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Sample records for human alveolar macrophage

  1. Peptide secreted by human alveolar macrophages releases neutrophil granule contents

    International Nuclear Information System (INIS)

    MacArthur, C.K.; Miller, E.J.; Cohen, A.B.

    1987-01-01

    A monoclonal antibody was developed against an 8000-kDa enzyme-releasing peptide (ERP) released from human alveolar macrophages. ERP was isolated on an immunoaffinity column containing the antibody bound to staphylococcal protein A-Sepharose, and by autoradiography. Release of ERP from the macrophages is not changed by plastic adherence, phagocytosis, calcium ionophore, or phorbol esters. The peptide was not antigenically similar to interferon-γ, tumor necrosis factor, or interleukin lα or 1β. The release of constituents from azurophilic and specific granules was the main identified biologic function of ERP. ERP was a more effective secretagogue in the untreated neutrophils and f-met-leu-phe was more effective in the cytochalasin B-treated neutrophils. Absorption of ERP from macrophage-conditioned medium removed a small amount of the chemotactic activity; however, the immunopurified peptide was not chemotactic or chemokinetic for neutrophils, and at high concentrations, it suppressed base line chemokinesis. Treatment of washed macrophages with trypsin released active ERP of approximately the same m.w. of spontaneously secreted ERP. These studies showed that human alveolar macrophages release a peptide which is a secretagogue for human neutrophils under conditions which may be encountered in the lungs during certain disease states. Proteolytic enzymes which are free in the lungs may release the peptide and lead to the secretion of neutrophil enzymes

  2. Cigarette smoking decreases global microRNA expression in human alveolar macrophages.

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    Joel W Graff

    Full Text Available Human alveolar macrophages are critical components of the innate immune system. Cigarette smoking-induced changes in alveolar macrophage gene expression are linked to reduced resistance to pulmonary infections and to the development of emphysema/COPD. We hypothesized that microRNAs (miRNAs could control, in part, the unique messenger RNA (mRNA expression profiles found in alveolar macrophages of cigarette smokers. Activation of macrophages with different stimuli in vitro leads to a diverse range of M1 (inflammatory and M2 (anti-inflammatory polarized phenotypes that are thought to mimic activated macrophages in distinct tissue environments. Microarray mRNA data indicated that smoking promoted an "inverse" M1 mRNA expression program, defined by decreased expression of M1-induced transcripts and increased expression of M1-repressed transcripts with few changes in M2-regulated transcripts. RT-PCR arrays identified altered expression of many miRNAs in alveolar macrophages of smokers and a decrease in global miRNA abundance. Stratification of human subjects suggested that the magnitude of the global decrease in miRNA abundance was associated with smoking history. We found that many of the miRNAs with reduced expression in alveolar macrophages of smokers were predicted to target mRNAs upregulated in alveolar macrophages of smokers. For example, miR-452 is predicted to target the transcript encoding MMP12, an important effector of smoking-related diseases. Experimental antagonism of miR-452 in differentiated monocytic cells resulted in increased expression of MMP12. The comprehensive mRNA and miRNA expression profiles described here provide insight into gene expression regulation that may underlie the adverse effects cigarette smoking has on alveolar macrophages.

  3. NOD2 enhances the innate response of alveolar macrophages to Mycobacterium tuberculosis in humans.

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    Juárez, Esmeralda; Carranza, Claudia; Hernández-Sánchez, Fernando; León-Contreras, Juan C; Hernández-Pando, Rogelio; Escobedo, Dante; Torres, Martha; Sada, Eduardo

    2012-04-01

    A role for the nucleotide-binding oligomerization domain 2 (NOD2) receptor in pulmonary innate immune responses has recently been explored. In the present study, we investigated the role that NOD2 plays in human alveolar macrophage innate responses and determined its involvement in the response to infection with virulent Mycobacterium tuberculosis. Our results showed that NOD2 was expressed in human alveolar macrophages, and significant amounts of IL-1β, IL-6, and TNF-α were produced upon ligand recognition with muramyldipeptide (MDP). NOD2 ligation induced the transcription and protein expression of the antimicrobial peptide LL37 and the autophagy enzyme IRGM in alveolar macrophages, demonstrating a novel function for this receptor in these cells. MDP treatment of alveolar macrophages improved the intracellular growth control of virulent M. tuberculosis; this was associated with a significant release of TNF-α and IL-6 and overexpression of bactericidal LL37. In addition, the autophagy proteins IRGM, LC3 and ATG16L1 were recruited to the bacteria-containing autophagosome after treatment with MDP. In conclusion, our results suggest that NOD2 can modulate the innate immune response of alveolar macrophages and play a role in the initial control of respiratory M. tuberculosis infections. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Benzo(a)pyrene activation and detoxification by human pulmonary alveolar macrophages and lymphocytes

    International Nuclear Information System (INIS)

    Marshall, M.V.; McLemore, T.L.; Martin, R.R.; Marshall, M.H.; Wray, N.P.; Busbee, D.L.; Cantrell, E.T.; Arnott, M.S.; Griffin, A.C.

    1980-01-01

    Comparisons of pulmonary alveolar macrophages and circulating lymphocytes from five smokers and five nonsmokers for their ability to metabolize benzo(a)pyrene as determined by high pressure liquid chromatography were carried out. Utilizing this approach, further investigation of activation and detoxification by several human cell types could provide the basis for more precise and comprehensive studies of carcinogen and drug metabolism in the human lung, and for a better assessment of cancer risk in selected populations

  5. The Alveolar Microenvironment of Patients Infected with Human Immunodeficiency Virus Does Not Modify Alveolar Macrophage Interactions with Streptococcus pneumoniae

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    Jagoe, R. Thomas; Jarman, Elizabeth R.; North, James C.; Pridmore, Alison; Musaya, Janelisa; French, Neil; Zijlstra, Eduard E.; Molyneux, Malcolm E.; Read, Robert C.

    2013-01-01

    We tested the hypothesis that HIV infection results in activation of alveolar macrophages and that this might be associated with impaired defense against pneumococcus. We compared alveolar macrophages and lymphocytes in 131 bronchoalveolar lavage samples from HIV-infected and healthy controls using inflammatory gene microarrays, flow cytometry, real-time PCR, and enzyme-linked immunosorbent assay (ELISA) to determine the pattern of macrophage activation associated with HIV infection and the effect of this activation on defense against pneumococcus. We used gamma interferon (IFN-γ) priming to mimic the cellular milieu in HIV-infected lungs. InnateDB and BioLayout 3D were used to analyze the interactions of the upregulated genes. Alveolar macrophages from HIV-infected adults showed increased gene expression and cytokine production in a classical pattern. Bronchoalveolar lavage from HIV-infected subjects showed excess CD8+ lymphocytes with activated phenotype. Toll-like receptor 4 (TLR4) expression was increased in macrophages from HIV-infected subjects, but function was similar between the groups; lung lavage fluid did not inhibit TLR function in transfected HeLa cells. Alveolar macrophages from HIV-infected subjects showed normal binding and internalization of opsonized pneumococci, with or without IFN-γ priming. Alveolar macrophages from HIV-infected subjects showed classical activation compared to that of healthy controls, but this does not alter macrophage interactions with pneumococci. PMID:23576675

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

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    Jones William

    2006-03-01

    Full Text Available Abstract Background Synthetic vitreous fibers (SVFs are inorganic noncrystalline materials widely used in residential and industrial settings for insulation, filtration, and reinforcement purposes. SVFs conventionally include three major categories: fibrous glass, rock/slag/stone (mineral wool, and ceramic fibers. Previous in vitro studies from our laboratory demonstrated length-dependent cytotoxic effects of glass fibers on rat alveolar macrophages which were possibly associated with incomplete phagocytosis of fibers ≥ 17 μm in length. The purpose of this study was to examine the influence of fiber length on primary human alveolar macrophages, which are larger in diameter than rat macrophages, using length-classified Manville Code 100 glass fibers (8, 10, 16, and 20 μm. It was hypothesized that complete engulfment of fibers by human alveolar macrophages could decrease fiber cytotoxicity; i.e. shorter fibers that can be completely engulfed might not be as cytotoxic as longer fibers. Human alveolar macrophages, obtained by segmental bronchoalveolar lavage of healthy, non-smoking volunteers, were treated with three different concentrations (determined by fiber number of the sized fibers in vitro. Cytotoxicity was assessed by monitoring cytosolic lactate dehydrogenase release and loss of function as indicated by a decrease in zymosan-stimulated chemiluminescence. Results Microscopic analysis indicated that human alveolar macrophages completely engulfed glass fibers of the 20 μm length. All fiber length fractions tested exhibited equal cytotoxicity on a per fiber basis, i.e. increasing lactate dehydrogenase and decreasing chemiluminescence in the same concentration-dependent fashion. Conclusion The data suggest that due to the larger diameter of human alveolar macrophages, compared to rat alveolar macrophages, complete phagocytosis of longer fibers can occur with the human cells. Neither incomplete phagocytosis nor length-dependent toxicity was

  7. Mycobacterium tuberculosis infection causes different levels of apoptosis and necrosis in human macrophages and alveolar epithelial cells.

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    Danelishvili, Lia; McGarvey, Jeffery; Li, Yong-Jun; Bermudez, Luiz E

    2003-09-01

    Mycobacterium tuberculosis interacts with macrophages and epithelial cells in the alveolar space of the lung, where it is able to invade and replicate in both cell types. M. tuberculosis-associated cytotoxicity to these cells has been well documented, but the mechanisms of host cell death are not well understood. We examined the induction of apoptosis and necrosis of human macrophages (U937) and type II alveolar epithelial cells (A549) by virulent (H37Rv) and attenuated (H37Ra) M. tuberculosis strains. Apoptosis was determined by both enzyme-linked immunosorbent assay (ELISA) and TdT-mediated dUTP nick end labelling (TUNEL) assay, whereas necrosis was evaluated by the release of lactate dehydrogenase (LDH). Both virulent and attenuated M. tuberculosis induced apoptosis in macrophages; however, the attenuated strain resulted in significantly more apoptosis than the virulent strain after 5 days of infection. In contrast, cytotoxicity of alveolar cells was the result of necrosis, but not apoptosis. Although infection with M. tuberculosis strains resulted in apoptosis of 14% of the cells on the monolayer, cell death associated with necrosis was observed in 59% of alveolar epithelial cells after 5 days of infection. Infection with M. tuberculosis suppressed apoptosis of alveolar epithelial cells induced by the kinase inhibitor, staurosporine. Because our findings suggest that M. tuberculosis can modulate the apoptotic response of macrophages and epithelial cells, we carried out an apoptosis pathway-specific cDNA microarray analysis of human macrophages and alveolar epithelial cells. Whereas the inhibitors of apoptosis, bcl-2 and Rb, were upregulated over 2.5-fold in infected (48 h) alveolar epithelial cells, the proapoptotic genes, bad and bax, were downregulated. The opposite was observed when U937 macrophages were infected with M. tuberculosis. Upon infection of alveolar epithelial cells with M. tuberculosis, the generation of apoptosis, as determined by the

  8. Morphometric Characterization of Rat and Human Alveolar Macrophage Cell Models and their Response to Amiodarone using High Content Image Analysis.

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    Hoffman, Ewelina; Patel, Aateka; Ball, Doug; Klapwijk, Jan; Millar, Val; Kumar, Abhinav; Martin, Abigail; Mahendran, Rhamiya; Dailey, Lea Ann; Forbes, Ben; Hutter, Victoria

    2017-12-01

    Progress to the clinic may be delayed or prevented when vacuolated or "foamy" alveolar macrophages are observed during non-clinical inhalation toxicology assessment. The first step in developing methods to study this response in vitro is to characterize macrophage cell lines and their response to drug exposures. Human (U937) and rat (NR8383) cell lines and primary rat alveolar macrophages obtained by bronchoalveolar lavage were characterized using high content fluorescence imaging analysis quantification of cell viability, morphometry, and phospholipid and neutral lipid accumulation. Cell health, morphology and lipid content were comparable (p content. Responses to amiodarone, a known inducer of phospholipidosis, required analysis of shifts in cell population profiles (the proportion of cells with elevated vacuolation or lipid content) rather than average population data which was insensitive to the changes observed. A high content image analysis assay was developed and used to provide detailed morphological characterization of rat and human alveolar-like macrophages and their response to a phospholipidosis-inducing agent. This provides a basis for development of assays to predict or understand macrophage vacuolation following inhaled drug exposure.

  9. 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

  10. Nucleotide-oligomerizing domain-1 (NOD1) receptor activation induces pro-inflammatory responses and autophagy in human alveolar macrophages.

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    Juárez, Esmeralda; Carranza, Claudia; Hernández-Sánchez, Fernando; Loyola, Elva; Escobedo, Dante; León-Contreras, Juan Carlos; Hernández-Pando, Rogelio; Torres, Martha; Sada, Eduardo

    2014-09-25

    Nucleotide-binding oligomerizing domain-1 (NOD1) is a cytoplasmic receptor involved in recognizing bacterial peptidoglycan fragments that localize to the cytosol. NOD1 activation triggers inflammation, antimicrobial mechanisms and autophagy in both epithelial cells and murine macrophages. NOD1 mediates intracellular pathogen clearance in the lungs of mice; however, little is known about NOD1's role in human alveolar macrophages (AMs) or its involvement in Mycobacterium tuberculosis (Mtb) infection. AMs, monocytes (MNs), and monocyte-derived macrophages (MDMs) from healthy subjects were assayed for NOD1 expression. Cells were stimulated with the NOD1 ligand Tri-DAP and cytokine production and autophagy were assessed. Cells were infected with Mtb and treated with Tri-DAP post-infection. CFUs counting determined growth control, and autophagy protein recruitment to pathogen localization sites was analyzed by immunoelectron microscopy. NOD1 was expressed in AMs, MDMs and to a lesser extent MNs. Tri-DAP stimulation induced NOD1 up-regulation and a significant production of IL1β, IL6, IL8, and TNFα in AMs and MDMs; however, the level of NOD1-dependent response in MNs was limited. Autophagy activity determined by expression of proteins Atg9, LC3, IRGM and p62 degradation was induced in a NOD1-dependent manner in AMs and MDMs but not in MNs. Infected AMs could be activated by stimulation with Tri-DAP to control the intracellular growth of Mtb. In addition, recruitment of NOD1 and the autophagy proteins IRGM and LC3 to the Mtb localization site was observed in infected AMs after treatment with Tri-DAP. NOD1 is involved in AM and MDM innate responses, which include proinflammatory cytokines and autophagy, with potential implications in the killing of Mtb in humans.

  11. Adherent Human Alveolar Macrophages Exhibit a Transient Pro-Inflammatory Profile That Confounds Responses to Innate Immune Stimulation

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    Tomlinson, Gillian S.; Booth, Helen; Petit, Sarah J.; Potton, Elspeth; Towers, Greg J.; Miller, Robert F.; Chain, Benjamin M.; Noursadeghi, Mahdad

    2012-01-01

    Alveolar macrophages (AM) are thought to have a key role in the immunopathogenesis of respiratory diseases. We sought to test the hypothesis that human AM exhibit an anti-inflammatory bias by making genome-wide comparisons with monocyte derived macrophages (MDM). Adherent AM obtained by bronchoalveolar lavage of patients under investigation for haemoptysis, but found to have no respiratory pathology, were compared to MDM from healthy volunteers by whole genome transcriptional profiling before and after innate immune stimulation. We found that freshly isolated AM exhibited a marked pro-inflammatory transcriptional signature. High levels of basal pro-inflammatory gene expression gave the impression of attenuated responses to lipopolysaccharide (LPS) and the RNA analogue, poly IC, but in rested cells pro-inflammatory gene expression declined and transcriptional responsiveness to these stimuli was restored. In comparison to MDM, both freshly isolated and rested AM showed upregulation of MHC class II molecules. In most experimental paradigms ex vivo adherent AM are used immediately after isolation. Therefore, the confounding effects of their pro-inflammatory profile at baseline need careful consideration. Moreover, despite the prevailing view that AM have an anti-inflammatory bias, our data clearly show that they can adopt a striking pro-inflammatory phenotype, and may have greater capacity for presentation of exogenous antigens than MDM. PMID:22768282

  12. Differential cell reaction upon Toll-like receptor 4 and 9 activation in human alveolar and lung interstitial macrophages

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    Meyerhans Andreas

    2010-09-01

    Full Text Available Abstract Background Investigations on pulmonary macrophages (MΦ mostly focus on alveolar MΦ (AM as a well-defined cell population. Characteristics of MΦ in the interstitium, referred to as lung interstitial MΦ (IM, are rather ill-defined. In this study we therefore aimed to elucidate differences between AM and IM obtained from human lung tissue. Methods Human AM and IM were isolated from human non-tumor lung tissue from patients undergoing lung resection. Cell morphology was visualized using either light, electron or confocal microscopy. Phagocytic activity was analyzed by flow cytometry as well as confocal microscopy. Surface marker expression was measured by flow cytometry. Toll-like receptor (TLR expression patterns as well as cytokine expression upon TLR4 or TLR9 stimulation were assessed by real time RT-PCR and cytokine protein production was measured using a fluorescent bead-based immunoassay. Results IM were found to be smaller and morphologically more heterogeneous than AM, whereas phagocytic activity was similar in both cell types. HLA-DR expression was markedly higher in IM compared to AM. Although analysis of TLR expression profiles revealed no differences between the two cell populations, AM and IM clearly varied in cell reaction upon activation. Both MΦ populations were markedly activated by LPS as well as DNA isolated from attenuated mycobacterial strains (M. bovis H37Ra and BCG. Whereas AM expressed higher amounts of inflammatory cytokines upon activation, IM were more efficient in producing immunoregulatory cytokines, such as IL10, IL1ra, and IL6. Conclusion AM appear to be more effective as a non-specific first line of defence against inhaled pathogens, whereas IM show a more pronounced regulatory function. These dissimilarities should be taken into consideration in future studies on the role of human lung MΦ in the inflammatory response.

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

    Lifescience Database Archive (English)

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

  14. Long-term persistence of human donor alveolar macrophages in lung transplant recipients

    DEFF Research Database (Denmark)

    Eguíluz-Gracia, Ibon; Schultz, Hans Henrik Lawaetz; Sikkeland, Liv I. B.

    2016-01-01

    and life span of human AMFs is scarce. METHODS: To follow the origin and longevity of AMFs in patients with lung transplantation for more than 100 weeks, we obtained transbronchial biopsies from 10 gender-mismatched patients with lung transplantation. These were subjected to combined in situ hybridisation...... transplantation we found that recipient monocytes seeded the alveoli early after transplantation, and showed subsequent phenotypical changes consistent with differentiation into proliferating mature AMFs. This resulted in a stable mixed chimerism between donor and recipient AMFs throughout the 2-year period...

  15. SP-A binding sites on bovine alveolar macrophages.

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    Plaga, S; Plattner, H; Schlepper-Schaefer, J

    1998-11-25

    Surfactant protein A (SP-A) binding to bovine alveolar macrophages was examined in order to characterize SP-A binding proteins on the cell surface and to isolate putative receptors from these cells that could be obtained in large amounts. Human SP-A, unlabeled or labeled with gold particles, was bound to freshly isolated macrophages and analyzed with ELISA or the transmission electron microscope. Binding of SP-A was inhibited by Ca2+ chelation, by an excess of unlabeled SP-A, or by the presence of 20 mg/ml mannan. We conclude that bovine alveolar macrophages expose binding sites for SP-A that are specific and that depend on Ca2+ and on mannose residues. For isolation of SP-A receptors with homologous SP-A as ligand we isolated SP-A from bovine lung lavage. SDS-PAGE analysis of the purified SP-A showed a protein of 32-36 kDa. Functional integrity of the protein was demonstrated. Bovine SP-A bound to Dynabeads was used to isolate SP-A binding proteins. From the fractionated and blotted proteins of the receptor preparation two proteins bound SP-A in a Ca2+-dependent manner, a 40-kDa protein showing mannose dependency and a 210-kDa protein, showing no mannose sensitivity. Copyright 1998 Academic Press.

  16. In Vitro Toxicity of Aluminum Nanoparticles in Rat Alveolar Macrophages

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    2006-03-01

    including intravenous, intramuscular , and subcutaneous injections, and including oral and ocular administration (Kreuter, 1991). NPs allow delivery of... NANOPARTICLES IN RAT ALVEOLAR MACROPHAGES THESIS Andrew J Wagner, 1st Lt, USAF AFIT/GES/ENV/06M-06 DEPARTMENT OF THE AIR FORCE AIR UNIVERSITY ORCE...TOXICITY OF ALUMINUM NANOPARTICLES IN RAT ALVEOLAR MACROPHAGES THESIS Presented to the Faculty Department of Systems and Engineering

  17. Low Levels of IGF-1 Contribute to Alveolar Macrophage Dysfunction in Cystic Fibrosis1

    OpenAIRE

    Bessich, Jamie L.; Nymon, Amanda B.; Moulton, Lisa A; Dorman, Dana; Ashare, Alix

    2013-01-01

    Alveolar macrophages are major contributors to lung innate immunity. Although alveolar macrophages from CFTR−/− mice have impaired function, no study has investigated primary alveolar macrophages in adults with cystic fibrosis (CF). CF patients have low levels of insulin-like growth factor 1 (IGF-1), and our prior studies demonstrate a relationship between IGF-1 and macrophage function. We hypothesize that reduced IGF-1 in CF leads to impaired alveolar macrophage function and chronic infectio...

  18. Mobility of macrophages and alveolar decontamination in different kinds of animals

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    Nolibe, D; Metivier, H; Masse, R

    1973-05-01

    From congress on alveolar macrophage; Lille, France (28 May The mobility of macrophages in relation to alveolar decontamination following the inhalation of toxic substances was studied in the dog, monkey, cat, rat, and guinea pig. The alveolar macroPhages showed a migration rate that varied from 30 to 10% in the rat and rabbit. The measurement of alveolar decontamination should take into consideration inter-species differences in macrophage mobility. (JSR)

  19. Role of Alveolar Macrophages in Chronic Obstructive Pulmonary Disease

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    Vlahos, Ross; Bozinovski, Steven

    2014-01-01

    Alveolar macrophages (AMs) represent a unique leukocyte population that responds to airborne irritants and microbes. This distinct microenvironment coordinates the maturation of long-lived AMs, which originate from fetal blood monocytes and self-renew through mechanisms dependent on GM-CSF and CSF-1 signaling. Peripheral blood monocytes can also replenish lung macrophages; however, this appears to occur in a stimuli specific manner. In addition to mounting an appropriate immune response durin...

  20. Regulation of cytokine production in human alveolar macrophages and airway epithelial cells in response to ambient air pollution particles: Further mechanistic studies

    International Nuclear Information System (INIS)

    Becker, Susanne; Mundandhara, Sailaja; Devlin, Robert B.; Madden, Michael

    2005-01-01

    In order to better understand how ambient air particulate matter (PM) affect lung health, the two main airway cell types likely to interact with inhaled particles, alveolar macrophages (AM) and airway epithelial cells have been exposed to particles in vitro and followed for endpoints of inflammation, and oxidant stress. Separation of Chapel Hill PM 10 into fine and coarse size particles revealed that the main proinflammatory response (TNF, IL-6, COX-2) in AM was driven by material present in the coarse PM, containing 90-95% of the stimulatory material in PM10. The particles did not affect expression of hemoxygenase-1 (HO-1), a sensitive marker of oxidant stress. Primary cultures of normal human bronchial epithelial cells (NHBE) also responded to the coarse fraction with higher levels of IL-8 and COX-2, than induced by fine or ultrafine PM. All size PM induced oxidant stress in NHBE, while fine PM induced the highest levels of HO-1 expression. The production of cytokines in AM by both coarse and fine particles was blocked by the toll like receptor 4 (TLR4) antagonist E5531 involved in the recognition of LPS and Gram negative bacteria. The NHBE were found to recognize coarse and fine PM through TLR2, a receptor with preference for recognition of Gram positive bacteria. Compared to ambient PM, diesel PM induced only a minimal cytokine response in both AM and NHBE. Instead, diesel suppressed LPS-induced TNF and IL-8 release in AM. Both coarse and fine ambient air PM were also found to inhibit LPS-induced TNF release while silica, volcanic ash or carbon black had no inhibitory effect. Diesel particles did not affect cytokine mRNA induction nor protein accumulation but interfered with the release of cytokine from the cells. Ambient coarse and fine PM, on the other hand, inhibited both mRNA induction and protein production. Exposure to coarse and fine PM decreased the expression of TLR4 in the macrophages. Particle-induced decrease in TLR4 and hyporesponsiveness to LPS

  1. Low Levels of IGF-1 Contribute to Alveolar Macrophage Dysfunction in Cystic Fibrosis1

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    Bessich, Jamie L.; Nymon, Amanda B.; Moulton, Lisa A; Dorman, Dana; Ashare, Alix

    2013-01-01

    Alveolar macrophages are major contributors to lung innate immunity. Although alveolar macrophages from CFTR−/− mice have impaired function, no study has investigated primary alveolar macrophages in adults with cystic fibrosis (CF). CF patients have low levels of insulin-like growth factor 1 (IGF-1), and our prior studies demonstrate a relationship between IGF-1 and macrophage function. We hypothesize that reduced IGF-1 in CF leads to impaired alveolar macrophage function and chronic infections. Serum and bronchoalveolar lavage (BAL) samples were obtained from 8 CF subjects and 8 healthy subjects. Macrophages were isolated from BAL fluid. We measured the ability of alveolar macrophages to kill Pseudomonas aeruginosa. Subsequently, macrophages were incubated with IGF-1 prior to inoculation with bacteria to determine the effect of IGF-1 on bacterial killing. We found a significant decrease in bacterial killing by CF alveolar macrophages compared to controls. CF subjects had lower serum and BAL IGF-1 levels compared to healthy controls. Exposure to IGF-1 enhanced alveolar macrophage macrophages in both groups. Finally, exposing healthy alveolar macrophages to CF BAL fluid decreased bacterial killing, and this was reversed by the addition of IGF-1, while IGF-1 blockade worsened bacterial killing. Our studies demonstrate that alveolar macrophage function is impaired in patients with CF. Reductions in IGF-1 levels in CF contribute to the impaired alveolar macrophage function. Exposure to IGF-1 ex vivo, results in improved function of CF alveolar macrophages. Further studies are needed to determine whether alveolar macrophage function can be enhanced in vivo with IGF-1 treatment. PMID:23698746

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

    International Nuclear Information System (INIS)

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

    2012-01-01

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

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

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    Miguel Pinilla-Vera

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

  4. Fenspiride and membrane transduction signals in rat alveolar macrophages.

    Science.gov (United States)

    Féray, J C; Mohammadi, K; Taouil, K; Brunet, J; Garay, R P; Hannaert, P

    1997-07-15

    Fenspiride inhibits the calcium signal evoked by the inflammatory peptide formyl-Met-Leu-Phe (fMLP) in peritoneal macrophages, but at concentrations (approximately 1 mM) far above the therapeutic range (approximately 1 microM). Here, in rat alveolar macrophages, high fenspiride concentrations (1 mM) were required to inhibit the calcium signals evoked by the calcium agonist Bay K8644 or by ionomycin. Moreover, fenspiride (1 mM) was a poor inhibitor of the cell membrane depolarization induced by gramicidine D. By contrast, fenspiride blocked Na+-H+ antiport activation by (i) fMLP with an IC50 = 3.1 +/- 1.9 nM and (ii) PMA (phorbol 12-myristate 13-acetate) with an IC50 = 9.2 +/- 3.1 nM. Finally, protein kinase C (PKC) activity of macrophage homogenate was not significantly modified by 10 or 100 microM fenspiride (at 100 microM: 2.57 +/- 1.60 vs. 2.80 +/- 1.71 pmol/10(6) cells/min). In conclusion, fenspiride inhibits fMLP- and PMA-induced pH signals in rat alveolar macrophages, probably by acting distally on the PKC transduction signal. This pH antagonistic action may be relevant for the antiinflammatory mechanism of fenspiride and requires further investigation.

  5. Domestic smoke exposure is associated with alveolar macrophage particulate load.

    Science.gov (United States)

    Fullerton, Duncan G; Jere, Khuzwayo; Jambo, Kondwani; Kulkarni, Neeta S; Zijlstra, Eduard E; Grigg, Jonathan; French, Neil; Molyneux, Malcolm E; Gordon, Stephen B

    2009-03-01

    Indoor air pollution is associated with impaired respiratory health. The pre-dominant indoor air pollutant to which two billion of the world's population is exposed is biomass fuel smoke. We tested the hypothesis that reported smoke exposure in men and women is associated with increased alveolar macrophage uptake of biomass smoke particulates. Healthy volunteers attending for research bronchoscopy in Malawi completed a questionnaire assessment of smoke exposure. Particulate matter visible in alveolar macrophages (AM) was quantified using digital image analysis. The geometric mean of the percentage area of the cytoplasm occupied by particulates in 50 cover-slip adherent AM was calculated and termed particulate load. In 57 subjects (40 men and 17 women) there was a significant difference between the particulate load in groups divided according to pre-dominant lighting form used at home (ANOVA P = 0.0009) and type of cooking fuel (P = 0.0078). Particulate load observed in macrophages is associated with the reported type of biomass fuel exposure. Macrophage function in relation to respiratory health should now be investigated in biomass smoke exposed subjects.

  6. Role of Alveolar Macrophages in Chronic Obstructive Pulmonary Disease

    Science.gov (United States)

    Vlahos, Ross; Bozinovski, Steven

    2014-01-01

    Alveolar macrophages (AMs) represent a unique leukocyte population that responds to airborne irritants and microbes. This distinct microenvironment coordinates the maturation of long-lived AMs, which originate from fetal blood monocytes and self-renew through mechanisms dependent on GM-CSF and CSF-1 signaling. Peripheral blood monocytes can also replenish lung macrophages; however, this appears to occur in a stimuli specific manner. In addition to mounting an appropriate immune response during infection and injury, AMs actively coordinate the resolution of inflammation through efferocytosis of apoptotic cells. Any perturbation of this process can lead to deleterious responses. In chronic obstructive pulmonary disease (COPD), there is an accumulation of airway macrophages that do not conform to the classic M1/M2 dichotomy. There is also a skewed transcriptome profile that favors expression of wound-healing M2 markers, which is reflective of a deficiency to resolve inflammation. Endogenous mediators that can promote an imbalance in inhibitory M1 vs. healing M2 macrophages are discussed, as they are the plausible mechanisms underlying why AMs fail to effectively resolve inflammation and restore normal lung homeostasis in COPD. PMID:25309536

  7. Reduced number and morphofunctional change of alveolar macrophages in MafB gene-targeted mice.

    Directory of Open Access Journals (Sweden)

    Michiko Sato-Nishiwaki

    Full Text Available Alveolar macrophages (AMs play an important role in the pathogenesis of chronic obstructive pulmonary disease (COPD. We previously demonstrated that the transcription factor, MafB, increased in the AMs of mice exposed to cigarette smoke, and in those of human patients with COPD. The aim of this study was to evaluate the role of MafB in AMs using newly established transgenic (TG mice that specifically express dominant negative (DN MafB in macrophages under the control of macrophage scavenger receptor (MSR enhancer-promoter. We performed cell differential analyses in bronchoalveolar lavage cells, morphological analyses with electron microscopy, and flow cytometry-based analyses of surface markers and a phagocytic capacity assay in macrophages. AM number in the TG mice was significantly decreased compared with wild-type (WT mice. Morphologically, the high electron density area in the nucleus increased, the shape of pseudopods on the AMs was altered, and actin filament was less localized in the pseudopods of AMs of TG mice, compared with WT mice. The expression of surface markers, F4/80 and CD11b, on peritoneal macrophages in TG mice was reduced compared with WT mice, while those on AMs remained unchanged. Phagocytic capacity was decreased in AMs from TG mice, compared with WT mice. In conclusion, MafB regulates the phenotype of macrophages with respect to the number of alveolar macrophages, the nuclear compartment, cellular shape, surface marker expression, and phagocytic function. MSR-DN MafB TG mice may present a useful model to clarify the precise role of MafB in macrophages.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-05-15

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

  9. Effects of vitamin D supplementation on alveolar macrophage gene expression: preliminary results of a randomized, controlled trial.

    Science.gov (United States)

    Gerke, Alicia K; Pezzulo, Alejandro A; Tang, Fan; Cavanaugh, Joseph E; Bair, Thomas B; Phillips, Emily; Powers, Linda S; Monick, Martha M

    2014-03-26

    Vitamin D deficiency has been implicated as a factor in a number of infectious and inflammatory lung diseases. In the lung, alveolar macrophages play a key role in inflammation and defense of infection, but there are little data exploring the immunomodulatory effects of vitamin D on innate lung immunity in humans. The objective of this study was to determine the effects of vitamin D supplementation on gene expression of alveolar macrophages. We performed a parallel, double-blind, placebo-controlled, randomized trial to determine the effects of vitamin D on alveolar macrophage gene expression. Vitamin D3 (1000 international units/day) or placebo was administered to adults for three months. Bronchoscopy was performed pre- and post-intervention to obtain alveolar macrophages. Messenger RNA was isolated from the macrophages and subjected to whole genome exon array analysis. The primary outcome was differential gene expression of the alveolar macrophage in response to vitamin D supplementation. Specific genes underwent validation by polymerase chain reaction methods. Fifty-eight subjects were randomized to vitamin D (n = 28) or placebo (n = 30). There was a marginal overall difference between treatment group and placebo group in the change of 25-hydroxyvitaminD levels (4.43 ng/ml vs. 0.2 ng/ml, p = 0.10). Whole genome exon array analysis revealed differential gene expression associated with change in serum vitamin D levels in the treated group. CCL8/MCP-2 was the top-regulated cytokine gene and was further validated. Although only a non-significant increased trend was seen in serum vitamin D levels, subjects treated with vitamin D supplementation had immune-related differential gene expression in alveolar macrophages. ClinicalTrials.org: NCT01967628.

  10. RNA sequencing provides exquisite insight into the manipulation of the alveolar macrophage by tubercle bacilli.

    Science.gov (United States)

    Nalpas, Nicolas C; Magee, David A; Conlon, Kevin M; Browne, John A; Healy, Claire; McLoughlin, Kirsten E; Rue-Albrecht, Kévin; McGettigan, Paul A; Killick, Kate E; Gormley, Eamonn; Gordon, Stephen V; MacHugh, David E

    2015-09-08

    Mycobacterium bovis, the agent of bovine tuberculosis, causes an estimated $3 billion annual losses to global agriculture due, in part, to the limitations of current diagnostics. Development of next-generation diagnostics requires a greater understanding of the interaction between the pathogen and the bovine host. Therefore, to explore the early response of the alveolar macrophage to infection, we report the first application of RNA-sequencing to define, in exquisite detail, the transcriptomes of M. bovis-infected and non-infected alveolar macrophages from ten calves at 2, 6, 24 and 48 hours post-infection. Differentially expressed sense genes were detected at these time points that revealed enrichment of innate immune signalling functions, and transcriptional suppression of host defence mechanisms (e.g., lysosome maturation). We also detected differentially expressed natural antisense transcripts, which may play a role in subverting innate immune mechanisms following infection. Furthermore, we report differential expression of novel bovine genes, some of which have immune-related functions based on orthology with human proteins. This is the first in-depth transcriptomics investigation of the alveolar macrophage response to the early stages of M. bovis infection and reveals complex patterns of gene expression and regulation that underlie the immunomodulatory mechanisms used by M. bovis to evade host defence mechanisms.

  11. [Macrophages in human semen].

    Science.gov (United States)

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

    2003-11-01

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

  12. Anthrax Lethal Toxin Impairs Innate Immune Functions of Alveolar Macrophages and Facilitates Bacillus anthracis Survival

    National Research Council Canada - National Science Library

    Ribot, Wilson J; Panchal, Rekha G; Brittingham, Katherine C; Ruthel, Gordon; Kenny, Tara A; Lane, Douglas; Curry, Bob; Hoover, Timothy A; Friedlander, Arthur M; Bavari, Sina

    2006-01-01

    Alveolar macrophages (AM) are very important for pulmonary innate immune responses against invading inhaled pathogens because they directly kill the organisms and initiate a cascade of innate and adaptive immune responses...

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

    Science.gov (United States)

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

    1995-01-01

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

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

    DEFF Research Database (Denmark)

    Oleksiewicz, Martin B.; Nielsen, Jens

    1999-01-01

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

  15. Arachidonic acid metabolism in silica-stimulated bovine alveolar macrophages

    International Nuclear Information System (INIS)

    Englen, M.D.

    1989-01-01

    The in vitro production of arachidonic acid (AA) metabolites in adherent bovine alveolar macrophages (BAM) incubated with silica was investigated. BAM were pre-labelled with 3 H-AA, and lipid metabolites released into the culture medium were analyzed by high performance liquid chromatography (HPLC). Lactate dehydrogenase (LDH) release was simultaneously assayed to provide an indication of cell injury. Increasing doses of silica selectively stimulated the 5-lipoxygenase pathway of AA metabolism, while cyclooxygenase metabolite output was suppressed. LDH release increased in a linear, dose-dependent fashion over the range of silica doses used. Moreover, within 15 min following addition of a high silica dose, a shift to the production of 5-lipoxygenase metabolites occurred, accompanied by a reduction in cyclooxygenase products. This rapid alteration in AA metabolism preceded cell injury. To examine the relationship between cytotoxicity and AA metabolite release by BAM exposed to silicas with different cytotoxic and fibrogenic activities, BAM were exposed to different doses of DQ-12, Minusil-5, and Sigma silicas, and carbonyl iron beads. The median effective dose (ED 50 ) of each particulate to stimulate the release of AA metabolites and LDH was calculated. The ED 50 values for DQ-12, Minusil-5, and Sigma silica showed that the relative cytotoxicities of the different silicas for BAM corresponded to the relative potencies of the silicas to elicit 5-lipoxygenase metabolites from BAM. These results indicate that the cytotoxic, and presumed fibrogenic potential, of a silica is correlated with the potency to stimulate the release of leukotrienes from AM

  16. Evidence for particle transport between alveolar macrophages in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Benson, J.M.; Nikula, K.J.; Guilmette, R.A.

    1995-12-01

    Recent studies at this Institute have focused on determining the role of alveolar macrophages (AMs) in the transport of particles within and form the lung. For those studies, AMs previously labeled using the nuclear stain Hoechst 33342 and polychromatic Fluoresbrite microspheres (1 {mu}m diameter, Polysciences, Inc., Warrington, PA) were instilled into lungs of recipient F344 rats. The fate of the donor particles and the doubly labeled AMs within recipient lungs was followed for 32 d. Within 2-4 d after instillation, the polychromatic microspheres were found in both donor and resident AMs, suggesting that particle transfer occurred between the donor and resident AMs. However, this may also have been an artifact resulting from phagocytosis of the microspheres form dead donor cells or from the fading or degradation of Hoechst 33342 within the donor cells leading to their misidentification as resident AMs. The results support the earlier findings that microspheres in donor AMs can be transferred to resident AMs within 2 d after instillation.

  17. Sulfite induces release of lipid mediators by alveolar macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Beck-Speier, I.; Dayal, N.; Maier, L. [GSF - National Research Center for Environment and Health, Neuherberg (Germany). Inst. for Inhalation Biology; Denzlinger, C. [Tuebingen Univ. (Germany). Dept. II, Medical Clinic; Haberl, C. [Tuebingen Univ. (Germany). Dept. III, Medical Clinic

    1998-03-01

    Air pollutants are supposed to modulate physiological responses of alveolar macrophages (AM). This study was addressed to the question whether at neutral pH sulfur(IV) species in comparison to sulfur(VI) species cause AM to release proinflammatory mediators and which pathways are involved in their generation. Supernatants obtained from canine AM treated with sulfite (0.1 mM to 2 mM) enhanced the respiratory burst of canine neutrophils, measured by lucigenin-dependent chemiluminescence, whereas supernatants derived from AM treated with sulfate (1 mM) did not. The neutrophil-stimulating activity released by sulfite-treated AM consisted of platelet-activating factor (PAF) and leukotriene B{sub 4} (LTB{sub 4}) as shown by desensitization of the platelet-activating factor (PAF) and leukotriene B{sub 4} (LTB{sub 4}) as shown by desensitization of the corresponding receptors. Inhibitors of phospholipase A{sub 2} substantially suppressed release of neutrophil-stimulating activity by sulfite-treated AM. Inhibition of 5-lipoxygenase in sulfite-treated AM also reduced neutrophil-stimulating activity, while inhibition of cyclooxygenase had no effect. In conclusion, sulfite induces AM to release lipid mediators via phospholipase A{sub 2}- and 5-lipoxygenase-dependent pathways. These mediators activate neutrophils via the receptors for PAF and LTB{sub 4}. (orig.)

  18. Macrophage-expressed IFN-β contributes to apoptotic alveolar epithelial cell injury in severe influenza virus pneumonia.

    Directory of Open Access Journals (Sweden)

    Katrin Högner

    2013-02-01

    Full Text Available Influenza viruses (IV cause pneumonia in humans with progression to lung failure and fatal outcome. Dysregulated release of cytokines including type I interferons (IFNs has been attributed a crucial role in immune-mediated pulmonary injury during severe IV infection. Using ex vivo and in vivo IV infection models, we demonstrate that alveolar macrophage (AM-expressed IFN-β significantly contributes to IV-induced alveolar epithelial cell (AEC injury by autocrine induction of the pro-apoptotic factor TNF-related apoptosis-inducing ligand (TRAIL. Of note, TRAIL was highly upregulated in and released from AM of patients with pandemic H1N1 IV-induced acute lung injury. Elucidating the cell-specific underlying signalling pathways revealed that IV infection induced IFN-β release in AM in a protein kinase R- (PKR- and NF-κB-dependent way. Bone marrow chimeric mice lacking these signalling mediators in resident and lung-recruited AM and mice subjected to alveolar neutralization of IFN-β and TRAIL displayed reduced alveolar epithelial cell apoptosis and attenuated lung injury during severe IV pneumonia. Together, we demonstrate that macrophage-released type I IFNs, apart from their well-known anti-viral properties, contribute to IV-induced AEC damage and lung injury by autocrine induction of the pro-apoptotic factor TRAIL. Our data suggest that therapeutic targeting of the macrophage IFN-β-TRAIL axis might represent a promising strategy to attenuate IV-induced acute lung injury.

  19. PPARγ regulates the expression of cholesterol metabolism genes in alveolar macrophages

    International Nuclear Information System (INIS)

    Baker, Anna D.; Malur, Anagha; Barna, Barbara P.; Kavuru, Mani S.; Malur, Achut G.; Thomassen, Mary Jane

    2010-01-01

    Peroxisome proliferator-activated receptor-gamma (PPARγ) is a nuclear transcription factor involved in lipid metabolism that is constitutively expressed in the alveolar macrophages of healthy individuals. PPARγ has recently been implicated in the catabolism of surfactant by alveolar macrophages, specifically the cholesterol component of surfactant while the mechanism remains unclear. Studies from other tissue macrophages have shown that PPARγ regulates cholesterol influx, efflux, and metabolism. PPARγ promotes cholesterol efflux through the liver X receptor-alpha (LXRα) and ATP-binding cassette G1 (ABCG1). We have recently shown that macrophage-specific PPARγ knockout (PPARγ KO) mice accumulate cholesterol-laden alveolar macrophages that exhibit decreased expression of LXRα and ABCG1 and reduced cholesterol efflux. We hypothesized that in addition to the dysregulation of these cholesterol efflux genes, the expression of genes involved in cholesterol synthesis and influx was also dysregulated and that replacement of PPARγ would restore regulation of these genes. To investigate this hypothesis, we have utilized a Lentivirus expression system (Lenti-PPARγ) to restore PPARγ expression in the alveolar macrophages of PPARγ KO mice. Our results show that the alveolar macrophages of PPARγ KO mice have decreased expression of key cholesterol synthesis genes and increased expression of cholesterol receptors CD36 and scavenger receptor A-I (SRA-I). The replacement of PPARγ (1) induced transcription of LXRα and ABCG1; (2) corrected suppressed expression of cholesterol synthesis genes; and (3) enhanced the expression of scavenger receptors CD36. These results suggest that PPARγ regulates cholesterol metabolism in alveolar macrophages.

  20. PPAR{gamma} regulates the expression of cholesterol metabolism genes in alveolar macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Baker, Anna D.; Malur, Anagha; Barna, Barbara P.; Kavuru, Mani S. [Department of Internal Medicine, Division of Pulmonary, Critical Care, and Sleep Medicine, East Carolina University (United States); Malur, Achut G. [Department of Microbiology and Immunology, East Carolina University (United States); Thomassen, Mary Jane, E-mail: thomassenm@ecu.edu [Department of Internal Medicine, Division of Pulmonary, Critical Care, and Sleep Medicine, East Carolina University (United States); Department of Microbiology and Immunology, East Carolina University (United States)

    2010-03-19

    Peroxisome proliferator-activated receptor-gamma (PPAR{gamma}) is a nuclear transcription factor involved in lipid metabolism that is constitutively expressed in the alveolar macrophages of healthy individuals. PPAR{gamma} has recently been implicated in the catabolism of surfactant by alveolar macrophages, specifically the cholesterol component of surfactant while the mechanism remains unclear. Studies from other tissue macrophages have shown that PPAR{gamma} regulates cholesterol influx, efflux, and metabolism. PPAR{gamma} promotes cholesterol efflux through the liver X receptor-alpha (LXR{alpha}) and ATP-binding cassette G1 (ABCG1). We have recently shown that macrophage-specific PPAR{gamma} knockout (PPAR{gamma} KO) mice accumulate cholesterol-laden alveolar macrophages that exhibit decreased expression of LXR{alpha} and ABCG1 and reduced cholesterol efflux. We hypothesized that in addition to the dysregulation of these cholesterol efflux genes, the expression of genes involved in cholesterol synthesis and influx was also dysregulated and that replacement of PPAR{gamma} would restore regulation of these genes. To investigate this hypothesis, we have utilized a Lentivirus expression system (Lenti-PPAR{gamma}) to restore PPAR{gamma} expression in the alveolar macrophages of PPAR{gamma} KO mice. Our results show that the alveolar macrophages of PPAR{gamma} KO mice have decreased expression of key cholesterol synthesis genes and increased expression of cholesterol receptors CD36 and scavenger receptor A-I (SRA-I). The replacement of PPAR{gamma} (1) induced transcription of LXR{alpha} and ABCG1; (2) corrected suppressed expression of cholesterol synthesis genes; and (3) enhanced the expression of scavenger receptors CD36. These results suggest that PPAR{gamma} regulates cholesterol metabolism in alveolar macrophages.

  1. Comparative morphology and morphometry of alveolar macrophages from six mammalian species

    International Nuclear Information System (INIS)

    Haley, P.J.; Muggenburg, B.A.; Weissman, D.N.; Bice, D.E.

    1988-01-01

    Pulmonary alveolar macrophages (PAM) were collected from normal, healthy mice, rats, dogs, cynomolgus monkeys, chimpanzees and humans and evaluated for morphologic and morphometric characteristics. The PAM of mice, rats, and dogs were morphologically similar to one another and had statistically similar frequency distributions for PAM size. The range of cell size for these three species was narrow. The PAM of nonhuman primates and humans were morphologically heterogenous with increased cytoplasmic vacuolation, irregular cell outlines and increased numbers of multi nucleated cells as compared to the PAM of rodents and dogs. The mean size of human PAMs was statistically greater than that for all other species evaluated, including nonhuman primates. These data indicate that significant differences in PAM morphology and size exist among species and that such differences may be important when selecting species for studies of PAM. (author)

  2. Depletion of alveolar macrophages in CD11c diphtheria toxin receptor mice produces an inflammatory response

    Science.gov (United States)

    Roberts, Lydia M; Ledvina, Hannah E; Tuladhar, Shraddha; Rana, Deepa; Steele, Shaun P; Sempowski, Gregory D; Frelinger, Jeffrey A

    2015-01-01

    Alveolar macrophages play a critical role in initiating the immune response to inhaled pathogens and have been shown to be the first cell type infected following intranasal inoculation with several pathogens, including Francisella tularensis. In an attempt to further dissect the role of alveolar macrophages in the immune response to Francisella, we selectively depleted alveolar macrophages using CD11c.DOG mice. CD11c.DOG mice express the diphtheria toxin receptor (DTR) under control of the full CD11c promoter. Because mice do not express DTR, tissue restricted expression of the primate DTR followed by treatment with diphtheria toxin (DT) has been widely used as a tool in immunology to examine the effect of acute depletion of a specific immune subset following normal development. We successfully depleted alveolar macrophages via intranasal administration of DT. However, alveolar macrophage depletion was accompanied by many other changes to the cellular composition and cytokine/chemokine milieu in the lung that potentially impact innate and adaptive immune responses. Importantly, we observed a transient influx of neutrophils in the lung and spleen. Our experience serves as a cautionary note to other researchers using DTR mice given the complex changes that occur following DT treatment that must be taken into account when analyzing data. PMID:26029367

  3. Trichinella spiralis infection enhances protein kinase C phosphorylation in guinea pig alveolar macrophages.

    Science.gov (United States)

    Dzik, J M; Zieliński, Z; Cieśla, J; Wałajtys-Rode, E

    2010-03-01

    To learn more about the signalling pathways involved in superoxide anion production in guinea pig alveolar macrophages, triggered by Trichinella spiralis infection, protein level and phosphorylation of mitogen activated protein (MAP) kinases and protein kinase C (PKC) were investigated. Infection with T. spiralis, the nematode having 'lung phase' during colonization of the host, enhances PKC phosphorylation in guinea pig alveolar macrophages. Isoenzymes beta and delta of PKC have been found significantly phosphorylated, although their location was not changed as a consequence of T. spiralis infection. Neither in macrophages from T. spiralis-infected guinea pig nor in platelet-activating factor (PAF)-stimulated macrophages from uninfected animals, participation of MAP kinases in respiratory burst activation was statistically significant. The parasite antigens seem to act through macrophage PAF receptors, transducing a signal for enhanced NADPH oxidase activity, as stimulating effect of newborn larvae homogenate on respiratory burst was abolished by specific PAF receptor antagonist CV 6209. A suppressive action of T. spiralis larvae on host alveolar macrophage innate immunological response was reflected by diminished protein level of ERK2 kinase and suppressed superoxide anion production, in spite of high level of PKC phosphorylation.

  4. STIMULATION OF OXIDANT PRODUCTION IN ALVEOLAR MACROPHAGES BY POLLUTANT AND LATEX PARTICLES

    Science.gov (United States)

    Air pollutant dusts as well as chemically defined particles were examined for their activating effect on oxidant production (O2- and H2O2) in guinea pig alveolar macrophages (AM). Oxidant production was measured as chemiluminescence of albumin-bound luminol. All particles examine...

  5. 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.

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

    International Nuclear Information System (INIS)

    Barbers, R.G.; Evans, M.J.; Gong, H. Jr.; Tashkin, D.P.

    1991-01-01

    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 [ 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 [ 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 [ 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

  7. Resident alveolar macrophages are susceptible to and permissive of Coxiella burnetii infection.

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    Matthew Calverley

    Full Text Available Coxiella burnetii, the causative agent of Q fever, is a zoonotic disease with potentially life-threatening complications in humans. Inhalation of low doses of Coxiella bacteria can result in infection of the host alveolar macrophage (AM. However, it is not known whether a subset of AMs within the heterogeneous population of macrophages in the infected lung is particularly susceptible to infection. We have found that lower doses of both phase I and phase II Nine Mile C. burnetii multiply and are less readily cleared from the lungs of mice compared to higher infectious doses. We have additionally identified AM resident within the lung prior to and shortly following infection, opposed to newly recruited monocytes entering the lung during infection, as being most susceptible to infection. These resident cells remain infected up to twelve days after the onset of infection, serving as a permissive niche for the maintenance of bacterial infection. A subset of infected resident AMs undergo a distinguishing phenotypic change during the progression of infection exhibiting an increase in surface integrin CD11b expression and continued expression of the surface integrin CD11c. The low rate of phase I and II Nine Mile C. burnetii growth in murine lungs may be a direct result of the limited size of the susceptible resident AM cell population.

  8. Importance of Bacterial Replication and Alveolar Macrophage-Independent Clearance Mechanisms during Early Lung Infection with Streptococcus pneumoniae

    Science.gov (United States)

    Camberlein, Emilie; Cohen, Jonathan M.; José, Ricardo; Hyams, Catherine J.; Callard, Robin; Chimalapati, Suneeta; Yuste, Jose; Edwards, Lindsey A.; Marshall, Helina; van Rooijen, Nico; Noursadeghi, Mahdad

    2015-01-01

    Although the importance of alveolar macrophages for host immunity during early Streptococcus pneumoniae lung infection is well established, the contribution and relative importance of other innate immunity mechanisms and of bacterial factors are less clear. We have used a murine model of S. pneumoniae early lung infection with wild-type, unencapsulated, and para-amino benzoic acid auxotroph mutant TIGR4 strains to assess the effects of inoculum size, bacterial replication, capsule, and alveolar macrophage-dependent and -independent clearance mechanisms on bacterial persistence within the lungs. Alveolar macrophage-dependent and -independent (calculated indirectly) clearance half-lives and bacterial replication doubling times were estimated using a mathematical model. In this model, after infection with a high-dose inoculum of encapsulated S. pneumoniae, alveolar macrophage-independent clearance mechanisms were dominant, with a clearance half-life of 24 min compared to 135 min for alveolar macrophage-dependent clearance. In addition, after a high-dose inoculum, successful lung infection required rapid bacterial replication, with an estimated S. pneumoniae doubling time of 16 min. The capsule had wide effects on early lung clearance mechanisms, with reduced half-lives of 14 min for alveolar macrophage-independent and 31 min for alveolar macrophage-dependent clearance of unencapsulated bacteria. In contrast, with a lower-dose inoculum, the bacterial doubling time increased to 56 min and the S. pneumoniae alveolar macrophage-dependent clearance half-life improved to 42 min and was largely unaffected by the capsule. These data demonstrate the large effects of bacterial factors (inoculum size, the capsule, and rapid replication) and alveolar macrophage-independent clearance mechanisms during early lung infection with S. pneumoniae. PMID:25583525

  9. Mycobacterium tuberculosis Infection and Innate Responses in a New Model of Lung Alveolar Macrophages.

    Science.gov (United States)

    Woo, Minjeong; Wood, Connor; Kwon, Doyoon; Park, Kyu-Ho Paul; Fejer, György; Delorme, Vincent

    2018-01-01

    Lung alveolar macrophages (AMs) are in the first line of immune defense against respiratory pathogens and play key roles in the pathogenesis of Mycobacterium tuberculosis ( Mtb ) in humans. Nevertheless, AMs are available only in limited amounts for in vitro studies, which hamper the detailed molecular understanding of host- Mtb interactions in these macrophages. The recent establishment of the self-renewing and primary Max Planck Institute (MPI) cells, functionally very close to lung AMs, opens unique opportunities for in vitro studies of host-pathogen interactions in respiratory diseases. Here, we investigated the suitability of MPI cells as a host cell system for Mtb infection. Bacterial, cellular, and innate immune features of MPI cells infected with Mtb were characterized. Live bacteria were readily internalized and efficiently replicated in MPI cells, similarly to primary murine macrophages and other cell lines. MPI cells were also suitable for the determination of anti-tuberculosis (TB) drug activity. The primary innate immune response of MPI cells to live Mtb showed significantly higher and earlier induction of the pro-inflammatory cytokines TNFα, interleukin 6 (IL-6), IL-1α, and IL-1β, as compared to stimulation with heat-killed (HK) bacteria. MPI cells previously showed a lack of induction of the anti-inflammatory cytokine IL-10 to a wide range of stimuli, including HK Mtb . By contrast, we show here that live Mtb is able to induce significant amounts of IL-10 in MPI cells. Autophagy experiments using light chain 3B immunostaining, as well as LysoTracker labeling of acidic vacuoles, demonstrated that MPI cells efficiently control killed Mtb by elimination through phagolysosomes. MPI cells were also able to accumulate lipid droplets in their cytoplasm following exposure to lipoproteins. Collectively, this study establishes the MPI cells as a relevant, versatile host cell model for TB research, allowing a deeper understanding of AMs functions in this

  10. Mycobacterium tuberculosis Infection and Innate Responses in a New Model of Lung Alveolar Macrophages

    Directory of Open Access Journals (Sweden)

    Minjeong Woo

    2018-03-01

    Full Text Available Lung alveolar macrophages (AMs are in the first line of immune defense against respiratory pathogens and play key roles in the pathogenesis of Mycobacterium tuberculosis (Mtb in humans. Nevertheless, AMs are available only in limited amounts for in vitro studies, which hamper the detailed molecular understanding of host-Mtb interactions in these macrophages. The recent establishment of the self-renewing and primary Max Planck Institute (MPI cells, functionally very close to lung AMs, opens unique opportunities for in vitro studies of host-pathogen interactions in respiratory diseases. Here, we investigated the suitability of MPI cells as a host cell system for Mtb infection. Bacterial, cellular, and innate immune features of MPI cells infected with Mtb were characterized. Live bacteria were readily internalized and efficiently replicated in MPI cells, similarly to primary murine macrophages and other cell lines. MPI cells were also suitable for the determination of anti-tuberculosis (TB drug activity. The primary innate immune response of MPI cells to live Mtb showed significantly higher and earlier induction of the pro-inflammatory cytokines TNFα, interleukin 6 (IL-6, IL-1α, and IL-1β, as compared to stimulation with heat-killed (HK bacteria. MPI cells previously showed a lack of induction of the anti-inflammatory cytokine IL-10 to a wide range of stimuli, including HK Mtb. By contrast, we show here that live Mtb is able to induce significant amounts of IL-10 in MPI cells. Autophagy experiments using light chain 3B immunostaining, as well as LysoTracker labeling of acidic vacuoles, demonstrated that MPI cells efficiently control killed Mtb by elimination through phagolysosomes. MPI cells were also able to accumulate lipid droplets in their cytoplasm following exposure to lipoproteins. Collectively, this study establishes the MPI cells as a relevant, versatile host cell model for TB research, allowing a deeper understanding of AMs functions

  11. Carbon black nanoparticles induce type II epithelial cells to release chemotaxins for alveolar macrophages

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    Donaldson Ken

    2005-12-01

    Full Text Available Abstract Background Alveolar macrophages are a key cell in dealing with particles deposited in the lungs and in determining the subsequent response to that particle exposure. Nanoparticles are considered a potential threat to the lungs and the mechanism of pulmonary response to nanoparticles is currently under intense scrutiny. The type II alveolar epithelial cell has previously been shown to release chemoattractants which can recruit alveolar macrophages to sites of particle deposition. The aim of this study was to assess the responses of a type II epithelial cell line (L-2 to both fine and nanoparticle exposure in terms of secretion of chemotactic substances capable of inducing macrophage migration. Results Exposure of type II cells to carbon black nanoparticles resulted in significant release of macrophage chemoattractant compared to the negative control and to other dusts tested (fine carbon black and TiO2 and nanoparticle TiO2 as measured by macrophage migration towards type II cell conditioned medium. SDS-PAGE analysis of the conditioned medium from particle treated type II cells revealed that a higher number of protein bands were present in the conditioned medium obtained from type II cells treated with nanoparticle carbon black compared to other dusts tested. Size-fractionation of the chemotaxin-rich supernatant determined that the chemoattractants released from the epithelial cells were between 5 and 30 kDa in size. Conclusion The highly toxic nature and reactive surface chemistry of the carbon black nanoparticles has very likely induced the type II cell line to release pro-inflammatory mediators that can potentially induce migration of macrophages. This could aid in the rapid recruitment of inflammatory cells to sites of particle deposition and the subsequent removal of the particles by phagocytic cells such as macrophages and neutrophils. Future studies in this area could focus on the exact identity of the substance(s released by the

  12. Wool and grain dusts stimulate TNF secretion by alveolar macrophages in vitro.

    Science.gov (United States)

    Brown, D M; Donaldson, K

    1996-06-01

    The aim of the study was to investigate the ability of two organic dusts, wool and grain, and their soluble leachates to stimulate secretion of tumour necrosis factor (TNF) by rat alveolar macrophages with special reference to the role of lipopolysaccharide (LPS). Rat alveolar macrophages were isolated by bronchoalveolar lavage (BAL) and treated in vitro with whole dust, dust leachates, and a standard LPS preparation. TNF production was measured in supernatants with the L929 cell line bioassay. Both wool and grain dust samples were capable of stimulating TNF release from rat alveolar macrophages in a dose-dependent manner. The standard LPS preparation caused a dose-dependent secretion of TNF. Leachates prepared from the dusts contained LPS and also caused TNF release but leachable LPS could not account for the TNF release and it was clear that non-LPS leachable activity was present in the grain dust and that wool dust particles themselves were capable of causing release of TNF. The role of LPS in wool dust leachates was further investigated by treating peritoneal macrophages from two strains of mice, LPS responders (C3H) and LPS non-responders (C3H/HEJ), with LPS. The non-responder mouse macrophages produced very low concentrations of TNF in response to the wool dust leachates compared with the responders. LPS and other unidentified leachable substances present on the surface of grain dust, and to a lesser extent on wool dust, are a trigger for TNF release by lung macrophages. Wool dust particles themselves stimulate TNF. TNF release from macrophages could contribute to enhancement of inflammatory responses and symptoms of bronchitis and breathlessness in workers exposed to organic dusts such as wool and grain.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2008-07-01

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

  14. Evaluation of the alveolar macrophage role in the pulmonary distribution of actinide oxides

    International Nuclear Information System (INIS)

    Guezingar-Liebard, Florence

    1999-01-01

    Actinide oxide inhalation is potentially a risk during the fuel fabrication process in the electronuclear industry. These particles can induce pulmonary lesions. The alveolar macrophage play an important role in the particle sequestration and transport but the actinide toxicity towards these cells is not well known. The aim of this work was to characterize the evolution of particle localisation in lungs after inhalation and to evaluate the role of macrophages in the lesion histo-genesis. We have used of a solid track detector to visualise alpha dose distribution within lung tissue. After 237 NpO 2 , MOX or PuO 2 inhalation by rats, different kinetics of clearance were observed for the sub-pleural and peri-bronchial areas compared to the others alveolar areas. For initial lung burdens that alter the lung clearance, particle aggregates were observed. Their kinetic and localisation vary depending on the aerosol, for a same global dose delivered to the lungs. This could be due to the different specific alpha activities of the particles and to the particle number deposited in the lung to obtain a similar burden but it could be also due to a chemical toxicity of neptunium higher than that of the others actinides. The flow cytometry methods developed allow us to measure apoptosis, phagocytosis and free radicals generation. After addition of soluble uranium to the culture medium, similar results were obtained using either alveolar macrophages extracted from rats or a macrophage cell line. This work confirms that alveolar macrophages are involved in the aggregate formation which induces heterogeneous dose distribution within the different lung tissues. (author) [fr

  15. Mechanisms underlying the redistribution of particles among the lung's alveolar macrophages during alveolar phase clearance

    Energy Technology Data Exchange (ETDEWEB)

    Lehnert, B.E.; Oritz, J.B.; Steinkamp, J.A.; Tietjen, G.L.; Sebring, R.J. (Los Alamos National Lab., NM (United States)); Oberdorster, G. (Rochester Univ., NY (United States))

    1991-01-01

    In order to obtain information about the particle redistribution phenomenon following the deposition of inhaled particles, as well as to obtain information about some of the mechanisms that may be operable in the redistribution of particles, lavaged lung free cell analyses and transmission electron microscopic (TEM) analyses of lung tissue and were performed using lungs from rats after they were subchronically exposed to aerosolized dioxide (TiO{sub 2}). TEM analyses indicated that the in situ autolysis of particle-containing Alveolar Macropages (AM) is one important mechanism involved in the redistribution of particles. Evidence was also obtained that indicated that the engulfment of one particle-containing phagocyte by another phagocyte also occurs. Another prominent mechanism of the particle redistribution phenomenon may be the in situ proliferation of particle-laden AM. We used the macrophage cell line J774A.1 as a surrogate for AM to investigate how different particulate loads in macrophages may affect their abilities to proliferate. These in vitro investigations indicated that the normal rate of proliferation of macrophages is essentially unaffected by the containment of relatively high particulate burdens. Overall, the results of our investigations suggest that in situ autolysis of particle-containing AM and the rephagocytosis of freed particles by other phagocytes, the phagocytosis of effete and disintegrating particle-containing phagocytes by other AM, and the in situ division of particle-containing AM are likely mechanisms that underlie the post-depositional redistribution of particles among the lung's AM during alveolar phase clearance. 19 refs., 8 figs., 2 tabs.

  16. The FGL2/fibroleukin prothrombinase is involved in alveolar macrophage activation in COPD through the MAPK pathway

    International Nuclear Information System (INIS)

    Liu, Yanling; Xu, Sanpeng; Xiao, Fei; Xiong, Yan; Wang, Xiaojin; Gao, Sui; Yan, Weiming; Ning, Qin

    2010-01-01

    Fibrinogen-like protein 2 (FGL2)/fibroleukin has been reported to play a vital role in the pathogenesis of some critical inflammatory diseases by possessing immunomodulatory activity through the mediation of 'immune coagulation' and the regulation of maturation and proliferation of immune cells. We observed upregulated FGL2 expression in alveolar macrophages from peripheral lungs of chronic obstructive pulmonary disease (COPD) patients and found a correlation between FGL2 expression and increased macrophage activation markers (CD11b and CD14). The role of FGL2 in the activation of macrophages was confirmed by the detection of significantly decreased macrophage activation marker (CD11b, CD11c, and CD71) expression as well as the inhibition of cell migration and inflammatory cytokine (IL-8 and MMP-9) production in an LPS-induced FGL2 knockdown human monocytic leukemia cell line (THP-1). Increased FGL2 expression co-localized with upregulated phosphorylated p38 mitogen-activated protein kinase (p38-MAPK) in the lung tissues from COPD patients. Moreover, FGL2 knockdown in THP-1 cells significantly downregulated LPS-induced phosphorylation of p38-MAPK while upregulating phosphorylation of c-Jun N-terminal kinase (JNK). Thus, we demonstrate that FGL2 plays an important role in macrophage activation in the lungs of COPD patients through MAPK pathway modulation.

  17. Recombinant guinea pig CCL5 (RANTES) differentially modulates cytokine production in alveolar and peritoneal macrophages.

    Science.gov (United States)

    Skwor, Troy A; Cho, Hyosun; Cassidy, Craig; Yoshimura, Teizo; McMurray, David N

    2004-12-01

    The CC chemokine ligand 5 (CCL5; regulated on activation, normal T expressed and secreted) is known to recruit and activate leukocytes; however, its role in altering the responses of host cells to a subsequent encounter with a microbial pathogen has rarely been studied. Recombinant guinea pig (rgp)CCL5 was prepared, and its influence on peritoneal and alveolar macrophage activation was examined by measuring cytokine and chemokine mRNA expression in cells stimulated with rgpCCL5 alone or exposed to rgpCCL5 prior to lipopolysaccharide (LPS) stimulation. Levels of mRNA for guinea pig tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-1beta, CCL2 (monocyte chemoattractant protein-1), and CXC chemokine ligand 8 (IL-8) were analyzed by reverse transcription followed by real-time polymerase chain reaction analysis using SYBR Green. Bioactive TNF-alpha protein concentration was measured using the L929 bioassay. Both macrophage populations displayed significant enhancement of all the genes and TNF-alpha protein levels when stimulated with rgpCCL5, except for CCL2 in alveolar macrophages. When peritoneal or alveolar macrophages were pretreated with rgpCCL5 for 2 h and then exposed to low concentrations of LPS, diminished cytokine and chemokine mRNA levels were apparent at 6 h compared with LPS alone. At the protein level, there was a reduction in TNF-alpha protein at 6 h in the CCL5-pretreated cells compared with LPS alone. These results further support a role for CCL5 in macrophage activation in addition to chemotactic properties and suggest a role in regulating the inflammatory response to LPS in the guinea pig by modulating the production of proinflammatory cytokines by macrophages.

  18. Effects of X irradiation on the cytoskeleton of rat alveolar macrophages in vitro

    International Nuclear Information System (INIS)

    Ladyman, S.J.; Townsend, K.M.S.; Edwards, C.

    1984-01-01

    The three-dimensional visualization of Triton X-100 resistant cytoskeletons has been used to demonstrate that an absorbed dose of 120 Gy from X rays causes a distinctive and reproducible alteration of the cytoskeleton of intact rat alveolar macrophages in vitro. The alteration has also been shown to be rapidly and completely ''repaired'' and to be apparently similar to alterations caused by colchicine but dissimilar to those caused by cytochalasin B. From these observations and those of other workers who have studied the irradiation of extracted microtubular proteins in vitro, the authors think it likely that microtubules rather than microfilaments are the radiosensitive component of the macrophage cytoskeleton

  19. Transcription analysis of the porcine alveolar macrophage response to Mycoplasma hyopneumoniae.

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    Li Bin

    Full Text Available Mycoplasma hyopneumoniae is considered the major causative agent of porcine respiratory disease complex, occurs worldwide and causes major economic losses to the pig industry. To gain more insights into the pathogenesis of this organism, the high throughput cDNA microarray assays were employed to evaluate host responses of porcine alveolar macrophages to M. hyopneumoniae infection. A total of 1033 and 1235 differentially expressed genes were identified in porcine alveolar macrophages in responses to exposure to M. hyopneumoniae at 6 and 15 hours post infection, respectively. The differentially expressed genes were involved in many vital functional classes, including inflammatory response, immune response, apoptosis, cell adhesion, defense response, signal transduction, protein folding, protein ubiquitination and so on. The pathway analysis demonstrated that the most significant pathways were the chemokine signaling pathway, Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, nucleotide-binding oligomerization domains (Nod-like receptor signaling pathway and apoptosis signaling pathway. The reliability of the data obtained from the microarray was verified by performing quantitative real-time PCR. The expression kinetics of chemokines was further analyzed. The present study is the first to document the response of porcine alveolar macrophages to M. hyopneumoniae infection. The data further developed our understanding of the molecular pathogenesis of M. hyopneumoniae.

  20. Transcription analysis of the porcine alveolar macrophage response to Mycoplasma hyopneumoniae.

    Science.gov (United States)

    Bin, Li; Luping, Du; Bing, Sun; Zhengyu, Yu; Maojun, Liu; Zhixin, Feng; Yanna, Wei; Haiyan, Wang; Guoqing, Shao; Kongwang, He

    2014-01-01

    Mycoplasma hyopneumoniae is considered the major causative agent of porcine respiratory disease complex, occurs worldwide and causes major economic losses to the pig industry. To gain more insights into the pathogenesis of this organism, the high throughput cDNA microarray assays were employed to evaluate host responses of porcine alveolar macrophages to M. hyopneumoniae infection. A total of 1033 and 1235 differentially expressed genes were identified in porcine alveolar macrophages in responses to exposure to M. hyopneumoniae at 6 and 15 hours post infection, respectively. The differentially expressed genes were involved in many vital functional classes, including inflammatory response, immune response, apoptosis, cell adhesion, defense response, signal transduction, protein folding, protein ubiquitination and so on. The pathway analysis demonstrated that the most significant pathways were the chemokine signaling pathway, Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, nucleotide-binding oligomerization domains (Nod)-like receptor signaling pathway and apoptosis signaling pathway. The reliability of the data obtained from the microarray was verified by performing quantitative real-time PCR. The expression kinetics of chemokines was further analyzed. The present study is the first to document the response of porcine alveolar macrophages to M. hyopneumoniae infection. The data further developed our understanding of the molecular pathogenesis of M. hyopneumoniae.

  1. Reactomes of porcine alveolar macrophages infected with porcine reproductive and respiratory syndrome virus.

    Directory of Open Access Journals (Sweden)

    Zhihua Jiang

    Full Text Available Porcine reproductive and respiratory syndrome (PRRS has devastated pig industries worldwide for many years. It is caused by a small RNA virus (PRRSV, which targets almost exclusively pig monocytes or macrophages. In the present study, five SAGE (serial analysis of gene expression libraries derived from 0 hour mock-infected and 6, 12, 16 and 24 hours PRRSV-infected porcine alveolar macrophages (PAMs produced a total 643,255 sequenced tags with 91,807 unique tags. Differentially expressed (DE tags were then detected using the Bayesian framework followed by gene/mRNA assignment, arbitrary selection and manual annotation, which determined 699 DE genes for reactome analysis. The DAVID, KEGG and REACTOME databases assigned 573 of the DE genes into six biological systems, 60 functional categories and 504 pathways. The six systems are: cellular processes, genetic information processing, environmental information processing, metabolism, organismal systems and human diseases as defined by KEGG with modification. Self-organizing map (SOM analysis further grouped these 699 DE genes into ten clusters, reflecting their expression trends along these five time points. Based on the number one functional category in each system, cell growth and death, transcription processes, signal transductions, energy metabolism, immune system and infectious diseases formed the major reactomes of PAMs responding to PRRSV infection. Our investigation also focused on dominant pathways that had at least 20 DE genes identified, multi-pathway genes that were involved in 10 or more pathways and exclusively-expressed genes that were included in one system. Overall, our present study reported a large set of DE genes, compiled a comprehensive coverage of pathways, and revealed system-based reactomes of PAMs infected with PRRSV. We believe that our reactome data provides new insight into molecular mechanisms involved in host genetic complexity of antiviral activities against PRRSV and

  2. YC-1 potentiates cAMP-induced CREB activation and nitric oxide production in alveolar macrophages

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    Hwang, Tsong-Long, E-mail: htl@mail.cgu.edu.tw [Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan, Taiwan (China); Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Kweishan, Taoyuan, Taiwan (China); Tang, Ming-Chi [Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan, Taiwan (China); Kuo, Liang-Mou [Department of General Surgery, Chang Gung Memorial Hospital at Chia-Yi, Taiwan (China); Chang, Wen-De; Chung, Pei-Jen; Chang, Ya-Wen; Fang, Yao-Ching [Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan, Taiwan (China)

    2012-04-15

    Alveolar macrophages play significant roles in the pathogenesis of several inflammatory lung diseases. Increases in exhaled nitric oxide (NO) are well documented to reflect disease severity in the airway. In this study, we investigated the effect of 3-(5′-hydroxymethyl-2′-furyl)-1-benzyl indazole (YC-1), a known activator of soluble guanylyl cyclase, on prostaglandin (PG)E{sub 1} (a stable PGE{sub 2} analogue) and forskolin (a adenylate cyclase activator) induced NO production and inducible NO synthase (iNOS) expression in rat alveolar macrophages (NR8383). YC-1 did not directly cause NO production or iNOS expression, but drastically potentiated PGE{sub 1}- or forskolin-induced NO production and iNOS expression in NR8383 alveolar macrophages. Combination treatment with YC-1 and PGE{sub 1} significantly increased phosphorylation of the cAMP response element-binding protein (CREB), but not nuclear factor (NF)-κB activation. The combined effect on NO production, iNOS expression, and CREB phosphorylation was reversed by a protein kinase (PK)A inhibitor (H89), suggesting that the potentiating functions were mediated through a cAMP/PKA signaling pathway. Consistent with this, cAMP analogues, but not the cGMP analogue, caused NO release, iNOS expression, and CREB activation. YC-1 treatment induced an increase in PGE{sub 1}-induced cAMP formation, which occurred through the inhibition of cAMP-specific phosphodiesterase (PDE) activity. Furthermore, the combination of rolipram (an inhibitor of PDE4), but not milronone (an inhibitor of PDE3), and PGE{sub 1} also triggered NO production and iNOS expression. In summary, YC-1 potentiates PGE{sub 1}-induced NO production and iNOS expression in alveolar macrophages through inhibition of cAMP PDE activity and activation of the cAMP/PKA/CREB signaling pathway. Highlights: ► YC-1 potentiated PGE1-induced iNOS expression in alveolar macrophages. ► The combination of YC-1 and PGE1 increased CREB but not NFκB activation.

  3. Alveolar macrophage release of tumor necrosis factor-alpha in chronic alcoholics without liver disease.

    Science.gov (United States)

    Omidvari, K; Casey, R; Nelson, S; Olariu, R; Shellito, J E

    1998-05-01

    Alcohol is an immunosuppressive drug, and chronic abuse has been associated with increased susceptibility to a variety of infections, including bacterial pneumonia and tuberculosis. Alveolar macrophages are the resident phagocytes of the lung and play a central role in lung host defenses against infection ranging from direct antibacterial activity to the release of proinflammatory cytokines such as tumor necrosis factor-alpha (TNFalpha). TNFalpha, in particular, plays a key role in the development of the early inflammatory response. In this study, we investigated the effects of chronic alcohol consumption on alveolar macrophage release of TNFalpha in vitro. We prospectively studied lipopolysaccharide (LPS)-stimulated release of TNFalpha from alveolar macrophages obtained from bronchoalveolar lavage fluid (BALF) in 22 alcoholic (18 smokers, 4 nonsmokers) and 7 nondrinking healthy volunteers (3 smokers, 4 nonsmokers). The total number of cells recovered by bronchoalveolar lavage (BAL) and their differential distribution were not significantly different in alcoholics versus controls (43 +/- 8 x 10(6) and 39 +/- 13 x 10(6), respectively). However, the total number of cells recovered from BALF was significantly higher in smokers (51 +/- 8 x 10(6)) than in nonsmokers (19 +/- 5 x 10(6)). Spontaneous (basal) release of TNFalpha by alveolar macrophages was the same in alcoholics and controls. In contrast, LPS-stimulated release of TNFalpha was significantly suppressed in alcoholics compared with that of controls (1343 +/- 271 vs. 3806 +/- 926 U TNF/ml/10(6) cells, respectively, p < 0.015). When controlled for smoking, LPS-stimulated TNFalpha production was suppressed in alcoholic nonsmokers (563 +/- 413 U TNF/ml/10(6)) compared with control nonsmokers (5113 +/- 1264 U TNF/ml/10(6)). LPS-stimulated TNFalpha production was also less in control smokers (2063 +/- 386 U TNF/ml/10(6) cells) than in control nonsmokers (5113 +/- 1264 U TNF/ml/10(6) cells). There was no difference

  4. Characterization of immortalized MARCO and SR-AI/II-deficient murine alveolar macrophage cell lines

    Directory of Open Access Journals (Sweden)

    Imrich Amy

    2008-05-01

    Full Text Available Abstract Background Alveolar macrophages (AM avidly bind and ingest unopsonized inhaled particles and bacteria through class A scavenger receptors (SRAs MARCO and SR-AI/II. Studies to characterize the function of these SRAs have used AMs from MARCO or SR-AI/II null mice, but this approach is limited by the relatively low yield of AMs. Moreover, studies using both MARCO and SR-AI/II-deficient (MS-/- mice have not been reported yet. Hence, we sought to develop continuous cell lines from primary alveolar macrophages from MS-/- mice. Results We used in vitro infection of the primary AMs with the J2 retrovirus carrying the v-raf and v-myc oncogenes. Following initial isolation in media supplemented with murine macrophage colony-stimulating factor (M-CSF, we subcloned three AM cell lines, designated ZK-1, ZK-2 and ZK-6. These cell lines grow well in RPMI-1640-10% FBS in the absence of M-CSF. These adherent but trypsin-sensitive cell lines have a doubling time of approximately 14 hours, exhibit typical macrophage morphology, and express macrophage-associated cell surface Mac-1 (CD11b and F4/80 antigens. The cell lines show robust Fc-receptor dependent phagocytosis of opsonized red blood cells. Similar to freshly isolated AMs from MS-/- mice, the cell lines exhibit decreased phagocytosis of unopsonized titanium dioxide (TiO2, fluorescent latex beads and bacteria (Staphylococcus aureus compared with the primary AMs from wild type (WT C57BL/6 mice. Conclusion Our results indicated that three contiguous murine alveolar macrophage cell lines with MS-/- (ZK1, ZK2 and ZK6 were established successfully. These cell lines demonstrated macrophage morphology and functional activity. Interestingly, similar to freshly isolated AMs from MS-/- mice, the cell lines have a reduced, but not absent, ability to bind and ingest particles, with an altered pattern of blockade by scavenger receptor inhibitors. These cell lines will facilitate in vitro studies to further define

  5. Alveolar macrophage accumulation rates, for 28 nm and 250 nm PSL, are mediated by separate mechanisms

    Energy Technology Data Exchange (ETDEWEB)

    Moss, O R; Wong, V A, E-mail: moss@thehamner.or [Hamner Institutes for Health Sciences, Research Triangle Park, NC 27509-2137 (United States)

    2009-02-01

    When macrophages accumulate 28 nm and 250 nm diameter polystyrene latex (PSL) beads, the accumulation rates should reflect differences in molecular and cellular function. We used a confocal microscope to measure the accumulation rates of nanoparticles by F344-rat-alveolar macrophages (approx25,000 cells adhered to a 0.7 cm{sup 2} surface). Over the cells were layered 0.1 ml of media, and 0.1 ml of media-with-beads. Fresh cells were introduced for each exposure scenario. The maximum possible individual macrophage exposures were as follows: 8x10{sup 6}, 8x10{sup 5}, and 8x10{sup 4} 28 nm beads per macrophage; and 8x10{sup 4} and 1.12x10{sup 4} 250 nm beads per macrophage. Accumulation rates were estimated over 23 minutes. The increase in bead accumulation-rate matched changes in bead-availability: 7x increase for 250 nm beads; 100x increase for 28 nm beads; and 700x increase for all bead availabilities. The maximum sustained 28 nm bead accumulation rate was > 30,000 /min (for 5 min). Increases in bead accumulation could be explained by two mechanisms: bead-diffusion; and, for the macrophage, macropinocytosis. Also for the highest concentrations of 28 nm beads, we saw a colligative threshold - possibly due to beads masking the cell surface or obstructing cellular mechanisms.

  6. Human macrophage hemoglobin-iron metabolism in vitro

    International Nuclear Information System (INIS)

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

    1982-01-01

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

  7. Toxicity of penicillic acid for rat alveolar macrophages in vitro

    International Nuclear Information System (INIS)

    Sorenson, W.G.; Simpson, J.

    1985-01-01

    Penicillic acid (PA) is a polyketide mycotoxin produced by several species of Aspergillus and Penicillium. This mycotoxin is toxic in experimental animals and has also been reported to be carcinogenic. The cytotoxicity of penicillic acid was studied in rat albeolar macrophages (AM) in vitro. The effects of penicillic acid on membrane integrity were studied by measuring cell volume changes and 51 Cr release. There was a significant decrease in adenosine triphosphate (ATP) in cell cultures exposed to 1.0 mM penicillic acid for 4 hr. Inhibition of the incorporation of [ 3 H]leucine into protein was both dose- and time-dependent and protein synthesis was inhibited significantly after 2 hr exposure to ≥0.1 mM penicillic acid. RNA synthesis was inhibited to a lesser extent than protein synthesis. There was significant inhibition of phagocytosis after 2 hr exposure at ≥0.3 mM penicillic acid and the ED 50 for phagocytosis was 0.09 mM. Thus phagocytosis was more sensitive to the toxic effects of penicillic acid than any other cellular process studied. The data suggest the possibility of a respiratory hazard to agricultural workers exposed to contaminated grain

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2009-01-15

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

  9. Alveolar macrophage phagocytosis is enhanced after blunt chest trauma and alters the posttraumatic mediator release.

    Science.gov (United States)

    Seitz, Daniel H; Palmer, Annette; Niesler, Ulrike; Fröba, Janine S; Heidemann, Vera; Rittlinger, Anne; Braumüller, Sonja T; Zhou, Shaoxia; Gebhard, Florian; Knöferl, Markus W

    2011-12-01

    Blunt chest trauma is known to induce a pulmonary invasion of short-lived polymorphonuclear neutrophils and apoptosis of alveolar epithelial type 2 (AT2) cells. Apoptotic cells are removed by alveolar macrophages (AMΦ). We hypothesized that chest trauma alters the phagocytic response of AMΦ as well as the mediator release of AMΦ during phagocytosis. To study this, male Sprague-Dawley rats were subjected to blunt chest trauma. Phagocytosis assays were performed in AMΦ isolated 2 or 24 h after trauma with apoptotic cells or opsonized beads. Phagocytosis of apoptotic AT2 cells by unstimulated AMΦ was significantly increased 2 h after trauma. At 24 h, AMΦ from traumatized animals, stimulated with phorbol-12-myristate-13-acetate, ingested significantly more apoptotic polymorphonuclear neutrophils than AMΦ from sham animals. Alveolar macrophages after trauma released significantly higher levels of tumor necrosis factor α, macrophage inflammatory protein 1α, and cytokine-induced neutrophil chemoattractant 1 when they incorporated latex beads, but significantly lower levels of interleukin 1β and macrophage inflammatory protein 1α when they ingested apoptotic cells. In vivo, phagocytosis of intratracheally instilled latex beads was decreased in traumatized rats. The bronchoalveolar lavage concentrations of the phagocytosis-supporting surfactant proteins A and D after blunt chest trauma were slightly decreased, whereas surfactant protein D mRNA expression in AT2 cells was significantly increased after 2 h. These findings indicate that chest trauma augments the phagocytosis of apoptotic cells by AMΦ. Phagocytosis of opsonized beads enhances and ingestion of apoptotic cells downregulates the immunologic response following lung contusion. Our data emphasize the important role of phagocytosis during posttraumatic inflammation after lung contusion.

  10. Neutrophil and Alveolar Macrophage-Mediated Innate Immune Control of Legionella pneumophila Lung Infection via TNF and ROS.

    Directory of Open Access Journals (Sweden)

    Pascal Ziltener

    2016-04-01

    Full Text Available Legionella pneumophila is a facultative intracellular bacterium that lives in aquatic environments where it parasitizes amoeba. However, upon inhalation of contaminated aerosols it can infect and replicate in human alveolar macrophages, which can result in Legionnaires' disease, a severe form of pneumonia. Upon experimental airway infection of mice, L. pneumophila is rapidly controlled by innate immune mechanisms. Here we identified, on a cell-type specific level, the key innate effector functions responsible for rapid control of infection. In addition to the well-characterized NLRC4-NAIP5 flagellin recognition pathway, tumor necrosis factor (TNF and reactive oxygen species (ROS are also essential for effective innate immune control of L. pneumophila. While ROS are essential for the bactericidal activity of neutrophils, alveolar macrophages (AM rely on neutrophil and monocyte-derived TNF signaling via TNFR1 to restrict bacterial replication. This TNF-mediated antibacterial mechanism depends on the acidification of lysosomes and their fusion with L. pneumophila containing vacuoles (LCVs, as well as caspases with a minor contribution from cysteine-type cathepsins or calpains, and is independent of NLRC4, caspase-1, caspase-11 and NOX2. This study highlights the differential utilization of innate effector pathways to curtail intracellular bacterial replication in specific host cells upon L. pneumophila airway infection.

  11. Alendronate inhalation ameliorates elastase-induced pulmonary emphysema in mice by induction of apoptosis of alveolar macrophages.

    Science.gov (United States)

    Ueno, Manabu; Maeno, Toshitaka; Nishimura, Satoshi; Ogata, Fusa; Masubuchi, Hiroaki; Hara, Kenichiro; Yamaguchi, Kouichi; Aoki, Fumiaki; Suga, Tatsuo; Nagai, Ryozo; Kurabayashi, Masahiko

    2015-03-10

    Alveolar macrophages play a crucial role in the pathogenesis of emphysema, for which there is currently no effective treatment. Bisphosphonates are widely used to treat osteoclast-mediated bone diseases. Here we show that delivery of the nitrogen-containing bisphosphonate alendronate via aerosol inhalation ameliorates elastase-induced emphysema in mice. Inhaled, but not orally ingested, alendronate inhibits airspace enlargement after elastase instillation, and induces apoptosis of macrophages in bronchoalveolar fluid via caspase-3- and mevalonate-dependent pathways. Cytometric analysis indicates that the F4/80(+)CD11b(high)CD11c(mild) population characterizing inflammatory macrophages, and the F4/80(+)CD11b(mild)CD11c(high) population defining resident alveolar macrophages take up substantial amounts of the bisphosphonate imaging agent OsteoSense680 after aerosol inhalation. We further show that alendronate inhibits macrophage migratory and phagocytotic activities and blunts the inflammatory response of alveolar macrophages by inhibiting nuclear factor-κB signalling. Given that the alendronate inhalation effectively induces apoptosis in both recruited and resident alveolar macrophages, we suggest this strategy may have therapeutic potential for the treatment of emphysema.

  12. Decreased sialidase activity in alveolar macrophages of guinea pigs exposed to coal mine dust.

    Science.gov (United States)

    Terzidis-Trabelsi, H; Lefèvre, J P; Bignon, J; Lambré, C R

    1992-01-01

    The origin of immune dysfunctions that are observed in pneumoconiotic miners still remains unknown. There is evidence that the carbohydrate moiety of membrane glycoconjugates is of primary importance in many functions of immunocompetent cells. The glycosylation, and especially the sialylation level of membrane components of various lymphocyte and macrophage subsets, vary depending on the state of cellular differentiation and activation. Sialidases, which may regulate the amount of sialic acids exposed on the cell membrane, can thus be considered as immunoregulatory enzymes. In this report, the sialidase activity has been measured in alveolar macrophages (AM) and in cell-free bronchoalveolar lavage fluid (BALF) from guinea pigs exposed for 4 months to coal mine dust at a concentration of 300 mg/m3. The samples were collected by bronchoalveolar lavage 2 months after cessation of exposure. The sialidase activity in the cell-free fluid and in the purified alveolar macrophages showed a 10-fold decrease (p less than 0.001). Kinetic parameters of the enzyme such as Km and optimum pH did not change. This changed activity was specific for sialidase, as two other lysosomal glycosidases, beta-galactosidase and N-acetylglucosaminidase, showed unchanged activities. These results suggest the possibility that, by inducing a decreased sialidase activity, exposure to coal mine dust may lead to a modified expression of AM membrane-associated sialic acids giving rise to altered immune functions (i. e., phagocytosis, antigen processing, response to cytokines, etc.). PMID:1396442

  13. Tachykinins activate guinea-pig alveolar macrophages: involvement of NK2 and NK1 receptors.

    OpenAIRE

    Brunelleschi, S.; Vanni, L.; Ledda, F.; Giotti, A.; Maggi, C. A.; Fantozzi, R.

    1990-01-01

    1. The effects of substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) were evaluated on superoxide anion (O2-.) production by guinea-pig alveolar macrophages (AM). 2. SP dose-dependently (ED50 = 0.7 nM) evoked O2-. production from guinea-pig AM; the N-terminal heptapeptide, SP(1-7), was ineffective. In the presence of thiorphan (10(-5) M), an enkephalinase inhibitor, the stimulating effects of SP were not significantly modified. NKA and NKB were both able to induce O2-. production fro...

  14. [Toxicity of chongqing acid fogwater on rabbit alveolar macrophages in vitro].

    Science.gov (United States)

    Shu, W Q; Zhuo, J B

    1992-07-01

    We collected acid fogwater on a fogday and observed its toxic effects on rabbits' pulmonary alveolar macrophages (AM) in vitro. The fogwater was diluted into 4 concentrations: 1, 1/10, 1/100, and 1/1000 of the original fogwater and the exposure time was 12 hours. The results showed that both the AM's viability and the phagocytic capacity were depressed significantly, but the AM's lysosomal enzyme--acid phosphatase activity was found to be stimulated to increase. All these changes were directly correlated with the degree of pollution of the fogwater. Of these three toxicity indices, the most sensitive one was the change of AM's phagocytic capacity.

  15. Further characterization of a highly attenuated Yersinia pestis CO92 mutant deleted for the genes encoding Braun lipoprotein and plasminogen activator protease in murine alveolar and primary human macrophages.

    Science.gov (United States)

    van Lier, Christina J; Tiner, Bethany L; Chauhan, Sadhana; Motin, Vladimir L; Fitts, Eric C; Huante, Matthew B; Endsley, Janice J; Ponnusamy, Duraisamy; Sha, Jian; Chopra, Ashok K

    2015-03-01

    We recently characterized the Δlpp Δpla double in-frame deletion mutant of Yersinia pestis CO92 molecularly, biologically, and immunologically. While Braun lipoprotein (Lpp) activates toll-like receptor-2 to initiate an inflammatory cascade, plasminogen activator (Pla) protease facilitates bacterial dissemination in the host. The Δlpp Δpla double mutant was highly attenuated in evoking bubonic and pneumonic plague, was rapidly cleared from mouse organs, and generated humoral and cell-mediated immune responses to provide subsequent protection to mice against a lethal challenge dose of wild-type (WT) CO92. Here, we further characterized the Δlpp Δpla double mutant in two murine macrophage cell lines as well as in primary human monocyte-derived macrophages to gauge its potential as a live-attenuated vaccine candidate. We first demonstrated that the Δpla single and the Δlpp Δpla double mutant were unable to survive efficiently in murine and human macrophages, unlike WT CO92. We observed that the levels of Pla and its associated protease activity were not affected in the Δlpp single mutant, and, likewise, deletion of the pla gene from WT CO92 did not alter Lpp levels. Further, our study revealed that both Lpp and Pla contributed to the intracellular survival of WT CO92 via different mechanisms. Importantly, the ability of the Δlpp Δpla double mutant to be phagocytized by macrophages, to stimulate production of tumor necrosis factor-α and interleukin-6, and to activate the nitric oxide killing pathways of the host cells remained unaltered when compared to the WT CO92-infected macrophages. Finally, macrophages infected with either the WT CO92 or the Δlpp Δpla double mutant were equally efficient in their uptake of zymosan particles as determined by flow cytometric analysis. Overall, our data indicated that although the Δlpp Δpla double mutant of Y. pestis CO92 was highly attenuated, it retained the ability to elicit innate and subsequent acquired immune

  16. Training modifies innate immune responses in blood monocytes and in pulmonary alveolar macrophages.

    Science.gov (United States)

    Frellstedt, Linda; Waldschmidt, Ingrid; Gosset, Philippe; Desmet, Christophe; Pirottin, Dimitri; Bureau, Fabrice; Farnir, Frédéric; Franck, Thierry; Dupuis-Tricaud, Marie-Capucine; Lekeux, Pierre; Art, Tatiana

    2014-07-01

    In humans, strenuous exercise causes increased susceptibility to respiratory infections associated with down-regulated expression of Toll-like receptors (TLRs) and costimulatory and antigen-presenting molecules. Lower airway diseases are also a common problem in sport and racing horses. Because innate immunity plays an essential role in lung defense mechanisms, we assessed the effect of acute exercise and training on innate immune responses in two different compartments. Blood monocytes and pulmonary alveolar macrophages (PAMs) were collected from horses in untrained, moderately trained, intensively trained, and deconditioned states before and after a strenuous exercise test. The cells were analyzed for TLR messenger ribonucleic acid (mRNA) expression by real-time PCR in vitro, and cytokine production after in vitro stimulation with TLR ligands was measured by ELISA. Our results showed that training, but not acute exercise, modified the innate immune responses in both compartments. The mRNA expression of TLR3 was down-regulated by training in both cell types, whereas the expression of TLR4 was up-regulated in monocytes. Monocytes treated with LPS and a synthetic diacylated lipoprotein showed increased cytokine secretion in trained and deconditioned subjects, indicating the activation of cells at the systemic level. The production of TNF-α and IFN-β in nonstimulated and stimulated PAMs was decreased in trained and deconditioned horses and might therefore explain the increased susceptibility to respiratory infections. Our study reports a dissociation between the systemic and the lung response to training that is probably implicated in the systemic inflammation and in the pulmonary susceptibility to infection.

  17. Edema toxin impairs anthracidal phospholipase A2 expression by alveolar macrophages.

    Directory of Open Access Journals (Sweden)

    Benoit Raymond

    2007-12-01

    Full Text Available Bacillus anthracis, the etiological agent of anthrax, is a spore-forming gram-positive bacterium. Infection with this pathogen results in multisystem dysfunction and death. The pathogenicity of B. anthracis is due to the production of virulence factors, including edema toxin (ET. Recently, we established the protective role of type-IIA secreted phospholipase A2 (sPLA2-IIA against B. anthracis. A component of innate immunity produced by alveolar macrophages (AMs, sPLA2-IIA is found in human and animal bronchoalveolar lavages at sufficient levels to kill B. anthracis. However, pulmonary anthrax is almost always fatal, suggesting the potential impairment of sPLA2-IIA synthesis and/or action by B. anthracis factors. We investigated the effect of purified ET and ET-deficient B. anthracis strains on sPLA2-IIA expression in primary guinea pig AMs. We report that ET inhibits sPLA2-IIA expression in AMs at the transcriptional level via a cAMP/protein kinase A-dependent process. Moreover, we show that live B. anthracis strains expressing functional ET inhibit sPLA2-IIA expression, whereas ET-deficient strains induced this expression. This stimulatory effect, mediated partly by the cell wall peptidoglycan, can be counterbalanced by ET. We conclude that B. anthracis down-regulates sPLA2-IIA expression in AMs through a process involving ET. Our study, therefore, describes a new molecular mechanism implemented by B. anthracis to escape innate host defense. These pioneering data will provide new molecular targets for future intervention against this deadly pathogen.

  18. Study of possible changes brought about by plutonium oxide in the acid phosphatase activity of alveolar macrophages of the rabbit

    International Nuclear Information System (INIS)

    Rouvroy, Huguette

    1970-06-01

    This report describes the various techniques used for determining the acid phosphatase activity of alveolar rabbit macrophages after inhalation of radioactive plutonium oxide particles, exposure of the animals, removal and sampling of the alveolar cells, and technical dosage. The results obtained are presented; they do not make it possible, in this particular case, to affirm that an important change in the enzymatic activity studied occurs. (author) [fr

  19. Alveolar Macrophages Play a Key Role in Cockroach-Induced Allergic Inflammation via TNF-α Pathway

    Science.gov (United States)

    Kim, Joo Young; Sohn, Jung Ho; Choi, Je-Min; Lee, Jae-Hyun; Hong, Chein-Soo; Lee, Joo-Shil; Park, Jung-Won

    2012-01-01

    The activity of the serine protease in the German cockroach allergen is important to the development of allergic disease. The protease-activated receptor (PAR)-2, which is expressed in numerous cell types in lung tissue, is known to mediate the cellular events caused by inhaled serine protease. Alveolar macrophages express PAR-2 and produce considerable amounts of tumor necrosis factor (TNF)-α. We determined whether the serine protease in German cockroach extract (GCE) enhances TNF-α production by alveolar macrophages through the PAR-2 pathway and whether the TNF-α production affects GCE-induced pulmonary inflammation. Effects of GCE on alveolar macrophages and TNF-α production were evaluated using in vitro MH-S and RAW264.6 cells and in vivo GCE-induced asthma models of BALB/c mice. GCE contained a large amount of serine protease. In the MH-S and RAW264.7 cells, GCE activated PAR-2 and thereby produced TNF-α. In the GCE-induced asthma model, intranasal administration of GCE increased airway hyperresponsiveness (AHR), inflammatory cell infiltration, productions of serum immunoglobulin E, interleukin (IL)-5, IL-13 and TNF-α production in alveolar macrophages. Blockade of serine proteases prevented the development of GCE induced allergic pathologies. TNF-α blockade also prevented the development of such asthma-like lesions. Depletion of alveolar macrophages reduced AHR and intracellular TNF-α level in pulmonary cell populations in the GCE-induced asthma model. These results suggest that serine protease from GCE affects asthma through an alveolar macrophage and TNF-α dependent manner, reflecting the close relation of innate and adaptive immune response in allergic asthma model. PMID:23094102

  20. Certain biochemical specificities of alveolar macrophages from gamma-irradiated guinea pigs

    International Nuclear Information System (INIS)

    Najdenski, H.M.; Velyanov, D.K.

    1991-01-01

    The changes in the metabolism of alveolar macrophages (aMas) from 0.5 Gy and 2 Gy gamma-irradiated guinea pigs have been studied. In view of predominantly aerobic metabolism of the aMas the investigations include oxygen uptake in the presence of substrate glucose as well as the activity of cytochrome oxidase and acid phosphatase. The results show that in the macrophages obtained from 2 Gy exposed guinea pigs there is simultaneous intensification of the oxygen uptake in the presence of glucose and of cytochrome oxidase activity by the 3rd day after irradiation. In the macrophages from the 0.5 Gy exposed guinea pigs there is also parallelism in the intensification of the respiration and cytochrome oxidase activity but on the 7th day of the investigation. In both doses applied the activity of the acid phosphatase of macrophages sharply increases to reach the maximum values between the 3rd and 7th days after irradiation. This discrepancy between the intracellular bactericidal effect and the respiration activity and the acid phosphatase of the aMas give grounds to support the view of Pavillard and Rowlei that the total metabolism of the aMas is not a limiting factor in relation to their intracellular killing effect on the absorbed bacteria. 3 figs., 6 refs

  1. Functional ability and fate of pulmonary alveolar macrophages after intratracheal instillation into rats

    International Nuclear Information System (INIS)

    Snipes, M.B.; Feddersen, D.; Mueller, H.L.; Guilmette, R.A.; Haley, P.J.

    1988-01-01

    Pulmonary alveolar macrophages (PAM) from donor rats were intratracheally instilled into recipient rats to determine if donor macrophages were functionally similar to the recipient's own macrophages. Recipient and donor (extrinsic) PAM were equivalent in their ability to phagocytize 1.7 μm and 3.9 μm latex microspheres in vivo and sensitized sheep red blood cells in vitro. Also, the extrinsic PAM appeared functionally equivalent to recipient PAM with respect to ability to translocate into interstitial tissue and migrate to the lung-associated lymph nodes (LALN). The recipient PAN appeared to phagocytize the extrinsic PAM, but the extrinsic PAM did not appear to phagocytize the recipient PAM. This could represent a different degree of physiological coordination of intrinsic and extrinsic PAM activities in the lung. Overall, results indicated that extrinsic PAM can live and function in the lungs of recipient rats, and perform most or all of the functions ascribed to recipient PAM. Results also support the hypothesis that PAM are able to move into the pulmonary interstitium and translocate to the LALM without the involvement of other pulmonary macrophages. (author)

  2. Identification of beta 2-adrenoceptors on guinea pig alveolar macrophages using (-)-3-[125I]iodocyanopindolol

    International Nuclear Information System (INIS)

    Leurs, R.; Beusenberg, F.D.; Bast, A.; Van Amsterdam, J.G.; Timmerman, H.

    1990-01-01

    The beta-adrenoceptor antagonist (-)-3-[ 125 I]iodocyanopindolol ([ 125 I]ICYP) binds with high affinity and in saturable way to membranes of guinea pig alveolar macrophages. The equilibrium dissociation constant for [ 125 I]ICYP is 24.3 +/- 1.2 pM, and the number of binding sites is 166.3 +/- 13.7 fmol/mg protein (N = 4, +/- SEM). Displacement studies with selective antagonists showed that [ 125 I]ICYP labels beta 2-adrenoceptors on guinea pig alveolar macrophages

  3. Radiation effects on chemiluminescence of resting and immunologically activated alveolar macrophages

    International Nuclear Information System (INIS)

    Benichou, G.; Dormont, D.; Herodin, F.; Pasquier, C.; Hopital Saint Antoine, 75 - Paris

    1986-01-01

    In resting cells, for low radiation doses, a transient activation of chemiluminescence was observed, maximal at 3 Gy. At 10 Gy, CL returned to the control value; greater doses (above 30 Gy) induced a progressive diminution of the response which was abolished at 100 Gy. Activated alveolar macrophages showed a 30% decrease of the chemiluminescence at 10 Gy. The respiratory burst induced by opsonized zymosan was abolished at 30 Gy. IgG anti-MHC(IgGαB 1 ) activated specifically the GP S2 alveolar macrophages by alloantibody bipolar bridging; by contrast IgG which are directed against non-specific allogeneic determinants (IgG α B 3 ) or specific F(ab') 2 (F(ab') 2 αB 1 ) are unable to stimulate the cells. For all the tested doses, irradiation had no effect on this activation mechanism. The results with the three doses tested (3 Gy, 10 Gy, 30 Gy) are comparable to those using the positive control cells. The same results are obtained with the class II antigens and their specific IgG. (UK)

  4. In vivo metabolism of pulmonary alveolar epithelial type II pneumonocytes and macrophages from Syrian hamsters

    International Nuclear Information System (INIS)

    Pfleger, R.C.; Waide, J.J.

    1976-01-01

    Young adult Syrian hamsters were injected intraperitoneally with 14 C-glycerol and 3 H-palmitate 17 hr before they were sacrificed and pulmonary alveolar epithelial type II cells and pulmonary alveolar macrophages (PAM) were isolated. Incorporation of the two labeled components into the cellular lipids showed that the 3 H-specific activity of the phospholipids from the type II cells was three times that of the PAM and the utilization of 14 C-glycerol into phosphatidyl choline (PC) was 50% greater than incorporation into the PC from PAMs. The PC from type II cells showed that 30% was disaturated and from PAMs 21% was disaturated. Another phosphatide, phosphatidyl glycerol contained about one-third of the molecules in disaturated form. These data are consistent with the view that both type II cells and PAMs can synthesize surface-active phospholipids but it is generally accepted that only the pulmonary alveolar epithelial type II cells excrete the disaturated phospholipids which comprise the surface-active components of pulmonary surfactant

  5. Increased iron sequestration in alveolar macrophages in chronic obstructive pulmonary disease.

    Directory of Open Access Journals (Sweden)

    Quentin Philippot

    Full Text Available Free iron in lung can cause the generation of reactive oxygen species, an important factor in chronic obstructive pulmonary disease (COPD pathogenesis. Iron accumulation has been implicated in oxidative stress in other diseases, such as Alzheimer's and Parkinson's diseases, but little is known about iron accumulation in COPD. We sought to determine if iron content and the expression of iron transport and/or storage genes in lung differ between controls and COPD subjects, and whether changes in these correlate with airway obstruction. Explanted lung tissue was obtained from transplant donors, GOLD 2-3 COPD subjects, and GOLD 4 lung transplant recipients, and bronchoalveolar lavage (BAL cells were obtained from non-smokers, healthy smokers, and GOLD 1-3 COPD subjects. Iron-positive cells were quantified histologically, and the expression of iron uptake (transferrin and transferrin receptor, storage (ferritin and export (ferroportin genes was examined by real-time RT-PCR assay. Percentage of iron-positive cells and expression levels of iron metabolism genes were examined for correlations with airflow limitation indices (forced expiratory volume in the first second (FEV1 and the ratio between FEV1 and forced vital capacity (FEV1/FVC. The alveolar macrophage was identified as the predominant iron-positive cell type in lung tissues. Furthermore, the quantity of iron deposit and the percentage of iron positive macrophages were increased with COPD and emphysema severity. The mRNA expression of iron uptake and storage genes transferrin and ferritin were significantly increased in GOLD 4 COPD lungs compared to donors (6.9 and 3.22 fold increase, respectively. In BAL cells, the mRNA expression of transferrin, transferrin receptor and ferritin correlated with airway obstruction. These results support activation of an iron sequestration mechanism by alveolar macrophages in COPD, which we postulate is a protective mechanism against iron induced oxidative

  6. Uranyl nitrate-exposed rat alveolar macrophages cell death: Influence of superoxide anion and TNF α mediators

    International Nuclear Information System (INIS)

    Orona, N.S.; Tasat, D.R.

    2012-01-01

    Uranium compounds are widely used in the nuclear fuel cycle, military and many other diverse industrial processes. Health risks associated with uranium exposure include nephrotoxicity, cancer, respiratory, and immune disorders. Macrophages present in body tissues are the main cell type involved in the internalization of uranium particles. To better understand the pathological effects associated with depleted uranium (DU) inhalation, we examined the metabolic activity, phagocytosis, genotoxicity and inflammation on DU-exposed rat alveolar macrophages (12.5–200 μM). Stability and dissolution of DU could differ depending on the dissolvent and in turn alter its biological action. We dissolved DU in sodium bicarbonate (NaHCO 3 100 mM) and in what we consider a more physiological vehicle resembling human internal media: sodium chloride (NaCl 0.9%). We demonstrate that uranyl nitrate in NaCl solubilizes, enters the cell, and elicits its cytotoxic effect similarly to when it is diluted in NaHCO 3 . We show that irrespective of the dissolvent employed, uranyl nitrate impairs cell metabolism, and at low doses induces both phagocytosis and generation of superoxide anion (O 2 − ). At high doses it provokes the secretion of TNFα and through all the range of doses tested, apoptosis. We herein suggest that at DU low doses O 2 − may act as the principal mediator of DNA damage while at higher doses the signaling pathway mediated by O 2 − may be blocked, prevailing damage to DNA by the TNFα route. The study of macrophage functions after uranyl nitrate treatment could provide insights into the pathophysiology of uranium‐related diseases. -- Highlights: ► Uranyl nitrate effect on cultured macrophages is linked to the doses and independent of its solubility. ► At low doses uranyl nitrate induces generation of superoxide anion. ► At high doses uranyl nitrate provokes secretion of TNFα. ► Uranyl nitrate induces apoptosis through all the range of doses tested.

  7. Curcumin enhances human macrophage control of Mycobacterium tuberculosis infection.

    Science.gov (United States)

    Bai, Xiyuan; Oberley-Deegan, Rebecca E; Bai, An; Ovrutsky, Alida R; Kinney, William H; Weaver, Michael; Zhang, Gong; Honda, Jennifer R; Chan, Edward D

    2016-07-01

    With the worldwide emergence of highly drug-resistant tuberculosis (TB), novel agents that have direct antimycobacterial effects or that enhance host immunity are urgently needed. Curcumin is a polyphenol responsible for the bright yellow-orange colour of turmeric, a spice derived from the root of the perennial herb Curcuma longa. Curcumin is a potent inducer of apoptosis-an effector mechanism used by macrophages to kill intracellular Mycobacterium tuberculosis (MTB). An in vitro human macrophage infection model was used to determine the effects of curcumin on MTB survival. We found that curcumin enhanced the clearance of MTB in differentiated THP-1 human monocytes and in primary human alveolar macrophages. We also found that curcumin was an inducer of caspase-3-dependent apoptosis and autophagy. Curcumin mediated these anti-MTB cellular functions, in part, via inhibition of nuclear factor-kappa B (NFκB) activation. Curcumin protects against MTB infection in human macrophages. The host-protective role of curcumin against MTB in macrophages needs confirmation in an animal model; if validated, the immunomodulatory anti-TB effects of curcumin would be less prone to drug resistance development. © 2016 Asian Pacific Society of Respirology.

  8. The Cytokine TGF-β Promotes the Development and Homeostasis of Alveolar Macrophages.

    Science.gov (United States)

    Yu, Xueyang; Buttgereit, Anne; Lelios, Iva; Utz, Sebastian G; Cansever, Dilay; Becher, Burkhard; Greter, Melanie

    2017-11-21

    Alveolar macrophages (AMs) derive from fetal liver monocytes, which colonize the lung during embryonic development and give rise to fully mature AMs perinatally. AM differentiation requires granulocyte macrophage colony-stimulating factor (GM-CSF), but whether additional factors are involved in AM regulation is not known. Here we report that AMs, in contrast to most other tissue macrophages, were also dependent on transforming growth factor-β receptor (TGF-βR) signaling. Conditional deletion of TGF-βR in mice at different time points halted the development and differentiation of AMs. In adult mice, TGF-β was also critical for AM homeostasis. The source of TGF-β was AMs themselves, indicative of an autocrine loop that promotes AM self-maintenance. Mechanistically, TGF-βR signaling resulted in upregulation of PPAR-γ, a signature transcription factor essential for the development of AMs. These findings reveal an additional layer of complexity regarding the guidance cues, which govern the genesis, maturation, and survival of AMs. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Retention of inhaled plutonium oxide. Elimination procedures by pulmonary lavage and effect of the alveolar macrophage

    International Nuclear Information System (INIS)

    Nolibe, Daniel.

    1977-03-01

    A large fraction of the plutonium particles, reaching the deeper lung are retained in the alveolar macrophages during several months. Cell function changes were measured in vivo and in vitro. Stimulation of macrophage mobility and phagocytosis or natural clearance processes were uneffective on PuO 2 excretion. In vivo pulmonary lavage was the only effective therapy. The procedures of in toto pulmonary lavage in order to obtain the highest number of macrophages are described. A study of the physiological and histological consequences showed no long-term pathology, lesions observed during 48 h after lavage were restored quickly. A single lavage eliminated 12-25% only of the lung burden. A procedure of ten repeated lavages (1 per week) eliminated 60-90% of the lung burden. The action of lavage seemed twofold: direct elimination in the rinsing liquid and faster pulmonary clearance with low lymph node overload. Survivals in treated animals kept for long-term observations were compatible with the lung burdens remaining after treatment. Demontration of an inhibiting effect on pulmonary fibrosis should indicate a larger utilization [fr

  10. Solubilization of 241AmO2 in alveolar macrophage cultures

    International Nuclear Information System (INIS)

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

    1980-01-01

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

  11. Exposure of alveolar macrophages to polybrominated diphenyl ethers suppresses the release of pro-inflammatory products in vitro.

    Science.gov (United States)

    Hennigar, Stephen R; Myers, Jay L; Tagliaferro, Anthony R

    2012-04-01

    Inhalation of chemical pollutants has been associated with a reduced immune response in humans. Inhalation of dust is a major route of exposure for one endocrine-disrupting chemical and suspected xenoestrogen, polybrominated diphenyl ethers (PBDEs); however, the impact of PBDEs on immune function is unclear. The objective of this study was to investigate the action of PBDEs on cytokine and eicosanoid release by alveolar macrophages and determine whether the effects are mediated via the estrogen receptor. The production of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β, IL-10 and prostaglandin E(2) (PGE(2)) by porcine alveolar macrophages exposed to different concentrations of the pentabrominated diphenyl ether mixture, DE-71, were measured; cells were also exposed to varying concentrations of 17β-estradiol and the selective estrogen receptor-modulating agent, tamoxifen. Cells exposed to PBDEs released significantly less pro-inflammatory cytokines (TNF-α and IL-6) and PGE(2) compared with controls; IL-1β and IL-10 were not detected in the culture medium. Cells exposed to 17β-estradiol released significantly less TNF-α compared with controls, an effect that was reversed by the addition of tamoxifen; tamoxifen had no effect on the inhibition of TNF-α release by PBDEs. Although the suppression of TNF-α with DE-71 was similar to that of estrogen, the inhibitory effects of DE-71 were not found to be dependent on the estrogen receptor. Findings of this study suggest that chronic exposure to PBDEs suppressed innate immunity in vitro. Whether the immunosuppressant effects of PBDEs occur in vivo, remains to be determined.

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

    Directory of Open Access Journals (Sweden)

    Schwarze Per E

    2006-06-01

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

  13. Toxicity of ozone and nitrogen dioxide to alveolar macrophages: comparative study revealing differences in their mechanism of toxic action

    NARCIS (Netherlands)

    Rietjens, I. M.; Poelen, M. C.; Hempenius, R. A.; Gijbels, M. J.; Alink, G. M.

    1986-01-01

    The toxicity of ozone and nitrogen dioxide is generally ascribed to their oxidative potential. In this study their toxic mechanism of action was compared using an intact cell model. Rat alveolar macrophages were exposed by means of gas diffusion through a Teflon film. In this in vitro system, ozone

  14. Atorvastatin protected from paraquat-induced cytotoxicity in alveolar macrophages via down-regulation of TLR-4

    NARCIS (Netherlands)

    Alizadeh-Tabrizi, Nazli; Malekinejad, Hassan; Varasteh, Soheil; Cheraghi, Hadi

    2017-01-01

    The current study designed to clarify the mechanism of paraquat-induced cytotoxicity and protective effects of Atorvastatin on freshly isolated alveolar macrophages (AMs). AMs were collected via bronchoalveolar lavage and exposed to various concentrations of paraquat in the presence and absence of

  15. HUMAL ALVEOLAR MACROPHAGE RESPONSES TO AIR POLLUTION PARTICULATES ARE ASSOCIATED WITH INSOLUBLE OCMPONENTS OF COARSE MATERIAL, INCLUDING PARTICULATE ENDOTOXIN

    Science.gov (United States)

    Inhalation of particulate matter in the ambient air has been shown to cause pulmonary morbidity and exacerbate asthma. Alveolar macrophage (AM) are essential for effective removal of inhaled particles and microbes in the lower airways. While some particles minimally effect AM...

  16. Profiling microRNA expression in bovine alveolar macrophages using RNA-seq.

    Science.gov (United States)

    Vegh, Peter; Foroushani, Amir B K; Magee, David A; McCabe, Matthew S; Browne, John A; Nalpas, Nicolas C; Conlon, Kevin M; Gordon, Stephen V; Bradley, Daniel G; MacHugh, David E; Lynn, David J

    2013-10-01

    MicroRNAs (miRNAs) are important regulators of gene expression and are known to play a key role in regulating both adaptive and innate immunity. Bovine alveolar macrophages (BAMs) help maintain lung homeostasis and constitute the front line of host defense against several infectious respiratory diseases, such as bovine tuberculosis. Little is known, however, about the role miRNAs play in these cells. In this study, we used a high-throughput sequencing approach, RNA-seq, to determine the expression levels of known and novel miRNAs in unchallenged BAMs isolated from lung lavages of eight different healthy Holstein-Friesian male calves. Approximately 80 million sequence reads were generated from eight BAM miRNA Illumina sequencing libraries, and 80 miRNAs were identified as being expressed in BAMs at a threshold of at least 100 reads per million (RPM). The expression levels of miRNAs varied over a large dynamic range, with a few miRNAs expressed at very high levels (up to 800,000RPM), and the majority lowly expressed. Notably, many of the most highly expressed miRNAs in BAMs have known roles in regulating immunity in other species (e.g. bta-let-7i, bta-miR-21, bta-miR-27, bta-miR-99b, bta-miR-146, bta-miR-147, bta-miR-155 and bta-miR-223). The most highly expressed miRNA in BAMs was miR-21, which has been shown to regulate the expression of antimicrobial peptides in Mycobacterium leprae-infected human monocytes. Furthermore, the predicted target genes of BAM-expressed miRNAs were found to be statistically enriched for roles in innate immunity. In addition to profiling the expression of known miRNAs, the RNA-seq data was also analysed to identify potentially novel bovine miRNAs. One putatively novel bovine miRNA was identified. To the best of our knowledge, this is the first RNA-seq study to profile miRNA expression in BAMs and provides an important reference dataset for investigating the regulatory roles miRNAs play in this important immune cell type. Copyright

  17. Ex vivo expansion of alveolar macrophages with Mycobacterium tuberculosis from the resected lungs of patients with pulmonary tuberculosis

    Science.gov (United States)

    Petrunina, Ekaterina; Umpeleva, Tatiana; Karskanova, Svetlana; Bayborodin, Sergey; Vakhrusheva, Diana; Kravchenko, Marionella; Skornyakov, Sergey

    2018-01-01

    Tuberculosis (TB), with the Mycobacterium tuberculosis (Mtb) as the causative agent, remains to be a serious world health problem. Traditional methods used for the study of Mtb in the lungs of TB patients do not provide information about the number and functional status of Mtb, especially if Mtb are located in alveolar macrophages. We have developed a technique to produce ex vivo cultures of cells from different parts of lung tissues surgically removed from patients with pulmonary TB and compared data on the number of cells with Mtb inferred by the proposed technique to the results of bacteriological and histological analyses used for examination of the resected lungs. The ex vivo cultures of cells obtained from the resected lungs of all patients were largely composed of CD14-positive alveolar macrophages, foamy or not, with or without Mtb. Lymphocytes, fibroblasts, neutrophils, and multinucleate Langhans giant cells were also observed. We found alveolar macrophages with Mtb in the ex vivo cultures of cells from the resected lungs of even those TB patients, whose sputum smears and lung tissues did not contain acid-fast Mtb or reveal growing Mtb colonies on dense medium. The detection of alveolar macrophages with Mtb in ex vivo culture as soon as 16–18 h after isolation of cells from the resected lungs of all TB patients suggests that the technique proposed for assessing the level of infection in alveolar macrophages of TB patients has higher sensitivity than do prolonged bacteriological or pathomorphological methods. The proposed technique allowed us to rapidly (in two days after surgery) determine the level of infection with Mtb in the cells of the resected lungs of TB patients and, by the presence or absence of Mtb colonies, including those with cording morphology, the functional status of the TB agent at the time of surgery. PMID:29401466

  18. Alveolar macrophages and type I IFN in airway homeostasis and immunity.

    Science.gov (United States)

    Divangahi, Maziar; King, Irah L; Pernet, Erwan

    2015-05-01

    Globally, respiratory infections cause more than 4 million deaths per year, with influenza and tuberculosis (TB) in particular being major causes of mortality and morbidity. Although immune cell activation is critical for killing respiratory pathogens, this response must be tightly regulated to effectively control and eliminate invading microorganisms while minimizing immunopathology and maintaining pulmonary function. The distinct microenvironment of the lung is constantly patrolled by alveolar macrophages (Mφ), which are essential for tissue homeostasis, early pathogen recognition, initiation of the local immune response, and resolution of inflammation. Here, we focus on recent advances that have provided insight into the relation between pulmonary Mφ, type I interferon (IFN) signaling, and the delicate balance between protective and pathological immune responses in the lung. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Chronic cigarette smoking enhances spontaneous release of tumour necrosis factor-α from alveolar macrophages of rats

    Directory of Open Access Journals (Sweden)

    G. P. Pessina

    1993-01-01

    Full Text Available Some biological effects of chronic cigarette smoking (two cigarettes for 2 h, daily for 4 months in rats were evaluated. During the smoking period, body weight of smoker rats was always significantly lower than that of control rats. Immediately after the last smoking session the carboxyhaemoglobin concentration in the blood was about 8.5% and the polymorphonuclear cells in the bronchoalveolar fluid increased significantly. At the same time, enzymatic analyses on the supernatants of bronchoalveolar fluid revealed a significant increase of β-glucuronidase in the smoker group. Alveolar macrophages, collected 0, 8 and 24 h after the last smoking session, significantly increased the generation of superoxide anion and, after incubation for 24 h at 37° C in a humidified atmosphere, released significantly high amounts of TNF-α. When challenged with lipopolysaccharide, alveolar macrophages of smoker rats released much more TNF-α but, in such a case, TNF-α release was about one half of that observed in the control group. Peritoneal macrophages of both control and smoker rats were unable either to generate high levels of superoxide anion or to release significant amounts of TNF-α. The results clearly demonstrated the activated state of alveolar macrophages and the resting state of peritoneal macrophages.

  20. Human Induced Pluripotent Stem Cell-Derived Macrophages Share Ontogeny with MYB-Independent Tissue-Resident Macrophages

    Directory of Open Access Journals (Sweden)

    Julian Buchrieser

    2017-02-01

    Full Text Available Tissue-resident macrophages, such as microglia, Kupffer cells, and Langerhans cells, derive from Myb-independent yolk sac (YS progenitors generated before the emergence of hematopoietic stem cells (HSCs. Myb-independent YS-derived resident macrophages self-renew locally, independently of circulating monocytes and HSCs. In contrast, adult blood monocytes, as well as infiltrating, gut, and dermal macrophages, derive from Myb-dependent HSCs. These findings are derived from the mouse, using gene knockouts and lineage tracing, but their applicability to human development has not been formally demonstrated. Here, we use human induced pluripotent stem cells (iPSCs as a tool to model human hematopoietic development. By using a CRISPR-Cas9 knockout strategy, we show that human iPSC-derived monocytes/macrophages develop in an MYB-independent, RUNX1-, and SPI1 (PU.1-dependent fashion. This result makes human iPSC-derived macrophages developmentally related to and a good model for MYB-independent tissue-resident macrophages, such as alveolar and kidney macrophages, microglia, Kupffer cells, and Langerhans cells.

  1. Studies on the binding and transport processes of americium-241 hydroxide polymers in rat lung and bovine alveolar macrophages

    International Nuclear Information System (INIS)

    Taya, A.

    1986-03-01

    The binding of Am-241 hydroxide polymers to the cell components of rat lung was investigated using differential centrifugation, density gradient centrifugation with different media, gel chromatography, free flow electrophoresis and electron microscopic autoradiography with Pu-241. The bovine alveolar macrophage cultures were introduced as an in vitro test system for Am-241 uptake. Form the biochemical and electron microscopic studies it can be concluded that Am-241 is taken up by pulmonary macrophages, where its first storage site is probably the lysosome. Then the Am-241 seems to be solubilized in the lysosomes and to be bound to the cytosolic ferritin of macrophages. Am-241 might be released from the cells and crosses the alveolar membranes as bound to transferrin or as low molecular weight form. (orig.) [de

  2. Activated alveolar macrophage and lymphocyte alveolitis in extrathoracic sarcoidosis without radiological mediastinopulmonary involvement

    International Nuclear Information System (INIS)

    Wallaert, B.; Ramon, P.; Fournier, E.C.; Prin, L.; Tonnel, A.B.; Voisin, C.

    1986-01-01

    Cellular characteristics of BAL were investigated in 18 patients with proved extrathoracic sarcoidosis (that is, sarcoidosis that affected the skin, eyes, parotid glands, stomach, nose, kidneys, or meninges) without clinical or radiological mediastinopulmonary involvement. Computed tomography of the thorax was performed on five patients: four patients were normal, and one had enlarged lymph nodes (these enlargements were not detectable on the patient's chest roentgenogram). The results of pulmonary function tests were normal in all patients. The total BAL cell count did not differ significantly between controls and patients. Abnormal percentages of alveolar lymphocytes (from 18 to 87%) were noted in 15 out of 18 patients. SACE levels were normal in 15 patients. No pulmonary gallium uptake was detected. The chemiluminescence of AM's, whether spontaneous or PMA induced, was increased in five out of seven patients. The percentages of T3+ lymphocytes in sarcoidosis patients did not significantly differ from those in controls. The T4+:T8+ ratio was normal in four patients and slightly increased in one. Follow-up of patients showed that alveolar lymphocytosis is as lasting as extrathoracic involvement. Our data demonstrate increased percentages of lymphocytes and activated AM's in the BAL of patients with extrathoracic sarcoidosis. This may be due to the initial involvement of the respiratory tract in extrathoracic sarcoidosis or to the diffusion of activated macrophages and lymphocytes from an extrathoracic site into the lung

  3. Metalloelastase in lungs and alveolar macrophages is modulated by extracellular substance P in mice.

    Science.gov (United States)

    Xu, J; Xu, F; Barrett, E

    2008-07-01

    Metalloelastase (MMP-12), mainly produced by macrophages, has been shown to play a key role in the pathogenesis of emphysema in animal models. Chronic cigarette smoke increases pulmonary MMP-12, which is closely correlated with an elevation of pulmonary substance P (SP). Because alveolar macrophages (AMs) contain the neurokinin-1 receptor (NK1R), we tested whether SP was able to trigger the upregulation of MMP-12 synthesis in AMs by acting on the NK1R. AMs isolated from bronchoalveolar lavage cells in C3H/HeN mice were cultured with control medium or SP that was coupled without or with NK1R antagonists (CP-99,994 or aprepitant) for 24 h. We found that SP significantly increased the mRNA of MMP-12 and NK1R by 11-fold and 82%, respectively, in AMs (PNCAP) to degenerate PCFs, respectively. Our results show that NCAP treatment significantly decreased mRNA and protein levels of SP associated with a reduction NK1R and MMP-12 in the lungs and AMs. These findings suggest that SP has a modulatory effect on pulmonary MMP-12 by acting on NK1R to trigger MMP-12 syntheses in the AMs.

  4. Arctigenin Induces an Activation Response in Porcine Alveolar Macrophage Through TLR6-NOX2-MAPKs Signaling Pathway

    Science.gov (United States)

    Lu, Zheng; Chang, Lingling; Du, Qian; Huang, Yong; Zhang, Xiujuan; Wu, Xingchen; Zhang, Jie; Li, Ruizhen; Zhang, Zelin; Zhang, Wenlong; Zhao, Xiaomin; Tong, Dewen

    2018-01-01

    Arctigenin (ARG), one of the most active ingredients abstracted from seeds of Arctium lappa L., has been proved to exert promising biological activities such as immunomodulatory, anti-viral, and anti-cancer etc. However, the mechanism behind its immunomodulatory function still remains elusive to be further investigated. In this study, we found that ARG had no significant effects on the cell proliferation in both porcine alveolar macrophage cell line (3D4/21) and primary porcine derived alveolar macrophage. It remarkably increased the expression and secretion of the two cytokines including tumor necrosis factor-alpha (TNF-α) and transforming growth factor beta1 (TGF-β1) in a dose-dependent manner with the concomitant enhancement of phagocytosis, which are the indicators of macrophage activation. ARG also elevated the intracellular reactive oxygen species (ROS) production by activating NOX2-based NADPH oxidase. Furthermore, inhibition of ROS generation by diphenyliodonium and apocynin significantly suppressed ARG-induced cytokine secretion and phagocytosis increase, indicating the requirement of ROS for the porcine alveolar macrophage activation. In addition, TLR6-My88 excitation, p38 MAPK and ERK1/2 phosphorylation were all involved in the process. As blocking TLR6 receptor dramatically attenuated the NOX2 oxidase activation, cytokine secretion and phagocytosis increase. Inhibiting ROS generation almost abolished p38 and ERK1/2 phosphorylation, and the cytokine secretion could also be remarkably reduced by p38 and ERK1/2 inhibitors (SB203580 and UO126). Our finding gave a new insight of understanding that ARG could improve the immune-function of porcine alveolar macrophages through TLR6-NOX2 oxidase-MAPKs signaling pathway.

  5. Arctigenin Induces an Activation Response in Porcine Alveolar Macrophage Through TLR6-NOX2-MAPKs Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Zheng Lu

    2018-05-01

    Full Text Available Arctigenin (ARG, one of the most active ingredients abstracted from seeds of Arctium lappa L., has been proved to exert promising biological activities such as immunomodulatory, anti-viral, and anti-cancer etc. However, the mechanism behind its immunomodulatory function still remains elusive to be further investigated. In this study, we found that ARG had no significant effects on the cell proliferation in both porcine alveolar macrophage cell line (3D4/21 and primary porcine derived alveolar macrophage. It remarkably increased the expression and secretion of the two cytokines including tumor necrosis factor-alpha (TNF-α and transforming growth factor beta1 (TGF-β1 in a dose-dependent manner with the concomitant enhancement of phagocytosis, which are the indicators of macrophage activation. ARG also elevated the intracellular reactive oxygen species (ROS production by activating NOX2-based NADPH oxidase. Furthermore, inhibition of ROS generation by diphenyliodonium and apocynin significantly suppressed ARG-induced cytokine secretion and phagocytosis increase, indicating the requirement of ROS for the porcine alveolar macrophage activation. In addition, TLR6-My88 excitation, p38 MAPK and ERK1/2 phosphorylation were all involved in the process. As blocking TLR6 receptor dramatically attenuated the NOX2 oxidase activation, cytokine secretion and phagocytosis increase. Inhibiting ROS generation almost abolished p38 and ERK1/2 phosphorylation, and the cytokine secretion could also be remarkably reduced by p38 and ERK1/2 inhibitors (SB203580 and UO126. Our finding gave a new insight of understanding that ARG could improve the immune-function of porcine alveolar macrophages through TLR6-NOX2 oxidase-MAPKs signaling pathway.

  6. Arctigenin Induces an Activation Response in Porcine Alveolar Macrophage Through TLR6-NOX2-MAPKs Signaling Pathway.

    Science.gov (United States)

    Lu, Zheng; Chang, Lingling; Du, Qian; Huang, Yong; Zhang, Xiujuan; Wu, Xingchen; Zhang, Jie; Li, Ruizhen; Zhang, Zelin; Zhang, Wenlong; Zhao, Xiaomin; Tong, Dewen

    2018-01-01

    Arctigenin (ARG), one of the most active ingredients abstracted from seeds of Arctium lappa L. , has been proved to exert promising biological activities such as immunomodulatory, anti-viral, and anti-cancer etc. However, the mechanism behind its immunomodulatory function still remains elusive to be further investigated. In this study, we found that ARG had no significant effects on the cell proliferation in both porcine alveolar macrophage cell line (3D4/21) and primary porcine derived alveolar macrophage. It remarkably increased the expression and secretion of the two cytokines including tumor necrosis factor-alpha (TNF-α) and transforming growth factor beta1 (TGF-β1) in a dose-dependent manner with the concomitant enhancement of phagocytosis, which are the indicators of macrophage activation. ARG also elevated the intracellular reactive oxygen species (ROS) production by activating NOX2-based NADPH oxidase. Furthermore, inhibition of ROS generation by diphenyliodonium and apocynin significantly suppressed ARG-induced cytokine secretion and phagocytosis increase, indicating the requirement of ROS for the porcine alveolar macrophage activation. In addition, TLR6-My88 excitation, p38 MAPK and ERK1/2 phosphorylation were all involved in the process. As blocking TLR6 receptor dramatically attenuated the NOX2 oxidase activation, cytokine secretion and phagocytosis increase. Inhibiting ROS generation almost abolished p38 and ERK1/2 phosphorylation, and the cytokine secretion could also be remarkably reduced by p38 and ERK1/2 inhibitors (SB203580 and UO126). Our finding gave a new insight of understanding that ARG could improve the immune-function of porcine alveolar macrophages through TLR6-NOX2 oxidase-MAPKs signaling pathway.

  7. Characterization of part of the toxic effects due to alpha irradiation and to the physico-chemical properties of some actinides. An in vitro study on the alveolar macrophage

    International Nuclear Information System (INIS)

    Lizon, Celine

    1999-01-01

    The aim of this work was to characterize the specific effects due to radiotoxicity of α irradiation and the chemical toxicity of actinides. This was performed on alveolar macrophages extracted from rats and primates by pulmonary lavage. This was done by an in vitro study using either α irradiation from electrodeposited sources, or soluble actinides and lanthanides added to the culture medium. Necrosis and apoptosis induction were quantified after vital staining. For each treatment, cells were studied 1 or 7 days after plating. After either α irradiation or exposure to elements, the main route of death induced was apoptosis. After α irradiation, alveolar macrophages are very radioresistant cells. The observed D0 was between 30 and 100 Gy, depending on the species studied and the time in culture at exposure. In fact, alveolar macrophages irradiated after 1 week in culture have show less radioresistance than those treated after 1 day. The chemical toxicity of Uranium and Neptunium was independent both of time in culture at exposure and the animal species. The threshold we observed were respectively at 5 10 -4 and 3 10 -6 M. Moreover, within the concentrations studied, Thorium have not shown any toxicity towards alveolar macrophages. 1 day after plating macrophages, lanthanides exerts a higher chemical toxicity than actinides (threshold : 5 10 -6 M, Gadolinium, 5 10 -5 M, Cerium). These toxicities decreases more than 10 times after exposure 7 days after plating or for primates cells. This phenomenon seems to be due to cell harvesting and/or to cell adaptation to culture. Preliminary results show an impairment of cytokines production, which could be specific of the toxic studied. This was observed at concentrations which appeared non toxic as regards to apoptosis induction. The use of primates alveolar macrophages allow us to extrapolate some of the obtained results to Human. (author) [fr

  8. Uranyl nitrate-exposed rat alveolar macrophages cell death: Influence of superoxide anion and TNF α mediators

    Energy Technology Data Exchange (ETDEWEB)

    Orona, N.S. [School of Science and Technology, National University of General Martín, Avda Gral Paz 5445 (1650) San Martín, Buenos Aires (Argentina); Tasat, D.R., E-mail: deborah.tasat@unsam.edu.ar [School of Science and Technology, National University of General Martín, Avda Gral Paz 5445 (1650) San Martín, Buenos Aires (Argentina); School of Dentistry, University of Buenos Aires, M. T. de Alvear 2142 (1122), Buenos Aires (Argentina)

    2012-06-15

    Uranium compounds are widely used in the nuclear fuel cycle, military and many other diverse industrial processes. Health risks associated with uranium exposure include nephrotoxicity, cancer, respiratory, and immune disorders. Macrophages present in body tissues are the main cell type involved in the internalization of uranium particles. To better understand the pathological effects associated with depleted uranium (DU) inhalation, we examined the metabolic activity, phagocytosis, genotoxicity and inflammation on DU-exposed rat alveolar macrophages (12.5–200 μM). Stability and dissolution of DU could differ depending on the dissolvent and in turn alter its biological action. We dissolved DU in sodium bicarbonate (NaHCO{sub 3} 100 mM) and in what we consider a more physiological vehicle resembling human internal media: sodium chloride (NaCl 0.9%). We demonstrate that uranyl nitrate in NaCl solubilizes, enters the cell, and elicits its cytotoxic effect similarly to when it is diluted in NaHCO{sub 3}. We show that irrespective of the dissolvent employed, uranyl nitrate impairs cell metabolism, and at low doses induces both phagocytosis and generation of superoxide anion (O{sub 2}{sup −}). At high doses it provokes the secretion of TNFα and through all the range of doses tested, apoptosis. We herein suggest that at DU low doses O{sub 2}{sup −} may act as the principal mediator of DNA damage while at higher doses the signaling pathway mediated by O{sub 2}{sup −} may be blocked, prevailing damage to DNA by the TNFα route. The study of macrophage functions after uranyl nitrate treatment could provide insights into the pathophysiology of uranium‐related diseases. -- Highlights: ► Uranyl nitrate effect on cultured macrophages is linked to the doses and independent of its solubility. ► At low doses uranyl nitrate induces generation of superoxide anion. ► At high doses uranyl nitrate provokes secretion of TNFα. ► Uranyl nitrate induces apoptosis through

  9. Arctigenin Induces an Activation Response in Porcine Alveolar Macrophage Through TLR6-NOX2-MAPKs Signaling Pathway

    OpenAIRE

    Zheng Lu; Lingling Chang; Qian Du; Yong Huang; Xiujuan Zhang; Xingchen Wu; Jie Zhang; Ruizhen Li; Zelin Zhang; Wenlong Zhang; Xiaomin Zhao; Dewen Tong

    2018-01-01

    Arctigenin (ARG), one of the most active ingredients abstracted from seeds of Arctium lappa L., has been proved to exert promising biological activities such as immunomodulatory, anti-viral, and anti-cancer etc. However, the mechanism behind its immunomodulatory function still remains elusive to be further investigated. In this study, we found that ARG had no significant effects on the cell proliferation in both porcine alveolar macrophage cell line (3D4/21) and primary porcine derived alveol...

  10. MCPIP1 Regulates Alveolar Macrophage Apoptosis and Pulmonary Fibroblast Activation After in vitro Exposure to Silica.

    Science.gov (United States)

    Wang, Xingang; Zhang, Yuxia; Zhang, Wei; Liu, Haijun; Zhou, Zewei; Dai, Xiaoniu; Cheng, Yusi; Fang, Shencun; Zhang, Yingming; Yao, Honghong; Chao, Jie

    2016-05-01

    Silicosis is a fatal and fibrotic pulmonary disease caused by the inhalation of silica. After arriving at the alveoli, silica is ingested by alveolar macrophages (AMOs), in which monocyte chemotactic protein-induced protein 1 (MCPIP1) plays an essential role in controlling macrophage-mediated inflammatory responses. However, the mechanism of action of MCPIP1 in silicosis is poorly understood. Primary rat AMOs were isolated and treated with SiO2 (50 µg/cm(2)). MCPIP1 and AMO activation/apoptosis markers were detected by immunoblotting. MCPIP1 was down-regulated using siRNA in AMOs. The effects of AMOs on fibroblast activation and migration were evaluated using a gel contraction assay, a scratch assay, and a nested collagen matrix migration model. After exposure to SiO2, MCPIP1 was significantly increased in rat AMOs. Activation and apoptosis markers in AMOs were up-regulated after exposure to SiO2 Following siRNA-mediated silencing of MCPIP1 mRNA, the markers of AMO activation and apoptosis were significantly decreased. Rat pulmonary fibroblasts (PFBs) cultured in conditional medium from AMOs treated with MCPIP1 siRNA and SiO2 showed significantly less activation and migration compared with those cultured in conditional medium from AMOs treated with control siRNA and SiO2 CONCLUSION: Our data suggest a vital role for MCPIP1 in AMO apoptosis and PFB activation/migration induced by SiO2. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  11. Injurious effects of wool and grain dusts on alveolar epithelial cells and macrophages in vitro.

    Science.gov (United States)

    Brown, D M; Donaldson, K

    1991-01-01

    Epidemiological studies of workers in wool textile mills have shown a direct relation between the concentration of wool dust in the air and respiratory symptoms. Injurious effects of wool dust on the bronchial epithelium could be important in causing inflammation and irritation. A pulmonary epithelial cell line in vitro was therefore used to study the toxic effects of wool dust. Cells of the A549 epithelial cell line were labelled with 51Cr and treated with whole wool dusts and extracts of wool, after which injury was assessed. Also, the effects of grain dust, which also causes a form of airway obstruction, were studied. The epithelial injury was assessed by measuring 51Cr release from cells as an indication of lysis, and by monitoring cells which had detached from the substratum. No significant injury to A549 cells was caused by culture with any of the dusts collected from the air but surface "ledge" dust caused significant lysis at some doses. Quartz, used as a toxic control dust, caused significant lysis at the highest concentration of 100 micrograms/well. To determine whether any injurious material was soluble the dusts were incubated in saline and extracts collected. No extracts caused significant injury to epithelial cells. A similar lack of toxicity was found when 51Cr labelled control alveolar macrophages were targets for injury. Significant release of radiolabel was evident when macrophages were exposed to quartz at concentrations of 10 and 20 micrograms/well, there being no significant injury with either wool or grain dusts. These data suggest that neither wool nor grain dust produce direct injury to epithelial cells, and further studies are necessary to explain inflammation leading to respiratory symptoms in wool and grain workers. PMID:2015211

  12. Mast cell granules modulate alveolar macrophage respiratory-burst activity and eicosanoid metabolism.

    Science.gov (United States)

    Rock, M J; Despot, J; Lemanske, R F

    1990-10-01

    Alveolar macrophages (AMs) and mast cells reside in the airway, and both have been demonstrated to contribute independently to allergic inflammatory responses through the generation of respiratory-burst metabolites and the release of biologically active mediators, respectively. Since mast cell granules (MCGs) contain mediators that could potentially interact with the AM respiratory burst, we investigated the effects of isolated MCGs on this important inflammatory pathway of the AM. MCGs and AMs were obtained by peritoneal and tracheoalveolar lavage, respectively, of Sprague-Dawley rats. First, the overall respiratory-burst activity was measured by luminal-enhanced chemiluminescence (CL), and second, the individual oxygen species contributing to CL (superoxide anion [O2-], hydrogen peroxide [H2O2], and hypochlorous acid) were measured. MCGs alone enhanced AM CL responses to an equivalent degree compared to zymosan-stimulated AMs. However, AMs preincubated with MCGs followed by zymosan stimulation significantly and synergistically enhanced the CL responses. This enhanced CL was not due to an increased production of O2-, H2O2, or hypochlorous acid; in fact, there were decreased measured amounts of O2- and H2O2 from zymosan-stimulated AMs in the presence of MCGs, most likely caused by the content of granules of superoxide dismutase and peroxidase, respectively. The lipoxygenase inhibitor, nordihydroguaiaretic acid, completely abolished the enhanced CL of AM preincubated with MCGs and subsequently stimulated by zymosan, but O2- production was not affected by nordihydroguaiaretic acid. Taken together, these results suggest that derivatives of arachidonic acid metabolism, most likely those of the lipoxygenase pathway, are responsible for the enhanced AM CL response observed in the presence of MCGs. Thus, mast cell-macrophage interactions may be important within the airway in enhancing the generation of mediators that contribute to tissue inflammation and bronchospasm.

  13. Evaluation of amniotic mesenchymal cell derivatives on cytokine production in equine alveolar macrophages: an in vitro approach to lung inflammation.

    Science.gov (United States)

    Zucca, Enrica; Corsini, Emanuela; Galbiati, Valentina; Lange-Consiglio, Anna; Ferrucci, Francesco

    2016-09-20

    Data obtained in both animal models and clinical trials suggest that cell-based therapies represent a potential therapeutic strategy for lung repair and remodeling. Recently, new therapeutic approaches based on the use of stem cell derivatives (e.g., conditioned medium (CM) and microvesicles (MVs)) to regenerate tissues and improve their functions were proposed. The aim of this study was to investigate the immunomodulatory effects of equine amniotic mesenchymal cell derivatives on lipopolysaccharide (LPS)-induced cytokine production in equine alveolar macrophages, which may be beneficial in lung inflammatory disorders such as recurrent airway obstruction (RAO) in horses. RAO shares many features with human asthma, including an increased number of cells expressing mRNA for interleukin (IL)-4 and IL-5 and a decreased expression of IFN-γ in bronchoalveolar lavage fluid (BALF) of affected horses. The release of TNF-α, IL-6, and TGF-β1 at different time points (1, 24, 48, and 72 h) was measured in equine alveolar macrophages stimulated or not with LPS (10 and 100 ng/mL) in the presence or absence of 10 % CM or 50 × 10(6) MVs/mL. Cytokines were measured using commercially available ELISA kits. For multiple comparisons, analysis of variance was used with Tukey post-hoc test. Differences were considered significant at p ≤ 0.05. Significant modulatory effects of CM on LPS-induced TNF-α release at 24 h, and of both CM and MVs on TNF-α release at 48 h were observed. A trend toward a modulatory effect of both CM and MVs on the release of TGF-β and possibly IL-6 was visible over time. Results support the potential use of CM and MVs in lung regenerative medicine, especially in situations in which TGF-β may be detrimental, such as respiratory allergy. Further studies should evaluate the potential clinical applications of CM and MVs in equine lung diseases, such as RAO and other inflammatory disorders.

  14. Cytogenetic effects of cigarette smoke on pulmonary alveolar macrophages of the rat

    International Nuclear Information System (INIS)

    Ritchideh, K.; Chen, B.T.; Mauderly, J.L.; Brooks, A.L.

    1988-01-01

    This study was part of a larger investigation of the health effects resulting from different methods of exposing rats to cigarette smoke. Cytogenetic effects of cigarette smoke on rat pulmonary alveolar macrophages (PAMs) were evaluated. Fischer 344/N, male rats (4/group) were randomly assigned to 5 different exposure groups: (1) nose-only sham-exposed control, (2) whole-body sham-exposed control, (3) nose-only intermittent, (4) nose-only continuous, and (5) whole-body continuous. Sham controls were exposed to clean air. PAMs were obtained by lung lavage and chromosomal damage was measured. Multiple comparison demonstrated no significant differences between smoke-exposed groups and their respective sham-exposed controls, between the sham-exposed groups, or among the three smoke exposed groups. Highly significant smoke-induced differences in both structural and numerical aberrations were observed when data for the respective control groups and exposed groups were pooled and compared. Results from this study demonstrate the clastogenicity of cigarette smoke on rat PAM. (author)

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

    International Nuclear Information System (INIS)

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

    1988-01-01

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

  16. Radiosensitivity of pulmonary alveolar macrophages in rats exposed to local X-irradiation

    International Nuclear Information System (INIS)

    Gong Yifen; Fei Lihua; Wu Dechang

    1987-01-01

    The radiosensitivity of pulmonary alveolar macrophages (PAMs) in rats exposed to local thoracic X-irradiatoin was studied. The percentages of mitotic and labeling cells were used as biological endpoints. The parameters of radiosensitivity of PAMs obtained on the second day after local exposure are as follows: D 0 = 0.68 Gy, Dq = 0.06 Gy, n = 1.1 for mitotic cells and D 0 = 1.04 Gy, Dq = 0.12 Gy, n = 1.12 for labeling cells. The parameters of radiosensitivity of PAMs in bronchical lavage obtained immediately after X-irradiation are: D 0 = 3.56 Gy, Dq = 0.77 Gy, n = 1.24 for labeling cells and D 0 = 3.69 Gy, Dq = 0.35 Gy, n = 1.1 for mitotic cells. The comparison of thses results indicates that the radiation effect on PAMs obtained immediately after X-irradiation is less severe than that of PAMs obtained 2 days later. It might be caused by the delay of cell cycle within 2 days after X-irradiation

  17. Cooperativity between CD8+ T cells, non-neutralizing antibodies, and alveolar macrophages is important for heterosubtypic influenza virus immunity.

    Directory of Open Access Journals (Sweden)

    Brian J Laidlaw

    2013-03-01

    Full Text Available Seasonal epidemics of influenza virus result in ∼36,000 deaths annually in the United States. Current vaccines against influenza virus elicit an antibody response specific for the envelope glycoproteins. However, high mutation rates result in the emergence of new viral serotypes, which elude neutralization by preexisting antibodies. T lymphocytes have been reported to be capable of mediating heterosubtypic protection through recognition of internal, more conserved, influenza virus proteins. Here, we demonstrate using a recombinant influenza virus expressing the LCMV GP33-41 epitope that influenza virus-specific CD8+ T cells and virus-specific non-neutralizing antibodies each are relatively ineffective at conferring heterosubtypic protective immunity alone. However, when combined virus-specific CD8 T cells and non-neutralizing antibodies cooperatively elicit robust protective immunity. This synergistic improvement in protective immunity is dependent, at least in part, on alveolar macrophages and/or other lung phagocytes. Overall, our studies suggest that an influenza vaccine capable of eliciting both CD8+ T cells and antibodies specific for highly conserved influenza proteins may be able to provide heterosubtypic protection in humans, and act as the basis for a potential "universal" vaccine.

  18. Zbtb7a induction in alveolar macrophages is implicated in anti-HLA-mediated lung allograft rejection.

    Science.gov (United States)

    Nayak, Deepak K; Zhou, Fangyu; Xu, Min; Huang, Jing; Tsuji, Moriya; Yu, Jinsheng; Hachem, Ramsey; Gelman, Andrew E; Bremner, Ross M; Smith, Michael A; Mohanakumar, Thalachallour

    2017-07-12

    Chronic rejection significantly limits long-term success of solid organ transplantation. De novo donor-specific antibodies (DSAs) to mismatched donor human leukocyte antigen after human lung transplantation predispose lung grafts to chronic rejection. We sought to delineate mediators and mechanisms of DSA pathogenesis and to define early inflammatory events that trigger chronic rejection in lung transplant recipients and obliterative airway disease, a correlate of human chronic rejection, in mouse. Induction of transcription factor zinc finger and BTB domain containing protein 7a (Zbtb7a) was an early response critical in the DSA-induced chronic rejection. A cohort of human lung transplant recipients who developed DSA and chronic rejection demonstrated greater Zbtb7a expression long before clinical diagnosis of chronic rejection compared to nonrejecting lung transplant recipients with stable pulmonary function. Expression of DSA-induced Zbtb7a was restricted to alveolar macrophages (AMs), and selective disruption of Zbtb7a in AMs resulted in less bronchiolar occlusion, low immune responses to lung-restricted self-antigens, and high protection from chronic rejection in mice. Additionally, in an allogeneic cell transfer protocol, antigen presentation by AMs was Zbtb7a-dependent where AMs deficient in Zbtb7a failed to induce antibody and T cell responses. Collectively, we demonstrate that AMs play an essential role in antibody-induced pathogenesis of chronic rejection by regulating early inflammation and lung-restricted humoral and cellular autoimmunity. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  19. The innate and adaptive immune response induced by alveolar macrophages exposed to ambient particulate matter

    International Nuclear Information System (INIS)

    Miyata, Ryohei; Eeden, Stephan F. van

    2011-01-01

    Emerging epidemiological evidence suggests that exposure to particulate matter (PM) air pollution increases the risk of cardiovascular events but the exact mechanism by which PM has adverse effects is still unclear. Alveolar macrophages (AM) play a major role in clearing and processing inhaled PM. This comprehensive review of research findings on immunological interactions between AM and PM provides potential pathophysiological pathways that interconnect PM exposure with adverse cardiovascular effects. Coarse particles (10 μm or less, PM 10 ) induce innate immune responses via endotoxin-toll-like receptor (TLR) 4 pathway while fine (2.5 μm or less, PM 2.5 ) and ultrafine particles (0.1 μm or less, UFP) induce via reactive oxygen species generation by transition metals and/or polyaromatic hydrocarbons. The innate immune responses are characterized by activation of transcription factors [nuclear factor (NF)-κB and activator protein-1] and the downstream proinflammatory cytokine [interleukin (IL)-1β, IL-6, and tumor necrosis factor-α] production. In addition to the conventional opsonin-dependent phagocytosis by AM, PM can also be endocytosed by an opsonin-independent pathway via scavenger receptors. Activation of scavenger receptors negatively regulates the TLR4-NF-κB pathway. Internalized particles are subsequently subjected to adaptive immunity involving major histocompatibility complex class II (MHC II) expression, recruitment of costimulatory molecules, and the modulation of the T helper (Th) responses. AM show atypical antigen presenting cell maturation in which phagocytic activity decreases while both MHC II and costimulatory molecules remain unaltered. PM drives AM towards a Th1 profile but secondary responses in a Th1- or Th-2 up-regulated milieu drive the response in favor of a Th2 profile.

  20. microRNA-124 negatively regulates TLR signaling in alveolar macrophages in response to mycobacterial infection.

    Science.gov (United States)

    Ma, Chunyan; Li, Yong; Li, Min; Deng, Guangcun; Wu, Xiaoling; Zeng, Jin; Hao, Xiujing; Wang, Xiaoping; Liu, Jing; Cho, William C S; Liu, Xiaoming; Wang, Yujiong

    2014-11-01

    The emerging roles of microRNAs (miRNAs) in regulating immune responses have attracted increasing attention in recent years; and the alveolar macrophages (AMs) are the main targets of mycobacterial infection, which play a pivotal role in the pathogenesis of Mycobacterium tuberculosis infection. However, the immunoregulatory role of miRNAs in AMs has not been fully demonstrated. In this study, we find that miR-124 is up-regulated in the peripheral leukocytes of patients with pulmonary tuberculosis; furthermore, the expression miR-124 can be induced upon Mycobacterium bovis Bacillus Calmette-Guerin (BCG) infection in both RAW264.7 AM cells in vitro and murine AMs in vivo. Mechanistically, miR-124 is able to modulate toll-like receptor (TLR) signaling activity in RAW264.7 cells in response to BCG infection. In this regard, multiple components of TLR signaling cascade, including the TLR6, myeloid differentiation factor 88 (MyD88), TNFR-associated factor 6 and tumor necrosis factor-α are directly targeted by miR-124. In addition, both overexpression of TLR signaling adaptor MyD88 and BCG infection are able to augment miR-124 transcription, while MyD88 expression silenced by small interfering RNA dramatically suppresses miR-124 expression in AMs in vitro. Moreover, the abundance of miR-124 transcript in murine AMs of MyD88 deficient mice is significantly less than that of their wild-type or heterozygous littermates; and the BCG infection fails to induce miR-124 expression in the lung of MyD88 deficient mouse. These results indicate a negative regulatory role of miR-124 in fine-tuning inflammatory response in AMs upon mycobacterial infection, in part through a mechanism by directly targeting TLR signaling. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. The innate and adaptive immune response induced by alveolar macrophages exposed to ambient particulate matter

    Energy Technology Data Exchange (ETDEWEB)

    Miyata, Ryohei; Eeden, Stephan F. van, E-mail: Stephan.vanEeden@hli.ubc.ca

    2011-12-15

    Emerging epidemiological evidence suggests that exposure to particulate matter (PM) air pollution increases the risk of cardiovascular events but the exact mechanism by which PM has adverse effects is still unclear. Alveolar macrophages (AM) play a major role in clearing and processing inhaled PM. This comprehensive review of research findings on immunological interactions between AM and PM provides potential pathophysiological pathways that interconnect PM exposure with adverse cardiovascular effects. Coarse particles (10 {mu}m or less, PM{sub 10}) induce innate immune responses via endotoxin-toll-like receptor (TLR) 4 pathway while fine (2.5 {mu}m or less, PM{sub 2.5}) and ultrafine particles (0.1 {mu}m or less, UFP) induce via reactive oxygen species generation by transition metals and/or polyaromatic hydrocarbons. The innate immune responses are characterized by activation of transcription factors [nuclear factor (NF)-{kappa}B and activator protein-1] and the downstream proinflammatory cytokine [interleukin (IL)-1{beta}, IL-6, and tumor necrosis factor-{alpha}] production. In addition to the conventional opsonin-dependent phagocytosis by AM, PM can also be endocytosed by an opsonin-independent pathway via scavenger receptors. Activation of scavenger receptors negatively regulates the TLR4-NF-{kappa}B pathway. Internalized particles are subsequently subjected to adaptive immunity involving major histocompatibility complex class II (MHC II) expression, recruitment of costimulatory molecules, and the modulation of the T helper (Th) responses. AM show atypical antigen presenting cell maturation in which phagocytic activity decreases while both MHC II and costimulatory molecules remain unaltered. PM drives AM towards a Th1 profile but secondary responses in a Th1- or Th-2 up-regulated milieu drive the response in favor of a Th2 profile.

  2. Ablation of the Leptin receptor in Myeloid Cells Impairs Pulmonary Clearance of Streptococcus Pneumoniae and Alveolar Macrophage Bactericidal Function.

    Science.gov (United States)

    Mancuso, Peter; Curtis, Jeffrey L; Freeman, Christine M; Peters-Golden, Marc; Weinberg, Jason B; Myers, Martin G

    2018-03-22

    Leptin is a pleiotropic hormone produced by white adipose tissue that regulates appetite and many physiologic functions including the immune response to infection. Genetic leptin deficiency in humans and mice impairs host defenses against respiratory tract infections. Since leptin deficiency is associated with obesity and other metabolic abnormalities, we generated mice that lack the leptin receptor (LepRb) in cells of the myeloid linage (LysM-LepRb-KO) to evaluate its impact in lean metabolically normal mice in a murine model of pneumococcal pneumonia. We observed higher lung and spleen bacterial burdens in LysM-LepRb-KO mice following an intratracheal challenge with S. pneumoniae. Although numbers of leukocytes recovered from bronchoalveolar lavage fluid did not differ between groups, we did observe higher levels of pulmonary IL-13 and TNFα in LysM-LepRb-KO mice 48 h post-infection. Phagocytosis and killing of ingested S. pneumoniae were also impaired in alveolar macrophages (AM)s from LysM-LepRb-KO mice in vitro, and was associated with reduced LTB4 and enhanced PGE2 synthesis in vitro. Pretreatment of AMs with LTB4 and the cyclooxygenase inhibitor, indomethacin, restored phagocytosis but not bacterial killing in vitro. These results, confirm our previous observations in leptin-deficient (ob/ob) and fasted mice, and demonstrate that decreased leptin action, as opposed to metabolic irregularities associated with obesity or starvation, are responsible for the defective host defense against pneumococcal pneumonia. They also provide novel targets for therapeutic intervention in humans with bacterial pneumonia.

  3. Autochthonous human alveolar echinococcosis in a Hungarian patient.

    Science.gov (United States)

    Dezsényi, Balázs; Strausz, Tamás; Makrai, Zita; Csomor, Judit; Danka, József; Kern, Peter; Rezza, Giovanni; Barth, Thomas F E; Casulli, Adriano

    2017-02-01

    Alveolar echinococcosis is a zoonotic parasitic disease causing a severe clinical condition and is known as the most deadly of all helminth infections. Moreover, this disease is also an increasing concern in Northern and Eastern Europe due to its spread in the wildlife animal host. An asymptomatic 70-year-old woman from south-western Hungary was diagnosed with multiple liver lesions. Imaging techniques (ultrasound, computed tomography and magnetic resonance imaging), serology (ELISA, indirect hemagglutination and Western blot), and conventional staining methods (hematoxylin-eosin and periodic acid-Schiff) were used for the detection of the disease. A histopathological re-evaluation of formalin-fixed paraffin block by immunohistochemical staining with the monoclonal antibody Em2G11 definitively confirmed the diagnosis of alveolar echinococcosis. To our knowledge, this is the first confirmed autochthonous case of human alveolar echinococcosis in Hungary. To what extent diagnostic difficulties may contribute to underestimate this zoonosis in Eastern Europe is unknown. Differential diagnosis with alveolar echinococcosis should be considered for patients with multiple, tumor-like cystic lesions of the liver, in countries where this parasite is emerging.

  4. The exhibition to ozone diminishes the adherence and increases the membrane permeability of macrophages alveolar of rate

    International Nuclear Information System (INIS)

    Garcia, J.

    2000-01-01

    Ozone gas is generated photochemically in areas with high levels of automotive or industrial emissions, and causes irritation and inflammation of the airways if inhaled. Rat alveolar macrophages were obtained by lung lavage from male Sprague Dawley rats and used as a model to assess ozone induced cell damage (0,594 ppm for up to 60 minutes). Ozone exposure caused loss of cell adherence to a polystyrene substrate and increased membrane permeability, as noted by increases in specific 51 Cr release and citoplasmic calcium levels. The results indicate that the cell membrane is a target for ozone damage. Elevations of cytoplasmic calcium could mediate other macrophage responses to ozone , including eicosanoid and nitric oxide production, with concomitant decreases in phagocytic ability and superoxide production. (Author) [es

  5. Cysteamine-mediated clearance of antibiotic-resistant pathogens in human cystic fibrosis macrophages.

    Directory of Open Access Journals (Sweden)

    Chandra L Shrestha

    Full Text Available Members of the Burkholderia cepacia complex are virulent, multi-drug resistant pathogens that survive and replicate intracellularly in patients with cystic fibrosis (CF. We have discovered that B. cenocepacia cannot be cleared from CF macrophages due to defective autophagy, causing continued systemic inflammation and infection. Defective autophagy in CF is mediated through constitutive reactive oxygen species (ROS activation of transglutaminase-2 (TG2, which causes the sequestration (accumulation of essential autophagy initiating proteins. Cysteamine is a TG2 inhibitor and proteostasis regulator with the potential to restore autophagy. Therefore, we sought to examine the impact of cysteamine on CF macrophage autophagy and bacterial killing. Human peripheral blood monocyte-derived macrophages (MDMs and alveolar macrophages were isolated from CF and non-CF donors. Macrophages were infected with clinical isolates of relevant CF pathogens. Cysteamine caused direct bacterial growth killing of live B. cenocepacia, B. multivorans, P. aeruginosa and MRSA in the absence of cells. Additionally, B. cenocepacia, B. multivorans, and P. aeruginosa invasion were significantly decreased in CF MDMs treated with cysteamine. Finally, cysteamine decreased TG2, p62, and beclin-1 accumulation in CF, leading to increased Burkholderia uptake into autophagosomes, increased macrophage CFTR expression, and decreased ROS and IL-1β production. Cysteamine has direct anti-bacterial growth killing and improves human CF macrophage autophagy resulting in increased macrophage-mediated bacterial clearance, decreased inflammation, and reduced constitutive ROS production. Thus, cysteamine may be an effective adjunct to antibiotic regimens in CF.

  6. Modulation of epithelial sodium channel in human alveolar epithelial ...

    African Journals Online (AJOL)

    Modulation of epithelial sodium channel in human alveolar epithelial cells by lipoxin A4 through AhR-cAMP-dependent pathway. Bi-Huan Cheng1,2, Li-Wei Pan2, Sheng-Rong Zhang3, Bin-Yu Ying2, Ben-Ji. Wang2, Guo-Liang Lin2 and Shi-Fang Ding1*. 1Department of Critical Care Medicine, Qilu Hospital of Shandong ...

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-01-15

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

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

    Directory of Open Access Journals (Sweden)

    Xiyuan Bai

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

  9. Genistein suppresses Prevotella intermedia lipopolysaccharide-induced inflammatory response in macrophages and attenuates alveolar bone loss in ligature-induced periodontitis.

    Science.gov (United States)

    Choi, Eun-Young; Bae, Seung Han; Ha, Min Hee; Choe, So-Hui; Hyeon, Jin-Yi; Choi, Jeom-Il; Choi, In Soon; Kim, Sung-Jo

    2016-02-01

    Genistein is a major isoflavone subclass of flavonoids found in soybean and a potent tyrosine kinase inhibitor. The present study aimed to assess the effect of genistein on the production of proinflammatory mediators in murine macrophages stimulated with lipopolysaccharide (LPS) isolated from Prevotella intermedia, a pathogen associated with different forms of periodontal disease, and to evaluate its possible influence on alveolar bone loss in ligature-induced periodontitis using micro-computed tomography (micro-CT) analysis as well. LPS was isolated from P. intermedia ATCC 25611 by using the standard hot phenol-water method. Culture supernatants were analyzed for nitric oxide (NO) and interleukin-6 (IL-6). Inducible NO synthase (iNOS) protein expression was evaluated by immunoblot analysis. Real-time PCR was carried out to measure iNOS and IL-6 mRNA expression. In addition, effect of genistein on alveolar bone loss was evaluated in a rat model of experimental periodontitis using micro-CT analysis. Genistein significantly attenuated P. intermedia LPS-induced production of iNOS-derived NO and IL-6 with attendant decrease in their mRNA expression in RAW264.7 cells. In addition, when genistein was administered to rats, decreases in alveolar bone height and bone volume fraction induced by ligature placement were significantly inhibited. Genistein administration also prevented ligature-induced alterations in the microstructural parameters of trabecular bone, including trabecular thickness, trabecular separation, bone mineral density and structure model index. While additional studies are required, we suggest that genistein could be utilized for the therapy of human periodontitis in the future. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Role of lysosomal enzymes released by alveolar macrophages in the pathogenesis of the acute phase of hypersensitivity pneumonitis

    Directory of Open Access Journals (Sweden)

    J. L. Pérez-Arellano

    1995-01-01

    Full Text Available Hydrolytic enzymes are the major constituents of alveolar macrophages (AM and have been shown to be involved in many aspects of the inflammatory pulmonary response. The aim of this study was to evaluate the role of lysosomal enzymes in the acute phase of hypersensitivity pneumonitis (HPs. An experimental study on AM lysosomal enzymes of an HP-guinea-pig model was performed. The results obtained both in vivo and in vitro suggest that intracellular enzymatic activity decrease is, at least partly, due to release of lysosomal enzymes into the medium. A positive but slight correlation was found between extracellular lysosomal activity and four parameters of lung lesion (lung index, bronchoalveolar fluid total (BALF protein concentration, BALF LDH and BALF alkaline phosphatase activities. All the above findings suggest that the AM release of lysosomal enzymes during HP is a factor involved, although possibly not the only one, in the pulmonary lesions appearing in this disease.

  11. Stimulation of alveolar macrophages by BCG vaccine enhances the process of lung fibrosis induced by bleomycin.

    Science.gov (United States)

    Chyczewska, E; Chyczewski, L; Bańkowski, E; Sułkowski, S; Nikliński, J

    1993-01-01

    It was found that the BCG vaccine injected subcutaneously to the rats enhances the process of lung fibrosis induced by bleomycin. Pretreatment of rats with this vaccine results in accumulation of activated macrophages in lung interstitium and in the bronchoalveolar spaces. It may be suggested that the activated macrophages release various cytokines which may stimulate the proliferation of fibroblasts and biosynthesis of extracellular matrix components.

  12. 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.

  13. Vasodilator-Stimulated Phosphoprotein Activity Is Required for Coxiella burnetii Growth in Human Macrophages.

    Directory of Open Access Journals (Sweden)

    Punsiri M Colonne

    2016-10-01

    Full Text Available Coxiella burnetii is an intracellular bacterial pathogen that causes human Q fever, an acute flu-like illness that can progress to chronic endocarditis and liver and bone infections. Humans are typically infected by aerosol-mediated transmission, and C. burnetii initially targets alveolar macrophages wherein the pathogen replicates in a phagolysosome-like niche known as the parasitophorous vacuole (PV. C. burnetii manipulates host cAMP-dependent protein kinase (PKA signaling to promote PV formation, cell survival, and bacterial replication. In this study, we identified the actin regulatory protein vasodilator-stimulated phosphoprotein (VASP as a PKA substrate that is increasingly phosphorylated at S157 and S239 during C. burnetii infection. Avirulent and virulent C. burnetii triggered increased levels of phosphorylated VASP in macrophage-like THP-1 cells and primary human alveolar macrophages, and this event required the Cα subunit of PKA. VASP phosphorylation also required bacterial protein synthesis and secretion of effector proteins via a type IV secretion system, indicating the pathogen actively triggers prolonged VASP phosphorylation. Optimal PV formation and intracellular bacterial replication required VASP activity, as siRNA-mediated depletion of VASP reduced PV size and bacterial growth. Interestingly, ectopic expression of a phospho-mimetic VASP (S239E mutant protein prevented optimal PV formation, whereas VASP (S157E mutant expression had no effect. VASP (S239E expression also prevented trafficking of bead-containing phagosomes to the PV, indicating proper VASP activity is critical for heterotypic fusion events that control PV expansion in macrophages. Finally, expression of dominant negative VASP (S157A in C. burnetii-infected cells impaired PV formation, confirming importance of the protein for proper infection. This study provides the first evidence of VASP manipulation by an intravacuolar bacterial pathogen via activation of PKA

  14. A method for comparison of animal and human alveolar dose and toxic effect of inhaled ozone

    International Nuclear Information System (INIS)

    Hatch, G.E.; Koren, H.; Aissa, M.

    1989-01-01

    Present models for predicting the pulmonary toxicity of O 3 in humans from the toxic effects observed in animals rely on dosimetric measurements of O 3 mass balance and species comparisons of mechanisms that protect tissue against O 3 . The goal of the study described was to identify a method to directly compare O 3 dose and effect in animals and humans using bronchoalveolar lavage fluid markers. The feasibility of estimating O 3 dose to alveoli of animals and humans was demonstrated through assay of reaction products of 18 O-labeled O 3 in lung surfactant and macrophage pellets of rabbits. The feasibility of using lung lavage fluid protein measurements to quantify the O 3 toxic response in humans was demonstrated by the finding of significantly increased lung lavage protein in 10 subjects exposed to 0.4 ppm O 3 for 2 h with intermittent periods of heavy exercise. The validity of using the lavage protein marker to quantify the response in animals has already been established. The positive results obtained in both the 18 O 3 and the lavage protein studies reported here suggest that it should be possible to obtain a direct comparison of both alveolar dose and toxic effect of O 3 to alveoli of animals or humans

  15. Human Induced Pluripotent Stem Cell-Derived Macrophages for Unraveling Human Macrophage Biology.

    Science.gov (United States)

    Zhang, Hanrui; Reilly, Muredach P

    2017-11-01

    Despite a substantial appreciation for the critical role of macrophages in cardiometabolic diseases, understanding of human macrophage biology has been hampered by the lack of reliable and scalable models for cellular and genetic studies. Human induced pluripotent stem cell (iPSC)-derived macrophages (IPSDM), as an unlimited source of subject genotype-specific cells, will undoubtedly play an important role in advancing our understanding of the role of macrophages in human diseases. In this review, we summarize current literature in the differentiation and characterization of IPSDM at phenotypic, functional, and transcriptomic levels. We emphasize the progress in differentiating iPSC to tissue resident macrophages, and in understanding the ontogeny of in vitro differentiated IPSDM that resembles primitive hematopoiesis, rather than adult definitive hematopoiesis. We review the application of IPSDM in modeling both Mendelian genetic disorders and host-pathogen interactions. Finally, we highlighted the potential areas of research using IPSDM in functional validation of coronary artery disease loci in genome-wide association studies, functional genomic analyses, drug testing, and cell therapeutics in cardiovascular diseases. © 2017 American Heart Association, Inc.

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

    Science.gov (United States)

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

    2018-02-05

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

  17. Formulation and Characterization of Pyrazinamide Polymeric Nanoparticles for Pulmonary Tuberculosis: Efficiency for Alveolar Macrophage Targeting

    OpenAIRE

    Varma, J. N. Ravi; Kumar, T. Santosh; Prasanthi, B.; Ratna, J. Vijaya

    2015-01-01

    Pyrazinamide, a highly specific agent against Mycobacterium tuberculosis is used as first-line drug to treat tuberculosis. The current work aims to formulate polymeric nanoparticles based drug delivery system to sustain the release profile and reduce the dosing frequency of pyrazinamide. Further aim was to target the macrophages within body fluid. These polymeric nanoparticles were prepared by simultaneous double-emulsion (W/O/W) solvent evaporation/diffusion technique. The prepared dispersio...

  18. Depletion of Alveolar Macrophages Does Not Prevent Hantavirus Disease Pathogenesis in Golden Syrian Hamsters

    Science.gov (United States)

    2016-05-20

    ANDV strain Chile -9717869 (27) was propagated in Vero E6 cells 122 (Vero C1008, ATCC CRL 1586). Preparation of twice-plaque-purified ANDV stock has...Research and Material Command, Military 537 Infectious Disease Research Program , Program Area T. Research reported in this publication 538 was also...prior to kidney, involvement, and diagnosed by viral 684 inclusions in lung macrophages. European journal of clinical microbiology & infectious

  19. Importance of the functional state of alveolar macrophages of the lungs for hygienic evaluation of protective reactions and cell damage due to atmospheric pollution

    Energy Technology Data Exchange (ETDEWEB)

    Tusl, M; Vyskocil, A; Duerrer, I; Aulika, B V; Litvinov, N N; Merkur' eva, R V

    1983-01-01

    Total number of cells, their viability and ability to adhesion were examined in surface alveolar macrophages isolated from rat livers after exposure to sulphur dioxide during 2, 4 and 6 weeks (0.05, 0.5, 1.0 and 5.0 mg/m3); to nitrogen oxide during 5, 8 and 15 hours, 28 and 56 days (19 mg/m3) and to carbon monoxide during 2, 28 and 56 days (0.01% or 10 MAC). In the experiment with exposure to sulphur dioxide, the activity of enzymes of varying localization in the macrophages - soluble in the cytoplasm (lactate dehydrogenase) and connected with subcellular structures - lysosomes (beta-galactosidase, beta-glucosidase and acid phosphatase) was tested by means of biochemical methods in parallel with cytological examinations. Low concentrations of various chemical contaminants of the atmospheric air (sulphur dioxide, nitrogen oxides, carbon monoxide) have an unfavourable biological effect on rats, manifest in the impairment of local immunity, i.e., decreased number of alveolar macrophages, disturbance of their viability and reduced ability of the macrophages to adhesion. At the same time, sulphur dioxide induces enzyme disorganization in lactate dehydrogenase and in a number of lysosomal enzymes of the macrophages. These results serve as a basis for the recommendation of cytobiochemical methods of elaborating methodological approaches to the regulation of environmental factors. Alveolar macrophages as a constituent part of the mononuclear phagocytic system ensuring local non-specific and specific resistance of the organism form one of the most important cellular mechanisms of protection of the organism against the harmful effect of environmental factors including chemical contaminants of the atmospheric air (1, 2).

  20. Platelet CLEC-2 protects against lung injury via effects of its ligand podoplanin on inflammatory alveolar macrophages in the mouse.

    Science.gov (United States)

    Lax, Siân; Rayes, Julie; Wichaiyo, Surasak; Haining, Elizabeth J; Lowe, Kate; Grygielska, Beata; Laloo, Ryan; Flodby, Per; Borok, Zea; Crandall, Edward D; Thickett, David R; Watson, Steve P

    2017-12-01

    There is no therapeutic intervention proven to prevent acute respiratory distress syndrome (ARDS). Novel mechanistic insights into the pathophysiology of ARDS are therefore required. Platelets are implicated in regulating many of the pathogenic processes that occur during ARDS; however, the mechanisms remain elusive. The platelet receptor CLEC-2 has been shown to regulate vascular integrity at sites of acute inflammation. Therefore the purpose of this study was to establish the role of CLEC-2 and its ligand podoplanin in a mouse model of ARDS. Platelet-specific CLEC-2-deficient, as well as alveolar epithelial type I cell (AECI)-specific or hematopoietic-specific podoplanin deficient, mice were established using cre-loxP strategies. Combining these with intratracheal (IT) instillations of lipopolysaccharide (LPS), we demonstrate that arterial oxygen saturation decline in response to IT-LPS in platelet-specific CLEC-2-deficient mice is significantly augmented. An increase in bronchoalveolar lavage (BAL) neutrophils and protein was also observed 48 h post-IT-LPS, with significant increases in pro-inflammatory chemokines detected in BAL of platelet-specific CLEC-2-deficient animals. Deletion of podoplanin from hematopoietic cells but not AECIs also reduces lung function and increases pro-inflammatory chemokine expression following IT-LPS. Furthermore, we demonstrate that following IT-LPS, platelets are present in BAL in aggregates with neutrophils, which allows for CLEC-2 interaction with podoplanin expressed on BAL inflammatory alveolar macrophages. Taken together, these data suggest that the platelet CLEC-2-podoplanin signaling axis regulates the severity of lung inflammation in mice and is a possible novel target for therapeutic intervention in patients at risk of developing ARDS. Copyright © 2017 the American Physiological Society.

  1. An interspecies comparison of the phagocytosis and dissolution of 241AmO2 particles by rat, dog and monkey alveolar macrophages in vitro

    International Nuclear Information System (INIS)

    Taya, A.; Carmack, D.B.; Muggenburg, B.A.; Mewhinney, J.A.

    1992-01-01

    Experiments were conducted to study the phagocytosis and dissolution of 241 AmO 2 particles by rat, dog and monkey alveolar macrophages (PAM) in vitro. The phagocytosis and dissolution of 241 AmO 2 particles were followed up to 20 and 72 h, respectively. Dog and monkey PAM took up 241 AmO 2 particles at similar rates, whereas rat PAM phagocytosed only 60% of the amount phagocytosed by dog and monkey PAM at 20h. The PAM of the three species dissolved 241 AmO 2 particles at similar rates; 8-10% was dissolved by 72h. The results of the 241 AmO 2 uptake in vitro may reflect in vivo situations, where the differences in uptake seen in vitro would probably diminish at later times after exposure. The dissolution results imply that the dissolution of 241 AmO 2 particles by alveolar macrophages of the three species might be species-independent. (author)

  2. Study on alveolar macrophage injure caused by uranium dust and its protection

    International Nuclear Information System (INIS)

    Zhou Liren; Xu Ying; Li Jianxiang; Zhang Yu; Chen Yuejin; Qiang Yizhong

    1995-10-01

    Dog's alveolar microphage (AM) obtained by lavage was cultured in vitro. The effects of uranium dust, quartz dust on peroxidation of AM and the effects of magnoliavinin C and V E on bio-membrane was observed. In addition the anti-oxidation effect of V E on the whole body was observed by means of experimental silicosis caused by single dust exposure to trachea. The results demonstrate that two kinds of dust all can induce membrane lipid peroxidation, magnoliavinin C and V E have marked anti-oxidation effect. The administration of V E in vivo demonstrates that V E has effect of inhibiting membrane unsaturated fatty acid peroxidation induced by these two kinds of dust in the ears stage of dust exposure and blocking the chain reaction of free radical so as to retard the pathological developing for silicosis. However it's effect is less than the combining effect of V E and phosphohydroxypipe quinoline. (6 tabs., 12 figs.)

  3. Effects of low molecular weight fungal compounds on inflammatory gene transcription and expression in mouse alveolar macrophages.

    Science.gov (United States)

    Rand, Thomas G; Dipenta, J; Robbins, C; Miller, J D

    2011-04-25

    The inflammatory potential and molecular mechanisms underscoring inflammatory responses of lung cells to compounds from fungi that grow on damp building materials is poorly understood in vitro. In this study we evaluated the effect of pure fungal compounds on potentiating acute inflammatory response in primary mouse alveolar macrophages (AMs) and tested the hypothesis that AM responses to low molecular weight fungal compounds exhibit temporal and compound specificity that mimic that observed in the whole lung. Transcriptional responses of 13 inflammation/respiratory burst-associated genes (KC=Cxcl1, Cxcl2, Cxcl5, Cxcl10, Ccl3, Ccl112, Ccl20, IL-1β, Il-6, ifi27 Tnfα, iNOS and Blvrb) were evaluated in mouse AMs exposed to a 1ml (10(-8)mol) dose of either pure atranone C, brevianimide, cladosporin, curdlan, LPS, neoechinulin A & B, sterigmatocystin or TMC-120A for 2h, 4h and 12h PE using customized reverse transcription (RT)-PCR based arrays. Multianalyte ELISA was used to measure expression of 6 pro-inflammatory cytokines common to the transcriptional assays (Cxcl1, Cxcl10, Ccl3, IL1β, Ifn-λ and Tnf-α) to determine whether gene expression corresponded to the transcription data. Compared to controls, all of these compounds induced significant (≥2.5-fold or ≤-2.5-fold change at p≤0.05) time- and compound-specific transcriptional gene alterations in treatment AMs. The highest number of transcribed genes were in LPS treatment AMs at 12h PE (12/13) followed by neoechinulin B at 4h PE (11/13). Highest fold change values (>30) were associated with KC, Cxcl2, Cxcl5 and IL1β genes in cells exposed to LPS. Compound exposures also induced significant (p≤0.05) time- and compound-specific pro-inflammatory responses manifest as differentially elevated Cxcl1, Cxcl10, Ccl3, Ifn-λ and Tnf-α concentrations in culture supernatant of treatment AMs. Dissimilarity in transcriptional responses in AMs and our in vivo model of lung disease is likely attributable to whole lung

  4. Evaluation of the concentration of marbofloxacin in alveolar macrophages and pulmonary epithelial lining fluid after administration in dogs.

    Science.gov (United States)

    Boothe, Harry W; Jones, Sarah A; Wilkie, W Scott; Boeckh, Albert; Stenstrom, Kristol K; Boothe, Dawn M

    2005-10-01

    To determine concentrations of marbofloxacin in alveolar macrophages (AMs) and epithelial lining fluid (ELF) and compare those concentrations with plasma concentrations in healthy dogs. 12 adult mixed-breed and purebred hounds. 10 dogs received orally administered marbofloxacin at a dosage of 2.75 mg/kg every 24 hours for 5 days. Two dogs served as nontreated controls. Fiberoptic bronchoscopy and bronchoalveolar lavage procedures were performed while dogs were anesthetized with propofol, approximately 6 hours after the fifth dose. The concentrations of marbofloxacin in plasma and bronchoalveolar fluid (cell and supernatant fractions) were determined by use of high-performance liquid chromatography with detection of fluorescence. Mean +/- SD plasma marbofloxacin concentrations 2 and 6 hours after the fifth dose were 2.36 +/- 0.52 microg/mL and 1.81 +/- 0.21 microg/mL, respectively. Mean +/- SD marbofloxacin concentration 6 hours after the fifth dose in AMs (37.43 +/- 24.61 microg/mL) was significantly greater than that in plasma (1.81 +/- 0.21 microg/mL) and ELF (0.82 +/- 0.34 microg/mL), resulting in a mean AM concentration-to-plasma concentration ratio of 20.4, a mean AM:ELF ratio of 60.8, and a mean ELF-to-plasma ratio of 0.46. Marbofloxacin was not detected in any samples from control dogs. Marbofloxacin concentrations in AMs were greater than the mean inhibitory concentrations of major bacterial pathogens in dogs. Results indicated that marbofloxacin accumulates in AMs at concentrations exceeding those reached in plasma and ELF The accumulation of marbofloxacin in AMs may facilitate treatment for susceptible intracellular pathogens or infections associated with pulmonary macrophage infiltration.

  5. Sonicated Protein Fractions of Mycoplasma hyopneumoniae Induce Inflammatory Responses and Differential Gene Expression in a Murine Alveolar Macrophage Cell Line.

    Science.gov (United States)

    Damte, Dereje; Lee, Seung-Jin; Birhanu, Biruk Tesfaye; Suh, Joo-Won; Park, Seung-Chun

    2015-12-28

    Mycoplasma hyopneumoniae is known to cause porcine enzootic pneumonia (EP), an important disease in swine production. The objective of this study was to examine the effects of sonicated protein fractions of M. hyopneumoniae on inflammatory response and gene expression in the murine alveolar macrophage MH-S cell line. The effects of sonicated protein fractions and intact M. hyopneumoniae on the gene expression of cytokines and iNOS were assessed using RT-PCR. The Annealing Control Primer (ACP)-based PCR method was used to screen differentially expressed genes. Increased transcription of interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, COX-2, and iNOS mRNA was observed after exposure to the supernatant (SPT), precipitant (PPT), and intact M. hyopneumoniae protein. A time-dependent analysis of the mRNA expression revealed an upregulation after 4 h for IL-6 and iNOS and after 12 h for IL-1β and TNF-α, for both SPT and PPT; the fold change in COX-2 expression was less. A dose- and time-dependent correlation was observed in nitrite (NO) production for both protein fractions; however, there was no significant difference between the effects of the two protein fractions. In a differential gene analysis, PCR revealed differential expression for nine gene bands after 3 h of stimulation - only one gene was downregulated, while the remaining eight were upregulated. The results of this study provide insights that help improve our understanding of the mechanisms underlying the pathogenesis of and macrophage defenses against M. hyopneumoniae assault, and suggest targets for future studies on therapeutic interventions for M. hyopneumoniae infections.

  6. Suppression and recovery of the alveolar macrophage phagocytic system during continuous exposure to 0. 5 ppm ozone

    Energy Technology Data Exchange (ETDEWEB)

    Gilmour, M.I.; Hmieleski, R.R.; Stafford, E.A.; Jakab, G.J. (Johns Hopkins University, School of Hygiene and Public Health, Baltimore, MD (USA))

    1991-05-01

    Short-term exposures to ozone (O3) are known to impair pulmonary antibacterial defenses and alveolar macrophage (AM) phagocytosis in a dose-related manner. To determine the effect of prolonged O3 exposure, Swiss mice were exposed continuously to 0.5 ppm O3. At 1, 3, 7, and 14 days, intrapulmonary killing was assessed by inhalation challenge with Staphylococcus aureus or Proteus mirabilis and by comparing the number of viable bacteria remaining in the lungs at 4 h between O3-exposed and control animals. To evaluate the effects of O3 on the functional capacity of the AMs, Fc-receptor mediated phagocytosis was assessed. Ozone exposure impaired the intrapulmonary killing of S. aureus at 1 and 3 days; however, with prolonged exposure, the bactericidal capacity of the lungs returned to normal. This trend of an initial suppression followed by recovery was reflected in the phagocytic capacity of the AMs. In contrast to S. aureus, when P. mirabilis was used as the challenge organism, O3 exposure had no suppressive effect on pulmonary bactericidal activity, which correlated with an increase in the phagocytic cell population in the lungs. Morphologic examination of the lavaged macrophages showed that after 1 day of O3 exposure, the AMs were more foamy, and contained significantly more vacuoles. There was also a significant increase in binucleated cells at 3 days. These studies demonstrate that continuous exposure to O3 modulates AM-dependent lung defenses and points to the importance of the challenge organism and exposure protocol in establishing the adverse effect of O3.

  7. Entry of porcine reproductive and respiratory syndrome virus into porcine alveolar macrophages via receptor-mediated endocytosis.

    Science.gov (United States)

    Nauwynck, H J; Duan, X; Favoreel, H W; Van Oostveldt, P; Pensaert, M B

    1999-02-01

    Porcine alveolar macrophages (AMphi) are the dominant cell type that supports the replication of porcine reproductive and respiratory syndrome virus (PRRSV) in vivo and in vitro. In order to determine the characteristics of the virus-receptor interaction, the attachment of PRRSV to cells was examined by using biotinylated virus in a series of flow cytometric assays. PRRSV bound specifically to AMphi in a dose-dependent manner. Binding of PRRSV to AMphi increased gradually and reached a maximum within 60 min at 4 degrees C. By confocal microscopy, it was shown that different degrees of PRRSV binding exist and that entry is by endocytosis. Virus uptake in vesicles is a clathrin-dependent process, as it was blocked by the addition of cytochalasin D and co-localization of PRRSV and clathrin was found. Furthermore, by the use of two weak bases, NH4Cl and chloroquine, it was demonstrated that PRRSV uses a low pH-dependent entry pathway. In the presence of these reagents, input virions accumulated in large vacuoles, indicating that uncoating was prevented. These results indicate that PRRSV entry into AMphi involves attachment to a specific virus receptor(s) followed by a process of endocytosis, by which virions are taken into the cell within vesicles by a clathrin-dependent pathway. A subsequent drop in pH is required for proper virus replication.

  8. Differential response to dexamethasone on the TXB2 release in guinea-pig alveolar macrophages induced by zymosan and cytokines

    Directory of Open Access Journals (Sweden)

    M. E. Salgueiro

    1997-01-01

    Full Text Available Glucocorticosteroids reduce the production of inflammatory mediators but this effect may depend on the stimulus. We have compared the time course of the effect of dexamethasone on the thromboxane B2 (TXB2 release induced by cytokine stimulation and zymosan in guinea-pig alveolar macrophages. Interleukin-1β (IL-1β, tumour necrosis factor-α (TNF-α and opsonized zymosan (OZ, all stimulate TXB2 release. High concentrations of dexamethasone (1–10 μM inhibit the TXB2 production induced by both cytokines and OZ, but the time course of this response is different. Four hours of incubation with dexamethasone reduce the basal TXB2 release and that induced by IL-1β and TNF-α, but do not modify the TXB2 release induced by OZ. However, this stimulus was reduced after 24 h incubation. Our results suggest that the antiinflammatory activity of glucocorticosteroids shows some dependence on stimulus and, therefore, may have more than one mechanism involved.

  9. Phagolysosomal pH and dissolution of cobalt oxide particles by alveolar macrophages

    International Nuclear Information System (INIS)

    Lundborg, M.; Johansson, A.; Camner, P.; Falk, R.; Kreyling, W.

    1992-01-01

    We studied phagolysosomal pH in rabbit macrophages (AM) incubated with 0.-15 μM chloroquine. There was a dose-related increase in pH with chloroquine concentration. Electron microscopy showed that chloroquine increased lysosomal size. In a second experiment we studied dissolution of radiolabeled cobalt oxide particles by rabbit AM, phagolysosomal pH, and lysosomal size. The cells were incubated for 2 days with 0, 2, 5, and 10 μM chloroquine. Size and pH increased with chloroquine concentration. Dissolution of cobalt particles by the AM did not clearly change with pH. In a third experiment, dissolution in acetate buffer was faster than in the AM, and the dissolution appeared to decrease faster with increasing pH than in the AM. A simple model for dissolution of a particle in a phagolysosome was proposed. This model predicts the types of difference in dissolution between AM and buffered saline. 19 refs., 3 figs., 3 tabs

  10. Human Alveolar Echinococcosis in Poland: 1990–2011

    Science.gov (United States)

    Nahorski, Wacław L.; Knap, Józef P.; Pawłowski, Zbigniew S.; Krawczyk, Marek; Polański, Jerzy; Stefaniak, Jerzy; Patkowski, Waldemar; Szostakowska, Beata; Pietkiewicz, Halina; Grzeszczuk, Anna; Felczak-Korzybska, Iwona; Gołąb, Elżbieta; Wnukowska, Natalia; Paul, Małgorzata; Kacprzak, Elżbieta; Sokolewicz-Bobrowska, Elżbieta; Niścigorska-Olsen, Jolanta; Czyrznikowska, Aleksandra; Chomicz, Lidia; Cielecka, Danuta; Myjak, Przemysław

    2013-01-01

    Background Alveolar echinococcosis (AE) caused by Echinococcus multilocularis infections is a dangerous old disease in the Northern Hemisphere. The aim of the paper was to collect and analyze data on human AE in Poland in the last two decades. Methodology/Principal Findings The sources of data were both the cases officially registered and detected by an active field and laboratory surveillance. The cases were verified by clinical, epidemiological, and laboratory criteria. Altogether 121 human cases of AE were detected. Among these 83 (68,6%) cases were classified as confirmed, 16 as probable and 22 as possible. During the two decades a continuous increase in detection rate was noticed. The cases were 6–82 years old at the time of diagnosis (mean - 47.7 years). Sex ratio M/F was 0.86/1.0. The AE was fatal in 23 (19%) patients (mean age at death - 54.1 years). Family agglomeration of AE was found in 4 foci, involving 9 patients. Seventy six of the cases were diagnosed in an advanced stage of disease. In all cases the liver was the primary location of AE. In 30 (24.8%) patients a spread to other organs was observed. Ninety four of the patients were treated with albendazole. In 73 (60%) patients a surgical operation was performed, including 15 liver transplantations. Conclusions/Significance The studies confirmed that AE is an emerging disease in Poland, which is the fourth country in Europe with over 120 cases detected. The results also indicate the need of a wider national programme for implementation of screening in the highest AE risk areas (north-eastern Poland) with an effort to increase the public awareness of the possibility of contracting E. multilocularis, and above all, training of the primary care physicians in the recognition of the risk of AE to allow for an early detection of this dangerous disease. PMID:23301116

  11. Nitrated Fatty Acids Reverse Cigarette Smoke-Induced Alveolar Macrophage Activation and Inhibit Protease Activity via Electrophilic S-Alkylation.

    Science.gov (United States)

    Reddy, Aravind T; Lakshmi, Sowmya P; Muchumarri, Ramamohan R; Reddy, Raju C

    2016-01-01

    Nitrated fatty acids (NFAs), endogenous products of nonenzymatic reactions of NO-derived reactive nitrogen species with unsaturated fatty acids, exhibit substantial anti-inflammatory activities. They are both reversible electrophiles and peroxisome proliferator-activated receptor γ (PPARγ) agonists, but the physiological implications of their electrophilic activity are poorly understood. We tested their effects on inflammatory and emphysema-related biomarkers in alveolar macrophages (AMs) of smoke-exposed mice. NFA (10-nitro-oleic acid or 12-nitrolinoleic acid) treatment downregulated expression and activity of the inflammatory transcription factor NF-κB while upregulating those of PPARγ. It also downregulated production of inflammatory cytokines and chemokines and of the protease cathepsin S (Cat S), a key mediator of emphysematous septal destruction. Cat S downregulation was accompanied by decreased AM elastolytic activity, a major mechanism of septal destruction. NFAs downregulated both Cat S expression and activity in AMs of wild-type mice, but only inhibited its activity in AMs of PPARγ knockout mice, pointing to a PPARγ-independent mechanism of enzyme inhibition. We hypothesized that this mechanism was electrophilic S-alkylation of target Cat S cysteines, and found that NFAs bind directly to Cat S following treatment of intact AMs and, as suggested by in silico modeling and calculation of relevant parameters, elicit S-alkylation of Cys25 when incubated with purified Cat S. These results demonstrate that NFAs' electrophilic activity, in addition to their role as PPARγ agonists, underlies their protective effects in chronic obstructive pulmonary disease (COPD) and support their therapeutic potential in this disease.

  12. Alveolar macrophages have a dual role in a rat model for trimellitic anhydride-induced occupational asthma

    International Nuclear Information System (INIS)

    Valstar, Dingena L.; Schijf, Marcel A.; Nijkamp, Frans P.; Storm, Gert; Arts, Josje H.E.; Kuper, C. Frieke; Bloksma, Nanne; Henricks, Paul A.J.

    2006-01-01

    Occupational exposure to low molecular weight chemicals, like trimellitic anhydride (TMA), can result in occupational asthma. Alveolar macrophages (AMs) are among the first cells to encounter inhaled compounds. These cells can produce many different mediators that have a putative role in asthma. In this study, we examined the role of AMs in lung function and airway inflammation of rats exposed to TMA. Female Brown Norway rats were sensitized by dermal application of TMA or received vehicle alone on days 0 and 7. One day before challenge, rats received intratracheally either empty or clodronate-containing liposomes to deplete the lungs of AMs. On day 21, all rats were challenged by inhalation of TMA in air. Lung function parameters were measured before, during, within 1 h after, and 24 h after challenge. IgE levels and parameters of inflammation and tissue damage were assessed 24 h after challenge. Sensitization with TMA led to decreased lung function parameters during and within 1 h after challenge as compared to non-sensitized rats. AM depletion alleviated the TMA-induced drop in lung function parameters and induced a faster recovery compared to sham-depleted TMA-sensitized rats. It also decreased the levels of serum IgE 24 h after challenge, but did not affect the sensitization-dependent increase in lung lavage fluid IL-6 and tissue TNF-α levels. In contrast, AM depletion augmented the TMA-induced tissue damage and inflammation 24 h after challenge. AMs seem to have a dual role in this model for TMA-induced occupational asthma since they potentiate the immediate TMA-induced decrease in lung function but tended to dampen the TMA-induced inflammatory reaction 24 h later

  13. SOLUBLE COMPONENTS OF UTAH VALLEY PARTICULATE POLLUTION ALTER ALVEOLAR MACROPHAGE FUNCTION IN VIVO AND IN VITRO

    Science.gov (United States)

    Water-soluble extracts of Utah Valley dust (UVD) have been found to cause inflammatory injury of the lung in both humans and rodents. The degree of lung damage found correlated with the metal content in the extracts. In the present study, extracts of a set of UVD PM(10) filters c...

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

    Science.gov (United States)

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

    2016-01-01

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

  15. Mucosa-Associated Lymphoid Tissue Lymphoma Translocation Protein 1 Positively Modulates Matrix Metalloproteinase-9 Production in Alveolar Macrophages upon Toll-Like Receptor 7 Signaling and Influenza Virus Infection

    Directory of Open Access Journals (Sweden)

    Yu-Hsiang Lee

    2017-09-01

    Full Text Available Influenza A virus (IAV infection causes significant morbidity and mortality worldwide. Matrix metalloproteinase-9 (MMP-9 degrades extracellular matrix and is involved in the pathology of influenza. It has been reported that MMP-9 mediates neutrophil migration in IAV infection. Whether alveolar macrophages, the first immune cells that encounter IAV, produce MMP-9, and the mechanism of its regulation have never been investigated. As Toll-like receptor 7 (TLR7 is one of the receptors in innate immune cells that recognize IAV, we used TLR7 agonists and IAV to stimulate alveolar macrophage MH-S cells, primary macrophages, and bone marrow neutrophils. Results showed that MMP-9 expression in macrophages is inducible by TLR7 agonists and IAV, yet, MMP-9 production by neutrophils is not inducible by either one of them. We hypothesized that MMP-9 production in macrophages is mediated through TLR7-NF-κB pathway and used microarray to analyze TLR7 agonist-induced NF-κB-related genes. Mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1, a positive regulator of NF-κB, is amongst the top highly induced genes. By use of MALT1 inhibitor (z-VRPR-fmk and alveolar macrophages from MALT1-deficient mice, we found that MMP-9 production is MALT1-dependent. While MALT1 can act as a paracaspase in lymphocytes through degrading various signaling proteins, we discovered that MALT1 functions to reduce a negative regulator of NF-κB, cylindromatosis (CYLD, in alveolar macrophages. IAV-induced MMP-9, TNF, and IL-6 in lungs of MALT1-deficient mice are significantly lower than in wild-type mice after intratracheal infection. MALT1-deficient mice also have less body weight loss and longer survival after infection. Taken together, we demonstrated a novel role of MALT1 in regulating alveolar macrophage MMP-9 production whose presence exacerbates the severity of influenza.

  16. Preincubation of macrophages alveolar of rate with vitamin C or E attenuate the damage to the plasmatic membrane caused by exhibition to ozone

    International Nuclear Information System (INIS)

    Garcia, J.

    2001-01-01

    The damaging effects of a 60 minute ozone exposure (0.594 ppm) on the cell membrane of rat alveolar macrophages was assessed by measuring specific release of 51 Cr label from the cells. Preincubation of the macrophages in the presence of vitamin C (sodium ascorbate) or vitamine E (DL α tocoferol) prior the ozone exposure significantly diminished 51 Cr release. The protective effect of vitamin E was dose dependent. A proposal accounting for the protective effect of vitamins E and C on the cell membrane is presented, and our findings are discussed in relation to recent reports showing that antioxidant supplementation contributes to preserve pulmonary function in ozone-exposed normal and asthmatic volunteers. (Author) [es

  17. Binding of radiolabeled asbestos fibers to guinea pig (gp) alveolar macrophages (AM)

    International Nuclear Information System (INIS)

    Giannotti, M.A.; Tewson, T.J.; Francsechini, M.P.; Scheule, R.K.; Holian, A.

    1990-01-01

    The mechanism by which fibrogenic particulates cause pulmonary fibrosis in humans is not understood, but is likely to involve the AM. Using two fibrogenic particulates, namely, chrysotile (CHR) and crocidolite (CRO) asbestos and gpAM as components of an in vitro model system, the authors have shown that CHR stimulates the gpAM to release superoxide anion, but CRO does not. To examine whether this difference in stimulatory abilities is a result of differences in cell-asbestos binding they have developed an efficient procedure that radiolabels asbestos fibers while retaining their bioactivity. The fibers are labeled with 68 Ge. The 68 Ge decays into 68 Ga, which then can be detected by its characteristic position emission. Both CHR and CRO asbestos were radiolabled successfully. Mild reaction conditions and short reaction times were found under which >90% of the added 68 Ge and 68 Ga bound to the fibers. The radiolabel was retained even after washing the fibers extensively with physiologic buffers. A density gradient procedure was developed to quantitate the binding of asbestos to gpAM in suspension. The binding of both fibers increased with time over one hr. Thus, these results indicate that although both CHR and CRO interact with the gpAM, only CHR interacts productively to stimulate superoxide anion release

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

    Directory of Open Access Journals (Sweden)

    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.

  19. Salmonella Typhimurium induces SPI-1 and SPI-2 regulated and strain dependent downregulation of MHC II expression on porcine alveolar macrophages

    Directory of Open Access Journals (Sweden)

    Van Parys Alexander

    2012-06-01

    Full Text Available Abstract Foodborne salmonellosis is one of the most important bacterial zoonotic diseases worldwide. Salmonella Typhimurium is the serovar most frequently isolated from persistently infected slaughter pigs in Europe. Circumvention of the host’s immune system by Salmonella might contribute to persistent infection of pigs. In the present study, we found that Salmonella Typhimurium strain 112910a specifically downregulated MHC II, but not MHC I, expression on porcine alveolar macrophages in a Salmonella pathogenicity island (SPI-1 and SPI-2 dependent way. Salmonella induced downregulation of MHC II expression and intracellular proliferation of Salmonella in macrophages were significantly impaired after opsonization with Salmonella specific antibodies prior to inoculation. Furthermore, the capacity to downregulate MHC II expression on macrophages differed significantly among Salmonella strains, independently of strain specific differences in invasion capacity, Salmonella induced cytotoxicity and altered macrophage activation status. The fact that strain specific differences in MHC II downregulation did not correlate with the extent of in vitro SPI-1 or SPI-2 gene expression indicates that other factors are involved in MHC II downregulation as well. Since Salmonella strain dependent interference with the pig’s immune response through downregulation of MHC II expression might indicate that certain Salmonella strains are more likely to escape serological detection, our findings are of major interest for Salmonella monitoring programs primarily based on serology.

  20. Changes in the rat lung after exposure to radon and its progeny: Effects on incorporation of bromodeoxyuridine in epithelial cells and on the incidence of nuclear aberrations in Alveolar macrophages

    International Nuclear Information System (INIS)

    Taya, A.; Morgan, A.; Baker, S.T.; Humphreys, J.A.H.; Collier, C.G.; Bisson, M.

    1994-01-01

    The aim of this study was to investigate some responses of cells in the rat respiratory tract as a function of time after inhalation exposure to various levels of radon and its progeny. Rats were exposed to a constant concentration of radon and its progeny to give cumulative exposure levels of 120, 225, 440 and 990 working level months (WLM). An additional unexposed group of rats served as controls. The end points selected for investigation were (a) the incorporation of bromodeoxyuridine (BrdU) in epithelial cells of the conducting airways and of the alveolar region of the respiratory tract and (b) the incidence of alveolar macrophages with nuclear aberrations. After exposure, the incidence of epithelial cells incorporating BrdU-the labeling index-increased in all regions of the respiratory tract examined, but the increase occurred later in alveolar than in airway epithelial cells. The highest labeling index was found in bronchial epithelial cells, which probably received the highest radiation dose. After an initial induction period, the incidence of alveolar macrophages with nuclear aberrations also increased. The possibility of using the labeling index of alveolar and airway epithelial cells, and/or the incidence of nuclear aberrations in alveolar macrophages, to estimate the radiation dose to various regions of the respiratory tract after exposure of rats to radon and its progeny is discussed. 22 refs., 3 figs., 1 tab

  1. Mycoplasma hyopneumoniae-derived lipid-associated membrane proteins induce apoptosis in porcine alveolar macrophage via increasing nitric oxide production, oxidative stress, and caspase-3 activation.

    Science.gov (United States)

    Bai, Fangfang; Ni, Bo; Liu, Maojun; Feng, Zhixin; Xiong, Qiyan; Xiao, Shaobo; Shao, Guoqing

    2013-09-15

    Mycoplasma hyopneumoniae is the primary etiological agent of enzootic pneumonia in swine. Lipid-associated membrane proteins (LAMP) of mycoplasma are the main pathogenicity factors in mycoplasma diseases. In this study, we investigated the effects of M. hyopneumoniae LAMP on porcine alveolar macrophage (PAM) 3D4/21 cell line. Apoptotic features, such as chromatin condensation and apoptotic bodies, were observed in LAMP-treated PAM 3D4/21 cells. Moreover, LAMP significantly increased the number of TUNEL positive apoptotic cells in PAM 3D4/21 cells compared with the untreated control. In addition, flow cytometric analysis using dual staining with annexin-V-FITC and propidium iodide (PI) showed that LAMP of M. hyopneumoniae induced a time-dependent apoptosis in PAM 3D4/21 cells. Moreover, increased levels of superoxide anion production and activated caspase-3 in PAM 3D4/21 cells were observed after exposure to LAMP. Increased production of nitric oxide (NO) was also confirmed in the cell supernatants. Besides, apoptotic rates increase and caspase-3 activation were suppressed by NOS inhibitor or antioxidant. It is suggested that LAMP of M. hyopneumoniae induced apoptosis in porcine alveolar macrophage via NO production, superoxide anion production, and caspase-3 activation. Copyright © 2013 Elsevier B.V. All rights reserved.

  2. Rhinovirus infection induces distinct transcriptome profiles in polarized human macrophages.

    Science.gov (United States)

    Rajput, Charu; Walsh, Megan P; Eder, Breanna N; Metitiri, Ediri E; Popova, Antonia P; Hershenson, Marc B

    2018-05-01

    Infections with rhinovirus (RV) cause asthma exacerbations. Recent studies suggest that macrophages play a role in asthmatic airway inflammation and the innate immune response to RV infection. Macrophages exhibit phenotypes based on surface markers and gene expression. We hypothesized that macrophage polarization state alters gene expression in response to RV infection. Cells were derived from human peripheral blood derived monocytes. M1 and M2 polarization was carried out by using IFN-γ and IL-4, respectively, and RNA was extracted for Affymetrix Human Gene ST2.1 exon arrays. Selected genes were validated by quantitative (q)PCR. Treatment of nonactivated (M0) macrophages with IFN-γ and IL-4 induced the expression of 252 and 153 distinct genes, respectively, including previously-identified M1 and M2 markers. RV infection of M0 macrophages induced upregulation of 232 genes; pathway analysis showed significant overrepresentation of genes involved in IFN-α/β signaling and cytokine signaling in the immune system. RV infection induced differential expression of 195 distinct genes in M1-like macrophages but only seven distinct genes in M2-like-polarized cells. In a secondary analysis, comparison between M0-, RV-infected, and M1-like-polarized, RV-infected macrophages revealed differential expression of 227 genes including those associated with asthma and its exacerbation. qPCR demonstrated increased expression of CCL8, CXCL10, TNFSF10, TNFSF18, IL6, NOD2, and GSDMD and reduced expression of VNN1, AGO1, and AGO2. Together, these data show that, in contrast to M2-like-polarized macrophages, gene expression of M1-like macrophages is highly regulated by RV.

  3. Virulent and avirulent strains of equine arteritis virus induce different quantities of TNF-α and other proinflammatory cytokines in alveolar and blood-derived equine macrophages

    International Nuclear Information System (INIS)

    Moore, Brian D.; Balasuriya, Udeni B.R.; Watson, Johanna L.; Bosio, Catharine M.; MacKay, Robert J.; MacLachlan, N. James

    2003-01-01

    Equine arteritis virus (EAV) infects endothelial cells (ECs) and macrophages in horses, and many of the clinical manifestations of equine viral arteritis (EVA) reflect vascular injury. To further evaluate the potential role of EAV-induced, macrophage-derived cytokines in the pathogenesis of EVA, we infected cultured equine alveolar macrophages (AMphi), blood monocyte-derived macrophages (BMphi), and pulmonary artery ECs with either a virulent (KY84) or an avirulent (CA95) strain of EAV. EAV infection of equine AMphi, BMphi, and ECs resulted in their activation with increased transcription of genes encoding proinflammatory mediators, including interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor (TNF)-α. Furthermore, the virulent KY84 strain of EAV induced significantly higher levels of mRNA encoding proinflammatory cytokines in infected AMphi and BMphi than did the avirulent CA95 strain. Treatment of equine ECs with the culture supernatants of EAV-infected AMphi and BMphi also resulted in EC activation with cell surface expression of E-selectin, whereas infection of ECs with purified EAV alone caused only minimal expression of E-selectin. The presence of TNF-α in the culture supernatants of EAV-infected equine AMphi, BMphi, and ECs was confirmed by bioassay, and the virulent KY84 strain of EAV induced significantly more TNF-α in all cell types than did the avirulent CA95 strain. Thus, the data indicate that EAV-induced, macrophage-derived cytokines may contribute to the pathogenesis of EVA in horses, and that the magnitude of the cytokine response of equine AMphi, BMphi, and ECs to EAV infection reflects the virulence of the infecting virus strain

  4. Modulation of human macrophage activity by Ascaris antigens is dependent on macrophage polarization state

    DEFF Research Database (Denmark)

    Almeida, Sara; Nejsum, Peter; Williams, Andrew R.

    2018-01-01

    Parasitic worms (helminths) are known to actively modulate host immune responses and inflammation. The aim of this study was to investigate if adult body fluid (ABF) from the helminth Ascaris suum has immunomodulatory effects on different subtypes of human monocyte-derived macrophages (Mɸ) in vitro...

  5. 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

  6. Transcriptomic profiling of primary alveolar epithelial cell differentiation in human and rat

    Directory of Open Access Journals (Sweden)

    Crystal N. Marconett

    2014-12-01

    Full Text Available Cell-type specific gene regulation is a key to gaining a full understanding of how the distinct phenotypes of differentiated cells are achieved and maintained. Here we examined how changes in transcriptional activation during alveolar epithelial cell (AEC differentiation determine phenotype. We performed transcriptomic profiling using in vitro differentiation of human and rat primary AEC. This model recapitulates in vitro an in vivo process in which AEC transition from alveolar type 2 (AT2 cells to alveolar type 1 (AT1 cells during normal maintenance and regeneration following lung injury. Here we describe in detail the quality control, preprocessing, and normalization of microarray data presented within the associated study (Marconett et al., 2013. We also include R code for reproducibility of the referenced data and easily accessible processed data tables.

  7. Innate immune response of alveolar macrophage to house dust mite allergen is mediated through TLR2/-4 co-activation.

    Directory of Open Access Journals (Sweden)

    Chia-Fang Liu

    Full Text Available House dust mite, Dermatophagoides pteronyssinus (Der p, is one of the major allergens responsible for allergic asthma. However, the putative receptors involved in the signalization of Der p to the innate immune cells are still poorly defined as well as the impact of their activation on the outcome of the allergen-induced cell response. We previously reported that the HDM activation of mouse alveolar macrophages (AM involves the TLR4/CD14 cell surface receptor complex. Here using a TLR ligand screening essay, we demonstrate that HDM protein extract engages the TLR2, in addition to the TLR4, in engineered TLR-transfected HEK cells but also in the MH-S mouse alveolar macrophage cell line model. Moreover we found that the concomitant recruitment of the MH-S cell's TLR2 and TLR4 receptors by the HDM extract activates the MyD88-dependent signaling pathway and leads to the secretion of the NF-κB regulated pro-inflammatory factors NO and TNF-α. However unlike with the canonical TLR4 ligand (i.e. the bacterial LPS mobilization of TLR4 by the HDM extract induces a reduced production of the IL-12 pro-inflammatory cytokine and fails to trigger the expression of the T-bet transcription factor. Finally we demonstrated that HDM extract down-regulates LPS induced IL-12 and T-bet expression through a TLR2 dependent mechanism. Therefore, we propose that the simultaneous engagement of the TLR2 and TLR4 receptors by the HDM extract results in a cross regulated original activation pattern of the AM which may contribute to the Th2 polarization of the allergen-induced immune response. The deciphering of these cross-regulation networks is of prime importance to open the way for original therapeutic strategies taking advantage of these receptors and their associated signaling pathways to treat allergic asthma.

  8. Moderate restriction of macrophage-tropic human immunodeficiency virus type 1 by SAMHD1 in monocyte-derived macrophages.

    Science.gov (United States)

    Taya, Kahoru; Nakayama, Emi E; Shioda, Tatsuo

    2014-01-01

    Macrophage-tropic human immunodeficiency virus type 1 (HIV-1) strains are able to grow to high titers in human monocyte-derived macrophages. However, it was recently reported that cellular protein SAMHD1 restricts HIV-1 replication in human cells of the myeloid lineage, including monocyte-derived macrophages. Here we show that degradation of SAMHD1 in monocyte-derived macrophages was associated with moderately enhanced growth of the macrophage-tropic HIV-1 strain. SAMHD1 degradation was induced by treating target macrophages with vesicular stomatitis virus glycoprotein-pseudotyped human immunodeficiency virus type 2 (HIV-2) particles containing viral protein X. For undifferentiated monocytes, HIV-2 particle treatment allowed undifferentiated monocytes to be fully permissive for productive infection by the macrophage-tropic HIV-1 strain. In contrast, untreated monocytes were totally resistant to HIV-1 replication. These results indicated that SAMHD1 moderately restricts even a macrophage-tropic HIV-1 strain in monocyte-derived macrophages, whereas the protein potently restricts HIV-1 replication in undifferentiated monocytes.

  9. MyD88 mediates in vivo effector functions of alveolar macrophages in acute lung inflammatory responses to carbon nanotube exposure

    Energy Technology Data Exchange (ETDEWEB)

    Frank, Evan A. [Division of Environmental Genetics and Molecular Toxicology, Department of Environmental Health, University of Cincinnati College of Medicine, Cincinnati, OH 45267 (United States); Birch, M. Eileen [National Institute for Occupational Safety and Health, Cincinnati, OH 45213 (United States); Yadav, Jagjit S., E-mail: Jagjit.Yadav@uc.edu [Division of Environmental Genetics and Molecular Toxicology, Department of Environmental Health, University of Cincinnati College of Medicine, Cincinnati, OH 45267 (United States)

    2015-11-01

    Carbon nanotubes (CNTs) are rapidly emerging as high-priority occupational toxicants. CNT powders contain fibrous particles that aerosolize readily in places of manufacture and handling, posing an inhalation risk for workers. Studies using animal models indicate that lung exposure to CNTs causes prolonged inflammatory responses and diffuse alveolar injury. The mechanisms governing CNT-induced lung inflammation are not fully understood but have been suggested to involve alveolar macrophages (AMs). In the current study, we sought to systematically assess the effector role of AMs in vivo in the induction of lung inflammatory responses to CNT exposures and investigate their cell type-specific mechanisms. Multi-wall CNTs characterized for various physicochemical attributes were used as the CNT type. Using an AM-specific depletion and repopulation approach in a mouse model, we unambiguously demonstrated that AMs are major effector cells necessary for the in vivo elaboration of CNT-induced lung inflammation. We further investigated in vitro AM responses and identified molecular targets which proved critical to pro-inflammatory responses in this model, namely MyD88 as well as MAPKs and Ca{sup 2} {sup +}/CamKII. We further demonstrated that MyD88 inhibition in donor AMs abrogated their capacity to reconstitute CNT-induced inflammation when adoptively transferred into AM-depleted mice. Taken together, this is the first in vivo demonstration that AMs act as critical effector cell types in CNT-induced lung inflammation and that MyD88 is required for this in vivo effector function. AMs and their cell type-specific mechanisms may therefore represent potential targets for future therapeutic intervention of CNT-related lung injury. - Highlights: • Demonstrated in vivo effector role of alveolar macrophages (AMs) in CNT toxicity • MyD88, MAPKs, and Ca{sup 2} {sup +}/CamKII are required for AM inflammatory responses in vitro. • MyD88 signaling is required for in vivo effector

  10. In-Depth Global Analysis of Transcript Abundance Levels in Porcine Alveolar Macrophages Following Infection with Porcine Reproductive and Respiratory Syndrome Virus

    Directory of Open Access Journals (Sweden)

    Laura C. Miller

    2010-01-01

    Full Text Available Porcine reproductive and respiratory syndrome virus (PRRSV is a major pathogen of swine worldwide and causes considerable economic loss. Identifying specific cell signaling or activation pathways that associate with variation in PRRSV replication and macrophage function may lead to identification of novel gene targets for the control of PRRSV infection. Serial Analysis of Gene Expression (SAGE was used to create and survey the transcriptome of in vitro mock-infected and PRRSV strain VR-2332-infected porcine alveolar macrophages (PAM at 0, 6, 12, 16, and 24 hours after infection. The transcriptome data indicated changes in transcript abundance occurring in PRRSV-infected PAMs over time after infection with more than 590 unique tags with significantly altered transcript abundance levels identified (P<.01. Strikingly, innate immune genes (whose transcript abundances are typically altered in response to other pathogens or insults including IL-8, CCL4, and IL-1β showed no or very little change at any time point following infection.

  11. Ionic channels and membrane hyperpolarization in human macrophages

    NARCIS (Netherlands)

    Ince, C.; van Duijn, B.; Ypey, D. L.; van Bavel, E.; Weidema, F.; Leijh, P. C.

    1987-01-01

    Microelectrode impalement of human macrophages evokes a transient hyperpolarizing response (HR) of the membrane potential. This HR was found to be dependent on the extracellular concentration of K+ but not on that of Na+ or Cl-. It was not influenced by low temperature (12 degrees C) or by 0.2 mM

  12. Cigarette smoke regulates the expression of TLR4 and IL-8 production by human macrophages

    Directory of Open Access Journals (Sweden)

    Rahman Irfan

    2009-05-01

    Full Text Available Abstract Background Toll-like receptors (TLRs are present on monocytes and alveolar macrophages that form the first line of defense against inhaled particles. The importance of those cells in the pathophysiology of chronic obstructive pulmonary disease (COPD has well been documented. Cigarette smoke contains high concentration of oxidants which can stimulate immune cells to produce reactive oxygen species, cytokines and chemokines. Methods In this study, we evaluated the effects of cigarette smoke medium (CSM on TLR4 expression and interleukin (IL-8 production by human macrophages investigating the involvement of ROS. Results and Discussion TLR4 surface expression was downregulated on short term exposure (1 h of CSM. The downregulation could be explained by internalization of the TLR4 and the upregulation by an increase in TLR4 mRNA. IL-8 mRNA and protein were also increased by CSM. CSM stimulation increased intracellular ROS-production and decreased glutathione (GSH levels. The modulation of TLR4 mRNA and surface receptors expression, IRAK activation, IκB-α degradation, IL-8 mRNA and protein, GSH depletion and ROS production were all prevented by antioxidants such as N-acetyl-L-cysteine (NAC. Conclusion TLR4 may be involved in the pathogenesis of lung emphysema and oxidative stress and seems to be a crucial contributor in lung inflammation.

  13. Angiogenic potential of human macrophages on electrospun bioresorbable vascular grafts

    Energy Technology Data Exchange (ETDEWEB)

    Garg, K; Sell, S A; Madurantakam, P; Bowlin, G L, E-mail: glbowlin@vcu.ed [Virginia Commonwealth University, Richmond, VA 23284 (United States)

    2009-06-15

    The aim of this study was to investigate macrophage interactions with electrospun scaffolds and quantify the expression of key angiogenic growth factors in vitro. This study will further help in evaluating the potential of these electrospun constructs as vascular grafts for tissue repair and regeneration in situ. Human peripheral blood macrophages were seeded in serum free media on electrospun (10 mm) discs of polydioxanone (PDO), elastin and PDO:elastin blends (50:50, 70:30 and 90:10). The growth factor secretion was analyzed by ELISA. Macrophages produced high levels of vascular endothelial growth factor and acidic fibroblast growth factor. Transforming growth factor beta-1 (TGF-beta1) secretion was relatively low and there was negligible production of basic fibroblast growth factor. Therefore, it can be anticipated that these scaffolds will support tissue regeneration and angiogenesis. (communication)

  14. Dopamine receptor activation increases HIV entry into primary human macrophages.

    Directory of Open Access Journals (Sweden)

    Peter J Gaskill

    Full Text Available Macrophages are the primary cell type infected with HIV in the central nervous system, and infection of these cells is a major component in the development of neuropathogenesis and HIV-associated neurocognitive disorders. Within the brains of drug abusers, macrophages are exposed to increased levels of dopamine, a neurotransmitter that mediates the addictive and reinforcing effects of drugs of abuse such as cocaine and methamphetamine. In this study we examined the effects of dopamine on HIV entry into primary human macrophages. Exposure to dopamine during infection increased the entry of R5 tropic HIV into macrophages, irrespective of the concentration of the viral inoculum. The entry pathway affected was CCR5 dependent, as antagonizing CCR5 with the small molecule inhibitor TAK779 completely blocked entry. The effect was dose-dependent and had a steep threshold, only occurring above 108 M dopamine. The dopamine-mediated increase in entry required dopamine receptor activation, as it was abrogated by the pan-dopamine receptor antagonist flupenthixol, and could be mediated through both subtypes of dopamine receptors. These findings indicate that the effects of dopamine on macrophages may have a significant impact on HIV pathogenesis. They also suggest that drug-induced increases in CNS dopamine may be a common mechanism by which drugs of abuse with distinct modes of action exacerbate neuroinflammation and contribute to HIV-associated neurocognitive disorders in infected drug abusers.

  15. Dopamine Receptor Activation Increases HIV Entry into Primary Human Macrophages

    Science.gov (United States)

    Gaskill, Peter J.; Yano, Hideaki H.; Kalpana, Ganjam V.; Javitch, Jonathan A.; Berman, Joan W.

    2014-01-01

    Macrophages are the primary cell type infected with HIV in the central nervous system, and infection of these cells is a major component in the development of neuropathogenesis and HIV-associated neurocognitive disorders. Within the brains of drug abusers, macrophages are exposed to increased levels of dopamine, a neurotransmitter that mediates the addictive and reinforcing effects of drugs of abuse such as cocaine and methamphetamine. In this study we examined the effects of dopamine on HIV entry into primary human macrophages. Exposure to dopamine during infection increased the entry of R5 tropic HIV into macrophages, irrespective of the concentration of the viral inoculum. The entry pathway affected was CCR5 dependent, as antagonizing CCR5 with the small molecule inhibitor TAK779 completely blocked entry. The effect was dose-dependent and had a steep threshold, only occurring above 108 M dopamine. The dopamine-mediated increase in entry required dopamine receptor activation, as it was abrogated by the pan-dopamine receptor antagonist flupenthixol, and could be mediated through both subtypes of dopamine receptors. These findings indicate that the effects of dopamine on macrophages may have a significant impact on HIV pathogenesis. They also suggest that drug-induced increases in CNS dopamine may be a common mechanism by which drugs of abuse with distinct modes of action exacerbate neuroinflammation and contribute to HIV-associated neurocognitive disorders in infected drug abusers. PMID:25268786

  16. Topography of the inferior alveolar nerve in human embryos and fetuses. An histomorphological study.

    Directory of Open Access Journals (Sweden)

    Sergey Lvovich Kabak

    2017-11-01

    Full Text Available The aim of this study is to establish the position of the inferior alveolar nerve in relation to the Meckel’s cartilage, the anlage of the mandibular body and primordia of the teeth, and also to trace the change in nerve trunk structure in the human prenatal ontogenesis. Serial sections (20µm from thirty-two 6-12 weeks-old entire human embryos and serial sections (10µm of six mandibles of 13-20 weeks-old human fetuses without developmental abnormalities were studied. Histological sections were impregnated with silver nitrate according to Bilshovsky-Buke and stained with hematoxylin and eosin. During embryonic development, the number of branches of the inferior alveolar nerve increases and its fascicular structure changes. In conclusion, the architecture of intraosseous canals in the body of the mandible, as well as the location of the foramina, is predetermined by the course and pattern of the vessel/nerve branching in the mandibular arch, even before the formation of bony trabeculae. Particularly, the formation of the incisive canal of the mandible can be explained by the presence of the incisive nerve as the extension of the inferior alveolar nerve. It has also been established that Meckel’s cartilage does not participate in mandibular canal morphogenesis.

  17. Adipose tissue macrophages impair preadipocyte differentiation in humans.

    Directory of Open Access Journals (Sweden)

    Li Fen Liu

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

  18. Avaliação da função de macrófagos alveolares em cavalos clinicamente sadios Evaluation of alveolar macrophage function in healthy horses

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    E. Mori

    2001-04-01

    Full Text Available Devido à importância dos macrófagos alveolares (MA nos mecanismos de defesa pulmonar, foram realizados estudos para avaliar a atividade desses fagócitos em cavalos hígidos. Foram realizados lavados broncoalveolares (LBA em cinco cavalos clinicamente sadios. A citologia foi realizada pela citocentrifugação das amostras e posterior confecção de lâminas coradas pelo método de Rosenfeld. Todas as amostras do LBA foram centrifugadas e a concentração celular foi ajustada para 2×10(6 células/ml, para a mensuração da atividade macrofágica (testes de espraiamento, fagocitose e liberação de peróxido de hidrogênio. A contagem diferencial das células presentes no LBA demonstrou a predominância de macrófagos (59,0± 6,9%. Os resultados obtidos nos testes de mensuração da atividade macrofágica foram: índice de espraiamento 25,1± 19,7%, índice de fagocitose 89,4± 6,2% e liberação de peróxido de hidrogênio 1,6± 0,3nmoles/2×10(5 células (sem PMA - phorbol 12-myristate 13-acetate e 1,8± 0,4nmoles/2×10(5 células (com PMA. Os resultados demonstraram um padrão de atividade para MA de cavalos hígidos, os quais apresentaram índices de ativação mesmo sem elicitação prévia, indicando que as técnicas utilizadas foram adequadas para tal propósito.Due to the importance of alveolar macrophages (AM in pulmonary defense mechanisms, studies were performed in order to evaluate the activity of these cells. Bronchoalveolar lavages (BAL were obtained from five healthy horses, and cytology was performed on glass slides after cytocentrifugation of the samples. Slides were stained by Rosenfeld. All BAL samples were centrifuged and cell concentration was adjusted to 2×10(6 cells/ml, for the measurement of AM activity (spreading, phagocytosis and hydrogen peroxide release tests. Differential counting of the BAL cells demonstrated that macrophages were the predominant type of cell (59.0± 6.9%. Measurement of AM activity presented the

  19. The concentrations of clinafloxacin in alveolar macrophages, epithelial lining fluid, bronchial mucosa and serum after administration of single 200 mg oral doses to patients undergoing fibre-optic bronchoscopy.

    Science.gov (United States)

    Honeybourne, D; Andrews, J M; Cunningham, B; Jevons, G; Wise, R

    1999-01-01

    The concentrations of clinafloxacin were measured in serum, bronchial mucosa, alveolar macrophages and epithelial lining fluid after single 200 mg oral doses of clinafloxacin had been administered to 15 subjects who were undergoing bronchoscopy. Concentrations were measured using a microbiological assay method. Mean concentrations in serum, bronchial mucosa, alveolar macrophages and epithelial lining fluid at a mean of 1.27 h post-dose were 1.54, 2.65, 15.60 and 2.71 mg/L respectively. These site concentrations exceeded the MIC90 for common respiratory pathogens and indicate that clinafloxacin is likely to be effective in the treatment of a wide range of respiratory tract infections.

  20. Key Role of Toll-Like Receptor 2 in the Inflammatory Response and Major Histocompatibility Complex Class II Downregulation in Brucella abortus-Infected Alveolar Macrophages

    Science.gov (United States)

    Ferrero, Mariana C.; Hielpos, M. Soledad; Carvalho, Natalia B.; Barrionuevo, Paula; Corsetti, Patricia P.; Giambartolomei, Guillermo H.; Oliveira, Sergio C.

    2014-01-01

    Alveolar macrophages (AM) seem to constitute the main cellular target of inhaled brucellae. Here, we show that Brucella abortus invades and replicates in murine AM without inducing cytotoxicity. B. abortus infection induced a statistically significant increase of tumor necrosis factor alpha (TNF-α), CXCL1 or keratinocyte chemoattractant (KC), interleukin-1β (IL-1β), IL-6, and IL-12 in AM from C57BL/6 mice and BALB/c mice, but these responses were generally weaker and/or delayed compared to those elicited in peritoneal macrophages. Studies using knockout mice for TLR2, TLR4, and TLR9 revealed that TNF-α and KC responses were mediated by TLR2 recognition. Brucella infection reduced in a multiplicity of infection-dependent manner the expression of major histocompatibility complex class II (MHC-II) molecules induced by gamma interferon (IFN-γ) in AM. The same phenomenon was induced by incubation with heat-killed B. abortus (HKBA) or the lipidated form of the 19-kDa outer membrane protein of Brucella (L-Omp19), and it was shown to be mediated by TLR2 recognition. In contrast, no significant downregulation of MHC-II was induced by either unlipidated Omp19 or Brucella LPS. In a functional assay, treatment of AM with either L-Omp19 or HKBA reduced the MHC-II-restricted presentation of OVA peptides to specific T cells. One week after intratracheal infection, viable B. abortus was detected in AM from both wild-type and TLR2 KO mice, but CFU counts were higher in the latter. These results suggest that B. abortus survives in AM after inhalatory infection in spite of a certain degree of immune control exerted by the TLR2-mediated inflammatory response. Both the modest nature of the latter and the modulation of MHC-II expression by the bacterium may contribute to such survival. PMID:24478078

  1. Motion and twisting of magnetic particles ingested by alveolar macrophages in non-smokers and smokers: Implementation of viscoelasticity

    International Nuclear Information System (INIS)

    Moeller, Winfried; Felten, Kathrin; Kohlhaeufl, Martin; Haeussinger, Karl; Kreyling, Wolfgang G.

    2007-01-01

    Ferrimagnetic iron oxide particles were inhaled by 17 healthy volunteers (9 non-smokers, 8 smokers), and the retained particles were magnetized and detected by a SQUID. Stochastic particle transport due to cytoskeletal reorganizations within macrophages (relaxation) and directed particle motion in a weak magnetic twisting field were investigated with respect to viscous and elastic properties of the cytoskeleton. Relaxation and cytoskeletal stiffness were not influenced by cigarette smoking. Relaxation and particle twisting revealed a non-Newtonian viscosity with a pure viscous and a viscoelastic compartment. Viscous and elastic data obtained from relaxation correlated with particle twisting, indicating that the proposed simple model is a reasonable approximation of cytoskeletal mechanical properties

  2. Soluble human leukocyte antigen G5 polarizes differentiation of macrophages toward a decidual macrophage-like phenotype.

    Science.gov (United States)

    Lee, Cheuk-Lun; Guo, YiFan; So, Kam-Hei; Vijayan, Madhavi; Guo, Yue; Wong, Vera H H; Yao, YuanQing; Lee, Kai-Fai; Chiu, Philip C N; Yeung, William S B

    2015-10-01

    What are the actions of soluble human leukocyte antigen G5 (sHLAG5) on macrophage differentiation? sHLAG5 polarizes the differentiation of macrophages toward a decidual macrophage-like phenotype, which could regulate fetomaternal tolerance and placental development. sHLAG5 is a full-length soluble isoform of human leukocyte antigen implicated in immune tolerance during pregnancy. Low or undetectable circulating level of sHLAG5 in first trimester of pregnancy is associated with pregnancy complications such as pre-eclampsia and spontaneous abortion. Decidual macrophages are located in close proximity to invasive trophoblasts, and are involved in regulating fetomaternal tolerance and placental development. Human peripheral blood monocytes were differentiated into macrophages by treatment with granulocyte macrophage colony-stimulating factor in the presence or absence of recombinant sHLAG5 during the differentiation process. The phenotypes and the biological activities of the resulting macrophages were compared. Recombinant sHLAG5 was produced in Escherichia coli BL21 and the protein identity was verified by tandem mass spectrometry. The expression of macrophage markers were analyzed by flow cytometry and quantitative PCR. Phagocytosis was determined by flow cytometry. Indoleamine 2,3-dioxygenase 1 expression and activity were measured by western blot analysis and kynurenine assay, respectively. Cell proliferation and cell cycling were determined by fluorometric cell proliferation assay and flow cytometry, respectively. Cytokine secretion was determined by cytokine array and ELISA kits. Intracellular cytokine expression was measured by flow cytometry. Cell invasion and migration were determined by trans-well invasion and migration assay, respectively. sHLAG5 drove the differentiation of macrophages with 'immuno-modulatory' characteristics, including reduced expression of M1 macrophage marker CD86 and increased expression of M2 macrophage marker CD163. sHLAG5-polarized

  3. Human immunodeficiency virus impairs reverse cholesterol transport from macrophages.

    Directory of Open Access Journals (Sweden)

    Zahedi Mujawar

    2006-10-01

    Full Text Available Several steps of HIV-1 replication critically depend on cholesterol. HIV infection is associated with profound changes in lipid and lipoprotein metabolism and an increased risk of coronary artery disease. Whereas numerous studies have investigated the role of anti-HIV drugs in lipodystrophy and dyslipidemia, the effects of HIV infection on cellular cholesterol metabolism remain uncharacterized. Here, we demonstrate that HIV-1 impairs ATP-binding cassette transporter A1 (ABCA1-dependent cholesterol efflux from human macrophages, a condition previously shown to be highly atherogenic. In HIV-1-infected cells, this effect was mediated by Nef. Transfection of murine macrophages with Nef impaired cholesterol efflux from these cells. At least two mechanisms were found to be responsible for this phenomenon: first, HIV infection and transfection with Nef induced post-transcriptional down-regulation of ABCA1; and second, Nef caused redistribution of ABCA1 to the plasma membrane and inhibited internalization of apolipoprotein A-I. Binding of Nef to ABCA1 was required for down-regulation and redistribution of ABCA1. HIV-infected and Nef-transfected macrophages accumulated substantial amounts of lipids, thus resembling foam cells. The contribution of HIV-infected macrophages to the pathogenesis of atherosclerosis was supported by the presence of HIV-positive foam cells in atherosclerotic plaques of HIV-infected patients. Stimulation of cholesterol efflux from macrophages significantly reduced infectivity of the virions produced by these cells, and this effect correlated with a decreased amount of virion-associated cholesterol, suggesting that impairment of cholesterol efflux is essential to ensure proper cholesterol content in nascent HIV particles. These results reveal a previously unrecognized dysregulation of intracellular lipid metabolism in HIV-infected macrophages and identify Nef and ABCA1 as the key players responsible for this effect. Our findings

  4. Murine iPSC-Derived Macrophages as a Tool for Disease Modeling of Hereditary Pulmonary Alveolar Proteinosis due to Csf2rb Deficiency

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    Adele Mucci

    2016-08-01

    Full Text Available Induced pluripotent stem cells (iPSCs represent an innovative source for the standardized in vitro generation of macrophages (Mφ. We here describe a robust and efficient protocol to obtain mature and functional Mφ from healthy as well as disease-specific murine iPSCs. With regard to morphology, surface phenotype, and function, our iPSC-derived Mφ (iPSC-Mφ closely resemble their counterparts generated in vitro from bone marrow cells. Moreover, when we investigated the feasibility of our differentiation system to serve as a model for rare congenital diseases associated with Mφ malfunction, we were able to faithfully recapitulate the pathognomonic defects in GM-CSF signaling and Mφ function present in hereditary pulmonary alveolar proteinosis (herPAP. Thus, our studies may help to overcome the limitations placed on research into certain rare disease entities by the lack of an adequate supply of disease-specific primary cells, and may aid the development of novel therapeutic approaches for herPAP patients.

  5. Ghrelin ameliorates the human alveolar epithelial A549 cell apoptosis induced by lipopolysaccharide

    International Nuclear Information System (INIS)

    Huang, Chunrong; Zheng, Haichong; He, Wanmei; Lu, Guifang; Li, Xia; Deng, Yubin; Zeng, Mian

    2016-01-01

    Ghrelin is a gastric acyl-peptide that plays an inhibitory role in cell apoptosis. Herein we investigate the protective effects of ghrelin in LPS-induced apoptosis of human alveolar epithelial A549 cells, along with the possible molecular mechanisms. LPS exposure impaired cell viability and increased apoptosis of A549 cells significantly in concentration- and time-dependent manners embodied in increased Bax and cleaved caspase-3 production, coupled with decreased Bcl-2 levels. Simultaneously, LPS remarkably decreased the expression of phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) and extracellular signal-regulated kinas (ERK) in A549 cells. However, ghrelin'pretreatment ameliorated LPS-caused alterations in the ratio of Bax/Bcl-2 and cleaved caspase-3 expression, whereas activated the PI3K/Akt and ERK signaling. These results demonstrate that ghrelin lightens LPS-induced apoptosis of human alveolar epithelial cells partly through activating the PI3K/Akt and ERK pathway and thereby might benefit alleviating septic ALI. -- Graphical abstract: Ghrelin ameliorates the human alveolar epithelial A549 cells apoptosis induced by lipopolysaccharide partly through activating the PI3K/Akt and ERK pathway. Display Omitted -- Highlights: •It has been observed that LPS insult significantly increased apoptosis in A549 cells. •Both Akt and ERK signaling are critical adapter molecules to mediate the ghrelin-mediated proliferative effect. •Ghrelin may have a therapeutic effect in the prevention of LPS-induced apoptosis.

  6. Ghrelin ameliorates the human alveolar epithelial A549 cell apoptosis induced by lipopolysaccharide

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Chunrong; Zheng, Haichong; He, Wanmei; Lu, Guifang; Li, Xia [Department of Medical Intensive Care Unit, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080 (China); Deng, Yubin, E-mail: dengyub@mail.sysu.edu.cn [Research Center of Translational Medicine, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080 (China); Zeng, Mian, E-mail: zengmian2004@163.com [Department of Medical Intensive Care Unit, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080 (China)

    2016-05-20

    Ghrelin is a gastric acyl-peptide that plays an inhibitory role in cell apoptosis. Herein we investigate the protective effects of ghrelin in LPS-induced apoptosis of human alveolar epithelial A549 cells, along with the possible molecular mechanisms. LPS exposure impaired cell viability and increased apoptosis of A549 cells significantly in concentration- and time-dependent manners embodied in increased Bax and cleaved caspase-3 production, coupled with decreased Bcl-2 levels. Simultaneously, LPS remarkably decreased the expression of phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) and extracellular signal-regulated kinas (ERK) in A549 cells. However, ghrelin'pretreatment ameliorated LPS-caused alterations in the ratio of Bax/Bcl-2 and cleaved caspase-3 expression, whereas activated the PI3K/Akt and ERK signaling. These results demonstrate that ghrelin lightens LPS-induced apoptosis of human alveolar epithelial cells partly through activating the PI3K/Akt and ERK pathway and thereby might benefit alleviating septic ALI. -- Graphical abstract: Ghrelin ameliorates the human alveolar epithelial A549 cells apoptosis induced by lipopolysaccharide partly through activating the PI3K/Akt and ERK pathway. Display Omitted -- Highlights: •It has been observed that LPS insult significantly increased apoptosis in A549 cells. •Both Akt and ERK signaling are critical adapter molecules to mediate the ghrelin-mediated proliferative effect. •Ghrelin may have a therapeutic effect in the prevention of LPS-induced apoptosis.

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

    Science.gov (United States)

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

    2015-08-01

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

  8. Legionella pneumophila transcriptome during intracellular multiplication in human macrophages

    Directory of Open Access Journals (Sweden)

    Sebastien P Faucher

    2011-04-01

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

  9. Human β-defensin 3 inhibits periodontitis development by suppressing inflammatory responses in macrophages.

    Science.gov (United States)

    Cui, Di; Lyu, Jinglu; Li, Houxuan; Lei, Lang; Bian, Tianying; Li, Lili; Yan, Fuhua

    2017-11-01

    Human β-defensin 3 (hBD3) is a cationic peptide with immunomodulatory effects on both innate and acquired immune responses. Periodontitis, an inflammatory disease that extends deep into periodontal tissues, causes the loss of supporting structures around the tooth. The present study assessed the effects of hBD3 as a monotherapy for periodontitis in mice and explored its potential mechanism. In vivo, hBD3 inhibited the levels of tumour necrosis factor (TNF)-α, interleukin-6, and matrix metalloprotease-9 in periodontium exposed to Porphyromonas gingivalis (P.g) in a mouse periodontitis model; reduced osteoclast formation and lower alveolar bone loss were also observed. In addition, hBD3 was related to the expression of polarization signature molecules in circulating monocytes. In vitro, hBD3 notably suppressed the production of TNF-α and interleukin-6 in RAW 264.7 cells stimulated by the lipopolysaccharide of P.g. Moreover, hBD3 attenuated polarization of RAW 264.7 cells into the M1 phenotype, with reduced activation of nuclear factor-κB signal transduction. In conclusion, hBD3 exhibits potent anti-periodontitis properties both in vitro and in vivo, and this effect may be correlated to inhibition of the nuclear factor-κB pathway and macrophage polarization. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  10. The Activin A-Peroxisome Proliferator-Activated Receptor Gamma Axis Contributes to the Transcriptome of GM-CSF-Conditioned Human Macrophages.

    Science.gov (United States)

    Nieto, Concha; Bragado, Rafael; Municio, Cristina; Sierra-Filardi, Elena; Alonso, Bárbara; Escribese, María M; Domínguez-Andrés, Jorge; Ardavín, Carlos; Castrillo, Antonio; Vega, Miguel A; Puig-Kröger, Amaya; Corbí, Angel L

    2018-01-01

    GM-CSF promotes the functional maturation of lung alveolar macrophages (A-MØ), whose differentiation is dependent on the peroxisome proliferator-activated receptor gamma (PPARγ) transcription factor. In fact, blockade of GM-CSF-initiated signaling or deletion of the PPARγ-encoding gene PPARG leads to functionally defective A-MØ and the onset of pulmonary alveolar proteinosis. In vitro , macrophages generated in the presence of GM-CSF display potent proinflammatory, immunogenic and tumor growth-limiting activities. Since GM-CSF upregulates PPARγ expression, we hypothesized that PPARγ might contribute to the gene signature and functional profile of human GM-CSF-conditioned macrophages. To verify this hypothesis, PPARγ expression and activity was assessed in human monocyte-derived macrophages generated in the presence of GM-CSF [proinflammatory GM-CSF-conditioned human monocyte-derived macrophages (GM-MØ)] or M-CSF (anti-inflammatory M-MØ), as well as in ex vivo isolated human A-MØ. GM-MØ showed higher PPARγ expression than M-MØ, and the expression of PPARγ in GM-MØ was found to largely depend on activin A. Ligand-induced activation of PPARγ also resulted in distinct transcriptional and functional outcomes in GM-MØ and M-MØ. Moreover, and in the absence of exogenous activating ligands, PPARγ knockdown significantly altered the GM-MØ transcriptome, causing a global upregulation of proinflammatory genes and significantly modulating the expression of genes involved in cell proliferation and migration. Similar effects were observed in ex vivo isolated human A-MØ, where PPARγ silencing led to enhanced expression of genes coding for growth factors and chemokines and downregulation of cell surface pathogen receptors. Therefore, PPARγ shapes the transcriptome of GM-CSF-dependent human macrophages ( in vitro derived GM-MØ and ex vivo isolated A-MØ) in the absence of exogenous activating ligands, and its expression is primarily regulated by activin A

  11. Organic extract of diesel exhaust particles stimulates expression of Ia and costimulatory molecules associated with antigen presentation in rat peripheral blood monocytes but not in alveolar macrophages

    International Nuclear Information System (INIS)

    Koike, Eiko; Kobayashi, Takahiro

    2005-01-01

    We hypothesized that diesel exhaust particles (DEP) induce the activation of antigen-presenting cells (APC) in lung. The present study was designed to clarify the following about DEP: (1) whether it affects the expression of Ia and B7 molecules in alveolar macrophages (AM) as a mature cell or in peripheral blood monocytes (PBM) as an immature cell (2) if it affects the antigen-presenting (AP) activity of PBM (3) what component of DEP is responsible for the effects, and (4) whether the effect of DEP is related to oxidative stress. DEP was extracted with methylene chloride. Cells were exposed to whole DEP, organic extract, or residual particles for 24 h. Cell-surface molecules were measured by flow cytometry. AP activity was assessed by antigen-specific T cell proliferation. Whole DEP or organic extract significantly increased the expression of Ia and B7 molecules on PBM but not on AM. No significant effect of residual particles was observed. A low concentration of organic extract also increased the AP activity of PBM. When the induction of an antioxidative enzyme was assessed, heme oxygenase-1 protein was found to be significantly increased by exposure to whole DEP, and the organic extract was more effective than the residual particles. Furthermore, the organic extract-induced expression of Ia antigen on PBM was reduced by the addition of an antioxidative agent. These results suggest that DEP may act on immature APC and enhance their AP activity and that the action contributing to oxidative stress may be mediated by organic compounds of DEP

  12. Cytokine release from alveolar macrophages exposed to ambient particulate matter: Heterogeneity in relation to size, city and season

    Directory of Open Access Journals (Sweden)

    Dybing Erik

    2005-08-01

    Full Text Available Abstract Background Several studies have demonstrated an association between exposure to ambient particulate matter (PM and respiratory and cardiovascular diseases. Inflammation seems to play an important role in the observed health effects. However, the predominant particle component(s that drives the inflammation is still not fully clarified. In this study representative coarse (2.5–10 μm and fine (0.1–2.5 μm particulate samples from a western, an eastern, a northern and a southern European city (Amsterdam, Lodz, Oslo and Rome were collected during three seasons (spring, summer and winter. All fractions were investigated with respect to cytokine-inducing potential in primary macrophages isolated from rat lung. The results were related to the physical and chemical parameters of the samples in order to disclose possible connections between inflammatory potential and specific characteristics of the particles. Results Compared on a gram-by gram basis, both site-specific and seasonal variations in the PM-induced cytokine responses were demonstrated. The samples collected in the eastern (Lodz and southern (Rome cities appeared to be the most potent. Seasonal variation was most obvious with the samples from Lodz, with the highest responses induced by the spring and summer samples. The site-specific or seasonal variation in cytokine release could not be attributed to variations in any of the chemical parameters. Coarse fractions from all cities were more potent to induce the inflammatory cytokines interleukin-6 and tumour necrosis factor-α than the corresponding fine fractions. Higher levels of specific elements such as iron and copper, some polycyclic aromatic hydrocarbons (PAHs and endotoxin/lipopolysaccaride seemed to be prevalent in the coarse fractions. However, variations in the content of these components did not reflect the variation in cytokine release induced by the different coarse fractions. Addition of polymyxin B did not affect

  13. Lysosomal Disorders Drive Susceptibility to Tuberculosis by Compromising Macrophage Migration

    Science.gov (United States)

    Berg, Russell D.; Levitte, Steven; O’Sullivan, Mary P.; O’Leary, Seónadh M.; Cambier, C.J.; Cameron, James; Takaki, Kevin K.; Moens, Cecilia B.; Tobin, David M.; Keane, Joseph; Ramakrishnan, Lalita

    2016-01-01

    Summary A zebrafish genetic screen for determinants of susceptibility to Mycobacterium marinum identified a hypersusceptible mutant deficient in lysosomal cysteine cathepsins that manifests hallmarks of human lysosomal storage diseases. Under homeostatic conditions, mutant macrophages accumulate undigested lysosomal material, which disrupts endocytic recycling and impairs their migration to, and thus engulfment of, dying cells. This causes a buildup of unengulfed cell debris. During mycobacterial infection, macrophages with lysosomal storage cannot migrate toward infected macrophages undergoing apoptosis in the tuberculous granuloma. The unengulfed apoptotic macrophages undergo secondary necrosis, causing granuloma breakdown and increased mycobacterial growth. Macrophage lysosomal storage similarly impairs migration to newly infecting mycobacteria. This phenotype is recapitulated in human smokers, who are at increased risk for tuberculosis. A majority of their alveolar macrophages exhibit lysosomal accumulations of tobacco smoke particulates and do not migrate to Mycobacterium tuberculosis. The incapacitation of highly microbicidal first-responding macrophages may contribute to smokers’ susceptibility to tuberculosis. PMID:27015311

  14. CRISPR/Cas9-Mediated Gene Editing in Human iPSC-Derived Macrophage Reveals Lysosomal Acid Lipase Function in Human Macrophages-Brief Report.

    Science.gov (United States)

    Zhang, Hanrui; Shi, Jianting; Hachet, Melanie A; Xue, Chenyi; Bauer, Robert C; Jiang, Hongfeng; Li, Wenjun; Tohyama, Junichiro; Millar, John; Billheimer, Jeffrey; Phillips, Michael C; Razani, Babak; Rader, Daniel J; Reilly, Muredach P

    2017-11-01

    To gain mechanistic insights into the role of LIPA (lipase A), the gene encoding LAL (lysosomal acid lipase) protein, in human macrophages. We used CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9) technology to knock out LIPA in human induced pluripotent stem cells and then differentiate to macrophage (human-induced pluripotent stem cells-derived macrophage [IPSDM]) to explore the human macrophage LIPA loss-of-function phenotypes. LIPA was abundantly expressed in monocyte-derived macrophages and was markedly induced on IPSDM differentiation to comparable levels as in human monocyte-derived macrophage. IPSDM with knockout of LIPA ( LIPA -/- ) had barely detectable LAL enzymatic activity. Control and LIPA -/- IPSDM were loaded with [ 3 H]-cholesteryl oleate-labeled AcLDL (acetylated low-density lipoprotein) followed by efflux to apolipoprotein A-I. Efflux of liberated [ 3 H]-cholesterol to apolipoprotein A-I was abolished in LIPA -/- IPSDM, indicating deficiency in LAL-mediated lysosomal cholesteryl ester hydrolysis. In cells loaded with [ 3 H]-cholesterol-labeled AcLDL, [ 3 H]-cholesterol efflux was, however, not different between control and LIPA -/- IPSDM. ABCA1 (ATP-binding cassette, subfamily A, member 1) expression was upregulated by AcLDL loading but to a similar extent between control and LIPA -/- IPSDM. In nonlipid loaded state, LIPA -/- IPSDM had high levels of cholesteryl ester mass compared with minute amounts in control IPSDM. Yet, with AcLDL loading, overall cholesteryl ester mass was increased to similar levels in both control and LIPA -/- IPSDM. LIPA -/- did not impact lysosomal apolipoprotein-B degradation or expression of IL1B , IL6 , and CCL5. CONCLUSIONS: LIPA -/- IPSDM reveals macrophage-specific hallmarks of LIPA deficiency. CRISPR/Cas9 and IPSDM provide important tools to study human macrophage biology and more broadly for future studies of disease-associated LIPA genetic variation in human

  15. Comparative analysis of the internalization of the macrophage receptor sialoadhesin in human and mouse primary macrophages and cell lines.

    Science.gov (United States)

    De Schryver, Marjorie; Leemans, Annelies; Pintelon, Isabel; Cappoen, Davie; Maes, Louis; Caljon, Guy; Cos, Paul; Delputte, Peter L

    2017-06-01

    Sialoadhesin (Sn) is a surface receptor expressed on resident macrophages with the ability to bind with sialic acids. During inflammation, an upregulation of Sn is observed. Upon binding of monoclonal antibodies to Sn, the receptor becomes internalized and this has been observed in multiple species. The latter characteristic, combined with the strong upregulation of Sn on inflammatory macrophages and the fact that Sn-positive macrophages contribute to certain inflammatory diseases, makes Sn an interesting entry portal for phenotype-modulating or cytotoxic drugs. Such drugs or toxins can be linked to Sn-specific antibodies which should enable their targeted uptake by macrophages. However, the activity of such drugs depends not only on their internalization but also on the intracellular trafficking and final fate in the endolysosomal system. Although information is available for porcine Sn, the detailed mechanisms of human and mouse Sn internalization and subsequent intracellular trafficking are currently unknown. To allow development of Sn-targeted therapies, differences across species and cellular background need to be characterized in more detail. In the current report, we show that internalization of human and mouse Sn is dynamin-dependent and clathrin-mediated, both in primary macrophages and CHO cell lines expressing a recombinant Sn. In primary macrophages, internalized Sn-specific F(ab') 2 fragments are located mostly in the early endosomes. With Fc containing Sn-specific antibodies, there is a slight shift towards lysosomal localization in mouse macrophages, possibly because of an interaction with Fc receptors. Surprisingly, in CHO cell lines expressing Sn, there is a predominant lysosomal localization. Our results show that the mechanism of Sn internalization and intracellular trafficking is concurrent in the tested species. The cellular background in which Sn is expressed and the type of antibody used can affect the intracellular fate, which in turn can

  16. MicroRNA profiling of the bovine alveolar macrophage response to Mycobacterium bovis infection suggests pathogen survival is enhanced by microRNA regulation of endocytosis and lysosome trafficking

    OpenAIRE

    BRADLEY, DANIEL

    2015-01-01

    PUBLISHED Mycobacterium bovis, the causative agent of bovine tuberculosis, a major problem for global agriculture, spreads via an airborne route and is taken up by alveolar macrophages (AM) in the lung. Here, we describe the first next-generation sequencing (RNA-seq) approach to temporally profile miRNA expression in primary bovine AMs post-infection with M. bovis. One, six, and forty miRNAs were identified as significantly differentially expressed at 2, 24 and 48 h post-infection, respect...

  17. Lung fibroblasts accelerate wound closure in human alveolar epithelial cells through hepatocyte growth factor/c-Met signaling.

    Science.gov (United States)

    Ito, Yoko; Correll, Kelly; Schiel, John A; Finigan, Jay H; Prekeris, Rytis; Mason, Robert J

    2014-07-01

    There are 190,600 cases of acute lung injury/acute respiratory distress syndrome (ALI/ARDS) each year in the United States, and the incidence and mortality of ALI/ARDS increase dramatically with age. Patients with ALI/ARDS have alveolar epithelial injury, which may be worsened by high-pressure mechanical ventilation. Alveolar type II (ATII) cells are the progenitor cells for the alveolar epithelium and are required to reestablish the alveolar epithelium during the recovery process from ALI/ARDS. Lung fibroblasts (FBs) migrate and proliferate early after lung injury and likely are an important source of growth factors for epithelial repair. However, how lung FBs affect epithelial wound healing in the human adult lung has not been investigated in detail. Hepatocyte growth factor (HGF) is known to be released mainly from FBs and to stimulate both migration and proliferation of primary rat ATII cells. HGF is also increased in lung tissue, bronchoalveolar lavage fluid, and serum in patients with ALI/ARDS. Therefore, we hypothesized that HGF secreted by FBs would enhance wound closure in alveolar epithelial cells (AECs). Wound closure was measured using a scratch wound-healing assay in primary human AEC monolayers and in a coculture system with FBs. We found that wound closure was accelerated by FBs mainly through HGF/c-Met signaling. HGF also restored impaired wound healing in AECs from the elderly subjects and after exposure to cyclic stretch. We conclude that HGF is the critical factor released from FBs to close wounds in human AEC monolayers and suggest that HGF is a potential strategy for hastening alveolar repair in patients with ALI/ARDS. Copyright © 2014 the American Physiological Society.

  18. Influenza H5N1 virus infection of polarized human alveolar epithelial cells and lung microvascular endothelial cells

    Directory of Open Access Journals (Sweden)

    Yuen Kit M

    2009-10-01

    Full Text Available Abstract Background Highly pathogenic avian influenza (HPAI H5N1 virus is entrenched in poultry in Asia and Africa and continues to infect humans zoonotically causing acute respiratory disease syndrome and death. There is evidence that the virus may sometimes spread beyond respiratory tract to cause disseminated infection. The primary target cell for HPAI H5N1 virus in human lung is the alveolar epithelial cell. Alveolar epithelium and its adjacent lung microvascular endothelium form host barriers to the initiation of infection and dissemination of influenza H5N1 infection in humans. These are polarized cells and the polarity of influenza virus entry and egress as well as the secretion of cytokines and chemokines from the virus infected cells are likely to be central to the pathogenesis of human H5N1 disease. Aim To study influenza A (H5N1 virus replication and host innate immune responses in polarized primary human alveolar epithelial cells and lung microvascular endothelial cells and its relevance to the pathogenesis of human H5N1 disease. Methods We use an in vitro model of polarized primary human alveolar epithelial cells and lung microvascular endothelial cells grown in transwell culture inserts to compare infection with influenza A subtype H1N1 and H5N1 viruses via the apical or basolateral surfaces. Results We demonstrate that both influenza H1N1 and H5N1 viruses efficiently infect alveolar epithelial cells from both apical and basolateral surface of the epithelium but release of newly formed virus is mainly from the apical side of the epithelium. In contrast, influenza H5N1 virus, but not H1N1 virus, efficiently infected polarized microvascular endothelial cells from both apical and basolateral aspects. This provides a mechanistic explanation for how H5N1 virus may infect the lung from systemic circulation. Epidemiological evidence has implicated ingestion of virus-contaminated foods as the source of infection in some instances and our

  19. CCR8 signaling influences Toll-like receptor 4 responses in human macrophages in inflammatory diseases.

    Science.gov (United States)

    Reimer, Martina Kvist; Brange, Charlotte; Rosendahl, Alexander

    2011-12-01

    CCR8 immunity is generally associated with Th2 responses in allergic diseases. In this study, we demonstrate for the first time a pronounced attenuated influx of macrophages in ovalbumin (OVA)-challenged CCR8 knockout mice. To explore whether macrophages in human inflamed lung tissue also were CCR8 positive, human lung tissue from patients with chronic obstructive pulmonary disease (COPD) was evaluated. Indeed, CCR8 expression was pronounced in invading monocytes/macrophages from lungs of patients with Global Initiative for Obstructive Lung Disease (GOLD) stage IV COPD. Given this expression pattern, the functional role of CCR8 on human macrophages was evaluated in vitro. Human peripheral blood monocytes expressed low levels of CCR8, while macrophage colony-stimulating factor (M-CSF)-derived human macrophages expressed significantly elevated surface levels of CCR8. Importantly, CCL1 directly regulated the expression of CD18 and CD49b and hence influenced the adhesion capacity of human macrophages. CCL1 drives chemotaxis in M-CSF-derived macrophages, and this could be completely inhibited by lipopolysaccharide (LPS). Whereas both CCL1 and LPS monotreatment inhibited spontaneous superoxide release in macrophages, CCL1 significantly induced superoxide release in the presence of LPS in a dose-dependent manner. Finally, CCL1 induced production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and could inhibit LPS-induced cytokine production in a dose-dependent manner. Our data demonstrate, for the first time, the presence of CCR8 on inflammatory macrophages in human COPD lung tissue. Importantly, the functional data from human macrophages suggest a potential cross talk between the CCR8 and the Toll-like receptor 4 (TLR4) pathways, both of which are present in COPD patients.

  20. Native low-density lipoprotein uptake by macrophage colony-stimulating factor-differentiated human macrophages is mediated by macropinocytosis and micropinocytosis.

    Science.gov (United States)

    Anzinger, Joshua J; Chang, Janet; Xu, Qing; Buono, Chiara; Li, Yifu; Leyva, Francisco J; Park, Bum-Chan; Greene, Lois E; Kruth, Howard S

    2010-10-01

    To examine the pinocytotic pathways mediating native low-density lipoprotein (LDL) uptake by human macrophage colony-stimulating factor-differentiated macrophages (the predominant macrophage phenotype in human atherosclerotic plaques). We identified the kinase inhibitor SU6656 and the Rho GTPase inhibitor toxin B as inhibitors of macrophage fluid-phase pinocytosis of LDL. Assessment of macropinocytosis by time-lapse microscopy revealed that both drugs almost completely inhibited macropinocytosis, although LDL uptake and cholesterol accumulation by macrophages were only partially inhibited (approximately 40%) by these agents. Therefore, we investigated the role of micropinocytosis in mediating LDL uptake in macrophages and identified bafilomycin A1 as an additional partial inhibitor (approximately 40%) of macrophage LDL uptake that targeted micropinocytosis. When macrophages were incubated with both bafilomycin A1 and SU6656, inhibition of LDL uptake was additive (reaching 80%), showing that these inhibitors target different pathways. Microscopic analysis of fluid-phase uptake pathways in these macrophages confirmed that LDL uptake occurs through both macropinocytosis and micropinocytosis. Our findings show that human macrophage colony-stimulating factor-differentiated macrophages take up native LDL by macropinocytosis and micropinocytosis, underscoring the importance of both pathways in mediating LDL uptake by these cells.

  1. Pulmonary alveolar macrophages (PAM) engulf and regain elastin particles and do not respond to some stimuli of neutrophil (PMN) elastinolysis

    International Nuclear Information System (INIS)

    Tricomi, S.M.; Hyers, T.M.; Yu, S.Y.; Liao, J.J.

    1986-01-01

    Elastin degradation by PMN and by PAM differs in the proteinases produced and in the method of cellular attack on the substrate. To further characterize the elastinolytic mechanisms of these two cells, 14 C-labelled bovine ligament elastin was dried onto 24-well culture plates and live cells were placed on the substrate in culture medium. Incubation times were 4 hours for PMN and 20 hours for PAM. Elastinolytic activity was determined by counting 14 C-elastin peptides in the supernatant. By lidocaine release of PAM from the surface, 14 C-elastin retained by the cell was measured. Studies on rabbit PAM showed that 40% of dpm remain associated with the cell at 20 hours. Transmission electron microscopy of human PAM confirmed that PAM can engulf and retain elastin particles at 4 and 24 hours of incubation when in close contact with the substrate. Of the number of dpm released by PMN in 4 hours, PAM in 20 hours released only 23% of that number into supernatant and retained 17% closely associated with the cell after lidocaine treatment. Platelet factor 4, a protein released by platelets upon aggregation which stimulates activity of PMN elastase on elastin, was shown to enhance elastinolysis by whole PMN by 57% at 10 μg/ml in this assay. Platelet factor 4 did not enhance elastinolysis by PAM at concentrations up to 100 μg/ml

  2. A multislice single breath-hold scheme for imaging alveolar oxygen tension in humans.

    Science.gov (United States)

    Hamedani, Hooman; Kadlecek, Stephen J; Emami, Kiarash; Kuzma, Nicholas N; Xu, Yinan; Xin, Yi; Mongkolwisetwara, Puttisarn; Rajaei, Jennia; Barulic, Amy; Wilson Miller, G; Rossman, Milton; Ishii, Masaru; Rizi, Rahim R

    2012-05-01

    Reliable, noninvasive, and high-resolution imaging of alveolar partial pressure of oxygen (p(A)O(2)) is a potentially valuable tool in the early diagnosis of pulmonary diseases. Several techniques have been proposed for regional measurement of p(A)O(2) based on the increased depolarization rate of hyperpolarized (3) He. In this study, we explore one such technique by applying a multislice p(A)O(2) -imaging scheme that uses interleaved-slice ordering to utilize interslice time-delays more efficiently. This approach addresses the low spatial resolution and long breath-hold requirements of earlier techniques, allowing p(A)O(2) measurements to be made over the entire human lung in 10-15 s with a typical resolution of 8.3 × 8.3 × 15.6 mm(3). PO(2) measurements in a glass syringe phantom were in agreement with independent gas analysis within 4.7 ± 4.1% (R = 0.9993). The technique is demonstrated in four human subjects (healthy nonsmoker, healthy former smoker, healthy smoker, and patient with COPD), each imaged six times on 3 different days during a 2-week span. Two independent measurements were performed in each session, consisting of 12 coronal slices. The overall p(A)O(2) mean across all subjects was 95.9 ± 12.2 Torr and correlated well with end-tidal O(2) (R = 0.805, P < 0.0001). The alveolar O(2) uptake rate was consistent with the expected range of 1-2 Torr/s. Repeatable visual features were observed in p(A)O(2) maps over different days, as were characteristic differences among the subjects and gravity-dependent effects. Copyright © 2011 Wiley Periodicals, Inc.

  3. Concentrations of garenoxacin in plasma, bronchial mucosa, alveolar macrophages and epithelial lining fluid following a single oral 600 mg dose in healthy adult subjects.

    Science.gov (United States)

    Andrews, J; Honeybourne, D; Jevons, G; Boyce, M; Wise, R; Bello, A; Gajjar, D

    2003-03-01

    A microbiological assay was used to measure concentrations of garenoxacin (BMS-284756) in plasma, bronchial mucosa (BM), alveolar macrophages (AM) and epithelial lining fluid (ELF), following a single 600 mg oral dose. Twenty-four healthy subjects were allocated into four nominal time intervals after the dose, 2.5-3.5, 4.5-5.5, 10.5-11.5 and 23.5-24.5 h. Mean concentrations in plasma, BM, AM and ELF, respectively, for the four nominal time windows were for 2.5-3.5 h 10.0 mg/L (S.D. 2.8), 7.0 mg/kg (S.D. 1.3), 106.1 mg/L (S.D. 60.3) and 9.2 mg/L (S.D. 3.6); 4.5-5.5 h 8.7 mg/L (S.D. 2.2), 6.0 mg/kg (S.D. 1.9), 158.6 mg/L (S.D. 137.4) and 14.3 mg/L (S.D. 8.2); 10.5-11.5 h 6.1 mg/L (S.D. 1.9), 4.0 mg/kg (S.D. 1.4), 76.0 mg/L (S.D. 47.7) and 7.9 mg/L (S.D. 4.6); and 23.5-24.5 h 2.1 mg/L (S.D. 0.5), 1.7 mg/kg (S.D. 0.7), 30.7 mg/L (S.D. 12.9) and 3.3 mg/L (S.D. 2.3). Concentrations at all sites exceeded MIC(90)s for the common respiratory pathogens Haemophilus influenzae (0.03 mg/L), Moraxella catarrhalis (0.015 mg/L) and Streptococcus pneumoniae (0.06 mg/L). These data suggest that garenoxacin should be effective in the treatment of community-acquired pneumonia and chronic obstructive pulmonary disease.

  4. Evidence for an intracellular niche for Bordetella pertussis in broncho-alveolar lavage cells of mice

    NARCIS (Netherlands)

    Hellwig, SMM; Hazenbos, WLW; van de Winkel, JGJ; Mooi, FR

    1999-01-01

    Bordetella pertussis can attach, invade and survive intracellularly in human macrophages in vitro. To study the significance of this bacterial feature in vivo, we analyzed the presence of viable bacteria in broncho-alveolar lavage (BAL) cells of mice infected with B, pertussis. We found B. pertussis

  5. Functionalized synchrotron in-line phase-contrast computed tomography: a novel approach for simultaneous quantification of structural alterations and localization of barium-labelled alveolar macrophages within mouse lung samples.

    Science.gov (United States)

    Dullin, Christian; dal Monego, Simeone; Larsson, Emanuel; Mohammadi, Sara; Krenkel, Martin; Garrovo, Chiara; Biffi, Stefania; Lorenzon, Andrea; Markus, Andrea; Napp, Joanna; Salditt, Tim; Accardo, Agostino; Alves, Frauke; Tromba, Giuliana

    2015-01-01

    Functionalized computed tomography (CT) in combination with labelled cells is virtually non-existent due to the limited sensitivity of X-ray-absorption-based imaging, but would be highly desirable to realise cell tracking studies in entire organisms. In this study we applied in-line free propagation X-ray phase-contrast CT (XPCT) in an allergic asthma mouse model to assess structural changes as well as the biodistribution of barium-labelled macrophages in lung tissue. Alveolar macrophages that were barium-sulfate-loaded and fluorescent-labelled were instilled intratracheally into asthmatic and control mice. Mice were sacrificed after 24 h, lungs were kept in situ, inflated with air and scanned utilizing XPCT at the SYRMEP beamline (Elettra Synchrotron Light Source, Italy). Single-distance phase retrieval was used to generate data sets with ten times greater contrast-to-noise ratio than absorption-based CT (in our setup), thus allowing to depict and quantify structural hallmarks of asthmatic lungs such as reduced air volume, obstruction of airways and increased soft-tissue content. Furthermore, we found a higher concentration as well as a specific accumulation of the barium-labelled macrophages in asthmatic lung tissue. It is believe that XPCT will be beneficial in preclinical asthma research for both the assessment of therapeutic response as well as the analysis of the role of the recruitment of macrophages to inflammatory sites.

  6. SILICA AND PM1648 MODIFY HUMAN ALVEOLAR MACROPHAGE ANTIGEN PRESENTING CELL ACTIVITY IN VITRO. (R826782)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  7. Gene expression patterns induced at different stages of rhinovirus infection in human alveolar epithelial cells.

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Etemadi

    Full Text Available Human rhinovirus (HRV is the common virus that causes acute respiratory infection (ARI and is frequently associated with lower respiratory tract infections (LRTIs. We aimed to investigate whether HRV infection induces a specific gene expression pattern in airway epithelial cells. Alveolar epithelial cell monolayers were infected with HRV species B (HRV-B. RNA was extracted from both supernatants and infected monolayer cells at 6, 12, 24 and 48 hours post infection (hpi and transcriptional profile was analyzed using Affymetrix GeneChip and the results were subsequently validated using quantitative Real-time PCR method. HRV-B infects alveolar epithelial cells which supports implication of the virus with LRTIs. In total 991 genes were found differentially expressed during the course of infection. Of these, 459 genes were up-regulated whereas 532 genes were down-regulated. Differential gene expression at 6 hpi (187 genes up-regulated vs. 156 down-regulated were significantly represented by gene ontologies related to the chemokines and inflammatory molecules indicating characteristic of viral infection. The 75 up-regulated genes surpassed the down-regulated genes (35 at 12 hpi and their enriched ontologies fell into discrete functional entities such as regulation of apoptosis, anti-apoptosis, and wound healing. At later time points of 24 and 48 hpi, predominated down-regulated genes were enriched for extracellular matrix proteins and airway remodeling events. Our data provides a comprehensive image of host response to HRV infection. The study suggests the underlying molecular regulatory networks genes which might be involved in pathogenicity of the HRV-B and potential targets for further validations and development of effective treatment.

  8. Human Adipose Tissue Macrophages Are Enhanced but Changed to an Anti-Inflammatory Profile in Obesity

    Directory of Open Access Journals (Sweden)

    Karen Fjeldborg

    2014-01-01

    Full Text Available Objective. Adipose tissue (AT macrophages are increased in obesity and associated with low grade inflammation. We aimed to characterize the phenotype of AT macrophages in humans in relation to obesity and insulin resistance. Design. Gene-expression levels of general macrophage markers (CD68 and CD14, proinflammatory markers/M1 (TNF-α, MCP-1, and IL-6, and anti-inflammatory markers/M2 (CD163, CD206, and IL-10 were determined by RT-PCR in subcutaneous AT samples from lean and obese subjects. Insulin resistance was determined by HOMA-IR. Results. All the macrophage markers were elevated in the AT from obese compared to lean subjects (P<0.001. To determine the phenotype of the macrophages the level of CD14 was used to adjust the total number of macrophages. The relative expression of CD163 and IL-10 was elevated, and TNF-α and IL-6 were reduced in AT from obese subjects (all P<0.05. In a multivariate regression analysis CD163 was the only macrophage marker significantly associated with HOMA-IR (β: 0.57; P<0.05. Conclusion. Obesity is associated with elevated numbers of macrophages in the AT. Unexpectedly, the macrophages change phenotype by obesity, with a preponderance of M2 and a decrement of M1 markers in AT from obese subjects. Moreover, CD163 was the only macrophage marker associated with HOMA-IR after multiple adjustments.

  9. Landscape and climatic characteristics associated with human alveolar echinococcosis in France, 1982 to 2007.

    Science.gov (United States)

    Piarroux, M; Gaudart, J; Bresson-Hadni, S; Bardonnet, K; Faucher, B; Grenouillet, F; Knapp, J; Dumortier, J; Watelet, J; Gerard, A; Beytout, J; Abergel, A; Wallon, M; Vuitton, D A; Piarroux, R

    2015-05-07

    Human alveolar echinococcosis (AE) is a severe hepatic disease caused by Echinococcus multilocularis. In France, the definitive and intermediate hosts of E. multilocularis (foxes and rodents, respectively) have a broader geographical distribution than that of human AE. In this two-part study, we describe the link between AE incidence in France between 1982 and 2007 and climatic and landscape characteristics. National-level analysis demonstrated a dramatic increase in AE risk in areas with very cold winters and high annual rainfall levels. Notably, 52% (207/401) of cases resided in French communes (smallest French administrative level) with a mountain climate. The mountain climate communes displayed a 133-fold (95% CI: 95-191) increase in AE risk compared with communes in which the majority of the population resides. A case-control study performed in the most affected areas confirmed the link between AE risk and climatic factors. This arm of the study also revealed that populations residing in forest or pasture areas were at high risk of developing AE. We therefore hypothesised that snow-covered ground may facilitate predators to track their prey, thus increasing E. multilocularis biomass in foxes. Such climatic and landscape conditions could lead to an increased risk of developing AE among humans residing in nearby areas.

  10. Cyclic GMP protects human macrophages against peroxynitrite-induced apoptosis.

    Science.gov (United States)

    Shaw, Catherine A; Webb, David J; Rossi, Adriano G; Megson, Ian L

    2009-05-07

    Nitric oxide (NO) can be both pro- and anti-apoptotic in various cell types, including macrophages. This apparent paradox may result from the actions of NO-related species generated in the microenvironment of the cell, for example the formation of peroxynitrite (ONOO-). In this study we have examined the ability of NO and ONOO- to evoke apoptosis in human monocyte-derived macrophages (MDMvarphi), and investigated whether preconditioning by cyclic guanosine monophosphate (cGMP) is able to limit apoptosis in this cell type. Characterisation of the NO-related species generated by (Z)-1- [2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA/NO) and 1,2,3,4-oxatriazolium, 5-amino-3-(3,4-dichlorophenyl)-, chloride (GEA-3162) was performed by electrochemistry using an isolated NO electrode and electron paramagnetic resonance (EPR) spectrometry. Mononuclear cells were isolated from peripheral blood of healthy volunteers and cultured to allow differentiation into MDMvarphi. Resultant MDMvarphi were treated for 24 h with DETA/NO (100 - 1000 muM) or GEA-3162 (10 - 300 muM) in the presence or absence of BAY 41-2272 (1 muM), isobutylmethylxanthine (IBMX; 1 muM), 1H- [1,2,4]oxadiazolo [4,3-a]quinoxalin-1-one (ODQ; 20 muM) or 8-bromo-cGMP (1 mM). Apoptosis in MDMvarphi was assessed by flow cytometric analysis of annexin V binding in combination with propidium iodide staining. Electrochemistry and EPR revealed that DETA/NO liberated free NO radical, whilst GEA-3162 concomitantly released NO and O2-, and is therefore a ONOO- generator. NO (DETA/NO) had no effect on cell viability, but ONOO- (GEA-3162) caused a concentration-dependent induction of apoptosis in MDMvarphi. Preconditioning of MDMvarphi with NO in combination with the phosphodiesterase inhibitor, 3-Isobutyl-1-methylxanthine (IBMX), or the NO-independent stimulator of soluble guanylate cyclase, BAY 41-2272, significantly attenuated ONOO--induced apoptosis in a cGMP-dependent manner. These results

  11. Cyclic GMP protects human macrophages against peroxynitrite-induced apoptosis

    Directory of Open Access Journals (Sweden)

    Rossi Adriano G

    2009-05-01

    Full Text Available Abstract Background Nitric oxide (NO can be both pro- and anti-apoptotic in various cell types, including macrophages. This apparent paradox may result from the actions of NO-related species generated in the microenvironment of the cell, for example the formation of peroxynitrite (ONOO-. In this study we have examined the ability of NO and ONOO- to evoke apoptosis in human monocyte-derived macrophages (MDMϕ, and investigated whether preconditioning by cyclic guanosine monophosphate (cGMP is able to limit apoptosis in this cell type. Methods Characterisation of the NO-related species generated by (Z-1- [2-(2-aminoethyl-N-(2-ammonioethylamino]diazen-1-ium-1,2-diolate (DETA/NO and 1,2,3,4-oxatriazolium, 5-amino-3-(3,4-dichlorophenyl-, chloride (GEA-3162 was performed by electrochemistry using an isolated NO electrode and electron paramagnetic resonance (EPR spectrometry. Mononuclear cells were isolated from peripheral blood of healthy volunteers and cultured to allow differentiation into MDMϕ. Resultant MDMϕ were treated for 24 h with DETA/NO (100 – 1000 μM or GEA-3162 (10 – 300 μM in the presence or absence of BAY 41–2272 (1 μM, isobutylmethylxanthine (IBMX; 1 μM, 1H- [1,2,4]oxadiazolo [4,3-a]quinoxalin-1-one (ODQ; 20 μM or 8-bromo-cGMP (1 mM. Apoptosis in MDMϕ was assessed by flow cytometric analysis of annexin V binding in combination with propidium iodide staining. Results Electrochemistry and EPR revealed that DETA/NO liberated free NO radical, whilst GEA-3162 concomitantly released NO and O2-, and is therefore a ONOO- generator. NO (DETA/NO had no effect on cell viability, but ONOO- (GEA-3162 caused a concentration-dependent induction of apoptosis in MDMϕ. Preconditioning of MDMϕ with NO in combination with the phosphodiesterase inhibitor, 3-Isobutyl-1-methylxanthine (IBMX, or the NO-independent stimulator of soluble guanylate cyclase, BAY 41–2272, significantly attenuated ONOO--induced apoptosis in a cGMP-dependent manner

  12. Radiological assessment of the inferior alveolar artery course in human corpse mandibles

    Energy Technology Data Exchange (ETDEWEB)

    Bertl, Kristina [Medical University of Vienna, Division of Oral Surgery, Bernhard Gottlieb School of Dentistry, Vienna (Austria); Malmoe University, Department of Periodontology, Faculty of Odontology, Malmoe (Sweden); Hirtler, Lena [Medical University of Vienna, Center for Anatomy and Cell Biology, Department of Systematic Anatomy, Vienna (Austria); Dobsak, Toni [Medical University of Vienna, Division of Oral Surgery, Bernhard Gottlieb School of Dentistry, Vienna (Austria); Medical University of Vienna, Karl Donath Laboratory for Hard Tissue and Biomaterial Research, Division of Oral Surgery, Vienna (Austria); Austrian Cluster for Tissue Regeneration, Vienna (Austria); Heimel, Patrick [Medical University of Vienna, Karl Donath Laboratory for Hard Tissue and Biomaterial Research, Division of Oral Surgery, Vienna (Austria); Austrian Cluster for Tissue Regeneration, Vienna (Austria); Ludwig Boltzmann Institute for Clinical and Experimental Traumatology, Vienna (Austria); Gahleitner, Andre [Medical University of Vienna, Division of Oral Surgery, Bernhard Gottlieb School of Dentistry, Vienna (Austria); Medical University of Vienna, Department of Diagnostic Radiology, Division of Osteoradiology, General Hospital, Vienna (Austria); Ulm, Christian [Medical University of Vienna, Division of Oral Surgery, Bernhard Gottlieb School of Dentistry, Vienna (Austria); Plenk, Hanns [Medical University of Vienna, Bone and Biomaterials Research, Institute for Histology and Embryology, Vienna (Austria)

    2015-04-01

    CT assessment of the entire course of the inferior alveolar artery (IAA) within the mandibular canal. After contrast medium injection (180 or 400 mg/ml iodine concentration) into the external carotid arteries of 15 fresh human cadaver heads, the main IAA's position in the canal (cranial, buccal, lingual or caudal) was assessed in dental CT images of partially edentulous mandibles. The course of the main IAA could be followed at both iodine concentrations. The higher concentration gave the expected better contrast, without creating artefacts, and improved visibility of smaller arteries, such as anastomotic sections, dental branches and the incisive branch. The main IAA changed its position in the canal more often than so far known (mean 4.3 times, SD 1.24, range 2-7), but with a similar bilateral course. A cranial position was most often detected (42 %), followed by lingual (36 %), caudal (16 %) and buccal (6 %). With this non-invasive radiologic method, the entire course of the main IAA in the mandibular canal could be followed simultaneously with other bone structures on both sides of human cadaver mandibles. This methodology allows one to amend existing anatomical and histological data, which are important for surgical interventions near the mandibular canal. (orig.)

  13. Radiological assessment of the inferior alveolar artery course in human corpse mandibles

    International Nuclear Information System (INIS)

    Bertl, Kristina; Hirtler, Lena; Dobsak, Toni; Heimel, Patrick; Gahleitner, Andre; Ulm, Christian; Plenk, Hanns

    2015-01-01

    CT assessment of the entire course of the inferior alveolar artery (IAA) within the mandibular canal. After contrast medium injection (180 or 400 mg/ml iodine concentration) into the external carotid arteries of 15 fresh human cadaver heads, the main IAA's position in the canal (cranial, buccal, lingual or caudal) was assessed in dental CT images of partially edentulous mandibles. The course of the main IAA could be followed at both iodine concentrations. The higher concentration gave the expected better contrast, without creating artefacts, and improved visibility of smaller arteries, such as anastomotic sections, dental branches and the incisive branch. The main IAA changed its position in the canal more often than so far known (mean 4.3 times, SD 1.24, range 2-7), but with a similar bilateral course. A cranial position was most often detected (42 %), followed by lingual (36 %), caudal (16 %) and buccal (6 %). With this non-invasive radiologic method, the entire course of the main IAA in the mandibular canal could be followed simultaneously with other bone structures on both sides of human cadaver mandibles. This methodology allows one to amend existing anatomical and histological data, which are important for surgical interventions near the mandibular canal. (orig.)

  14. Functionalized synchrotron in-line phase-contrast computed tomography: a novel approach for simultaneous quantification of structural alterations and localization of barium-labelled alveolar macrophages within mouse lung samples

    Energy Technology Data Exchange (ETDEWEB)

    Dullin, Christian, E-mail: christian.dullin@med.uni-goettingen.de [University Medical Center Göttingen, Robert Koch Strasse 40, 37075 Göttingen (Germany); Monego, Simeone dal [Cluster in Biomedicine, AREA Science Park Basovizza, Trieste (Italy); Larsson, Emanuel [Elettra Sincrotrone Trieste, Strada Statale 14, km 163.5 in AREA Science Park, 34149 Basovizza (Trieste) (Italy); University of Trieste, Trieste (Italy); Linköping University, SE-581 83 Linkoeping (Sweden); Mohammadi, Sara [Elettra Sincrotrone Trieste, Strada Statale 14, km 163.5 in AREA Science Park, 34149 Basovizza (Trieste) (Italy); Krenkel, Martin [University of Göttingen, Göttingen (Germany); Garrovo, Chiara; Biffi, Stefania [IRCCS Burlo Garofolo, Trieste (Italy); Lorenzon, Andrea [Cluster in Biomedicine, AREA Science Park Basovizza, Trieste (Italy); Markus, Andrea [University Medical Center Göttingen, Robert Koch Strasse 40, 37075 Göttingen (Germany); Napp, Joanna [University Medical Center Göttingen, Robert Koch Strasse 40, 37075 Göttingen (Germany); Max Planck Institute for Experimental Medicine, Hermann-Rein-Strasse 3, 37075 Göttingen (Germany); University Medical Center Göttingen, Robert Koch Strasse 40, 37075 Göttingen (Germany); Salditt, Tim [University of Göttingen, Göttingen (Germany); Accardo, Agostino [University of Trieste, Trieste (Italy); Alves, Frauke [University Medical Center Göttingen, Robert Koch Strasse 40, 37075 Göttingen (Germany); Max Planck Institute for Experimental Medicine, Hermann-Rein-Strasse 3, 37075 Göttingen (Germany); University Medical Center Göttingen, Robert Koch Strasse 40, 37075 Göttingen (Germany); Tromba, Giuliana [Elettra Sincrotrone Trieste, Strada Statale 14, km 163.5 in AREA Science Park, 34149 Basovizza (Trieste) (Italy)

    2015-01-01

    This study presents an approach to increase the sensitivity of lung computed tomography (CT) imaging by utilizing in-line phase contrast CT in combination with single-distance phase-retrieval algorithms and a dedicated image-processing regime. As demonstrated here, functional CT imaging can be achieved for the assessment of both structural alterations in asthmatic mouse lung tissue and the accumulation pattern of instilled barium-sulfate-labelled macrophages in comparison with healthy controls. Functionalized computed tomography (CT) in combination with labelled cells is virtually non-existent due to the limited sensitivity of X-ray-absorption-based imaging, but would be highly desirable to realise cell tracking studies in entire organisms. In this study we applied in-line free propagation X-ray phase-contrast CT (XPCT) in an allergic asthma mouse model to assess structural changes as well as the biodistribution of barium-labelled macrophages in lung tissue. Alveolar macrophages that were barium-sulfate-loaded and fluorescent-labelled were instilled intratracheally into asthmatic and control mice. Mice were sacrificed after 24 h, lungs were kept in situ, inflated with air and scanned utilizing XPCT at the SYRMEP beamline (Elettra Synchrotron Light Source, Italy). Single-distance phase retrieval was used to generate data sets with ten times greater contrast-to-noise ratio than absorption-based CT (in our setup), thus allowing to depict and quantify structural hallmarks of asthmatic lungs such as reduced air volume, obstruction of airways and increased soft-tissue content. Furthermore, we found a higher concentration as well as a specific accumulation of the barium-labelled macrophages in asthmatic lung tissue. It is believe that XPCT will be beneficial in preclinical asthma research for both the assessment of therapeutic response as well as the analysis of the role of the recruitment of macrophages to inflammatory sites.

  15. T3 Regulates a Human Macrophage-Derived TSH-β Splice Variant: Implications for Human Bone Biology.

    Science.gov (United States)

    Baliram, R; Latif, R; Morshed, S A; Zaidi, M; Davies, T F

    2016-09-01

    TSH and thyroid hormones (T3 and T4) are intimately involved in bone biology. We have previously reported the presence of a murine TSH-β splice variant (TSH-βv) expressed specifically in bone marrow-derived macrophages and that exerted an osteoprotective effect by inducing osteoblastogenesis. To extend this observation and its relevance to human bone biology, we set out to identify and characterize a TSH-β variant in human macrophages. Real-time PCR analyses using human TSH-β-specific primers identified a 364-bp product in macrophages, bone marrow, and peripheral blood mononuclear cells that was sequence verified and was homologous to a human TSH-βv previously reported. We then examined TSH-βv regulation using the THP-1 human monocyte cell line matured into macrophages. After 4 days, 46.1% of the THP-1 cells expressed the macrophage markers CD-14 and macrophage colony-stimulating factor and exhibited typical morphological characteristics of macrophages. Real-time PCR analyses of these cells treated in a dose-dependent manner with T3 showed a 14-fold induction of human TSH-βv mRNA and variant protein. Furthermore, these human TSH-βv-positive cells, induced by T3 exposure, had categorized into both M1 and M2 macrophage phenotypes as evidenced by the expression of macrophage colony-stimulating factor for M1 and CCL-22 for M2. These data indicate that in hyperthyroidism, bone marrow resident macrophages have the potential to exert enhanced osteoprotective effects by oversecreting human TSH-βv, which may exert its local osteoprotective role via osteoblast and osteoclast TSH receptors.

  16. Hyperglycemia induces mixed M1/M2 cytokine profile in primary human monocyte-derived macrophages.

    Science.gov (United States)

    Moganti, Kondaiah; Li, Feng; Schmuttermaier, Christina; Riemann, Sarah; Klüter, Harald; Gratchev, Alexei; Harmsen, Martin C; Kzhyshkowska, Julia

    2017-10-01

    Hyperglycaemia is a key factor in diabetic pathology. Macrophages are essential regulators of inflammation which can be classified into two major vectors of polarisation: classically activated macrophages (M1) and alternatively activated macrophages (M2). Both types of macrophages play a role in diabetes, where M1 and M2-produced cytokines can have detrimental effects in development of diabetes-associated inflammation and diabetic vascular complications. However, the effect of hyperglycaemia on differentiation and programming of primary human macrophages was not systematically studied. We established a unique model to assess the influence of hyperglycaemia on M1 and M2 differentiation based on primary human monocyte-derived macrophages. The effects of hyperglycaemia on the gene expression and secretion of prototype M1 cytokines TNF-alpha and IL-1beta, and prototype M2 cytokines IL-1Ra and CCL18 were quantified by RT-PCR and ELISA. Hyperglycaemia stimulated production of TNF-alpha, IL-1beta and IL-1Ra during macrophage differentiation. The effect of hyperglycaemia on TNF-alpha was acute, while the stimulating effect on IL-1beta and IL-1Ra was constitutive. Expression of CCL18 was supressed in M2 macrophages by hyperglycaemia. However the secreted levels remained to be biologically significant. Our data indicate that hyperglycaemia itself, without additional metabolic factors induces mixed M1/M2 cytokine profile that can support of diabetes-associated inflammation and development of vascular complications. Copyright © 2016 Elsevier GmbH. All rights reserved.

  17. Cytotoxicity and gene expression profiling of polyhexamethylene guanidine hydrochloride in human alveolar A549 cells.

    Science.gov (United States)

    Jung, Ha-Na; Zerin, Tamanna; Podder, Biswajit; Song, Ho-Yeon; Kim, Yong-Sik

    2014-06-01

    In Korea, lung disease of children and pregnant women associated with humidifier disinfectant use has become a major concern. A common sterilizer is polyhexamethylene guanidine (PHMG), a member of the guanidine family of antiseptics. This study was done to elucidate the putative cytotoxic effect of PHMG and the PHMG-mediated altered gene expression in human alveolar epithelial A549 cells in vitro. Cell viability analyses revealed the potent cytotoxicity of PHMG, with cell death evident at as low as 5 μg/mL. Death was dose- and time-dependent, and was associated with formation of intracellular reactive oxygen species, and apoptosis significantly, at even 2 μg/mL concentration. The gene expression profile in A549 cells following 24 h exposure to 5 μg/mL of PHMG was investigated using DNA microarray analysis. Changes in gene expression relevant to the progression of cell death included induction of genes related to apoptosis, autophagy, fibrosis, and cell cycle. However, the expressions of genes encoding antioxidant and detoxifying enzymes were down-regulated or not affected. The altered expression of selected genes was confirmed by quantitative reverse transcription-polymerase chain reaction and Western blot analyses. The collective data suggest that PHMG confers cellular toxicity through the generation of intracellular reactive oxygen species and alteration of gene expression. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Dysregulated Functions of Lung Macrophage Populations in COPD.

    Science.gov (United States)

    Kapellos, Theodore S; Bassler, Kevin; Aschenbrenner, Anna C; Fujii, Wataru; Schultze, Joachim L

    2018-01-01

    Chronic obstructive pulmonary disease (COPD) is a diverse respiratory disease characterised by bronchiolitis, small airway obstruction, and emphysema. Innate immune cells play a pivotal role in the disease's progression, and in particular, lung macrophages exploit their prevalence and strategic localisation to orchestrate immune responses. To date, alveolar and interstitial resident macrophages as well as blood monocytes have been described in the lungs of patients with COPD contributing to disease pathology by changes in their functional repertoire. In this review, we summarise recent evidence from human studies and work with animal models of COPD with regard to altered functions of each of these myeloid cell populations. We primarily focus on the dysregulated capacity of alveolar macrophages to secrete proinflammatory mediators and proteases, induce oxidative stress, engulf microbes and apoptotic cells, and express surface and intracellular markers in patients with COPD. In addition, we discuss the differences in the responses between alveolar macrophages and interstitial macrophages/monocytes in the disease and propose how the field should advance to better understand the implications of lung macrophage functions in COPD.

  19. Dysregulated Functions of Lung Macrophage Populations in COPD

    Science.gov (United States)

    Bassler, Kevin; Aschenbrenner, Anna C.

    2018-01-01

    Chronic obstructive pulmonary disease (COPD) is a diverse respiratory disease characterised by bronchiolitis, small airway obstruction, and emphysema. Innate immune cells play a pivotal role in the disease's progression, and in particular, lung macrophages exploit their prevalence and strategic localisation to orchestrate immune responses. To date, alveolar and interstitial resident macrophages as well as blood monocytes have been described in the lungs of patients with COPD contributing to disease pathology by changes in their functional repertoire. In this review, we summarise recent evidence from human studies and work with animal models of COPD with regard to altered functions of each of these myeloid cell populations. We primarily focus on the dysregulated capacity of alveolar macrophages to secrete proinflammatory mediators and proteases, induce oxidative stress, engulf microbes and apoptotic cells, and express surface and intracellular markers in patients with COPD. In addition, we discuss the differences in the responses between alveolar macrophages and interstitial macrophages/monocytes in the disease and propose how the field should advance to better understand the implications of lung macrophage functions in COPD. PMID:29670919

  20. Distinct Properties of Human M-CSF and GM-CSF Monocyte-Derived Macrophages to Simulate Pathological Lung Conditions In Vitro: Application to Systemic and Inflammatory Disorders with Pulmonary Involvement.

    Science.gov (United States)

    Lescoat, Alain; Ballerie, Alice; Augagneur, Yu; Morzadec, Claudie; Vernhet, Laurent; Fardel, Olivier; Jégo, Patrick; Jouneau, Stéphane; Lecureur, Valérie

    2018-03-17

    Macrophages play a central role in the pathogenesis of inflammatory and fibrotic lung diseases. However, alveolar macrophages (AM) are poorly available in humans to perform in vitro studies due to a limited access to broncho-alveolar lavage (BAL). In this study, to identify the best alternative in vitro model for human AM, we compared the phenotype of AM obtained from BAL of patients suffering from three lung diseases (lung cancers, sarcoidosis and Systemic Sclerosis (SSc)-associated interstitial lung disease) to human blood monocyte-derived macrophages (MDMs) differentiated with M-CSF or GM-CSF. The expression of eight membrane markers was evaluated by flow cytometry. Globally, AM phenotype was closer to GM-CSF MDMs. However, the expression levels of CD163, CD169, CD204, CD64 and CD36 were significantly higher in SSc-ILD than in lung cancers. Considering the expression of CD204 and CD36, the phenotype of SSc-AM was closer to MDMs, from healthy donors or SSc patients, differentiated by M-CSF rather than GM-CSF. The comparative secretion of IL-6 by SSc-MDMs and SSc-AM is concordant with these phenotypic considerations. Altogether, these results support the M-CSF MDM model as a relevant in vitro alternative to simulate AM in fibrotic disorders such as SSc.

  1. Potential risk factors associated with human alveolar echinococcosis: Systematic review and meta-analysis.

    Science.gov (United States)

    Conraths, Franz J; Probst, Carolina; Possenti, Alessia; Boufana, Belgees; Saulle, Rosella; La Torre, Giuseppe; Busani, Luca; Casulli, Adriano

    2017-07-01

    Human alveolar echinococcosis (AE) is a severe zoonotic disease caused by the metacestode stage of Echinococcus multilocularis. AE is commonly associated with a long incubation period that may last for more than ten years. The objective of this systematic literature review was to identify and summarize the current knowledge on statistically relevant potential risk factors (PRFs) associated with AE in humans. Six bibliographic databases were searched, generating a total of 1,009 publications. Following the removal of duplicate records and the exclusion of papers that failed to meet the criteria of a previously agreed a priori protocol, 23 publications were retained; however, 6 of these did not contain data in a format that allowed their inclusion in the meta-analysis. The remaining 17 publications (6 case-control and 11 cross-sectional studies) were meta-analysed to investigate associations between AE and PRFs. Pooled odds ratios (OR) were used as a measure of effect and separately analysed for case-control and cross-sectional studies. In the case-control studies, the following PRFs for human AE showed higher odds of outcome: "dog ownership", "cat ownership", "have a kitchen garden", "occupation: farmer", "haymaking in meadows not adjacent to water", "went to forests for vocational reasons", "chewed grass" and "hunting / handling foxes". In the cross-sectional studies, the following PRFs showed higher odds of outcome: "dog ownership", "play with dogs", "gender: female", "age over 20 years", "ethnic group: Tibetan", "low income", "source of drinking water other than well or tap", "occupation: herding" and "low education". Our meta-analysis confirmed that the chance of AE transmission through ingestion of food and water contaminated with E. multilocularis eggs exists, but showed also that food- and water-borne PRFs do not significantly increase the risk of infection. This systematic review analysed international peer-reviewed articles that have over the years

  2. Potential risk factors associated with human alveolar echinococcosis: Systematic review and meta-analysis.

    Directory of Open Access Journals (Sweden)

    Franz J Conraths

    2017-07-01

    Full Text Available Human alveolar echinococcosis (AE is a severe zoonotic disease caused by the metacestode stage of Echinococcus multilocularis. AE is commonly associated with a long incubation period that may last for more than ten years. The objective of this systematic literature review was to identify and summarize the current knowledge on statistically relevant potential risk factors (PRFs associated with AE in humans.Six bibliographic databases were searched, generating a total of 1,009 publications. Following the removal of duplicate records and the exclusion of papers that failed to meet the criteria of a previously agreed a priori protocol, 23 publications were retained; however, 6 of these did not contain data in a format that allowed their inclusion in the meta-analysis. The remaining 17 publications (6 case-control and 11 cross-sectional studies were meta-analysed to investigate associations between AE and PRFs. Pooled odds ratios (OR were used as a measure of effect and separately analysed for case-control and cross-sectional studies. In the case-control studies, the following PRFs for human AE showed higher odds of outcome: "dog ownership", "cat ownership", "have a kitchen garden", "occupation: farmer", "haymaking in meadows not adjacent to water", "went to forests for vocational reasons", "chewed grass" and "hunting / handling foxes". In the cross-sectional studies, the following PRFs showed higher odds of outcome: "dog ownership", "play with dogs", "gender: female", "age over 20 years", "ethnic group: Tibetan", "low income", "source of drinking water other than well or tap", "occupation: herding" and "low education". Our meta-analysis confirmed that the chance of AE transmission through ingestion of food and water contaminated with E. multilocularis eggs exists, but showed also that food- and water-borne PRFs do not significantly increase the risk of infection.This systematic review analysed international peer-reviewed articles that have over the

  3. Progress in the pharmacological treatment of human cystic and alveolar echinococcosis: Compounds and therapeutic targets

    Science.gov (United States)

    Siles-Lucas, Mar; Casulli, Adriano; Cirilli, Roberto

    2018-01-01

    Human cystic and alveolar echinococcosis are helmintic zoonotic diseases caused by infections with the larval stages of the cestode parasites Echinococcus granulosus and E. multilocularis, respectively. Both diseases are progressive and chronic, and often fatal if left unattended for E. multilocularis. As a treatment approach, chemotherapy against these orphan and neglected diseases has been available for more than 40 years. However, drug options were limited to the benzimidazoles albendazole and mebendazole, the only chemical compounds currently licensed for treatment in humans. To compensate this therapeutic shortfall, new treatment alternatives are urgently needed, including the identification, development, and assessment of novel compound classes and drug targets. Here is presented a thorough overview of the range of compounds that have been tested against E. granulosus and E. multilocularis in recent years, including in vitro and in vivo data on their mode of action, dosage, administration regimen, therapeutic outcomes, and associated clinical symptoms. Drugs covered included albendazole, mebendazole, and other members of the benzimidazole family and their derivatives, including improved formulations and combined therapies with other biocidal agents. Chemically synthetized molecules previously known to be effective against other infectious and non-infectious conditions such as anti-virals, antibiotics, anti-parasites, anti-mycotics, and anti-neoplastics are addressed. In view of their increasing relevance, natural occurring compounds derived from plant and fungal extracts are also discussed. Special attention has been paid to the recent application of genomic science on drug discovery and clinical medicine, particularly through the identification of small inhibitor molecules tackling key metabolic enzymes or signalling pathways. PMID:29677189

  4. Progress in the pharmacological treatment of human cystic and alveolar echinococcosis: Compounds and therapeutic targets.

    Directory of Open Access Journals (Sweden)

    Mar Siles-Lucas

    2018-04-01

    Full Text Available Human cystic and alveolar echinococcosis are helmintic zoonotic diseases caused by infections with the larval stages of the cestode parasites Echinococcus granulosus and E. multilocularis, respectively. Both diseases are progressive and chronic, and often fatal if left unattended for E. multilocularis. As a treatment approach, chemotherapy against these orphan and neglected diseases has been available for more than 40 years. However, drug options were limited to the benzimidazoles albendazole and mebendazole, the only chemical compounds currently licensed for treatment in humans. To compensate this therapeutic shortfall, new treatment alternatives are urgently needed, including the identification, development, and assessment of novel compound classes and drug targets. Here is presented a thorough overview of the range of compounds that have been tested against E. granulosus and E. multilocularis in recent years, including in vitro and in vivo data on their mode of action, dosage, administration regimen, therapeutic outcomes, and associated clinical symptoms. Drugs covered included albendazole, mebendazole, and other members of the benzimidazole family and their derivatives, including improved formulations and combined therapies with other biocidal agents. Chemically synthetized molecules previously known to be effective against other infectious and non-infectious conditions such as anti-virals, antibiotics, anti-parasites, anti-mycotics, and anti-neoplastics are addressed. In view of their increasing relevance, natural occurring compounds derived from plant and fungal extracts are also discussed. Special attention has been paid to the recent application of genomic science on drug discovery and clinical medicine, particularly through the identification of small inhibitor molecules tackling key metabolic enzymes or signalling pathways.

  5. Activated prostaglandin D2 receptors on macrophages enhance neutrophil recruitment into the lung

    Science.gov (United States)

    Jandl, Katharina; Stacher, Elvira; Bálint, Zoltán; Sturm, Eva Maria; Maric, Jovana; Peinhaupt, Miriam; Luschnig, Petra; Aringer, Ida; Fauland, Alexander; Konya, Viktoria; Dahlen, Sven-Erik; Wheelock, Craig E.; Kratky, Dagmar; Olschewski, Andrea; Marsche, Gunther; Schuligoi, Rufina; Heinemann, Akos

    2016-01-01

    Background Prostaglandin (PG) D2 is an early-phase mediator in inflammation, but its action and the roles of the 2 D-type prostanoid receptors (DPs) DP1 and DP2 (also called chemoattractant receptor–homologous molecule expressed on TH2 cells) in regulating macrophages have not been elucidated to date. Objective We investigated the role of PGD2 receptors on primary human macrophages, as well as primary murine lung macrophages, and their ability to influence neutrophil action in vitro and in vivo. Methods In vitro studies, including migration, Ca2+ flux, and cytokine secretion, were conducted with primary human monocyte-derived macrophages and neutrophils and freshly isolated murine alveolar and pulmonary interstitial macrophages. In vivo pulmonary inflammation was assessed in male BALB/c mice. Results Activation of DP1, DP2, or both receptors on human macrophages induced strong intracellular Ca2+ flux, cytokine release, and migration of macrophages. In a murine model of LPS-induced pulmonary inflammation, activation of each PGD2 receptor resulted in aggravated airway neutrophilia, tissue myeloperoxidase activity, cytokine contents, and decreased lung compliance. Selective depletion of alveolar macrophages abolished the PGD2-enhanced inflammatory response. Activation of PGD2 receptors on human macrophages enhanced the migratory capacity and prolonged the survival of neutrophils in vitro. In human lung tissue specimens both DP1 and DP2 receptors were located on alveolar macrophages along with hematopoietic PGD synthase, the rate-limiting enzyme of PGD2 synthesis. Conclusion For the first time, our results show that PGD2 markedly augments disease activity through its ability to enhance the proinflammatory actions of macrophages and subsequent neutrophil activation. PMID:26792210

  6. Viral infection of human lung macrophages increases PDL1 expression via IFNβ.

    Directory of Open Access Journals (Sweden)

    Karl J Staples

    Full Text Available Lung macrophages are an important defence against respiratory viral infection and recent work has demonstrated that influenza-induced macrophage PDL1 expression in the murine lung leads to rapid modulation of CD8+ T cell responses via the PD1 receptor. This PD1/PDL1 pathway may downregulate acute inflammatory responses to prevent tissue damage. The aim of this study was to investigate the mechanisms of PDL1 regulation by human macrophages in response to viral infection. Ex-vivo viral infection models using influenza and RSV were established in human lung explants, isolated lung macrophages and monocyte-derived macrophages (MDM and analysed by flow cytometry and RT-PCR. Incubation of lung explants, lung macrophages and MDM with X31 resulted in mean cellular infection rates of 18%, 18% and 29% respectively. Viral infection significantly increased cell surface expression of PDL1 on explant macrophages, lung macrophages and MDM but not explant epithelial cells. Infected MDM induced IFNγ release from autologous CD8+ T cells, an effect enhanced by PDL1 blockade. We observed increases in PDL1 mRNA and IFNβ mRNA and protein release by MDM in response to influenza infection. Knockdown of IFNβ by siRNA, resulted in a 37.5% reduction in IFNβ gene expression in response to infection, and a significant decrease in PDL1 mRNA. Furthermore, when MDM were incubated with IFNβ, this cytokine caused increased expression of PDL1 mRNA. These data indicate that human macrophage PDL1 expression modulates CD8+ cell IFNγ release in response to virus and that this expression is regulated by autologous IFNβ production.

  7. Tie2 signaling cooperates with TNF to promote the pro-inflammatory activation of human macrophages independently of macrophage functional phenotype.

    Science.gov (United States)

    García, Samuel; Krausz, Sarah; Ambarus, Carmen A; Fernández, Beatriz Malvar; Hartkamp, Linda M; van Es, Inge E; Hamann, Jörg; Baeten, Dominique L; Tak, Paul P; Reedquist, Kris A

    2014-01-01

    Angiopoietin (Ang) -1 and -2 and their receptor Tie2 play critical roles in regulating angiogenic processes during development, homeostasis, tumorigenesis, inflammation and tissue repair. Tie2 signaling is best characterized in endothelial cells, but a subset of human and murine circulating monocytes/macrophages essential to solid tumor formation express Tie2 and display immunosuppressive properties consistent with M2 macrophage polarization. However, we have recently shown that Tie2 is strongly activated in pro-inflammatory macrophages present in rheumatoid arthritis patient synovial tissue. Here we examined the relationship between Tie2 expression and function during human macrophage polarization. Tie2 expression was observed under all polarization conditions, but was highest in IFN-γ and IL-10 -differentiated macrophages. While TNF enhanced expression of a common restricted set of genes involved in angiogenesis and inflammation in GM-CSF, IFN-γ and IL-10 -differentiated macrophages, expression of multiple chemokines and cytokines, including CXCL3, CXCL5, CXCL8, IL6, and IL12B was further augmented in the presence of Ang-1 and Ang-2, via Tie2 activation of JAK/STAT signaling. Conditioned medium from macrophages stimulated with Ang-1 or Ang-2 in combination with TNF, sustained monocyte recruitment. Our findings suggest a general role for Tie2 in cooperatively promoting the inflammatory activation of macrophages, independently of polarization conditions.

  8. Human Studies of Vertical and Horizontal Alveolar Ridge Augmentation Comparing Different Types of Bone Graft Materials: A Systematic Review.

    Science.gov (United States)

    Chavda, Suraj; Levin, Liran

    2018-02-01

    Alveolar ridge augmentation can be completed with various types of bone augmentation materials (autogenous, allograft, xenograft, and alloplast). Currently, autogenous bone is labeled as the "gold standard" because of faster healing times and integration between native and foreign bone. No systematic review has currently determined whether there is a difference in implant success between various bone augmentation materials. The purpose of this article was to systematically review comparative human studies of vertical and horizontal alveolar ridge augmentation comparing different types of bone graft materials (autogenous, allograft, xenograft, and alloplast). A MEDLINE search was conducted under the 3 search concepts of bone augmentation, dental implants, and alveolar ridge augmentation. Studies pertaining to socket grafts or sinus lifts were excluded. Case reports, small case series, and review papers were excluded. A bias assessment tool was applied to the final articles. Overall, 219 articles resulted from the initial search, and 9 articles were included for final analysis. There were no discernible differences in implant success between bone augmentation materials. Generally, patients preferred nonautogenous bone sources as there were fewer hospital days, less pain, and better recovery time. Two articles had industrial support; however, conclusions of whether that support influenced the outcomes could not be determined. Future comparative studies should compare nonautogenous bone sources and have longer follow-up times.

  9. Differential Regulation of Gene Expression of Alveolar Epithelial Cell Markers in Human Lung Adenocarcinoma-Derived A549 Clones

    Directory of Open Access Journals (Sweden)

    Hiroshi Kondo

    2015-01-01

    Full Text Available Stem cell therapy appears to be promising for restoring damaged or irreparable lung tissue. However, establishing a simple and reproducible protocol for preparing lung progenitor populations is difficult because the molecular basis for alveolar epithelial cell differentiation is not fully understood. We investigated an in vitro system to analyze the regulatory mechanisms of alveolus-specific gene expression using a human alveolar epithelial type II (ATII cell line, A549. After cloning A549 subpopulations, each clone was classified into five groups according to cell morphology and marker gene expression. Two clones (B7 and H12 were further analyzed. Under serum-free culture conditions, surfactant protein C (SPC, an ATII marker, was upregulated in both H12 and B7. Aquaporin 5 (AQP5, an ATI marker, was upregulated in H12 and significantly induced in B7. When the RAS/MAPK pathway was inhibited, SPC and thyroid transcription factor-1 (TTF-1 expression levels were enhanced. After treatment with dexamethasone (DEX, 8-bromoadenosine 3′5′-cyclic monophosphate (8-Br-cAMP, 3-isobutyl-1-methylxanthine (IBMX, and keratinocyte growth factor (KGF, surfactant protein B and TTF-1 expression levels were enhanced. We found that A549-derived clones have plasticity in gene expression of alveolar epithelial differentiation markers and could be useful in studying ATII maintenance and differentiation.

  10. CCR5 Signal Transduction in Macrophages by Human Immunodeficiency Virus and Simian Immunodeficiency Virus Envelopes

    OpenAIRE

    Arthos, James; Rubbert, Andrea; Rabin, Ronald L.; Cicala, Claudia; Machado, Elizabeth; Wildt, Kathryne; Hanbach, Meredith; Steenbeke, Tavis D.; Swofford, Ruth; Farber, Joshua M.; Fauci, Anthony S.

    2000-01-01

    The capacity of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelopes to transduce signals through chemokine coreceptors on macrophages was examined by measuring the ability of recombinant envelope proteins to mobilize intracellular calcium stores. Both HIV and SIV envelopes mobilized calcium via interactions with CCR5. The kinetics of these responses were similar to those observed when macrophages were treated with MIP-1β. Distinct differences in the capacity o...

  11. MicroRNA profiling of the bovine alveolar macrophage response to Mycobacterium bovis infection suggests pathogen survival is enhanced by microRNA regulation of endocytosis and lysosome trafficking.

    Science.gov (United States)

    Vegh, Peter; Magee, David A; Nalpas, Nicolas C; Bryan, Kenneth; McCabe, Matthew S; Browne, John A; Conlon, Kevin M; Gordon, Stephen V; Bradley, Daniel G; MacHugh, David E; Lynn, David J

    2015-01-01

    Mycobacterium bovis, the causative agent of bovine tuberculosis, a major problem for global agriculture, spreads via an airborne route and is taken up by alveolar macrophages (AM) in the lung. Here, we describe the first next-generation sequencing (RNA-seq) approach to temporally profile miRNA expression in primary bovine AMs post-infection with M. bovis. One, six, and forty miRNAs were identified as significantly differentially expressed at 2, 24 and 48 h post-infection, respectively. The differential expression of three miRNAs (bta-miR-142-5p, bta-miR-146a, and bta-miR-423-3p) was confirmed by RT-qPCR. Pathway analysis of the predicted mRNA targets of differentially expressed miRNAs suggests that these miRNAs preferentially target several pathways that are functionally relevant for mycobacterial pathogenesis, including endocytosis and lysosome trafficking, IL-1 signalling and the TGF-β pathway. Over-expression studies using a bovine macrophage cell-line (Bomac) reveal the targeting of two key genes in the innate immune response to M. bovis, IL-1 receptor-associated kinase 1 (IRAK1) and TGF-β receptor 2 (TGFBR2), by miR-146. Taken together, our study suggests that miRNAs play a key role in tuning the complex interplay between M. bovis survival strategies and the host immune response.

  12. p53 alterations in atypical alveolar hyperplasia of the human lung

    NARCIS (Netherlands)

    Slebos, R. J.; Baas, I. O.; Clement, M. J.; Offerhaus, G. J.; Askin, F. B.; Hruban, R. H.; Westra, W. H.

    1998-01-01

    Atypical alveolar hyperplasia (AAH) is a potential precursor lesion from which lung adenocarcinomas arise and may be a good target for studying the early events of lung tumorigenesis. We have previously shown that AAHs are neoplastic epithelial proliferations that often harbor activating mutations

  13. Differential replication of avian influenza H9N2 viruses in human alveolar epithelial A549 cells

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    Peiris Malik

    2010-03-01

    Full Text Available Abstract Avian influenza virus H9N2 isolates cause a mild influenza-like illness in humans. However, the pathogenesis of the H9N2 subtypes in human remains to be investigated. Using a human alveolar epithelial cell line A549 as host, we found that A/Quail/Hong Kong/G1/97 (H9N2/G1, which shares 6 viral "internal genes" with the lethal A/Hong Kong/156/97 (H5N1/97 virus, replicates efficiently whereas other H9N2 viruses, A/Duck/Hong Kong/Y280/97 (H9N2/Y280 and A/Chicken/Hong Kong/G9/97 (H9N2/G9, replicate poorly. Interestingly, we found that there is a difference in the translation of viral protein but not in the infectivity or transcription of viral genes of these H9N2 viruses in the infected cells. This difference may possibly be explained by H9N2/G1 being more efficient on viral protein production in specific cell types. These findings suggest that the H9N2/G1 virus like its counterpart H5N1/97 may be better adapted to the human host and replicates efficiently in human alveolar epithelial cells.

  14. Transcriptomic analysis of human polarized macrophages: more than one role of alternative activation?

    Directory of Open Access Journals (Sweden)

    Eleonora Derlindati

    Full Text Available Macrophages are a heterogeneous cell population which in response to the cytokine milieu polarize in either classically activated macrophages (M1 or alternatively activated macrophages (M2. This plasticity makes macrophages essential in regulating inflammation, immune response and tissue remodeling and a novel therapeutic target in inflammatory diseases such as atherosclerosis. The aim of the study was to describe the transcriptomic profiles of differently polarized human macrophages to generate new hypotheses on the biological function of the different macrophage subtypes.Polarization of circulating monocytes/macrophages of blood donors was induced in vitro by IFN-γ and LPS (M1, by IL-4 (M2a, and by IL-10 (M2c. Unstimulated cells (RM served as time controls. Gene expression profile of M1, M2a, M2c and RM was assessed at 6, 12 and 24h after polarization with Whole Human Genome Agilent Microarray technique. When compared to RM, M1 significantly upregulated pathways involved in immunity and inflammation, whereas M2a did the opposite. Conversely, decreased and increased expression of mitochondrial metabolism, consistent with insulin resistant and insulin sensitive patterns, was seen in M1 and M2a, respectively. The time sequence in the expression of some pathways appeared to have some specific bearing on M1 function. Finally, canonical and non-canonical Wnt genes and gene groups, promoting inflammation and tissue remodeling, were upregulated in M2a compared to RM.Our data in in vitro polarized human macrophages: 1. confirm and extend known inflammatory and anti-inflammatory gene expression patterns; 2. demonstrate changes in mitochondrial metabolism associated to insulin resistance and insulin sensitivity in M1 and M2a, respectively; 3. highlight the potential relevance of gene expression timing in M1 function; 4. unveil enhanced expression of Wnt pathways in M2a suggesting a potential dual (pro-inflammatory and anti-inflammatory role of M2a in

  15. The impact of splenectomy on human coronary artery atherosclerosis and vascular macrophage distribution.

    Science.gov (United States)

    Li, Yu; Stone, James R

    Splenectomy can potentially impact atherosclerosis through multiple mechanisms including altered lipid homeostasis, increased coagulation, and altered macrophage recruitment to the plaque. In patients, splenectomy has been associated with increased rates of coronary artery events, while in experimental mice, splenectomy causes increased atherosclerosis but reduces systemic monocyte supply. In this study, the direct impact of splenectomy on human coronary artery atherosclerotic plaque severity and macrophage content was investigated. Coronary artery atherosclerotic plaque severity was determined at autopsy in 18 long-term (≥10 years) splenectomy patients and 90 matched control patients. Coronary artery macrophage content was evaluated in mild atherosclerotic plaques of 11 mid- to long-term (≥1 year) splenectomy patients and 11 matched control patients. Splenectomy was associated with reduced coronary artery atherosclerosis (P=.03). The association was most pronounced for the subgroup of patients who had undergone splenectomy 20 years or more prior to death (P=.02). There was no difference in the density of macrophages in the plaque, media, or adventitia upon comparing splenectomy and control patients. In the control group, there was no correlation between the macrophage densities in the three arterial layers. However, in the splenectomy patients, there was a strong correlation in the macrophage densities across the plaque, media, and adventitia (P≤.0002), with resulting slopes that were significantly greater than seen in the control patients (P=.0007-.011). These findings indicate that, in humans, splenectomy is associated with lower coronary artery atherosclerotic plaque severity and altered coronary artery macrophage distribution. These results suggest that the spleen can modulate the recruitment of macrophages into human coronary arteries and the progression of atherosclerosis. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Dual Transcriptome Profiling of Leishmania-Infected Human Macrophages Reveals Distinct Reprogramming Signatures.

    Science.gov (United States)

    Fernandes, Maria Cecilia; Dillon, Laura A L; Belew, Ashton Trey; Bravo, Hector Corrada; Mosser, David M; El-Sayed, Najib M

    2016-05-10

    Macrophages are mononuclear phagocytes that constitute a first line of defense against pathogens. While lethal to many microbes, they are the primary host cells of Leishmania spp. parasites, the obligate intracellular pathogens that cause leishmaniasis. We conducted transcriptomic profiling of two Leishmania species and the human macrophage over the course of intracellular infection by using high-throughput RNA sequencing to characterize the global gene expression changes and reprogramming events that underlie the interactions between the pathogen and its host. A systematic exclusion of the generic effects of large-particle phagocytosis revealed a vigorous, parasite-specific response of the human macrophage early in the infection that was greatly tempered at later time points. An analogous temporal expression pattern was observed with the parasite, suggesting that much of the reprogramming that occurs as parasites transform into intracellular forms generally stabilizes shortly after entry. Following that, the parasite establishes an intracellular niche within macrophages, with minimal communication between the parasite and the host cell later during the infection. No significant difference was observed between parasite species transcriptomes or in the transcriptional response of macrophages infected with each species. Our comparative analysis of gene expression changes that occur as mouse and human macrophages are infected by Leishmania spp. points toward a general signature of the Leishmania-macrophage infectome. Little is known about the transcriptional changes that occur within mammalian cells harboring intracellular pathogens. This study characterizes the gene expression signatures of Leishmania spp. parasites and the coordinated response of infected human macrophages as the pathogen enters and persists within them. After accounting for the generic effects of large-particle phagocytosis, we observed a parasite-specific response of the human macrophages early in

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  18. The effects of exogenous fatty acids and niacin on human monocyte-macrophage plasticity.

    Science.gov (United States)

    Montserrat-de la Paz, Sergio; Rodriguez, Dolores; Cardelo, Magdalena P; Naranjo, Maria C; Bermudez, Beatriz; Abia, Rocio; Muriana, Francisco J G; Lopez, Sergio

    2017-08-01

    Macrophage plasticity allows adapting to different environments, having a dual activity in inflammatory-related diseases. Our hypothesis is that the type of dietary fatty acids into human postprandial triglyceride-rich lipoproteins (TRLs), alone or in combination with niacin (vitamin B3), could modulate the plasticity of monocytes-macrophages. We isolated TRLs at the postprandial peak from blood samples of healthy volunteers after the ingestion of a meal rich in saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs) or MUFAs plus omega-3 long-chain polyunsaturated fatty acids (LCPUFAs). Autologous monocytes isolated at fasting were first induced to differentiate into naïve macrophages. We observed that postprandial TRL-MUFAs, particularly in combination with niacin, enhance competence to monocytes to differentiate and polarise into M2 macrophages. Postprandial TRL-SFAs made polarised macrophages prone to an M1 phenotype. In contrast to dietary SFAs, dietary MUFAs in the meals plus immediate-release niacin primed circulating monocytes for a reduced postprandial pro-inflammatory profile. Our study underlines a role of postprandial TRLs as a metabolic entity in regulating the plasticity of the monocyte-macrophage lineage and also brings an understanding of the mechanisms by which dietary fatty acids are environmental factors fostering the innate immune responsiveness in humans. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    Directory of Open Access Journals (Sweden)

    Carlos Hernández

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

  20. Estrogen Signaling Contributes to Sex Differences in Macrophage Polarization during Asthma.

    Science.gov (United States)

    Keselman, Aleksander; Fang, Xi; White, Preston B; Heller, Nicola M

    2017-09-01

    Allergic asthma is a chronic Th2 inflammation in the lungs that constricts the airways and presents as coughing and wheezing. Asthma mostly affects boys in childhood and women in adulthood, suggesting that shifts in sex hormones alter the course of the disease. Alveolar macrophages have emerged as major mediators of allergic lung inflammation in animal models as well as humans. Whether sex differences exist in macrophage polarization and the molecular mechanism(s) that drive differential responses are not well understood. We found that IL-4-stimulated bone marrow-derived and alveolar macrophages from female mice exhibited greater expression of M2 genes in vitro and after allergen challenge in vivo. Alveolar macrophages from female mice exhibited greater expression of the IL-4Rα and estrogen receptor (ER) α compared with macrophages from male mice following allergen challenge. An ERα-specific agonist enhanced IL-4-induced M2 gene expression in macrophages from both sexes, but more so in macrophages from female mice. Furthermore, IL-4-stimulated macrophages from female mice exhibited more transcriptionally active histone modifications at M2 gene promoters than did macrophages from male mice. We found that supplementation of estrogen into ovariectomized female mice enhanced M2 polarization in vivo upon challenge with allergen and that macrophage-specific deletion of ERα impaired this M2 polarization. The effects of estrogen are long-lasting; bone marrow-derived macrophages from ovariectomized mice implanted with estrogen exhibited enhanced IL-4-induced M2 gene expression compared with macrophages from placebo-implanted littermates. Taken together, our findings suggest that estrogen enhances IL-4-induced M2 gene expression and thereby contributes to sex differences observed in asthma. Copyright © 2017 by The American Association of Immunologists, Inc.

  1. Human macrophages support persistent transcription from unintegrated HIV-1 DNA

    International Nuclear Information System (INIS)

    Kelly, Jeremy; Beddall, Margaret H.; Yu Dongyang; Iyer, Subashini R.; Marsh, Jon W.; Wu Yuntao

    2008-01-01

    Retroviruses require integration of their RNA genomes for both stability and productive viral replication. In HIV infection of non-dividing, resting CD4 T cells, where integration is greatly impeded, the reverse transcribed HIV DNA has limited biological activity and a short half-life. In metabolically active and proliferating T cells, unintegrated DNA rapidly diminishes with cell division. HIV also infects the non-dividing but metabolically active macrophage population. In an in vitro examination of HIV infection of macrophages, we find that unintegrated viral DNA not only has an unusual stability, but also maintains biological activity. The unintegrated linear DNA, 1-LTR, and 2-LTR circles are stable for at least 30 days. Additionally, there is persistent viral gene transcription, which is selective and skewed towards viral early genes such as nef and tat with highly diminished rev and vif. One viral early gene product Nef was measurably synthesized. We also find that independent of integration, the HIV infection process in macrophages leads to generation of numerous chemokines

  2. Comparative 'omics analyses differentiate Mycobacterium tuberculosis and Mycobacterium bovis and reveal distinct macrophage responses to infection with the human and bovine tubercle bacilli

    Science.gov (United States)

    Malone, Kerri M.; Rue-Albrecht, Kévin; Magee, David A.; Conlon, Kevin; Schubert, Olga T.; Nalpas, Nicolas C.; Browne, John A.; Smyth, Alicia; Gormley, Eamonn; Aebersold, Ruedi; MacHugh, David E.; Gordon, Stephen V.

    2018-01-01

    Members of the Mycobacterium tuberculosis complex (MTBC) are the causative agents of tuberculosis in a range of mammals, including humans. A key feature of MTBC pathogens is their high degree of genetic identity yet distinct host tropism. Notably, while Mycobacterium bovis is highly virulent and pathogenic for cattle, the human pathogen M. tuberculosis is attenuated in cattle. Previous research also suggests that host preference amongst MTBC members has a basis in host innate immune responses. To explore MTBC host tropism, we present in-depth profiling of the MTBC reference strains M. bovis AF2122/97 and M. tuberculosis H37Rv at both the global transcriptional and the translational level via RNA-sequencing and SWATH MS. Furthermore, a bovine alveolar macrophage infection time course model was used to investigate the shared and divergent host transcriptomic response to infection with M. tuberculosis H37Rv or M. bovis AF2122/97. Significant differential expression of virulence-associated pathways between the two bacilli was revealed, including the ESX-1 secretion system. A divergent transcriptional response was observed between M. tuberculosis H37Rv and M. bovis AF2122/97 infection of bovine alveolar macrophages, in particular cytosolic DNA-sensing pathways at 48 h post-infection, and highlights a distinct engagement of M. bovis with the bovine innate immune system. The work presented here therefore provides a basis for the identification of host innate immune mechanisms subverted by virulent host-adapted mycobacteria to promote their survival during the early stages of infection. PMID:29557774

  3. Modulation of cytokine expression in human macrophages by endocrine-disrupting chemical Bisphenol-A

    International Nuclear Information System (INIS)

    Liu, Yanzhen; Mei, Chenfang; Liu, Hao; Wang, Hongsheng; Zeng, Guoqu; Lin, Jianhui; Xu, Meiying

    2014-01-01

    Highlights: • Effects of BPA on the cytokines expression of human macrophages were investigated. • BPA increased pro-inflammation cytokines TNF-α and IL-6 production. • BPA decreased anti-inflammation IL-10 and TGF-β production. • ERα/β/ERK/NF-κB signaling involved in BPA-mediated cytokines expression. - Abstract: Exposure to environmental endocrine-disrupting chemical Bisphenol-A (BPA) is often associated with dysregulated immune homeostasis, but the mechanisms remain unclear. In the present study, the effects of BPA on the cytokines responses of human macrophages were investigated. Treatment with BPA increased pro-inflammation cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) production, but decreased anti-inflammation cytokines interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production in THP1 macrophages, as well as in primary human macrophages. BPA effected cytokines expression through estrogen receptor α/β (ERα/β)-dependent mechanism with the evidence of ERα/β antagonist reversed the expression of cytokines. We also identified that activation of extracellular regulated protein kinases (ERK)/nuclear factor κB (NF-κB) signal cascade marked the effects of BPA on cytokines expression. Our results indicated that BPA effected inflammatory responses of macrophages via modulating of cytokines expression, and provided a new insight into the link between exposure to BPA and human health

  4. Modulation of cytokine expression in human macrophages by endocrine-disrupting chemical Bisphenol-A

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yanzhen; Mei, Chenfang [State Key Laboratory of Applied Microbiology Southern China, Guangzhou 510070 (China); Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Institute of Microbiology, Guangzhou 510070 (China); Liu, Hao [Affiliated Cancer Hospital and Cancer Research Institute, Guangzhou Medical University, Guangzhou 510095 (China); Wang, Hongsheng [Department of Microbial and Biochemical Pharmacy, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006 (China); Zeng, Guoqu; Lin, Jianhui [State Key Laboratory of Applied Microbiology Southern China, Guangzhou 510070 (China); Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Institute of Microbiology, Guangzhou 510070 (China); Xu, Meiying, E-mail: xumy@gdim.cn [State Key Laboratory of Applied Microbiology Southern China, Guangzhou 510070 (China); Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Institute of Microbiology, Guangzhou 510070 (China)

    2014-09-05

    Highlights: • Effects of BPA on the cytokines expression of human macrophages were investigated. • BPA increased pro-inflammation cytokines TNF-α and IL-6 production. • BPA decreased anti-inflammation IL-10 and TGF-β production. • ERα/β/ERK/NF-κB signaling involved in BPA-mediated cytokines expression. - Abstract: Exposure to environmental endocrine-disrupting chemical Bisphenol-A (BPA) is often associated with dysregulated immune homeostasis, but the mechanisms remain unclear. In the present study, the effects of BPA on the cytokines responses of human macrophages were investigated. Treatment with BPA increased pro-inflammation cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) production, but decreased anti-inflammation cytokines interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production in THP1 macrophages, as well as in primary human macrophages. BPA effected cytokines expression through estrogen receptor α/β (ERα/β)-dependent mechanism with the evidence of ERα/β antagonist reversed the expression of cytokines. We also identified that activation of extracellular regulated protein kinases (ERK)/nuclear factor κB (NF-κB) signal cascade marked the effects of BPA on cytokines expression. Our results indicated that BPA effected inflammatory responses of macrophages via modulating of cytokines expression, and provided a new insight into the link between exposure to BPA and human health.

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

    Science.gov (United States)

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

    2013-01-01

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

  6. A new data management system for the French National Registry of human alveolar echinococcosis cases

    Directory of Open Access Journals (Sweden)

    Charbonnier Amandine

    2014-01-01

    Full Text Available Alveolar echinococcosis (AE is an endemic zoonosis in France due to the cestode Echinococcus multilocularis. The French National Reference Centre for Alveolar Echinococcosis (CNR-EA, connected to the FrancEchino network, is responsible for recording all AE cases diagnosed in France. Administrative, epidemiological and medical information on the French AE cases may currently be considered exhaustive only on the diagnosis time. To constitute a reference data set, an information system (IS was developed thanks to a relational database management system (MySQL language. The current data set will evolve towards a dynamic surveillance system, including follow-up data (e.g. imaging, serology and will be connected to environmental and parasitological data relative to E. multilocularis to better understand the pathogen transmission pathway. A particularly important goal is the possible interoperability of the IS with similar European and other databases abroad; this new IS could play a supporting role in the creation of new AE registries.

  7. Variations in alveolar partial pressure for carbon dioxide and oxygen have additive not synergistic acute effects on human pulmonary vasoconstriction.

    Science.gov (United States)

    Croft, Quentin P P; Formenti, Federico; Talbot, Nick P; Lunn, Daniel; Robbins, Peter A; Dorrington, Keith L

    2013-01-01

    The human pulmonary vasculature constricts in response to hypercapnia and hypoxia, with important consequences for homeostasis and adaptation. One function of these responses is to direct blood flow away from poorly-ventilated regions of the lung. In humans it is not known whether the stimuli of hypercapnia and hypoxia constrict the pulmonary blood vessels independently of each other or whether they act synergistically, such that the combination of hypercapnia and hypoxia is more effective than the sum of the responses to each stimulus on its own. We independently controlled the alveolar partial pressures of carbon dioxide (Paco 2) and oxygen (Pao 2) to examine their possible interaction on human pulmonary vasoconstriction. Nine volunteers each experienced sixteen possible combinations of four levels of Paco 2 (+6, +1, -4 and -9 mmHg, relative to baseline) with four levels of Pao 2 (175, 100, 75 and 50 mmHg). During each of these sixteen protocols Doppler echocardiography was used to evaluate cardiac output and systolic tricuspid pressure gradient, an index of pulmonary vasoconstriction. The degree of constriction varied linearly with both Paco 2 and the calculated haemoglobin oxygen desaturation (1-So2). Mixed effects modelling delivered coefficients defining the interdependence of cardiac output, systolic tricuspid pressure gradient, ventilation, Paco 2 and So2. No interaction was observed in the effects on pulmonary vasoconstriction of carbon dioxide and oxygen (p>0.64). Direct effects of the alveolar gases on systolic tricuspid pressure gradient greatly exceeded indirect effects arising from concurrent changes in cardiac output.

  8. 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

  9. Anti-inflammatory effects of octadecylamine-functionalized nanodiamond on primary human macrophages.

    Science.gov (United States)

    Pentecost, A E; Witherel, C E; Gogotsi, Y; Spiller, K L

    2017-09-26

    Chronic inflammatory disorders such as rheumatoid arthritis are characterized by excessive pro-inflammatory or "M1" activation of macrophages, the primary cells of the innate immune system. Current treatments include delivery of glucocorticoids (e.g. dexamethasone - Dex), which reduce pro-inflammatory M1 behaviour in macrophages. However, these treatments have many off-target effects on cells other than macrophages, resulting in broad immunosuppression. To limit such side effects, drug-incorporated nano- and microparticles may be used to selectively target macrophages via phagocytosis, because of their roles as highly effective phagocytes in the body. In this study, surface-modified nanodiamond (ND) was explored as a platform for the delivery of dexamethasone to macrophages because of ND's rich surface chemistry, which contributes to ND's high potential as a versatile drug delivery platform. After finding that octadecylamine-functionalized nanodiamond (ND-ODA) enhanced adsorption of Dex compared to carboxylated ND, the effects of Dex, ND-ODA, and Dex-adsorbed ND-ODA on primary human macrophage gene expression were characterized. Surprisingly, even in the absence of Dex, ND-ODA had strong anti-inflammatory effects, as determined by multiplex gene expression via NanoString and by protein secretion analysis via ELISA. ND-ODA also inhibited expression of M2a markers yet increased the expression of M2c markers and phagocytic receptors. Interestingly, the adsorption of Dex to ND-ODA further increased some anti-inflammatory effects, but abrogated the effect on phagocytic receptors, compared to its individual components. Overall, the ability of ND-ODA to promote anti-inflammatory and pro-phagocytic behaviour in macrophages, even in the absence of loaded drugs, suggests its potential for use as an anti-inflammatory therapeutic to directly target macrophages through phagocytosis.

  10. Human CD68 promoter GFP transgenic mice allow analysis of monocyte to macrophage differentiation in vivo.

    Science.gov (United States)

    Iqbal, Asif J; McNeill, Eileen; Kapellos, Theodore S; Regan-Komito, Daniel; Norman, Sophie; Burd, Sarah; Smart, Nicola; Machemer, Daniel E W; Stylianou, Elena; McShane, Helen; Channon, Keith M; Chawla, Ajay; Greaves, David R

    2014-10-09

    The recruitment of monocytes and their differentiation into macrophages at sites of inflammation are key events in determining the outcome of the inflammatory response and initiating the return to tissue homeostasis. To study monocyte trafficking and macrophage differentiation in vivo, we have generated a novel transgenic reporter mouse expressing a green fluorescent protein (GFP) under the control of the human CD68 promoter. CD68-GFP mice express high levels of GFP in both monocyte and embryo-derived tissue resident macrophages in adult animals. The human CD68 promoter drives GFP expression in all CD115(+) monocytes of adult blood, spleen, and bone marrow; we took advantage of this to directly compare the trafficking of bone marrow-derived CD68-GFP monocytes to that of CX3CR1(GFP) monocytes in vivo using a sterile zymosan peritonitis model. Unlike CX3CR1(GFP) monocytes, which downregulate GFP expression on differentiation into macrophages in this model, CD68-GFP monocytes retain high-level GFP expression for 72 hours after differentiation into macrophages, allowing continued cell tracking during resolution of inflammation. In summary, this novel CD68-GFP transgenic reporter mouse line represents a powerful resource for analyzing monocyte mobilization and monocyte trafficking as well as studying the fate of recruited monocytes in models of acute and chronic inflammation. © 2014 by The American Society of Hematology.

  11. CCR8 Signaling Influences Toll-Like Receptor 4 Responses in Human Macrophages in Inflammatory Diseases ▿

    Science.gov (United States)

    Kvist Reimer, Martina; Brange, Charlotte; Rosendahl, Alexander

    2011-01-01

    CCR8 immunity is generally associated with Th2 responses in allergic diseases. In this study, we demonstrate for the first time a pronounced attenuated influx of macrophages in ovalbumin (OVA)-challenged CCR8 knockout mice. To explore whether macrophages in human inflamed lung tissue also were CCR8 positive, human lung tissue from patients with chronic obstructive pulmonary disease (COPD) was evaluated. Indeed, CCR8 expression was pronounced in invading monocytes/macrophages from lungs of patients with Global Initiative for Obstructive Lung Disease (GOLD) stage IV COPD. Given this expression pattern, the functional role of CCR8 on human macrophages was evaluated in vitro. Human peripheral blood monocytes expressed low levels of CCR8, while macrophage colony-stimulating factor (M-CSF)-derived human macrophages expressed significantly elevated surface levels of CCR8. Importantly, CCL1 directly regulated the expression of CD18 and CD49b and hence influenced the adhesion capacity of human macrophages. CCL1 drives chemotaxis in M-CSF-derived macrophages, and this could be completely inhibited by lipopolysaccharide (LPS). Whereas both CCL1 and LPS monotreatment inhibited spontaneous superoxide release in macrophages, CCL1 significantly induced superoxide release in the presence of LPS in a dose-dependent manner. Finally, CCL1 induced production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and could inhibit LPS-induced cytokine production in a dose-dependent manner. Our data demonstrate, for the first time, the presence of CCR8 on inflammatory macrophages in human COPD lung tissue. Importantly, the functional data from human macrophages suggest a potential cross talk between the CCR8 and the Toll-like receptor 4 (TLR4) pathways, both of which are present in COPD patients. PMID:21976223

  12. Degalactosylated/desialylated human serum containing GcMAF induces macrophage phagocytic activity and in vivo antitumor activity.

    Science.gov (United States)

    Kuchiike, Daisuke; Uto, Yoshihiro; Mukai, Hirotaka; Ishiyama, Noriko; Abe, Chiaki; Tanaka, Daichi; Kawai, Tomohito; Kubo, Kentaro; Mette, Martin; Inui, Toshio; Endo, Yoshio; Hori, Hitoshi

    2013-07-01

    The group-specific component protein-derived macrophage-activating factor (GcMAF) has various biological activities, such as macrophage activation and antitumor activity. Clinical trials of GcMAF have been carried out for metastatic breast cancer, prostate cancer, and metastatic colorectal cancer. In this study, despite the complicated purification process of GcMAF, we used enzymatically-treated human serum containing GcMAF with a considerable macrophage-stimulating activity and antitumor activity. We detected GcMAF in degalactosylated/desialylated human serum by western blotting using an anti-human Gc globulin antibody, and Helix pomatia agglutinin lectin. We also found that GcMAF-containing human serum significantly enhanced the phagocytic activity of mouse peritoneal macrophages and extended the survival time of mice bearing Ehrlich ascites tumors. We demonstrated that GcMAF-containing human serum can be used as a potential macrophage activator for cancer immunotherapy.

  13. In vitro differentiation of human monocytes to macrophages: change of PDE profile and its relationship to suppression of tumour necrosis factor-α release by PDE inhibitors

    Science.gov (United States)

    Gantner, Florian; Kupferschmidt, Rochus; Schudt, Christian; Wendel, Albrecht; Hatzelmann, Armin

    1997-01-01

    During in vitro culture in 10% human AB serum, human peripheral blood monocytes acquire a macrophage-like phenotype. The underlying differentiation was characterized by increased activities of the macrophage marker enzymes unspecific esterase (NaF-insensitive form) and acid phosphatase, as well as by a down-regulation in surface CD14 expression. In parallel, a dramatic change in the phosphodiesterase (PDE) profile became evident within a few days that strongly resembled that previously described for human alveolar macrophages. Whereas PDE1 and PDE3 activities were augmented, PDE4 activity, which represented the major cyclic AMP-hydrolysing activity of peripheral blood monocytes, rapidly declined. Monocytes and monocyte-derived macrophages responded to lipopolysaccharide (LPS) with the release of tumour necrosis factor-α (TNF). In line with the change in CD14 expression, the EC50 value of LPS for induction of TNF release increased from approximately 0.1 ng ml−1 in peripheral blood monocytes to about 2 ng ml−1 in macrophages. Both populations of cells were equally susceptible towards inhibition of TNF release by cyclic AMP elevating agents such as dibutyryl cyclic AMP, prostaglandin E2 (PGE2) or forskolin, which all led to a complete abrogation of TNF production in a concentration-dependent manner and which were more efficient than the glucocorticoid dexamethasone. In monocytes, PDE4 selective inhibitors (rolipram, RP73401) suppressed TNF formation by 80%, whereas motapizone, a PDE3 selective compound, exerted a comparatively weak effect (10–15% inhibition). Combined use of PDE3 plus PDE4 inhibitors resulted in an additive effect and fully abrogated LPS-induced TNF release as did the mixed PDE3/4 inhibitor tolafentrine. In monocyte-derived macrophages, neither PDE3- nor PDE4-selective drugs markedly affected TNF generation when used alone (<15% inhibition), whereas in combination, they led to a maximal inhibition of TNF formation by about 40–50

  14. Foamy macrophages and the progression of the human TB granuloma

    Science.gov (United States)

    Russell, David G.; Cardona, Pere-Joan; Kim, Mi-Jeong; Allain, Sophie; Altare, Frédéric

    2009-01-01

    The progression of tuberculosis from a latent, sub-clinical infection to active disease that culminates in transmission of infectious bacilli is determined locally at the level of the granuloma. This progression takes place even in the face of a robust immune response that, while it contains infection, is unable to eliminate the bacterium. The factors or environmental conditions that influence this progression remain to be determined. Recent advances have indicated that pathogen-induced dysregulation of host lipid synthesis and sequestration plays a critical role in this transition. The foamy macrophage appears to be a key player in both sustaining persistent bacteria and contributing to the tissue pathology that leads to cavitation and release of infectious bacilli. PMID:19692995

  15. Cloning of a cDNA encoding chitotriosidase, a human chitinase produced by macrophages

    NARCIS (Netherlands)

    Boot, R. G.; Renkema, G. H.; Strijland, A.; van Zonneveld, A. J.; Aerts, J. M.

    1995-01-01

    We have recently observed that chitotriosidase, a chitinolytic enzyme, is secreted by activated human macrophages and is markedly elevated in plasma of Gaucher disease patients (Hollak, C. E. M., van Weely, S., van Oers, M. H. J., and Aerts, J. M. F. G. (1994) J. Clin. Invest. 93, 1288-1292). Here,

  16. Cytotoxicity and inflammation in human alveolar epithelial cells following exposure to occupational levels of gold and silver nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Bachand, George D., E-mail: gdbacha@sandia.gov [Sandia National Laboratories, Center for Integrated Nanotechnologies (United States); Allen, Amy [Sandia National Laboratories, Department of Analytical Science (United States); Bachand, Marlene [Sandia National Laboratories, Department of Nanobiology (United States); Achyuthan, Komandoor E. [Sandia National Laboratories, Department of Biosensors and Nanomaterials (United States); Seagrave, Jean Clare [Lovelace Respiratory Research Institute, Applied Life Science and Toxicology Division (United States); Brozik, Susan M. [Sandia National Laboratories, Department of Biosensors and Nanomaterials (United States)

    2012-10-15

    While inhalation represents one of the most likely routes of exposure, the toxicity and response of nanoparticles at concentrations expected from such an exposure are not well understood. Here we characterized the in vitro response of human A549 adenocarcinomic alveolar epithelial cells following exposure to gold (AuNP) and silver (AgNP) nanoparticles at levels approximating an occupational exposure. Changes in neither oxidative stress nor cytotoxicity were significantly affected by exposure to AgNPs and AuNPs, regardless of NP type (Ag vs. Au), concentration, surface ligand (citrate or tannic acid), or size. An inflammatory response was, however, observed in response to 20 nm AgNPs and 20 nm AuNPs, where significant differences in the release of interleukin (IL)-8 but not IL-6 were observed. Additional data demonstrated that increased IL-8 secretion was strongly dependent on both nanoparticle size and concentration. Overall these data suggest that, while not acutely toxic, occupational exposure to AuNPs and AgNPs may trigger a significant inflammatory response in alveolar epithelium. Moreover, the differential responses in IL-8 and IL-6 secretion suggest that NPs may induce a response pathway that is distinct from those commonly elicited by allergens and pathogens.

  17. 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.

  18. Macrophage-specific nanotechnology-driven CD163 overexpression in human macrophages results in an M2 phenotype under inflammatory conditions.

    Science.gov (United States)

    Alvarado-Vazquez, Perla Abigail; Bernal, Laura; Paige, Candler A; Grosick, Rachel L; Moracho Vilrriales, Carolina; Ferreira, David Wilson; Ulecia-Morón, Cristina; Romero-Sandoval, E Alfonso

    2017-08-01

    M1 macrophages release proinflammatory factors during inflammation. They transit to an M2 phenotype and release anti-inflammatory factors to resolve inflammation. An imbalance in the transition from M1 to M2 phenotype in macrophages contributes to the development of persistent inflammation. CD163, a member of the scavenger receptor cysteine-rich family, is an M2 macrophage marker. The functional role of CD163 during the resolution of inflammation is not completely known. We postulate that CD163 contributes to the transition from M1 to M2 phenotype in macrophages. We induced CD163 gene in THP-1 and primary human macrophages using polyethylenimine nanoparticles grafted with a mannose ligand (Man-PEI). This nanoparticle specifically targets cells of monocytic origin via mannose receptors. Cells were challenged with a single or a double stimulation of lipopolysaccharide (LPS). A CD163 or empty plasmid was complexed with Man-PEI nanoparticles for cell transfections. Quantitative RT-PCR, immunocytochemistry, and ELISAs were used for molecular assessments. CD163-overexpressing macrophages displayed reduced levels of tumor necrosis factor-alpha (TNF)-α and monocytes chemoattractant protein (MCP)-1 after a single stimulation with LPS. Following a double stimulation paradigm, CD163-overexpressing macrophages showed an increase of interleukin (IL)-10 and IL-1ra and a reduction of MCP-1. This anti-inflammatory phenotype was partially blocked by an anti-CD163 antibody (effects on IL-10 and IL-1ra). A decrease in the release of TNF-α, IL-1β, and IL-6 was observed in CD163-overexpressing human primary macrophages. The release of IL-6 was blocked by an anti-CD163 antibody in the CD163-overexpressing group. Our data show that the induction of the CD163 gene in human macrophages under inflammatory conditions produces changes in cytokine secretion in favor of an anti-inflammatory phenotype. Targeting macrophages to induce CD163 using cell-directed nanotechnology is an attractive

  19. Efeitos do estresse agudo de contenção, do estresse crônico de natação e da administração de glutamina sobre a liberação de superóxido por macrófagos alveolares de ratos Effects of acute restraint stress, chronic swim stress and glutamine administration on the release of superoxide from alveolar macrophages of rats

    Directory of Open Access Journals (Sweden)

    Elizabeth do Nascimento

    2007-08-01

    Full Text Available OBJETIVO: Avaliar a liberação de ânion superóxido por macrófagos alveolares em ratos submetidos ou não ao estresse agudo, ao exercício físico de natação e à suplementação com glutamina. MÉTODOS: Quarenta e dois ratos machos da linhagem Wistar com idade em torno de 62 (desvio-padrão=3 dias de idade foram divididos em grupos controle, treino, estresse e glutamina. Após a intervenção, macrófagos alveolares foram coletados e estimulados com acetato de formol miristato para a avaliação da liberação de ânion superóxido. RESULTADOS: Em comparação à primeira hora (controle=26,2, desvio-padrão=4,2; treino=28,7, desvio-padrão=5,1; estresse=20,3, desvio-padrão=4,4; glutamina=26,2, desvio-padrão=4,2, houve aumento (pOBJECTIVE: To assess the release of superoxide anion from alveolar macrophages of rats submitted or not to acute restraint stress, forced swimming and glutamine supplementation. METHODS: Forty-two male Wistar rats aging roughly 62 days (standard deviation=3 were randomly divided into four groups: control, training, stress and glutamine. After the intervention, alveolar macrophages were collected and stimulated with phorbol myristate acetate to assess the release of superoxide anion. RESULTS: When compared with the first hour (control=26.2, standard deviation=4.2; training=28.7, standard deviation=5.1; stress=20.3 , standard deviation=4.4; glutamine=26.2, standard deviation=4.2, the release of superoxide increased (p<0.001 in all experimental groups in the second hour (control=38.4, standard deviation=4.9; training=40.7, standard deviation=6.1; stress=30.2, standard deviation=5.6; glutamine=39.2, standard deviation=5.2 of observation. Training and glutamine supplementation did not induce differences in the release of superoxide from alveolar macrophages when compared with the control group. Only the rats submitted to stress showed a reduction in the release of superoxide in both the first (20.3, standard deviation

  20. Micro-computed tomography assessment of human alveolar bone: bone density and three-dimensional micro-architecture.

    Science.gov (United States)

    Kim, Yoon Jeong; Henkin, Jeffrey

    2015-04-01

    Micro-computed tomography (micro-CT) is a valuable means to evaluate and secure information related to bone density and quality in human necropsy samples and small live animals. The aim of this study was to assess the bone density of the alveolar jaw bones in human cadaver, using micro-CT. The correlation between bone density and three-dimensional micro architecture of trabecular bone was evaluated. Thirty-four human cadaver jaw bone specimens were harvested. Each specimen was scanned with micro-CT at resolution of 10.5 μm. The bone volume fraction (BV/TV) and the bone mineral density (BMD) value within a volume of interest were measured. The three-dimensional micro architecture of trabecular bone was assessed. All the parameters in the maxilla and the mandible were subject to comparison. The variables for the bone density and the three-dimensional micro architecture were analyzed for nonparametric correlation using Spearman's rho at the significance level of p architecture parameters were consistently higher in the mandible, up to 3.3 times greater than those in the maxilla. The most linear correlation was observed between BV/TV and BMD, with Spearman's rho = 0.99 (p = .01). Both BV/TV and BMD were highly correlated with all micro architecture parameters with Spearman's rho above 0.74 (p = .01). Two aspects of bone density using micro-CT, the BV/TV and BMD, are highly correlated with three-dimensional micro architecture parameters, which represent the quality of trabecular bone. This noninvasive method may adequately enhance evaluation of the alveolar bone. © 2013 Wiley Periodicals, Inc.

  1. A methodological approach to assessing alveolar ridge preservation procedures in humans: soft tissue profile.

    Science.gov (United States)

    Vanhoutte, Vanessa; Rompen, Eric; Lecloux, Geoffrey; Rues, Stefan; Schmitter, Marc; Lambert, France

    2014-03-01

    The aesthetic results of implant restoration in the anterior maxilla are particularly related to the soft tissue profile. Although socket preservation techniques appear to reduce bone remodelling after tooth extraction, there is still few investigations assessing the external soft tissue profile after such procedures. The goal of this study was to describe an accurate technique to evaluate soft tissue contour changes after performing socket preservation procedures. The secondary objective was to apply the newly developed measuring method to a specific socket preservation using a "saddled" connective tissue graft combined with the insertion of slowly resorbable biomaterials into the socket. A total of 14 patients needing tooth replacement in the aesthetic region were included to receive a socket preservation procedure using a connective tissue graft. Impressions were taken before the tooth extraction (baseline) and at 2, 4, and 12 weeks after the procedure. The corresponding plaster casts were scanned, and the evolution of the soft tissue profile in relation to the baseline situation was assessed using imaging software. The measuring technique allowed assessing the soft tissue profiles accurately at different levels of the alveolar process. The insertion of a saddled connective tissue appeared to compensate for the horizontal and vertical bone remodelling after a socket preservation procedure in most regions of the alveolar crest. After 12 weeks, the only significant change was located in the more cervical and central region of the alveolar process and reached a median drop of 0.62 mm from baseline. Within the limitations of this study, we found that a saddled connective tissue graft combined with a socket preservation procedure could almost completely counteract the bone remodelling in terms of the external soft tissue profile. The minor changes found in the cervical region might disappear with the emergence profile of the prosthodontic components. The described

  2. The Reactive Oxygen Species in Macrophage Polarization: Reflecting Its Dual Role in Progression and Treatment of Human Diseases

    Science.gov (United States)

    Tan, Hor-Yue; Li, Sha; Hong, Ming; Wang, Xuanbin

    2016-01-01

    High heterogeneity of macrophage is associated with its functions in polarization to different functional phenotypes depending on environmental cues. Macrophages remain in balanced state in healthy subject and thus macrophage polarization may be crucial in determining the tissue fate. The two distinct populations, classically M1 and alternatively M2 activated, representing the opposing ends of the full activation spectrum, have been extensively studied for their associations with several disease progressions. Accumulating evidences have postulated that the redox signalling has implication in macrophage polarization and the key roles of M1 and M2 macrophages in tissue environment have provided the clue for the reasons of ROS abundance in certain phenotype. M1 macrophages majorly clearing the pathogens and ROS may be crucial for the regulation of M1 phenotype, whereas M2 macrophages resolve inflammation which favours oxidative metabolism. Therefore how ROS play its role in maintaining the homeostatic functions of macrophage and in particular macrophage polarization will be reviewed here. We also review the biology of macrophage polarization and the disturbance of M1/M2 balance in human diseases. The potential therapeutic opportunities targeting ROS will also be discussed, hoping to provide insights for development of target-specific delivery system or immunomodulatory antioxidant for the treatment of ROS-related diseases. PMID:27143992

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

    International Nuclear Information System (INIS)

    Pons, I.

    1997-09-01

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

  4. CSF1R inhibition prevents radiation pulmonary fibrosis by depletion of interstitial macrophages.

    Science.gov (United States)

    Meziani, Lydia; Mondini, Michele; Petit, Benoît; Boissonnas, Alexandre; Thomas de Montpreville, Vincent; Mercier, Olaf; Vozenin, Marie-Catherine; Deutsch, Eric

    2018-03-01

    Radiation-induced lung fibrosis (RIF) is a delayed side-effect of chest radiotherapy, frequently associated with macrophage infiltration.We aimed to characterise the role of pulmonary macrophages in RIF using human lung biopsies from patients receiving radiotherapy for thorax malignancies and a RIF model developed in C57BL/6 mice after 16-Gy thorax irradiation.High numbers of macrophages (both interstitial and alveolar) were detected in clinical and preclinical RIF. In the preclinical model, upregulation of T-helper (Th)2 cytokines was measured, whereas Th1 cytokines were downregulated in RIF tissue lysate. Bronchoalveolar lavage demonstrated upregulation of both types of cytokines. At steady state, tissue-infiltrating macrophages (IMs) expressed 10-fold more arginase (Arg)-1 than alveolar macrophages (AMs), and a 40-fold upregulation of Arg-1 was found in IMs isolated from RIF. IMs, but not AMs, were able to induce myofibroblast activation in vitro In addition, whereas depletion of AMs using Clodrosome didn't affect RIF score, depletion of IMs using a clinically available colony-stimulating factor receptor-1 (CSF1R) neutralising antibody was antifibrotic.These findings suggest differential contributions of alveolar versus interstitial macrophages in RIF, highlighting the fibrogenic role of IMs. The CSF1/CSF1R pathway was identified as a new therapeutic target to inhibit RIF. Copyright ©ERS 2018.

  5. Enhanced M1 macrophage polarization in human helicobacter pylori-associated atrophic gastritis and in vaccinated mice.

    Directory of Open Access Journals (Sweden)

    Marianne Quiding-Järbrink

    Full Text Available BACKGROUND: Infection with Helicobacter pylori triggers a chronic gastric inflammation that can progress to atrophy and gastric adenocarcinoma. Polarization of macrophages is a characteristic of both cancer and infection, and may promote progression or resolution of disease. However, the role of macrophages and their polarization during H. pylori infection has not been well defined. METHODOLOGY/PRINCIPAL FINDINGS: By using a mouse model of infection and gastric biopsies from 29 individuals, we have analyzed macrophage recruitment and polarization during H. pylori infection by flow cytometry and real-time PCR. We found a sequential recruitment of neutrophils, eosinophils and macrophages to the gastric mucosa of infected mice. Gene expression analysis of stomach tissue and sorted macrophages revealed that gastric macrophages were polarized to M1 after H. pylori infection, and this process was substantially accelerated by prior vaccination. Human H. pylori infection was characterized by a mixed M1/M2 polarization of macrophages. However, in H. pylori-associated atrophic gastritis, the expression of inducible nitric oxide synthase was markedly increased compared to uncomplicated gastritis, indicative of an enhanced M1 macrophage polarization in this pre-malignant lesion. CONCLUSIONS/SIGNIFICANCE: These results show that vaccination of mice against H. pylori amplifies M1 polarization of gastric macrophages, and that a similar enhanced M1 polarization is present in human H. pylori-induced atrophic gastritis.

  6. Genetic programs expressed in resting and IL-4 alternatively activated mouse and human macrophages : similarities and differences

    NARCIS (Netherlands)

    Martinez, Fernando O.; Helming, Laura; Milde, Ronny; Varin, Audrey; Melgert, Barbro N.; Draijer, Christina; Thomas, Benjamin; Fabbri, Marco; Crawshaw, Anjali; Ho, Ling Pei; Ten Hacken, Nick H.; Jimenez, Viviana Cobos; Kootstra, Neeltje A.; Hamann, Jorg; Greaves, David R.; Locati, Massimo; Mantovani, Alberto; Gordon, Siamon

    2013-01-01

    The molecular repertoire of macrophages in health and disease can provide novel biomarkers for diagnosis, prognosis, and treatment. Th2-IL-4-activated macrophages (M2) have been associated with important diseases in mice, yet no specific markers are available for their detection in human tissues.

  7. Genetic programs expressed in resting and IL-4 alternatively activated mouse and human macrophages: similarities and differences

    NARCIS (Netherlands)

    Martinez, Fernando O.; Helming, Laura; Milde, Ronny; Varin, Audrey; Melgert, Barbro N.; Draijer, Christina; Thomas, Benjamin; Fabbri, Marco; Crawshaw, Anjali; Ho, Ling Pei; ten Hacken, Nick H.; Cobos Jiménez, Viviana; Kootstra, Neeltje A.; Hamann, Jörg; Greaves, David R.; Locati, Massimo; Mantovani, Alberto; Gordon, Siamon

    2013-01-01

    The molecular repertoire of macrophages in health and disease can provide novel biomarkers for diagnosis, prognosis, and treatment. Th2-IL-4-activated macrophages (M2) have been associated with important diseases in mice, yet no specific markers are available for their detection in human tissues.

  8. Acute Exposure to Electronic and Combustible Cigarette Aerosols: Effects in an Animal Model and in Human Alveolar Cells.

    Science.gov (United States)

    Husari, Ahmad; Shihadeh, Alan; Talih, Soha; Hashem, Yasmine; El Sabban, Marwan; Zaatari, Ghazi

    2016-05-01

    Smoking electronic cigarettes (ECIG) is promoted as a safer alternative to smoking combustible cigarettes. This study investigates the effects of ECIG aerosol and cigarette smoke (CS) in an animal model and in human alveolar cell cultures (A549). Mice were divided into Control, ECIG, and CS. Animals were exposed for 6h/d to either lab air, ECIG or CS, for of 3 days. Total particulate matter exposure for the ECIG was set at higher levels compared to CS. Lung injury was determined by: (1) measurement of wet-to-dry ratio; (2) albumin concentration in the bronchoalveolar lavage fluid; (3) transcriptional expression of inflammatory mediators IL-1β, IL-6, TNF-α; (4) oxidative stress; (5) assessment of cell death; and (6) lung histopathology. Human alveolar cell cultures were treated with various concentrations of ECIG and CS aerosol extracts and the effects on cell proliferation were evaluated. Wet-to-dry ratio was higher in CS when compared to ECIG. Albumin leak in bronchoalveolar lavage fluid was evident in CS but not in ECIG. ECIG exposure was only associated with a significant increase in IL-1β. In contrast, CS exposure resulted in significant increases in IL-1β, IL-6, TNF-α expression, and oxidative stress. TUNEL staining demonstrated significant cell death in CS but not in ECIG. At the cellular level, ECIG and CS extracts reduced cell proliferation, however, CS exhibited effects at lower concentrations. Despite higher exposure conditions, ECIG exhibited less toxic effects on lungs of experimental animals and on A549 cell cultures when compared to CS. © The Author 2015. Published by Oxford University Press on behalf of the Society for Research on Nicotine and Tobacco. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  9. CXCL9 Regulates TGF-β1-Induced Epithelial to Mesenchymal Transition in Human Alveolar Epithelial Cells.

    Science.gov (United States)

    O'Beirne, Sarah L; Walsh, Sinead M; Fabre, Aurélie; Reviriego, Carlota; Worrell, Julie C; Counihan, Ian P; Lumsden, Robert V; Cramton-Barnes, Jennifer; Belperio, John A; Donnelly, Seamas C; Boylan, Denise; Marchal-Sommé, Joëlle; Kane, Rosemary; Keane, Michael P

    2015-09-15

    Epithelial to mesenchymal cell transition (EMT), whereby fully differentiated epithelial cells transition to a mesenchymal phenotype, has been implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). CXCR3 and its ligands are recognized to play a protective role in pulmonary fibrosis. In this study, we investigated the presence and extent of EMT and CXCR3 expression in human IPF surgical lung biopsies and assessed whether CXCR3 and its ligand CXCL9 modulate EMT in alveolar epithelial cells. Coexpression of the epithelial marker thyroid transcription factor-1 and the mesenchymal marker α-smooth muscle actin and CXCR3 expression was examined by immunohistochemical staining of IPF surgical lung biopsies. Epithelial and mesenchymal marker expression was examined by quantitative real-time PCR, Western blotting, and immunofluorescence in human alveolar epithelial (A549) cells treated with TGF-β1 and CXCL9, with Smad2, Smad3, and Smad7 expression and cellular localization examined by Western blotting. We found that significantly more cells were undergoing EMT in fibrotic versus normal areas of lung in IPF surgical lung biopsy samples. CXCR3 was expressed by type II pneumocytes and fibroblasts in fibrotic areas in close proximity to cells undergoing EMT. In vitro, CXCL9 abrogated TGF-β1-induced EMT. A decrease in TGF-β1-induced phosphorylation of Smad2 and Smad3 occurred with CXCL9 treatment. This was associated with increased shuttling of Smad7 from the nucleus to the cytoplasm where it inhibits Smad phosphorylation. This suggests a role for EMT in the pathogenesis of IPF and provides a novel mechanism for the inhibitory effects of CXCL9 on TGF-β1-induced EMT. Copyright © 2015 by The American Association of Immunologists, Inc.

  10. Vitamin D Is Required for IFN-γ–Mediated Antimicrobial Activity of Human Macrophages

    Science.gov (United States)

    Fabri, Mario; Stenger, Steffen; Shin, Dong-Min; Yuk, Jae-Min; Liu, Philip T.; Realegeno, Susan; Lee, Hye-Mi; Krutzik, Stephan R.; Schenk, Mirjam; Sieling, Peter A.; Teles, Rosane; Montoya, Dennis; Iyer, Shankar S.; Bruns, Heiko; Lewinsohn, David M.; Hollis, Bruce W.; Hewison, Martin; Adams, John S.; Steinmeyer, Andreas; Zügel, Ulrich; Cheng, Genhong; Jo, Eun-Kyeong; Bloom, Barry R.; Modlin, Robert L.

    2012-01-01

    Control of tuberculosis worldwide depends on our understanding of human immune mechanisms, which combat the infection. Acquired T cell responses are critical for host defense against microbial pathogens, yet the mechanisms by which they act in humans remain unclear. We report that T cells, by the release of interferon-γ (IFN-γ), induce autophagy, phagosomal maturation, the production of antimicrobial peptides such as cathelicidin, and antimicrobial activity against Mycobacterium tuberculosis in human macrophages via a vitamin D–dependent pathway. IFN-γ induced the antimicrobial pathway in human macrophages cultured in vitamin D–sufficient sera, but not in sera from African-Americans that have lower amounts of vitamin D and who are more susceptible to tuberculosis. In vitro supplementation of vitamin D–deficient serum with 25-hydroxyvitamin D3 restored IFN-γ–induced antimicrobial peptide expression, autophagy, phagosome-lysosome fusion, and antimicrobial activity. These results suggest a mechanism in which vitamin D is required for acquired immunity to overcome the ability of intracellular pathogens to evade macrophage-mediated antimicrobial responses. The present findings underscore the importance of adequate amounts of vitamin D in all human populations for sustaining both innate and acquired immunity against infection. PMID:21998409

  11. Detection of active alveolar bone destruction in human periodontal disease by analysis of radiopharmaceutical uptake after a single injection of 99m-Tc-methylene diphosphonate

    International Nuclear Information System (INIS)

    Jeffcoat, M.K.; Williams, R.C.; Holman, B.L.; English, R.; Goldhaber, P.

    1986-01-01

    Previous studies have shown that, following a single injection of 99m-Tc-MDP, measurement of bone-seeking radiopharmaceutical uptake can detect ''active'' alveolar bone loss due to periodontal disease in beagle dogs, as determined by radiographs taken at the time of, and several months after, the nuclear medicine procedure. The efficacy of this diagnostic test, however, had not been assessed in human periodontal disease. The ability of a single boneseeking radiopharmaceutical uptake examination to detect ''active'' alveolar bone loss due to periodontal disease in human patients was assessed by comparing a single uptake measurement to the rate of bone loss determined from serial radiographs taken over a 6-month period. Uptake was expressed as a ratio of the cpm from the alveolar bone divided by the cpm from the non-tooth supporting bone of the nuchal crest. High uptake ratios were associated with ''active'' loss and low uptake ratios were associated with little if any change in alveolar bone height (p<0.001). The nuclear medicine examination was an accurate detector of periodontal disease activity in nearly 80% of the individual teeth studied. These data indicate that high bone-seeking radiopharmaceutical uptake ratios may be pathognomonic of active bone loss in human periodontal disease. (author)

  12. Detection of active alveolar bone destruction in human periodontal disease by analysis of radiopharmaceutical uptake after a single injection of 99m-Tc-methylene diphosphonate

    Energy Technology Data Exchange (ETDEWEB)

    Jeffcoat, M.K.; Williams, R.C.; Holman, B.L.; English, R.; Goldhaber, P.

    1986-01-01

    Previous studies have shown that, following a single injection of 99m-Tc-MDP, measurement of bone-seeking radiopharmaceutical uptake can detect ''active'' alveolar bone loss due to periodontal disease in beagle dogs, as determined by radiographs taken at the time of, and several months after, the nuclear medicine procedure. The efficacy of this diagnostic test, however, had not been assessed in human periodontal disease. The ability of a single boneseeking radiopharmaceutical uptake examination to detect ''active'' alveolar bone loss due to periodontal disease in human patients was assessed by comparing a single uptake measurement to the rate of bone loss determined from serial radiographs taken over a 6-month period. Uptake was expressed as a ratio of the cpm from the alveolar bone divided by the cpm from the non-tooth supporting bone of the nuchal crest. High uptake ratios were associated with ''active'' loss and low uptake ratios were associated with little if any change in alveolar bone height (p<0.001). The nuclear medicine examination was an accurate detector of periodontal disease activity in nearly 80% of the individual teeth studied. These data indicate that high bone-seeking radiopharmaceutical uptake ratios may be pathognomonic of active bone loss in human periodontal disease.

  13. A very rare cause of dyspnea with a unique presentation on a computed tomography scan of the chest: macrophage activation syndrome

    International Nuclear Information System (INIS)

    Brandao-Neto, Rodrigo Antonio; Santana, Alfredo Nicodemos Cruz; Danilovic, Debora Lucia Seguro; Mendonca, Berenice Bilharinho de; Bernardi, Fabiola Del Carlo; Barbas, Carmen Silvia Valente

    2008-01-01

    Macrophage activation syndrome is a rare and potentially life-threatening disease. It occurs due to immune dysregulation manifested as excessive macrophage proliferation, typically causing hepatosplenomegaly, pancytopenia and hepatic dysfunction. Here, we report an unusual case of macrophage activation syndrome presenting as dyspnea, as well as (reported here for the first time) high resolution computed tomography findings of an excavated nodule, diffuse ground glass opacities and consolidations (mimicking severe pneumonia or alveolar hemorrhage). The patient was successfully treated with human immunoglobulin. We recommend that macrophage activation syndrome be considered in the differential diagnosis of respiratory failure. Rapid diagnosis and treatment are essential to achieving favorable outcomes in patients with this syndrome. (author)

  14. Efficient internalization of mesoporous silica particles of different sizes by primary human macrophages without impairment of macrophage clearance of apoptotic or antibody-opsonized target cells

    International Nuclear Information System (INIS)

    Witasp, Erika; Kupferschmidt, Natalia; Bengtsson, Linnea; Hultenby, Kjell; Smedman, Christian; Paulie, Staffan; Garcia-Bennett, Alfonso E.; Fadeel, Bengt

    2009-01-01

    Macrophage recognition and ingestion of apoptotic cell corpses, a process referred to as programmed cell clearance, is of considerable importance for the maintenance of tissue homeostasis and in the resolution of inflammation. Moreover, macrophages are the first line of defense against microorganisms and other foreign materials including particles. However, there is sparse information on the mode of uptake of engineered nanomaterials by primary macrophages. In this study, mesoporous silica particles with cubic pore geometries and covalently fluorescein-grafted particles were synthesized through a novel route, and their interactions with primary human monocyte-derived macrophages were assessed. Efficient and active internalization of mesoporous silica particles of different sizes was observed by transmission electron microscopic and flow cytometric analysis and studies using pharmacological inhibitors suggested that uptake occurred through a process of endocytosis. Moreover, uptake of silica particles was independent of serum factors. The silica particles with very high surface areas due to their porous structure did not impair cell viability or function of macrophages, including the ingestion of different classes of apoptotic or opsonized target cells. The current findings are relevant to the development of mesoporous materials for drug delivery and other biomedical applications.

  15. 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.

  16. S-carboxymethylcysteine inhibits adherence of Streptococcus pneumoniae to human alveolar epithelial cells.

    Science.gov (United States)

    Sumitomo, Tomoko; Nakata, Masanobu; Yamaguchi, Masaya; Terao, Yutaka; Kawabata, Shigetada

    2012-01-01

    Streptococcus pneumoniae is a major pathogen of respiratory infections that utilizes platelet-activating factor receptor (PAFR) for firm adherence to host cells. The mucolytic agent S-carboxymethylcysteine (S-CMC) has been shown to exert inhibitory effects against infection by several respiratory pathogens including S. pneumoniae in vitro and in vivo. Moreover, clinical studies have implicated the benefits of S-CMC in preventing exacerbation of chronic obstructive pulmonary disease, which is considered to be related to respiratory infections. In this study, to assess whether the potency of S-CMC is attributable to inhibition of pneumococcal adherence to host cells, an alveolar epithelial cell line stimulated with interleukin-1α was used as a model of inflamed epithelial cells. Despite upregulation of PAFR by inflammatory activation, treatment with S-CMC efficiently inhibited pneumococcal adherence to host epithelial cells. In order to gain insight into the inhibitory mechanism, the effects of S-CMC on PAFR expression were also investigated. Following treatment with S-CMC, PAFR expression was reduced at both mRNA and post-transcriptional levels. Interestingly, S-CMC was also effective in inhibiting pneumococcal adherence to cells transfected with PAFR small interfering RNAs. These results indicate S-CMC as a probable inhibitor targeting numerous epithelial receptors that interact with S. pneumoniae.

  17. Effect of laser phototherapy on human alveolar bone repair: micro tomographic and histomorphometrical analysis

    Science.gov (United States)

    Romão, Marcia M. A.; Marques, Márcia M.; Cortes, Arthur R. G.; Horliana, Anna C. R. T.; Moreira, Maria S.; Lascala, Cesar A.

    2015-06-01

    The immediate dental implant placement in the molars region is critical, because of the high amount of bone loss and the discrepancy between the alveolar crest thickness and the dental implant platform. Laser phototherapy (LPT) improves bone repair thus could accelerate the implant placement. Twenty patients were selected for the study. Ten patients were submitted to LPT with GaAlAs diode laser (808nm) during molar extraction, immediately after, 24h, 48h, 72h, 96h and 7 days. The irradiations were applied in contact and punctual mode (100mW, 0.04cm2, 0.75J/cm2, 30s per point, 3J per point). The control group (n=10) received the same treatment; however with the power of the laser off. Forty days later samples of the tissue formed inside the sockets were obtained for further microtomography (microCTs) and histomorphometry analyses. Data were compared by the Student t test, whereas those from the different microCT parameters were compared by the Pearson correlation test (pPearson correlation test it was possible to infer that the lased group presented a more homogeneous trabecular configuration, which would allow earlier dental implant placement.

  18. Ocimum basilicum ethanolic extract decreases cholesterol synthesis and lipid accumulation in human macrophages.

    Science.gov (United States)

    Bravo, Elena; Amrani, Souliman; Aziz, Mohammed; Harnafi, Hicham; Napolitano, Mariarosaria

    2008-12-01

    Macrophage lipid accumulation induced by low density lipoproteins (LDL) plays a pivotal role in atherosclerotic plaque development. Previous work showed that Ocimum basilicum extract, used as hypocholesterolemic agent by traditional medicine in Morocco, has hypolipidemic activity in rat acute hyperlipimidemia. This study investigated the effects of ethanolic extract of O. basilicum on lipid accumulation in human macrophages. As modification of LDL increase atherogenicity of the particles we evaluated the effects of the extract on LDL oxidation. The extract caused a dose-related increase of LDL-resistance to Cu(2+)-induced oxidation. Furthermore, at the dose of 60 microg/ml, significantly decreases the accumulation of macrophage lipid droplets induced by modified LDL evaluated as by red-oil staining. Cholesterol esterification and triacylglycerol synthesis in the cells were not affected. Macrophage treatment with 60 microg/ml, but not 20 microg/ml, of the extract reduced newly synthesized unesterified cholesterol by about 60% and decreased scavenger receptors activity by about 20-30%, evaluated by the internalization of cholesterol carried by [(3)H]CE-aggregated-LDL. The results suggest that O. basilicum ethanolic extract has the capability to reduce foam cell formation through the reduction of cholesterol synthesis and the modulation of the activity of surface scavenger receptors.

  19. Exosomes derived from human macrophages suppress endothelial cell migration by controlling integrin trafficking.

    Science.gov (United States)

    Lee, Hee Doo; Kim, Yeon Hyang; Kim, Doo-Sik

    2014-04-01

    Integrin trafficking, including internalization, recycling, and lysosomal degradation, is crucial for the regulation of cellular functions. Exosomes, nano-sized extracellular vesicles, are believed to play important roles in intercellular communications. This study demonstrates that exosomes released from human macrophages negatively regulate endothelial cell migration through control of integrin trafficking. Macrophage-derived exosomes promote internalization of integrin β1 in primary HUVECs. The internalized integrin β1 persistently accumulates in the perinuclear region and is not recycled back to the plasma membrane. Experimental results indicate that macrophage-derived exosomes stimulate trafficking of internalized integrin β1 to lysosomal compartments with a corresponding decrease in the integrin destined for recycling endosomes, resulting in proteolytic degradation of the integrin. Moreover, ubiquitination of HUVEC integrin β1 is enhanced by the exosomes, and exosome-mediated integrin degradation is blocked by bafilomycin A, a lysosomal degradation inhibitor. Macrophage-derived exosomes were also shown to effectively suppress collagen-induced activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase signaling pathway and HUVEC migration, which are both dependent on integrin β1. These observations provide new insight into the functional significance of exosomes in the regulation of integrin trafficking. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Zinc and zinc transporters in macrophages and their roles in efferocytosis in COPD.

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    Rhys Hamon

    Full Text Available Our previous studies have shown that nutritional zinc restriction exacerbates airway inflammation accompanied by an increase in caspase-3 activation and an accumulation of apoptotic epithelial cells in the bronchioles of the mice. Normally, apoptotic cells are rapidly cleared by macrophage efferocytosis, limiting any secondary necrosis and inflammation. We therefore hypothesized that zinc deficiency is not only pro-apoptotic but also impairs macrophage efferocytosis. Impaired efferocytic clearance of apoptotic epithelial cells by alveolar macrophages occurs in chronic obstructive pulmonary disease (COPD, cigarette-smoking and other lung inflammatory diseases. We now show that zinc is a factor in impaired macrophage efferocytosis in COPD. Concentrations of zinc were significantly reduced in the supernatant of bronchoalveolar lavage fluid of patients with COPD who were current smokers, compared to healthy controls, smokers or COPD patients not actively smoking. Lavage zinc was positively correlated with AM efferocytosis and there was decreased efferocytosis in macrophages depleted of Zn in vitro by treatment with the membrane-permeable zinc chelator TPEN. Organ and cell Zn homeostasis are mediated by two families of membrane ZIP and ZnT proteins. Macrophages of mice null for ZIP1 had significantly lower intracellular zinc and efferocytosis capability, suggesting ZIP1 may play an important role. We investigated further using the human THP-1 derived macrophage cell line, with and without zinc chelation by TPEN to mimic zinc deficiency. There was no change in ZIP1 mRNA levels by TPEN but a significant 3-fold increase in expression of another influx transporter ZIP2, consistent with a role for ZIP2 in maintaining macrophage Zn levels. Both ZIP1 and ZIP2 proteins were localized to the plasma membrane and cytoplasm in normal human lung alveolar macrophages. We propose that zinc homeostasis in macrophages involves the coordinated action of ZIP1 and ZIP2

  1. Influence of ER leak on resting cytoplasmic Ca2+ and receptor-mediated Ca2+ signalling in human macrophage.

    Science.gov (United States)

    Layhadi, Janice A; Fountain, Samuel J

    2017-06-03

    Mechanisms controlling endoplasmic reticulum (ER) Ca 2+ homeostasis are important regulators of resting cytoplasmic Ca 2+ concentration ([Ca 2+ ] cyto ) and receptor-mediated Ca 2+ signalling. Here we investigate channels responsible for ER Ca 2+ leak in THP-1 macrophage and human primary macrophage. In the absence of extracellular Ca 2+ we employ ionomycin action at the plasma membrane to stimulate ER Ca 2+ leak. Under these conditions ionomycin elevates [Ca 2+ ] cyto revealing a Ca 2+ leak response which is abolished by thapsigargin. IP 3 receptors (Xestospongin C, 2-APB), ryanodine receptors (dantrolene), and translocon (anisomycin) inhibition facilitated ER Ca 2+ leak in model macrophage, with translocon inhibition also reducing resting [Ca 2+ ] cyto . In primary macrophage, translocon inhibition blocks Ca 2+ leak but does not influence resting [Ca 2+ ] cyto . We identify a role for translocon-mediated ER Ca 2+ leak in receptor-mediated Ca 2+ signalling in both model and primary human macrophage, whereby the Ca 2+ response to ADP (P2Y receptor agonist) is augmented following anisomycin treatment. In conclusion, we demonstrate a role of ER Ca 2+ leak via the translocon in controlling resting cytoplasmic Ca 2+ in model macrophage and receptor-mediated Ca 2+ signalling in model macrophage and primary macrophage. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Unlike PPARγ, PPARα or PPARβ/δ activation does not promote human monocyte differentiation toward alternative macrophages

    International Nuclear Information System (INIS)

    Bouhlel, Mohamed Amine; Brozek, John; Derudas, Bruno; Zawadzki, Christophe; Jude, Brigitte; Staels, Bart; Chinetti-Gbaguidi, Giulia

    2009-01-01

    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 PPARγ promotes the differentiation of monocytes into anti-inflammatory M2 macrophages in humans and mice, while a role for PPARβ/δ in this process has been reported only in mice and no data are available for PPARα. Here, we show that in contrast to PPARγ, expression of PPARα and PPARβ/δ 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 PPARγ, PPARα or PPARβ/δ activation does not influence human monocyte differentiation into M2 macrophages in vitro. Thus, PPARα and PPARβ/δ do not appear to modulate the alternative differentiation of human macrophages.

  3. Amiodarone increases the accumulation of DEA in a human alveolar epithelium-derived cell line.

    Science.gov (United States)

    Seki, Satoru; Itagaki, Shirou; Kobayashi, Masaki; Hirano, Takeshi; Iseki, Ken

    2008-07-01

    Amiodarone (AMD)-induced pulmonary toxicity (AIPT) is the most life-threatening side-effect of AMD treatment. N-Monodesethylamiodarone (DEA), an active metabolite of AMD, also exhibits cytotoxicity and tends to accumulate in the lung more intensively than AMD. In this study, we characterized the mechanism of DEA accumulation using A549 cells as a model of the alveolar epithelium. Typical ATP-depletion compounds caused an approximately 30% increase in the accumulation of DEA in A549 cells, although these effects were less than those in Caco-2 cells. Triiodothyronine (T(3)), which exhibited an inhibitory effect on DEA efflux in Caco-2 cells, did not affect the accumulation of DEA in A549 cells. On the other hand, 100 microM AMD caused an approximately 200% increase in DEA content in A549 cells, although AMD accumulation was not affected by 100 microM DEA. Since the reducing effect of AMD on cellular ATP levels and that of FCCP were similar, the mechanism by which DEA accumulation is increased by AMD might be different from the ATP-dependent DEA efflux mechanism. The decrease in cell viability by DEA in the presence of AMD (IC(50) value of DEA for A549 cell viability: 25.4+/-2.4 microM) was more pronounced than that by DEA alone (IC(50) value: 11.5+/-3.0 microM). This further DEA accumulation by AMD might be a factor responsible for the greater accumulation of DEA than that of AMD in the lung in long-term AMD-treated patients.

  4. Anti-Inflammatory Effects of Lactobacillus Rahmnosus and Bifidobacterium Breve on Cigarette Smoke Activated Human Macrophages.

    Science.gov (United States)

    Mortaz, Esmaeil; Adcock, Ian M; Ricciardolo, Fabio L M; Varahram, Mohammad; Jamaati, Hamidreza; Velayati, Ali Akbar; Folkerts, Gert; Garssen, Johan

    2015-01-01

    Chronic obstructive pulmonary disease (COPD) is a major global health problem with cigarette smoke (CS) as the main risk factor for its development. Airway inflammation in COPD involves the increased expression of inflammatory mediators such as CXCL-8 and IL-1β which are important mediators for neutrophil recruitment. Macrophages are an important source of these mediators in COPD. Lactobacillus rhamnosus (L. rhamnosus) and Befidobacterium breve (B. breve) attenuate the development of 'allergic asthma' in animals but their effects in COPD are unknown. To determine the anti-inflammatory effects of L. rhamnosus and B. breve on CS and Toll-like receptor (TLR) activation. We stimulated the human macrophage cell line THP-1 with CS extract in the presence and absence of L. rhamnosus and B. breve and measured the expression and release of inflammatory mediators by RT-qPCR and ELISA respectively. An activity assay and Western blotting were used to examine NF-κB activation. Both L. rhamnosus and B. breve were efficiently phagocytized by human macrophages. L. rhamnosus and B. breve significantly suppressed the ability of CS to induce the expression of IL-1β, IL-6, IL-10, IL-23, TNFα, CXCL-8 and HMGB1 release (all p<0.05) in human THP-1 macrophages. Similar suppression of TLR4- and TLR9-induced CXCL8 expression was also observed (p<0.05). The effect of L. rhamnosus and B. breve on inflammatory mediator release was associated with the suppression of CS-induced NF-κB activation (p<0.05). This data indicate that these probiotics may be useful anti-inflammatory agents in CS-associated disease such as COPD.

  5. Anti-Inflammatory Effects of Lactobacillus Rahmnosus and Bifidobacterium Breve on Cigarette Smoke Activated Human Macrophages.

    Directory of Open Access Journals (Sweden)

    Esmaeil Mortaz

    Full Text Available Chronic obstructive pulmonary disease (COPD is a major global health problem with cigarette smoke (CS as the main risk factor for its development. Airway inflammation in COPD involves the increased expression of inflammatory mediators such as CXCL-8 and IL-1β which are important mediators for neutrophil recruitment. Macrophages are an important source of these mediators in COPD. Lactobacillus rhamnosus (L. rhamnosus and Befidobacterium breve (B. breve attenuate the development of 'allergic asthma' in animals but their effects in COPD are unknown.To determine the anti-inflammatory effects of L. rhamnosus and B. breve on CS and Toll-like receptor (TLR activation.We stimulated the human macrophage cell line THP-1 with CS extract in the presence and absence of L. rhamnosus and B. breve and measured the expression and release of inflammatory mediators by RT-qPCR and ELISA respectively. An activity assay and Western blotting were used to examine NF-κB activation.Both L. rhamnosus and B. breve were efficiently phagocytized by human macrophages. L. rhamnosus and B. breve significantly suppressed the ability of CS to induce the expression of IL-1β, IL-6, IL-10, IL-23, TNFα, CXCL-8 and HMGB1 release (all p<0.05 in human THP-1 macrophages. Similar suppression of TLR4- and TLR9-induced CXCL8 expression was also observed (p<0.05. The effect of L. rhamnosus and B. breve on inflammatory mediator release was associated with the suppression of CS-induced NF-κB activation (p<0.05.This data indicate that these probiotics may be useful anti-inflammatory agents in CS-associated disease such as COPD.

  6. Human macrophage scavenger receptors: Primary structure, expression, and localization in atherosclerotic lesions

    International Nuclear Information System (INIS)

    Matsumoto, Akiyo; Itakura, Hiroshige; Kodama, Tatsuhiko; Naito, Makoto; Takahashi, Kiyoshi; Ikemoto, Shinji; Asaoka, Hitoshi; Hayakawa, Ikuho; Kanamori, Hiroshi; Takaku, Fumimaro; Aburatani, Hiroyuki; Suzuki, Hiroshi; Kobari, Yukage; Miyai, Tatsuya; Cohen, E.H.; Wydro, R.; Housman, D.E.

    1990-01-01

    Two types of cDNAs for human macrophage scavenger receptors were cloned from a cDNA library derived from the phorbol ester-treated human monocytic cell line THP-1. The type I and type II human scavenger receptors encoded by these cDNAs are homologous (73% and 71% amino acid identity) to their previously characterized bovine counterparts and consist of six domains: cytoplasmic (I), membrane-spanning (II), spacer (III), α-helical coiled-coil (IV), collagen-like (V), and a type-specific C-terminal (VI). The receptor gene is located on human chromosome 8. The human receptors expressed in CHO-K1 cells mediated endocytosis of modified low density lipoproteins. Two mRNAs, 4.0 and 3.2 kilobases, have been detected in human liver, placenta, and brain. Immunohistochemical studies using an anti-peptide antibody which recognizes human scavenger receptors indicated the presence of the scavenger receptors in the macrophages of lipid-rich atherosclerotic lesions, suggesting the involvement of scavenger receptors in atherogenesis

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

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    Weibin Zha

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

  8. Matrix metalloproteinase-12 gene regulation by a PPAR alpha agonist in human monocyte-derived macrophages

    International Nuclear Information System (INIS)

    Souissi, Imen Jguirim; Billiet, Ludivine; Cuaz-Perolin, Clarisse; Slimane, Mohamed-Naceur; Rouis, Mustapha

    2008-01-01

    MMP-12, a macrophage-specific matrix metalloproteinase with large substrate specificity, has been reported to be highly expressed in mice, rabbits and human atherosclerotic lesions. Increased MMP-12 from inflammatory macrophages is associated with several degenerative diseases such as atherosclerosis. In this manuscript, we show that IL-1β, a proinflammatory cytokine found in atherosclerotic plaques, increases both mRNA and protein levels of MMP-12 in human monocyte-derived macrophages (HMDM). Since peroxisome proliferator-activated receptors (PPARs), such as PPARα and PPARγ, are expressed in macrophages and because PPAR activation exerts an anti-inflammatory effect on vascular cells, we have investigated the effect of PPARα and γ isoforms on MMP-12 regulation in HMDM. Our results show that MMP-12 expression (mRNA and protein) is down regulated in IL-1β-treated macrophages only in the presence of a specific PPARα agonist, GW647, in a dose-dependent manner. In contrast, this inhibitory effect was abolished in IL-1β-stimulated peritoneal macrophages isolated from PPARα -/- mice and treated with the PPARα agonist, GW647. Moreover, reporter gene transfection experiments using different MMP-12 promoter constructs showed a reduction of the promoter activities by ∼ 50% in IL-1β-stimulated PPARα-pre-treated cells. However, MMP-12 promoter analysis did not reveal the presence of a PPRE response element. The IL-1β effect is known to be mediated through the AP-1 binding site. Mutation of the AP-1 site, located at - 81 in the MMP-12 promoter region relative to the transcription start site, followed by transfection analysis, gel shift and ChIP experiments revealed that the inhibitory effect was the consequence of the protein-protein interaction between GW 647-activated PPARα and c-Fos or c-Jun transcription factors, leading to inhibition of their binding to the AP-1 motif. These studies suggest that PPARα agonists may be used therapeutically, not only for lipid

  9. CCR2 and CXCR3 agonistic chemokines are differently expressed and regulated in human alveolar epithelial cells type II

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    Prasse Antje

    2005-07-01

    Full Text Available Abstract The attraction of leukocytes from circulation to inflamed lungs depends on the activation of both the leukocytes and the resident cells within the lung. In this study we determined gene expression and secretion patterns for monocyte chemoattractant protein-1 (MCP-1/CCL2 and T-cell specific CXCR3 agonistic chemokines (Mig/CXCL9, IP-10/CXCL10, and I-TAC/CXCL11 in TNF-α-, IFN-γ-, and IL-1β-stimulated human alveolar epithelial cells type II (AEC-II. AEC-II constitutively expressed high level of CCL2 mRNA in vitro and in situ , and released CCL2 protein in vitro . Treatment of AEC-II with proinflammatory cytokines up-regulated both CCL2 mRNA expression and release of immunoreactive CCL2, whereas IFN-γ had no effect on CCL2 release. In contrast, CXCR3 agonistic chemokines were not detected in freshly isolated AEC-II or in non-stimulated epithelial like cell line A549. IFN-γ, alone or in combination with IL-1β and TNF-α resulted in an increase in CXCL10, CXCL11, and CXCL9 mRNA expression and generation of CXCL10 protein by AEC-II or A549 cells. CXCL10 gene expression and secretion were induced in dose-dependent manner after cytokine-stimulation of AEC-II with an order of potency IFN-γ>>IL-1β ≥ TNF-α. Additionally, we localized the CCL2 and CXCL10 mRNAs in human lung tissue explants by in situ hybridization, and demonstrated the selective effects of cytokines and dexamethasone on CCL2 and CXCL10 expression. These data suggest that the regulation of the CCL2 and CXCL10 expression exhibit significant differences in their mechanisms, and also demonstrate that the alveolar epithelium contributes to the cytokine milieu of the lung, with the ability to respond to locally generated cytokines and to produce potent mediators of the local inflammatory response.

  10. 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.

  11. Cytoskeletal stability and metabolic alterations in primary human macrophages in long-term microgravity.

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    Svantje Tauber

    Full Text Available The immune system is one of the most affected systems of the human body during space flight. The cells of the immune system are exceptionally sensitive to microgravity. Thus, serious concerns arise, whether space flight associated weakening of the immune system ultimately precludes the expansion of human presence beyond the Earth's orbit. For human space flight, it is an urgent need to understand the cellular and molecular mechanisms by which altered gravity influences and changes the functions of immune cells. The CELLBOX-PRIME (= CellBox-Primary Human Macrophages in Microgravity Environment experiment investigated for the first time microgravity-associated long-term alterations in primary human macrophages, one of the most important effector cells of the immune system. The experiment was conducted in the U.S. National Laboratory on board of the International Space Station ISS using the NanoRacks laboratory and Biorack type I standard CELLBOX EUE type IV containers. Upload and download were performed with the SpaceX CRS-3 and the Dragon spaceship on April 18th, 2014 / May 18th, 2014. Surprisingly, primary human macrophages exhibited neither quantitative nor structural changes of the actin and vimentin cytoskeleton after 11 days in microgravity when compared to 1g controls. Neither CD18 or CD14 surface expression were altered in microgravity, however ICAM-1 expression was reduced. The analysis of 74 metabolites in the cell culture supernatant by GC-TOF-MS, revealed eight metabolites with significantly different quantities when compared to 1g controls. In particular, the significant increase of free fucose in the cell culture supernatant was associated with a significant decrease of cell surface-bound fucose. The reduced ICAM-1 expression and the loss of cell surface-bound fucose may contribute to functional impairments, e.g. the activation of T cells, migration and activation of the innate immune response. We assume that the surprisingly small

  12. Evaluation of a nanotechnology-based approach to induce gene-expression in human THP-1 macrophages under inflammatory conditions.

    Science.gov (United States)

    Bernal, Laura; Alvarado-Vázquez, Abigail; Ferreira, David Wilson; Paige, Candler A; Ulecia-Morón, Cristina; Hill, Bailey; Caesar, Marina; Romero-Sandoval, E Alfonso

    2017-02-01

    Macrophages orchestrate the initiation and resolution of inflammation by producing pro- and anti-inflammatory products. An imbalance in these mediators may originate from a deficient or excessive immune response. Therefore, macrophages are valid therapeutic targets to restore homeostasis under inflammatory conditions. We hypothesize that a specific mannosylated nanoparticle effectively induces gene expression in human macrophages under inflammatory conditions without undesirable immunogenic responses. THP-1 macrophages were challenged with lipopolysaccharide (LPS, 5μg/mL). Polyethylenimine (PEI) nanoparticles grafted with a mannose receptor ligand (Man-PEI) were used as a gene delivery method. Nanoparticle toxicity, Man-PEI cellular uptake rate and gene induction efficiency (GFP, CD14 or CD68) were studied. Potential immunogenic responses were evaluated by measuring the production of tumor necrosis factor-alpha (TNF-α), Interleukin (IL)-6 and IL-10. Man-PEI did not produce cytotoxicity, and it was effectively up-taken by THP-1 macrophages (69%). This approach produced a significant expression of GFP (mRNA and protein), CD14 and CD68 (mRNA), and transiently and mildly reduced IL-6 and IL-10 levels in LPS-challenged macrophages. Our results indicate that Man-PEI is suitable for inducing an efficient gene overexpression in human macrophages under inflammatory conditions with limited immunogenic responses. Our promising results set the foundation to test this technology to induce functional anti-inflammatory genes. Copyright © 2016 Elsevier GmbH. All rights reserved.

  13. Enhanced uptake of multiple sclerosis-derived myelin by THP-1 macrophages and primary human microglia.

    Science.gov (United States)

    Hendrickx, Debbie A E; Schuurman, Karianne G; van Draanen, Michael; Hamann, Jörg; Huitinga, Inge

    2014-03-31

    The pathological hallmark of multiple sclerosis (MS) is myelin phagocytosis. It remains unclear why microglia and macrophages demyelinate axons in MS, but previously found or yet-unknown changes in the myelin of MS patients could contribute to this process. We therefore studied whether myelin from normal-appearing white matter (NAWM) of MS donors is phagocytosed more efficiently than myelin from control donors. Myelin was isolated from 11 MS and 12 control brain donors and labeled with the pH-sensitive fluorescent dye pHrodo to quantify uptake in lysosomes. Phagocytosis by differentiated THP-1 macrophages and by primary human microglia was quantified with flow cytometry. Whereas myelin uptake by THP-1 macrophages reached a plateau after approximately 24 hours, uptake by primary human microglia showed an almost linear increase over a 72-hour period. Data were statistically analyzed with the Mann-Whitney U test. MS-derived myelin was phagocytosed more efficiently by THP-1 macrophages after 6-hour incubation (P = 0.001 for the percentage of myelin-phagocytosing cells and P = 0.0005 for total myelin uptake) and after 24-hour incubation (P = 0.0006 and P = 0.0001, respectively), and by microglia after 24-hour incubation (P = 0.0106 for total myelin uptake). This enhanced uptake was not due to differences in the oxidation status of the myelin. Interestingly, myelin phagocytosis correlated negatively with the age of myelin donors, whereas the age of microglia donors showed a positive trend with myelin phagocytosis. Myelin isolated from normal-appearing white matter of MS donors was phagocytosed more efficiently than was myelin isolated from control brain donors by both THP-1 macrophages and primary human microglia. These data indicate that changes in MS myelin might precede phagocyte activation and subsequent demyelination in MS. Identifying these myelin changes responsible for enhancing phagocytic ability could be an interesting therapeutic target to

  14. Disrupted epithelial/macrophage crosstalk via Spinster homologue 2-mediated S1P signaling may drive defective macrophage phagocytic function in COPD.

    Directory of Open Access Journals (Sweden)

    Hai B Tran

    Full Text Available We have previously established a link between impaired phagocytic capacity and deregulated S1P signaling in alveolar macrophages from COPD subjects. We hypothesize that this defect may include a disruption of epithelial-macrophage crosstalk via Spns2-mediated intercellular S1P signaling.Primary alveolar macrophages and bronchial epithelial cells from COPD subjects and controls, cell lines, and a mouse model of chronic cigarette smoke exposure were studied. Cells were exposed to 10% cigarette smoke extract, or vehicle control. Spns2 expression and subcellular localization was studied by immunofluorescence, confocal microscopy and RT-PCR. Phagocytosis was assessed by flow-cytometry. Levels of intra- and extracellular S1P were measured by S1P [3H]-labeling.Spns2 expression was significantly increased (p<0.05 in alveolar macrophages from current-smokers/COPD patients (n = 5 compared to healthy nonsmokers (n = 8 and non-smoker lung transplant patients (n = 4. Consistent with this finding, cigarette smoke induced a significant increase in Spns2 expression in both human alveolar and THP-1 macrophages. In contrast, a remarkable Spns2 down-regulation was noted in response to cigarette smoke in 16HBE14o- cell line (p<0.001 in 3 experiments, primary nasal epithelial cells (p<0.01 in 2 experiments, and in smoke-exposed mice (p<0.001, n = 6 animals per group. Spns2 was localized to cilia in primary bronchial epithelial cells. In both macrophage and epithelial cell types, Spns2 was also found localized to cytoplasm and the nucleus, in line with a predicted bipartile Nuclear Localization Signal at the position aa282 of the human Spns2 sequence. In smoke-exposed mice, alveolar macrophage phagocytic function positively correlated with Spns2 protein expression in bronchial epithelial cells.Our data suggest that the epithelium may be the major source for extracellular S1P in the airway and that there is a possible disruption of epithelial/macrophage cross talk via

  15. Human alveolar bone cell proliferation, expression of osteoblastic phenotype, and matrix mineralization on porous titanium produced by powder metallurgy.

    Science.gov (United States)

    Rosa, Adalberto Luiz; Crippa, Grasiele Edilaine; de Oliveira, Paulo Tambasco; Taba, Mario; Lefebvre, Louis-Philippe; Beloti, Marcio Mateus

    2009-05-01

    This study aimed at investigating the influence of the porous titanium (Ti) structure on the osteogenic cell behaviour. Porous Ti discs were fabricated by the powder metallurgy process with the pore size typically between 50 and 400 microm and a porosity of 60%. Osteogenic cells obtained from human alveolar bone were cultured until subconfluence and subcultured on dense Ti (control) and porous Ti for periods of up to 17 days. Cultures grown on porous Ti exhibited increased cell proliferation and total protein content, and lower levels of alkaline phosphatase (ALP) activity than on dense Ti. In general, gene expression of osteoblastic markers-runt-related transcription factor 2, collagen type I, alkaline phosphatase, bone morphogenetic protein-7, and osteocalcin was lower at day 7 and higher at day 17 in cultures grown on porous Ti compared with dense Ti, a finding consistent with the enhanced growth rate for such cultures. The amount of mineralized matrix was greater on porous Ti compared with the dense one. These results indicate that the porous Ti is an appropriate substrate for osteogenic cell adhesion, proliferation, and production of a mineralized matrix. Because of the three-dimensional environment it provides, porous Ti should be considered an advantageous substrate for promoting desirable implant surface-bone interactions.

  16. Apoptotic death of Listeria monocytogenes-infected human macrophages induced by lactoferricin B, a bovine lactoferrin-derived peptide.

    Science.gov (United States)

    Longhi, C; Conte, M P; Ranaldi, S; Penta, M; Valenti, P; Tinari, A; Superti, F; Seganti, L

    2005-01-01

    Listeria monocytogenes, an intracellular facultative food-borne pathogen, was reported to induce apoptosis in vitro and in vivo in a variety of cell types with the exception of murine macrophages. These cells represent the predominant compartment of bacterial multiplication and die as a result of necrosis. In this study we showed that human non-activated and IFN-gamma-activated macrophagic-like (THP-1) cells infected with L. monocytogenes, mainly die by necrosis rather than by an apoptotic process. Two natural products derived from bovine milk, lactoferrin and its derivative peptide lactoferricin B, are capable of regulating the fate of infected human macrophages. Bovine lactoferrin treatment of macrophages protects them from L. monocytogenes-induced death whereas lactoferricin B, its derivative peptide, determines a shifting of the equilibrium from necrosis to apoptosis.

  17. Human mesenchymal stromal cell-secreted lactate induces M2-macrophage differentiation by metabolic reprogramming

    Science.gov (United States)

    Civini, Sara; Pacelli, Consiglia; Dieng, Mame Massar; Lemieux, William; Jin, Ping; Bazin, Renée; Patey, Natacha; Marincola, Francesco M.; Moldovan, Florina; Zaouter, Charlotte; Trudeau, Louis-Eric; Benabdhalla, Basma; Louis, Isabelle; Beauséjour, Christian; Stroncek, David; Le Deist, Françoise; Haddad, Elie

    2016-01-01

    Human mesenchymal stromal cells (MSC) have been shown to dampen immune response and promote tissue repair, but the underlying mechanisms are still under investigation. Herein, we demonstrate that umbilical cord-derived MSC (UC-MSC) alter the phenotype and function of monocyte-derived dendritic cells (DC) through lactate-mediated metabolic reprogramming. UC-MSC can secrete large quantities of lactate and, when present during monocyte-to-DC differentiation, induce instead the acquisition of M2-macrophage features in terms of morphology, surface markers, migratory properties and antigen presentation capacity. Microarray expression profiling indicates that UC-MSC modify the expression of metabolic-related genes and induce a M2-macrophage expression signature. Importantly, monocyte-derived DC obtained in presence of UC-MSC, polarize naïve allogeneic CD4+ T-cells into Th2 cells. Treatment of UC-MSC with an inhibitor of lactate dehydrogenase strongly decreases lactate concentration in culture supernatant and abrogates the effect on monocyte-to-DC differentiation. Metabolic analysis further revealed that UC-MSC decrease oxidative phosphorylation in differentiating monocytes while strongly increasing the spare respiratory capacity proportional to the amount of secreted lactate. Because both MSC and monocytes are recruited in vivo at the site of tissue damage and inflammation, we propose the local increase of lactate concentration induced by UC-MSC and the consequent enrichment in M2-macrophage generation as a mechanism to achieve immunomodulation. PMID:27070086

  18. Interaction of differentiated human adipocytes with macrophages leads to trogocytosis and selective IL-6 secretion.

    Science.gov (United States)

    Sárvári, A K; Doan-Xuan, Q-M; Bacsó, Z; Csomós, I; Balajthy, Z; Fésüs, L

    2015-01-22

    Obesity leads to adipose tissue inflammation that is characterized by increased release of proinflammatory molecules and the recruitment of activated immune cells. Although macrophages are present in the highest number among the immune cells in obese adipose tissue, not much is known about their direct interaction with adipocytes. We have introduced an ex vivo experimental system to characterize the cellular interactions and the profile of secreted cytokines in cocultures of macrophages and human adipocytes differentiated from either mesenchymal stem cells or a preadipocyte cell line. As observed by time-lapse microscopy, flow, and laser-scanning cytometry, macrophages phagocytosed bites of adipocytes (trogocytosis), which led to their de novo, phagocytosis and NF-κB-dependent synthesis, then release of interleukin (IL)-6 and monocyte chemoattractant protein (MCP)-1. IL-6 secretion was not accompanied by secretion of other proinflammatory cytokines, such as tumor necrosis factor (TNF)-α and IL-8, except MCP-1. LPS-induced release of TNF-α, IL-8 and MCP-1 was decreased in the presence of the differentiated adipocytes but the IL-6 level did not subside suggesting that phagocytosis-dependent IL-6 secretion may have significant regulatory function in the inflamed adipose tissue.

  19. Quantification of sterol-specific response in human macrophages using automated imaged-based analysis.

    Science.gov (United States)

    Gater, Deborah L; Widatalla, Namareq; Islam, Kinza; AlRaeesi, Maryam; Teo, Jeremy C M; Pearson, Yanthe E

    2017-12-13

    The transformation of normal macrophage cells into lipid-laden foam cells is an important step in the progression of atherosclerosis. One major contributor to foam cell formation in vivo is the intracellular accumulation of cholesterol. Here, we report the effects of various combinations of low-density lipoprotein, sterols, lipids and other factors on human macrophages, using an automated image analysis program to quantitatively compare single cell properties, such as cell size and lipid content, in different conditions. We observed that the addition of cholesterol caused an increase in average cell lipid content across a range of conditions. All of the sterol-lipid mixtures examined were capable of inducing increases in average cell lipid content, with variations in the distribution of the response, in cytotoxicity and in how the sterol-lipid combination interacted with other activating factors. For example, cholesterol and lipopolysaccharide acted synergistically to increase cell lipid content while also increasing cell survival compared with the addition of lipopolysaccharide alone. Additionally, ergosterol and cholesteryl hemisuccinate caused similar increases in lipid content but also exhibited considerably greater cytotoxicity than cholesterol. The use of automated image analysis enables us to assess not only changes in average cell size and content, but also to rapidly and automatically compare population distributions based on simple fluorescence images. Our observations add to increasing understanding of the complex and multifactorial nature of foam-cell formation and provide a novel approach to assessing the heterogeneity of macrophage response to a variety of factors.

  20. Tfe3 expression is closely associated to macrophage terminal differentiation of human hematopoietic myeloid precursors

    International Nuclear Information System (INIS)

    Zanocco-Marani, Tommaso; Vignudelli, Tatiana; Gemelli, Claudia; Pirondi, Sara; Testa, Anna; Montanari, Monica; Parenti, Sandra; Tenedini, Elena; Grande, Alexis; Ferrari, Sergio

    2006-01-01

    The MItf-Tfe family of basic helix-loop-helix leucine zipper (bHLH-Zip) transcription factors encodes four family members: MItf, Tfe3, TfeB and TfeC. In vitro, each protein of the family binds DNA in a homo- or heterodimeric form with other family members. Tfe3 is involved in chromosomal translocations recurrent in different tumors and it has been demonstrated, by in vivo studies, that it plays, redundantly with MItf, an important role in the process of osteoclast formation, in particular during the transition from mono-nucleated to multi-nucleated osteoclasts. Since mono-nucleated osteoclasts derive from macrophages we investigated whether Tfe3 might play a role upstream during hematopoietic differentiation. Here we show that Tfe3 is able to induce mono-macrophagic differentiation of U937 cells, in association with a decrease of cell proliferation and an increase of apoptosis. We also show that Tfe3 does not act physiologically during commitment of CD34+ hematopoietic stem cells (HSCs), since it is not able to direct HSCs toward a specific lineage as observed by clonogenic assay, but is a strong actor of terminal differentiation since it allows human primary myeloblasts' maturation toward the macrophage lineage

  1. Imatinib and Nilotinib Off-Target Effects on Human NK Cells, Monocytes, and M2 Macrophages.

    Science.gov (United States)

    Bellora, Francesca; Dondero, Alessandra; Corrias, Maria Valeria; Casu, Beatrice; Regis, Stefano; Caliendo, Fabio; Moretta, Alessandro; Cazzola, Mario; Elena, Chiara; Vinti, Luciana; Locatelli, Franco; Bottino, Cristina; Castriconi, Roberta

    2017-08-15

    Tyrosine kinase inhibitors (TKIs) are used in the clinical management of hematological neoplasms. Moreover, in solid tumors such as stage 4 neuroblastomas (NB), imatinib showed benefits that might depend on both on-target and immunological off-target effects. We investigated the effects of imatinib and nilotinib on human NK cells, monocytes, and macrophages. High numbers of monocytes died upon exposure to TKI concentrations similar to those achieved in patients. Conversely, NK cells were highly resistant to the TKI cytotoxic effect, were properly activated by immunostimulatory cytokines, and degranulated in the presence of NB cells. In NB, neither drug reduced the expression of ligands for activating NK receptors or upregulated that of HLA class I, B7-H3, PD-L1, and PD-L2, molecules that might limit NK cell function. Interestingly, TKIs modulated the chemokine receptor repertoire of immune cells. Acting at the transcriptional level, they increased the surface expression of CXCR4, an effect observed also in NK cells and monocytes of patients receiving imatinib for chronic myeloid leukemia. Moreover, TKIs reduced the expression of CXCR3 (in NK cells) and CCR1 (in monocytes). Monocytes also decreased the expression of M-CSFR, and low numbers of cells underwent differentiation toward macrophages. M0 and M2 macrophages were highly resistant to TKIs and maintained their phenotypic and functional characteristics. Importantly, also in the presence of TKIs, the M2 immunosuppressive polarization was reverted by TLR engagement, and M1-oriented macrophages fully activated autologous NK cells. Our results contribute to better interpreting the off-target efficacy of TKIs in tumors and to envisaging strategies aimed at facilitating antitumor immune responses. Copyright © 2017 by The American Association of Immunologists, Inc.

  2. Viral Inhibition of Bacterial Phagocytosis by Human Macrophages: Redundant Role of CD36.

    Directory of Open Access Journals (Sweden)

    Grace E Cooper

    Full Text Available Macrophages are essential to maintaining lung homoeostasis and recent work has demonstrated that influenza-infected lung macrophages downregulate their expression of the scavenger receptor CD36. This receptor has also been shown to be involved in phagocytosis of Streptococcus pneumoniae, a primary agent associated with pneumonia secondary to viral infection. The aim of this study was to investigate the role of CD36 in the effects of viral infection on macrophage phagocytic function. Human monocyte-derived macrophages (MDM were exposed to H3N2 X31 influenza virus, M37 respiratory syncytial virus (RSV or UV-irradiated virus. No infection of MDM was seen upon exposure to UV-irradiated virus but incubation with live X31 or M37 resulted in significant levels of viral detection by flow cytometry or RT-PCR respectively. Infection resulted in significantly diminished uptake of S. pneumoniae by MDM and significantly decreased expression of CD36 at both the cell surface and mRNA level. Concurrently, there was a significant increase in IFNβ gene expression in response to infection and we observed a significant decrease in bacterial phagocytosis (p = 0.031 and CD36 gene expression (p = 0.031 by MDM cultured for 24 h in 50IU/ml IFNβ. Knockdown of CD36 by siRNA resulted in decreased phagocytosis, but this was mimicked by transfection reagent alone. When MDM were incubated with CD36 blocking antibodies no effect on phagocytic ability was observed. These data indicate that autologous IFNβ production by virally-infected cells can inhibit bacterial phagocytosis, but that decreased CD36 expression by these cells does not play a major role in this functional deficiency.

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

    Science.gov (United States)

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

    2017-07-18

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

  4. Mycobacterium tuberculosis decreases human macrophage IFN-γ responsiveness through miR-132 and miR-26a.

    Science.gov (United States)

    Ni, Bin; Rajaram, Murugesan V S; Lafuse, William P; Landes, Michelle B; Schlesinger, Larry S

    2014-11-01

    IFN-γ-activated macrophages play an essential role in controlling intracellular pathogens; however, macrophages also serve as the cellular home for the intracellular pathogen Mycobacterium tuberculosis. Based on previous evidence that M. tuberculosis can modulate host microRNA (miRNA) expression, we examined the miRNA expression profile of M. tuberculosis-infected primary human macrophages. We identified 31 differentially expressed miRNAs in primary human macrophages during M. tuberculosis infection by NanoString and confirmed our findings by quantitative real-time RT-PCR. In addition, we determined a role for two miRNAs upregulated upon M. tuberculosis infection, miR-132 and miR-26a, as negative regulators of transcriptional coactivator p300, a component of the IFN-γ signaling cascade. Knockdown expression of miR-132 and miR-26a increased p300 protein levels and improved transcriptional, translational, and functional responses to IFN-γ in human macrophages. Collectively, these data validate p300 as a target of miR-132 and miR-26a, and demonstrate a mechanism by which M. tuberculosis can limit macrophage responses to IFN-γ by altering host miRNA expression. Copyright © 2014 by The American Association of Immunologists, Inc.

  5. Osteogenesis differentiation of human periodontal ligament cells by CO2 laser-treatment stimulating macrophages via BMP2 signalling pathway

    International Nuclear Information System (INIS)

    Hsieh, Wen-Hui; Chen, Yi-Jyun; Hung, Chi-Jr; Huang, Tsui-Hsien; Kao, Chia-Tze; Shie, Ming-You

    2014-01-01

    Immune reactions play an important role in determining the biostimulation of bone formation, either in new bone formation or inflammatory fibrous tissue encapsulation. Macrophage cell, the important effector cells in the immune reaction, which are indispensable for osteogenesis and their heterogeneity and plasticity, render macrophages a primer target for immune system modulation. However, there are very few studies about the effects of macrophage cells on laser treatment-regulated osteogenesis. In this study, we used CO 2 laser as a model biostimulation to investigate the role of macrophage cells on the CO 2 laser stimulated osteogenesis. Bone morphogenetic protein 2 (BMP2) was also significantly up regulated by the CO 2 laser stimulation, indicating that macrophage may participate in the CO 2 laser stimulated osteogenesis. Interestingly, when laser treatment macrophage-conditioned medium were applied to human periodontal ligament cells (hPDLs), the osteogenesis differentiation of hPDLs was significantly enhanced, indicating the important role of macrophages in CO 2 laser-induced osteogenesis. These findings provided valuable insights into the mechanism of CO 2 laser-stimulated osteogenic differentiation, and a strategy to optimize the evaluation system for the in vitro osteogenesis capacity of laser treatment. (paper)

  6. Concentrations in plasma, epithelial lining fluid, alveolar macrophages and bronchial mucosa after a single intravenous dose of 1.6 mg/kg of iclaprim (AR-100) in healthy men.

    Science.gov (United States)

    Andrews, J; Honeybourne, D; Ashby, J; Jevons, G; Fraise, A; Fry, P; Warrington, S; Hawser, S; Wise, R

    2007-09-01

    A validated microbiological assay was used to measure concentrations of iclaprim (AR-100) in plasma, bronchial mucosa (BM), alveolar macrophages (AM) and epithelial lining fluid (ELF) after a single 1.6 mg/kg intravenous 60 min iv infusion of iclaprim. Male volunteers were randomly allocated to three nominal sampling time intervals 1-2 h (Group A), 3-4 h (Group B) and 5.5-7.0 h (Group C) after the start of the drug infusion. Mean iclaprim concentrations in plasma, BM, AM and ELF, respectively, were for Group A 0.59 mg/L (SD 0.18), 0.51 mg/kg (SD 0.17), 24.51 mg/L (SD 21.22) and 12.61 mg/L (SD 7.33); Group B 0.24 mg/L (SD 0.05), 0.35 mg/kg (SD 0.17), 7.16 mg/L (SD 1.91) and 6.38 mg/L (SD 5.17); and Group C 0.14 mg/L (SD 0.05), no detectable level in BM, 5.28 mg/L (SD 2.30) and 2.66 mg/L (SD 2.08). Iclaprim concentrations in ELF and AM exceeded the MIC(90) for penicillin-susceptible Streptococcus pneumoniae (MIC90 0.06 mg/L), penicillin-intermediate S. pneumoniae (MIC90 2 mg/L), penicillin-resistant S. pneumoniae (MIC90 4 mg/L) for 7, 7 and 4 h, respectively, and Chlamydia pneumoniae (MIC90 0.5 mg/L) for 7 h. Mean iclaprim concentrations in ELF exceeded the MIC90 for Haemophilus influenzae (MIC90 4 mg/L) and Moraxella catarrhalis (MIC90 8 mg/L) for up to 4 and 2 h, respectively; in AM the MIC90 was exceeded for up to 7 h. Furthermore, the MIC90 for methicillin-resistant Staphylococcus aureus of 0.12 mg/L was exceeded at all sites for up to 7 h. These data suggest that iclaprim reaches lung concentrations that should be effective in the treatment of community-acquired pneumonia.

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

    Directory of Open Access Journals (Sweden)

    Stephen H-F Macdonald

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

  8. CCL20 and Beta-Defensin 2 Production by Human Lung Epithelial Cells and Macrophages in Response to Brucella abortus Infection

    Science.gov (United States)

    Fernández, Andrea G.; Bonetto, Josefina; Giambartolomei, Guillermo H.; Fossati, Carlos A.; Baldi, Pablo C.

    2015-01-01

    Both CCL20 and human β-defensin 2 (hBD2) interact with the same membrane receptor and display chemotactic and antimicrobial activities. They are produced by airway epithelia in response to infectious agents and proinflammatory cytokines. Whereas Brucella spp. can infect humans through inhalation, their ability to induce CCL20 and hBD2 in lung cells is unknown. Here we show that B. abortus induces CCL20 expression in human alveolar (A549) or bronchial (Calu-6) epithelial cell lines, primary alveolar epithelial cells, primary human monocytes, monocyte-derived macrophages and the monocytic cell line THP-1. CCL20 expression was mainly mediated by JNK1/2 and NF-kB in both Calu-6 and THP-1 cells. CCL20 secretion was markedly induced in A549, Calu-6 and THP-1 cells by heat-killed B. abortus or a model Brucella lipoprotein (L-Omp19) but not by the B. abortus lipopolysaccharide (LPS). Accordingly, CCL20 production by B. abortus-infected cells was strongly TLR2-dependent. Whereas hBD2 expression was not induced by B. abortus infection, it was significantly induced in A549 cells by conditioned media from B. abortus-infected THP-1 monocytes (CMB). A similar inducing effect was observed on CCL20 secretion. Experiments using blocking agents revealed that IL-1β, but not TNF-α, was involved in the induction of hBD2 and CCL20 secretion by CMB. In the in vitro antimicrobial assay, the lethal dose (LD) 50 of CCL20 for B. abortus (>50 μg/ml) was markedly higher than that against E. coli (1.5 μg/ml) or a B. abortus mutant lacking the O polysaccharide in its LPS (8.7 ug/ml). hBD2 did not kill any of the B. abortus strains at the tested concentrations. These results show that human lung epithelial cells secrete CCL20 and hBD2 in response to B. abortus and/or to cytokines produced by infected monocytes. Whereas these molecules do not seem to exert antimicrobial activity against this pathogen, they could recruit immune cells to the infection site. PMID:26448160

  9. Soluble immune complexes shift the TLR-induced cytokine production of distinct polarized human macrophage subsets towards IL-10.

    Directory of Open Access Journals (Sweden)

    Carmen A Ambarus

    Full Text Available BACKGROUND: Costimulation of murine macrophages with immune complexes (ICs and TLR ligands leads to alternative activation. Studies on human myeloid cells, however, indicate that ICs induce an increased pro-inflammatory cytokine production. This study aimed to clarify the effect of ICs on the pro- versus anti-inflammatory profile of human polarized macrophages. MATERIALS AND METHODS: Monocytes isolated from peripheral blood of healthy donors were polarized for four days with IFN-γ, IL-4, IL-10, GM-CSF, M-CSF, or LPS, in the presence or absence of heat aggregated gamma-globulins (HAGGs. Phenotypic polarization markers were measured by flow cytometry. Polarized macrophages were stimulated with HAGGs or immobilized IgG alone or in combination with TLR ligands. TNF, IL-6, IL-10, IL-12, and IL-23 were measured by Luminex and/or RT-qPCR. RESULTS: HAGGs did not modulate the phenotypic polarization and the cytokine production of macrophages. However, HAGGs significantly altered the TLR-induced cytokine production of all polarized macrophage subsets, with the exception of MΦ(IL-4. In particular, HAGGs consistently enhanced the TLR-induced IL-10 production in both classically and alternatively polarized macrophages (M1 and M2. The effect of HAGGs on TNF and IL-6 production was less pronounced and depended on the polarization status, while IL-23p19 and IL-12p35 expression was not affected. In contrast with HAGGs, immobilized IgG induced a strong upregulation of not only IL-10, but also TNF and IL-6. CONCLUSION: HAGGs alone do not alter the phenotype and cytokine production of in vitro polarized human macrophages. In combination with TLR-ligands, however, HAGGs but not immobilized IgG shift the cytokine production of distinct macrophage subsets toward IL-10.

  10. Alveolar recruitment of ficolin-3 in response to acute pulmonary inflammation in humans

    DEFF Research Database (Denmark)

    Plovsing, Ronni R; Berg, Ronan M G; Munthe-Fog, Lea

    2016-01-01

    acute lung and systemic inflammation induce recruitment of lectins in humans. METHODS: Fifteen healthy volunteers received LPS intravenously (IV) or in a lung subsegment on two different occasions. Volunteers were evaluated by consecutive blood samples and by bronchoalveolar lavage 2, 4, 6, 8, or 24h...... acute phase response with an increase in CRP (precruitment...

  11. First-in-human study and clinical case reports of the alveolar bone regeneration with the secretome from human mesenchymal stem cells.

    Science.gov (United States)

    Katagiri, Wataru; Osugi, Masashi; Kawai, Takamasa; Hibi, Hideharu

    2016-01-15

    Secreted growth factors and cytokines in the conditioned medium from bone marrow-derived mesenchymal stem cells (MSC-CM) have several effects on cell behavior. Our previous studies revealed that MSC-CM enhances bone regeneration by increasing cell mobilization, angiogenesis, and osteogenesis in vitro and in vivo. This clinical study was undertaken to evaluate the safety and use of MSC-CM for alveolar bone regeneration in eight patients who were diagnosed as needing bone augmentation prior to dental implant placement. The protocol of this clinical study was approved by the ethics committee of Nagoya University Hospital. MSC-CM was prepared from conditioned medium from commercially available human bone marrow-derived MSCs. Patients were treated with beta-tricalcium phosphate (β-TCP) or an atelocollagen sponge soaked with MSC-CM. Clinical and radiographic assessments were performed during the follow-up period. Histological assessments were also performed in some cases. Clinical and histological data from patients who underwent the SFE procedure without MSC-CM were also used retrospectively as reference controls. MSC-CM contained several cytokines such as insulin-like growth factor-1, vascular endothelial growth factor, transforming growth factor-β1, and hepatocyte growth factor in relatively low amounts. No systemic or local complications were reported throughout the study. Radiographic evaluation revealed early bone formation in all cases. Histological evaluation also supported the radiographic findings. Furthermore, infiltration of inflammatory cells was scarce throughout the specimens. MSC-CM was used safely and with less inflammatory signs and appears to have great osteogenic potential for regenerative medicine of bone. This is the first in-human clinical study of alveolar bone regeneration using MSC-CM.

  12. Ebola virion attachment and entry into human macrophages profoundly effects early cellular gene expression.

    Directory of Open Access Journals (Sweden)

    Victoria Wahl-Jensen

    2011-10-01

    Full Text Available Zaire ebolavirus (ZEBOV infections are associated with high lethality in primates. ZEBOV primarily targets mononuclear phagocytes, which are activated upon infection and secrete mediators believed to trigger initial stages of pathogenesis. The characterization of the responses of target cells to ZEBOV infection may therefore not only further understanding of pathogenesis but also suggest possible points of therapeutic intervention. Gene expression profiles of primary human macrophages exposed to ZEBOV were determined using DNA microarrays and quantitative PCR to gain insight into the cellular response immediately after cell entry. Significant changes in mRNA concentrations encoding for 88 cellular proteins were observed. Most of these proteins have not yet been implicated in ZEBOV infection. Some, however, are inflammatory mediators known to be elevated during the acute phase of disease in the blood of ZEBOV-infected humans. Interestingly, the cellular response occurred within the first hour of Ebola virion exposure, i.e. prior to virus gene expression. This observation supports the hypothesis that virion binding or entry mediated by the spike glycoprotein (GP(1,2 is the primary stimulus for an initial response. Indeed, ZEBOV virions, LPS, and virus-like particles consisting of only the ZEBOV matrix protein VP40 and GP(1,2 (VLP(VP40-GP triggered comparable responses in macrophages, including pro-inflammatory and pro-apoptotic signals. In contrast, VLP(VP40 (particles lacking GP(1,2 caused an aberrant response. This suggests that GP(1,2 binding to macrophages plays an important role in the immediate cellular response.

  13. Triglyceride-rich lipoprotein regulates APOB48 receptor gene expression in human THP-1 monocytes and macrophages.

    Science.gov (United States)

    Bermudez, Beatriz; Lopez, Sergio; Varela, Lourdes M; Ortega, Almudena; Pacheco, Yolanda M; Moreda, Wenceslao; Moreno-Luna, Rafael; Abia, Rocio; Muriana, Francisco J G

    2012-02-01

    The postprandial metabolism of dietary fats implies that the production of TG-rich lipoproteins (TRL) contributes to the progression of plaque development. TRL and their remnants cause rapid receptor-mediated monocyte/macrophage lipid engorgement via the cell surface apoB48 receptor (apoB48R). However, the mechanistic basis for apoB48 receptor (APOB48R) regulation by postprandial TRL in monocytes and macrophages is not well established. In this study, we investigated the effects of postprandial TRL from healthy volunteers on the expression of APOB48R mRNA and lipid uptake in human THP-1 monocytes and THP-1-derived macrophages. The expression of APOB48R mRNA was upregulated in THP-1 monocytes, but downregulated in THP-1-derived macrophages when treated with postprandial TRL (P < 0.05), in a dose- and time-dependent manner. TG and free cholesterol were dramatically increased in THP-1-derived macrophages (140 and 50%, respectively; P < 0.05) and in THP-1 monocytes (160 and 95%, respectively; P < 0.05). This lipid accumulation was severely decreased (~50%; P < 0.05) in THP-1-derived macrophages by small interfering RNA (siRNA) targeting of APOB48R. Using PPAR and retinoid X receptor (RXR) agonists, antagonists, and siRNA, our data indicate that PPARα, PPARγ, and RXRα are involved in postprandial TRL-induced APOB48R transcriptional regulation. Co-incubation with acyl-CoA synthetase or acyl-CoA:cholesterol acyltransferase inhibitors potentiated the effects of postprandial TRL on the expression of APOB48R mRNA in THP-1 monocytes and THP-1-derived macrophages. Our findings collectively suggest that APOB48R represents a molecular target of postprandial TRL via PPAR-dependent pathways in human THP-1 monocytes and macrophages and advance a potentially important link between postprandial metabolism of dietary fats and atherogenesis.

  14. Toxicity of silver nanoparticles in human macrophages: uptake, intracellular distribution and cellular responses

    Science.gov (United States)

    Haase, A.; Tentschert, J.; Jungnickel, H.; Graf, P.; Mantion, A.; Draude, F.; Plendl, J.; Goetz, M. E.; Galla, S.; Mašić, A.; Thuenemann, A. F.; Taubert, A.; Arlinghaus, H. F.; Luch, A.

    2011-07-01

    Silver nanoparticles (SNP) are among the most commercialized nanoparticles worldwide. They can be found in many diverse products, mostly because of their antibacterial properties. Despite its widespread use only little data on possible adverse health effects exist. It is difficult to compare biological data from different studies due to the great variety in sizes, coatings or shapes of the particles. Here, we applied a novel synthesis approach to obtain SNP, which are covalently stabilized by a small peptide. This enables a tight control of both size and shape. We applied these SNP in two different sizes of 20 or 40 nm (Ag20Pep and Ag40Pep) and analyzed responses of THP-1-derived human macrophages. Similar gold nanoparticles with the same coating (Au20Pep) were used for comparison and found to be non-toxic. We assessed the cytotoxicity of particles and confirmed their cellular uptake via transmission electron microscopy and confocal Raman microscopy. Importantly a majority of the SNP could be detected as individual particles spread throughout the cells. Furthermore we studied several types of oxidative stress related responses such as induction of heme oxygenase I or formation of protein carbonyls. In summary, our data demonstrate that even low doses of SNP exerted adverse effects in human macrophages.

  15. Toxicity of silver nanoparticles in human macrophages: uptake, intracellular distribution and cellular responses

    International Nuclear Information System (INIS)

    Haase, A; Tentschert, J; Jungnickel, H; Goetz, M E; Luch, A; Graf, P; Mantion, A; Thuenemann, A F; Draude, F; Galla, S; Arlinghaus, H F; Plendl, J; Masic, A; Taubert, A

    2011-01-01

    Silver nanoparticles (SNP) are among the most commercialized nanoparticles worldwide. They can be found in many diverse products, mostly because of their antibacterial properties. Despite its widespread use only little data on possible adverse health effects exist. It is difficult to compare biological data from different studies due to the great variety in sizes, coatings or shapes of the particles. Here, we applied a novel synthesis approach to obtain SNP, which are covalently stabilized by a small peptide. This enables a tight control of both size and shape. We applied these SNP in two different sizes of 20 or 40 nm (Ag20Pep and Ag40Pep) and analyzed responses of THP-1-derived human macrophages. Similar gold nanoparticles with the same coating (Au20Pep) were used for comparison and found to be non-toxic. We assessed the cytotoxicity of particles and confirmed their cellular uptake via transmission electron microscopy and confocal Raman microscopy. Importantly a majority of the SNP could be detected as individual particles spread throughout the cells. Furthermore we studied several types of oxidative stress related responses such as induction of heme oxygenase I or formation of protein carbonyls. In summary, our data demonstrate that even low doses of SNP exerted adverse effects in human macrophages.

  16. Inhibition of transglutaminase 2 reduces efferocytosis in human macrophages: Role of CD14 and SR-AI receptors.

    Science.gov (United States)

    Eligini, S; Fiorelli, S; Tremoli, E; Colli, S

    2016-10-01

    Transglutaminase 2 (TGM2), a member of the transglutaminase family of enzymes, is a multifunctional protein involved in numerous events spanning from cell differentiation, to signal transduction, apoptosis, and wound healing. It is expressed in a variety of cells, macrophages included. Macrophage TGM2 promotes the clearance of apoptotic cells (efferocytosis) and emerging evidence suggests that defective efferocytosis contributes to the consequences of inflammation-associated diseases, including atherosclerotic lesion progression and its sequelae. Of interest, active TGM2 identified in human atherosclerotic lesions plays critical roles in plaque stability through effects on matrix cross-linking and TGFβ activity. This study explores the mechanisms by which TGM2 controls efferocytosis in human macrophages. Herein we show that TGM2 increases progressively during monocyte differentiation towards macrophages and controls their efferocytic potential as well as morphology and viability. Two experimental approaches that took advantage of the inhibition of TGM2 activity and protein silencing give proof that TGM2 reduction significantly impairs macrophage efferocytosis. Among the mechanisms involved we highlighted a role of the receptors CD14 and SR-AI whose levels were markedly reduced by TGM2 inhibition. Conversely, CD36 receptor and αvβ3 integrin levels were not influenced. Of note, lipid accumulation and IL-10 secretion were reduced in macrophages displaying defective efferocytosis. Overall, our data define a crucial role of TGM2 activity during macrophage differentiation via mechanisms involving CD14 and SR-AI receptors and show that TGM2 inhibition triggers a pro-inflammatory phenotype. Copyright © 2016 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

  17. Macrophage immunoregulatory pathways in tuberculosis.

    Science.gov (United States)

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

    2014-12-01

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

  18. Human Subcutaneous Tissue Response to Glucose Sensors: Macrophages Accumulation Impact on Sensor Accuracy.

    Science.gov (United States)

    Rigla, Mercedes; Pons, Belén; Rebasa, Pere; Luna, Alexis; Pozo, Francisco Javier; Caixàs, Assumpta; Villaplana, Maria; Subías, David; Bella, Maria Rosa; Combalia, Neus

    2018-04-01

    Subcutaneous (s.c.) glucose sensors have become a key component in type 1 diabetes management. However, their usability is limited by the impact of foreign body response (FBR) on their duration, reliability, and accuracy. Our study gives the first description of human acute and subacute s.c. response to glucose sensors, showing the changes observed in the sensor surface, the inflammatory cells involved in the FBR and their relationship with sensor performance. Twelve obese patients (seven type 2 diabetes) underwent two abdominal biopsies comprising the surrounding area where they had worn two glucose sensors: the first one inserted 7 days before and the second one 24 h before biopsy procedure. Samples were processed and studied to describe tissue changes by two independent pathologists (blind regarding sensor duration). Macrophages quantification was studied by immunohistochemistry methods in the area surrounding the sensor (CD68, CD163). Sensor surface changes were studied by scanning electron microscopy. Seven-day continuous glucose monitoring records were considered inaccurate when mean absolute relative difference was higher than 10%. Pathologists were able to correctly classify all the biopsies regarding sensor duration. Acute response (24 h) was characterized by the presence of neutrophils while macrophages were the main cell involved in subacute inflammation. The number of macrophages around the insertion hole was higher for less accurate sensors compared with those performing more accurately (32.6 ± 14 vs. 10.6 ± 1 cells/0.01 mm 2 ; P sensor-tissue interface is related with decrease in accuracy of the glucose measure.

  19. Pharmacological inhibition of dynamin II reduces constitutive protein secretion from primary human macrophages.

    Directory of Open Access Journals (Sweden)

    Maaike Kockx

    Full Text Available Dynamins are fission proteins that mediate endocytic and exocytic membrane events and are pharmacological therapeutic targets. These studies investigate whether dynamin II regulates constitutive protein secretion and show for the first time that pharmacological inhibition of dynamin decreases secretion of apolipoprotein E (apoE and several other proteins constitutively secreted from primary human macrophages. Inhibitors that target recruitment of dynamin to membranes (MiTMABs or directly target the GTPase domain (Dyngo or Dynole series, dose- and time- dependently reduced the secretion of apoE. SiRNA oligo's targeting all isoforms of dynamin II confirmed the involvement of dynamin II in apoE secretion. Inhibition of secretion was not mediated via effects on mRNA or protein synthesis. 2D-gel electrophoresis showed that inhibition occurred after apoE was processed and glycosylated in the Golgi and live cell imaging showed that inhibited secretion was associated with reduced post-Golgi movement of apoE-GFP-containing vesicles. The effect was not restricted to macrophages, and was not mediated by the effects of the inhibitors on microtubules. Inhibition of dynamin also altered the constitutive secretion of other proteins, decreasing the secretion of fibronectin, matrix metalloproteinase 9, Chitinase-3-like protein 1 and lysozyme but unexpectedly increasing the secretion of the inflammatory mediator cyclophilin A. We conclude that pharmacological inhibitors of dynamin II modulate the constitutive secretion of macrophage apoE as a class effect, and that their capacity to modulate protein secretion may affect a range of biological processes.

  20. Genome-wide analysis reveals loci encoding anti-macrophage factors in the human pathogen Burkholderia pseudomallei K96243.

    Directory of Open Access Journals (Sweden)

    Andrea J Dowling

    2010-12-01

    Full Text Available Burkholderia pseudomallei is an important human pathogen whose infection biology is still poorly understood. The bacterium is endemic to tropical regions, including South East Asia and Northern Australia, where it causes melioidosis, a serious disease associated with both high mortality and antibiotic resistance. B. pseudomallei is a Gram-negative facultative intracellular pathogen that is able to replicate in macrophages. However despite the critical nature of its interaction with macrophages, few anti-macrophage factors have been characterized to date. Here we perform a genome-wide gain of function screen of B. pseudomallei strain K96243 to identify loci encoding factors with anti-macrophage activity. We identify a total of 113 such loci scattered across both chromosomes, with positive gene clusters encoding transporters and secretion systems, enzymes/toxins, secondary metabolite, biofilm, adhesion and signal response related factors. Further phenotypic analysis of four of these regions shows that the encoded factors cause striking cellular phenotypes relevant to infection biology, including apoptosis, formation of actin 'tails' and multi-nucleation within treated macrophages. The detailed analysis of the remaining host of loci will facilitate genetic dissection of the interaction of this important pathogen with host macrophages and thus further elucidate this critical part of its infection cycle.

  1. Differentially activated macrophages orchestrate myogenic precursor cell fate during human skeletal muscle regeneration

    DEFF Research Database (Denmark)

    Saclier, Marielle; Yacoub-Youssef, Houda; Mackey, Abigail

    2013-01-01

    , we explored both in vitro and in vivo, in human, the interactions of differentially activated MPs with myogenic precursor cells (MPCs) during adult myogenesis and skeletal muscle regeneration. We showed in vitro that through the differential secretion of cytokines and growth factors, proinflammatory...... anti-inflammatory markers. These data demonstrate for the first time in human that MPs sequentially orchestrate adult myogenesis during regeneration of damaged skeletal muscle. These results support the emerging concept that inflammation, through MP activation, controls stem cell fate and coordinates......Macrophages (MPs) exert either beneficial or deleterious effects on tissue repair, depending on their activation/polarization state. They are crucial for adult skeletal muscle repair, notably by acting on myogenic precursor cells. However, these interactions have not been fully characterized. Here...

  2. 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...

  3. 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

    assays, CFT073 was able to survive within primary mouse bone marrow-derived macrophages (BMM) up to 24h post-infection. Three additional well-characterized clinical UPEC isolates associated with distinct UTI symptomatologies displayed variable long-term survival within BMM. UPEC strains UTI89 and VR50...... 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...

  4. A familial case of alveolar capillary dysplasia with misalignment of pulmonary veins supports paternal imprinting of FOXF1 in human

    Science.gov (United States)

    Alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV) is a rare developmental lung disorder that is uniformly lethal. Affected infants die within the first few weeks of their life despite aggressive treatment, although a few cases of late manifestation and longer survival have ...

  5. Gene expression profiles of human dendritic cells interacting with Aspergillus fumigatus in a bilayer model of the alveolar epithelium/endothelium interface.

    Directory of Open Access Journals (Sweden)

    Charles Oliver Morton

    Full Text Available The initial stages of the interaction between the host and Aspergillus fumigatus at the alveolar surface of the human lung are critical in the establishment of aspergillosis. Using an in vitro bilayer model of the alveolus, including both the epithelium (human lung adenocarcinoma epithelial cell line, A549 and endothelium (human pulmonary artery epithelial cells, HPAEC on transwell membranes, it was possible to closely replicate the in vivo conditions. Two distinct sub-groups of dendritic cells (DC, monocyte-derived DC (moDC and myeloid DC (mDC, were included in the model to examine immune responses to fungal infection at the alveolar surface. RNA in high quantity and quality was extracted from the cell layers on the transwell membrane to allow gene expression analysis using tailored custom-made microarrays, containing probes for 117 immune-relevant genes. This microarray data indicated minimal induction of immune gene expression in A549 alveolar epithelial cells in response to germ tubes of A. fumigatus. In contrast, the addition of DC to the system greatly increased the number of differentially expressed immune genes. moDC exhibited increased expression of genes including CLEC7A, CD209 and CCL18 in the absence of A. fumigatus compared to mDC. In the presence of A. fumigatus, both DC subgroups exhibited up-regulation of genes identified in previous studies as being associated with the exposure of DC to A. fumigatus and exhibiting chemotactic properties for neutrophils, including CXCL2, CXCL5, CCL20, and IL1B. This model closely approximated the human alveolus allowing for an analysis of the host pathogen interface that complements existing animal models of IA.

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

    International Nuclear Information System (INIS)

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

    1987-01-01

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

  7. Extracts of human atherosclerotic lesions modify LDL inducing enhanced macrophage uptake

    International Nuclear Information System (INIS)

    Hoff, H.F.; O'Neill, J.

    1986-01-01

    Both an LDL-like fraction isolated from human aortic plaques and LDL incubated with cultured aortic endothelial or smooth muscle cells have been shown to be internalized by macrophages in vitro in an unregulated fashion leading to foam cell formation. Lipid peroxidation induced by free radicals released from cells was shown to be responsible for cell-modified LDL. The authors incubated LDL with a supernatant fraction of leached, i.e. non-homogenized, extracts of aortic plaques for one hour at 37 0 C, to determine whether extracellular components present in arteries were also capable of modifying LDL. Extract-treated LDL showed the following changes relative to untreated LDL: 1) increased electrophretic mobility, 2) altered pattern of B-100 on SDS-PAGE, i.e. presence of a doublet with higher M/sub r/ than B-100, and 3) enhanced uptake by cultured mouse peritoneal macrophages as measured by increased degradation of 125 I-LDL, and increased stimulation of cholesterol esterification using 14 C-oleate. Extracts from homogenized plaques and grossly normal intima induced similar changes. The modification was tissue specific in that extracts of arteries but not of liver, muscle or skin modified LDL. Protease degradation of LDL during incubation was probably not responsible since inhibitors did not prevent modification. It is possible that products of lipid peroxidation present in extracellular lipid of arteries may propagate free radicals or be incorporated into LDL, leading to modifications similar to those found in cell-modified LDL

  8. Intramacrophage survival of uropathogenic Escherichia coli: Differences between diverse clinical isolates and between mouse and human macrophages

    KAUST Repository

    Bokil, Nilesh J.

    2011-11-01

    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 pathogens target macrophages, we assessed the interactions between UPEC and macrophages. Colonization of the mouse bladder by UPEC strain CFT073 resulted in increased expression of myeloid-restricted genes, consistent with the recruitment of inflammatory macrophages to the site of infection. In in vitro assays, CFT073 was able to survive within primary mouse bone marrow-derived macrophages (BMM) up to 24h post-infection. Three additional well-characterized clinical UPEC isolates associated with distinct UTI symptomatologies displayed variable long-term survival within BMM. UPEC strains UTI89 and VR50, 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 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 suggest that some UPEC isolates may subvert macrophage anti-microbial pathways, and that host species differences may impact on intracellular UPEC survival. © 2011 Elsevier GmbH.

  9. Th1-like human T-cell clones recognizing Leishmania gp63 inhibit Leishmania major in human macrophages

    DEFF Research Database (Denmark)

    Kemp, M; Hey, A S; Bendtzen, K

    1994-01-01

    The major surface protease of Leishmania major, gp63, has been suggested as a vaccine candidate for cutaneous leishmaniasis. In this study gp63 was purified from L. major promastigotes. A panel of human T-cell clones recognizing this protein were generated from individuals who had previously had...... resembling Th1 cells. Autologous mononuclear cells and Epstein-Barr virus-transformed B cell lines were equally efficient in presenting the antigen to the T cells. The gp63 reactive T cells induced resistance to infection in cultured human macrophages by L. major. The data confirm that human CD4+ T cells...... recognizing gp63 can take part in the host defence against L. major infections....

  10. Decreased expression of liver X receptor-α in macrophages infected with Chlamydia pneumoniae in human atherosclerotic arteries in situ.

    Science.gov (United States)

    Bobryshev, Yuri V; Orekhov, Alexander N; Killingsworth, Murray C; Lu, Jinhua

    2011-01-01

    In in vitro experiments, Chlamydia pneumoniae has been shown to infect macrophages and to accelerate foam cell formation. It has been hypothesized that the C. pneumoniae infection affects foam cell formation by suppressing the expression of liver X receptors (LXR), but whether such an event occurs in human atherosclerosis is not known. In this study we examined carotid artery segments, obtained by endarterectomy, in which the presence of C. pneumoniae was confirmed by both polymerase chain reaction and immunohistochemistry. The expression of LXR-α in macrophages infected with C. pneumoniae and macrophages that were not infected was compared using a quantitative immunohistochemical analysis. The analysis revealed a 2.2-fold reduction in the expression of LXR-α in C. pneumoniae-infected cells around the lipid cores in atherosclerotic plaques. In the cytoplasm of laser-capture microdissected cells that were immunopositive for C. pneumoniae, electron microscopy demonstrated the presence of structures with the appearance of elementary, reticulate and aberrant bodies of C. pneumoniae. We conclude that LXR-α expression is reduced in C. pneumoniae-infected macrophages in human atherosclerotic lesions which supports the hypothesis that C. pneumoniae infection might suppress LXR expression in macrophages transforming into foam cells. Copyright © 2011 S. Karger AG, Basel.

  11. Improved gene expression in resting macrophages using an oligopeptide derived from Vpr of human immunodeficiency virus type-1

    International Nuclear Information System (INIS)

    Mizoguchi, Izuru; Ooe, Yoshihiro; Hoshino, Shigeki; Shimura, Mari; Kasahara, Tadashi; Kano, Shigeyuki; Ohta, Toshiko; Takaku, Fumimaro; Nakayama, Yasuhide; Ishizaka, Yukihito

    2005-01-01

    Vpr, an accessory gene product of human immunodeficiency virus type-1, is thought to transport a viral DNA from the cytoplasm to the nucleus in resting macrophages. Previously, we reported that a peptide encompassing amino acids 52-78 of Vpr (C45D18) promotes the nuclear trafficking of recombinant proteins that are conjugated with C45D18. Here, we present evidence that C45D18, when conjugated with a six-branched cationic polymer of poly(N,N-dimethylaminopropylacrylamide)-block-oligo(4-aminostyrene) (SV: star vector), facilitates gene expression in resting macrophages. Although there was no difference between SV alone and C45D18-SV with respect to gene transduction into growing cells, C45D18-SV resulted in more than 40-fold greater expression of the exogenous gene upon transduction into chemically differentiated macrophages and human quiescent monocyte-derived macrophages. The data suggest that C45D18 contributes to improving the ability of a non-viral vector to transduce macrophages with exogenous genes and we discuss its further application

  12. Preservation Analysis of Macrophage Gene Coexpression Between Human and Mouse Identifies PARK2 as a Genetically Controlled Master Regulator of Oxidative Phosphorylation in Humans

    Directory of Open Access Journals (Sweden)

    Veronica Codoni

    2016-10-01

    Full Text Available Macrophages are key players involved in numerous pathophysiological pathways and an in-depth characterization of their gene regulatory networks can help in better understanding how their dysfunction may impact on human diseases. We here conducted a cross-species network analysis of macrophage gene expression data between human and mouse to identify conserved networks across both species, and assessed whether such networks could reveal new disease-associated regulatory mechanisms. From a sample of 684 individuals processed for genome-wide macrophage gene expression profiling, we identified 27 groups of coexpressed genes (modules. Six modules were found preserved (P < 10−4 in macrophages from 86 mice of the Hybrid Mouse Diversity Panel. One of these modules was significantly [false discovery rate (FDR = 8.9 × 10−11] enriched for genes belonging to the oxidative phosphorylation (OXPHOS pathway. This pathway was also found significantly (FDR < 10−4 enriched in susceptibility genes for Alzheimer, Parkinson, and Huntington diseases. We further conducted an expression quantitative trait loci analysis to identify SNP that could regulate macrophage OXPHOS gene expression in humans. This analysis identified the PARK2 rs192804963 as a trans-acting variant influencing (minimal P-value = 4.3 × 10−8 the expression of most OXPHOS genes in humans. Further experimental work demonstrated that PARK2 knockdown expression was associated with increased OXPHOS gene expression in THP1 human macrophages. This work provided strong new evidence that PARK2 participates to the regulatory networks associated with oxidative phosphorylation and suggested that PARK2 genetic variations could act as a trans regulator of OXPHOS gene macrophage expression in humans.

  13. Volcanic ash activates the NLRP3 inflammasome in murine and human macrophages

    Science.gov (United States)

    Damby, David; Horwell, Claire J.; Baxter, Peter J.; Kueppers, Ulrich; Schnurr, Max; Dingwell, Donald B.; Duewell, Peter

    2018-01-01

    Volcanic ash is a heterogeneous mineral dust that is typically composed of a mixture of amorphous (glass) and crystalline (mineral) fragments. It commonly contains an abundance of the crystalline silica (SiO2) polymorph cristobalite. Inhalation of crystalline silica can induce inflammation by stimulating the NLRP3 inflammasome, a cytosolic receptor complex that plays a critical role in driving inflammatory immune responses. Ingested material results in the assembly of NLRP3, ASC, and caspase-1 with subsequent secretion of the interleukin-1 family cytokine IL-1β. Previous toxicology work suggests that cristobalite-bearing volcanic ash is minimally reactive, calling into question the reactivity of volcanically derived crystalline silica, in general. In this study, we target the NLRP3 inflammasome as a crystalline silica responsive element to clarify volcanic cristobalite reactivity. We expose immortalized bone marrow-derived macrophages of genetically engineered mice and primary human peripheral blood mononuclear cells (PBMCs) to ash from the Soufrière Hills volcano as well as representative, pure-phase samples of its primary componentry (volcanic glass, feldspar, cristobalite) and measure NLRP3 inflammasome activation. We demonstrate that respirable Soufrière Hills volcanic ash induces the activation of caspase-1 with subsequent release of mature IL-1β in a NLRP3 inflammasome-dependent manner. Macrophages deficient in NLRP3 inflammasome components are incapable of secreting IL-1β in response to volcanic ash ingestion. Cellular uptake induces lysosomal destabilization involving cysteine proteases. Furthermore, the response involves activation of mitochondrial stress pathways leading to the generation of reactive oxygen species. Considering ash componentry, cristobalite is the most reactive pure-phase with other components inducing only low-level IL-1β secretion. Inflammasome activation mediated by inhaled ash and its potential relevance in chronic pulmonary

  14. [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.

  15. Human mesenchymal stem cells alter macrophage phenotype and promote regeneration via homing to the kidney following ischemia-reperfusion injury

    NARCIS (Netherlands)

    Wise, Andrea F; Williams, Timothy M; Kiewiet, Mensiena B G; Payne, Natalie L; Siatskas, Christopher; Samuel, Chrishan S; Ricardo, Sharon D

    2014-01-01

    Mesenchymal stem cells (MSCs) ameliorate injury and accelerate repair in many organs, including the kidney, although the reparative mechanisms and interaction with macrophages have not been elucidated. This study investigated the reparative potential of human bone marrow-derived MSCs and traced

  16. Organic UV filters exposure induces the production of inflammatory cytokines in human macrophages.

    Science.gov (United States)

    Ao, Junjie; Yuan, Tao; Gao, Li; Yu, Xiaodan; Zhao, Xiaodong; Tian, Ying; Ding, Wenjin; Ma, Yuning; Shen, Zhemin

    2018-09-01

    Organic ultraviolet (UV) filters, found in many personal care products, are considered emerging contaminants due to growing concerns about potential long-term deleterious effects. We investigated the immunomodulatory effects of four commonly used organic UV filters (2-hydroxy-4-methoxybenzophenone, BP-3; 4-methylbenzylidene camphor, 4-MBC; 2-ethylhexyl 4-methoxycinnamate, EHMC; and butyl-methoxydibenzoylmethane, BDM) on human macrophages. Our results indicated that exposure to these four UV filters significantly increased the production of various inflammatory cytokines in macrophages, particular tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). After exposure to the UV filters, a significant 1.1-1.5 fold increase were found in TNF-α and IL-6 mRNA expression. In addition, both the p38 MAPK and the NF-κB signaling pathways were enhanced 2 to 10 times in terms of phosphorylation after exposure to the UV filters, suggesting that these pathways are involved in the release of TNF-α and IL-6. Molecular docking analysis predicted that all four UV filter molecules would efficiently bind transforming growth factor beta-activated kinase 1 (TAK1), which is responsible for the activation of the p38 MAPK and NF-κB pathways. Our results therefore demonstrate that exposure to the four organic UV filters investigated may alter human immune system function. It provides new clue for the development of asthma or allergic diseases in terms of the environmental pollutants. Copyright © 2018 Elsevier B.V. All rights reserved.

  17. Involvement of macrophage migration inhibitory factor and its receptor (CD74) in human breast cancer.

    Science.gov (United States)

    Richard, Vincent; Kindt, Nadège; Decaestecker, Christine; Gabius, Hans-Joachim; Laurent, Guy; Noël, Jean-Christophe; Saussez, Sven

    2014-08-01

    Macrophage migration inhibitory factor (MIF) and its receptor CD74 appear to be involved in tumorigenesis. We evaluated, by immunohistochemical staining, the tissue expression and distribution of MIF and CD74 in serial sections of human invasive breast cancer tumor specimens. The serum MIF level was also determined in breast cancer patients. We showed a significant increase in serum MIF average levels in breast cancer patients compared to healthy individuals. MIF tissue expression, quantified by a modified Allred score, was strongly increased in carcinoma compared to tumor-free specimens, in the cancer cells and in the peritumoral stroma, with fibroblasts the most intensely stained. We did not find any significant correlation with histoprognostic factors, except for a significant inverse correlation between tumor size and MIF stromal positivity. CD74 staining was heterogeneous and significantly decreased in cancer cells but increased in the surrounding stroma, namely in lymphocytes, macrophages and vessel endothelium. There was no significant variation according to classical histoprognostic factors, except that CD74 stromal expression was significantly correlated with triple-negative receptor (TRN) status and the absence of estrogen receptors. In conclusion, our data support the concept of a functional role of MIF in human breast cancer. In addition to auto- and paracrine effects on cancer cells, MIF could contribute to shape the tumor microenvironment leading to immunomodulation and angiogenesis. Interfering with MIF effects in breast tumors in a therapeutic perspective remains an attractive but complex challenge. Level of co-expression of MIF and CD74 could be a surrogate marker for efficacy of anti-angiogenic drugs, particularly in TRN breast cancer tumor.

  18. Responses of murine and human macrophages to leptospiral infection: a study using comparative array analysis.

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    Feng Xue

    Full Text Available Leptospirosis is a re-emerging tropical infectious disease caused by pathogenic Leptospira spp. The different host innate immune responses are partially related to the different severities of leptospirosis. In this study, we employed transcriptomics and cytokine arrays to comparatively calculate the responses of murine peritoneal macrophages (MPMs and human peripheral blood monocytes (HBMs to leptospiral infection. We uncovered a series of different expression profiles of these two immune cells. The percentages of regulated genes in several biological processes of MPMs, such as antigen processing and presentation, membrane potential regulation, and the innate immune response, etc., were much greater than those of HBMs (>2-fold. In MPMs and HBMs, the caspase-8 and Fas-associated protein with death domain (FADD-like apoptosis regulator genes were significantly up-regulated, which supported previous results that the caspase-8 and caspase-3 pathways play an important role in macrophage apoptosis during leptospiral infection. In addition, the key component of the complement pathway, C3, was only up-regulated in MPMs. Furthermore, several cytokines, e.g. interleukin 10 (IL-10 and tumor necrosis factor alpha (TNF-alpha, were differentially expressed at both mRNA and protein levels in MPMs and HBMs. Some of the differential expressions were proved to be pathogenic Leptospira-specific regulations at mRNA level or protein level. Though it is still unclear why some animals are resistant and others are susceptible to leptospiral infection, this comparative study based on transcriptomics and cytokine arrays partially uncovered the differences of murine resistance and human susceptibility to leptospirosis. Taken together, these findings will facilitate further molecular studies on the innate immune response to leptospiral infection.

  19. Responses of Murine and Human Macrophages to Leptospiral Infection: A Study Using Comparative Array Analysis

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    Yang, Yingchao; Zhao, Jinping; Yang, Yutao; Cao, Yongguo; Hong, Cailing; Liu, Yuan; Sun, Lan; Huang, Minjun; Gu, Junchao

    2013-01-01

    Leptospirosis is a re-emerging tropical infectious disease caused by pathogenic Leptospira spp. The different host innate immune responses are partially related to the different severities of leptospirosis. In this study, we employed transcriptomics and cytokine arrays to comparatively calculate the responses of murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) to leptospiral infection. We uncovered a series of different expression profiles of these two immune cells. The percentages of regulated genes in several biological processes of MPMs, such as antigen processing and presentation, membrane potential regulation, and the innate immune response, etc., were much greater than those of HBMs (>2-fold). In MPMs and HBMs, the caspase-8 and Fas-associated protein with death domain (FADD)-like apoptosis regulator genes were significantly up-regulated, which supported previous results that the caspase-8 and caspase-3 pathways play an important role in macrophage apoptosis during leptospiral infection. In addition, the key component of the complement pathway, C3, was only up-regulated in MPMs. Furthermore, several cytokines, e.g. interleukin 10 (IL-10) and tumor necrosis factor alpha (TNF-alpha), were differentially expressed at both mRNA and protein levels in MPMs and HBMs. Some of the differential expressions were proved to be pathogenic Leptospira-specific regulations at mRNA level or protein level. Though it is still unclear why some animals are resistant and others are susceptible to leptospiral infection, this comparative study based on transcriptomics and cytokine arrays partially uncovered the differences of murine resistance and human susceptibility to leptospirosis. Taken together, these findings will facilitate further molecular studies on the innate immune response to leptospiral infection. PMID:24130911

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

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    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.

  1. Chlamydia pneumoniae hides inside apoptotic neutrophils to silently infect and propagate in macrophages.

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    Jan Rupp

    Full Text Available BACKGROUND: Intracellular pathogens have developed elaborate strategies for silent infection of preferred host cells. Chlamydia pneumoniae is a common pathogen in acute infections of the respiratory tract (e.g. pneumonia and associated with chronic lung sequelae in adults and children. Within the lung, alveolar macrophages and polymorph nuclear neutrophils (PMN are the first line of defense against bacteria, but also preferred host phagocytes of chlamydiae. METHODOLOGY/PRINCIPAL FINDINGS: We could show that C. pneumoniae easily infect and hide inside neutrophil granulocytes until these cells become apoptotic and are subsequently taken up by macrophages. C. pneumoniae infection of macrophages via apoptotic PMN results in enhanced replicative activity of chlamydiae when compared to direct infection of macrophages, which results in persistence of the pathogen. Inhibition of the apoptotic recognition of C. pneumoniae infected PMN using PS- masking Annexin A5 significantly lowered the transmission of chlamydial infection to macrophages. Transfer of apoptotic C. pneumoniae infected PMN to macrophages resulted in an increased TGF-ss production, whereas direct infection of macrophages with chlamydiae was characterized by an enhanced TNF-alpha response. CONCLUSIONS/SIGNIFICANCE: Taken together, our data suggest that C. pneumoniae uses neutrophil granulocytes to be silently taken up by long-lived macrophages, which allows for efficient propagation and immune protection within the human host.

  2. Experimental Evolution of Mycobacterium tuberculosis in Human Macrophages Results in Low-Frequency Mutations Not Associated with Selective Advantage.

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    Valentina Guerrini

    Full Text Available Isolates of the human pathogen Mycobacterium tuberculosis recovered from clinical samples exhibit genetic heterogeneity. Such variation may result from the stressful environment encountered by the pathogen inside the macrophage, which is the host cell tubercle bacilli parasitize. To study the evolution of the M. tuberculosis genome during growth inside macrophages, we developed a model of intracellular culture in which bacteria were serially passaged in macrophage-like THP-1 cells for about 80 bacterial generations. Genome sequencing of single bacterial colonies isolated before and after the infection cycles revealed that M. tuberculosis developed mutations at a rate of about 5.7 × 10-9 / bp/ generation, consistent with mutation rates calculated during in vivo infection. Analysis of mutant growth in macrophages and in mice showed that the mutations identified after the cyclic infection conferred no advantage to the mutants relative to wild-type. Furthermore, activity testing of the recombinant protein harboring one of these mutations showed that the presence of the mutation did not affect the enzymatic activity. The serial infection protocol developed in this work to study M. tuberculosis genome microevolution can be applied to exposure to stressors to determine their effect on genome remodeling during intra-macrophage growth.

  3. Virulence of Mycobacterium avium Subsp. hominissuis Human Isolates in an in vitro Macrophage Infection Model

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    Laura Rindi

    2018-01-01

    Full Text Available Background: Mycobacterium avium subsp. hominissuis (MAH is an environmental opportunistic pathogen for humans and swine worldwide; in humans, the vast majority of MAH infections is due to strains belonging to specific genotypes, such as the internal transcribed spacer (ITS-sequevars Mav-A and Mav-B that mostly cause pulmonary infections in elderly patients and severe disseminated infections in acquired immunodeficiency syndrome patients, respectively. To test whether the different types of infections in distinct patients' populations might reflect a different virulence of the infecting genotypes, MAH human isolates, genotyped by ITS sequencing and MIRU-VNTR minisatellite analysis, were studied for the capacity to infect and replicate in human macrophages in vitro. Methods: Cultures of human peripheral blood mononuclear cells and phagocytic human leukemic cell line THP-1 cells were infected with each MAH isolate and intracellular colony-forming units (CFU were determined. Results: At 2 h after infection, i.e., immediately after cell entry, the numbers of intracellular bacteria did not differ between Mav-A and Mav-B organisms in both phagocytic cell types. At 5 days, Mav-A organisms, sharing highly related VNTR-MIRU genotypes, yielded numbers of intracellular CFUs significantly higher than Mav-B organisms in both phagocytic cell types. MIRU-VNTR-based minimum spanning tree analysis of the MAH isolates showed a divergent phylogenetic pathway of Mav-A and Mav-B organisms. Conclusion: Mav-A and Mav-B sequevars might have evolved different pathogenetic properties that might account for their association with different human infections.

  4. The differential role of human macrophage in triggering secondary bystander effects after either gamma-ray or carbon beam irradiation.

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    Dong, Chen; He, Mingyuan; Tu, Wenzhi; Konishi, Teruaki; Liu, Weili; Xie, Yuexia; Dang, Bingrong; Li, Wenjian; Uchihori, Yukio; Hei, Tom K; Shao, Chunlin

    2015-07-10

    The abscopal effect could be an underlying factor in evaluating prognosis of radiotherapy. This study established an in vitro system to examine whether tumor-generated bystander signals could be transmitted by macrophages to further trigger secondary cellular responses after different irradiations, where human lung cancer NCI-H446 cells were irradiated with either γ-rays or carbon ions and co-cultured with human macrophage U937 cells, then these U937 cells were used as a bystander signal transmitter and co-cultured with human bronchial epithelial cells BEAS-2B. Results showed that U937 cells were only activated by γ-irradiated NCI-H446 cells so that the secondary injuries in BEAS-2B cells under carbon ion irradiation were weaker than γ-rays. Both TNF-α and IL-1α were involved in the γ-irradiation induced secondary bystander effect but only TNF-α contributed to the carbon ion induced response. Further assay disclosed that IL-1α but not TNF-α was largely responsible for the activation of macrophages and the formation of micronucleus in BEAS-2B cells. These data suggest that macrophages could transfer secondary bystander signals and play a key role in the secondary bystander effect of photon irradiation, while carbon ion irradiation has conspicuous advantage due to its reduced secondary injury. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  5. Specific Kv1.3 blockade modulates key cholesterol-metabolism-associated molecules in human macrophages exposed to ox-LDL.

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    Yang, Yong; Wang, Yan-Fu; Yang, Xiao-Fang; Wang, Zhao-Hui; Lian, Yi-Tian; Yang, Ying; Li, Xiao-Wei; Gao, Xiang; Chen, Jian; Shu, Yan-Wen; Cheng, Long-Xian; Liao, Yu-Hua; Liu, Kun

    2013-01-01

    Cholesterol-metabolism-associated molecules, including scavenger receptor class A (SR-A), lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), CD36, ACAT1, ABCA1, ABCG1, and scavenger receptor class B type I, can modulate cholesterol metabolism in the transformation from macrophages to foam cells. Voltage-gated potassium channel Kv1.3 has increasingly been demonstrated to play an important role in the modulation of macrophage function. Here, we investigate the role of Kv1.3 in modulating cholesterol-metabolism-associated molecules in human acute monocytic leukemia cell-derived macrophages (THP-1 macrophages) and human monocyte-derived macrophages exposed to oxidized LDL (ox-LDL). Human Kv1.3 and Kv1.5 channels (hKv1.3 and hKv1.5) are expressed in macrophages and form a heteromultimeric channel. The hKv1.3-E314 antibody that we had generated as a specific hKv1.3 blocker inhibited outward delayed rectifier potassium currents, whereas the hKv1.5-E313 antibody that we had generated as a specific hKv1.5 blocker failed. Accordingly, the hKv1.3-E314 antibody reduced percentage of cholesterol ester and enhanced apoA-I-mediated cholesterol efflux in THP-1 macrophages and human monocyte-derived macrophages exposed to ox-LDL. The hKv1.3-E314 antibody downregulated SR-A, LOX-1, and ACAT1 expression and upregulated ABCA1 expression in THP-1 macrophages and human monocyte-derived macrophages. Our results reveal that specific Kv1.3 blockade represents a novel strategy modulating cholesterol metabolism in macrophages, which benefits the treatment of atherosclerotic lesions.

  6. Specific Kv1.3 blockade modulates key cholesterol-metabolism-associated molecules in human macrophages exposed to ox-LDL[S

    Science.gov (United States)

    Yang, Yong; Wang, Yan-Fu; Yang, Xiao-Fang; Wang, Zhao-Hui; Lian, Yi-Tian; Yang, Ying; Li, Xiao-Wei; Gao, Xiang; Chen, Jian; Shu, Yan-Wen; Cheng, Long-Xian; Liao, Yu-Hua; Liu, Kun

    2013-01-01

    Cholesterol-metabolism-associated molecules, including scavenger receptor class A (SR-A), lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), CD36, ACAT1, ABCA1, ABCG1, and scavenger receptor class B type I, can modulate cholesterol metabolism in the transformation from macrophages to foam cells. Voltage-gated potassium channel Kv1.3 has increasingly been demonstrated to play an important role in the modulation of macrophage function. Here, we investigate the role of Kv1.3 in modulating cholesterol-metabolism-associated molecules in human acute monocytic leukemia cell-derived macrophages (THP-1 macrophages) and human monocyte-derived macrophages exposed to oxidized LDL (ox-LDL). Human Kv1.3 and Kv1.5 channels (hKv1.3 and hKv1.5) are expressed in macrophages and form a heteromultimeric channel. The hKv1.3-E314 antibody that we had generated as a specific hKv1.3 blocker inhibited outward delayed rectifier potassium currents, whereas the hKv1.5-E313 antibody that we had generated as a specific hKv1.5 blocker failed. Accordingly, the hKv1.3-E314 antibody reduced percentage of cholesterol ester and enhanced apoA-I-mediated cholesterol efflux in THP-1 macrophages and human monocyte-derived macrophages exposed to ox-LDL. The hKv1.3-E314 antibody downregulated SR-A, LOX-1, and ACAT1 expression and upregulated ABCA1 expression in THP-1 macrophages and human monocyte-derived macrophages. Our results reveal that specific Kv1.3 blockade represents a novel strategy modulating cholesterol metabolism in macrophages, which benefits the treatment of atherosclerotic lesions. PMID:23099443

  7. Edaravone protects rats and human pulmonary alveolar epithelial cells against hyperoxia injury: heme oxygenase-1 and PI3K/Akt pathway may be involved.

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    Cao, Huifang; Feng, Ying; Ning, Yunye; Zhang, Zinan; Li, Weihao; Li, Qiang

    2015-01-01

    Hyperoxic acute lung injury (HALI) is a clinical syndrome as a result of prolonged supplement of high concentrations of oxygen. As yet, no specific treatment is available for HALI. The present study aims to investigate the effects of edaravone on hyperoxia-induced oxidative injury and the underlying mechanism. We treated rats and human pulmonary alveolar epithelial cells with hyperoxia and different concentration of edaravone, then examined the effects of edaravone on cell viability, cell injury and two oxidative products. The roles of heme oxygenase-1 (HO-1) and PI3K/Akt pathway were explored using Western blot and corresponding inhibitors. The results showed that edaravone reduced lung biochemical alterations induced by hyperoxia and mortality of rats, dose-dependently alleviated cell mortality, cell injury, and peroxidation of cellular lipid and DNA oxidative damage. It upregulated cellular HO-1 expression and activity, which was reversed by PI3K/Akt pathway inhibition. The administration of zinc protoporphyrin-IX, a HO-1 inhibitor, and LY249002, a PI3K/Akt pathway inhibitor, abolished the protective effects of edaravone in cells. This study indicates that edaravone protects rats and human pulmonary alveolar epithelial cells against hyperoxia-induced injury and the antioxidant effect may be related to upregulation of HO-1, which is regulated by PI3K/Akt pathway.

  8. Small airway epithelial cells exposure to printer-emitted engineered nanoparticles induces cellular effects on human microvascular endothelial cells in an alveolar-capillary co-culture model.

    Science.gov (United States)

    Sisler, Jennifer D; Pirela, Sandra V; Friend, Sherri; Farcas, Mariana; Schwegler-Berry, Diane; Shvedova, Anna; Castranova, Vincent; Demokritou, Philip; Qian, Yong

    2015-01-01

    The printer is one of the most common office equipment. Recently, it was reported that toner formulations for printing equipment constitute nano-enabled products (NEPs) and contain engineered nanomaterials (ENMs) that become airborne during printing. To date, insufficient research has been performed to understand the potential toxicological properties of printer-emitted particles (PEPs) with several studies using bulk toner particles as test particles. These studies demonstrated the ability of toner particles to cause chronic inflammation and fibrosis in animal models. However, the toxicological implications of inhalation exposures to ENMs emitted from laser printing equipment remain largely unknown. The present study investigates the toxicological effects of PEPs using an in vitro alveolar-capillary co-culture model with Human Small Airway Epithelial Cells (SAEC) and Human Microvascular Endothelial Cells (HMVEC). Our data demonstrate that direct exposure of SAEC to low concentrations of PEPs (0.5 and 1.0 µg/mL) caused morphological changes of actin remodeling and gap formations within the endothelial monolayer. Furthermore, increased production of reactive oxygen species (ROS) and angiogenesis were observed in the HMVEC. Analysis of cytokine and chemokine levels demonstrates that interleukin (IL)-6 and MCP-1 may play a major role in the cellular communication observed between SAEC and HMVEC and the resultant responses in HMVEC. These data indicate that PEPs at low, non-cytotoxic exposure levels are bioactive and affect cellular responses in an alveolar-capillary co-culture model, which raises concerns for potential adverse health effects.

  9. The β-hemolysin and intracellular survival of Streptococcus agalactiae in human macrophages.

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    Sagar, Anubha; Klemm, Carolin; Hartjes, Lara; Mauerer, Stefanie; van Zandbergen, Ger; Spellerberg, Barbara

    2013-01-01

    S. agalactiae (group B streptococci, GBS) is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches.

  10. The β-Hemolysin and Intracellular Survival of Streptococcus agalactiae in Human Macrophages

    Science.gov (United States)

    Sagar, Anubha; Klemm, Carolin; Hartjes, Lara; Mauerer, Stefanie; van Zandbergen, Ger; Spellerberg, Barbara

    2013-01-01

    S. agalactiae (group B streptococci, GBS) is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches. PMID:23593170

  11. The β-hemolysin and intracellular survival of Streptococcus agalactiae in human macrophages.

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    Anubha Sagar

    Full Text Available S. agalactiae (group B streptococci, GBS is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches.

  12. C–C Chemokines Released by Lipopolysaccharide (LPS)-stimulated Human Macrophages Suppress HIV-1 Infection in Both Macrophages and T Cells

    Science.gov (United States)

    Verani, Alessia; Scarlatti, Gabriella; Comar, Manola; Tresoldi, Eleonora; Polo, Simona; Giacca, Mauro; Lusso, Paolo; Siccardi, Antonio G.; Vercelli, Donata

    1997-01-01

    Human immunodeficiency virus-1 (HIV-1) expression in monocyte-derived macrophages (MDM) infected in vitro is known to be inhibited by lipopolysaccharide (LPS). However, the mechanisms are incompletely understood. We show here that HIV-1 suppression is mediated by soluble factors released by MDM stimulated with physiologically significant concentrations of LPS. LPS-conditioned supernatants from MDM inhibited HIV-1 replication in both MDM and T cells. Depletion of C–C chemokines (RANTES, MIP-1α, and MIP-1β) neutralized the ability of LPS-conditioned supernatants to inhibit HIV-1 replication in MDM. A combination of recombinant C–C chemokines blocked HIV-1 infection as effectively as LPS. Here, we report an inhibitory effect of C–C chemokines on HIV replication in primary macrophages. Our results raise the possibility that monocytes may play a dual role in HIV infection: while representing a reservoir for the virus, they may contribute to the containment of the infection by releasing factors that suppress HIV replication not only in monocytes but also in T lymphocytes. PMID:9120386

  13. Changes in the topology of gene expression networks by human immunodeficiency virus type 1 (HIV-1) integration in macrophages.

    Science.gov (United States)

    Soto-Girón, María Juliana; García-Vallejo, Felipe

    2012-01-01

    One key step of human immunodeficiency virus type 1 (HIV-1) infection is the integration of its viral cDNA. This process is mediated through complex networks of host-virus interactions that alter several normal cell functions of the host. To study the complexity of disturbances in cell gene expression networks by HIV-1 integration, we constructed a network of human macrophage genes located close to chromatin regions rich in proviruses. To perform the network analysis, we selected 28 genes previously identified as the target of cDNA integration and their transcriptional profiles were obtained from GEO Profiles (NCBI). A total of 2770 interactions among the 28 genes located around the HIV-1 proviruses in human macrophages formed a highly dense main network connected to five sub-networks. The overall network was significantly enriched by genes associated with signal transduction, cellular communication and regulatory processes. To simulate the effects of HIV-1 integration in infected macrophages, five genes with the most number of interaction in the normal network were turned off by putting in zero the correspondent expression values. The HIV-1 infected network showed changes in its topology and alteration in the macrophage functions reflected in a re-programming of biosynthetic and general metabolic process. Understanding the complex virus-host interactions that occur during HIV-1 integration, may provided valuable genomic information to develop new antiviral treatments focusing on the management of some specific gene expression networks associated with viral integration. This is the first gene network which describes the human macrophages genes interactions related with HIV-1 integration. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Granulocyte macrophage colony stimulating factor (GM-CSF biological actions on human dermal fibroblasts

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    S Montagnani

    2009-12-01

    Full Text Available Fibroblasts are involved in all pathologies characterized by increased ExtraCellularMatrix synthesis, from wound healing to fibrosis. Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF is a cytokine isolated as an hemopoietic growth factor but recently indicated as a differentiative agent on endothelial cells. In this work we demonstrated the expression of the receptor for GM-CSF (GMCSFR on human normal skin fibroblasts from healthy subjects (NFPC and on a human normal fibroblast cell line (NHDF and we try to investigate the biological effects of this cytokine. Human normal fibroblasts were cultured with different doses of GM-CSF to study the effects of this factor on GMCSFR expression, on cell proliferation and adhesion structures. In addition we studied the production of some Extra-Cellular Matrix (ECM components such as Fibronectin, Tenascin and Collagen I. The growth rate of fibroblasts from healthy donors (NFPC is not augmented by GM-CSF stimulation in spite of increased expression of the GM-CSFR. On the contrary, the proliferation of normal human dermal fibroblasts (NHDF cell line seems more influenced by high concentration of GM-CSF in the culture medium. The adhesion structures and the ECM components appear variously influenced by GM-CSF treatment as compared to fibroblasts cultured in basal condition, but newly only NHDF cells are really induced to increase their synthesis activity. We suggest that the in vitro treatment with GM-CSF can shift human normal fibroblasts towards a more differentiated state, due or accompanied by an increased expression of GM-CSFR and that such “differentiation” is an important event induced by such cytokine.

  15. alpha-MSH and its receptors in regulation of tumor necrosis factor-alpha production by human monocyte/macrophages.

    Science.gov (United States)

    Taherzadeh, S; Sharma, S; Chhajlani, V; Gantz, I; Rajora, N; Demitri, M T; Kelly, L; Zhao, H; Ichiyama, T; Catania, A; Lipton, J M

    1999-05-01

    The hypothesis that macrophages contain an autocrine circuit based on melanocortin [ACTH and alpha-melanocyte-stimulating hormone (alpha-MSH)] peptides has major implications for neuroimmunomodulation research and inflammation therapy. To test this hypothesis, cells of the THP-1 human monocyte/macrophage line were stimulated with lipopolysaccharide (LPS) in the presence and absence of alpha-MSH. The inflammatory cytokine tumor necrosis factor (TNF)-alpha was inhibited in relation to alpha-MSH concentration. Similar inhibitory effects on TNF-alpha were observed with ACTH peptides that contain the alpha-MSH amino acid sequence and act on melanocortin receptors. Nuclease protection assays indicated that expression of the human melanocortin-1 receptor subtype (hMC-1R) occurs in THP-1 cells; Southern blots of RT-PCR product revealed that additional subtypes, hMC-3R and hMC-5R, also occur. Incubation of resting macrophages with antibody to hMC-1R increased TNF-alpha concentration; the antibody also markedly reduced the inhibitory influence of alpha-MSH on TNF-alpha in macrophages treated with LPS. These results in cells known to produce alpha-MSH at rest and to increase secretion of the peptide when challenged are consistent with an endogenous regulatory circuit based on melanocortin peptides and their receptors. Targeting of this neuroimmunomodulatory circuit in inflammatory diseases in which myelomonocytic cells are prominent should be beneficial.

  16. Increased platelet reactivity is associated with circulating platelet-monocyte complexes and macrophages in human atherosclerotic plaques.

    Directory of Open Access Journals (Sweden)

    Bert Rutten

    Full Text Available Platelet reactivity, platelet binding to monocytes and monocyte infiltration play a detrimental role in atherosclerotic plaque progression. We investigated whether platelet reactivity was associated with levels of circulating platelet-monocyte complexes (PMCs and macrophages in human atherosclerotic carotid plaques.Platelet reactivity was determined by measuring platelet P-selectin expression after platelet stimulation with increasing concentrations of adenosine diphosphate (ADP, in two independent cohorts: the Circulating Cells cohort (n = 244 and the Athero-Express cohort (n = 91. Levels of PMCs were assessed by flow cytometry in blood samples of patients who were scheduled for percutaneous coronary intervention (Circulating Cells cohort. Monocyte infiltration was semi-quantitatively determined by histological examination of atherosclerotic carotid plaques collected during carotid endarterectomy (Athero-Express cohort.We found increased platelet reactivity in patients with high PMCs as compared to patients with low PMCs (median (interquartile range: 4153 (1585-11267 area under the curve (AUC vs. 9633 (3580-21565 AUC, P<0.001. Also, we observed increased platelet reactivity in patients with high macrophage levels in atherosclerotic plaques as compared to patients with low macrophage levels in atherosclerotic plaques (mean ± SD; 8969 ± 3485 AUC vs. 7020 ± 3442 AUC, P = 0.02. All associations remained significant after adjustment for age, sex and use of drugs against platelet activation.Platelet reactivity towards ADP is associated with levels of PMCs and macrophages in human atherosclerotic carotid plaques.

  17. Peroxisome Proliferator-Activated Receptor γ Induces the Expression of Tissue Factor Pathway Inhibitor-1 (TFPI-1 in Human Macrophages

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    G. Chinetti-Gbaguidi

    2016-01-01

    Full Text Available Tissue factor (TF is the initiator of the blood coagulation cascade after interaction with the activated factor VII (FVIIa. Moreover, the TF/FVIIa complex also activates intracellular signalling pathways leading to the production of inflammatory cytokines. The TF/FVIIa complex is inhibited by the tissue factor pathway inhibitor-1 (TFPI-1. Peroxisome proliferator-activated receptor gamma (PPARγ is a transcription factor that, together with PPARα and PPARβ/δ, controls macrophage functions. However, whether PPARγ activation modulates the expression of TFP1-1 in human macrophages is not known. Here we report that PPARγ activation increases the expression of TFPI-1 in human macrophages in vitro as well as in vivo in circulating peripheral blood mononuclear cells. The induction of TFPI-1 expression by PPARγ ligands, an effect shared by the activation of PPARα and PPARβ/δ, occurs also in proinflammatory M1 and in anti-inflammatory M2 polarized macrophages. As a functional consequence, treatment with PPARγ ligands significantly reduces the inflammatory response induced by FVIIa, as measured by variations in the IL-8, MMP-2, and MCP-1 expression. These data identify a novel role for PPARγ in the control of TF the pathway.

  18. GABA and Topiramate Inhibit the Formation of Human Macrophage-Derived Foam Cells by Modulating Cholesterol-Metabolism-Associated Molecules

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    Ying Yang

    2014-04-01

    Full Text Available Aims: γ-aminobutyric acid (GABA, the principal inhibitory neurotransmitter, acts on GABA receptors to play an important role in the modulation of macrophage functions. The present study examined the effects of GABA and a GABA receptor agonist on modulating cholesterol-metabolism-associated molecules in human monocyte-derived macrophages (HMDMs. Methods: ORO stain, HPLC, qRT-PCR, Western blot and EMSA were carried out using HMDMs exposed to ox-LDL with or without GABAergic agents as the experimental model. Results: GABA and topiramate reduced the percentage of cholesterol ester in lipid-laden HMDMs by down-regulating SR-A, CD36 and LOX-1 expression and up-regulating ABCA1, ABCG1 and SR-BI expression in lipid-laden HMDMs. The production of TNF-a was decreased in GABA-and topiramate-treated lipid-laden HMDMs, and levels of interleukin (IL-6 did not change. The activation of two signaling pathways, p38MAPK and NF-γB, was repressed by GABA and topiramate in lipid-laden HMDMs. Conclusion: GABA and topiramate inhibit the formation of human macrophage-derived foam cells and may be a possibility for macrophage targeted therapy of atherosclerotic lesions.

  19. Acceleration of Alveolar Ridge Augmentation Using a Low Dose of Recombinant Human Bone Morphogenetic Protein-2 Loaded on a Resorbable Bioactive Ceramic.

    Science.gov (United States)

    Fahmy, Rania A; Mahmoud, Naguiba; Soliman, Samia; Nouh, Samir R; Cunningham, Larry; El-Ghannam, Ahmed

    2015-12-01

    The aim of the present study was to evaluate the effect of a porous silica-calcium phosphate composite (SCPC50) loaded with and without recombinant human bone morphogenetic protein-2 (rhBMP-2) on alveolar ridge augmentation in saddle-type defects. Micro-granules of SCPC50 resorbable bioactive ceramic were coated with rhBMP-2 10 mg and then implanted into a saddle-type defect (12 × 7 mm) in a dog mandible and covered with a collagen membrane. Control groups included defects grafted with SCPC50 granules without rhBMP-2 and un-grafted defects. Bone healing was evaluated at 8 and 16 weeks using histologic and histomorphometric techniques. The increase in bone height and total defect fill were assessed for each specimen using the ImageJ 1.46 program. The release kinetics of rhBMP-2 was determined in vitro. The height of the bone in the grafted defects and the total defect fill were statistically analyzed. SCPC50 enhanced alveolar ridge augmentation as indicated by the increased vertical bone height, bone surface area, and bone volume after 16 weeks. SCPC50-rhBMP-2 provided a sustained release profile of a low effective dose (BMP-2 4.6 ± 1.34 pg/mL per hour) during the 1- to 21-day period. The slow rate of release of rhBMP-2 from SCPC50 accelerated synchronized complete bone regeneration and graft material resorption in 8 weeks. Successful rapid reconstruction of the alveolar ridge by SCPC50 and SCPC50-rhBMP-2 occurred without any adverse excessive bone formation, inflammation, or fluid-filled voids. Results of this study suggest that SCPC50 is an effective graft material to preserve the alveolar ridge after tooth extraction. Coating SCPC50-rhBMP-2 further accelerated bone regeneration and a considerable increase in vertical bone height. These findings make SCPC50 the primary choice as a carrier for rhBMP-2. SCPC50-rhBMP-2 can serve as an alternative to autologous bone grafting. Published by Elsevier Inc.

  20. Cholesterol Oxidase Binds TLR2 and Modulates Functional Responses of Human Macrophages

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    Katarzyna Bednarska

    2014-01-01

    Full Text Available Cholesterol oxidase (ChoD is considered to be an important virulence factor for Mycobacterium tuberculosis (Mtb, but its influence on macrophage activity is unknown. Here we used Nocardia erythropolis ChoD, which is very similar to the Mtb enzyme (70% identity at the amino-acid level, to evaluate the impact of bacterial ChoD on the activity of THP-1-derived macrophages in vitro. We found that ChoD decreased the surface expression of Toll-like receptor type 2 (TLR2 and complement receptor 3 (CR3 on these macrophages. Flow cytometry and confocal microscopy showed that ChoD competed with lipoteichoic acid for ligand binding sites on TLR2 but not on CR3, suggesting that ChoD signaling is mediated via TLR2. Binding of ChoD to the membrane of macrophages had diverse effects on the activity of macrophages, activating p38 mitogen activated kinase and stimulating production of a large amount of interleukin-10. Moreover, ChoD primed macrophages to enhance the production of reactive oxygen species in response to the phorbol myristate acetate, which was reduced by “switching off” TLR-derived signaling through interleukin-1 receptor-associated kinases 1 and 4 inhibition. Our study revealed that ChoD interacts directly with macrophages via TLR2 and influences the biological activity of macrophages during the development of the initial response to infection.

  1. Biocompatibility and Inflammatory Potential of Titanium Alloys Cultivated with Human Osteoblasts, Fibroblasts and Macrophages

    Science.gov (United States)

    Markhoff, Jana; Krogull, Martin; Schulze, Christian; Rotsch, Christian; Hunger, Sandra; Bader, Rainer

    2017-01-01

    The biomaterials used to maintain or replace functions in the human body consist mainly of metals, ceramics or polymers. In orthopedic surgery, metallic materials, especially titanium and its alloys, are the most common, due to their excellent mechanical properties, corrosion resistance, and biocompatibility. Aside from the established Ti6Al4V alloy, shape memory materials such as nickel-titanium (NiTi) have risen in importance, but are also discussed because of the adverse effects of nickel ions. These might be reduced by specific surface modifications. In the present in vitro study, the osteoblastic cell line MG-63 as well as primary human osteoblasts, fibroblasts, and macrophages were cultured on titanium alloys (forged Ti6Al4V, additive manufactured Ti6Al4V, NiTi, and Diamond-Like-Carbon (DLC)-coated NiTi) to verify their specific biocompatibility and inflammatory potential. Additive manufactured Ti6Al4V and NiTi revealed the highest levels of metabolic cell activity. DLC-coated NiTi appeared as a suitable surface for cell growth, showing the highest collagen production. None of the implant materials caused a strong inflammatory response. In general, no distinct cell-specific response could be observed for the materials and surface coating used. In summary, all tested titanium alloys seem to be biologically appropriate for application in orthopedic surgery. PMID:28772412

  2. Biocompatibility and Inflammatory Potential of Titanium Alloys Cultivated with Human Osteoblasts, Fibroblasts and Macrophages

    Directory of Open Access Journals (Sweden)

    Jana Markhoff

    2017-01-01

    Full Text Available The biomaterials used to maintain or replace functions in the human body consist mainly of metals, ceramics or polymers. In orthopedic surgery, metallic materials, especially titanium and its alloys, are the most common, due to their excellent mechanical properties, corrosion resistance, and biocompatibility. Aside from the established Ti6Al4V alloy, shape memory materials such as nickel-titanium (NiTi have risen in importance, but are also discussed because of the adverse effects of nickel ions. These might be reduced by specific surface modifications. In the present in vitro study, the osteoblastic cell line MG-63 as well as primary human osteoblasts, fibroblasts, and macrophages were cultured on titanium alloys (forged Ti6Al4V, additive manufactured Ti6Al4V, NiTi, and Diamond-Like-Carbon (DLC-coated NiTi to verify their specific biocompatibility and inflammatory potential. Additive manufactured Ti6Al4V and NiTi revealed the highest levels of metabolic cell activity. DLC-coated NiTi appeared as a suitable surface for cell growth, showing the highest collagen production. None of the implant materials caused a strong inflammatory response. In general, no distinct cell-specific response could be observed for the materials and surface coating used. In summary, all tested titanium alloys seem to be biologically appropriate for application in orthopedic surgery.

  3. Transforming growth factor beta-1 expression in macrophages of human chronic periapical diseases.

    Science.gov (United States)

    Liang, Z-Z; Li, J; Huang, S-G

    2017-03-30

    The objective of this study was to observe the distribution of macrophages (MPs) expressing transforming growth factor beta-1 (TGF-β1) in tissue samples from patients with different human chronic periapical diseases. In this study, samples were collected from 75 volunteers, who were divided into three groups according to classified standards, namely, healthy control (N = 25), periapical granuloma (N = 25), and periapical cyst (N = 25). The samples were fixed in 10% buffered formalin for more than 48 h, dehydrated, embedded, and stained with hematoxylin and eosin for histopathology. Double immunofluorescence was conducted to analyze the expression of TGF-β-CD14 double-positive MPs in periapical tissues. The number of double-positive cells (cells/mm 2 ) were significantly higher in the chronic periapical disease tissues (P periapical cyst group than in the periapical granuloma group (P periapical diseases. The TGF-β1-CD14 double-positive cells might play an important role in the pathology of human chronic periapical diseases.

  4. Expression of functional toll-like receptor-2 and -4 on alveolar epithelial cells.

    Science.gov (United States)

    Armstrong, Lynne; Medford, Andrew R L; Uppington, Kay M; Robertson, John; Witherden, Ian R; Tetley, Teresa D; Millar, Ann B

    2004-08-01

    The recognition of potentially harmful microorganisms involves the specific recognition of pathogen-associated molecular patterns (PAMPs) and the family of Toll-like receptors (TLRs) is known to play a central role in this process. TLR-4 is the major recognition receptor for lipopolysaccharide (LPS), a component of gram-negative bacterial cell walls, whereas TLR-2 responds to bacterial products from gram-positive organisms. Although resident alveolar macrophages are the first line of defense against microbial attack, it is now understood that the alveolar epithelium also plays a pivotal role in the innate immunity of the lung. The purpose of the current study was to determine whether human primary type II alveolar epithelial cells (ATII) express functional TLR-2 and TLR-4 and how they may be regulated by inflammatory mediators. We have used reverse transcriptase-polymerase chain reaction and flow cytometry to determine basal and inducible expression on ATII. We have used highly purified preparations of the gram-positive bacterial product lipoteichoic acid (LTA) and LPS to look at the functional consequences of TLR-2 and TLR-4 ligation, respectively, in terms of interleukin-8 release. We have shown that human primary ATII cells express mRNA and protein for both TLR-2 and TLR-4, which can be modulated by incubation with LPS and tumor necrosis factor. Furthermore, we have demonstrated that these receptors are functional. This suggests that ATII have the potential to contribute significantly to the host defense of the human alveolus against bacteria.

  5. The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema.

    Science.gov (United States)

    Tan, Li Hui; Bahmed, Karim; Lin, Chih-Ru; Marchetti, Nathaniel; Bolla, Sudhir; Criner, Gerard J; Kelsen, Steven; Madesh, Muniswamy; Kosmider, Beata

    2018-02-23

    Emphysema is characterized by irreversibly enlarged airspaces and destruction of alveolar walls. One of the factors contributing to this disease pathogenesis is an elevation in extracellular matrix (ECM) degradation in the lung. Alveolar type II (ATII) cells produce and secrete pulmonary surfactants and proliferate to restore the epithelium after damage. We isolated ATII cells from control non-smokers, smokers and patients with emphysema to determine the role of NFE2 (nuclear factor, erythroid-derived 2). NFE2 is a heterodimer composed of two subunits, a 45 kDa (p45 NFE2) and 18 kDa (p18 NFE2) polypeptides. Low expression of p45 NFE2 in patients with emphysema correlated with a high ECM degradation. Moreover, we found that NFE2 knockdown increased cell death induced by cigarette smoke extract. We also studied the cross talk between p45 NFE2 and DJ-1. DJ-1 protein is a redox-sensitive chaperone that protects cells from oxidative stress. We detected that cigarette smoke significantly increased p45 NFE2 levels in DJ-1 KO mice compared to wild-type mice. Our results indicate that p45 NFE2 expression is induced by exposure to cigarette smoke, has a cytoprotective activity against cell injury, and its downregulation in human primary ATII cells may contribute to emphysema pathogenesis.

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

    Science.gov (United States)

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

    2014-03-10

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

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

    Directory of Open Access Journals (Sweden)

    Bonnie van Wilgenburg

    Full Text Available Human macrophages are specialised hosts for HIV-1, dengue virus, Leishmania and Mycobacterium tuberculosis. Yet macrophage research is hampered by lack of appropriate cell models for modelling infection by these human pathogens, because available myeloid cell lines are, by definition, not terminally differentiated like tissue macrophages. We describe here a method for deriving monocytes and macrophages from human Pluripotent Stem Cells which improves on previously published protocols in that it uses entirely defined, feeder- and serum-free culture conditions and produces very consistent, pure, high yields across both human Embryonic Stem Cell (hESC and multiple human induced Pluripotent Stem Cell (hiPSC lines over time periods of up to one year. Cumulatively, up to ∼3×10(7 monocytes can be harvested per 6-well plate. The monocytes produced are most closely similar to the major blood monocyte (CD14(+, CD16(low, CD163(+. Differentiation with M-CSF produces macrophages that are highly phagocytic, HIV-1-infectable, and upon activation produce a pro-inflammatory cytokine profile similar to blood monocyte-derived macrophages. Macrophages are notoriously hard to genetically manipulate, as they recognise foreign nucleic acids; the lentivector system described here overcomes this, as pluripotent stem cells can be relatively simply genetically manipulated for efficient transgene expression in the differentiated cells, surmounting issues of transgene silencing. Overall, the method we describe here is an efficient, effective, scalable system for the reproducible production and genetic modification of human macrophages, facilitating the interrogation of human macrophage biology.

  8. Differentiation and cytokine synthesis of human alveolar osteoblasts compared to osteoblast-like cells (MG63) in response to titanium surfaces.

    Science.gov (United States)

    Rausch-fan, Xiaohui; Qu, Zhe; Wieland, Marco; Matejka, Michael; Schedle, Andreas

    2008-01-01

    The aim of this study was to investigate the influence of different implant surface topographies and chemistries on the expression of differentiation/proliferation markers on MG63 cells and primary human alveolar osteoblasts. Hydrophobic acid-etched (A) and hydrophobic coarse-grit-blasted, acid-etched (SLA) surfaces and hydrophilic acid-etched (modA) and hydrophilic coarse-grit-blasted (modSLA) surfaces were produced. Thereby, modA and modSLA surfaces were rinsed under nitrogen protection and stored in a sealed glass tube containing isotonic NaCl solution at pH 4-6. Tissue culture plates without specimens served as controls. The behavior of MG63 cells and primary human alveolar osteoblasts (AOB) grown on all surfaces was compared through determination of alkaline phosphatase (ALP) activity, cell proliferation ((3)H-thymidin incorporation, MTT colorimetric assay) and expression of osteocalcin (OC), osteoprotegerin (OPG), transforming growth factor-beta1 (TGF-beta(1)) and vascular endothelial growth factor (VEGF), detected with commercial available test kits. Proliferation of MG63 and primary cells was highest on controls, followed by A surfaces, modA and SLA surfaces being almost on the same level and lowest on modSLA surfaces. modSLA surfaces exhibited highest ALP and OC production, followed by SLA, modA and A surfaces. Proliferation and OC production were comparable for MG63 cells and AOB. OPG, TGF-beta(1) and VEGF produced on primary cells showed a slightly different rank order on different surfaces compared to MG63 cells. modSLA still showed the highest production of OPG, TGF-beta(1) and VEGF, but was followed by modA, SLA and A. Statistical significance was checked by ANOVA (pmodA surfaces showed enhanced expression of OPG, TGF-beta(1) and VEGF on MG63 cells compared to primary human alveolar osteoblasts. Overall, the lowest proliferation rates and the highest expressions of differentiation markers and growth factor productions were observed on modSLA.

  9. Exposure of Monocytes to Lipoarabinomannan Promotes Their Differentiation into Functionally and Phenotypically Immature Macrophages

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    Leslie Chávez-Galán

    2015-01-01

    Full Text Available Lipoarabinomannan (LAM is a lipid virulence factor secreted by Mycobacterium tuberculosis (Mtb, the etiologic agent of tuberculosis. LAM can be measured in the urine or serum of tuberculosis patients (TB-patients. Circulating monocytes are the precursor cells of alveolar macrophages and might be exposed to LAM in patients with active TB. We speculated that exposing monocytes to LAM could produce phenotypically and functionally immature macrophages. To test our hypothesis, human monocytes were stimulated with LAM (24–120 hours and various readouts were measured. The study showed that when monocytes were exposed to LAM, the frequency of CD68+, CD33+, and CD86+ macrophages decreased, suggesting that monocyte differentiation into mature macrophages was affected. Regarding functionality markers, TLR2+ and TLR4+ macrophages also decreased, but the percentage of MMR+ expression did not change. LAM-exposed monocytes generated macrophages that were less efficient in producing proinflammatory cytokines such as TNF-α and IFN-γ; however, their phagocytic capacity was not modified. Taken together, these data indicate that LAM exposure influenced monocyte differentiation and produced poorly functional macrophages with a different phenotype. These results may help us understand how mycobacteria can limit the quality of the innate and adaptive immune responses.

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

    Science.gov (United States)

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

    1990-01-01

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

  11. Classification of M1/M2-polarized human macrophages by label-free hyperspectral reflectance confocal microscopy and multivariate analysis.

    Science.gov (United States)

    Bertani, Francesca R; Mozetic, Pamela; Fioramonti, Marco; Iuliani, Michele; Ribelli, Giulia; Pantano, Francesco; Santini, Daniele; Tonini, Giuseppe; Trombetta, Marcella; Businaro, Luca; Selci, Stefano; Rainer, Alberto

    2017-08-21

    The possibility of detecting and classifying living cells in a label-free and non-invasive manner holds significant theranostic potential. In this work, Hyperspectral Imaging (HSI) has been successfully applied to the analysis of macrophagic polarization, given its central role in several pathological settings, including the regulation of tumour microenvironment. Human monocyte derived macrophages have been investigated using hyperspectral reflectance confocal microscopy, and hyperspectral datasets have been analysed in terms of M1 vs. M2 polarization by Principal Components Analysis (PCA). Following PCA, Linear Discriminant Analysis has been implemented for semi-automatic classification of macrophagic polarization from HSI data. Our results confirm the possibility to perform single-cell-level in vitro classification of M1 vs. M2 macrophages in a non-invasive and label-free manner with a high accuracy (above 98% for cells deriving from the same donor), supporting the idea of applying the technique to the study of complex interacting cellular systems, such in the case of tumour-immunity in vitro models.

  12. Human umbilical cord blood-derived f-macrophages retain pluripotentiality after thrombopoietin expansion

    International Nuclear Information System (INIS)

    Zhao Yong; Mazzone, Theodore

    2005-01-01

    We have previously characterized a new type of stem cell from human peripheral blood, termed fibroblast-like macrophage (f-MΦ). Here, using umbilical cord blood as a source, we identified cells with similar characteristics including expression of surface markers (CD14, CD34, CD45, CD117, and CD163), phagocytosis, and proliferative capacity. Further, thrombopoietin (TPO) significantly stimulated the proliferation of cord blood-derived f-MΦ (CB f-MΦ) at low dosage without inducing a megakaryocytic phenotype. Additional experiments demonstrated that TPO-expanded cord blood-derived f-MΦ (TCB f-MΦ) retained their surface markers and differentiation ability. Treatment with vascular endothelial cell growth factor (VEGF) gave rise to endothelial-like cells, expressing Flt-1, Flk-1, von Willebrand Factor (vWF), CD31, acetylated low density lipoprotein internalization, and the ability to form endothelial-like cell chains. In the presence of lipopolyssacharide (LPS) and 25 mM glucose, the TCB f-MΦ differentiated to express insulin mRNA, C-peptide, and insulin. In vitro functional analysis demonstrated that these insulin-positive cells could release insulin in response to glucose and other secretagogues. These findings demonstrate a potential use of CB f-MΦ and may lead to develop new therapeutic strategy for treating dominant disease

  13. Identification of the Common Origins of Osteoclasts, Macrophages, and Dendritic Cells in Human Hematopoiesis

    Directory of Open Access Journals (Sweden)

    Yanling Xiao

    2015-06-01

    Full Text Available Osteoclasts (OCs originate from the myeloid cell lineage, but the successive steps in their lineage commitment are ill-defined, especially in humans. To clarify OC origin, we sorted cell populations from pediatric bone marrow (BM by flow cytometry and assessed their differentiation potential in vitro. Within the CD11b−CD34+c-KIT+ BM cell population, OC-differentiation potential was restricted to FLT3+ cells and enriched in an IL3 receptor (Rαhigh subset that constituted less than 0.5% of total BM. These IL3Rαhigh cells also generated macrophages (MΦs and dendritic cells (DCs but lacked granulocyte (GR-differentiation potential, as demonstrated at the clonal level. The IL3Rαlow subset was re-defined as common progenitor of GR, MΦ, OC, and DC (GMODP and gave rise to the IL3Rαhigh subset that was identified as common progenitor of MΦ, OC, and DC (MODP. Unbiased transcriptome analysis of CD11b−CD34+c-KIT+FLT3+ IL3Rαlow and IL3Rαhigh subsets corroborated our definitions of the GMODP and MODP and their developmental relationship.

  14. Soluble ICAM-1 activates lung macrophages and enhances lung injury

    DEFF Research Database (Denmark)

    Schmal, H; Czermak, B J; Lentsch, A B

    1998-01-01

    production of TNF-alpha and the CXC chemokine, macrophage inflammatory protein-2 (MIP-2). Alveolar macrophages exhibited cytokine responses to both sICAM-1 and immobilized sICAM-1, while rat PBMCs failed to demonstrate similar responses. Exposure of alveolar macrophages to sICAM-1 resulted in NFkappa......B activation (which was blocked by the presence of the aldehyde peptide inhibitor of 28S proteosome and by genistein, a tyrosine kinase inhibitor). As expected, cross-linking of CD18 on macrophages with Ab resulted in generation of TNF-alpha and MIP-2. This response was also inhibited in the presence...... of TNF-alpha and MIP-2 and increased neutrophil recruitment. Therefore, through engagement of beta2 integrins, sICAM-1 enhances alveolar macrophage production of MIP-2 and TNF-alpha, the result of which is intensified lung injury after intrapulmonary disposition of immune complexes....

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

    DEFF Research Database (Denmark)

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

    2003-01-01

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

  16. Is length an appropriate estimator to characterize pulmonary alveolar capillaries? A critical evaluation in the human lung

    DEFF Research Database (Denmark)

    Mühlfeld, Christian; Weibel, Ewald R.; Hahn, Ute

    2010-01-01

    Stereological estimations of total capillary length have been used to characterize changes in the alveolar capillary network (ACN) during developmental processes or pathophysiological conditions. Here, we analyzed whether length estimations are appropriate to describe the 3D nature of the ACN. Semi...... resulted in a mean of 2,746 km (SD: 722 km). Because of the geometry of the ACN both approaches carry an unpredictable bias. The bias incurred by the design-based approach is proportional to the ratio between radius and length of the capillary segments in the ACN, the number of branching points...... and the winding of the capillaries. The model-based approach is biased because of the real noncylindrical shape of capillaries and the network structure. In conclusion, the estimation of the total length of capillaries in the ACN cannot be recommended as the geometry of the ACN does not fulfill the requirements...

  17. Proteinosis alveolar pulmonar Pulmonary alveolar proteinosis

    Directory of Open Access Journals (Sweden)

    Concepción Sánchez Infante

    2011-12-01

    Full Text Available La proteinosis alveolar pulmonar es una enfermedad respiratoria crónica, caracterizada por alteración en el metabolismo del surfactante, lo que determina su acumulación anormal en el espacio alveolar. Es una enfermedad extremadamente rara. Se han reportado solamente 500 casos en la literatura. Se describió por primera vez en 1958. Se presenta un caso de proteinosis alveolar pulmonar en un lactante de 2 meses, con desnutrición proteico energética, que ingresa por dificultad respiratoria e hipoxemia, y, con imágenes radiológicas de tipo retículo-nodulillar, en vidrio deslustrado, en el cual se plantea inicialmente el diagnóstico de bronconeumonía. Ante la evolución desfavorable y no respuesta al tratamiento, se realizó un estudio para descartar enfermedades pulmonares crónicas. El paciente fallece y se confirma el diagnóstico por anatomía patológica. Se realiza una revisión del tema.The pulmonary alveolar proteinosis is a chronic respiratory disease characterized by surfactant metabolism alteration determining its abnormal accumulation in the alveolar space. It is a disease very rare and in literature only 500 cases have been reported; it was described for the first time in 1958. This is a case presentation of pulmonary alveolar proteinosis in an infant aged 2 months with energetic protein malnutrition admitted due to respiratory difficulty and hypoxemia and with radiologic images of the reticulonodulillary, in frosting glass, where initially is made the diagnosis of bronchopneumonia. In the face of unfavorable evolution and no response to treatment, a study was conducted to rule out chronic pulmonary diseases. Patient died confirming the diagnosis according to the pathologic anatomy. A review on subject is carried out.

  18. DJ-1 Modulates Nuclear Erythroid 2-Related Factor-2-Mediated Protection in Human Primary Alveolar Type II Cells in Smokers.

    Science.gov (United States)

    Bahmed, Karim; Messier, Elise M; Zhou, Wenbo; Tuder, Rubin M; Freed, Curt R; Chu, Hong Wei; Kelsen, Steven G; Bowler, Russell P; Mason, Robert J; Kosmider, Beata

    2016-09-01

    Cigarette smoke (CS) is a main source of oxidative stress and a key risk factor for emphysema, which consists of alveolar wall destruction. Alveolar type (AT) II cells are in the gas exchange regions of the lung. We isolated primary ATII cells from deidentified organ donors whose lungs were not suitable for transplantation. We analyzed the cell injury obtained from nonsmokers, moderate smokers, and heavy smokers. DJ-1 protects cells from oxidative stress and induces nuclear erythroid 2-related factor-2 (Nrf2) expression, which activates the antioxidant defense system. In ATII cells isolated from moderate smokers, we found DJ-1 expression by RT-PCR, and Nrf2 and heme oxygenase (HO)-1 translocation by Western blotting and immunocytofluorescence. In ATII cells isolated from heavy smokers, we detected Nrf2 and HO-1 cytoplasmic localization. Moreover, we found high oxidative stress, as detected by 4-hydroxynonenal (4-HNE) (immunoblotting), inflammation by IL-8 and IL-6 levels by ELISA, and apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay in ATII cells obtained from heavy smokers. Furthermore, we detected early DJ-1 and late Nrf2 expression after ATII cell treatment with CS extract. We also overexpressed DJ-1 by adenovirus construct and found that this restored Nrf2 and HO-1 expression and induced nuclear translocation in heavy smokers. Moreover, DJ-1 overexpression also decreased ATII cell apoptosis caused by CS extract in vitro. Our results indicate that DJ-1 activates the Nrf2-mediated antioxidant defense system. Furthermore, DJ-1 overexpression can restore the impaired Nrf2 pathway, leading to ATII cell protection in heavy smokers. This suggests a potential therapeutic strategy for targeting DJ-1 in CS-related lung diseases.

  19. Alteration of human macrophages microRNA expression profile upon infection with Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Lucinda Furci

    2013-01-01

    Conclusions: This study signifies the miRNA host response upon intracellular mycobacterial infection in macrophages, providing new aspects of regulation in host-pathogen interactions, at post-transcriptional levels.

  20. PLASMA AND LUNG MACROPHAGE CAROTENOID RESPONSIVENESS TO SUPPLEMENTATION AND OZONE EXPOSURE IN HUMANS

    Science.gov (United States)

    OBJECTIVE:: To examine the effect of ozone exposure and vegetable juice supplementation on plasma and lung macrophage concentrations of carotenoids. DESIGN:: A randomized trial. SETTING:: Subjects were exposed to ambient air prior to antioxidant supplementation and to ozone after...

  1. Extracellular traps are associated with human and mouse neutrophil and macrophage mediated killing of larval Strongyloides stercoralis.

    Science.gov (United States)

    Bonne-Année, Sandra; Kerepesi, Laura A; Hess, Jessica A; Wesolowski, Jordan; Paumet, Fabienne; Lok, James B; Nolan, Thomas J; Abraham, David

    2014-06-01

    Neutrophils are multifaceted cells that are often the immune system's first line of defense. Human and murine cells release extracellular DNA traps (ETs) in response to several pathogens and diseases. Neutrophil extracellular trap (NET) formation is crucial to trapping and killing extracellular pathogens. Aside from neutrophils, macrophages and eosinophils also release ETs. We hypothesized that ETs serve as a mechanism of ensnaring the large and highly motile helminth parasite Strongyloides stercoralis thereby providing a static target for the immune response. We demonstrated that S. stercoralis larvae trigger the release of ETs by human neutrophils and macrophages. Analysis of NETs revealed that NETs trapped but did not kill larvae. Induction of NETs was essential for larval killing by human but not murine neutrophils and macrophages in vitro. In mice, extracellular traps were induced following infection with S. stercoralis larvae and were present in the microenvironment of worms being killed in vivo. These findings demonstrate that NETs ensnare the parasite facilitating larval killing by cells of the immune system. Copyright © 2014 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  2. MIR144* inhibits antimicrobial responses against Mycobacterium tuberculosis in human monocytes and macrophages by targeting the autophagy protein DRAM2.

    Science.gov (United States)

    Kim, Jin Kyung; Lee, Hye-Mi; Park, Ki-Sun; Shin, Dong-Min; Kim, Tae Sung; Kim, Yi Sak; Suh, Hyun-Woo; Kim, Soo Yeon; Kim, In Soo; Kim, Jin-Man; Son, Ji-Woong; Sohn, Kyung Mok; Jung, Sung Soo; Chung, Chaeuk; Han, Sang-Bae; Yang, Chul-Su; Jo, Eun-Kyeong

    2017-02-01

    Autophagy is an important antimicrobial effector process that defends against Mycobacterium tuberculosis (Mtb), the human pathogen causing tuberculosis (TB). MicroRNAs (miRNAs), endogenous noncoding RNAs, are involved in various biological functions and act as post-transcriptional regulators to target mRNAs. The process by which miRNAs affect antibacterial autophagy and host defense mechanisms against Mtb infections in human monocytes and macrophages is largely uncharacterized. In this study, we show that Mtb significantly induces the expression of MIR144*/hsa-miR-144-5p, which targets the 3'-untranslated region of DRAM2 (DNA damage regulated autophagy modulator 2) in human monocytes and macrophages. Mtb infection downregulated, whereas the autophagy activators upregulated, DRAM2 expression in human monocytes and macrophages by activating AMP-activated protein kinase. In addition, overexpression of MIR144* decreased DRAM2 expression and formation of autophagosomes in human monocytes, whereas inhibition of MIR144* had the opposite effect. Moreover, the levels of MIR144* were elevated, whereas DRAM2 levels were reduced, in human peripheral blood cells and tissues in TB patients, indicating the clinical significance of MIR144* and DRAM2 in human TB. Notably, DRAM2 interacted with BECN1 and UVRAG, essential components of the autophagic machinery, leading to displacement of RUBCN from the BECN1 complex and enhancement of Ptdlns3K activity. Furthermore, MIR144* and DRAM2 were critically involved in phagosomal maturation and enhanced antimicrobial effects against Mtb. Our findings identify a previously unrecognized role of human MIR144* in the inhibition of antibacterial autophagy and the innate host immune response to Mtb. Additionally, these data reveal that DRAM2 is a key coordinator of autophagy activation that enhances antimicrobial activity against Mtb.

  3. Interaction of differentiated human adipocytes with macrophages leads to trogocytosis and selective IL-6 secretion

    OpenAIRE

    Sárvári, Anitta Kinga; Doan-Xuan, Quang-Minh; Bacsó, Zsolt; Csomós, István; Balajthy, Zoltán; Fésüs, László

    2015-01-01

    Obesity leads to adipose tissue inflammation that is characterized by increased release of proinflammatory molecules and the recruitment of activated immune cells. Although macrophages are present in the highest number among the immune cells in obese adipose tissue, not much is known about their direct interaction with adipocytes. We have introduced an ex vivo experimental system to characterize the cellular interactions and the profile of secreted cytokines in cocultures of macrophages and h...

  4. The humanized anti-human AMHRII mAb 3C23K exerts an anti-tumor activity against human ovarian cancer through tumor-associated macrophages.

    Science.gov (United States)

    Bougherara, Houcine; Némati, Fariba; Nicolas, André; Massonnet, Gérald; Pugnière, Martine; Ngô, Charlotte; Le Frère-Belda, Marie-Aude; Leary, Alexandra; Alexandre, Jérôme; Meseure, Didier; Barret, Jean-Marc; Navarro-Teulon, Isabelle; Pèlegrin, André; Roman-Roman, Sergio; Prost, Jean-François; Donnadieu, Emmanuel; Decaudin, Didier

    2017-11-21

    Müllerian inhibiting substance, also called anti-Müllerian hormone (AMH), inhibits proliferation and induces apoptosis of AMH type II receptor-positive tumor cells, such as human ovarian cancers (OCs). On this basis, a humanized glyco-engineered monoclonal antibody (3C23K) has been developed. The aim of this study was therefore to experimentally confirm the therapeutic potential of 3C23K in human OCs. We first determined by immunofluorescence, immunohistochemistry and cytofluorometry analyses the expression of AMHRII in patient's tumors and found that a majority (60 to 80% depending on the detection technique) of OCs were positive for this marker. We then provided evidence that the tumor stroma of OC is enriched in tumor-associated macrophages and that these cells are responsible for 3C23K-induced killing of tumor cells through ADCP and ADCC mechanisms. In addition, we showed that 3C23K reduced macrophages induced-T cells immunosuppression. Finally, we evaluated the therapeutic efficacy of 3C23K alone and in combination with a carboplatin-paclitaxel chemotherapy in a panel of OC Patient-Derived Xenografts. In those experiments, we showed that 3C23K significantly increased the proportion and the quality of chemotherapy-based in vivo responses. Altogether, our data support the potential interest of AMHRII targeting in human ovarian cancers and the evaluation of 3C23K in further clinical trials.

  5. Effect of neonatal malnutrition on expression of nitric oxide synthase enzyme, production of free radicals and in vitro viability of alveolar macrophages infected with methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

    Science.gov (United States)

    de Morais, Natália Gomes; da Costa, Thacianna Barreto; Pedrosa, Amanda Lúcia Farias; de Castro, Maria Carolina Accioly Brelaz; da Gonçalves de Albuquerque, Suênia Cunha; Pereira, Valéria Rêgo Alves; de Paiva Cavalcanti, Milena; de Castro, Célia Maria Machado Barbosa

    2016-02-01

    Evaluate the effects of neonatal malnutrition on the microbicidal response and viability of in vitro macrophages infected with Staphylococcus aureus sensitive/resistant to methicillin. Male Wistar rats (n = 24) were divided into two distinct groups: nourished (rats breast-fed by mothers undergoing diet with 17% casein) and malnourished (rats breast-fed by mothers undergoing diet with 8% casein). Macrophages were recovered after surgical tracheostomy procedure by collecting bronchoalveolar lavage. Four systems were established: negative control, composed only by phagocytes; positive control, macrophages plus lipopolysaccharide; and two test systems, macrophages plus Staphylococcus aureus sensitive and resistant to methicillin. Plates were incubated at 37 °C for 24 h. After this period, tests for the analysis of cell viability and microbicidal response were performed. In the statistical analysis, the Student's t and ANOVA tests were used, accepting p resistant Staphylococcus aureus. However, increased production of superoxide anion in the malnourished group was detected. Neonatal malnutrition focusing on critical periods of development promoted lower expression of iNOS, nitric oxide production, cell viability, and exacerbated reactive oxygen species production. The high levels of reactive oxygen species may favor the onset of serious and systemic infections with fatal outcome if associated with methicillin-resistant Staphylococcus aureus.

  6. Neisseria gonorrhoeae Modulates Immunity by Polarizing Human Macrophages to a M2 Profile.

    Directory of Open Access Journals (Sweden)

    María Carolina Ortiz

    Full Text Available Current data suggest that Neisseria gonorrhoeae is able to suppress the protective immune response at different levels, such as B and T lymphocytes and antigen-presenting cells. The present report is focused on gonococcus evasion mechanism on macrophages (MФ and its impact in the subsequent immune response. In response to various signals MФ may undergo classical-M1 (M1-MФ or alternative-M2 (M2-MФ activation. Until now there are no reports of the gonococcus effects on human MФ polarization. We assessed the phagocytic ability of monocyte-derived MФ (MDM upon gonococcal infection by immunofluorescence and gentamicin protection experiments. Then, we evaluated cytokine profile and M1/M2 specific-surface markers on MФ challenged with N. gonorrhoeae and their proliferative effect on T cells. Our findings lead us to suggest N. gonorrhoeae stimulates a M2-MФ phenotype in which some of the M2b and none of the M1-MФ-associated markers are induced. Interestingly, N. gonorrhoeae exposure leads to upregulation of a Programmed Death Ligand 1 (PD-L1, widely known as an immunosuppressive molecule. Moreover, functional results showed that N. gonorrhoeae-treated MФ are unable to induce proliferation of human T-cells, suggesting a more likely regulatory phenotype. Taken together, our data show that N. gonorroheae interferes with MФ polarization. This study has important implications for understanding the mechanisms of clearance versus long-term persistence of N. gonorroheae infection and might be applicable for the development of new therapeutic strategies.

  7. Uptake of [18F]fluorodeoxyglucose in human monocyte-macrophages in vitro

    International Nuclear Information System (INIS)

    Deichen, Jan Thiess; Prante, Olaf; Gack, Michaela; Schmiedehausen, Kristin; Kuwert, Torsten

    2003-01-01

    The fact that fluorine-18 fluorodeoxyglucose ([ 18 F]FDG) accumulates in inflammatory lesions as well as in tumours reduces the diagnostic specificity of positron emission tomography (PET) in oncology. The aim of this study was to characterise the uptake of [ 18 F]FDG in isolated human monocyte-macrophages (HMMs) in vitro in comparison with that in human glioblastoma (GLI) and pancreatic carcinoma cells (PAN). The purity of HMM preparations was determined by immunohistochemical staining and their functional integrity was assessed by long-term incubation with iodine-131 acetylated bovine serum albumin. [ 18 F]FDG uptake in HMMs was quantified as percent of whole [ 18 F]FDG activity per well (% ID) or as % ID in relation to total protein mass. [ 18 F]FDG uptake in HMMs significantly increased with culture duration, yielding 7.5%±0.9% (% ID/100 μg) at day 14. Stimulation by lipopolysaccharide further enhanced [ 18 F]FDG uptake in HMMs by a factor of 2. [ 18 F]FDG uptake significantly decreased with increasing glucose concentration in the medium. Radio-thin layer chromatography of intracellular metabolites revealed that [ 18 F]FDG was trapped by HMMs mainly as [ 18 F]FDG-6-phosphate and [ 18 F]FDG-1,6-diphosphate. [ 18 F]FDG uptake was in the range of uptake values measured in GLI and PAN. By accumulating [ 18 F]FDG in a manner analogous to uptake by tumour cells, activated HMMs may contribute to the [ 18 F]FDG uptake values measured by PET in neoplasms. (orig.)

  8. G-CSF regulates macrophage phenotype and associates with poor overall survival in human triple-negative breast cancer

    Science.gov (United States)

    Hollmén, Maija; Karaman, Sinem; Schwager, Simon; Lisibach, Angela; Christiansen, Ailsa J.; Maksimow, Mikael; Varga, Zsuzsanna; Jalkanen, Sirpa; Detmar, Michael

    2016-01-01

    ABSTRACT Tumor-associated macrophages (TAMs) have been implicated in the promotion of breast cancer growth and metastasis, and a strong infiltration by TAMs has been associated with estrogen receptor (ER)-negative tumors and poor prognosis. However, the molecular mechanisms behind these observations are unclear. We investigated macrophage activation in response to co-culture with several breast cancer cell lines (T47D, MCF-7, BT-474, SKBR-3, Cal-51 and MDA-MB-231) and found that high granulocyte colony-stimulating factor (G-CSF) secretion by the triple-negative breast cancer (TNBC) cell line MDA-MB-231 gave rise to immunosuppressive HLA-DRlo macrophages that promoted migration of breast cancer cells via secretion of TGF-α. In human breast cancer samples (n = 548), G-CSF was highly expressed in TNBC (p CSF blockade in the 4T1 mammary tumor model promoted maturation of MHCIIhi blood monocytes and TAMs and significantly reduced lung metastasis, anti-CSF-1R treatment promoted MHCIIloF4/80hiMRhi anti-inflammatory TAMs and enhanced lung metastasis in the presence of high G-CSF levels. Combined anti-G-CSF and anti-CSF-1R therapy significantly increased lymph node metastases, possibly via depletion of the so-called “gate-keeper” subcapsular sinus macrophages. These results indicate that G-CSF promotes the anti-inflammatory phenotype of tumor-induced macrophages when CSF-1R is inhibited and therefore caution against the use of M-CSF/CSF-1R targeting agents in tumors with high G-CSF expression. PMID:27141367

  9. The Omega-3 Fatty Acid Docosahexaenoic Acid Modulates Inflammatory Mediator Release in Human Alveolar Cells Exposed to Bronchoalveolar Lavage Fluid of ARDS Patients

    Directory of Open Access Journals (Sweden)

    Paolo Cotogni

    2015-01-01

    Full Text Available Background. This study investigated whether the 1 : 2 ω-3/ω-6 ratio may reduce proinflammatory response in human alveolar cells (A549 exposed to an ex vivo inflammatory stimulus (bronchoalveolar lavage fluid (BALF of acute respiratory distress syndrome (ARDS patients. Methods. We exposed A549 cells to the BALF collected from 12 ARDS patients. After 18 hours, fatty acids (FA were added as docosahexaenoic acid (DHA, ω-3 and arachidonic acid (AA, ω-6 in two ratios (1 : 2 or 1 : 7. 24 hours later, in culture supernatants were evaluated cytokines (TNF-α, IL-6, IL-8, and IL-10 and prostaglandins (PGE2 and PGE3 release. The FA percentage content in A549 membrane phospholipids, content of COX-2, level of PPARγ, and NF-κB binding activity were determined. Results. The 1 : 2 DHA/AA ratio reversed the baseline predominance of ω-6 over ω-3 in the cell membranes (P < 0.001. The proinflammatory cytokine release was reduced by the 1 : 2 ratio (P < 0.01 to <0.001 but was increased by the 1 : 7 ratio (P < 0.01. The 1 : 2 ratio reduced COX-2 and PGE2 (P < 0.001 as well as NF-κB translocation into the nucleus (P < 0.01, while it increased activation of PPARγ and IL-10 release (P < 0.001. Conclusion. This study demonstrated that shifting the FA supply from ω-6 to ω-3 decreased proinflammatory mediator release in human alveolar cells exposed to BALF of ARDS patients.

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

    Science.gov (United States)

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

    2014-01-01

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

  11. Alveolar proteinosis associated with aluminium dust inhalation.

    Science.gov (United States)

    Chew, R; Nigam, S; Sivakumaran, P

    2016-08-01

    Secondary alveolar proteinosis is a rare lung disease which may be triggered by a variety of inhaled particles. The diagnosis is made by detection of anti-granulocyte-macrophage colony-stimulating factor antibodies in bronchoalveolar lavage fluid, which appears milky white and contains lamellar bodies. Aluminium has been suggested as a possible cause, but there is little evidence in the literature to support this assertion. We report the case of a 46-year-old former boilermaker and boat builder who developed secondary alveolar proteinosis following sustained heavy aluminium exposure. The presence of aluminium was confirmed both by histological examination and metallurgical analysis of a mediastinal lymph node. Despite cessation of exposure to aluminium and treatment with whole-lung lavage which normally results in improvements in both symptoms and lung function, the outcome was poor and novel therapies are now being used for this patient. It may be that the natural history in aluminium-related alveolar proteinosis is different, with the metal playing a mediating role in the disease process. Our case further supports the link between aluminium and secondary alveolar proteinosis and highlights the need for measures to prevent excessive aluminium inhalation in relevant industries. © The Author 2016. Published by Oxford University Press on behalf of the Society of Occupational Medicine. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  12. Pulmonary Alveolar Proteinosis in Setting of Inhaled Toxin Exposure and Chronic Substance Abuse

    Directory of Open Access Journals (Sweden)

    Meirui Li

    2018-01-01

    Full Text Available Pulmonary alveolar proteinosis (PAP is a rare lung disorder in which defects in alveolar macrophage maturation or function lead to the accumulation of proteinaceous surfactant in alveolar space, resulting in impaired gas exchange and hypoxemia. PAP is categorized into three types: hereditary, autoimmune, and secondary. We report a case of secondary PAP in a 47-year-old man, whose risk factors include occupational exposure to inhaled toxins, especially aluminum dust, the use of anabolic steroids, and alcohol abuse, which in mice leads to alveolar macrophage dysfunction through a zinc-dependent mechanism that inhibits granulocyte macrophage-colony stimulating factor (GM-CSF receptor signalling. Although the rarity and vague clinical presentation of PAP can pose diagnostic challenges, clinician awareness of PAP risk factors may facilitate the diagnostic process and lead to more prompt treatment.

  13. Systematic Review and Meta-Analysis of Recombinant Human Bone Morphogenetic Protein-2 in Localized Alveolar Ridge and Maxillary Sinus Augmentation.

    Science.gov (United States)

    Kelly, Mick P; Vaughn, Olushola L Akinshemoyin; Anderson, Paul A

    2016-05-01

    Recombinant human bone morphogenetic protein-2 (rhBMP-2) is approved by the Food and Drug Administration as a viable alternative to bone graft in spinal fusion and maxillary sinus lift. The research questions for meta-analysis were: Is rhBMP-2 an effective bone graft substitute in localized alveolar ridge augmentation and maxillary sinus floor augmentation? What are the potential adverse events? A search of MEDLINE from January 1980 to January 2014 using PubMed, the Cochrane Database of Systematic Reviews and Controlled Trials, CINAHL, and EMBASE was performed. Searches were performed from Medical Subject Headings. The quality of each study included was graded by Review Manager software. The primary outcome variable was bone formation measured as change in bone height on computed tomogram. A systematic review of adverse events also was performed. A random-effects model was chosen. Continuous variables were calculated using the standardized mean difference and 95% confidence intervals (CIs) comparing improvement from baseline of the experimental group with that of the control group. Change in bone height was calculated using logarithmic odds ratio. Test of significance used the Z statistic with a P value of .05. Ten studies met the criteria for systematic review; 8 studies were included in the meta-analysis. Five studies assessed localized alveolar ridge augmentation and resulted in an overall standardized mean difference of 0.56 (CI, 0.20-0.92) in favor of BMP; this result was statistically important. Three studies assessed maxillary sinus floor augmentation and resulted in an overall standardized mean difference of -0.50 (CI, -0.93 to -0.09), which was meaningfully different in favor of the control group. Adverse events were inconsistently reported, ranging from no complications to widespread adverse events. For localized alveolar ridge augmentation, this meta-analysis showed that rhBMP-2 substantially increases bone height. However, rhBMP-2 does not perform as

  14. Environmental Legionella spp. collected in urban test sites of South East Queensland, Australia, are virulent to human macrophages in vitro.

    Science.gov (United States)

    Lawrence, Amba; Eglezos, Sofroni; Huston, Wilhelmina

    2016-01-01

    Legionellae are frequent contaminants of potable water supplies, resulting in sporadic infections and occasional outbreaks. Isolates of Legionella were collected from urban test sites within South East Queensland and evaluated for their virulence potential in vitro. Two strains (from the species Legionella londiniensis and Legionella quinlivanii) were demonstrated to have the ability to infect human macrophages, while a strain from the species Legionella anisa did not maintain an infection over the same time course. This suggests that the spectrum of urban environmentally associated Legionella with potential to cause human disease might be greater than currently considered. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  15. Nanoparticles as Antituberculosis Drugs Carriers: Effect on Activity Against Mycobacterium tuberculosis in Human Monocyte-Derived Macrophages

    International Nuclear Information System (INIS)

    Anisimova, Y.V.; Gelperina, S.I.; Peloquin, C.A.; Heifets, L.B.

    2000-01-01

    This is the first report evaluating the nanoparticle delivery system for three antituberculosis drugs: isoniazid, rifampin, and streptomycin. The typical particle size is 250 nm. We studied accumulation of these drugs in human monocytes as well as their antimicrobial activity against Mycobacterium tuberculosis residing in human monocyte-derived macrophages. Nanoparticle encapsulation increased the intracellular accumulation (cell-association) of all three tested drugs, but it enhanced the antimicrobial activity of isoniazid and streptomycin only. On the other hand, the activity of encapsulated rifampin against intracellular bacteria was not higher than that of the free drug

  16. CD1 molecule expression on human monocytes induced by granulocyte-macrophage colony-stimulating factor.

    Science.gov (United States)

    Kasinrerk, W; Baumruker, T; Majdic, O; Knapp, W; Stockinger, H

    1993-01-15

    In this paper we demonstrate that granulocyte-macrophage CSF (GM-CSF) specifically induces the expression of CD1 molecules, CD1a, CD1b and CD1c, upon human monocytes. CD1 molecules appeared upon monocytes on day 1 of stimulation with rGM-CSF, and expression was up-regulated until day 3. Monocytes cultured in the presence of LPS, FMLP, PMA, recombinant granulocyte-CSF, rIFN-gamma, rTNF-alpha, rIL-1 alpha, rIL-1 beta, and rIL-6 remained negative. The induction of CD1 molecules by rGM-CSF was restricted to monocytes, since no such effect was observed upon peripheral blood granulocytes, PBL, and the myeloid cell lines Monomac1, Monomac6, MV4/11, HL60, U937, THP1, KG1, and KG1A. CD1a mRNA was detectable in rGM-CSF-induced monocytes but not in those freshly isolated. SDS-PAGE and immunoblotting analyses of CD1a mAb VIT6 immunoprecipitate from lysate of rGM-CSF-activated monocytes revealed an appropriate CD1a polypeptide band of 49 kDa associated with beta 2-microglobulin. Expression of CD1 molecules on monocytes complements the distribution of these structures on accessory cells, and their specific induction by GM-CSF strengthens the suggestion that CD1 is a family of crucial structures required for interaction between accessory cells and T cells.

  17. 15-Lipoxygenases regulate the production of chemokines in human lung macrophages.

    Science.gov (United States)

    Abrial, C; Grassin-Delyle, S; Salvator, H; Brollo, M; Naline, E; Devillier, P

    2015-09-01

    15-Lipoxygenase (15-LOX) activity is associated with inflammation and immune regulation. The objectives of the present study were to investigate the expression of 15-LOX-1 and 15-LOX-2 and evaluate the enzymes' roles in the polarization of human lung macrophages (LMs) in response to LPS and Th2 cytokines (IL-4/-13). LMs were isolated from patients undergoing surgery for carcinoma. The cells were cultured with a 15-LOX inhibitor (PD146176 or ML351), a COX inhibitor (indomethacin), a 5-LOX inhibitor (MK886) or vehicle and then stimulated with LPS (10 ng · mL(-1)), IL-4 (10 ng · mL(-1)) or IL-13 (50 ng · mL(-1)) for 24 h. Levels of ALOX15 (15-LOX-1) and ALOX15B (15-LOX-2) transcripts were determined by real-time quantitative PCR. Immunoassays were used to measure levels of LPS-induced cytokines (TNF-α, CCL2, CCL3, CCL4, CXCL1, CXCL8 and CXCL10) and Th2 cytokine-induced chemokines (CCL13, CCL18 and CCL22) in the culture supernatant. Stimulation of LMs with LPS was associated with increased expression of ALOX15B, whereas stimulation with IL-4/IL-13 induced the expression of ALOX15. PD146176 and ML351 (10 μM) reduced the release of the chemokines induced by LPS and Th2 cytokines. The effects of these 15-LOX inhibitors were maintained in the presence of indomethacin and MK886. Furthermore, indomethacin revealed the inhibitory effect of PD146176 on TNF-α release. Inhibition of the 15-LOX pathways is involved in the down-regulation of the in vitro production of chemokines in LMs. Our results suggest that the 15-LOX pathways have a role in the pathogenesis of inflammatory lung disorders and may thus constitute a potential drug target. © 2015 The British Pharmacological Society.

  18. A mature macrophage is a principal HIV-1 cellular reservoir in humanized mice after treatment with long acting antiretroviral therapy.

    Science.gov (United States)

    Araínga, Mariluz; Edagwa, Benson; Mosley, R Lee; Poluektova, Larisa Y; Gorantla, Santhi; Gendelman, Howard E

    2017-03-09

    Despite improved clinical outcomes seen following antiretroviral therapy (ART), resting CD4+ T cells continue to harbor latent human immunodeficiency virus type one (HIV-1). However, such cells are not likely the solitary viral reservoir and as such defining where and how others harbor virus is imperative for eradication measures. To such ends, we used HIV-1 ADA -infected NOD.Cg-Prkdc scid Il2rg tm1Wjl /SzJ mice reconstituted with a human immune system to explore two long-acting ART regimens investigating their abilities to affect viral cell infection and latency. At 6 weeks of infection animals were divided into four groups. One received long-acting (LA) cabotegravir (CAB) and rilpivirine (RVP) (2ART), a second received LA CAB, lamivudine, abacavir and RVP (4ART), a third were left untreated and a fourth served as an uninfected control. After 4 weeks of LA ART treatment, blood, spleen and bone marrow (BM) cells were collected then phenotypically characterized. CD4+ T cell subsets, macrophages and hematopoietic progenitor cells were analyzed for HIV-1 nucleic acids by droplet digital PCR. Plasma viral loads were reduced by two log 10 or to undetectable levels in the 2 and 4ART regimens, respectively. Numbers and distributions of CD4+ memory and regulatory T cells, macrophages and hematopoietic progenitor cells were significantly altered by HIV-1 infection and by both ART regimens. ART reduced viral DNA and RNA in all cell and tissue compartments. While memory cells were the dominant T cell reservoir, integrated HIV-1 DNA was also detected in the BM and spleen macrophages in both regimen-treated mice. Despite vigorous ART regimens, HIV-1 DNA and RNA were easily detected in mature macrophages supporting their potential role as an infectious viral reservoir.

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

    Directory of Open Access Journals (Sweden)

    Eyayu Belay

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

  20. A Comparative Analysis of Recombinant Human Bone Morphogenetic Protein-2 with a Demineralized Bone Matrix versus Iliac Crest Bone Graft for Secondary Alveolar Bone Grafts in Patients with Cleft Lip and Palate: Review of 501 Cases.

    Science.gov (United States)

    Hammoudeh, Jeffrey A; Fahradyan, Artur; Gould, Daniel J; Liang, Fan; Imahiyerobo, Thomas; Urbinelli, Leo; Nguyen, JoAnna T; Magee, William; Yen, Stephen; Urata, Mark M

    2017-08-01

    Alveolar cleft reconstruction using iliac crest bone graft is considered standard of care for children with complete cleft lip and palate at the time of mixed dentition. Harvesting bone may result in donor-site morbidity and additional operating time and length of hospitalization. Recombinant human bone morphogenetic protein (rhBMP)-2 with a demineralized bone matrix is an alternative bone source for alveolar cleft reconstruction. The authors investigated the outcomes of rhBMP-2/demineralized bone matrix versus iliac crest bone graft for alveolar cleft reconstruction by reviewing postoperative surgical complications and cleft closure. A retrospective chart review was conducted for 258 rhBMP-2/demineralized bone matrix procedures (mean follow-up, 2.9 years) and 243 iliac crest bone graft procedures (mean follow-up, 4.1 years) on 414 patients over a 12-year period. The authors compared complications, canine eruption, and alveolar cleft closure between the two groups. In the rhBMP-2/demineralized bone matrix group, one patient required prolonged intubation because of intraoperative airway swelling not thought to be caused by rhBMP-2, 36 reported facial swelling and one required outpatient steroids as treatment, and 12 had dehiscence; however, half of these complications resolved without intervention. Twenty-three of the 228 rhBMP-2/demineralized bone matrix patients and 28 of the 242 iliac crest bone graft patients required repeated surgery for alveolar cleft repair. Findings for canine tooth eruption into the cleft site through the graft were similar between the groups. The rhBMP-2/demineralized bone matrix appears to be an acceptable alternative for alveolar cleft repair. The authors found no increase in serious adverse events with the use of this material. Local complications, such as swelling and minor wound dehiscence, predominantly improved without intervention. Therapeutic, III.

  1. Importance of Physical and Physiological Parameters in Simulated Particle Transport in the Alveolar Zone of the Human Lung

    Directory of Open Access Journals (Sweden)

    Dogan Ciloglu

    2017-01-01

    Full Text Available The trajectory and deposition efficiency of micron-sized (1–5 µm particles, inhaled into the pulmonary system, are accurately determined with the aid of a newly developed model and modified simulation techniques. This alveolar model, which has a simple but physiologically appropriate geometry, and the utilized fluid structure interaction (FSI methods permit the precise simulation of tissue wall deformation and particle fluid interactions. The relation between tissue movement and airflow in the alveolated duct is solved by a two-way fluid structure interaction simulation technique, using ANSYS Workbench (Release 16.0, ANSYS INC., Pittsburgh, PA, USA, 2015. The dynamic transport of particles and their deposition are investigated as a function of aerodynamic particle size, tissue visco-elasticity, tidal breathing period, gravity orientation and particle–fluid interactions. It is found that the fluid flows and streamlines differ between the present flexible model and rigid models, and the two-way coupling particle trajectories vary relative to one-way particle coupling. In addition, the results indicate that modelling the two-way coupling particle system is important because the two-way discrete phase method (DPM approach despite its complexity provides more extensive particle interactions and is more reliable than transport results from the one-way DPM approach. The substantial difference between the results of the two approaches is likely due to particle–fluid interactions, which re-suspend the sediment particles in the airway stream and hence pass from the current generation.

  2. Downregulation of host tryptophan-aspartate containing coat (TACO gene restricts the entry and survival of Leishmania donovani in human macrophage model

    Directory of Open Access Journals (Sweden)

    Venkateswara Reddy Gogulamudi

    2015-10-01

    Full Text Available Leishmania are obligate intracellular protozoan parasites of mammalian hosts. Promastigotes of Leishmania are internalized by macrophages and transformed into amastigotes in phagosomes, and replicate in phagolysosomes. Phagosomal maturation arrest is known to play a central role in the survival of pathogenic Leishmania within activated macrophages. Recently, tryptophan-aspartate containing coat (TACO gene has been recognized as playing a crucial role in the survival of Mycobacterium tuberculosis within human macrophages by arresting the phagosome maturation process. We postulated that a similar association of TACO gene with phagosomes would prevent the vacuole from maturation in the case of Leishmania. In this study we attempted to define the effect of TACO gene downregulation on the uptake/survival of Leishmania donovani intracellularly, by treatment with Vitamin D3/Retinoic acid (RA & Chenodeoxycholic acid (CDCA/Retinoic acid (RA combinations in human THP-1 macrophages (in vitro. Treatment with these molecules downregulated the TACO gene in macrophages, resulting in reduced parasite load and marked reduction of disease progression in L. donovani infected macrophages. Taken together, these results suggest that TACO gene downregulation may play a role in subverting macrophage machinery in establishing the L.donovani replicative niche inside the host. Our study is the first to highlight the importantrole of the TACO gene in Leishmania entry, and to identify TACO gene downregulation as potential drug target against leishmaniasis.

  3. Targeted Delivery of Glucan Particle Encapsulated Gallium Nanoparticles Inhibits HIV Growth in Human Macrophages

    Directory of Open Access Journals (Sweden)

    Ernesto R. Soto

    2016-01-01

    Full Text Available Glucan particles (GPs are hollow, porous 3–5 μm microspheres derived from the cell walls of Baker’s yeast (Saccharomyces cerevisiae. The 1,3-β-glucan outer shell provides for receptor-mediated uptake by phagocytic cells expressing β-glucan receptors. GPs have been used for macrophage-targeted delivery of a wide range of payloads (DNA, siRNA, protein, small molecules, and nanoparticles encapsulated inside the hollow GPs or bound to the surface of chemically derivatized GPs. Gallium nanoparticles have been proposed as an inhibitory agent against HIV infection. Here, macrophage targeting of gallium using GPs provides for more efficient delivery of gallium and inhibition of HIV infection in macrophages compared to free gallium nanoparticles.

  4. Macrophage specific overexpression of the human macrophage scavenger receptor in transgenic mice, using a 180-kb yeast artificial chromosome, leads to enhanced foam cell formation of isolated peritoneal macrophages

    NARCIS (Netherlands)

    de Winther, M. P.; van Dijk, K. W.; van Vlijmen, B. J.; Gijbels, M. J.; Heus, J. J.; Wijers, E. R.; van den Bos, A. C.; Breuer, M.; Frants, R. R.; Havekes, L. M.; Hofker, M. H.

    1999-01-01

    Macrophage scavenger receptors class A (MSR) are thought to play an important role in atherogenesis by mediating the unrestricted uptake of modified lipoproteins by macrophages in the vessel wall leading to foam cell formation. To investigate the in vivo role of the MSR in this process, a transgenic

  5. The toxicity of rifampicin polylactic acid nanoparticles against Mycobacterium bovis BCG and human macrophage THP-1 cell line

    International Nuclear Information System (INIS)

    Erokhina, M; Rybalkina, E; Lepekha, L; Barsegyan, G; Onishchenko, G

    2015-01-01

    Tuberculosis is rapidly becoming a major health problem. The rise in tuberculosis incidence stimulates efforts to develop more effective delivery systems for the existing antituberculous drugs while decreasing the side effects. The nanotechnology may provide novel drug delivery tools allowing controlled drug release. Rifampicin is one of the main antituberculous drugs, characterized by high toxicity, and Poly (L-lactic acid) (PLLA) is a biodegradable polymer used for the preparation of encapsulated drugs. The aim of our work was to evaluate the toxicity of rifampicin-PLLA nanoparticles against Mycobacterium bovis BCG using human macrophage THP-1 cell line. Our data demonstrate that rifampicin-PLLA is effective against M. bovis BCG in the infected macrophages. The drug is inducing the dysfunction of mitochondria and apoptosis in the macrophages and is acting as a potential substrate of Pgp thereby modulating cell chemosensitivity. The severity of the toxic effects of the rifampicin-PLLA nanoparticles is increasing in a dose-dependent manner. We suggest that free rifampicin induces death of M. bovis BCG after PLLA degradation and diffusion from phago-lysosomes to cytoplasm causing mitochondria dysfunction and affecting the Pgp activity. (paper)

  6. Haemophilus ducreyi infection induces activation of the NLRP3 inflammasome in nonpolarized but not in polarized human macrophages.

    Science.gov (United States)

    Li, Wei; Katz, Barry P; Bauer, Margaret E; Spinola, Stanley M

    2013-08-01

    Recognition of microbial infection by certain intracellular pattern recognition receptors leads to the formation of a multiprotein complex termed the inflammasome. Inflammasome assembly activates caspase-1 and leads to cleavage and secretion of the proinflammatory cytokines interleukin-1 beta (IL-1β) and IL-18, which help control many bacterial pathogens. However, excessive inflammation mediated by inflammasome activation can also contribute to immunopathology. Here, we investigated whether Haemophilus ducreyi, a Gram-negative bacterium that causes the genital ulcer disease chancroid, activates inflammasomes in experimentally infected human skin and in monocyte-derived macrophages (MDM). Although H. ducreyi is predominantly extracellular during human infection, several inflammasome-related components were transcriptionally upregulated in H. ducreyi-infected skin. Infection of MDM with live, but not heat-killed, H. ducreyi induced caspase-1- and caspase-5-dependent processing and secretion of IL-1β. Blockage of H. ducreyi uptake by cytochalasin D significantly reduced the amount of secreted IL-1β. Knocking down the expression of the inflammasome components NLRP3 and ASC abolished IL-1β production. Consistent with NLRP3-dependent inflammasome activation, blocking ATP signaling, K(+) efflux, cathepsin B activity, and lysosomal acidification all inhibited IL-1β secretion. However, inhibition of the production and function of reactive oxygen species did not decrease IL-1β production. Polarization of macrophages to classically activated M1 or alternatively activated M2 cells abrogated IL-1β secretion elicited by H. ducreyi. Our study data indicate that H. ducreyi induces NLRP3 inflammasome activation via multiple mechanisms and suggest that the heterogeneity of macrophages within human lesions may modulate inflammasome activation during human infection.

  7. Long-term follow-up of metabolic activity in human alveolar echinococcosis using FDG-PET

    International Nuclear Information System (INIS)

    Reuter, S.; Gruener, B.; Kern, P.; Buck, A.K.; Blumstein, N.; Reske, S.N.

    2008-01-01

    Aim: [ 18 F]fluoro-deoxyglucose positron-emission-tomography (FDG-PET) detects metabolic activity in alveolar echinococcosis (AE). The slow changes in metabolic and morphological characteristics require long-term follow-up of patients. This is the first study to evaluate metabolic activity over may years, hereby assessing the utility of FDG-PET for the evaluation of disease progression and response to treatment. Patients, methods: 15 patients received a follow-up FDG-PET combined with computed tomography (integrated PET/CT) with a median of 6.5 years after the first PET in 1999. Number and location of enhanced metabolic activity in the area of AE lesions was determined. Quantification of intensity of metabolic activity was assessed by calculation of mean standardized uptake values. Results: AE lesions in 11/15 patients had been metabolically inactive initially, but only two showed permanent inactivity over the course of 81 months. Interestingly, in two patients metabolic activity was newly detected after 80 and 82 months. Benzimidazole treatment was intermittently discontinued in seven cases. Persisting activity at FDG-PET demanded continued benzimidazole treatment in four patients. Neither treatment duration, lesional size, calcifications nor regressive changes correlated with metabolic activity. Conclusion: treatment responses are heterogeneous and vary from progressive disease despite treatment to long-term inactive disease with discontinued treatment. Lack of metabolic activity indicates suppressed parasite activity and is not equivalent to parasite death. However, metabolic activity may remain suppressed for years, allowing for temporary treatment discontinuation. Relapses are reliably detected with PET and restarting benzimidazole treatment prevents parasite expansion. (orig.)

  8. Xylitol, an anticaries agent, exhibits potent inhibition of inflammatory responses in human THP-1-derived macrophages infected with Porphyromonas gingivalis.

    Science.gov (United States)

    Park, Eunjoo; Na, Hee Sam; Kim, Sheon Min; Wallet, Shannon; Cha, Seunghee; Chung, Jin

    2014-06-01

    Xylitol is a well-known anticaries agent and has been used for the prevention and treatment of dental caries. In this study, the anti-inflammatory effects of xylitol are evaluated for possible use in the prevention and treatment of periodontal infections. Cytokine expression was stimulated in THP-1 (human monocyte cell line)-derived macrophages by live Porphyromonas gingivalis, and enzyme-linked immunosorbent assay and a commercial multiplex assay kit were used to determine the effects of xylitol on live P. gingivalis-induced production of cytokine. The effects of xylitol on phagocytosis and the production of nitric oxide were determined using phagocytosis assay, viable cell count, and Griess reagent. The effects of xylitol on P. gingivalis adhesion were determined by immunostaining, and costimulatory molecule expression was examined by flow cytometry. Live P. gingivalis infection increased the production of representative proinflammatory cytokines, such as tumor necrosis factor-α and interleukin (IL)-1β, in a multiplicity of infection- and time-dependent manner. Live P. gingivalis also enhanced the release of cytokines and chemokines, such as IL-12 p40, eotaxin, interferon γ-induced protein 10, monocyte chemotactic protein-1, and macrophage inflammatory protein-1. The pretreatment of xylitol significantly inhibited the P. gingivalis-induced cytokines production and nitric oxide production. In addition, xylitol inhibited the attachment of live P. gingivalis on THP-1-derived macrophages. Furthermore, xylitol exerted antiphagocytic activity against both Escherichia coli and P. gingivalis. These findings suggest that xylitol acts as an anti-inflammatory agent in THP-1-derived macrophages infected with live P. gingivalis, which supports its use in periodontitis.

  9. Cathepsin D Specifically Cleaves the Chemokines Macrophage Inflammatory Protein-1α, Macrophage Inflammatory Protein-1β, and SLC That Are Expressed in Human Breast Cancer

    Science.gov (United States)

    Wolf, Marlene; Clark-Lewis, Ian; Buri, Caroline; Langen, Hanno; Lis, Maddalena; Mazzucchelli, Luca

    2003-01-01

    Cathepsin D (Cath-D) expression in human primary breast cancer has been associated with a poor prognosis. In search of a better understanding of the Cath-D substrates possibly involved in cancer invasiveness and metastasis, we investigated the potential interactions between this protease and chemokines. Here we report that purified Cath-D, as well as culture supernatants from the human breast carcinoma cell lines MCF-7 and T47D, selectively degrade macrophage inflammatory protein (MIP)-1α (CCL3), MIP-1β (CCL4), and SLC (CCL21). Proteolysis was totally blocked by the protease inhibitor pepstatin A, and specificity of Cath-D cleavage was demonstrated using a large chemokine panel. Whereas MIP-1α and MIP-1β degradation was rapid and complete, cleavage of SLC was slow and not complete. Mass spectrometry analysis showed that Cath-D cleaves the Leu58 to Trp59 bond of SLC producing two functionally inactive fragments. Analysis of Cath-D proteolysis of a series of monocyte chemoattractant protein-3/MIP-1β hybrids indicated that processing of MIP-1β might start by cleaving off amino acids located in the C-terminal domain. In situ hybridization studies revealed MIP-1α, MIP-1β, and Cath-D gene expression mainly in the stromal compartment of breast cancers whereas SLC transcripts were found in endothelial cells of capillaries and venules within the neoplastic tissues. Cath-D production in the breast carcinoma cell lines MCF-7 and T47D, as assessed by enzyme-linked immunosorbent assay of culture supernatants and cell lysates, was not affected by stimulation with chemokines such as interleukin-8 (CXCL8), SDF-1 (CXCL12), and SLC. These data suggest that inactivation of chemokines by Cath-D possibly influences regulatory mechanisms in the tumoral extracellular microenvironment that in turn may affect the generation of the antitumoral immune response, the migration of cancer cells, or both processes. PMID:12651610

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-12-01

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

  11. Human macrophages primed with angiogenic factors show dynamic plasticity, irrespective of extracellular matrix components

    NARCIS (Netherlands)

    Ploeger, Diana T. A.; van Putten, Sander M.; Koerts, Jasper A.; van Luyn, Marja J. A.; Harmsen, Martin C.

    Macrophages are important in inflammation as well as in tissue repair processes. They can be activated by various stimuli and classified into two major groups: M1 (classically activated) or M2 (alternatively activated). Inflammation, angiogenesis and matrix remodeling play a major role in tissue

  12. Modeling receptor-mediated endocytosis of polymer-functionalized iron oxide nanoparticles by human macrophages

    Czech Academy of Sciences Publication Activity Database

    Lunov, O.; Zablotskyy, Vitaliy A.; Syrovets, T.; Röcker, C.; Tron, K.; Nienhaus, G.U.; Simmet, T.

    2011-01-01

    Roč. 32, č. 2 (2011), s. 547-555 ISSN 0142-9612 Institutional research plan: CEZ:AV0Z10100522 Keywords : macrophage * nanoparticle * bioabsorption * modeling * antisense * MRI (magnetic resonance imaging) Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 7.404, year: 2011 http://www.sciencedirect.com/science/article/pii/S014296121001149X

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

    International Nuclear Information System (INIS)

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

    2013-01-01

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

  14. Placental macrophage contact potentiates the complete replicative cycle of human cytomegalovirus in syncytiotrophoblast cells: role of interleukin-8 and transforming growth factor-beta1.

    Science.gov (United States)

    Bácsi, A; Aranyosi, J; Beck, Z; Ebbesen, P; Andirkó, I; Szabó, J; Lampé, L; Kiss, J; Gergely, L; Tóth, F D

    1999-10-01

    Although syncytiotrophoblast (ST) cells can be infected by human cytomegalovirus (HCMV), in vitro studies have indicated that ST cells do not support the complete viral reproductive cycle, or HCMV replication may occur in less than 3% of ST cells. The present study tested the possibility that placental macrophages might enhance activation of HCMV carried in ST cells and, further, that infected ST cells would be capable of transmitting virus to neighboring macrophages. For this purpose, we studied HCMV replication in ST cells grown alone or cocultured with uninfected placental macrophages. Our results demonstrated that HCMV gene expression in ST cells was markedly upregulated by coculture with macrophages, resulting in release of substantial amounts of infectious virus from HCMV-infected ST cells. After having become permissive for viral replication, ST cells delivered HCMV to the cocultured macrophages, as evidenced by detection of virus-specific antigens in these cells. The stimulatory effect of coculture on HCMV gene expression in ST cells was mediated by marked interleukin-8 (IL-8) and transforming growth factor-beta1 (TGF-beta1) release from macrophages, an effect caused by contact between the different placental cells. Our findings indicate an interactive role for the ST layer and placental macrophages in the dissemination of HCMV among placental tissue. Eventually, these interactions may contribute to the transmission of HCMV from mother to the fetus.

  15. Functional characterization and phenotypic monitoring of human hematopoietic stem cell expansion and differentiation of monocytes and macrophages by whole-cell mass spectrometry

    Directory of Open Access Journals (Sweden)

    Guido Vogel

    2018-01-01

    Full Text Available The different facets of macrophages allow them to play distinct roles in tissue homeostasis, tissue repair and in response to infections. Individuals displaying dysregulated macrophage functions are proposed to be prone to inflammatory disorders or infections. However, this being a cause or a consequence of the pathology remains often unclear. In this context, we isolated and expanded CD34+ HSCs from healthy blood donors and derived them into CD14+ myeloid progenitors which were further enriched and differentiated into macrophages. Aiming for a comprehensive phenotypic profiling, we generated whole-cell mass spectrometry (WCMS fingerprints of cell samples collected along the different stages of the differentiation process to build a predictive model using a linear discriminant analysis based on principal components. Through the capacity of the model to accurately predict sample's identity of a validation set, we demonstrate that WCMS profiles obtained from bona fide blood monocytes and respectively derived macrophages mirror profiles obtained from equivalent HSC derivatives. Finally, HSC-derived macrophage functionalities were assessed by quantifying cytokine and chemokine responses to a TLR agonist in a 34-plex luminex assay and by measuring their capacity to phagocytise mycobacteria. These functional read-outs could not discriminate blood monocytes-derived from HSC-derived macrophages. To conclude, we propose that this method opens new avenues to distinguish the impact of human genetics on the dysregulated biological properties of macrophages in pathological conditions.

  16. Dual prognostic significance of tumour-associated macrophages in human pancreatic adenocarcinoma treated or untreated with chemotherapy.

    Science.gov (United States)

    Di Caro, Giuseppe; Cortese, Nina; Castino, Giovanni Francesco; Grizzi, Fabio; Gavazzi, Francesca; Ridolfi, Cristina; Capretti, Giovanni; Mineri, Rossana; Todoric, Jelena; Zerbi, Alessandro; Allavena, Paola; Mantovani, Alberto; Marchesi, Federica

    2016-10-01

    Tumour-associated macrophages (TAMs) play key roles in tumour progression. Recent evidence suggests that TAMs critically modulate the efficacy of anticancer therapies, raising the prospect of their targeting in human cancer. In a large retrospective cohort study involving 110 patients with pancreatic ductal adenocarcinoma (PDAC), we assessed the density of CD68-TAM immune reactive area (%IRA) at the tumour-stroma interface and addressed their prognostic relevance in relation to postsurgical adjuvant chemotherapy (CTX). In vitro, we dissected the synergism of CTX and TAMs. In human PDAC, TAMs predominantly exhibited an immunoregulatory profile, characterised by expression of scavenger receptors (CD206, CD163) and production of interleukin 10 (IL-10). Surprisingly, while the density of TAMs associated to worse prognosis and distant metastasis, CTX restrained their protumour prognostic significance. High density of TAMs at the tumour-stroma interface positively dictated prognostic responsiveness to CTX independently of T-cell density. Accordingly, in vitro, gemcitabine-treated macrophages became tumoricidal, activating a cytotoxic gene expression programme, inhibiting their protumoural effect and switching to an antitumour phenotype. In patients with human PDAC, neoadjuvant CTX was associated to a decreased density of CD206(+) and IL-10(+) TAMs at the tumour-stroma interface. Overall, our data highlight TAMs as critical determinants of prognostic responsiveness to CTX and provide clinical and in vitro evidence that CTX overall directly re-educates TAMs to restrain tumour progression. These results suggest that the quantification of TAMs could be exploited to select patients more likely to respond to CTX and provide the basis for novel strategies aimed at re-educating macrophages in the context of CTX. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

  17. Human-induced pluripotent stem cell-derived macrophages and their immunological function in response to tuberculosis infection.

    Science.gov (United States)

    Hong, Danping; Ding, Jiongyan; Li, Ouyang; He, Quan; Ke, Minxia; Zhu, Mengyi; Liu, Lili; Ou, Wen-Bin; He, Yulong; Wu, Yuehong

    2018-02-26

    Induced pluripotent stem cells (iPS) represent an innovative source for the standardized in vitro generation of macrophages (Mφ). Mφ show great promise in disease pathogenesis, particularly tuberculosis. However, there is no information about human iPS-derived (hiPS) macrophages (hiPS-Mφ) in response to tuberculosis infection. In the present study, macrophages derived from hiPS were established via embryoid body (EB) formation by using feeder-free culture conditions, and the human monocyte cell line THP-1 (THP-1-Mφ) was used as control. iPS-Mφ were characterized by using morphology, Giemsa staining, nonspecific esterase staining (α-NAE), phagocytosis, and surface phenotype. Additionally, after treatment with Bacillus Calmette-Guérin (BCG) for 24 h, cell apoptosis was detected by using an Annexin V-FITC Apoptosis Detection assay. The production of nitric oxide (NO), expression of tumor necrosis factor alpha (TNF-α), activity of apoptosis-related protein cysteine-3 (Caspase-3) and expression of B-cell lymphoma-2 (Bcl-2) were analyzed. With respect to morphology, surface phenotype, and function, the iPS-Mφ closely resembled their counterparts generated in vitro from a human monocyte cell line. iPS-Mφ exhibited the typically morphological characteristics of macrophages, such as round, oval, fusiform and irregular characteristics. The cells were Giemsa-stained-positive, α-NAE-positive, and possessed phagocytic ability. iPS-Mφ express high levels of CD14, CD11b, CD40, CD68, and major histocompatibility complex II (MHC-II). Moreover, with regard to the apoptotic rate, the production of NO, expression of TNF-α, and activity of Caspase-3 and Bcl-2, iPS-Mφ closely resemble that of their counterparts generated in vitro from human monocyte cell line in response to BCG infection. The rate of apoptosis of BCG-treated iPS-Mφ was 37.77 ± 7.94% compared to that of the untreated group at 4.97 ± 1.60% (P immunological function in response to Bacillus Calmette

  18. Development of an Intracellular Screen for New Compounds Able To Inhibit Mycobacterium tuberculosis Growth in Human Macrophages.

    Science.gov (United States)

    Sorrentino, Flavia; Gonzalez del Rio, Ruben; Zheng, Xingji; Presa Matilla, Jesus; Torres Gomez, Pedro; Martinez Hoyos, Maria; Perez Herran, Maria Esther; Mendoza Losana, Alfonso; Av-Gay, Yossef

    2016-01-01

    Here we describe the development and validation of an intracellular high-throughput screening assay for finding new antituberculosis compounds active in human macrophages. The assay consists of a luciferase-based primary identification assay, followed by a green fluorescent protein-based secondary profiling assay. Standard tuberculosis drugs and 158 previously recognized active antimycobacterial compounds were used to evaluate assay robustness. Data show that the assay developed is a short and valuable tool for the discovery of new antimycobacterial compounds. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  19. Reactive oxygen species mediated DNA damage in human lung alveolar epithelial (A549) cells from exposure to non-cytotoxic MFI-type zeolite nanoparticles.

    Science.gov (United States)

    Bhattacharya, Kunal; Naha, Pratap C; Naydenova, Izabela; Mintova, Svetlana; Byrne, Hugh J

    2012-12-17

    Increasing utilization of engineered nanoparticles in the field of electronics and biomedical applications demands an assessment of risk associated with deliberate or accidental exposure. Metal based nanoparticles are potentially most important of all the nanoparticles in terms of health risks. Microporous alumino-silicates and pure silicates named as zeolites and zeo-type materials with variety of structures, chemical compositions, particle sizes and morphologies have a significant number of industrial uses such as in catalysis, sorption and ion-exchange processes. In particular, the nanosized particles due to their unique properties are used in hybrid organic-inorganic materials for photography, photonics, electronics, labeling, imaging, and sensing. The aim of the current study is to investigate pure silica MFI-type zeolites nanoparticles with sizes of 50nm and 100nm (samples MFI-50 and MFI-100) under suspended conditions and their toxicological effects on human lung alveolar (A549) cells under in vitro conditions. Live cell imaging showed that the nanoparticles precipitated from the colloidal suspension of cell culture media as large agglomerates, coming in contact with the cell surface through sedimentation. A cellular proliferative capacity test showed the zeolite nanoparticles to exhibit no significant cytotoxicity below a concentration of 100μg/ml. However, both the MFI-50 and MFI-100 nanoparticles induced high intracellular reactive oxygen species (ROS) generation and elevated mitochondrial membrane potential in the A549 cells over the measured time period of 12h and at concentrations up to ≤50μg/ml. DNA fragmentation analysis using the comet assay showed that the MFI-50 and MFI-100 nanoparticles cause genotoxicity in a concentration dependent manner. Furthermore, the rate at which maximum genomic damage was caused by MFI-100 nanoparticles in the A549 cells was found to be high as compared to the MFI-50 nanoparticles. However, the damage caused by the

  20. Increased alveolar soluble Annexin V promotes lung inflammation and fibrosis

    OpenAIRE

    Buckley, S.; Shi, W.; Xu, W.; Frey, M.R.; Moats, R.; Pardo, A.; Selman, M.; Warburton, D.

    2015-01-01

    The causes underlying the self-perpetuating nature of idiopathic pulmonary fibrosis (IPF), a progressive and usually lethal disease, remain unknown. We hypothesized that alveolar soluble Annexin V contributes to lung fibrosis, based on the observation that human IPF BALF containing high Annexin V levels promoted fibroblast involvement in alveolar epithelial wound healing that was reduced when Annexin V was depleted from the BALF.

  1. 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

  2. The Effects of First-Line Anti-Tuberculosis Drugs on the Actions of Vitamin D in Human Macrophages.

    Science.gov (United States)

    Chesdachai, Supavit; Zughaier, Susu M; Hao, Li; Kempker, Russell R; Blumberg, Henry M; Ziegler, Thomas R; Tangpricha, Vin

    2016-12-01

    Tuberculosis (TB) is a major global health problem. Patients with TB have a high rate of vitamin D deficiency, both at diagnosis and during the course of treatment with anti-tuberculosis drugs. Although data on the efficacy of vitamin D supplementation on Mycobacterium tuberculosis (Mtb) clearance is uncertain from randomized controlled trials (RCTs), vitamin D enhances the expression of the anti-microbial peptide