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Sample records for alveolar macrophages infected

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

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    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

  2. 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.

  3. Innate immune response of human alveolar macrophages during influenza A infection.

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    Jieru Wang

    Full Text Available Alveolar macrophages (AM are one of the key cell types for initiating inflammatory and immune responses to influenza virus in the lung. However, the genome-wide changes in response to influenza infection in AM have not been defined. We performed gene profiling of human AM in response to H1N1 influenza A virus PR/8 using Affymetrix HG-U133 Plus 2.0 chips and verified the changes at both mRNA and protein levels by real-time RT-PCR and ELISA. We confirmed the response with a contemporary H3N2 influenza virus A/New York/238/2005 (NY/238. To understand the local cellular response, we also evaluated the impact of paracrine factors on virus-induced chemokine and cytokine secretion. In addition, we investigated the changes in the expression of macrophage receptors and uptake of pathogens after PR/8 infection. Although macrophages fail to release a large amount of infectious virus, we observed a robust induction of type I and type III interferons and several cytokines and chemokines following influenza infection. CXCL9, 10, and 11 were the most highly induced chemokines by influenza infection. UV-inactivation abolished virus-induced cytokine and chemokine response, with the exception of CXCL10. The contemporary influenza virus NY/238 infection of AM induced a similar response as PR/8. Inhibition of TNF and/or IL-1β activity significantly decreased the secretion of the proinflammatory chemokines CCL5 and CXCL8 by over 50%. PR/8 infection also significantly decreased mRNA levels of macrophage receptors including C-type lectin domain family 7 member A (CLEC7A, macrophage scavenger receptor 1 (MSR1, and CD36, and reduced uptake of zymosan. In conclusion, influenza infection induced an extensive proinflammatory response in human AM. Targeting local components of innate immune response might provide a strategy for controlling influenza A infection-induced proinflammatory response in vivo.

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

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    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

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

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    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.

  6. Alveolar macrophages infected with Ames or Sterne strain of Bacillus anthracis elicit differential molecular expression patterns.

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    Felicia D Langel

    Full Text Available Alveolar macrophages (AMs phagocytose Bacillus anthracis following inhalation and induce the production of pro-inflammatory cytokines and chemokines to mediate the activation of innate immunity. Ames, the virulent strain of B. anthracis, contains two plasmids that encode the antiphagocytic poly-γ-d-glutamic acid capsule and the lethal toxin. The attenuated Sterne strain of B. anthracis, which lacks the plasmid encoding capsule, is widely adapted as a vaccine strain. Although differences in the outcome of infection with the two strains may have originated from the presence or absence of an anti-phagocytic capsule, the disease pathogenesis following infection will be manifested via the host responses, which is not well understood. To gain understanding of the host responses at cellular level, a microarray analysis was performed using primary rhesus macaque AMs infected with either Ames or Sterne spores. Notably, 528 human orthologs were identified to be differentially expressed in AMs infected with either strain of the B. anthracis. Meta-analyses revealed genes differentially expressed in response to B. anthracis infection were also induced upon infections with multiple pathogens such as Francisella Novicida or Staphylococcus aureus. This suggests the existence of a common molecular signature in response to pathogen infections. Importantly, the microarray and protein expression data for certain cytokines, chemokines and host factors provide further insights on how cellular processes such as innate immune sensing pathways, anti-apoptosis versus apoptosis may be differentially modulated in response to the virulent or vaccine strain of B. anthracis. The reported differences may account for the marked difference in pathogenicity between these two strains.

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

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    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.

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

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    Ziltener, Pascal; Reinheckel, Thomas; Oxenius, Annette

    2016-01-01

    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. PMID:27105352

  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. Microarray analysis of the effect of Streptococcus equi subsp. zooepidemicus M-like protein in infecting porcine pulmonary alveolar macrophage.

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    Ma, Zhe; Zhang, Hui; Yi, Li; Fan, Hongjie; Lu, Chengping

    2012-01-01

    Streptococcus equi subsp. zooepidemicus (S. zooepidemicus), which belongs to Lancefield group C streptococci, is an important pathogen of domesticated species, causing septicemia, meningitis and mammitis. M-like protein (SzP) is an important virulence factor of S. zooepidemicus and contributes to bacterial infection and antiphagocytosis. To increase our knowledge of the mechanism of SzP in infection, we profiled the response of porcine pulmonary alveolar macrophage (PAM) to infection with S. zooepidemicus ATCC35246 wild strain (WD) and SzP-knockout strain (KO) using the Roche NimbleGen Porcine Genome Expression Array. We found SzP contributed to differential expression of 446 genes, with upregulation of 134 genes and downregulation of 312 genes. Gene Ontology category and KEGG pathway were analyzed for relationships among differentially expressed genes. These genes were represented in a variety of functional categories, including genes involved in immune response, regulation of chemokine production, signal transduction and regulation of apoptosis. The reliability of the data obtained from the microarray was verified by performing quantitative real-time PCR on 12 representative genes. The data will contribute to understanding of SzP mediated mechanisms of S. zooepidemicus pathogenesis.

  11. Microarray analysis of the effect of Streptococcus equi subsp. zooepidemicus M-like protein in infecting porcine pulmonary alveolar macrophage.

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    Zhe Ma

    Full Text Available Streptococcus equi subsp. zooepidemicus (S. zooepidemicus, which belongs to Lancefield group C streptococci, is an important pathogen of domesticated species, causing septicemia, meningitis and mammitis. M-like protein (SzP is an important virulence factor of S. zooepidemicus and contributes to bacterial infection and antiphagocytosis. To increase our knowledge of the mechanism of SzP in infection, we profiled the response of porcine pulmonary alveolar macrophage (PAM to infection with S. zooepidemicus ATCC35246 wild strain (WD and SzP-knockout strain (KO using the Roche NimbleGen Porcine Genome Expression Array. We found SzP contributed to differential expression of 446 genes, with upregulation of 134 genes and downregulation of 312 genes. Gene Ontology category and KEGG pathway were analyzed for relationships among differentially expressed genes. These genes were represented in a variety of functional categories, including genes involved in immune response, regulation of chemokine production, signal transduction and regulation of apoptosis. The reliability of the data obtained from the microarray was verified by performing quantitative real-time PCR on 12 representative genes. The data will contribute to understanding of SzP mediated mechanisms of S. zooepidemicus pathogenesis.

  12. Modulation of alveolar macrophage innate response in proinflammatory-, pro-oxidant-, and infection- models by mint extract and chemical constituents: Role of MAPKs.

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    Yadav, Niket; Chandra, Harish

    2018-01-01

    There is a continuing need for discovering novel primary or adjunct therapeutic agents to treat inflammatory conditions and infections. Natural products have inspired the discovery of several modern therapeutics; however, there is a paucity of mechanistic information on their mode of action. This study investigated the therapeutic potential and mode of action of corn mint's (Mentha arvensis) leaf extract (ME) in alveolar macrophages (AMs) challenged with model pro-inflammatory (LPS), pro- oxidant (LPS or H 2 O 2 ), and infection (Mycobacterium) agents and contribution of its dominant constituents rosmarinic acid, l-menthol, and l-menthone. LPS-induced inflammatory response in the murine AM cell line MH-S was significantly reduced in terms of pro-inflammatory cytokines (TNF-α, IL-1α) and nitric oxide (NO) when pre- or post-treated with ME. The ME pretreatment of macrophages led to a significant increase (P≤0.05) in phagocytic activity toward Mycobacterium smegmatis and a greater pathogen clearance in 24h in both ME pre-treated (P≤0.05) and post-treated cells. Significant attenuation (P≤0.01) of reactive oxygen species (ROS) production in LPS- or H 2 O 2- treated macrophages by pretreatment with whole mint extract (ME) was accounted for in part by the mint constituents rosmarinic acid and l-menthone. Attenuation of pro-inflammatory response by ME pretreatment coincided with the significant reduction in total and phosphorylated JNK1/2, decrease in total p38, and increase in phospho-ERK1/2 thereby implying a role of differential modulation of MAPKs. Taken together, the results demonstrate that corn mint leaf components cause potent anti-inflammatory, anti-oxidant, and anti-infection effects in AMs via suppression of the production of cytokines/soluble mediators and ROS and increased pathogen clearance, respectively. To our knowledge, this is the first report on the mode of action of corn mint targeting the alveolar macrophages and on the potential role of MAPKs

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

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    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

  14. Aerosol-based efficient delivery of clarithromycin, a macrolide antimicrobial agent, to lung epithelial lining fluid and alveolar macrophages for treatment of respiratory infections.

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    Togami, Kohei; Chono, Sumio; Morimoto, Kazuhiro

    2012-04-01

    Macrolide antimicrobial agents are generally given by the oral route for the treatment of respiratory infections caused by pathogenic microorganisms infected in lung epithelial lining fluid (ELF) and alveolar macrophages (AMs). However, because macrolides distribute to many different tissues via the blood after oral administration, systemic side effects are frequently induced. In contrast with oral administration, aerosolization may be an efficient method for delivering macrolides directly to ELF and AMs. In this study, the efficacy of aerosol-based delivery of clarithromycin (CAM), as a model macrolide, for the treatment of respiratory infections was evaluated by comparison with oral administration. The aerosol formulation of CAM (0.2 mg/kg) was administered to rat lungs using a Liquid MicroSprayer(®). The oral formulation of CAM (50 mg/kg) was used for comparison. Time courses of concentrations of CAM in ELF and AMs following administration were obtained, and then the bioavailability (BA) was calculated. In addition, the area under the concentrations of CAM in ELF and AMs-time curve/minimum inhibitory concentration at which 90% of isolates ratio [area under the curve (AUC/MIC(90))] were calculated to estimate the antibacterial effects in ELF and AMs. The BA of CAM in ELF and AMs following administration of aerosol formulation were markedly greater than that following administration of oral formulation. This indicates that the aerosol formulation is more effective in delivering CAM to ELF and AMs, compared with the oral formulation, despite a low dose. The AUC/MIC(90) of CAM in ELF and AMs were markedly higher than the effective values. This indicates that the aerosol formulation could be useful for the treatment of respiratory infections caused by pathogenic microorganisms infected in ELF and AMs. This study suggests that aerosol formulation of macrolides is an effective pulmonary drug delivery system for the treatment of respiratory infections.

  15. Aerosol-based efficient delivery of telithromycin, a ketolide antimicrobial agent, to lung epithelial lining fluid and alveolar macrophages for treatment of respiratory infections.

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    Togami, Kohei; Chono, Sumio; Seki, Toshinobu; Morimoto, Kazuhiro

    2010-07-01

    The efficacy of aerosol-based delivery of telithromycin (TEL), as a model antimicrobial agent, for the treatment of respiratory infections was evaluated by comparison with oral administration. The aerosol formulation (0.2 mg/kg) was administered to rat lungs using a Liquid MicroSprayer. The time courses of the concentration of TEL in lung epithelial lining fluid (ELF) and alveolar macrophages (AMs) following administration of an aerosol formulation to rat lungs were markedly higher than that following the administration of an oral formulation (50 mg/kg). The time course of the concentrations of TEL in plasma following administration of the aerosol formulation was markedly lower than that in ELF and AMs. These results indicate that the aerosol formulation is more effective in delivering TEL to ELF and AMs, compared to the oral formulation, despite a low dose and it avoids distribution of TEL to the blood. In addition, the antibacterial effects of TEL in ELF and AMs following administration of the aerosol formulation were estimated by pharmacokinetics/pharmacodynamics analysis. The concentrations of TEL in ELF and the AMs time curve/minimum inhibitory concentration of TEL ratio were markedly higher than the effective values. This study indicates that an antibiotic aerosol formulation may be an effective pulmonary drug delivery system for the treatment of respiratory infections.

  16. Rapid host defense against Aspergillus fumigatus involves alveolar macrophages with a predominance of alternatively activated phenotype.

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    Shikha Bhatia

    Full Text Available The ubiquitous fungus Aspergillus fumigatus is associated with chronic diseases such as invasive pulmonary aspergillosis in immunosuppressed patients and allergic bronchopulmonary aspergillosis (ABPA in patients with cystic fibrosis or severe asthma. Because of constant exposure to this fungus, it is critical for the host to exercise an immediate and decisive immune response to clear fungal spores to ward off disease. In this study, we observed that rapidly after infection by A. fumigatus, alveolar macrophages predominantly express Arginase 1 (Arg1, a key marker of alternatively activated macrophages (AAMs. The macrophages were also found to express Ym1 and CD206 that are also expressed by AAMs but not NOS2, which is expressed by classically activated macrophages. The expression of Arg1 was reduced in the absence of the known signaling axis, IL-4Rα/STAT6, for AAM development. While both Dectin-1 and TLR expressed on the cell surface have been shown to sense A. fumigatus, fungus-induced Arg1 expression in CD11c(+ alveolar macrophages was not dependent on either Dectin-1 or the adaptor MyD88 that mediates intracellular signaling by most TLRs. Alveolar macrophages from WT mice efficiently phagocytosed fungal conidia, but those from mice deficient in Dectin-1 showed impaired fungal uptake. Depletion of macrophages with clodronate-filled liposomes increased fungal burden in infected mice. Collectively, our studies suggest that alveolar macrophages, which predominantly acquire an AAM phenotype following A. fumigatus infection, have a protective role in defense against this fungus.

  17. Radionuclide study of the action of cadmium on alveolar macrophages

    International Nuclear Information System (INIS)

    Reulet, Maryse.

    1976-01-01

    The experimental toxicity of cadmium was studied on the lung, using cadmium sulfate, cadmium acetate and the radioactive isotope cadmium 109 in chloride or acetate form. The results are given in the following order: part one is devoted to the results of investigations on chronic cadmium poisoning and the role of alveolar macrophages in this poisoning; in part two the uptake of cadmium on alveolar macrophages is studied with cadmium 109, administered intraperitoneally; in part three the action of cadmium on the phospholipid metabolism of alveolar macrophages is examined. The cadmium, as sulfate or acetate, is administered in several ways: by intraperitoneal injection; or by inhalation of cadmium dusts or aerosols. The effect of cadmium on the oxidative metabolism of alveolar macrophages is studied in part four. This work is carried out 'in vitro' and 'in vivo' after cadmium oxide dusting of the air [fr

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

    International Nuclear Information System (INIS)

    Chuan Junlan; Li Yanzhen; Yang Likai; Sun Xun; Zhang Qiang; Gong Tao; Zhang Zhirong

    2013-01-01

    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.

  19. In utero infection with PRRS virus modulates cellular functions of blood monocytes and alveolar lung macrophages in piglets

    DEFF Research Database (Denmark)

    Riber, Ulla; Nielsen, Jens; Lind, Peter

    2004-01-01

    The putative immunosuppressive effect of PRRS virus (PRRSV) on innate immune responses was studied in piglets infected in utero with PRRSV. Phagocytosis and oxidative burst capacities in 2-, 4- and 6-week-old in utero infected piglets were investigated and compared with age-matched control piglets...

  20. 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...

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

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    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

  2. Differential gene expression profiling of Actinobacillus pleuropneumoniae during induction of primary alveolar macrophage apoptosis in piglets

    NARCIS (Netherlands)

    Wang, Lei; Qin, Wanhai; Ruidong, Zhai; Liu, Shiting; Zhang, Hu; Sun, Changjiang; Feng, Xin; Gu, Jingmin; Du, Chongtao; Han, Wenyu; Langford, P. R.; Lei, Liancheng

    2015-01-01

    Actinobacillus pleura pneumoniae (A. pleuropneumoniae) is the causative agent of porcine pleuropneumonia, a disease that causes serious problems for the swine industry. Successful infection by this bacterium requires breaking the first line of defence in the lungs, the primary alveolar macrophages

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

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    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...... analysis of cell size and membrane integrity) led to 40% reduction in the total number of phagocytozing cells. However, viable/uninfected macrophages in PRRSV-infected cultures exhibited normal phagocytic ability at 48 h, indicating that no soluble phagocytosis-suppressive mediators were induced by PRRSV...... 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....

  4. Entry and elimination of marine mammal Brucella spp. by hooded seal (Cystophora cristata alveolar macrophages in vitro.

    Directory of Open Access Journals (Sweden)

    Anett K Larsen

    Full Text Available A high prevalence of Brucellapinnipedialis serology and bacteriology positive animals has been found in the Northeast Atlantic stock of hooded seal (Cystophoracristata; however no associated gross pathological changes have been identified. Marine mammal brucellae have previously displayed different infection patterns in human and murine macrophages. To investigate if marine mammal Brucella spp. are able to invade and multiply in cells originating from a presumed host species, we infected alveolar macrophages from hooded seal with a B. pinnipedialis hooded seal isolate. Hooded seal alveolar macrophages were also challenged with B. pinnipedialis reference strain (NCTC 12890 from harbor seal (Phocavitulina, B. ceti reference strain (NCTC 12891 from harbor porpoise (Phocoenaphocoena and a B. ceti Atlantic white-sided dolphin (Lagenorhynchusacutus isolate (M83/07/1, to evaluate possible species-specific differences. Brucella suis 1330 was included as a positive control. Alveolar macrophages were obtained by post mortem bronchoalveolar lavage of euthanized hooded seals. Phenotyping of cells in the lavage fluid was executed by flow cytometry using the surface markers CD14 and CD18. Cultured lavage cells were identified as alveolar macrophages based on morphology, expression of surface markers and phagocytic ability. Alveolar macrophages were challenged with Brucella spp. in a gentamicin protection assay. Following infection, cell lysates from different time points were plated and evaluated quantitatively for colony forming units. Intracellular presence of B. pinnipedialis hooded seal isolate was verified by immunocytochemistry. Our results show that the marine mammal brucellae were able to enter hooded seal alveolar macrophages; however, they did not multiply intracellularly and were eliminated within 48 hours, to the contrary of B. suis that showed the classical pattern of a pathogenic strain. In conclusion, none of the four marine mammal strains

  5. 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

  6. Virus-induced enhancement of arachidonate metabolism by bovine alveolar macrophages in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Laegreid, W.W.; Taylor, S.M.; Leid, R.W.; Silflow, R.M.; Evermann, J.R.; Breeze, R.G.; Liggitt, H.D.

    1989-04-01

    Virus infection of alveolar macrophages both in vivo and in vitro has been associated with a variety of changes in cellular function. Some of these changes are identical to the effects that arachidonate-derived mediators, prostaglandins, leukotrienes, and hydroxyeicosatetraenoic acids, have on macrophage function. Virus infection of macrophages has been previously shown to increase the output of some arachidonate metabolites, most notably PGE2. However, the effect of virus infection on arachidonate metabolism in general has not been well described. In our experiments, primary cultures of alveolar macrophages obtained from normal cattle by bronchoalveolar lavage, were infected in vitro with parainfluenza type 3 virus. At days 0 to 4 post-infection (p.i.) these cells were labelled with 3H-arachidonic acid and stimulated with either serum-coated zymosan, the calcium ionophore A23187, or phorbol myristate acetate. The complete spectrum of arachidonate-derived metabolites was determined by reverse-phase high performance liquid chromatography with UV and on-line radiometric monitoring of column eluant. The total output of metabolites of arachidonic acid by virus-infected alveolar macrophages was increased over that of noninfected controls (with all stimuli tested) by day 4 p.i. (P less than or equal to 0.05). The production of metabolites by the cyclooxygenase, 12- and 5-lipoxygenase enzyme systems was significantly increased, as was the release of 3H-arachidonate. The lack of stimulus specificity and the increases in arachidonate release suggest that greater substrate availability, due either to increased phospholipase activity or direct virus-membrane interaction, may be responsible for the virus-induced enhancement of metabolite output.

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

    Directory of Open Access Journals (Sweden)

    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.

  8. 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.

  9. 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.

  10. Alveolar macrophage dysregulation in Hermansky-Pudlak syndrome type 1.

    Science.gov (United States)

    Rouhani, Farshid N; Brantly, Mark L; Markello, Thomas C; Helip-Wooley, Amanda; O'Brien, Kevin; Hess, Richard; Huizing, Marjan; Gahl, William A; Gochuico, Bernadette R

    2009-12-01

    Individuals with Hermansky-Pudlak syndrome type 1 (HPS-1), an autosomal recessive disorder characterized by defective biogenesis of lysosome-related organelles, develop an accelerated form of progressive fibrotic lung disease. The etiology of pulmonary fibrosis associated with HPS-1 is unknown. To investigate the potential pathogenesis of pulmonary fibrosis in HPS-1, lung cells and proteins from individuals with HPS-1 were studied. Forty-one subjects with HPS-1 with and without pulmonary fibrosis were evaluated with pulmonary function tests, high-resolution computed tomography scan, and bronchoscopy. Bronchoalveolar lavage cells and analytes were analyzed. Concentrations of total bronchoalveolar lavage cells and alveolar macrophages were significantly higher in epithelial lining fluid from subjects with HPS-1 with and without pulmonary fibrosis compared with healthy research volunteers. Concentrations of cytokines and chemokines (i.e., monocyte chemoattractant protein-1, macrophage inflammatory protein-1alpha, and granulocyte-macrophage colony-stimulating factor) in alveolar epithelial lining fluid were significantly higher in subjects with HPS-1 with and without pulmonary fibrosis compared with healthy research volunteers (P system in which to study the pathogenesis and treatment of HPS pulmonary fibrosis.

  11. Surfactant Protein D Binds to Coxiella burnetii and Results in a Decrease in Interactions with Murine Alveolar Macrophages.

    Directory of Open Access Journals (Sweden)

    Kelly A Soltysiak

    Full Text Available Coxiella burnetii is a Gram-negative, obligate intracellular bacterium and the causative agent of Q fever. Infections are usually acquired after inhalation of contaminated particles, where C. burnetii infects its cellular target cells, alveolar macrophages. Respiratory pathogens encounter the C-type lectin surfactant protein D (SP-D during the course of natural infection. SP-D is a component of the innate immune response in the lungs and other mucosal surfaces. Many Gram-negative pulmonary pathogens interact with SP-D, which can cause aggregation, bactericidal effects and aid in bacterial clearance. Here we show that SP-D binds to C. burnetii in a calcium-dependent manner with no detectable bacterial aggregation or bactericidal effects. Since SP-D interactions with bacteria often alter macrophage interactions, it was determined that SP-D treatment resulted in a significant decrease in C. burnetii interactions to a mouse alveolar macrophage model cell line MH-S indicating SP-D causes a significant decrease in phagocytosis. The ability of SP-D to modulate macrophage activation by C. burnetii was tested and it was determined that SP-D does not alter the correlates measured for macrophage activation. Taken together these studies support those demonstrating limited activation of alveolar macrophages with C. burnetii and demonstrate interactions with SP-D participate in reduction of phagocyte attachment and phagocytosis.

  12. SIV Infection of Lung Macrophages.

    Directory of Open Access Journals (Sweden)

    Yue Li

    Full Text Available HIV-1 depletes CD4+ T cells in the blood, lymphatic tissues, gut and lungs. Here we investigated the relationship between depletion and infection of CD4+ T cells in the lung parenchyma. The lungs of 38 Indian rhesus macaques in early to later stages of SIVmac251 infection were examined, and the numbers of CD4+ T cells and macrophages plus the frequency of SIV RNA+ cells were quantified. We showed that SIV infected macrophages in the lung parenchyma, but only in small numbers except in the setting of interstitial inflammation where large numbers of SIV RNA+ macrophages were detected. However, even in this setting, the number of macrophages was not decreased. By contrast, there were few infected CD4+ T cells in lung parenchyma, but CD4+ T cells were nonetheless depleted by unknown mechanisms. The CD4+ T cells in lung parenchyma were depleted even though they were not productively infected, whereas SIV can infect large numbers of macrophages in the setting of interstitial inflammation without depleting them. These observations point to the need for future investigations into mechanisms of CD4+ T cell depletion at this mucosal site, and into mechanisms by which macrophage populations are maintained despite high levels of infection. The large numbers of SIV RNA+ macrophages in lungs in the setting of interstitial inflammation indicates that lung macrophages can be an important source for SIV persistent infection.

  13. Interaction of rat alveolar macrophages with dental composite dust.

    Science.gov (United States)

    Van Landuyt, K L; Cokic, S M; Asbach, C; Hoet, P; Godderis, L; Reichl, F X; Van Meerbeek, B; Vennemann, A; Wiemann, M

    2016-11-26

    Dental composites have become the standard filling material to restore teeth, but during the placement of these restorations, high amounts of respirable composite dust (nano-sized particles may be released in the breathing zone of the patient and dental operator. Here we tested the respirable fraction of several composite particles for their cytotoxic effect using an alveolar macrophage model system. ​METHODS: Composite dust was generated following a clinical protocol, and the dust particles were collected under sterile circumstances. Dust was dispersed in fluid, and 5-μm-filtered to enrich the respirable fractions. Quartz DQ12 and corundum were used as positive and negative control, respectively. Four concentrations (22.5 μg/ml, 45 μg/ml, 90 μg/ml and 180 μg/ml) were applied to NR8383 alveolar macrophages. Light and electron microscopy were used for subcellular localization of particles. Culture supernatants were tested for release of lactate dehydrogenase, glucuronidase, TNF-α, and H 2 O 2 . Characterization of the suspended particles revealed numerous nano-sized particles but also many high volume particles, most of which could be removed by filtering. Even at the highest concentration (180 μg/ml), cells completely cleared settled particles from the bottom of the culture vessel. Accordingly, a mixture of nano- and micron-scaled particles was observed inside cells where they were confined to phagolysosomes. The filtered particle fractions elicited largely uniform dose-dependent responses, which were elevated compared to the control only at the highest concentration, which equaled a mean cellular dose of 120 pg/cell. A low inflammatory potential was identified due to dose-dependent release of H 2 O 2 and TNF-α. However, compared to the positive control, the released levels of H 2 O 2 and TNF-α were still moderate, but their release profiles depended on the type of composite. Alveolar macrophages are able to phagocytize respirable composite dust

  14. 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

  15. Gene expression profiling of human alveolar macrophages infected by B. anthracis spores demonstrates TNF-α and NF-κb are key components of the innate immune response to the pathogen

    Directory of Open Access Journals (Sweden)

    Hurst Robert E

    2009-09-01

    Full Text Available Abstract Background Bacillus anthracis, the etiologic agent of anthrax, has recently been used as an agent of bioterrorism. The innate immune system initially appears to contain the pathogen at the site of entry. Because the human alveolar macrophage (HAM plays a key role in lung innate immune responses, studying the HAM response to B. anthracis is important in understanding the pathogenesis of the pulmonary form of this disease. Methods In this paper, the transcriptional profile of B. anthracis spore-treated HAM was compared with that of mock-infected cells, and differentially expressed genes were identified by Affymetrix microarray analysis. A portion of the results were verified by Luminex protein analysis. Results The majority of genes modulated by spores were upregulated, and a lesser number were downregulated. The differentially expressed genes were subjected to Ingenuity Pathway analysis, the Database for Annotation, Visualization and Integrated Discovery (DAVID analysis, the Promoter Analysis and Interaction Network Toolset (PAINT and Oncomine analysis. Among the upregulated genes, we identified a group of chemokine ligand, apoptosis, and, interestingly, keratin filament genes. Central hubs regulating the activated genes were TNF-α, NF-κB and their ligands/receptors. In addition to TNF-α, a broad range of cytokines was induced, and this was confirmed at the level of translation by Luminex multiplex protein analysis. PAINT analysis revealed that many of the genes affected by spores contain the binding site for c-Rel, a member of the NF-κB family of transcription factors. Other transcription regulatory elements contained in many of the upregulated genes were c-Myb, CP2, Barbie Box, E2F and CRE-BP1. However, many of the genes are poorly annotated, indicating that they represent novel functions. Four of the genes most highly regulated by spores have only previously been associated with head and neck and lung carcinomas. Conclusion The

  16. Mannose-functionalized solid lipid nanoparticles are effective in targeting alveolar macrophages.

    Science.gov (United States)

    Costa, Ana; Sarmento, Bruno; Seabra, Vítor

    2018-03-01

    Mannose receptor is highly expressed on alveolar macrophages, being a potential target to promote the specific local drug delivery of anti-tuberculosis agents through the use of functionalized nanocarriers. In this work, isoniazid (Isn)-loaded solid lipid nanoparticles (SLN), reinforced with stearylamine (SA) were produced by double emulsion technique and further surface-functionalized with mannose in a straightforward chemical approach. Upon pre-formulation assessment, SLN close to 500 nm average size, positively charged and with association efficiency of ISN close to 50% were obtained. Functionalization with mannose was performed after SLN production and confirmed by Fourier transform infrared spectroscopy (FTIR). Both functionalized and non-functionalized SLN demonstrated to devoid of toxicity when tested in human lung epithelial cell line (NCI-H441) and differentiated THP-1 (dTHP-1), reducing the intrinsic cytotoxicity of Isn when incorporated into SLN. Uptake studies were conducted on same macrophage-like cells and the results showed that fluorescent mannosylated SLN (M-SLN) were more efficient in be internalized comparatively to SLN. Moreover, the uptake of M-SLN was reduced when cells were pre-incubated with mannose, demonstrating the receptor-dependence internalization of functionalized SLN. These functionalized nanocarriers may represent a useful platform to target alveolar macrophages for delivering anti-infective drugs. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Characterization of CD44 expressed on alveolar macrophages in patients with diffuse panbronchiolitis

    Science.gov (United States)

    Katoh, S; Matsubara, Y; Taniguchi, H; Fukushima, K; Mukae, H; Kadota, J; Matsukura, S; Kohno, S

    2001-01-01

    Interleukin (IL)-8 may play an important role in neutrophil infiltration in the airways of patients with diffuse panbronchiolitis (DPB). Furthermore, alveolar macrophages could produce IL-8 subsequent to CD44-hyaluronic acid (HA) interaction. The purpose of this study was to evaluate the contribution of CD44 expressed on alveolar macrophages to the pathogenesis of DPB. We examined the concentration of soluble CD44 (sCD44) in bronchoalveolar lavage fluid (BALF) and CD44 expression on macrophages in BALF from patients with DPB before and after low-dose, long-term macrolide therapy. We also assessed the HA-binding ability of alveolar macrophages as a functional analysis of the CD44 molecule. The sCD44 concentration in BALF was significantly lower in patients with DPB than in healthy volunteers. Percentages of alveolar macrophages expressing low CD44 (CD44 low+) and HA-nonbinding alveolar macrophages were higher in patients with DPB compared with healthy volunteers. Furthermore, macrolide therapy normalized CD44 expression and HA-binding ability of macrophages in BALF from DPB patients. Our findings suggest that alveolar macrophage dysfunction could result from abnormalities of CD44 expression in patients with DPB and that these events could contribute to the pathogenesis of DPB. PMID:11737075

  18. 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...

  19. Ultrastructural types of alveolar macrophages in bronchoalveolar lavages from patients with pulmonary sarcoidosis.

    Science.gov (United States)

    Burkhardt, Olaf; Lode, Hartmut; Welte, Tobias; Merker, Hans-Joachim

    2007-01-01

    By routine applied quantitative BAL methods are particularly helpful for the diagnosis of pulmonary sarcoidosis. Here the morphology of the alveolar cells does not play a role. However, morphological and especially electron microscopic investigations might contribute to the clarification of the aetiology of this disease. In a prospective study we investigated the bronchoalveolar lavages (BALs) from 10 patients with recently histologically diagnosed, untreated pulmonary sarcoidosis. Commonly applied cytological and immunological BAL diagnostic techniques were accompanied by morphological investigations of alveolar cells, especially alveolar macrophages, using light and electron microscopy. All patients showed lymphocytic alveolitis with an increased number of CD4 positive lymphocytes as well as an increased CD4/CD8 ratio. A striking light microscopic finding was the great morphological variety of the alveolar macrophages. Electron microscopy revealed typical lymphocytes, neutrophils, and eosinophils as well as three different types of alveolar macrophages in all 10 patients: type I (approx. 30%) with a normal macrophage morphology, a vacuole-rich type II (approx. 30%) with myelin-like structures and type III (approx. 40%) with electron-dense inclusions. The occurrence of intracellular myelin figures in type II macrophages is a hint for increased phagocytotic processes of surfactant with or without its overproduction in the sense of a secondary alveolar proteinosis. Numerous electron-dense inclusions in type III also indicate an increased macrophage activity that leads to an increased release of cytokines, which in turn can trigger an inflammatory reaction.

  20. 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

  1. 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.)

  2. Antioxidant properties of taurine in rat alveolar macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Castranova, V.; Banks, M.A.; Porter, D.W.; Martin, W.G. (National Inst. for Occupational Safety and Health, Morgantown, WV (United States) West Virginia Univ., Morgantown (United States))

    1990-02-26

    Isolated rat alveolar macrophages (RAM) which had taken-up and accumulated extracellular (0-500 {mu}M) taurine (TAU) were exposed to 0.45 {plus minus} 0.05 ppm ozone for 30 minutes in a modified tissue culture flask containing TAU-supplemented medium. Recovered cells were assayed for oxidant damage and media analyzed for leakage of intracellular components. Cell viability significantly increased, while recovery of cells decreased (possibly due to increased adherence) with increasing TAU. At 100 {mu}M (rat plasma TAU level), TAU protected against the ozone-induced increase in zymosan-stimulated chemiluminescence, diminished leakages of lipid peroxidation products and protein into the medium, and partially restored the ozone-inactivated Na{sup +}/K{sup +} ATPase activity of RAM. Efflux of oxidized glutathione was maximized and K{sup +} leakage was minimized by the addition of 250 {mu}M TAU. At 250-500 {mu}M TAU, leakages of lipid peroxidation products and protein were enhanced, while the intracellular TAU content dramatically increased. These results indicate that TAU has both direct and indirect antioxidant properties at low levels and pro-oxidant properties at high levels in RAM.

  3. 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.

  4. 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.

  5. 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.

  6. 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

  7. Evaluation of alveolar macrophage function after experimental infection with equine herpesvirus-1 in horses Avaliação da função dos macrófagos alveolares após infecção experimental em cavalos por herpesvírus eqüino tipo 1

    Directory of Open Access Journals (Sweden)

    E. Mori

    2003-06-01

    Full Text Available The role of the pulmonary alveolar macrophages (PAM in the lung defense mechanism was evaluated in horses infected with equine hespesvirus-1 (EHV-1. Five adult horses were exposed to 10(6.6 TCID50 EHV-1 by intranasal instillation. Cytology of bronchoalveolar lavage (BAL was performed using cytocentrifugation of samples and slides stained by Rosenfeld. Cell concentration was adjusted to 2´10(6 cells/ml, for the measurement of macrophage activity - spreading, phagocytosis of zymosan particles and release of hydrogen peroxide (H2O2. All animals were positive in virus isolation on the second, third and fifth days post-inoculation (DPI. Seroconversion was observed on the 14th DPI. Lymphocytosis was observed by BAL cytology on the 16th DPI. Measurement of macrophage activity demonstrated a marked increase in the spreading rate, on the 23rd and 30th DPI. Phagocytosis was decreased on the second DPI, and returned to levels similar to those observed before inoculation on the 23rd DPI. The amount of H2O2 released by PAM declined on day 2, but, by day 16, they returned to values similar to those observed before inoculation. The decline in PAM activity in the acute phase of disease is indirect evidence that these cells have an important role in lung defense mechanisms against this agent.O papel dos macrófagos alveolares (MA nos mecanismos de defesa pulmonar foi estudado em cavalos infectados pelo herpesvírus eqüino tipo 1 (EHV-1. Cinco cavalos adultos foram inoculados com 10(6,6 TCID50 do EHV-1, por instilação intranasal. A citologia do lavado broncoalveolar (LBA foi feita usando-se citocentrifugação das amostras e confecção de lâminas coradas por Rosenfeld. A concentração celular foi ajustada para 2´10(6 células/ml, para mensuração da atividade macrofágica - espraiamento, fagocitose de partículas de zymosan e liberação de peróxido de hidrogênio (H2O2. Observou-se soroconversão no 14º dia pós-inoculação (DPI e isolamento viral

  8. Evaluation of Inflammatory Cytokine Secretion by Human Alveolar Macrophages

    Directory of Open Access Journals (Sweden)

    J. E. Losa García

    1999-01-01

    Full Text Available The alveolar macrophage (AM secretes interleukin 1β (IL-1β, tumour necrosis factor-α (TNF-α, interleukin-6 (IL-6 and interleukin-8 (IL-8, all of them inflammatory cytokines involved in the pathogenesis of many lung diseases. The aim of the present work was to evaluate the basal and stimulated secretion of these cytokines by human AMs. Human AMs were collected by bronchoalveolar lavage (BAL from four healthy controls and 13 patients with diffuse interstitial lung disease (five cases of sarcoidosis, three of hypersensitivity pneumonitis and five of idiopathic pulmonary fibrosis. AMs were cultured in the presence or absence of different concentrations of lipopolysaccharide (LPS, phorbolmyristate and gammainterferon. IL-1β, TNF-α, IL-6 and IL-8 levels were measured in BAL fluid and culture supernatant using specific enzyme-linked immunosorbent assays. The substance found to stimulate the secretion of inflammatory cytokines to the greatest extent was LPS at a concentration of 10 μg/ml. Regarding the secretion of IL-1β, four observations were of interest: basal secretion was very low; LPS exerted a potent stimulatory effect; considerable within-group variability was observed; and there were no significant differences in the comparisons among groups. With respect to TNF-α secretion, the results were similar. The only striking finding was the higher basal secretion of this cytokine with respect to that of IL-1β. Regarding the secretion of IL-6, the same pattern followed by TNF-α was found. However, it should be stressed that the increase induced by LPS was smaller than in the two previous cytokines. Regarding the secretion of IL-8, three findings were patent: the strong basal secretion of this cytokine; the moderate increase induced by LPS; and the existence of significant differences among the different groups with respect to the stimulated secretion of this cytokine, which reached maximum values in patients with idiopathic pulmonary

  9. 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.

  10. EFFECTS OF DIESEL EXHAUST PARTICLES ON HUMAN ALVEOLAR MACROPHAGE RESPONSIVENESS TO LIPOPOLYSACCHARIDE

    Science.gov (United States)

    Effects of diesel exhaust particles on human alveolar macrophage responsiveness to lipopolysaccharideS. Mundandhara1 , S. Becker2 and M. Madden2, 1UNC Center for Environmental Medicine, Asthma, and Lung Biology, 2US EPA, NHEERL, HSD, Chapel Hill, NC, USEpidemiological...

  11. A Potent Tartrate Resistant Acid Phosphatase Inhibitor to Study the Function of TRAP in Alveolar Macrophages

    NARCIS (Netherlands)

    Boorsma, Carian E; van der Veen, T. Anienke; Putri, Kurnia S S; de Almeida, Andreia; Draijer, Christina; Mauad, Thais; Fejer, Gyorgy; Brandsma, Corry-Anke; van den Berge, Maarten; Bossé, Yohan; Sin, Don; Hao, Ke; Reithmeier, Anja; Andersson, Göran; Olinga, Peter; Timens, Wim; Casini, Angela; Melgert, Barbro N

    2017-01-01

    The enzyme tartrate resistant acid phosphatase (TRAP, two isoforms 5a and 5b) is highly expressed in alveolar macrophages, but its function there is unclear and potent selective inhibitors of TRAP are required to assess functional aspects of the protein. We found higher TRAP activity/expression in

  12. 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.

  13. Carbon Nanotube-Induced Pulmonary Granulomatous Disease: Twist1 and Alveolar Macrophage M1 Activation

    Directory of Open Access Journals (Sweden)

    Barbara P. Barna

    2013-12-01

    Full Text Available Sarcoidosis, a chronic granulomatous disease of unknown cause, has been linked to several environmental risk factors, among which are some that may favor carbon nanotube formation. Using gene array data, we initially observed that bronchoalveolar lavage (BAL cells from sarcoidosis patients displayed elevated mRNA of the transcription factor, Twist1, among many M1-associated genes compared to healthy controls. Based on this observation we hypothesized that Twist1 mRNA and protein expression might become elevated in alveolar macrophages from animals bearing granulomas induced by carbon nanotube instillation. To address this hypothesis, wild-type and macrophage-specific peroxisome proliferator-activated receptor gamma (PPARγ knock out mice were given oropharyngeal instillation of multiwall carbon nanotubes (MWCNT. BAL cells obtained 60 days later exhibited significantly elevated Twist1 mRNA expression in granuloma-bearing wild-type or PPARγ knock out alveolar macrophages compared to sham controls. Overall, Twist1 expression levels in PPARγ knock out mice were higher than those of wild-type. Concurrently, BAL cells obtained from sarcoidosis patients and healthy controls validated gene array data: qPCR and protein analysis showed significantly elevated Twist1 in sarcoidosis compared to healthy controls. In vitro studies of alveolar macrophages from healthy controls indicated that Twist1 was inducible by classical (M1 macrophage activation stimuli (LPS, TNFα but not by IL-4, an inducer of alternative (M2 macrophage activation. Findings suggest that Twist1 represents a PPARγ-sensitive alveolar macrophage M1 biomarker which is induced by inflammatory granulomatous disease in the MWCNT model and in human sarcoidosis.

  14. 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

  15. 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)

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

    Directory of Open Access Journals (Sweden)

    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

  17. In vitro toxicity of gallium arsenide in alveolar macrophages evaluated by magnetometry, cytochemistry and morphology.

    Science.gov (United States)

    Okada, M; Karube, H; Niitsuya, M; Aizawa, Y; Okayasu, I; Kotani, M

    1999-12-01

    Gallium arsenide (GaAs), a chemical compound of gallium and arsenic, causes various toxic effects including pulmonary diseases in animals. Since the toxicity is not completely investigated, GaAs has been used in workplaces as the material of various semiconductor products. The present study was conducted to clarify the toxicity of GaAs particles in the alveolar macrophages of hamsters using magnetometry, enzyme release assays and morphological examinations. Alveolar macrophages obtained from hamsters by tracheobronchial lavage and adhered to the disks in the bottom of wells were exposed to ferrosoferric oxide and GaAs particles. Ferrosoferric oxide particles were magnetized externally and the remanent magnetic field was measured. Relaxation, a fast decline of the remanent magnetic fields radiated from the alveolar macrophages, was delayed and decay constants were decreased dose-dependently due to exposure to GaAs. Because the relaxation is thought to be associated with cytoskeleton, the exposure of GaAs may have impaired the motor function of them. Enzyme release assay and morphological findings indicated the damage to the macrophages. Thus the cytotoxicity causes cytostructural changes and cell death. According to DNA electrophoresis and the TUNEL method, necrotic changes occur more frequently than apoptotic changes.

  18. 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

  19. 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.

  20. Early activation of the alveolar macrophage is critical to the development of lung ischemia-reperfusion injury.

    NARCIS (Netherlands)

    Naidu, BV; Krishnadasan, B; Farivar, AS; Woolley, SM; Thomas, R; Rooijen, van N.; Verrier, ED; Mulligan, MS

    2003-01-01

    .006) and marked reductions in bronchoalveolar lavage fluid leukocyte accumulation. Alveolar macrophage-depleted animals also demonstrated marked reductions of the elaboration of multiple proinflammatory chemokines and cytokines in the lavage effluent and nuclear transcription factors in lung

  1. 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.

  2. 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.

  3. 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

  4. Effects of inhaled alpha-emitting actinides on mouse alveolar macrophages.

    Science.gov (United States)

    Morgan, A; Talbot, R J

    1992-01-01

    The effects of inhaled alpha-emitting actinides on the alveolar macrophage (AM) population of the rodent lung are reviewed and, in particular, of the effects of 239PuO2 on murine AM. The effects discussed include changes the AM pool size, macrophage diameter, mobility, phagocytic competence, and enzyme content. Finally, similarities in the dose-response relationships for the induction of nuclear aberrations by alpha emitters and in the induction of lung tumors by the same materials are noted. PMID:1327735

  5. Effect of gamma irradiation on chemoluminescence of alveolar macrophages in Guinea Pig

    International Nuclear Information System (INIS)

    Benichou, G.; Dormont, D.; Herodin, F.; Pasquier, C.

    1988-01-01

    The capacity of Guinea Pig alveolar macrophages to produce oxygenated free radicals after in vitro gamma irradiation was studied. We observed an increase of resting cell chemiluminescence with 3 Gy, then an important decrease from 3 Gy to 30 Gy. Chemiluminescence of zymosan activated macrophages was reduced at 10 Gy and 30 Gy, while activation by IgG anti-MHC antigens was not affected by the same radiation doses. These results could be of great interest for the study pulmonary defense mechanisms after irradiation [fr

  6. 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

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  8. Chlamydia pneumoniae hides inside apoptotic neutrophils to silently infect and propagate in macrophages.

    Directory of Open Access Journals (Sweden)

    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.

  9. 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

  10. 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.

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

    Science.gov (United States)

    Moss, O. R.; Wong, V. A.

    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 (~25,000 cells adhered to a 0.7 cm2 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: 8x106, 8x105, and 8x104 28 nm beads per macrophage; and 8x104 and 1.12x104 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.

  12. Ceramide 1-phosphate inhibits serine palmitoyltransferase and blocks apoptosis in alveolar macrophages.

    Science.gov (United States)

    Granado, María H; Gangoiti, Patricia; Ouro, Alberto; Arana, Lide; Gómez-Muñoz, Antonio

    2009-04-01

    We previously reported that incubation of bone-marrow derived macrophages in the absence of macrophage-colony stimulating factor (M-CSF), a cytokine that is essential for their growth and survival, resulted in stimulation of acid sphingomyelinase, accumulation of ceramides, and induction of apoptosis [A. Gomez-Munoz et al. 2004. Ceramide 1-phosphate blocks apoptosis through inhibition of acid sphingomyelinase in macrophages. J Lipid Res 45: 99-105]. Here, we show that alveolar NR8383 macrophages, which are not dependent on M-CSF for viability, undergo apoptosis when they are incubated in the absence of serum. NR8383 cells showed increased levels of ceramides under apoptotic conditions, but in contrast to bone marrow macrophage acid and neutral sphingomyelinases were only slightly activated. We found that the major mechanism for ceramide generation in NR8383 macrophages was stimulation of their synthesis de novo. This action involved activation of serine palmitoyltransferase (SPT), the key regulatory enzyme of this pathway. A relevant finding was that ceramide 1-phosphate (C1P) inhibited SPT activity and ceramide accumulation leading to inhibition of apoptosis. Furthermore, C1P enhanced the activity of antiapoptotic protein kinase B and its downstream effector nuclear factor kappa B. These observations add a new dimension to the understanding of the pro-survival actions of C1P in mammalian cells.

  13. Metabolism of (/sup 3/H)benzo(a)pyrene by cultured human bronchus and cultured human pulmonary alveolar macrophages

    DEFF Research Database (Denmark)

    Autrup, Herman; Harris, C.C.; Stoner, G.D.

    1978-01-01

    The metabolism of (/sup 3/H)benzo(a)pyrene by cultured human bronchial epithelium and pulmonary alveolar macrophages was studied. Explants of bronchus were prepared and pulmonary alveolar macrophages were isolated from peripheral lung by trypsinization and by differential adhesion to plastic tissue...... culture dishes. After 7 days in culture the bronchus explant and the macrophages were exposed to (/sup 3/H)benzo(a)pyrene, and the binding to cellular macromolecules was studied. Aryl hydrocarbon hydroxylase activity was determined by the release of tritiated water into the culture medium from metabolized...

  14. 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.

  15. Mouse adenovirus type 1 infection of macrophages

    NARCIS (Netherlands)

    Ashley, S.L.; Welton, A.R.; Harwood, K.M.; Rooijen, van N.; Spindler, K.R.

    2009-01-01

    Mouse adenovirus type 1 (MAV-1) causes acute and persistent infections in mice, with high levels of virus found in the brain, spinal cord and spleen in acute infections. MAV-1 infects endothelial cells throughout the mouse, and monocytes/macrophages have also been implicated as targets of the virus.

  16. SPI-1-encoded type III secretion system of Salmonella enterica is required for the suppression of porcine alveolar macrophage cytokine expression

    Directory of Open Access Journals (Sweden)

    Pavlova Barbora

    2011-01-01

    Full Text Available Abstract Genes localized at Salmonella pathogenicity island-1 (SPI-1 are involved in Salmonella enterica invasion of host non-professional phagocytes. Interestingly, in macrophages, SPI-1-encoded proteins, in addition to invasion, induce cell death via activation of caspase-1 which also cleaves proIL-1β and proIL-18, precursors of 2 proinflammatory cytokines. In this study we were therefore interested in whether SPI-1-encoded type III secretion system (T3SS may influence proinflammatory response of macrophages. To test this hypothesis, we infected primary porcine alveolar macrophages with wild-type S. Typhimurium and S. Enteritidis and their isogenic SPI-1 deletion mutants. ΔSPI1 mutants of both serovars invaded approx. 5 times less efficiently than the wild-type strains and despite this, macrophages responded to the infection with ΔSPI1 mutants by increased expression of proinflammatory cytokines IL-1β, IL-8, TNFα, IL-23α and GM-CSF. Identical macrophage responses to that induced by the ΔSPI1 mutants were also observed to the infection with sipB but not the sipA mutant. The hilA mutant exhibited an intermediate phenotype between the ΔSPI1 mutant and the wild-type S. Enteritidis. Our results showed that the SPI-1-encoded T3SS is required not only for cell invasion but in macrophages also for the suppression of early proinflammatory cytokine expression.

  17. [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.

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

    Directory of Open Access Journals (Sweden)

    Jones William

    2006-03-01

    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

  19. Hemosiderin in sputum macrophages may predict infective exacerbations of chronic obstructive pulmonary disease: a retrospective observational study.

    Science.gov (United States)

    Mohan, Sindu; Ho, Terence; Kjarsgaard, Melanie; Radford, Katherine; Borhan, A S M; Thabane, Lehana; Nair, Parameswaran

    2017-04-12

    Infective exacerbations of COPD are common and are accompanied by neutrophilic bronchitis in sputum. Increased respiratory iron content has been associated with respiratory tract infection, though it is unclear if this represents a predisposing factor for infection or the sequelae of inflammation. Iron overload, as assessed in the airways, may be an important biomarker for recurrent infective exacerbations of COPD. The purpose of our study was to determine if hemosiderin in sputum macrophages is related to infective exacerbations of COPD. We undertook a retrospective observational study of 54 consecutive patients who presented with an exacerbation of COPD and had sputum examined including assessment for hemosiderin in alveolar macrophages. The relation between infective exacerbations in the previous two years and the percent of hemosiderin-positive macrophages was analyzed with linear regression. To account for the non-parametric distribution of infective exacerbations, negative binomial regression modelling was used to account for other covariates. The percent of hemosiderin positive alveolar macrophages (hemosiderin index), analyzed parametrically and non-parametrically, demonstrated a significant correlation with increasing numbers of infective exacerbations in the previous two years. In a multivariate regression analysis, hemosiderin index was an independent predictor of infective exacerbations. COPD patients with raised hemosiderin index (≥20%) had higher levels of sputum IL-6 compared to patients with lower levels (hemosiderin index in sputum alveolar macrophages measured at the time of AECOPD may be related to the frequency of infective exacerbations of COPD.

  20. 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

  1. Differential and strain-specific triggering of bovine alveolar macrophage effector functions by mycoplasmas.

    Science.gov (United States)

    Jungi, T W; Krampe, M; Sileghem, M; Griot, C; Nicolet, J

    1996-12-01

    Mycoplasma strains being considered as pathogenic or non-pathogenic for cattle were tested on their capacity to activate bovine alveolar macrophages in vitro. Of particular interest was the behaviour of Mycoplasma mycoides ssp. mycoides small colony type (M.m.m. SC), the causative agent of contagious bovine pleuropneumonia (CBPP). Increases in procoagulant activity (PCA), tumor necrosis factor-alpha- (TNF-alpha) and nitrogen monoxide (NO) generation were tested. To minimize an influence of macrophage activation by mycoplasma growth media, mycoplasmas were cultured on embryonic calf nose epithelial cells. The three macrophage functions tested were not correlated, but were differentially induced in strain-specific manner. Four out of seven strains induced PCA, regardless of pathogenicity, and all strains promoted moderate NO generation at high concentrations. All tested M.m.m. SC strains (Afadé, L2 and PG1), and the pathogenic M. bovis, induced TNF-alpha production at low concentrations (10(6) colony forming units per ml). M.sp. serogroup 7 and the non-pathogenic M. bovirhinis and Acholeplasma laidlawii did not induce TNF-alpha up to 10(8) cfu/ml. Thus, strain-specific differences are reflected in differential macrophage activation patterns. The findings are consistent with an important role for TNF-alpha in pathogenesis of CBPP.

  2. 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)

  3. Modulation of the effects of alveolar macrophages on lung fibroblast collagen production rate

    Energy Technology Data Exchange (ETDEWEB)

    Clark, J.G.; Greenberg, J.

    1987-01-01

    Alveolar macrophages (AM) may function as effector cells that can either stimulate or inhibit lung fibroblast collagen production. However, conditions that determine the predominant effect of AM on fibroblasts are not well understood. To delineate factors that modulate the effects of AM on lung fibroblasts, we studied the interaction of AM products and fibroblasts in vitro. The AM were obtained by bronchoalveolar lavage of hamsters with bleomycin-induced pulmonary fibrosis. Conditioned medium (CM) from the AM cultures was incubated in varying amounts with lung fibroblast (IMR-90) cultures. After metabolic labeling with (/sup 3/H)proline, fibroblast collagen production based on procollagen-specific radioactivity was determined. Macrophage CM in concentrations greater than 5% suppressed collagen production, an event attributed to the macrophage-derived suppressive factor that we have previously characterized. Macrophages were also determined to produce PGE2 in culture. Authentic PGE2 at concentrations found in CM was found to suppress fibroblast collagen production, indicating that AM-derived PGE2 contributes to the suppressive activity in CM. To examine possible stimulatory factors in CM, the fibroblasts were preincubated with indomethacin. This approach was based on our previous observation that AM-derived suppressive factor increases endogenous fibroblast PGE2 and that its activity can be blocked by indomethacin. Macrophage CM in a concentration of 20% did not suppress the collagen production of indomethacin-treated fibroblasts. However, CM concentrations of 5 and 10% increased collagen production (173 and 143% of control values, respectively), indicating the presence of stimulatory factor(s) in macrophage-conditioned medium.

  4. Electron-microscopic studies of alveolar macrophages from gamma-ray irradiated guinea pigs

    International Nuclear Information System (INIS)

    Krystev, Kh.; Najdenski, Kh.M.; Velyanov, D.K.; Radoevska, S.A.

    1988-01-01

    The alveolar macrophages (AM) were obtained from whole body gamma-irradiated guinea pigs (0.5 Gy and 2 Gy; 92.5 rad/min). The cell suspension contained granulocytes, lymphocytes and disintegrating epithelial and white blood cells, as well as two types of microphages: large (possessing nuclei of saddle bag-like or highly folded form) and small (with spherical or eggshaped nuclei). Eleven electronograms were presented showing all ultrastuctural changes of both small and large AM. The morphological differences between the small and large alveolar macrophages were slight. Marked changes were observed in the large AM on day 1 following 0.5 Gy irradiation: a considerable increase in dimensions of phagosomes turning in digestive vacuoles, lamellarly limited and containing osmiophilic, irregularly formed, densely or lamellarly arranged matter; folded nuclei with slightly vacuolized cytoplasm. The ultrastructural changes in the AM of sublethal dose (2 Gy) irradiated animals were stronger and regenerative processes in them were less possible. On day 30 after irradiation several damaged AM were observed with large digestive vacuoles of fine-grain content, vacuolized endoplasmic reticulum, entirely lyzed nuclear chromatin and free nuclei without cytoplasm. All observations were a convincing indication that guinea pigs AM were more radiosensitive than that obtained from rats and mice

  5. 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)

  6. Functional alterations of alveolar macrophages subjected to smoke exposure and antioxidant lazaroids.

    Science.gov (United States)

    Wang, S; Lantz, R C; Vermeulen, M W; Chen, G J; Breceda, V; Robledo, R F; Hays, A M; Young, S; Witten, M L

    1999-08-01

    Acute inhalation of diesel fuel-polycarbonate plastic (DFPP) smoke causes severe lung injury, leading to acute respiratory distress syndrome (ARDS) and death. It has been reported that the initiation of acute lung injury is associated with the activation of pulmonary alveolar macrophages (PAM). To further explore the pathogenesis, alveolar macrophages (AM) of New Zealand rabbits ventilated and exposed to a 60 tidal volume of DFPP smoke in vivo were recovered at 1 h post-smoke. Smoke exposure induced significant increases in both mRNA and protein levels for PAM tumor necrosis factor-alpha (TNF-alpha), when compared to smoke control. Smoke also induced a biphasic response (inhibited at 2 h, enhanced at 24 h after cell isolation) in the production of superoxide (O2-) by PAM. However, aerosolized lazaroid, U75412E (1.6 mg/kg body weight), significantly attenuated smoke-induced expression in AM TNF-alpha at the protein level but not at the mRNA level, and smoke-induced changes in AM production of O2-. This study suggests that highly expressing AM TNF-alpha following smoke may be a key contributor to the cascade that establishes an acute injury process and exacerbates oxidant-derived cell injury. Whereas, the lazaroid may ameliorate smoke-induced lung injury by attenuating AM TNF-alpha release, in addition to its primary antioxidative mechanism.

  7. 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

  8. Immunomodulation of the response to excretory/secretory antigens of Fasciola hepatica by Anapsos in Balb/C mice and rat alveolar macrophages.

    Science.gov (United States)

    López-Abán, Julio; Andrade, Maria do Amparo; Nogal-Ruiz, Juan José; Martínez-Fernández, Antonio R; Muro, Antonio

    2007-04-01

    It is known that excretory/secretory antigens of Fasciola hepatica (ESFh) trigger a Th2-like immune response. Anapsos (A) is an aqueous hydrosoluble extract obtained from the rhizomes of the fern Polypodium leucotomos that has shown immunomodulator effects in some parasitic infections and immunological disorders. In this work we assess the effect of Anapsos and ESFh and Quillaja saponaria extract (Qs) on BALB/c mice and rat alveolar macrophages. Anapsos modulates the response of mice immunized with ESFh, decreasing IgG antibodies in A+ESFh- and A+Qs+ESFh-treated mice and triggering high levels of gammaIFN in spleen cell culture in comparison with ESFh- and Qs + ESFh--treated groups. Moreover, Anapsos showed statistically significant inhibitory effects on the nitrite production by rat alveolar macrophages prestimulated with lipopolysaccharide (LPS) as well as ESFh antigen in comparison with macrophages stimulated only with LPS. The application of ESFh and Anapsos combined avoids this inhibitory effect. Thus, Anapsos modulates the immune response against ESFh in naive mice and on the nitrite production in prestimulated rat aveolar macrophages.

  9. 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

  10. 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

  11. 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

  12. Role of macrophages in early host resistance to respiratory Acinetobacter baumannii infection.

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    Hongyu Qiu

    Full Text Available Acinetobacter baumannii is an emerging bacterial pathogen that causes nosocomial pneumonia and other infections. Although it is recognized as an increasing threat to immunocompromised patients, the mechanism of host defense against A. baumannii infection remains poorly understood. In this study, we examined the potential role of macrophages in host defense against A. baumannii infection using in vitro macrophage culture and the mouse model of intranasal (i.n. infection. Large numbers of A. baumannii were taken up by alveolar macrophages in vivo as early as 4 h after i.n. inoculation. By 24 h, the infection induced significant recruitment and activation (enhanced expression of CD80, CD86 and MHC-II of macrophages into bronchoalveolar spaces. In vitro cell culture studies showed that A. baumannii were phagocytosed by J774A.1 (J774 macrophage-like cells within 10 minutes of co-incubation, and this uptake was microfilament- and microtubule-dependent. Moreover, the viability of phagocytosed bacteria dropped significantly between 24 and 48 h after co-incubation. Infection of J774 cells by A. baumannii resulted in the production of large amounts of proinflammatory cytokines and chemokines, and moderate amounts of nitric oxide (NO. Prior treatment of J774 cells with NO inhibitors significantly suppressed their bactericidal efficacy (P<0.05. Most importantly, in vivo depletion of alveolar macrophages significantly enhanced the susceptibility of mice to i.n. A. baumannii challenge (P<0.01. These results indicate that macrophages may play an important role in early host defense against A. baumannii infection through the efficient phagocytosis and killing of A. baumannii to limit initial pathogen replication and the secretion of proinflammatory cytokines and chemokines for the rapid recruitment of other innate immune cells such as neutrophils.

  13. 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

  14. 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

  15. 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.

  16. Macrophages as drivers of an opportunistic infection

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    Annette C. Vergunst

    2017-09-01

    Full Text Available Opportunistic pathogens are a worldwide cause of mortality and morbidity, and infections with intrinsically antibiotic-resistant pathogens have a large clinical, social and economic impact. Bacteria belonging to the Burkholderia cepacia complex (Bcc, ubiquitous in natural and industrial environments, are notorious pathogens for individuals with cystic fibrosis (CF. In addition, Burkholderia cenocepacia is emerging as the culprit of non-CF related, sometimes fatal infections. Knowledge of the underlying infection mechanism of these pathogens is important for efficient treatment, however, to date not much is known about the lifestyle of Bcc bacteria during infection. In our recent study published in PLoS Pathogens, we provide experimental evidence that macrophages are critically important for proliferation of B. cenocepacia, and are major drivers of fatal pro-inflammatory infections in zebrafish larvae. This is in agreement with recent clinical information showing that B. cenocepacia is mainly localised in phagocytes in infected CF lungs. A predominant intramacrophage stage and a host-detrimental role for macrophages have major implications for treatment strategies of both CF and non-CF infections. Intracellular survival of bacteria traditionally classified as extracellular, including Staphylococcus aureus and Pseudomonas aeruginosa, is an emerging theme. Our finding that macrophages are essential for proliferation of B. cenocepacia in the host suggests a new paradigm for Bcc infections and urges the development of novel anti-infectious therapies to efficiently disarm these intrinsically antibiotic resistant facultative intracellular pathogens.

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

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    Schwarze Per E

    2006-06-01

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

  18. 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

  19. 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

  20. Macrophage Migration Inhibitory Factor in Protozoan Infections

    Directory of Open Access Journals (Sweden)

    Marcelo T. Bozza

    2012-01-01

    Full Text Available Macrophage migration inhibitory factor (MIF is a cytokine that plays a central role in immune and inflammatory responses. In the present paper, we discussed the participation of MIF in the immune response to protozoan parasite infections. As a general trend, MIF participates in the control of parasite burden at the expense of promoting tissue damage due to increased inflammation.

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

    Science.gov (United States)

    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.

  3. 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.

  4. 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

  5. DNA repair in baboon alveolar macrophages: a system for assessing biohazardous materials

    Energy Technology Data Exchange (ETDEWEB)

    Meltz, M.L.

    1976-06-01

    The repair of DNA in the alveolar macrophages of the baboon has been investigated after treatment of the cells in vitro with ultraviolet light and with the alkylating agent, methyl methanesulfonate. Repair replication has been observed to occur after ultraviolet-light irradiation of macrophages attached to plastic dishes. The two different techniques used for measuring repair replication were a standard density-gradient procedure which separates normal-density, repair-replicated, preexisting DNA from semiconservatively synthesized DNA containing sufficient (/sup 3/H)bromodeoxyuridine to cause a density shift; and an alternative procedure applicable to nonproliferating cell systems wherein residual semiconservative synthesis is inhibited by hydroxyurea, and the increased incorporation of the DNA precursor (/sup 3/H)thymidine due to repair replication is measured. The latter technique was applied to the investigation of the concentration dependence of repair replication both after and during the treatment of the cells with the alkylating agent, methyl methanesulfonate. The labeling after treatment measures the repair of damage still present at the end of the treatment interval, while the labeling during treatment allows for greater sensitivity and detection of all repair occurring during the treatment period.

  6. 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.

  7. Lipid-Laden Alveolar Macrophages and pH Monitoring in Gastroesophageal Reflux-Related Respiratory Symptoms

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    R. Kitz

    2012-01-01

    Full Text Available Lipid-laden alveolar macrophages and pH monitoring have been used in the diagnosis of chronic aspiration in children with gastroesophageal reflux (GER. This study was conducted to prove a correlation between the detection of alimentary pulmonary fat phagocytosis and an increasing amount of proximal gastroesophageal reflux. It was assumed that proximal gastroesophageal reflux better correlates with aspiration than distal GER. Patients from 6 months to 16 years with unexplained recurrent wheezy bronchitis and bronchial hyperreactivity, or recurrent pneumonia with chronic cough underwent 24-hour double-channel pH monitoring and bronchoscopy with bronchoalveolar lavage (BAL. Aspiration of gastric content was determined by counting lipid laden alveolar macrophages from BAL specimens. There were no correlations between any pH-monitoring parameters and counts of lipid-laden macrophages in the whole study population, even when restricting analysis to those with abnormal reflux index expressing clinically significant GER. Quantifying lipid-laden alveolar macrophages from BAL in children with gastroesophageal-related respiratory disorders does not have an acceptable specificity to prove chronic aspiration as an underlying etiology. Therefore, research for other markers of pulmonary aspiration is needed.

  8. [Role of oxidative stress in endoplasmic reticulum stress? induced apoptosis of alveolar macrophages triggered by quartz dust].

    Science.gov (United States)

    Song, Jing; Lu, Xiaoting; Li, Qiuying; Liu, Chengyun; Liu, Ying

    2014-07-01

    To investigate the role of oxidative stress in the endoplasmic reticulum stress-induced apoptosis of alveolar macrophages triggered by quartz dust. Seventy-two healthy adult Wistar rats were randomly divided into control group, quartz dust group, quartz dust plus N-acetyl cysteine (NAC) group, and NAC group, with 18 rats in each group. One milliliter of sterile saline (for the control and NAC groups) or 1 ml of saline with 5%ultrafine quartz dust (for dust group and dust plus NAC group) was given to each rat by non-exposed endotracheal infusion. From the second day after dust infusion, rats in dust plus NAC group and NAC group received intragastric administration of NAC (100 mg/kg). In each week, the treatment with NAC lasted for 5 consecutive days, followed by 2 days' interval. For each group, 6 rats were randomly selected on the 14th, 28th, or 56th day after dust exposure; they were sacrificed by bloodletting from the femoral artery, and the lungs were collected. Bronchoalveolar lavage fluid was collected to separate macrophages. The protein expression of caspase-12 in alveolar macrophages, the apoptosis rate and reactive oxygen species (ROS) content of alveolar macrophages, and the protein carbonyl content of alveolar macrophages were determined by Western blot, flow cytometry, and colorimetry, respectively. Increased protein expression of caspase-12, apoptosis rate, and content of ROS and protein carbonyl were discovered on the 14th day in the dust group, in comparison with the control group (P quartz dust. Oxidative damage of protein in the endoplasmic reticulum may play an important role in the process.

  9. 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.

  10. Ameobal pathogen mimivirus infects macrophages through phagocytosis.

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    Eric Ghigo

    2008-06-01

    Full Text Available Mimivirus, or Acanthamoeba polyphaga mimivirus (APMV, a giant double-stranded DNA virus that grows in amoeba, was identified for the first time in 2003. Entry by phagocytosis within amoeba has been suggested but not demonstrated. We demonstrate here that APMV was internalized by macrophages but not by non-phagocytic cells, leading to productive APMV replication. Clathrin- and caveolin-mediated endocytosis pathways, as well as degradative endosome-mediated endocytosis, were not used by APMV to invade macrophages. Ultrastructural analysis showed that protrusions were formed around the entering virus, suggesting that macropinocytosis or phagocytosis was involved in APMV entry. Reorganization of the actin cytoskeleton and activation of phosphatidylinositol 3-kinases were required for APMV entry. Blocking macropinocytosis and the lack of APMV colocalization with rabankyrin-5 showed that macropinocytosis was not involved in viral entry. Overexpression of a dominant-negative form of dynamin-II, a regulator of phagocytosis, inhibited APMV entry. Altogether, our data demonstrated that APMV enters macrophages through phagocytosis, a new pathway for virus entry in cells. This reinforces the paradigm that intra-amoebal pathogens have the potential to infect macrophages.

  11. Key Role of the Scavenger Receptor MARCO in Mediating Adenovirus Infection and Subsequent Innate Responses of Macrophages.

    Science.gov (United States)

    Maler, Mareike D; Nielsen, Peter J; Stichling, Nicole; Cohen, Idan; Ruzsics, Zsolt; Wood, Connor; Engelhard, Peggy; Suomalainen, Maarit; Gyory, Ildiko; Huber, Michael; Müller-Quernheim, Joachim; Schamel, Wolfgang W A; Gordon, Siamon; Jakob, Thilo; Martin, Stefan F; Jahnen-Dechent, Willi; Greber, Urs F; Freudenberg, Marina A; Fejer, György

    2017-08-01

    The scavenger receptor MARCO is expressed in several subsets of naive tissue-resident macrophages and has been shown to participate in the recognition of various bacterial pathogens. However, the role of MARCO in antiviral defense is largely unexplored. Here, we investigated whether MARCO might be involved in the innate sensing of infection with adenovirus and recombinant adenoviral vectors by macrophages, which elicit vigorous immune responses in vivo Using cells derived from mice, we show that adenovirus infection is significantly more efficient in MARCO-positive alveolar macrophages (AMs) and in AM-like primary macrophage lines (Max Planck Institute cells) than in MARCO-negative bone marrow-derived macrophages. Using antibodies blocking ligand binding to MARCO, as well as gene-deficient and MARCO-transfected cells, we show that MARCO mediates the rapid adenovirus transduction of macrophages. By enhancing adenovirus infection, MARCO contributes to efficient innate virus recognition through the cytoplasmic DNA sensor cGAS. This leads to strong proinflammatory responses, including the production of interleukin-6 (IL-6), alpha/beta interferon, and mature IL-1α. These findings contribute to the understanding of viral pathogenesis in macrophages and may open new possibilities for the development of tools to influence the outcome of infection with adenovirus or adenovirus vectors. IMPORTANCE Macrophages play crucial roles in inflammation and defense against infection. Several macrophage subtypes have been identified with differing abilities to respond to infection with both natural adenoviruses and recombinant adenoviral vectors. Adenoviruses are important respiratory pathogens that elicit vigorous innate responses in vitro and in vivo The cell surface receptors mediating macrophage type-specific adenovirus sensing are largely unknown. The scavenger receptor MARCO is expressed on some subsets of naive tissue-resident macrophages, including lung alveolar macrophages

  12. Alveolar macrophage graded hemosiderin score from bronchoalveolar lavage in horses with exercise-induced pulmonary hemorrhage and controls.

    Science.gov (United States)

    Doucet, Michèle Y; Viel, Laurent

    2002-01-01

    The objective of this study was to determine if a quantitative scoring system for evaluation of hemosiderin content of alveolar macrophages obtained by bronchoalevolar lavage provides a more sensitive test for the detection of exercise-induced pulmonary hemorrhage (EIPH) in horses than does endoscopy of the lower airways. A sample population composed of 74 Standardbred racehorses aged 2-5 years was used. Horses were grouped as either control (EIPH-negative) or EIPH-positive based on history and repeated postexertional endoscopic evaluation of the bronchial airways. Bronchoalveolar lavage was performed and cytocentrifuge slides were stained with Perl's Prussian blue. Alveolar macrophages were scored for hemosiderin content by a method described by Golde and associates to obtain the total hemosiderin score (THS). Test performance criteria were determined with a contingency table. All subjects had some degree of hemosiderin in the alveolar macrophages, regardless of group. The distribution of cells among the different grades followed a significantly different pattern for the control group versus horses with EIPH (P hemosiderin by means of the Golde scoring system shows promise as a more sensitive approach than repeated postexertional endoscopy alone to detect EIPH.

  13. [Role of calcium activated-potassium channels in the injury to rat alveolar macrophages induced by quartz].

    Science.gov (United States)

    Li, Jun; Sun, Jingzhi; Yang, Li; Zhao, Jing; Wang, Zhenglun; Yang, Lei

    2014-01-01

    To investigate the role of calcium activated-potassium channels (KCa) in the injury to rat alveolar macrophages induced by quartz. The experiments were conducted on a rat alveolar macrophage cell line (NR8383) in vitro, where crystal silica (100 üg/ml) and amorphous silica (100 üg/ml) were used as the test substances and the cells without any treatment as negative controls. At first the effects of two kinds of quartz were compared. Then KCa special inhibitors (Paxilline for BK, Tram-34 for IK, Apamin for SK) were added in different doses to the in vitro test system with 100 üg/ml crystal quartz as matrix, to observe the function of such channels. Cell viability, lactate dehydrogenase (LDH), interleukin-1β (IL-1β) and tumor necrosis factor-a (TNF-α) were tested. Comparing to the negative control group, cell viability reduced, LDH leakage, IL-1β and TNF-α release increased significantly in the amorphous quartz group, furthermore, the effects by crystal quartz were much more serious than those by amorphous quartz, with a statistical significance (P quartz group, IK blockers (Tram-34) led to increase in cell viability significantly, with a statistical significance (P quartz in the rat alveolar macrophages cell line in vitro, which might serve as a signal in the early regulation of inflammatory responses by quartz.

  14. Three-dimensional characteristics of alveolar macrophages in vitro observed by dark field microscopy

    Science.gov (United States)

    Swarat, Dominic; Wiemann, Martin; Lipinski, Hans-Gerd

    2014-05-01

    Alveolar macrophages (AM) are cells from immune defense inside the lung. They engulf particles in vacuoles from the outer membrane. Volume and surface are important parameters to characterize the particle uptake. AM change their shape within a few seconds, therefore it is hard to obtain by confocal laser scanning microscopy, which is commonly used to generate 3D-images. So we used an intensified dark field microscopy (DFM) as an alternative method to generate contrast rich AM gray tone image slices used for 3D-reconstructions of AM cells by VTK software applications. From these 3D-reconstructions approximate volume and surface data of the AM were obtained and compared to values found in the literature. Finally, simple geometrical 3D-models of the AM were created and compared to real data. Averaged volume and surface data from the DFM images are close to values found in the literature. Furthermore, calculation of volume and surface data from DFM images could be done faster if simplified geometrical 3D-models of the cells were used.

  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. Investigation of fine chalk dust particles' chemical compositions and toxicities on alveolar macrophages in vitro.

    Science.gov (United States)

    Zhang, Yuexia; Yang, Zhenhua; Li, Ruijin; Geng, Hong; Dong, Chuan

    2015-02-01

    The aim of the study is to investigate chemical compositions of fine chalk dust particles (chalk PM2.5) and examine their adverse effects on alveolar macrophages (AMs) in vitro. Morphologies and element concentrations of individual chalk particles were analyzed by using the quantitative energy-dispersive electron probe X-ray microanalysis (ED-EPMA). The oxidative response of AMs and the potential to generate nitric oxide (NO) by luminol-dependent chemiluminescence (CL) and nitrate reductase method were assessed 4h following the treatment of AMs with differing dosages of fine chalk particles, respectively. Oxidative stress and cytotoxicity elicited by chalk PM2.5 were also examined. The results showed that fine chalk particles were mainly composed of gypsum, calcite, dolomite and a little amount of organic adhesives. Exposure to chalk PM2.5 at 100 μg mL(-1) or 300 μg mL(-1) significantly increased intracellular catalase, malondialdehyde, and NO levels and decreased superoxide dismutase level in AMs, leading to leakage of lactate dehydrogenase (LDH) and reduction of the cell viability. Furthermore, luminol-dependent CL from respiratory burst in AMs was enhanced. It was suggested that chalk PM2.5 could make oxidative damages on AMs and result in cytotoxicity, being likely attributed to excessive reactive oxygen species or reactive nitrogen species induced by mixture of fine gypsum and calcite/dolomite particles. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. 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)

  18. 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

  19. Protein tyrosine kinase but not protein kinase C inhibition blocks receptor induced alveolar macrophage activation

    Directory of Open Access Journals (Sweden)

    K. Pollock

    1993-01-01

    Full Text Available The selective enzyme inhibitors genistein and Ro 31-8220 were used to assess the importance of protein tyrosine kinase (PTK and protein kinase C (PKC, respectively, in N-formyl-methionyl-leucyl-phenylalanine (FMLP induced generation of superoxide anion and thromboxane B2 (TXB2 in guinea-pig alveolar macrophages (AM. Genistein (3–100 μM dose dependently inhibited FMLP (3 nM induced superoxide generation in non-primed AM and TXB2 release in non-primed or in lipopolysaccharide (LPS (10 ng/ml primed AM to a level > 80% but had litle effect up to 100 μM on phorbol myristate acetate (PMA (10 nM induced superoxide release. Ro 31-8220 inhibited PMA induced superoxide generation (IC50 0.21 ± 0.10 μM but had no effect on or potentiated (at 3 and 10 μM FMLP responses in non-primed AM. In contrast, when present during LPS priming as well as during FMLP challenge Ro 31-8220 (10 μM inhibited primed TXB2 release by > 80%. The results indicate that PTK activation is required for the generation of these inflammatory mediators by FMLP in AM. PKC activation appears to be required for LPS priming but not for transducing the FMLP signal; rather, PKC activation may modulate the signal by a negative feedback mechanism.

  20. Pulmonary alveolar proteinosis

    Science.gov (United States)

    PAP; Alveolar proteinosis; Pulmonary alveolar phospholipoproteinosis; Alveolar lipoproteinosis phospholipidosis ... PAP is unknown. In others, it occurs with lung infection or an immune problem. It also can ...

  1. HIV Infection of Macrophages: Implications for Pathogenesis and Cure

    Directory of Open Access Journals (Sweden)

    Kiera Leigh Clayton

    2017-05-01

    Full Text Available Although CD4+ T cells represent the major reservoir of persistent HIV and SIV infection, accumulating evidence suggests that macrophages also contribute. However, investigations of the role of macrophages are often underrepresented at HIV pathogenesis and cure meetings. This was the impetus for a scientific workshop dedicated to this area of study, held in Cambridge, MA in January 2017. The workshop brought together experts in the fields of HIV/SIV immunology/virology, macrophage biology and immunology, and animal models of HIV/SIV infection to facilitate discussions regarding the role of macrophages as a physiologically relevant viral reservoir, and the implications of macrophage infection for HIV pathogenesis and cure strategies. An emerging consensus that infected macrophages likely persist in the setting of combination antiretroviral therapy, driving persistent inflammation and contributing to the viral reservoir, indicate the importance of addressing macrophages as well as CD4+ T cells with future therapeutic strategies.

  2. The effect of oxygen and paraquat on the 14C-glucose oxidation of rabbit alveolar macrophages and lung slices

    International Nuclear Information System (INIS)

    Roussouw, D.J.; Engelbrecht, F.M.

    1979-01-01

    In this study, we measured the effects of different concentrations of paraquat (0,01 mM and 1,0 mM) on the 1- 14 CO 2 and 6- 14 CO 2 production of rabbit lung slices and isolated alveolar macrophages, in 20% and 95% oxygen phases respectively. A 95% oxygen phase induced an increase in the 6- 14 C-glucose oxidation of control lung slices over a 3-hour period, while the increased activity of the pentose pathway over the first 2 hours started to decline during the third hour of incubation. Paraquat (1,0mM) in 20% oxygen caused a consistent increase in the 6- 14 CO 2 production by lung slices, but in a 95% oxygen phase gradually inhibited the 6- 14 C-glucose oxidation over a period of 3 hours. The pentose phosphate pathway was highly significantly stimulated by 1,0 mM paraquat in 20% and 95% oxygen over 3 hours. When isolated alveolar macrophages (viability 95%) were incubated in a 20% and 95% oxygen phase respectively, both the 6- 14 C-glucose and 1- 14 C-glucose oxidation rates were significantly inhibited by 1,0 mM paraquat after 1 hour. Our results confirmed the initial increase in glycolytic metabolism induced by paraquat, but also indicated that the 6- 14 CO 2 production was significantly inhibited by paraquat when lung slices were incubated in a 95% oxygen phase. The fact that the glucose metabolism in alveolar macrophages is more sensitive to paraquat exposure than that of cells in lung slices may be related to the genesis of the intra-alveolar pulmonary lesions described in the literature

  3. 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.

  4. 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.

  5. 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.

  6. The macrophage in HIV-1 infection: From activation to deactivation?

    Directory of Open Access Journals (Sweden)

    Varin Audrey

    2010-04-01

    Full Text Available Abstract Macrophages play a crucial role in innate and adaptative immunity in response to microorganisms and are an important cellular target during HIV-1 infection. Recently, the heterogeneity of the macrophage population has been highlighted. Classically activated or type 1 macrophages (M1 induced in particular by IFN-γ display a pro-inflammatory profile. The alternatively activated or type 2 macrophages (M2 induced by Th-2 cytokines, such as IL-4 and IL-13 express anti-inflammatory and tissue repair properties. Finally IL-10 has been described as the prototypic cytokine involved in the deactivation of macrophages (dM. Since the capacity of macrophages to support productive HIV-1 infection is known to be modulated by cytokines, this review shows how modulation of macrophage activation by cytokines impacts the capacity to support productive HIV-1 infection. Based on the activation status of macrophages we propose a model starting with M1 classically activated macrophages with accelerated formation of viral reservoirs in a context of Th1 and proinflammatory cytokines. Then IL-4/IL-13 alternatively activated M2 macrophages will enter into the game that will stop the expansion of the HIV-1 reservoir. Finally IL-10 deactivation of macrophages will lead to immune failure observed at the very late stages of the HIV-1 disease.

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

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yanling; Xu, Sanpeng; Xiao, Fei; Xiong, Yan; Wang, Xiaojin; Gao, Sui; Yan, Weiming [Department and Institute of Infectious Disease, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030 (China); Ning, Qin, E-mail: qning@tjh.tjmu.edu.cn [Department and Institute of Infectious Disease, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030 (China)

    2010-05-28

    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.

  8. Activation of Alveolar Macrophages with Interferon-γ Promotes Antioxidant Defenses via the Nrf2-ARE Pathway

    Science.gov (United States)

    Staitieh, Bashar S; Egea, Eduardo E; Fan, Xian; Azih, Nnamdi; Neveu, Wendy; Guidot, David M

    2015-01-01

    Macrophage phenotype and function is dependent on the underlying microenvironment. Many diseases are accompanied by abnormal shifts in macrophage polarization state that limit the ability of the cells to become innate immune effectors. Previous work in the field suggests that chronic alcohol ingestion, which is associated with a shift away from innate immune effector macrophages, is also associated with a deficient response to oxidative stress. We therefore hypothesized that the optimal response to oxidative stress was dependent on the ability of the macrophage to become an innate immune effector cell. To investigate this hypothesis, we first confirmed that we could reproducibly polarize NR8383 cells (a rat alveolar macrophage cell line) into the prototypical M1 and M2 states (using IFN-γ and IL-4, respectively). We then tested the polarized cells for their ability to scavenge reactive oxygen species generated by glucose oxidase (GOX) using the Amplex red assay and found that IFN-γ-polarized cells had greater scavenging capacity. To elucidate the mechanism of the enhanced response to oxidative stress, we then assessed key components of the anti-oxidant response; specifically, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), the master transcription factor responsible for the cellular response to oxidative stress, and one of its downstream effectors, glutamate-cysteine ligase catalytic subunit (GCLC). We found that both proteins were significantly upregulated in the IFN-γ-polarized cells. To confirm that Nrf2 is an integral component of this improved anti-oxidant response, we transfected IFN-γ-polarized cells with either silencing RNA to Nrf2 or control silencing RNA and found that hydrogen peroxide scavenging was significantly impaired in the si-Nrf2-treated cells. Further, transfecting untreated cells with si-Nrf2 polarized them toward the M2 phenotype in the absence of IL-4, suggesting a mechanistic role for Nrf2 in macrophage polarization. We then

  9. Role of Monocyte/Macrophages during HIV/SIV Infection in Adult and Pediatric Acquired Immune Deficiency Syndrome

    Directory of Open Access Journals (Sweden)

    Kristen M. Merino

    2017-12-01

    Full Text Available Monocytes/macrophages are a diverse group of cells that act as first responders in innate immunity and then as mediators for adaptive immunity to help clear infections. In performing these functions, however, the macrophage inflammatory responses can also contribute to pathogenesis. Various monocyte and tissue macrophage subsets have been associated with inflammatory disorders and tissue pathogeneses such as occur during HIV infection. Non-human primate research of simian immunodeficiency virus (SIV has been invaluable in better understanding the pathogenesis of HIV infection. The question of HIV/SIV-infected macrophages serving as a viral reservoir has become significant for achieving a cure. In the rhesus macaque model, SIV-infected macrophages have been shown to promote pathogenesis in several tissues resulting in cardiovascular, metabolic, and neurological diseases. Results from human studies illustrated that alveolar macrophages could be an important HIV reservoir and humanized myeloid-only mice supported productive HIV infection and viral persistence in macrophages during ART treatment. Depletion of CD4+ T cells is considered the primary cause for terminal progression, but it was reported that increasing monocyte turnover was a significantly better predictor in SIV-infected adult macaques. Notably, pediatric cases of HIV/SIV exhibit faster and more severe disease progression than adults, yet neonates have fewer target T cells and generally lack the hallmark CD4+ T cell depletion typical of adult infections. Current data show that the baseline blood monocyte turnover rate was significantly higher in neonatal macaques compared to adults and this remained high with disease progression. In this review, we discuss recent data exploring the contribution of monocytes and macrophages to HIV/SIV infection and progression. Furthermore, we highlight the need to further investigate their role in pediatric cases of infection.

  10. Key Role of the Scavenger Receptor MARCO in Mediating Adenovirus Infection and Subsequent Innate Responses of Macrophages

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    Mareike D. Maler

    2017-08-01

    Full Text Available The scavenger receptor MARCO is expressed in several subsets of naive tissue-resident macrophages and has been shown to participate in the recognition of various bacterial pathogens. However, the role of MARCO in antiviral defense is largely unexplored. Here, we investigated whether MARCO might be involved in the innate sensing of infection with adenovirus and recombinant adenoviral vectors by macrophages, which elicit vigorous immune responses in vivo. Using cells derived from mice, we show that adenovirus infection is significantly more efficient in MARCO-positive alveolar macrophages (AMs and in AM-like primary macrophage lines (Max Planck Institute cells than in MARCO-negative bone marrow-derived macrophages. Using antibodies blocking ligand binding to MARCO, as well as gene-deficient and MARCO-transfected cells, we show that MARCO mediates the rapid adenovirus transduction of macrophages. By enhancing adenovirus infection, MARCO contributes to efficient innate virus recognition through the cytoplasmic DNA sensor cGAS. This leads to strong proinflammatory responses, including the production of interleukin-6 (IL-6, alpha/beta interferon, and mature IL-1α. These findings contribute to the understanding of viral pathogenesis in macrophages and may open new possibilities for the development of tools to influence the outcome of infection with adenovirus or adenovirus vectors.

  11. 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

  12. Tissue factor deficiency increases alveolar hemorrhage and death in influenza A virus-infected mice.

    Science.gov (United States)

    Antoniak, S; Tatsumi, K; Hisada, Y; Milner, J J; Neidich, S D; Shaver, C M; Pawlinski, R; Beck, M A; Bastarache, J A; Mackman, N

    2016-06-01

    Essentials H1N1 Influenza A virus (IAV) infection is a hemostatic challenge for the lung. Tissue factor (TF) on lung epithelial cells maintains lung hemostasis after IAV infection. Reduced TF-dependent activation of coagulation leads to alveolar hemorrhage. Anticoagulation might increase the risk for hemorrhages into the lung during severe IAV infection. Background Influenza A virus (IAV) infection is a common respiratory tract infection that causes considerable morbidity and mortality worldwide. Objective To investigate the effect of genetic deficiency of tissue factor (TF) in a mouse model of IAV infection. Methods Wild-type mice, low-TF (LTF) mice and mice with the TF gene deleted in different cell types were infected with a mouse-adapted A/Puerto Rico/8/34 H1N1 strain of IAV. TF expression was measured in the lungs, and bronchoalveolar lavage fluid (BALF) was collected to measure extracellular vesicle TF, activation of coagulation, alveolar hemorrhage, and inflammation. Results IAV infection of wild-type mice increased lung TF expression, activation of coagulation and inflammation in BALF, but also led to alveolar hemorrhage. LTF mice and mice with selective deficiency of TF in lung epithelial cells had low basal levels of TF and failed to increase TF expression after infection; these two strains of mice had more alveolar hemorrhage and death than controls. In contrast, deletion of TF in either myeloid cells or endothelial cells and hematopoietic cells did not increase alveolar hemorrhage or death after IAV infection. These results indicate that TF expression in the lung, particularly in epithelial cells, is required to maintain alveolar hemostasis after IAV infection. Conclusion Our study indicates that TF-dependent activation of coagulation is required to limit alveolar hemorrhage and death after IAV infection. © 2016 International Society on Thrombosis and Haemostasis.

  13. Regulation of major histocompatibility complex class II antigens on human alveolar macrophages by granulocyte–macrophage colony-stimulating factor in the presence of glucocorticoids

    Science.gov (United States)

    Caulfield, J J; Fernandez, M H; Sousa, A R; Lane, S J; Lee, T H; Hawrylowicz, C M

    1999-01-01

    Alveolar macrophages (AM) present antigen poorly to CD4+ T cells and respond weakly to interferon-γ (IFN-γ) for up-regulation of major histocompatibility complex (MHC) class II and costimulatory molecule expression. In atopic asthma, however, AM exhibit enhanced antigen-presenting cell (APC) activity. Since granulocyte–macrophage colony-stimulating factor (GM-CSF) is increased in the airways of asthmatic patients, we have investigated its role in modulating the APC function of AM. The effects of glucocorticoids were also studied since earlier studies showed optimal induction of MHC antigens on monocytes by GM-CSF in their presence. GM-CSF in the presence, but not the absence, of dexamethasone enhanced the expression of HLA-DR, -DP and -DQ antigens by AM. However AM and monocytes differed in the optimal concentration of steroid required to mediate this effect (10−10 m and 10−7 m, respectively). Induction of MHC antigens was glucocorticoid specific and independent of IFN-γ. These studies suggest the existence of an IFN-γ-independent pathway of macrophage activation, which may be important in regulating APC function within the lung. PMID:10469240

  14. A co-culture system with an organotypic lung slice and an immortal alveolar macrophage cell line to quantify silica-induced inflammation.

    Directory of Open Access Journals (Sweden)

    Falk Hofmann

    Full Text Available There is growing evidence that amorphous silica nanoparticles cause toxic effects on lung cells in vivo as well as in vitro and induce inflammatory processes. The phagocytosis of silica by alveolar macrophages potentiates these effects. To understand the underlying molecular mechanisms of silica toxicity, we applied a co-culture system including the immortal alveolar epithelial mouse cell line E10 and the macrophage cell line AMJ2-C11. In parallel we exposed precision-cut lung slices (lacking any blood cells as well as residual alveolar macrophages of wild type and P2rx7-/- mice with or without AMJ2-C11 cells to silica nanoparticles. Exposure of E10 cells as well as slices of wild type mice resulted in an increase of typical alveolar epithelial type 1 cell proteins like T1α, caveolin-1 and -2 and PKC-β1, whereas the co-culture with AMJ2-C11 showed mostly a slightly lesser increase of these proteins. In P2rx7-/- mice most of these proteins were slightly decreased. ELISA analysis of the supernatant of wild type and P2rx7-/- mice precision-cut lung slices showed decreased amounts of IL-6 and TNF-α when incubated with nano-silica. Our findings indicate that alveolar macrophages influence the early inflammation of the lung and also that cell damaging reagents e.g. silica have a smaller impact on P2rx7-/- mice than on wild type mice. The co-culture system with an organotypic lung slice is a useful tool to study the role of alveolar macrophages during lung injury at the organoid level.

  15. Macrophages Subvert Adaptive Immunity to Urinary Tract Infection.

    Science.gov (United States)

    Mora-Bau, Gabriela; Platt, Andrew M; van Rooijen, Nico; Randolph, Gwendalyn J; Albert, Matthew L; Ingersoll, Molly A

    2015-07-01

    Urinary tract infection (UTI) is one of the most common bacterial infections with frequent recurrence being a major medical challenge. Development of effective therapies has been impeded by the lack of knowledge of events leading to adaptive immunity. Here, we establish conclusive evidence that an adaptive immune response is generated during UTI, yet this response does not establish sterilizing immunity. To investigate the underlying deficiency, we delineated the naïve bladder immune cell compartment, identifying resident macrophages as the most populous immune cell. To evaluate their impact on the establishment of adaptive immune responses following infection, we measured bacterial clearance in mice depleted of either circulating monocytes, which give rise to macrophages, or bladder resident macrophages. Surprisingly, mice depleted of resident macrophages, prior to primary infection, exhibited a nearly 2-log reduction in bacterial burden following secondary challenge compared to untreated animals. This increased bacterial clearance, in the context of a challenge infection, was dependent on lymphocytes. Macrophages were the predominant antigen presenting cell to acquire bacteria post-infection and in their absence, bacterial uptake by dendritic cells was increased almost 2-fold. These data suggest that bacterial uptake by tissue macrophages impedes development of adaptive immune responses during UTI, revealing a novel target for enhancing host responses to bacterial infection of the bladder.

  16. Atividade antioxidante de macrófagos alveolares em ratos endotoxêmicos Anti-oxidative activity of alveolar macrophages in endotoxemic rats

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    Juliana Félix de Melo

    2010-04-01

    Full Text Available Avaliou-se o efeito da endotoxemia sobre a atividade antioxidante de macrófagos alveolares em ratos da linhagem Wistar. Foram utilizados 24 ratos machos, com idade entre 90 e 120 dias, os quais foram divididos em dois grupos: controle e endotoxêmico. O grupo endotoxêmico foi submetido à injeção intraperitonial de lipopolissacarídio na dose de 1mg/kg de peso corporal. Após 24 h, coletou-se sangue para contagem total e diferencial de leucócitos; lavado broncoalveolar para contagem total e diferencial dos leucócitos e, a partir de macrófagos isolados deste lavado, foram realizadas as dosagens de superóxido e superóxido dismutase. A endotoxemia aumentou a contagem total de leucócitos e o número de neutrófilos no sangue periférico, no lavado broncoalveolar, e aumentou a produção de superóxido sem modificar a produção da superóxido dismutase. Esses resultados sugerem que a endotoxemia induz a uma resposta inflamatória no pulmão. Contudo, não altera a atividade antioxidante em ratos adultos. Tal fato potencializa a resposta contra agentes infecciosos pelo hospedeiro, mas também pode contribuir na patogênese de injúria pulmonar.The effects of endotoxemia on the antioxidant activity in alveolar macrophages of Wistar rats were evaluated. Twenty-four male rats, 90-120 days of age, were separated into 2 groups: control and endotoxemic. To the endotoxemic animals was administered, intraperitoneally, a lipopolyssaccaride at dosage of 1mg/kg body weight. Twenty-four hours after this procedure, blood was collected for total and differential leukocytes counts. In addition, bronchoalveolar lavage was collected for total and differential leukocyte counting. From this lavage macrophages were isolated for the dosage of superoxide and superoxide dismutase. The endotoxemia increased the total leukocyte counts and the number of neutrophils in the peripheral blood and bronchoalveolar lavage of the rats. There was an increased superoxide

  17. Sex differences in the response of the alveolar macrophage proteome to treatment with exogenous surfactant protein-A

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    Phelps David S

    2012-07-01

    Full Text Available Abstract Background Male wild type (WT C57BL/6 mice are less capable of clearing bacteria and surviving from bacterial pneumonia than females. However, if an oxidative stress (acute ozone exposure occurs before infection, the advantage shifts to males who then survive at higher rates than females. We have previously demonstrated that survival in surfactant protein-A (SP-A knockout (KO mice compared to WT was significantly reduced. Because the alveolar macrophage (AM is pivotal in host defense we hypothesized that SP-A and circulating sex hormones are responsible for these sex differences. We used 2D-DIGE to examine the relationship of sex and SP-A on the AM proteome. The role of SP-A was investigated by treating SP-A KO mice with exogenous SP-A for 6 and 18 hr and studying its effects on the AM proteome. Results We found: 1 less variance between KO males and females than between the WT counterparts by principal component analysis, indicating that SP-A plays a role in sex differences; 2 fewer changes in females when the total numbers of significantly changing protein spots or identified whole proteins in WT or 18 hr SP-A-treated males or females were compared to their respective KO groups; 3 more proteins with functions related to chaperones or protease balance and Nrf2-regulated proteins changed in response to SP-A in females than in males; and 4 the overall pattern of SP-A induced changes in actin-related proteins were similar in both sexes, although males had more significant changes. Conclusions Although there seems to be an interaction between sex and the effect of SP-A, it is unclear what the responsible mechanisms are. However, we found that several of the proteins that were expressed at significantly higher levels in females than in males in WT and/or in KO mice are known to interact with the estrogen receptor and may thus play a role in the SP-A/sex interaction. These include major vault protein, chaperonin subunit 2 (beta (CCT2, and Rho

  18. Stimulation of interleukin-1 and -6 production in alveolar macrophages by the neotropical liana, Uncaria tomentosa (uña de gato)

    Science.gov (United States)

    Lemaire, I; Assinewe, V; Cano, P; Awang, D V; Arnason, J T

    1999-02-01

    Two extracts of different collections of the traditional medicine uña de gato (Uncaria tomentosa) from Peru were characterized by High Pressure Liquid Chromatography as containing approximately 6 mg/g total oxindole content prior to studies with alveolar macrophages. The plant preparations greatly stimulated IL-1 and IL-6 production by rat macrophages in a dose dependent manner in the range of 0.025-0.1 mg/ml. They were also able to enhance IL-1 and -6 in lipopolysaccharide-stimulated macrophages. The results suggest a strong immunostimulant action of this plant.

  19. 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.

  20. 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.

  1. Protective phenotypes of club cells and alveolar macrophages are favored as part of endotoxin-mediated prevention of asthma

    Science.gov (United States)

    García, Luciana N; Leimgruber, Carolina; Uribe Echevarría, Elisa M; Acosta, Patricio L; Brahamian, Jorge M; Polack, Fernando P; Miró, María S; Quintar, Amado A; Sotomayor, Claudia E

    2014-01-01

    Atopic asthma is a chronic allergic disease that involves T-helper type 2 (Th2)-inflammation and airway remodeling. Bronchiolar club cells (CC) and alveolar macrophages (AM) are sentinel cells of airway barrier against inhaled injuries, where allergy induces mucous metaplasia of CC and the alternative activation of AM, which compromise host defense mechanisms and amplify Th2-inflammation. As there is evidence that high levels of environmental endotoxin modulates asthma, the goal of this study was to evaluate if the activation of local host defenses by Lipopolysaccharide (LPS) previous to allergy development can contribute to preserving CC and AM protective phenotypes. Endotoxin stimulus before allergen exposition reduced hallmarks of allergic inflammation including eosinophil influx, Interleukin-4 and airway hyperreactivity, while the T-helper type 1 related cytokines IL-12 and Interferon-γ were enhanced. This response was accompanied by the preservation of the normal CC phenotype and the anti-allergic proteins Club Cell Secretory Protein (CCSP) and Surfactant-D, thereby leading to lower levels of CC metaplasia and preventing the increase of the pro-Th2 cytokine Thymic stromal lymphopoietin. In addition, classically activated alveolar macrophages expressing nitric oxide were promoted over the alternatively activated ones that expressed arginase-1. We verified that LPS induced a long-term overexpression of CCSP and the innate immune markers Toll-like receptor 4, and Tumor Necrosis Factor-α, changes that were preserved in spite of the allergen challenge. These results demonstrate that LPS pre-exposition modifies the local bronchioalveolar microenvironment by inducing natural anti-allergic mechanisms while reducing local factors that drive Th2 type responses, thus modulating allergic inflammation. PMID:25504013

  2. The antifibrotic effects of alveolar macrophages 5-HT2C receptors blockade on bleomycin-induced pulmonary fibrosis in rats.

    Science.gov (United States)

    Elaidy, Samah M; Essawy, Soha S

    2016-12-01

    The most widespread chronic fibrosing lung disease is idiopathic pulmonary fibrosis. Lung serotonin (5-HT) content is increased during pulmonary fibrosis with the implication of 5-HT2 receptors in the pathogenesis. Serotonin plays important roles in alveolar macrophages function through 5-HT2C receptors activation. Numerous studies described the important role of 5-HT2A/B receptor blockers in suppressing different types of fibrosis as idiopathic pulmonary fibrosis. The current study pointed to examine the antifibrotic effects of RS-102221 and/or terguride through in vivo model of pulmonary fibrosis. Induction of pulmonary fibrosis was through a single dose of intra-tracheal bleomycin instillation (5mg/kg dissolved in phosphate buffer saline) in adult male albino Wistar rats. Next day of bleomycin instillation, intraperitoneal injection of RS-102221 (0.5mg/kg/d) and/or terguride (1.2mg/kg/d) were administered and continued for 14 days. Noticeable increase in 5-HT2C receptors expression was observed in fibrotic lung tissues with marked allocation belonging to alveolar macrophages. Either RS-102221 or terguride reduced the increments in lung water contents, grading of lung fibrosis, lung tumor necrosis factor-alpha (TNF-α), transforming growth factor-beta1 (TGF- β1 ) and vascular endothelial growth factor (VEGF) levels in lung injury and fibrosis-induced by bleomycin. Moreover, collagen content and myofibroblasts-alpha smooth muscle actin (α-SМA) were significantly decreased. Additionally, the simultaneous administration of RS-102221 with terguride had a synergistic outcome compared to that obtained by individual monotherapy. These findings suggested the potential use of 5-HT2A/B/C antagonists as anti-fibrotic agents in lung fibrosis. Copyright © 2016. Published by Elsevier Urban & Partner Sp. z o.o.

  3. [Reference values of hemosiderin-laden alveolar macrophages in bronchoalveolar lavage in children].

    Science.gov (United States)

    Ahrens, P; Kitz, R; Weimer, B; Hofmann, D

    1999-05-01

    Pulmonary haemosiderosis is a rare disease of unknown etiology, mainly affecting children and adolescents. Pulmonary haemosiderosis may occur in association with several respiratory or other disease (Lupus erythematosus, Goodpastures syndrome). Delay in diagnosing can lead to fatal complications. BAL appears to be the method of choice to detect haemosiderin-laden macrophages. No reference values are available for children to date. 64 bronchoalveolar lavages were performed to establish reference values of haemosiderin-laden macrophages in children. Less haemosiderin was found in children compared to adults, and hence a new haemosiderin score was established for paediatric patients. Compared to healthy children, no elevated haemosiderine levels were found in children with chronic pulmonary disease such as bronchiectasis or chronic aspiration caused by gastro-oesophageal reflux. Therefore even a mildly increased amount of haemosiderin-laden macrophages in children requires medical attention.

  4. Astrocytes Resist HIV-1 Fusion but Engulf Infected Macrophage Material

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    Rebecca A. Russell

    2017-02-01

    Full Text Available HIV-1 disseminates to diverse tissues and establishes long-lived viral reservoirs. These reservoirs include the CNS, in which macrophage-lineage cells, and as suggested by many studies, astrocytes, may be infected. Here, we have investigated astrocyte infection by HIV-1. We confirm that astrocytes trap and internalize HIV-1 particles for subsequent release but find no evidence that these particles infect the cell. Astrocyte infection was not observed by cell-free or cell-to-cell routes using diverse approaches, including luciferase and GFP reporter viruses, fixed and live-cell fusion assays, multispectral flow cytometry, and super-resolution imaging. By contrast, we observed intimate interactions between HIV-1-infected macrophages and astrocytes leading to signals that might be mistaken for astrocyte infection using less stringent approaches. These results have implications for HIV-1 infection of the CNS, viral reservoir formation, and antiretroviral therapy.

  5. Expression of matrix metalloproteinase-1 in alveolar macrophages, type II pneumocytes, and airways in smokers: relationship to lung function and emphysema.

    Science.gov (United States)

    Wallace, Alison M; Loy, Leanna B; Abboud, Raja T; D'Armiento, Jeanine M; Coxson, Harvey O; Muller, Nestor L; Kalloger, Steve; Li, Xing; Mark Elliott, W; English, John C; Finley, Richard J; Paré, Peter D

    2014-08-01

    An imbalance between proteolytic enzymes and their inhibitors is thought to be involved in the pathogenesis of chronic obstructive pulmonary disease. Matrix metalloproteinase-1, also known as interstitial collagenase, has been implicated as a potentially important proteinase in the genesis of chronic obstructive pulmonary disease and, more specifically, emphysema. We performed quantitative immunohistochemical assessment of matrix metalloproteinase-1 expression in the resected lung of 20 smokers/ex-smokers who had varying severity of airflow obstruction and emphysema and compared this with the lungs of 5 nonsmokers. Emphysema was measured using a morphometric measure of the lungs' surface area/volume ratio and with qualitative and quantitative computed tomography (CT) measures of emphysema. There were significantly more matrix metalloproteinase-1-expressing alveolar macrophages and type II pneumocytes as well as a greater percentage of small airways that stained positively for matrix metalloproteinase-1 in the lungs of smokers than in those of nonsmokers (p lung surface area/volume ratio and to qualitative estimates of emphysema on CT. These findings suggest that cigarette smoking increases expression of matrix metalloproteinase-1 in alveolar macrophages as well as in alveolar and small airway epithelial cells. Smokers who develop emphysema have increased alveolar macrophage expression of matrix metalloproteinase-1.

  6. Innate immune response to a H3N2 subtype swine influenza virus in newborn porcine trachea cells, alveolar macrophages, and precision-cut lung slices.

    Science.gov (United States)

    Delgado-Ortega, Mario; Melo, Sandrine; Punyadarsaniya, Darsaniya; Ramé, Christelle; Olivier, Michel; Soubieux, Denis; Marc, Daniel; Simon, Gaëlle; Herrler, Georg; Berri, Mustapha; Dupont, Joëlle; Meurens, François

    2014-04-09

    Viral respiratory diseases remain of major importance in swine breeding units. Swine influenza virus (SIV) is one of the main known contributors to infectious respiratory diseases. The innate immune response to swine influenza viruses has been assessed in many previous studies. However most of these studies were carried out in a single-cell population or directly in the live animal, in all its complexity. In the current study we report the use of a trachea epithelial cell line (newborn pig trachea cells - NPTr) in comparison with alveolar macrophages and lung slices for the characterization of innate immune response to an infection by a European SIV of the H3N2 subtype. The expression pattern of transcripts involved in the recognition of the virus, interferon type I and III responses, and the host-response regulation were assessed by quantitative PCR in response to infection. Some significant differences were observed between the three systems, notably in the expression of type III interferon mRNA. Then, results show a clear induction of JAK/STAT and MAPK signaling pathways in infected NPTr cells. Conversely, PI3K/Akt signaling pathways was not activated. The inhibition of the JAK/STAT pathway clearly reduced interferon type I and III responses and the induction of SOCS1 at the transcript level in infected NPTr cells. Similarly, the inhibition of MAPK pathway reduced viral replication and interferon response. All together, these results contribute to an increased understanding of the innate immune response to H3N2 SIV and may help identify strategies to effectively control SIV infection.

  7. The Role of Alveolar Macrophage Beta-2 Adrenergic Receptors in Acute Lung Injury

    Science.gov (United States)

    2017-10-01

    which are required for pro-inflammatory response against influenza suggested by attenuation of inflammation with inhibitors of glycolysis (Hk2) and...increased expression of glycolytic metabolic genes and Irg1 and exhibited increased glycolysis (measured by Seahorse XF24). Importantly, inhibition...of glycolysis or carbonic anhydrase attenuated influenza induced IL-6 response from macrophages. These results suggested influenza induced

  8. Potential Link between the Sphingosine-1-Phosphate (S1P System and Defective Alveolar Macrophage Phagocytic Function in Chronic Obstructive Pulmonary Disease (COPD.

    Directory of Open Access Journals (Sweden)

    Jameel Barnawi

    Full Text Available We previously reported that alveolar macrophages from patients with chronic obstructive pulmonary disease (COPD are defective in their ability to phagocytose apoptotic cells, with a similar defect in response to cigarette smoke. The exact mechanisms for this defect are unknown. Sphingolipids including ceramide, sphingosine and sphingosine-1-phosphate (S1P are involved in diverse cellular processes and we hypothesised that a comprehensive analysis of this system in alveolar macrophages in COPD may help to delineate the reasons for defective phagocytic function.We compared mRNA expression of sphingosine kinases (SPHK1/2, S1P receptors (S1PR1-5 and S1P-degrading enzymes (SGPP1, SGPP2, SGPL1 in bronchoalveolar lavage-derived alveolar macrophages from 10 healthy controls, 7 healthy smokers and 20 COPD patients (10 current- and 10 ex-smokers using Real-Time PCR. Phagocytosis of apoptotic cells was investigated using flow cytometry. Functional associations were assessed between sphingosine signalling system components and alveolar macrophage phagocytic ability in COPD. To elucidate functional effects of increased S1PR5 on macrophage phagocytic ability, we performed the phagocytosis assay in the presence of varying concentrations of suramin, an antagonist of S1PR3 and S1PR5. The effects of cigarette smoking on the S1P system were investigated using a THP-1 macrophage cell line model.We found significant increases in SPHK1/2 (3.4- and 2.1-fold increases respectively, S1PR2 and 5 (4.3- and 14.6-fold increases respectively, and SGPL1 (4.5-fold increase in COPD vs. controls. S1PR5 and SGPL1 expression was unaffected by smoking status, suggesting a COPD "disease effect" rather than smoke effect per se. Significant associations were noted between S1PR5 and both lung function and phagocytosis. Cigarette smoke extract significantly increased mRNA expression of SPHK1, SPHK2, S1PR2 and S1PR5 by THP-1 macrophages, confirming the results in patient

  9. Potential Link between the Sphingosine-1-Phosphate (S1P) System and Defective Alveolar Macrophage Phagocytic Function in Chronic Obstructive Pulmonary Disease (COPD).

    Science.gov (United States)

    Barnawi, Jameel; Tran, Hai; Jersmann, Hubertus; Pitson, Stuart; Roscioli, Eugene; Hodge, Greg; Meech, Robyn; Haberberger, Rainer; Hodge, Sandra

    2015-01-01

    We previously reported that alveolar macrophages from patients with chronic obstructive pulmonary disease (COPD) are defective in their ability to phagocytose apoptotic cells, with a similar defect in response to cigarette smoke. The exact mechanisms for this defect are unknown. Sphingolipids including ceramide, sphingosine and sphingosine-1-phosphate (S1P) are involved in diverse cellular processes and we hypothesised that a comprehensive analysis of this system in alveolar macrophages in COPD may help to delineate the reasons for defective phagocytic function. We compared mRNA expression of sphingosine kinases (SPHK1/2), S1P receptors (S1PR1-5) and S1P-degrading enzymes (SGPP1, SGPP2, SGPL1) in bronchoalveolar lavage-derived alveolar macrophages from 10 healthy controls, 7 healthy smokers and 20 COPD patients (10 current- and 10 ex-smokers) using Real-Time PCR. Phagocytosis of apoptotic cells was investigated using flow cytometry. Functional associations were assessed between sphingosine signalling system components and alveolar macrophage phagocytic ability in COPD. To elucidate functional effects of increased S1PR5 on macrophage phagocytic ability, we performed the phagocytosis assay in the presence of varying concentrations of suramin, an antagonist of S1PR3 and S1PR5. The effects of cigarette smoking on the S1P system were investigated using a THP-1 macrophage cell line model. We found significant increases in SPHK1/2 (3.4- and 2.1-fold increases respectively), S1PR2 and 5 (4.3- and 14.6-fold increases respectively), and SGPL1 (4.5-fold increase) in COPD vs. controls. S1PR5 and SGPL1 expression was unaffected by smoking status, suggesting a COPD "disease effect" rather than smoke effect per se. Significant associations were noted between S1PR5 and both lung function and phagocytosis. Cigarette smoke extract significantly increased mRNA expression of SPHK1, SPHK2, S1PR2 and S1PR5 by THP-1 macrophages, confirming the results in patient-derived macrophages

  10. 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...

  11. 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

  12. Macrophage Activation by Ursolic and Oleanolic Acids during Mycobacterial Infection

    Directory of Open Access Journals (Sweden)

    Sonia López-García

    2015-08-01

    Full Text Available Oleanolic (OA and ursolic acids (UA are triterpenes that are abundant in vegetables, fruits and medicinal plants. They have been described as active moieties in medicinal plants used for the treatment of tuberculosis. In this study, we analyzed the effects of these triterpenes on macrophages infected in vitro with Mycobacterium tuberculosis (MTB. We evaluated production of nitric oxide (NO, reactive oxygen species (ROS, and cytokines (TNF-α and TGF-β as well as expression of cell membrane receptors (TGR5 and CD36 in MTB-infected macrophages following treatment with OA and UA. Triterpenes caused reduced MTB growth in macrophages, stimulated production of NO and ROS in the early phase, stimulated TNF-α, suppressed TGF-β and caused over-expression of CD36and TGR5 receptors. Thus, our data suggest immunomodulatory properties of OA and UA on MTB infected macrophages. In conclusion, antimycobacterial effects induced by these triterpenes may be attributable to the conversion of macrophages from stage M2 (alternatively activated to M1 (classically activated.

  13. 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.

  14. Toll-like receptor 2 expression is decreased on alveolar macrophages in cigarette smokers and COPD patients

    Directory of Open Access Journals (Sweden)

    Zabel Peter

    2005-07-01

    Full Text Available Abstract Backround Cigarette smoke exposure including biologically active lipopolysaccharide (LPS in the particulate phase of cigarette smoke induces activation of alveolar macrophages (AM and alveolar epithelial cells leading to production of inflammatory mediators. This represents a crucial mechanism in the pathogenesis of chronic obstructive pulmonary disease (COPD. Respiratory pathogens are a major cause of exacerbations leading to recurrent cycles of injury and repair. The interaction between pathogen-associated molecular patterns and the host is mediated by pattern recognition receptors (PRR's. In the present study we characterized the expression of Toll-like receptor (TLR- 2, TLR4 and CD14 on human AM compared to autologous monocytes obtained from patients with COPD, healthy smokers and non-smokers. Methods The study population consisted of 14 COPD patients without evidence for acute exacerbation, 10 healthy smokers and 17 healthy non-smokers stratified according to age. The expression of TLR2, TLR4 and CD14 surface molecules on human AM compared to autologous monocytes was assessed ex vivo using FACS analysis. In situ hybridization was performed on bronchoalveolar lavage (BAL cells by application of the new developed HOPE-fixative. Results The expression of TLR2, TLR4 and CD14 on AM from COPD patients, smokers and non-smokers was reduced as compared to autologous monocytes. Comparing AM we detected a reduced expression of TLR2 in COPD patients and smokers. In addition TLR2 mRNA and protein expression was increased after LPS stimulation on non-smokers AM in contrast to smokers and COPD patients. Conclusion Our data suggest a smoke related change in the phenotype of AM's and the cellular response to microbial stimulation which may be associated with impairment of host defenses in the lower respiratory tract.

  15. An in vitro alveolar macrophage assay for the assessment of inflammatory cytokine expression induced by atmospheric particulate matter.

    Science.gov (United States)

    Sijan, Zana; Antkiewicz, Dagmara S; Heo, Jongbae; Kado, Norman Y; Schauer, James J; Sioutas, Constantinos; Shafer, Martin M

    2015-07-01

    Exposures to air pollution in the form of particulate matter (PM) can result in excess production of reactive oxygen species (ROS) in the respiratory system, potentially causing both localized cellular injury and triggering a systemic inflammatory response. PM-induced inflammation in the lung is modulated in large part by alveolar macrophages and their biochemical signaling, including production of inflammatory cytokines, the primary mechanism via which inflammation is initiated and sustained. We developed a robust, relevant, and flexible method employing a rat alveolar macrophage cell line (NR8383) which can be applied to routine samples of PM from air quality monitoring sites to gain insight into the drivers of PM toxicity that lead to oxidative stress and inflammation. Method performance was characterized using extracts of ambient and vehicular engine exhaust PM samples. Our results indicate that the reproducibility and the sensitivity of the method are satisfactory and comparisons between PM samples can be made with good precision. The average relative percent difference for all genes detected during 10 different exposures was 17.1%. Our analysis demonstrated that 71% of genes had an average signal to noise ratio (SNR) ≥ 3. Our time course study suggests that 4 h may be an optimal in vitro exposure time for observing short-term effects of PM and capturing the initial steps of inflammatory signaling. The 4 h exposure resulted in the detection of 57 genes (out of 84 total), of which 86% had altered expression. Similarities and conserved gene signaling regulation among the PM samples were demonstrated through hierarchical clustering and other analyses. Overlying the core congruent patterns were differentially regulated genes that resulted in distinct sample-specific gene expression "fingerprints." Consistent upregulation of Il1f5 and downregulation of Ccr7 was observed across all samples, while TNFα was upregulated in half of the samples and downregulated in

  16. 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)

  17. 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

    . A fraction of the AMFs proliferated locally, demonstrating that at least a subset of human AMFs have the capacity to self-renew. Lungs of humanised mice were found to abundantly contain populations of human AMFs expressing markers compatible with a monocyte origin. Moreover, in patients with lung....... CONCLUSIONS: The finding that human AMFs are maintained in the lung parenchyma for several years indicates that pulmonary macrophage transplantation can be a feasible therapeutic option for patients with diseases caused by dysfunctional AMFs. Moreover, in a lung transplantation setting, long-term persistence...

  18. Characteristics of Suppressor Macrophages Induced by Mycobacterial and Protozoal Infections in relation to Alternatively Activated M2 Macrophages

    Directory of Open Access Journals (Sweden)

    Haruaki Tomioka

    2012-01-01

    Full Text Available In the advanced stages of mycobacterial infections, host immune systems tend to change from a Th1-type to Th2-type immune response, resulting in the abrogation of Th1 cell- and macrophage-mediated antimicrobial host protective immunity. Notably, this type of immune conversion is occasionally associated with the generation of certain types of suppressor macrophage populations. During the course of Mycobacterium tuberculosis (MTB and Mycobacterium avium-intracellulare complex (MAC infections, the generation of macrophages which possess strong suppressor activity against host T- and B-cell functions is frequently encountered. This paper describes the immunological properties of M1- and M2-type macrophages generated in tumor-bearing animals and those generated in hosts with certain microbial infections. In addition, this paper highlights the immunological and molecular biological characteristics of suppressor macrophages generated in hosts with mycobacterial infections, especially MAC infection.

  19. 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

    .... Although several factors contribute to inhalational anthrax, we hypothesized that unimpeded infection of Bacillus anthracis is directly linked to disabling the innate immune functions contributed by AM...

  20. Silica Induces Changes in Cytosolic Free Calcium, Cytosolic pH, and Plasma Membrane Potential in Bovine Alveolar Macrophages

    Directory of Open Access Journals (Sweden)

    Attila Tárnok

    1997-01-01

    Full Text Available The mineral‐dust induced activation of pulmonary phagocytes is thought to be involved in the induction of severe lung diseases. The activation of bovine alveolar macrophages (BAM by silica was investigated by flow cytometry. Short‐term incubation (10 min of BAM with silica gel and quartz dust particles induced increases in the cytosolic free calcium concentration ([Ca2+]i, decreases in intracellular pH (pHi, and increases in plasma membrane potential (PMP. The extent of these changes was concentration dependent, related to the type of dust and was due to Ca2+ influx from the extracellular medium. An increase in [Ca2+]i was inhibited, when extracellular Ca2+ was removed. Furthermore the calcium signal was quenched by Mn2+ and diminished by the calcium channel blocker verapamil. The protein kinase C specific inhibitor bisindolylmaleimide II (GF 109203 X did not inhibit the silica‐induced [Ca2+]i rise. In contrast, silica‐induced cytosolic acidification and depolarization were inhibited by GF 109203 X but not by removal of extracellular calcium. Addition of TiO2 particles or heavy metal‐containing dusts had no effect on any of the three parameters. Our data suggest the existence of silica‐activated transmembrane ion exchange mechanisms in BAM, which might be involved in the specific cytotoxicity of silica by Ca2+‐dependent and independent pathways.

  1. Low pH Environmental Stress Inhibits LPS and LTA-Stimulated Proinflammatory Cytokine Production in Rat Alveolar Macrophages

    Directory of Open Access Journals (Sweden)

    Stanley F. Fernandez

    2013-01-01

    Full Text Available Gastric aspiration increases the risks for developing secondary bacterial pneumonia. Cytokine elaboration through pathogen recognition receptors (PRRs is an important mechanism in initiating innate immune host response. Effects of low pH stress, a critical component of aspiration pathogenesis, on the PRR pathways were examined, specifically toll-like receptor-2 (TLR2 and TLR4, using isolated rat alveolar macrophages (aMØs. We assessed the ability of aMØs after brief exposure to acidified saline to elaborate proinflammatory cytokines in response to lipopolysaccharide (LPS and lipoteichoic acid (LTA stimulation, known ligands of TLR4 and TLR2, respectively. Low pH stress reduced LPS- and LTA-mediated cytokine release (CINC-1, MIP-2, TNF-, MCP-1, and IFN-. LPS and LTA increased intracellular Ca2+ concentrations while Ca2+ chelation by BAPTA decreased LPS- and LTA-mediated cytokine responses. BAPTA blocked the effects of low pH stress on most of LPS-stimulated cytokines but not of LTA-stimulated responses. In vivo mouse model demonstrates suppressed E. coli and S. pneumoniae clearance following acid aspiration. In conclusion, low pH stress inhibits antibacterial cytokine response of aMØs due to impaired TLR2 (MyD88 pathway and TLR4 signaling (MyD88 and TRIF pathways. The role of Ca2+ in low pH stress-induced signaling is complex but appears to be distinct between LPS- and LTA-mediated responses.

  2. 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.

  3. Macrophage depletion prior to Neospora caninum infection results in severe neosporosis in mice.

    Science.gov (United States)

    Abe, Chisa; Tanaka, Sachi; Ihara, Fumiaki; Nishikawa, Yoshifumi

    2014-08-01

    We observed that murine macrophages showed greater activation and increased interleukin 6 (IL-6), IL-12p40, and interferon gamma (IFN-γ) production during Neospora caninum infection. Many macrophages migrated to the site of infection. Furthermore, macrophage-depleted mice exhibited increased sensitivity to N. caninum infection. This study indicates that macrophages are required for achieving protective immunity against N. caninum. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  4. Induction of Programmed Cell Death in Human Alveolar Epithelial Cells Infected with Influenza Virus

    Directory of Open Access Journals (Sweden)

    Sh Shahsavandi

    2015-11-01

    Full Text Available Introduction: Avian influenza viruses are considered as a serious threat to human and animal health. An increase in expression of proinflammatory cytokines and type I IFN genes, as well as host cell death responses contribute to the pathogenesis of influenza infection. Hence, this study aimed to evaluate the growth dynamics of subacute avian influenza virus in human respiratory alveolar epithelium cells (A549. Methods: The A549 cell cultures were infected at MOIs 0.1 and 2.0 viral doses in the presence and absence of trypsin. The virus growth kinetics were elucidated by the plaque assay and the cell viability was determined by MTT at various times after the infection. The induction quality of programmed cell death as well as the signal transduction pathway of death were assessed by genomic DNA fragmentation and western blotting respectively. Results: The study findings indicated that although the H9N2 virus replication did produce a marked cytopathic effect on the alveolar cells, which led to a reduction in the cell viability, the viral titers were increased in the infected cells. The virus replication of in these cells indicated repression of host defense mechanism as well as activation of cell death. The induction of apoptosis in A549 cells was correlated with the increased virus titers as well as virus replication (p< 0.05. Conclusion: H9N2 avian influenza virus were demonstrated to induce apoptosis in human alveolar epithelial cells via the intrinsic pathway in a dose-dependent manner.

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

    Directory of Open Access Journals (Sweden)

    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.

  6. 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.

  7. Comparison of plasma, epithelial lining fluid, and alveolar macrophage concentrations of solithromycin (CEM-101) in healthy adult subjects.

    Science.gov (United States)

    Rodvold, Keith A; Gotfried, Mark H; Still, J Gordon; Clark, Kay; Fernandes, Prabhavathi

    2012-10-01

    The steady-state concentrations of solithromycin in plasma were compared with concomitant concentrations in epithelial lining fluid (ELF) and alveolar macrophages (AM) obtained from intrapulmonary samples during bronchoscopy and bronchoalveolar lavage (BAL) in 30 healthy adult subjects. Subjects received oral solithromycin at 400 mg once daily for five consecutive days. Bronchoscopy and BAL were carried out once in each subject at either 3, 6, 9, 12, or 24 h after the last administered dose of solithromycin. Drug concentrations in plasma, ELF, and AM were assayed by a high-performance liquid chromatography-tandem mass spectrometry method. Solithromycin was concentrated extensively in ELF (range of mean [± standard deviation] concentrations, 1.02 ± 0.83 to 7.58 ± 6.69 mg/liter) and AM (25.9 ± 20.3 to 101.7 ± 52.6 mg/liter) in comparison with simultaneous plasma concentrations (0.086 ± 0.070 to 0.730 ± 0.692 mg/liter). The values for the area under the concentration-time curve from 0 to 24 h (AUC(0-24) values) based on mean and median ELF concentrations were 80.3 and 63.2 mg · h/liter, respectively. The ratio of ELF to plasma concentrations based on the mean and median AUC(0-24) values were 10.3 and 10.0, respectively. The AUC(0-24) values based on mean and median concentrations in AM were 1,498 and 1,282 mg · h/L, respectively. The ratio of AM to plasma concentrations based on the mean and median AUC(0-24) values were 193 and 202, respectively. Once-daily oral dosing of solithromycin at 400 mg produced steady-state concentrations that were significantly (P solithromycin administration.

  8. Human alveolar macrophage responses to air pollution particulates are associated with insoluble components of coarse material, including particulate endotoxin.

    Science.gov (United States)

    Soukup, J M; Becker, S

    2001-02-15

    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 function others inhibit antimicrobial activity or cause cytokine and growth factor production leading to inflammation and tissue remodeling. This study has investigated the effects of water soluble (s) and insoluble (is) components of Chapel Hill, North Carolina ambient particulate matter in the size ranges 0.1-2.5 microm (PM2.5) and 2.5-10 microm (PM10) diameter, on human AM IL-6, TNFalpha, and MCP-1 cytokine production and host defense mechanisms including phagocytosis and oxidant production. Cytokines were found to be induced by isPM10 to a much higher extent (>50-fold) than sPM10, which in turn stimulated production better than isPM2.5, while sPM2.5 was inactive. Previous studies have indicated that endotoxin (ETOX) is a component of sPM10 responsible for cytokine production. Here, it is shown that inhibition of isPM10-induced cytokine production was partially achieved with polymyxin B and LPS-binding protein (LBP), but not with a metal chelator, implicating ETOX as a cytokine-inducing moiety also in isPM10. In addition to inducing cytokines, exposure to isPM10, but not the other PM fractions, also inhibited phagocytosis and oxidant generation in response to yeast. This inhibition was ETOX independent. The decrease in host defenses may be the result of apoptosis in the AM population, which was also found to be specifically caused by isPM10. These results show that the functional capacity of AM is selectively modulated by insoluble components of coarse PM, including the biocontaminant ETOX.

  9. Macrophage activation and HIV infection: can the Trojan horse turn into a fortress?

    Science.gov (United States)

    Herbein, G; Coaquette, A; Perez-Bercoff, D; Pancino, G

    2002-12-01

    Macrophages are infected early during HIV infection and are thought to play the role of a Trojan horse by spreading infection in tissues. Most recent studies point out to a more complex role for macrophages in HIV infection: macrophages could contribute to both host defense and viral persistence and pathogenesis. Infected macrophages are a reservoir for HIV and modulate apoptosis of T cells present in their vicinity. Also, a functional impairment of HIV-infected macrophages may play a role in AIDS pathogenesis. Nevertheless, both activation and differentiation of monocyte/macrophages can interfere with susceptibility of these cells to infection. Therefore, a wide variety of stimuli result in HIV suppression through macrophage activation. At present times, a dynamic view on the role of macrophages in HIV infection arises which indicates that macrophages are a target for the virus and at the same time regulate its replication. Therefore, macrophages are at the cross-road between protection and pathogenesis in HIV infection due to their involvement both as a viral target and a key modulator of non-specific and specific immune responses. Future studies will help unravel the cellular and molecular mechanisms that underlie HIV-macrophage interactions and might result in new vaccine and/or therapeutic strategies.

  10. In Vitro Study of Mutagenesis Induced by Crocidolite-Exposed Alveolar Macrophages NR8383 in Cocultured Big Blue Rat2 Embryonic Fibroblasts

    International Nuclear Information System (INIS)

    Guichard, Y.; Gate, L.; Darne, C.; Bottin, M.C.; Langlais, C.

    2010-01-01

    Asbestos-induced mutagenicity in the lung may involve reactive oxygen/nitrogen species (ROS/RNS) released by alveolar macrophages. With the aim of proposing an alternative in vitro mutagenesis test, a co culture system of rat alveolar macrophages (NR8383) and transgenic Big Blue Rat 2 embryonic fibroblasts was developed and tested with a crocidolite sample. Crocidolite exposure induced no detectable increase in ROS production from NR8383, contrasting with the oxidative burst that occurred following a brief exposure (1 hour) to zymosan, a known macrophage activator. In separated co cultures, crocidolite and zymosan induced different changes in the gene expressions involved in cellular inflammation in NR8383 and Big Blue. In particular, both particles induced up-regulation of iNOS expression in Big Blue, suggesting the formation of potentially genotoxic nitrogen species. However, crocidolite exposure in separated or mixed co cultures induced no mutagenic effects whereas an increase in Big Blue mutants was detected after exposure to zymosan in mixed co cultures. NR8383 activation by crocidolite is probably insufficient to induce in vitro mutagenic events. The mutagenesis assay based on the co culture of NR8383 and Big Blue cannot be used as an alternative in vitro method to assess the mutagenic properties of asbestos fibres.

  11. Response of a phagocyte cell system to products of macrophage breakdown as a probable mechanism of alveolar phagocytosis adaptation to deposition of particles of different cytotoxicity.

    Science.gov (United States)

    Privalova, L I; Katsnelson, B A; Osipenko, A B; Yushkov, B N; Babushkina, L G

    1980-04-01

    The adaptation of the alveolar phagocytosis response to the quantitative and qualitative features of dust deposited during inhalation consists not only in enhanced recruitment of alveolar macrophages (AM), but also in adding a more or less pronounced neutrophil leukocyte (NL) recruitment as an auxiliary participant of particle clearance. The NL contribution to clearance is especially typical for response to cytotoxic particles (quartz, in particular). An important feature of the adaptation considered is the limitation of the number of AM and NL recruited when an efficient clearance can be achieved by a lesser number of cells due to increased AM reistance to the damaging actin of phagocytized particles. The main mechanism providing the adequacy of the alveolar phagocytosis response is its self-regulation thrugh the products of macrophage breakdown (PMB). In a series of experiments with intraperitoneal and intratracheal injections of syngenetic PMB into rats and mice, it was shown that these products stimulate respiration and migration of phagocytic cells, their dose-dependent attraction to the site of PMB formation with the predominant NL contribution, increasing with the increase of amount of PMB, the AM and NL precursor cells recruitment from reserve pools, and the replenishment of these reserves in the process of hemopoiesis. At least some of the above effects are connected with the action of the lipid components of PMB. The action of specialized regulative systems of the organism can modify the response to PMB, judging by the results obtained by hydrocortisone injection. Autocontrol of alveolar phagocytosis requires great care in attempts at artificial stimulation of this process, as an excessive cell recruitment may promote the retention of particles in lungs.

  12. Aspirin inhibits lipopolysaccharide-induced COX-2 expression and PGE2 production in porcine alveolar macrophages by modulating protein kinase C and protein tyrosine phosphatase activity

    OpenAIRE

    Duan, Yuzhong; Chen, Fanglin; Zhang, Anmei; Zhu, Bo; Sun, Jianguo; Xie, Qichao; Chen, Zhengtang

    2014-01-01

    Aspirin has been demonstrated to be effective in inhibiting COX-2 and PGE2 in Alveolar macrophages (AMs). However, the mechanisms have not been fully understood. In the present study, we found that pretreatment with aspirin inhibited LPS-induced COX-2 and PGE2 upregulation, IκBα degradation, NFκB activation and the increase of PKC activity, but elevated LPS-induced the decrease of PTP activity. The PKC inhibitor calphostin C dramatically reduced the COX-2 mRNA and PGE2 levels, but the PTP inh...

  13. Studies on the biological behaviour of uranium-plutonium mixed oxide aerosols: Inhalation experiments with rats and in vitro studies with alveolar macrophages

    International Nuclear Information System (INIS)

    Mueller, H.L.; Drosselmeyer, E.; Hotz, G.; Seidel, A.; Thiele, H.; Pickering, S.; Ray, I.C.

    1987-02-01

    The retention of spherical and of irregularly shaped (U, Pu) mixed oxides in rat lung was analyzed after inhalation and intratracheal instillation. Their biological behaviour was relatively independent of particle shape and application route with only a few percent of radioactivity being transferred to other organs. In vivo and in vitro uptake and intracellular distribution in rat and bovine alveolar macrophages were analyzed as dependent on various parameters. In addition, detailed electron microscopic studies were performed demonstrating particles within membrane limited vacuoles as well as lying free in the cytoplasm. Under in vitro conditions the uptake process was finished after a few hours. After differential centrifugation of lung or macrophage homogenates the particles sedimented in the first (1000 g) fraction. (orig.) [de

  14. 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

  15. 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.

  16. Synthetic Macromolecular Antibiotic Platform for Inhalable Therapy against Aerosolized Intracellular Alveolar Infections.

    Science.gov (United States)

    Das, Debobrato; Chen, Jasmin; Srinivasan, Selvi; Kelly, Abby M; Lee, Brian; Son, Hye-Nam; Radella, Frank; West, T Eoin; Ratner, Daniel M; Convertine, Anthony J; Skerrett, Shawn J; Stayton, Patrick S

    2017-06-05

    Lung-based intracellular bacterial infections remain one of the most challenging infectious disease settings. For example, the current standard for treating Franciscella tularensis pneumonia (tularemia) relies on administration of oral or intravenous antibiotics that poorly achieve and sustain pulmonary drug bioavailability. Inhalable antibiotic formulations are approved and in clinical development for upper respiratory infections, but sustained drug dosing from inhaled antibiotics against alveolar intracellular infections remains a current unmet need. To provide an extended therapy against alveolar intracellular infections, we have developed a macromolecular therapeutic platform that provides sustained local delivery of ciprofloxacin with controlled dosing profiles. Synthesized using RAFT polymerization, these macromolecular prodrugs characteristically have high drug loading (16-17 wt % drug), tunable hydrolysis kinetics mediated by drug linkage chemistry (slow-releasing alkyllic vs fast-releasing phenolic esters), and, in general, represent new fully synthetic nanotherapeutics with streamlined manufacturing profiles. In aerosolized and completely lethal F.t. novicida mouse challenge models, the fast-releasing ciprofloxacin macromolecular prodrug provided high cure efficiencies (75% survival rate under therapeutic treatment), and the importance of release kinetics was demonstrated by the inactivity of the similar but slow-releasing prodrug system. Pharmacokinetics and biodistribution studies further demonstrated that the efficacious fast-releasing prodrug retained drug dosing in the lung above the MIC over a 48 h period with corresponding C max /MIC and AUC 0-24h /MIC ratios being greater than 10 and 125, respectively; the thresholds for optimal bactericidal efficacy. These findings identify the macromolecular prodrug platform as a potential therapeutic system to better treat alveolar intracellular infections such as F. tularensis, where positive patient outcomes

  17. Brucella infection inhibits macrophages apoptosis via Nedd4-dependent degradation of calpain2.

    Science.gov (United States)

    Cui, Guimei; Wei, Pan; Zhao, Yuxi; Guan, Zhenhong; Yang, Li; Sun, Wanchun; Wang, Shuangxi; Peng, Qisheng

    2014-11-07

    The calcium-dependent protease calpain2 is involved in macrophages apoptosis. Brucella infection-induced up-regulation of intracellular calcium level is an essential factor for the intracellular survival of Brucella within macrophages. Here, we hypothesize that calcium-dependent E3 ubiquitin ligase Nedd4 ubiquitinates calpain2 and inhibits Brucella infection-induced macrophage apoptosis via degradation of calpain2.Our results reveal that Brucella infection induces increases in Nedd4 activity in an intracellular calcium dependent manner. Furthermore, Brucella infection-induced degradation of calpain2 is mediated by Nedd4 ubiquitination of calpain2. Brucella infection-induced calpain2 degradation inhibited macrophages apoptosis. Treatment of Brucella infected macrophages with calcium chelator BAPTA or Nedd4 knock-down decreased Nedd4 activity, prevented calpain2 degradation, and resulted in macrophages apoptosis. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Macrophage origin limits functional plasticity in helminth-bacterial co-infection.

    Directory of Open Access Journals (Sweden)

    Dominik Rückerl

    2017-03-01

    Full Text Available Rapid reprogramming of the macrophage activation phenotype is considered important in the defense against consecutive infection with diverse infectious agents. However, in the setting of persistent, chronic infection the functional importance of macrophage-intrinsic adaptation to changing environments vs. recruitment of new macrophages remains unclear. Here we show that resident peritoneal macrophages expanded by infection with the nematode Heligmosomoides polygyrus bakeri altered their activation phenotype in response to infection with Salmonella enterica ser. Typhimurium in vitro and in vivo. The nematode-expanded resident F4/80high macrophages efficiently upregulated bacterial induced effector molecules (e.g. MHC-II, NOS2 similarly to newly recruited monocyte-derived macrophages. Nonetheless, recruitment of blood monocyte-derived macrophages to Salmonella infection occurred with equal magnitude in co-infected animals and caused displacement of the nematode-expanded, tissue resident-derived macrophages from the peritoneal cavity. Global gene expression analysis revealed that although nematode-expanded resident F4/80high macrophages made an anti-bacterial response, this was muted as compared to newly recruited F4/80low macrophages. However, the F4/80high macrophages adopted unique functional characteristics that included enhanced neutrophil-stimulating chemokine production. Thus, our data provide important evidence that plastic adaptation of MΦ activation does occur in vivo, but that cellular plasticity is outweighed by functional capabilities specific to the tissue origin of the cell.

  19. B cell, CD8 + T cell and gamma delta T cell infiltration alters alveolar immune cell homeostasis in HIV-infected Malawian adults [version 2; referees: 1 approved, 2 approved with reservations

    Directory of Open Access Journals (Sweden)

    Andrew Mwale

    2017-12-01

    Full Text Available Background: HIV infection is associated with increased risk to lower respiratory tract infections (LRTI. However, the impact of HIV infection on immune cell populations in the lung is not well defined. We sought to comprehensively characterise the impact of HIV infection on immune cell populations in the lung. Methods: Twenty HIV-uninfected controls and 17 HIV-1 infected ART-naïve adults were recruited from Queen Elizabeth Central Hospital, Malawi. Immunophenotyping of lymphocyte and myeloid cell populations was done on bronchoalveolar lavage fluid and peripheral blood cells. Results: We found that the numbers of CD8 + T cells, B cells and gamma delta T cells were higher in BAL fluid of HIV-infected adults compared to HIV-uninfected controls (all p<0.05. In contrast, there was no difference in the numbers of alveolar CD4 + T cells in HIV-infected adults compared to HIV-uninfected controls (p=0.7065. Intermediate monocytes were the predominant monocyte subset in BAL fluid (HIV-, 63%; HIV+ 81%, while the numbers of classical monocytes was lower in HIV-infected individuals compared to HIV-uninfected adults (1 × 10 5 vs. 2.8 × 10 5 cells/100ml of BAL fluid, p=0.0001. The proportions of alveolar macrophages and myeloid dendritic cells was lower in HIV-infected adults compared to HIV-uninfected controls (all p<0.05. Conclusions: Chronic HIV infection is associated with broad alteration of immune cell populations in the lung, but does not lead to massive depletion of alveolar CD4 + T cells. Disruption of alveolar immune cell homeostasis likely explains in part the susceptibility for LRTIs in HIV-infected adults.

  20. Concurrent infection with porcine reproductive and respiratory syndrome virus and Haemophilus parasuis in two types of porcine macrophages: apoptosis, production of ROS and formation of multinucleated giant cells.

    Science.gov (United States)

    Kavanová, Lenka; Matiašková, Katarína; Levá, Lenka; Štěpánová, Hana; Nedbalcová, Kateřina; Matiašovic, Ján; Faldyna, Martin; Salát, Jiří

    2017-05-04

    Porcine reproductive and respiratory syndrome (PRRS) is one of the most significant and economically important infectious diseases affecting swine worldwide and can predispose pigs to secondary bacterial infections caused by, e.g. Haemophilus parasuis. The aim of the presented study was to compare susceptibility of two different types of macrophages which could be in contact with both pathogens during infection with PRRS virus (PRRSV) and in co-infection with H. parasuis. Alveolar macrophages (PAMs) as resident cells provide one of the first lines of defence against microbes invading lung tissue. On the other hand, monocyte derived macrophages (MDMs) represent inflammatory cells accumulating at the site of inflammation. While PAMs were relatively resistant to cytopathogenic effect caused by PRRSV, MDMs were much more sensitive to PRRSV infection. MDMs infected with PRRSV increased expression of pro-apoptotic Bad, Bax and p53 mRNA. Increased mortality of MDMs may be also related to a higher intensity of ROS production after infection with PRRSV. In addition, MDMs (but not PAMs) infected with H. parasuis alone formed multinucleated giant cells (MGC); these cells were not observed in MDMs infected with both pathogens. Higher sensitivity of MDMs to PRRSV infection, which is associated with limited MDMs survival and restriction of MGC formation, could contribute to the development of multifactorial respiratory disease of swine.

  1. Critical Role of Airway Macrophages in Modulating Disease Severity during Influenza Virus Infection of Mice ▿

    Science.gov (United States)

    Tate, Michelle D.; Pickett, Danielle L.; van Rooijen, Nico; Brooks, Andrew G.; Reading, Patrick C.

    2010-01-01

    Airway macrophages provide a first line of host defense against a range of airborne pathogens, including influenza virus. In this study, we show that influenza viruses differ markedly in their abilities to infect murine macrophages in vitro and that infection of macrophages is nonproductive and no infectious virus is released. Virus strain BJx109 (H3N2) infected macrophages with high efficiency and was associated with mild disease following intranasal infection of mice. In contrast, virus strain PR8 (H1N1) was poor in its ability to infect macrophages and highly virulent for mice. Depletion of airway macrophages by clodronate-loaded liposomes led to the development of severe viral pneumonia in BJx109-infected mice but did not modulate disease severity in PR8-infected mice. The severe disease observed in macrophage-depleted mice infected with BJx109 was associated with exacerbated virus replication in the airways, leading to severe airway inflammation, pulmonary edema, and vascular leakage, indicative of lung injury. Thymic atrophy, lymphopenia, and dysregulated cytokine and chemokine production were additional systemic manifestations associated with severe disease. Thus, airway macrophages play a critical role in limiting lung injury and associated disease caused by BJx109. Furthermore, the inability of PR8 to infect airway macrophages may be a critical factor contributing to its virulence for mice. PMID:20504924

  2. 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.

  3. Down regulation of macrophage IFNGR1 exacerbates systemic L. monocytogenes infection.

    Directory of Open Access Journals (Sweden)

    Emily M Eshleman

    2017-05-01

    Full Text Available Interferons (IFNs target macrophages to regulate inflammation and resistance to microbial infections. The type II IFN (IFNγ acts on a cell surface receptor (IFNGR to promote gene expression that enhance macrophage inflammatory and anti-microbial activity. Type I IFNs can dampen macrophage responsiveness to IFNγ and are associated with increased susceptibility to numerous bacterial infections. The precise mechanisms responsible for these effects remain unclear. Type I IFNs silence macrophage ifngr1 transcription and thus reduce cell surface expression of IFNGR1. To test how these events might impact macrophage activation and host resistance during bacterial infection, we developed transgenic mice that express a functional FLAG-tagged IFNGR1 (fGR1 driven by a macrophage-specific promoter. Macrophages from fGR1 mice expressed physiologic levels of cell surface IFNGR1 at steady state and responded equivalently to WT C57Bl/6 macrophages when treated with IFNγ alone. However, fGR1 macrophages retained cell surface IFNGR1 and showed enhanced responsiveness to IFNγ in the presence of type I IFNs. When fGR1 mice were infected with the bacterium Listeria monocytogenes their resistance was significantly increased, despite normal type I and II IFN production. Enhanced resistance was dependent on IFNγ and associated with increased macrophage activation and antimicrobial function. These results argue that down regulation of myeloid cell IFNGR1 is an important mechanism by which type I IFNs suppress inflammatory and anti-bacterial functions of macrophages.

  4. 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.

  5. 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

  6. 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

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

    LENUS (Irish Health Repository)

    Lawlor, Ciaran

    2016-01-01

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

  8. Cigarette smoke extract-treated mast cells promote alveolar macrophage infiltration and polarization in experimental chronic obstructive pulmonary disease.

    Science.gov (United States)

    Li, Hong; Yang, Tian; Ning, Qian; Li, Feiyan; Chen, Tianjun; Yao, Yan; Sun, Zhongmin

    2015-01-01

    Cigarette smoking is the main cause of chronic obstructive pulmonary disease (COPD) and may modulate the immune response of exposed individuals. Mast cell function can be altered by cigarette smoking, but the role of smoking in COPD remains poorly understood. The current study aimed to explore the role of cigarette smoke extract (CSE)-treated mast cells in COPD pathogenesis. Cytokine and chemokine expression as well as degranulation of bone marrow-derived mast cells (BMMCs) were detected in cells exposed to immunoglobulin E (IgE) and various doses of CSE. Adoptive transfer of CSE-treated BMMCs into C57BL/6J mice was performed, and macrophage infiltration and polarization were evaluated by fluorescence-activated cell sorting (FACS). Furthermore, a coculture system of BMMCs and macrophages was established to examine macrophage phenotype transition. The role of protease serine member S31 (Prss31) was also investigated in the co-culture system and in COPD mice. CSE exposure suppressed cytokine expression and degranulation in BMMCs, but promoted the expressions of chemokines and Prss31. Adoptive transfer of CSE-treated BMMCs induced macrophage infiltration and M2 polarization in the mouse lung. Moreover, CSE-treated BMMCs triggered macrophage M2 polarization via Prss31 secretion. Recombinant Prss31 was shown to activate interleukin (IL)-13/IL-13Rα/Signal transducers and activators of transcription (Stat) 6 signaling in macrophages. Additionally, a positive correlation was found between Prss31 expression and the number of M2 macrophages in COPD mice. In conclusion, CSE-treated mast cells may induce macrophage infiltration and M2 polarization via Prss31 expression, and potentially contribute to COPD progression.

  9. Changes to cholesterol trafficking in macrophages by Leishmania parasites infection.

    Science.gov (United States)

    Semini, Geo; Paape, Daniel; Paterou, Athina; Schroeder, Juliane; Barrios-Llerena, Martin; Aebischer, Toni

    2017-08-01

    Leishmania spp. are protozoan parasites that are transmitted by sandfly vectors during blood sucking to vertebrate hosts and cause a spectrum of diseases called leishmaniases. It has been demonstrated that host cholesterol plays an important role during Leishmania infection. Nevertheless, little is known about the intracellular distribution of this lipid early after internalization of the parasite. Here, pulse-chase experiments with radiolabeled cholesteryl esterified to fatty acids bound to low-density lipoproteins indicated that retention of this source of cholesterol is increased in parasite-containing subcellular fractions, while uptake is unaffected. This is correlated with a reduction or absence of detectable NPC1 (Niemann-Pick disease, type C1), a protein responsible for cholesterol efflux from endocytic compartments, in the Leishmania mexicana habitat and infected cells. Filipin staining revealed a halo around parasites within parasitophorous vacuoles (PV) likely representing free cholesterol accumulation. Labeling of host cell membranous cholesterol by fluorescent cholesterol species before infection revealed that this pool is also trafficked to the PV but becomes incorporated into the parasites' membranes and seems not to contribute to the halo detected by filipin. This cholesterol sequestration happened early after infection and was functionally significant as it correlated with the upregulation of mRNA-encoding proteins required for cholesterol biosynthesis. Thus, sequestration of cholesterol by Leishmania amastigotes early after infection provides a basis to understand perturbation of cholesterol-dependent processes in macrophages that were shown previously by others to be necessary for their proper function in innate and adaptive immune responses. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  10. 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.

  11. Interactions between human NK cells and macrophages in response to Salmonella infection.

    Science.gov (United States)

    Lapaque, Nicolas; Walzer, Thierry; Méresse, Stéphane; Vivier, Eric; Trowsdale, John

    2009-04-01

    NK cells play a key role in host resistance to a range of pathogenic microorganisms, particularly during the initial stages of infection. NK cell interactions with cells infected with viruses and parasites have been studied extensively, but human bacterial infections have not been given the same attention. We studied crosstalk between human NK cells and macrophages infected with intracellular Salmonella. These macrophages activated NK cells, resulting in secretion of IFN-gamma and degranulation. Reciprocally, NK cell activation led to a dramatic reduction in numbers of intramacrophagic live bacteria. We identified three elements in the interaction of NK cells with infected macrophages. First, communication between NK cells and infected macrophages was contact-dependent. The second requirement was IL-2- and/or IL-15-dependent priming of NK cells to produce IFN-gamma. The third was activation of NK cells by IL-12 and IL-18, which were secreted by the Salmonella-infected macrophages. Adhesion molecules and IL-12Rbeta2 were enriched in the contact zone between NK cells and macrophages, consistent with contact- and IL-12/IL-18-dependent NK activation. Our results suggest that, in humans, bacterial clearance is consistent with a model invoking a "ménage à trois" involving NK cells, IL-2/IL-15-secreting cells, and infected macrophages.

  12. Quantitative Analysis of Proteome Modulations in Alveolar Epithelial Type II Cells in Response to PulmonaryAspergillus fumigatusInfection.

    Science.gov (United States)

    Seddigh, Pegah; Bracht, Thilo; Molinier-Frenkel, Válerie; Castellano, Flavia; Kniemeyer, Olaf; Schuster, Marc; Weski, Juliane; Hasenberg, Anja; Kraus, Andreas; Poschet, Gernot; Hager, Thomas; Theegarten, Dirk; Opitz, Christiane A; Brakhage, Axel A; Sitek, Barbara; Hasenberg, Mike; Gunzer, Matthias

    2017-12-01

    The ubiquitous mold Aspergillus fumigatus threatens immunosuppressed patients as inducer of lethal invasive aspergillosis. A. fumigatus conidia are airborne and reach the alveoli, where they encounter alveolar epithelial cells (AEC). Previous studies reported the importance of the surfactant-producing AEC II during A. fumigatus infection via in vitro experiments using cell lines. We established a negative isolation protocol yielding untouched primary murine AEC II with a purity >90%, allowing ex vivo analyses of the cells, which encountered the mold in vivo By label-free proteome analysis of AEC II isolated from mice 24h after A. fumigatus or mock infection we quantified 2256 proteins and found 154 proteins to be significantly differentially abundant between both groups (ANOVA p value ≤ 0.01, ratio of means ≥1.5 or ≤0.67, quantified with ≥2 peptides). Most of these proteins were higher abundant in the infected condition and reflected a comprehensive activation of AEC II on interaction with A. fumigatus This was especially represented by proteins related to oxidative phosphorylation, hence energy production. However, the most strongly induced protein was the l-amino acid oxidase (LAAO) Interleukin 4 induced 1 (IL4I1) with a 42.9 fold higher abundance (ANOVA p value 2.91 -10 ). IL4I1 has previously been found in B cells, macrophages, dendritic cells and rare neurons. Increased IL4I1 abundance in AEC II was confirmed by qPCR, Western blot and immunohistology. Furthermore, A. fumigatus infected lungs showed high levels of IL4I1 metabolic products. Importantly, higher IL4I1 abundance was also confirmed in lung tissue from human aspergilloma. Because LAAO are key enzymes for bactericidal product generation, AEC II might actively participate in pathogen defense. We provide insights into proteome changes of primary AEC II thereby opening new avenues to analyze the molecular changes of this central lung cell on infectious threats. Data are available via Proteome

  13. Changes in Macrophage Gene Expression Associated with Leishmania (Viannia braziliensis Infection.

    Directory of Open Access Journals (Sweden)

    Clemencia Ovalle-Bracho

    Full Text Available Different Leishmania species cause distinct clinical manifestations of the infectious disease leishmaniasis. It is fundamentally important to understand the mechanisms governing the interaction between Leishmania and its host cell. Little is known about this interaction between Leishmania (Viannia braziliensis and human macrophages. In this study, we aimed to identify differential gene expression between non-infected and L. (V braziliensis-infected U937-derived macrophages. We deployed a whole human transcriptome microarray analysis using 72 hours post-infection samples and compared those samples with their non-infected counterparts. We found that 218 genes were differentially expressed between infected and non-infected macrophages. A total of 71.6% of these genes were down-regulated in the infected macrophages. Functional enrichment analyses identified the steroid and sterol/cholesterol biosynthetic processes between regulatory networks down-regulated in infected macrophages. RT-qPCR further confirmed this down-regulation in genes belonging to these pathways. These findings contrast with those from studies involving other Leishmania species at earlier infection stages, where gene up-regulation for this metabolic pathway has been reported. Sterol biosynthesis could be an important biological process associated with the expression profile of macrophages infected by L. (V. braziliensis. Differential transcriptional results suggest a negative regulation of the genetic regulatory network involved in cholesterol biosynthesis.

  14. Low Dose BCG Infection as a Model for Macrophage Activation Maintaining Cell Viability

    Directory of Open Access Journals (Sweden)

    Leslie Chávez-Galán

    2016-01-01

    Full Text Available Mycobacterium bovis BCG, the current vaccine against tuberculosis, is ingested by macrophages promoting the development of effector functions including cell death and microbicidal mechanisms. Despite accumulating reports on M. tuberculosis, mechanisms of BCG/macrophage interaction remain relatively undefined. In vivo, few bacilli are sufficient to establish a mycobacterial infection; however, in vitro studies systematically use high mycobacterium doses. In this study, we analyze macrophage/BCG interactions and microenvironment upon infection with low BCG doses and propose an in vitro model to study cell activation without affecting viability. We show that RAW macrophages infected with BCG at MOI 1 activated higher and sustained levels of proinflammatory cytokines and transcription factors while MOI 0.1 was more efficient for early stimulation of IL-1β, MCP-1, and KC. Both BCG infection doses induced iNOS and NO in a dose-dependent manner and maintained nuclear and mitochondrial structures. Microenvironment generated by MOI 1 induced macrophage proliferation but not MOI 0.1 infection. In conclusion, BCG infection at low dose is an efficient in vitro model to study macrophage/BCG interactions that maintains macrophage viability and mitochondrial structures. This represents a novel model that can be applied to BCG research fields including mycobacterial infections, cancer immunotherapy, and prevention of autoimmunity and allergies.

  15. Inhibitory effects of Zanthoxylum rhoifolium Lam. (Rutaceae) against the infection and infectivity of macrophages by Leishmania amazonensis

    OpenAIRE

    MELO NETO, BERNARDO; LEITÃO, JOSEANA M.S.R.; OLIVEIRA, LUCIANO G.C.; SANTOS, SÉRGIO E.M.; CARNEIRO, SABRINA M.P.; RODRIGUES, KLINGER A.F.; CHAVES, MARIANA H.; ARCANJO, DANIEL D.R.; CARVALHO, FERNANDO A.A.

    2016-01-01

    ABSTRACT Zanthoxylum rhoifolium Lam. (Rutaceae) has been traditionally used in the treatment of microbial infections and parasitic diseases. In the present study, the antileishmanial effect induced by the ethanol extract of stem barks from Z. rhoifolium (ZR-EEtOH) and its n-hexane fraction (ZR-FHEX) on infection and infectivity of murine macrophages by promastigote forms of Leishmania amazonensis were investigated. In different set of experiments, macrophages or promastigotes were pretreated ...

  16. Aspirin inhibits lipopolysaccharide-induced COX-2 expression and PGE2 production in porcine alveolar macrophages by modulating protein kinase C and protein tyrosine phosphatase activity.

    Science.gov (United States)

    Duan, Yuzhong; Chen, Fanglin; Zhang, Anmei; Zhu, Bo; Sun, Jianguo; Xie, Qichao; Chen, Zhengtang

    2014-01-01

    Aspirin has been demonstrated to be effective in inhibiting COX-2 and PGE(2) in Alveolar macrophages (AMs). However, the mechanisms have not been fully understood. In the present study, we found that pretreatment with aspirin inhibited LPS-induced COX-2 and PGE(2) upregulation, IκBα degradation, NFκB activation and the increase of PKC activity, but elevated LPS-induced the decrease of PTP activity. The PKC inhibitor calphostin C dramatically reduced the COX-2 mRNA and PGE(2) levels, but the PTP inhibitor peroxovanadium (POV) significantly increased the COX-2 mRNA and PGE(2) levels. Furthermore, the PTP inhibitor mitigated the inhibitory effect of aspirin on COX-2 and PGE(2) upregulation and NF-κB activation, whereas the PKC inhibitor enhanced the inhibitory effects of aspirin on the production of COX-2 and PGE(2). Our data indicate a novel mechanism by which aspirin acts as a potent anti-inflammatory agent in alveolus macrophages and ALI.

  17. Inhibitory effects of Zanthoxylum rhoifolium Lam. (Rutaceae against the infection and infectivity of macrophages by Leishmania amazonensis

    Directory of Open Access Journals (Sweden)

    BERNARDO MELO NETO

    2016-01-01

    Full Text Available ABSTRACT Zanthoxylum rhoifolium Lam. (Rutaceae has been traditionally used in the treatment of microbial infections and parasitic diseases. In the present study, the antileishmanial effect induced by the ethanol extract of stem barks from Z. rhoifolium (ZR-EEtOH and its n-hexane fraction (ZR-FHEX on infection and infectivity of murine macrophages by promastigote forms of Leishmania amazonensis were investigated. In different set of experiments, macrophages or promastigotes were pretreated with ZR-EEtOH or ZR-FHEX at non-lethal concentrations for 24 hours, and then macrophages were submitted to infection by promastigotes. Moreover, their effects on activation of macrophages, as well as on the DNA content, size and number of promastigotes by flow cytometry were also evaluated. The infection rate and the number of internalized amastigote forms were markedly decreased after pretreatment of macrophages or promastigotes when compared with non-treated cells. The increase in phagocytic capability and nitrite content was also observed. Furthermore, the decrease of DNA content, size and number of promastigotes was also observed. In conclusion, ZR-EEtOH and ZR-FHEX promoted a markedly significant antileishmanial effect and reduction of infection of macrophages, probably underlying defense mechanisms activation in macrophages. These findings reinforce the potential application of Z. rhoifolium in the treatment of leishmaniasis.

  18. Inhibitory effects of Zanthoxylum rhoifolium Lam. (Rutaceae) against the infection and infectivity of macrophages by Leishmania amazonensis.

    Science.gov (United States)

    Melo, Bernardo; Leitão, Joseana M S R; Oliveira, Luciano G C; Santos, Sérgio E M; Carneiro, Sabrina M P; Rodrigues, Klinger A F; Chaves, Mariana H; Arcanjo, Daniel D R; Carvalho, Fernando A A

    2016-01-01

    Zanthoxylum rhoifolium Lam. (Rutaceae) has been traditionally used in the treatment of microbial infections and parasitic diseases. In the present study, the antileishmanial effect induced by the ethanol extract of stem barks from Z. rhoifolium (ZR-EEtOH) and its n-hexane fraction (ZR-FHEX) on infection and infectivity of murine macrophages by promastigote forms of Leishmania amazonensis were investigated. In different set of experiments, macrophages or promastigotes were pretreated with ZR-EEtOH or ZR-FHEX at non-lethal concentrations for 24 hours, and then macrophages were submitted to infection by promastigotes. Moreover, their effects on activation of macrophages, as well as on the DNA content, size and number of promastigotes by flow cytometry were also evaluated. The infection rate and the number of internalized amastigote forms were markedly decreased after pretreatment of macrophages or promastigotes when compared with non-treated cells. The increase in phagocytic capability and nitrite content was also observed. Furthermore, the decrease of DNA content, size and number of promastigotes was also observed. In conclusion, ZR-EEtOH and ZR-FHEX promoted a markedly significant antileishmanial effect and reduction of infection of macrophages, probably underlying defense mechanisms activation in macrophages. These findings reinforce the potential application of Z. rhoifolium in the treatment of leishmaniasis.

  19. Interim endodontic therapy for alveolar socket bone regeneration of infected hopeless teeth prior to implant therapy.

    Science.gov (United States)

    Rass, Marwan Abou

    2010-01-01

    patients' dental care. When treated, these hopeless teeth served many preventive, biologic, and esthetic functions. The infections of the alveolar sockets were eliminated, the alveolar bone defects were repaired through normal bone regeneration, and sockets anatomies were maintained or restored. Furthermore, the patients were spared maxillary sinus surgery and the possible complications resulting from major GBR and GTR procedures. In summary, the interim treatment facilitated tooth extraction and immediate implant placement.

  20. L-Arginine is not the limiting factor for nitric oxide synthesis by human alveolar macrophages in vitro

    NARCIS (Netherlands)

    Muijsers, RBR; ten Hacken, NHT; Van Ark, [No Value; Folkerts, G; Nijkamp, FP; Postma, DS

    2001-01-01

    Unlike murine mononuclear phagocytes, human macrophages do not release high amounts of nitric oxide (NO) in vitro despite the presence of nitric oxide synthase (NOS). To determine whether this limited NO synthesis in vitro is due to limited availability of the NOS substrate L-arginine, and putative

  1. CD4 depletion in SIV-infected macaques results in macrophage and microglia infection with rapid turnover of infected cells.

    Directory of Open Access Journals (Sweden)

    Luca Micci

    2014-10-01

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

  2. Novel elvitegravir nanoformulation approach to suppress the viral load in HIV-infected macrophages

    Directory of Open Access Journals (Sweden)

    Yuqing Gong

    2017-12-01

    Conclusions: PLGA-based EVG nanoformulation increased the intracellular uptake of EVG, as well as enhanced viral suppression in HIV-infected macrophages, suggesting its potential for improved HIV treatment in monocytic cells.

  3. Immuno-Pharmacological Targeting of Virus-Containing Compartments in HIV-1-Infected Macrophages.

    Science.gov (United States)

    Graziano, Francesca; Vicenzi, Elisa; Poli, Guido

    2016-07-01

    In addition to CD4 T lymphocytes, HIV-1 infects tissue macrophages that can actively accumulate infectious virions in vacuolar subcellular structures mostly connected to the plasma membrane and recently termed virus-containing compartments (VCCs). The VCC-associated HIV-1 reservoir of infected macrophages can be either increased or depleted by immunologic and pharmacologic agents, at least in vitro, thus suggesting that these factors (or related molecules) could be effective in curtailing the macrophage-associated HIV-1 reservoir in infected individuals receiving combination antiretroviral therapy (cART). Here we review evidence on the pathogenic role of tissue macrophages as long-term viral reservoirs in vivo and upon in vitro infection with a particular emphasis on the immuno-pharmacological modulation of VCCs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Alteration of human macrophages microRNA expression profile upon infection with Mycobacterium tuberculosis

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    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.

  5. ABO Blood Groups Influence Macrophage-mediated Phagocytosis of Plasmodium falciparum-infected Erythrocytes

    Science.gov (United States)

    Branch, Donald R.; Hult, Annika K.; Olsson, Martin L.; Liles, W. Conrad; Cserti-Gazdewich, Christine M.; Kain, Kevin C.

    2012-01-01

    Erythrocyte polymorphisms associated with a survival advantage to Plasmodium falciparum infection have undergone positive selection. There is a predominance of blood group O in malaria-endemic regions, and several lines of evidence suggest that ABO blood groups may influence the outcome of P. falciparum infection. Based on the hypothesis that enhanced innate clearance of infected polymorphic erythrocytes is associated with protection from severe malaria, we investigated whether P. falciparum-infected O erythrocytes are more efficiently cleared by macrophages than infected A and B erythrocytes. We show that human macrophages in vitro and mouse monocytes in vivo phagocytose P. falciparum-infected O erythrocytes more avidly than infected A and B erythrocytes and that uptake is associated with increased hemichrome deposition and high molecular weight band 3 aggregates in infected O erythrocytes. Using infected A1, A2, and O erythrocytes, we demonstrate an inverse association of phagocytic capacity with the amount of A antigen on the surface of infected erythrocytes. Finally, we report that enzymatic conversion of B erythrocytes to type as O before infection significantly enhances their uptake by macrophages to observed level comparable to that with infected O wild-type erythrocytes. These data provide the first evidence that ABO blood group antigens influence macrophage clearance of P. falciparum-infected erythrocytes and suggest an additional mechanism by which blood group O may confer resistance to severe malaria. PMID:23071435

  6. Neutrophil Migration into the Infected Uroepithelium Is Regulated by the Crosstalk between Resident and Helper Macrophages

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    Kristina Zec

    2016-02-01

    Full Text Available The antibacterial defense against infections depends on the cooperation between distinct phagocytes of the innate immune system, namely macrophages and neutrophils. However, the mechanisms driving this cooperation are incompletely understood. In this study we describe the crosstalk between Ly6C+ and Ly6C− macrophage-subtypes and neutrophils in the context of urinary tract infection (UTI with uropathogenic E. coli (UPEC. Ly6C− macrophages acted as tissue resident sentinels and attracted circulating phagocytes by chemokines. Ly6C+ macrophages produced tumor necrosis factor (TNF that licensed Ly6C− macrophages to release preformed CXCL2, which in turn caused matrix metalloproteinases (MMP-9 secretion by neutrophils to enable transepithelial migration.

  7. A comparison of two distinct murine macrophage gene expression profiles in response to Leishmania amazonensis infection

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    Probst Christian M

    2012-02-01

    Full Text Available Abstract Background The experimental murine model of leishmaniasis has been widely used to characterize the immune response against Leishmania. CBA mice develop severe lesions, while C57BL/6 present small chronic lesions under L. amazonensis infection. Employing a transcriptomic approach combined with biological network analysis, the gene expression profiles of C57BL/6 and CBA macrophages, before and after L. amazonensis infection in vitro, were compared. These strains were selected due to their different degrees of susceptibility to this parasite. Results The genes expressed by C57BL/6 and CBA macrophages, before and after infection, differ greatly, both with respect to absolute number as well as cell function. Uninfected C57BL/6 macrophages express genes involved in the deactivation pathway of macrophages at lower levels, while genes related to the activation of the host immune inflammatory response, including apoptosis and phagocytosis, have elevated expression levels. Several genes that participate in the apoptosis process were also observed to be up-regulated in C57BL/6 macrophages infected with L. amazonensis, which is very likely related to the capacity of these cells to control parasite infection. By contrast, genes involved in lipid metabolism were found to be up-regulated in CBA macrophages in response to infection, which supports the notion that L. amazonensis probably modulates parasitophorous vacuoles in order to survive and multiply in host cells. Conclusion The transcriptomic profiles of C57BL/6 macrophages, before and after infection, were shown to be involved in the macrophage pathway of activation, which may aid in the control of L. amazonensis infection, in contrast to the profiles of CBA cells.

  8. 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

  9. Transcriptional immunoresponse of tissue-specific macrophages in swine after infection with African swine fever virus

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    Kowalczyk Andrzej

    2015-12-01

    Full Text Available Macrophages and cytokines are important in the control of inflammation and regulation of the immune response. However, they can also contribute to immunopathology in the host after viral infection and the regulatory network can be subverted by infectious agents, including viruses, some of which produce cytokine analogues or have mechanisms that inhibit cytokine function. African swine fever virus (ASFV encodes a number of proteins which modulate cytokine and chemokine induction, host transcription factor activation, stress responses, and apoptosis. The aim of this review is to elucidate the mechanisms of immune responses to ASFV in different subpopulations of porcine macrophages. A transcriptional immune response in different resident tissue macrophages following ASFV infection was presented in many publications. ASFV-susceptible porcine macrophages can be of several origins, such as peripheral blood, lungs, bone marrow, etc. blood monocytes, blood macrophages, and lung macrophages have demonstrated a modulation of phenotype. Monocyte-derived macrophages could express surface markers not found on their monocyte precursors. Moreover, they can undergo further differentiation after infection and during inflammation. When viruses infect such cells, immunological activity can be seriously impaired or modified.

  10. Peroxisome proliferator-activated receptor-γ-mediated polarization of macrophages in Neospora caninum infection.

    Science.gov (United States)

    He, Xuexiu; Gong, Pengtao; Wei, Zhengkai; Liu, Weijian; Wang, Weili; Li, Jianhua; Yang, Zhengtao; Zhang, Xichen

    2017-07-01

    Neospora caninum is an apicomplexan parasite closely related Toxoplasma gondii, which causes neurological disease and abortion in multiple animal species. Macrophage polarization plays an important role in host immune responses to parasites infection, such as Toxoplasma gondii, Leishmania, Trypanosoma cruzi. However, the dynamics of macrophage polarization, as well as the possible mechanism that regulate macrophage polarization, during N. caninum infection remains unclear. The M1 and M2-phenotypic markers of peritoneal macrophages from mice infected with tachyzoites of Nc-1 were analyzed by flow cytometry (FCM) analysis. Then J774A.1 cells were respectively treated with GW9662 and RGZ, and stimulated by tachyzoites of Nc-1. M1 and M2-phenotypic markers were determined by FCM and ELISA. And the activations of PPAR-γ and NF-κB were determined by Western blotting. In this study, our data showed that macrophages were preferentially differentiated into the M1 type during the acute stage of N. caninum infection, while the level of M2 macrophages significantly increased during the chronic stage of infection. In vitro study, compared with the GW9662 group and RGZ group, N. caninum can promote M2-polarized phenotype through up-regulate the activity of PPAR-γ and inhibting NF-κB activation. In conclusion, this study demonstrated that macrophages are plastic since M1 differentiated macrophages can express M2 markers with N. caninum infection through up-regulating the activity of PPAR-γ and inhibting NF-κB activation and may be providing new insights for the prevention and treatment of N. caninum infection. Copyright © 2017. Published by Elsevier Inc.

  11. Type I Interferon Induced Epigenetic Regulation of Macrophages Suppresses Innate and Adaptive Immunity in Acute Respiratory Viral Infection.

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    Danielle N Kroetz

    2015-12-01

    Full Text Available Influenza A virus (IAV is an airborne pathogen that causes significant morbidity and mortality each year. Macrophages (Mϕ are the first immune population to encounter IAV virions in the lungs and are required to control infection. In the present study, we explored the mechanism by which cytokine signaling regulates the phenotype and function of Mϕ via epigenetic modification of chromatin. We have found that type I interferon (IFN-I potently upregulates the lysine methyltransferase Setdb2 in murine and human Mϕ, and in turn Setdb2 regulates Mϕ-mediated immunity in response to IAV. The induction of Setdb2 by IFN-I was significantly impaired upon inhibition of the JAK-STAT signaling cascade, and chromatin immunoprecipitation revealed that both STAT1 and interferon regulatory factor 7 bind upstream of the transcription start site to induce expression. The generation of Setdb2LacZ reporter mice revealed that IAV infection results in systemic upregulation of Setdb2 in myeloid cells. In the lungs, alveolar Mϕ expressed the highest level of Setdb2, with greater than 70% lacZ positive on day 4 post-infection. Silencing Setdb2 activity in Mϕ in vivo enhanced survival in lethal IAV infection. Enhanced host protection correlated with an amplified antiviral response and less obstruction to the airways. By tri-methylating H3K9, Setdb2 silenced the transcription of Mx1 and Isg15, antiviral effectors that inhibit IAV replication. Accordingly, a reduced viral load in knockout mice on day 8 post-infection was linked to elevated Isg15 and Mx1 transcript in the lungs. In addition, Setdb2 suppressed the expression of a large number of other genes with proinflammatory or immunomodulatory function. This included Ccl2, a chemokine that signals through CCR2 to regulate monocyte recruitment to infectious sites. Consistently, knockout mice produced more CCL2 upon IAV infection and this correlated with a 2-fold increase in the number of inflammatory monocytes and

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

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    Aruna Amarasinghe

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

  13. NLRP3 inflammasome activation in murine macrophages caused by Neospora caninum infection.

    Science.gov (United States)

    Wang, Xiaocen; Gong, Pengtao; Zhang, Xu; Wang, Jielin; Tai, Lixin; Wang, Xu; Wei, Zhengkai; Yang, Yongjun; Yang, Zhengtao; Li, Jianhua; Zhang, Xichen

    2017-05-30

    Neospora caninum is an intracellular parasite that causes significant economic losses in cattle industry. Understanding the host resistance mechanisms in the innate immune response to neosporosis could facilitate the exploration of approaches for controlling N. caninum infection. The NLR inflammasome is a multiprotein platform in the cell cytoplasm and plays critical roles in the host response against microbes. Neospora caninum-infected wild-type (WT) macrophages and Nlrp3 -/- macrophages, and inhibitory approaches were used to investigate inflammasome activation and its role in N. caninum infection. Inflammasome RT Profiler PCR Arrays were used to identify the primary genes involved in N. caninum infection. The expression of the sensor protein NLRP3, processing of caspase-1, secretion of IL-1β and cell death were detected. Neospora caninum replication in macrophages was also assessed. Many NLR molecules participated in the recognition of N. caninum, especially the sensor protein NLRP3, and further study revealed that the NLRP3 distribution became punctate in the cell cytoplasm, which facilitated inflammasome activation. Inflammasome activation-mediated caspase-1 processing and IL-1β cleavage in response to N. caninum infection were observed and were correlated with the time of infection and number of infecting parasites. LDH-related cell death was also observed, and this death was regarded as beneficial for the clearance of N. caninum. Treatment of N. caninum-infected macrophages with caspase-1, pan-caspase and NLRP3 inhibitors led to the impaired release of active IL-1β and a failure to restrict parasite replication. And Neospora caninum infected peritoneal macrophages from Nlrp3-deficient mice displayed greatly decreased release of active IL-1β and the failure of caspase-1 cleavage. The NLRP3 inflammasome can be activated in N. caninum-infected macrophages, and plays a protective role in the host response to control N. caninum.

  14. 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.

  15. 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

  16. 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

    2018-03-20

    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.

  17. Chronic filarial infection provides protection against bacterial sepsis by functionally reprogramming macrophages.

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    Fabian Gondorf

    2015-01-01

    Full Text Available Helminths immunomodulate their hosts and induce a regulatory, anti-inflammatory milieu that prevents allergies and autoimmune diseases. Helminth immunomodulation may benefit sepsis outcome by preventing exacerbated inflammation and severe pathology, but the influence on bacterial clearance remains unclear. To address this, mice were chronically infected with the filarial nematode Litomosoides sigmodontis (L.s. and the outcome of acute systemic inflammation caused by i.p. Escherichia coli injection was determined. L.s. infection significantly improved E. coli-induced hypothermia, bacterial clearance and sepsis survival and correlated with reduced concentrations of associated pro-inflammatory cytokines/chemokines and a less pronounced pro-inflammatory macrophage gene expression profile. Improved sepsis outcome in L.s.-infected animals was mediated by macrophages, but independent of the alternatively activated macrophage subset. Endosymbiotic Wolbachia bacteria that are present in most human pathogenic filariae, as well as L.s., signal via TLR2 and modulate macrophage function. Here, gene expression profiles of peritoneal macrophages from L.s.-infected mice revealed a downregulation of genes involved in TLR signaling, and pulsing of macrophages in vitro with L.s. extract reduced LPS-triggered activation. Subsequent transfer improved sepsis outcome in naïve mice in a Wolbachia- and TLR2-dependent manner. In vivo, phagocytosis was increased in macrophages from L.s.-infected wild type, but not TLR2-deficient animals. In association, L.s. infection neither improved bacterial clearance in TLR2-deficient animals nor ameliorated E. coli-induced hypothermia and sepsis survival. These results indicate that chronic L.s. infection has a dual beneficial effect on bacterial sepsis, reducing pro-inflammatory immune responses and improving bacterial control. Thus, helminths and their antigens may not only improve the outcome of autoimmune and allergic diseases

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

    International Nuclear Information System (INIS)

    Okamoto, Mika; Wang, Xin; Baba, Masanori

    2005-01-01

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

  19. Murine macrophage inflammatory cytokine production and immune activation in response to Vibrio parahaemolyticus infection

    Science.gov (United States)

    Vibrio parahaemolyticus is the most common cause of bacterial seafood-related illness in the United States. Currently, there is a dearth of literature regarding immunity to infection with this pathogen. Here we studied V. parahaemolyticus-infected RAW 264.7 murine macrophage detecting both pro- and...

  20. Alternate splicing of transcripts shape macrophage response to Mycobacterium tuberculosis infection.

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    Haroon Kalam

    2017-03-01

    Full Text Available Transcriptional reprogramming of macrophages upon Mycobacterium tuberculosis (Mtb infection is widely studied; however, the significance of alternate splicing (AS in shaping cellular responses to mycobacterial infections is not yet appreciated. Alternate splicing can influence transcript stability or structure, function and localization of corresponding proteins thereby altering protein stoichiometry and physiological consequences. Using comprehensive analysis of a time-series RNA-seq data obtained from human macrophages infected with virulent or avirulent strains of Mtb, we show extensive remodeling of alternate splicing in macrophage transcriptome. The global nature of this regulation was evident since genes belonging to functional classes like trafficking, immune response, autophagy, redox and metabolism showed marked departure in the pattern of splicing in the infected macrophages. The systemic perturbation of splicing machinery in the infected macrophages was apparent as genes involved at different stages of spliceosome assembly were also regulated at the splicing level. Curiously there was a considerable increase in the expression of truncated/non-translatable variants of several genes, specifically upon virulent infections. Increased expression of truncated transcripts correlated with a decline in the corresponding protein levels. We verified the physiological relevance for one such candidate gene RAB8B; whose truncated variant gets enriched in H37Rv infected cells. Upon tweaking relative abundance of longer or shorter variants of RAB8B transcripts by specialized transduction, mycobacterial targeting to lysosomes could be promoted or blocked respectively, which also resulted in corresponding changes in the bacterial survival. Our results show RAB8B recruitment to the mycobacterial phagosomes is required for phagosome maturation. Thus the abundance of truncated RAB8B variant helps virulent Mtb survival by limiting the RAB8B levels in the

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

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    Erico Vinícius de Souza Carmo

    2010-11-01

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

  2. Proteomic Investigation of the Time Course Responses of RAW 264.7 Macrophages to Infection with Salmonella enterica

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    Shi, Liang; Chowdhury, Saiful M.; Smallwood, Heather S.; Yoon, Hyunjin; Mottaz-Brewer, Heather M.; Norbeck, Angela D.; McDermott, Jason E.; Clauss, Therese RW; Heffron, Fred; Smith, Richard D.; Adkins, Joshua N.

    2009-08-01

    Macrophages plan important roles in controlling Salmonella-mediated systemic infection. To investigate the responses of macrophages to Salmonella infection, we infected RAW 264.7 macrophages with Salmonella enterica serovar Typhimurium (STM) and then performed a comparative liquid chromatography-tandem mass spectrometry [LC-MS(/MS)]-based proteomics analysis of the infected macrophages. A total of 1006 macrophage and 115 STM proteins were indentified from this study. Most of STM proteins were found at late stage of the time course of infection, consistent with the fact that STM proliferates inside RAW 264.7 macrophages. Majority of the identified macrophage proteins were house keeping-related, including cytoplasmic superoxide dismutase 1 (SOD1), whose peptide abundances were relatively constant during the time course of infection. Compared to those in no infection control, the peptide abundances of 244 macrophage proteins (or 24% of total indentified macrophage proteins) changed considerably after STM infection. The functions of these STM infection-affected macrophage proteins were diverse and ranged from production of antibacterial nitric oxide (i.e., inducible nitric oxide synthase or iNOS) or production of prostaglandin H2 (i.e., prostaglandin-endoperoxide synthase 2, also know as cyclooxygenase-2 or COX-2) to regulation of intracellular traffic (e.g., sorting nexin or SNX 5, 6 and 9), demonstrating a global impact of STM infection on macrophage proteome. Western-blot analysis not only confirmed the LC-MS(/MS) results of SOD1, COX-2 and iNOS, but also revealed that the protein abundances of mitochondrial SOD2 increased after STM infection, indicating an infection-induced oxidative stress in mitochondria.

  3. 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

  4. Real-time visualization of HIV-1 GAG trafficking in infected macrophages.

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    Karine Gousset

    2008-03-01

    Full Text Available HIV-1 particle production is driven by the Gag precursor protein Pr55(Gag. Despite significant progress in defining both the viral and cellular determinants of HIV-1 assembly and release, the trafficking pathway used by Gag to reach its site of assembly in the infected cell remains to be elucidated. The Gag trafficking itinerary in primary monocyte-derived macrophages is especially poorly understood. To define the site of assembly and characterize the Gag trafficking pathway in this physiologically relevant cell type, we have made use of the biarsenical-tetracysteine system. A small tetracysteine tag was introduced near the C-terminus of the matrix domain of Gag. The insertion of the tag at this position did not interfere with Gag trafficking, virus assembly or release, particle infectivity, or the kinetics of virus replication. By using this in vivo detection system to visualize Gag trafficking in living macrophages, Gag was observed to accumulate both at the plasma membrane and in an apparently internal compartment that bears markers characteristic of late endosomes or multivesicular bodies. Significantly, the internal Gag rapidly translocated to the junction between the infected macrophages and uninfected T cells following macrophage/T-cell synapse formation. These data indicate that a population of Gag in infected macrophages remains sequestered internally and is presented to uninfected target cells at a virological synapse.

  5. Siglecs facilitate HIV-1 infection of macrophages through adhesion with viral sialic acids.

    Directory of Open Access Journals (Sweden)

    Zhongcheng Zou

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 infects macrophages effectively, despite relatively low levels of cell surface-expressed CD4. Although HIV-1 infections are defined by viral tropisms according to chemokine receptor usage (R5 and X4, variations in infection are common within both R5- and X4-tropic viruses, indicating additional factors may contribute to viral tropism.Using both solution and cell surface binding experiments, we showed that R5- and X4-tropic HIV-1 gp120 proteins recognized a family of I-type lectin receptors, the Sialic acid-binding immunoglobulin-like lectins (Siglec. The recognition was through envelope-associated sialic acids that promoted viral adhesion to macrophages. The sialic acid-mediated viral-host interaction facilitated both R5-tropic pseudovirus and HIV-1(BaL infection of macrophages. The high affinity Siglec-1 contributed the most to HIV-1 infection and the variation in Siglec-1 expression on primary macrophages from different donors was associated statistically with sialic acid-facilitated viral infection. Furthermore, envelope-associated sialoglycan variations on various strains of R5-tropic viruses also affected infection.Our study showed that sialic acids on the viral envelope facilitated HIV-1 infection of macrophages through interacting with Siglec receptors, and the expression of Siglec-1 correlated with viral sialic acid-mediated host attachment. This glycan-mediated viral adhesion underscores the importance of viral sialic acids in HIV infection and pathogenesis, and suggests a novel class of antiviral compounds targeting Siglec receptors.

  6. Effect of Chaiqinchengqi decoction on inositol requiring enzyme 1α in alveolar macrophages of dogs with acute necrotising pancreatitis induced by sodium taurocholate.

    Science.gov (United States)

    Jia, Guo; Xiaoxiang, Wang; Ruijie, Luo; Xiaoxin, Zhang; Xiaonan, Yang; Qing, Xia; Ping, Xue

    2015-08-01

    To investigate the effect of Chaiqinchengqi decoction (CQCQD) on inositol requiring enzyme lα (IRElα) in alveolar macrophages (AMs) of the dog model of acute necrotising pancreatitis (ANP) induced by sodium taurocholate. Fifteen beagle dogs were randomised into a control group, ANP group and CQCQD group (n = 5 per group). ANP was induced by a retrograde duct injection of 50 mg/kg of 5% sodium taurocholate. The dogs in the control group received injections of the same volume of saline as the sodium taurocholate. After the models were induced, the dogs in the CQCQD group were administered 10 mL/kg CQCQD every 2 h for 6 h. Two hours after the last administration of either CQCQD or saline, they were sacrificed by anaesthesia. AMs were collected to determine the IRElα and interleukin-1β (IL-1β) mRNA and protein expression, and pancreatic tissues were collected for histopathology analysis. Compared with the ANP group, the mRNA and protein expression of IREl a and the protein expression of IL-1β of AMs in the CQCQD group were significantly down-regulated, and the pancreatic histopathology score of the CQCQD group also was lower. There was no significant difference in the mRNA expression of IL-1β of AMs between the two groups. The CQCQD-induced down-regulation of the IL-1β protein expression may involve the down-regulation of the mRNA and protein expression of IRElα in AMs.

  7. Respiratory tract responses to dust: Relationships between dust burden, lung injury, alveolar macrophage fibronectin release, and the development of pulmonary fibrosis

    Energy Technology Data Exchange (ETDEWEB)

    Driscoll, K.E.; Maurer, J.K.; Lindenschmidt, R.C.; Romberger, D.; Rennard, S.I.; Crosby, L. (Procter Gamble Company, Cincinnati, OH (USA))

    1990-10-01

    A multidisciplinary approach was used to investigate the responses of the respiratory tract to silica (SiO2) or titanium dioxide (TiO2). Rats were intratracheally instilled with 5-100 mg/kg of dust and bronchoalveolar lavage fluid (BALF) lactate dehydrogenase (LDH) and total protein (TP) and ex vivo alveolar macrophage (AM) fibronectin release assessed on Days 7, 14, and 28 after exposure. Lung dust burdens were determined on Days 1, 7, and 28 after instillation. Both dusts elicited dose-related increases in BALF LDH and TP, a response which was more pronounced and progressive with SiO2. All doses of SiO2 elicited persistent increases in AM fibronectin release. TiO2 stimulated persistent increases in AM fibronectin release at greater than or equal to 50 mg/kg, with transient or no effect at less than or equal to 10 mg/kg. Increased SiO2 retention was observed for all doses and TiO2 retention was increased only at doses greater than or equal to 50 mg/kg. In vitro exposure of naive AM to SiO2 or TiO2 did not stimulate AM fibronectin release. Histopathology demonstrated fibrosis at all SiO2 doses; only TiO2 doses greater than or equal to 50 mg/kg resulted in fibrosis. These results reveal an association between increased dust retention, lung injury, activation of AM fibronectin release, and the development of fibrosis. The magnitude and temporal pattern of responses clearly differentiated SiO2 from TiO2. The correlation of BALF markers of lung injury and increased AM fibronectin release with the development of fibrosis supports the use of these parameters as predictive biomarkers of dust-induced interstitial lung disease.

  8. 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

  9. Classical and alternative macrophages have impaired function during acute and chronic HIV-1 infection.

    Science.gov (United States)

    Galvão-Lima, Leonardo J; Espíndola, Milena S; Soares, Luana S; Zambuzi, Fabiana A; Cacemiro, Maira; Fontanari, Caroline; Bollela, Valdes R; Frantz, Fabiani G

    Three decades after HIV recognition and its association with AIDS development, many advances have emerged - especially related to prevention and treatment. Undoubtedly, the development of Highly Active Antiretroviral Therapy (HAART) dramatically changed the future of the syndrome that we know today. In the present study, we evaluate the impact of Highly Active Antiretroviral Therapy on macrophage function and its relevance to HIV pathogenesis. PBMCs were isolated from blood samples and monocytes (CD14+ cells) were purified. Monocyte-Derived Macrophages (MDMs) were activated on classical (M GM-CSF+IFN-γ ) or alternative (M IL-4+IL13 ) patterns using human recombinant cytokines for six days. After this period, Monocyte-Derived Macrophages were stimulated with TLR2/Dectin-1 or TLR4 agonists and we evaluated the influence of HIV-1 infection and Highly Active Antiretroviral Therapy on the release of cytokines/chemokines by macrophages. The data were obtained using Monocyte-Derived Macrophages derived from HIV naïve or from patients on regular Highly Active Antiretroviral Therapy. Classically Monocyte-Derived Macrophages obtained from HIV-1 infected patients on Highly Active Antiretroviral Therapy released higher levels of IL-6 and IL-12 even without PAMPs stimuli when compared to control group. On the other hand, alternative Monocyte-Derived Macrophages derived from HIV-1 infected patients on Highly Active Antiretroviral Therapy released lower levels of IL-6, IL-10, TNF-α, IP-10 and RANTES after LPS stimuli when compared to control group. Furthermore, healthy individuals have a complex network of cytokines/chemokines released by Monocyte-Derived Macrophages after PAMP stimuli, which was deeply affected in MDMs obtained from naïve HIV-1 infected patients and only partially restored in MDMs derived from HIV-1 infected patients even on regular Highly Active Antiretroviral Therapy. Our therapy protocols were not effective in restoring the functional alterations induced

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

    Czech Academy of Sciences Publication Activity Database

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

    2009-01-01

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

  11. Regulation of L-type Voltage Gated Calcium Channel CACNA1S in Macrophages upon Mycobacterium tuberculosis Infection

    OpenAIRE

    Antony, Cecil; Mehto, Subhash; Tiwari, Brijendra K.; Singh, Yogendra; Natarajan, Krishnamurthy

    2015-01-01

    We demonstrated earlier the inhibitory role played by Voltage Gated Calcium Channels (VGCCs) in regulating Mycobacterium tuberculosis (M. tb) survival and pathogenesis. In this report, we investigated mechanisms and key players that regulate the surface expression of VGCC-CACNA1S by Rv2463 and M. tb infection in macrophages. Our earlier work identified Rv2463 to be expressed at early times post infection in macrophages that induced suppressor responses to dendritic cells and macrophages. Our ...

  12. Inhibition of ecto-ATPase activities impairs HIV-1 infection of macrophages.

    Science.gov (United States)

    Schachter, Julieta; Delgado, Kelly Valcárcel; Barreto-de-Souza, Victor; Bou-Habib, Dumith Chequer; Persechini, Pedro Muanis; Meyer-Fernandes, José Roberto

    2015-05-01

    Nucleotides and nucleosides are secreted into extracellular media at different concentrations as a consequence of different physiologic and pathological conditions. Ecto-nucleotidases, enzymes present on the surface of most cells, hydrolyze these extracellular nucleotides and reduce the concentration of them, thus affecting the activation of different nucleotide and nucleoside receptors. Also, ecto-nucleotidases are present in a number of microorganisms and play important roles in host-pathogen interactions. Here, we characterized the ecto-ATPase activities present on the surface of HIV-1 particle and human macrophages as well. We found that the kinetic properties of HIV-1 and macrophage ecto-ATPases are similar, suggesting that the enzyme is the same. This ecto-ATPase activity was increased in macrophages infected in vitro with HIV-1. Using three different non-related ecto-ATPase inhibitors-POM-1, ARL67156 and BG0-we showed that the inhibition of these macrophage and viral ecto-ATPase activities impairs HIV-1 infection. In addition, we also found that elevated extracellular concentrations of ATP inhibit HIV-1 production by infected macrophages. Copyright © 2014 Elsevier GmbH. All rights reserved.

  13. Mycobacterium tuberculosis induces an atypical cell death mode to escape from infected macrophages.

    Directory of Open Access Journals (Sweden)

    Jinhee Lee

    Full Text Available BACKGROUND: Macrophage cell death following infection with Mycobacterium tuberculosis plays a central role in tuberculosis disease pathogenesis. Certain attenuated strains induce extrinsic apoptosis of infected macrophages but virulent strains of M. tuberculosis suppress this host response. We previously reported that virulent M. tuberculosis induces cell death when bacillary load exceeds ∼20 per macrophage but the precise nature of this demise has not been defined. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed the characteristics of cell death in primary murine macrophages challenged with virulent or attenuated M. tuberculosis complex strains. We report that high intracellular bacillary burden causes rapid and primarily necrotic death via lysosomal permeabilization, releasing hydrolases that promote Bax/Bak-independent mitochondrial damage and necrosis. Cell death was independent of cathepsins B or L and notable for ultrastructural evidence of damage to lipid bilayers throughout host cells with depletion of several host phospholipid species. These events require viable bacteria that can respond to intracellular cues via the PhoPR sensor kinase system but are independent of the ESX1 system. CONCLUSIONS/SIGNIFICANCE: Cell death caused by virulent M. tuberculosis is distinct from classical apoptosis, pyroptosis or pyronecrosis. Mycobacterial genes essential for cytotoxicity are regulated by the PhoPR two-component system. This atypical death mode provides a mechanism for viable bacilli to exit host macrophages for spreading infection and the eventual transition to extracellular persistence that characterizes advanced pulmonary tuberculosis.

  14. HIV-1 infection of macrophages is dependent on evasion of innate immune cellular activation.

    Science.gov (United States)

    Tsang, Jhen; Chain, Benjamin M; Miller, Robert F; Webb, Benjamin L J; Barclay, Wendy; Towers, Greg J; Katz, David R; Noursadeghi, Mahdad

    2009-11-13

    The cellular innate immune response to HIV-1 is poorly characterized. In view of HIV-1 tropism for macrophages, which can be activated via pattern recognition receptors to trigger antimicrobial defences, we investigated innate immune responses to HIV-1 by monocyte-derived macrophages. In a model of productive HIV-1 infection, cellular innate immune responses to HIV-1 were investigated, at the level of transcription factor activation, specific gene expression and genome-wide transcriptional profiling. In addition, the viral determinants of macrophage responses and the physiological effect of innate immune cellular activation on HIV-1 replication were assessed. Productive HIV-1 infection did not activate nuclear factor-kappaB and interferon regulatory factor 3 transcription factors or interferon gene expression (IFN) and caused remarkably small changes to the host-cell transcriptome, with no evidence of inflammatory or IFN signatures. Evasion of IFN induction was not dependent on HIV-1 envelope-mediated cellular entry, inhibition by accessory proteins or reverse transcription of ssRNA that may reduce innate immune cellular activation by viral RNA. Furthermore, IFNbeta priming did not sensitize responses to HIV-1. Importantly, exogenous IFNbeta or stimulation with the RNA analogue poly I:C to simulate innate immune activation invoked HIV-1 restriction. We conclude that macrophages lack functional pattern recognition receptors for this virus and that HIV-1 tropism for macrophages helps to establish a foothold in the host without triggering innate immune cellular activation, which would otherwise block viral infection effectively.

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

    Science.gov (United States)

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

    2017-05-26

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

  16. Pulmonary alveolar proteinosis in a child from an informal settlement

    African Journals Online (AJOL)

    clinical, histopathological, biochemical and genetic data.[3]. Surfactant homeostasis is critical for lung function and is tightly regulated, in part by pulmonary granulocyte-macrophage colony- stimulating factor (GM-CSF), which is required for surfactant clearance by alveolar macrophages and alveolar macrophage maturation.

  17. CD44 enhances macrophage phagocytosis and plays a protective role in Streptococcus equi subsp. zooepidemicus infection.

    Science.gov (United States)

    Fu, Qiang; Wei, Zigong; Xiao, Pingping; Chen, Yaosheng; Liu, Xiaohong

    2017-01-01

    Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is an important pathogen associated with a wide range of diseases in many mammalian species. CD44 is a transmembrane adhesion molecule involved in innate and adaptive immune responses. The aim of this study was to determine the protective role of CD44 during S. zooepidemicus infection. CD44-deficient mice exhibited reduced macrophage accumulation in the bronchoalveolar space and enhanced bacterial outgrowth and dissemination, which resulted in reduced mouse survival. An in vitro analysis revealed that CD44 can directly bind to S. zooepidemicus. Additionally, S. zooepidemicus interacted with macrophage-associated CD44, as reflected by the reduced uptake of S. zooepidemicus by CD44-deficient macrophages. These data suggest that CD44 contributes to effective antibacterial defense during S. zooepidemicus infection, thereby limiting the accompanying injury and death. Copyright © 2016. Published by Elsevier B.V.

  18. B Cell IgD Deletion Prevents Alveolar Bone Loss Following Murine Oral Infection

    Directory of Open Access Journals (Sweden)

    Pamela J. Baker

    2009-01-01

    and CD4+ T cells in immune normal mice compared to IgD deficient mice. These data suggest that IgD is an important mediator of alveolar bone resorption, possibly through antigen-specific coactivation of B cells and CD4+ T cells.

  19. The effect of hyperglycaemia on in vitro cytokine production and macrophage infection with Mycobacterium tuberculosis

    NARCIS (Netherlands)

    Lachmandas, E.; Vrieling, F.; Wilson, L.G.; Joosten, S.A.; Netea, M.G.; Ottenhoff, T.H.; Crevel, R. van

    2015-01-01

    Type 2 diabetes mellitus is an established risk factor for tuberculosis but the underlying mechanisms are largely unknown. We examined the effects of hyperglycaemia, a hallmark of diabetes, on the cytokine response to and macrophage infection with Mycobacterium tuberculosis. Increasing in vitro

  20. Modulation of Stat-1 in Human Macrophages Infected with Different Species of Intracellular Pathogenic Bacteria

    Directory of Open Access Journals (Sweden)

    Giuditta Fiorella Schiavano

    2016-01-01

    Full Text Available The infection of human macrophages by pathogenic bacteria induces different signaling pathways depending on the type of cellular receptors involved in the microorganism entry and on their mechanism(s of survival and replication in the host cell. It was reported that Stat proteins play an important role in this process. In the present study, we investigate the changes in Stat-1 activation (phosphorylation in p-tyr701 after uptake of two Gram-positive (Listeria monocytogenes and Staphylococcus aureus and two Gram-negative bacteria (Salmonella typhimurium and Legionella pneumophila characterized by their varying abilities to enter, survive, and replicate in human macrophages. Comparing the results obtained with Gram-negative and Gram-positive bacteria, Stat-1 activation in macrophages does not seem to be related to LPS content. The p-tyr701Stat-1 expression levels were found to be independent of the internalized bacterial number and IFN-γ release. On the contrary, Jak/Stat-1 pathway activation only occurs when an active infection has been established in the host macrophage, and it is plausible that the differences in the expression levels of p-tyr701Stat-1 could be due to different survival mechanisms or to differences in bacteria life cycles within macrophages.

  1. MicroRNA expression profile in human macrophages in response to Leishmania major infection.

    Directory of Open Access Journals (Sweden)

    Julien Lemaire

    Full Text Available Leishmania (L. are intracellular protozoan parasites able to survive and replicate in the hostile phagolysosomal environment of infected macrophages. They cause leishmaniasis, a heterogeneous group of worldwide-distributed affections, representing a paradigm of neglected diseases that are mainly embedded in impoverished populations. To establish successful infection and ensure their own survival, Leishmania have developed sophisticated strategies to subvert the host macrophage responses. Despite a wealth of gained crucial information, these strategies still remain poorly understood. MicroRNAs (miRNAs, an evolutionarily conserved class of endogenous 22-nucleotide non-coding RNAs, are described to participate in the regulation of almost every cellular process investigated so far. They regulate the expression of target genes both at the levels of mRNA stability and translation; changes in their expression have a profound effect on their target transcripts.We report in this study a comprehensive analysis of miRNA expression profiles in L. major-infected human primary macrophages of three healthy donors assessed at different time-points post-infection (three to 24 h. We show that expression of 64 out of 365 analyzed miRNAs was consistently deregulated upon infection with the same trends in all donors. Among these, several are known to be induced by TLR-dependent responses. GO enrichment analysis of experimentally validated miRNA-targeted genes revealed that several pathways and molecular functions were disturbed upon parasite infection. Finally, following parasite infection, miR-210 abundance was enhanced in HIF-1α-dependent manner, though it did not contribute to inhibiting anti-apoptotic pathways through pro-apoptotic caspase-3 regulation.Our data suggest that alteration in miRNA levels likely plays an important role in regulating macrophage functions following L. major infection. These results could contribute to better understanding of the

  2. Minocycline differentially modulates macrophage mediated peripheral immune response following Japanese encephalitis virus infection.

    Science.gov (United States)

    Dutta, Kallol; Mishra, Manoj Kumar; Nazmi, Arshed; Kumawat, Kanhaiya Lal; Basu, Anirban

    2010-11-01

    Japanese encephalitis virus (JEV) is a neurotropic flavivirus that is the causative agent of a major mosquito-borne encephalitis in the world. Evasion of peripheral immune system facilitates the entry of the virus into the central nervous system (CNS) where it causes extensive neuronal inflammatory damage that leads to death or severe neuropschychiatric sequel in survivors. It has been proposed that after entry into the body, the virus is carried into the CNS by peripheral immune cells that act as Trojan horses. In this study we investigate whether macrophages can be considered as such a Trojan horse. We also investigate the role of minocycline, a synthetic tetracycline, in such processes. Minocycline has been found to be broadly protective in neurological disease models featuring inflammation and cell death but there has been no report of it having any modulatory role in peripheral macrophage-mediated immune response against viral infection. Persistence of internalized virus within macrophages was visualized by immunofluorescent staining. Cytotoxicity assay revealed that there was no significant cell death after 24 h and 72 h infection with JEV. Proinflammatory cytokine levels were elevated in cells that were infected with JEV but it was abrogated following minocycline treatment. Reactive oxygen species level was also increased after JEV infection. Nitric oxide level was found to increase after 72 h post infection but remained unchanged after 24h. The cellular levels of signaling molecules such as PI3 kinase, phophoAkt and phospho p38MAP kinase were found to be altered after JEV infection and minocycline treatment. JEV infection also affected the VEGF-MMP pathway. Increased activity of MMP-9 was detected from JEV-infected macrophage culture supernatants after 72 h; minocycline treatment resulted in reduced activity. Thus it seems that minocycline dampens peripheral immune reactions by decreasing proinflammatory cytokine release from infected macrophages and the

  3. The Role of Mcl-1 in S. aureus-Induced Cytoprotection of Infected Macrophages

    Directory of Open Access Journals (Sweden)

    Joanna Koziel

    2013-01-01

    cytoprotection of infected cells leading to apoptosis. Increased MCL1 expression in infected cells was associated with enhanced NFκB activation and subsequent IL-6 secretion, since the inhibition of both NFκB and IL-6 signalling pathways abrogated Mcl-1 induction and cytoprotection. Finally, we confirmed our observation in vivo in murine model of septic arthritis showing the association between the severity of arthritis and Mcl-1 expression. Therefore, we propose that S. aureus is hijacking the Mcl-1-dependent inhibition of apoptosis to prevent the elimination of infected host cells, thus allowing the intracellular persistence of the pathogen, its dissemination by infected macrophages, and the progression of staphylococci diseases.

  4. Involvement of Pro-Inflammatory Macrophages in Liver Pathology of Pirital Virus-Infected Syrian Hamsters

    Directory of Open Access Journals (Sweden)

    Corey L. Campbell

    2018-05-01

    Full Text Available New World arenaviruses cause fatal hemorrhagic disease in South America. Pirital virus (PIRV, a mammarenavirus hosted by Alston’s cotton rat (Sigmodon alstoni, causes a disease in Syrian golden hamsters (Mesocricetus auratus (biosafety level-3, BSL-3 that has many pathologic similarities to the South American hemorrhagic fevers (BSL-4 and, thus, is considered among the best small-animal models for human arenavirus disease. Here, we extend in greater detail previously described clinical and pathological findings in Syrian hamsters and provide evidence for a pro-inflammatory macrophage response during PIRV infection. The liver was the principal target organ of the disease, and signs of Kupffer cell involvement were identified in mortally infected hamster histopathology data. Differential expression analysis of liver mRNA revealed signatures of the pro-inflammatory response, hematologic dysregulation, interferon pathway and other host response pathways, including 17 key transcripts that were also reported in two non-human primate (NHP arenavirus liver-infection models, representing both Old and New World mammarenavirus infections. Although antigen presentation may differ among rodent and NHP species, key hemostatic and innate immune-response components showed expression parallels. Signatures of pro-inflammatory macrophage involvement in PIRV-infected livers included enrichment of Ifng, Nfkb2, Stat1, Irf1, Klf6, Il1b, Cxcl10, and Cxcl11 transcripts. Together, these data indicate that pro-inflammatory macrophage M1 responses likely contribute to the pathogenesis of acute PIRV infection.

  5. Transcriptomic signature of Leishmania infected mice macrophages: a metabolic point of view.

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    Imen Rabhi

    Full Text Available We analyzed the transcriptional signatures of mouse bone marrow-derived macrophages at different times after infection with promastigotes of the protozoan parasite Leishmania major. Ingenuity Pathway Analysis revealed that the macrophage metabolic pathways including carbohydrate and lipid metabolisms were among the most altered pathways at later time points of infection. Indeed, L. major promastiogtes induced increased mRNA levels of the glucose transporter and almost all of the genes associated with glycolysis and lactate dehydrogenase, suggesting a shift to anaerobic glycolysis. On the other hand, L. major promastigotes enhanced the expression of scavenger receptors involved in the uptake of Low-Density Lipoprotein (LDL, inhibited the expression of genes coding for proteins regulating cholesterol efflux, and induced the synthesis of triacylglycerides. These data suggested that Leishmania infection disturbs cholesterol and triglycerides homeostasis and may lead to cholesterol accumulation and foam cell formation. Using Filipin and Bodipy staining, we showed cholesterol and triglycerides accumulation in infected macrophages. Moreover, Bodipy-positive lipid droplets accumulated in close proximity to parasitophorous vacuoles, suggesting that intracellular L. major may take advantage of these organelles as high-energy substrate sources. While the effect of infection on cholesterol accumulation and lipid droplet formation was independent on parasite development, our data indicate that anaerobic glycolysis is actively induced by L. major during the establishment of infection.

  6. Galectin-1 promotes HIV-1 infectivity in macrophages through stabilization of viral adsorption

    International Nuclear Information System (INIS)

    Mercier, Simon; St-Pierre, Christian; Pelletier, Isabelle; Ouellet, Michel; Tremblay, Michel J.; Sato, Sachiko

    2008-01-01

    Following primary infection with human immunodeficiency virus type-1 (HIV-1), macrophages are thought to play an important role, as they are one of the first target cells the virus encounters and can also sustain a significant production of viruses over extended periods of time. While the interaction between the primary cellular receptor CD4 and the virus-encoded external envelope glycoprotein gp120 initiates the infection process, it has been suggested that various host factors are exploited by HIV-1 to facilitate adsorption onto the cell surface. Macrophages and other cells found at the infection site can secrete a soluble mammalian lectin, galectin-1, which binds to β-galactoside residues through its carbohydrate recognition domain. Being a dimer, galectin-1 can cross-link ligands expressed on different constituents to mediate adhesion between cells or between cells and pathogens. We report here that galectin-1, but not galectin-3, increased HIV-1 infectivity in monocyte-derived macrophages (MDMs). This phenomenon was likely due to an enhancement of virus adsorption kinetics, which facilitates HIV-1 entry. The fusion inhibitors T-20 and TAK779 remained effective at reducing infection even in the presence of galectin-1, indicating that the galectin-1-mediated effect is occurring at a step prior to fusion. Together, our data suggest that galectin-1 can facilitate HIV-1 infection in MDMs by promoting early events of the virus replicative cycle (i.e. adsorption)

  7. 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

  8. 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.

  9. [Inclusion Bodies are Formed in SFTSV-infected Human Macrophages].

    Science.gov (United States)

    Jin, Cong; Song, Jingdong; Han, Ying; Li, Chuan; Qiu, Peihong; Liang, Mifang

    2016-01-01

    The severe fever with thrombocytopenia syndrome virus (SFTSV) is a new member in the genus Phlebovirus of the family Bunyaviridae identified in China. The SFTSV is also the causative pathogen of an emerging infectious disease: severe fever with thrombocytopenia syndrome. Using immunofluorescent staining and confocal microscopy, the intracellular distribution of nucleocapsid protein (NP) in SFTSV-infected THP-1 cells was investigated with serial doses of SFTSV at different times after infection. Transmission electron microscopy was used to observe the ultrafine intracellular structure of SFTSV-infected THP-1 cells at different times after infection. SFTSV NP could form intracellular inclusion bodies in infected THP-1 cells. The association between NP-formed inclusion bodies and virus production was analyzed: the size of the inclusion body formed 3 days after infection was correlated with the viral load in supernatants collected 7 days after infection. These findings suggest that the inclusion bodies formed in SFTSV-infected THP-1 cells could be where the SFTSV uses host-cell proteins and intracellular organelles to produce new viral particles.

  10. 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.

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    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.

  11. Antibody-dependent infection of human macrophages by severe acute respiratory syndrome coronavirus.

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    Yip, Ming Shum; Leung, Nancy Hiu Lan; Cheung, Chung Yan; Li, Ping Hung; Lee, Horace Hok Yeung; Daëron, Marc; Peiris, Joseph Sriyal Malik; Bruzzone, Roberto; Jaume, Martial

    2014-05-06

    Public health risks associated to infection by human coronaviruses remain considerable and vaccination is a key option for preventing the resurgence of severe acute respiratory syndrome coronavirus (SARS-CoV). We have previously reported that antibodies elicited by a SARS-CoV vaccine candidate based on recombinant, full-length SARS-CoV Spike-protein trimers, trigger infection of immune cell lines. These observations prompted us to investigate the molecular mechanisms and responses to antibody-mediated infection in human macrophages. We have used primary human immune cells to evaluate their susceptibility to infection by SARS-CoV in the presence of anti-Spike antibodies. Fluorescence microscopy and real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR) were utilized to assess occurrence and consequences of infection. To gain insight into the underlying molecular mechanism, we performed mutational analysis with a series of truncated and chimeric constructs of fragment crystallizable γ receptors (FcγR), which bind antibody-coated pathogens. We show here that anti-Spike immune serum increased infection of human monocyte-derived macrophages by replication-competent SARS-CoV as well as Spike-pseudotyped lentiviral particles (SARS-CoVpp). Macrophages infected with SARS-CoV, however, did not support productive replication of the virus. Purified anti-viral IgGs, but not other soluble factor(s) from heat-inactivated mouse immune serum, were sufficient to enhance infection. Antibody-mediated infection was dependent on signaling-competent members of the human FcγRII family, which were shown to confer susceptibility to otherwise naïve ST486 cells, as binding of immune complexes to cell surface FcγRII was necessary but not sufficient to trigger antibody-dependent enhancement (ADE) of infection. Furthermore, only FcγRII with intact cytoplasmic signaling domains were competent to sustain ADE of SARS-CoVpp infection, thus providing additional

  12. Expression of hsa Let-7a MicroRNA of Macrophages Infected by Leishmania Major

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    Nooshin Hashemi

    2016-10-01

    Full Text Available Leishmaniasis is a vector-born disease caused by species of the genus Leishmania and is transmitted from host to host through the bite of an infected sandfly. MicroRNAs (miRNAs are non-coding small RNAs with 22-nucleotide length. They are involved in some biological and cellular processes. We aimed to evaluate the expression of let-7a in human macrophages miRNA when are infected by Leishmania major. We also evaluated the impact of Leishmania major infection on the expression of let-7a at two different times, 24 and 48 hours, after infection. Blood samples were collected from ten healthy volunteers with no history of leishmaniasis. Development of macrophages from peripheral monocytes and infection with stationary phase of Leishmania major promastigotes were done through serial cultures under 5% CO2 environment and 37C. To measure the expression levels of let-7a real-time PCR was performed with specific related primers using the SYBR® Green master mix Kit™. The real-time PCR showed let-7a was expressed in cells infected with parasites after 24 and 48h post-infection. Comparison of let-7a miRNA expression after 24 and 48 h revealed that let-7a miRNAs were down-regulated at 48 h post-infection more than 24h after infection. The results of this study suggest that according to the main function of miRNA in repression of mRNA translation it could be possible to manipulate host cells in order to alter miRNA levels and regulate macrophage functions after establishment of intracellular parasites such as Leishmania.

  13. Macrophage Expression of Inflammatory Genes in Response to EMCV Infection

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    Zachary R. Shaheen

    2015-08-01

    Full Text Available The expression and production of type 1 interferon is the classic cellular response to virus infection. In addition to this antiviral response, virus infection also stimulates the production of proinflammatory mediators. In this review, the pathways controlling the induction of inflammatory genes and the roles that these inflammatory mediators contribute to host defense against viral pathogens will be discussed. Specific focus will be on the role of the chemokine receptor CCR5, as a signaling receptor controlling the activation of pathways leading to virus-induced inflammatory gene expression.

  14. Mycobacterium leprae-Infected Macrophages Preferentially Primed Regulatory T Cell Responses and Was Associated with Lepromatous Leprosy.

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    Degang Yang

    2016-01-01

    Full Text Available The persistence of Mycobacterium leprae (M. leprae infection is largely dependent on the types of host immune responses being induced. Macrophage, a crucial modulator of innate and adaptive immune responses, could be directly infected by M. leprae. We therefore postulated that M. leprae-infected macrophages might have altered immune functions.Here, we treated monocyte-derived macrophages with live or killed M. leprae, and examined their activation status and antigen presentation. We found that macrophages treated with live M. leprae showed committed M2-like function, with decreased interleukin 1 beta (IL-1beta, IL-6, tumor necrosis factor alpha (TNF-alpha and MHC class II molecule expression and elevated IL-10 and CD163 expression. When incubating with naive T cells, macrophages treated with live M. leprae preferentially primed regulatory T (Treg cell responses with elevated FoxP3 and IL-10 expression, while interferon gamma (IFN-gamma expression and CD8+ T cell cytotoxicity were reduced. Chromium release assay also found that live M. leprae-treated macrophages were more resistant to CD8+ T cell-mediated cytotoxicity than sonicated M. leprae-treated monocytes. Ex vivo studies showed that the phenotype and function of monocytes and macrophages had clear differences between L-lep and T-lep patients, consistent with the in vitro findings.Together, our data demonstrate that M. leprae could utilize infected macrophages by two mechanisms: firstly, M. leprae-infected macrophages preferentially primed Treg but not Th1 or cytotoxic T cell responses; secondly, M. leprae-infected macrophages were more effective at evading CD8+ T cell-mediated cytotoxicity.

  15. Lipid Droplet Formation, Their Localization and Dynamics during Leishmania major Macrophage Infection.

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    Sameh Rabhi

    Full Text Available Leishmania, the causative agent of vector-borne diseases, known as leishmaniases, is an obligate intracellular parasite within mammalian hosts. The outcome of infection depends largely on the activation status of macrophages, the first line of mammalian defense and the major target cells for parasite replication. Understanding the strategies developed by the parasite to circumvent macrophage defense mechanisms and to survive within those cells help defining novel therapeutic approaches for leishmaniasis. We previously showed the formation of lipid droplets (LDs in L. major infected macrophages. Here, we provide novel insights on the origin of the formed LDs by determining their cellular distribution and to what extent these high-energy sources are directed to the proximity of Leishmania parasites. We show that the ability of L. major to trigger macrophage LD accumulation is independent of parasite viability and uptake and can also be observed in non-infected cells through paracrine stimuli suggesting that LD formation is from cellular origin. The accumulation of LDs is demonstrated using confocal microscopy and live-cell imagin in parasite-free cytoplasmic region of the host cell, but also promptly recruited to the proximity of Leishmania parasites. Indeed LDs are observed inside parasitophorous vacuole and in parasite cytoplasm suggesting that Leishmania parasites besides producing their own LDs, may take advantage of these high energy sources. Otherwise, these LDs may help cells defending against parasitic infection. These metabolic changes, rising as common features during the last years, occur in host cells infected by a large number of pathogens and seem to play an important role in pathogenesis. Understanding how Leishmania parasites and different pathogens exploit this LD accumulation will help us define the common mechanism used by these different pathogens to manipulate and/or take advantage of this high-energy source.

  16. Changes in lymphocyte and macrophage subsets due to morphine and ethanol treatment during a retrovirus infection causing murine AIDS

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    Watson, R.R.; Prabhala, R.H.; Darban, H.R.; Yahya, M.D.; Smith, T.L.

    1988-01-01

    The number of lymphocytes of various subsets were not significantly changed by the ethanol exposure except those showing activation markers which were reduced. The percentage of peripheral blood cells showing markers for macrophage functions and their activation were significantly reduced after binge use of ethanol. Ethanol retarded suppression of cells by retroviral infection. However by 25 weeks of infection there was a 8.6% survival in the ethanol fed mice infected with retrovirus which was much less than virally infected controls. Morphine treatment also increased the percentage of cells with markers for macrophages and activated macrophages in virally infected mice, while suppressing them in uninfected mice. The second and third morphine injection series suppressed lymphocyte T-helper and T-suppressor cells, but not total T cells. However, suppression by morphine was significantly less during retroviral disease than suppression caused by the virus only. At 25 weeks of infection 44.8% of morphine treated, infected mice survived.

  17. Viable group A streptococci in macrophages during acute soft tissue infection.

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    Pontus Thulin

    2006-03-01

    Full Text Available Group A streptococcal severe soft tissue infections, such as necrotizing fasciitis, are rapidly progressive infections associated with high mortality. Group A streptococcus is typically considered an extracellular pathogen, but has been shown to reside intracellularly in host cells.We characterized in vivo interactions between group A streptococci (GAS and cells involved in innate immune responses, using human biopsies (n = 70 collected from 17 patients with soft tissue infections. Immunostaining and in situ image analysis revealed high amounts of bacteria in the biopsies, even in those collected after prolonged antibiotic therapy. Viability of the streptococci was assessed by use of a bacterial viability stain, which demonstrated viable bacteria in 74% of the biopsies. GAS were present both extracellularly and intracellularly within phagocytic cells, primarily within macrophages. Intracellular GAS were predominantly noted in biopsies from newly involved tissue characterized by lower inflammation and bacterial load, whereas purely extracellular GAS or a combination of intra- and extracellular GAS dominated in severely inflamed tissue. The latter tissue was also associated with a significantly increased amount of the cysteine protease streptococcal pyrogenic exotoxin SpeB. In vitro studies confirmed that macrophages serve as reservoirs for viable GAS, and infection with a speB-deletion mutant produced significantly lower frequencies of cells with viable GAS following infection as compared to the wild-type bacteria.This is the first study to demonstrate that GAS survive intracellularly in macrophages during acute invasive infections. This intracellular presence may have evolved as a mechanism to avoid antibiotic eradication, which may explain our finding that high bacterial load is present even in tissue collected after prolonged intravenous antibiotic therapy. This new insight into the pathogenesis of streptococcal soft tissue infections

  18. Viable Group A Streptococci in Macrophages during Acute Soft Tissue Infection.

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    2006-01-01

    Full Text Available BACKGROUND: Group A streptococcal severe soft tissue infections, such as necrotizing fasciitis, are rapidly progressive infections associated with high mortality. Group A streptococcus is typically considered an extracellular pathogen, but has been shown to reside intracellularly in host cells. METHODS AND FINDINGS: We characterized in vivo interactions between group A streptococci (GAS and cells involved in innate immune responses, using human biopsies (n = 70 collected from 17 patients with soft tissue infections. Immunostaining and in situ image analysis revealed high amounts of bacteria in the biopsies, even in those collected after prolonged antibiotic therapy. Viability of the streptococci was assessed by use of a bacterial viability stain, which demonstrated viable bacteria in 74% of the biopsies. GAS were present both extracellularly and intracellularly within phagocytic cells, primarily within macrophages. Intracellular GAS were predominantly noted in biopsies from newly involved tissue characterized by lower inflammation and bacterial load, whereas purely extracellular GAS or a combination of intra- and extracellular GAS dominated in severely inflamed tissue. The latter tissue was also associated with a significantly increased amount of the cysteine protease streptococcal pyrogenic exotoxin SpeB. In vitro studies confirmed that macrophages serve as reservoirs for viable GAS, and infection with a speB-deletion mutant produced significantly lower frequencies of cells with viable GAS following infection as compared to the wild-type bacteria. CONCLUSIONS: This is the first study to demonstrate that GAS survive intracellularly in macrophages during acute invasive infections. This intracellular presence may have evolved as a mechanism to avoid antibiotic eradication, which may explain our finding that high bacterial load is present even in tissue collected after prolonged intravenous antibiotic therapy. This new insight into the pathogenesis

  19. Role of macrophages in the altered epithelial function during a type 2 immune response induced by enteric nematode infection.

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    Luigi Notari

    Full Text Available Parasitic enteric nematodes induce a type 2 immune response characterized by increased production of Th2 cytokines, IL-4 and IL-13, and recruitment of alternatively activated macrophages (M2 to the site of infection. Nematode infection is associated with changes in epithelial permeability and inhibition of sodium-linked glucose absorption, but the role of M2 in these effects is unknown. Clodronate-containing liposomes were administered prior to and during nematode infection to deplete macrophages and prevent the development of M2 in response to infection with Nippostrongylus brasiliensis. The inhibition of epithelial glucose absorption that is associated with nematode infection involved a macrophage-dependent reduction in SGLT1 activity, with no change in receptor expression, and a macrophage-independent down-regulation of GLUT2 expression. The reduced transport of glucose into the enterocyte is compensated partially by an up-regulation of the constitutive GLUT1 transporter consistent with stress-induced activation of HIF-1α. Thus, nematode infection results in a "lean" epithelial phenotype that features decreased SGLT1 activity, decreased expression of GLUT2 and an emergent dependence on GLUT1 for glucose uptake into the enterocyte. Macrophages do not play a role in enteric nematode infection-induced changes in epithelial barrier function. There is a greater contribution, however, of paracellular absorption of glucose to supply the energy demands of host resistance. These data provide further evidence of the ability of macrophages to alter glucose metabolism of neighboring cells.

  20. The Nuclear Receptor LXR Limits Bacterial Infection of Host Macrophages through a Mechanism that Impacts Cellular NAD Metabolism

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    Jonathan Matalonga

    2017-01-01

    Full Text Available Macrophages exert potent effector functions against invading microorganisms but constitute, paradoxically, a preferential niche for many bacterial strains to replicate. Using a model of infection by Salmonella Typhimurium, we have identified a molecular mechanism regulated by the nuclear receptor LXR that limits infection of host macrophages through transcriptional activation of the multifunctional enzyme CD38. LXR agonists reduced the intracellular levels of NAD+ in a CD38-dependent manner, counteracting pathogen-induced changes in macrophage morphology and the distribution of the F-actin cytoskeleton and reducing the capability of non-opsonized Salmonella to infect macrophages. Remarkably, pharmacological treatment with an LXR agonist ameliorated clinical signs associated with Salmonella infection in vivo, and these effects were dependent on CD38 expression in bone-marrow-derived cells. Altogether, this work reveals an unappreciated role for CD38 in bacterial-host cell interaction that can be pharmacologically exploited by activation of the LXR pathway.

  1. Differential expression of miRNAs by macrophages infected with virulent and avirulent Mycobacterium tuberculosis.

    Science.gov (United States)

    Das, Kishore; Saikolappan, Sankaralingam; Dhandayuthapani, Subramanian

    2013-12-01

    MicroRNAs (miRNAs) are small non-coding RNAs which post-transcriptionally regulate a wide range of biological processes that include cellular differentiation, development, immunity and apoptosis. There is a growing body of evidences that bacteria modulate immune responses by altering the expression of host miRNAs. Since macrophages are immune cells associated with innate and adaptive immunity, we investigated whether Mycobacterium tuberculosis infection affects miRNAs of macrophages. THP-1 macrophages infected with virulent (H37Rv) and avirulent (H37Ra) strains of M. tuberculosis were analyzed for changes in miRNAs' expression using microarray. This revealed that nine miRNA genes (miR-30a, miR-30e, miR-155, miR-1275, miR-3665, miR-3178, miR-4484, miR-4668-5p and miR-4497) were differentially expressed between THP-1cells infected with M. tuberculosis H37Rv and M. tuberculosis H37Ra strains. Additional characterization of these genes is likely to provide insights into their role in the pathogenesis of tuberculosis. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Isolation of F. novicida-Containing Phagosome from Infected Human Monocyte Derived Macrophages

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    Valentina Marecic

    2017-07-01

    Full Text Available Francisella is a gram-negative bacterial pathogen, which causes tularemia in humans and animals. A crucial step of Francisella infection is its invasion of macrophage cells. Biogenesis of the Francisella-containing phagosome (FCP is arrested for ~15 min at the endosomal stage, followed by gradual bacterial escape into the cytosol, where the microbe proliferates. The crucial step in pathogenesis of tularemia is short and transient presence of the bacterium within phagosome. Isolation of FCPs for further studies has been challenging due to the short period of time of bacterial residence in it and the characteristics of the FCP. Here, we will for the first time present the method for isolation of the FCPs from infected human monocytes-derived macrophages (hMDMs. For elimination of lysosomal compartment these organelles were pre-loaded with dextran coated colloidal iron particles prior infection and eliminated by magnetic separation of the post-nuclear supernatant (PNS. We encountered the challenge that mitochondria has similar density to the FCP. To separate the FCP in the PNS from mitochondria, we utilized iodophenylnitrophenyltetrazolium, which is converted by the mitochondrial succinate dehydrogenase into formazan, leading to increased density of the mitochondria and allowing separation by the discontinuous sucrose density gradient ultracentrifugation. The purity of the FCP preparation and its acquisition of early endosomal markers was confirmed by Western blots, confocal and transmission electron microscopy. Our strategy to isolate highly pure FCPs from macrophages should facilitate studies on the FCP and its biogenesis.

  3. Trojan Horse macrophages: studies with the murine lactate dehydrogenase-elevating virus and implications for sexually transmitted virus infection.

    Science.gov (United States)

    Cafruny, W A; Bradley, S E

    1996-12-01

    Previous studies have suggested that monocytes or macrophages may mediate internal virus spread. For the present study, the tissue distribution and infectious potential of dye-labelled and/or lactate dehydrogenase-elevating virus (LDV)-infected murine macrophages were determined. Murine peritoneal macrophages were labelled with the fluorescent carbocyanine tracking dye Dil, injected into mice, and the tissue distribution of Dil-labelled cells was determined by fluorescence analysis of frozen sections. Mice receiving intravenous (i.v.) or intraperitoneal injections of Dil-labelled macrophages displayed rapid and broad tissue distribution of the labelled cells. Intravaginal injection of Dil-labelled macrophages resulted in penetration into the placentas, but not the fetuses, of pregnant mice. When macrophages were LDV-infected and Dil-labelled prior to i.v. injection into pregnant mice, they homed to various tissues including the placenta, but were not found in fetuses. Intravaginal injection of LDV-infected macrophages resulted in systemic LDV infection, even though the free-virus dose was less than the minimum infectious dose by this route. Neither polyclonal nor monoclonal IgG anti-LDV antibodies protected mice from vaginal infection with cell-associated virus, and LDV-immune complexes were themselves infectious by the vaginal route. These results show that exogenous macrophages are widely distributed following parenteral injection, penetrate locally to placentas after intravaginal injection, and are capable of acting vaginally as relatively efficient virus infection-delivery vehicles. Thus, 'Trojan Horse' macrophages are potentially infectious vehicles both for internal virus spread and for animal-to-animal transmission.

  4. Rhadinovirus host entry by co-operative infection.

    Science.gov (United States)

    Lawler, Clara; Milho, Ricardo; May, Janet S; Stevenson, Philip G

    2015-03-01

    Rhadinoviruses establish chronic infections of clinical and economic importance. Several show respiratory transmission and cause lung pathologies. We used Murid Herpesvirus-4 (MuHV-4) to understand how rhadinovirus lung infection might work. A primary epithelial or B cell infection often is assumed. MuHV-4 targeted instead alveolar macrophages, and their depletion reduced markedly host entry. While host entry was efficient, alveolar macrophages lacked heparan - an important rhadinovirus binding target - and were infected poorly ex vivo. In situ analysis revealed that virions bound initially not to macrophages but to heparan⁺ type 1 alveolar epithelial cells (AECs). Although epithelial cell lines endocytose MuHV-4 readily in vitro, AECs did not. Rather bound virions were acquired by macrophages; epithelial infection occurred only later. Thus, host entry was co-operative - virion binding to epithelial cells licensed macrophage infection, and this in turn licensed AEC infection. An antibody block of epithelial cell binding failed to block host entry: opsonization provided merely another route to macrophages. By contrast an antibody block of membrane fusion was effective. Therefore co-operative infection extended viral tropism beyond the normal paradigm of a target cell infected readily in vitro; and macrophage involvement in host entry required neutralization to act down-stream of cell binding.

  5. Nicotine Impairs Macrophage Control of Mycobacterium tuberculosis.

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

    2017-09-01

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

  6. Blood and milk polymorphonuclear leukocyte and monocyte/macrophage functions in naturally caprine arthritis encephalitis virus infection in dairy goats.

    Science.gov (United States)

    Santos, Bruna Parapinski; Souza, Fernando Nogueira; Blagitz, Maiara Garcia; Batista, Camila Freitas; Bertagnon, Heloísa Godoi; Diniz, Soraia Araújo; Silva, Marcos Xavier; Haddad, João Paulo Amaral; Della Libera, Alice Maria Melville Paiva

    2017-06-01

    The exact influence of caprine arthritis encephalitis virus (CAEV) infection on blood and milk polymorphonuclear leukocytes (PMNLs) and monocyte/macrophages of goats remains unclear. Thus, the present study sought to explore the blood and milk PMNL and monocyte/macrophage functions in naturally CAEV-infected goats. The present study used 18 healthy Saanen goats that were segregated according to sera test outcomes into serologically CAEV negative (n=8; 14 halves) and positive (n=10; 14 halves) groups. All milk samples from mammary halves with milk bacteriologically positive outcomes, somatic cell count ≥2×10 6 cellsmL -1 , and abnormal secretions in the strip cup test were excluded. We evaluated the percentage of blood and milk PMNLs and monocyte/macrophages, the viability of PMNLs and monocyte/macrophages, the levels of intracellular reactive oxygen species (ROS) and the nonopsonized phagocytosis of Staphylococcus aureus and Escherichia coli by flow cytometry. In the present study, a higher percentage of milk macrophages (CD14 + ) and milk polymorphonuclear leukocytes undergoing late apoptosis or necrosis (Annexin-V + /Propidium iodide + ) was observed in CAEV-infected goats; we did not find any further alterations in blood and milk PMNL and monocyte/macrophage functions. Thus, regarding our results, the goats naturally infected with CAEV did not reveal pronounced dysfunctions in blood and milk polymorphonuclear leukocytes and monocytes/macrophages. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. CP and CP-PGN protect mice against MRSA infection by inducing M1 macrophages.

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    Zhang, Yang; Li, Xiang-Xiang; Ma, Yuan; Xu, Jie; Zhao, Li-Na; Qian, Xue-Feng; Zhang, Xian-Feng; Shi, Jin-Fang; Han, Qing-Zhen

    2017-12-04

    Corynebacterium pyruviciproducens (C. pyruviciproducens, CP), as a newly discovered immunomodulator, has been confirmed to have a stronger immunoregulation than Propionibacterium acnes (P. acnes) of the traditional immune adjuvant, by previous experiments with model antigen ovalbumin and sheep red blood cells. Here, it was designed to assess its ability to resist methicillin-resistant Staphylococcus aureus (MRSA), since MRSA as a vital gram positive pathogen is characterized by high morbidity and mortality. In this report, it was indicated that C. pyruviciproducens and its peptidoglycan (CP-PGN) could help to be against bloodstream infection of MRSA with raised survival rate, decreased bacteria load and alleviated systemic inflammation, and these effects of CP-PGN were more pronounced. However, the whole CP was inclined to prevent localized abdominal infection of MRSA from progressing to a systemic infection. And they showed the potential as a therapeutic drug alone or combined with vancomycin. The diversity of capacity of activating macrophages induced by CP and CP-PGN may result in distinct resistance to MRSA in different infection models. Furthermore, both CP and CP-PGN induced M1 macrophages. In conclusion, CP and its PGN could act as promising immune agents to treat and prevent MRSA infection.

  8. Kinetics of liver macrophages (Kupffer cells) in SIV-infected macaques

    International Nuclear Information System (INIS)

    Ahsan, Muhammad H.; Gill, Amy F.; Alvarez, Xavier; Lackner, Andrew A.; Veazey, Ronald S.

    2013-01-01

    Since the liver drains antigens from the intestinal tract, and since the intestinal tract is a major site of viral replication, we examined the dynamics of liver macrophages (Kupffer cells) throughout SIV infection. Absolute numbers of Kupffer cells increased in the livers in acute infection, and in animals with AIDS. Significantly higher percentages of proliferating (BrdU+) Kupffer cells were detected in acute infection and in AIDS with similar trends in blood monocytes. Significantly higher percentages of apoptotic (AC3+) Kupffer cells were also found in acute and AIDS stages. However, productively infected cells were not detected in liver of 41/42 animals examined, despite abundant infected cells in gut and lymph nodes of all animals. Increased rates of Kupffer cell proliferation resulting in an increase in Kupffer cells without productive infection indicate SIV infection affects Kupffer cells, but the liver does not appear to be a major site of productive viral replication. - Highlights: • Kupffer cells increase in the liver of SIV-infected macaques. • Increased proliferation and apoptosis of Kupffer cells occurs in SIV infection. • Productively infected cells are rarely detected in the liver. • The liver is not a major site for SIV replication

  9. HIV-associated fulminating herpes zoster infection with alveolar necrosis and tooth exfoliation: a case report.

    Science.gov (United States)

    Chindia, M L

    1997-04-01

    This paper presents a case of HIV-associated fulminating herpes zoster infection (HZI) that culminated in right mandibular necrosis and tooth exfoliation. The occurrence of such infection in immunosuppression and the impending clinical features are briefly reviewed and discussed.

  10. Caspase-1-dependent and -independent cell death pathways in Burkholderia pseudomallei infection of macrophages.

    Directory of Open Access Journals (Sweden)

    Antje Bast

    2014-03-01

    Full Text Available The cytosolic pathogen Burkholderia pseudomallei and causative agent of melioidosis has been shown to regulate IL-1β and IL-18 production through NOD-like receptor NLRP3 and pyroptosis via NLRC4. Downstream signalling pathways of those receptors and other cell death mechanisms induced during B. pseudomallei infection have not been addressed so far in detail. Furthermore, the role of B. pseudomallei factors in inflammasome activation is still ill defined. In the present study we show that caspase-1 processing and pyroptosis is exclusively dependent on NLRC4, but not on NLRP3 in the early phase of macrophage infection, whereas at later time points caspase-1 activation and cell death is NLRC4- independent. In the early phase we identified an activation pathway involving caspases-9, -7 and PARP downstream of NLRC4 and caspase-1. Analyses of caspase-1/11-deficient infected macrophages revealed a strong induction of apoptosis, which is dependent on activation of apoptotic initiator and effector caspases. The early activation pathway of caspase-1 in macrophages was markedly reduced or completely abolished after infection with a B. pseudomallei flagellin FliC or a T3SS3 BsaU mutant. Studies using cells transfected with the wild-type and mutated T3SS3 effector protein BopE indicated also a role of this protein in caspase-1 processing. A T3SS3 inner rod protein BsaK mutant failed to activate caspase-1, revealed higher intracellular counts, reduced cell death and IL-1β secretion during early but not during late macrophage infection compared to the wild-type. Intranasal infection of BALB/c mice with the BsaK mutant displayed a strongly decreased mortality, lower bacterial loads in organs, and reduced levels of IL-1β, myeloperoxidase and neutrophils in bronchoalveolar lavage fluid. In conclusion, our results indicate a major role for a functional T3SS3 in early NLRC4-mediated caspase-1 activation and pyroptosis and a contribution of late caspase-1

  11. Expression of Nocardia brasiliensis superoxide dismutase during the early infection of murine peritoneal macrophages.

    Science.gov (United States)

    Revol, Agnès; Espinoza-Ruiz, Marisol; Medina-Villanueva, Igor; Salinas-Carmona, Mario Cesar

    2006-12-01

    Nocardia brasiliensis is the main agent of actinomycetoma in Mexico, but little is known about its virulence and molecular pathogenic pathways. These facultative intracellular bacteria are able to survive and divide within the host phagocytic cells, in part by neutralizing the reactive oxygen intermediates. Superoxide dismutase (SOD) participates in the intracellular survival of several bacterial species and, in particular, constitutes one of Nocardia asteroides virulence factors. To clarify SOD participation in the N. brasiliensis early infective process, we report its isolation and the consequent comparison of its transcript level. A 630 bp polymerase chain reaction fragment that included most of the coding sequence of N. brasiliensis sodA was cloned. A competitive assay was developed, allowing comparison of bacterial sod expression in exponential culture and 1 h after infecting peritoneal macrophages from BALB/c mice. At that time, there were viable bacteria in the macrophages. The intracellular bacteria presented a clear decrease in their sod transcript amount, although their 16S rRNA (used as an internal control) and hsp levels were maintained or slightly increased, respectively. These results indicate that sodA transcription is not maintained within the SOS bacterial response induced by phagosomal conditions. Further kinetics will be necessary to precisely define sod transcriptional regulation during N. brasiliensis intra-macrophage growth.

  12. Mammalian Target of Rapamycin Inhibition in Trypanosoma cruzi-Infected Macrophages Leads to an Intracellular Profile That Is Detrimental for Infection

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    Jorge David Rojas Márquez

    2018-02-01

    Full Text Available The causative agent of Chagas’ disease, Trypanosoma cruzi, affects approximately 10 million people living mainly in Latin America, with macrophages being one of the first cellular actors confronting the invasion during T. cruzi infection and their function depending on their proper activation and polarization into distinct M1 and M2 subtypes. Macrophage polarization is thought to be regulated not only by cytokines and growth factors but also by environmental signals. The metabolic checkpoint kinase mammalian target of rapamycin (mTOR-mediated sensing of environmental and metabolic cues influences macrophage polarization in a complex and as of yet incompletely understood manner. Here, we studied the role of the mTOR pathway in macrophages during T. cruzi infection. We demonstrated that the parasite activated mTOR, which was beneficial for its replication since inhibition of mTOR in macrophages by different inhibitors decreased parasite replication. Moreover, in rapamycin pretreated and infected macrophages, we observed a decreased arginase activity and expression, reduced IL-10 and increased interleukin-12 production, compared to control infected macrophages treated with DMSO. Surprisingly, we also found a reduced iNOS activity and expression in these macrophages. Therefore, we investigated possible alternative mechanisms involved in controlling parasite replication in rapamycin pretreated and infected macrophages. Although, cytoplasmic ROS and the enzyme indoleamine 2, 3-dioxygenase (IDO were not involved, we observed a significant increase in IL-6, TNF-α, and IL-1β production. Taking into account that IL-1β is produced by activation of the cytoplasmic receptor NLRP3, which is one of the main components of the inflammasome, we evaluated NLRP3 expression during mTOR inhibition and T. cruzi infection. We observed that rapamycin-pretreated and infected macrophages showed a significant increase in NLRP3 expression and produced higher levels of

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

    DEFF Research Database (Denmark)

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

    2010-01-01

    that siderocalin expression is upregulated following M.tb infection of mouse macrophage cell lines and primary murine alveolar macrophages. Furthermore, siderocalin added exogenously as a recombinant protein or overexpressed in the RAW264.7 macrophage cell line inhibited the intracellular growth of the pathogen......Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show....... A variant form of siderocalin, which is expressed only in the macrophage cytosol, inhibited intracellular M.tb growth as effectively as the normal, secreted form, an observation that provides mechanistic insight into how siderocalin might influence iron acquisition by the bacteria in the phagosome. Our...

  14. Sympathetic glial cells and macrophages develop different responses to Trypanosoma cruzi infection or lipopolysaccharide stimulation.

    Science.gov (United States)

    de Almeida-Leite, Camila Megale; Silva, Isabel Cristina Costa; Galvão, Lúcia Maria da Cunha; Arantes, Rosa Maria Esteves

    2014-07-01

    Nitric oxide (NO) participates in neuronal lesions in the digestive form of Chagas disease and the proximity of parasitised glial cells and neurons in damaged myenteric ganglia is a frequent finding. Glial cells have crucial roles in many neuropathological situations and are potential sources of NO. Here, we investigate peripheral glial cell response to Trypanosoma cruzi infection to clarify the role of these cells in the neuronal lesion pathogenesis of Chagas disease. We used primary glial cell cultures from superior cervical ganglion to investigate cell activation and NO production after T. cruzi infection or lipopolysaccharide (LPS) exposure in comparison to peritoneal macrophages. T. cruzi infection was greater in glial cells, despite similar levels of NO production in both cell types. Glial cells responded similarly to T. cruzi and LPS, but were less responsive to LPS than macrophages were. Our observations contribute to the understanding of Chagas disease pathogenesis, as based on the high susceptibility of autonomic glial cells to T. cruzi infection with subsequent NO production. Moreover, our findings will facilitate future research into the immune responses and activation mechanisms of peripheral glial cells, which are important for understanding the paradoxical responses of this cell type in neuronal lesions and neuroprotection.

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

    Science.gov (United States)

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

    2017-09-01

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

  16. Regulation of Viral Replication, Apoptosis and Pro-Inflammatory Responses by 17-AAG during Chikungunya Virus Infection in Macrophages

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    Tapas K. Nayak

    2017-01-01

    Full Text Available Chikungunya virus (CHIKV infection has re-emerged as a major public health concern due to its recent worldwide epidemics and lack of control measures. Although CHIKV is known to infect macrophages, regulation of CHIKV replication, apoptosis and immune responses towards macrophages are not well understood. Accordingly, the Raw264.7 cells, a mouse macrophage cell line, were infected with CHIKV and viral replication as well as new viral progeny release was assessed by flow cytometry and plaque assay, respectively. Moreover, host immune modulation and apoptosis were studied through flow cytometry, Western blot and ELISA. Our current findings suggest that expression of CHIKV proteins were maximum at 8 hpi and the release of new viral progenies were remarkably increased around 12 hpi. The induction of Annexin V binding, cleaved caspase-3, cleaved caspase-9 and cleaved caspase-8 in CHIKV infected macrophages suggests activation of apoptosis through both intrinsic and extrinsic pathways. The pro-inflammatory mediators (TNF and IL-6 MHC-I/II and B7.2 (CD86 were also up-regulated during infection over time. Further, 17-AAG, a potential HSP90 inhibitor, was found to regulate CHIKV infection, apoptosis and pro-inflammatory cytokine/chemokine productions of host macrophages significantly. Hence, the present findings might bring new insight into the therapeutic implication in CHIKV disease biology.

  17. Regulation of Viral Replication, Apoptosis and Pro-Inflammatory Responses by 17-AAG during Chikungunya Virus Infection in Macrophages.

    Science.gov (United States)

    Nayak, Tapas K; Mamidi, Prabhudutta; Kumar, Abhishek; Singh, Laishram Pradeep K; Sahoo, Subhransu S; Chattopadhyay, Soma; Chattopadhyay, Subhasis

    2017-01-06

    Chikungunya virus (CHIKV) infection has re-emerged as a major public health concern due to its recent worldwide epidemics and lack of control measures. Although CHIKV is known to infect macrophages, regulation of CHIKV replication, apoptosis and immune responses towards macrophages are not well understood. Accordingly, the Raw264.7 cells, a mouse macrophage cell line, were infected with CHIKV and viral replication as well as new viral progeny release was assessed by flow cytometry and plaque assay, respectively. Moreover, host immune modulation and apoptosis were studied through flow cytometry, Western blot and ELISA. Our current findings suggest that expression of CHIKV proteins were maximum at 8 hpi and the release of new viral progenies were remarkably increased around 12 hpi. The induction of Annexin V binding, cleaved caspase-3, cleaved caspase-9 and cleaved caspase-8 in CHIKV infected macrophages suggests activation of apoptosis through both intrinsic and extrinsic pathways. The pro-inflammatory mediators (TNF and IL-6) MHC-I/II and B7.2 (CD86) were also up-regulated during infection over time. Further, 17-AAG, a potential HSP90 inhibitor, was found to regulate CHIKV infection, apoptosis and pro-inflammatory cytokine/chemokine productions of host macrophages significantly. Hence, the present findings might bring new insight into the therapeutic implication in CHIKV disease biology.

  18. Protective Role of Toll-like Receptor 3-Induced Type I Interferon in Murine Coronavirus Infection of Macrophages

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    Sonia Navas-Martin

    2012-05-01

    Full Text Available Toll-like Receptors (TLRs sense viral infections and induce production of type I interferons (IFNs, other cytokines, and chemokines. Viral recognition by TLRs and other pattern recognition receptors (PRRs has been proven to be cell-type specific. Triggering of TLRs with selected ligands can be beneficial against some viral infections. Macrophages are antigen-presenting cells that express TLRs and have a key role in the innate and adaptive immunity against viruses. Coronaviruses (CoVs are single-stranded, positive-sense RNA viruses that cause acute and chronic infections and can productively infect macrophages. Investigation of the interplay between CoVs and PRRs is in its infancy. We assessed the effect of triggering TLR2, TLR3, TLR4, and TLR7 with selected ligands on the susceptibility of the J774A.1 macrophage cell line to infection with murine coronavirus (mouse hepatitis virus, [MHV]. Stimulation of TLR2, TLR4, or TLR7 did not affect MHV production. In contrast, pre-stimulation of TLR3 with polyinosinic-polycytidylic acid (poly I:C hindered MHV infection through induction of IFN-β in macrophages. We demonstrate that activation of TLR3 with the synthetic ligand poly I:C mediates antiviral immunity that diminishes (MHV-A59 or suppresses (MHV-JHM, MHV-3 virus production in macrophages.

  19. 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

  20. Macrophage migration inhibitory factor in cerebrospinal fluid from patients with central nervous system infection

    DEFF Research Database (Denmark)

    Ostergaard, Christian; Benfield, Thomas

    2009-01-01

    .01). Among patients with purulent meningitis, CSF MIF levels were significantly higher in patients infected with pneumococci as compared to infection due to meningococci (11569 ng/L (8615-21935) vs. 5006 ng/L (1717-10905) respectively, P=0.02), in patients requiring assisted ventilation (10493 ng/L (5961-22725......ABSTRACT: INTRODUCTION: Macrophage Migration Inhibitory Factor (MIF) plays an essential pathophysiological role in septic shock; however, its role in central nervous system infection (CNS) remains to be defined. METHODS: The aim of the present study was to investigate cerebrospinal fluid (CSF......) levels of MIF in 171 patients clinically suspected of having meningitis on admission. Of these, 31 were found to have purulent meningitis with a known aetiology, 20 to have purulent meningitis with an unknown aetiology, 59 to have lymphocytic meningitis, and 11 to have encephalitis, whereas 50 were...

  1. Modulation of macrophage cytokine profiles during solid tumor progression: susceptibility to Candida albicans infection

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    Venturini James

    2009-06-01

    Full Text Available Abstract Background In order to attain a better understanding of the interactions between opportunist fungi and their hosts, we investigated the cytokine profile associated with the inflammatory response to Candida albicans infection in mice with solid Ehrlich tumors of different degrees. Methods Groups of eight animals were inoculated intraperitoneally with 5 × 106 C. albicans 7, 14 or 21 days after tumor implantation. After 24 or 72 hours, the animals were euthanized and intraperitoneal lavage fluid was collected. Peritoneal macrophages were cultivated and the levels of IFN-γ, TNF-α, IL-12, IL-10 and IL-4 released into the supernatants were measured by ELISA. Kidney, liver and spleen samples were evaluated for fungal dissemination. Tumor-free animals and animals that had only been subjected to C. albicans infection were used as control groups. Results Our results demonstrated that the mice produced more IFN-γ and TNF-α and less IL-10, and also exhibited fungal clearance, at the beginning of tumor evolution. With the tumor progression, this picture changed: IL-10 production increased and IFN-γ and TNF-α release decreased; furthermore, there was extensive fungal dissemination. Conclusion Our results indicate that solid tumors can affect the production of macrophage cytokines and, in consequence, affect host resistance to opportunistic infections.

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

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    Devyn D Gillette

    2014-02-01

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

  3. Macrophage-tropic variants initiate human immunodeficiency virus type 1 infection after sexual, parenteral, and vertical transmission

    NARCIS (Netherlands)

    van't Wout, A. B.; Kootstra, N. A.; Mulder-Kampinga, G. A.; Albrecht-van Lent, N.; Scherpbier, H. J.; Veenstra, J.; Boer, K.; Coutinho, R. A.; Miedema, F.; Schuitemaker, H.

    1994-01-01

    Macrophage-tropic, non-syncytium-inducing, HIV-1 variants predominate in the asymptomatic phase of infection and may be responsible for establishing infection in an individual exposed to the mixture of HIV-1 variants. Here, genotypical and phenotypical characteristics of virus populations, present

  4. Intracellular activity of tedizolid phosphate and ACH-702 versus Mycobacterium tuberculosis infected macrophages.

    Science.gov (United States)

    Molina-Torres, Carmen A; Barba-Marines, Alejandra; Valles-Guerra, Orestes; Ocampo-Candiani, Jorge; Cavazos-Rocha, Norma; Pucci, Michael J; Castro-Garza, Jorge; Vera-Cabrera, Lucio

    2014-04-04

    Due to the emergency of multidrug-resistant strains of Mycobacterium tuberculosis, is necessary the evaluation of new compounds. Tedizolid, a novel oxazolidinone, and ACH-702, a new isothiazoloquinolone, were tested against M. tuberculosis infected THP-1 macrophages. These two compounds significantly decreased the number of intracellular mycobacteria at 0.25X, 1X, 4X and 16X the MIC value. The drugs were tested either in nanoparticules or in free solution. Tedizolid and ACH-702 have a good intracellular killing activity comparable to that of rifampin or moxifloxacin.

  5. Differential regulation of caspase-1 activation, pyroptosis, and autophagy via Ipaf and ASC in Shigella-infected macrophages.

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    Toshihiko Suzuki

    2007-08-01

    Full Text Available Shigella infection, the cause of bacillary dysentery, induces caspase-1 activation and cell death in macrophages, but the precise mechanisms of this activation remain poorly understood. We demonstrate here that caspase-1 activation and IL-1beta processing induced by Shigella are mediated through Ipaf, a cytosolic pattern-recognition receptor of the nucleotide-binding oligomerization domain (NOD-like receptor (NLR family, and the adaptor protein apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC. We also show that Ipaf was critical for pyroptosis, a specialized form of caspase-1-dependent cell death induced in macrophages by bacterial infection, whereas ASC was dispensable. Unlike that observed in Salmonella and Legionella, caspase-1 activation induced by Shigella infection was independent of flagellin. Notably, infection of macrophages with Shigella induced autophagy, which was dramatically increased by the absence of caspase-1 or Ipaf, but not ASC. Autophagy induced by Shigella required an intact bacterial type III secretion system but not VirG protein, a bacterial factor required for autophagy in epithelial-infected cells. Treatment of macrophages with 3-methyladenine, an inhibitor of autophagy, enhanced pyroptosis induced by Shigella infection, suggesting that autophagy protects infected macrophages from pyroptosis. Thus, Ipaf plays a critical role in caspase-1 activation induced by Shigella independently of flagellin. Furthermore, the absence of Ipaf or caspase-1, but not ASC, regulates pyroptosis and the induction of autophagy in Shigella-infected macrophages, providing a novel function for NLR proteins in bacterial-host interactions.

  6. Distinct Macrophage Fates after in vitro Infection with Different Species of Leishmania: Induction of Apoptosis by Leishmania (Leishmania amazonensis, but Not by Leishmania (Viannia guyanensis.

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    Jarina Pena DaMata

    Full Text Available Leishmania is an intracellular parasite in vertebrate hosts, including man. During infection, amastigotes replicate inside macrophages and are transmitted to healthy cells, leading to amplification of the infection. Although transfer of amastigotes from infected to healthy cells is a crucial step that may shape the outcome of the infection, it is not fully understood. Here we compare L. amazonensis and L. guyanensis infection in C57BL/6 and BALB/c mice and investigate the fate of macrophages when infected with these species of Leishmania in vitro. As previously shown, infection of mice results in distinct outcomes: L. amazonensis causes a chronic infection in both strains of mice (although milder in C57BL/6, whereas L. guyanensis does not cause them disease. In vitro, infection is persistent in L. amazonensis-infected macrophages whereas L. guyanensis growth is controlled by host cells from both strains of mice. We demonstrate that, in vitro, L. amazonensis induces apoptosis of both C57BL/6 and BALB/c macrophages, characterized by PS exposure, DNA cleavage into nucleosomal size fragments, and consequent hypodiploidy. None of these signs were seen in macrophages infected with L. guyanensis, which seem to die through necrosis, as indicated by increased PI-, but not Annexin V-, positive cells. L. amazonensis-induced macrophage apoptosis was associated to activation of caspases-3, -8 and -9 in both strains of mice. Considering these two species of Leishmania and strains of mice, macrophage apoptosis, induced at the initial moments of infection, correlates with chronic infection, regardless of its severity. We present evidence suggestive that macrophages phagocytize L. amazonensis-infected cells, which has not been verified so far. The ingestion of apoptotic infected macrophages by healthy macrophages could be a way of amastigote spreading, leading to the establishment of infection.

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

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    Maura E Casey

    2015-02-01

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

  8. Placement of endosseous implant in infected alveolar socket with large fenestration defect: A comparative case report

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    Balaji Anitha

    2010-01-01

    Full Text Available Placement of endosseous implants into infected bone is often deferred or avoided due to fear of failure. However, with the development of guided bone regeneration [GBR], some implantologists have reported successful implant placement in infected sockets, even those with fenestration defects. We had the opportunity to compare the osseointegration of an immediate implant placed in an infected site associated with a large buccal fenestration created by the removal of a root stump with that of a delayed implant placed 5 years after extraction. Both implants were placed in the same patient, in the same dental quadrant by the same implantologist. GBR was used with the fenestration defect being filled with demineralized bone graftFNx01 and covered with collagen membraneFNx08. Both implants were osseointegrated and functional when followed up after 12 months.

  9. Interleukin-4 activated macrophages mediate immunity to filarial helminth infection by sustaining CCR3-dependent eosinophilia.

    Science.gov (United States)

    Turner, Joseph D; Pionnier, Nicolas; Furlong-Silva, Julio; Sjoberg, Hanna; Cross, Stephen; Halliday, Alice; Guimaraes, Ana F; Cook, Darren A N; Steven, Andrew; Van Rooijen, Nico; Allen, Judith E; Jenkins, Stephen J; Taylor, Mark J

    2018-03-01

    Eosinophils are effectors in immunity to tissue helminths but also induce allergic immunopathology. Mechanisms of eosinophilia in non-mucosal tissues during infection remain unresolved. Here we identify a pivotal function of tissue macrophages (Mϕ) in eosinophil anti-helminth immunity using a BALB/c mouse intra-peritoneal Brugia malayi filarial infection model. Eosinophilia, via C-C motif chemokine receptor (CCR)3, was necessary for immunity as CCR3 and eosinophil impairments rendered mice susceptible to chronic filarial infection. Post-infection, peritoneal Mϕ populations proliferated and became alternatively-activated (AAMϕ). Filarial AAMϕ development required adaptive immunity and interleukin-4 receptor-alpha. Depletion of Mϕ prior to infection suppressed eosinophilia and facilitated worm survival. Add back of filarial AAMϕ in Mϕ-depleted mice recapitulated a vigorous eosinophilia. Transfer of filarial AAMϕ into Severe-Combined Immune Deficient mice mediated immunological resistance in an eosinophil-dependent manner. Exogenous IL-4 delivery recapitulated tissue AAMϕ expansions, sustained eosinophilia and mediated immunological resistance in Mϕ-intact SCID mice. Co-culturing Brugia with filarial AAMϕ and/or filarial-recruited eosinophils confirmed eosinophils as the larvicidal cell type. Our data demonstrates that IL-4/IL-4Rα activated AAMϕ orchestrate eosinophil immunity to filarial tissue helminth infection.

  10. Murine macrophage response from peritoneal cavity requires signals mediated by chemokine receptor CCR-2 during Staphylococcus aureus infection.

    Science.gov (United States)

    Nandi, Ajeya; Bishayi, Biswadev

    2016-02-01

    C-C chemokine receptor-2 (CCR-2) is a cognate receptor for monocyte chemotactic protein-1 (MCP-1), and recent studies revealed that MCP-1-CCR-2 signaling is involved in several inflammatory diseases characterized by macrophage infiltration. Currently, there is no study on the involvement of CCR-2 in the killing of S. aureus by macrophages of Swiss albino mice, and its substantial role in host defense against S. aureus infection in murine macrophages is still unclear. Therefore, the present study was aimed to investigate the functional and interactive role of CCR-2 and MCP-1 in regulating peritoneal macrophage responses with respect to acute S. aureus infection. We found that phagocytosis of S. aureus can serve as an important stimulus for MCP-1 production by peritoneal macrophages, which is dependent directly or indirectly on cytokines, reactive oxygen species and nitric oxide. Neutralization of CCR-2 in macrophages leads to increased production of IL-10 and decreased production of IFN-γ and IL-6. In CCR-2 blocked macrophages, pretreatment with specific blocker of NF-κB or p38-MAPK causes elevation in MCP-1 level and subsequent downregulation of CCR-2 itself. We speculate that CCR-2 is involved in S. aureus-induced MCP-1 production via NF-κB or p38-MAPK signaling. We also hypothesized that unnaturally high level of MCP-1 that build up upon CCR-2 neutralization might allow promiscuous binding to one or more other chemokine receptors, a situation that would not occur in CCR-2 non-neutralized condition. This may be the plausible explanation for such observed Th-2 response in CCR-2 blocked macrophages infected with S. aureus in the present study.

  11. Value of Tc99m-DTPA alveolar permeability in lung involvement detection of patients with HIV infection

    International Nuclear Information System (INIS)

    Massardo Vega, Teresa; Jofre Manieu, Maria Josefina; Cabello Araya, Hernan; Sepulveda Carvajal, Cecilia; Ruiz Carmona, Mauricio; Moyano Schlegel, Leonor; Fica Cubillos, Alberto; Alay Perez, Rita

    2001-01-01

    We studied 35 HIV patients in order to know the value of Tc99mDTPA in the assessment of pulmonary lung involvement, especially pneumocystis carinii (PC) infection. Lung DTPA clearance measures increased alveolar permeability. Twenty patients with respiratory symptoms were included, 4 with systemic symptoms and also 11 asymptomatics, with similar immune condition (CD4 lymphocytes <400) as a control group. Smoking habit was suspended prior the test. Clinical follow up, chest film, induced sputum and/or fibrobronchoscopy were obtained. There was histological confirmation of PC presence or absence in 16 symptomatics and 3 asymptomatics. DTPA sensitivity for PC detection was 78%, specificity 40% and accuracy 58%; the values were 85%, 60% and 79%, respectively, for inflammatory lung processes. There were 4/6 cases false positive for PC detection with respiratory features explaining DTPA abnormalities. Concluding, Tc99m-DTPA is sensitive but not specific for detecting PC pneumonia but its value is higher for pulmonary inflammatory processes (Au)

  12. Protozoa and human macrophages infection by Legionella pneumophila environmental strains belonging to different serogroups.

    Science.gov (United States)

    Messi, Patrizia; Patrizia, Messi; Bargellini, Annalisa; Annalisa, Bargellini; Anacarso, Immacolata; Immacolata, Anacarso; Marchesi, Isabella; Isabella, Marchesi; de Niederhäusern, Simona; Bondi, Moreno; Moreno, Bondi

    2013-02-01

    Three Legionella pneumophila strains isolated from municipal hot tap water during a multicentric Italian survey and belonging to serogroups 1, 6, 9 and the reference strain Philadelphia-1 were studied to determine the intracellular replication capability and the cytopathogenicity in human monocyte cell line U937 and in an Acanthamoeba polyphaga strain. Our results show that both serogroups 1 and Philadelphia-1 were able to multiply into macrophages inducing cytopathogenicity, while serogroup 6 and ever more serogroup 9 were less efficient in leading to death of the infected macrophages. Both serogroups 1 and 6 displayed a quite good capability of intracellular replication in A. polyphaga, although serogroup 1 was less cytopathogenic than serogroup 6. Serogroup 9, like Philadelphia-1 strain, showed a reduced efficiency of infection and replication and a low cytopathogenicity towards the protozoan. Our study suggests that bacterial pathogenesis is linked to the difference in the virulence expression of L. pneumophila serogroups in both hosts, as demonstrated by the fact that only L. pneumophila serogroup 1 shows the contextual expression of the two virulence traits. Serogroup 6 proves to be a good candidate as pathogen since it shows a good capacity for intracellular replication in protozoan.

  13. Virulence of Mycobacterium avium Subsp. hominissuis Human Isolates in an in vitro Macrophage Infection Model

    Directory of Open Access Journals (Sweden)

    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.

  14. Soluble CD40 Ligand in Sera of Subjects Exposed to Leishmania infantum Infection Reduces the Parasite Load in Macrophages.

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    Fabrícia Alvisi de Oliveira

    Full Text Available While CD40L is typically a membrane glycoprotein expressed on activated T cells and platelets that binds and activates CD40 on the surface on antigen presenting cells, a soluble derivative (sCD40L that appears to retain its biological activity after cleavage from cell membrane also exists. We recently reported that sCD40L is associated with clinical resolution of visceral leishmaniasis and protection against the disease. In the present study we investigated if this sCD40L is functional and exerts anti-parasitic effect in L. infantum-infected macrophages.Macrophages from normal human donors were infected with L. infantum promastigotes and incubated with either sera from subjects exposed to L. infantum infection, monoclonal antibodies against human CD40L, or an isotype control antibody. We then evaluated infection by counting the number of infected cells and the number of parasites in each cell. We also measured a variety of immune modulatory cytokines in these macrophage culture supernatants by Luminex assay. The addition of sCD40L, either recombinant or from infected individuals' serum, decreased both the number of infected macrophages and number of intracellular parasites. Moreover, this treatment increased the production of IL-12, IL-23, IL-27, IL-15, and IL1β such that negative correlations between the levels of these cytokines with both the infection ratio and number of intracellular parasites were observed.sCD40L from sera of subjects exposed to L. infantum is functional and improves both the control of parasite and production of inflamatory cytokines of infected macrophages. Although the mechanisms involved in parasite killing are still unclear and require further exploration, these findings indicate a protective role of sCD40L in visceral leishmaniasis.

  15. Responses of murine and human macrophages to leptospiral infection: a study using comparative array analysis.

    Directory of Open Access Journals (Sweden)

    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.

  16. Infection Related Inferior Alveolar Nerve Paresthesia in the Lower Premolar Teeth

    Directory of Open Access Journals (Sweden)

    Rachele Censi

    2016-01-01

    Full Text Available Introduction. The aim of this paper was to describe two cases of IAN infection-induced paresthesia and to discuss the most appropriate treatment solutions. Methods. For two patients, periapical lesions that induced IAN paresthesia were revealed. In the first case, the tooth was previously endodontically treated, whereas in the second case the lesion was due to pulp necrosis. Results. For the first patient, a progressive healing was observed only after the tooth extraction. In the second patient, the paresthesia had resolved after endodontic treatment. Conclusions. The endodontic-related paresthesia is a rare complication that can be the result of a combination of etiopathogenic mechanisms such as mechanical pressure on the nerve fibers due to the expanding infectious process and the production of microbial toxins. Paresthesia resulting from periapical lesions usually subsides through elimination of infection by root canal treatment. However, if there are no signs of enhancement, the immediate extraction of the tooth is the treatment of choice in order to prevent irreversible paresthesia because it was demonstrated that there is a correlation between the duration of mechanical or chemical irritation and the risk of permanent paresthesia.

  17. Losartan and enalapril decrease viral absorption and interleukin 1 beta production by macrophages in an experimental dengue virus infection.

    Science.gov (United States)

    Hernández-Fonseca, Juan Pablo; Durán, Anyelo; Valero, Nereida; Mosquera, Jesús

    2015-11-01

    The role of angiotensin II (Ang II) in dengue virus infection remains unknown. The aim of this study was to determine the effect of losartan, an antagonist of the angiotensin II type 1 receptor (AT1 receptor), and enalapril, an inhibitor of angiotensin I-converting enzyme (ACE), on viral antigen expression and IL-1β production in peritoneal macrophages infected with dengue virus type 2. Mice treated with losartan or enalapril and untreated controls were infected intraperitoneally with the virus, and macrophages were analyzed. Infection resulted in increased IL-1β production and a high percentage of cells expressing viral antigen, and this was decreased by treatment with anti-Ang II drugs, suggesting a role for Ang II in dengue virus infection.

  18. Macrophage Colony Stimulating Factor Derived from CD4+ T Cells Contributes to Control of a Blood-Borne Infection.

    Science.gov (United States)

    Fontana, Mary F; de Melo, Gabrielly L; Anidi, Chioma; Hamburger, Rebecca; Kim, Chris Y; Lee, So Youn; Pham, Jennifer; Kim, Charles C

    2016-12-01

    Dynamic regulation of leukocyte population size and activation state is crucial for an effective immune response. In malaria, Plasmodium parasites elicit robust host expansion of macrophages and monocytes, but the underlying mechanisms remain unclear. Here we show that myeloid expansion during P. chabaudi infection is dependent upon both CD4+ T cells and the cytokine Macrophage Colony Stimulating Factor (MCSF). Single-cell RNA-Seq analysis on antigen-experienced T cells revealed robust expression of Csf1, the gene encoding MCSF, in a sub-population of CD4+ T cells with distinct transcriptional and surface phenotypes. Selective deletion of Csf1 in CD4+ cells during P. chabaudi infection diminished proliferation and activation of certain myeloid subsets, most notably lymph node-resident CD169+ macrophages, and resulted in increased parasite burden and impaired recovery of infected mice. Depletion of CD169+ macrophages during infection also led to increased parasitemia and significant host mortality, confirming a previously unappreciated role for these cells in control of P. chabaudi. This work establishes the CD4+ T cell as a physiologically relevant source of MCSF in vivo; probes the complexity of the CD4+ T cell response during type 1 infection; and delineates a novel mechanism by which T helper cells regulate myeloid cells to limit growth of a blood-borne intracellular pathogen.

  19. Contrasting transcriptional responses of a virulent and an attenuated strain of Mycobacterium tuberculosis infecting macrophages.

    Directory of Open Access Journals (Sweden)

    Alice H Li

    2010-06-01

    Full Text Available H37Rv and H37Ra are well-described laboratory strains of Mycobacterium tuberculosis derived from the same parental strain, H37, that show dramatically different pathogenic phenotypes.In this study, the transcriptomes of the two strains during axenic growth in broth and during intracellular growth within murine bone-marrow macrophages were compared by whole genome expression profiling. We identified and compared adaptations of either strain upon encountering an intracellular environment, and also contrasted the transcriptomes of the two strains while inside macrophages. In the former comparison, both strains induced genes that would facilitate intracellular survival including those involved in mycobactin synthesis and fatty acid metabolism. However, this response was stronger and more extensive for H37Rv than for H37Ra. This was manifested as the differential expression of a greater number of genes and an increased magnitude of expression for these genes in H37Rv. In comparing intracellular transcriptional signatures, fifty genes were found to be differentially expressed between the strains. Of these fifty, twelve were under control of the PhoPR regulon. Further differences between strains included genes whose products were members of the ESAT-6 family of proteins, or were associated with their secretion.Along with the recent identification of single nucleotide polymorphisms in H37Ra when compared to H37Rv, our demonstration of differential expression of PhoP-regulated and ESX-1 region-related genes during macrophage infection further highlights the significance of these genes in the attenuation of H37Ra.

  20. Regulation of macrophage accessory cell activity by mycobacteria. I. Ia expression in normal and irradiated mice infected with Mycobacterium mycroti

    International Nuclear Information System (INIS)

    Kaye, P.M.; Feldmann, M.

    1986-01-01

    CBA/Ca mice were infected by either the intravenous or intraperitoneal route with Mycobacterium microti and the subsequent changes in local macrophage populations examined. Following infection, the number of macrophages increased and they showed greater expression of both MHC Class II molecules. This response was not dependent on viability of the mycobacteria, in contrast to reports with other microorganisms such as Listeria. Studies in sublethally irradiated mice indicated that persistent antigen could give rise to a response after a period of host recovery which was radiation dose dependent. This procedure also highlighted differences in the regulation of different murine class II antigens in vivo, as seen by delayed re-expression of I-E antigens. Macrophage accessory cell function, as assessed by an in vitro T cell proliferation assay, correlated with Ia expression after fixation, but not after indomethacin treatment; this highlights the diverse nature of regulatory molecules produced by these cells. (author)

  1. An incomplete TCA cycle increases survival of Salmonella Typhimurium during infection of resting and activated murine macrophages.

    Science.gov (United States)

    Bowden, Steven D; Ramachandran, Vinoy K; Knudsen, Gitte M; Hinton, Jay C D; Thompson, Arthur

    2010-11-08

    In comparison to the comprehensive analyses performed on virulence gene expression, regulation and action, the intracellular metabolism of Salmonella during infection is a relatively under-studied area. We investigated the role of the tricarboxylic acid (TCA) cycle in the intracellular replication of Salmonella Typhimurium in resting and activated macrophages, epithelial cells, and during infection of mice. We constructed deletion mutations of 5 TCA cycle genes in S. Typhimurium including gltA, mdh, sdhCDAB, sucAB, and sucCD. We found that the mutants exhibited increased net intracellular replication in resting and activated murine macrophages compared to the wild-type. In contrast, an epithelial cell infection model showed that the S. Typhimurium ΔsucCD and ΔgltA strains had reduced net intracellular replication compared to the wild-type. The glyoxylate shunt was not responsible for the net increased replication of the TCA cycle mutants within resting macrophages. We also confirmed that, in a murine infection model, the S. Typhimurium ΔsucAB and ΔsucCD strains are attenuated for virulence. Our results suggest that disruption of the TCA cycle increases the ability of S. Typhimurium to survive within resting and activated murine macrophages. In contrast, epithelial cells are non-phagocytic cells and unlike macrophages cannot mount an oxidative and nitrosative defence response against pathogens; our results show that in HeLa cells the S. Typhimurium TCA cycle mutant strains show reduced or no change in intracellular levels compared to the wild-type. The attenuation of the S. Typhimurium ΔsucAB and ΔsucCD mutants in mice, compared to their increased net intracellular replication in resting and activated macrophages suggest that Salmonella may encounter environments within the host where a complete TCA cycle is advantageous.

  2. Immune modulation with sulfasalazine attenuates immunopathogenesis but enhances macrophage-mediated fungal clearance during Pneumocystis pneumonia.

    Directory of Open Access Journals (Sweden)

    Jing Wang

    2010-08-01

    Full Text Available Although T cells are critical for host defense against respiratory fungal infections, they also contribute to the immunopathogenesis of Pneumocystis pneumonia (PcP. However, the precise downstream effector mechanisms by which T cells mediate these diverse processes are undefined. In the current study the effects of immune modulation with sulfasalazine were evaluated in a mouse model of PcP-related Immune Reconstitution Inflammatory Syndrome (PcP-IRIS. Recovery of T cell-mediated immunity in Pneumocystis-infected immunodeficient mice restored host defense, but also initiated the marked pulmonary inflammation and severe pulmonary function deficits characteristic of IRIS. Sulfasalazine produced a profound attenuation of IRIS, with the unexpected consequence of accelerated fungal clearance. To determine whether macrophage phagocytosis is an effector mechanism of T cell-mediated Pneumocystis clearance and whether sulfasalazine enhances clearance by altering alveolar macrophage phagocytic activity, a novel multispectral imaging flow cytometer-based method was developed to quantify the phagocytosis of Pneumocystis in vivo. Following immune reconstitution, alveolar macrophages from PcP-IRIS mice exhibited a dramatic increase in their ability to actively phagocytose Pneumocystis. Increased phagocytosis correlated temporally with fungal clearance, and required the presence of CD4(+ T cells. Sulfasalazine accelerated the onset of the CD4(+ T cell-dependent alveolar macrophage phagocytic response in PcP-IRIS mice, resulting in enhanced fungal clearance. Furthermore, sulfasalazine promoted a TH2-polarized cytokine environment in the lung, and sulfasalazine-enhanced phagocytosis of Pneumocystis was associated with an alternatively activated alveolar macrophage phenotype. These results provide evidence that macrophage phagocytosis is an important in vivo effector mechanism for T cell-mediated Pneumocystis clearance, and that macrophage phenotype can be altered

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

    Science.gov (United States)

    2017-10-01

    monocyte-derived alveolar macrophages based on single cell molecular classification in patients with pulmonary fibrosis. We have recruited a planned number...of patients into the study and performed transcriptional profiling by means of RNA-seq on single cell and bulk sorted populations or alveolar...macrophages. Major findings: In agreement with our hypothesis, using single cell RNA-seq, we found substantial heterogeneity of alveolar macrophages in the

  4. Construction of genetic markers for the study of Salmonella typhimurium infection of murine macrophages

    DEFF Research Database (Denmark)

    Jelsbak, Lotte; Olsen, John Elmerdahl

    were constructed using Red-mediated recombination (1) and the promoters were selected from the recently published expressional data of Salmonella infection of murine macrophages (2). As reporter proteins we both use a stable GFPmut3 variant as well as an unstable GFP variant (3). Using these fusions...

  5. 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

  6. MD-2 regulates LPS-induced NLRP3 inflammasome activation and IL-1beta secretion by a MyD88/NF-κB-dependent pathway in alveolar macrophages cell line.

    Science.gov (United States)

    Luo, Man; Hu, Lijuan; Li, Dandan; Wang, Yanying; He, Yuting; Zhu, Lei; Ren, Weiying

    2017-10-01

    Myeloid differentiation protein 2 (MD-2) is required in the recognition of lipopolysaccharide (LPS) by toll-like receptor 4 (TLR4), and participates in LPS-induced alveolar macrophage (AM) inflammation during acute lung injury (ALI). Activation of the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome aggravates inflammation in LPS-induced ALI. However, there is currently little known about the relationship between MD-2 signaling and the NLRP3 inflammasome. This study showed that NLRP3 expression, IL-1beta (IL-1β) secretion, and pyroptosis were up-regulated after LPS stimulation in the NR8383 AM cell-line. MD-2 gene knock-down reduced LPS-induced mRNA and protein expression of NLRP3 and IL-1β secretion in NR8383 cells, and inhibited the MyD88/NF-κB signaling pathway. Conversely, over-expression of MD-2 not only heightened NLRP3, MyD88, and NF-κB p65 protein expression, it also aggravated the LPS-induced inflammatory response. Furthermore, the NF-κB inhibitor SN50 had a beneficial role in decreasing NLRP3 and caspase-1 mRNA and protein expression. The observations suggest that MD-2 helps to regulate LPS-induced NLRP3 inflammasome activation and the inflammatory response in NR8383 cells, and likely does so by affecting MyD88/NF-κB signaling. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

    Science.gov (United States)

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

    2015-06-01

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

  8. Distinct characteristics of endometrial and decidual macrophages and regulation of their permissivity to HIV-1 infection by SAMHD1.

    Science.gov (United States)

    Quillay, Héloïse; El Costa, Hicham; Marlin, Romain; Duriez, Marion; Cannou, Claude; Chrétien, Fabrice; Fernandez, Hervé; Lebreton, Anne; Ighil, Julien; Schwartz, Olivier; Barré-Sinoussi, Françoise; Nugeyre, Marie-Thérèse; Menu, Elisabeth

    2015-01-15

    In order to develop strategies to prevent HIV-1 (human immunodeficiency virus type 1) transmission, it is crucial to better characterize HIV-1 target cells in the female reproductive tract (FRT) mucosae and to identify effective innate responses. Control of HIV-1 infection in the decidua (the uterine mucosa during pregnancy) can serve as a model to study natural mucosal protection. Macrophages are the main HIV-1 target cells in the decidua. Here we report that in vitro, macrophages and T cells are the main HIV-1 targets in the endometrium in nonpregnant women. As reported for decidual macrophages (dM), endometrial macrophages (eM) were found to have an M2-like phenotype (CD68+ CD163+ CD206+ IL-10high). However, eM and dM may belong to different subpopulations, as they differently express certain markers and secrete different amounts of proinflammatory and anti-inflammatory cytokines. We observed strong expression of the SAMHD1 restriction factor and weak expression of its inactive form (pSAMHD1, phosphorylated at residue Thr592) in both eM and dM. Infection of macrophages from both tissues was enhanced in the presence of the viral protein Vpx, suggesting a role for SAMHD1 in the restriction of HIV-1 infection. This study and further comparisons of the decidua with FRT mucosae in nonpregnant women should help to identify mechanisms of mucosal protection against HIV-1 infection. The female reproductive tract mucosae are major portals of HIV-1 entry into the body. The decidua (uterine mucosa during pregnancy) can serve as a model for studying natural mucosal protection against HIV-1 transmission. A comparison of target cells and innate responses in the decidua versus the endometrium in nonpregnant women could help to identify protective mechanisms. Here, we report for the first time that macrophages are one of the main HIV-1 target cells in the endometrium and that infection of macrophages from both the endometrium and the decidua is restricted by SAMHD1. These

  9. SARS-CoV replicates in primary human alveolar type II cell cultures but not in type I-like cells

    Science.gov (United States)

    Mossel, Eric C.; Wang, Jieru; Jeffers, Scott; Edeen, Karen E.; Wang, Shuanglin; Cosgrove, Gregory P.; Funk, C. Joel; Manzer, Rizwan; Miura, Tanya A.; Pearson, Leonard D.; Holmes, Kathryn V.; Mason, Robert J.

    2008-01-01

    Severe acute respiratory syndrome (SARS) is a disease characterized by diffuse alveolar damage. We isolated alveolar type II cells and maintained them in a highly differentiated state. Type II cell cultures supported SARS-CoV replication as evidenced by RT-PCR detection of viral subgenomic RNA and an increase in virus titer. Virus titers were maximal by 24 hours and peaked at approximately 105 pfu/mL. Two cell types within the cultures were infected. One cell type was type II cells, which were positive for SP-A, SP-C, cytokeratin, a type II cell-specific monoclonal antibody, and Ep-CAM. The other cell type was composed of spindle-shaped cells that were positive for vimentin and collagen III and likely fibroblasts. Viral replication was not detected in type I-like cells or macrophages. Hence, differentiated adult human alveolar type II cells were infectible but alveolar type I-like cells and alveolar macrophages did not support productive infection. PMID:18022664

  10. 17-AAG kills intracellular Leishmania amazonensis while reducing inflammatory responses in infected macrophages.

    Science.gov (United States)

    Petersen, Antonio Luis de Oliveira Almeida; Guedes, Carlos Eduardo Sampaio; Versoza, Carolina Leite; Lima, José Geraldo Bomfim; de Freitas, Luiz Antônio Rodrigues; Borges, Valéria Matos; Veras, Patrícia Sampaio Tavares

    2012-01-01

    Leishmaniasis is a neglected endemic disease with a broad spectrum of clinical manifestations. Pentavalent antimonials have been the treatment of choice for the past 70 years and, due to the emergence of resistant cases, the efficacy of these drugs has come under scrutiny. Second-line drugs are less efficacious, cause a range of side effects and can be costly. The formulation of new generations of drugs, especially in developing countries, has become mandatory. We investigated the anti-leishmanial effect of 17-(allylamino)-17-demethoxygeldanamycin (17-AAG), an HSP90 inhibitor, in vitro. This inhibitor is currently in clinical trials for cancer treatment; however, its effects against intracellular Leishmania remain untested. Macrophages infected with L. amazonensis were treated with 17-AAG (25-500 nM) and parasite load was quantified using optical microscopy. Parasite load declined in 17-AAG-treated macrophages in a dose- and time-dependent manner. Intracellular parasite death became irreversible after 4 h of treatment with 17-AAG, and occurred independent of nitric oxide (NO) and superoxide (O(2) (-)) production. Additionally, intracellular parasite viability was severely reduced after 48 h of treatment. Interestingly, treatment with 17-AAG reduced pro-inflammatory mediator production, including TNF-α, IL-6 and MCP-1, yet IL-12 remained unaffected. Electron microscopy revealed morphological alterations, such as double-membrane vacuoles and myelin figures at 24 and 48 h after 17-AAG treatment. The HSP90 inhibitor, 17-AAG, possesses high potency under low dosage and reduces both pro-inflammatory and oxidative molecule production. Therefore, further studies are warranted to investigate this inhibitor's potential in the development of new generations of anti-leishmanials.

  11. Physico-chemical properties of quartz from industrial manufacturing and its cytotoxic effects on alveolar macrophages: The case of green sand mould casting for iron production.

    Science.gov (United States)

    Di Benedetto, Francesco; Gazzano, Elena; Tomatis, Maura; Turci, Francesco; Pardi, Luca A; Bronco, Simona; Fornaciai, Gabriele; Innocenti, Massimo; Montegrossi, Giordano; Muniz Miranda, Maurizio; Zoleo, Alfonso; Capacci, Fabio; Fubini, Bice; Ghigo, Dario; Romanelli, Maurizio

    2016-07-15

    Industrial processing of materials containing quartz induces physico-chemical modifications that contribute to the variability of quartz hazard in different plants. Here, modifications affecting a quartz-rich sand during cast iron production, have been investigated. Composition, morphology, presence of radicals associated to quartz and reactivity in free radical generation were studied on a raw sand and on a dust recovered after mould dismantling. Additionally, cytotoxicity of the processed dust and ROS and NO generation were evaluated on MH-S macrophages. Particle morphology and size were marginally affected by casting processing, which caused only a slight increase of the amount of respirable fraction. The raw sand was able to catalyze OH and CO2(-) generation in cell-free test, even if in a lesser extent than the reference quartz (Min-U-Sil), and shows hAl radicals, conventionally found in any quartz-bearing raw materials. Enrichment in iron and extensive coverage with amorphous carbon were observed during processing. They likely contributed, respectively, to increasing the ability of processed dust to release CO2- and to suppressing OH generation respect to the raw sand. Carbon coverage and repeated thermal treatments during industrial processing also caused annealing of radiogenic hAl defects. Finally, no cellular responses were observed with the respirable fraction of the processed powder. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Extracellular Vesicles Released from Mycobacterium tuberculosis-Infected Neutrophils Promote Macrophage Autophagy and Decrease Intracellular Mycobacterial Survival

    Science.gov (United States)

    Alvarez-Jiménez, Violeta D.; Leyva-Paredes, Kahiry; García-Martínez, Mariano; Vázquez-Flores, Luis; García-Paredes, Víctor Gabriel; Campillo-Navarro, Marcia; Romo-Cruz, Israel; Rosales-García, Víctor Hugo; Castañeda-Casimiro, Jessica; González-Pozos, Sirenia; Hernández, José Manuel; Wong-Baeza, Carlos; García-Pérez, Blanca Estela; Ortiz-Navarrete, Vianney; Estrada-Parra, Sergio; Serafín-López, Jeanet; Wong-Baeza, Isabel; Chacón-Salinas, Rommel; Estrada-García, Iris

    2018-01-01

    Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis (Mtb). In the lungs, macrophages and neutrophils are the first immune cells that have contact with the infecting mycobacteria. Neutrophils are phagocytic cells that kill microorganisms through several mechanisms, which include the lytic enzymes and antimicrobial peptides that are found in their lysosomes, and the production of reactive oxygen species. Neutrophils also release extracellular vesicles (EVs) (100–1,000 nm in diameter) to the extracellular milieu; these EVs consist of a lipid bilayer surrounding a hydrophilic core and participate in intercellular communication. We previously demonstrated that human neutrophils infected in vitro with Mtb H37Rv release EVs (EV-TB), but the effect of these EVs on other cells relevant for the control of Mtb infection, such as macrophages, has not been completely analyzed. In this study, we characterized the EVs produced by non-stimulated human neutrophils (EV-NS), and the EVs produced by neutrophils stimulated with an activator (PMA), a peptide derived from bacterial proteins (fMLF) or Mtb, and observed that the four EVs differed in their size. Ligands for toll-like receptor (TLR) 2/6 were detected in EV-TB, and these EVs favored a modest increase in the expression of the co-stimulatory molecules CD80, a higher expression of CD86, and the production of higher amounts of TNF-α and IL-6, and of lower amounts of TGF-β, in autologous human macrophages, compared with the other EVs. EV-TB reduced the amount of intracellular Mtb in macrophages, and increased superoxide anion production in these cells. TLR2/6 ligation and superoxide anion production are known inducers of autophagy; accordingly, we found that EV-TB induced higher expression of the autophagy-related marker LC3-II in macrophages, and the co-localization of LC3-II with Mtb inside infected macrophages. The intracellular mycobacterial load increased when autophagy was inhibited with

  13. Extracellular Vesicles Released from Mycobacterium tuberculosis-Infected Neutrophils Promote Macrophage Autophagy and Decrease Intracellular Mycobacterial Survival

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    Violeta D. Alvarez-Jiménez

    2018-02-01

    Full Text Available Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis (Mtb. In the lungs, macrophages and neutrophils are the first immune cells that have contact with the infecting mycobacteria. Neutrophils are phagocytic cells that kill microorganisms through several mechanisms, which include the lytic enzymes and antimicrobial peptides that are found in their lysosomes, and the production of reactive oxygen species. Neutrophils also release extracellular vesicles (EVs (100–1,000 nm in diameter to the extracellular milieu; these EVs consist of a lipid bilayer surrounding a hydrophilic core and participate in intercellular communication. We previously demonstrated that human neutrophils infected in vitro with Mtb H37Rv release EVs (EV-TB, but the effect of these EVs on other cells relevant for the control of Mtb infection, such as macrophages, has not been completely analyzed. In this study, we characterized the EVs produced by non-stimulated human neutrophils (EV-NS, and the EVs produced by neutrophils stimulated with an activator (PMA, a peptide derived from bacterial proteins (fMLF or Mtb, and observed that the four EVs differed in their size. Ligands for toll-like receptor (TLR 2/6 were detected in EV-TB, and these EVs favored a modest increase in the expression of the co-stimulatory molecules CD80, a higher expression of CD86, and the production of higher amounts of TNF-α and IL-6, and of lower amounts of TGF-β, in autologous human macrophages, compared with the other EVs. EV-TB reduced the amount of intracellular Mtb in macrophages, and increased superoxide anion production in these cells. TLR2/6 ligation and superoxide anion production are known inducers of autophagy; accordingly, we found that EV-TB induced higher expression of the autophagy-related marker LC3-II in macrophages, and the co-localization of LC3-II with Mtb inside infected macrophages. The intracellular mycobacterial load increased when autophagy was inhibited

  14. Insulin-Like Growth Factor-I Induces Arginase Activity in Leishmania amazonensis Amastigote-Infected Macrophages through a Cytokine-Independent Mechanism

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    Celia Maria Vieira Vendrame

    2014-01-01

    Full Text Available Leishmania (Leishmania amazonensis exhibits peculiarities in its interactions with hosts. Because amastigotes are the primary form associated with the progression of infection, we studied the effect of insulin-like growth factor (IGF-I on interactions between L. (L. amazonensis amastigotes and macrophages. Upon stimulation of infected macrophages with IGF-I, we observed decreased nitric oxide production but increased arginase expression and activity, which lead to increased parasitism. However, stimulation of amastigote-infected macrophages with IGF-I did not result in altered cytokine levels compared to unstimulated controls. Because IGF-I is present in tissue fluids and also within macrophages, we examined the possible effect of this factor on phosphatidylserine (PS exposure on amastigotes, seen previously in tissue-derived amastigotes leading to increased parasitism. Stimulation with IGF-I induced PS exposure on amastigotes but not on promastigotes. Using a PS-liposome instead of amastigotes, we observed that the PS-liposome but not the control phosphatidylcholine-liposome led to increased arginase activity in macrophages, and this process was not blocked by anti-TGF-β antibodies. Our results suggest that in L. (L. amazonensis amastigote-infected macrophages, IGF-I induces arginase activity directly in amastigotes and in macrophages through the induction of PS exposure on amastigotes in the latter, which could lead to the alternative activation of macrophages through cytokine-independent mechanisms.

  15. Burkholderia cenocepacia type VI secretion system mediates escape of type II secreted proteins into the cytoplasm of infected macrophages.

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    Roberto Rosales-Reyes

    Full Text Available Burkholderia cenocepacia is an opportunistic pathogen that survives intracellularly in macrophages and causes serious respiratory infections in patients with cystic fibrosis. We have previously shown that bacterial survival occurs in bacteria-containing membrane vacuoles (BcCVs resembling arrested autophagosomes. Intracellular bacteria stimulate IL-1β secretion in a caspase-1-dependent manner and induce dramatic changes to the actin cytoskeleton and the assembly of the NADPH oxidase complex onto the BcCV membrane. A Type 6 secretion system (T6SS is required for these phenotypes but surprisingly it is not required for the maturation arrest of the BcCV. Here, we show that macrophages infected with B. cenocepacia employ the NLRP3 inflammasome to induce IL-1β secretion and pyroptosis. Moreover, IL-1β secretion by B. cenocepacia-infected macrophages is suppressed in deletion mutants unable to produce functional Type VI, Type IV, and Type 2 secretion systems (SS. We provide evidence that the T6SS mediates the disruption of the BcCV membrane, which allows the escape of proteins secreted by the T2SS into the macrophage cytoplasm. This was demonstrated by the activity of fusion derivatives of the T2SS-secreted metalloproteases ZmpA and ZmpB with adenylcyclase. Supporting this notion, ZmpA and ZmpB are required for efficient IL-1β secretion in a T6SS dependent manner. ZmpA and ZmpB are also required for the maturation arrest of the BcCVs and bacterial intra-macrophage survival in a T6SS-independent fashion. Our results uncover a novel mechanism for inflammasome activation that involves cooperation between two bacterial secretory pathways, and an unanticipated role for T2SS-secreted proteins in intracellular bacterial survival.

  16. HIV-1 Infection of Macrophages Dysregulates Innate Immune Responses to Mycobacterium tuberculosis by Inhibition of Interleukin-10

    Science.gov (United States)

    Tomlinson, Gillian S.; Bell, Lucy C. K.; Walker, Naomi F.; Tsang, Jhen; Brown, Jeremy S.; Breen, Ronan; Lipman, Marc; Katz, David R.; Miller, Robert F.; Chain, Benjamin M.; Elkington, Paul T. G.; Noursadeghi, Mahdad

    2014-01-01

    Human immunodeficiency virus (HIV)–1 and Mycobacterium tuberculosis (M. tuberculosis) both target macrophages, which are key cells in inflammatory responses and their resolution. Therefore, we tested the hypothesis that HIV-1 may modulate macrophage responses to coinfection with M. tuberculosis. HIV-1 caused exaggerated proinflammatory responses to M. tuberculosis that supported enhanced virus replication, and were associated with deficient stimulus-specific induction of anti-inflammatory interleukin (IL)–10 and attenuation of mitogen-activated kinase signaling downstream of Toll-like receptor 2 and dectin-1 stimulation. Our in vitro data were mirrored by lower IL-10 and higher proinflammatory IL-1β in airway samples from HIV-1–infected patients with pulmonary tuberculosis compared with those with non-tuberculous respiratory tract infections. Single-round infection of macrophages with HIV-1 was sufficient to attenuate IL-10 responses, and antiretroviral treatment of replicative virus did not affect this phenotype. We propose that deficient homeostatic IL-10 responses may contribute to the immunopathogenesis of active tuberculosis and propagation of virus infection in HIV-1/M. tuberculosis coinfection. PMID:24265436

  17. Regulation of L-type Voltage Gated Calcium Channel CACNA1S in Macrophages upon Mycobacterium tuberculosis Infection.

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    Cecil Antony

    Full Text Available We demonstrated earlier the inhibitory role played by Voltage Gated Calcium Channels (VGCCs in regulating Mycobacterium tuberculosis (M. tb survival and pathogenesis. In this report, we investigated mechanisms and key players that regulate the surface expression of VGCC-CACNA1S by Rv2463 and M. tb infection in macrophages. Our earlier work identified Rv2463 to be expressed at early times post infection in macrophages that induced suppressor responses to dendritic cells and macrophages. Our results in this study demonstrate a role of MyD88 independent TLR pathway in mediating CACNA1S expression. Dissecting the role for second messengers, we show that calcium homeostasis plays a key role in CACNA1S expression during M. tb infection. Using siRNAs against molecular sensors of calcium regulation, we show an involvement of ER associated Stromal Interaction Molecules 1 and 2 (STIM1 and STIM2, and transcription factor pCREB, towards CACNA1S expression that also involved the MyD88 independent pathway. Interestingly, reactive oxygen species played a negative role in M. tb mediated CACNA1S expression. Further, a cross-regulation of ROS and pCREB was noted that governed CACNA1S expression. Characterizing the mechanisms governing CACNA1S expression would improve our understanding of the regulation of VGCC expression and its role in M. tb pathogenesis during M. tb infection.

  18. Regulation of L-type Voltage Gated Calcium Channel CACNA1S in Macrophages upon Mycobacterium tuberculosis Infection.

    Science.gov (United States)

    Antony, Cecil; Mehto, Subhash; Tiwari, Brijendra K; Singh, Yogendra; Natarajan, Krishnamurthy

    2015-01-01

    We demonstrated earlier the inhibitory role played by Voltage Gated Calcium Channels (VGCCs) in regulating Mycobacterium tuberculosis (M. tb) survival and pathogenesis. In this report, we investigated mechanisms and key players that regulate the surface expression of VGCC-CACNA1S by Rv2463 and M. tb infection in macrophages. Our earlier work identified Rv2463 to be expressed at early times post infection in macrophages that induced suppressor responses to dendritic cells and macrophages. Our results in this study demonstrate a role of MyD88 independent TLR pathway in mediating CACNA1S expression. Dissecting the role for second messengers, we show that calcium homeostasis plays a key role in CACNA1S expression during M. tb infection. Using siRNAs against molecular sensors of calcium regulation, we show an involvement of ER associated Stromal Interaction Molecules 1 and 2 (STIM1 and STIM2), and transcription factor pCREB, towards CACNA1S expression that also involved the MyD88 independent pathway. Interestingly, reactive oxygen species played a negative role in M. tb mediated CACNA1S expression. Further, a cross-regulation of ROS and pCREB was noted that governed CACNA1S expression. Characterizing the mechanisms governing CACNA1S expression would improve our understanding of the regulation of VGCC expression and its role in M. tb pathogenesis during M. tb infection.

  19. Ion-abrasion scanning electron microscopy reveals surface-connected tubular conduits in HIV-infected macrophages.

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    Adam E Bennett

    2009-09-01

    Full Text Available HIV-1-containing internal compartments are readily detected in images of thin sections from infected cells using conventional transmission electron microscopy, but the origin, connectivity, and 3D distribution of these compartments has remained controversial. Here, we report the 3D distribution of viruses in HIV-1-infected primary human macrophages using cryo-electron tomography and ion-abrasion scanning electron microscopy (IA-SEM, a recently developed approach for nanoscale 3D imaging of whole cells. Using IA-SEM, we show the presence of an extensive network of HIV-1-containing tubular compartments in infected macrophages, with diameters of approximately 150-200 nm, and lengths of up to approximately 5 microm that extend to the cell surface from vesicular compartments that contain assembling HIV-1 virions. These types of surface-connected tubular compartments are not observed in T cells infected with the 29/31 KE Gag-matrix mutant where the virus is targeted to multi-vesicular bodies and released into the extracellular medium. IA-SEM imaging also allows visualization of large sheet-like structures that extend outward from the surfaces of macrophages, which may bend and fold back to allow continual creation of viral compartments and virion-lined channels. This potential mechanism for efficient virus trafficking between the cell surface and interior may represent a subversion of pre-existing vesicular machinery for antigen capture, processing, sequestration, and presentation.

  20. The co-transcriptome of uropathogenic Escherichia coli-infected mouse macrophages reveals new insights into host-pathogen interactions

    KAUST Repository

    Mavromatis, Charalampos Harris

    2015-01-24

    Urinary tract infections (UTI) are among the most common infections in humans. Uropathogenic Escherichia coli (UPEC) can invade and replicate within bladder epithelial cells, and some UPEC strains can also survive within macrophages. To understand the UPEC transcriptional programme associated with intramacrophage survival, we performed host–pathogen co-transcriptome analyses using RNA sequencing. Mouse bone marrow-derived macrophages (BMMs) were challenged over a 24 h time course with two UPEC reference strains that possess contrasting intramacrophage phenotypes: UTI89, which survives in BMMs, and 83972, which is killed by BMMs. Neither of these strains caused significant BMM cell death at the low multiplicity of infection that was used in this study. We developed an effective computational framework that simultaneously separated, annotated and quantified the mammalian and bacterial transcriptomes. Bone marrow-derived macrophages responded to the two UPEC strains with a broadly similar gene expression programme. In contrast, the transcriptional responses of the UPEC strains diverged markedly from each other. We identified UTI89 genes up-regulated at 24 h post-infection, and hypothesized that some may contribute to intramacrophage survival. Indeed, we showed that deletion of one such gene (pspA) significantly reduced UTI89 survival within BMMs. Our study provides a technological framework for simultaneously capturing global changes at the transcriptional level in co-cultures, and has generated new insights into the mechanisms that UPEC use to persist within the intramacrophage environment.

  1. HIV-1 Infection of T Cells and Macrophages Are Differentially Modulated by Virion-Associated Hck: A Nef-Dependent Phenomenon

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    Gilda Tachedjian

    2013-09-01

    Full Text Available The proline repeat motif (PxxP of Nef is required for interaction with the SH3 domains of macrophage-specific Src kinase Hck. However, the implication of this interaction for viral replication and infectivity in macrophages and T lymphocytes remains unclear. Experiments in HIV-1 infected macrophages confirmed the presence of a Nef:Hck complex which was dependent on the Nef proline repeat motif. The proline repeat motif of Nef also enhanced both HIV-1 infection and replication in macrophages, and was required for incorporation of Hck into viral particles. Unexpectedly, wild-type Hck inhibited infection of macrophages, but Hck was shown to enhance infection of primary T lymphocytes. These results indicate that the interaction between Nef and Hck is important for Nef-dependent modulation of viral infectivity. Hck-dependent enhancement of HIV-1 infection of T cells suggests that Nef-Hck interaction may contribute to the spread of HIV-1 infection from macrophages to T cells by modulating events in the producer cell, virion and target cell.

  2. Macrophage and T-cell gene expression in a model of early infection with the protozoan Leishmania chagasi.

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    Nicholas A Ettinger

    2008-06-01

    Full Text Available Visceral leishmaniasis is a potentially fatal infectious disease caused by the protozoan parasite Leishmania infantum/chagasi in the New World, or by L. donovani or L. infantum/chagasi in the Old World. Infection leads to a variety of outcomes ranging from asymptomatic infection to active disease, characterized by fevers, cachexia, hepatosplenomegaly and suppressed immune responses. We reasoned that events occurring during the initial few hours when the parasite encounters cells of the innate and adaptive immune systems are likely to influence the eventual immune response that develops. Therefore, we performed gene expression analysis using Affymetrix U133Plus2 microarray chips to investigate a model of early infection with human monocyte-derived macrophages (MDMs challenged with wild-type L. chagasi parasites, with or without subsequent co-culture with Leishmania-naïve, autologous T-cells. Microarray data generated from total RNA were analyzed with software from the Bioconductor Project and functional clustering and pathway analysis were performed with DAVID and Gene Set Enrichment Analysis (GSEA, respectively. Many transcripts were down-regulated by infection in cultures containing macrophages alone, and the pattern indicated a lack of a classically activated phenotype. By contrast, the addition of autologous Leishmania-naïve T cells to infected macrophages resulted in a pattern of gene expression including many markers of type 1 immune cytokine activation (IFN-gamma, IL-6, IL-1alpha, IL-1beta. There was simultaneous up-regulation of a few markers of immune modulation (IL-10 cytokine accumulation; TGF-beta Signaling Pathway. We suggest that the initial encounter between L. chagasi and cells of the innate and adaptive immune system stimulates primarily type 1 immune cytokine responses, despite a lack of classical macrophage activation. This local microenvironment at the site of parasite inoculation may determine the initial course of immune T

  3. MicroRNAs play big roles in modulating macrophages response toward mycobacteria infection.

    Science.gov (United States)

    Abdalla, Abualgasim Elgaili; Duan, Xiangke; Deng, Wanyan; Zeng, Jie; Xie, Jianping

    2016-11-01

    Macrophages are crucial player in the defense against multiple intracellular pathogens. Mycobacterium tuberculosis, the causative agent of tuberculosis which inflicted around one third of global population, can replicate and persist within macrophages. MicroRNAs, endogenous, small noncoding RNA, can regulate the expression of macrophages genes required for appropriate signaling. Mycobacteria can manipulate the expression of macrophages microRNAs to subvert cell response for its survival and persistence. This review summarized the progress of microRNAs in mycobacterial pathogenesis. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Antibiotic-Mediated Inhibition of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV Infection: A Novel Quinolone Function Which Potentiates the Antiviral Cytokine Response in MARC-145 Cells and Pig Macrophages

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    William A. Cafruny

    2008-01-01

    Full Text Available Porcine reproductive and respiratory syndrome virus (PRRSV is an economically significant agent for which there currently are no effective treatments. Development of antiviral agents for PRRSV as well as many other viruses has been limited by toxicity of known antiviral compounds. In contrast, antibiotics for non-virus microbial infections have been widely useful, in part because of their acceptable toxicity in animals. We report here the discovery that the quinolonecontaining compound Plasmocin™, as well as the quinolones nalidixic acid and ciprofloxacin, have potent anti-PRRSV activity in vitro. PRRSV replication was inhibited by these antibiotics in both cultured MARC-145 cells and cultured primary alveolar porcine macrophages (PAMs. Furthermore, sub-optimal concentrations of nalidixic acid synergized with antiviral cytokines (AK-2 or IFN-γ to quantitatively and qualitatively inhibit PRRSV replication in MARC-145 cells or PAMs. The antiviral activity of Plasmocin and nalidixic acid correlated with reduced actin expression in MARC-145 cells. Replication of the related lactate dehydrogenase-elevating virus (LDV was also inhibited in primary mouse macrophages by Plasmocin. These results are significant to the development of antiviral strategies with potentially reduced toxicity, and provide a model system to better understand regulation of arterivirus replication.

  5. Antibiotic-Mediated Inhibition of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV Infection: A Novel Quinolone Function Which Potentiates the Antiviral Cytokine Response in MARC-145 Cells and Pig Macrophages

    Directory of Open Access Journals (Sweden)

    William A. Cafruny

    2008-01-01

    Full Text Available Porcine reproductive and respiratory syndrome virus (PRRSV is an economically significant agent for which there currently are no effective treatments. Development of antiviral agents for PRRSV as well as many other viruses has been limited by toxicity of known antiviral compounds. In contrast, antibiotics for non-virus microbial infections have been widely useful, in part because of their acceptable toxicity in animals. We report here the discovery that the quinolone-containing compound Plasmocin ™ , as well as the quinolones nalidixic acid and ciprofloxacin, have potent anti-PRRSV activity in vitro. PRRSV replication was inhibited by these antibiotics in both cultured MARC-145 cells and cultured primary alveolar porcine macrophages (PAMs. Furthermore, sub-optimal concentrations of nalidixic acid synergized with antiviral cytokines (AK-2 or IFN-γ to quantitatively and qualitatively inhibit PRRSV replication in MARC-145 cells or PAMs. The antiviral activity of Plasmocin and nalidixic acid correlated with reduced actin expression in MARC-145 cells. Replication of the related lactate dehydrogenase-elevating virus (LDV was also inhibited in primary mouse macrophages by Plasmocin. These results are significant to the development of antiviral strategies with potentially reduced toxicity, and provide a model system to better understand regulation of arterivirus replication.

  6. Coarse(PM(2.5-10)), fine(PM(2.5)), and ultrafine air pollution particles induce/increase immune costimulatory receptors on human blood-derived monocytes but not on alveolar macrophages.

    Science.gov (United States)

    Becker, Susanne; Soukup, Joleen

    2003-05-09

    Diesel particles have been shown to possess adjuvant activity and influence the development of allergic sensitization. Also, more heterogeneous mixtures of pollution particles have been shown to affect host defenses and development of immunity in animal models. In the present study it was determined whether freshly collected particulate matter (PM(10)) in the size ranges 2.5-10 micro m (PM(2.5-10), coarse), 0.1-2.5 micro m (PM(2.5), fine), and micro m (ultrafine) in diameter affected the development of antigen presenting cells by evaluating the expression of surface receptors involved in T-cell interaction on both human alveolar macrophages (AM) and blood-derived monocytes (Mo). A Mo-AM coculture was exposed to 50 micro g/ml of particles and expression of HLA-DR, CD40, CD80, and CD86 on each cell type was assessed by flow cytometry. Mo upregulated the expression of all four receptors in response to each of the particle fractions, while expression was unaffected in AM. The cells were also exposed to two model air pollution particles, diesel dust and volcanic ash, neither of which affected receptor expression. Furthermore, Mo and AM were separately exposed to the three PM size fractions and supernatants assessed for the T-helper (CD4(+)) lymphocyte chemoattractant interleukin-16 (IL-16). AM, but not Mo, produced IL-16, and this chemoattractant was released only in response to PM(2.5-10). These data suggest that a wide size range of pollution particles contain materials that may promote antigen presentation by Mo, while the capability to specifically recruit CD4(+) lymphocytes is contained in AM stimulated with the coarse PM fraction.

  7. 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.

  8. Serial bronchoscopic lung lavage in pulmonary alveolar proteinosis under local anesthesia

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    K Rennis Davis

    2015-01-01

    Full Text Available Pulmonary alveolar proteinosis (PAP is a rare disease, characterized by alveolar accumulation of surfactant composed of proteins and lipids due to defective surfactant clearance by alveolar macrophages. Mainstay of treatment is whole lung lavage, which requires general anesthesia. Herein, we report a case of primary PAP, successfully treated with serial bronchoscopic lung lavages under local anesthesia.

  9. Lipoxin Inhibits Fungal Uptake by Macrophages and Reduces the Severity of Acute Pulmonary Infection Caused by Paracoccidioides brasiliensis

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    Laura R. R. Ribeiro

    2015-01-01

    Full Text Available Cysteinyl leukotrienes (CysLTs and lipoxins (LXs are lipid mediators that control inflammation, with the former inducing and the latter inhibiting this process. Because the role played by these mediators in paracoccidioidomycosis was not investigated, we aimed to characterize the role of CysLT in the pulmonary infection developed by resistant (A/J and susceptible (B10.A mice. 48 h after infection, elevated levels of pulmonary LTC4 and LXA4 were produced by both mouse strains, but higher levels were found in the lungs of susceptible mice. Blocking the CysLTs receptor by MTL reduced fungal loads in B10.A, but not in A/J mice. In susceptible mice, MLT treatment led to reduced influx of PMN leukocytes, increased recruitment of monocytes, predominant synthesis of anti-inflammatory cytokines, and augmented expression of 5- and 15-lipoxygenase mRNA, suggesting a prevalent LXA4 activity. In agreement, MTL-treated macrophages showed reduced fungal burdens associated with decreased ingestion of fungal cells. Furthermore, the addition of exogenous LX reduced, and the specific blockade of the LX receptor increased the fungal loads of B10.A macrophages. This study showed for the first time that inhibition of CysLTs signaling results in less severe pulmonary paracoccidioidomycosis that occurs in parallel with elevated LX activity and reduced infection of macrophages.

  10. Probing host pathogen cross-talk by transcriptional profiling of both Mycobacterium tuberculosis and infected human dendritic cells and macrophages.

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    Ludovic Tailleux

    2008-01-01

    Full Text Available Transcriptional profiling using microarrays provides a unique opportunity to decipher host pathogen cross-talk on the global level. Here, for the first time, we have been able to investigate gene expression changes in both Mycobacterium tuberculosis, a major human pathogen, and its human host cells, macrophages and dendritic cells.In addition to common responses, we could identify eukaryotic and microbial transcriptional signatures that are specific to the cell type involved in the infection process. In particular M. tuberculosis shows a marked stress response when inside dendritic cells, which is in accordance with the low permissivity of these specialized phagocytes to the tubercle bacillus and to other pathogens. In contrast, the mycobacterial transcriptome inside macrophages reflects that of replicating bacteria. On the host cell side, differential responses to infection in macrophages and dendritic cells were identified in genes involved in oxidative stress, intracellular vesicle trafficking and phagosome acidification.This study provides the proof of principle that probing the host and the microbe transcriptomes simultaneously is a valuable means to accessing unique information on host pathogen interactions. Our results also underline the extraordinary plasticity of host cell and pathogen responses to infection, and provide a solid framework to further understand the complex mechanisms involved in immunity to M. tuberculosis and in mycobacterial adaptation to different intracellular environments.

  11. Local and Systemic Functional Responses of Mouse Macrophages to Intravaginal Infection with Type 2 Herpes Simplex Virus and Vaccination.

    Science.gov (United States)

    Zaitseva, L G; Bekhalo, V A; Kireeva, I V; Shaposhnikova, G M; Nagurskaya, E V; Barinskii, I F; Nesterenko, V G

    2017-05-01

    Activity of cathepsin D and phagocytosis of macrophages from vaginal lavage fluid, peritoneal exudation, and spleen were studied in mice of sensitive (DBA/2) and resistant (BALB/c) lines after intravaginal infection with type 2 herpes simplex virus and vaccination. Activity of cathepsin D and intensity of phagocytosis (irrespective of the macrophage source) and their ratio in BALB/c mice in early terms after infection were close to the control levels taken as a unit. In DBA/2 mice, these parameters and their balance were shifted and changes in cathepsin D activity depended on the time after challenge. Activities of cellular and extracellular cathepsin D increased sharply on day 1 postinfection under conditions of local virus interaction with the vaginal mucosa and activation of the pathological process. Later, after generalization of the infection, activity of cathepsin D decreased, while phagocytosis increased in all the studied macrophage populations. Vaccination corrected the cathepsin D/phagocytosis imbalance and created conditions for rapid elimination of the virus.

  12. Emergence of quinolone-resistant, topoisomerase-mutant Brucella after treatment with fluoroquinolones in a macrophage experimental infection model.

    Science.gov (United States)

    Rodríguez Tarazona, Elisa; García Rodríguez, José Ángel; Muñoz Bellido, Juan Luis

    2015-04-01

    To determine the activity of fluoroquinolones (FQ) and the selection of FQ-resistant mutants in a macrophage experimental infection model (MEIM). Canine macrophages were inoculated with Brucella melitensis ATCC 23457 (WT), achieving intracellular counts of around 105 CFU/mL. Cell cultures were incubated in the presence of ciprofloxacin (CIP), levofloxacin (LEV), moxifloxacin (MOX), and doxycycline (DOX). After cell lysis, surviving microorganisms were plated for count purposes, and plated onto antibiotics-containing media for mutant selection. Topoisomerases mutations were detected by PCR and sequencing. Bacterial counts after cell lysis were 14.3% (CIP), 65.3% (LEV), and 75% (MOX) lower compared to the control. Quinolone-resistant mutants emerged in cell cultures containing CIP and LEV with a frequency of around 0.5×10(-3). All mutants showed an Ala87Val change in GyrA. Mutants had FQs MICs around 10×WT. The ability of these mutants for infecting new macrophages and the intracellular lysis after antibiotic exposure did not change significantly. No 2nd step FQ-resistant mutants were selected from 1st step mutants. Intracellular activity of FQs is low against WT and gyrA-mutant Brucella. FQs easily select gyrA mutants in MEIM. The ability of mutants for infecting new macrophages remains unchanged. In this MEIM, 2nd step mutants do not emerge. Copyright © 2014 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  13. Alveolar inflammation in cystic fibrosis

    DEFF Research Database (Denmark)

    Ulrich, Martina; Worlitzsch, Dieter; Viglio, Simona

    2010-01-01

    BACKGROUND: In infected lungs of the cystic fibrosis (CF) patients, opportunistic pathogens and mutated cystic fibrosis transmembrane conductance regulator protein (CFTR) contribute to chronic airway inflammation that is characterized by neutrophil/macrophage infiltration, cytokine release...

  14. Innate invariant NKT cells recognize Mycobacterium tuberculosis-infected macrophages, produce interferon-gamma, and kill intracellular bacteria.

    Directory of Open Access Journals (Sweden)

    Isabel Sada-Ovalle

    2008-12-01

    Full Text Available Cellular immunity to Mycobacterium tuberculosis (Mtb requires a coordinated response between the innate and adaptive arms of the immune system, resulting in a type 1 cytokine response, which is associated with control of infection. The contribution of innate lymphocytes to immunity against Mtb remains controversial. We established an in vitro system to study this question. Interferon-gamma is produced when splenocytes from uninfected mice are cultured with Mtb-infected macrophages, and, under these conditions, bacterial replication is suppressed. This innate control of bacterial replication is dependent on CD1d-restricted invariant NKT (iNKT cells, and their activation requires CD1d expression by infected macrophages as well as IL-12 and IL-18. We show that iNKT cells, even in limiting quantities, are sufficient to restrict Mtb replication. To determine whether iNKT cells contribute to host defense against tuberculosis in vivo, we adoptively transferred iNKT cells into mice. Primary splenic iNKT cells obtained from uninfected mice significantly reduce the bacterial burden in the lungs of mice infected with virulent Mtb by the aerosol route. Thus, iNKT cells have a direct bactericidal effect, even in the absence of synthetic ligands such as alpha-galactosylceramide. Our finding that iNKT cells protect mice against aerosol Mtb infection is the first evidence that CD1d-restricted NKT cells mediate protection against Mtb in vivo.

  15. Role of Granulocyte-Macrophage Colony-Stimulating Factor Production by T Cells duringMycobacterium tuberculosisInfection.

    Science.gov (United States)

    Rothchild, Alissa C; Stowell, Britni; Goyal, Girija; Nunes-Alves, Cláudio; Yang, Qianting; Papavinasasundaram, Kadamba; Sassetti, Christopher M; Dranoff, Glenn; Chen, Xinchun; Lee, Jinhee; Behar, Samuel M

    2017-10-24

    Mice deficient for granulocyte-macrophage colony-stimulating factor (GM-CSF -/- ) are highly susceptible to infection with Mycobacterium tuberculosis , and clinical data have shown that anti-GM-CSF neutralizing antibodies can lead to increased susceptibility to tuberculosis in otherwise healthy people. GM-CSF activates human and murine macrophages to inhibit intracellular M. tuberculosis growth. We have previously shown that GM-CSF produced by iNKT cells inhibits growth of M. tuberculosis However, the more general role of T cell-derived GM-CSF during infection has not been defined and how GM-CSF activates macrophages to inhibit bacterial growth is unknown. Here we demonstrate that, in addition to nonconventional T cells, conventional T cells also produce GM-CSF during M. tuberculosis infection. Early during infection, nonconventional iNKT cells and γδ T cells are the main source of GM-CSF, a role subsequently assumed by conventional CD4 + T cells as the infection progresses. M. tuberculosis -specific T cells producing GM-CSF are also detected in the peripheral blood of infected people. Under conditions where nonhematopoietic production of GM-CSF is deficient, T cell production of GM-CSF is protective and required for control of M. tuberculosis infection. However, GM-CSF is not required for T cell-mediated protection in settings where GM-CSF is produced by other cell types. Finally, using an in vitro macrophage infection model, we demonstrate that GM-CSF inhibition of M. tuberculosis growth requires the expression of peroxisome proliferator-activated receptor gamma (PPARγ). Thus, we identified GM-CSF production as a novel T cell effector function. These findings suggest that a strategy augmenting T cell production of GM-CSF could enhance host resistance against M. tuberculosis IMPORTANCE Mycobacterium tuberculosis is the bacterium that causes tuberculosis, the leading cause of death by any infection worldwide. T cells are critical components of the immune

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

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    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.

  17. miR-126-5p by direct targeting of JNK-interacting protein-2 (JIP-2) plays a key role in Theileria-infected macrophage virulence

    KAUST Repository

    Haidar, Malak

    2018-03-23

    Theileria annulata is an apicomplexan parasite that infects and transforms bovine macrophages that disseminate throughout the animal causing a leukaemia-like disease called tropical theileriosis. Using deep RNAseq of T. annulata-infected B cells and macrophages we identify a set of microRNAs induced by infection, whose expression diminishes upon loss of the hyper-disseminating phenotype of virulent transformed macrophages. We describe how infection-induced upregulation of miR-126-5p ablates JIP-2 expression to release cytosolic JNK to translocate to the nucleus and trans-activate AP-1-driven transcription of mmp9 to promote tumour dissemination. In non-disseminating attenuated macrophages miR-126-5p levels drop, JIP-2 levels increase, JNK1 is retained in the cytosol leading to decreased c-Jun phosphorylation and dampened AP-1-driven mmp9 transcription. We show that variation in miR-126-5p levels depends on the tyrosine phosphorylation status of AGO2 that is regulated by Grb2-recruitment of PTP1B. In attenuated macrophages Grb2 levels drop resulting in less PTP1B recruitment, greater AGO2 phosphorylation, less miR-126-5p associated with AGO2 and a consequent rise in JIP-2 levels. Changes in miR-126-5p levels therefore, underpin both the virulent hyper-dissemination and the attenuated dissemination of T. annulata-infected macrophages.

  18. Macrophagic control of the response to uropathogenic E. coli infection by regulation of iron retention in an IL‐6‐dependent manner

    Science.gov (United States)

    Owusu‐Boaitey, Nana; Bauckman, Kyle A.; Zhang, Tingxuan

    2016-01-01

    Abstract Introduction Uropathogenic Escherichia coli (UPEC), the causative agent of over 85% of urinary tract infections (UTIs), elaborate a number of siderophores to chelate iron from the host. On the other hand, the host immune imperative is to limit the availability of iron to the bacteria. Little is known regarding the mechanisms underlying this host‐iron‐UPEC interaction. Our objective was to determine whether macrophages, in response to UPEC infection, retain extracellular siderophore‐bound and free iron, thus limiting the ability of UPEC to access iron. Methods Quantitative PCR, immunoblotting analysis, and gene expression analysis of wild type and IL‐6‐deficient macrophages was performed. Results We found that (1) macrophages upon UPEC infection increased expression of lipocalin 2, a siderophore‐binding molecule, of Dmt1, a molecule that facilitates macrophage uptake of free iron, and of the intracellular iron cargo molecule ferritin, and decreased expression of the iron exporter ferroportin; (2) bladder macrophages regulate expression of genes involved in iron retention upon UPEC infection; (3) IL‐6, a cytokine known to play an important role in regulating host iron homeostasis as well as host defense to UPEC, regulates this process, in part by promoting production of lipocalin 2; and finally, (4) inhibition of IL‐6 signaling genetically and by neutralizing antibodies against the IL‐6 receptor, promoted intra‐macrophagic UPEC growth in the presence of excess iron. Conclusions Together, our study suggests that macrophages retain siderophore‐bound and free iron in response to UPEC and IL‐6 signaling is necessary for macrophages to limit the growth of UPEC in the presence of excess iron. IL‐6 signaling and iron regulation is one mechanism by which macrophages may mediate UPEC clearance. PMID:27980776

  19. Modulation of monocyte/macrophage-derived cytokine and chemokine profile by persistent Hepatitis C virus (HCV infection leads to chronic inflammation

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    Penelope Mavromara

    2012-02-01

    Full Text Available HCV infection presents a major public health problem, with more than 170 million people infected worldwide. Chronicity and persistence of infection constitute the hallmark of the disease. Although HCV is a hepatotropic virus, subsets of immune cells have been found to be permissive to infection and viral replication. Peripheral blood monocytes, attracted to the site of infection and differentiated into macrophages, and resident hepatic macrophages, known as Kupffer cells, are important mediators of innate immunity, through production of several chemokines and cytokines in addition to their phagocytic activity. HCV proteins have been shown to modulate the cytokine and chemokine production profile of monocytes/macrophages, as it is suggested by both in vitro and clinical studies. This modified expression profile appears crucial for the establishment of aberrant inflammation that leads to liver cirrhosis and hepatocellular carcinoma.

  20. The Local Inflammatory Responses to Infection of the Peritoneal Cavity in Humans: Their Regulation by Cytokines, Macrophages, and Other Leukocytes

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    Marien Willem Johan Adriaan Fieren

    2012-01-01

    Full Text Available Studies on infection-induced inflammatory reactions in humans rely largely on findings in the blood compartment. Peritoneal leukocytes from patients treated with peritoneal dialysis offer a unique opportunity to study in humans the inflammatory responses taking place at the site of infection. Compared with peritoneal macrophages (pM from uninfected patients, pM from infected patients display ex vivo an upregulation and downregulation of proinflammatory and anti-inflammatory mediators, respectively. Pro-IL-1 processing and secretion rather than synthesis proves to be increased in pM from infectious peritonitis suggesting up-regulation of caspase-1 in vivo. A crosstalk between pM, γ T cells, and neutrophils has been found to be involved in augmented TNF expression and production during infection. The recent finding in experimental studies that alternatively activated macrophages (M2 increase by proliferation rather than recruitment may have significant implications for the understanding and treatment of chronic inflammatory conditions such as encapsulating peritoneal sclerosis (EPS.

  1. Protection against type 1 diabetes upon Coxsackievirus B4 infection and iNKT-cell stimulation: role of suppressive macrophages.

    Science.gov (United States)

    Ghazarian, Liana; Diana, Julien; Beaudoin, Lucie; Larsson, Pär G; Puri, Raj K; van Rooijen, Nico; Flodström-Tullberg, Malin; Lehuen, Agnès

    2013-11-01

    Invariant natural killer T (iNKT) cells belong to the innate immune system and exercise a dual role as potent regulators of autoimmunity and participate in responses against different pathogens. They have been shown to prevent type 1 diabetes development and to promote antiviral responses. Many studies in the implication of environmental factors on the etiology of type 1 diabetes have suggested a link between enteroviral infections and the development of this disease. This study of the pancreatropic enterovirus Coxsackievirus B4 (CVB4) shows that although infection accelerated type 1 diabetes development in a subset of proinsulin 2-deficient NOD mice, the activation of iNKT cells by a specific agonist, α-galactosylceramide, at the time of infection inhibited the disease. Diabetes development was associated with the infiltration of pancreatic islets by inflammatory macrophages, producing high levels of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α and activation of anti-islet T cells. On the contrary, macrophages infiltrating the islets after CVB4 infection and iNKT-cell stimulation expressed a number of suppressive enzymes, among which indoleamine 2,3-dioxygenase was sufficient to inhibit anti-islet T-cell response and to prevent diabetes. This study highlights the critical interaction between virus and the immune system in the acceleration or prevention of type 1 diabetes.

  2. Fungal infection control by garlic extracts (Allium sativum L.) and modulation of peritoneal macrophages activity in murine model of sporotrichosis.

    Science.gov (United States)

    Burian, J P; Sacramento, L V S; Carlos, I Z

    2017-11-01

    Garlic (Allium sativum L.) is grown all over the world as seasoning and medicinal vegetable since 3,000 BC. Allicin is the main component of garlic, being attributed to it the most of its biological activities, such as bactericidal, antifungal and antiviral actions. However, other compounds of garlic present antioxidant, hypocholesterolemic, vasodilator activities, protective action against different types of cancer, and immunomodulatory. Fungal infections are important causes of morbidity and mortality in people mainly in immunosuppressed ones. Sporothrix schenckii, the causing agent of Sporotrichosis (most common subcutaneous mycosis in Latin America), is dimorphic fungus, of saprophytic life in soil or plants, infecting people and animals mainly through skin injuries and bruises. The main of this work was to evaluate the influence of garlic consuming on immune modulation of healthy and infected Swiss mice in induced way by S. schenckii, since these animals functioning of peritoneal macrophages as well as the nitric oxide and cytokines' production (IL-1β, IL-10 and IL-12) and to evaluate the antifungal potential of garlic with S. schenckii through minimum inhibitory concentration test and colony-forming units. The results showed that garlic offers antifungal potential with S. schenckii. The oral taking of garlic extracts influences the releasing of cytokines by macrophages, regular consuming shows anti-inflammatory effect, and its acute use may take to an inflammatory response. Mice that consumed garlic responded more effectively to fight against the infection.

  3. Macrophages as target cells for Mayaro virus infection: involvement of reactive oxygen species in the inflammatory response during virus replication

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    MARIANA G. CAVALHEIRO

    Full Text Available ABSTRACT Alphaviruses among the viruses that cause arthritis, consisting in a public health problem worldwide by causing localized outbreaks, as well as large epidemics in humans. Interestingly, while the Old World alphaviruses are arthritogenic, the New World alphaviruses cause encephalitis. One exception is Mayaro virus (MAYV, which circulates exclusively in South America but causes arthralgia and is phylogenetically related to the Old World alphaviruses. Although MAYV-induced arthritis in humans is well documented, the molecular and cellular factors that contribute to its pathogenesis are completely unknown. In this study, we demonstrated for the first time that macrophages, key players in arthritis development, are target cells for MAYV infection, which leads to cell death through apoptosis. We showed that MAYV replication in macrophage induced the expression of TNF, a cytokine that would contribute to pathogenesis of MAYV fever, since TNF promotes an inflammatory profile characteristic of arthritis. We also found a significant increase in the production of reactive oxygen species (ROS at early times of infection, which coincides with the peak of virus replication and precedes TNF secretion. Treatment of the cells with antioxidant agents just after infection completely abolished TNF secretion, indicating an involvement of ROS in inflammation induced during MAYV infection.

  4. Fungal infection control by garlic extracts (Allium sativum L. and modulation of peritoneal macrophages activity in murine model of sporotrichosis

    Directory of Open Access Journals (Sweden)

    J. P. Burian

    2017-05-01

    Full Text Available Abstract Garlic (Allium sativum L. is grown all over the world as seasoning and medicinal vegetable since 3,000 BC. Allicin is the main component of garlic, being attributed to it the most of its biological activities, such as bactericidal, antifungal and antiviral actions. However, other compounds of garlic present antioxidant, hypocholesterolemic, vasodilator activities, protective action against different types of cancer, and immunomodulatory. Fungal infections are important causes of morbidity and mortality in people mainly in immunosuppressed ones. Sporothrix schenckii, the causing agent of Sporotrichosis (most common subcutaneous mycosis in Latin America, is dimorphic fungus, of saprophytic life in soil or plants, infecting people and animals mainly through skin injuries and bruises. The main of this work was to evaluate the influence of garlic consuming on immune modulation of healthy and infected Swiss mice in induced way by S. schenckii, since these animals functioning of peritoneal macrophages as well as the nitric oxide and cytokines’ production (IL-1β, IL-10 and IL-12 and to evaluate the antifungal potential of garlic with S. schenckii through minimum inhibitory concentration test and colony-forming units. The results showed that garlic offers antifungal potential with S. schenckii. The oral taking of garlic extracts influences the releasing of cytokines by macrophages, regular consuming shows anti-inflammatory effect, and its acute use may take to an inflammatory response. Mice that consumed garlic responded more effectively to fight against the infection.

  5. Vaccination reduces macrophage infiltration in bronchus-associated lymphoid tissue in pigs infected with a highly virulent Mycoplasma hyopneumoniae strain

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    Vranckx Katleen

    2012-03-01

    Full Text Available Abstract Background Mycoplasma hyopneumoniae is the causative agent of enzootic pneumonia and is responsible for significant economic losses to the pig industry. To better understand the mode of action of a commercial, adjuvanted, inactivated whole cell vaccine and the influence of diversity on the efficacy of vaccination, we investigated samples from vaccinated and non-vaccinated pigs experimentally infected with either a low (LV or a highly virulent (HV M. hyopneumoniae strain. Non-vaccinated and sham-infected control groups were included. Lung tissue samples collected at 4 and 8 weeks post infection (PI were immunohistochemically tested for the presence of T-lymphocytes, B-lymphocytes and macrophages in the bronchus-associated lymphoid tissue (BALT. The number of M. hyopneumoniae organisms in bronchoalveolar lavage (BAL fluid was determined using quantitative PCR at 4 and 8 weeks PI. Serum antibodies against M. hyopneumoniae were determined at 0, 2, 4, 6 and 8 weeks PI. Results The immunostaining revealed a lower density of macrophages in the BALT of the vaccinated groups compared to the non-vaccinated groups. The highest number of M. hyopneumoniae organisms in the BAL fluid was measured at 4 weeks PI for the HV strain and at 8 weeks PI for the LV strain. Vaccination reduced the number of organisms non-significantly, though for the HV strain the reduction was clinically more relevant than for the LV strain. At the level of the individual pigs, a higher lung lesion score was associated with more M. hyopneumoniae organisms in the lungs and a higher density of the investigated immune cells in the BALT. Conclusions In conclusion, the infiltration of macrophages after infection with M. hyopneumoniae is reduced by vaccination. The M. hyopneumoniae replication in the lungs is also reduced in vaccinated pigs, though the HV strain is inhibited more than the LV strain.

  6. Vaccination reduces macrophage infiltration in bronchus-associated lymphoid tissue in pigs infected with a highly virulent Mycoplasma hyopneumoniae strain.

    Science.gov (United States)

    Vranckx, Katleen; Maes, Dominiek; Marchioro, Silvana B; Villarreal, Iris; Chiers, Koen; Pasmans, Frank; Haesebrouck, Freddy

    2012-03-12

    Mycoplasma hyopneumoniae is the causative agent of enzootic pneumonia and is responsible for significant economic losses to the pig industry. To better understand the mode of action of a commercial, adjuvanted, inactivated whole cell vaccine and the influence of diversity on the efficacy of vaccination, we investigated samples from vaccinated and non-vaccinated pigs experimentally infected with either a low (LV) or a highly virulent (HV) M. hyopneumoniae strain. Non-vaccinated and sham-infected control groups were included. Lung tissue samples collected at 4 and 8 weeks post infection (PI) were immunohistochemically tested for the presence of T-lymphocytes, B-lymphocytes and macrophages in the bronchus-associated lymphoid tissue (BALT). The number of M. hyopneumoniae organisms in bronchoalveolar lavage (BAL) fluid was determined using quantitative PCR at 4 and 8 weeks PI. Serum antibodies against M. hyopneumoniae were determined at 0, 2, 4, 6 and 8 weeks PI. The immunostaining revealed a lower density of macrophages in the BALT of the vaccinated groups compared to the non-vaccinated groups. The highest number of M. hyopneumoniae organisms in the BAL fluid was measured at 4 weeks PI for the HV strain and at 8 weeks PI for the LV strain. Vaccination reduced the number of organisms non-significantly, though for the HV strain the reduction was clinically more relevant than for the LV strain. At the level of the individual pigs, a higher lung lesion score was associated with more M. hyopneumoniae organisms in the lungs and a higher density of the investigated immune cells in the BALT. In conclusion, the infiltration of macrophages after infection with M. hyopneumoniae is reduced by vaccination. The M. hyopneumoniae replication in the lungs is also reduced in vaccinated pigs, though the HV strain is inhibited more than the LV strain.

  7. Transcriptional Analysis of Murine Macrophages Infected with Different Toxoplasma Strains Identifies Novel Regulation of Host Signaling Pathways

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    Melo, Mariane B.; Nguyen, Quynh P.; Cordeiro, Cynthia; Hassan, Musa A.; Yang, Ninghan; McKell, Renée; Rosowski, Emily E.; Julien, Lindsay; Butty, Vincent; Dardé, Marie-Laure; Ajzenberg, Daniel; Fitzgerald, Katherine; Young, Lucy H.; Saeij, Jeroen P. J.

    2013-01-01

    Most isolates of Toxoplasma from Europe and North America fall into one of three genetically distinct clonal lineages, the type I, II and III lineages. However, in South America these strains are rarely isolated and instead a great variety of other strains are found. T. gondii strains differ widely in a number of phenotypes in mice, such as virulence, persistence, oral infectivity, migratory capacity, induction of cytokine expression and modulation of host gene expression. The outcome of toxoplasmosis in patients is also variable and we hypothesize that, besides host and environmental factors, the genotype of the parasite strain plays a major role. The molecular basis for these differences in pathogenesis, especially in strains other than the clonal lineages, remains largely unexplored. Macrophages play an essential role in the early immune response against T. gondii and are also the cell type preferentially infected in vivo. To determine if non-canonical Toxoplasma strains have unique interactions with the host cell, we infected murine macrophages with 29 different Toxoplasma strains, representing global diversity, and used RNA-sequencing to determine host and parasite transcriptomes. We identified large differences between strains in the expression level of known parasite effectors and large chromosomal structural variation in some strains. We also identified novel strain-specifically regulated host pathways, including the regulation of the type I interferon response by some atypical strains. IFNβ production by infected cells was associated with parasite killing, independent of interferon gamma activation, and dependent on endosomal Toll-like receptors in macrophages and the cytoplasmic receptor retinoic acid-inducible gene 1 (RIG-I) in fibroblasts. PMID:24367253

  8. Histone deacetylase inhibitor MS-275 augments expression of a subset of IFN-γ-regulated genes in Toxoplasma gondii-infected macrophages but does not improve parasite control.

    Science.gov (United States)

    Sumpf, Kristina; Nast, Roswitha; Downie, Bryan; Salinas, Gabriela; Lüder, Carsten G K

    2017-09-01

    Toxoplasma gondii is a ubiquitous apicomplexan parasite of mammals and birds and an important pathogen of humans. IFN-γ is the major mediator of host resistance against T. gondii but intriguingly, parasite-infected host cells including macrophages are severely impaired to respond to IFN-γ due to defective transcriptional activation of target genes. Here, we tested the possibility that the impaired responsiveness of T. gondii-infected macrophages to IFN-γ can be restored by inhibiting histone deacetylases (HDACs) using the class I-specific inhibitor MS-275. Treatment of RAW264.7 cells with MS-275 indeed increased MHC class II surface expression in infected and non-infected cells and largely abolished the inhibition of IFN-γ-regulated MHC class II expression exerted by T. gondii. Genome-wide transcriptome profiling revealed that MS-275 increased mean mRNA levels of IFN-γ-regulated genes particularly in non-infected macrophages. Transcript levels of 33% of IFN-γ secondary response genes but only those of a few primary response genes were also increased by MS-275 in T. gondii-infected cells. Importantly, the unresponsiveness of parasite-infected cells to IFN-γ was however not abolished by MS-275. Furthermore, MS-275 also up-regulated several anti-inflammatory cytokines or signaling molecules in T. gondii-infected macrophages. It additionally regulated expression of more than 2500 genes in non-infected macrophages expression of which was surprisingly counteracted by prior infection with T. gondii. FACS analysis and immunofluorescence microscopy revealed that MS-275 did not considerably diminish the number of parasite-positive cells or the intracellular replication in macrophages stimulated or not with IFN-γ. Thus, a supportive therapy using MS-275 appears inappropriate for treatment of toxoplasmosis. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. 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.

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    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.

  10. Modeling of HIV-1 infection: insights to the role of monocytes/macrophages, latently infected T4 cells, and HAART regimes.

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    Qiang Li

    Full Text Available A novel dynamic model covering five types of cells and three connected compartments, peripheral blood (PB, lymph nodes (LNs, and the central nervous system (CNS, is here proposed. It is based on assessment of the biological principles underlying the interactions between the human immunodeficiency virus type I (HIV-1 and the human immune system. The simulated results of this model matched the three well-documented phases of HIV-1 infection very closely and successfully described the three stages of LN destruction that occur during HIV-1 infection. The model also showed that LNs are the major location of viral replication, creating a pool of latently infected T4 cells during the latency period. A detailed discussion of the role of monocytes/macrophages is made, and the results indicated that infected monocytes/macrophages could determine the progression of HIV-1 infection. The effects of typical highly active antiretroviral therapy (HAART drugs on HIV-1 infection were analyzed and the results showed that efficiency of each drug but not the time of the treatment start contributed to the change of the turnover of the disease greatly. An incremental count of latently infected T4 cells was made under therapeutic simulation, and patients were found to fail to respond to HAART therapy in the presence of certain stimuli, such as opportunistic infections. In general, the dynamics of the model qualitatively matched clinical observations very closely, indicating that the model may have benefits in evaluating the efficacy of different drug therapy regimens and in the discovery of new monitoring markers and therapeutic schemes for the treatment of HIV-1 infection.

  11. Fusobacterium nucleatum and Tannerella forsythia Induce Synergistic Alveolar Bone Loss in a Mouse Periodontitis Model

    Science.gov (United States)

    Settem, Rajendra P.; El-Hassan, Ahmed Taher; Honma, Kiyonobu; Stafford, Graham P.

    2012-01-01

    Tannerella forsythia is strongly associated with chronic periodontitis, an inflammatory disease of the tooth-supporting tissues, leading to tooth loss. Fusobacterium nucleatum, an opportunistic pathogen, is thought to promote dental plaque formation by serving as a bridge bacterium between early- and late-colonizing species of the oral cavity. Previous studies have shown that F. nucleatum species synergize with T. forsythia during biofilm formation and pathogenesis. In the present study, we showed that coinfection of F. nucleatum and T. forsythia is more potent than infection with either species alone in inducing NF-κB activity and proinflammatory cytokine secretion in monocytic cells and primary murine macrophages. Moreover, in a murine model of periodontitis, mixed infection with the two species induces synergistic alveolar bone loss, characterized by bone loss which is greater than the additive alveolar bone losses induced by each species alone. Further, in comparison to the single-species infection, mixed infection caused significantly increased inflammatory cell infiltration in the gingivae and osteoclastic activity in the jaw bones. These data show that F. nucleatum subspecies and T. forsythia synergistically stimulate the host immune response and induce alveolar bone loss in a murine experimental periodontitis model. PMID:22547549

  12. Herpes simplex virus type 2 infection increases human immunodeficiency virus type 1 entry into human primary macrophages

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    Palù Giorgio

    2011-04-01

    Full Text Available Abstract Epidemiological and clinical data indicate that genital ulcer disease (GUD pathogens are associated with an increased risk of human immunodeficiency virus type 1 (HIV-1 acquisition and/or transmission. Among them, genital herpes simplex virus type 2 (HSV-2 seems to play a relevant role. Indeed, the ability of HSV-2 to induce massive infiltration at the genital level of cells which are potential targets for HIV-1 infection may represent one of the mechanisms involved in this process. Here we show that infection of human primary macrophages (MDMs by HSV-2 results in an increase of CCR5 expression levels on cell surface and allows higher efficiency of MDMs to support entry of R5 HIV-1 strains. This finding could strengthen, at the molecular level, the evidence linking HSV-2 infection to an increased susceptibility to HIV-1 acquisition.

  13. Herpes simplex virus type 2 infection increases human immunodeficiency virus type 1 entry into human primary macrophages.

    Science.gov (United States)

    Sartori, Elena; Calistri, Arianna; Salata, Cristiano; Del Vecchio, Claudia; Palù, Giorgio; Parolin, Cristina

    2011-04-12

    Epidemiological and clinical data indicate that genital ulcer disease (GUD) pathogens are associated with an increased risk of human immunodeficiency virus type 1 (HIV-1) acquisition and/or transmission. Among them, genital herpes simplex virus type 2 (HSV-2) seems to play a relevant role. Indeed, the ability of HSV-2 to induce massive infiltration at the genital level of cells which are potential targets for HIV-1 infection may represent one of the mechanisms involved in this process. Here we show that infection of human primary macrophages (MDMs) by HSV-2 results in an increase of CCR5 expression levels on cell surface and allows higher efficiency of MDMs to support entry of R5 HIV-1 strains. This finding could strengthen, at the molecular level, the evidence linking HSV-2 infection to an increased susceptibility to HIV-1 acquisition.

  14. Long-term effects of neonatal malnutrition on microbicide response, production of cytokines, and survival of macrophages infected by Staphylococcus aureus sensitive/resistant to methicillin

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    Natália Gomes de Morais

    2014-10-01

    Full Text Available OBJECTIVE: To assess microbicide function and macrophage viability after in vitro cellular infection by methicillin-sensitive/resistant Staphylococcus aureus in nourished rats and rats subjected to neonatal malnutrition. METHODS: Male Wistar rats (n=40 were divided in two groups: Nourished (rats suckled by dams consuming a 17% casein diet and Malnourished (rats suckled by dams consuming an 8% casein diet. Macrophages were recovered after tracheotomy, by bronchoalveolar lavage. After mononuclear cell isolation, four systems were established: negative control composed exclusively of phagocytes; positive control composed of macrophages plus lipopolysaccharide; and two testing systems, macrophages plus methicillin-sensitive Staphylococcus aureus and macrophages plus methicillin-resistant Staphylococcus aureus. The plates were incubated in a humid atmosphere at 37 degrees Celsius containing 5% CO2 for 24 hours. After this period tests the microbicidal response, cytokine production, and cell viability were analyzed. The statistical analysis consisted of analysis of variance (p<0.05. RESULTS: Malnutrition reduced weight gain, rate of phagocytosis, production of superoxide anion and nitric oxide, and macrophage viability. Production of nitrite and interleukin 18, and viability of macrophages infected with methicillin-resistant Staphylococcus aureus were lower. CONCLUSION: The neonatal malnutrition model compromised phagocyte function and reduced microbicidal response and cell viability. Interaction between malnutrition and the methicillin-resistant strain decreased the production of inflammatory mediators by effector cells of the immune response, which may compromise the immune system's defense ability.

  15. Persistent Arthralgia Induced by Chikungunya Virus Infection is Associated with Interleukin-6 and Granulocyte Macrophage Colony-Stimulating Factor

    Science.gov (United States)

    Chow, Angela; Her, Zhisheng; Ong, Edward K. S.; Chen, Jin-miao; Dimatatac, Frederico; Kwek, Dyan J. C.; Barkham, Timothy; Yang, Henry; Rénia, Laurent; Leo, Yee-Sin

    2011-01-01

    Background. Chikungunya virus (CHIKV) infection induces arthralgia. The involvement of inflammatory cytokines and chemokines has been suggested, but very little is known about their secretion profile in CHIKV-infected patients. Methods. A case-control longitudinal study was performed that involved 30 adult patients with laboratory-confirmed Chikungunya fever. Their profiles of clinical disease, viral load, and immune mediators were investigated. Results. When patients were segregated into high viral load and low viral load groups during the acute phase, those with high viremia had lymphopenia, lower levels of monocytes, neutrophilia, and signs of inflammation. The high viral load group was also characterized by a higher production of pro-inflammatory cytokines, such as interferon-α and interleukin (IL)–6, during the acute phase. As the disease progressed to the chronic phase, IL-17 became detectable. However, persistent arthralgia was associated with higher levels of IL-6 and granulocyte macrophage colony-stimulating factor, whereas patients who recovered fully had high levels of Eotaxin and hepatocyte growth factor. Conclusions. The level of CHIKV viremia during the acute phase determined specific patterns of pro-inflammatory cytokines, which were associated with disease severity. At the chronic phase, levels of IL-6, and granulocyte macrophage colony-stimulating factor found to be associated with persistent arthralgia provide a possible explanation for the etiology of arthralgia that plagues numerous CHIKV-infected patients. PMID:21288813

  16. Novel incision design and primary flap closure reduces the incidence of alveolar osteitis and infection in impacted mandibular third molar surgery.

    Science.gov (United States)

    Elo, Jeffrey A; Sun, Ho-Hyun Brian; Dong, Fanglong; Tandon, Rahul; Singh, Hardev M

    2016-08-01

    To measure the influence of flap design on alveolar osteitis (AO) and postoperative complications following mandibular third molar surgery. This study was designed as a randomized single-blind, split-mouth clinical trial. The predictor variable was flap type. Envelope flap, modified triangular flap (MTF), and two related experimental flaps (second molar mesial papilla-sparing marginal incision with distobuccal release with double-pass single-layered primary closure [MPMI-2 X] and single-pass single-layered primary closure [MPMI-1 X]) were used. The primary outcome variable was AO. The secondary outcome variables were wound dehiscence and infection. Bivariate and logistic analyses were computed. P value third molars participated. No sites (0 of 196) treated with MPMI-2 X developed AO, and only two sites (2 of 196) treated with MPMI-2 X developed postoperative infection. Both MPMI-1 X and MPMI-2 X were associated with decreased odds of complications compared with MTF and envelope flap. MPMI-2 X sites were significantly less likely than MTF sites to experience complications for both sides. MPMI-2 X is a reliable technique to reduce complications, such as AO, wound dehiscence, and infection in mandibular third molar surgery. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Circulating and broncho-alveolar interleukin-6 in relation to body temperature in an experimental model of bovine Chlamydia psittaci infection.

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    Annette Prohl

    Full Text Available In rodent models of experimentally induced fever, the important role of interleukin-6 (IL-6 as a circulating endogenous pyrogen is well established. Studies employing larger animal species and real infections are scarce. Therefore, we assessed bioactive IL-6 in peripheral blood and in broncho-alveolar lavage fluid (BALF of calves after intra-bronchial inoculation with vital Chlamydia psittaci (Cp, with inactivated Cp, or with BGM cells. Only calves inoculated with vital Cp developed fever (peak at 2-3 days after challenge and significantly increased IL-6 activity. Controls inoculated with either inactivated Cp or BGM cells also expressed increased bioactive IL-6, but no fever developed. Activity of IL-6 in BALF was significantly higher compared to blood serum. This experimental model of Cp infection revealed no apparent relation between IL-6 in blood and body temperature, but did reveal a relation between IL-6 and other markers of inflammation in BALF. We conclude that a local inflammatory response in the lungs of infected calves caused fever, which developed by mechanisms including other mediators besides IL-6.

  18. Expression of bacterial virulence factors and cytokines during in vitro macrophage infection by enteroinvasive Escherichia coli and Shigella flexneri: a comparative study

    Directory of Open Access Journals (Sweden)

    Silvia Y Bando

    2010-09-01

    Full Text Available Enteroinvasive Escherichia coli (EIEC and Shigellaspp cause bacillary dysentery in humans by invading and multiplying within epithelial cells of the colonic mucosa. Although EIEC and Shigellashare many genetic and biochemical similarities, the illness caused by Shigellais more severe. Thus, genomic and structure-function molecular studies on the biological interactions of these invasive enterobacteria with eukaryotic cells have focused on Shigella rather than EIEC. Here we comparatively studied the interactions of EIEC and of Shigella flexneriwith cultured J774 macrophage-like cells. We evaluated several phenotypes: (i bacterial escape from macrophages after phagocytosis, (ii macrophage death induced by EIEC and S. flexneri, (iii macrophage cytokine expression in response to infection and (iv expression of plasmidial (pINV virulence genes. The results showed thatS. flexneri caused macrophage killing earlier and more intensely than EIEC. Both pathogens induced significant macrophage production of TNF, IL-1 and IL-10 after 7 h of infection. Transcription levels of the gene invasion plasmid antigen-C were lower in EIEC than in S. flexneri throughout the course of the infection; this could explain the diminished virulence of EIEC compared to S. flexneri.

  19. Simultaneous RNA-seq based transcriptional profiling of intracellular Brucella abortus and B. abortus-infected murine macrophages.

    Science.gov (United States)

    Hop, Huynh Tan; Arayan, Lauren Togonon; Reyes, Alisha Wehdnesday Bernardo; Huy, Tran Xuan Ngoc; Min, WonGi; Lee, Hu Jang; Son, Jee Soo; Kim, Suk

    2017-12-01

    Brucella is a zoonotic pathogen that survives within macrophages; however the replicative mechanisms involved are not fully understood. We describe the isolation of sufficient Brucella abortus RNA from primary host cell environment using modified reported methods for RNA-seq analysis, and simultaneously characterize the transcriptional profiles of intracellular B. abortus and bone marrow-derived macrophages (BMM) from BALB/c mice at 24 h (replicative phase) post-infection. Our results revealed that 25.12% (801/3190) and 16.16% (515/3190) of the total B. abortus genes were up-regulated and down-regulated at >2-fold, respectively as compared to the free-living B. abortus. Among >5-fold differentially expressed genes, the up-regulated genes are mostly involved in DNA, RNA manipulations as well as protein biosynthesis and secretion while the down-regulated genes are mainly involved in energy production and metabolism. On the other hand, the host responses during B. abortus infection revealed that 14.01% (6071/43,346) of BMM genes were reproducibly transcribed at >5-fold during infection. Transcription of cytokines, chemokines and transcriptional factors, such as tumor necrosis factor (Tnf), interleukin-1α (Il1α), interleukin-1β (Il1β), interleukin-6 (Il6), interleukin-12 (Il12), chemokine C-X-C motif (CXCL) family, nuclear factor kappa B (Nf-κb), signal transducer and activator of transcription 1 (Stat1), that may contribute to host defense were markedly induced while transcription of various genes involved in cell proliferation and metabolism were suppressed upon B. abortus infection. In conclusion, these data suggest that Brucella modulates gene expression in hostile intracellular environment while simultaneously alters the host pathways that may lead to the pathogen's intracellular survival and infection. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

    Science.gov (United States)

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

    2014-08-09

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

  1. Protein Phosphatase, Mg2+/Mn2+-dependent 1A controls the innate antiviral and antibacterial response of macrophages during HIV-1 and Mycobacterium tuberculosis infection

    Science.gov (United States)

    Sun, Jim; Schaaf, Kaitlyn; Duverger, Alexandra; Wolschendorf, Frank; Speer, Alexander; Wagner, Frederic; Niederweis, Michael; Kutsch, Olaf

    2016-01-01

    Co-infection with HIV-1 and Mycobacterium tuberculosis (Mtb) is a major public health issue. While some research has described how each pathogen accelerates the course of infection of the other pathogen by compromising the immune system, very little is known about the molecular biology of HIV-1/Mtb co-infection at the host cell level. This is somewhat surprising, as both pathogens are known to replicate and persist in macrophages. We here identify Protein Phosphatase, Mg2+/Mn2+-dependent 1A (PPM1A) as a molecular link between Mtb infection and increased HIV-1 susceptibility of macrophages. We demonstrate that both Mtb and HIV-1 infection induce the expression of PPM1A in primary human monocyte/macrophages and THP-1 cells. Genetic manipulation studies revealed that increased PPMA1 expression rendered THP-1 cells highly susceptible to HIV-1 infection, while depletion of PPM1A rendered them relatively resistant to HIV-1 infection. At the same time, increased PPM1A expression abrogated the ability of THP-1 cells to respond to relevant bacterial stimuli with a proper cytokine/chemokine secretion response, blocked their chemotactic response and impaired their ability to phagocytose bacteria. These data suggest that PPM1A, which had previously been shown to play a role in the antiviral response to Herpes Simplex virus infection, also governs the antibacterial response of macrophages to bacteria, or at least to Mtb infection. PPM1A thus seems to play a central role in the innate immune response of macrophages, implying that host directed therapies targeting PPM1A could be highly beneficial, in particular for HIV/Mtb co-infected patients. PMID:27004401

  2. 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)

  3. Alendronate augments interleukin-1β release from macrophages infected with periodontal pathogenic bacteria through activation of caspase-1

    International Nuclear Information System (INIS)

    Deng Xue; Tamai, Riyoko; Endo, Yasuo; Kiyoura, Yusuke

    2009-01-01

    Nitrogen-containing bisphosphonates (NBPs) are anti-bone-resorptive drugs with inflammatory side effects that include osteomyelitis and osteonecrosis of the jaw. Oral bacteria have been considered to be a trigger for these NBP-associated jaw bone diseases. The present study examined the effects of alendronate (a typical NBP) and clodronate (a non-NBP) on the production of proinflammatory cytokines by macrophages infected with Porphyromonas gingivalis and Tannerella forsythia, which are important pathogens of periodontal diseases. Pretreatment with alendronate augmented IL-1β, but not TNFα, production by macrophages infected with P. gingivalis or T. forsythia. This augmentation of IL-1β production was inhibited by clodronate. Furthermore, caspase-1, a promoter of IL-1β production, was activated by treatment with alendronate, and caspase-1 inhibitor reduced the production of IL-1β induced by alendronate and P. gingivalis. These results suggest that NBPs augment periodontal pathogenic bacteria-induced IL-1β release via caspase-1 activation, and this phenomenon may contribute to the development of NBP-associated inflammatory side effects including jaw osteomyelitis. Co-treatment with clodronate may prevent and/or reduce these inflammatory effects induced by NBPs

  4. Prevention of pathogenic Escherichia coli infection in mice and stimulation of macrophage activation in rats by an oral administration of probiotic Lactobacillus casei I-5.

    Science.gov (United States)

    Ishida-Fujii, Keiko; Sato, Rieko; Goto, Shingo; Yang, Xiao-Ping; Kuboki, Hiroshi; Hirano, Shin-Ichi; Sato, Michikatsu

    2007-04-01

    Lactobacillus casei I-5 isolated from an alcohol fermentation broth enhanced immunity and prevented pathogenic infection as a probiotic. Mice fed with I-5 cells for 11 days prior to an intraperitoneal challenge with pathogenic Escherichia coli Juhl exhibited a high survival rate compared with the control group. Rats fed with I-5 cells for 10 days significantly increased the phagocytosis of peritoneal macrophages. In a cell culture system employing peritoneal macrophages from rats, the I-5 administration activated NF-kappaB stimulated by LPS. It also enhanced LPS-stimulated IL-12 and TNF-alpha production, but not IL-6 production. These results show that L. casei I-5 effectively prevented infection by pathogenic E. coli possibly through the activation of peritoneal macrophages. The strain would be useful to prevent pathogenic microbial infections in humans and farm animals.

  5. DMPD: Innate immune responses during infection. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15576198 Innate immune responses during infection. Ulevitch RJ, Mathison JC, da Sil...ses during infection. PubmedID 15576198 Title Innate immune responses during infection. Authors Ulevitch RJ, Math

  6. Antibodies against glycolipids enhance antifungal activity of macrophages and reduce fungal burden after infection with Paracoccidioides brasiliensis

    Directory of Open Access Journals (Sweden)

    Renata Amelia eBueno

    2016-02-01

    Full Text Available Paracoccidioidomycosis is a fungal disease endemic in Latin America. Polyclonal antibodies to acidic glycosphingolipids (GSLs from Paracoccidioides brasiliensis opsonized yeast forms in vitro increasing phagocytosis and reduced the fungal burden of infected animals. Antibodies to GSL were active in both prophylactic and therapeutic protocols using a murine intratracheal infection model. Pathological examination of the lungs of animals treated with antibodies to GSL showed well-organized granulomas and minimally damaged parenchyma compared to the untreated control. Murine peritoneal macrophages activated by IFN-γ and incubated with antibodies against acidic GSLs more effectively phagocytosed and killed P. brasiliensis yeast cells as well as produced more nitric oxide compared to controls. The present work discloses a novel target of protective antibodies against P. brasiliensis adding to other well-studied mediators of the immune response to this fungus.

  7. Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis

    Directory of Open Access Journals (Sweden)

    Michell Stephen L

    2011-01-01

    Full Text Available Abstract Background Burkholderia pseudomallei is the causative agent of melioidosis, a tropical disease of humans with a variable and often fatal outcome. In murine models of infection, different strains exhibit varying degrees of virulence. In contrast, two related species, B. thailandensis and B. oklahomensis, are highly attenuated in mice. Our aim was to determine whether virulence in mice is reflected in macrophage or wax moth larvae (Galleria mellonella infection models. Results B. pseudomallei strains 576 and K96243, which have low median lethal dose (MLD values in mice, were able to replicate and induce cellular damage in macrophages and caused rapid death of G. mellonella. In contrast, B. pseudomallei strain 708a, which is attenuated in mice, showed reduced replication in macrophages, negligible cellular damage and was avirulent in G. mellonella larvae. B. thailandensis isolates were less virulent than B. pseudomallei in all of the models tested. However, we did record strain dependent differences. B. oklahomensis isolates were the least virulent isolates. They showed minimal ability to replicate in macrophages, were unable to evoke actin-based motility or to form multinucleated giant cells and were markedly attenuated in G. mellonella compared to B. thailandensis. Conclusions We have shown that the alternative infection models tested here, namely macrophages and Galleria mellonella, are able to distinguish between strains of B. pseudomallei, B. thailandensis and B. oklahomensis and that these differences reflect the observed virulence in murine infection models. Our results indicate that B. oklahomensis is the least pathogenic of the species investigated. They also show a correlation between isolates of B. thailandensis associated with human infection and virulence in macrophage and Galleria infection models.

  8. Differential MicroRNA Expression in Human Macrophages with Mycobacterium tuberculosis Infection of Beijing/W and Non-Beijing/W Strain Types.

    Directory of Open Access Journals (Sweden)

    Lin Zheng

    Full Text Available The role of microRNAs in association with Mycobacterium tuberculosis (MTB infection and the immunology regulated by microRNAs upon MTB infection have not been fully unravelled. We examined the microRNA profiles of THP-1 macrophages upon the MTB infection of Beijing/W and non-Beijing/W clinical strains. We also studied the microRNA profiles of the host macrophages by microarray in a small cohort with active MTB disease, latent infection (LTBI, and from healthy controls.The results revealed that 14 microRNAs differentiated infections of Beijing/W from non-Beijing/W strains (P<0.05. A unique signature of 11 microRNAs in human macrophages was identified to differentiate active MTB disease from LTBI and healthy controls. Pathway analyses of these differentially expressed miRNAs suggest that the immune-regulatory interactions involving TGF-β signalling pathway take part in the dysregulation of critical TB processes in the macrophages, resulting in active expression of both cell communication and signalling transduction systems.We showed for the first time that the Beijing/W TB strains repressed a number of miRNAs expressions which may reflect their virulence characteristics in altering the host response. The unique signatures of 11 microRNAs may deserve further evaluation as candidates for biomarkers in the diagnosis of MTB and Beijing/W infections.

  9. Differential MicroRNA Expression in Human Macrophages with Mycobacterium tuberculosis Infection of Beijing/W and Non-Beijing/W Strain Types.

    Science.gov (United States)

    Zheng, Lin; Leung, Eric; Lee, Nelson; Lui, Grace; To, Ka-Fai; Chan, Raphael C Y; Ip, Margaret

    2015-01-01

    The role of microRNAs in association with Mycobacterium tuberculosis (MTB) infection and the immunology regulated by microRNAs upon MTB infection have not been fully unravelled. We examined the microRNA profiles of THP-1 macrophages upon the MTB infection of Beijing/W and non-Beijing/W clinical strains. We also studied the microRNA profiles of the host macrophages by microarray in a small cohort with active MTB disease, latent infection (LTBI), and from healthy controls. The results revealed that 14 microRNAs differentiated infections of Beijing/W from non-Beijing/W strains (PmicroRNAs in human macrophages was identified to differentiate active MTB disease from LTBI and healthy controls. Pathway analyses of these differentially expressed miRNAs suggest that the immune-regulatory interactions involving TGF-β signalling pathway take part in the dysregulation of critical TB processes in the macrophages, resulting in active expression of both cell communication and signalling transduction systems. We showed for the first time that the Beijing/W TB strains repressed a number of miRNAs expressions which may reflect their virulence characteristics in altering the host response. The unique signatures of 11 microRNAs may deserve further evaluation as candidates for biomarkers in the diagnosis of MTB and Beijing/W infections.

  10. Effect of Macrophage Migration Inhibitory Factor (MIF) in Human Placental Explants Infected with Toxoplasma gondii Depends on Gestational Age

    Science.gov (United States)

    de Oliveira Gomes, Angelica; de Oliveira Silva, Deise Aparecida; Silva, Neide Maria; de Freitas Barbosa, Bellisa; Franco, Priscila Silva; Angeloni, Mariana Bodini; Fermino, Marise Lopes; Roque-Barreira, Maria Cristina; Bechi, Nicoletta; Paulesu, Luana Ricci; dos Santos, Maria Célia; Mineo, José Roberto; Ferro, Eloisa Amália Vieira

    2011-01-01

    Because macrophage migration inhibitory factor (MIF) is a key cytokine in pregnancy and has a role in inflammatory response and pathogen defense, the objective of the present study was to investigate the effects of MIF in first- and third-trimester human placental explants infected with Toxoplasma gondii. Explants were treated with recombinant MIF, IL-12, interferon-γ, transforming growth factor-β1, or IL-10, followed by infection with T. gondii RH strain tachyzoites. Supernatants of cultured explants were assessed for MIF production. Explants were processed for morphologic analysis, immunohistochemistry, and real-time PCR analysis. Comparison of infected and stimulated explants versus noninfected control explants demonstrated a significant increase in MIF release in first-trimester but not third-trimester explants. Tissue parasitism was higher in third- than in first-trimester explants. Moreover, T. gondii DNA content was lower in first-trimester explants treated with MIF compared with untreated explants. However, in third-trimester explants, MIF stimulus decreased T. gondii DNA content only at the highest concentration of the cytokine. In addition, high expression of MIF receptor was observed in first-trimester placental explants, whereas MIF receptor expression was low in third-trimester explants. In conclusion, MIF was up-regulated and demonstrated to be important for control of T. gondii infection in first-trimester explants, whereas lack of MIF up-regulation in third-trimester placentas may be involved in higher susceptibility to infection at this gestational age. PMID:21641401

  11. DMPD: Role of Nods in bacterial infection. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17379560 Role of Nods in bacterial infection. Bourhis LL, Werts C. Microbes Infect.... 2007 Apr;9(5):629-36. Epub 2007 Jan 27. (.png) (.svg) (.html) (.csml) Show Role of Nods in bacterial infection.... PubmedID 17379560 Title Role of Nods in bacterial infection. Authors Bourhis LL, Werts C. Publication M

  12. Role of macrophage inflammatory protein-1alpha in T-cell-mediated immunity to viral infection

    DEFF Research Database (Denmark)

    Madsen, Andreas N; Nansen, Anneline; Christensen, Jan P

    2003-01-01

    The immune response to lymphocytic choriomeningitis virus in mice lacking macrophage inflammatory protein-1alpha (MIP-1alpha) was evaluated. Generation of virus-specific effector T cells is unimpaired in MIP-1alpha-deficient mice. Furthermore, MIP-1alpha is not required for T-cell-mediated virus...... control or virus-induced T-cell-dependent inflammation. Thus, MIP-1alpha is not mandatory for T-cell-mediated antiviral immunity....

  13. HIV-1 infection induces changes in expression of cellular splicing factors that regulate alternative viral splicing and virus production in macrophages

    Directory of Open Access Journals (Sweden)

    Purcell Damian FJ

    2008-02-01

    Full Text Available Abstract Background Macrophages are important targets and long-lived reservoirs of HIV-1, which are not cleared of infection by currently available treatments. In the primary monocyte-derived macrophage model of infection, replication is initially productive followed by a decline in virion output over ensuing weeks, coincident with a decrease in the levels of the essential viral transactivator protein Tat. We investigated two possible mechanisms in macrophages for regulation of viral replication, which appears to be primarily regulated at the level of tat mRNA: 1 differential mRNA stability, used by cells and some viruses for the rapid regulation of gene expression and 2 control of HIV-1 alternative splicing, which is essential for optimal viral replication. Results Following termination of transcription at increasing times after infection in macrophages, we found that tat mRNA did indeed decay more rapidly than rev or nef mRNA, but with similar kinetics throughout infection. In addition, tat mRNA decayed at least as rapidly in peripheral blood lymphocytes. Expression of cellular splicing factors in uninfected and infected macrophage cultures from the same donor showed an inverse pattern over time between enhancing factors (members of the SR family of RNA binding proteins and inhibitory factors (members of the hnRNP family. While levels of the SR protein SC35 were greatly up-regulated in the first week or two after infection, hnRNPs of the A/B and H groups were down-regulated. Around the peak of virus production in each culture, SC35 expression declined to levels in uninfected cells or lower, while the hnRNPs increased to control levels or above. We also found evidence for increased cytoplasmic expression of SC35 following long-term infection. Conclusion While no evidence of differential regulation of tat mRNA decay was found in macrophages following HIV-1 infection, changes in the balance of cellular splicing factors which regulate alternative

  14. Increased survival and proliferation of the epidemic strain Mycobacterium abscessus subsp. massiliense CRM0019 in alveolar epithelial cells.

    Science.gov (United States)

    Ribeiro, Giovanni Monteiro; Matsumoto, Cristianne Kayoko; Real, Fernando; Teixeira, Daniela; Duarte, Rafael Silva; Mortara, Renato Arruda; Leão, Sylvia Cardoso; de Souza Carvalho-Wodarz, Cristiane

    2017-09-13

    Outbreaks of infections caused by rapidly growing mycobacteria have been reported worldwide generally associated with medical procedures. Mycobacterium abscessus subsp. massiliense CRM0019 was obtained during an epidemic of postsurgical infections and was characterized by increased persistence in vivo. To better understand the successful survival strategies of this microorganism, we evaluated its infectivity and proliferation in macrophages (RAW and BMDM) and alveolar epithelial cells (A549). For that, we assessed the following parameters, for both M. abscessus CRM0019 as well as the reference strain M. abscessus ATCC 19977: internalization, intracellular survival for up 3 days, competence to subvert lysosome fusion and the intracellular survival after cell reinfection. CRM0019 and ATCC 19977 strains showed the same internalization rate (approximately 30% after 6 h infection), in both A549 and RAW cells. However, colony forming units data showed that CRM0019 survived better in A549 cells than the ATCC 19977 strain. Phagosomal characteristics of CRM0019 showed the bacteria inside tight phagosomes in A549 cells, contrasting to the loosely phagosomal membrane in macrophages. This observation holds for the ATCC 19977 strain in both cell types. The competence to subvert lysosome fusion was assessed by acidification and acquisition of lysosomal protein. For M. abscessus strains the phagosomes were acidified in all cell lines; nevertheless, the acquisition of lysosomal protein was reduced by CRM0019 compared to the ATCC 19977 strain, in A549 cells. Conversely, in macrophages, both M. abscessus strains were located in mature phagosomes, however without bacterial death. Once recovered from macrophages M. abscessus could establish a new intracellular infection. Nevertheless, only CRM0019 showed a higher growth rate in A549, increasing nearly 10-fold after 48 and 72 h. M. abscessus CRM0019 creates a protective and replicative niche in alveolar epithelial cells mainly by

  15. Rare Lung Diseases II: Pulmonary Alveolar Proteinosis

    Directory of Open Access Journals (Sweden)

    Stephen C Juvet

    2008-01-01

    Full Text Available The present article is the second in a series on rare lung diseases. It focuses on pulmonary alveolar proteinosis (PAP, a disorder in which lipoproteinaceous material accumulates in the alveolar space. PAP was first described in 1958, and for many years the nature of the material accumulating in the lungs was unknown. Major insights into PAP have been made in the past decade, and these have led to the notion that PAP is an autoimmume disorder in which autoantibodies interfere with signalling through the granulocyte-macrophage colony-stimulating factor receptor, leading to macrophage and neutrophil dysfunction. This has spurred new therapeutic approaches to this disorder. The discussion of PAP will begin with a case report, then will highlight the classification of PAP and review recent insights into the pathogenesis of PAP. The approach to therapy and the prognosis of PAP will also be discussed.

  16. HIV aspartyl peptidase inhibitors interfere with cellular proliferation, ultrastructure and macrophage infection of Leishmania amazonensis.

    Directory of Open Access Journals (Sweden)

    Lívia O Santos

    Full Text Available BACKGROUND: Leishmania is the etiologic agent of leishmanisais, a protozoan disease whose pathogenic events are not well understood. Current therapy is suboptimal due to toxicity of the available therapeutic agents and the emergence of drug resistance. Compounding these problems is the increase in the number of cases of Leishmania-HIV coinfection, due to the overlap between the AIDS epidemic and leishmaniasis. METHODOLOGY/PRINCIPAL FINDINGS: In the present report, we have investigated the effect of HIV aspartyl peptidase inhibitors (PIs on the Leishmania amazonensis proliferation, ultrastructure, interaction with macrophage cells and expression of classical peptidases which are directly involved in the Leishmania pathogenesis. All the HIV PIs impaired parasite growth in a dose-dependent fashion, especially nelfinavir and lopinavir. HIV PIs treatment caused profound changes in the leishmania ultrastructure as shown by transmission electron microscopy, including cytoplasm shrinking, increase in the number of lipid inclusions and some cells presenting the nucleus closely wrapped by endoplasmic reticulum resembling an autophagic process, as well as chromatin condensation which is suggestive of apoptotic death. The hydrolysis of HIV peptidase substrate by L. amazonensis extract was inhibited by pepstatin and HIV PIs, suggesting that an aspartyl peptidase may be the intracellular target of the inhibitors. The treatment with HIV PIs of either the promastigote forms preceding the interaction with macrophage cells or the amastigote forms inside macrophages drastically reduced the association indexes. Despite all these beneficial effects, the HIV PIs induced an increase in the expression of cysteine peptidase b (cpb and the metallopeptidase gp63, two well-known virulence factors expressed by Leishmania spp. CONCLUSIONS/SIGNIFICANCE: In the face of leishmaniasis/HIV overlap, it is critical to further comprehend the sophisticated interplays among Leishmania

  17. Toll-like receptor 4 is involved in the cell cycle modulation and required for effective human cytomegalovirus infection in THP-1 macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Arcangeletti, Maria-Cristina, E-mail: mariacristina.arcangeletti@unipr.it [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy); Germini, Diego; Rodighiero, Isabella [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy); Mirandola, Prisco [Department of Biomedical, Biotechnological and Translational Sciences, University of Parma, Parma (Italy); De Conto, Flora; Medici, Maria-Cristina [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy); Gatti, Rita [Department of Biomedical, Biotechnological and Translational Sciences, University of Parma, Parma (Italy); Chezzi, Carlo; Calderaro, Adriana [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy)

    2013-05-25

    Suitable host cell metabolic conditions are fundamental for the effective development of the human cytomegalovirus (HCMV) lytic cycle. Indeed, several studies have demonstrated the ability of this virus to interfere with cell cycle regulation, mainly by blocking proliferating cells in G1 or G1/S. In the present study, we demonstrate that HCMV deregulates the cell cycle of THP-1 macrophages (a cell line irreversibly arrested in G0) by pushing them into S and G2 phases. Moreover, we show that HCMV infection of THP-1 macrophages leads to Toll-like receptor 4 (TLR4) activation. Since various studies have indicated TLR4 to be involved in promoting cell proliferation, here we investigate the possible role of TLR4 in the observed HCMV-induced cell cycle perturbation. Our data strongly support TLR4 as a mediator of HCMV-triggered cell cycle activation in THP-1 macrophages favouring, in turn, the development of an efficient viral lytic cycle. - Highlights: ► We studied HCMV infection impact on THP-1 macrophage cell cycle. ► We analysed the role played by Toll-like receptor (TLR) 4 upon HCMV infection. ► HCMV pushes THP-1 macrophages (i.e. resting cells) to re-enter the cell cycle. ► TLR4 pathway inhibition strongly affects the effectiveness of HCMV replication. ► TLR4 pathway inhibition significantly decreases HCMV-induced cell cycle re-entry.

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

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Zhao-Hua [Division of Monoclonal Antibodies, Center for Drug Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States); Kumari, Namita; Nekhai, Sergei [Center for Sickle Cell Disease, Department of Medicine, Howard University, Washington, DC (United States); Clouse, Kathleen A. [Division of Monoclonal Antibodies, Center for Drug Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States); Wahl, Larry M. [National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD (United States); Yamada, Kenneth M. [Laboratory of Cell and Development Biology, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD (United States); Dhawan, Subhash, E-mail: subhash.dhawan@fda.hhs.gov [Viral Immunology Section, Laboratory of Molecular Virology, Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States)

    2013-06-07

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

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

    International Nuclear Information System (INIS)

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

    2013-01-01

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

  20. Macrophage migration inhibitory factor regulates interleukin-6 production by facilitating nuclear factor-kappa B activation during Vibrio vulnificus infection

    Directory of Open Access Journals (Sweden)

    Choi Pui-Ching

    2010-10-01

    Full Text Available Abstract Background Patients infected with Vibrio vulnificus (V. vulnificus show severe inflammatory responses characterised by the upregulation of proinflammatory cytokines. Macrophage migration inhibitory factor (MIF, an upstream proinflammatory regulator, increases the inflammation caused by sepsis. Whether MIF regulates responses to V. vulnificus infection and the actual mechanism by which V. vulnificus initiates these MIF-modulated proinflammatory cytokines remain unclear. Results MIF increased inflammation during V. vulnificus infection in vivo. In V. vulnificus-infected mice, MIF was produced earlier than tumour necrosis factor (TNF-α and interleukin (IL-6 and was expressed in a time-dependent manner. ISO-1 ((S, R-3-(4-hydroxyphenyl-4,5-dihydro-5-isoxazole acetic acid methyl ester, a small-molecule inhibitor of MIF, significantly decreased IL-6, IL-8, and TNF-α production in a time- and dose-dependent manner in human peripheral blood cells infected with V. vulnificus. The induction of IL-6, IL-8, and TNF-α production by V. vulnificus infection was mediated via the NF-κB- and p38 MAPK-regulated pathways but not via the Akt pathway. ISO-1-treated human peripheral blood cells showed lower V. vulnificus-induced NF-κB activation, IL-6 mRNA expression, and IκB phosphorylation, but they did not show lower p38 MAPK activation. Conclusions We conclude that MIF regulates V. vulnificus-induced IL-6 production via NF-κB activation and that p38 MAPK activation in V. vulnificus infection is not MIF dependent.

  1. Natural killer cell-intrinsic type I IFN signaling controls Klebsiella pneumoniae growth during lung infection

    Science.gov (United States)

    Borroni, Martina; Kavirayani, Anoop; Przybyszewska, Kornelia N.; Ingram, Rebecca J.; Lienenklaus, Stefan

    2017-01-01

    Klebsiella pneumoniae is a significant cause of nosocomial pneumonia and an alarming pathogen owing to the recent isolation of multidrug resistant strains. Understanding of immune responses orchestrating K. pneumoniae clearance by the host is of utmost importance. Here we show that type I interferon (IFN) signaling protects against lung infection with K. pneumoniae by launching bacterial growth-controlling interactions between alveolar macrophages and natural killer (NK) cells. Type I IFNs are important but disparate and incompletely understood regulators of defense against bacterial infections. Type I IFN receptor 1 (Ifnar1)-deficient mice infected with K. pneumoniae failed to activate NK cell-derived IFN-γ production. IFN-γ was required for bactericidal action and the production of the NK cell response-amplifying IL-12 and CXCL10 by alveolar macrophages. Bacterial clearance and NK cell IFN-γ were rescued in Ifnar1-deficient hosts by Ifnar1-proficient NK cells. Consistently, type I IFN signaling in myeloid cells including alveolar macrophages, monocytes and neutrophils was dispensable for host defense and IFN-γ activation. The failure of Ifnar1-deficient hosts to initiate a defense-promoting crosstalk between alveolar macrophages and NK cell was circumvented by administration of exogenous IFN-γ which restored endogenous IFN-γ production and restricted bacterial growth. These data identify NK cell-intrinsic type I IFN signaling as essential driver of K. pneumoniae clearance, and reveal specific targets for future therapeutic exploitations. PMID:29112952

  2. [Macrophages in asthma].

    Science.gov (United States)

    Medina Avalos, M A; Orea Solano, M

    1997-01-01

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

  3. 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

  4. DMPD: Innate immune response to viral infection. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18694646 Innate immune response to viral infection. Koyama S, Ishii KJ, Coban C, Ak...ira S. Cytokine. 2008 Sep;43(3):336-41. Epub 2008 Aug 9. (.png) (.svg) (.html) (.csml) Show Innate immune response to viral infection.... PubmedID 18694646 Title Innate immune response to viral infection. Authors Koyama

  5. DMPD: Toll-like receptors regulation of viral infection and disease. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18280610 Toll-like receptors regulation of viral infection and disease. Thompson JM...how Toll-like receptors regulation of viral infection and disease. PubmedID 18280610 Title Toll-like recepto...rs regulation of viral infection and disease. Authors Thompson JM, Iwasaki A. Pub

  6. DMPD: Is HIV infection a TNF receptor signalling-driven disease? [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18178131 Is HIV infection a TNF receptor signalling-driven disease? Herbein G, Khan... KA. Trends Immunol. 2008 Feb;29(2):61-7. (.png) (.svg) (.html) (.csml) Show Is HIV infection a TNF receptor signalling-driven dise...ase? PubmedID 18178131 Title Is HIV infection a TNF receptor signalling-driven diseas

  7. Lymphocytes and macrophages are infected by Theileria equi, but T cells and B cells are not required to establish infection in vivo.

    Directory of Open Access Journals (Sweden)

    Joshua D Ramsay

    Full Text Available Theileria equi has a biphasic life cycle in horses, with a period of intraleukocyte development followed by patent erythrocytic parasitemia that causes acute and sometimes fatal hemolytic disease. Unlike Theileria spp. that infect cattle (Theileria parva and Theileria annulata, the intraleukocyte stage (schizont of Theileria equi does not cause uncontrolled host cell proliferation or other significant pathology. Nevertheless, schizont-infected leukocytes are of interest because of their potential to alter host cell function and because immune responses directed against this stage could halt infection and prevent disease. Based on cellular morphology, Theileria equi has been reported to infect lymphocytes in vivo and in vitro, but the specific phenotype of schizont-infected cells has yet to be defined. To resolve this knowledge gap in Theileria equi pathogenesis, peripheral blood mononuclear cells were infected in vitro and the phenotype of infected cells determined using flow cytometry and immunofluorescence microscopy. These experiments demonstrated that the host cell range of Theileria equi was broader than initially reported and included B lymphocytes, T lymphocytes and monocyte/macrophages. To determine if B and T lymphocytes were required to establish infection in vivo, horses affected with severe combined immunodeficiency (SCID, which lack functional B and T lymphocytes, were inoculated with Theileria equi sporozoites. SCID horses developed patent erythrocytic parasitemia, indicating that B and T lymphocytes are not necessary to complete the Theileria equi life cycle in vivo. These findings suggest that the factors mediating Theileria equi leukocyte invasion and intracytoplasmic differentiation are common to several leukocyte subsets and are less restricted than for Theileria annulata and Theileria parva. These data will greatly facilitate future investigation into the relationships between Theileria equi leukocyte tropism and pathogenesis

  8. Effects of ischemia on lung macrophages.

    Directory of Open Access Journals (Sweden)

    Aigul Moldobaeva

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

  9. An Interferon-Related Signature in the Transcriptional Core Response of Human Macrophages to Mycobacterium tuberculosis Infection

    Science.gov (United States)

    Jin, Wen; Mei, Jian; Gicquel, Brigitte; Du, Yanzhi; Wang, Kankan; Gao, Qian; Neyrolles, Olivier; Zhang, Ji

    2012-01-01

    The W-Beijing family of Mycobacterium tuberculosis (Mtb) strains is known for its high-prevalence and -virulence, as well as for its genetic diversity, as recently reported by our laboratories and others. However, little is known about how the immune system responds to these strains. To explore this issue, here we used reverse engineering and genome-wide expression profiling of human macrophage-like THP-1 cells infected by different Mtb strains of the W-Beijing family, as well as by the reference laboratory strain H37Rv. Detailed data mining revealed that host cell transcriptome responses to H37Rv and to different strains of the W-Beijing family are similar and overwhelmingly induced during Mtb infections, collectively typifying a robust gene expression signature (“THP1r2Mtb-induced signature”). Analysis of the putative transcription factor binding sites in promoter regions of genes in this signature identified several key regulators, namely STATs, IRF-1, IRF-7, and Oct-1, commonly involved in interferon-related immune responses. The THP1r2Mtb-induced signature appeared to be highly relevant to the interferon-inducible signature recently reported in active pulmonary tuberculosis patients, as revealed by cross-signature and cross-module comparisons. Further analysis of the publicly available transcriptome data from human patients showed that the signature appears to be relevant to active pulmonary tuberculosis patients and their clinical therapy, and be tuberculosis specific. Thus, our results provide an additional layer of information at the transcriptome level on mechanisms involved in host macrophage response to Mtb, which may also implicate the robustness of the cellular defense system that can effectively fight against genetic heterogeneity in this pathogen. PMID:22675550

  10. Thalidomide treatment modulates macrophage pro-inflammatory function and cytokine levels in Klebsiella pneumoniae B5055 induced pneumonia in BALB/c mice.

    Science.gov (United States)

    Kumar, Vijay; Harjai, Kusum; Chhibber, Sanjay

    2010-07-01

    Lung innate immune response plays an important role in the clearance of pathogens from lungs, however, profound activation of innate immune cells (alveolar macrophages or neutrophils) can lead to development of acute lung inflammation or injury by producing various pro-inflammatory molecules (IL-1, TNF-alpha and H2O2 etc.). Present study is designed to investigate the immunomodulatory action of thalidomide in Klebsiella pneumoniae B5055 induced acute lung infection in BALB/c mice. Acute lung inflammation was induced by intranasal instillation of K. pneumoniae B5055 into mice without any anaesthesia and treated with thalidomide (30 mg/kg/day/po) or normal saline orally using a treatment schedule shown to modulate pro-inflammatory innate immune response. Thalidomide treatment modulated pro-inflammatory function of alveolar macrophages by significantly (ppneumonia caused by gram negative bacterial infection. 2010 Elsevier B.V. All rights reserved.

  11. Effects of interferon gamma and specific polyclonal antibody on the infection of murine peritoneal macrophages and murine macrophage cell line PMJ2-R with Encephalitozoon cuniculi

    Czech Academy of Sciences Publication Activity Database

    Jelínek, Jiří; Salát, Jiří; Sak, Bohumil; Kopecký, Jan

    2007-01-01

    Roč. 54, č. 3 (2007), s. 172-176 ISSN 0015-5683 R&D Projects: GA ČR GP524/03/D167 Institutional research plan: CEZ:AV0Z60220518 Keywords : microsporidia * Encephalitozoon cuniculi * antibody * macrophage s * interferon gamma (IFN-gamma) Subject RIV: EC - Immunology Impact factor: 1.000, year: 2007

  12. A Novel Strategy for TNF-Alpha Production by 2-APB Induced Downregulated SOCE and Upregulated HSP70 in O. tsutsugamushi-Infected Human Macrophages.

    Directory of Open Access Journals (Sweden)

    Ching-Ying Wu

    Full Text Available Orientia (O. tsutsugamushi-induced scrub typhus is endemic across many regions of Asia and the Western Pacific, where an estimated 1 million cases occur each year; the majority of patients infected with O. tsutsugamushi end up with a cytokine storm from a severe inflammatory response. Previous reports have indicated that blocking tumor necrosis factor (TNF-α reduced cell injury from a cytokine storm. Since TNF-α production is known to be associated with intracellular Ca2+ elevation, we examined the effect of store-operated Ca2+ entry (SOCE inhibitors on TNF-α production in O. tsutsugamushi-infected macrophages. We found that 2-aminoethoxydiphenyl borate (2-APB, but not SKF96365, facilitates the suppression of Ca2+ mobilization via the interruption of Orai1 expression in O. tsutsugamushi-infected macrophages. Due to the decrease of Ca2+ elevation, the expression of TNF-α and its release from macrophages was repressed by 2-APB. In addition, a novel role of 2-APB was found in macrophages that causes the upregulation of heat shock protein 70 (HSP70 expression associated with ERK activation; upregulated TNF-α production in the case of knockdown HSP70 was inhibited with 2-APB treatment. Furthermore, elevated HSP70 formation unexpectedly did not help the cell survival of O. tsutsugamushi-infected macrophages. In conclusion, the parallelism between downregulated Ca2+ mobilization via SOCE and upregulated HSP70 after treatment with 2-APB against TNF-α production was found to efficiently attenuate an O. tsutsugamushi-induced severe inflammatory response.

  13. Thymus vulgaris L. and thymol assist murine macrophages (RAW 264.7) in the control of in vitro infections by Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans.

    Science.gov (United States)

    de Oliveira, Jonatas Rafael; Figueira, Leandro Wagner; Sper, Fábia Lugli; Meccatti, Vanessa Marques; Camargo, Samira Esteves Afonso; de Oliveira, Luciane Dias

    2017-08-01

    Microorganisms are capable to combat defense cells by means of strategies that contribute to their stabilization and proliferation in invaded tissues. Frequently antimicrobial-resistant strains appear; therefore, alternative methods to control them must be investigated, for example, the use of plant products. The capacity of the thyme extract (Thymus vulgaris L.) and phytocompound thymol in the control of in vitro infections by Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans in murine macrophages (RAW 264.7) was evaluated. Minimal inhibitory concentrations (MIC) of the plant products were used. The effect of these MIC were analyzed in the assays of phagocytosis and immunoregulation by analysis of the production of cytokines (IL-1β, TNF-α, and IL-10) and nitric oxide (NO). The plant products effectively assisted the macrophages in the phagocytosis of microorganisms, presenting significant reductions of S. aureus and P. aeruginosa. The macrophages also regulated the production of inflammatory mediators in the infections by S. aureus, P. aeruginosa, and C. albicans. In addition, thyme provided a satisfactory effect in response to the bacterial infections, regarding generation of NO. Thus, the effectiveness of the thyme and thymol to control in vitro infections by S. aureus, P. aeruginosa, and C. albicans was observed. Phagocytosis of S. aureus by RAW 264.7 was enhanced with thymol Thyme enhanced the phagocytosis of P. aeruginosa by RAW 264.7 Plant products provided immunoregulation of inflammatory cytokines Production of nitric oxide was improved with the treatments in bacterial infections.

  14. Cholesterol Corrects Altered Conformation of MHC-II Protein in Leishmania donovani Infected Macrophages: Implication in Therapy

    Science.gov (United States)

    Chakrabarti, Saikat; Roy, Syamal

    2016-01-01

    Background Previously we reported that Kala-azar patients show progressive decrease in serum cholesterol as a function of splenic parasite burden. Splenic macrophages (MΦ) of Leishmania donovani (LD) infected mice show decrease in membrane cholesterol, while LD infected macrophages (I-MΦ) show defective T cell stimulating ability that could be corrected by liposomal delivery of cholesterol. T helper cells recognize peptide antigen in the context of class II MHC molecule. It is known that the conformation of a large number of membrane proteins is dependent on membrane cholesterol. In this investigation we tried to understand the influence of decreased membrane cholesterol in I-MΦ on the conformation of MHC-II protein and peptide-MHC-II stability, and its bearing on the antigen specific T-cell activation. Methodology/Principal Findings MΦ of CBA/j mice were infected with Leishmania donovani (I-MΦ). Two different anti-Aκ mAbs were used to monitor the status of MHC-II protein under parasitized condition. One of them (11.5–2) was conformation specific, whereas the other one (10.2.16) was not. Under parasitized condition, the binding of 11.5–2 decreased significantly with respect to the normal counterpart, whereas that of 10.2.16 remained unaltered. The binding of 11.5–2 was restored to normal upon liposomal delivery of cholesterol in I-MΦ. By molecular dynamics (MD) simulation studies we found that there was considerable conformational fluctuation in the transmembrane domain of the MHC-II protein in the presence of membrane cholesterol than in its absence, which possibly influenced the distal peptide binding groove. This was evident from the faster dissociation of the cognate peptide from peptide-MHC complex under parasitized condition, which could be corrected by liposomal delivery of cholesterol in I-MΦ. Conclusion The decrease in membrane cholesterol in I-MΦ may lead to altered conformation of MHC II, and this may contribute to a faster dissociation of

  15. Primary alveolar echinococcosis: course of larval development and antibody responses in intermediate host rodents with different genetic backgrounds after oral infection with eggs of Echinococcus multilocularis.

    Science.gov (United States)

    Matsumoto, Jun; Kouguchi, Hirokazu; Oku, Yuzaburo; Yagi, Kinpei

    2010-09-01

    We investigated parasite establishment, subsequent larval development and antibody responses in gerbils, cotton rats and 4 inbred mouse strains until 16 weeks post inoculation (p.i.) with 200 eggs of Echinococcus multilocularis. The rate of parasite establishment in the liver determined at 4 weeks p.i. was highest in DBA/2, followed by AKR/N, C57BL/10 and C57BL/6 mice, whereas gerbils harboured few parasite foci. The accurate number of liver lesions in cotton rats could not be determined due to rapid growth and advanced multivesiculation of the parasite observed at 2 weeks p.i. The course of larval development was most advanced in DBA/2 mice with mature protoscolex formation at 16 weeks p.i., followed by AKR/N harbouring metacestodes with sparsely distributed immature protoscoleces. On the other hand, C57BL/6 and C57BL/10 mice had infertile metacestodes without any protoscolex formation. The parasite growth in mice was totally slower than those in gerbils and cotton rats. Specific IgG and IgM responses against 3 types of native crude antigens of larval E. multilocularis were evaluated using somatic extracts of and vesicle fluid of metacestode, and somatic extracts from purified protoscoleces. The 4 mouse strains demonstrated basically similar kinetics with apparent IgG and IgM increases at 9 weeks p.i. and thereafter, except C57BL/10, exhibited higher levels of IgM against crude antigens at some time point of infection. On the other hand, a follow-up determination of specific IgG and IgM levels against recombinant antigens from larval E. multilocularis revealed that each mouse strain showed different antibody-level kinetics. The findings in the present study demonstrate that the course of host-parasite interactions in primary alveolar echinococcosis, caused by larval E. multilocularis, clearly varies among intermediate host rodents with different genetic backgrounds.

  16. MicroRNA induction in human macrophages associated with infection with ancient and modern TB strains

    Directory of Open Access Journals (Sweden)

    L Furci

    2015-01-01

    Conclusion: In this study it was observed that the genetic diversity among Mtb strains and, in particular between ancient and modern strains, reflects on several aspects of host-pathogen interaction. In particular, the modulation of specific cellular microRNAs upon MTBC infection suggests a potential role for these microRNAs in the outcome of infection and, to a major extent, to the different epidemiological success of Mtb strains.

  17. Genetic-and-epigenetic Interspecies Networks for Cross-talk Mechanisms in Human Macrophages and Dendritic Cells During MTB Infection

    Directory of Open Access Journals (Sweden)

    Cheng-Wei Li

    2016-10-01

    Full Text Available Tuberculosis is caused by Mycobacterium tuberculosis (Mtb infection. Mtb is one of the oldest human pathogens, and evolves mechanisms implied in human evolution. The lungs are the first organ exposed to aerosol-transmitted Mtb during gaseous exchange. Therefore, the guards of the immune system in the lungs, such as macrophages (Mϕs and dendritic cells (DCs, are the most important defense against Mtb infection. There have been several studies discussing the functions of Mϕs and DCs during Mtb infection, but the genome-wide pathways and networks are still incomplete. Furthermore, the immune response induced by Mϕs and DCs varies. Therefore, we analyzed the cross-talk genome-wide genetic-and-epigenetic interspecies networks (GWGEINs between Mϕs vs. Mtb and DCs vs. Mtb to determine the varying mechanisms of both the host and pathogen as it relates to Mϕs and DCs during early Mtb infection.First, we performed database mining to construct candidate cross-talk GWGEIN between human cells and Mtb. Then we constructed dynamic models to characterize the molecular mechanisms, including intraspecies gene/microRNA (miRNA regulation networks (GRNs, intraspecies protein-protein interaction networks (PPINs, and the interspecies PPIN of the cross-talk GWGEIN. We applied a system identification method and a system order detection scheme to dynamic models to identify the real cross-talk GWGEINs using the microarray data of Mϕs, DCs and Mtb.After identifying the real cross-talk GWGEINs, the principal network projection (PNP method was employed to construct host-pathogen core networks (HPCNs between Mϕs vs. Mtb and DCs vs. Mtb during infection process. Thus, we investigated the underlying cross-talk mechanisms between the host and the pathogen to determine how the pathogen counteracts host defense mechanisms in Mϕs and DCs during Mtb H37Rv early infection. Based on our findings, we propose Rv1675c as a potential drug target because of its important defensive

  18. CD36 Provides Host Protection Against Klebsiella pneumoniae Intrapulmonary Infection by Enhancing Lipopolysaccharide Responsiveness and Macrophage Phagocytosis.

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    Olonisakin, Tolani F; Li, Huihua; Xiong, Zeyu; Kochman, Elizabeth J K; Yu, Minting; Qu, Yanyan; Hulver, Mei; Kolls, Jay K; St Croix, Claudette; Doi, Yohei; Nguyen, Minh-Hong; Shanks, Robert M Q; Mallampalli, Rama K; Kagan, Valerian E; Ray, Anuradha; Silverstein, Roy L; Ray, Prabir; Lee, Janet S

    2016-12-15

    Klebsiella pneumoniae remains an important cause of intrapulmonary infection and invasive disease worldwide. K. pneumoniae can evade serum killing and phagocytosis primarily through the expression of a polysaccharide capsule, but its pathogenicity is also influenced by host factors. We examined whether CD36, a scavenger receptor that recognizes pathogen and modified self ligands, is a host determinant of K. pneumoniae pathogenicity. Despite differences in serum sensitivity and virulence of 3 distinct K. pneumoniae (hypermucoviscous K1, research K2, and carbapenemase-producing ST258) strains, the absence of CD36 significantly increased host susceptibility to acute intrapulmonary infection by K. pneumoniae, regardless of strain. We demonstrate that CD36 enhances LPS responsiveness to K. pneumoniae to increase downstream cytokine production and macrophage phagocytosis that is independent of polysaccharide capsular antigen. Our study provides new insights into host determinants of K. pneumoniae pathogenicity and raises the possibility that functional mutations in CD36 may predispose individuals to K. pneumoniae syndromes. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  19. Pan-genomic analysis of bovine monocyte-derived macrophage gene expression in response to in vitro infection with Mycobacterium avium subspecies paratuberculosis

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    MacHugh David E

    2012-03-01

    Full Text Available Abstract Mycobacterium avium subspecies paratuberculosis is the causative agent of Johne’s disease, an intestinal disease of ruminants with major economic consequences. Infectious bacilli are phagocytosed by host macrophages upon exposure where they persist, resulting in lengthy subclinical phases of infection that can lead to immunopathology and disease dissemination. Consequently, analysis of the macrophage transcriptome in response to M. avium subsp. paratuberculosis infection can provide valuable insights into the molecular mechanisms that underlie Johne’s disease. Here, we investigate pan-genomic gene expression in bovine monocyte-derived macrophages (MDM purified from seven age-matched females, in response to in vitro infection with M. avium subsp. paratuberculosis (multiplicity of infection 2:1 at intervals of 2 hours, 6 hours and 24 hours post-infection (hpi. Differentially expressed genes were identified by comparing the transcriptomes of the infected MDM to the non-infected control MDM at each time point (adjusted P-value threshold ≤ 0.10. 1050 differentially expressed unique genes were identified 2 hpi, with 974 and 78 differentially expressed unique genes detected 6 and 24 hpi, respectively. Furthermore, in the infected MDM the number of upregulated genes exceeded the number of downregulated genes at each time point, with the fold-change in expression for the upregulated genes markedly higher than that for the downregulated genes. Inspection and systems biology analysis of the differentially expressed genes revealed an enrichment of genes involved in the inflammatory response, cell signalling pathways and apoptosis. The transcriptional changes associated with cellular signalling and the inflammatory response may reflect different immuno-modulatory mechanisms that underlie host-pathogen interactions during infection.

  20. Identification of latent tuberculosis infection-related microRNAs in human U937 macrophages expressing Mycobacterium tuberculosis Hsp16.3.

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    Meng, Qing-Lin; Liu, Fei; Yang, Xing-Yuan; Liu, Xiao-Mei; Zhang, Xia; Zhang, Chun; Zhang, Zong-De

    2014-02-12

    Latent tuberculosis infection (LTBI) relies on a homeostasis of macrophages and Mycobacterium tuberculosis (Mtb). The small heat shock protein, Mtb Hsp16.3 (also known as latency-associated antigen), plays an important role in Mtb persistence within macrophages. However, the mechanism of LTBI remains elusive. The aim of this study was to delineate LTBI-related miRNA expression in U937 macrophages expressing Mtb Hsp16.3 protein. U937 macrophages were infected with an integrase-deficient Lentivirus vector to transiently express Mtb Hsp16.3, and green fluorescent protein (GFP) as a control. We used a microRNA (miRNA) microarray chip containing more than 1000 probes to identify the significant differentially expressed miRNAs in the infected U937 cells, and employed real-time quantitative polymerase chain reaction (qRT-PCR) for validation. Furthermore, we confirmed these candidate LTBI-related miRNAs in peripheral blood mononuclear cells from subjects with LTBI and in healthy control individuals. Functional annotation prediction of miRNA target genes and pathway enrichment analyses were used to explore the putative links between these miRNAs and LTBI. Analysis of the miRNA expression profile identified 149 miRNAs that were differentially expressed in U937 macrophages expressing Mtb Hsp16.3 compared with the control expressing GFP. The expression level of seven miRNAs (miR-424-5p, miR-493-5p, miR-296-5p, miR-27b-3p, miR-377-5p, miR-3680-5p, miR-191-5p) were validated by qRT-PCR. The expression level of four miRNAs (miR-424-5p, miR-27b-3p, miR-377-5p, miR-3680-5p) in the peripheral blood mononuclear cells samples from LTBI and healthy participants reflected the altered patterns observed in the microarray profile. The bioinformatic analyses suggest that the miRNAs may regulate Mtb latent infection by affecting the development of macrophage cells. The results suggest that miRNA expression may play a considerable role in the pathogenesis of LTBI, and this would increase our

  1. Proteinosis alveolar pulmonar Pulmonary alveolar proteinosis

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    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.

  2. The co‐transcriptome of uropathogenic E scherichia coli‐infected mouse macrophages reveals new insights into host–pathogen interactions

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    Mavromatis, Charalampos (Harris); Bokil, Nilesh J.; Totsika, Makrina; Kakkanat, Asha; Schaale, Kolja; Cannistraci, Carlo V.; Ryu, Taewoo; Beatson, Scott A.; Ulett, Glen C.; Schembri, Mark A.; Sweet, Matthew J.

    2015-01-01

    Summary Urinary tract infections (UTI) are among the most common infections in humans. Uropathogenic E scherichia coli (UPEC) can invade and replicate within bladder epithelial cells, and some UPEC strains can also survive within macrophages. To understand the UPEC transcriptional programme associated with intramacrophage survival, we performed host–pathogen co‐transcriptome analyses using RNA sequencing. Mouse bone marrow‐derived macrophages (BMMs) were challenged over a 24 h time course with two UPEC reference strains that possess contrasting intramacrophage phenotypes: UTI89, which survives in BMMs, and 83972, which is killed by BMMs. Neither of these strains caused significant BMM cell death at the low multiplicity of infection that was used in this study. We developed an effective computational framework that simultaneously separated, annotated and quantified the mammalian and bacterial transcriptomes. Bone marrow‐derived macrophages responded to the two UPEC strains with a broadly similar gene expression programme. In contrast, the transcriptional responses of the UPEC strains diverged markedly from each other. We identified UTI89 genes up‐regulated at 24 h post‐infection, and hypothesized that some may contribute to intramacrophage survival. Indeed, we showed that deletion of one such gene (pspA) significantly reduced UTI89 survival within BMMs. Our study provides a technological framework for simultaneously capturing global changes at the transcriptional level in co‐cultures, and has generated new insights into the mechanisms that UPEC use to persist within the intramacrophage environment. PMID:25410299

  3. The absence of MyD88 has no effect on the induction of alternatively activated macrophage during Fasciola hepatica infection

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    Luo HongLin

    2011-11-01

    Full Text Available Abstract Background Alternatively activated macrophages (AAMϕ play important roles in allergies and responses to parasitic infections. However, whether signaling through toll-like receptors (TLRs plays any role in AAMϕ induction when young Fasciola hepatica penetrates the liver capsule and migrates through the liver tissue is still unclear. Results The data show that the lack of myeloid differentiation factor 88 (MyD88 has no effect on the AAMϕ derived from the bone marrow (BMMϕ in vitro and does not impair the mRNA expression of arginase-1, resistin-like molecule (RELMα, and Ym1 in BMMϕs. The Th2 cytokine production bias in splenocytes was not significantly altered in F. hepatica-infected mice in the absence of MyD88 in vitro and in the pleural cavity lavage in vivo. In addition, MyD88-deficiency has no effect on the arginase production of the F. hepatica elicited macrophages (Fe Mϕs, production of RELMα and Ym1 proteins and mRNA expression of Ym1 and RELMα of macrophages in the peritoneal cavity 6 weeks post F. hepatica infection. Conclusions The absence of MyD88 has no effect on presence of AAMϕ 6 weeks post F. hepatica infection.

  4. Alveolar proteinosis associated with aluminium dust inhalation.

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    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.

  5. Infective Larvae of Brugia malayi Induce Polarization of Host Macrophages that Helps in Immune Evasion

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    Aditi Sharma

    2018-02-01

    Full Text Available Filarial parasites suppress, divert, or polarize the host immune response to aid their survival. However, mechanisms that govern the polarization of host MΦs during early filarial infection are not completely understood. In this study, we infected BALB/c mice with infective larvae stage-3 of Brugia malayi (Bm-L3 and studied their effect on the polarization of splenic MΦs. Results showed that MΦs displayed M2-phenotype by day 3 p.i. characterized by upregulated IL-4, but reduced IL-12 and Prostaglandin-D2 secretion. Increased arginase activity, higher arginase-1 but reduced NOS2 expression and poor phagocytic and antigen processing capacity was also observed. M2 MΦs supported T-cell proliferation and characteristically upregulated p-ERK but downregulated NF-κB-p65 and NF-κB-p50/105. Notably, Bm-L3 synergized with host regulatory T-cells (Tregs and polarized M2 MΦs to regulatory MΦs (Mregs by day 7 p.i., which secreted copious amounts of IL-10 and prostaglandin-E2. Mregs also showed upregulated expression levels of MHC-II, CD80, and CD86 and exhibited increased antigen-processing capacity but displayed impaired activation of NF-κB-p65 and NF-κB-p50/105. Neutralization of Tregs by anti-GITR + anti-CD25 antibodies checked the polarization of M2 MΦs to Mregs, decreased accumulation of regulatory B cells and inflammatory monocytes, and reduced secretion of IL-10, but enhanced IL-4 production and percentages of eosinophils, which led to Bm-L3 killing. In summary, we report hitherto undocumented effects of early Bm-L3 infection on the polarization of splenic MΦs and show how infective larvae deftly utilize the functional plasticity of host MΦs to establish themselves inside the host.

  6. Liposomes or traditional adjuvants: induction of bactericidal activity by the macrophage infectivity potentiator protein (Mip) of Neisseria meningitidis.

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    Costoya, Liliana; Marzoa, Juan; Ferreirós, Carlos; Criado, Maria Teresa

    2017-08-01

    Currently, one of the main approaches to achieve a vaccine for serogroup B Neisseria meningitidis is based on outer membrane proteins with low antigenic variability among strains. Since these proteins tend to be minor components of the outer membrane, recombinant production is required to obtain them in sufficient amounts for evaluation and development of vaccines. In this study, we analysed the ability of recombinant macrophage infectivity potentiator (rMip) protein to induce protective bactericidal activity in mice. The rMip protein was cloned from N. meningitidis strain H44/76 and was used to immunise mice, and the sera obtained were tested against the homologous and several heterologous N. meningitidis strains. The sera were obtained using the rMip alone, with adjuvant Al(OH) 3 , or after inclusion into liposomes. Bactericidal activity was variable depending on the strain, although high titres were seen against strains H44/76 and NmP27. Liposomes enhanced fourfold the reactivity against the homologous strain. The results presented suggest that the rMip protein should be considered a promising candidate for the improvement of future protein-based vaccines. © 2017 APMIS. Published by John Wiley & Sons Ltd.

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

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    Nan Li

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

  8. The Ag85B protein of the BCG vaccine facilitates macrophage uptake but is dispensable for protection against aerosol Mycobacterium tuberculosis infection.

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    Prendergast, Kelly A; Counoupas, Claudio; Leotta, Lisa; Eto, Carolina; Bitter, Wilbert; Winter, Nathalie; Triccas, James A

    2016-05-17

    Defining the function and protective capacity of mycobacterial antigens is crucial for progression of tuberculosis (TB) vaccine candidates to clinical trials. The Ag85B protein is expressed by all pathogenic mycobacteria and is a component of multiple TB vaccines under evaluation in humans. In this report we examined the role of the BCG Ag85B protein in host cell interaction and vaccine-induced protection against virulent Mycobacterium tuberculosis infection. Ag85B was required for macrophage infection in vitro, as BCG deficient in Ag85B expression (BCG:(Δ85B)) was less able to infect RAW 264.7 macrophages compared to parental BCG, while an Ag85B-overexpressing BCG strain (BCG:(oex85B)) demonstrated improved uptake. A similar pattern was observed in vivo after intradermal delivery to mice, with significantly less BCG:(Δ85B) present in CD64(hi)CD11b(hi) macrophages compared to BCG or BCG:(oex85B). After vaccination of mice with BCG:(Δ85B) or parental BCG and subsequent aerosol M. tuberculosis challenge, similar numbers of activated CD4(+) and CD8(+) T cells were detected in the lungs of infected mice for both groups, suggesting the reduced macrophage uptake observed by BCG:(Δ85B) did not alter host immunity. Further, vaccination with both BCG:(Δ85B) and parental BCG resulted in a comparable reduction in pulmonary M. tuberculosis load. These data reveal an unappreciated role for Ag85B in the interaction of mycobacteria with host cells and indicates that single protective antigens are dispensable for protective immunity induced by BCG. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Pulmonary Alveolar Proteinosis in Setting of Inhaled Toxin Exposure and Chronic Substance Abuse

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    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.

  10. Does Administration of Oral Versus Intravenous Antibiotics for Third Molar Removal Have an Effect on the Incidence of Alveolar Osteitis or Postoperative Surgical Site Infections?

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    Reiland, Matthew D; Ettinger, Kyle S; Lohse, Christine M; Viozzi, Christopher F

    2017-09-01

    To compare the incidence of postoperative alveolar osteitis (AO) and surgical site infections (SSIs) in 2 separate cohorts of patients undergoing elective third molar removal: those who received postoperative oral (PO) antibiotics and those who received perioperative intravenous (IV) antibiotics. A retrospective cohort study of all patients 14 to 30 years old undergoing elective outpatient third molar removal under a single surgeon's service over a 12-year period was completed. Patients undergoing third molar removal during the first 72 months received postoperative PO antibiotics alone. Patients undergoing third molar removal during the second 72 months received perioperative IV antibiotics alone. The primary predictor variable for the study was the antibiotic regimen used at the time of third molar removal. The primary outcome variable was the postoperative development of AO or SSI. Covariates included age and gender. Univariable and multivariable regression models assessed for associations between the antibiotic regimen used and the presence of AO and SSI. The study sample consisted of 1,895 patients (1,020 patients receiving postoperative PO antibiotics and 875 patients receiving perioperative IV antibiotics). Of patients receiving postoperative PO antibiotics, 6.4% developed AO an average of 5.7 days after the procedure and 2.6% developed an SSI an average of 23.2 days after the procedure. Of patients receiving perioperative IV antibiotics, 5.5% developed AO an average of 6.2 days after the procedure and 3.3% developed an SSI an average of 18.2 days after the procedure. No statistically significant associations between the antibiotic regimen used and the presence of AO or SSI were identified in univariable (P = 0.42 for AO, P = 0.32 for SSI) or multivariable (P = 0.65 for AO, P = 0.26 for SSI) analyses. In the postoperative PO antibiotic cohort, older age (P third molar removal. If the surgeon chooses to use antibiotics in the setting of third molar

  11. Nitric Oxide Modulates Macrophage Responses toMycobacterium tuberculosisInfection through Activation of HIF-1α and Repression of NF-κB.

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    Braverman, Jonathan; Stanley, Sarah A

    2017-09-01

    IFN-γ is essential for control of Mycobacterium tuberculosis infection in vitro and in vivo. However, the mechanisms by which IFN-γ controls infection remain only partially understood. One of the crucial IFN-γ target genes required for control of M. tuberculosis is inducible NO synthase (iNOS). Although NO produced by iNOS is thought to have direct bactericidal activity against M. tuberculosis , the role of NO as a signaling molecule has been poorly characterized in the context M. tuberculosis infection. In this study, we found that iNOS broadly regulates the macrophage transcriptome during M. tuberculosis infection, activating antimicrobial pathways while also limiting inflammatory cytokine production. The transcription factor hypoxia inducible factor-1α (HIF-1α) was recently shown to be critical for IFN-γ-mediated control of M. tuberculosis infection. We found that HIF-1α function requires NO production, and that HIF-1α and iNOS are linked by a positive feedback loop that amplifies macrophage activation. Furthermore, we found that NO inhibits NF-κB activity to prevent hyperinflammatory responses. Thus, NO activates robust microbicidal programs while also limiting damaging inflammation. IFN-γ signaling must carefully calibrate an effective immune response that does not cause excessive tissue damage, and this study identifies NO as a key player in establishing this balance during M. tuberculosis infection. Copyright © 2017 by The American Association of Immunologists, Inc.

  12. Overexpression of sICAM-1 in the Alveolar Epithelial Space Results in an Exaggerated Inflammatory Response and Early Death in Gram Negative Pneumonia

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    Curtis Jeffery L

    2011-01-01

    Full Text Available Abstract Background A sizeable body of data demonstrates that membrane ICAM-1 (mICAM-1 plays a significant role in host defense in a site-specific fashion. On the pulmonary vascular endothelium, mICAM-1 is necessary for normal leukocyte recruitment during acute inflammation. On alveolar epithelial cells (AECs, we have shown previously that the presence of normal mICAM-1 is essential for optimal alveolar macrophage (AM function. We have also shown that ICAM-1 is present in the alveolar space as a soluble protein that is likely produced through cleavage of mICAM-1. Soluble intercellular adhesion molecule-1 (sICAM-1 is abundantly present in the alveolar lining fluid of the normal lung and could be generated by proteolytic cleavage of mICAM-1, which is highly expressed on type I AECs. Although a growing body of data suggesting that intravascular sICAM-1 has functional effects, little is known about sICAM-1 in the alveolus. We hypothesized that sICAM-1 in the alveolar space modulates the innate immune response and alters the response to pulmonary infection. Methods Using the surfactant protein C (SPC promoter, we developed a transgenic mouse (SPC-sICAM-1 that constitutively overexpresses sICAM-1 in the distal lung, and compared the responses of wild-type and SPC-sICAM-1 mice following intranasal inoculation with K. pneumoniae. Results SPC-sICAM-1 mice demonstrated increased mortality and increased systemic dissemination of organisms compared with wild-type mice. We also found that inflammatory responses were significantly increased in SPC-sICAM-1 mice compared with wild-type mice but there were no difference in lung CFU between groups. Conclusions We conclude that alveolar sICAM-1 modulates pulmonary inflammation. Manipulating ICAM-1 interactions therapeutically may modulate the host response to Gram negative pulmonary infections.

  13. Whole transcriptome analysis reveals differential gene expression profile reflecting macrophage polarization in response to influenza A H5N1 virus infection.

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    Zhang, Na; Bao, Yun-Juan; Tong, Amy Hin-Yan; Zuyderduyn, Scott; Bader, Gary D; Malik Peiris, J S; Lok, Si; Lee, Suki Man-Yan

    2018-02-23

    Avian influenza A H5N1 virus can cause lethal disease in humans. The virus can trigger severe pneumonia and lead to acute respiratory distress syndrome. Data from clinical, in vitro and in vivo suggest that virus-induced cytokine dysregulation could be a contributory factor to the pathogenesis of human H5N1 disease. However, the precise mechanism of H5N1 infection eliciting the unique host response are still not well understood. To obtain a better understanding of the molecular events at the earliest time points, we used RNA-Seq to quantify and compare the host mRNA and miRNA transcriptomes induced by the highly pathogenic influenza A H5N1 (A/Vietnam/3212/04) or low virulent H1N1 (A/Hong Kong/54/98) viruses in human monocyte-derived macrophages at 1-, 3-, and 6-h post infection. Our data reveals that two macrophage populations corresponding to M1 (classically activated) and M2 (alternatively activated) macrophage subtypes respond distinctly to H5N1 virus infection when compared to H1N1 virus or mock infection, a distinction that could not be made from previous microarray studies. When this confounding variable is considered in our statistical model, a clear set of dysregulated genes and pathways emerges specifically in H5N1 virus-infected macrophages at 6-h post infection, whilst was not found with H1N1 virus infection. Furthermore, altered expression of genes in these pathways, which have been previously implicated in viral host response, occurs specifically in the M1 subtype. We observe a significant up-regulation of genes in the RIG-I-like receptor signaling pathway. In particular, interferons, and interferon-stimulated genes are broadly affected. The negative regulators of interferon signaling, the suppressors of cytokine signaling, SOCS-1 and SOCS-3, were found to be markedly up-regulated in the initial round of H5N1 virus replication. Elevated levels of these suppressors could lead to the eventual suppression of cellular antiviral genes, contributing to

  14. Inefficient HIV-1 trans infection of CD4+ T cells by macrophages from HIV-1 nonprogressors is associated with altered membrane cholesterol and DC-SIGN.

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    DeLucia, Diana C; Rinaldo, Charles R; Rappocciolo, Giovanna

    2018-04-11

    Professional antigen presenting cells (APC: myeloid dendritic cells (DC) and macrophages (MΦ); B lymphocytes) mediate highly efficient HIV-1 infection of CD4 + T cells, termed trans infection, that could contribute to HIV-1 pathogenesis. We have previously shown that lower cholesterol content in DC and B lymphocytes is associated with a lack of HIV-1 trans infection in HIV-1 infected nonprogressors (NP). Here we assessed whether HIV-1 trans infection mediated by another major APC, MΦ, is deficient in NP due to altered cholesterol metabolism. When comparing healthy HIV-1 seronegatives (SN), rapid progressors (PR), and NP, we found that monocyte-derived MΦ from NP did not mediate HIV-1 trans infection of autologous CD4 + T cells, in contrast to efficient trans infection mediated by SN and PR MΦ. MΦ trans infection efficiency was directly associated with the number of DC-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN)-expressing MΦ. Significantly fewer NP MΦ expressed DC-SIGN. Unesterified (free) cholesterol in MΦ cell membranes and lipid rafting was significantly lower in NP than PR, as well as virus internalization in early endosomes. Furthermore, simvastatin (SIMV), decreased the subpopulation of DC-SIGN + MΦ, as well as MΦ cis and trans infection. Notably, SIMV decreased cell membrane cholesterol and led to lipid raft dissociation, effectively mimicking the incompetent APC trans infection environment characteristic of NP. Our data support that DC-SIGN and membrane cholesterol are central to MΦ trans infection, and a lack of these limits HIV-1 disease progression. Targeting the ability of MΦ to drive HIV-1 dissemination in trans could enhance HIV-1 therapeutic strategies. IMPORTANCE Despite the success of combination anti-retroviral therapy, neither a vaccine nor a cure for HIV infection has been developed, demonstrating a need for novel prophylactic and therapeutic strategies. Here we show that efficiency of macrophage (M

  15. Impaired Chromatin Remodelling at STAT1-Regulated Promoters Leads to Global Unresponsiveness of Toxoplasma gondii-Infected Macrophages to IFN-γ

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    Lang, Christine; Hildebrandt, Anke; Brand, Franziska; Opitz, Lennart; Dihazi, Hassan; Lüder, Carsten G. K.

    2012-01-01

    Intracellular pathogens including the apicomplexan and opportunistic parasite Toxoplasma gondii profoundly modify their host cells in order to establish infection. We have shown previously that intracellular T. gondii inhibit up-regulation of regulatory and effector functions in murine macrophages (MΦ) stimulated with interferon (IFN)-γ, which is the cytokine crucial for controlling the parasites' replication. Using genome-wide transcriptome analysis we show herein that infection with T. gondii leads to global unresponsiveness of murine macrophages to IFN-γ. More than 61% and 89% of the transcripts, which were induced or repressed by IFN-γ in non-infected MΦ, respectively, were not altered after stimulation of T. gondii-infected cells with IFN-γ. These genes are involved in a variety of biological processes, which are mostly but not exclusively related to immune responses. Analyses of the underlying mechanisms revealed that IFN-γ-triggered nuclear translocation of STAT1 still occurred in Toxoplasma-infected MΦ. However, STAT1 bound aberrantly to oligonucleotides containing the IFN-γ-responsive gamma-activated site (GAS) consensus sequence. Conversely, IFN-γ did not induce formation of active GAS-STAT1 complexes in nuclear extracts from infected MΦ. Mass spectrometry of protein complexes bound to GAS oligonucleotides showed that T. gondii-infected MΦ are unable to recruit non-muscle actin to IFN-γ-responsive DNA sequences, which appeared to be independent of stimulation with IFN-γ and of STAT1 binding. IFN-γ-induced recruitment of BRG-1 and acetylation of core histones at the IFN-γ-regulated CIITA promoter IV, but not β-actin was diminished by >90% in Toxoplasma-infected MΦ as compared to non-infected control cells. Remarkably, treatment with histone deacetylase inhibitors restored the ability of infected macrophages to express the IFN-γ regulated genes H2-A/E and CIITA. Taken together, these results indicate that Toxoplasma-infected MΦ are

  16. In vitro dose and duration dependent approaches for the assessment of ameliorative effects of nanoconjugated vancomycin against Staphylococcus aureus infection induced oxidative stress in murine peritoneal macrophages.

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    Chakraborty, Subhankari Prasad; Pramanik, Panchanan; Roy, Somenath

    2016-02-01

    The present study was aimed to evaluate the in vitro ameliorative effect of nanoconjugated vancomycin (NV) against vancomycin sensitive and resistant strains of Staphylococcus aureus infection-induced oxidative stress in murine peritoneal macrophage. Peritoneal macrophages from mice were treated with VSSA and VRSA (5 × 10(6) CFU/mL), VSSA + NV (5-250 μg/ml) and VRSA + NV (5-250 μg/ml) for 18 h, having 3 h interval in culture media; and the superoxide anion generation, lipid peroxidation, protein oxidation, antioxidant enzymes status and glutathione enzymes activity were monitored. The significantly increased free radical generation, lipid peroxidation, protein carbonyls and oxidized glutathione levels were observed in VSSA and VRSA treated group as compared to control group; where as reduced glutathione level, antioxidant enzymes status and glutathione dependent enzymes were decreased significantly. All these changes come near to control in NV treated group in a dose and duration dependent fashion. Among the different doses and duration intervals of NV, maximum ameliorative effect was observed by 100 μg/ml for 12 h treatment which does not produce any damage to the cell. These findings suggest the potential use and beneficial role of nanoconjugated vancomycin as a modulator of S. aureus infection-induced cellular damage in murine peritoneal macrophage. Copyright © 2015. Published by Elsevier Ltd.

  17. Antagonism of miR-328 increases the antimicrobial function of macrophages and neutrophils and rapid clearance of non-typeable Haemophilus influenzae (NTHi from infected lung.

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    Hock L Tay

    2015-04-01

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

  18. Distinct regulation of host responses by ERK and JNK MAP kinases in swine macrophages infected with pandemic (H1N1 2009 influenza virus.

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    Wei Gao

    Full Text Available Swine influenza is an acute respiratory disease in pigs caused by swine influenza virus (SIV. Highly virulent SIV strains cause mortality of up to 10%. Importantly, pigs have long been considered "mixing vessels" that generate novel influenza viruses with pandemic potential, a constant threat to public health. Since its emergence in 2009 and subsequent pandemic spread, the pandemic (H1N1 2009 (H1N1pdm has been detected in pig farms, creating the risk of generating new reassortants and their possible infection of humans. Pathogenesis in SIV or H1N1pdm-infected pigs remains poorly characterized. Proinflammatory and antiviral cytokine responses are considered correlated with the intensity of clinical signs, and swine macrophages are found to be indispensible in effective clearance of SIV from pig lungs. In this study, we report a unique pattern of cytokine responses in swine macrophages infected with H1N1pdm. The roles of mitogen-activated protein (MAP kinases in the regulation of the host responses were examined. We found that proinflammatory cytokines IL-6, IL-8, IL-10, and TNF-α were significantly induced and their induction was ERK1/2-dependent. IFN-β and IFN-inducible antiviral Mx and 2'5'-OAS were sharply induced, but the inductions were effectively abolished when ERK1/2 was inhibited. Induction of CCL5 (RANTES was completely inhibited by inhibitors of ERK1/2 and JNK1/2, which appeared also to regulate FasL and TNF-α, critical for apoptosis in pig macrophages. We found that NFκB was activated in H1N1pdm-infected cells, but the activation was suppressed when ERK1/2 was inhibited, indicating there is cross-talk between MAP kinase and NFκB responses in pig macrophages. Our data suggest that MAP kinase may activate NFκB through the induction of RIG-1, which leads to the induction of IFN-β in swine macrophages. Understanding host responses and their underlying mechanisms may help identify venues for effective control of SIV and assist in

  19. Mapping of the Co-Transcriptomes of UPEC-Infected Macrophages Reveals New Insights into the Molecular Basis of Host-Pathogen Interactions in Human and Mouse

    KAUST Repository

    Mavromatis, Charalampos Harris

    2014-01-01

    Urinary tract infections (UTI) are among the most common infections in humans. Uropathogenic Escherichia coli (UPEC), the main causative agent of UTIs, can invade and replicate within bladder epithelial cells, and recent evidence demonstrated that some UPEC strains also survive within macrophages. To understand the mechanisms of host subversion that enable UPEC to survive within macrophages, and the contribution of macrophages to UPEC-mediated pathology, I performed hostpathogen co-transcriptome analyses using RNA sequencing. I developed an effective computational framework that simultaneously separated, annotated, and quantified the mammalian and bacterial transcriptomes. First, mouse bone morrow-derived macrophages (BMM) were challenged over a 24 h time course with UPEC reference strains, UTI89 (cystitis strain), 83972 and VR50 (asymptomatic bacteriuria strains) that possess contrasting intramacrophage phenotypes. My results showed that BMM responded to the three different UPEC strains with broadly similar gene expression programs. In contrast to the conserved pattern of BMM responses, the transcriptional responses of the different UPEC strains diverged markedly from each other. Hypothesizing that genes upregulated at 24 h post-infection may contribute to intramacrophage survival, I identified UTI89 genes upregulated at this time point, and showed that deletion of one of these genes (pspA) compromised intramacrophage survival of UPEC strain UTI89. Second, human monocyte-derived macrophages (HMDM) and BMM were challenged over a 24 h course with the UPEC strain EC958, a globally disseminated, multi-drug resistant strain. My analysis identified extensive divergence in UPEC-regulated orthologous gene expression between HMDM and BMM, and I validated both known and novel genes in the context of differential regulation. On the contrary, the transcriptional response of EC958 showed a broad conservation across both mammalian intramacrophage environments. My study thus

  20. Ehrlichia chaffeensis infection in the reservoir host (white-tailed deer and in an incidental host (dog is impacted by its prior growth in macrophage and tick cell environments.

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    Arathy D S Nair

    Full Text Available Ehrlichia chaffeensis, transmitted from Amblyomma americanum ticks, causes human monocytic ehrlichiosis. It also infects white-tailed deer, dogs and several other vertebrates. Deer are its reservoir hosts, while humans and dogs are incidental hosts. E. chaffeensis protein expression is influenced by its growth in macrophages and tick cells. We report here infection progression in deer or dogs infected intravenously with macrophage- or tick cell-grown E. chaffeensis or by tick transmission in deer. Deer and dogs developed mild fever and persistent rickettsemia; the infection was detected more frequently in the blood of infected animals with macrophage inoculum compared to tick cell inoculum or tick transmission. Tick cell inoculum and tick transmission caused a drop in tick infection acquisition rates compared to infection rates in ticks fed on deer receiving macrophage inoculum. Independent of deer or dogs, IgG antibody response was higher in animals receiving macrophage inoculum against macrophage-derived Ehrlichia antigens, while it was significantly lower in the same animals against tick cell-derived Ehrlichia antigens. Deer infected with tick cell inoculum and tick transmission caused a higher antibody response to tick cell cultured bacterial antigens compared to the antibody response for macrophage cultured antigens for the same animals. The data demonstrate that the host cell-specific E. chaffeensis protein expression influences rickettsemia in a host and its acquisition by ticks. The data also reveal that tick cell-derived inoculum is similar to tick transmission with reduced rickettsemia, IgG response and tick acquisition of E. chaffeensis.

  1. Use of a capture-based pathogen transcript enrichment strategy for RNA-Seq analysis of the Francisella tularensis LVS transcriptome during infection of murine macrophages.

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    Zachary W Bent

    Full Text Available Francisella tularensis is a zoonotic intracellular pathogen that is capable of causing potentially fatal human infections. Like all successful bacterial pathogens, F. tularensis rapidly responds to changes in its environment during infection of host cells, and upon encountering different microenvironments within those cells. This ability to appropriately respond to the challenges of infection requires rapid and global shifts in gene expression patterns. In this study, we use a novel pathogen transcript enrichment strategy and whole transcriptome sequencing (RNA-Seq to perform a detailed characterization of the rapid and global shifts in F. tularensis LVS gene expression during infection of murine macrophages. We performed differential gene expression analysis on all bacterial genes at two key stages of infection: phagosomal escape, and cytosolic replication. By comparing the F. tularensis transcriptome at these two stages of infection to that of the bacteria grown in culture, we were able to identify sets of genes that are differentially expressed over the course of infection. This analysis revealed the temporally dynamic expression of a number of known and putative transcriptional regulators and virulence factors, providing insight into their role during infection. In addition, we identified several F. tularensis genes that are significantly up-regulated during infection but had not been previously identified as virulence factors. These unknown genes may make attractive therapeutic or vaccine targets.

  2. Establishment and evaluation of a stable cattle type II alveolar epithelial cell line.

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    Feng Su

    Full Text Available Macrophages and dendritic cells are recognized as key players in the defense against mycobacterial infection. Recent research has confirmed that alveolar epithelial cells (AECs also play important roles against mycobacterium infections. Thus, establishing a stable cattle AEC line for future endogenous immune research on bacterial invasion is necessary. In the present study, we first purified and immortalized type II AECs (AEC II cells by transfecting them with a plasmid containing the human telomerase reverse trancriptase gene. We then tested whether or not the immortalized cells retained the basic physiological properties of primary AECs by reverse-transcription polymerase chain reaction and Western blot. Finally, we tested the secretion capacity of immortalized AEC II cells upon stimulation by bacterial invasion. The cattle type II alveolar epithelial cell line (HTERT-AEC II that we established retained lung epithelial cell characteristics: the cells were positive for surfactants A and B, and they secreted tumor necrosis factor-α and interleukin-6 in response to bacterial invasion. Thus, the cell line we established is a potential tool for research on the relationship between AECs and Mycobacterium tuberculosis.

  3. Microparticles released from Mycobacterium tuberculosis-infected human macrophages contain increased levels of the type I interferon inducible proteins including ISG15.

    Science.gov (United States)

    Hare, Nathan J; Chan, Brian; Chan, Edwina; Kaufman, Kimberley L; Britton, Warwick J; Saunders, Bernadette M

    2015-09-01

    Microparticles (MPs) are small membranous particles (100-1000 nm) released under normal steady-state conditions and are thought to provide a communication network between host cells. Previous studies demonstrated that Mycobacterium tuberculosis (M. tb) infection of macrophages increased the release of MPs, and these MPs induced a proinflammatory response from uninfected macrophages in vitro and in vivo following their transfer into uninfected mice. To determine how M. tb infection modulates the protein composition of the MPs, and if this contributes to their proinflammatory properties, we compared the proteomes of MPs derived from M. tb-infected (TBinf-MP) and uninfected human THP-1 monocytic cells. MP proteins were analyzed by GeLC-MS/MS with spectral counting revealing 68 proteins with statistically significant differential abundances. The 42 proteins increased in abundance in TBinf-MPs included proteins associated with immune function (7), lysosomal/endosomal maturation (4), vesicular formation (12), nucleosome proteins (4), and antigen processing (9). Prominent among these were the type I interferon inducible proteins, ISG15, IFIT1, IFIT2, and IFIT3. Exposure of uninfected THP-1 cells to TBinf-MPs induced increased gene expression of isg15, ifit1, ifit2, and ifit3 and the release of proinflammatory cytokines. These proteins may regulate the proinflammatory potential of the MPs and provide candidate biomarkers for M. tb infection. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. MicroRNA-206 regulates the secretion of inflammatory cytokines and MMP9 expression by targeting TIMP3 in Mycobacterium tuberculosis-infected THP-1 human macrophages.

    Science.gov (United States)

    Fu, Xiangdong; Zeng, Lihong; Liu, Zhi; Ke, Xue; Lei, Lin; Li, Guobao

    2016-08-19

    Tuberculosis (TB) is a serious disease that is characterized by Mycobacterium tuberculosis (M.tb)-triggered immune system impairment and lung tissue damage shows limited treatment options. MicroRNAs (miRNAs) are regulators of gene expression that play critical roles in many human diseases, and can be up- or downregulated by M.tb infection in macrophage. Recently, tissue inhibitor of matrix metalloproteinase (TIMP) 3 has been found to play roles in regulating macrophage inflammation. Here, we found that TIMP3 expression was regulated by miR-206 in M.tb-infected THP-1 human macrophages. In THP-1 cells infected with M.tb, the miR-206 level was significantly upregulated and the expression of TIMP3 was markedly decreased when the secretion of inflammatory cytokines was increased. Inhibition of miR-206 markedly suppressed inflammatory cytokine secretion and upregulated the expression of TIMP3. In contrast, the upregulation of miR-206 promoted the matrix metalloproteinase (MMP) 9 levels and inhibited TIMP3 levels. Using a dual-luciferase reporter assay, a direct interaction between miR-206 and the 3'-untranslated region (UTR) of TIMP3 was confirmed. SiTIMP3, the small interfering RNA (siRNA) specific for TIMP3, significantly attenuated the suppressive effects of miR-206-inhibitor on inflammatory cytokine secretion and MMP9 expression. Our data suggest that miR-206 may function as an inflammatory regulator and drive the expression of MMP9 in M.tb-infected THP-1 cells by targeting TIMP3, indicating that miR-206 is a potential therapeutic target for patients with TB. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Pulmonary alveolar microlithiasis

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    Surender Kashyap

    2013-01-01

    Full Text Available Pulmonary alveolar microlithiasis (PAM is a rare, chronic lung disease with bilateral intra-alveolar calcium and phosphate deposition throughout the lung parenchyma with predominance to lower and midzone. Although, etiology and pathogenesis of PAM is not fully understood, the mutation in SLC34A2 gene that encodes a sodium-phosphate co-transporter in alveolar type II cells resulting in the accumulation and forming of microliths rich in calcium phosphate (due to impaired clearance are considered to be the cause of the disease. Chest radiograph and high-resolution CT of thorax are nearly pathognomonic for diagnosing PAM. HRCT demonstrates diffuse micronodules showing slight perilobular predominance resulting in calcification of interlobular septa. Patients with PAM are asymptomatic till development of hypoxemia and cor-pulmonale. No therapy has been proven to be beneficial except lung transplantation.

  6. TLR3 signaling in macrophages is indispensable for the protective immunity of invariant natural killer T cells against enterovirus 71 infection.

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    Kai Zhu

    2015-01-01

    Full Text Available Enterovirus 71 (EV71 is the most virulent pathogen among enteroviruses that cause hand, foot and mouth disease in children but rarely in adults. The mechanisms that determine the age-dependent susceptibility remain largely unclear. Here, we found that the paucity of invariant natural killer T (iNKT cells together with immaturity of the immune system was related to the susceptibility of neonatal mice to EV71 infection. iNKT cells were crucial antiviral effector cells to protect young mice from EV71 infection before their adaptive immune systems were fully mature. EV71 infection led to activation of iNKT cells depending on signaling through TLR3 but not other TLRs. Surprisingly, iNKT cell activation during EV71 infection required TLR3 signaling in macrophages, but not in dendritic cells (DCs. Mechanistically, interleukin (IL-12 and endogenous CD1d-restricted antigens were both required for full activation of iNKT cells. Furthermore, CD1d-deficiency led to dramatically increased viral loads in central nervous system and more severe disease in EV71-infected mice. Altogether, our results suggest that iNKT cells may be involved in controlling EV71 infection in children when their adaptive immune systems are not fully developed, and also imply that iNKT cells might be an intervention target for treating EV71-infected patients.

  7. TLR3 signaling in macrophages is indispensable for the protective immunity of invariant natural killer T cells against enterovirus 71 infection.

    Science.gov (United States)

    Zhu, Kai; Yang, Juhao; Luo, Kaiming; Yang, Chunhui; Zhang, Na; Xu, Ruifeng; Chen, Jianxia; Jin, Mingfei; Xu, Bin; Guo, Nining; Wang, Jianrong; Chen, Zuolong; Cui, Ying; Zhao, Hui; Wang, Yan; Deng, Chaoyang; Bai, Li; Ge, Baoxue; Qin, Cheng-Feng; Shen, Hao; Yang, Chun-Fu; Leng, Qibin

    2015-01-01

    Enterovirus 71 (EV71) is the most virulent pathogen among enteroviruses that cause hand, foot and mouth disease in children but rarely in adults. The mechanisms that determine the age-dependent susceptibility remain largely unclear. Here, we found that the paucity of invariant natural killer T (iNKT) cells together with immaturity of the immune system was related to the susceptibility of neonatal mice to EV71 infection. iNKT cells were crucial antiviral effector cells to protect young mice from EV71 infection before their adaptive immune systems were fully mature. EV71 infection led to activation of iNKT cells depending on signaling through TLR3 but not other TLRs. Surprisingly, iNKT cell activation during EV71 infection required TLR3 signaling in macrophages, but not in dendritic cells (DCs). Mechanistically, interleukin (IL)-12 and endogenous CD1d-restricted antigens were both required for full activation of iNKT cells. Furthermore, CD1d-deficiency led to dramatically increased viral loads in central nervous system and more severe disease in EV71-infected mice. Altogether, our results suggest that iNKT cells may be involved in controlling EV71 infection in children when their adaptive immune systems are not fully developed, and also imply that iNKT cells might be an intervention target for treating EV71-infected patients.