrophages in vitro. Cells were exposed to Gram-negative (Escherichia coli and Gram-positive (Lactobacillus acidophilus bacteria for 24 h, then allowed to rest, and then re-challenged with the same stimuli. The presence of biologically active IL-1β in cell supernatants was calculated as the ratio between free IL-1β (i.e., the cytokine that is not bound/inhibited by sIL-1R2 and its receptor antagonist IL-1Ra. As expected, we observed that the responsiveness of tissue-like/deactivated macrophages to bacterial stimuli was lower than that of monocytes. After resting and re-stimulation, a memory effect was evident for the production of inflammatory cytokines, whereas production of alarm signals (chemokines was minimally affected. We observed a high variability in the innate memory response among individual donors. This is expected since innate memory largely depends on the previous history of exposure or infections, which is different in different subjects. Overall, innate memory appeared to limit the amount of active IL-1β produced by macrophages in response to a bacterial challenge, while enhancing the responsiveness of monocytes. The functional re-programming of mononuclear phagocytes through modulation of innate memory may provide innovative approaches in the management

  13. Canine Distemper Virus Infection Leads to an Inhibitory Phenotype of Monocyte-Derived Dendritic Cells In Vitro with Reduced Expression of Co-Stimulatory Molecules and Increased Interleukin-10 Transcription

    Science.gov (United States)

    Herder, Vanessa; Stein, Veronika M.; Tipold, Andrea; Urhausen, Carola; Günzel-Apel, Anne-Rose; Rohn, Karl; Baumgärtner, Wolfgang; Beineke, Andreas

    2014-01-01

    Canine distemper virus (CDV) exhibits a profound lymphotropism that causes immunosuppression and increased susceptibility of affected dogs to opportunistic infections. Similar to human measles virus, CDV is supposed to inhibit terminal differentiation of dendritic cells (DCs), responsible for disturbed repopulation of lymphoid tissues and diminished antigen presenting function in dogs. In order to testify the hypothesis that CDV-infection leads to an impairment of professional antigen presenting cells, canine DCs have been generated from peripheral blood monocytes in vitro and infected with CDV. Virus infection was confirmed and quantified by transmission electron microscopy, CDV-specific immunofluorescence, and virus titration. Flow cytometric analyses revealed a significant down-regulation of the major histocompatibility complex class II and co-stimulatory molecules CD80 and CD86 in CDV-infected DCs, indicative of disturbed antigen presenting capacity. Molecular analyses revealed an increased expression of the immune inhibitory cytokine interleukin-10 in DCs following infection. Results of the present study demonstrate that CDV causes phenotypical changes and altered cytokine expression of DCs, which represent potential mechanisms to evade host immune responses and might contribute to immune dysfunction and virus persistence in canine distemper. PMID:24769532

  14. Canine distemper virus infection leads to an inhibitory phenotype of monocyte-derived dendritic cells in vitro with reduced expression of co-stimulatory molecules and increased interleukin-10 transcription.

    Directory of Open Access Journals (Sweden)

    Visar Qeska

    Full Text Available Canine distemper virus (CDV exhibits a profound lymphotropism that causes immunosuppression and increased susceptibility of affected dogs to opportunistic infections. Similar to human measles virus, CDV is supposed to inhibit terminal differentiation of dendritic cells (DCs, responsible for disturbed repopulation of lymphoid tissues and diminished antigen presenting function in dogs. In order to testify the hypothesis that CDV-infection leads to an impairment of professional antigen presenting cells, canine DCs have been generated from peripheral blood monocytes in vitro and infected with CDV. Virus infection was confirmed and quantified by transmission electron microscopy, CDV-specific immunofluorescence, and virus titration. Flow cytometric analyses revealed a significant down-regulation of the major histocompatibility complex class II and co-stimulatory molecules CD80 and CD86 in CDV-infected DCs, indicative of disturbed antigen presenting capacity. Molecular analyses revealed an increased expression of the immune inhibitory cytokine interleukin-10 in DCs following infection. Results of the present study demonstrate that CDV causes phenotypical changes and altered cytokine expression of DCs, which represent potential mechanisms to evade host immune responses and might contribute to immune dysfunction and virus persistence in canine distemper.

  15. Impact of individual intravenous iron preparations on the differentiation of monocytes towards macrophages and dendritic cells

    Science.gov (United States)

    Fell, Lisa H.; Seiler-Mußler, Sarah; Sellier, Alexander B.; Rotter, Björn; Winter, Peter; Sester, Martina; Fliser, Danilo; Heine, Gunnar H.; Zawada, Adam M.

    2016-01-01

    Background Treatment of iron deficiency with intravenous (i.v.) iron is a first-line strategy to improve anaemia of chronic kidney disease. Previous in vitro experiments demonstrated that different i.v. iron preparations inhibit differentiation of haematopoietic stem cells to monocytes, but their effect on monocyte differentiation to macrophages and mature dendritic cells (mDCs) has not been assessed. We investigated substance-specific effects of iron sucrose (IS), sodium ferric gluconate (SFG), ferric carboxymaltose (FCM) and iron isomaltoside 1000 (IIM) on monocytic differentiation to M1/M2 macrophages and mDCs. Methods Via flow cytometry and microRNA (miRNA) expression analysis, we morphologically and functionally characterized monocyte differentiation to M1/M2 macrophages and mDCs after monocyte stimulation with IS, SFG, FCM and IIM (0.133, 0.266 and 0.533 mg/mL, respectively). To assess potential clinical implications, we compared monocytic phagocytosis capacity in dialysis patients who received either 500 mg IS or IIM. Results Phenotypically, IS and SFG dysregulated the expression of macrophage (e.g. CD40, CD163) and mDC (e.g. CD1c, CD141) surface markers. Functionally, IS and SFG impaired macrophage phagocytosis capacity. Phenotypic and functional alterations were less pronounced with FCM, and virtually absent with IIM. In miRNA expression analysis of mDCs, IS dysregulated miRNAs such as miR-146b-5p and miR-155-5p, which are linked to Toll-like receptor and mitogen-activated protein kinase signalling pathways. In vivo, IS reduced monocytic phagocytosis capacity within 1 h after infusion, while IIM did not. Conclusions This study demonstrates that less stable i.v. iron preparations specifically affect monocyte differentiation towards macrophages and mDCs. PMID:27190361

  16. A Higher Frequency of CD14+ CD169+ Monocytes/Macrophages in Patients with Colorectal Cancer.

    Directory of Open Access Journals (Sweden)

    Chenguang Li

    Full Text Available Monocytes and macrophages can infiltrate into tumor microenvironment and regulate the progression of tumors. This study aimed at determining the frequency of different subsets of circulating monocytes and tumor infiltrating macrophages (TIMs in patients with colorectal cancer (CRC.The frequency of different subsets of circulating monocytes was characterized in 46 CRC patients and 22 healthy controls (HC by flow cytometry. The frequency of different subsets of macrophages was analyzed in TIMs from 30 tumor tissues and in lamina propria mononuclear cells (LPMCs from 12 non-tumor tissues. The concentrations of plasma cytokines and carcinoembryonic antigen (CEA were determined. The potential association of these measures with the values of clinical parameters was analyzed.In comparison with that in the HC, the percentages of circulating CD14+ CD169+, CD14+ CD169+ CD163+ and CD14+ CD169+ CD206+ monocytes and TIMs CD14+ CD169+ as well as IL-10+ CD14+ CD169+, but not IL-12+ CD14+ CD169+ macrophages were significantly increased, accompanied by higher levels of plasma IL-10 in the CRC patients. The percentages of CD14+ CD169+ circulating monocytes and TIM macrophages were associated with the stage of disease and correlated positively with the levels of plasma IL-10 and CEA in CRC patients.Our data suggest that an increase in the frequency of CD14+ CD169+ cells may be associated with the development and progression of CRC and is concomitant rise of both, pro-tumor (M2-like, IL-10 producing and anti-tumor (M1-like, IL-12 producing monocytes and infiltrating macrophages. The frequency of CD14+ CD169+ circulating monocytes and infiltrating macrophages may serve as a biomarker for evaluating the pathogenic degrees of CRC.

  17. Silver nanoparticles impede phorbol myristate acetate-induced monocyte-macrophage differentiation and autophagy

    Science.gov (United States)

    Xu, Yingying; Wang, Liming; Bai, Ru; Zhang, Tianlu; Chen, Chunying

    2015-09-01

    Monocytes/macrophages are important constituents of the innate immune system. Monocyte-macrophage differentiation is not only crucial for innate immune responses, but is also related to some cardiovascular diseases. Silver nanoparticles (AgNPs) are one of the most widely used nanomaterials because of their broad-spectrum antimicrobial properties. However, the effect of AgNPs on the functions of blood monocytes is scarcely reported. Here, we report the impedance effect of AgNPs on THP-1 monocyte differentiation, and that this effect was mediated by autophagy blockade and lysosomal impairment. Firstly, AgNPs inhibit phorbol 12-myristate 13-acetate (PMA)-induced monocyte differentiation by down-regulating both expression of surface marker CD11b and response to lipopolysaccharide (LPS) stimulation. Secondly, autophagy is activated during PMA-induced THP-1 monocyte differentiation, and the autophagy inhibitor chloroquine (CQ) can inhibit this process. Thirdly, AgNPs block the degradation of the autophagy substrate p62 and induce autophagosome accumulation, which demonstrates the blockade of autophagic flux. Fourthly, lysosomal impairments including alkalization and decrease of lysosomal membrane stability were observed in AgNP-treated THP-1 cells. In conclusion, we demonstrate that the impedance of monocyte-macrophage differentiation by AgNPs is mediated by autophagy blockade and lysosomal dysfunction. Our results suggest that crosstalk exists in different biological effects induced by AgNPs.

  18. The role of HIV and monocytes/macrophages in adipose tissue biology.

    Science.gov (United States)

    Shikuma, Cecilia M; Gangcuangco, Louie Mar A; Killebrew, Deirdre A; Libutti, Daniel E; Chow, Dominic C; Nakamoto, Beau K; Liang, Chin Yuan; Milne, Cris I P; Ndhlovu, Lishomwa C; Barbour, Jason D; Shiramizu, Bruce T; Gerschenson, Mariana

    2014-02-01

    To assess the role of HIV and monocytes/macrophages in adipose tissue dysregulation. Cross-sectional study in 5 groups: HIV seronegative, HIV+ antiretroviral therapy (ART)-naive, HIV+ nonlipoatrophic on zidovudine- and/or stavudine-containing ART, HIV+ lipoatrophic on similar ART, and HIV+ on abacavir- or tenofovir-containing ART. HIV DNA in circulating monocyte subsets was quantitated by real-time polymerase chain reaction. Biopsied subcutaneous fat was examined for macrophage content by CD68 staining. Isolated adipocytes and macrophages were cultured and the supernatant assayed for secretory products by Luminex multiplex cytokine technology. Sixty-nine subjects were enrolled. Lipoatrophic subjects had higher median HIV DNA levels (270.5 copies/10 cells) in circulating peripheral CD14CD16 co-expressing monocyte subsets compared with subjects who were ART-naive (25.0 copies), nonlipoatrophic (15.0 copies), or on abacavir/tenofovir (57.5 copies), P adipocytes and adipose macrophage content were marginal. Although adipocyte secretory products were similar, HIV-infected subjects had higher adipose macrophage-derived interleukin (IL)-12p40, IL-6, IL-8, and monocyte inflammatory protein 1 alpha and lower eotaxin and interferon gamma levels than HIV seronegative subjects (P adipose macrophage secretory products were comparable between subjects naive with ART versus those on ART. Circulating HIV-infected and proinflammatory CD14CD16 monocyte subsets contribute to the pathogenesis of HIV-associated lipoatrophy. Among HIV-infected individuals, macrophages, rather than adipocytes, are the primary source of low-grade inflammation in subcutaneous adipose tissue. HIV infection modifies these macrophages to a more proinflammatory phenotype, and these changes are not substantially mitigated by the use of ART.

  19. Molecular mechanisms of HIV-1 persistence in the monocyte-macrophage lineage

    Directory of Open Access Journals (Sweden)

    Rohr Olivier

    2010-04-01

    Full Text Available Abstract The introduction of the highly active antiretroviral therapy (HAART has greatly improved survival. However, these treatments fail to definitively cure the patients and unveil the presence of quiescent HIV-1 reservoirs like cells from monocyte-macrophage lineage. A purge, or at least a significant reduction of these long lived HIV-1 reservoirs will be needed to raise the hope of the viral eradication. This review focuses on the molecular mechanisms responsible for viral persistence in cells of the monocyte-macrophage lineage. Controversy on latency and/or cryptic chronic replication will be specifically evoked. In addition, since HIV-1 infected monocyte-macrophage cells appear to be more resistant to apoptosis, this obstacle to the viral eradication will be discussed. Understanding the intimate mechanisms of HIV-1 persistence is a prerequisite to devise new and original therapies aiming to achieve viral eradication.

  20. The interplay between monocytes/macrophages and CD4+ T cell subsets in rheumatoid arthritis

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    Ceri A. Roberts

    2015-11-01

    Full Text Available Rheumatoid arthritis (RA is a chronic inflammatory disease characterized by inflammation of the synovial lining (synovitis. The inflammation in the RA joint is associated with and driven by immune cell infiltration, synovial hyperproliferation and excessive production of pro-inflammatory mediators, such as TNFα, IFNγ, IL-1β, IL-6 and IL-17, eventually resulting in damage to the cartilage and underlying bone. The RA joint harbors a wide range of immune cell types, including monocytes, macrophages and CD4+ T cells (both pro-inflammatory and regulatory. The interplay between CD14+ myeloid cells and CD4+ T cells can significantly influence CD4+ T cell function and conversely, effector vs. regulatory CD4+ T cell subsets can exert profound effects on monocyte/macrophage function. In this review, we will discuss how the interplay between CD4+ T cells and monocytes/macrophages may contribute to the immunopathology of RA.

  1. Hemoglobin induces monocyte recruitment and CD163-macrophage polarization in abdominal aortic aneurysm.

    Science.gov (United States)

    Rubio-Navarro, Alfonso; Amaro Villalobos, Juan Manuel; Lindholt, Jes S; Buendía, Irene; Egido, Jesús; Blanco-Colio, Luis Miguel; Samaniego, Rafael; Meilhac, Olivier; Michel, Jean Baptiste; Martín-Ventura, José Luis; Moreno, Juan Antonio

    2015-12-15

    Increased hemoglobin (Hb) accumulation was reported in abdominal aortic aneurysms (AAAs). CD163 is a macrophage receptor involved in tissue Hb clearance, however its role in AAA has not been reported. We investigated the role of Hb on monocyte recruitment and differentiation towards CD163 expressing macrophages ex vivo, in vitro and in human AAA. CD163 mRNA and protein expression was significantly higher in human AAA (n=7) vs. healthy wall (n=6). CD163 was predominantly found in adventitia of AAA, coinciding with areas rich in hemosiderin and adjacent to neoangiogenic microvessels. Dual CD14/CD163 expression was observed in recently infiltrated monocytes surrounding microvessels. A higher release of soluble CD163 was observed in the conditioned medium from AAA (AAA-CM, n=10), mainly in the adventitial layer. Similar to Hb, AAA-CM induced CD163-dependent monocyte chemotaxis, especially on circulating monocytes from AAA patients. Hb or AAA-CM promoted differentiation towards CD163(high)/HLA-DR(low)-expressing macrophages, with enhanced Hb uptake, increased anti-inflammatory IL-10 secretion and decreased pro-inflammatory IL-12p40 release. All these effects were partially suppressed when Hb was removed from AAA-CM. Separate analysis on circulating monocytes reported increased percentage of pre-infiltrating CD14(++)CD16(+) monocytes in patients with AAA (n=21), as compared to controls (n=14). A significant increase in CD163 expression in CD14(++)CD16(+) monocyte subpopulation was observed in AAA patients. The presence of Hb in the adventitial AAA-wall promotes the migration and differentiation of activated circulating monocytes in AAA patients, explaining the existence of a protective CD163-macrophage phenotype that could take up the Hb present in the AAA-wall, avoiding its injurious effects. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  2. Macrophage Differentiation from Monocytes Is Influenced by the Lipid Oxidation Degree of Low Density Lipoprotein

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    Jin-Won Seo

    2015-01-01

    Full Text Available LDL plays an important role in atherosclerotic plaque formation and macrophage differentiation. However, there is no report regarding the oxidation degree of LDL and macrophage differentiation. Our study has shown that the differentiation into M1 or M2 macrophages is related to the lipid oxidation level of LDL. Based on the level of lipid peroxidation, LDL is classified into high-oxidized LDL (hi-oxLDL and low-oxidized LDL (low-oxLDL. The differentiation profiles of macrophages were determined by surface receptor expression and cytokine secretion profiles. Low-oxLDL induced CD86 expression and production of TNF-α and IL-12p40 in THP-1 cells, indicating an M1 macrophage phenotype. Hi-oxLDL induced mannose receptor expression and production of IL-6 and monocyte chemoattractant protein-1, which mostly match the phenotype of M2 macrophages. Further supporting evidence for an M2 polarization by hi-oxLDL was the induction of LOX-1 in THP-1 cells treated with hi-oxLDL but not with low-oxLDL. Similar results were obtained in primary human monocytes. Therefore, our results strongly suggest that the oxidation degree of LDL influences the differentiation of monocytes into M1 or M2 macrophages and determines the inflammatory fate in early stages of atherosclerosis.

  3. Anti-myeloperoxidase antibodies attenuate the monocyte response to LPS and shape macrophage development

    Science.gov (United States)

    Popat, Reena J.; Hakki, Seran; Coughlan, Alice M.; Watson, Julie; Little, Mark A.; Spickett, Corinne M.; Lavender, Paul; Afzali, Behdad; Kemper, Claudia; Robson, Michael G.

    2017-01-01

    Anti-neutrophil cytoplasmic antibody (ANCA) vasculitis is characterized by the presence of autoantibodies to myeloperoxidase and proteinase-3, which bind monocytes in addition to neutrophils. While a pathological effect on neutrophils is acknowledged, the impact of ANCA on monocyte function is less well understood. Using IgG from patients we investigated the effect of these autoantibodies on monocytes and found that anti-myeloperoxidase antibodies (MPO-ANCA) reduced both IL-10 and IL-6 secretion in response to LPS. This reduction in IL-10 and IL-6 depended on Fc receptors and enzymatic myeloperoxidase and was accompanied by a significant reduction in TLR-driven signaling pathways. Aligning with changes in TLR signals, oxidized phospholipids, which function as TLR4 antagonists, were increased in monocytes in the presence of MPO-ANCA. We further observed that MPO-ANCA increased monocyte survival and differentiation to macrophages by stimulating CSF-1 production. However, this was independent of myeloperoxidase enzymatic activity and TLR signaling. Macrophages differentiated in the presence of MPO-ANCA secreted more TGF-β and further promoted the development of IL-10– and TGF-β–secreting CD4+ T cells. Thus, MPO-ANCA may promote inflammation by reducing the secretion of antiinflammatory IL-10 from monocytes, and MPO-ANCA can alter the development of macrophages and T cells to potentially promote fibrosis. PMID:28138552

  4. Methamphetamine Enhances HIV-1 Infectivity in Monocyte Derived Dendritic Cells

    OpenAIRE

    2008-01-01

    The US is currently experiencing an epidemic of methamphetamine (Meth) use as a recreational drug. Recent studies also show a high prevalence of HIV-1 infection among Meth users. We report that Meth enhances HIV-1 infectivity of dendritic cells as measured by multinuclear activation of a galactosidase indicator (MAGI) cell assay, p24 assay, and LTR-RU5 amplification. Meth induces increased HIV-1 infection in association with an increase in the HIV-1 coreceptors, CXCR4 and CCR5, and infection ...

  5. Galectin-2 induces a proinflammatory, anti-arteriogenic phenotype in monocytes and macrophages.

    Directory of Open Access Journals (Sweden)

    Cansu Yıldırım

    Full Text Available Galectin-2 is a monocyte-expressed carbohydrate-binding lectin, for which increased expression is genetically determined and associated with decreased collateral arteriogenesis in obstructive coronary artery disease patients. The inhibiting effect of galectin-2 on arteriogenesis was confirmed in vivo, but the mechanism is largely unknown. In this study we aimed to explore the effects of galectin-2 on monocyte/macrophage phenotype in vitro and vivo, and to identify the receptor by which galectin-2 exerts these effects. We now show that the binding of galectin-2 to different circulating human monocyte subsets is dependent on monocyte surface expression levels of CD14. The high affinity binding is blocked by an anti-CD14 antibody but not by carbohydrates, indicating a specific protein-protein interaction. Galectin-2 binding to human monocytes modulated their transcriptome by inducing proinflammatory cytokines and inhibiting pro-arteriogenic factors, while attenuating monocyte migration. Using specific knock-out mice, we show that galectin-2 acts through the CD14/toll-like receptor (TLR-4 pathway. Furthermore, galectin-2 skews human macrophages to a M1-like proinflammatory phenotype, characterized by a reduced motility and expression of an anti-arteriogenic cytokine/growth factor repertoire. This is accompanied by a switch in surface protein expression to CD40-high and CD206-low (M1. In a murine model we show that galectin-2 administration, known to attenuate arteriogenesis, leads to increased numbers of CD40-positive (M1 and reduced numbers of CD206-positive (M2 macrophages surrounding actively remodeling collateral arteries. In conclusion galectin-2 is the first endogenous CD14/TLR4 ligand that induces a proinflammatory, non-arteriogenic phenotype in monocytes/macrophages. Interference with CD14-Galectin-2 interaction may provide a new intervention strategy to stimulate growth of collateral arteries in genetically compromised cardiovascular

  6. Functional role of monocytes and macrophages for the inflammatory response in acute liver injury

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    Henning W Zimmermann

    2012-10-01

    Full Text Available Different etiologies such as drug toxicity, acute viral hepatitis B or acetaminophen poisoning can cause acute liver injury (ALI or even acute liver failure (ALF. Excessive cell death of hepatocytes in the liver is known to result in a strong hepatic inflammation. Experimental murine models of liver injury highlighted the importance of hepatic macrophages, so-called Kupffer cells, for initiating and driving this inflammatory response by releasing proinflammatory cytokines and chemokines including tumor necrosis factor (TNF, interleukin-6 (IL-6, IL-1-beta or monocyte chemoattractant protein 1 (MCP-1, CCL2 as well as activating other non-parenchymal liver cells, e.g. endothelial or hepatic stellate cells (HSC. Many of these proinflammatory mediators can trigger hepatocytic cell death pathways, e.g. via caspase activation, but also activate protective signaling pathways, e.g. via nuclear factor kappa B (NF-kB. Recent studies in mice demonstrated that these macrophage actions largely depend on the recruitment of monocytes into the liver, namely of the inflammatory Ly6c+ (Gr1+ monocyte subset as precursors of tissue macrophages. The chemokine receptor CCR2 and its ligand MCP-1/CCL2 promote monocyte subset infiltration upon liver injury. In contrast, the chemokine receptor CX3CR1 and its ligand fractalkine (CX3CL1 are important negative regulators of monocyte infiltration by controlling their survival and differentiation into functionally diverse macrophage subsets upon injury. The recently identified cellular and molecular pathways for monocyte subset recruitment, macrophage differentiation and interactions with other hepatic cell types in the injured liver may therefore represent interesting novel targets for future therapeutic approaches in ALF.

  7. Age-dependent alterations of monocyte subsets and monocyte-related chemokine pathways in healthy adults

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    Trautwein Christian

    2010-06-01

    Full Text Available Abstract Background Recent experimental approaches have unraveled essential migratory and functional differences of monocyte subpopulations in mice. In order to possibly translate these findings into human physiology and pathophysiology, human monocyte subsets need to be carefully revisited in health and disease. In analogy to murine studies, we hypothesized that human monocyte subsets dynamically change during ageing, potentially influencing their functionality and contributing to immunosenescence. Results Circulating monocyte subsets, surface marker and chemokine receptor expression were analyzed in 181 healthy volunteers (median age 42, range 18-88. Unlike the unaffected total leukocyte or total monocyte counts, non-classical CD14+CD16+ monocytes significantly increased with age, but displayed reduced HLA-DR and CX3CR1 surface expression in the elderly. Classical CD14++CD16- monocyte counts did not vary dependent on age. Serum MCP-1 (CCL2, but not MIP1α (CCL3, MIP1β (CCL4 or fractalkine (CX3CL1 concentrations increased with age. Monocyte-derived macrophages from old or young individuals did not differ with respect to cytokine release in vitro at steady state or upon LPS stimulation. Conclusions Our study demonstrates dynamic changes of circulating monocytes during ageing in humans. The expansion of the non-classical CD14+CD16+ subtype, alterations of surface protein and chemokine receptor expression as well as circulating monocyte-related chemokines possibly contribute to the preserved functionality of the monocyte pool throughout adulthood.

  8. Human monocyte differentiation stage affects response to arachidonic acid.

    Science.gov (United States)

    Escobar-Alvarez, Elizabeth; Pelaez, Carlos A; García, Luis F; Rojas, Mauricio

    2010-01-01

    AA-induced cell death mechanisms acting on human monocytes and monocyte-derived macrophages (MDM), U937 promonocytes and PMA-differentiated U937 cells were studied. Arachidonic acid induced apoptosis and necrosis in monocytes and U937 cells but only apoptosis in MDM and U937D cells. AA increased both types of death in Mycobacterium tuberculosis-infected cells and increased the percentage of TNFalpha+ cells and reduced IL-10+ cells. Experiments blocking these cytokines indicated that AA-mediated death was TNFalpha- and IL-10-independent. The differences in AA-mediated cell death could be explained by high ROS, calpain and sPLA-2 production and activity in monocytes. Blocking sPLA-2 in monocytes and treatment with antioxidants favored M. tuberculosis control whereas AA enhanced M. tuberculosis growth in MDM. Such evidence suggested that AA-modulated effector mechanisms depend on mononuclear phagocytes' differentiation stage.

  9. Glycoengineering of therapeutic antibodies enhances monocyte/macrophage-mediated phagocytosis and cytotoxicity.

    Science.gov (United States)

    Herter, Sylvia; Birk, Martina C; Klein, Christian; Gerdes, Christian; Umana, Pablo; Bacac, Marina

    2014-03-01

    Therapeutic Abs possess several clinically relevant mechanisms of action including perturbation of tumor cell signaling, activation of complement-dependent cytotoxicity, Ab-dependent cellular cytotoxicity (ADCC), Ab-dependent cellular phagocytosis (ADCP), and induction of adaptive immunity. In view of the important role of phagocytic lineage cells in the mechanism of action of therapeutic Abs, we analyzed FcγR receptor-dependent effector functions of monocytes and macrophages triggered by glycoengineered (GE) Abs (having enhanced FcγRIIIa [CD16a] binding affinity) versus their wild-type (WT) counterparts under different experimental conditions. We first defined the precise FcγR repertoire on classical and nonclassical intermediate monocytes--M1 and M2c macrophage populations. We further show that WT and GE Abs display comparable binding and induce similar effector functions (ADCC and ADCP) in the absence of nonspecific, endogenous IgGs. However, in the presence of these IgGs (i.e., in a situation that more closely mimics physiologic conditions), GE Abs display significantly superior binding and promote stronger monocyte and macrophage activity. These data show that in addition to enhancing CD16a-dependent NK cell cytotoxicity, glycoengineering also enhances monocyte and macrophage phagocytic and cytotoxic activities through enhanced binding to CD16a under conditions that more closely resemble the physiologic setting.

  10. Hemoglobin induces monocyte recruitment and CD163-macrophage polarization in abdominal aortic aneurysm

    DEFF Research Database (Denmark)

    Rubio-Navarro, Alfonso; Amaro Villalobos, Juan Manuel; Lindholt, Jes S

    2015-01-01

    BACKGROUND: Increased hemoglobin (Hb) accumulation was reported in abdominal aortic aneurysms (AAAs). CD163 is a macrophage receptor involved in tissue Hb clearance, however its role in AAA has not been reported. We investigated the role of Hb on monocyte recruitment and differentiation towards C...

  11. Increased Expression of Visfatin in Monocytes and Macrophages in Male Acute Myocardial Infarction Patients

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    Cheng-An Chiu

    2012-01-01

    Full Text Available We demonstrated that visfatin expressed in monocytes and neutrophils and increased their reactivity in male acute ST-segment elevation myocardial infarction patients. Furthermore, visfatin was strongly appeared in lipid rich coronary rupture plaques and macrophages. These results suggest another explanation about leukocytes mediated visfatin that may play a pathogenesis role in coronary vulnerable plaques rupture.

  12. Monocytes and macrophages in pregnancy and pre-eclampsia

    NARCIS (Netherlands)

    Faas, Marijke M.; Spaans, Floor; De Vos, Paul

    2014-01-01

    Preeclampsia is an important complication in pregnancy, characterized by hypertension and proteinuria in the second half of pregnancy. Generalized activation of the inflammatory response is thought to play a role in the pathogenesis of pre-eclampsia. Monocytes may play a central role in this

  13. Monocytes and macrophages in pregnancy and pre-eclampsia

    NARCIS (Netherlands)

    Faas, Marijke M.; Spaans, Floor; De Vos, Paul

    2014-01-01

    Preeclampsia is an important complication in pregnancy, characterized by hypertension and proteinuria in the second half of pregnancy. Generalized activation of the inflammatory response is thought to play a role in the pathogenesis of pre-eclampsia. Monocytes may play a central role in this inflamm

  14. Functional activity of monocytes and macrophages in HTLV-1 infected subjects.

    Directory of Open Access Journals (Sweden)

    Camila F Amorim

    2014-12-01

    Full Text Available The Human T lymphotropic virus type-1 (HTLV-1 infects predominantly T cells, inducing proliferation and lymphocyte activation. Additionally, HTLV-1 infected subjects are more susceptible to other infections caused by other intracellular agents. Monocytes/macrophages are important cells in the defense against intracellular pathogens. Our aims were to determine the frequency of monocytes subsets, expression of co-stimulatory molecules in these cells and to evaluate microbicidal ability and cytokine and chemokine production by macrophages from HTLV-1 infected subjects. Participants were 23 HTLV-1 carriers (HC, 22 HAM/TSP patients and 22 healthy subjects (HS not infected with HTLV-1. The frequencies of monocyte subsets and expression of co-stimulatory molecules were determined by flow cytometry. Macrophages were infected with L. braziliensis or stimulated with LPS. Microbicidal activity of macrophages was determined by optic microscopy. Cytokines/chemokines from macrophage supernatants were measured by ELISA. HAM/TSP patients showed an increase frequency of intermediate monocytes, but expression of co-stimulatory molecules was similar between the groups. Macrophages from HTLV-1 infected individuals were infected with L. braziliensis at the same ratio than macrophages from HS, and all the groups had the same ability to kill Leishmania parasites. However, macrophages from HTLV-1 infected subjects produced more CXCL9 and CCL5, and less IL-10 than cells from HS. While there was no correlation between IFN-γ and cytokine/chemokine production by macrophages, there was a correlation between proviral load and TNF and CXCL10. These data showed a dissociation between the inflammatory response and microbicidal ability of macrophages from HTLV-1 infected subjects. While macrophages ability to kill an intracellular pathogen did not differ among HTLV-1 infected subjects, these cells secreted high amount of chemokines even in unstimulated cultures. Moreover the

  15. Nf1+/- monocytes/macrophages induce neointima formation via CCR2 activation.

    Science.gov (United States)

    Bessler, Waylan K; Kim, Grace; Hudson, Farlyn Z; Mund, Julie A; Mali, Raghuveer; Menon, Keshav; Kapur, Reuben; Clapp, D Wade; Ingram, David A; Stansfield, Brian K

    2016-03-15

    Persons with neurofibromatosis type 1 (NF1) have a predisposition for premature and severe arterial stenosis. Mutations in the NF1 gene result in decreased expression of neurofibromin, a negative regulator of p21(Ras), and increases Ras signaling. Heterozygous Nf1 (Nf1(+/-)) mice develop a marked arterial stenosis characterized by proliferating smooth muscle cells (SMCs) and a predominance of infiltrating macrophages, which closely resembles arterial lesions from NF1 patients. Interestingly, lineage-restricted inactivation of a single Nf1 allele in monocytes/macrophages is sufficient to recapitulate the phenotype observed in Nf1(+/-) mice and to mobilize proinflammatory CCR2+ monocytes into the peripheral blood. Therefore, we hypothesized that CCR2 receptor activation by its primary ligand monocyte chemotactic protein-1 (MCP-1) is critical for monocyte infiltration into the arterial wall and neointima formation in Nf1(+/-) mice. MCP-1 induces a dose-responsive increase in Nf1(+/-) macrophage migration and proliferation that corresponds with activation of multiple Ras kinases. In addition, Nf1(+/-) SMCs, which express CCR2, demonstrate an enhanced proliferative response to MCP-1 when compared with WT SMCs. To interrogate the role of CCR2 activation on Nf1(+/-) neointima formation, we induced neointima formation by carotid artery ligation in Nf1(+/-) and WT mice with genetic deletion of either MCP1 or CCR2. Loss of MCP-1 or CCR2 expression effectively inhibited Nf1(+/-) neointima formation and reduced macrophage content in the arterial wall. Finally, administration of a CCR2 antagonist significantly reduced Nf1(+/-) neointima formation. These studies identify MCP-1 as a potent chemokine for Nf1(+/-) monocytes/macrophages and CCR2 as a viable therapeutic target for NF1 arterial stenosis.

  16. Adrenergic regulation of monocyte chemotactic protein 1 leads to enhanced macrophage recruitment and ovarian carcinoma growth

    Science.gov (United States)

    Armaiz-Pena, Guillermo N.; Gonzalez-Villasana, Vianey; Nagaraja, Archana S.; Rodriguez-Aguayo, Cristian; Sadaoui, Nouara C.; Stone, Rebecca L.; Matsuo, Koji; Dalton, Heather J.; Previs, Rebecca A.; Jennings, Nicholas B.; Dorniak, Piotr; Hansen, Jean M.; Arevalo, Jesusa M.G.; Cole, Steve W.; Lutgendorf, Susan K.; Sood, Anil K.; Lopez-Berestein, Gabriel

    2015-01-01

    Increased adrenergic signaling facilitates tumor progression, but the underlying mechanisms remain poorly understood. We examined factors responsible for stress-mediated effects on monocyte/macrophage recruitment into the tumor microenvironment, and the resultant effects on tumor growth. In vitro, MCP1 was significantly increased after catecholamine exposure, which was mediated by cAMP and PKA. Tumor samples from mice subjected to daily restraint stress had elevated MCP1 gene and protein levels, increased CD14+ cells, and increased infiltration of CD68+ cells. hMCP1 siRNA-DOPC nanoparticles significantly abrogated daily restraint stress-induced tumor growth and inhibited infiltration of CD68+ and F4/80+ cells. In ovarian cancer patients, elevated peripheral blood monocytes and tumoral macrophages were associated with worse overall survival. Collectively, we demonstrate that increased adrenergic signaling is associated with macrophage infiltration and mediated by tumor cell-derived MCP1 production. PMID:25738355

  17. Methylome of fetal and maternal monocytes and macrophages at the feto-maternal interface.

    Science.gov (United States)

    Kim, Sun Young; Romero, Roberto; Tarca, Adi L; Bhatti, Gaurav; Kim, Chong Jai; Lee, JoonHo; Elsey, Amelia; Than, Nandor Gabor; Chaiworapongsa, Tinnakorn; Hassan, Sonia S; Kang, Gyeong Hoon; Kim, Jung-Sun

    2012-07-01

    Decidual macrophages (dMφ) of the mother and placental macrophages (Hofbauer cells, HC) of the fetus are deployed at a critical location: the feto-maternal interface. This study was conducted to compare the DNA methylome of maternal and fetal monocytes, dMφ, and HC and thereby to determine the immunobiological importance of DNA methylation in pregnancy. Paired samples were obtained from normal pregnant women at term not in labor and their neonates. Maternal monocytes (MMo) and fetal monocytes (FMo) were isolated from the peripheral blood of mothers and fetal cord blood, respectively. dMφ and HC were obtained from the decidua of fetal membranes and placentas, respectively. DNA methylation profiling was performed using the Illumina Infinium Human Methylation27 BeadChip. Quantitative real-time PCR and Western Blot were performed for validation experiments. (i) Significant differences in DNA methylation were found in each comparison (MMo versus FMo, 65 loci; dMφ versus HC, 266 loci; MMo versus dMφ, 199 loci; FMo versus HC, 1030 loci). (ii) Many of the immune response-related genes were hypermethylated in fetal cells (FMo and HC) compared to maternal cells (MMo and dMφ). (iii) Genes encoding markers of classical macrophage activation were hypermethylated, and genes encoding alternative macrophage activation were hypomethylated in dMφ and HC compared to MMo and FMo, respectively. (iv) mRNA expressions of DNMT1, DNMT3A, and DNMT3B were significantly lower in dMφ than in HC. (v) 5-azacytidine treatment increased expression of INCA1 in dMφ. The findings herein indicate that DNA methylation patterns change during monocyte-macrophage differentiation at the feto-maternal interface. It is also suggested that DNA methylation is an important component of the biological machinery conferring an anti-inflammatory phenotype to macrophages at the feto-maternal interface. © 2012 John Wiley & Sons A/S.

  18. Human monocytes/macrophages are a target of Neisseria meningitidis Adhesin A (NadA).

    Science.gov (United States)

    Franzoso, Susanna; Mazzon, Cristina; Sztukowska, Maryta; Cecchini, Paola; Kasic, Tihana; Capecchi, Barbara; Tavano, Regina; Papini, Emanuele

    2008-05-01

    Specific surface proteins of Neisseria meningitidis have been proposed to stimulate leukocytes during tissue invasion and septic shock. In this study, we demonstrate that the adhesin N. meningitidis Adhesin A (NadA) involved in the colonization of the respiratory epithelium by hypervirulent N. meningitidis B strains also binds to and activates human monocytes/macrophages. Expression of NadA on the surface on Escherichia coli does not increase bacterial-monocyte association, but a NadA-positive strain induced a significantly higher amount of TNF-alpha and IL-8 compared with the parental NadA-negative strain, suggesting that NadA has an intrinsic stimulatory action on these cells. Consistently, highly pure, soluble NadA(Delta351-405), a proposed component of an antimeningococcal vaccine, efficiently stimulates monocytes/macrophages to secrete a selected pattern of cytokines and chemotactic factors characterized by high levels of IL-8, IL-6, MCP-1, and MIP-1alpha and low levels of the main vasoactive mediators TNF-alpha and IL-1. NadA(Delta351-405) also inhibited monocyte apoptosis and determined its differentiation into a macrophage-like phenotype.

  19. Derivation of multipotent progenitors from human circulating CD14+ monocytes.

    Science.gov (United States)

    Seta, Noriyuki; Kuwana, Masataka

    2010-07-01

    Circulating CD14(+) monocytes are originated from hematopoietic stem cells in the bone marrow and believed to be committed precursors for phagocytes, such as macrophages. Recently, we have reported a primitive cell population termed monocyte-derived multipotential cells (MOMCs), which has a fibroblast-like morphology in culture and a unique phenotype positive for CD14, CD45, CD34, and type I collagen. MOMCs are derived from circulating CD14(+) monocytes, but circulating precursors for MOMCs still remain undetermined. Comparative analysis of gene expression profiles of MOMCs and other monocyte-derived cells has revealed that embryonic stem cell markers, Nanog and Oct-4, are specifically expressed by MOMCs. In vitro generation of MOMCs requires binding to fibronectin and exposure to soluble factors derived from activated platelets. MOMCs contain progenitors with capacity to differentiate into a variety of nonphagocytes, including bone, cartilage, fat, skeletal and cardiac muscle, neuron, and endothelium, indicating that circulating monocytes are more multipotent than previously thought. In addition, MOMCs are capable of promoting ex vivo expansion of human hematopoietic progenitor cells through direct cell-to-cell contact and secretion of a variety of hematopoietic growth factors. These findings obtained from the research on MOMCs indicate that CD14(+) monocytes in circulation are involved in a variety of physiologic functions other than innate and acquired immune responses, such as repair and regeneration of the damaged tissue.

  20. CXCR3+ monocytes/macrophages are required for establishment of pulmonary metastases

    Science.gov (United States)

    Butler, Kiah L.; Clancy-Thompson, Eleanor; Mullins, David W.

    2017-01-01

    We present a new foundational role for CXCR3+ monocytes/macrophages in the process of tumor engraftment in the lung. CXCR3 is associated with monocytic and lymphocytic infiltration of inflamed or tumor-bearing lung. Although the requirement for tumor-expressed CXCR3 in metastatic engraftment has been demonstrated, the role of monocyte-expressed CXCR3 had not been appreciated. In a murine model of metastatic-like melanoma, engraftment was coordinate with CXCR3+ monocyte/macrophage accumulation in the lungs and was sensitive to pharmacologic inhibition of CXCR3 signaling. Tumor engraftment to lung was impaired in CXCR3−/− mice, and transient reconstitution with circulating CXCR3-replete monocytes was sufficient to restore engraftment. These data illustrate the paradoxical pro-tumor role for CXCR3 in lung immunobiology wherein the CXCR3 axis drives both the anti-tumor effector cell chemoattraction and pro-tumor infiltration of the lungs and suggests a potential therapeutic target for lung-tropic metastasizing cancers. PMID:28358049

  1. Enhanced alveolar monocytic phagocyte (macrophage) proliferation in tobacco and marijuana smokers

    Energy Technology Data Exchange (ETDEWEB)

    Barbers, R.G.; Evans, M.J.; Gong, H. Jr.; Tashkin, D.P. (Univ. of California-Los Angeles School of Medicine (USA))

    1991-05-01

    We tested the hypothesis that enhanced cell division accounted for the augmented numbers of monocytic phagocytes with characteristics attributed to alveolar macrophages (AM) found in the lungs of habitual tobacco (T) and marijuana (M) smokers. The monocytic phagocytes, that is, alveolar macrophages, were obtained by bronchoalveolar lavage (BAL) from 12 nonsmoking subjects; 10 subjects who smoked T only (TS); 13 subjects who smoked M only (MS); and 6 smokers of both T and M (MTS). The replication of these cells was determined by measuring the incorporation of ({sup 3}H)thymidine into the DNA of dividing cells and visually counting 2,000 cells on autoradiographically prepared cytocentrifuge cell preparations. This study demonstrated that the number of ({sup 3}H)thymidine-labeled monocytic phagocytes with characteristics of alveolar macrophages from either TS or MS have a higher proliferative index compared to cells (macrophages) from nonsmokers, p less than 0.05 by one-way ANOVA. The total number of BAL macrophages that are in mitosis in TS (17.90 +/- 4.50 labeled AM x 10(3)/ml) or MTS (10.50 +/- 4.20 labeled AM x 10(3)/ml) are 18- and 10-fold greater, respectively, than the number obtained from nonsmokers (1.01 +/- 0.18 labeled AM x 10(3)/ml). Interestingly, the number of ({sup 3}H)thymidine-labeled macrophages from MS (2.90 +/- 0.66 labeled AM x 10(3)/ml) are also greater than the number obtained from nonsmokers, although this is not statistically significant. The stimulus augmenting alveolar macrophage replication is as yet unknown but may likely be found in the T or M smoke.

  2. Tacaribe virus but not junin virus infection induces cytokine release from primary human monocytes and macrophages.

    Directory of Open Access Journals (Sweden)

    Allison Groseth

    Full Text Available The mechanisms underlying the development of disease during arenavirus infection are poorly understood. However, common to all hemorrhagic fever diseases is the involvement of macrophages as primary target cells, suggesting that the immune response in these cells may be of paramount importance during infection. Thus, in order to identify features of the immune response that contribute to arenavirus pathogenesis, we have examined the growth kinetics and cytokine profiles of two closely related New World arenaviruses, the apathogenic Tacaribe virus (TCRV and the hemorrhagic fever-causing Junin virus (JUNV, in primary human monocytes and macrophages. Both viruses grew robustly in VeroE6 cells; however, TCRV titres were decreased by approximately 10 fold compared to JUNV in both monocytes and macrophages. Infection of both monocytes and macrophages with TCRV also resulted in the release of high levels of IL-6, IL-10 and TNF-α, while levels of IFN-α, IFN-β and IL-12 were not affected. However, we could show that the presence of these cytokines had no direct effect on growth of either TCRV of JUNV in macrophages. Further analysis also showed that while the production of IL-6 and IL-10 are dependent on viral replication, production of TNF-α also occurs after exposure to UV-inactivated TCRV particles and is thus independent of productive virus infection. Surprisingly, JUNV infection did not have an effect on any of the cytokines examined indicating that, in contrast to other viral hemorrhagic fever viruses, macrophage-derived cytokine production is unlikely to play an active role in contributing to the cytokine dysregulation observed in JUNV infected patients. Rather, these results suggest that an early, controlled immune response by infected macrophages may be critical for the successful control of infection of apathogenic viruses and prevention of subsequent disease, including systemic cytokine dysregulation.

  3. Brugia malayi Microfilariae Induce a Regulatory Monocyte/Macrophage Phenotype That Suppresses Innate and Adaptive Immune Responses

    Science.gov (United States)

    Venugopal, Gopinath; Rao, Gopala B.; Lucius, Richard; Srikantam, Aparna; Hartmann, Susanne

    2014-01-01

    Background Monocytes and macrophages contribute to the dysfunction of immune responses in human filariasis. During patent infection monocytes encounter microfilariae in the blood, an event that occurs in asymptomatically infected filariasis patients that are immunologically hyporeactive. Aim To determine whether blood microfilariae directly act on blood monocytes and in vitro generated macrophages to induce a regulatory phenotype that interferes with innate and adaptive responses. Methodology and principal findings Monocytes and in vitro generated macrophages from filaria non-endemic normal donors were stimulated in vitro with Brugia malayi microfilarial (Mf) lysate. We could show that monocytes stimulated with Mf lysate develop a defined regulatory phenotype, characterised by expression of the immunoregulatory markers IL-10 and PD-L1. Significantly, this regulatory phenotype was recapitulated in monocytes from Wuchereria bancrofti asymptomatically infected patients but not patients with pathology or endemic normals. Monocytes from non-endemic donors stimulated with Mf lysate directly inhibited CD4+ T cell proliferation and cytokine production (IFN-γ, IL-13 and IL-10). IFN-γ responses were restored by neutralising IL-10 or PD-1. Furthermore, macrophages stimulated with Mf lysate expressed high levels of IL-10 and had suppressed phagocytic abilities. Finally Mf lysate applied during the differentiation of macrophages in vitro interfered with macrophage abilities to respond to subsequent LPS stimulation in a selective manner. Conclusions and significance Conclusively, our study demonstrates that Mf lysate stimulation of monocytes from healthy donors in vitro induces a regulatory phenotype, characterized by expression of PD-L1 and IL-10. This phenotype is directly reflected in monocytes from filarial patients with asymptomatic infection but not patients with pathology or endemic normals. We suggest that suppression of T cell functions typically seen in lymphatic

  4. Brugia malayi microfilariae induce a regulatory monocyte/macrophage phenotype that suppresses innate and adaptive immune responses.

    Directory of Open Access Journals (Sweden)

    Noëlle Louise O'Regan

    2014-10-01

    Full Text Available Monocytes and macrophages contribute to the dysfunction of immune responses in human filariasis. During patent infection monocytes encounter microfilariae in the blood, an event that occurs in asymptomatically infected filariasis patients that are immunologically hyporeactive.To determine whether blood microfilariae directly act on blood monocytes and in vitro generated macrophages to induce a regulatory phenotype that interferes with innate and adaptive responses.Monocytes and in vitro generated macrophages from filaria non-endemic normal donors were stimulated in vitro with Brugia malayi microfilarial (Mf lysate. We could show that monocytes stimulated with Mf lysate develop a defined regulatory phenotype, characterised by expression of the immunoregulatory markers IL-10 and PD-L1. Significantly, this regulatory phenotype was recapitulated in monocytes from Wuchereria bancrofti asymptomatically infected patients but not patients with pathology or endemic normals. Monocytes from non-endemic donors stimulated with Mf lysate directly inhibited CD4+ T cell proliferation and cytokine production (IFN-γ, IL-13 and IL-10. IFN-γ responses were restored by neutralising IL-10 or PD-1. Furthermore, macrophages stimulated with Mf lysate expressed high levels of IL-10 and had suppressed phagocytic abilities. Finally Mf lysate applied during the differentiation of macrophages in vitro interfered with macrophage abilities to respond to subsequent LPS stimulation in a selective manner.Conclusively, our study demonstrates that Mf lysate stimulation of monocytes from healthy donors in vitro induces a regulatory phenotype, characterized by expression of PD-L1 and IL-10. This phenotype is directly reflected in monocytes from filarial patients with asymptomatic infection but not patients with pathology or endemic normals. We suggest that suppression of T cell functions typically seen in lymphatic filariasis is caused by microfilaria-modulated monocytes in an IL

  5. Regulation of ICAM-1 in Cells of the Monocyte/Macrophage System in Microgravity

    Directory of Open Access Journals (Sweden)

    Katrin Paulsen

    2015-01-01

    Full Text Available Cells of the immune system are highly sensitive to altered gravity, and the monocyte as well as the macrophage function is proven to be impaired under microgravity conditions. In our study, we investigated the surface expression of ICAM-1 protein and expression of ICAM-1 mRNA in cells of the monocyte/macrophage system in microgravity during clinostat, parabolic flight, sounding rocket, and orbital experiments. In murine BV-2 microglial cells, we detected a downregulation of ICAM-1 expression in clinorotation experiments and a rapid and reversible downregulation in the microgravity phase of parabolic flight experiments. In contrast, ICAM-1 expression increased in macrophage-like differentiated human U937 cells during the microgravity phase of parabolic flights and in long-term microgravity provided by a 2D clinostat or during the orbital SIMBOX/Shenzhou-8 mission. In nondifferentiated U937 cells, no effect of microgravity on ICAM-1 expression could be observed during parabolic flight experiments. We conclude that disturbed immune function in microgravity could be a consequence of ICAM-1 modulation in the monocyte/macrophage system, which in turn could have a strong impact on the interaction with T lymphocytes and cell migration. Thus, ICAM-1 can be considered as a rapid-reacting and sustained gravity-regulated molecule in mammalian cells.

  6. Regulation of ICAM-1 in cells of the monocyte/macrophage system in microgravity.

    Science.gov (United States)

    Paulsen, Katrin; Tauber, Svantje; Dumrese, Claudia; Bradacs, Gesine; Simmet, Dana M; Gölz, Nadine; Hauschild, Swantje; Raig, Christiane; Engeli, Stephanie; Gutewort, Annett; Hürlimann, Eva; Biskup, Josefine; Unverdorben, Felix; Rieder, Gabriela; Hofmänner, Daniel; Mutschler, Lisa; Krammer, Sonja; Buttron, Isabell; Philpot, Claudia; Huge, Andreas; Lier, Hartwin; Barz, Ines; Engelmann, Frank; Layer, Liliana E; Thiel, Cora S; Ullrich, Oliver

    2015-01-01

    Cells of the immune system are highly sensitive to altered gravity, and the monocyte as well as the macrophage function is proven to be impaired under microgravity conditions. In our study, we investigated the surface expression of ICAM-1 protein and expression of ICAM-1 mRNA in cells of the monocyte/macrophage system in microgravity during clinostat, parabolic flight, sounding rocket, and orbital experiments. In murine BV-2 microglial cells, we detected a downregulation of ICAM-1 expression in clinorotation experiments and a rapid and reversible downregulation in the microgravity phase of parabolic flight experiments. In contrast, ICAM-1 expression increased in macrophage-like differentiated human U937 cells during the microgravity phase of parabolic flights and in long-term microgravity provided by a 2D clinostat or during the orbital SIMBOX/Shenzhou-8 mission. In nondifferentiated U937 cells, no effect of microgravity on ICAM-1 expression could be observed during parabolic flight experiments. We conclude that disturbed immune function in microgravity could be a consequence of ICAM-1 modulation in the monocyte/macrophage system, which in turn could have a strong impact on the interaction with T lymphocytes and cell migration. Thus, ICAM-1 can be considered as a rapid-reacting and sustained gravity-regulated molecule in mammalian cells.

  7. Short Communication: HIV Controller T Cells Effectively Inhibit Viral Replication in Alveolar Macrophages.

    Science.gov (United States)

    Walker-Sperling, Victoria E; Merlo, Christian A; Buckheit, Robert W; Lambert, Allison; Tarwater, Patrick; Kirk, Greg D; Drummond, M Bradley; Blankson, Joel N

    Macrophages are targets of HIV-1 infection, and control of viral replication within these cells may be an important component of a T-cell-based vaccine. Although several studies have analyzed the ability of CD8(+) T cells to inhibit viral replication in monocyte-derived macrophages, the effect of T cells on HIV-1-infected tissue macrophages is less clear. We demonstrate here that both CD4(+) and CD8(+) T-cell effectors from HIV controllers are capable of suppressing viral replication in bronchoalveolar lavage-derived alveolar macrophages. These findings have implications for HIV-1 vaccine and eradication strategies.

  8. Ly6C- Monocytes Regulate Parasite-Induced Liver Inflammation by Inducing the Differentiation of Pathogenic Ly6C+ Monocytes into Macrophages.

    Directory of Open Access Journals (Sweden)

    Yannick Morias

    2015-05-01

    Full Text Available Monocytes consist of two well-defined subsets, the Ly6C+ and Ly6C- monocytes. Both CD11b+ myeloid cells populations have been proposed to infiltrate tissues during inflammation. While infiltration of Ly6C+ monocytes is an established pathogenic factor during hepatic inflammation, the role of Ly6C- monocytes remains elusive. Mice suffering experimental African trypanosome infection die from systemic inflammatory response syndrome (SIRS that is initiated by phagocytosis of parasites by liver myeloid cells and culminates in apoptosis/necrosis of liver myeloid and parenchymal cells that reduces host survival. C57BL/6 mice are considered as trypanotolerant to Trypanosoma congolense infection. We have reported that in these animals, IL-10, produced among others by myeloid cells, limits the liver damage caused by pathogenic TNF-producing Ly6C+ monocytes, ensuring prolonged survival. Here, the heterogeneity and dynamics of liver myeloid cells in T. congolense-infected C57/BL6 mice was further dissected. Moreover, the contribution of Ly6C- monocytes to trypanotolerance was investigated. By using FACS analysis and adoptive transfer experiments, we found that the accumulation of Ly6C- monocytes and macrophages in the liver of infected mice coincided with a drop in the pool of Ly6C+ monocytes. Pathogenic TNF mainly originated from Ly6C+ monocytes while Ly6C- monocytes and macrophages were major and equipotent sources of IL-10 within myeloid cells. Moreover, Nr4a1 (Nur77 transcription factor-dependent Ly6C- monocytes exhibited IL-10-dependent and cell contact-dependent regulatory properties contributing to trypanotolerance by suppressing the production of TNF by Ly6C+ monocytes and by promoting the differentiation of the latter cells into macrophages. Thus, Ly6C- monocytes can dampen liver damage caused by an extensive Ly6C+ monocyte-associated inflammatory immune response in T. congolense trypanotolerant animals. In a more general context, Ly6C- or Ly6C

  9. CD14-dependent monocyte isolation enhances phagocytosis of listeria monocytogenes by proinflammatory, GM-CSF-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Caroline Neu

    Full Text Available Macrophages are an important line of defence against invading pathogens. Human macrophages derived by different methods were tested for their suitability as models to investigate Listeria monocytogenes (Lm infection and compared to macrophage-like THP-1 cells. Human primary monocytes were isolated by either positive or negative immunomagnetic selection and differentiated in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF or macrophage colony-stimulating factor (M-CSF into pro- or anti-inflammatory macrophages, respectively. Regardless of the isolation method, GM-CSF-derived macrophages (GM-Mφ stained positive for CD206 and M-CSF-derived macrophages (M-Mφ for CD163. THP-1 cells did not express CD206 or CD163 following incubation with PMA, M- or GM-CSF alone or in combination. Upon infection with Lm, all primary macrophages showed good survival at high multiplicities of infection whereas viability of THP-1 was severely reduced even at lower bacterial numbers. M-Mφ generally showed high phagocytosis of Lm. Strikingly, phagocytosis of Lm by GM-Mφ was markedly influenced by the method used for isolation of monocytes. GM-Mφ derived from negatively isolated monocytes showed low phagocytosis of Lm whereas GM-Mφ generated from positively selected monocytes displayed high phagocytosis of Lm. Moreover, incubation with CD14 antibody was sufficient to enhance phagocytosis of Lm by GM-Mφ generated from negatively isolated monocytes. By contrast, non-specific phagocytosis of latex beads by GM-Mφ was not influenced by treatment with CD14 antibody. Furthermore, phagocytosis of Lactococcus lactis, Escherichia coli, human cytomegalovirus and the protozoan parasite Leishmania major by GM-Mφ was not enhanced upon treatment with CD14 antibody indicating that this effect is specific for Lm. Based on these observations, we propose macrophages derived by ex vivo differentiation of negatively selected human primary monocytes as the most

  10. Intracellular survival of Clostridium chauvoei in bovine macrophages.

    Science.gov (United States)

    Pires, Prhiscylla Sadanã; Santos, Renato Lima; da Paixão, Tatiane Alves; de Oliveira Bernardes, Laura Cristina; de Macêdo, Auricélio Alves; Gonçalves, Luciana Aramuni; de Oliveira Júnior, Carlos Augusto; Silva, Rodrigo Otávio Silveira; Lobato, Francisco Carlos Faria

    2017-02-01

    Clostridium chauvoei is the etiological agent of blackleg, a severe disease of domestic ruminants, causing myonecrosis and serious toxemia with high mortality. Despite the known importance of this agent, studies evaluating its pathogenesis of blackleg are scarce, and many are based on an unproven hypothesis that states that macrophages are responsible for carrying C. chauvoei spores from the intestines to muscles in the early stages of blackleg. Therefore, the present study aimed to investigate the survival of C. chauvoei vegetative cells or spores after phagocytosis by a murine macrophage cell line (RAW 264.7) and bovine monocyte-derived macrophages and to profile inflammatory and anti-inflammatory cytokine transcripts of bovine macrophages infected with C. chauvoei vegetative cells or spores. Both vegetative cells and spores of C. chauvoei remain viable after internalization by murine and bovine macrophages. Bovine macrophages infected with vegetative cells showed a pro-inflammatory profile, while those infected with spores displayed an anti-inflammatory profile. Together, these results corroborate the classical hypothesis that macrophages may play a role in the early pathogenesis of blackleg. Moreover, this is the first study to evaluate the infection kinetics and cytokine profile of bovine monocyte-derived macrophages infected with a Clostridium species.

  11. Influence of phthalates on cytokine production in monocytes and macrophages: a systematic review of experimental trials.

    Directory of Open Access Journals (Sweden)

    Juliana Frohnert Hansen

    Full Text Available Phthalates are a group of endocrine disrupting chemicals suspected to influence the immune system. The aim of this systematic review is to summarise the present knowledge on the influence of phthalates on monocyte and macrophage production and secretion of cytokines, an influence which could affect both pro- and anti-inflammatory abilities of these cells.A systematic search was performed in Medline, Embase and Toxline in June 2013, last updated 3rd of August 2014. Criteria used to select studies were described and published beforehand online on Prospero (http://www.crd.york.ac.uk/NIHR_PROSPERO, registration number CRD42013004236. In vivo, ex vivo and in vitro studies investigating the influence of phthalates on cytokine mRNA expression and cytokine secretion in animals and humans were included. A total of 11 reports, containing 12 studies, were found eligible for inclusion. In these, a total of four different phthalate diesters, six primary metabolites (phthalate monoesters and seven different cytokines were investigated. Though all studies varied greatly in study design and species sources, four out of five studies that investigated di-2-ethylhexyl phthalate found an increased tumour necrosis factor-α secretion/production from monocytes or macrophages. A summary of cytokine measurements was not possible since few studies were comparable in study design and due to insufficient reporting of raw data for most of the included studies.Results from this review have suggested that at least one phthalate (di-2-ethylhexyl phthalate has the ability to enhance tumour necrosis factor-α production/secretion from monocytes/macrophages in vitro, but also observed ex vivo. Influence of other phthalates on other cytokines has only been investigated in few studies. Thus, in vitro studies on primary human monocytes/macrophages as well as more in vivo studies are needed to confirm or dispute these findings.

  12. Activation of caprine arthritis-encephalitis virus expression during maturation of monocytes to macrophages.

    OpenAIRE

    Narayan, O; Kennedy-Stoskopf, S; Sheffer, D; Griffin, D E; Clements, J E

    1983-01-01

    Lentiviruses, which cause arthritis-encephalitis and maedi-visna in goats and sheep, respectively, cause persistent infections in these animals. The viruses replicate productively at low levels in macrophages in diseased organs such as the "maedi lung" and nonproductively in other cell types such as leukocytes in peripheral blood. Nonproductive infections become productive during in vitro cultivation of the cells. This study showed that monocytes were the only cells in the peripheral blood le...

  13. Role of monocytes and macrophages in experimental and human acute liver failure

    Institute of Scientific and Technical Information of China (English)

    Lucia; A; Possamai; Charalambos; Gustav; Antoniades; Quentin; M; Anstee; Alberto; Quaglia; Diego; Vergani; Mark; Thursz; Julia; Wendon

    2010-01-01

    Acute liver failure (ALF) is a devastating clinical syndrome characterised by progressive encephalopathy, coagulopathy, and circulatory dysfunction, which commonly leads to multiorgan failure and death. Central to the pathogenesis of ALF is activation of the immune system with mobilisation of cellular effectors and massive production of cytokines. As key components of the innate immune system, monocytes and macrophages are postulated to play a central role in the initiation, progression and resolution of AL...

  14. Critical Role for Monocytes/Macrophages in Rapid Progression to AIDS in Pediatric Simian Immunodeficiency Virus-Infected Rhesus Macaques.

    Science.gov (United States)

    Sugimoto, Chie; Merino, Kristen M; Hasegawa, Atsuhiko; Wang, Xiaolei; Alvarez, Xavier A; Wakao, Hiroshi; Mori, Kazuyasu; Kim, Woong-Ki; Veazey, Ronald S; Didier, Elizabeth S; Kuroda, Marcelo J

    2017-09-01

    Infant humans and rhesus macaques infected with the human or simian immunodeficiency virus (HIV or SIV), respectively, express higher viral loads and progress more rapidly to AIDS than infected adults. Activated memory CD4(+) T cells in intestinal tissues are major primary target cells for SIV/HIV infection, and massive depletion of these cells is considered a major cause of immunodeficiency. Monocytes and macrophages are important cells of innate immunity and also are targets of HIV/SIV infection. We reported previously that a high peripheral blood monocyte turnover rate was predictive for the onset of disease progression to AIDS in SIV-infected adult macaques. The purpose of this study was to determine if earlier or higher infection of monocytes/macrophages contributes to the more rapid progression to AIDS in infants. We observed that uninfected infant rhesus macaques exhibited higher physiologic baseline monocyte turnover than adults. Early after SIV infection, the monocyte turnover further increased, and it remained high during progression to AIDS. A high percentage of terminal deoxynucleotidyltransferase dUTP nick end label (TUNEL)-positive macrophages in the lymph nodes (LNs) and intestine corresponded with an increasing number of macrophages derived from circulating monocytes (bromodeoxyuridine positive [BrdU(+)] CD163(+)), suggesting that the increased blood monocyte turnover was required to rapidly replenish destroyed tissue macrophages. Immunofluorescence analysis further demonstrated that macrophages were a significant portion of the virus-producing cells found in LNs, intestinal tissues, and lungs. The higher baseline monocyte turnover in infant macaques and subsequent macrophage damage by SIV infection may help explain the basis of more rapid disease progression to AIDS in infants.IMPORTANCE HIV infection progresses much more rapidly in pediatric cases than in adults; however, the mechanism for this difference is unclear. Using the rhesus macaque model

  15. RNY (YRNA)-derived small RNAs regulate cell death and inflammation in monocytes/macrophages.

    Science.gov (United States)

    Hizir, Zoheir; Bottini, Silvia; Grandjean, Valerie; Trabucchi, Michele; Repetto, Emanuela

    2017-01-05

    The recent discovery of new classes of small RNAs has opened unknown territories to explore new regulations of physiopathological events. We have recently demonstrated that RNY (or Y RNA)-derived small RNAs (referred to as s-RNYs) are an independent class of clinical biomarkers to detect coronary artery lesions and are associated with atherosclerosis burden. Here, we have studied the role of s-RNYs in human and mouse monocytes/macrophages and have shown that in lipid-laden monocytes/macrophages s-RNY expression is timely correlated to the activation of both NF-κB and caspase 3-dependent cell death pathways. Loss- or gain-of-function experiments demonstrated that s-RNYs activate caspase 3 and NF-κB signaling pathways ultimately promoting cell death and inflammatory responses. As, in atherosclerosis, Ro60-associated s-RNYs generated by apoptotic macrophages are released in the blood of patients, we have investigated the extracellular function of the s-RNY/Ro60 complex. Our data demonstrated that s-RNY/Ro60 complex induces caspase 3-dependent cell death and NF-κB-dependent inflammation, when added to the medium of cultured monocytes/macrophages. Finally, we have shown that s-RNY function is mediated by Toll-like receptor 7 (TLR7). Indeed using chloroquine, which disrupts signaling of endosome-localized TLRs 3, 7, 8 and 9 or the more specific TLR7/9 antagonist, the phosphorothioated oligonucleotide IRS954, we blocked the effect of either intracellular or extracellular s-RNYs. These results position s-RNYs as relevant novel functional molecules that impacts on macrophage physiopathology, indicating their potential role as mediators of inflammatory diseases, such as atherosclerosis.

  16. 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.

  17. Induction of monocyte-derived dendritic cell differentiation by asthmatic serum in a transendothelial trafficking model%支气管哮喘患者血清对内皮穿越模型中单核细胞分化为树突状细胞的作用

    Institute of Scientific and Technical Information of China (English)

    周林福; 王文璐; 李红岩; 张明顺; 季晓辉; 何韶衡; 黄茂; 殷凯生

    2011-01-01

    新的体外实验平台.%Objective To explore the effect of asthmatic and healthy serum on differentiation and function of monocyte-derived dendritic cells (MDDC) in a transendothelial trafficking model. Methods The sera and peripheral blood mononuclear cells (PBMC) were separated from 12 asthmatic patients and 12 healthy volunteers, and monocytes were selected from PBMC using magnetic beads. The trypsin-digested human umbilical vein endothelial cells (HUVEC) at passage 2 from 5 healthy lying-in women were used to construct the transendothelial trafficking model under asthmatic or healthy serum, wherein MDDC were identified by silver nitrate staining and scanning electron microscopy. Nuclear factor κB (NF-κB) activity was determined by electrophoretic mobility shift assay. Flow cytometry, ELISA and mixed leukocyte reaction were relevantly utilized to detect the phenotype, cytokine and T cell proliferation. Results ( 1 ) Monocytes traversed through HUVEC monolayer after 2 h, and reverse-transmigrated to develop into DC 48 h later. ( 2 ) The healthy serum stimulated monocytes into immature MDDC with lower CD14 [ ( 20 ±5)%] (F=49.01, P<0.05), and higher HLA-DR, CD80, CD86 and CD83 [(43 ±4)%, (17.9 ±3.5)%, (43 ± 11)% and (6.7 ± 1.8)%, respectively] (F= 10.35 -40.17, all P<0.05) than monocytes did before transmigration at 0 h [ CD14(81 ±6)%, HLA-DR (24 ±5)%, CD80(2. 8 ±2. 0)%,CD86( 14 ±4)% and CD83(0. 9 ±0. 8)%, respectively]. (3) The asthmatic serum stimulated monocytes into mature MDDC, characteristic of dendrites, with similar HLA-DR and CD86 [ ( 55 ± 6 ) % and ( 59 ±12)%] (F=15.29 and 35.97, all P >0.05), higher CD80 and CD83 [(49.7 ± 10.2)% and (30.2 ±6. 8) % ] ( F = 4. 01 and 20. 68, all P < 0. 05), accompanied by increased levels of NF-κB activity, IL-12 p70 and T cell proliferation [ ( 100 ± 11 )%, (568 ±43) ng/L and (2033 ± 198) cpm, respectively] (F=49. 23 - 350. 84, all P < 0. 05 ) relative to the healthy serum-stimulated immature MDDC [ ( 12 ± 3 ) %,(220 ± 35) ng

  18. Genetics of SLE: Functional Relevance for Monocytes/Macrophages in Disease

    Directory of Open Access Journals (Sweden)

    Jennifer C. Byrne

    2012-01-01

    Full Text Available Genetic studies in the last 5 years have greatly facilitated our understanding of how the dysregulation of diverse components of the innate immune system contributes to pathophysiology of SLE. A role for macrophages in the pathogenesis of SLE was first proposed as early as the 1980s following the discovery that SLE macrophages were defective in their ability to clear apoptotic cell debris, thus prolonging exposure of potential autoantigens to the adaptive immune response. More recently, there is an emerging appreciation of the contribution both monocytes and macrophages play in orchestrating immune responses with perturbations in their activation or regulation leading to immune dysregulation. This paper will focus on understanding the relevance of genes identified as being associated with innate immune function of monocytes and macrophages and development of SLE, particularly with respect to their role in (1 immune complex (IC recognition and clearance, (2 nucleic acid recognition via toll-like receptors (TLRs and downstream signalling, and (3 interferon signalling. Particular attention will be paid to the functional consequences these genetic associations have for disease susceptibility or pathogenesis.

  19. Gene expression study of monocytes/macrophages during early foreign body reaction and identification of potential precursors of myofibroblasts.

    Directory of Open Access Journals (Sweden)

    Lindsay Mesure

    Full Text Available Foreign body reaction (FBR, initiated by adherence of macrophages to biomaterials, is associated with several complications. Searching for mechanisms potentially useful to overcome these complications, we have established the signaling role of monocytes/macrophages in the development of FBR and the presence of CD34(+ cells that potentially differentiate into myofibroblasts. Therefore, CD68(+ cells were in vitro activated with fibrinogen and also purified from the FBR after 3 days of implantation in rats. Gene expression profiles showed a switch from monocytes and macrophages attracted by fibrinogen to activated macrophages and eventually wound-healing macrophages. The immature FBR also contained a subpopulation of CD34(+ cells, which could be differentiated into myofibroblasts. This study showed that macrophages are the clear driving force of FBR, dependent on milieu, and myofibroblast deposition and differentiation.

  20. Δ(9)-Tetrahydrocannabinol treatment during human monocyte differentiation reduces macrophage susceptibility to HIV-1 infection.

    Science.gov (United States)

    Williams, Julie C; Appelberg, Sofia; Goldberger, Bruce A; Klein, Thomas W; Sleasman, John W; Goodenow, Maureen M

    2014-06-01

    The major psychoactive component of marijuana, Δ(9)-tetrahydrocannabinol (THC), also acts to suppress inflammatory responses. Receptors for THC, CB1, CB2, and GPR55, are differentially expressed on multiple cell types including monocytes and macrophages, which are important modulators of inflammation in vivo and target cells for HIV-1 infection. Use of recreational and medicinal marijuana is increasing, but the consequences of marijuana exposure on HIV-1 infection are unclear. Ex vivo studies were designed to investigate effects on HIV-1 infection in macrophages exposed to THC during or following differentiation. THC treatment of primary human monocytes during differentiation reduced HIV-1 infection of subsequent macrophages by replication competent or single cycle CCR5 using viruses. In contrast, treatment of macrophages with THC immediately prior to or continuously following HIV-1 exposure failed to alter infection. Specific receptor agonists indicated that the THC effect during monocyte differentiation was mediated primarily through CB2. THC reduced the number of p24 positive cells with little to no effect on virus production per infected cell, while quantitation of intracellular viral gag pinpointed the THC effect to an early event in the viral life cycle. Cells treated during differentiation with THC displayed reduced expression of CD14, CD16, and CD163 and donor dependent increases in mRNA expression of selected viral restriction factors, suggesting a fundamental alteration in phenotype. Ultimately, the mechanism of THC suppression of HIV-1 infection was traced to a reduction in cell surface HIV receptor (CD4, CCR5 and CXCR4) expression that diminished entry efficiency.

  1. Δ9-tetrahydrocannabinol treatment during human monocyte differentiation reduces macrophage susceptibility to HIV-1 infection

    Science.gov (United States)

    Williams, Julie C.; Appelberg, Sofia; Goldberger, Bruce A.; Klein, Thomas W.; Sleasman, John W.; Goodenow, Maureen M.

    2014-01-01

    The major psychoactive component of marijuana, Δ9-tetrahydrocannabinol (THC), also acts to suppress inflammatory responses. Receptors for THC, CB1, CB2, and GPR55, are differentially expressed on multiple cell types including monocytes and macrophages, which are important modulators of inflammation in vivo and target cells for HIV-1 infection. Use of recreational and medicinal marijuana is increasing, but the consequences of marijuana exposure on HIV-1 infection are unclear. Ex vivo studies were designed to investigate effects on HIV-1 infection in macrophages exposed to THC during or following differentiation. THC treatment of primary human monocytes during differentiation reduced HIV-1 infection of subsequent macrophages by replication competent or single cycle CCR5 using viruses. In contrast, treatment of macrophages with THC immediately prior to or continuously following HIV-1 exposure failed to alter infection. Specific receptor agonists indicated that the THC effect during monocyte differentiation was mediated primarily through CB2. THC reduced the number of p24 positive cells with little to no effect on virus production per infected cell, while quantitation of intracellular viral gag pinpointed the THC effect to an early event in the viral life cycle. Cells treated during differentiation with THC displayed reduced expression of CD14, CD16, and CD163 and donor dependent increases in mRNA expression of selected viral restriction factors, suggesting a fundamental alteration in phenotype. Ultimately, the mechanism of THC suppression of HIV-1 infection was traced to a reduction in cell surface HIV receptor (CD4, CCR5 and CXCR4) expression that diminished entry efficiency. PMID:24562630

  2. Functional TRAIL receptors in monocytes and tumor-associated macrophages: A possible targeting pathway in the tumor microenvironment

    Science.gov (United States)

    Liguori, Manuela; Buracchi, Chiara; Pasqualini, Fabio; Bergomas, Francesca; Pesce, Samantha; Sironi, Marina; Grizzi, Fabio; Mantovani, Alberto

    2016-01-01

    Despite the accepted dogma that TRAIL kills only tumor cells and spares normal ones, we show in this study that mononuclear phagocytes are susceptible to recombinant TRAIL via caspase-dependent apoptosis. Human resting monocytes and in vitro-differentiated macrophages expressed substantial levels of the functional TRAIL receptors (TRAIL-R1 and TRAIL-R2), while neutrophils and lymphocytes mostly expressed the non-signaling decoy receptor (TRAIL-R3). Accordingly, exclusively monocytes and macrophages activated caspase-8 and underwent apoptosis upon recombinant TRAIL treatment. TRAIL-Rs were up-regulated by anti-inflammatory agents (IL-10, glucocorticoids) and by natural compounds (Apigenin, Quercetin, Palmitate) and their treatment resulted in increased TRAIL-induced apoptosis. In mice, the only signaling TRAIL-R (DR5) was preferentially expressed by blood monocytes rather than neutrophils or lymphocytes. In both mice and humans, Tumor-Associated Macrophages (TAM) expressed functional TRAIL-R, while resident macrophages in normal tissues did not. As a proof of principle, we treated mice bearing a murine TRAIL-resistant fibrosarcoma with recombinant TRAIL. We observed significant decrease of circulating monocytes and infiltrating TAM, as well as reduced tumor growth and lower metastasis formation. Overall, these findings demonstrate that human and murine monocytes/macrophages are, among leukocytes, uniquely susceptible to TRAIL-mediated killing. This differential susceptibility to TRAIL could be exploited to selectively target macrophages in tumors. PMID:27191500

  3. Intravital live cell triggered imaging system reveals monocyte patrolling and macrophage migration in atherosclerotic arteries

    Science.gov (United States)

    McArdle, Sara; Chodaczek, Grzegorz; Ray, Nilanjan; Ley, Klaus

    2015-02-01

    Intravital multiphoton imaging of arteries is technically challenging because the artery expands with every heartbeat, causing severe motion artifacts. To study leukocyte activity in atherosclerosis, we developed the intravital live cell triggered imaging system (ILTIS). This system implements cardiac triggered acquisition as well as frame selection and image registration algorithms to produce stable movies of myeloid cell movement in atherosclerotic arteries in live mice. To minimize tissue damage, no mechanical stabilization is used and the artery is allowed to expand freely. ILTIS performs multicolor high frame-rate two-dimensional imaging and full-thickness three-dimensional imaging of beating arteries in live mice. The external carotid artery and its branches (superior thyroid and ascending pharyngeal arteries) were developed as a surgically accessible and reliable model of atherosclerosis. We use ILTIS to demonstrate Cx3cr1GFP monocytes patrolling the lumen of atherosclerotic arteries. Additionally, we developed a new reporter mouse (Apoe-/-Cx3cr1GFP/+Cd11cYFP) to image GFP+ and GFP+YFP+ macrophages "dancing on the spot" and YFP+ macrophages migrating within intimal plaque. ILTIS will be helpful to answer pertinent open questions in the field, including monocyte recruitment and transmigration, macrophage and dendritic cell activity, and motion of other immune cells.

  4. SMAD-PI3K-Akt-mTOR pathway mediates BMP-7 polarization of monocytes into M2 macrophages.

    Directory of Open Access Journals (Sweden)

    Crystal Rocher

    Full Text Available Previously we demonstrated that bone morphogenetic protein-7 (BMP-7 treatment polarizes monocytes into M2 macrophages and increases the expression of anti-inflammatory cytokines. Despite these findings, the mechanisms for the observed BMP-7 induced monocyte polarization into M2 macrophages are completely unknown. In this study, we demonstrate the mechanisms involved in the polarization of monocytes into M2 macrophages. Apoptotic conditioned media (ACM was generated to mimic the stressed conditions, inducing monocyte polarization. Monocytes were treated with ACM along with BMP-7 and/or its inhibitor, follistatin, for 48 hours. Furthermore, an inhibitor of the PI3K pathway, LY-294002, was also studied. Our data show that BMP-7 induces polarization of monocytes into M2 macrophages while significantly increasing the expression of anti-inflammatory markers, arginase-1 and IL-10, and significantly (p<0.05 decreasing the expression of pro-inflammatory markers iNOS, IL-6, TNF-α and MCP-1; (p<0.05. Moreover, addition of the PI3K inhibitor, LY-294002, significantly (p<0.05 decreases upregulation of IL-10 and arginase-1, suggesting involvement of the PI3K pathway in M2 macrophage polarization. Next, following BMP-7 treatment, a significant (p<0.05 increase in p-SMAD1/5/8 and p-PI3K expression resulting in downstream activation of p-Akt and p-mTOR was observed. Furthermore, expression of p-PTEN, an inhibitor of the PI3K pathway, was significantly (p<0.05 increased in the ACM group. However, BMP-7 treatment inhibited its expression, suggesting involvement of the PI3K-Akt-mTOR pathway. In conclusion, we demonstrate that BMP-7 polarizes monocytes into M2 macrophages and enhances anti-inflammatory cytokine expression which is mediated by the activated SMAD-PI3K-Akt-mTOR pathway.

  5. Tracking the Spatial and Functional Gradient of Monocyte-To-Macrophage Differentiation in Inflamed Lung.

    Science.gov (United States)

    Sen, Debasish; Jones, Stephen M; Oswald, Erin M; Pinkard, Henry; Corbin, Kaitlin; Krummel, Matthew F

    2016-01-01

    Myeloid-derived cells such as monocytes, dendritic cells (DCs), and macrophages are at the heart of the immune effector function in an inflammatory response. But because of the lack of an efficient imaging system to trace these cells live during their migration and maturation in their native environment at sub-cellular resolution, our knowledge is limited to data available from specific time-points analyzed by flow cytometry, histology, genomics and other immunological methods. Here, we have developed a ratiometric imaging method for measuring monocyte maturation in inflamed mouse lungs in situ using real-time using 2-photon imaging and complementary methods. We visualized that while undifferentiated monocytes were predominantly found only in the vasculature, a semi-differentiated monocyte/macrophage population could enter the tissue and resembled more mature and differentiated populations by morphology and surface phenotype. As these cells entered and differentiated, they were already selectively localized near inflamed airways and their entry was associated with changes in motility and morphology. We were able to visualize these during the act of differentiation, a process that can be demonstrated in this way to be faster on a per-cell basis under inflammatory conditions. Finally, our in situ analyses demonstrated increases, in the differentiating cells, for both antigen uptake and the ability to mediate interactions with T cells. This work, while largely confirming proposed models for in situ differentiation, provides important in situ data on the coordinated site-specific recruitment and differentiation of these cells and helps elaborate the predominance of immune pathology at the airways. Our novel imaging technology to trace immunogenic cell maturation in situ will complement existing information available on in situ differentiation deduced from other immunological methods, and assist better understanding of the spatio-temporal cellular behavior during an

  6. Mesenchymal stem/stromal cells precondition lung monocytes/macrophages to produce tolerance against allo- and autoimmunity in the eye.

    Science.gov (United States)

    Ko, Jung Hwa; Lee, Hyun Ju; Jeong, Hyun Jeong; Kim, Mee Kum; Wee, Won Ryang; Yoon, Sun-Ok; Choi, Hosoon; Prockop, Darwin J; Oh, Joo Youn

    2016-01-01

    Intravenously administered mesenchymal stem/stromal cells (MSCs) engraft only transiently in recipients, but confer long-term therapeutic benefits in patients with immune disorders. This suggests that MSCs induce immune tolerance by long-lasting effects on the recipient immune regulatory system. Here, we demonstrate that i.v. infusion of MSCs preconditioned lung monocytes/macrophages toward an immune regulatory phenotype in a TNF-α-stimulated gene/protein (TSG)-6-dependent manner. As a result, mice were protected against subsequent immune challenge in two models of allo- and autoimmune ocular inflammation: corneal allotransplantation and experimental autoimmune uveitis (EAU). The monocytes/macrophages primed by MSCs expressed high levels of MHC class II, B220, CD11b, and IL-10, and exhibited T-cell-suppressive activities independently of FoxP3(+) regulatory T cells. Adoptive transfer of MSC-induced B220(+)CD11b(+) monocytes/macrophages prevented corneal allograft rejection and EAU. Deletion of monocytes/macrophages abrogated the MSC-induced tolerance. However, MSCs with TSG-6 knockdown did not induce MHC II(+)B220(+)CD11b(+) cells, and failed to attenuate EAU. Therefore, the results demonstrate a mechanism of the MSC-mediated immune modulation through induction of innate immune tolerance that involves monocytes/macrophages.

  7. Activation of farnesoid X receptor downregulates monocyte chemoattractant protein-1 in murine macrophage.

    Science.gov (United States)

    Li, Liangpeng; Zhang, Qian; Peng, Jiahe; Jiang, Chanjui; Zhang, Yan; Shen, Lili; Dong, Jinyu; Wang, Yongchao; Jiang, Yu

    2015-11-27

    Farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily, which plays important roles in bile acids/lipid homeostasis and inflammation. Monocyte chemoattractant protein-1 (MCP-1) contributes to macrophage infiltration into body tissues during inflammation. Here we investigated whether FXR can regulate MCP-1 expression in murine macrophage. FXR activation down regulate MCP-1 mRNA and protein levels in ANA-1 and Raw264.7 cells. Luciferase reporter assay, Gel shift and Chromatin immunoprecipitation assays have revealed that the activated FXR bind to the FXR element located in -738 bp ∼  -723 bp in MCP-1 promoter. These results suggested that FXR may serve as a novel target for regulating MCP-1 levels for the inflammation related diseases therapies.

  8. Induction of HO-1 in tissue macrophages and monocytes in fatal falciparum malaria and sepsis

    Directory of Open Access Journals (Sweden)

    Liomba N

    2003-11-01

    Full Text Available Abstract Background As well as being inducible by haem, haemoxygenase -1 (HO-1 is also induced by interleukin-10 and an anti-inflammatory prostaglandin, 15d PGJ2, the carbon monoxide thus produced mediating the anti-inflammatory effects of these molecules. The cellular distribution of HO-1, by immunohistochemistry, in brain, lung and liver in fatal falciparum malaria, and in sepsis, is reported. Methods Wax sections were stained, at a 1:1000 dilution of primary antibody, for HO-1 in tissues collected during paediatric autopsies in Blantyre, Malawi. These comprised 37 acutely ill comatose patients, 32 of whom were diagnosed clinically as cerebral malaria and the other 5 as bacterial diseases with coma. Another 3 died unexpectedly from an alert state. Other control tissues were from Australian adults. Results Apart from its presence in splenic red pulp macrophages and microhaemorrhages, staining for HO-1 was confined to intravascular monocytes and certain tissue macrophages. Of the 32 clinically diagnosed cerebral malaria cases, 11 (category A cases had negligible histological change in the brain and absence of or scanty intravascular sequestration of parasitized erythrocytes. Of these 11 cases, eight proved at autopsy to have other pathological changes as well, and none of these eight showed HO-1 staining within the brain apart from isolated moderate staining in one case. Two of the three without another pathological diagnosis showed moderate staining of scattered monocytes in brain vessels. Six of these 11 (category A cases exhibited strong lung staining, and the Kupffer cells of nine of them were intensely stained. Of the seven (category B cases with no histological changes in the brain, but appreciable sequestered parasitised erythrocytes present, one was without staining, and the other six showed strongly staining, rare or scattered monocytes in cerebral vessels. All six lung sections not obscured by neutrophils showed strong staining of

  9. 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

    for the resolution of inflammation. Clin Exp Immunol. 2005 Dec;142(3):481-9. 2. Mantovani A, Sica A, Sozzani S, Allavena P, Vecchi A, Locati M. The chemokine system in diverse forms of macrophage activation and polarization. Trends Immunol. 2004 Dec;25(12):677-86. 3. Weaver LK, Hintz-Goldstein KA, Pioli PA, Wardwell...

  10. Research on the Effects of the Fluconazole resistance and Fluconazole susceptible Candida Albicans Strains in RVVC on the Shapes and the Surface Molecules of the Human Peripheral Blood Monocyte-Derived DCs%RVVC白念珠菌氟康唑敏感株和耐药株对外周血DCs分化成熟的影响

    Institute of Scientific and Technical Information of China (English)

    宣晓梅; 刘静; 沈丽; 刘瑞琴; 李艳佳; 刘丽娟; 李英涛; 于亮

    2012-01-01

    目的 探讨复发性外阴阴道念珠菌病(RVVC)患者阴道白念珠菌对氟康唑敏感情况及其对人外周血单核细胞源树突状细胞(DCs)形态及表面分子的影响.方法 科玛嘉念珠菌显色培养基及芽管试验分离、鉴定白念珠菌;MIC法判断敏感菌及耐药菌;体外培养正常人外周血单核细胞源DCs,实验组以不同剂量氟康唑敏感菌悬液、耐药菌悬液刺激,对照组以同等剂量标准菌悬液刺激后,倒置显微镜观察细胞形态变化,流式细胞仪检测细胞表面分子的表达.结果 RVVC患者阴道分泌物共分离出白念珠菌108株,其中对氟康唑敏感67株(62.04%),耐药41株(37.96%);各实验组与对照组DCs细胞形态无明显差别.DCs表面分子CD80,CD86表达量均与白念珠菌悬液剂量呈正相关;加入标准菌株悬液的对照组DCs表面分子CD80,CD86表达明显高于同等剂量加入敏感菌悬液和耐药菌悬液的实验组,且加入敏感菌悬液的实验1组DCs表面分子CD80,CD86表达明显高于同等剂量加入耐药菌悬液的实验2组.结论 在一定剂量范围内,白念珠菌可促进DCs表面分子CD80,CD86表达,使DCs进一步成熟,氟康唑敏感菌株优于耐药菌株,但两者均不及标准菌株.%Objective To investigate the effects of the fluconazole resistance and fluconazole susceptible Candida albicans strains in recurrent vulvovaginal candidiasis( RVVC) on the Shapes and the Surface Molecules of the Human Peripheral Blood Monocyte-Derived dendritic cells ( DCs ). Methods Candida albicans in RVVC were separated and were judged by the CHROMagar Monilia colouration nutritive medium and the genn-tube-formming tests, and then were judged the fluconazole-resistance or fluconazole-usceptible strains by MIC tests. The dendritic cells were obtained by culture in vitro. Then after they were stimulated by the different doses of Candida albicans including the fluconazole-esistance and fluconazole-usceptible strains

  11. 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

  12. Human recombinant macrophage inflammatory protein-1 alpha and -beta and monocyte chemotactic and activating factor utilize common and unique receptors on human monocytes.

    Science.gov (United States)

    Wang, J M; Sherry, B; Fivash, M J; Kelvin, D J; Oppenheim, J J

    1993-04-01

    The human macrophage inflammatory proteins-1 alpha and -beta (MIP-1 alpha and -beta), which are also known as LD78 and ACT2, respectively, are distinct but highly related members of the chemoattractant cytokine (chemokine) family. rMIP-1 alpha and -beta labeled with 125I specifically bind to human peripheral blood monocytes, the monocytic cell line THP-1, peripheral blood T cells, and the YT cell line. Steady state binding experiments revealed approximately 3000 high affinity binding sites/cell for MIP-1 alpha on human monocytes and on THP-1 cells, with Kd values of 383 pM and 450 pM, respectively. Human MIP-1 alpha and -beta had nearly identical affinities for the binding sites and each competed equally well for binding. Human monocyte chemotactic and activating factor (MCAF), a member of the same chemokine family, consistently displaced about 25% of human MIP-1 alpha and -beta binding on monocytes but not on YT cells, which did not bind MCAF. On the other hand, human rMIP-1 alpha and -beta partially inhibited binding of radiolabeled MCAF to monocytes. Both MIP-1 alpha and -beta were chemotactic for human monocytes. Preincubation of monocytes with human rMIP-1 alpha or -beta markedly reduced cell migration towards the other cytokine, whereas preincubation with human rMCAF only partially desensitized the monocyte chemotaxis response to human rMIP-1 alpha or -beta. These data suggest the existence of three subtypes of receptors, i.e., one unique receptor shared by MIP-1 alpha and -beta, a second unique receptor for MCAF, and a third species that recognizes both MCAF and MIP-1 peptides.

  13. 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.

  14. Monocyte/macrophage lineage commitment and distribution are affected by the lack of regulatory T cells in scurfy mice.

    Science.gov (United States)

    Skuljec, Jelena; Cabanski, Maciej; Surdziel, Ewa; Lachmann, Nico; Brennig, Sebastian; Pul, Refik; Jirmo, Adan C; Habener, Anika; Visic, Julia; Dalüge, Kathleen; Hennig, Christian; Moritz, Thomas; Happle, Christine; Hansen, Gesine

    2016-07-01

    Foxp3(+) regulatory T (Treg) cells play a pivotal role in maintaining immunological tolerance. Loss-of-function mutations in the Foxp3 gene result in multiorgan inflammation known as immunodysregulation, polyendocrinopathy, enteropathy, X-linked syndrome in humans and scurfy (Sf) disease in mice. While the impact of missing Treg cells on adaptive immune cells is well documented, their role in regulation of myeloid cells remains unclear. Here we report that Sf mice exhibit an altered composition of stem and progenitor cells, characterized by increased numbers of myeloid precursors and higher efficiency of macrophage generation ex vivo. The proportion of monocytes/macrophages in the bone marrow, blood, and spleen was significantly elevated in Sf mice, which was accompanied with tissue-specific monocyte expression of homing receptor and phagocytic activity. Sf mice displayed high levels of M-CSF and other inflammatory cytokines, including monocyte-recruiting chemokines. Adoptive transfer of WT CD4(+) cells and in vivo neutralization of M-CSF normalized frequencies of monocyte subsets and their progenitors and reduced high levels of monocyte-related cytokines in Sf mice, while Treg cell transfer to RAG2(-/-) mice had no effect on myelopoiesis and monocyte/macrophage counts. Our findings illustrate that deregulated myelopoiesis in Sf mice is mainly caused by the inflammatory reaction resulting from the lack of Treg cells.

  15. Hoxb8 conditionally immortalised macrophage lines model inflammatory monocytic cells with important similarity to dendritic cells.

    Science.gov (United States)

    Rosas, Marcela; Osorio, Fabiola; Robinson, Matthew J; Davies, Luke C; Dierkes, Nicola; Jones, Simon A; Reis e Sousa, Caetano; Taylor, Philip R

    2011-02-01

    We have examined the potential to generate bona fide macrophages (MØ) from conditionally immortalised murine bone marrow precursors. MØ can be derived from Hoxb8 conditionally immortalised macrophage precursor cell lines (MØP) using either M-CSF or GM-CSF. When differentiated in GM-CSF (GM-MØP) the resultant cells resemble GM-CSF bone marrow-derived dendritic cells (BMDC) in morphological phenotype, antigen phenotype and functional responses to microbial stimuli. In spite of this high similarity between the two cell types and the ability of GM-MØP to effectively present antigen to a T-cell hybridoma, these cells are comparatively poor at priming the expansion of IFN-γ responses from naïve CD4(+) T cells. The generation of MØP from transgenic or genetically aberrant mice provides an excellent opportunity to study the inflammatory role of GM-MØP, and reduces the need for mouse colonies in many studies. Hence differentiation of conditionally immortalised MØPs in GM-CSF represents a unique in vitro model of inflammatory monocyte-like cells, with important differences from bone marrow-derived dendritic cells, which will facilitate functional studies relating to the many 'sub-phenotypes' of inflammatory monocytes.

  16. Drug induced increases in CNS dopamine alter monocyte, macrophage and T cell functions: implications for HAND

    Science.gov (United States)

    Gaskill, Peter J.; Calderon, Tina M.; Coley, Jacqueline S.; Berman, Joan W.

    2013-01-01

    Central nervous system (CNS) complications resulting from HIV infection remain a major public health problem as individuals live longer due to the success of combined antiretroviral therapy (cART). As many as 70% of HIV infected people have HIV associated neurocognitive disorders (HAND). Many HIV infected individuals abuse drugs, such as cocaine, heroin or methamphetamine, that may be important cofactors in the development of HIV CNS disease. Despite different mechanisms of action, all drugs of abuse increase extracellular dopamine in the CNS. The effects of dopamine on HIV neuropathogenesis are not well understood, and drug induced increases in CNS dopamine may be a common mechanism by which different types of drugs of abuse impact the development of HAND. Monocytes and macrophages are central to HIV infection of the CNS and to HAND. While T cells have not been shown to be a major factor in HIV-associated neuropathogenesis, studies indicate that T cells may play a larger role in the development of HAND in HIV infected drug abusers. Drug induced increases in CNS dopamine may dysregulate functions of, or increase HIV infection in, monocytes, macrophages and T cells in the brain. Thus, characterizing the effects of dopamine on these cells is important for understanding the mechanisms that mediate the development of HAND in drug abusers. PMID:23456305

  17. Monocyte / macrophage inflammatory response pathways to combat Francisella infection: possible therapeutic targets?

    Directory of Open Access Journals (Sweden)

    Devyn D Gillette

    2014-02-01

    Full Text Available Francisella tularensis can bypass and suppress host immune responses, even to the point of manipulating immune cell phenotypes and intercellular inflammatory networks. Strengthening these responses such that immune cells more readily identify and destroy the bacteria is likely to become a viable (and perhaps necessary strategy for combating infections with Francisella, especially given the likelihood of antibiotic resistance in the foreseeable future. Monocytes and macrophages offer a niche wherein Francisella can invade and replicate, resulting in substantially higher bacterial load that can overcome the host. As such, understanding their responses to Francisella may uncover potential avenues of therapy that could promote a lowering of bacterial burden and clearance of infection. These response pathways include Toll-like Receptor 2 (TLR2, the caspase-1 inflammasome, Interferons, NADPH oxidase, Phosphatidylinositide 3-kinase (PI3K and the Ras pathway. In this review we summarize the literature pertaining to the roles of these pathways during Francisella infection, with an emphasis on monocyte / macrophage responses. The therapeutic targeting of one or more such pathways may ultimately become a valuable tool for the treatment of tularemia, and several possibilities are discussed.

  18. Monocyte Subpopulations in Angiogenesis

    Science.gov (United States)

    Dalton, Heather J.; Armaiz-Pena, Guillermo; Gonzalez-Villasana, Vianey; Lopez-Berestein, Gabriel; Bar-Eli, Menashe; Sood, Anil K.

    2014-01-01

    Growing understanding of the role of the tumor microenvironment in angiogenesis has brought monocyte-derived cells into focus. Monocyte subpopulations are an increasingly attractive therapeutic target in many pathologic states, including cancer. Before monocyte-directed therapies can be fully harnessed for clinical use, understanding of monocyte-driven angiogenesis in tissue development and homeostasis, as well as malignancy, is required. Here, we provide an overview of the mechanisms by which monocytic subpopulations contribute to angiogenesis in tissue and tumor development, highlight gaps in our existing knowledge, and discuss opportunities to exploit these cells for clinical benefit. PMID:24556724

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

    Science.gov (United States)

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

    2015-05-01

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

  20. Modulation of the expression of chondroitin sulfate proteoglycan in stimulated human monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Uhlin-Hansen, L.; Eskeland, T.; Kolset, S.O. (Univ. of Tromso (Norway))

    1989-09-05

    Proteoglycan biosynthesis was studied in human monocytes and monocyte-derived macrophages (MDM) after exposure to typical activators of the monocyte/macrophage system: interferon-gamma (IFN-gamma), lipopolysaccharide (LPS), and phorbol 12-myristate 13-acetate (PMA). By morphological examination, both monocytes and MDM were stimulated by these activators. Treatment with IFN-gamma resulted in a slight decrease in the expression of (35S)chondroitin sulfate proteoglycan (CSPG) in both monocytes and MDM, whereas LPS treatment increased the (35S)CSPG expression 1.8 and 2.2 times, respectively. PMA, in contrast, decreased the CSPG expression 0.4 times in monocytes, whereas MDM were stimulated to increase the biosynthesis 1.9 times. An increase in the sulfate density of the chondroitin sulfate chains was evident following differentiation of monocytes into MDM due to the expression of disulfated disaccharide units of the chondroitin sulfate E type (CS-E). However, monocytes exposed to PMA did also express disaccharides of the chondroitin sulfate E type. Furthermore, the expression of CS-E in MDM was increased 2 times following PMA treatment. An inactive phorbol ester, phorbol 12,13-diacetate, did not affect the expression of CS-E in either monocytes or MDM when compared with control cultures, suggesting that protein kinase C-dependent signal pathways may be involved in the regulation of sulfation of CSPG. Exposure to LPS or IFN-gamma did not lead to any changes in the sulfation of the chondroitin sulfate chains.

  1. Dysfunctional CFTR alters the bactericidal activity of human macrophages against Pseudomonas aeruginosa.

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    Paola Del Porto

    Full Text Available Chronic inflammation of the lung, as a consequence of persistent bacterial infections by several opportunistic pathogens represents the main cause of mortality and morbidity in cystic fibrosis (CF patients. Mechanisms leading to increased susceptibility to bacterial infections in CF are not completely known, although the involvement of cystic fibrosis transmembrane conductance regulator (CFTR in microbicidal functions of macrophages is emerging. Tissue macrophages differentiate in situ from infiltrating monocytes, additionally, mature macrophages from different tissues, although having a number of common activities, exhibit variation in some molecular and cellular functions. In order to highlight possible intrinsic macrophage defects due to CFTR dysfunction, we have focused our attention on in vitro differentiated macrophages from human peripheral blood monocytes. Here we report on the contribution of CFTR in the bactericidal activity against Pseudomonas aeruginosa of monocyte derived human macrophages. At first, by real time PCR, immunofluorescence and patch clamp recordings we demonstrated that CFTR is expressed and is mainly localized to surface plasma membranes of human monocyte derived macrophages (MDM where it acts as a cAMP-dependent chloride channel. Next, we evaluated the bactericidal activity of P. aeruginosa infected macrophages from healthy donors and CF patients by antibiotic protection assays. Our results demonstrate that control and CF macrophages do not differ in the phagocytic activity when infected with P. aeruginosa. Rather, although a reduction of intracellular live bacteria was detected in both non-CF and CF cells, the percentage of surviving bacteria was significantly higher in CF cells. These findings further support the role of CFTR in the fundamental functions of innate immune cells including eradication of bacterial infections by macrophages.

  2. Activation of farnesoid X receptor downregulates monocyte chemoattractant protein-1 in murine macrophage

    Energy Technology Data Exchange (ETDEWEB)

    Li, Liangpeng; Zhang, Qian; Peng, Jiahe; Jiang, Chanjui; Zhang, Yan; Shen, Lili; Dong, Jinyu; Wang, Yongchao; Jiang, Yu, E-mail: yujiang0207@163.com

    2015-11-27

    Farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily, which plays important roles in bile acids/lipid homeostasis and inflammation. Monocyte chemoattractant protein-1 (MCP-1) contributes to macrophage infiltration into body tissues during inflammation. Here we investigated whether FXR can regulate MCP-1 expression in murine macrophage. FXR activation down regulate MCP-1 mRNA and protein levels in ANA-1 and Raw264.7 cells. Luciferase reporter assay, Gel shift and Chromatin immunoprecipitation assays have revealed that the activated FXR bind to the FXR element located in −738 bp ∼  −723 bp in MCP-1 promoter. These results suggested that FXR may serve as a novel target for regulating MCP-1 levels for the inflammation related diseases therapies. - Highlights: • FXR is expressed in murine macrophage cell line. • FXR down regulates MCP-1 expression. • FXR binds to the DR4 in MCP-1 promoter.

  3. Effect of monocyte chemoattractant protein-1 on chemotactic gene expression by macrophage cell line U937

    Institute of Scientific and Technical Information of China (English)

    BIAN Guang-xing; GUO Bao-yu; MIAO Hong; QIU Lei; CAO Dong-mei; DAO Shu-yan; ZHANG Ran

    2004-01-01

    Objective: To study the chemotactic superfamily genes expression profiling of macrophage line U937 treated with monocyte chemoattractant protein-1 (MCP-1) using gene chip technique. Methods: Total RNA from macrophage line U937 (as control) and U937 with MCP-1 was extracted, made reverse transcript to cDNA and tested with gene expression chip HO2 human. Results: Some chemotactic-related gene expressions were changed in all analyzed genes. Regulated upon activation, normal T cell expressed and secreted (RANTES) was up-regulated over 2-fold and 7 chemotactic-related genes (CCR2, CCR5, CCL16, GROβ, GROγ, IL-8 and granulocyte chemotactic protein 2) were down-regulated over 2-fold inMCP-1 treated U937 cells at mRNA level. Conclusion: MCP-1 can influence some chemokines and receptors expression in macrophage in vitro, in which MCP-1 mainly down-regulates the chemotactic genes expression of those influencing neutrophilic granulocyte (GROβ, GROγ, IL-8 and granulocyte chemotactic protein 2). Another novel finding is that it can also down-regulate the mRNA level of CCR5, which plays a critical role in many disorders and illnesses.

  4. Maturation Phenotype of Peripheral Blood Monocyte/Macrophage After Stimulation with Lipopolysaccharides in Irritable Bowel Syndrome

    Science.gov (United States)

    Rodríguez-Fandiño, Oscar A; Hernández-Ruiz, Joselín; López-Vidal, Yolanda; Charúa-Guindic, Luis; Escobedo, Galileo; Schmulson, Max J

    2017-01-01

    Background/Aims Abnormal immune regulation and increased intestinal permeability augmenting the passage of bacterial molecules that can activate immune cells, such as monocytes/macrophages, have been reported in irritable bowel syndrome (IBS). The aim was to compare the maturation phenotype of monocytes/macrophages (CD14+) from IBS patients and controls in the presence or absence of Escherichia coli lipopolysaccharides (LPS), in vitro. Methods Mononuclear cells were isolated from peripheral blood of 20 Rome II-IBS patients and 19 controls and cultured with or without LPS for 72 hours. The maturation phenotype was examined by flow cytometry as follows: M1-Early (CD11c+CD206−), M2-Advanced (CD11c−CD206+CX3CR1+); expression of membrane markers was reported as mean fluorescence intensity (MFI). The Mann-Whitney test was used and significance was set at P < 0.05. Results In CD14+ cells, CD11c expression decreased with vs without LPS both in IBS (MFI: 8766.0 ± 730.2 vs 12 920.0 ± 949.2, P < 0.001) and controls (8233.0 ± 613.9 vs 13 750.0 ± 743.3, P < 0.001). M1-Early cells without LPS, showed lower CD11c expression in IBS than controls (MFI: 11 540.0 ± 537.5 vs 13 860.0 ± 893.7, P = 0.040), while both groups showed less CD11c in response to LPS (P < 0.01). Furthermore, the percentage of “Intermediate” (CD11c+CD206+CX3CR1+) cells without LPS, was higher in IBS than controls (IBS = 9.5 ± 1.5% vs C = 4.9 ± 1.4%, P < 0.001). Finally, fractalkine receptor (CX3CR1) expression on M2-Advanced cells was increased when treated with LPS in controls but not in IBS (P < 0.001). Conclusions The initial phase of monocyte/macrophage maturation appears to be more advanced in IBS compared to controls. However, the decreased CX3CR1 in patients with IBS, compared to controls, when stimulated with LPS suggests a state of immune activation in IBS. PMID:28044051

  5. Strong inhibition of TNF-alpha production and inhibition of IL-8 and COX-2 mRNA expression in monocyte-derived macrophages by RWJ 67657, a p38 mitogen-activated protein kinase (MAPK) inhibitor

    NARCIS (Netherlands)

    Westra, J; Doornbos-van der Meer, B; de Boer, Peter; van Leeuwen, MA; van Rijswijk, Martin; Limburg, PC

    2004-01-01

    In inflammatory processes, the p38 mitogen-activated protein kinase ( MAPK) signal transduction route regulates production and expression of cytokines and other inflammatory mediators. Tumor necrosis factor alpha (TNF-alpha) is a pivotal cytokine in rheumatoid arthritis and its production in macroph

  6. Stimulation of PBMC and Monocyte-derived-Macrophages via Toll-Like Receptor (TLRs Activates Innate Immune Pathways in HIV-Infected Patients on Virally-Suppressive Combination Antiretroviral Therapy (cART

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    Esther Merlini

    2016-12-01

    Full Text Available In HIV-infected cART-treated patients, immune activation and microbial translocation persist and associate with inadequate CD4 recovery and morbidity/mortality. We analyzed whether alterations in the TLR pathway could be responsible for the immune hyper-activation seen in these patients.PBMC/MDM of 28 HIV+ untreated and 35 cART treated patients with HIV-RNA<40cp/mL (20 Full Responders: CD4≥350; 15 Immunological Non Responders:CD4<350 as well as of 16 healthy controls were stimulated with a panel of TLR agonists. We measured: CD4/CD8/CD14/CD38/HLA-DR/Ki67/AnnexV/CD69/TLR4/8 (Flow Cytometry; PBMC expression of 84 TLR pathway genes (qPCR; PBMC/MDM cytokine release (Multiplex; plasma LPS/sCD14 (LAL/ELISA. PBMC/MDM from cART patients responded weakly to LPS stimulation but released high amounts of pro-inflammatory cytokines. MDM from these patients were characterized by a reduced expression of HLA-DR+MDM and failed to expand activated HLA-DR+CD38+ T-lymphocytes. PBMC/MDM from cART patients responded more robustly to ssRNA stimulation; this resulted in a significant expansion of activated CD38+CD8 and the release of amounts of pro-inflammatory cytokines comparable to those seen in untreated viremic patients. Despite greater constitutive TLR pathway gene expression, PBMC from Immunological Non Responders seemed to up-regulate only type I IFN genes following TLR stimulation, whereas PBMC from Full Responders showed a broader response. Systemic exposure to microbial antigens drives immune activation during cART by triggering TLRs. Bacterial stimulation modifies MDM function/pro-inflammatory profile in cART patients without affecting T-lymphocytes; this suggests translocating bacteria as selective stimulus to chronic innate activation during cART. High constitutive TLR activation is seen in patients lacking CD4 recovery, suggesting an exhausted immune milieu, anergic to further antigen encounters.

  7. [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.

  8. Modulation of neutrophil and monocyte function by recombinant human granulocyte macrophage colony-stimulating factor in patients with lymphoma

    DEFF Research Database (Denmark)

    Kharazmi, A; Nielsen, H; Hovgaard, D;

    1991-01-01

    Granulocyte macrophage colony-stimulating factor (GM-CSF) has been shown to inhibit the chemotaxis and enhance the oxidative burst response of human neutrophils in vitro. The present study describes the effect of recombinant GM-CSF on the neutrophil and monocyte function in patients with lymphoma...

  9. Soluble CD163, a product of monocyte/macrophage activation, is inversely associated with haemoglobin levels in placental malaria.

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    Caroline Lin Lin Chua

    Full Text Available In Plasmodium falciparum malaria, activation of monocytes and macrophages (monocytes/macrophages can result in the production of various inflammatory mediators that contribute to immunopathology. Soluble CD163 (sCD163 is a specific marker of monocyte/macrophage activation typically found at increased levels during various inflammatory conditions and can be associated with poor clinical outcomes. To better understand the relationships between levels of sCD163 and clinical parameters in women with placental malaria, we measured plasma sCD163 levels in maternal peripheral and placental blood compartments at delivery and determined their correlations with birth weight and maternal haemoglobin concentrations. sCD163 levels were negatively correlated with birth weight only in the placental compartment (r = -0.145, p = 0.03 and were inversely correlated with maternal haemoglobin concentrations, both in peripheral blood (r = -0.238, p = 0.0004 and in placental blood (r = -0.259, p = 0.0001. These inverse relationships suggest a potential role for monocyte/macrophage activation in the pathogenesis of malaria in pregnancy, particularly in relation to malaria-associated anaemia.

  10. Arteriogenesis depends on circulating monocytes and macrophage accumulation and is severely depressed in op/op mice

    NARCIS (Netherlands)

    C.E. Bergmann; I.E. Hoefer; B. Meder; H. Roth; N. van Royen; S.M. Breit; M.M. Jost; S. Aharinejad; S. Hartmann; I.R. Buschmann

    2006-01-01

    It has been suggested that monocytes/macrophages represent the pivotal cell type during early adaptive growth of pre-existent arterial anastomoses toward functional collateral arteries (arteriogenesis) upon arterial occlusion. This hypothesis was supported by previous studies providing evidence that

  11. Mature dendritic cells derived from human monocytes within 48 hours: a novel strategy for dendritic cell differentiation from blood precursors.

    Science.gov (United States)

    Dauer, Marc; Obermaier, Bianca; Herten, Jan; Haerle, Carola; Pohl, Katrin; Rothenfusser, Simon; Schnurr, Max; Endres, Stefan; Eigler, Andreas

    2003-04-15

    It is widely believed that generation of mature dendritic cells (DCs) with full T cell stimulatory capacity from human monocytes in vitro requires 5-7 days of differentiation with GM-CSF and IL-4, followed by 2-3 days of activation. Here, we report a new strategy for differentiation and maturation of monocyte-derived DCs within only 48 h of in vitro culture. Monocytes acquire immature DC characteristics by day 2 of culture with GM-CSF and IL-4; they down-regulate CD14, increase dextran uptake, and respond to the inflammatory chemokine macrophage inflammatory protein-1alpha. To accelerate DC development and maturation, monocytes were incubated for 24 h with GM-CSF and IL-4, followed by activation with proinflammatory mediators for another 24 h (FastDC). FastDC expressed mature DC surface markers as well as chemokine receptor 7 and secreted IL-12 (p70) upon CD40 ligation in the presence of IFN-gamma. The increase in intracellular calcium in response to 6Ckine showed that chemokine receptor 7 expression was functional. When FastDC were compared with mature monocyte-derived DCs generated by a standard 7-day protocol, they were equally potent in inducing Ag-specific T cell proliferation and IFN-gamma production as well as in priming autologous naive T cells using tetanus toxoid as a model Ag. These findings indicate that FastDC are as effective as monocyte-derived DCs in stimulating primary, Ag-specific, Th 1-type immune responses. Generation of FastDC not only reduces labor, cost, and time required for in vitro DC development, but may also represent a model more closely resembling DC differentiation from monocytes in vivo.

  12. Human Macrophage Response to L. (Viannia) panamensis: Microarray Evidence for an Early Inflammatory Response

    Science.gov (United States)

    Rojas, Ricardo; Ettinger, Nicholas A.; Tikhonova, Irina; Alexander, Neal D.; Valderrama, Liliana; Hager, Janet; Wilson, Mary E.; Lin, Aiping; Zhao, Hongyu; Saravia, Nancy G.; McMahon-Pratt, Diane

    2012-01-01

    Background Previous findings indicate that susceptibility to Leishmania (Viannia) panamensis infection of monocyte-derived macrophages from patients and asymptomatically infected individuals were associated with the adaptive immune response and clinical outcome. Methodology/Principal Findings To understand the basis for this difference we examined differential gene expression of human monocyte-derived macrophages following exposure to L. (V.) panamensis. Gene activation profiles were determined using macrophages from healthy volunteers cultured with or without stationary phase promastigotes of L. (V.) panamensis. Significant changes in expression (>1.5-fold change; p<0.05; up- or down-regulated) were identified at 0.5, 4 and 24 hours. mRNA abundance profiles varied over time, with the highest level of activation occurring at earlier time points (0.5 and 4 hrs). In contrast to observations for other Leishmania species, most significantly changed mRNAs were up- rather than down-regulated, especially at early time points. Up-regulated transcripts over the first 24 hours belonged to pathways involving eicosanoid metabolism, oxidative stress, activation of PKC through G protein coupled receptors, or mechanism of gene regulation by peroxisome proliferators via PPARα. Additionally, a marked activation of Toll-receptor mediated pathways was observed. Comparison with published microarray data from macrophages infected with L. (Leishmania) chagasi indicate differences in the regulation of genes involved in signaling, motility and the immune response. Conclusions Results show that the early (0.5 to 24 hours) human monocyte-derived macrophage response to L. (Viannia) panamensis is not quiescent, in contrast to published reports examining later response times (48–96 hours). Early macrophage responses are important for the developing cellular response at the site of infection. The kinetics and the mRNA abundance profiles induced by L. (Viannia) panamensis illustrate the

  13. Effects of ischemia on lung macrophages.

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    Aigul Moldobaeva

    Full Text Available Angiogenesis after pulmonary ischemia is initiated by reactive O(2 species and is dependent on CXC chemokine growth factors, and its magnitude is correlated with the number of lavaged macrophages. After complete obstruction of the left pulmonary artery in mice, the left lung is isolated from the peripheral circulation until 5-7 days later, when a new systemic vasculature invades the lung parenchyma. Consequently, this model offers a unique opportunity to study the differentiation and/or proliferation of monocyte-derived cells within the lung. In this study, we questioned whether macrophage subpopulations were differentially expressed and which subset contributed to growth factor release. We characterized the change in number of all macrophages (MHCII(int, CD11C+, alveolar macrophages (MHCII(int, CD11C+, CD11B- and mature lung macrophages (MHCII(int, CD11C+, CD11B+ in left lungs from mice immediately (0 h or 24 h after left pulmonary artery ligation (LPAL. In left lung homogenates, only lung macrophages increased 24 h after LPAL (vs. 0 h; p<0.05. No changes in proliferation were seen in any subset by PCNA expression (0 h vs. 24 h lungs. When the number of monocytic cells was reduced with clodronate liposomes, systemic blood flow to the left lung 14 days after LPAL decreased by 42% (p<0.01 compared to vehicle controls. Furthermore, when alveolar macrophages and lung macrophages were sorted and studied in vitro, only lung macrophages secreted the chemokine MIP-2α (ELISA. These data suggest that ischemic stress within the lung contributes to the differentiation of immature monocytes to lung macrophages within the first 24 h after LPAL. Lung macrophages but not alveolar macrophages increase and secrete the proangiogenic chemokine MIP-2α. Overall, an increase in the number of lung macrophages appears to be critical for neovascularization in the lung, since clodronate treatment decreased their number and attenuated functional angiogenesis.

  14. How mouse macrophages sense what is going on

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    Klaus eLey

    2016-06-01

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

  15. Possible impact of microglial cells and the monocyte-macrophage system on suicidal behavior.

    Science.gov (United States)

    Steiner, Johann; Gos, Tomasz; Bogerts, Bernhard; Bielau, Hendrik; Drexhage, Hemmo A; Bernstein, Hans-Gert

    2013-11-01

    Immune dysfunction, including monocytosis, increased blood levels of interleukin-1 (IL-1), interleukin-6 (IL- 6) and tumor necrosis factor-alpha (TNF-alpha), as well as an increased microglial density in certain brain areas, have been described in schizophrenia and depression. Interestingly, similar immune alterations have been observed in suicide patients regardless of their underlying psychiatric diagnosis. This review summarizes relevant data from previous studies that have examined peripheral blood, cerebrospinal fluid and human brains (using postmortem histology and in vivo positron emission tomography) to investigate immune mechanisms in suicidal patients. We discuss whether the observed findings indicate that microgliosis and monocyte-macrophage system activation may be a useful marker of disease acuity/severity or whether they instead indicate a distinct neurobiology of suicide. Notably, pathophysiological mechanisms could change during the long-term course of psychiatric diseases. Therefore, different patterns of immune activation may be observed when comparing newly diseased patients with those who are chronically ill.

  16. Monocyte/macrophage-derived soluble CD163: A novel biomarker in multiple myeloma

    DEFF Research Database (Denmark)

    Andersen, Morten Nørgaard; Abildgaard, Niels; Maniecki, Maciej B

    2014-01-01

    fluids (soluble CD163, sCD163). In this study, we examined serum sCD163 as a biomarker in patients with newly diagnosed multiple myeloma. METHODS: Peripheral blood (n = 104) and bone marrow (n = 17) levels of sCD163 were measured using an enzyme-linked immunosorbent assay. RESULTS: At diagnosis, high s......CD163 was associated with higher stage according to the International Staging System (ISS) and with other known prognostic factors in multiple myeloma (creatinine, C-reactive protein, and beta-2 microglobulin). Soluble CD163 decreased upon high-dose treatment, and in a multivariate survival analysis...... in bone marrow samples than in the matched blood samples, which indicate a localized production of sCD163 within the bone marrow microenvironment. CONCLUSIONS: Soluble CD163 was found to be a prognostic marker in patients with multiple myeloma. This may indicate that macrophages and/or monocytes have...

  17. Flagella from five Cronobacter species induce pro-inflammatory cytokines in macrophage derivatives from human monocytes.

    Directory of Open Access Journals (Sweden)

    Ariadnna Cruz-Córdova

    Full Text Available Cronobacter spp. are opportunistic pathogens linked to lie-threatening infections in neonates and contaminated powdered infant formula that has been epidemiologically associated with these cases. Clinical symptoms of Cronobacter include necrotizing enterocolitis, bacteremia, and meningitis. Flagella from C. sakazakii are involved in biofilm formation and its adhesion to epithelial cells. We investigated the role of flagella from C. sakazakii ST1 and ST4, C. malonaticus, C. muytjensii, C. turicensis and C. dublinensis during the activation of cytokines (IL-8, TNF-α, and IL-10 in macrophage derivatives from human monocytes, which has not been extensively studied. The production and identity of flagella from the five Cronobacter species were visualized and recognized with anti-flagella antibodies by immunogold labeling through transmission electron microscopy. Purified flagella were dissociated into monomers in 12% SDS-PAGE Coomassie blue-stained gels showing a band of ∼28 kDa and, in addition, mass spectrometry revealed the presence of several peptides that correspond to flagellin. Flagella (100 ng induced the release of IL-8 (3314-6025 pg/ml, TNF-α (39-359 pg/ml, and IL-10 (2-96 pg/ml, in macrophage isolates from human monocytes and similar results were obtained when flagella were dissociated into monomers. Inhibition assays using three dilutions of anti-flagella antibodies (1∶10, 1∶100, and 1∶200 suppressed the secretion of IL-8, TNF-α, and IL-10 between 95-100% using 100 ng of protein. A transfection assay using 293-hTLR5 cells showed IL-8 release of 197 pg/ml and suppression in the secretion of IL-8 when anti-hTLR5-IgA antibodies were used at different concentrations. These observations suggest that flagella and flagellin are involved in an inflammatory response dependent on TLR5 recognition, which could contribute to the pathogenesis of the bacteria.

  18. Flagella from Five Cronobacter Species Induce Pro-Inflammatory Cytokines in Macrophage Derivatives from Human Monocytes

    Science.gov (United States)

    Cruz-Córdova, Ariadnna; Rocha-Ramírez, Luz M.; Ochoa, Sara A.; Gónzalez-Pedrajo, Bertha; Espinosa, Norma; Eslava, Carlos; Hernández-Chiñas, Ulises; Mendoza-Hernández, Guillermo; Rodríguez-Leviz, Alejandra; Valencia-Mayoral, Pedro; Sadowinski-Pine, Stanislaw; Hernández-Castro, Rigoberto; Estrada-García, Iris; Muñoz-Hernández, Onofre; Rosas, Irma; Xicohtencatl-Cortes, Juan

    2012-01-01

    Cronobacter spp. are opportunistic pathogens linked to lie-threatening infections in neonates and contaminated powdered infant formula that has been epidemiologically associated with these cases. Clinical symptoms of Cronobacter include necrotizing enterocolitis, bacteremia, and meningitis. Flagella from C. sakazakii are involved in biofilm formation and its adhesion to epithelial cells. We investigated the role of flagella from C. sakazakii ST1 and ST4, C. malonaticus, C. muytjensii, C. turicensis and C. dublinensis during the activation of cytokines (IL-8, TNF-α, and IL-10) in macrophage derivatives from human monocytes, which has not been extensively studied. The production and identity of flagella from the five Cronobacter species were visualized and recognized with anti-flagella antibodies by immunogold labeling through transmission electron microscopy. Purified flagella were dissociated into monomers in 12% SDS-PAGE Coomassie blue-stained gels showing a band of ∼28 kDa and, in addition, mass spectrometry revealed the presence of several peptides that correspond to flagellin. Flagella (100 ng) induced the release of IL-8 (3314–6025 pg/ml), TNF-α (39–359 pg/ml), and IL-10 (2–96 pg/ml), in macrophage isolates from human monocytes and similar results were obtained when flagella were dissociated into monomers. Inhibition assays using three dilutions of anti-flagella antibodies (1∶10, 1∶100, and 1∶200) suppressed the secretion of IL-8, TNF-α, and IL-10 between 95–100% using 100 ng of protein. A transfection assay using 293-hTLR5 cells showed IL-8 release of 197 pg/ml and suppression in the secretion of IL-8 when anti-hTLR5-IgA antibodies were used at different concentrations. These observations suggest that flagella and flagellin are involved in an inflammatory response dependent on TLR5 recognition, which could contribute to the pathogenesis of the bacteria. PMID:23284883

  19. Dynamical optical imaging monocytes/macrophages migration and activation in contact hypersensitivity (Conference Presentation)

    Science.gov (United States)

    Zhang, Zhihong

    2017-02-01

    Inflammatory monocytes/macrophages (Mon/Mφ) play an important role in cutaneous allergic inflammation. However, their migration and activation in dermatitis and how they accelerate the inflammatory reaction are largely unknown. Optical molecular imaging is the most promising tool for investigating the function and motility of immune cells in vivo. We have developed a multi-scale optical imaging approach to evaluate the spatio-temporal dynamic behavior and properties of immune cells from the whole field of organs to the cellular level at the inflammatory site in delayed type hypersensitivity reaction. Here, we developed some multi-color labeling mouse models based on the endogenous labeling with fluorescent proteins and the exogenous labeling with fluorescent dyes. We investigated the cell movement, cell interaction and function of immunocytes (e.g. Mon/Mφ, DC, T cells and neutrophils) in the skin allergy inflammation (e.g., contact hypersensitivity) by using intravital microscopy. The long-term imaging data showed that after inflammatory Mon/Mφ transendothelial migration in dermis, they migrating in interstitial space of dermis. Depletion of blood monocyte with clodronate liposome extremely reduced the inflammatory reaction. Our finding provided further insight into inflammatory Mon/Mφ mediating the inflammatory cascade through functional migration in allergic contact dermatitis.

  20. Generation of novel bone forming cells (monoosteophils from the cathelicidin-derived peptide LL-37 treated monocytes.

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    Zhifang Zhang

    Full Text Available BACKGROUND: Bone generation and maintenance involve osteoblasts, osteoclasts, and osteocytes which originate from unique precursors and rely on key growth factors for differentiation. However, an incomplete understanding of bone forming cells during wound healing has led to an unfilled clinical need such as nonunion of bone fractures. Since circulating monocytes are often recruited to sites of injury and may differentiate into various cell types including osteoclasts, we investigated the possibility that circulating monocytes in the context of tissue injury may also contribute to bone repair. In particular, we hypothesized that LL-37 (produced from hCAP-18, cathelicidin, which recruits circulating monocytes during injury, may play a role in bone repair. METHODS AND FINDINGS: Treatment of monocytes from blood with LL-37 for 6 days resulted in their differentiation to large adherent cells. Growth of LL-37-differentiated monocytes on osteologic discs reveals bone-like nodule formation by scanning electron microscopy (SEM. In vivo transplantation studies in NOD/SCID mice show that LL-37-differentiated monocytes form bone-like structures similar to endochondral bone formation. Importantly, LL-37-differentiated monocytes are distinct from conventional monocyte-derived osteoclasts, macrophages, and dendritic cells and do not express markers of the mesenchymal stem cells (MSC lineage, distinguishing them from the conventional precursors of osteoblasts. Furthermore, LL-37 differentiated monocytes express intracellular proteins of both the osteoblast and osteoclast lineage including osteocalcin (OC, osteonectin (ON, bone sialoprotein II (BSP II, osteopontin (OP, RANK, RANKL, MMP-9, tartrate resistant acid phosphatase (TRAP, and cathepsin K (CK. CONCLUSION: Blood derived monocytes treated with LL-37 can be differentiated into a novel bone forming cell that functions both in vitro and in vivo. We propose the name monoosteophil to indicate their monocyte

  1. Prostaglandin E2 suppresses beta1-integrin expression via E-prostanoid receptor in human monocytes/macrophages.

    Science.gov (United States)

    Hasegawa, Shunji; Ichiyama, Takashi; Kohno, Fumitaka; Korenaga, Yuno; Ohsaki, Ayami; Hirano, Reiji; Haneda, Yasuhiro; Fukano, Reiji; Furukawa, Susumu

    2010-01-01

    Beta1-integrins mediate cell attachment to different extracellular matrix proteins, intracellular proteins, and intercellular adhesions. Recently, it has been reported that prostaglandin E2 (PGE2) has anti-inflammatory properties such as inhibition of the expression of adhesion molecules or production of chemokines. However, the effect of PGE2 on the expression of beta1-integrin remains unknown. In this study, we investigated the effects of PGE2 on the expression of beta1-integrin in the human monocytic cell line THP-1 and in CD14+ monocytes/macrophages in human peripheral blood. For this, we examined the role of four subtypes of PGE2 receptors and E-prostanoid (EP) receptors on PGE2-mediated inhibition. We found that PGE2 significantly inhibited the expression of beta1-integrin, mainly through EP4 receptors in THP-1 cells and CD14+ monocytes/macrophages in human peripheral blood. We suggest that PGE2 has anti-inflammatory effects, leading to the inhibited expression of beta1-integrin in human monocytes/macrophages, and that the EP4 receptor may play an important role in PGE2-mediated inhibition. Copyright (c) 2010 Elsevier Inc. All rights reserved.

  2. Monocyte/macrophage-derived soluble CD163: a novel biomarker in multiple myeloma.

    Science.gov (United States)

    Andersen, Morten N; Abildgaard, Niels; Maniecki, Maciej B; Møller, Holger J; Andersen, Niels F

    2014-07-01

    Macrophages play an important role in cancer by suppression of adaptive immunity and promotion of angiogenesis and metastasis. Tumor-associated macrophages strongly express the hemoglobin scavenger receptor CD163, which can also be found as a soluble protein in serum and other body fluids (soluble CD163, sCD163). In this study, we examined serum sCD163 as a biomarker in patients with newly diagnosed multiple myeloma. Peripheral blood (n = 104) and bone marrow (n = 17) levels of sCD163 were measured using an enzyme-linked immunosorbent assay. At diagnosis, high sCD163 was associated with higher stage according to the International Staging System (ISS) and with other known prognostic factors in multiple myeloma (creatinine, C-reactive protein, and beta-2 microglobulin). Soluble CD163 decreased upon high-dose treatment, and in a multivariate survival analysis including the covariates treatment modality and age at diagnosis, higher levels of sCD163 were associated with poor outcome (HR = 1.82; P = 0.010). The prognostic significance of sCD163 was lost when including ISS stage in the model (HR = 1.51; P = 0.085). Soluble CD163 values were significantly higher in bone marrow samples than in the matched blood samples, which indicate a localized production of sCD163 within the bone marrow microenvironment. Soluble CD163 was found to be a prognostic marker in patients with multiple myeloma. This may indicate that macrophages and/or monocytes have an important role in the bone marrow microenvironment of myeloma patients, supporting myeloma cell proliferation and survival. We propose the serum sCD163 value 1.8 mg/L as a cutoff concentration for survival analysis in patients with multiple myeloma, which should be validated in future studies. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. In Lysinuric Protein Intolerance system y+L activity is defective in monocytes and in GM-CSF-differentiated macrophages

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    Mariani Francesca

    2010-11-01

    Full Text Available Abstract Background In the recessive aminoaciduria Lysinuric Protein Intolerance (LPI, mutations of SLC7A7/y+LAT1 impair system y+L transport activity for cationic amino acids. A severe complication of LPI is a form of Pulmonary Alveolar Proteinosis (PAP, in which alveolar spaces are filled with lipoproteinaceous material because of the impaired surfactant clearance by resident macrophages. The pathogenesis of LPI-associated PAP remains still obscure. The present study investigates for the first time the expression and function of y+LAT1 in monocytes and macrophages isolated from a patient affected by LPI-associated PAP. A comparison with mesenchymal cells from the same subject has been also performed. Methods Monocytes from peripheral blood were isolated from a 21-year-old patient with LPI. Alveolar macrophages and fibroblastic-like mesenchymal cells were obtained from a whole lung lavage (WLL performed on the same patient. System y+L activity was determined measuring the 1-min uptake of [3H]-arginine under discriminating conditions. Gene expression was evaluated through qRT-PCR. Results We have found that: 1 system y+L activity is markedly lowered in monocytes and alveolar macrophages from the LPI patient, because of the prevailing expression of SLC7A7/y+LAT1 in these cells; 2 on the contrary, fibroblasts isolated from the same patient do not display the transport defect due to compensation by the SLC7A6/y+LAT2 isoform; 3 in both normal and LPI monocytes, GM-CSF induces the expression of SLC7A7, suggesting that the gene is a target of the cytokine; 4 GM-CSF-induced differentiation of LPI monocytes is comparable to that of normal cells, demonstrating that GM-CSF signalling is unaltered; 5 general and respiratory conditions of the patient, along with PAP-associated parameters, markedly improved after GM-CSF therapy through aerosolization. Conclusions Monocytes and macrophages, but not fibroblasts, derived from a LPI patient clearly display the

  4. Monocyte-macrophage membrane possesses free radicals scavenging activity: stimulation by polyphenols or by paraoxonase 1 (PON1).

    Science.gov (United States)

    Rosenblat, M; Elias, A; Volkova, N; Aviram, M

    2013-04-01

    In the current study, we analysed free radicals scavenging activity of monocytes-macrophages in the absence or presence of antioxidants such as polyphenols or paraoxonase 1 (PON1). THP-1 human monocytic cell line, murine J774A.1 macrophages, as well as human primary monocytes have the capability to scavenge free radicals, as measured by the 1-diphenyl-2-picryl-hydrazyl (DPPH) assay. This effect (which could be attributed to the cell's membrane) was cell number and incubation time dependent. Upon incubation of J774A.1 macrophages with acetylated LDL (Ac-LDL), with VLDL, or with the radical generator, AAPH, the cells' lipid peroxides content, and paraoxonase 2 (PON2) activity were significantly increased. While non-treated cells decreased DPPH absorbance by 65%, the Ac-LDL-, VLDL- or AAPH-treated cells, decreased it by only 33%, 30%, or 45%, respectively. We next analysed the effect of J774A.1 macrophage enrichment with antioxidants, such as polyphenols or PON1 on the cells' free radicals scavenging activity. Non-treated cells decreased DPPH absorbance by 50%, whereas vitamin E-, punicalagin- or PJ-treated cells significantly further decreased it, by 75%. Similarly, in PON1-treated cells DPPH absorbance was further decreased by 63%, in association with 23% increment in PON1 catalytic activity. In cells under oxidative stress [treated with AAPH-, or with oxidized LDL], PON1 activity was decreased by 31% or 40%, as compared to the activity observed in PON1 incubated with non-treated cells. We conclude that monocytes-macrophages possess free radicals scavenging activity, which is decreased under atherogenic conditions, and increased upon cell enrichment with potent antioxidants such as nutritional polyphenols, or PON1.

  5. The monocytic lineage specific soluble CD163 is a plasma marker of coronary atherosclerosis

    DEFF Research Database (Denmark)

    Aristoteli, Lina Panayiota; Møller, Holger Jon; Bailey, Brian

    2006-01-01

    BACKGROUND: CD163 is a monocyte-macrophage lineage specific scavenger receptor that mediates the uptake and clearance of haptoglobin-haemoglobin complexes, and soluble CD163 (sCD163) is also present in plasma. As atherosclerosis involves infiltration by monocyte-derived macrophages, we investigated...... whether sCD163 may act as a marker of coronary atherosclerosis (CAD). METHODS AND RESULTS: Clinical features were identified and plasma was collected from 147 consecutive patients presenting for coronary angiography. Patients were classified as having CAD+, or being free of CAD- haemodynamically...... significant (>50% luminal diameter) coronary stenoses in one or more major coronary arteries (1, 2 or 3 vessel disease), and sCD163 concentration was measured by ELISA. Plasma sCD163 was non-parametrically distributed, being significantly higher in CAD+ patients (median 2.47 mg/L, 25th-75th percentile, 1...

  6. Dimethyphenylpiperazinium, a nicotinic receptor agonist, downregulates inflammation in monocytes/macrophages through PI3K and PLC chronic activation.

    Science.gov (United States)

    Blanchet, Marie-Renée; Israël-Assayag, Evelyne; Daleau, Pascal; Beaulieu, Marie-Josée; Cormier, Yvon

    2006-10-01

    Activation of nicotinic acetylcholine receptors (nAChRs) on inflammatory cells induces anti-inflammatory effects. The intracellular mechanisms that regulate this effect are still poorly understood. In neuronal cells, nAChRs are associated with phosphatidylinositol 3-kinase (PI3K). This enzyme, which can activate phospholipase C (PLC), is also present in monocytes. The aim of this study was to assess the role of these proteins in the signaling pathways involved in the anti-inflammatory effect of dimethylphenylpiperazinium (DMPP), a synthetic nAChR agonist, on monocytes and macrophages. The results indicate that PI3K is associated with alpha3, -4, and -5 nAChR subunits in monocytes. The PI3K inhibitors wortmannin and LY294002 abrogated the inhibitory effect of DMPP on LPS-induced TNF release by monocytes. Treatment with DMPP for 24 and 48 h provoked a mild PLC phosphorylation, which was blocked by the nAChR antagonist mecamylamine and reversed by PI3K inhibitors. Treatment of monocytes and alveolar macrophages with DMPP reduced the inositol 1,4,5-trisphosphate (IP3)-dependent intracellular calcium mobilization induced by platelet-activating factor (PAF), an effect that was reversed by mecamylamine in alveolar macrophages. DMPP did not have any effect on PAF receptor expression. DMPP also inhibited the thapsigargin-provoked calcium release, indicating that the endoplasmic reticulum calcium stores might be depleted by treatment with the nAChR agonist. Taken together, these results suggest that PI3K and PLC activation is involved in the anti-inflammatory effect of DMPP. PLC limited, but constant activation could induce, the depletion of intracellular calcium stores, leading to the anti-inflammatory effect of DMPP.

  7. Regulation of MMP-9 by a WIN-binding site in the monocyte-macrophage system independent from cannabinoid receptors.

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    Svantje Tauber

    Full Text Available The cannabinoid system is known to be involved in the regulation of inflammatory processes. Therefore, drugs targeting cannabinoid receptors are considered as candidates for anti-inflammatory and tissue protective therapy. We demonstrated that the prototypical cannabinoid agonist R(+WIN55,212-2 (WIN reduced the secretion of matrix metalloproteinase-9 (MMP-9 in a murine model of cigarette-smoke induced lung inflammation. In experiments using primary cells and cell lines of the monocyte-macrophage-system we found that binding of the cannabinoid-receptor agonist WIN to a stereo-selective, specific binding site in cells of the monocyte-macrophage-system induced a significant down-regulation of MMP-9 secretion and disturbance of intracellular processing, which subsequently down-regulated MMP-9 mRNA expression via a ERK1/2-phosphorylation-dependent pathway. Surprisingly, the anti-inflammatory effect was independent from classical cannabinoid receptors. Our experiments supposed an involvement of TRPV1, but other yet unidentified sites are also possible. We conclude that cannabinoid-induced control of MMP-9 in the monocyte-macrophage system via a cannabinoid-receptor independent pathway represents a general option for tissue protection during inflammation, such as during lung inflammation and other diseases associated with inflammatory tissue damage.

  8. Nanoporosity of Alumina Surfaces Induces Different Patterns of Activation in Adhering Monocytes/Macrophages

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    Natalia Ferraz

    2010-01-01

    Full Text Available The present study shows that alumina nanotopography affects monocyte/macrophage behavior. Human mononuclear cells cultured on alumina membranes with pore diameters of 20 and 200 nm were evaluated in terms of cell adhesion, viability, morphology, and release of proinflammatory cytokines. After 24 hours, cell adhesion was assessed by means of light microscopy and cell viability by measuring LDH release. The inflammatory response was evaluated by quantifying interleukin-1β and tumour necrosis factor-α. Finally, scanning electron microscopy was used to study cell morphology. Results showed pronounced differences in cell number, morphology, and cytokine release depending on the nanoporosity. Few but highly activated cells were found on the 200 nm porous alumina, while relatively larger number of cells were found on the 20 nm porous surface. However, despite their larger number, the cells adhering on the 20 nm surface exhibited reduced pro-inflammatory activity. The data of this paper implies that nanotopography could be exploited for controlling the inflammatory response to implants.

  9. Pathology of African swine fever: the role of monocyte-macrophage.

    Science.gov (United States)

    Gómez-Villamandos, J C; Bautista, M J; Sánchez-Cordón, P J; Carrasco, L

    2013-04-01

    African swine fever (ASF) is a viral hemorrhagic disease with different clinical and lesional changes depending of virulence of strains/isolates and immunological status of pigs. In acute and subacute forms of ASF, severe vascular changes are present, with hemorrhages in different organs (mainly melena, epistaxis, erythema, renal petechiaes and diffuse hemorrhages in lymph nodes), pulmonary edema, disseminate intravascular coagulation and thrombocytopenia. Lymphopenia and monocytopenia are developed during acute and subacute ASF. Lymphopenia is associated with lymphoid depletion in primary and secondary lymphoid organs, which is caused by apoptosis. All these lesions are not related to viral replication in endothelial cells or lymphocytes. Monocytes-macrophages show viral replication and cytophatic effect, including hemadsorption. The more significant changes in these cells are increased number and secretory activation (increased levels of proinflammatory cytokines) in targets organs. Proinflammatory activation is the initial cause of clinical and lesional pictures in ASF, including fever and changes in levels of acute phase proteins. Levels of IFN-β and -γ are increased from initial phase of acute ASF. Anti-inflammatory response, represented by increased level of IL-10, is observed also, although in the final phase of acute ASF only.

  10. Unsaturated fatty acids prevent activation of NLRP3 inflammasome in human monocytes/macrophages[S

    Science.gov (United States)

    L'homme, Laurent; Esser, Nathalie; Riva, Laura; Scheen, André; Paquot, Nicolas; Piette, Jacques; Legrand-Poels, Sylvie

    2013-01-01

    The NLRP3 inflammasome is involved in many obesity-associated diseases, such as type 2 diabetes, atherosclerosis, and gouty arthritis, through its ability to induce interleukin (IL)-1β release. The molecular link between obesity and inflammasome activation is still unclear, but free fatty acids have been proposed as one triggering event. Here we reported opposite effects of saturated fatty acids (SFAs) compared with unsaturated fatty acids (UFAs) on NLRP3 inflammasome in human monocytes/macrophages. Palmitate and stearate, both SFAs, triggered IL-1β secretion in a caspase-1/ASC/NLRP3-dependent pathway. Unlike SFAs, the UFAs oleate and linoleate did not lead to IL-1β secretion. In addition, they totally prevented the IL-1β release induced by SFAs and, with less efficiency, by a broad range of NLRP3 inducers, including nigericin, alum, and monosodium urate. UFAs did not affect the transcriptional effect of SFAs, suggesting a specific effect on the NLRP3 activation. These results provide a new anti-inflammatory mechanism of UFAs by preventing the activation of the NLRP3 inflammasome and, therefore, IL-1β processing. By this way, UFAs might play a protective role in NLRP3-associated diseases. PMID:24006511

  11. Liver macrophages in healthy and diseased liver.

    Science.gov (United States)

    Abdullah, Zeinab; Knolle, Percy A

    2017-04-01

    Kupffer cells, the largest tissue resident macrophage population, are key for the maintenance of liver integrity and its restoration after injury and infections, as well as the local initiation and resolution of innate and adaptive immunity. These important roles of Kupffer cells were recently identified in healthy and diseased liver revealing diverse functions and phenotypes of hepatic macrophages. High-level phenotypic and genomic analysis revealed that Kupffer cells are not a homogenous population and that the hepatic microenvironment actively shapes both phenotype and function of liver macrophages. Compared to macrophages from other organs, hepatic macrophages bear unique properties that are instrumental for their diverse roles in local immunity as well as liver regeneration. The diverse and, in part, contradictory roles of hepatic macrophages in anti-tumor and inflammatory immune responses as well as regulatory and regenerative processes have been obscured by the lack of appropriate technologies to specifically target or ablate Kupffer cells or monocyte-derived hepatic macrophages. Future studies will need to dissect the exact role of the hepatic macrophages with distinct functional properties linked to their differentiation status and thereby provide insight into the functional plasticity of hepatic macrophages.

  12. Macrophage models of Gaucher disease for evaluating disease pathogenesis and candidate drugs.

    Science.gov (United States)

    Aflaki, Elma; Stubblefield, Barbara K; Maniwang, Emerson; Lopez, Grisel; Moaven, Nima; Goldin, Ehud; Marugan, Juan; Patnaik, Samarjit; Dutra, Amalia; Southall, Noel; Zheng, Wei; Tayebi, Nahid; Sidransky, Ellen

    2014-06-11

    Gaucher disease is caused by an inherited deficiency of glucocerebrosidase that manifests with storage of glycolipids in lysosomes, particularly in macrophages. Available cell lines modeling Gaucher disease do not demonstrate lysosomal storage of glycolipids; therefore, we set out to develop two macrophage models of Gaucher disease that exhibit appropriate substrate accumulation. We used these cellular models both to investigate altered macrophage biology in Gaucher disease and to evaluate candidate drugs for its treatment. We generated and characterized monocyte-derived macrophages from 20 patients carrying different Gaucher disease mutations. In addition, we created induced pluripotent stem cell (iPSC)-derived macrophages from five fibroblast lines taken from patients with type 1 or type 2 Gaucher disease. Macrophages derived from patient monocytes or iPSCs showed reduced glucocerebrosidase activity and increased storage of glucocerebroside and glucosylsphingosine in lysosomes. These macrophages showed efficient phagocytosis of bacteria but reduced production of intracellular reactive oxygen species and impaired chemotaxis. The disease phenotype was reversed with a noninhibitory small-molecule chaperone drug that enhanced glucocerebrosidase activity in the macrophages, reduced glycolipid storage, and normalized chemotaxis and production of reactive oxygen species. Macrophages differentiated from patient monocytes or patient-derived iPSCs provide cellular models that can be used to investigate disease pathogenesis and facilitate drug development.

  13. DMPD: LPS induction of gene expression in human monocytes. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 11257452 LPS induction of gene expression in human monocytes. Guha M, Mackman N. Ce...ll Signal. 2001 Feb;13(2):85-94. (.png) (.svg) (.html) (.csml) Show LPS induction of gene expression in human... monocytes. PubmedID 11257452 Title LPS induction of gene expression in human monocytes. Authors Guha M, Ma

  14. The dysregulation of the monocyte/macrophage effector function induced by isopropanol is mediated by the defective activation of distinct members of the AP-1 family of transcription factors.

    Science.gov (United States)

    Carignan, Damien; Désy, Olivier; de Campos-Lima, Pedro O

    2012-01-01

    Isopropanol is the second most common cause of short-chain alcohol acute intoxication. Nonethanolic short-chain alcohols mediate their immunomodulatory effect by interfering with nuclear factor of activated T cells (NFAT) activation with or without additional activator protein-1 (AP-1) involvement. In the present study, we examined the immunomodulation induced by isopropanol in conditions that are not reliant on NFAT: the inflammatory cytokine response of lipopolysaccharide (LPS)-stimulated monocytes. Our hypothesis was that isopropanol acute exposure would have an attenuated effect or no consequence in this setting. To our surprise, the impairment of AP-1 activation was sufficient to mediate a severe and dose-dependent phenotype in human monocytes in vitro at alcohol concentrations as low as 0.16% (or 26 mM). There were three outcomes: interleukin (IL)-1β/IL-8 were unaltered; IL-6 was upregulated; and tumor necrosis factor alpha (TNF-α)/CCL2 were downregulated. The effector function of human monocyte-derived macrophages was also compromised. Our results showed that Toll-like receptor 4 early signaling was preserved, as isopropanol did not change the kinase activity of the IL-1 receptor-associated kinase 1 in LPS-stimulated cells. The nuclear factor-κB signaling cascade and the p38/c-Jun N-terminal kinase modules of the mitogen-activated protein kinase pathway were alcohol insensitive. Conversely, the activation of extracellular signal-regulated protein kinase and, ultimately, of c-Fos and JunB were impaired. The alcohol-induced cytokine dysregulation was confirmed in a mouse model of isopropanol intoxication in which the production of TNF-α in response to LPS challenge was virtually abolished. The magnitude of this alcohol effect was sufficiently high to rescue animals from LPS-induced toxic shock. Our data contribute to the dismal body of information on the immunotoxicology of isopropanol, one of the most ubiquitous chemicals to which the general population

  15. Porphyromonas gingivalis-mediated signaling through TLR4 mediates persistent HIV infection of primary macrophages

    Science.gov (United States)

    Agosto, Luis M.; Hirnet, Juliane B.; Michaels, Daniel H.; Shaik-Dasthagirisaheb, Yazdani B.; Gibson, Frank C.; Viglianti, Gregory; Henderson, Andrew J.

    2016-01-01

    Periodontal infections contribute to HIV-associated co-morbidities in the oral cavity and provide a model to interrogate the dysregulation of macrophage function, inflammatory disease progression, and HIV replication during co-infections. We investigated the effect of Porphyromonas gingivalis on the establishment of HIV infection in monocyte-derived macrophages. HIV replication in macrophages was significantly repressed in the presence of P. gingivalis. This diminished viral replication was due partly to a decrease in the expression of integrated HIV provirus. HIV repression depended upon signaling through TLR4 as knock-down of TLR4 with siRNA rescued HIV expression. Importantly, HIV expression was reactivated upon removal of P. gingivalis. Our observations suggest that exposure of macrophages to Gram-negative bacteria influence the establishment and maintenance of HIV persistence in macrophages through a TLR4-dependent mechanism. PMID:27639573

  16. Evidence That Ly6C(hi) Monocytes are Protective in Acute Ischemic Stroke by Promoting M2 Macrophage Polarization.

    Science.gov (United States)

    Chu, Hannah X; Broughton, Brad R S; Kim, Hyun Ah; Lee, Seyoung; Drummond, Grant R; Sobey, Christopher G

    2015-07-01

    Ly6C(hi) monocytes are generally thought to exert a proinflammatory role in acute tissue injury, although their impact after injuries to the central nervous system is poorly defined. CC chemokine receptor 2 is expressed on Ly6C(hi) monocytes and plays an essential role in their extravasation and transmigration into the brain after cerebral ischemia. We used a selective CC chemokine receptor 2 antagonist, INCB3344, to assess the effect of Ly6C(hi) monocytes recruited into the brain early after ischemic stroke. Male C57Bl/6J mice underwent occlusion of the middle cerebral artery for 1 hour followed by 23 hours of reperfusion. Mice were administered either vehicle (dimethyl sulfoxide/carboxymethylcellulose) or INCB3344 (10, 30 or 100 mg/kg IP) 1 hour before ischemia and at 2 and 6 hours after ischemia. At 24 hours, we assessed functional outcomes, infarct volume, and quantified the immune cells in blood and brain by flow cytometry or immunofluorescence. Gene expression of selected inflammatory markers was assessed by quantitative polymerase chain reaction. Ly6C(hi) monocytes were increased 3-fold in the blood and 10-fold in the brain after stroke, and these increases were selectively prevented by INCB3344 in a dose-dependent manner. Mice treated with INCB3344 exhibited markedly worse functional outcomes and larger infarct volumes, in association with reduced M2 polarization and increased peroxynitrite production in macrophages, compared with vehicle-treated mice. Our data suggest that Ly6C(hi) monocytes exert an acute protective effect after ischemic stroke to limit brain injury and functional deficit that involves promotion of M2 macrophage polarization. © 2015 American Heart Association, Inc.

  17. Induction of Monocyte Chemoattractant Proteins in Macrophages via the Production of Granulocyte-macrophage Colony Stimulating Factor by Breast Cancer Cells

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    Teizo eYoshimura

    2016-01-01

    Full Text Available Monocyte chemoattractant protein-1 (MCP-1/CCL2 plays an important role in the initiation and progression of cancer. We previously reported that in 4T1 murine breast cancer, non-tumor stromal cells, including macrophages, were the major source of MCP-1. In the present study, we analyzed the potential mechanisms by which MCP-1 is upregulated in macrophages infiltrating 4T1 tumors. We found that cell-free culture supernatants of 4T1 cells (4T1-sup markedly upregulated MCP-1 production by peritoneal inflammatory macrophages. 4T1-sup also upregulated other MCPs, such as MCP-3/CCL7 and MCP-5/CCL12, but modestly neutrophil chemotactic chemokines, such as KC/CXCL1 or MIP-2/CXCL2. Physicochemical analysis indicated that an approximately 2 to 3 kDa 4T1 cell product was responsible for the capacity of 4T1-sup to upregulate MCP-1 expression by macrophages. A neutralizing antibody against granulocyte-macrophage-colony stimulating factor (GM-CSF, but not macrophage-colony stimulating factor, almost completely abrogated MCP-1-inducing activity of 4T1-sup, and recombinant GM-CSF potently up-regulated MCP-1 production by macrophages. The expression levels of GM-CSF in 4T1 tumors in vivo were higher than other tumors, such as Lewis lung carcinoma. Treatment of mice with anti-GM-CSF antibody significantly reduced the growth of 4T1 tumors at the injection sites but did not reduce MCP-1 production or lung metastasis in tumor-bearing mice. These results indicate that 4T1 cells have the capacity to directly up-regulate MCP-1 production by macrophages by releasing GM-CSF; however, other mechanisms are also involved in increased MCP-1 levels in the 4T1 tumor microenvironment.

  18. The role of monocytes in ischemic stroke pathobiology: New avenues to explore

    Directory of Open Access Journals (Sweden)

    Ayman eElAli

    2016-02-01

    Full Text Available Ischemic stroke accounts for the majority of stroke cases and constitutes a major cause of death and disability in the industrialized world. Inflammation has been reported to constitute a major component of ischemic stroke pathobiology. In the acute phase of ischemic stroke, microglia, the resident macrophages of the brain, are activated, followed by several infiltration waves of different circulating immune cells into the brain. Among these circulating immune cells, monocytes have been shown to play a particularly important role. Following their infiltration, monocytes differentiate into potent phagocytic cells, monocyte-derived macrophages (MDMs in the ischemic brain. Initially, the presence of these cells was considered as marker of an exacerbated inflammatory response that contributes to brain damage. However, the recent reports are suggesting a more complex and multiphasic roles of these cells in ischemic stroke pathobiology. Monocytes constitute a heterogeneous group of cells, which comprises two major subsets in rodent and three major subsets in human. In both species, two equivalent subsets exist, the pro-inflammatory subset and the anti-inflammatory subset. Recent data have demonstrated that ischemic stroke differentially regulate monocyte subsets, which directly affect ischemic stroke pathobiology and may have direct implications in ischemic stroke therapies. Here we review the recent findings that addressed the role of different monocyte subsets in ischemic stroke pathobiology, and the implications on therapies.

  19. The role of macrophages in skin homeostasis.

    Science.gov (United States)

    Yanez, Diana A; Lacher, Richard K; Vidyarthi, Aurobind; Colegio, Oscar R

    2017-04-01

    The skin and its appendages comprise the largest and fastest growing organ in the body. It performs multiple tasks and maintains homeostatic control, including the regulation of body temperature and protection from desiccation and from pathogen invasion. The skin can perform its functions with the assistance of different immune cell populations. Monocyte-derived cells are imperative for the completion of these tasks. The comprehensive role of macrophages and Langerhans cells in establishing and maintaining skin homeostasis remains incompletely defined. However, over the past decade, innovations in mouse genetics have allowed for advancements in the field. In this review, we explore different homeostatic roles of macrophages and Langerhans cells, including wound repair, follicle regeneration, salt balance, and cancer regression and progression in the skin. The understanding of the precise functions of myeloid-derived cells in the skin under basal conditions can help develop specific therapies that aid in skin and hair follicle regeneration and cutaneous cancer prevention.

  20. Uptake of cerium oxide nanoparticles and its influence on functions of mouse leukemic monocyte macrophages

    Science.gov (United States)

    Zhou, Xiangyan; Wang, Bing; Jiang, Pengfei; Chen, Yiqi; Mao, Zhengwei; Gao, Changyou

    2015-01-01

    Exposure of the CeO2 nanoparticles (NPs) causes a public concern on their potential health risk due to their wide applications in the fields of fuel additive, commodities, pharmaceutical, and other industries. In this study, the interactions between two commercial CeO2 NPs (D-CeO2 from Degussa and PC-CeO2 from PlasmaChem) and mouse leukemic monocyte macrophage Raw264.7 cells were investigated to provide a fast and in-depth understanding of the biological influences of the NPs. Both types of the CeO2 NPs had a negative surface charge around -12 mV and showed a tendency to form aggregates with sizes of 191 ± 5.9 and 60.9 ± 2.8 nm in cell culture environment, respectively. The cellular uptake of the CeO2 NPs increased along with the increase of feeding dosage and prolongation of the culture time. The PC-CeO2 NPs had a faster uptake rate and reached higher cellular loading amount at the highest feeding concentration (200 µg/mL). In general, both types of the CeO2 NPs had rather small cytotoxicity even with a dosage as high as 200 µg/mL. The D-CeO2 NPs showed a relative stronger cytotoxicity especially at higher concentrations and longer incubation time. The NPs were dispersed in vacuoles (most likely endosomes and lysosomes) and cytoplasm. Although both types of the CeO2 NPs could suppress the production of reactive oxygen species, they impaired the mitochondria membrane potential to some extent. The cytoskeleton organization was altered and consequently the cell adhesion ability decreased after uptake of both types of the CeO2 NPs.

  1. Uptake of cerium oxide nanoparticles and its influence on functions of mouse leukemic monocyte macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Xiangyan; Wang, Bing; Jiang, Pengfei; Chen, Yiqi; Mao, Zhengwei, E-mail: zwmao@zju.edu.cn; Gao, Changyou [Zhejiang University, MOE Key Laboratory of Macromolecular Synthesis and Functionalization, Department of Polymer Science and Engineering (China)

    2015-01-15

    Exposure of the CeO{sub 2} nanoparticles (NPs) causes a public concern on their potential health risk due to their wide applications in the fields of fuel additive, commodities, pharmaceutical, and other industries. In this study, the interactions between two commercial CeO{sub 2} NPs (D-CeO{sub 2} from Degussa and PC-CeO{sub 2} from PlasmaChem) and mouse leukemic monocyte macrophage Raw264.7 cells were investigated to provide a fast and in-depth understanding of the biological influences of the NPs. Both types of the CeO{sub 2} NPs had a negative surface charge around −12 mV and showed a tendency to form aggregates with sizes of 191 ± 5.9 and 60.9 ± 2.8 nm in cell culture environment, respectively. The cellular uptake of the CeO{sub 2} NPs increased along with the increase of feeding dosage and prolongation of the culture time. The PC-CeO{sub 2} NPs had a faster uptake rate and reached higher cellular loading amount at the highest feeding concentration (200 µg/mL). In general, both types of the CeO{sub 2} NPs had rather small cytotoxicity even with a dosage as high as 200 µg/mL. The D-CeO{sub 2} NPs showed a relative stronger cytotoxicity especially at higher concentrations and longer incubation time. The NPs were dispersed in vacuoles (most likely endosomes and lysosomes) and cytoplasm. Although both types of the CeO{sub 2} NPs could suppress the production of reactive oxygen species, they impaired the mitochondria membrane potential to some extent. The cytoskeleton organization was altered and consequently the cell adhesion ability decreased after uptake of both types of the CeO{sub 2} NPs.

  2. HIV/SIV infection primes monocytes and dendritic cells for apoptosis.

    Directory of Open Access Journals (Sweden)

    Mireille Laforge

    2011-06-01

    Full Text Available Subversion or exacerbation of antigen-presenting cells (APC death modulates host/pathogen equilibrium. We demonstrated during in vitro differentiation of monocyte-derived macrophages and monocyte-derived dendritic cells (DCs that HIV sensitizes the cells to undergo apoptosis in response to TRAIL and FasL, respectively. In addition, we found that HIV-1 increased the levels of pro-apoptotic Bax and Bak molecules and decreased the levels of anti-apoptotic Mcl-1 and FLIP proteins. To assess the relevance of these observations in the context of an experimental model of HIV infection, we investigated the death of APC during pathogenic SIV-infection in rhesus macaques (RMs. We demonstrated increased apoptosis, during the acute phase, of both peripheral blood DCs and monocytes (CD14(+ from SIV(+RMs, associated with a dysregulation in the balance of pro- and anti-apoptotic molecules. Caspase-inhibitor and death receptors antagonists prevented apoptosis of APCs from SIV(+RMs. Furthermore, increased levels of FasL in the sera of pathogenic SIV(+RMs were detected, compared to non-pathogenic SIV infection of African green monkey. We suggest that inappropriate apoptosis of antigen-presenting cells may contribute to dysregulation of cellular immunity early in the process of HIV/SIV infection.

  3. Low molecular weight hyaluronan activates cytosolic phospholipase A2α and eicosanoid production in monocytes and macrophages.

    Science.gov (United States)

    Sokolowska, Milena; Chen, Li-Yuan; Eberlein, Michael; Martinez-Anton, Asuncion; Liu, Yueqin; Alsaaty, Sara; Qi, Hai-Yan; Logun, Carolea; Horton, Maureen; Shelhamer, James H

    2014-02-14

    Hyaluronan (HA) is the major glycosaminoglycan in the extracellular matrix. During inflammation, there is an increased breakdown of HA, resulting in the accumulation of low molecular weight (LMW) HA and activation of monocytes and macrophages. Eicosanoids, derived from the cytosolic phospholipase A2 group IVA (cPLA2α) activation, are potent lipid mediators also attributed to acute and chronic inflammation. The aim of this study was to determine the effect of LMW HA on cPLA2α activation, arachidonic acid (AA) release, and subsequent eicosanoid production and to examine the receptors and downstream mechanisms involved in these processes in monocytes and differently polarized macrophages. LMW HA was a potent stimulant of AA release in a time- and dose-dependent manner, induced cPLA2α, ERK1/2, p38, and JNK phosphorylation, as well as activated COX2 expression and prostaglandin (PG) E2 production in primary human monocytes, murine RAW 264.7, and wild-type bone marrow-derived macrophages. Specific cPLA2α inhibitor blocked HA-induced AA release and PGE2 production in all of these cells. Using CD44, TLR4, TLR2, MYD88, RHAMM or STAB2 siRNA-transfected macrophages and monocytes, we found that AA release, cPLA2α, ERK1/2, p38, and JNK phosphorylation, COX2 expression, and PGE2 production were activated by LMW HA through a TLR4/MYD88 pathway. Likewise, PGE2 production and COX2 expression were blocked in Tlr4(-/-) and Myd88(-/-) mice, but not in Cd44(-/-) mice, after LMW HA stimulation. Moreover, we demonstrated that LMW HA activated the M1 macrophage phenotype with the unique cPLA2α/COX2(high) and COX1/ALOX15/ALOX5/LTA4H(low) gene and PGE2/PGD2/15-HETE(high) and LXA4(low) eicosanoid profile. These findings reveal a novel link between HA-mediated inflammation and lipid metabolism.

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

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    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

  5. Monocytes/macrophages infected with Toxoplasma gondii do not increase co-stimulatory molecules while maintaining their migratory ability.

    Science.gov (United States)

    Seipel, Daniele; Ribeiro-Gomes, Flavia Lima; Barcelos, Michelle Willmen; Ramalho, André Villaça; Kanashiro, Milton M; Kipnis, Thereza Liberman; Arnholdt, Andrea Cristina Veto

    2009-09-01

    Toxoplasma gondii is an obligate intracellular parasite that is able to disseminate into deep tissues and cross biological barriers, reaching immunoprivileged sites such as the brain and retina. The parasite is able to infect macrophages and dendritic cells and use them for dispersal throughout the body, but the activation state of those cells is unknown. We investigated the ability of human and murine cells from monocytic/macrophage lineages that had not previously been exposed to inflammatory cytokines to up-regulate co-stimulatory and adhesion molecules upon infection. Toxoplasma gondii-infected human monocytes (freshly isolated and THP1 lineage) were unable to up-regulate CD86, CD83, CD40 or CD1a. CD80 expression increased in infected cells but expression of l-selectin and beta2 integrin was unaltered. We evaluated the ability of infected macrophages from wild type C57/BL/6 or CD14(-/-) mice to migrate in 8 mum transwells. Infected cells from CD14(-/-) mice were more likely to de-adhere than infected cells from wild type mice but they did not show any increase in migratory ability. The non-stimulatory profile of these infected cells may contribute to parasite spread throughout the lymphatic circulation in the initial phases of infection.

  6. Identification and characterization of a non-interferon antileishmanial macrophage activating factor (antileishmanial MAF).

    Science.gov (United States)

    Van Niel, A; Zacks, S E; David, J R; Remold, H G; Weiser, W Y

    1988-01-01

    A non-interferon lymphokine elaborated from PHA and Con A-stimulated human T-cell hybridoma, T-CEMA, has been found to activate monocyte-derived macrophages for the intracellular killing of L. donovani (antileishmanial MAF). This T-cell hybridoma derived antileishmanial MAF which has an apparent mw of 65,000 and pI of 5.3-5.6, contains neither antiviral activity nor colony stimulating activity. Furthermore, antileishmanial MAF is not neutralized by anti-MIF, anti-IFN-gamma or anti-GM-CSF antibodies.

  7. The glial scar-monocyte interplay: a pivotal resolution phase in spinal cord repair.

    Directory of Open Access Journals (Sweden)

    Ravid Shechter

    Full Text Available The inflammatory response in the injured spinal cord, an immune privileged site, has been mainly associated with the poor prognosis. However, recent data demonstrated that, in fact, some leukocytes, namely monocytes, are pivotal for repair due to their alternative anti-inflammatory phenotype. Given the pro-inflammatory milieu within the traumatized spinal cord, known to skew monocytes towards a classical phenotype, a pertinent question is how parenchymal-invading monocytes acquire resolving properties essential for healing, under such unfavorable conditions. In light of the spatial association between resolving (interleukin (IL-10 producing monocytes and the glial scar matrix chondroitin sulfate proteoglycan (CSPG, in this study we examined the mutual relationship between these two components. By inhibiting the de novo production of CSPG following spinal cord injury, we demonstrated that this extracellular matrix, mainly known for its ability to inhibit axonal growth, serves as a critical template skewing the entering monocytes towards the resolving phenotype. In vitro cell culture studies demonstrated that this matrix alone is sufficient to induce such monocyte polarization. Reciprocal conditional ablation of the monocyte-derived macrophages concentrated at the lesion margins, using diphtheria toxin, revealed that these cells have scar matrix-resolving properties. Replenishment of monocytic cell populations to the ablated mice demonstrated that this extracellular remodeling ability of the infiltrating monocytes requires their expression of the matrix-degrading enzyme, matrix metalloproteinase 13 (MMP-13, a property that was found here to be crucial for functional recovery. Altogether, this study demonstrates that the glial scar-matrix, a known obstacle to regeneration, is a critical component skewing the encountering monocytes towards a resolving phenotype. In an apparent feedback loop, monocytes were found to regulate scar resolution. This

  8. Macrophage Inflammatory Protein-1alpha mediates Matrix Metalloproteinase-9 enhancement in human adherent monocytes fed with malarial pigment

    Institute of Scientific and Technical Information of China (English)

    Giuliana Giribaldi; Elena Valente; Amina Khadjavi; Manuela Polimeni; Mauro Prato

    2011-01-01

    Objective:To investigate the role of macrophage inflammatory protein-1alpha (MIP-1alpha) in the detrimental enhancement of matrix metalloproteinase-9 (MMP-9)expression, release and activity induced by phagocytosis of malarial pigment (haemozoin,HZ) in human monocytes. Methods: Human adherent monocytes were unfed/fed with nativeHZ for 2 h. After 24 hours, MIP-1alpha production was evaluated by ELISA in cell supernatants. Alternatively,HZ-unfed/fed monocytes were treated in presence/absence of anti-humanMIP-1alpha blocking antibodies or recombinant humanMIP-1alpha for15 h (RNA studies) or 24 h (protein studies); therefore,MMP-9mRNA expression was evaluated in cell lysates by Real TimeRT-PCR, whereas proMMP-9and activeMMP-9protein release were measured in cell supernatants by Western blotting and gelatin zymography.Results: Phagocytosis ofHZ by human monocytes increased production ofMIP-1alpha, mRNA expression ofMMP-9and protein release of proMMP-9 and activeMMP-9. All theHZ-enhancing effects onMMP-9 were abrogated by anti-humanMIP-1alpha blocking antibodies and mimicked by recombinant humanMIP-1alpha.Conclusions:The present work suggests a role for MIP-1alpha in theHZ-dependent enhancement ofMMP-9 expression, release and activity observed in human monocytes, highlighting new detrimental effects ofHZ-triggered proinflammatory response by phagocytic cells in falciparum malaria.

  9. Human lung-resident macrophages express CB1 and CB2 receptors whose activation inhibits the release of angiogenic and lymphangiogenic factors.

    Science.gov (United States)

    Staiano, Rosaria I; Loffredo, Stefania; Borriello, Francesco; Iannotti, Fabio Arturo; Piscitelli, Fabiana; Orlando, Pierangelo; Secondo, Agnese; Granata, Francescopaolo; Lepore, Maria Teresa; Fiorelli, Alfonso; Varricchi, Gilda; Santini, Mario; Triggiani, Massimo; Di Marzo, Vincenzo; Marone, Gianni

    2016-04-01

    Macrophages are pivotal effector cells in immune responses and tissue remodeling by producing a wide spectrum of mediators, including angiogenic and lymphangiogenic factors. Activation of cannabinoid receptor types 1 and 2 has been suggested as a new strategy to modulate angiogenesis in vitro and in vivo. We investigated whether human lung-resident macrophages express a complete endocannabinoid system by assessing their production of endocannabinoids and expression of cannabinoid receptors. Unstimulated human lung macrophage produce 2-arachidonoylglycerol,N-arachidonoyl-ethanolamine,N-palmitoyl-ethanolamine, and N-oleoyl-ethanolamine. On LPS stimulation, human lung macrophages selectively synthesize 2-arachidonoylglycerol in a calcium-dependent manner. Human lung macrophages express cannabinoid receptor types 1 and 2, and their activation induces ERK1/2 phosphorylation and reactive oxygen species generation. Cannabinoid receptor activation by the specific synthetic agonists ACEA and JWH-133 (but not the endogenous agonist 2-arachidonoylglycerol) markedly inhibits LPS-induced production of vascular endothelial growth factor-A, vascular endothelial growth factor-C, and angiopoietins and modestly affects IL-6 secretion. No significant modulation of TNF-α or IL-8/CXCL8 release was observed. The production of vascular endothelial growth factor-A by human monocyte-derived macrophages is not modulated by activation of cannabinoid receptor types 1 and 2. Given the prominent role of macrophage-assisted vascular remodeling in many tumors, we identified the expression of cannabinoid receptors in lung cancer-associated macrophages. Our results demonstrate that cannabinoid receptor activation selectively inhibits the release of angiogenic and lymphangiogenic factors from human lung macrophage but not from monocyte-derived macrophages. Activation of cannabinoid receptors on tissue-resident macrophages might be a novel strategy to modulate macrophage-assisted vascular remodeling

  10. Inflammatory cytokine regulation by LPS and lymphoid cells in human gamma-irradiated monocytes/macrophages; Regulation des cytokines de l`inflammation en presence de LPS ou de lymphocytes dans les monocytes/macrophages humains irradies

    Energy Technology Data Exchange (ETDEWEB)

    Pons, I.; Gras, G.; Dormont, D. [Centre de Recherches du Service de Sante des Armees, La Tronche, 38 - Grenoble (France)]|[Centre de Recherches du Service de Sante des Armees - Centre d`Etudes Nucleaires de Fontenay-aux-Roses, 92 (France)]|[Paris-5 Univ., 75 (France)

    1997-12-31

    We have investigated the inflammatory cytokine regulation after ionizing radiation of monocytes/macrophages. We have not evidenced any significant induction of tumour necrosis factor-{alpha}(TNF{alpha}) after irradiation alone. For one donor only out of eight, interleukin-1{beta}(IL-l{beta}) gene expression was affected by {gamma}-irradiation, with a 2-3-fold increase in level, while for two other donors, interleukin-6 (IL-6) mRNA expression was 5-14 fold increased. For one of the eight donors tested, monocytes/macrophages responded to 10 Gy {gamma}-rays by releasing inflammatory cytokines. In the presence of LPS, a significant increase of IL-1{beta} mRNA expression was detected in 10 Gy {gamma}-irradiated cells treated with 1 {mu}g/ml LPS. In most cases, combination of LPS treatment and 10 Gy irradiation down-regulated cytokine secretion except for a TNF{alpha} induction at 6 h post-irradiation. In the presence of lymphoid cells, IL-6 mRNA level was increased in irradiated cells at 24 h. Increases of IL-1{beta} and IL-6 releases were detected at 24 h post-irradiation too. (authors)

  11. Transcriptional immunoresponse of tissue-specific macrophages in swine after infection with African swine fever virus

    Directory of Open Access Journals (Sweden)

    Kowalczyk Andrzej

    2015-12-01

    Full Text Available Macrophages and cytokines are important in the control of inflammation and regulation of the immune response. However, they can also contribute to immunopathology in the host after viral infection and the regulatory network can be subverted by infectious agents, including viruses, some of which produce cytokine analogues or have mechanisms that inhibit cytokine function. African swine fever virus (ASFV encodes a number of proteins which modulate cytokine and chemokine induction, host transcription factor activation, stress responses, and apoptosis. The aim of this review is to elucidate the mechanisms of immune responses to ASFV in different subpopulations of porcine macrophages. A transcriptional immune response in different resident tissue macrophages following ASFV infection was presented in many publications. ASFV-susceptible porcine macrophages can be of several origins, such as peripheral blood, lungs, bone marrow, etc. blood monocytes, blood macrophages, and lung macrophages have demonstrated a modulation of phenotype. Monocyte-derived macrophages could express surface markers not found on their monocyte precursors. Moreover, they can undergo further differentiation after infection and during inflammation. When viruses infect such cells, immunological activity can be seriously impaired or modified.

  12. Exenatide (a GLP-1 agonist) improves the antioxidative potential of in vitro cultured human monocytes/macrophages.

    Science.gov (United States)

    Bułdak, Łukasz; Łabuzek, Krzysztof; Bułdak, Rafał Jakub; Machnik, Grzegorz; Bołdys, Aleksandra; Okopień, Bogusław

    2015-09-01

    Macrophages are dominant cells in the pathogenesis of atherosclerosis. They are also a major source of reactive oxygen species (ROS). Oxidative stress, which is particularly high in subjects with diabetes, is responsible for accelerated atherosclerosis. Novel antidiabetic drugs (e.g., glucagon-like peptide-1 (GLP-1) agonists) were shown to reduce ROS level. Therefore, we conceived a study to evaluate the influence of exenatide, a GLP-1 agonist, on redox status in human monocytes/macrophages cultured in vitro, which may explain the beneficial effects of incretin-based antidiabetic treatment. Human macrophages obtained from 10 healthy volunteers were in vitro subjected to the treatment with GLP-1 agonist (exenatide) in the presence of lipopolysaccharide (LPS), antagonist of GLP-1 receptors (exendin 9-39), or protein kinase A inhibitor (H89). Afterwards, reactive oxygen species, malondialdehyde level, NADPH oxidase, and antioxidative enzymes [superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase] expression was evaluated. Finally, we estimated the activity of the abovementioned enzymes in the presence of H89. According to our findings, exenatide reduced ROS and malondialdyhyde (MDA) level by decreasing the expression of ROS-generating NADPH oxidase and by increasing the expression and activities of SOD and GSH-Px. We also showed that this effect was significantly inhibited by exendin 9-39 (a GLP-1 antagonist) and blocked by H89. Exenatide improved the antioxidative potential and reduced oxidative stress in cultured human monocytes/macrophages, and this finding may be responsible for the pleiotropic effects of incretin-based therapies. This effect relied on the stimulation of GLP-1 receptor.

  13. Recruitment of beneficial M2 macrophages to injured spinal cord is orchestrated by remote brain choroid plexus.

    Science.gov (United States)

    Shechter, Ravid; Miller, Omer; Yovel, Gili; Rosenzweig, Neta; London, Anat; Ruckh, Julia; Kim, Ki-Wook; Klein, Eugenia; Kalchenko, Vyacheslav; Bendel, Peter; Lira, Sergio A; Jung, Steffen; Schwartz, Michal

    2013-03-21

    Monocyte-derived macrophages are essential for recovery after spinal cord injury, but their homing mechanism is poorly understood. Here, we show that although of common origin, the homing of proinflammatory (M1) and the "alternatively activated" anti-inflammatory (M2) macrophages to traumatized spinal cord (SC) was distinctly regulated, neither being through breached blood-brain barrier. The M1 macrophages (Ly6c(hi)CX3CR1(lo)) derived from monocytes homed in a CCL2 chemokine-dependent manner through the adjacent SC leptomeninges. The resolving M2 macrophages (Ly6c(lo)CX3CR1(hi)) derived from monocytes trafficked through a remote blood-cerebrospinal-fluid (CSF) barrier, the brain-ventricular choroid plexus (CP), via VCAM-1-VLA-4 adhesion molecules and epithelial CD73 enzyme for extravasation and epithelial transmigration. Blockage of these determinants, or mechanical CSF flow obstruction, inhibited M2 macrophage recruitment and impaired motor-function recovery. The CP, along with the CSF and the central canal, provided an anti-inflammatory supporting milieu, potentially priming the trafficking monocytes. Overall, our finding demonstrates that the route of monocyte entry to central nervous system provides an instructional environment to shape their function.

  14. Dyslipidemic Diet-Induced Monocyte “Priming” and Dysfunction in Non-Human Primates Is Triggered by Elevated Plasma Cholesterol and Accompanied by Altered Histone Acetylation

    Directory of Open Access Journals (Sweden)

    John D. Short

    2017-08-01

    Full Text Available Monocytes and the recruitment of monocyte-derived macrophages into sites of inflammation play a key role in atherogenesis and other chronic inflammatory diseases linked to cardiometabolic syndrome and obesity. Previous studies from our group have shown that metabolic stress promotes monocyte priming, i.e., enhanced adhesion and accelerated chemotaxis of monocytes in response to chemokines, both in vitro and in dyslipidemic LDLR−/− mice. We also showed that metabolic stress-induced monocyte dysfunction is, at least to a large extent caused by the S-glutathionylation, inactivation, and subsequent degradation of mitogen-activated protein kinase phosphatase 1. Here, we analyzed the effects of a Western-style, dyslipidemic diet (DD, which was composed of high levels of saturated fat, cholesterol, and simple sugars, on monocyte (dysfunction in non-human primates (NHPs. We found that similar to mice, a DD enhances monocyte chemotaxis in NHP within 4 weeks, occurring concordantly with the onset of hypercholesterolemia but prior to changes in triglycerides, blood glucose, monocytosis, or changes in monocyte subset composition. In addition, we identified transitory decreases in the acetylation of histone H3 at the lysine residues 18 and 23 in metabolically primed monocytes, and we found that monocyte priming was correlated with the acetylation of histone H3 at lysine 27 after an 8-week DD regimen. Our data show that metabolic stress promotes monocyte priming and hyper-chemotactic responses in NHP. The histone modifications accompanying monocyte priming in primates suggest a reprogramming of the epigenetic landscape, which may lead to dysregulated responses and functionalities in macrophages derived from primed monocytes that are recruited to sites of inflammation.

  15. 雷公藤红素对 CD1+4单核细胞来源的树突细胞分化及成熟的影响%Effect of celastrol on differentiation and maturation of CD1+4 monocyte-derived dendritic cells

    Institute of Scientific and Technical Information of China (English)

    王瑄; 刘郁莹; 彭美玉; 薛振毅

    2014-01-01

    Objective To Make the differentiation and activation models of dendritic cells ( DCs ) and to investigate the effect of celastrol on the differentiation and maturation of human DCs in vitro .Methods Fresh human buffy coat was obtained , and peripheral blood mononuclear cells were isolated .Purified CDl4+monocytes were analyzed by FACSCalibur to confirm the purity of CD1+4 cells.The purified CD1+4 cells were cultured in complete 1640 medium supplemented with GM-CSF and IL-4 for 5 days and 7 days.For DC maturation, cells were stimulated with 5?g/mL lipopolysaccharide (LPS) at day 5.The control group was not added celastrol , but the celastrol group was added 10 and 20 nM celastrol, respectively. Then immature DCs ( iDC ) and mature DCs ( mDC ) were collected to detect the mean fluorescent intensity ( MFI ) of CD80 , CD83 , CD86 , HLA-DR and the absorb capacity of FITC-dextran by flow cytometry .Results Compared with the control group, the MFI of CD80 , CD83 , CD86 and HLA-DR of iDC and mDC in the celastrol group was lower , and the treat-ment of 20 nM was lower than the treatment of 10 nM, all P<0.05.The MFI of FITC-dextran in the negative control group (10.1 ±2.1) was the lowest, and was the highest in control group (243.7 ±31.3);in addition, the MFI of FITC-dextran in the 20 nM celastrol group (72.4 ±10.2) was lower than that of the 10 nM celastrol group (186.3 ±22.6), all P<0.05.Conclusions Celastrol inhibits CD1+4 monocytes differentiating into DCs , further inhibits phenotypic maturation of LPS-induced DCs , and suppresses the antigen-presenting function of iDC .%目的:制作免疫树突细胞( DC)分化和激活模型,观察雷公藤红素对人DC体外分化和成熟的影响。方法采集浓缩人白细胞,淋巴细胞分离液分离单个核细胞,利用免疫磁珠法分离CD1+4细胞,流式细胞分选仪确认分选纯度。将分选的CD1+4细胞在含IL-4和GM-CSF的完全1640培养基中分别培养5 d(分化)和7 d(5 d

  16. Tracking Monocyte Recruitment and Macrophage Accumulation in Atherosclerotic Plaque Progression Using a Novel hCD68GFP/ApoE−/− Reporter Mouse—Brief Report

    Science.gov (United States)

    Iqbal, Asif J.; Jones, Daniel; Patel, Jyoti; Coutinho, Patricia; Taylor, Lewis; Greaves, David R.; Channon, Keith M.

    2017-01-01

    Objective— To create a model of atherosclerosis using green fluorescent protein (GFP)–targeted monocytes/macrophages, allowing analysis of both endogenous GFP+ and adoptively transferred GFP+ myeloid cells in arterial inflammation. Approach and Results— hCD68GFP reporter mice were crossed with ApoE−/− mice. Expression of GFP was localized to macrophages in atherosclerotic plaques and in angiotensin II–induced aortic aneurysms and correlated with galectin 3 and mCD68 expression. Flow cytometry confirmed GFP+ expression in CD11b+/CD64+, CD11c+/MHC-IIHI, and CD11b+/F4/80+ myeloid cells. Adoptive transfer of GFP+ monocytes demonstrated monocyte recruitment to both adventitia and atherosclerotic plaque, throughout the aortic root, within 72 hours. We demonstrated the biological utility of hCD68GFP monocytes by comparing the recruitment of wild-type and CCR2−/− monocytes to sites of inflammation. Conclusions— hCD68GFP/ApoE−/− mice provide a new approach to study macrophage accumulation in atherosclerotic plaque progression and to identify cells recruited from adoptively transferred monocytes. PMID:27908893

  17. Monocyte/macrophage androgen receptor suppresses cutaneous wound healing in mice by enhancing local TNF-alpha expression.

    Science.gov (United States)

    Lai, Jiann-Jyh; Lai, Kuo-Pao; Chuang, Kuang-Hsiang; Chang, Philip; Yu, I-Chen; Lin, Wen-Jye; Chang, Chawnshang

    2009-12-01

    Cutaneous wounds heal more slowly in elderly males than in elderly females, suggesting a role for sex hormones in the healing process. Indeed, androgen/androgen receptor (AR) signaling has been shown to inhibit cutaneous wound healing. AR is expressed in several cell types in healing skin, including keratinocytes, dermal fibroblasts, and infiltrating macrophages, but the exact role of androgen/AR signaling in these different cell types remains unclear. To address this question, we generated and studied cutaneous wound healing in cell-specific AR knockout (ARKO) mice. General and myeloid-specific ARKO mice exhibited accelerated wound healing compared with WT mice, whereas keratinocyte- and fibroblast-specific ARKO mice did not. Importantly, the rate of wound healing in the general ARKO mice was dependent on AR and not serum androgen levels. Interestingly, although dispensable for wound closure, keratinocyte AR promoted re-epithelialization, while fibroblast AR suppressed it. Further analysis indicated that AR suppressed wound healing by enhancing the inflammatory response through a localized increase in TNF-alpha expression. Furthermore, AR enhanced local TNF-alpha expression via multiple mechanisms, including increasing the inflammatory monocyte population, enhancing monocyte chemotaxis by upregulating CCR2 expression, and enhancing TNF-alpha expression in macrophages. Finally, targeting AR by topical application of a compound (ASC-J9) that degrades AR protein resulted in accelerated healing, suggesting a potential new therapeutic approach that may lead to better treatment of wound healing.

  18. HIV-1 regulation of latency in the monocyte-macrophage lineage and in CD4+ T lymphocytes.

    Science.gov (United States)

    Redel, Laetitia; Le Douce, Valentin; Cherrier, Thomas; Marban, Céline; Janossy, Andrea; Aunis, Dominique; Van Lint, Carine; Rohr, Olivier; Schwartz, Christian

    2010-04-01

    The introduction in 1996 of the HAART raised hopes for the eradication of HIV-1. Unfortunately, the discovery of latent HIV-1 reservoirs in CD4+ T cells and in the monocyte-macrophage lineage proved the optimism to be premature. The long-lived HIV-1 reservoirs constitute a major obstacle to the eradication of HIV-1. In this review, we focus on the establishment and maintenance of HIV-1 latency in the two major targets for HIV-1: the CD4+ T cells and the monocyte-macrophage lineage. Understanding the cell-type molecular mechanisms of establishment, maintenance, and reactivation of HIV-1 latency in these reservoirs is crucial for efficient therapeutic intervention. A complete viral eradication, the holy graal for clinicians, might be achieved by strategic interventions targeting latently and productively infected cells. We suggest that new approaches, such as the combination of different kinds of proviral activators, may help to reduce dramatically the size of latent HIV-1 reservoirs in patients on HAART.

  19. Differential roles of the protein corona in the cellular uptake of nanoporous polymer particles by monocyte and macrophage cell lines.

    Science.gov (United States)

    Yan, Yan; Gause, Katelyn T; Kamphuis, Marloes M J; Ang, Ching-Seng; O'Brien-Simpson, Neil M; Lenzo, Jason C; Reynolds, Eric C; Nice, Edouard C; Caruso, Frank

    2013-12-23

    Many biomolecules, mainly proteins, adsorb onto polymer particles to form a dynamic protein corona in biological environments. The protein corona can significantly influence particle-cell interactions, including internalization and pathway activation. In this work, we demonstrate the differential roles of a given protein corona formed in cell culture media in particle uptake by monocytes and macrophages. By exposing disulfide-stabilized poly(methacrylic acid) nanoporous polymer particles (PMASH NPPs) to complete cell growth media containing 10% fetal bovine serum, a protein corona, with the most abundant component being bovine serum albumin, was characterized. Upon adsorption onto the PMASH NPPs, native bovine serum albumin (BSA) was found to undergo conformational changes. The denatured BSA led to a significant decrease in internalization efficiency in human monocytic cells, THP-1, compared with the bare particles, due to reduced cell membrane adhesion. In contrast, the unfolded BSA on the NPPs triggered class A scavenger receptor-mediated phagocytosis in differentiated macrophage-like cells (dTHP-1) without a significant impact on the overall internalization efficiency. Taken together, this work demonstrates the disparate effects of a given protein corona on particle-cell interactions, highlighting the correlation between protein corona conformation in situ and relevant biological characteristics for biological functionalities.

  20. Infiltration Pattern of Blood Monocytes into the Central Nervous System during Experimental Herpes Simplex Virus Encephalitis.

    Directory of Open Access Journals (Sweden)

    Rafik Menasria

    Full Text Available The kinetics and distribution of infiltrating blood monocytes into the central nervous system and their involvement in the cerebral immune response together with resident macrophages, namely microglia, were evaluated in experimental herpes simplex virus 1 (HSV-1 encephalitis (HSE. To distinguish microglia from blood monocyte-derived macrophages, chimeras were generated by conditioning C57BL/6 recipient mice with chemotherapy regimen followed by transplantation of bone morrow-derived cells that expressed the green fluorescent protein. Mice were infected intranasally with a sub-lethal dose of HSV-1 (1.2 x 10(6 plaque forming units. Brains were harvested prior to and on days 4, 6, 8 and 10 post-infection for flow cytometry and immunohistochemistry analysis. The amounts of neutrophils (P < 0.05 and "Ly6C hi" inflammatory monocytes (P < 0.001 significantly increased in the CNS compared to non-infected controls on day 6 post-infection, which corresponded to more severe clinical signs of HSE. Levels decreased on day 8 for both leukocytes subpopulations (P < 0.05 for inflammatory monocytes compared to non-infected controls to reach baseline levels on day 10 following infection. The percentage of "Ly6C low" patrolling monocytes significantly increased (P < 0.01 at a later time point (day 8, which correlated with the resolution phase of HSE. Histological analysis demonstrated that blood leukocytes colonized mostly the olfactory bulb and the brainstem, which corresponded to regions where HSV-1 particles were detected. Furthermore, infiltrating cells from the monocytic lineage could differentiate into activated local tissue macrophages that express the microglia marker, ionized calcium-binding adaptor molecule 1. The lack of albumin detection in the brain parenchyma of infected mice showed that the infiltration of blood leukocytes was not necessarily related to a breakdown of the blood-brain barrier but could be the result of a functional recruitment. Thus

  1. Infiltration Pattern of Blood Monocytes into the Central Nervous System during Experimental Herpes Simplex Virus Encephalitis

    Science.gov (United States)

    Menasria, Rafik; Canivet, Coraline; Piret, Jocelyne; Boivin, Guy

    2015-01-01

    The kinetics and distribution of infiltrating blood monocytes into the central nervous system and their involvement in the cerebral immune response together with resident macrophages, namely microglia, were evaluated in experimental herpes simplex virus 1 (HSV-1) encephalitis (HSE). To distinguish microglia from blood monocyte-derived macrophages, chimeras were generated by conditioning C57BL/6 recipient mice with chemotherapy regimen followed by transplantation of bone morrow-derived cells that expressed the green fluorescent protein. Mice were infected intranasally with a sub-lethal dose of HSV-1 (1.2x106 plaque forming units). Brains were harvested prior to and on days 4, 6, 8 and 10 post-infection for flow cytometry and immunohistochemistry analysis. The amounts of neutrophils (P<0.05) and «Ly6Chi» inflammatory monocytes (P<0.001) significantly increased in the CNS compared to non-infected controls on day 6 post-infection, which corresponded to more severe clinical signs of HSE. Levels decreased on day 8 for both leukocytes subpopulations (P<0.05 for inflammatory monocytes compared to non-infected controls) to reach baseline levels on day 10 following infection. The percentage of «Ly6Clow» patrolling monocytes significantly increased (P<0.01) at a later time point (day 8), which correlated with the resolution phase of HSE. Histological analysis demonstrated that blood leukocytes colonized mostly the olfactory bulb and the brainstem, which corresponded to regions where HSV-1 particles were detected. Furthermore, infiltrating cells from the monocytic lineage could differentiate into activated local tissue macrophages that express the microglia marker, ionized calcium-binding adaptor molecule 1. The lack of albumin detection in the brain parenchyma of infected mice showed that the infiltration of blood leukocytes was not necessarily related to a breakdown of the blood-brain barrier but could be the result of a functional recruitment. Thus, our findings suggest

  2. Modulation of neutrophil and monocyte function by recombinant human granulocyte macrophage colony-stimulating factor in patients with lymphoma.

    Science.gov (United States)

    Kharazmi, A; Nielsen, H; Hovgaard, D; Borregaard, N; Nissen, N I

    1991-04-01

    Granulocyte macrophage colony-stimulating factor (GM-CSF) has been shown to inhibit the chemotaxis and enhance the oxidative burst response of human neutrophils in vitro. The present study describes the effect of recombinant GM-CSF on the neutrophil and monocyte function in patients with lymphoma undergoing GM-CSF treatment. Patients with either Hodgkin's or non-Hodgkin's lymphoma were treated with various dosages (2-16 micrograms kg-1 body weight per day for 5 days) of rhGM-CSF by intravenous or subcutaneous route. Prior to and on day 5 of rhGM-CSF treatment, neutrophil and monocyte chemotaxis and chemiluminescence responses to f-Met-Leu-Phe, zymosan activated serum (ZAS) and opsonized zymosan (OZ) were determined. It was observed that chemotactic response of neutrophils to f-Met-Leu-Phe and ZAS was reduced, whereas the chemiluminescence response of both cell types to f-Met-Leu-Phe and zymosan was enhanced by up to 43-fold. rhGM-CSF treatment did not affect degranulation of the neutrophils as measured by release of vitamin B12 binding protein. Degree of modulation of neutrophil and monocyte function by rhGM-CSF was independent of rhGM-CSF dosages administered. These data suggest that phagocytic defence system may be enhanced by GM-CSF treatment and that this cytokine may be a useful therapeutic adjunct in compromised patients.

  3. Constitutive activity of NF-kappa B in myeloid cells drives pathogenicity of monocytes and macrophages during autoimmune neuroinflammation

    Directory of Open Access Journals (Sweden)

    Ellrichmann Gisa

    2012-01-01

    Full Text Available Abstract The NF-κB/REL-family of transcription factors plays a central role in coordinating the expression of a wide variety of genes controlling immune responses including autoimmunity of the central nervous system (CNS. The inactive form of NF-κB consists of a heterodimer which is complexed with its inhibitor, IκB. Conditional knockout-mice for IκBα in myeloid cells (lysMCreIκBαfl/fl have been generated and are characterized by a constitutive activation of NF-κB proteins allowing the study of this transcription factor in myelin-oligodendrocyte-glycoprotein induced experimental autoimmune encephalomyelitis (MOG-EAE, a well established experimental model for autoimmune demyelination of the CNS. In comparison to controls, lysMCreIκBαfl/fl mice developed a more severe clinical course of EAE. Upon histological analysis on day 15 p.i., there was an over two fold increased infiltration of T-cells and macrophages/microglia. In addition, lysMCreIκBαfl/fl mice displayed an increased expression of the NF-κB dependent factor inducible nitric oxide synthase in inflamed lesions. These changes in the CNS are associated with increased numbers of CD11b positive splenocytes and a higher expression of Ly6c on monocytes in the periphery. Well in accordance with these changes in the myeloid cell compartment, there was an increased production of the monocyte cytokines interleukin(IL-12 p70, IL-6 and IL-1beta in splenocytes. In contrast, production of the T-cell associated cytokines interferon gamma (IFN-gamma and IL-17 was not influenced. In summary, myeloid cell derived NF-κB plays a crucial role in autoimmune inflammation of the CNS and drives a pathogenic role of monocytes and macrophages independently from T-cells.

  4. Saturated fatty acids activate caspase-4/5 in human monocytes, triggering IL-1β and IL-18 release.

    Science.gov (United States)

    Pillon, Nicolas J; Chan, Kenny L; Zhang, Shitian; Mejdani, Marios; Jacobson, Maya R; Ducos, Alexandre; Bilan, Philip J; Niu, Wenyan; Klip, Amira

    2016-11-01

    Obesity is associated with metabolic tissue infiltration by monocyte-derived macrophages. Saturated fatty acids contribute to proinflammatory gene induction in tissue-embedded immune cells. However, it is unknown how circulating monocytes, the macrophage precursors, react to high-fat environments. In macrophages, saturated fatty acids activate inflammatory pathways and, notably, prime caspase-associated inflammasomes. Inflammasome-activated IL-1β contributes to type 2 diabetes. We hypothesized that 1) human monocytes from obese patients show caspase activation, and 2) fatty acids trigger this response and consequent release of IL-1β/IL-18. Human peripheral blood monocytes were sorted by flow cytometry, and caspase activity was measured with a FLICA dye-based assay. Blood monocytes from obese individuals exhibited elevated caspase activity. To explore the nature and consequence of this activity, human THP1 monocytes were exposed to saturated or unsaturated fatty acids. Caspase activity was revealed by isoform-specific cleavage and enzymatic activity; cytokine expression/release was measured by qPCR and ELISA. Palmitate, but not palmitoleate, increased caspase activity in parallel to the release of IL-1β and IL-18. Palmitate induced eventual monocyte cell death with features of pyroptosis (an inflammation-linked cell death program involving caspase-4/5), scored through LDH release, vital dye influx, cell volume changes, and nuclear morphology. Notably, selective gene silencing or inhibition of caspase-4/5 reduced palmitate-induced release of IL-1β and IL-18. In summary, monocytes from obese individuals present elevated caspase activity. Mechanistically, palmitate activates a pyroptotic program in monocytes through caspase-4/5, causing inflammatory cytokine release, additional to inflammasomes. These caspases represent potential, novel, therapeutic targets to taper obesity-associated inflammation.

  5. Transcription of innate immunity genes and cytokine secretion by canine macrophages resistant or susceptible to intracellular survival of Leishmania infantum.

    Science.gov (United States)

    Turchetti, Andréia Pereira; da Costa, Luciana Fachini; Romão, Everton de Lima; Fujiwara, Ricardo Toshio; da Paixão, Tatiane Alves; Santos, Renato Lima

    2015-01-15

    In this study we assessed the basal transcription of genes associated with innate immunity (i.e. Nramp1, NOD1, NOD2, TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, and TLR9) in canine monocyte-derived macrophages from Leishmania-free dogs. Additionally, secretion of cytokines (IL-10, IL-12, TNF-α and IFN-γ) and nitric oxide in culture supernatants of macrophages with higher or lower resistance to intracellular survival of Leishmania infantum was also measured. Constitutive transcription of TLR9 and NOD2 were negligible; NOD1, TLR1, and TLR7 had low levels of transcription, whereas Nramp1 and TLR2, 3, 4, 5, and 6 had higher levels of constitutive transcription in canine monocyte-derived macrophages. There were no significant differences in transcription between macrophages with higher or lower resistance to intracellular survival of L. infantum. Secretion of TNF-α was higher in more resistant macrophages (designated as resistant) at 24h after infection when compared to less resistant macrophages (designated as susceptible), as well as the secretion of IFN-γ at 72 h post infection. Secretion of IL-10 was lower in resistant macrophages at 24h after infection. No detectable production of nitric oxide was observed. Interestingly, there was a negative correlation between NOD2 transcript levels and intracellular survival of L. infantum in resistant macrophages. This study demonstrated that decreased intracellular survival of L. infantum in canine macrophages was associated with increased production of TNF-α and IFN-γ and decreased production of IL-10; and that constitutive transcription of Nramp1, TLR and NLR does not interfere in intracellular survival of L. infantum.

  6. DMPD: The oxidation of lipoproteins by monocytes-macrophages. Biochemical andbiological mechanisms. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available logical mechanisms. Chisolm GM 3rd, Hazen SL, Fox PL, Cathcart MK. J Biol Chem. 1999 Sep 10;274(37):25959-62...onocytes-macrophages. Biochemical andbiological mechanisms. Authors Chisolm GM 3rd, Hazen SL, Fox PL, Cathcart

  7. Monocyte Trafficking, Engraftment, and Delivery of Nanoparticles and an Exogenous Gene into the Acutely Inflamed Brain Tissue - Evaluations on Monocyte-Based Delivery System for the Central Nervous System.

    Directory of Open Access Journals (Sweden)

    Hsin-I Tong

    Full Text Available The ability of monocytes and monocyte-derived macrophages (MDM to travel towards chemotactic gradient, traverse tissue barriers, and accumulate precisely at diseased sites makes them attractive candidates as drug carriers and therapeutic gene delivery vehicles targeting the brain, where treatments are often hampered by the blockade of the blood brain barrier (BBB. This study was designed to fully establish an optimized cell-based delivery system using monocytes and MDM, by evaluating their homing efficiency, engraftment potential, as well as carriage and delivery ability to transport nano-scaled particles and exogenous genes into the brain, following the non-invasive intravenous (IV cell adoptive transfer in an acute neuroinflammation mouse model induced by intracranial injection of Escherichia coli lipopolysaccharides. We demonstrated that freshly isolated monocytes had superior inflamed-brain homing ability over MDM cultured in the presence of macrophage colony stimulating factor. In addition, brain trafficking of IV infused monocytes was positively correlated with the number of adoptive transferred cells, and could be further enhanced by transient disruption of the BBB with IV administration of Mannitol, Bradykinin or Serotonin right before cell infusion. A small portion of transmigrated cells was detected to differentiate into IBA-1 positive cells with microglia morphology in the brain. Finally, with the use of superparamagnetic iron oxide nanoparticles SHP30, the ability of nanoscale agent-carriage monocytes to enter the inflamed brain region was validated. In addition, lentiviral vector DHIV-101 was used to introduce green fluorescent protein (GFP gene into monocytes, and the exogenous GFP gene was detected in the brain at 48 hours following IV infusion of the transduced monocytes. All together, our study has set up the optimized conditions for the more-in-depth tests and development of monocyte-mediated delivery, and our data supported

  8. Macrophage colony-stimulating factor (CSF1) controls monocyte production and maturation and the steady-state size of the liver in pigs.

    Science.gov (United States)

    Sauter, Kristin A; Waddell, Lindsey A; Lisowski, Zofia M; Young, Rachel; Lefevre, Lucas; Davis, Gemma M; Clohisey, Sara M; McCulloch, Mary; Magowan, Elizabeth; Mabbott, Neil A; Summers, Kim M; Hume, David A

    2016-09-01

    Macrophage colony-stimulating factor (CSF1) is an essential growth and differentiation factor for cells of the macrophage lineage. To explore the role of CSF1 in steady-state control of monocyte production and differentiation and tissue repair, we previously developed a bioactive protein with a longer half-life in circulation by fusing pig CSF1 with the Fc region of pig IgG1a. CSF1-Fc administration to pigs expanded progenitor pools in the marrow and selectively increased monocyte numbers and their expression of the maturation marker CD163. There was a rapid increase in the size of the liver, and extensive proliferation of hepatocytes associated with increased macrophage infiltration. Despite the large influx of macrophages, there was no evidence of liver injury and no increase in circulating liver enzymes. Microarray expression profiling of livers identified increased expression of macrophage markers, i.e., cytokines such as TNF, IL1, and IL6 known to influence hepatocyte proliferation, alongside cell cycle genes. The analysis also revealed selective enrichment of genes associated with portal, as opposed to centrilobular regions, as seen in hepatic regeneration. Combined with earlier data from the mouse, this study supports the existence of a CSF1-dependent feedback loop, linking macrophages of the liver with bone marrow and blood monocytes, to mediate homeostatic control of the size of the liver. The results also provide evidence of safety and efficacy for possible clinical applications of CSF1-Fc.

  9. Regulation of nutrition-associated receptors in blood monocytes of normal weight and obese humans.

    Science.gov (United States)

    Pivovarova, Olga; Hornemann, Silke; Weimer, Sandra; Lu, Ye; Murahovschi, Veronica; Zhuk, Sergei; Seltmann, Anne-Cathrin; Malashicheva, Anna; Kostareva, Anna; Kruse, Michael; Busjahn, Andreas; Rudovich, Natalia; Pfeiffer, Andreas F H

    2015-03-01

    Obesity, type 2 diabetes and associated metabolic diseases are characterized by low-grade systemic inflammation which involves interplay of nutrition and monocyte/macrophage functions. We suggested that some factors such as nutrient components, neuropeptides involved in the control of gastrointestinal functions, and gastrointestinal hormones might influence immune cell functions and in this way contribute to the disease pathogenesis. The aim of this study was to investigate the mRNA expression of twelve nutrition-associated receptors in peripheral blood mononuclear cells (PBMC), isolated monocytes and monocyte-derived macrophages and their regulation under the switching from the high-carbohydrate low-fat diet to the low-carbohydrate high-fat (LC/HFD) isocaloric diet in healthy humans. The mRNA expression of receptors for short chain fatty acids (GPR41, GPR43), bile acids (TGR5), incretins (GIPR, GLP1R), cholecystokinin (CCKAR), neuropeptides VIP and PACAP (VIPR1, VIPR2), and neurotensin (NTSR1) was detected in PBMC and monocytes, while GPR41, GPR43, GIPR, TGR5, and VIPR1 were found in macrophages. Correlations of the receptor expression in monocytes with a range of metabolic and inflammatory markers were found. In non-obese subjects, the dietary switch to LC/HFD induced the increase of GPR43 and VIPR1 expression in monocytes. No significant differences of receptor expression between normal weight and moderately obese subjects were found. Our study characterized for the first time the expression pattern of nutrition-associated receptors in human blood monocytes and its dietary-induced changes linking metabolic responses to nutrition with immune functions in health and metabolic diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. Suppression of cell division-associated genes by Helicobacter pylori attenuates proliferation of RAW264.7 monocytic macrophage cells.

    Science.gov (United States)

    Tan, Grace Min Yi; Looi, Chung Yeng; Fernandez, Keith Conrad; Vadivelu, Jamuna; Loke, Mun Fai; Wong, Won Fen

    2015-06-16

    Helicobacter pylori at multiplicity of infection (MOI ≥ 50) have been shown to cause apoptosis in RAW264.7 monocytic macrophage cells. Because chronic gastric infection by H. pylori results in the persistence of macrophages in the host's gut, it is likely that H. pylori is present at low to moderate, rather than high numbers in the infected host. At present, the effect of low-MOI H. pylori infection on macrophage has not been fully elucidated. In this study, we investigated the genome-wide transcriptional regulation of H. pylori-infected RAW264.7 cells at MOI 1, 5 and 10 in the absence of cellular apoptosis. Microarray data revealed up- and down-regulation of 1341 and 1591 genes, respectively. The expression of genes encoding for DNA replication and cell cycle-associated molecules, including Aurora-B kinase (AurkB) were down-regulated. Immunoblot analysis verified the decreased expression of AurkB and downstream phosphorylation of Cdk1 caused by H. pylori infection. Consistently, we observed that H. pylori infection inhibited cell proliferation and progression through the G1/S and G2/M checkpoints. In summary, we suggest that H. pylori disrupts expression of cell cycle-associated genes, thereby impeding proliferation of RAW264.7 cells, and such disruption may be an immunoevasive strategy utilized by H. pylori.

  11. Recombinant glycoprotein 63 (Gp63) of Trypanosoma carassii suppresses antimicrobial responses of goldfish (Carassius auratus L.) monocytes and macrophages.

    Science.gov (United States)

    Oladiran, Ayoola; Belosevic, Miodrag

    2012-06-01

    We previously reported that proteins secreted by Trypanosoma carassii play a role in evasion of fish host immune responses. To further understand how these parasites survive in the host, we cloned and expressed T. carassii glycoprotein 63 (Tcagp63), and generated a rabbit polyclonal antibody to the recombinant protein (rTcagp63). Tcagp63 was similar to gp63 of other trypanosomes and grouped with Trypanosoma cruzi and Trypanosoma brucei gp63 in phylogenetic analysis. We showed that rTcagp63 down-regulated Aeromonas salmonicida and recombinant goldfish TNFα2-induced production of reactive oxygen and nitrogen intermediates. Macrophages treated with rTcagp63 also exhibited significant reduction in the expression of inducible nitric oxide synthase (iNOS)-A, TNFα-1 and TNFα-2. Recombinant Tcagp63 bound to and was internalised by goldfish macrophages. The Tcagp63 may act by altering the signalling events important in downstream monocyte/macrophage antimicrobial and other cytokine-induced functions. We believe that this is the first report on downregulation of antimicrobial responses by trypanosome gp63.

  12. Granulocyte macrophage colony stimulating factor is elevated in alveolar macrophages from sheep naturally infected with maedi-visna virus and stimulates maedi-visna virus replication in macrophages in vitro.

    Science.gov (United States)

    Zhang, Z; Harkiss, G D; Hopkins, J; Woodall, C J

    2002-08-01

    Infection by maedi-visna virus, a lentivirus of sheep, leads to chronic inflammatory reactions of various tissues. In this report we have analysed the role of specific cytokines in the disease process. A significant increase in expression of interleukin-6, interleukin-10, granulocyte macrophage-colony stimulating factor (GM-CSF) and transforming growth factor-beta1 mRNA was observed in alveolar macrophages isolated from the lungs of naturally infected animals when compared with lungs of seronegative controls. Levels of GM-CSF mRNA expression in alveolar macrophages correlated with the presence of lung lesions, but there was no correlation of interleukin-10, interleukin-6, tumour necrosis factor-alpha and transforming growth factor-beta1 mRNA levels in alveolar macrophages from animals with pulmonary lesions. In vitro investigation showed that GM-CSF in the range 0.1-10 ng/ml induced a significant increase in viral p25 production after 7 days in acutely infected blood monocyte-derived macrophages. The production of p25 peaked between 7 and 14 days exposure to 10 ng/ml of GM-CSF. Quantitative polymerase chain reaction showed that the level of viral DNA in monocyte-derived macrophages was dose-dependent following GM-CSF treatment in the range 0.1-100 ng/ml after 7 days. Viral mRNA expression was also enhanced. These findings indicate a role for GM-CSF in the pathogenesis of lymphoid interstitial pneumonia in infected animals.

  13. Bole of macrophage colony-stimulating factor in the differentiation and expansion of monocytes and dendritic cells from CD34(+) progenitor cells

    NARCIS (Netherlands)

    Kamps, AWA; Smit, JW; Vellenga, E

    1999-01-01

    The present study focused on whether it is possible to expand monocytic cells from CD34(+) progenitor cells by using macrophage colony-stimulating factor (M-CSF) in the absence and presence of mast cell growth factor (MGF) and IL-6. It was demonstrated that CD34(+) cells differentiate without expans

  14. Monocyte differentiation in localized juvenile periodontitis is skewed toward the dendritic cell phenotype.

    Science.gov (United States)

    Barbour, Suzanne E; Ishihara, Yuichi; Fakher, Mohammed; Al-Darmaki, Salma; Caven, Timothy H; Shelburne, C P; Best, Al M; Schenkein, Harvey A; Tew, John G

    2002-06-01

    Several lines of evidence indicate that the monocytes of subjects with localized juvenile periodontitis (LJP) are functionally distinct from cells of age- and race-matched nonperiodontitis (NP) subjects. Among the abnormalities are the propensity to secrete large amounts of prostaglandin E(2) and the induction of immunoglobulin G2 (IgG2) antibodies. The experiments described here were performed to further characterize the LJP monocytes and to determine if these cells mature differently than NP monocytes. When adherent monocytes from LJP subjects were cultured in the presence of human serum, both macrophages and cells with the morphology of immature monocyte-derived dendritic cells (MDDC) were observed. Within 4 days the prevalence of the immature MDDC was approximately twofold higher in LJP cultures than in NP cultures. In addition to their dendritic morphology, these cells were CD11c(+) and CD14(-) or CD14(low) and stimulated potent autologous mixed leukocyte reactions, consistent with differentiation to the MDDC phenotype. Like LJP monocytes, cultures of MDDC generated with interleukin-4 and granulocyte-macrophage colony-stimulating factor selectively induced IgG2 in cultures of pokeweed mitogen-stimulated NP leukocytes. Together, these data suggest that the monocytes of LJP subjects have a propensity to differentiate into MDDC and that this differentiation may be related to the high levels of IgG2 that are observed in the sera of LJP subjects. As high levels of circulating IgG2 are correlated with less severe disease, the propensity of LJP monocytes to differentiate into MDDC may have important implications for both the host response against oral pathogens and the progression of LJP.

  15. Human macrophage responses to clinical isolates from the Mycobacterium tuberculosis complex discriminate between ancient and modern lineages.

    Directory of Open Access Journals (Sweden)

    Damien Portevin

    2011-03-01

    Full Text Available The aim of the present study was to determine whether there is a correlation between phylogenetic relationship and inflammatory response amongst a panel of clinical isolates representative of the global diversity of the human Mycobacterium tuberculosis Complex (MTBC. Measurement of cytokines from infected human peripheral blood monocyte-derived macrophages revealed a wide variation in the response to different strains. The same pattern of high or low response to individual strains was observed for different pro-inflammatory cytokines and chemokines, and was conserved across multiple human donors. Although each major phylogenetic lineage of MTBC included strains inducing a range of cytokine responses, we found that overall inflammatory phenotypes differed significantly across lineages. In particular, comparison of evolutionarily modern lineages demonstrated a significant skewing towards lower early inflammatory response. The differential response to ancient and modern lineages observed using GM-CSF derived macrophages was also observed in autologous monocyte-derived dendritic cells and murine bone marrow-derived macrophages, but not in human unfractionated peripheral blood mononuclear cells. We hypothesize that the reduced immune responses to modern lineages contribute to more rapid disease progression and transmission, which might be a selective advantage in the context of expanding human populations. In addition to the lineage effects, the large strain-to-strain variation in innate immune responses elicited by MTBC will need to be considered in tuberculosis vaccine development.

  16. Translation of Pur-α is targeted by cellular miRNAs to modulate the differentiation-dependent susceptibility of monocytes to HIV-1 infection.

    Science.gov (United States)

    Shen, Chan-Juan; Jia, Yan-Hui; Tian, Ren-Rong; Ding, Ming; Zhang, Chiyu; Wang, Jian-Hua

    2012-11-01

    The postentry restriction of HIV-1 replication in monocytes can be relieved when they differentiate to dendritic cells (DCs) or macrophages. Multiple mechanisms have been proposed to interpret the differentiation-dependent susceptibility of monocytes to HIV-1 infection, and the absence of host-cell-encoded essential factors for HIV-1 completing the life cycle may provide an explanation. We have analyzed the gene expression profile in monocytes by mRNA microarray and compared it with that of differentiated DCs. We demonstrated that purine-rich element binding protein α (Pur-α), a host-cell-encoded ubiquitous, sequence-specific DNA- and RNA-binding protein, showed inadequate expression in monocytes, and the translation of Pur-α mRNA was repressed by cell-expressed microRNA (miRNA). These Pur-α-targeted miRNAs modulated the differentiation-dependent susceptibility of monocytes/DCs to HIV-1 infection, because rescue of Pur-α expression by transfection of miRNA inhibitors relieved the restriction of HIV-1 infection in monocytes, and ectopic input of miRNA mimics significantly reduced HIV-1 infection of monocyte-derived DCs (MDDCs). Collectively, our data emphasized that inadequate host factors contribute to HIV-1 restriction in monocytes, and cellular miRNAs modulate differentiation-dependent susceptibility of host cells to HIV-1 infection. Elaboration of HIV-1 restriction in host cells facilitates our understanding of viral pathogenesis and the search for a new antiviral strategy.

  17. The MAPK ERK5, but not ERK1/2, inhibits the progression of monocytic phenotype to the functioning macrophage

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Xuening [Department of Pathology and Laboratory Medicine, Rutgers, NJ Medical School, 185 South Orange Ave, Newark, NJ 07103 (United States); Pesakhov, Stella [Department of Clinical Biochemistry and Pharmacology, Faculty of Health Sciences, Ben-Gurion University of the Negev, PO Box 653, 84105 Beer-Sheva (Israel); Harrison, Jonathan S [Department of Medicine, Rutgers, Robert Wood Johnson Medical School, New Brunswick, NJ 08903 (United States); Kafka, Michael; Danilenko, Michael [Department of Clinical Biochemistry and Pharmacology, Faculty of Health Sciences, Ben-Gurion University of the Negev, PO Box 653, 84105 Beer-Sheva (Israel); Studzinski, George P, E-mail: studzins@njms.rutgers.edu [Department of Pathology and Laboratory Medicine, Rutgers, NJ Medical School, 185 South Orange Ave, Newark, NJ 07103 (United States)

    2015-01-01

    Intracellular signaling pathways present targets for pharmacological agents with potential for treatment of neoplastic diseases, with some disease remissions already recorded. However, cellular compensatory mechanisms usually negate the initial success. For instance, attempts to interrupt aberrant signaling downstream of the frequently mutated ras by inhibiting ERK1/2 has shown only limited usefulness for cancer therapy. Here, we examined how ERK5, that overlaps the functions of ERK1/2 in cell proliferation and survival, functions in a manner distinct from ERK1/2 in human AML cells induced to differentiate by 1,25D-dihydroxyvitamin D{sub 3} (1,25D). Using inhibitors of ERK1/2 and of MEK5/ERK5 at concentrations specific for each kinase in HL60 and U937 cells, we observed that selective inhibition of the kinase activity of ERK5, but not of ERK1/2, in the presence of 1,25D resulted in macrophage-like cell morphology and enhancement of phagocytic activity. Importantly, this was associated with increased expression of the macrophage colony stimulating factor receptor (M-CSFR), but was not seen when M-CSFR expression was knocked down. Interestingly, inhibition of ERK1/2 led to activation of ERK5 in these cells. Our results support the hypothesis that ERK5 negatively regulates the expression of M-CSFR, and thus has a restraining function on macrophage differentiation. The addition of pharmacological inhibitors of ERK5 may influence trials of differentiation therapy of AML. - Highlights: • ERK5 has at least some functions in AML cells which are distinct from those of ERK1/2. • ERK5 activity negatively controls the expression of M-CSFR. • ERK5 retards the progression of differentiation from monocyte to functional macrophage.

  18. Role of CC chemokines (macrophage inflammatory protein-1 beta, monocyte chemoattractant protein-1, RANTES) in acute lung injury in rats

    DEFF Research Database (Denmark)

    Bless, N M; Huber-Lang, M; Guo, R F

    2000-01-01

    were cloned, the proteins were expressed, and neutralizing Abs were developed. mRNA and protein expression for MIP-1 beta and MCP-1 were up-regulated during the inflammatory response, while mRNA and protein expression for RANTES were constitutive and unchanged during the inflammatory response...... that in chemokine-dependent inflammatory responses in lung CC chemokines do not necessarily demonstrate redundant function.......The role of the CC chemokines, macrophage inflammatory protein-1 beta (MIP-1 beta), monocyte chemotactic peptide-1 (MCP-1), and RANTES, in acute lung inflammatory injury induced by intrapulmonary deposition of IgG immune complexes injury in rats was determined. Rat MIP-1 beta, MCP-1, and RANTES...

  19. Pathogenic bacteria and TNF do not induce production of macrophage migration inhibitory factor (MIF) by human monocytes.

    Science.gov (United States)

    Temple, Suzanna E L; Cheong, Karey Y; Price, Patricia; Waterer, Grant W

    2009-06-01

    Elevated serum macrophage migration inhibitory factor (MIF) is associated with severe sepsis, but it is not clear whether bacteria stimulate synthesis of MIF by blood leukocytes directly or via induction of TNF. Here we assess production of MIF mRNA and protein by blood leukocytes from healthy human subjects (n=28) following exposure to bacteria commonly associated with sepsis (Escherichia coli and Streptococcus pneumoniae). Bacteria did not increase levels of MIF mRNA or secreted protein. CD14(+) monocytes were the main cell type producing MIF before and after stimulation. Exposure of leukocytes to TNF did not induce MIF. Hence elevated levels of serum MIF observed in sepsis may not reflect MIF produced by blood leukocytes stimulated directly by bacteria or TNF.

  20. Unlike PPAR{gamma}, PPAR{alpha} or PPAR{beta}/{delta} activation does not promote human monocyte differentiation toward alternative macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Bouhlel, Mohamed Amine [Univ Lille Nord de France, F-59000 Lille (France); Inserm U545, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France); Brozek, John [Genfit, Loos (France); Derudas, Bruno [Univ Lille Nord de France, F-59000 Lille (France); Inserm U545, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France); Zawadzki, Christophe; Jude, Brigitte [Inserm ERI-9 and Equipe d' Accueil 2693, IFR114, Universite de Lille, Lille (France); Staels, Bart, E-mail: bart.staels@pasteur-lille.fr [Univ Lille Nord de France, F-59000 Lille (France); Inserm U545, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France); Chinetti-Gbaguidi, Giulia [Univ Lille Nord de France, F-59000 Lille (France); Inserm U545, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France)

    2009-08-28

    Macrophages adapt their response to micro-environmental signals. While Th1 cytokines promote pro-inflammatory M1 macrophages, Th2 cytokines promote an 'alternative' anti-inflammatory M2 macrophage phenotype. Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors expressed in macrophages where they control the inflammatory response. It has been shown that PPAR{gamma} promotes the differentiation of monocytes into anti-inflammatory M2 macrophages in humans and mice, while a role for PPAR{beta}/{delta} in this process has been reported only in mice and no data are available for PPAR{alpha}. Here, we show that in contrast to PPAR{gamma}, expression of PPAR{alpha} and PPAR{beta}/{delta} overall does not correlate with the expression of M2 markers in human atherosclerotic lesions, whereas a positive correlation with genes of lipid metabolism exists. Moreover, unlike PPAR{gamma}, PPAR{alpha} or PPAR{beta}/{delta} activation does not influence human monocyte differentiation into M2 macrophages in vitro. Thus, PPAR{alpha} and PPAR{beta}/{delta} do not appear to modulate the alternative differentiation of human macrophages.

  1. 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.

  2. Support of HUVEC proliferation by pro-angiogenic intermediate CD163+ monocytes/macrophages: a co-culture experiment.

    Science.gov (United States)

    Mayer, A; Hiebl, B; Lendlein, A; Jung, F

    2011-01-01

    So called intermediate (MO2) monocytes/macrophages possess anti-inflammatory properties and express the MO lineage marker CD163. On a hydrophilic, acrylamide-based hydrogel human intermediate (CD14++ CD16+) CD163++ monocytes/macrophages (aMO2) which were angiogenically stimulated, maintained a pro-angiogenic and non-inflammatory status for at least 14 days. Here we explored, whether this aMO2 subset can positively influence the proliferation of human umbilical venous endothelial cells (HUVECs) without switching back into a pro-inflammatory (MO1) phenotype. aMO2 or HUVEC were seeded alone on glass cover slips (0.5 × 10(5) cells / 1.33 cm(2)) in a HUVEC specific cell culture medium (EGM-2) for 3 hrs, 24 hrs and 72 hrs or under co-culture conditions (0.5 × 10(5) HUVEC + 0.25 × 10(5) aMO2 / 1.33 cm(2)) in EGM-2 for the same time window as well (n = 6 each). Under co-culture conditions the numbers of adherent HUVEC per unit area were significantly higher (p HUVEC/mm(2)) compared to control mono-cultures (473 ± 76 HUVEC/mm(2)) after 72 hrs of cultivation and showed their typically spread morphology. The aMO2 remained in their subset status and secreted VEGF-A165 without release of pro-inflammatory cytokines until the end of the 72 hrs cultivation time period, thereby supporting the HUVEC proliferation. These in vitro results might indicate that this MO subset can be used as cellular delivery system for pro-angiogenic and non-inflammatory mediators to support the endothelialisation of biomaterials like e.g. cPnBA.

  3. Transmembrane oligomeric form of Vibrio cholerae cytolysin triggers TLR2/TLR6-dependent proinflammatory responses in monocytes and macrophages.

    Science.gov (United States)

    Khilwani, Barkha; Mukhopadhaya, Arunika; Chattopadhyay, Kausik

    2015-02-15

    Vibrio cholerae cytolysin (VCC) kills target eukaryotic cells by forming transmembrane oligomeric β-barrel pores. Once irreversibly converted into the transmembrane oligomeric form, VCC acquires an unusual structural stability and loses its cytotoxic property. It is therefore possible that, on exertion of its cytotoxic activity, the oligomeric form of VCC retained in the disintegrated membrane fractions of the lysed cells would survive within the host cellular milieu for a long period, without causing any further cytotoxicity. Under such circumstances, VCC oligomers may potentially be recognized by the host immune cells. Based on such a hypothesis, in the present study we explored the interaction of the transmembrane oligomeric form of VCC with the monocytes and macrophages of the innate immune system. Our study shows that the VCC oligomers assembled in the liposome membranes elicit potent proinflammatory responses in monocytes and macrophages, via stimulation of the toll-like receptor (TLR)2/TLR6-dependent signalling cascades that involve myeloid differentiation factor 88 (MyD88)/interleukin-1-receptor-associated kinase (IRAK)1/tumour-necrosis-factor-receptor-associated factor (TRAF)6. VCC oligomer-mediated proinflammatory responses critically depend on the activation of the transcription factor nuclear factor-κB. Proinflammatory responses induced by the VCC oligomers also require activation of the mitogen-activated protein kinase (MAPK) family member c-Jun N-terminal kinase, which presumably acts via stimulation of the transcription factor activator protein-1. Notably, the role of the MAPK p38 could not be documented in the process.

  4. Cytometric analysis of surface molecules of leucocytes and phagocytic activity of granulocytes and monocytes/macrophages in cows with pyometra.

    Science.gov (United States)

    Brodzki, P; Kostro, K; Brodzki, A; Niemczuk, K; Lisiecka, U

    2014-10-01

    Pyometra is a serious problem in dairy cow herds, causing large economic losses due to infertility. The development of pyometra depends mainly on the immunological status of the cow. The aim of the study was a comparative evaluation of selected indicators involving non-specific and specific immunity in cows with pyometra and in cows without inflammation of the uterus. The study was performed in 20 cows, which were divided into two groups: pyometra group and healthy group, each comprising 10 cows, based on the results of cytological and ultrasonographic tests. A flow cytometric analysis was performed for the surface molecules CD4, CD8, CD14, CD21, CD25 and CD4(+) CD25(+) on leucocytes, and the phagocytic activity was determined from granulocytes and monocytes/macrophages in the peripheral blood and uterine washings, respectively. It was demonstrated that the percentage of phagocytic granulocytes and monocytes/macrophages in both the peripheral blood and uterine washings was significantly lower in cows with pyometra compared with the healthy group (p < 0.001). Significantly (p ≤ 0.001) lower percentage of CD4(+) , CD14(+) , CD25(+) and CD4(+) CD25(+) phenotype leucocytes was also observed in the peripheral blood of cows from the pyometra group, along with a significantly higher (p < 0.001) percentage of CD8(+) and CD21(+) lymphocytes as compared to the healthy group. The results of work indicate that disfunction of cell immunity coexisting with pyometra may be caused by a bacterial infection and the presence of blocking agents (IL-10), released by the increasing number of CD8(+) lymphocytes what leads to the advanced inflammation of uterus.

  5. C-Myb(+) erythro-myeloid progenitor-derived fetal monocytes give rise to adult tissue-resident macrophages.

    Science.gov (United States)

    Hoeffel, Guillaume; Chen, Jinmiao; Lavin, Yonit; Low, Donovan; Almeida, Francisca F; See, Peter; Beaudin, Anna E; Lum, Josephine; Low, Ivy; Forsberg, E Camilla; Poidinger, Michael; Zolezzi, Francesca; Larbi, Anis; Ng, Lai Guan; Chan, Jerry K Y; Greter, Melanie; Becher, Burkhard; Samokhvalov, Igor M; Merad, Miriam; Ginhoux, Florent

    2015-04-21

    Although classified as hematopoietic cells, tissue-resident macrophages (MFs) arise from embryonic precursors that seed the tissues prior to birth to generate a self-renewing population, which is maintained independently of adult hematopoiesis. Here we reveal the identity of these embryonic precursors using an in utero MF-depletion strategy and fate-mapping of yolk sac (YS) and fetal liver (FL) hematopoiesis. We show that YS MFs are the main precursors of microglia, while most other MFs derive from fetal monocytes (MOs). Both YS MFs and fetal MOs arise from erythro-myeloid progenitors (EMPs) generated in the YS. In the YS, EMPs gave rise to MFs without monocytic intermediates, while EMP seeding the FL upon the establishment of blood circulation acquired c-Myb expression and gave rise to fetal MOs that then seeded embryonic tissues and differentiated into MFs. Thus, adult tissue-resident MFs established from hematopoietic stem cell-independent embryonic precursors arise from two distinct developmental programs.

  6. Effect of Polarization on Airway Epithelial Conditioning of Monocyte-Derived Dendritic Cells

    DEFF Research Database (Denmark)

    Papazian, Dick; Chhoden, Tashi; Arge, Maria

    2015-01-01

    to sample allergens administered to the apical side. Allergen uptake depended on both polarization and the nature of the allergen. AEC conditioning led to decreased birch allergen-specific proliferation of autologous T cells and a trend toward decreased secretion of the Th2-specific cytokines IL-5 and IL-13...

  7. Activation and Genetic Modification of Human Monocyte-Derived Dendritic Cells using Attenuated Salmonella typhimurium

    Directory of Open Access Journals (Sweden)

    Agnieszka Michael

    2010-01-01

    Full Text Available Live attenuated bacterial vectors, such as Salmonella typhimurium, have shown promise as delivery vehicles for DNA. We have examined two new strains of S. typhimurium and their impact on dendritic cell maturation (CD12-sifA/aroC mutant and WT05-ssaV/aroC, both in TML background. Strain WT05 matured dendritic cells in a more efficient way; caused higher release of cytokines TNF-α, IL-12, IL-1β; and was efficient for gene transfer. These findings suggest that the genetic background of the attenuation can influence the pattern of inflammatory immune response to Salmonella infection.

  8. Effects of titanium(iv) ions on human monocyte-derived dendritic cells.

    Science.gov (United States)

    Chan, Erwin Ph; Mhawi, Amir; Clode, Peta; Saunders, Martin; Filgueira, Luis

    2009-03-01

    Orthopaedic metal implants composed of titanium are routinely used in bone fracture repair and for joint replacement therapies. A considerable fraction of implant recipients are unable to benefit due to implant failure resulting from aseptic loosening, while others may experience cutaneous sensitivity to titanium after implantation. An adaptive immune reactivity towards titanium ions, originating from the biocorrosion of the implants, could play a role. As an initiator of the adaptive immune response, dendritic cells (DC) were studied for uptake and characteristics after titanium exposure. Energy filtered transmission electron microscopy showed uptake of titanium(iv) (Ti(iv)) ions by DCs in vitro and co-localisation with phosphorus-rich cell structures of the DC membranes (phospholipids), cytoplasm (ribosomes and phosphorylated proteins) and the nucleus (DNA). DC maturation and function were investigated by measuring cell surface marker expression by flow cytometry. After exposure, DCs showed a decrease in MHC class II (HLA-DR), co-stimulatory molecules (CD40, CD80 & CD86) and chemokine receptors (CCR) 6 and CCR7 but an increase in CCR4 after Ti(iv) treatment. However, Ti(iv) treated DCs had an increased stimulatory capacity towards allogenic lymphocytes. A Ti(iv) concentration dependant increase of IL-12p70 was observed amidst decrease of the other measured cytokines (TGF-β1 and TGF-β2). Hence, Ti(iv) alters DC properties, resulting in an enhanced T lymphocyte reactivity and deviation towards a Th1 type immune response. This effect may be responsible for the inflammatory side effects of titanium implants seen in patients.

  9. Modulation of monocyte/macrophage-derived cytokine and chemokine profile by persistent Hepatitis C virus (HCV infection leads to chronic inflammation

    Directory of Open Access Journals (Sweden)

    Penelope Mavromara

    2012-02-01

    Full Text Available HCV infection presents a major public health problem, with more than 170 million people infected worldwide. Chronicity and persistence of infection constitute the hallmark of the disease. Although HCV is a hepatotropic virus, subsets of immune cells have been found to be permissive to infection and viral replication. Peripheral blood monocytes, attracted to the site of infection and differentiated into macrophages, and resident hepatic macrophages, known as Kupffer cells, are important mediators of innate immunity, through production of several chemokines and cytokines in addition to their phagocytic activity. HCV proteins have been shown to modulate the cytokine and chemokine production profile of monocytes/macrophages, as it is suggested by both in vitro and clinical studies. This modified expression profile appears crucial for the establishment of aberrant inflammation that leads to liver cirrhosis and hepatocellular carcinoma.

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

    Directory of Open Access Journals (Sweden)

    Noemi Cifani

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

  11. Effects of beauvericin, enniatin b and moniliformin on human dendritic cells and macrophages: an in vitro study.

    Science.gov (United States)

    Ficheux, A S; Sibiril, Y; Parent-Massin, D

    2013-09-01

    The aim of this study was to assess the in vitro effects of emerging mycotoxins beauvericin, enniatin B and moniliformin on human dendritic cells and macrophages. Beauvericin and enniatin B were cytotoxic on these cells. IC50 were equal to 1.0 μM, 2.9 μM and 2.5 μM beauvericin for immature dendritic cells, mature dendritic cells and macrophages, respectively. IC50 were equal to 1.6 μM, 2.6 μM and 2.5 μM for immature dendritic cells, mature dendritic cells and macrophages exposed to enniatin B, respectively. Effects on the differentiation process of monocytes into macrophages or into immature dendritic cells as well as effects on dendritic cells maturation have been studied. The differentiation process of monocytes into immature dendritic cells was not disturbed in the presence of beauvericin. Dendritic cells exposed to beauvericin during the maturation process presented a decrease of CCR7 expression and an increase of IL-10 secretion. Monocytes exposed to beauvericin during the differentiation process into macrophages presented a decrease of endocytosis ability. The differentiation process of monocytes into immature dendritic cells was not disturbed in the presence of enniatin B. Dendritic cells exposed to enniatin B during the maturation process presented a decrease of expression of the maturation makers CD80, CD86 and CCR7 and an increase of IL-10 secretion. Monocytes exposed to enniatin B during the differentiation process into macrophages presented a decrease of endocytosis ability and an increase of CD71. CD1a expression and endocytosis capacity were decreased on immature dendritic cells exposed to moniliformin. Monocytes-derived macrophages exposed to moniliformin during the differentiation process presented a decrease of endocytosis ability, and a decrease of CD71 and HLA-DR expression. According to these results, immunological disorders could be observed on human after ingestion of these alimentary toxins.

  12. Gomesin acts in the immune system and promotes myeloid differentiation and monocyte/macrophage activation in mouse.

    Science.gov (United States)

    Buri, Marcus V; Dias, Carol C; Barbosa, Christiano M V; Nogueira-Pedro, Amanda; Ribeiro-Filho, Antonio C; Miranda, Antonio; Paredes-Gamero, Edgar J

    2016-11-01

    Due to the cytotoxic effect of antimicrobial peptides (AMP) against several microorganism and tumor cells has been proposed their association with the immune system. However, just a few reports have shown this relationship. In this study, mice were treated with gomesin, a β-hairpin AMP that exhibit high cytotoxicity against bacterial and tumor cells. Different effects in the immune system were observed, such as, decrease of CD3(+) in T lymphocytes (Control: 17.7±1.4%; Gomesin: 7.67±1.2%) and in hematopoietic progenitors and increase of hematopoietic stem cell (Control: 0.046±0.004%; Gomesin: 0.067±0.003%), B220(+) B lymphocytes (Control: 38.63±1.5%; Gomesin: 47.83±0.48%), and Mac-1(+)F4/80(+) macrophages (Control: 11.76±3.4%; Gomesin: 27.13±4.0%). Additionally, macrophage increase was accompanied by an increase of macrophage phagocytosis (Control 20.85±1.53; Gomesin 31.32±1 Geometric mean), interleukin 6 (Control: 47.24±1.9ng/mL; Gomesin: 138.68±33.68ng/mL) and monocyte chemoattractant protein-1 (Control: 0.872±0.093ng/mL; Gomesin: 1.83±0.067ng/mL). Thus, this report showed immunomodulatory activity of gomesin in the immune system of mice. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Effects of age and macrophage lineage on intracellular survival and cytokine induction after infection with Rhodococcus equi.

    Science.gov (United States)

    Berghaus, Londa J; Giguère, Steeve; Sturgill, Tracy L

    2014-07-15

    Rhodococcus equi, a facultative intracellular pathogen of macrophages, causes life-threatening pneumonia in foals and in people with underlying immune deficiencies. As a basis for this study, we hypothesized that macrophage lineage and age would affect intracellular survival of R. equi and cytokine induction after infection. Monocyte-derived and bronchoalveolar macrophages from 10 adult horses and from 10 foals (sampled at 1-3 days, 2 weeks, 1 month, 3 months, and 5 months of age) were infected ex vivo with virulent R. equi. Intracellular R. equi were quantified and mRNA expression of IL-1β, IL-4, IL-6, IL-8, IL-10, IL-12 p40, IL-18, IFN-γ, and TNF-α was measured. Intracellular replication of R. equi was significantly (Pequi was significantly (P=0.002) higher in 3-month-old foals than in 3-day old foals, 2-week-old foals, 1-month-old foals, and adult horses. Expression of IL-4 mRNA was significantly higher in monocyte-derived macrophages whereas expression of IL-6, IL-18, and TNF-α was significantly higher in bronchoalveolar macrophages. Induction of IL-1β, IL-10, IL-12 p40, and IL-8 mRNA in bronchoalveolar macrophages of 1-3-day old foals was significantly higher than in older foals or adult horses. Preferential intracellular survival of R. equi in bronchoalveolar macrophages of juvenile horses may play a role in the pulmonary tropism of the pathogen and in the window of age susceptibility to infection. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Recruitment of CCR6-expressing Th17 cells by CCL20 secreted from plasmin-stimulated macrophages

    Institute of Scientific and Technical Information of China (English)

    Qun Li; Yves Laumonnier; Tatiana Syrovets; Thomas Simmet

    2013-01-01

    In the present study,monocyte-derived human macrophages were differentiated from buffy coats.Na(i)ve CD4+ T-cells enriched from peripheral blood mononuclear cells using anti-CD4 magnetic beads and the autoMACS separation system were polarized under T-helper 17 (Th17)-promoting conditions for 6 days to get Th17 cells.The frequency of Th17 cell differentiation and the expression of C-C chemokine receptor type 6 (CCR6) on Th17 cells were investigated by flow cytometry.Plasmin-triggered induction of macrophage inflammatory protein-3alpha/C-C chemokine ligand 20 (CCL20) genes in macrophages was assessed by reverse transcription-polymerase chain reaction,and secreted protein levels were measured by enzymelinked immunosorbent assay.Th17 cell migration induced by CCL20 secreted from plasmin-stimulated macrophages was tested in vitro by chemotaxis using a transwell system.These results demonstrate that plasmin triggers the expression of chemokine CCL20 messenger RNA and the release of CCL20 protein in human monocyte-derived macrophages,which critically depend on the proteolytic activity of plasmin and activation of p38 mitogen-activated protein kinase and nuclear factor-kappaB signaling pathways.Expression of CCR6 was detected on 87.23 ± 8.6% of Th17 cells in vitro.Similar to chemotaxis triggered by recombinant human CCL20,supernatants collected from plasmin-stimulated macrophage-induced chemotactic migration of Th17 cells,which could be inhibited by an anti-CCL20 neutralizing antibody.These results suggest that plasmin generated in inflamed tissues might elicit production of chemokine CCL20 by human macrophages leading to the recruitmentof CCR6 positive Th17 cells to the inflammatory sites.

  15. OSCAR-collagen signaling in monocytes plays a proinflammatory role and may contribute to the pathogenesis of rheumatoid arthritis

    DEFF Research Database (Denmark)

    Schultz, Heidi Schiøler; Guo, Li; Keller, Pernille;

    2016-01-01

    functional maturation of monocyte-derived dendritic cells. OSCAR is upregulated on monocytes from rheumatoid arthritis (RA) patients with active disease, and these monocytes show an increased proosteoclastogenic potential. In the current study, we have addressed a functional role for an OSCAR...... fluid of RA patients plated on ColII secreted TNF-α and IL-8 in an OSCAR-dependent manner. Global RNA profiling showed that components of multiple signaling pathways relevant to RA pathogenesis are regulated at the transcriptional level by OSCAR in monocytes. Thus, OSCAR can play a proinflammatory role...... in monocyte-derived cells and may contribute crucially on multiple levels to RA pathogenesis....

  16. 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.

  17. Human monocytes and macrophages express NADPH oxidase 5; a potential source of reactive oxygen species in atherosclerosis.

    Science.gov (United States)

    Manea, Adrian; Manea, Simona-Adriana; Gan, Ana Maria; Constantin, Alina; Fenyo, Ioana Madalina; Raicu, Monica; Muresian, Horia; Simionescu, Maya

    2015-05-22

    Monocytes (Mon) and Mon-derived macrophages (Mac) orchestrate important oxidative and inflammatory reactions in atherosclerosis by secreting reactive oxygen species (ROS) due, in large part, to the upregulated NADPH oxidases (Nox). The Nox enzymes have been extensively investigated in human Mon and Mac. However, the expression and functional significance of the Nox5 subtypes is not known. We aimed at elucidating whether Nox5 is expressed in human Mon and Mac, and examine its potential role in atherosclerosis. Human monocytic THP-1 cell line and CD14(+) Mon were employed to search for Nox5 expression. RT-PCR, Western blot, lucigenin-enhanced chemiluminescence and dihydroethidium assays were utilized to examine Nox5 in these cells. We found that Nox5 transcription variants and proteins are constitutively expressed in THP-1 cells and primary CD14(+) Mon. Silencing of Nox5 protein expression by siRNA reduced the Ca(2+)-dependent Nox activity and the formation of ROS in Mac induced by A23187, a selective Ca(2+) ionophore. Exposure of Mac to increasing concentrations of IFNγ (5-100 ng/ml) or oxidized LDL (5-100 μg/ml) resulted in a dose-dependent increase in Nox5 protein expression and elevation in intracellular Ca(2+) concentration. Immunohistochemical staining revealed that Nox5 is present in CD68(+) Mac-rich area within human carotid artery atherosclerotic plaques. To the best of our knowledge, this is the first evidence that Nox5 is constitutively expressed in human Mon. Induction of Nox5 expression in IFNγ- and oxidized LDL-exposed Mac and the presence of Nox5 in Mac-rich atheroma are indicative of the implication of Nox5 in atherogenesis.

  18. Adipocyte progenitor cells initiate monocyte chemoattractant protein-1-mediated macrophage accumulation in visceral adipose tissue

    Directory of Open Access Journals (Sweden)

    Jennifer L. Kaplan

    2015-11-01

    Conclusions: This study provides the first in vivo evidence, to our knowledge, that committed AdPCs in VAT are the initial source of obesity-induced MCP-1 and identifies the helix-loop-helix transcription factor Id3 as a critical regulator of p21Cip1 expression, AdPC proliferation, MCP-1 expression and M1 macrophage accumulation in VAT. Inhibition of Id3 and AdPC expansion, as well as CD44 expression in human AdPCs, may serve as unique therapeutic targets for the regulation of adipose tissue inflammation.

  19. A unique anti-CD115 monoclonal antibody which inhibits osteolysis and skews human monocyte differentiation from M2-polarized macrophages toward dendritic cells.

    Science.gov (United States)

    Haegel, Hélène; Thioudellet, Christine; Hallet, Rémy; Geist, Michel; Menguy, Thierry; Le Pogam, Fabrice; Marchand, Jean-Baptiste; Toh, Myew-Ling; Duong, Vanessa; Calcei, Alexandre; Settelen, Nathalie; Preville, Xavier; Hennequi, Marie; Grellier, Benoit; Ancian, Philippe; Rissanen, Jukka; Clayette, Pascal; Guillen, Christine; Rooke, Ronald; Bonnefoy, Jean-Yves

    2013-01-01

    Cancer progression has been associated with the presence of tumor-associated M2-macrophages (M2-TAMs) able to inhibit anti-tumor immune responses. It is also often associated with metastasis-induced bone destruction mediated by osteoclasts. Both cell types are controlled by the CD115 (CSF-1R)/colony-stimulating factor-1 (CSF-1, M-CSF) pathway, making CD115 a promising target for cancer therapy. Anti-human CD115 monoclonal antibodies (mAbs) that inhibit the receptor function have been generated in a number of laboratories. These mAbs compete with CSF-1 binding to CD115, dramatically affecting monocyte survival and preventing osteoclast and macrophage differentiation, but they also block CD115/CSF-1 internalization and degradation, which could lead to potent rebound CSF-1 effects in patients after mAb treatment has ended. We thus generated and selected a non-ligand competitive anti-CD115 mAb that exerts only partial inhibitory effects on CD115 signaling without blocking the internalization or the degradation of the CD115/CSF-1 complex. This mAb, H27K15, affects monocyte survival only minimally, but downregulates osteoclast differentiation and activity. Importantly, it inhibits monocyte differentiation to CD163(+)CD64(+) M2-polarized suppressor macrophages, skewing their differentiation toward CD14(-)CD1a(+) dendritic cells (DCs). In line with this observation, H27K15 also drastically inhibits monocyte chemotactic protein-1 secretion and reduces interleukin-6 production; these two molecules are known to be involved in M2-macrophage recruitment. Thus, the non-depleting mAb H27K15 is a promising anti-tumor candidate, able to inhibit osteoclast differentiation, likely decreasing metastasis-induced osteolysis, and able to prevent M2 polarization of TAMs while inducing DCs, hence contributing to the creation of more efficient anti-tumor immune responses.

  20. Serum concentrations and peripheral secretion of the beta chemokines monocyte chemoattractant protein 1 and macrophage inflammatory protein 1α in alcoholic liver disease

    OpenAIRE

    Fisher, N; Neil, D.; Williams, A.; Adams, D.

    1999-01-01

    BACKGROUND—Alcoholic liver disease is associated with increased hepatic expression of monocyte chemoattractant protein 1 (MCP-1) and macrophage inflammatory protein 1α (MIP-1α).
AIMS—To determine whether concentrations of chemokines in the peripheral circulation reflect disease activity, and whether chemokine secretion is restricted to the liver or is part of a systemic inflammatory response in alcoholic liver disease.
PATIENTS—Fifty one patients with alcoholic liver disease and 12 healthy co...

  1. Alkali treatment of microrough titanium surfaces affects macrophage/monocyte adhesion, platelet activation and architecture of blood clot formation

    Directory of Open Access Journals (Sweden)

    V Milleret

    2011-05-01

    Full Text Available Titanium implants are most commonly used for bone augmentation and replacement due to their favorable osseointegration properties. Here, hyperhydrophilic sand-blasted and acid-etched (SBA titanium surfaces were produced by alkali treatment and their responses to partially heparinized whole human blood were analyzed. Blood clot formation, platelet activation and activation of the complement system was analyzed revealing that exposure time between blood and the material surface is crucial as increasing exposure time results in higher amount of activated platelets, more blood clots formed and stronger complement activation. In contrast, the number of macrophages/monocytes found on alkali-treated surfaces was significantly reduced as compared to untreated SBA Ti surfaces. Interestingly, when comparing untreated to modified SBA Ti surfaces very different blood clots formed on their surfaces. On untreated Ti surfaces blood clots remain thin (below 15 mm, patchy and non-structured lacking large fibrin fiber networks whereas blood clots on differentiated surfaces assemble in an organized and layered architecture of more than 30 mm thickness. Close to the material surface most nucleated cells adhere, above large amounts of non-nucleated platelets remain entrapped within a dense fibrin fiber network providing a continuous cover of the entire surface. These findings might indicate that, combined with findings of previous in vivo studies demonstrating that alkali-treated SBA Ti surfaces perform better in terms of osseointegration, a continuous and structured layer of blood components on the blood-facing surface supports later tissue integration of an endosseous implant.

  2. Dysregulated Type I Interferon and Inflammatory Monocyte-Macrophage Responses Cause Lethal Pneumonia in SARS-CoV-Infected Mice.

    Science.gov (United States)

    Channappanavar, Rudragouda; Fehr, Anthony R; Vijay, Rahul; Mack, Matthias; Zhao, Jincun; Meyerholz, David K; Perlman, Stanley

    2016-02-10

    Highly pathogenic human respiratory coronaviruses cause acute lethal disease characterized by exuberant inflammatory responses and lung damage. However, the factors leading to lung pathology are not well understood. Using mice infected with SARS (severe acute respiratory syndrome)-CoV, we show that robust virus replication accompanied by delayed type I interferon (IFN-I) signaling orchestrates inflammatory responses and lung immunopathology with diminished survival. IFN-I remains detectable until after virus titers peak, but early IFN-I administration ameliorates immunopathology. This delayed IFN-I signaling promotes the accumulation of pathogenic inflammatory monocyte-macrophages (IMMs), resulting in elevated lung cytokine/chemokine levels, vascular leakage, and impaired virus-specific T cell responses. Genetic ablation of the IFN-αβ receptor (IFNAR) or IMM depletion protects mice from lethal infection, without affecting viral load. These results demonstrate that IFN-I and IMM promote lethal SARS-CoV infection and identify IFN-I and IMMs as potential therapeutic targets in patients infected with pathogenic coronavirus and perhaps other respiratory viruses. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Carbon nanohorns allow acceleration of osteoblast differentiation via macrophage activation

    Science.gov (United States)

    Hirata, Eri; Miyako, Eijiro; Hanagata, Nobutaka; Ushijima, Natsumi; Sakaguchi, Norihito; Russier, Julie; Yudasaka, Masako; Iijima, Sumio; Bianco, Alberto; Yokoyama, Atsuro

    2016-07-01

    Carbon nanohorns (CNHs), formed by a rolled graphene structure and terminating in a cone, are promising nanomaterials for the development of a variety of biological applications. Here we demonstrate that alkaline phosphatase activity is dramatically increased by coculture of human monocyte derived macrophages (hMDMs) and human mesenchymal stem cells (hMSCs) in the presence of CNHs. CNHs were mainly localized in the lysosome of macrophages more than in hMSCs during coculturing. At the same time, the amount of Oncostatin M (OSM) in the supernatant was also increased during incubation with CNHs. Oncostatin M (OSM) from activated macrophage has been reported to induce osteoblast differentiation and matrix mineralization through STAT3. These results suggest that the macrophages engulfed CNHs and accelerated the differentiation of mesenchymal stem cells into the osteoblast via OSM release. We expect that the proof-of-concept on the osteoblast differentiation capacity by CNHs will allow future studies focused on CNHs as ideal therapeutic materials for bone regeneration.Carbon nanohorns (CNHs), formed by a rolled graphene structure and terminating in a cone, are promising nanomaterials for the development of a variety of biological applications. Here we demonstrate that alkaline phosphatase activity is dramatically increased by coculture of human monocyte derived macrophages (hMDMs) and human mesenchymal stem cells (hMSCs) in the presence of CNHs. CNHs were mainly localized in the lysosome of macrophages more than in hMSCs during coculturing. At the same time, the amount of Oncostatin M (OSM) in the supernatant was also increased during incubation with CNHs. Oncostatin M (OSM) from activated macrophage has been reported to induce osteoblast differentiation and matrix mineralization through STAT3. These results suggest that the macrophages engulfed CNHs and accelerated the differentiation of mesenchymal stem cells into the osteoblast via OSM release. We expect that the

  4. Contrasting Roles of Islet Resident Immunoregulatory Macrophages and Dendritic Cells in Experimental Autoimmune Type 1 Diabetes.

    Directory of Open Access Journals (Sweden)

    Thomas B Thornley

    Full Text Available The innate immune system critically shapes diabetogenic adaptive immunity during type 1 diabetes (T1D pathogenesis. While the role of tissue-infiltrating monocyte-derived macrophages in T1D is well established, the role of their tissue-resident counterparts remains undefined. We now demonstrate that islet resident macrophages (IRMs from non-autoimmune mice have an immunoregulatory phenotype and powerfully induce FoxP3+ Tregs in vitro. The immunoregulatory phenotype and function of IRMs is compromised by TLR4 activation in vitro. Moreover, as T1D approaches in NOD mice, the immunoregulatory phenotype of IRMs is diminished as is their relative abundance compared to immunostimulatory DCs. Our findings suggest that maintenance of IRM abundance and their immunoregulatory phenotype may constitute a novel therapeutic strategy to prevent and/or cure T1D.

  5. DMPD: Monocyte CD14: a multifunctional receptor engaged in apoptosis from both sides. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 10380893 Monocyte CD14: a multifunctional receptor engaged in apoptosis from both s...ides. Heidenreich S. J Leukoc Biol. 1999 Jun;65(6):737-43. (.png) (.svg) (.html) (.csml) Show Monocyte CD14: a multi...functional receptor engaged in apoptosis from both sides. PubmedID 10380893 Title Monocyte CD14: a multi

  6. Mycobacterium leprae-Infected Macrophages Preferentially Primed Regulatory T Cell Responses and Was Associated with Lepromatous Leprosy.

    Directory of Open Access Journals (Sweden)

    Degang Yang

    2016-01-01

    Full Text Available The persistence of Mycobacterium leprae (M. leprae infection is largely dependent on the types of host immune responses being induced. Macrophage, a crucial modulator of innate and adaptive immune responses, could be directly infected by M. leprae. We therefore postulated that M. leprae-infected macrophages might have altered immune functions.Here, we treated monocyte-derived macrophages with live or killed M. leprae, and examined their activation status and antigen presentation. We found that macrophages treated with live M. leprae showed committed M2-like function, with decreased interleukin 1 beta (IL-1beta, IL-6, tumor necrosis factor alpha (TNF-alpha and MHC class II molecule expression and elevated IL-10 and CD163 expression. When incubating with naive T cells, macrophages treated with live M. leprae preferentially primed regulatory T (Treg cell responses with elevated FoxP3 and IL-10 expression, while interferon gamma (IFN-gamma expression and CD8+ T cell cytotoxicity were reduced. Chromium release assay also found that live M. leprae-treated macrophages were more resistant to CD8+ T cell-mediated cytotoxicity than sonicated M. leprae-treated monocytes. Ex vivo studies showed that the phenotype and function of monocytes and macrophages had clear differences between L-lep and T-lep patients, consistent with the in vitro findings.Together, our data demonstrate that M. leprae could utilize infected macrophages by two mechanisms: firstly, M. leprae-infected macrophages preferentially primed Treg but not Th1 or cytotoxic T cell responses; secondly, M. leprae-infected macrophages were more effective at evading CD8+ T cell-mediated cytotoxicity.

  7. [Spectrophotometric determination of protein content in THP-1 monocytes/macrophages - description of the method].

    Science.gov (United States)

    Wolska, Jolanta; Janda, Katarzyna; Gutowska, Izabela

    2015-01-01

    Proteins are the basic building block of tissue, and are part of enzymes and hormones regulating many important life processes. Changes in their concentration control the metabolic processes of the cell. Quantitative determination of the protein content is divided into indirect methods (e.g. Kjeldahl method) and direct methods (buret method, Lowry, immunoenzymatic, formol method, based on incorporation of dye in the range of ultraviolet spectrophotometry, and based on the phenomenon of selective absorption of radiation in the infrared range). One of the methods for the determination of protein content is the spectrophotometric method described by Bradford. The protein concentration assay procedure utilizes the phenomenon of formation of the dye (Coomassie Brillant Blue G-250)-protein and colour intensity is proportional to the protein content in the solution. The aim of this study was to verify the usefulness of this method for determining the protein content in THP-1 cells cultured with extracts of nettle fruit stalks (Urtica dioica L.). Aqueous and alcohol extracts at two concentrations were used. It has been shown that the spectrophotometric determination of protein content by the Bradford method is an effective and accurate method for determining the concentration of protein in THP-1 macrophages. The results indicate that this method can be recommended for the determination of the protein content in other cell cultures.

  8. Alveolar macrophage-epithelial cell interaction following exposure to atmospheric particles induces the release of mediators involved in monocyte mobilization and recruitment

    Directory of Open Access Journals (Sweden)

    Mukae Hiroshi

    2005-08-01

    Full Text Available Abstract Background Studies from our laboratory have shown that human alveolar macrophages (AM and bronchial epithelial cells (HBEC exposed to ambient particles (PM10 in vitro increase their production of inflammatory mediators and that supernatants from PM10-exposed cells shorten the transit time of monocytes through the bone marrow and promote their release into the circulation. Methods The present study concerns co-culture of AM and HBEC exposed to PM10 (EHC-93 and the production of mediators involved in monocyte kinetics measured at both the mRNA and protein levels. The experiments were also designed to determine the role of the adhesive interaction between these cells via the intercellular adhesion molecule (ICAM-1 in the production of these mediators. Results AM/HBEC co-cultures exposed to 100 μg/ml of PM10 for 2 or 24 h increased their levels of granulocyte-macrophage colony-stimulating factor (GM-CSF, M-CSF, macrophage inflammatory protein (MIP-1β, monocyte chemotactic protein (MCP-1, interleukin (IL-6 and ICAM-1 mRNA, compared to exposed AM or HBEC mono-cultures, or control non-exposed co-cultures. The levels of GM-CSF, M-CSF, MIP-1β and IL-6 increased in co-cultured supernatants collected after 24 h exposure compared to control cells (p 10-induced increase in co-culture mRNA expression. Conclusion We conclude that an ICAM-1 independent interaction between AM and HBEC, lung cells that process inhaled particles, increases the production and release of mediators that enhance bone marrow turnover of monocytes and their recruitment into tissues. We speculate that this interaction amplifies PM10-induced lung inflammation and contributes to both the pulmonary and systemic morbidity associated with exposure to air pollution.

  9. Macrophages from chickens selected for high antibody response produced more nitric oxide and have greater phagocytic capacity.

    Science.gov (United States)

    Guimarães, Marco Cesar Cunegundes; Guillermo, Landi Veivi Costilla; Matta, Marcos Fernando de Rezende; Soares, Sandro Gomes; DaMatta, Renato Augusto

    2011-04-15

    Macrophages are fundamental cells of the innate immune system, which, through phagocytosis and nitric oxide production, eliminate pathogens. The aim of the present study was to determine if macrophages from chicken families divergently selected to high and low antibodies response differ in nitric oxide production and phagocytic capacity. Blood monocytes derived macrophages were activated with lipopolysaccharide and supernatant from chicken spleen lymphocytes cultured with Concanavalin A (containing chicken interferon). Nitric oxide production was evaluated in culture supernatants. Phagocytic capacity of activated and non-activated macrophages was assayed using yeasts and IgY opsonized sheep red blood cells. Activated and non-activated macrophages from the high antibodies response family produced higher nitric oxide levels, internalized more yeast and significantly more opsonized sheep red blood cells than macrophages from the low antibodies response family. Moreover, activated macrophages became more elongated and widely spread. These findings indicate that macrophages from the high antibodies response family were more active suggesting that the differences in antibody response also depend on macrophage function.

  10. DYSFUNCTION OF MONOCYTES AND DENDRITIC CELLS IN PATIENTS WITH PREMATURE OVARIAN FAILURE

    NARCIS (Netherlands)

    HOEK, A; VAN KASTEREN, Y; DE HAAN-MEULMAN, M; SCHOEMAKER, J; DREXHAGE, HA

    1993-01-01

    PROBLEM: Due to the presence of ovarian antibodies it has been suggested that premature ovarian failure (POF) belongs to the autoimmune endocrinopathies. Monocytes and the monocyte-derived dendritic cells play a prominent role in the initial stages of endocrine autoimmune reactions: the accumulation

  11. The Ron Receptor Tyrosine Kinase Regulates Macrophage Heterogeneity and Plays a Protective Role in Diet-Induced Obesity, Atherosclerosis, and Hepatosteatosis.

    Science.gov (United States)

    Yu, Shan; Allen, Joselyn N; Dey, Adwitia; Zhang, Limin; Balandaram, Gayathri; Kennett, Mary J; Xia, Mingcan; Xiong, Na; Peters, Jeffrey M; Patterson, Andrew; Hankey-Giblin, Pamela A

    2016-07-01

    Obesity is a chronic inflammatory disease mediated in large part by the activation of inflammatory macrophages. This chronic inflammation underlies a whole host of diseases including atherosclerosis, hepatic steatosis, insulin resistance, type 2 diabetes, and cancer, among others. Macrophages are generally classified as either inflammatory or alternatively activated. Some tissue-resident macrophages are derived from yolk sac erythromyeloid progenitors and fetal liver progenitors that seed tissues during embryogenesis and have the ability to repopulate through local proliferation. These macrophages tend to be anti-inflammatory in nature and are generally involved in tissue remodeling, repair, and homeostasis. Alternatively, during chronic inflammation induced by obesity, bone marrow monocyte-derived macrophages are recruited to inflamed tissues, where they produce proinflammatory cytokines and exacerbate inflammation. The extent to which these two populations of macrophages are plastic in their phenotype remains controversial. We have demonstrated previously that the Ron receptor tyrosine kinase is expressed on tissue-resident macrophages, where it limits inflammatory macrophage activation and promotes a repair phenotype. In this study, we demonstrate that Ron is expressed in a subpopulation of macrophages during chronic inflammation induced by obesity that exhibit a repair phenotype as determined by the expression of arginase 1. In addition, we demonstrate that the Ron receptor plays a protective role in the progression of diet-induced obesity, hepatosteatosis, and atherosclerosis. These results suggest that altering macrophage heterogeneity in vivo could have the potential to alleviate obesity-associated diseases. Copyright © 2016 by The American Association of Immunologists, Inc.

  12. Monocyte-macrophage differentiation of acute myeloid leukemia cell lines by small molecules identified through interrogation of the Connectivity Map database.

    Science.gov (United States)

    Manzotti, Gloria; Parenti, Sandra; Ferrari-Amorotti, Giovanna; Soliera, Angela Rachele; Cattelani, Sara; Montanari, Monica; Cavalli, Daniel; Ertel, Adam; Grande, Alexis; Calabretta, Bruno

    2015-01-01

    The transcription factor C/EBPα is required for granulocytic differentiation of normal myeloid progenitors and is frequently inactivated in acute myeloid leukemia (AML) cells. Ectopic expression of C/EBPα in AML cells suppresses proliferation and induces differentiation suggesting that restoring C/EBPα expression/activity in AML cells could be therapeutically useful. Unfortunately, current approaches of gene or protein delivery in leukemic cells are unsatisfactory. However, "drug repurposing" is becoming a very attractive strategy to identify potential new uses for existing drugs. In this study, we assessed the biological effects of candidate C/EBPα-mimetics identified by interrogation of the Connectivity Map database. We found that amantadine, an antiviral and anti-Parkinson agent, induced a monocyte-macrophage-like differentiation of HL60, U937, Kasumi-1 myeloid leukemia cell lines, as indicated by morphology and differentiation antigen expression, when used in combination with suboptimal concentration of all trans retinoic acid (ATRA) or Vit D3. The effect of amantadine depends, in part, on increased activity of the vitamin D receptor (VDR), since it induced VDR expression and amantadine-dependent monocyte-macrophage differentiation of HL60 cells was blocked by expression of dominant-negative VDR. These results reveal a new function for amantadine and support the concept that screening of the Connectivity Map database can identify small molecules that mimic the effect of transcription factors required for myelo-monocytic differentiation.

  13. Elucidation of adhesion-dependent spontaneous apoptosis in macrophages using phase separated PEG/polyurethane films

    Science.gov (United States)

    Zachman, Angela L.; Page, Jonathan M.; Prabhakar, Gayathri; Guelcher, Scott A.; Sung, Hak-Joon

    2013-01-01

    Circulating monocytes undergo spontaneous apoptosis when there is no activation stimulus, which is critical to population control for proper host response to implants. As activation and apoptosis of monocytes/macrophages are regulated by cell–cell and cell–matrix interactions, their regulatory mechanism was investigated in this study using polyethylene glycol (PEG)-containing polyurethane films in which PEG-rich and polyester-rich domains were phase separated. Human blood monocyte-derived macrophages (HBMs) preferentially adhered to PEG domains (cell–matrix interaction) due to the low molecular weight (600 g mol−1), resulting in increased HBM density (cell–cell interaction). As both cell–cell and cell–matrix interactions were promoted, HBM apoptosis increased, while their activation as measured by phagocytosis, intracellular reactive oxygen species (ROS) level and matrix metalloproteinase-9 production decreased compared to PEG-free films. When cell seeding density and cell-adhesive gelatin coating on silicone films were controlled, a cooperative role of cell–matrix (adhesion) and cell–cell (density) interactions in inducing HBM apoptosis was observed. Expression of the macrophage adhesion molecule CD11b caused apoptosis in this context, which was mediated by tissue necrosis factor-α signaling but down-regulated by the ROS inhibitor diphenylene iodonium and the anti-inflammatory peptide Ac-SDKP, suggesting a new concept for the design of biomaterials that allows for cell adhesion without excessive inflammatory activation. PMID:23128157

  14. Ultrafine carbon particles down-regulate CYP1B1 expression in human monocytes

    Directory of Open Access Journals (Sweden)

    Ziegler-Heitbrock Loems

    2009-10-01

    Full Text Available Abstract Background Cytochrome P450 monoxygenases play an important role in the defence against inhaled toxic compounds and in metabolizing a wide range of xenobiotics and environmental contaminants. In ambient aerosol the ultrafine particle fraction which penetrates deeply into the lungs is considered to be a major factor for adverse health effects. The cells mainly affected by inhaled particles are lung epithelial cells and cells of the monocyte/macrophage lineage. Results In this study we have analyzed the effect of a mixture of fine TiO2 and ultrafine carbon black Printex 90 particles (P90 on the expression of cytochrome P450 1B1 (CYP1B1 in human monocytes, macrophages, bronchial epithelial cells and epithelial cell lines. CYP1B1 expression is strongly down-regulated by P90 in monocytes with a maximum after P90 treatment for 3 h while fine and ultrafine TiO2 had no effect. CYP1B1 was down-regulated up to 130-fold and in addition CYP1A1 mRNA was decreased 13-fold. In vitro generated monocyte-derived macrophages (MDM, epithelial cell lines, and primary bronchial epithelial cells also showed reduced CYP1B1 mRNA levels. Benzo[a]pyrene (BaP is inducing CYB1B1 but ultrafine P90 can still down-regulate gene expression at 0.1 μM of BaP. The P90-induced reduction of CYP1B1 was also demonstrated at the protein level using Western blot analysis. Conclusion These data suggest that the P90-induced reduction of CYP gene expression may interfere with the activation and/or detoxification capabilities of inhaled toxic compounds.

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

    Science.gov (United States)

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

    2012-01-01

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

  16. Mycobacterium avium Subspecies paratuberculosis Recombinant Proteins Modulate Antimycobacterial Functions of Bovine Macrophages.

    Science.gov (United States)

    Bannantine, John P; Stabel, Judith R; Laws, Elizabeth; D Cardieri, Maria Clara; Souza, Cleverson D

    2015-01-01

    It has been shown that Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) activates the Mitogen Activated Protein Kinase (MAPK) p38 pathway, yet it is unclear which components of M. paratuberculosis are involved in the process. Therefore, a set of 42 M. paratuberculosis recombinant proteins expressed from coding sequences annotated as lipoproteins were screened for their ability to induce IL-10 expression, an indicator of MAPKp38 activation, in bovine monocyte-derived macrophages. A recombinant lipoprotein, designated as MAP3837c, was among a group of 6 proteins that strongly induced IL-10 gene transcription in bovine macrophages, averaging a 3.1-fold increase compared to non-stimulated macrophages. However, a parallel increase in expression of IL-12 and TNF-α was only observed in macrophages exposed to a subset of these 6 proteins. Selected recombinant proteins were further analyzed for their ability to enhance survival of M. avium within bovine macrophages as measured by recovered viable bacteria and nitrite production. All 6 IL-10 inducing MAP recombinant proteins along with M. paratuberculosis cells significantly enhanced phosphorylation of MAPK-p38 in bovine macrophages. Although these proteins are likely not post translationally lipidated in E. coli and thus is a limitation in this study, these results form the foundation of how the protein component of the lipoprotein interacts with the immune system. Collectively, these data reveal M. paratuberculosis proteins that might play a role in MAPK-p38 pathway activation and hence in survival of this organism within bovine macrophages.

  17. Monocyte to macrophage differentiation-associated (MMD) targeted by miR-140-5p regulates tumor growth in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Li, Weina, E-mail: liweina228@163.com [Department of Biomedical Engineering, Fourth Military Medical University, Xi’an 710032 (China); He, Fei, E-mail: hesili1027@163.com [Department of Hepatic Surgery, Xijing Hospital, Fourth Military Medical University, Xi’an 710032 (China)

    2014-07-18

    Highlights: • Expression of MMD is increased in lung cancer tissues. • Knockdown of MMD inhibits growth of A549 and LLC cells in vitro and in vivo. • MMD is a direct functional target of miR-140-5p. • MiR-140-5p/MMD axis regulates Erk1/2 signaling. - Abstract: Monocyte to macrophage differentiation-associated (MMD) is identified in macrophages as a gene associated with the differentiation from monocytes to macrophages. Recent microarray analysis for non-small cell lung cancer (NSCLC) suggests that MMD is an important signature associated with relapse and survival among patients with NSCLC. Therefore, we speculate that MMD likely plays a role in lung cancer. In this study, we found that the protein level of MMD was increased in lung cancer compared to benign lung tissues, and knockdown of MMD inhibited the growth of A549 and Lewis lung cancer cells (LLC) in vitro and in vivo. Integrated analysis demonstrated that MMD was a direct functional target of miR-140-5p. Furthermore, we found that miR-140-5p/MMD axis could affect the cell proliferation of lung cancer cells by regulating Erk signaling. Together, our results highlight the significance of miR-140-5p/MMD axis in lung cancer, and miR-140-5p/MMD axis could serve as new molecular targets for the therapy against lung cancer.

  18. The in vitro fungicidal activity of human macrophages against Penicillium marneffei is suppressed by dexamethasone.

    Science.gov (United States)

    Ma, Tuan; Chen, Renqiong; Li, Xiqing; Lu, Changming; Xi, Liyan

    2015-09-01

    Penicillium marneffei (P. marneffei) is a pathogenic fungus that can persist in macrophages and cause a life-threatening systemic mycosis in immunocompromised hosts. To elucidate the mechanisms underlying this opportunistic fungal infection, we established the co-culture system of P. marneffei conidia and human monocyte-derived macrophages (MDM) for investigating the interactions between them. And, we impaired the immune state of MDM by the addition of dexamethasone (DEX). Compared with immunocompetent MDM without DEX treatment in response to P. marneffei, DEX could damage MDM function in initiating the innate immune response through decreasing TNF-α production and the proportion of P. marneffei conidia in mature phagolysosomes, while the red pigment secretion by P. marneffei conidia was promoted by DEX following MDM lysis. Our data provide the evidence that DEX-treated MDM have a low fungicidal activity against P. marneffei that causes penicilliosis in immunocompromised hosts.

  19. Intramacrophage survival of uropathogenic Escherichia coli: Differences between diverse clinical isolates and between mouse and human macrophages

    DEFF Research Database (Denmark)

    Bokil, Nilesh J.; Totsika, Makrina; Carey, Alison J.;

    2011-01-01

    Uropathogenic E. coli (UPEC) are the primary cause of urinary tract infections. Recent studies have demonstrated that UPEC can invade and replicate within epithelial cells, suggesting that this bacterial pathogen may occupy an intracellular niche within the host. Given that many intracellular...... or initial uptake of bacteria. E. coli UTI89 localized to a Lamp1+ vesicular compartment within BMM. In contrast to survival within mouse BMM, intracellular bacterial loads of VR50 were low in both human monocyte-derived macrophages (HMDM) and in human T24 bladder epithelial cells. Collectively, these data......, originally isolated from patients with cystitis and asymptomatic bacteriuria respectively, showed elevated bacterial loads in BMM at 24h post-infection as compared to CFT073 and the asymptomatic bacteriuria strain 83972. These differences did not correlate with differential effects on macrophage survival...

  20. Glucagon-like peptide-1 (GLP-1) induces M2 polarization of human macrophages via STAT3 activation.

    Science.gov (United States)

    Shiraishi, Daisuke; Fujiwara, Yukio; Komohara, Yoshihiro; Mizuta, Hiroshi; Takeya, Motohiro

    2012-08-24

    It is known that glucagon-like peptide-1 (GLP-1) is a hormone secreted postprandially from the L-cells of the small intestine and regulates glucose homeostasis. GLP-1 is now used for the treatment of diabetes because of its beneficial role against insulin resistance. The GLP-1 receptor (GLP-1R) is expressed on many cell types, including macrophages, and GLP-1 suppresses the development of atherosclerosis by inhibiting macrophage function. However, there have so far been few studies that have investigated the significance of GLP-1/GLP-1R signaling in macrophage activation. In the present study, we examined the effect of GLP-1 and exenatide, a GLP-1R agonist, on human monocyte-derived macrophage (HMDM) activation. We found that GLP-1 induced signal transducer and activator of transcription 3 (STAT3) activation. Silencing of GLP-1R suppressed the GLP-1-induced STAT3 activation. In addition, alternatively activated (M2) macrophage-related molecules, such as IL-10, CD163, and CD204 in HMDM, were significantly upregulated by GLP-1. Furthermore, the co-culture of 3T3-L1 adipocytes with GLP-1-treated RAW 264.7 macrophages increased the secretion of adiponectin compared to co-culture of the 3T3-L1 adipocytes with untreated RAW 264.7 macrophages. Our results demonstrate that GLP-1 induces macrophage polarization toward the M2 phenotype, which may contribute to the protective effects of GLP-1 against diabetes and cardiovascular diseases.

  1. Monocyte/macrophage and protein interactions with non-fouling plasma polymerized tetraglyme and chemically modified polystyrene surfaces: In vitro and in vivo studies

    Science.gov (United States)

    Shen, Mingchao

    2001-07-01

    Biomaterials become encapsulated by fibrous tissues after implantation in soft tissues. Monocytes and macrophages are believed to play important roles in this response. The hypothesis tested in this dissertation is that material surface chemistry determines the amount of adsorbed proteins, which mediate monocyte adhesion, activation, and the foreign body response. On chemically modified polystyrene surfaces, monocyte adhesion in vitro was promoted by preadsorbed fibrinogen, fibronectin, and IgG, and increased with increasing amount of adsorbed fibrinogen. Adsorbed proteins and material surface chemistry mediated monocyte activation. TNFalpha release, procoagulant activity, and multinucleated foreign body giant cell (FBGC) formation was at least two-fold higher on IgG than other protein adsorbed surfaces. Adsorbed IgG and fibrinogen triggered monocyte intracellular calcium changes. FBGC formation was the highest on the hydrophobic polystyrene surface. Materials that greatly reduce non-specific protein adsorption may reduce the foreign body response to implanted materials. Radio-frequency plasma polymerized tetraglyme (CH3O(CH2CH2O)4CH 3) surfaces contained PEO-like chemical species and reduced fibrinogen adsorption to less than 10 ng/cm2. Monocyte adhesion to tetraglyme in vitro was also greatly reduced. Monocyte adhesion correlated linearly to the amount of adsorbed fibrinogen on a series of tetraglyme surfaces deposited at different plasma powers. Multivariate analysis using partial least squares regression identified the key surface spectra variables from electron spectroscopy for chemical analysis (ESCA) and time of flight secondary ion mass spectrometry (ToF-SIMS) that contributed to the non-fouling properties of tetraglyme. However, leukocyte adhesion to surfaces implanted subcutaneously in mice for 1 or 28 days did not correlate with protein adsorption and was higher on tetraglyme than the FEP control. Fibrous encapsulation to tetraglyme implanted for 28 days

  2. HIV-1 RNAs are Not Part of the Argonaute 2 Associated RNA Interference Pathway in Macrophages.

    Directory of Open Access Journals (Sweden)

    Valentina Vongrad

    Full Text Available MiRNAs and other small noncoding RNAs (sncRNAs are key players in post-transcriptional gene regulation. HIV-1 derived small noncoding RNAs (sncRNAs have been described in HIV-1 infected cells, but their biological functions still remain to be elucidated. Here, we approached the question whether viral sncRNAs may play a role in the RNA interference (RNAi pathway or whether viral mRNAs are targeted by cellular miRNAs in human monocyte derived macrophages (MDM.The incorporation of viral sncRNAs and/or their target RNAs into RNA-induced silencing complex was investigated using photoactivatable ribonucleoside-induced cross-linking and immunoprecipitation (PAR-CLIP as well as high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP, which capture Argonaute2-bound miRNAs and their target RNAs. HIV-1 infected monocyte-derived macrophages (MDM were chosen as target cells, as they have previously been shown to express HIV-1 sncRNAs. In addition, we applied small RNA deep sequencing to study differential cellular miRNA expression in HIV-1 infected versus non-infected MDMs.PAR-CLIP and HITS-CLIP data demonstrated the absence of HIV-1 RNAs in Ago2-RISC, although the presence of a multitude of HIV-1 sncRNAs in HIV-1 infected MDMs was confirmed by small RNA sequencing. Small RNA sequencing revealed that 1.4% of all sncRNAs were of HIV-1 origin. However, neither HIV-1 derived sncRNAs nor putative HIV-1 target sequences incorporated into Ago2-RISC were identified suggesting that HIV-1 sncRNAs are not involved in the canonical RNAi pathway nor is HIV-1 targeted by this pathway in HIV-1 infected macrophages.

  3. MIF inhibition reverts the gene expression profile of human melanoma cell line-induced MDSCs to normal monocytes

    Directory of Open Access Journals (Sweden)

    Sabine Waigel

    2016-03-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are potently immunosuppressive innate immune cells that accumulate in advanced cancer patients and actively inhibit anti-tumor T lymphocyte responses [1]. Increased numbers of circulating MDSCs directly correlate with melanoma patient morbidity and reduced anti-tumor immune responses [2,3]. Previous studies have revealed that monocyte-derived macrophage migration inhibitory factor (MIF is necessary for the immune suppressive function of MDSCs in mouse models of melanoma [4,5]. To investigate whether MIF participates in human melanoma-induced MDSC differentiation and/or suppressive function, we have established an in vitro MDSC induction model using primary, normal human monocytes co-cultured with human melanoma cell lines in the presence or absence of the MIF antagonist—4-IPP [4,6–9]. To identify potential mechanistic effectors, we have performed transcriptome analyses on cultured monocytes and on melanoma-induced MDSCs obtained from either untreated or 4-IPP-treated A375:monocyte co-cultures. Here, we present a detailed protocol, which can facilitate easy reproduction of the microarray results (NCBI GEO accession number GSE73333 published by Yaddanapudi et al. (2015 in Cancer Immunology Research [10].

  4. Function of microglia and macrophages in secondary damage after spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    Xiang Zhou; Xijing He; Yi Ren

    2014-01-01

    Spinal cord injury (SCI) is a devastating type of neurological trauma with limited therapeutic op-portunities. The pathophysiology of SCI involves primary and secondary mechanisms of injury. Among all the secondary injury mechanisms, the inlfammatory response is the major contrib-utor and results in expansion of the lesion and further loss of neurologic function. Meanwhile, the inlfammation directly and indirectly dominates the outcomes of SCI, including not only pain and motor dysfunction, but also preventingneuronal regeneration. Microglia and macrophages play very important roles in secondary injury. Microglia reside in spinal parenchyma and survey the microenvironment through the signals of injury or infection. Macrophages are derived from monocytes recruited to injured sites from the peripheral circulation. Activated resident microglia and monocyte-derived macrophages induce and magnify immune and inlfammatory responses not only by means of their secretory moleculesand phagocytosis, but also through their inlfuence on astrocytes, oligodendrocytes and demyelination. In this review, we focus on the roles of mi-croglia and macrophages in secondary injury and how they contribute to the sequelae of SCI.

  5. Mycobacterium tuberculosis-Induced Polarization of Human Macrophage Orchestrates the Formation and Development of Tuberculous Granulomas In Vitro.

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    Zikun Huang

    Full Text Available The tuberculous granuloma is an elaborately organized structure and one of the main histological hallmarks of tuberculosis. Macrophages, which are important immunologic effector and antigen-presenting cells, are the main cell type found in the tuberculous granuloma and have high plasticity. Macrophage polarization during bacterial infection has been elucidated in numerous recent studies; however, macrophage polarization during tuberculous granuloma formation and development has rarely been reported. It remains to be clarified whether differences in the activation status of macrophages affect granuloma formation. In this study, the variation in macrophage polarization during the formation and development of tuberculous granulomas was investigated in both sections of lung tissues from tuberculosis patients and an in vitro tuberculous granuloma model. The roles of macrophage polarization in this process were also investigated. Mycobacterium tuberculosis (M. tuberculosis infection was found to induce monocyte-derived macrophage polarization. In the in vitro tuberculous granuloma model, macrophage transformation from M1 to M2 was observed over time following M. tuberculosis infection. M2 macrophages were found to predominate in both necrotic and non-necrotic granulomas from tuberculosis patients, while both M1 and M2 polarized macrophages were found in the non-granulomatous lung tissues. Furthermore, it was found that M1 macrophages promote granuloma formation and macrophage bactericidal activity in vitro, while M2 macrophages inhibit these effects. The findings of this study provide insights into the mechanism by which M. tuberculosis circumvents the host immune system as well as a theoretical foundation for the development of novel tuberculosis therapies based on reprogramming macrophage polarization.

  6. Monocyte/macrophage and T-cell infiltrates in peritoneum of patients with ovarian cancer or benign pelvic disease

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    Ma Qing

    2006-07-01

    Full Text Available Abstract Background We previously showed that tumor-free peritoneum of patients with epithelial ovarian cancer (EOC exhibited enhanced expression of several inflammatory response genes compared to peritoneum of benign disease. Here, we examined peritoneal inflammatory cell patterns to determine their concordance with selected enhanced genes. Methods Expression patterns of selected inflammatory genes were mined from our previously published data base. Bilateral pelvic peritoneal and subjacent stromal specimens were obtained from 20 women with EOC and 7 women with benign pelvic conditions. Sections were first stained by indirect immunoperoxidase and numbers of monocytes/macrophages (MO/MA, T cells, B cells, and NK cells counted. Proportions of CD68+ cells and CD3+ cells that coexpressed MO/MA differentiation factors (CD163, CCR1, CXCR8, VCAM1, and phosphorylated cytosolic phospholipase A2 [pcPLA2], which had demonstrated expression in EOC peritoneal samples, were determined by multicolor immunofluorescence. Results MO/MA were present on both sides of the pelvic peritoneum in EOC patients, with infiltration of the subjacent stroma and mesothelium. CD68+ MO/MA, the most commonly represented population, and CD3+ T cells were present more often in EOC than in benign pelvic tumors. NK cells, B cells, and granulocytes were rare. CXCL8 (IL-8 and the chemokine receptor CCR1 were coexpressed more frequently on MO/MA than on CD3+ cells contrasting with CD68+/CD163+ cells that coexpressed CXCL8 less often. An important activated enzyme in the eicosanoid pathway, pcPLA2, was highly expressed on both CD68+ and CD163+ cells. The adherence molecule Vascular Cell Adhesion Molecule-1 (VCAM1 was expressed on CD31+ endothelial cells and on a proportion of CD68+ MO/MA but rarely on CD3+ cells. Conclusion The pelvic peritoneum in EOC exhibits a general pattern of chronic inflammation, represented primarily by differentiated MO/MA, and distinct from that in benign

  7. Cellular internalization and cytotoxicity of the antimicrobial proline-rich peptide Bac7(1-35) in monocytes/macrophages, and its activity against phagocytosed Salmonella typhimurium.

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    Pelillo, Chiara; Benincasa, Monica; Scocchi, Marco; Gennaro, Renato; Tossi, Alessandro; Pacor, Sabrina

    2014-04-01

    Bac7(1-35) is an active fragment of the bovine cathelicidin antimicrobial peptide Bac7, which selectively inactivates Gram-negative bacteria both in vitro and in mice infected with Salmonella typhimurium. It has a non-lytic mechanism of action, is rapidly internalized by susceptible bacteria and mammalian cells and likely acts by binding to internal targets. In this study we show that Bac7(1-35) accumulates selectively within primed macrophages with respect to resting monocytes. Confocal microscopy analysis showed that the peptide mainly distributes in the cytoplasm and perinuclear region of macrophages within 3 hours of incubation, without affecting cell viability. Cytotoxicity studies showed that the peptide does not induce necrotic or apoptotic damage up to concentrations 50-100-fold higher than minimal inhibitory concentrations (MIC). Moreover, Bac7(1-35) did not affect the ability of macrophages to engulf S. typhimurium, a species that may proliferate within this cell type. Conversely, when added to macrophages after phagocytosis, Bac7(1-35) caused a significant reduction in the number of recovered bacteria, indicating that it can kill the engulfed microorganisms directly and/or indirectly, via activation of the defense response of the cells.

  8. The membrane-bound ectopeptidase CPM as a marker of macrophage maturation in vitro and in vivo.

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    Rehli, M; Krause, S W; Andreesen, R

    2000-01-01

    During terminal maturation of human blood monocytes into macrophages, a multitude of phenotypic and functional changes occurs: cells increase in size, they enhance their capacity for phagocytosis and tumor cytotoxicity but decrease their ability for T-lymphocyte stimulation. The pattern of secreted cytokines is shifted as is the profile of surface antigens. We recently identified carboxypeptidase M (CPM) as a macrophage maturation-associated antigen detected by mAb MAX. 1/MAX. 11. CPM, a phosphoinositol-linked ectopeptidase, is able to process a multitude of different substrates, among them immunologically important peptides like bradykinin, anaphylatoxins and enkephalins. It was previously shown to be expressed in placenta, lung, and kidney. CPM as detected by MAX. 1/11 shows a strong expression on monocyte-derived macrophages in vitro and on macrophages in vivo accompanying T-lymphocyte activation like during allogeneic transplant rejection or allergic alveolitis. In contrast, its expression is suppressed on macrophages by some types of tumor cells. CPM expression seems to correlate with macrophage cytotoxic functions. However, the biological importance of CPM expression in human macrophages in vivo is difficult to predict. A wide range of biologically active peptides are cleaved by CPM, and the relevance of CPM peptide processing during an immune reaction is only poorly understood. The generation and analysis of CPM-deficient animals might improve our understanding of CPM function. Therefore we cloned a cDNA for the murine homologue of CPM. However, expression of mCPM was undetectable in murine primary macrophages and macrophage cell-lines, suggesting that CPM expression and function is not conserved between human and mouse macrophages.

  9. Macrophage-restricted interleukin-10 receptor deficiency, but not IL-10 deficiency, causes severe spontaneous colitis.

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    Zigmond, Ehud; Bernshtein, Biana; Friedlander, Gilgi; Walker, Catherine R; Yona, Simon; Kim, Ki-Wook; Brenner, Ori; Krauthgamer, Rita; Varol, Chen; Müller, Werner; Jung, Steffen

    2014-05-15

    Interleukin-10 (IL-10) is a pleiotropic anti-inflammatory cytokine produced and sensed by most hematopoietic cells. Genome-wide association studies and experimental animal models point at a central role of the IL-10 axis in inflammatory bowel diseases. Here we investigated the importance of intestinal macrophage production of IL-10 and their IL-10 exposure, as well as the existence of an IL-10-based autocrine regulatory loop in the gut. Specifically, we generated mice harboring IL-10 or IL-10 receptor (IL-10Rα) mutations in intestinal lamina propria-resident chemokine receptor CX3CR1-expressing macrophages. We found macrophage-derived IL-10 dispensable for gut homeostasis and maintenance of colonic T regulatory cells. In contrast, loss of IL-10 receptor expression impaired the critical conditioning of these monocyte-derived macrophages and resulted in spontaneous development of severe colitis. Collectively, our results highlight IL-10 as a critical homeostatic macrophage-conditioning agent in the colon and define intestinal CX3CR1(hi) macrophages as a decisive factor that determines gut health or inflammation.

  10. The effect of squalane-dissolved fullerene-C60 on adipogenesis-accompanied oxidative stress and macrophage activation in a preadipocyte-monocyte co-culture system.

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    Xiao, Li; Aoshima, Hisae; Saitoh, Yasukazu; Miwa, Nobuhiko

    2010-08-01

    Effects of squalane-dissolved fullerene-C60 (Sql-fullerene) on macrophage activation and adipose conversion with oxidative stress were studied using an inflammatory adipose-tissue equivalent (ATE) and OP9 mouse stromal preadipocyte-U937 lymphoma cell co-culture systems. Differentiation of OP9 cells was initiated by insulin-rich serum replacement (SR) as an adipogenic stimulant, and then followed by accumulation of intracellular lipid droplets and reactive oxygen species (ROS), both of which were significantly inhibited by Sql-fullerene. In the OP9-U937 cell co-culture system, U937 cells rapidly differentiated to macrophage-like cells during SR-induced adipogenesis in OP9 cells. The ROS accumulation was in the co-culture more marked than in OP9 cells alone, suggesting that the interaction between adipocytes and monocytes/macrophages promotes inflammatory responses. Sql-fullerene significantly inhibited macrophage activation and low-grade adipogenesis in the OP9-U937 co-culture system. We developed a three-dimensional inflammatory adipose-tissue model "ATE" consisting of, characteristically, U937 cells in the culture-wells, and, in addition, mounted a culture insert containing OP9 cells-populated collagen gel. ATE is enabled with suitable stimulation to represent the pathology of inflammatory disorders, such as macrophage infiltration in adipose tissue. Five-day culturing of ATE in SR medium occurred U937 macrophage migration and intracellular oil-droplet accumulation that were significantly inhibited by Sql-fullerene. Our results suggest that Sql-fullerene might be explored as a potential medicine for the treatment of metabolic syndrome or other obesity-related disorders.

  11. Fibroblasts and monocyte macrophages contract and degrade three-dimensional collagen gels in extended co-culture

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    Ertl Ronald F

    2001-09-01

    Full Text Available Abstract Background Inflammatory cells are believed to play a prominent role during tissue repair and remodeling. Since repair processes develop and mature over extended time frames, the present study was designed to evaluate the effect of monocytes and fibroblasts in prolonged culture in three-dimensional collagen gels. Methods Blood monocytes from healthy donors and human fetal lung fibroblasts were cast into type I collagen gels and maintained in floating cultures for three weeks. Results Fibroblast-mediated gel contraction was initially inhibited by the presence of monocytes (P P P 2 production was significantly increased by co-culture and its presence attenuated collagen degradation. Conclusion The current study, therefore, demonstrates that interaction between monocytes and fibroblasts can contract and degrade extracellular matrix in extended culture.

  12. Haemophilus ducreyi-induced interleukin-10 promotes a mixed M1 and M2 activation program in human macrophages.

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    Li, Wei; Katz, Barry P; Spinola, Stanley M

    2012-12-01

    During microbial infection, macrophages are polarized to classically activated (M1) or alternatively activated (M2) cells in response to microbial components and host immune mediators. Proper polarization of macrophages is critical for bacterial clearance. To study the role of macrophage polarization during Haemophilus ducreyi infection, we analyzed a panel of macrophage surface markers in skin biopsy specimens of pustules obtained from experimentally infected volunteers. Lesional macrophages expressed markers characteristic of both M1 and M2 polarization. Monocyte-derived macrophages (MDM) also expressed a mixed M1 and M2 profile of surface markers and cytokines/chemokines upon infection with H. ducreyi in vitro. Endogenous interleukin 10 (IL-10) produced by infected MDM downregulated and enhanced expression of several M1 and M2 markers, respectively. Bacterial uptake, mediated mainly by class A scavenger receptors, and activation of mitogen-activated protein kinase and phosphoinositide 3-kinase signaling pathways were required for H. ducreyi-induced IL-10 production in MDM. Compared to M1 cells, IL-10-polarized M2 cells displayed enhanced phagocytic activity against H. ducreyi and similar bacterial killing. Thus, IL-10-modulated macrophage polarization may contribute to H. ducreyi clearance during human infection.

  13. Platelet-activating factor increases reactive oxygen species-mediated microbicidal activity of human macrophages infected with Leishmania (Viannia) braziliensis.

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    Borges, Arissa Felipe; Morato, Camila Imai; Gomes, Rodrigo Saar; Dorta, Miriam Leandro; de Oliveira, Milton Adriano Pelli; Ribeiro-Dias, Fátima

    2017-09-29

    Platelet-activating factor (PAF) is produced by macrophages during inflammation and infections. We evaluated whether PAF is able to modulate the infection of human macrophages by Leishmania braziliensis, the main Leishmania sp. in Brazil. Monocyte-derived macrophages were incubated with promastigote forms in absence or presence of exogenous PAF. We observed that the treatment of macrophages with low concentrations of PAF prior to infection increased the phagocytosis of L. braziliensis. More importantly, exogenous PAF reduced the parasitism when it was added before, during or after infection. In addition, treatment with a PAF antagonist (PCA 4248) resulted in a significant increase of macrophage infection in a concentration-dependent manner, suggesting that endogenous PAF is important to control L. braziliensis infection. Mechanistically, while exogenous PAF increased production of reactive oxygen species (ROS) treatment with PCA 4248 reduced oxidative burst during L. braziliensis infection. The microbicidal effects of exogenous PAF were abolished when macrophages were treated with apocynin, an NADPH oxidase inhibitor. The data show that PAF promotes the production of ROS induced by L. braziliensis, suggesting that this lipid mediator may be relevant to control L. braziliensis infection in human macrophages. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  14. Human macrophages chronically exposed to LPS can be reactivated by stimulation with MDP to acquire an antimicrobial phenotype.

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    Guzmán-Beltrán, Silvia; Torres, Martha; Arellano, Monserrat; Juárez, Esmeralda

    2017-02-21

    Macrophages are important in host defense and can differentiate into functionally distinct subsets named classically (M1) or alternatively (M2) activated. In several inflammatory disorders, macrophages become tolerized to prevent deleterious consequences. This tolerization reduces the ability of macrophages to respond to bacterial components (e.g., LPS) maintaining a low level of inflammation but compromising the ability of macrophages to mount an effective immune response during subsequent pathogen encounters. In this study, we aimed to reactivate human monocyte-derived macrophages chronically exposed to LPS by re-stimulation with muramyl dipeptide (MDP). We observed an undefined profile of cell surface marker expression during endotoxin tolerance and absence of TNFα production. Stimulating macrophages chronically exposed to LPS with LPS+MDP restored TNFα, production together with an increased production of IL1, IL6, IFNγ, IL4, IL5 and IL10. These results suggest that macrophages chronically exposed to LPS possess a mixed M1-M2 phenotype with sufficient antimicrobial and homeostatic potential.

  15. A primary human macrophage-enteroid co-culture model to investigate mucosal gut physiology and host-pathogen interactions

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    Noel, Gaelle; Baetz, Nicholas W.; Staab, Janet F.; Donowitz, Mark; Kovbasnjuk, Olga; Pasetti, Marcela F.; Zachos, Nicholas C.

    2017-01-01

    Integration of the intestinal epithelium and the mucosal immune system is critical for gut homeostasis. The intestinal epithelium is a functional barrier that secludes luminal content, senses changes in the gut microenvironment, and releases immune regulators that signal underlying immune cells. However, interactions between epithelial and innate immune cells to maintain barrier integrity and prevent infection are complex and poorly understood. We developed and characterized a primary human macrophage-enteroid co-culture model for in-depth studies of epithelial and macrophage interactions. Human intestinal stem cell-derived enteroid monolayers co-cultured with human monocyte-derived macrophages were used to evaluate barrier function, cytokine secretion, and protein expression under basal conditions and following bacterial infection. Macrophages enhanced barrier function and maturity of enteroid monolayers as indicated by increased transepithelial electrical resistance and cell height. Communication between the epithelium and macrophages was demonstrated through morphological changes and cytokine production. Intraepithelial macrophage projections, efficient phagocytosis, and stabilized enteroid barrier function revealed a coordinated response to enterotoxigenic and enteropathogenic E. coli infections. In summary, we have established the first primary human macrophage-enteroid co-culture system, defined conditions that allow for a practical and reproducible culture model, and demonstrated its suitability to study gut physiology and host responses to enteric pathogens. PMID:28345602

  16. Simian Immunodeficiency Virus Targeting of CXCR3(+) CD4(+) T Cells in Secondary Lymphoid Organs Is Associated with Robust CXCL10 Expression in Monocyte/Macrophage Subsets.

    Science.gov (United States)

    Fujino, Masayuki; Sato, Hirotaka; Okamura, Tomotaka; Uda, Akihiko; Takeda, Satoshi; Ahmed, Nursarat; Shichino, Shigeyuki; Shiino, Teiichiro; Saito, Yohei; Watanabe, Satoru; Sugimoto, Chie; Kuroda, Marcelo J; Ato, Manabu; Nagai, Yoshiyuki; Izumo, Shuji; Matsushima, Kouji; Miyazawa, Masaaki; Ansari, Aftab A; Villinger, Francois; Mori, Kazuyasu

    2017-07-01

    Glycosylation of Env defines pathogenic properties of simian immunodeficiency virus (SIV). We previously demonstrated that pathogenic SIVmac239 and a live-attenuated, quintuple deglycosylated Env mutant (Δ5G) virus target CD4(+) T cells residing in different tissues during acute infection. SIVmac239 and Δ5G preferentially infected distinct CD4(+) T cells in secondary lymphoid organs (SLOs) and within the lamina propria of the small intestine, respectively (C. Sugimoto et al., J Virol 86:9323-9336, 2012, https://doi.org/10.1128/JVI.00948-12). Here, we studied the host responses relevant to SIV targeting of CXCR3(+) CCR5(+) CD4(+) T cells in SLOs. Genome-wide transcriptome analyses revealed that Th1-polarized inflammatory responses, defined by expression of CXCR3 chemokines, were distinctly induced in the SIVmac239-infected animals. Consistent with robust expression of CXCL10, CXCR3(