Significant Correlation between TLR2 Agonist Activity and TNF-α Induction in J774.A1 Macrophage Cells by Different Medicinal Mushroom Products.

PubMed

Coy, Catherine; Standish, Leanna J; Bender, Geoff; Lu, Hailing

2015-01-01

In the US market, there is a variety of mushroom preparations available, even within the same species of mushroom. Nonetheless, little is known about whether species or the various extraction methods affect biological activity and potency of the immune modulatory activity of mushroom extracts. After discovering that protein-bound polysaccharide-K, a hot water extract from Trametes versicolor, was a potent Toll-like receptor (TLR)-2 agonist that stimulates both innate and adaptive immunity, this study was initiated to evaluate whether other medicinal mushroom products also have TLR2 agonist activity and immune-enhancing potential as measured by the induction of tumor necrosis factor (TNF)-α in J774.A1 murine macrophage cells. Furthermore, the products were divided by extraction method and species to determine whether these factors affect their immunomodulatory activity. The results showed that the majority (75%) of mushroom products tested had TLR2 agonist activity and that there was a significant correlation between TLR2 agonist activity and TNF-α induction potential in the mushroom products analyzed. In addition, the data demonstrated that hot water mushroom extracts are more potent than ground mushroom products in activating TLR2 and inducing TNF-α. These data provide evidence that extraction methods may affect the biological activity of mushroom products; thus, further studies are warranted to investigate the structural differences between various mushroom products.

Protective effects of a standard extract of Mangifera indica L. (VIMANG) against mouse ear edemas and its inhibition of eicosanoid production in J774 murine macrophages.

PubMed

Garrido, G; González, D; Lemus, Y; Delporte, C; Delgado, R

2006-06-01

A standard aqueous extract of Mangifera indica L., used in Cuba as antioxidant under the brand name VIMANG, was tested in vivo for its anti-inflammatory activity, using commonly accepted assays. The standard extract of M. indica, administered orally (50-200mg/kg body wt.), reduced ear edema induced by arachidonic acid (AA) and phorbol myristate acetate (PMA) in mice. In the PMA model, M. indica extract also reduced myeloperoxidase (MPO) activity. In vitro studies were performed using macrophage cell line J774 stimulated with pro-inflammatory stimuli lipopolysaccharide-interferon gamma (LPS-IFNgamma) or calcium ionophore A23187 to determine prostaglandin PGE(2) or leukotriene LTB(4) release, respectively. The extract inhibited the induction of PGE(2) and LTB(4) with IC(50) values of 21.7 and 26.0microg/ml, respectively. Mangiferin (a glucosylxanthone isolated from the extract) also inhibited these AA metabolites (PGE(2), IC(50) value=17.2microg/ml and LTB(4), IC(50) value=2.1microg/ml). These results represent an important contribution to the elucidation of the mechanism involved in the anti-inflammatory and anti-nociceptive effects reported for the standard extract of M. indica VIMANG.

Anti-inflammatory effects of shea butter through inhibition of iNOS, COX-2, and cytokines via the Nf-κB pathway in LPS-activated J774 macrophage cells.

PubMed

Verma, Nandini; Chakrabarti, Rina; Das, Rakha H; Gautam, Hemant K

2012-01-12

Shea butter is traditionally used in Africa for its anti-inflammatory and analgesic effects. In this study we explored the anti-inflammatory activities of the methanolic extract of shea butter (SBE) using lipopolysaccharide (LPS)-induced murine macrophage cell line J774. It was observed that SBE significantly reduced the levels of LPS-induced nitric oxide, Tumor necrosis factor-α (TNF-α), interleukins, 1β (IL-1β), and -12 (IL-12) in the culture supernatants in a dose dependent manner. Expression of pro-inflammatory enzymes, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) were also inhibited by SBE. These anti-inflammatory effects were due to an inhibitory action of SBE on LPS-induced iNOS, COX-2, TNF-α, IL-1β, and IL-12 mRNA expressions. Moreover, SBE efficiently suppressed IκB phosphorylation and NF-κB nuclear translocation induced by LPS. These findings explain the molecular bases of shea butter's bioactivity against various inflammatory conditions and substantiate it as a latent source of novel therapeutic agents.

Quantitative measurements of intercellular adhesion between a macrophage and cancer cells using a cup-attached AFM chip.

PubMed

Kim, Hyonchol; Yamagishi, Ayana; Imaizumi, Miku; Onomura, Yui; Nagasaki, Akira; Miyagi, Yohei; Okada, Tomoko; Nakamura, Chikashi

2017-07-01

Intercellular adhesion between a macrophage and cancer cells was quantitatively measured using atomic force microscopy (AFM). Cup-shaped metal hemispheres were fabricated using polystyrene particles as a template, and a cup was attached to the apex of the AFM cantilever. The cup-attached AFM chip (cup-chip) approached a murine macrophage cell (J774.2), the cell was captured on the inner concave of the cup, and picked up by withdrawing the cup-chip from the substrate. The cell-attached chip was advanced towards a murine breast cancer cell (FP10SC2), and intercellular adhesion between the two cells was quantitatively measured. To compare cell adhesion strength, the work required to separate two adhered cells (separation work) was used as a parameter. Separation work was almost 2-fold larger between a J774.2 cell and FP10SC2 cell than between J774.2 cell and three additional different cancer cells (4T1E, MAT-LyLu, and U-2OS), two FP10SC2 cells, or two J774.2 cells. FP10SC2 was established from 4T1E as a highly metastatic cell line, indicates separation work increased as the malignancy of cancer cells became higher. One possible explanation of the strong adhesion of macrophages to cancer cells observed in this study is that the measurement condition mimicked the microenvironment of tumor-associated macrophages (TAMs) in vivo, and J774.2 cells strongly expressed CD204, which is a marker of TAMs. The results of the present study, which were obtained by measuring cell adhesion strength quantitatively, indicate that the fabricated cup-chip is a useful tool for measuring intercellular adhesion easily and quantitatively. Copyright © 2017 Elsevier B.V. All rights reserved.

Activation of macrophages stimulated by the bengkoang fiber extract through toll-like receptor 4.

PubMed

Kumalasari, Ika Dyah; Nishi, Kosuke; Putra, Agus Budiawan Naro; Sugahara, Takuya

2014-07-25

Bengkoang (Pachyrhizus erosus (L.) Urban) is an edible root tuber containing fairly large amounts of carbohydrates and crude fibers. Our previous studies showed that the bengkoang fiber extract (BFE) stimulates activation of macrophages, leading to induction of phagocytotic activity and cytokine production. In the present study we investigated the mechanism underlying activation of murine macrophages by BFE. BFE increased production of TNF-α, IL-6, and nitric oxide by J774.1 cells. In addition BFE also facilitated the gene expression levels of inducible nitric oxide synthase. We examined the effect of a TLR4 inhibitor on cytokine production to investigate the membrane receptor of macrophage activation by BFE. Treatment of J774.1 cells with the TLR4 inhibitor significantly inhibited production of IL-6 and TNF-α, suggesting that TLR4 is the target membrane receptor for BFE. The main signal molecules located downstream of TLR4 such as JNK, p38, ERK, and NF-κB were activated by BFE treatment. The immunostimulatory effect of BFE was cancelled by the pectinase treatment, suggesting that the active ingredient in BFE is pectin-like molecules. Overall results suggested that BFE activates J774.1 cells via the MAPK and NF-κB signaling pathways.

Fc-receptor induced cell spreading during frustrated phagocytosis in J774A.1 macrophages

NASA Astrophysics Data System (ADS)

Kovari, Daniel; Curtis, Jennifer; Wei, Wenbin

2014-03-01

Phagocytosis is the process where by cells engulf foreign particles. It is the primary mechanism through which macrophages and neutrophils (white blood cells) eliminate pathogens and debris from the body. The behavior is the result of a cascade of chemical and mechanical cues, which result in the actin-driven expansion of the cell's membrane around its target. For macrophages undergoing Fc-mediated phagocytosis, we show that above a minimum threshold the spreading rate and maximum cell-target contact area are independent of the target's opsonin density. Qualitatively, macrophage phagocytic spreading is similar to the spreading of other cell types (e.g. fibroblasts, lymphocytes, and Dict.d.). Early spreading is most likely the result of ``passive'' alignment of the cell to the target surface. This is followed by an active expansion period driven by actin. Finally upon reaching a maximum contact area, typically 2-3 times the size of ``non-activated'' cells, macrophages often undergo a period of rapid contraction not reported in other cell types. We hypothesize that this, as yet unexplained, transition may be specific to the chemical and mechanical machinery associated with phagocytosis. This work was funded by NSF grant PHYS 0848797 and NSF grant DMR 0820382.

Fibronectin induces macrophage migration through a SFK-FAK/CSF-1R pathway.

PubMed

Digiacomo, Graziana; Tusa, Ignazia; Bacci, Marina; Cipolleschi, Maria Grazia; Dello Sbarba, Persio; Rovida, Elisabetta

2017-07-04

Integrins, following binding to proteins of the extracellular matrix (ECM) including collagen, laminin and fibronectin (FN), are able to transduce molecular signals inside the cells and to regulate several biological functions such as migration, proliferation and differentiation. Besides activation of adaptor molecules and kinases, integrins transactivate Receptor Tyrosine Kinases (RTK). In particular, adhesion to the ECM may promote RTK activation in the absence of growth factors. The Colony-Stimulating Factor-1 Receptor (CSF-1R) is a RTK that supports the survival, proliferation, and motility of monocytes/macrophages, which are essential components of innate immunity and cancer development. Macrophage interaction with FN is recognized as an important aspect of host defense and wound repair. The aim of the present study was to investigate on a possible cross-talk between FN-elicited signals and CSF-1R in macrophages. FN induced migration in BAC1.2F5 and J774 murine macrophage cell lines and in human primary macrophages. Adhesion to FN determined phosphorylation of the Focal Adhesion Kinase (FAK) and Src Family Kinases (SFK) and activation of the SFK/FAK complex, as witnessed by paxillin phosphorylation. SFK activity was necessary for FAK activation and macrophage migration. Moreover, FN-induced migration was dependent on FAK in either murine macrophage cell lines or human primary macrophages. FN also induced FAK-dependent/ligand-independent CSF-1R phosphorylation, as well as the interaction between CSF-1R and β1. CSF-1R activity was necessary for FN-induced macrophage migration. Indeed, genetic or pharmacological inhibition of CSF-1R prevented FN-induced macrophage migration. Our results identified a new SFK-FAK/CSF-1R signaling pathway that mediates FN-induced migration of macrophages.

Endotoxin contamination of apolipoprotein A-I: effect on macrophage proliferation--a cautionary tale.

PubMed

Jin, Xueting; Xu, Qing; Champion, Keith; Kruth, Howard S

2015-05-01

This technical report addresses the problem of endotoxin contamination of apolipoprotein reagents. Using a bromodeoxyuridine incorporation cell proliferation assay, we observed that human plasma ApoA-I as low as 1 μg/ml resulted in a >90% inhibition in macrophage proliferation. However, not all ApoA-I from different sources showed this effect. We considered the possibility that endotoxin contamination of the apolipoproteins contributed to the differential inhibition of macrophage cell proliferation. Endotoxin alone very potently inhibited macrophage proliferation (0.1 ng/ml inhibited macrophage proliferation>90%). Measurement of endotoxin levels in the apolipoprotein products, including an analysis of free versus total endotoxin, the latter which included endotoxin that was masked due to binding to protein, suggested that free endotoxin mediated inhibition of macrophage proliferation. Despite the use of an advanced endotoxin removal procedure and agents commonly used to inhibit endotoxin action, the potency of endotoxin precluded successful elimination of endotoxin effect. Our findings show that endotoxin contamination can significantly influence apparent apolipoprotein-mediated cell effects (or effects of any other biological products), especially when these products are tested on highly endotoxin-sensitive cells, such as macrophages. Published by Elsevier Ireland Ltd.

Characterization of lipopolysaccharide-stimulated cytokine expression in macrophages and monocytes

USDA-ARS?s Scientific Manuscript database

Inflammation plays a pivotal role in several chronic human conditions and diseases including atherosclerosis, ischemic heart disease, cancer, obesity, diabetes, and autoimmune diseases. In vitro cell culture models such as exposure of mouse macrophage J774A.1 and human monocyte THP-1 cells to bacter...

Endotoxin Contamination of Apolipoprotein A-I: Effect on Macrophage Proliferation – A Cautionary Tale

PubMed Central

Jin, Xueting; Xu, Qing; Champion, Keith; Kruth, Howard S.

2015-01-01

This technical report addresses the problem of endotoxin contamination of apolipoprotein reagents. Using a bromodeoxyuridine incorporation cell proliferation assay, we observed that human plasma ApoA-I as low as 1 μg/ml resulted in a >90% inhibition in macrophage proliferation. However, not all ApoA-I from different sources showed this effect. We considered the possibility that endotoxin contamination of the apolipoproteins contributed to the differential inhibition of macrophage cell proliferation. Endotoxin alone very potently inhibited macrophage proliferation (0.1 ng/ml inhibited macrophage proliferation >90%). Measurement of endotoxin levels in the apolipoprotein products, including an analysis of free versus total endotoxin, the latter which included endotoxin that was masked due to binding to protein, suggested that free endotoxin mediated inhibition of macrophage proliferation. Despite the use of an advanced endotoxin removal procedure and agents commonly used to inhibit endotoxin action, the potency of endotoxin precluded successful elimination of endotoxin effect. Our findings show that endotoxin contamination can significantly influence apparent apolipoprotein-mediated cell effects (or effects of any other biological products), especially when these products are tested on highly endotoxin-sensitive cells, such as macrophages. PMID:25778625

Macrophage-derived microvesicles promote proliferation and migration of Schwann cell on peripheral nerve repair

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhan, Chuan, E-mail: zhchuansy@163.com; Ma, Cheng-bin; Yuan, Hong-mou

Background: Macrophages have been implicated in peripheral nerve regeneration. However, whether macrophages-derived microvesicles (MVs) are involved in this process remains unknown. In the present study, the effects of macrophages-derived MVs on proliferation and migration of Schwann cells (SCs) were evaluated in both in vitro and in vivo. Methods: Human monocytic leukaemia cell line (THP-1) was successfully driven to M1 and M2 phenotypes by delivery of either IFN-γ or IL-4, respectively. SCs incubated with M1 or M2 macrophages-derived MVs, the cell migration and proliferation were assessed, and expression levels of nerve growth factor (NGF) and Laminin were measured. A rat model of sciaticmore » nerve was established and the effects of macrophages-derived MVs on nerve regeneration were investigated. Results: M2-derived MVs elevated migration, proliferation, NFG and Laminin protein levels of SCs compared with M1-or M0-derived MVs. The relative expression levels of miR-223 were also increased in M2 macrophages and M2-derived MVs. Transfected M2 macrophages with miR-223 inhibitor then co-incubated with SCs, an inhibition of cell migration and proliferation and a down-regulated levels of NFG and Laminin protein expression were observed. In vivo, M2-derived MVs significantly increased the infiltration and axon number of SCs. Conclusion: M2-derived MVs promoted proliferation and migration of SCs in vitro and in vivo, which provided a therapeutic strategy for nerve regeneration. - Highlights: • M2 macrophages-derived MVs elevated migration and proliferation of SCs. • M2 macrophages-derived MVs up-regulated NFG and Laminin expression of SCs. • MiR-223 expression was increased in M2 macrophages-derived MVs. • MiR-223 inhibitor reduced migration and proliferation of SCs co-incubated with MVs. • MiR-223 inhibitor down-regulated NFG and Laminin levels of SCs co-incubated with MVs.« less

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

PubMed

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

2013-04-01

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

Macrophages clear refrigerator storage-damaged RBCs and subsequently secrete cytokines in vivo, but not in vitro, in a murine model

PubMed Central

Wojczyk, Boguslaw S.; Kim, Nina; Bandyopadhyay, Sheila; Francis, Richard O.; Zimring, James C.; Hod, Eldad A.; Spitalnik, Steven L.

2014-01-01

BACKGROUND In mice, refrigerator-stored red blood cells (RBCs) are cleared by extravascular hemolysis and induce cytokine production. To enhance understanding of this phenomenon, we sought to model it in vitro. STUDY DESIGN AND METHODS Ingestion of refrigerator-stored murine RBCs and subsequent cytokine production were studied using J774A.1 mouse macrophage cells and primary murine splenic macrophages. Wild-type and Ccl2-GFP-reporter mice were used for RBC clearance in vivo. RESULTS Although J774A.1 cells and primary macrophages preferentially ingested refrigerator-stored RBCs in vitro, as compared to freshly-isolated RBCs, neither produced increased cytokines following erythrophagocytosis. In contrast, phagocytosis of refrigerator-stored RBCs in vivo induced increases in circulating monocyte chemoattractant protein-1 (MCP-1) and keratinocyte chemoattractant (KC), and correspondingly increased mRNA levels in mouse spleen and liver. In the spleen, these were predominantly expressed by CD11b+ cells. Using Ccl2-GFP-reporter mice, the predominant splenic population responsible for MCP-1 mRNA production were tissue-resident macrophages (i.e., CD45+, CD11b+, F4/80+, Ly6c+, CD11clow cells). CONCLUSION J774A.1 cells and primary macrophages selectively ingested refrigerator-stored RBCs by phagocytosis. Although cytokine expression was not enhanced, this approach could be used to identify the relevant receptor-ligand combination(s). In contrast, cytokine levels increased following phagocytosis of refrigerator-stored RBCs in vivo. These were primarily cleared in the liver and spleen, which demonstrated increased MCP-1 and KC mRNA expression. Finally, in mouse spleen, tissue-resident macrophages were predominantly involved in MCP-1 mRNA production. The differences between cytokine production in vitro and in vivo are not yet well understood. PMID:25041478

Relaxation of ferromagnetic nanoparticles in macrophages: In vitro and in vivo studies

NASA Astrophysics Data System (ADS)

Möller, Winfried; Takenaka, Shinji; Buske, Norbert; Felten, Kathrin; Heyder, Joachim

2005-05-01

The relaxation characteristics of magnetic nanoparticles (CoFe 2O 4) were investigated in J774A.1 macrophages and after voluntary inhalation. In dry form 25% of the particles showed Néel relaxation. Relaxation in macrophages occurred within minutes and could be inhibited by fixation, showing Brownian relaxation and intracellular transport processes. Relaxation in the lung happened similarly, but was dependent on the time after deposition. The particles were cleared from the lung within 2 weeks.

Homocysteine elicits an M1 phenotype in murine macrophages through an EMMPRIN-mediated pathway.

PubMed

Winchester, Lee J; Veeranki, Sudhakar; Givvimani, Srikanth; Tyagi, Suresh C

2015-07-01

Hyperhomocysteinemia (HHcy) is associated with inflammatory diseases and is known to increase the production of reactive oxygen species (ROS), matrix metalloproteinase (MMP)-9, and inducible nitric oxide synthase, and to decrease endothelial nitric oxide production. However, the impact of HHcy on macrophage phenotype differentiation is not well-established. It has been documented that macrophages have 2 distinct phenotypes: the "classically activated/destructive" (M1), and the "alternatively activated/constructive" (M2) subtypes. We hypothesize that HHcy increases M1 macrophage differentiation through extracellular matrix metalloproteinase inducer (EMMPRIN), a known inducer of matrix metalloproteinases. murine J774A.1 and Raw 264.7 macrophages were treated with 100 and 500 μmol/L Hcy, respectively, for 24 h. Samples were analyzed using Western blotting and immunocytochemistry. Homocysteine treatment increased cluster of differentiation 40 (CD40; M1 marker) in J774A.1 and Raw 264.7 macrophages. MMP-9 was induced in both cell lines. EMMPRIN protein expression was also increased in both cell lines. Blocking EMMPRIN function by pre-treating cells with anti-EMMPRIN antibody, with or without Hcy, resulted in significantly lower expression of CD40 in both cell lines by comparison with the controls. A DCFDA assay demonstrated increased ROS production in both cell lines with Hcy treatment when compared with the controls. Our results suggest that HHcy results in an increase of the M1 macrophage phenotype. This effect seems to be at least partially mediated by EMMPRIN induction.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lehnert, B.E.; Oritz, J.B.; Steinkamp, J.A.

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 ofmore » 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.« less

Macrophages clear refrigerator storage-damaged red blood cells and subsequently secrete cytokines in vivo, but not in vitro, in a murine model.

PubMed

Wojczyk, Boguslaw S; Kim, Nina; Bandyopadhyay, Sheila; Francis, Richard O; Zimring, James C; Hod, Eldad A; Spitalnik, Steven L

2014-12-01

In mice, refrigerator-stored red blood cells (RBCs) are cleared by extravascular hemolysis and induce cytokine production. To enhance understanding of this phenomenon, we sought to model it in vitro. Ingestion of refrigerator-stored murine RBCs and subsequent cytokine production were studied using J774A.1 mouse macrophage cells and primary murine splenic macrophages. Wild-type and Ccl2-GFP reporter mice were used for RBC clearance in vivo. Although J774A.1 cells and primary macrophages preferentially ingested refrigerator-stored RBCs in vitro, compared to freshly isolated RBCs, neither produced increased cytokines after erythrophagocytosis. In contrast, phagocytosis of refrigerator-stored RBCs in vivo induced increases in circulating monocyte chemoattractant protein-1 (MCP-1) and keratinocyte chemoattractant (KC) and correspondingly increased mRNA levels in mouse spleen and liver. In the spleen, these were predominantly expressed by CD11b+ cells. Using Ccl2-GFP reporter mice, the predominant splenic population responsible for MCP-1 mRNA production was tissue-resident macrophages (i.e., CD45+, CD11b+, F4/80+, Ly6c+, and CD11c(low) cells). J774A.1 cells and primary macrophages selectively ingested refrigerator-stored RBCs by phagocytosis. Although cytokine expression was not enhanced, this approach could be used to identify the relevant receptor-ligand combination(s). In contrast, cytokine levels increased after phagocytosis of refrigerator-stored RBCs in vivo. These were primarily cleared in the liver and spleen, which demonstrated increased MCP-1 and KC mRNA expression. Finally, in mouse spleen, tissue-resident macrophages were predominantly involved in MCP-1 mRNA production. The differences between cytokine production in vitro and in vivo are not yet well understood. © 2014 AABB.

Hyphal growth of phagocytosed Fusarium oxysporum causes cell lysis and death of murine macrophages.

PubMed

Schäfer, Katja; Bain, Judith M; Di Pietro, Antonio; Gow, Neil A R; Erwig, Lars P

2014-01-01

Fusarium oxysporum is an important plant pathogen and an opportunistic pathogen of humans. Here we investigated phagocytosis of F. oxysporum by J774.1 murine cell line macrophages using live cell video microscopy. Macrophages avidly migrated towards F. oxysporum germlings and were rapidly engulfed after cell-cell contact was established. F. oxysporum germlings continued hyphal growth after engulfment by macrophages, leading to associated macrophage lysis and escape. Macrophage killing depended on the multiplicity of infection. After engulfment, F. oxysporum inhibited macrophages from completing mitosis, resulting in large daughter cells fused together by means of a F. oxysporum hypha. These results shed new light on the initial stages of Fusarium infection and the innate immune response of the mammalian host.

UPLC-ESI-QTOF-MS2 characterisation of Cola nitida resin fractions with inhibitory effects on NO and TNF-α released by LPS-activated J774 macrophage and on Trypanosoma cruzi and Leishmania amazonensis.

PubMed

Frankenberger, Larissa; Mora, Tamara D; de Siqueira, Carolina D; Filippin-Monteiro, Fabiola B; de Moraes, Milene H; Biavatti, Maique W; Steindel, Mario; Sandjo, Louis P

2018-05-29

The resin of Cola nitida is used in western Cameroon as incense for spiritual protection and during ritual ceremonies. This plant secretion has never been investigated although previous chemical and biological studies on other resins have drawn many attentions. The resin fractions which revealed inhibitory effect on nitric oxide (NO) and tumour necrosis factor alpha (TNF-α) released by lipopolysaccharide (LPS)-activated J774 macrophage as well as on intracellular forms of Leishmania amazonensis and Trypanosoma cruzi amastigote were chemically characterised. Moreover, their antiparasitic activities were compared to those of semi-synthetic triterpenes. The anti-inflammatory activity was evaluated by measuring the nitrite production and the TNF-α concentration in the supernatants of LPS-activated macrophages by antigen capture enzyme-linked immunosorbent assay. Moreover, the antiparasitic assay was performed by infecting the host cells (THP-1) in a ratio parasite/cell 10:1 (L. amazonensis) and 2:1 (T. cruzi) and then exposed to the samples. The resin was separated in vacuo by liquid chromatography because of its sticky behaviour and the chemical profiles of the obtained fractions (F1-F4) were established by dereplication based on UPLC-ESI-MS 2 data while semi-synthetic triterpenes were prepared from α-amyrin by oxidation reactions. Fractions F1-F4 inhibited NO and TNF-α almost similarly. However, only F1, F3 and F4 showed promising antiparasitic activities while F2 was moderately active against both parasites. Hence, F1-F4 were exclusively composed of pentacyclic triterpenes bearing oleanane and ursane skeletons. Semi-synthetic compounds revealed no to moderate antiparasitic activity compared to the fractions. Although it will be difficult to prove the interaction resin-spirit, interesting bioactivities were found in the resin fractions. Copyright © 2018 John Wiley & Sons, Ltd.

The Impairment of Macrophage-to-Feces Reverse Cholesterol Transport during Inflammation Does Not Depend on Serum Amyloid A

PubMed Central

de Beer, Maria C.; Wroblewski, Joanne M.; Noffsinger, Victoria P.; Meyer, Jason M.; van der Westhuyzen, Deneys R.

2013-01-01

Studies suggest that inflammation impairs reverse cholesterol transport (RCT). We investigated whether serum amyloid A (SAA) contributes to this impairment using an established macrophage-to-feces RCT model. Wild-type (WT) mice and mice deficient in SAA1.1 and SAA2.1 (SAAKO) were injected intraperitoneally with 3H-cholesterol-labeled J774 macrophages 4 hr after administration of LPS or buffered saline. 3H-cholesterol in plasma 4 hr after macrophage injection was significantly reduced in both WT and SAAKO mice injected with LPS, but this was not associated with a reduced capacity of serum from LPS-injected mice to promote macrophage cholesterol efflux in vitro. Hepatic accumulation of 3H-cholesterol was unaltered in either WT or SAAKO mice by LPS treatment. Radioactivity present in bile and feces of LPS-injected WT mice 24 hr after macrophage injection was reduced by 36% (P < 0.05) and 80% (P < 0.001), respectively. In contrast, in SAAKO mice, LPS did not significantly reduce macrophage-derived 3H-cholesterol in bile, and fecal excretion was reduced by only 45% (P < 0.05). Injection of cholesterol-loaded allogeneic J774 cells, but not syngeneic bone-marrow-derived macrophages, transiently induced SAA in C57BL/6 mice. Our study confirms reports that acute inflammation impairs steps in the RCT pathway and establishes that SAA plays only a minor role in this impairment. PMID:23431457

The Impairment of Macrophage-to-Feces Reverse Cholesterol Transport during Inflammation Does Not Depend on Serum Amyloid A.

PubMed

de Beer, Maria C; Wroblewski, Joanne M; Noffsinger, Victoria P; Ji, Ailing; Meyer, Jason M; van der Westhuyzen, Deneys R; de Beer, Frederick C; Webb, Nancy R

2013-01-01

Studies suggest that inflammation impairs reverse cholesterol transport (RCT). We investigated whether serum amyloid A (SAA) contributes to this impairment using an established macrophage-to-feces RCT model. Wild-type (WT) mice and mice deficient in SAA1.1 and SAA2.1 (SAAKO) were injected intraperitoneally with (3)H-cholesterol-labeled J774 macrophages 4 hr after administration of LPS or buffered saline. (3)H-cholesterol in plasma 4 hr after macrophage injection was significantly reduced in both WT and SAAKO mice injected with LPS, but this was not associated with a reduced capacity of serum from LPS-injected mice to promote macrophage cholesterol efflux in vitro. Hepatic accumulation of (3)H-cholesterol was unaltered in either WT or SAAKO mice by LPS treatment. Radioactivity present in bile and feces of LPS-injected WT mice 24 hr after macrophage injection was reduced by 36% (P < 0.05) and 80% (P < 0.001), respectively. In contrast, in SAAKO mice, LPS did not significantly reduce macrophage-derived (3)H-cholesterol in bile, and fecal excretion was reduced by only 45% (P < 0.05). Injection of cholesterol-loaded allogeneic J774 cells, but not syngeneic bone-marrow-derived macrophages, transiently induced SAA in C57BL/6 mice. Our study confirms reports that acute inflammation impairs steps in the RCT pathway and establishes that SAA plays only a minor role in this impairment.

Hyphal Growth of Phagocytosed Fusarium oxysporum Causes Cell Lysis and Death of Murine Macrophages

PubMed Central

Schäfer, Katja; Bain, Judith M.

2014-01-01

Fusarium oxysporum is an important plant pathogen and an opportunistic pathogen of humans. Here we investigated phagocytosis of F. oxysporum by J774.1 murine cell line macrophages using live cell video microscopy. Macrophages avidly migrated towards F. oxysporum germlings and were rapidly engulfed after cell-cell contact was established. F. oxysporum germlings continued hyphal growth after engulfment by macrophages, leading to associated macrophage lysis and escape. Macrophage killing depended on the multiplicity of infection. After engulfment, F. oxysporum inhibited macrophages from completing mitosis, resulting in large daughter cells fused together by means of a F. oxysporum hypha. These results shed new light on the initial stages of Fusarium infection and the innate immune response of the mammalian host. PMID:25025395

Effect of low extracellular pH on NF-κB activation in macrophages.

PubMed

Gerry, A B; Leake, D S

2014-04-01

Many diseases, including atherosclerosis, involve chronic inflammation. The master transcription factor for inflammation is NF-κB. Inflammatory sites have a low extracellular pH. Our objective was to demonstrate the effect of pH on NF-κB activation and cytokine secretion. Mouse J774 macrophages or human THP-1 or monocyte-derived macrophages were incubated at pH 7.0-7.4 and inflammatory cytokine secretion and NF-κB activity were measured. A pH of 7.0 greatly decreased pro-inflammatory cytokine secretion (TNF or IL-6) by J774 macrophages, but not THP-1 or human monocyte-derived macrophages. Upon stimulation of mouse macrophages, the levels of IκBα, which inhibits NF-κB, fell but low pH prevented its later increase, which normally restores the baseline activity of NF-κB, even though the levels of mRNA for IκBα were increased. pH 7.0 greatly increased and prolonged NF-κB binding to its consensus promoter sequence, especially the anti-inflammatory p50:p50 homodimers. Human p50 was overexpressed using adenovirus in THP-1 macrophages and monocyte-derived macrophages to see if it would confer pH sensitivity to NF-κB activity in human cells. Overexpression of p50 increased p50:p50 DNA-binding and in THP-1 macrophages inhibited considerably TNF and IL-6 secretion, but there was still no effect of pH on p50:p50 DNA binding or cytokine secretion. A modest decrease in pH can sometimes have marked effects on NF-κB activation and cytokine secretion and might be one reason to explain why mice normally develop less atherosclerosis than do humans. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Macrophage Biocompatibility of CoCr Wear Particles Produced under Polarization in Hyaluronic Acid Aqueous Solution

PubMed Central

Perez-Maceda, Blanca Teresa; López-Fernández, María Encarnación; Díaz, Iván; Kavanaugh, Aaron; Billi, Fabrizio; Escudero, María Lorenza; García-Alonso, María Cristina; Lozano, Rosa María

2018-01-01

Macrophages are the main cells involved in inflammatory processes and in the primary response to debris derived from wear of implanted CoCr alloys. The biocompatibility of wear particles from a high carbon CoCr alloy produced under polarization in hyaluronic acid (HA) aqueous solution was evaluated in J774A.1 mouse macrophages cultures. Polarization was applied to mimic the electrical interactions observed in living tissues. Wear tests were performed in a pin-on-disk tribometer integrating an electrochemical cell in phosphate buffer solution (PBS) and in PBS supplemented with 3 g/L HA, an average concentration that is generally found in synovial fluid, used as lubricant solution. Wear particles produced in 3 g/L HA solution showed a higher biocompatibility in J774A.1 macrophages in comparison to those elicited by particles obtained in PBS. A considerable enhancement in macrophages biocompatibility in the presence of 3 g/L of HA was further observed by the application of polarization at potentials having current densities typical of injured tissues suggesting that polarization produces an effect on the surface of the metallic material that leads to the production of wear particles that seem to be macrophage-biocompatible and less cytotoxic. The results showed the convenience of considering the influence of the electric interactions in the chemical composition of debris detached from metallic surfaces under wear corrosion to get a better understanding of the biological effects caused by the wear products. PMID:29738506

Artemisia asiatica Nakai Attenuates the Expression of Proinflammatory Mediators in Stimulated Macrophages Through Modulation of Nuclear Factor-κB and Mitogen-Activated Protein Kinase Pathways

PubMed Central

Kim, Eun-Kyung; Tang, Yujiao; Cha, Kwang-Suk; Choi, Heeri; Lee, Chun Bok; Yoon, Jin-Hwan; Kim, Sang Bae; Kim, Jong-Shik; Kim, Jong Moon; Han, Weon Cheol; Choi, Suck-Jun; Lee, Sangmin; Choi, Eun-Ju; Kim, Sang-Hyun

2015-01-01

Abstract The present study aimed to examine the anti-inflammatory effects and potential mechanism of action of Artemisia asiatica Nakai (A. asiatica Nakai) extract in activated murine macrophages. A. asiatica Nakai extract showed dose-dependent suppression of lipopolysaccharide (LPS)-induced nitric oxide, inducible nitric oxide synthase, and cyclooxygenase-2 activity. It also showed dose-dependent inhibition of nuclear factor-κB (NF-κB) translocation from the cytosol to the nucleus and as an inhibitor of NF-κB-alpha phosphorylation. The extract's inhibitory effects were found to be mediated through NF-κB inhibition and phosphorylation of extracellular signal-regulated kinase 1/2 and p38 in LPS-stimulated J774A.1 murine macrophages, suggesting a potential mechanism for the anti-inflammatory activity of A. asiatica Nakai. To our knowledge, this is the first report of the anti-inflammatory effects of A. asiatica Nakai on J774A.1 murine macrophages; these results may help develop functional foods possessing an anti-inflammatory activity. PMID:26061361

Atheroprotection through SYK inhibition fails in established disease when local macrophage proliferation dominates lesion progression.

PubMed

Lindau, Alexandra; Härdtner, Carmen; Hergeth, Sonja P; Blanz, Kelly Daryll; Dufner, Bianca; Hoppe, Natalie; Anto-Michel, Nathaly; Kornemann, Jan; Zou, Jiadai; Gerhardt, Louisa M S; Heidt, Timo; Willecke, Florian; Geis, Serjosha; Stachon, Peter; Wolf, Dennis; Libby, Peter; Swirski, Filip K; Robbins, Clinton S; McPheat, William; Hawley, Shaun; Braddock, Martin; Gilsbach, Ralf; Hein, Lutz; von zur Mühlen, Constantin; Bode, Christoph; Zirlik, Andreas; Hilgendorf, Ingo

2016-03-01

Macrophages in the arterial intima sustain chronic inflammation during atherogenesis. Under hypercholesterolemic conditions murine Ly6C(high) monocytes surge in the blood and spleen, infiltrate nascent atherosclerotic plaques, and differentiate into macrophages that proliferate locally as disease progresses. Spleen tyrosine kinase (SYK) may participate in downstream signaling of various receptors that mediate these processes. We tested the effect of the SYK inhibitor fostamatinib on hypercholesterolemia-associated myelopoiesis and plaque formation in Apoe(-/-) mice during early and established atherosclerosis. Mice consuming a high cholesterol diet supplemented with fostamatinib for 8 weeks developed less atherosclerosis. Histologic and flow cytometric analysis of aortic tissue showed that fostamatinib reduced the content of Ly6C(high) monocytes and macrophages. SYK inhibition limited Ly6C(high) monocytosis through interference with GM-CSF/IL-3 stimulated myelopoiesis, attenuated cell adhesion to the intimal surface, and blocked M-CSF stimulated monocyte to macrophage differentiation. In Apoe(-/-) mice with established atherosclerosis, however, fostamatinib treatment did not limit macrophage accumulation or lesion progression despite a significant reduction in blood monocyte counts, as lesional macrophages continued to proliferate. Thus, inhibition of hypercholesterolemia-associated monocytosis, monocyte infiltration, and differentiation by SYK antagonism attenuates early atherogenesis but not established disease when local macrophage proliferation dominates lesion progression.

Proliferating cellular nuclear antigen expression as a marker of perivascular macrophages in simian immunodeficiency virus encephalitis.

PubMed

Williams, Kenneth; Schwartz, Annette; Corey, Sarah; Orandle, Marlene; Kennedy, William; Thompson, Brendon; Alvarez, Xavier; Brown, Charlie; Gartner, Suzanne; Lackner, Andrew

2002-08-01

Brain perivascular macrophages are a major target of simian immunodeficiency virus (SIV) infection in rhesus macaques and HIV infection in humans. Perivascular macrophages are distinct from parenchymal microglia in their location, morphology, expression of myeloid markers, and turnover in the CNS. In contrast to parenchymal microglia, perivascular macrophages are continuously repopulated by blood monocytes, which undergo maturation to macrophages on entering the central nervous system (CNS). We studied differences in monocyte/macrophages in vivo that might account for preferential infection of perivascular macrophages by SIV. In situ hybridization for SIV and proliferating cellular nuclear antigen (PCNA) immunohistochemistry demonstrated that SIV-infected and PCNA-positive cells were predominantly found in perivascular cuffs of viremic animals and in histopathological lesions that characterize SIV encephalitis (SIVE) in animals with AIDS. Multilabel techniques including double-label immunohistochemistry and combined in situ hybridization and immunofluorescence confocal microscopy revealed numerous infected perivascular macrophages that were PCNA-positive. Outside the CNS, SIV-infected, PCNA-expressing macrophage subpopulations were found in the small intestine and lung of animals with AIDS. While PCNA is used as a marker of cell proliferation it is also strongly expressed in non-dividing cells undergoing DNA synthesis and repair. Therefore, more specific markers for cell proliferation including Ki-67, topoisomerase IIalpha, and bromodeoxyuridine (BrdU) incorporation were used which indicated that PCNA-positive cells within SIVE lesions were not proliferating. These observations are consistent with perivascular macrophages as terminally differentiated, non-dividing cells and underscores biological differences that could potentially define mechanisms of preferential, productive infection of perivascular macrophages in the rhesus macaque model of neuroAIDS. These

SSN 774 Virginia Class Submarine (SSN 774)

DTIC Science & Technology

2015-12-01

Selected Acquisition Report ( SAR ) RCS: DD-A&T(Q&A)823-516 SSN 774 Virginia Class Submarine (SSN 774) As of FY 2017 President’s Budget Defense...Acquisition Management Information Retrieval (DAMIR) March 8, 2016 11:22:44 UNCLASSIFIED SSN 774 December 2015 SAR March 8, 2016 11:22:44 UNCLASSIFIED 2...Document OSD - Office of the Secretary of Defense O&S - Operating and Support PAUC - Program Acquisition Unit Cost SSN 774 December 2015 SAR March 8

Pomegranate Juice Polyphenols Induce Macrophage Death via Apoptosis as Opposed to Necrosis Induced by Free Radical Generation: A Central Role for Oxidative Stress.

PubMed

Rom, Oren; Volkova, Nina; Nandi, Sukhendu; Jelinek, Raz; Aviram, Michael

2016-08-01

At high concentrations, polyphenols induce cell death, and the polyphenols-rich pomegranate juice (PJ), known for its antioxidative/antiatherogenic properties, can possibly affect cell death, including macrophage death involved in atherogenesis. In the present study, apoptotic/necrotic macrophage death was analyzed in J774A.1 macrophages and in peritoneal macrophages isolated from atherosclerotic apoE-/- mice treated with PJ. The effects of PJ were compared with those of the free radical generator 2, 2'-azobis (2-amidinopropane) dihydrochloride (AAPH). Both PJ and AAPH significantly increased J774A.1 macrophage death; however, flow cytometric and microscopic analyses using annexin V/propidium iodide revealed that PJ increased the early apoptosis of the macrophage dose dependently (up to 2.5-fold, P < 0.01), whereas AAPH caused dose-dependent increases in late apoptosis/necrosis (up to 12-fold, P < 0.001). Unlike PJ, AAPH-induced macrophage death was associated with increased intracellular oxidative stress (up to 7-fold, P < 0.001) and with lipid stress demonstrated by triglyceride accumulation (up to 3-fold, P < 0.01) and greater chromatic vesicle response to culture medium (up to 5-fold, P < 0.001). Accordingly, recombinant paraoxonase 1, which hydrolyzes oxidized lipids, attenuated macrophage death induced by AAPH, but not by PJ. Similar apoptotic and oxidative effects were found in macrophages from apoE-/- mice treated with PJ or AAPH. As macrophage apoptotic/necrotic death has considerable impact on atherosclerosis progression, these findings may provide novel mechanisms for the antiatherogenicity of PJ.

Garlic compounds modulate macrophage and T-lymphocyte functions.

PubMed

Lau, B H; Yamasaki, T; Gridley, D S

1991-06-01

Organosulfur compounds of garlic have been shown to inhibit growth of animal tumors and to modulate the activity of diverse chemical carcinogens. There is also evidence that garlic may modulate antitumor immunity. In this study, we determined the effects of an aqueous garlic extract and a protein fraction isolated from the extract on the chemiluminescent oxidative burst of the murine J774 macrophage cell line and thioglycollate-elicited peritoneal macrophages obtained from BALB/c mice. T-lymphocyte activity was determined using mouse splenocytes incubated with phytohemagglutinin, labeled with [3H]-thymidine and assayed for lymphoproliferation. Significant dose-related augmentation of oxidative burst was observed with garlic extract and the protein fraction. The protein fraction also enhanced the T-lymphocyte blastogenesis. The data suggest that garlic compounds may serve as biological response modifiers by augmenting macrophage and T-lymphocyte functions.

Mouse macrophages primed with alendronate down-regulate monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1alpha (MIP-1alpha) production in response to Toll-like receptor (TLR) 2 and TLR4 agonist via Smad3 activation.

PubMed

Masuda, Takahiro; Deng, Xue; Tamai, Riyoko

2009-08-01

Alendronate is one of the nitrogen-containing bisphosphonates (NBPs) used as anti-bone resorptive drugs. However, NBPs have inflammatory side effects including osteomyelitis and osteonecrosis of the jaw. In the present study, we examined the effects of alendronate on chemokine production by the macrophage-like cell line, J774.1, when incubated with Pam(3)CSK(4) (a Toll-like receptor (TLR) 2 agonist) and Lipid A (a TLR4 agonist). Pretreatment of J774.1 cells with alendronate decreased the production of TLR ligand-induced monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1alpha (MIP-1alpha) but did not influence nuclear factor-kappaB (NF-kappaB) activation. While this agent induced caspase-8 activation, a caspase-8 inhibitor did not affect the decrease in MCP-1 production by alendronate and TLR ligands. Thus, the alendronate-mediated decrease in chemokine production was independent of NF-kappaB and caspase-8 activation. Although transforming growth factor-beta1 (TGF-beta1) is known to inhibit chemokine production by various cell types via Smad3 activation, pretreatment with alendronate did not increase TGF-beta1 production by J774.1 cells incubated in the presence or absence of TLR ligands. However, alendronate directly activated Smad3. These results suggest that by down-regulating MCP-1 and MIP-1alpha production via Smad3, long-term use of alendronate might inhibit normal activation and migration of osteoclasts and cause osteonecrosis.

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

PubMed

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

2018-02-05

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

Homocysteine enhances MMP-9 production in murine macrophages via ERK and Akt signaling pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Seung Jin; Lee, Yi Sle; Seo, Kyo Won

2012-04-01

Homocysteine (Hcy) at elevated levels is an independent risk factor of cardiovascular diseases, including atherosclerosis. In the present study, we investigated the effect of Hcy on the production of matrix metalloproteinases (MMP) in murine macrophages. Among the MMP known to regulate the activities of collagenase and gelatinase, Hcy exclusively increased the gelatinolytic activity of MMP-9 in J774A.1 cells as well as in mouse peritoneal macrophages. Furthermore, this activity was found to be correlated with Western blot findings in J774A.1 cells, which showed that MMP-9 expression was concentration- and time-dependently increased by Hcy. Inhibition of the ERK and Akt pathways ledmore » to a significant decrease in Hcy-induced MMP-9 expression, and combined treatment with inhibitors of the ERK and Akt pathways showed an additive effects. Activity assays for ERK and Akt showed that Hcy increased the phosphorylation of both, but these phosphorylation were not affected by inhibitors of the Akt and ERK pathways. In line with these findings, the molecular inhibition of ERK and Akt using siRNA did not affect the Hcy-induced phosphorylation of Akt and ERK, respectively. Taken together, these findings suggest that Hcy enhances MMP-9 production in murine macrophages by separately activating the ERK and Akt signaling pathways. -- Highlights: ► Homocysteine (Hcy) induced MMP-9 production in murine macrophages. ► Hcy induced MMP-9 production through ERK and Akt signaling pathways. ► ERK and Akt signaling pathways were activated by Hcy in murine macrophages. ► ERK and Akt pathways were additively act on Hcy-induced MMP-9 production. ► Hcy enhances MMP-9 production in macrophages via activation of ERK and Akt signaling pathways in an independent manner.« less

Plasminogen promotes macrophage phagocytosis in mice

PubMed Central

Ganapathy, Swetha; Settle, Megan; Plow, Edward F.

2014-01-01

The phagocytic function of macrophages plays a pivotal role in eliminating apoptotic cells and invading pathogens. Evidence implicating plasminogen (Plg), the zymogen of plasmin, in phagocytosis is extremely limited with the most recent in vitro study showing that plasmin acts on prey cells rather than on macrophages. Here, we use apoptotic thymocytes and immunoglobulin opsonized bodies to show that Plg exerts a profound effect on macrophage-mediated phagocytosis in vitro and in vivo. Plg enhanced the uptake of these prey by J774A.1 macrophage-like cells by 3.5- to fivefold Plg receptors and plasmin proteolytic activity were required for phagocytosis of both preys. Compared with Plg+/+ mice, Plg−/− mice exhibited a 60% delay in clearance of apoptotic thymocytes by spleen and an 85% reduction in uptake by peritoneal macrophages. Phagocytosis of antibody-mediated erythrocyte clearance by liver Kupffer cells was reduced by 90% in Plg−/− mice compared with Plg+/+ mice. A gene array of splenic and hepatic tissues from Plg−/− and Plg+/+ mice showed downregulation of numerous genes in Plg−/− mice involved in phagocytosis and regulation of phagocytic gene expression was confirmed in macrophage-like cells. Thus, Plg may play an important role in innate immunity by changing expression of genes that contribute to phagocytosis. PMID:24876560

Truncated EphA2 likely potentiates cell adhesion via integrins as well as infiltration and/or lodgment of a monocyte/macrophage cell line in the red pulp and marginal zone of the mouse spleen, where ephrin-A1 is prominently expressed in the vasculature.

PubMed

Konda, Naoko; Saeki, Noritaka; Nishino, Shingo; Ogawa, Kazushige

2017-03-01

We previously established a J774.1 monocyte/macrophage subline expressing a truncated EphA2 construct lacking the kinase domain. We demonstrated that following ephrin-A1 stimulation, endogenous EphA2 promotes cell adhesion through interaction with integrins and integrin ligands such as ICAM1 and that truncated EphA2 potentiates the adhesion and becomes associated with the integrin/integrin ligand complex. Based on these findings, we hypothesized that the EphA/ephrin-A system, particularly EphA2/ephrin-A1, regulates transendothelial migration/tissue infiltration of monocytes/macrophages, because ephrin-A1 is widely recognized to be upregulated in inflammatory vasculatures. To evaluate whether this hypothesis is applicable in the spleen, we screened for EphA2/ephrin-A1 expression and reexamined the cellular properties of the J774.1 subline. We found that ephrin-A1 was expressed in the vasculature of the marginal zone and the red pulp and that its expression was upregulated in response to phagocyte depletion; further, CD115, F4/80, and CXCR4 were expressed in J774.1 cells, which serve as a usable substitute for monocytes/macrophages. Moreover, following ephrin-A1 stimulation, truncated EphA2 did not detectably interfere with the phosphorylation of endogenous EphA2, and it potentiated cell adhesion possibly through modulation of integrin avidity. Accordingly, by intravenously injecting mice with equal numbers of J774.1 and the subline cells labeled with distinct fluorochromes, we determined that truncated EphA2 markedly potentiated preferential cell infiltration into the red pulp and the marginal zone. Thus, modulation of EphA2 signaling might contribute to effective transplantation of tissue-specific resident macrophages and/or monocytes.

Staphylococcus haemolyticus strains target mitochondria and induce caspase-dependent apoptosis of macrophages.

PubMed

Krzymińska, Sylwia; Szczuka, Ewa; Kaznowski, Adam

2012-11-01

The aim of this study was to investigate the interaction of Staphylococcus haemolyticus strains with a macrophage cell line. Infection with the strains resulted in macrophage injury. All strains exhibited cytotoxic effects towards J774 cells. Moreover, the bacteria triggered apoptosis of the cells. The lowest apoptotic index did not exceed 21 %, whereas the highest reached 70 % at 24 h and 85 % at 48 h after infection. Incubation with the bacteria caused loss of mitochondrial membrane potential (ΔΨm) in macrophages. The pro-apoptotic activity of the strains was blocked by a pan-caspase inhibitor z-VAD-fmk, indicating the involvement of caspases in the bacteria-mediated cell death. We observed that the induction of macrophage apoptosis could constitute an important mechanism of pathogenesis by which S. haemolyticus strains evade host immune defences and cause disease.

Inactivation of p27kip1 Promoted Nonspecific Inflammation by Enhancing Macrophage Proliferation in Islet Transplantation.

PubMed

Li, Yang; Ding, Xiaoming; Fan, Ping; Guo, Jian; Tian, Xiaohui; Feng, Xinshun; Zheng, Jin; Tian, Puxun; Ding, Chenguang; Xue, Wujun

2016-11-01

Islet transplantation suffers from low efficiency caused by nonspecific inflammation-induced graft loss after transplantation. This study reports increased islet loss and enhanced inflammatory response in p27-deficient mice (p27-/-) and proposes a possible mechanism. Compared with wild type, p27-/- mice showed more severe functional injury of islet, with increased serum levels of inflammatory cytokines IL-1 and TNF-α, inducing macrophage proliferation. Furthermore, the increased number, proapoptotic proteins, and nuclear factor-kappa b (NF-κB) phosphorylation status of the infiltrating macrophages were accompanied by increased TNF-α mRNA level of islet graft site in p27-/- mice. Moreover, in vitro, we found that macrophages were still activated and cocultured with islet and promoted islet loss even blocking the direct effect of TNF-α on islets. Malondialdehyde (MDA, an end product of lipid peroxidation) in islet and media were increased after cocultured with macrophages. p27 deficiency also increased macrophage proliferation and islet injury. Therefore, p27 inactivation promotes injury islet graft loss via the elevation of proliferation and inflammatory cytokines secretion in infiltrating macrophages which induced nonspecific inflammation independent of TNF-α/nuclear factor-kappa b pathway. This potentially represents a promising therapeutic target in improving islet graft survival.

Effect of low-level laser therapy on the modulation of the mitochondrial activity of macrophages

PubMed Central

Souza, Nadhia H. C.; Ferrari, Raquel A. M.; Silva, Daniela F. T.; Nunes, Fabio D.; Bussadori, Sandra K.; Fernandes, Kristianne P. S.

2014-01-01

BACKGROUND: Macrophages play a major role among the inflammatory cells that invade muscle tissue following an injury. Low-level laser therapy (LLLT) has long been used in clinical practice to accelerate the muscle repair process. However, little is known regarding its effect on macrophages. OBJECTIVE: This study evaluated the effect of LLLT on the mitochondrial activity (MA) of macrophages. METHOD: J774 macrophages were treated with lipopolysaccharide (LPS) and interferon - gamma (IFN-γ) (activation) for 24 h to simulate an inflammatory process, then irradiated with LLLT using two sets of parameters (780 nm; 70 mW; 3 J/cm2 and 660 nm; 15 mW; 7.5 J/cm2). Non-activated/non-irradiated cells composed the control group. MA was evaluated by the cell mitochondrial activity (MTT) assay (after 1, 3 and 5 days) in three independent experiments. The data were analyzed statistically. RESULTS: After 1 day of culture, activated and 780 nm irradiated macrophages showed lower MA than activated macrophages, but activated and 660 nm irradiated macrophages showed MA similar to activated cells. After 3 days, activated and irradiated (660 nm and 780 nm) macrophages showed greater MA than activated macrophages, and after 5 days, the activated and irradiated (660 nm and 780 nm) macrophages showed similar MA to the activated macrophages. CONCLUSIONS: These results show that 660 nm and 780 nm LLLT can modulate the cellular activation status of macrophages in inflammation, highlighting the importance of this resource and of the correct determination of its parameters in the repair process of skeletal muscle. PMID:25076002

Manipulation of NF-KappaBetta Activity in the Macrophage Lineage as a Novel Therapeutic Approach

DTIC Science & Technology

2007-05-01

Sadikot, J. W. Christman, and T. S. Blackwell. 2003. Bioluminescent detection of endotoxin effects on HIV-1 LTR-driven transcription in vivo. J...differences in proliferation rates, expression of downstream gene expression and effects mediated by altered macrophages on associated epithelial...kappaB activity within macrophages has significant effects on mammary ductal development. 15. SUBJECT TERMS NF-kappaB, macrophages, mammary ductal

Boron Induces Lymphocyte Proliferation and Modulates the Priming Effects of Lipopolysaccharide on Macrophages.

PubMed

Routray, Indusmita; Ali, Shakir

2016-01-01

Chemical mediators of inflammation (CMI) are important in host defense against infection. The reduced capacity of host to induce the secretion of these mediators following infection is one of the factors in host susceptibility to infection. Boron, which has been suggested for its role in infection, is reported in this study to increase lymphocyte proliferation and the secretion of CMI by the lipopolysaccharide (LPS)-stimulated peritoneal macrophages in BALB/c mice. Boron was administered to mice orally as borax at different doses for 10 consecutive days, followed by the stimulation of animals with ovalbumin and isolation of splenocytes for proliferation assay. The lymphocyte subsets were determined by flow cytometry in spleen cell suspension. The mediators of inflammation, TNF-α, IL-6, IL-1β and nitric oxide (NO), were measured in culture supernatant of LPS-primed macrophages isolated from borax treated mice. TNF and ILs were measured by ELISA. NO was determined by Griess test. The expression of inducible nitric oxide synthase (iNOS) in macrophages was studied by confocal microscopy. Results showed a significant increase in T and B cell populations, as indicated by an increase in CD4 and CD19, but not CD8, cells. Boron further stimulated the secretion of TNF-α, IL-6, IL-1β, NO and the expression of iNOS by the LPS-primed macrophages. The effect was dose dependent and most significant at a dose level of 4.6 mg/kg b. wt. Taken together, the study concludes that boron at physiological concentration induces lymphocyte proliferation and increases the synthesis and secretion of pro-inflammatory mediators by the LPS-primed macrophages, more specifically the M1 macrophages, possibly acting through Toll-like receptor. The study implicates boron as a regulator of the immune and inflammatory reactions and macrophage polarization, thus playing an important role in augmenting host defense against infection, with possible role in cancer and other diseases.

Boron Induces Lymphocyte Proliferation and Modulates the Priming Effects of Lipopolysaccharide on Macrophages

PubMed Central

Routray, Indusmita; Ali, Shakir

2016-01-01

Chemical mediators of inflammation (CMI) are important in host defense against infection. The reduced capacity of host to induce the secretion of these mediators following infection is one of the factors in host susceptibility to infection. Boron, which has been suggested for its role in infection, is reported in this study to increase lymphocyte proliferation and the secretion of CMI by the lipopolysaccharide (LPS)-stimulated peritoneal macrophages in BALB/c mice. Boron was administered to mice orally as borax at different doses for 10 consecutive days, followed by the stimulation of animals with ovalbumin and isolation of splenocytes for proliferation assay. The lymphocyte subsets were determined by flow cytometry in spleen cell suspension. The mediators of inflammation, TNF-α, IL-6, IL-1β and nitric oxide (NO), were measured in culture supernatant of LPS-primed macrophages isolated from borax treated mice. TNF and ILs were measured by ELISA. NO was determined by Griess test. The expression of inducible nitric oxide synthase (iNOS) in macrophages was studied by confocal microscopy. Results showed a significant increase in T and B cell populations, as indicated by an increase in CD4 and CD19, but not CD8, cells. Boron further stimulated the secretion of TNF-α, IL-6, IL-1β, NO and the expression of iNOS by the LPS-primed macrophages. The effect was dose dependent and most significant at a dose level of 4.6 mg/kg b. wt. Taken together, the study concludes that boron at physiological concentration induces lymphocyte proliferation and increases the synthesis and secretion of pro-inflammatory mediators by the LPS-primed macrophages, more specifically the M1 macrophages, possibly acting through Toll-like receptor. The study implicates boron as a regulator of the immune and inflammatory reactions and macrophage polarization, thus playing an important role in augmenting host defense against infection, with possible role in cancer and other diseases. PMID:26934748

MIF-driven activation of macrophages induces killing of intracellular Trypanosoma cruzi dependent on endogenous production of tumor necrosis factor, nitric oxide and reactive oxygen species.

PubMed

Cutrullis, Romina A; Petray, Patricia B; Corral, Ricardo S

2017-02-01

The proinflammatory cytokine macrophage migration inhibitory factor (MIF) is a key player in innate immunity. MIF has been considered critical for controlling acute infection by the protozoan Trypanosoma cruzi, but the underlying mechanisms are poorly understood. Our study aimed to analyze whether MIF could favor microbicidal activity of the macrophage, a site where T. cruzi grows and the initial effector cell against this parasite. Using murine macrophages infected in vitro, we examined the effect of MIF on their parasiticidal ability and attempted to identify inflammatory agents involved in MIF-induced protection. Our findings show that MIF is readily secreted from peritoneal macrophages upon T. cruzi infection. MIF activates both primary and J774 phagocytes boosting the endogenous production of tumor necrosis factor-alpha via mitogen-activated protein kinase p38 signaling, as well as the release of nitric oxide and reactive oxygen species, leading to enhanced pathogen elimination. MIF can also potentiate the effect of interferon-gamma on T. cruzi killing by J774 and mouse peritoneal macrophages, rendering these cells more competent in reducing intracellular parasite burden. The present results unveil a novel innate immune pathway that contributes to host defense and broaden our understanding of the regulation of inflammatory mediators implicated in early parasite containment that is decisive for resistance to T. cruzi infection. Copyright © 2016 Elsevier GmbH. All rights reserved.

Alterations of zinc homeostasis in response to Cryptococcus neoformans in a murine macrophage cell line.

PubMed

Dos Santos, Francine Melise; Piffer, Alícia Corbellini; Schneider, Rafael de Oliveira; Ribeiro, Nicole Sartori; Garcia, Ane Wichine Acosta; Schrank, Augusto; Kmetzsch, Lívia; Vainstein, Marilene Henning; Staats, Charley Christian

2017-05-01

To evaluate alterations of zinc homeostasis in macrophages exposed to Cryptococcus neoformans. Materials & methods: Using a fluorescent zinc probe-based flow cytometry and atomic absorption spectrometry, zinc levels were evaluated in J774.A1 cell lines exposed to C. neoformans H99 cells. The transcription profile of macrophage zinc related homeostasis genes - metallothioneins and zinc transporters (ZnTs) of the SLC30 and SLC39 (Zrt-Irt-protein) families - was analyzed by quantitative PCR. Macrophage intracellular labile zinc levels decreased following exposure to C. neoformans. A significant decrease in transcription levels was detected in specific ZnTs from both the Zrt-Irt-protein and ZnT families, especially 24 h after infection. These findings suggest that macrophages may exhibit zinc depletion in response to C. neoformans infection.

Activation of macrophages and interference with CD4+ T-cell stimulation by Mycobacterium avium subspecies paratuberculosis and Mycobacterium avium subspecies avium

PubMed Central

Lage, Susanne Zur; Goethe, Ralph; Darji, Ayub; Valentin-Weigand, Peter; Weiss, Siegfried

2003-01-01

Mycobacterium avium subspecies paratuberculosis (M. ptb) and M. avium subspecies avium (M. avium) are closely related but exhibit significant differences in their interaction with the host immune system. The macrophage line, J774, was infected with M. ptb and M. avium and analysed for cytokine production and stimulatory capacity towards antigen-specific CD4+ T cells. Under all conditions J774 cells were activated to produce proinflammatory cytokines. No influence on the expression of major histocompatibility complex (MHC) class II, intracellular adhesion molecule-1 (ICAM-1), B7.1, B7.2 or CD40 was found. However, the antigen-specific stimulatory capacity of J774 cells for a CD4+ T-cell line was significantly inhibited after infection with M. ptb, but not with M. avium. When a T-cell hybridoma expressing a T-cell receptor identical to that of the T-cell line was used, this inhibition was not observed, suggesting that costimulation which is essential for the CD4+ T-cell line is influenced by the pathogenic bacterium M. ptb. PMID:12519304

Biotin deficiency up-regulates TNF-alpha production in murine macrophages.

PubMed

Kuroishi, Toshinobu; Endo, Yasuo; Muramoto, Koji; Sugawara, Shunji

2008-04-01

Biotin, a water-soluble vitamin of the B complex, functions as a cofactor of carboxylases that catalyze an indispensable cellular metabolism. Although significant decreases in serum biotin levels have been reported in patients with chronic inflammatory diseases, the biological roles of biotin in inflammatory responses are unclear. In this study, we investigated the effects of biotin deficiency on TNF-alpha production. Mice were fed a basal diet or a biotin-deficient diet for 8 weeks. Serum biotin levels were significantly lower in biotin-deficient mice than biotin-sufficient mice. After i.v. administration of LPS, serum TNF-alpha levels were significantly higher in biotin-deficient mice than biotin-sufficient mice. A murine macrophage-like cell line, J774.1, was cultured in a biotin-sufficient or -deficient medium for 4 weeks. Cell proliferation and biotinylation of intracellular proteins were decreased significantly in biotin-deficient cells compared with biotin-sufficient cells. Significantly higher production and mRNA expression of TNF-alpha were detected in biotin-deficient J774.1 cells than biotin-sufficient cells in response to LPS and even without LPS stimulation. Intracellular TNF-alpha expression was inhibited by actinomycin D, indicating that biotin deficiency up-regulates TNF-alpha production at the transcriptional level. However, the expression levels of TNF receptors, CD14, and TLR4/myeloid differentiation protein 2 complex were similar between biotin-sufficient and -deficient cells. No differences were detected in the activities of the NF-kappaB family or AP-1. The TNF-alpha induction by biotin deficiency was down-regulated by biotin supplementation in vitro and in vivo. These results indicate that biotin deficiency may up-regulate TNF-alpha production or that biotin excess down-regulates TNF-alpha production, suggesting that biotin status may influence inflammatory diseases.

A study to evaluate the effect of nootropic drug-piracetam on DNA damage in leukocytes and macrophages.

PubMed

Singh, Sarika; Goswami, Poonam; Swarnkar, Supriya; Singh, Sheelendra Pratap; Wahajuddin; Nath, Chandishwar; Sharma, Sharad

2011-11-27

Piracetam is a nootropic drug that protects neurons in neuropathological and age-related diseases and the activation and modulation of peripheral blood cells in patients with neuropathological conditions is well known. Therefore, in the present study, in vivo, ex vivo, and in vitro tests were conducted to investigate the effect of piracetam on leukocytes and macrophages. Lipopolysaccharide (LPS) causes oxidative DNA damage; thus, in the present study, LPS was used as a tool to induce DNA damage. In vivo experiments were conducted on Sprague Dawley rats, and piracetam (600mg/kg, oral) was provided for five consecutive days. On the fifth day, a single injection of LPS (10mg/kg, i.p.) was administered. Three hours after LPS injection, blood leukocytes and peritoneal macrophages were collected and processed, and a variety of different assays were conducted. Ex vivo treatments were performed on isolated rat blood leukocytes, and in vitro experiments were conducted on rat macrophage cell line J774A.1. Cell viability and the level of reactive oxygen species (ROS), mitochondrial membrane potential (MMP) and DNA damage were estimated in untreated (control) and piracetam-, LPS- and LPS+piracetam-treated leukocytes and macrophages. In vivo experiments revealed that rats pretreated with piracetam were significantly protected against LPS-induced increases in ROS levels and DNA damage. Ex vivo isolated leukocytes and J774A.1 cells treated with LPS exhibited augmented ROS levels and DNA damage, which were attenuated with piracetam treatment. Thus, the present study revealed the salutary effect of piracetam against LPS-induced oxidative stress and DNA damage in leukocytes and macrophages. Copyright © 2011 Elsevier B.V. All rights reserved.

Impact of Glutathione Peroxidase-1 Deficiency on Macrophage Foam Cell Formation and Proliferation: Implications for Atherogenesis

PubMed Central

Degreif, Adriana; Rossmann, Heidi; Canisius, Antje; Lackner, Karl J.

2013-01-01

Clinical and experimental evidence suggests a protective role for the antioxidant enzyme glutathione peroxidase-1 (GPx-1) in the atherogenic process. GPx-1 deficiency accelerates atherosclerosis and increases lesion cellularity in ApoE−/− mice. However, the distribution of GPx-1 within the atherosclerotic lesion as well as the mechanisms leading to increased macrophage numbers in lesions is still unknown. Accordingly, the aims of the present study were (1) to analyze which cells express GPx-1 within atherosclerotic lesions and (2) to determine whether a lack of GPx-1 affects macrophage foam cell formation and cellular proliferation. Both in situ-hybridization and immunohistochemistry of lesions of the aortic sinus of ApoE−/− mice after 12 weeks on a Western type diet revealed that both macrophages and – even though to a less extent – smooth muscle cells contribute to GPx-1 expression within atherosclerotic lesions. In isolated mouse peritoneal macrophages differentiated for 3 days with macrophage-colony-stimulating factor (MCSF), GPx-1 deficiency increased oxidized low density-lipoprotein (oxLDL) induced foam cell formation and led to increased proliferative activity of peritoneal macrophages. The MCSF- and oxLDL-induced proliferation of peritoneal macrophages from GPx-1−/−ApoE−/− mice was mediated by the p44/42 MAPK (p44/42 mitogen-activated protein kinase), namely ERK1/2 (extracellular-signal regulated kinase 1/2), signaling pathway as demonstrated by ERK1/2 signaling pathways inhibitors, Western blots on cell lysates with primary antibodies against total and phosphorylated ERK1/2, MEK1/2 (mitogen-activated protein kinase kinase 1/2), p90RSK (p90 ribosomal s6 kinase), p38 MAPK and SAPK/JNK (stress-activated protein kinase/c-Jun N-terminal kinase), and immunohistochemistry of mice atherosclerotic lesions with antibodies against phosphorylated ERK1/2, MEK1/2 and p90RSK. Representative effects of GPx-1 deficiency on both macrophage proliferation

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

PubMed

Park, Sung Bum; Park, Ji Seon; Jung, Won Hoon; Kim, Hee Youn; Kwak, Hyun Jung; Ahn, Jin Hee; Choi, Kyoung-Jin; Na, Yoon-Ju; Choi, Sunhwa; Dal Rhee, Sang; Kim, Ki Young

2016-08-01

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts inactive cortisone to the active cortisol. 11β-HSD1 may be involved in the resolution of inflammation. In the present study, we investigate the anti-inflammatory effects of 2-(3-benzoyl)-4-hydroxy-1,1-dioxo-2H-1,2-benzothiazine-2-yl-1-phenylethanone (KR-66344), a selective 11β-HSD1 inhibitor, in lipopolysaccharide (LPS)-activated C57BL/6J mice and macrophages. LPS increased 11β-HSD1 activity and expression in macrophages, which was inhibited by KR-66344. In addition, KR-66344 increased survival rate in LPS treated C57BL/6J mice. HO-1 mRNA expression level was increased by KR-66344, and this effect was reversed by the HO competitive inhibitor, ZnPP, in macrophages. Moreover, ZnPP reversed the suppression of ROS formation and cell death induced by KR-66344. ZnPP also suppressed animal survival rate in LPS plus KR-66344 treated C57BL/6J mice. In the spleen of LPS-treated mice, KR-66344 prevented cell death via suppression of inflammation, followed by inhibition of ROS, iNOS and COX-2 expression. Furthermore, LPS increased NFκB-p65 and MAPK phosphorylation, and these effects were abolished by pretreatment with KR-66344. Taken together, KR-66344 protects against LPS-induced animal death and spleen injury by inhibition of inflammation via induction of HO-1 and inhibition of 11β-HSD1 activity. Thus, we concluded that the selective 11β-HSD1 inhibitor may provide a novel strategy in the prevention/treatment of inflammatory disorders in patients. Copyright © 2016 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

Low-dose Norfloxacin-treated leptospires induce less IL-1β release in J774A.1 cells following discrepant leptospiral gene expression.

PubMed

Cao, Yongguo; Xie, Xufeng; Zhang, Wenlong; Wu, Dianjun; Tu, Changchun

2018-06-01

Currently, accumulating evidence is challenging subtherapeutic therapy. Low-dose Norfloxacin (Nor) has been reported to suppress the immune response and worsen leptospirosis. In this study, we investigated the influence of low-dose Nor (0.03 μg/ml, 0.06 μg/ml, 0.125 μg/ml) on leptospiral gene expression and analyzed the immunomodulatory effects of low-dose Nor-treated leptospires in J774A.1 cells. To study the expression profiles of low-dose Nor-treated leptospires, we chose LipL71/LipL21 as reference genes determined by the geNorm applet in this experiment. The results showed that low-dose Nor up-regulated the expression of FlaB and inhibited the expression of 16S rRNA, LipL32, LipL41, Loa22, KdpA, and KdpB compared with the untreated leptospires. These results indicated that low-dose Nor could regulate leptospiral gene expression. Using RT-PCR, the gene expression of IL-1β and TNF-α in J774A.1 cells was detected. Nor-treated leptospires induced higher expression levels of both IL-1β and TNF-α. However, when analyzed by ELISA, the release of mature IL-1β was reduced compared with that observed in cells induced with no Nor-treated leptospires, although the TNF-α protein level showed no significant change. Our study indicated that the gene expression of leptospires could be modulated by low-dose Nor, which induced less IL-1β release in J774A.1 cells. Copyright © 2018 Elsevier Ltd. All rights reserved.

In situ proliferation contributes to accumulation of tumor-associated macrophages in spontaneous mammary tumors.

PubMed

Tymoszuk, Piotr; Evens, Hanneke; Marzola, Vanessa; Wachowicz, Katarzyna; Wasmer, Marie-Helene; Datta, Sebak; Müller-Holzner, Elisabeth; Fiegl, Heidi; Böck, Günther; van Rooijen, Nico; Theurl, Igor; Doppler, Wolfgang

2014-08-01

Infiltration of a neoplasm with tumor-associated macrophages (TAMs) is considered an important negative prognostic factor and is functionally associated with tumor vascularization, accelerated growth, and dissemination. However, the ontogeny and differentiation pathways of TAMs are only incompletely characterized. Here, we report that intense local proliferation of fully differentiated macrophages rather than low-pace recruitment of blood-borne precursors drives TAM accumulation in a mouse model of spontaneous mammary carcinogenesis, the MMTVneu strain. TAM differentiation and expansion is regulated by CSF1, whose expression is directly controlled by STAT1 at the gene promoter level. These findings appear to be also relevant for human breast cancer, in which an interrelationship between STAT1, CSF1, and macrophage marker expression was identified. We propose that, akin to various MU subtypes in nonmalignant tissues, local proliferation and CSF1 play a vital role in the homeostasis of TAMs. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

OK-432 Suppresses Proliferation and Metastasis by Tumor Associated Macrophages in Bladder Cancer.

PubMed

Tian, Yuan-Feng; Tang, Kun; Guan, Wei; Yang, Tao; Xu, Hua; Zhuang, Qian-Yuan; Ye, Zhang-Qun

2015-01-01

OK-432, a Streptococcus-derived anticancer immunotherapeutic agent, has been applied in clinic for many years and achieved great progress in various cancers. In the present study, we investigated its anticancer effect on bladder cancer through tumor associated macrophages (TAMs). MTS assay validated OK-432 could inhibit proliferation in both T24 and EJ bladder cell lines. OK-432 also induced apoptosis of bladder cancer cells in vitro. Consequently, we demonstrated that OK-432 could suppress the bladder cancer cells migration and invasion by altering the EMT-related factors. Furthermore, using SD rat model, we revealed that OK-432 inhibited tumor growth, suppressed PCNA expression and inhibited metastasis in vivo. Taken together, these findings strongly suggest that OK-432 inhibits cell proliferation and metastasis through inducing macrophages to secret cytokines in bladder cancer.

Nanomaterial induction of oxidative stress in lung epithelial cells and macrophages

NASA Astrophysics Data System (ADS)

Wang, Lin; Pal, Anoop K.; Isaacs, Jacqueline A.; Bello, Dhimiter; Carrier, Rebecca L.

2014-09-01

Oxidative stress in the lung epithelial A549 cells and macrophages J774A.1 due to contact with commercially important nanomaterials [i.e., nano-silver (nAg), nano-alumina (nAl2O3), single-wall carbon nanotubes (CNT), and nano-titanium oxide anatase (nTiO2)] was evaluated. Nanomaterial-induced intracellular oxidative stress was analyzed by both H2DCFDA fluorescein probe and GSH depletion, extracellular oxidative stress was assessed by H2HFF fluorescein probes, and the secretion of chemokine IL-8 by A549 cells due to elevation of cellular oxidative stress was also monitored, in order to provide a comprehensive in vitro study on nanomaterial-induced oxidative stress in lung. In addition, results from this study were also compared with an acellular "ferric reducing ability of serum" (FRAS) assay and a prokaryotic cell-based assay in evaluating oxidative damage caused by the same set of nanomaterials, for comparison purposes. In general, it was found that nanomaterial-induced oxidative stress is highly cell-type dependent. In A549 lung epithelial cells, nAg appeared to induce highest level of oxidative stress and cell death followed by CNT, nTiO2, and nAl2O3. Different biological oxidative damage (BOD) assays' (i.e., H2DCFA, GSH, and IL-8 release) results generally agreed with each other, and the same trends of nanomaterial-induced BOD were also observed in acellular FRAS and prokaryotic E. coli K12-based assay. In macrophage J774A.1 cells, nAl2O3 and nTiO2 appeared to induce highest levels of oxidative stress. These results suggest that epithelial and macrophage cell models may provide complimentary information when conducting cell-based assays to evaluate nanomaterial-induced oxidative damage in lung.

Anti-atherogenic activity of wild grape (Vitis thunbergii) extract antagonizing smooth muscle cell proliferation and migration promoted by neighboring macrophages.

PubMed

Kang, Sang-Wook; Kim, Min Soo; Kim, Hyun-Sung; Lee, Yong-Jin; Kang, Young-Hee

2012-06-01

The proliferation and migration of vascular smooth muscle cells (SMCs) play critical roles in intimal thickening and neointimal hyperplasia in early-phase atherosclerosis. This study tested whether wild grape extract (WGE) suppressed the proliferation and migration of human aortic SMCs induced by neighboring macrophages. Cellular expression of fibrogenic connective tissue growth factor (CTGF) and secretion of collagen IV and matrix metalloproteinase (MMP)-2 were determined in SMCs exposed to THP-1-differentiated macrophage-conditioned media. Proliferation was enhanced in SMCs exposed to macrophage-conditioned media collected during the early stage of differentiation, which was attenuated by treatment with ≥ 10 µg/ml WGE. Increased secretion of CTGF and collagen IV macrophage-conditioned media was suppressed in WGE-supplemented SMCs. TGF-β1-promoted production of CTGF and collagen IV was suppressed by blocking TGF-β receptors of R1 and R2 in SMCs. WGE repressed macrophage-conditioned media-upregulated MMP-2 secretion, indicating that WGE had an ability to encumber plaque rupture within atherosclerotic lesions. In addition, ≥ 1 µg/ml WGE ameliorated the migration of SMCs promoted by neighboring macrophages. These results demonstrate that WGE retarded neointimal hyperplasia and thickening within atherosclerotic plaques largely comprising of macrophages and SMCs. Therefore, WGE may be developed as an anti-proliferative and anti-migratory agent targeting SMCs in the proximity of newly differentiated and resident macrophages.

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

PubMed

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

2016-12-27

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

P2Y receptors and atherosclerosis in apolipoprotein E-deficient mice

PubMed Central

Guns, Pieter-Jan DF; Hendrickx, Jan; Van Assche, Tim; Fransen, Paul; Bult, Hidde

2010-01-01

Background and purpose: P2Y nucleotide receptors are involved in the regulation of vascular tone, smooth muscle cell (SMC) proliferation and inflammatory responses. The present study investigated whether they are involved in atherosclerosis. Experimental approach: mRNA of P2Y receptors was quantified (RT-PCR) in atherosclerotic and plaque-free aorta segments of apolipoprotein E-deficient (apoE–/–) mice. Macrophage activation was assessed in J774 macrophages, and effects of non-selective purinoceptor antagonists on atherosclerosis were evaluated in cholesterol-fed apoE–/– mice. Key results: P2Y6 receptor mRNA was consistently elevated in segments with atherosclerosis, whereas P2Y2 receptor expression remained unchanged. Expression of P2Y1 or P2Y4 receptor mRNA was low or undetectable, and not influenced by atherosclerosis. P2Y6 mRNA expression was higher in cultured J774 macrophages than in cultured aortic SMCs. Furthermore, immunohistochemical staining of plaques demonstrated P2Y6-positive macrophages, but few SMCs, suggesting that macrophage recruitment accounted for the increase in P2Y6 receptor mRNA during atherosclerosis. In contrast to ATP, the P2Y6-selective agonist UDP increased mRNA expression and activity of inducible nitric oxide synthase and interleukin-6 in J774 macrophages; this effect was blocked by suramin (100–300 µM) or pyridoxal-phosphate-6-azophenyl-2′-4′-disulphonic acid (PPADS, 10–30 µM). Finally, 4-week treatment of cholesterol-fed apoE–/– mice with suramin or PPADS (50 and 25 mg·kg−1·day−1 respectively) reduced plaque size, without changing plaque composition (relative SMC and macrophage content) or cell replication. Conclusions and implications: These results suggest involvement of nucleotide receptors, particularly P2Y6 receptors, during atherosclerosis, and warrant further research with selective purinoceptor antagonists or P2Y6 receptor-deficient mice. PMID:20050854

The receptor tyrosine kinase MerTK activates phospholipase C γ2 during recognition of apoptotic thymocytes by murine macrophages

PubMed Central

Todt, Jill C.; Hu, Bin; Curtis, Jeffrey L.

2008-01-01

Apoptotic leukocytes must be cleared efficiently by macrophages (Mø). Apoptotic cell phagocytosis by Mø requires the receptor tyrosine kinase (RTK) MerTK (also known as c-Mer and Tyro12), the phosphatidylserine receptor (PS-R), and the classical protein kinase C (PKC) isoform βII, which translocates to Mø membrane and cytoskeletal fractions in a PS-R-dependent fashion. How these molecules cooperate to induce phagocytosis is unknown. Because the phosphatidylinositol-specific phospholipase (PI-PLC) PLC γ2 is downstream of RTKs in some cell types and can activate classical PKCs, we hypothesized that MerTK signals via PLC γ2. To test this hypothesis, we examined the interaction of MerTK and PLC γ2 in resident murine PMø and in the murine Mø cell line J774A.1 (J774) following exposure to apoptotic thymocytes. We found that, as with PMø, J774 phagocytosis of apoptotic thymocytes was inhibited by antibody against MerTK. Western blotting and immunoprecipitation showed that exposure to apoptotic cells produced three time-dependent changes in PMø and J774: (1) tyrosine phosphorylation of MerTK; (2) association of PLC γ2 with MerTK; and (3) tyrosine phosphorylation of PLC γ2. Phosphorylation of PLC γ2 and its association with MerTK was also induced by cross-linking MerTK using antibody. A PI-PLC appears to be required for phagocytosis of apoptotic cells because the PI-PLC inhibitor Et-18-OCH3 and the PLC inhibitor U73122, but not the inactive control U73343, blocked phagocytosis without impairing adhesion. On apoptotic cell adhesion to Mø, MerTK signals at least in part via PLC γ2. PMID:14704368

Cytotoxic activity and composition of petroleum ether extract from Magydaris tomentosa (Desf.) W. D. J. Koch (Apiaceae).

PubMed

Autore, Giuseppina; Marzocco, Stefania; Formisano, Carmen; Bruno, Maurizio; Rosselli, Sergio; Jemia, Mariem Ben; Senatore, Felice

2015-01-16

The petroleum ether extract of Magydaris tomentosa flowers (Desf.) W. D. J. Koch has been analyzed by GC-MS. It is mainly constituted by furanocoumarins such as xanthotoxin, xanthotoxol, isopimpinellin, and bergaptene. Other coumarins such as 7-methoxy-8-(2-formyl-2-methylpropyl) coumarin and osthole also occurred. The antiproliferative activity of Magydaris tomentosa flower extract has been evaluated in vitro on murine monocye/macrophages (J774A.1), human melanoma (A375) and human breast cancer (MCF-7) tumor cell lines, showing a major activity against the latter.

Rebamipide delivered by brushite cement enhances osteoblast and macrophage proliferation.

PubMed

Pujari-Palmer, Michael; Pujari-Palmer, Shiuli; Engqvist, Håkan; Karlsson Ott, Marjam

2015-01-01

Many of the bioactive agents capable of stimulating osseous regeneration, such as bone morphogenetic protein-2 (BMP-2) or prostaglandin E2 (PGE2), are limited by rapid degradation, a short bioactive half-life at the target site in vivo, or are prohibitively expensive to obtain in large quantities. Rebamipide, an amino acid modified hydroxylquinoline, can alter the expression of key mediators of bone anabolism, cyclo-oxygenase 2 (COX-2), BMP-2 and vascular endothelial growth factor (VEGF), in diverse cell types such as mucosal and endothelial cells or chondrocytes. The present study investigates whether Rebamipide enhances proliferation and differentiation of osteoblasts when delivered from brushite cement. The reactive oxygen species (ROS) quenching ability of Rebampide was tested in macrophages as a measure of bioactivity following drug release incubation times, up to 14 days. Rebamipide release from brushite occurs via non-fickian diffusion, with a rapid linear release of 9.70% ± 0.37% of drug per day for the first 5 days, and an average of 0.5%-1% per day thereafter for 30 days. Rebamipide slows the initial and final cement setting time by up to 3 and 1 minute, respectively, but does not significantly reduce the mechanical strength below 4% (weight percentage). Pre-osteoblast proliferation increases by 24% upon exposure to 0.4 uM Rebamipide, and by up to 73% when Rebamipide is delivered via brushite cement. Low doses of Rebamipide do not adversely affect peak alkaline phosphatase activity in differentiating pre-osteoblasts. Rebamipide weakly stimulates proliferation in macrophages at low concentrations (118 ± 7.4% at 1 uM), and quenches ROS by 40-60%. This is the first investigation of Rebamipide in osteoblasts.

Evaluation of the leishmanicidal and cytotoxic potential of essential oils derived from ten colombian plants.

PubMed

Sanchez-Suarez, Jf; Riveros, I; Delgado, G

2013-01-01

The leishmanicidal and cytotoxic activity of ten essential oils obtained from ten plant specimens were evaluated. Essential oils were obtained by the steam distillation of plant leaves without any prior processing. Cytotoxicity was tested on J774 macrophages and leishmanicidal activity was assessed against four species of Leishmania associated with cutaneous leishmaniasis. Seven essential oils exhibited activity against Leishmania parasites, five of which were toxic against J774 macrophages. Selectivity indices of >6 and 13 were calculated for the essential oils of Ocimum basilicum and Origanum vulgare, respectively. The essential oil of Ocimum basilicum was active against promastigotes of Leishmania and innocuous to J774 macrophages at concentrations up to 1600 µg/mL and should be further investigated for leishmanicidal activity in others in vitro and in vivo experimental models.

Wallerian degeneration in C57BL/6J and A/J mice: differences in time course of neurofilament and myelin breakdown, macrophage recruitment and iNOS expression

PubMed Central

de la Hoz, Cristiane L R; Oliveira, Alexandre L R; de S Queiroz, Luciano; Langone, Francesco

2003-01-01

The lower regeneration potential reported for C57BL/6J mice strain after peripheral nerve lesion may result from alterations in crucial events during Wallerian degeneration. We analysed neurofilament and myelin breakdown, macrophage recruitment, NADPH-diaphorase reaction and inducible nitric oxide synthase (iNOS) expression in transected sciatic nerves of C57BL/6J and A/J mice. The neurofilament volume density was lower in C57BL/6J strain mice at 1 and 3 days after lesion, and later equalled the density observed in A/J. C57BL/6J mice presented a high number of cells containing myelin debris, 3 and 5 days after the lesion. In both strains iNOS immunoreactivity was intense in macrophages and less evident in Schwann cells. However, a delay in macrophage recruitment and a lower percentage of iNOS-expressing macrophages on the third day were observed in C57BL/6J mice. NADPH-diaphorase reaction disclosed a similar pattern for both strains until the seventh day. However, at 5 days, cells with slender processes involving ellipsoid segments showed a well-defined cytoplasmic labelling in C57BL/6J whereas in A/J most of these cells exhibited a more granular and disperse labelling. We propose that these differences between A/J and C57BL/6J strains during Wallerian degeneration may be implicated in the lower regeneration potential observed in the latter. PMID:14686692

Macrophage sphingolipids are essential for the entry of mycobacteria.

PubMed

Viswanathan, Gopinath; Jafurulla, Md; Kumar, G Aditya; Raghunand, Tirumalai R; Chattopadhyay, Amitabha

2018-07-01

Mycobacteria are intracellular pathogens that can invade and survive within host macrophages. Mycobacterial infections remain a major cause of mortality and morbidity worldwide, with serious concerns of emergence of multi and extensively drug-resistant tuberculosis. While significant advances have been made in identifying mycobacterial virulence determinants, the detailed molecular mechanism of internalization of mycobacteria into host cells remains poorly understood. Although several studies have highlighted the crucial role of sphingolipids in mycobacterial growth, persistence and establishment of infection, the role of sphingolipids in the entry of mycobacteria into host cells is not known. In this work, we explored the role of host membrane sphingolipids in the entry of Mycobacterium smegmatis into J774A.1 macrophages. Our results show that metabolic depletion of sphingolipids in host macrophages results in a significant reduction in the entry of M. smegmatis. Importantly, the entry of Escherichia coli into host macrophages under similar conditions remained invariant, implying the specificity of the requirement of sphingolipids in mycobacterial entry. To the best of our knowledge, our results constitute the first report demonstrating the role of host macrophage sphingolipids in the entry of mycobacteria. Our results could help in the development of novel therapeutic strategies targeting sphingolipid-mediated entry of mycobacteria into host cells. Copyright © 2018 Elsevier B.V. All rights reserved.

Rebamipide Delivered by Brushite Cement Enhances Osteoblast and Macrophage Proliferation

PubMed Central

Pujari-Palmer, Michael; Pujari-Palmer, Shiuli; Engqvist, Håkan; Karlsson Ott, Marjam

2015-01-01

Many of the bioactive agents capable of stimulating osseous regeneration, such as bone morphogenetic protein-2 (BMP-2) or prostaglandin E2 (PGE2), are limited by rapid degradation, a short bioactive half-life at the target site in vivo, or are prohibitively expensive to obtain in large quantities. Rebamipide, an amino acid modified hydroxylquinoline, can alter the expression of key mediators of bone anabolism, cyclo-oxygenase 2 (COX-2), BMP-2 and vascular endothelial growth factor (VEGF), in diverse cell types such as mucosal and endothelial cells or chondrocytes. The present study investigates whether Rebamipide enhances proliferation and differentiation of osteoblasts when delivered from brushite cement. The reactive oxygen species (ROS) quenching ability of Rebampide was tested in macrophages as a measure of bioactivity following drug release incubation times, up to 14 days. Rebamipide release from brushite occurrs via non-fickian diffusion, with a rapid linear release of 9.70% ±0.37% of drug per day for the first 5 days, and an average of 0.5%-1% per day thereafter for 30 days. Rebamipide slows the initial and final cement setting time by up to 3 and 1 minute, respectively, but does not significantly reduce the mechanical strength below 4% (weight percentage). Pre-osteoblast proliferation increases by 24% upon exposure to 0.4uM Rebamipide, and by up to 73% when Rebamipide is delivered via brushite cement. Low doses of Rebamipide do not adversely affect peak alkaline phosphatase activity in differentiating pre-osteoblasts. Rebamipide weakly stimulates proliferation in macrophages at low concentrations (118 ±7.4% at 1uM), and quenches ROS by 40-60%. This is the first investigation of Rebamipide in osteoblasts. PMID:26023912

Differential activation of Fyn kinase distinguishes saturated and unsaturated fats in mouse macrophages

PubMed Central

Tarabra, Elena; An Lee, Ting-Wen; Zammit, Victor A.; Vatish, Manu; Yamada, Eijiro; Pessin, Jeffrey E.; Bastie, Claire C.

2017-01-01

Diet-induced obesity is associated with increased adipose tissue activated macrophages. Yet, how macrophages integrate fatty acid (FA) signals remains unclear. We previously demonstrated that Fyn deficiency (fynKO) protects against high fat diet-induced adipose tissue macrophage accumulation. Herein, we show that inflammatory markers and reactive oxygen species are not induced in fynKO bone marrow-derived macrophages exposed to the saturated FA palmitate, suggesting that Fyn regulates macrophage function in response to FA signals. Palmitate activates Fyn and re-localizes Fyn into the nucleus of RAW264.7, J774 and wild-type bone marrow-derived macrophages. Similarly, Fyn activity is increased in cells of adipose tissue stromal vascular fraction of high fat-fed control mice, with Fyn protein being located in the nucleus of these cells. We demonstrate that Fyn modulates palmitate-dependent oxidative stress in macrophages. Moreover, Fyn catalytic activity is necessary for its nuclear re-localization and downstream effects, as Fyn pharmacological inhibition abolishes palmitate-induced Fyn nuclear redistribution and palmitate-dependent increase of oxidative stress markers. Importantly, mono-or polyunsaturated FAs do not activate Fyn, and fail to re-localize Fyn to the nucleus. Together these data demonstrate that macrophages integrate nutritional FA signals via a differential activation of Fyn that distinguishes, at least partly, the effects of saturated versus unsaturated fats. PMID:29156823

Differential activation of Fyn kinase distinguishes saturated and unsaturated fats in mouse macrophages.

PubMed

Tarabra, Elena; An Lee, Ting-Wen; Zammit, Victor A; Vatish, Manu; Yamada, Eijiro; Pessin, Jeffrey E; Bastie, Claire C

2017-10-17

Diet-induced obesity is associated with increased adipose tissue activated macrophages. Yet, how macrophages integrate fatty acid (FA) signals remains unclear. We previously demonstrated that Fyn deficiency ( fynKO ) protects against high fat diet-induced adipose tissue macrophage accumulation. Herein, we show that inflammatory markers and reactive oxygen species are not induced in fynKO bone marrow-derived macrophages exposed to the saturated FA palmitate, suggesting that Fyn regulates macrophage function in response to FA signals. Palmitate activates Fyn and re-localizes Fyn into the nucleus of RAW264.7, J774 and wild-type bone marrow-derived macrophages. Similarly, Fyn activity is increased in cells of adipose tissue stromal vascular fraction of high fat-fed control mice, with Fyn protein being located in the nucleus of these cells. We demonstrate that Fyn modulates palmitate-dependent oxidative stress in macrophages. Moreover, Fyn catalytic activity is necessary for its nuclear re-localization and downstream effects, as Fyn pharmacological inhibition abolishes palmitate-induced Fyn nuclear redistribution and palmitate-dependent increase of oxidative stress markers. Importantly, mono-or polyunsaturated FAs do not activate Fyn, and fail to re-localize Fyn to the nucleus. Together these data demonstrate that macrophages integrate nutritional FA signals via a differential activation of Fyn that distinguishes, at least partly, the effects of saturated versus unsaturated fats.

Evaluation of the Leishmanicidal and Cytotoxic Potential of Essential Oils Derived From Ten Colombian Plants

PubMed Central

Sanchez-Suarez, JF; Riveros, I; Delgado, G

2013-01-01

Background The leishmanicidal and cytotoxic activity of ten essential oils obtained from ten plant specimens were evaluated. Methods Essential oils were obtained by the steam distillation of plant leaves without any prior processing. Cytotoxicity was tested on J774 macrophages and leishmanicidal activity was assessed against four species of Leishmania associated with cutaneous leishmaniasis. Results Seven essential oils exhibited activity against Leishmania parasites, five of which were toxic against J774 macrophages. Selectivity indices of >6 and 13 were calculated for the essential oils of Ocimum basilicum and Origanum vulgare, respectively. Conclusion The essential oil of Ocimum basilicum was active against promastigotes of Leishmania and innocuous to J774 macrophages at concentrations up to 1600 µg/mL and should be further investigated for leishmanicidal activity in others in vitro and in vivo experimental models. PMID:23682270

Anti-inflammatory activity of horseradish (Armoracia rusticana) root extracts in LPS-stimulated macrophages.

PubMed

Marzocco, Stefania; Calabrone, Luana; Adesso, Simona; Larocca, Marilena; Franceschelli, Silvia; Autore, Giuseppina; Martelli, Giuseppe; Rossano, Rocco

2015-12-01

Horseradish (Armoracia rusticana) is a perennial crop belonging to the Brassicaceae family. Horseradish root is used as a condiment due to its extremely pungent flavour, deriving from the high content of glucosinolates and their breakdown products such as isothiocyanates and other sulfur compounds. Horseradish also has a long history in ethnomedicine. In this study the anti-inflammatory potential of three accessions of Armoracia rusticana on lipopolysaccharide from E. coli treated J774A.1 murine macrophages was evaluated. Our results demonstrate that Armoracia rusticana reduced nitric oxide, tumor necrosis factor-α and interleukin-6 release and nitric oxide synthase and cyclooxygenase-2 expression in macrophages, acting on nuclear transcription factor NF-κB p65 activation. Moreover Armoracia rusticana reduced reactive oxygen species release and increased heme-oxygenase-1 expression, thus contributing to the cytoprotective cellular effect during inflammation.

The thymus of the hairless rhino-j (hr/hr-j) mice

PubMed Central

SAN JOSE, I.; GARCÍA-SUÁREZ, O.; HANNESTAD, J.; CABO, R.; GAUNA, L.; REPRESA, J.; VEGA, J. A.

2001-01-01

The hairless (hr) gene is expressed in a large number of tissues, primarily the skin, and a mutation in the hr gene is responsible for the typical cutaneous phenotype of hairless mice. Mutant hr mouse strains show immune defects involving especially T cells and macrophages, as well as an age-related immunodeficiency and an accelerated atrophy of the thymus. These data suggest that the hr mutation causes a defect of this organ, although hr transcripts have not been detected in fetal or adult mice thymus. The present study analyses the thymus of young (3 mo) and adult (9 mo) homozygous hr-rh-j mice (a strain of hairless mice) by means of structural techniques and immunohistochemistry to selectively identify thymic epithelial cells, dendritic cells, and macrophages. There were structural alterations in the thymus of both young and adult rh-rh-j mice, which were more severe in older animals. These alterations consisted of relative cortical atrophy, enlargement of blood vessels, proliferation of perivascular connective tissue, and the appearance of cysts. hr-rh-j mice also showed a decrease in the number of epithelial and dendritic cells, and macrophages. Taken together, present results strongly suggest degeneration and accelerated age-dependent regression of the thymus in hr-rh-j mice, which could explain at least in part the immune defects reported in hairless mouse strains. PMID:11327202

The scavenger receptor SR-A I/II (CD204) signals via the receptor tyrosine kinase Mertk during apoptotic cell uptake by murine macrophages

PubMed Central

Todt, Jill C.; Hu, Bin; Curtis, Jeffrey L.

2008-01-01

Apoptotic cells (AC) must be cleared by macrophages (Mø) to resolve inflammation effectively. Mertk and scavenger receptor A (SR-A) are two of many receptors involved in AC clearance. As SR-A lacks enzymatic activity or evident intracellular signaling motifs, yet seems to signal in some cell types, we hypothesized that SR-A signals via Mer receptor tyrosine kinase (Mertk), which contains a multisubstrate docking site. We induced apoptosis in murine thymocytes by dexamethasone and used Western blotting and immunoprecipitation to analyze the interaction of Mertk and SR-A in the J774A.1 (J774) murine Mø cell line and in peritoneal Mø of wild-type mice and SR-A−/− mice. Phagocytosis (but not adhesion) of AC by J774 was inhibited by anti-SR-A or function-blocking SR-A ligands. In resting J774, SR-A was associated minimally with unphosphorylated (monomeric) Mertk; exposure to AC induced a time-dependent increase in association of SR-A with Mertk in a direct or indirect manner. Anti-SR-A inhibited AC-induced phosphorylation of Mertk and of phospholipase Cγ2, essential steps in AC ingestion. Relative to tissue Mø of wild-type mice, AC-induced Mertk phosphorylation was reduced and delayed in tissue Mø of SR-A−/− mice, as was in vitro AC ingestion at early time-points. Thus, during AC uptake by murine Mø, SR-A is essential for optimal phosphorylation of Mertk and subsequent signaling required for AC ingestion. These data support the Mertk/SR-A complex as a potential target to manipulate AC clearance and hence, resolution of inflammation and infections. PMID:18511575

YopJ-induced caspase-1 activation in Yersinia-infected macrophages: independent of apoptosis, linked to necrosis, dispensable for innate host defense.

PubMed

Zheng, Ying; Lilo, Sarit; Mena, Patricio; Bliska, James B

2012-01-01

Yersinia outer protein J (YopJ) is a type III secretion system (T3SS) effector of pathogenic Yersinia (Yersinia pestis, Yersinia enterocolitica and Yersinia pseudotuberculosis) that is secreted into host cells. YopJ inhibits survival response pathways in macrophages, causing cell death. Allelic variation of YopJ is responsible for differential cytotoxicity in Yersinia strains. YopJ isoforms in Y. enterocolitica O:8 (YopP) and Y. pestis KIM (YopJ(KIM)) strains have high cytotoxic activity. In addition, YopJ(KIM)-induced macrophage death is associated with caspase-1 activation and interleukin-1β (IL-1β secretion. Here, the mechanism of YopJ(KIM)-induced cell death, caspase-1 activation, and IL-1β secretion in primary murine macrophages was examined. Caspase-3/7 activity was low and the caspase-3 substrate poly (ADP-ribose) polymerase (PARP) was not cleaved in Y. pestis KIM5-infected macrophages. In addition, cytotoxicity and IL-1β secretion were not reduced in the presence of a caspase-8 inhibitor, or in B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax)/Bcl-2 homologous antagonist/killer (Bak) knockout macrophages, showing that YopJ(KIM)-mediated cell death and caspase-1 activation occur independent of mitochondrial-directed apoptosis. KIM5-infected macrophages released high mobility group protein B1 (HMGB1), a marker of necrosis, and microscopic analysis revealed that necrotic cells contained active caspase-1, indicating that caspase-1 activation is associated with necrosis. Inhibitor studies showed that receptor interacting protein 1 (RIP1) kinase and reactive oxygen species (ROS) were not required for cytotoxicity or IL-β release in KIM5-infected macrophages. IL-1β secretion was reduced in the presence of cathepsin B inhibitors, suggesting that activation of caspase-1 requires cathepsin B activity. Ectopically-expressed YopP caused higher cytotoxicity and secretion of IL-1β in Y. pseudotuberculosis-infected macrophages than YopJ(KIM). Wild-type and

The Effect of Garlic Extract on Expression of INFγ And Inos Genes in Macrophages Infected with Leishmania major

PubMed Central

Gharavi, MJ; Nobakht, M; Khademvatan, SH; Bandani, E; Bakhshayesh, M; Roozbehani, M

2011-01-01

Background The study was aimed to show the effect of molecular mechanism of Aqueous Garlic Extract (AGE) on expression of IFNγ and iNOS genes in Leishmania major. Methods Leishmania major promastigotes (MRHO/IR/75/ER) were added to the in-vitro cultured J774 cell line, the cells were incubated for 72 hours. Various concentrations of garlic extract (9.25, 18.5, 37, 74, 148 mg/ml) were added to the infected cells. MTT assay was applied for cellular proliferation. After 72 hours of incubation, supernatants were collected and total RNA was extracted from the infected cells. The express of IFNγ and iNOS genes were studied by RT-PCR method. Results The colorimetric MTT assay after 3 days of incubation showed cytotoxic effect of garlic extract with an IC50 of 37 mg/ml. In addition, IFNγ and iNOS genes expression by RT-PCR indicated that garlic extract lead to over expression of these genes in J774 cell line infected with L. major. Conclusion Garlic extract exerts cytotoxic effect on infected J774 cell line. In addition, the hypothesis that garlic can improve cellular immunity with raising the expression of IFNγ and of iNOS genes confirmed. PMID:22347300

Silica-induced initiation of circular ZC3H4 RNA/ZC3H4 pathway promotes the pulmonary macrophage activation.

PubMed

Yang, Xiyue; Wang, Jing; Zhou, Zewei; Jiang, Rong; Huang, Jie; Chen, Lulu; Cao, Zhouli; Chu, Han; Han, Bing; Cheng, Yusi; Chao, Jie

2018-06-01

Phagocytosis of silicon dioxide (SiO 2 ) into lung cells causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Circular RNAs (circRNAs) are a subclass of noncoding RNAs that are present within mammalian cells; however, researchers have not determined whether circRNAs are involved in the pathophysiologic process of silicosis. To elucidate the role of these RNAs in SiO 2 -induced inflammation in pulmonary macrophages, we investigated the upstream molecular mechanisms and functional effects of circRNAs on cell apoptosis, proliferation, and migration. Primary cultures of alveolar macrophages from healthy donors and from patients and the RAW264.7 macrophage cell line were used to explore the functions of circZC3H4 RNA in macrophage activation. The experimental results indicated the following: 1) SiO 2 concomitantly increased circZC3H4 RNA expression and increased ZC3H4 protein levels; 2) circular ZC3H4 (circZC3H4) RNA and ZC3H4 protein participated in SiO 2 -induced macrophage activation; and 3) SiO 2 -activated macrophages promoted fibroblast proliferation and migration via the circZC3H4 RNA/ZC3H4 pathway. The up-regulation of the ZC3H4 protein was confirmed in tissue samples from patients with silicosis. Our study elucidates a link between SiO 2 -induced macrophage activation and the circZC3H4 RNA/ZC3H4 pathway, thereby providing novel insight into the potential use of ZC3H4 to develop novel therapeutic strategies for silicosis.-Yang, X., Wang, J., Zhou, Z., Jiang, R., Huang, J., Chen, L., Cao, Z., Chu, H., Han, B., Cheng, Y., Chao, J. Silica-induced initiation of circular ZC3H4 RNA/ZC3H4 pathway promotes the pulmonary macrophage activation.

30 CFR 774.9 - Information collection.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 30 Mineral Resources 3 2011-07-01 2011-07-01 false Information collection. 774.9 Section 774.9... OTHER ACTIONS BASED ON OWNERSHIP, CONTROL, AND VIOLATION INFORMATION § 774.9 Information collection. (a) The collections of information contained in part 774 have been approved by the Office of Management...

30 CFR 774.9 - Information collection.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 30 Mineral Resources 3 2012-07-01 2012-07-01 false Information collection. 774.9 Section 774.9... OTHER ACTIONS BASED ON OWNERSHIP, CONTROL, AND VIOLATION INFORMATION § 774.9 Information collection. (a) The collections of information contained in part 774 have been approved by the Office of Management...

Peroxisome proliferator-activated receptor-gamma agonist rosiglitazone attenuates postincisional pain by regulating macrophage polarization

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hasegawa-Moriyama, Maiko, E-mail: hase-mai@m3.kufm.kagoshima-u.ac.jp; Ohnou, Tetsuya; Godai, Kohei

Highlights: Black-Right-Pointing-Pointer Rosiglitazone attenuated postincisional pain. Black-Right-Pointing-Pointer Rosiglitazone alters macrophage polarization to F4/80{sup +}CD206{sup +} M2 macrophages at the incisional sites. Black-Right-Pointing-Pointer Transplantation of rosiglitazone-treated macrophages produced analgesic effects. -- Abstract: Acute inflammation triggered by macrophage infiltration to injured tissue promotes wound repair and may induce pain hypersensitivity. Peroxisome proliferator-activated receptor {gamma} (PPAR){gamma} signaling is known to regulate heterogeneity of macrophages, which are often referred to as classically activated (M1) and alternatively activated (M2) macrophages. M1 macrophages have considerable antimicrobial activity and produce a wide variety of proinflammatory cytokines. In contrast, M2 macrophages are involved in anti-inflammatory and homeostatic functionsmore » linked to wound healing and tissue repair. Although it has been suggested that PPAR{gamma} agonists attenuate pain hypersensitivity, the molecular mechanism of macrophage-mediated effects of PPAR{gamma} signaling on pain development has not been explored. In this study, we investigated the link between the phenotype switching of macrophage polarization induced by PPAR{gamma} signaling and the development of acute pain hypersensitivity. Local administration of rosiglitazone significantly ameliorated hypersensitivity to heat and mechanical stimuli, and paw swelling. Consistent with the down-regulation of nuclear factor {kappa}B (NF{kappa}B) phosphorylation by rosiglitazone at the incisional sites, the number of F4/80{sup +}iNOS{sup +} M1 macrophages was decreased whereas numbers of F4/80{sup +}CD206{sup +} M2 macrophages were increased in rosiglitazone-treated incisional sites 24 h after the procedure. In addition, gene induction of anti-inflammatory M2-macrophage-associated markers such as arginase1, FIZZ1 and interleukin (IL)-10 were significantly increased

Nanospheres Encapsulating Anti-Leishmanial Drugs for Their Specific Macrophage Targeting, Reduced Toxicity, and Deliberate Intracellular Release

PubMed Central

Shukla, Anil Kumar; Patra, Sanjukta

2012-01-01

Abstract The current work focuses on the study of polymeric, biodegradable nanoparticles (NPs) for the encapsulation of doxorubicin and mitomycin C (anti-leishmanial drugs), and their efficient delivery to macrophages, the parasite's home. The biodegradable polymer methoxypoly-(ethylene glycol)-b-poly (lactic acid) (MPEG-PLA) was used to prepare polymeric NPs encapsulating doxorubicin and mitomycin C. The morphology, mean diameter, and surface area of spherical NPs were determined by transmission electron microscopy (TEM), field emission scanning electron microscopy (FESEM), and BET surface area analysis. X-ray diffraction was performed to validate drug encapsulation. An in vitro release profile of the drugs suggested a fairly slow release. These polymeric NPs were efficiently capable of releasing drug inside macrophages at a slower pace than the free drug, which was monitored by epi-fluorescence microscopy. Encapsulation of doxorubicin and mitomycin C into NPs also decreases cellular toxicity in mouse macrophages (J774.1A). PMID:22925019

[Phagocytosis and intracellular proliferation of Nocardia asteroides (strain Weipheld) in cell structures in vitro. 2. Peritoneal macrophages of guinea-pigs (author's transl)].

PubMed

Splino, M; Mĕrka, V; Kyntera, F

1976-08-01

The study deals with the phagocytosis of Nocardia asteroides (strain Weipheld) and the subsequent intracellular proliferation in peritoneal macrophage cells. Normal, two-stage-immunized and long-term cortison-treated guinea-pig (28 mg cortison / kg weight / day during 30 days) macrophages were used. Further, the cytotoxic effect of Nocardia upon the cells in the peritoneal washing liquid in vitro and the influence of the normal, immune and antimacrophage serum upon the phagocytosis and the intracellular proliferation were studied. Among the cells obtained from the peritoneal washing liquid macrophages were most frequently subject to phagocytosis, leukocytes to a lesser degree. The normal macrophages phagocytized in 14.56% (Fig. 1), macrophages of two-stage-immunized guinea-pigs in 18.21% (Fig. 2) and macrophages from cortison treated guinea-pigs in 12.48% of cases. Intracellular observation showed phagocytized germs after 3 min. of exposure. The course of the intracellular proliferation of Nocardia can be seen in Fig. 3. The phagocytosis index increases slowly in all three groups of macrophages; least so in the immunized macrophages (1.30-after 8 hours). The highest values were obtained in the macrophages of cortison treated guinea-pigs (2.02-after 8 hours). Within 8 hours of exposure the filaments of Nocardia grew through the cell membrane of phagocytizing cells (Figs. 4 A, 4 B). Fig. 5 shows the course of the cytopathogenic effect of Nocardia upon the cells. After 1 hr. the number of dead cells increased from 0.30% to 1.9-3.8%; after 4 hrs. it reached 8.15-9.80%; after 8 hrs. 10.1-14.80%. The highest values were observed in cells from cortison treated guinea-pigs (14.80%). After addition of normal serum (time of phagocytosis 60 min.) normal peritoneal macrophages phagocytized in 13.30% of cases; immune serum stimulated phagocytosis (16.21%); antimacrophage serum significantly reduced phagocytosis (4.10%). The phagocytosis index in peritoneal macrophages with

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

PubMed

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

2017-01-01

Avian leucosis virus subgroup J (ALV-J) can cause lifelong infection and can escape from the host immune defenses in chickens. Since macrophages act as the important defense line against invading pathogens in host innate immunity, we investigated the function and innate immune responses of chicken primary monocyte-derived macrophages (MDM) after ALV-J infection in this study. Our results indicated that ALV-J was stably maintained in MDM cells but that the viral growth rate was significantly lower than that in DF-1 cells. We also found that ALV-J infection significantly increased nitric oxide (NO) production, but had no effect on MDM phagocytic capacity. Interestingly, infection with ALV-J rapidly promoted the expression levels of Myxovirus resistance 1 (Mx) (3 h, 6 h), ISG12 (6 h), and interleukin-1β (IL-1β) (3 h, 12 h) at an early infection stage, whereas it sharply decreased the expression of Mx (24 h, 36 h), ISG12 (36 h), and made little change on IL-1β (24 h, 36 h) production at a late infection stage in MDM cells. Moreover, the protein levels of interferon-β (IFN-β) and interleukin-6 (IL-6) had sharply increased in infected MDM cells from 3 to 36 h post infection (hpi) of ALV-J. And, the protein level of interleukin-10 (IL-10) was dramatically decreased at 36 hpi in MDM cells infected with ALV-J. These results demonstrate that ALV-J can induce host innate immune responses and we hypothesize that macrophages play an important role in host innate immune attack and ALV-J immune escape. © The Author 2016. Published by Oxford University Press on behalf of Poultry Science Association.

Photodynamic therapy mediates innate immune responses via fibroblast-macrophage interactions.

PubMed

Zulaziz, N; Azhim, A; Himeno, N; Tanaka, M; Satoh, Y; Kinoshita, M; Miyazaki, H; Saitoh, D; Shinomiya, N; Morimoto, Y

2015-10-01

Antibacterial photodynamic therapy (PDT) has come to attract attention as an alternative therapy for drug-resistant bacteria. Recent reports revealed that antibacterial PDT induces innate immune response and stimulates abundant cytokine secretion as a part of inflammatory responses. However, the underlying mechanism how antibacterial PDT interacts with immune cells responsible for cytokine secretion has not been well outlined. In this study, we aimed to clarify the difference in gene expression and cytokine secretion between combined culture of fibroblasts and macrophages and their independent cultures. SCRC-1008, mouse fibroblast cell line and J774, mouse macrophage-like cell line were co-cultured and PDT treatments with different parameters were carried out. After various incubation periods (1-24 h), cells and culture medium were collected, and mRNA and protein levels for cytokines were measured using real-time PCR and ELISA, respectively. Our results showed that fibroblasts and macrophages interact with each other to mediate the immune response. We propose that fibroblasts initially respond to PDT by expressing Hspa1b, which regulates the NF-κB pathway via Tlr2 and Tlr4. Activation of the NF-κB pathway then results in an enhanced secretion of pro-inflammatory cytokines (TNF-α, IL-6 and IL-1β) and neutrophil chemoattractant MIP-2 and KC from macrophages.

7 CFR 774.9 - Environmental requirements.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 7 Agriculture 7 2013-01-01 2013-01-01 false Environmental requirements. 774.9 Section 774.9 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.9 Environmental requirements. The...

7 CFR 774.9 - Environmental requirements.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 7 2011-01-01 2011-01-01 false Environmental requirements. 774.9 Section 774.9 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.9 Environmental requirements. The...

7 CFR 774.9 - Environmental requirements.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Environmental requirements. 774.9 Section 774.9 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.9 Environmental requirements. The...

7 CFR 774.9 - Environmental requirements.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 7 Agriculture 7 2014-01-01 2014-01-01 false Environmental requirements. 774.9 Section 774.9 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.9 Environmental requirements. The...

7 CFR 774.9 - Environmental requirements.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 7 Agriculture 7 2012-01-01 2012-01-01 false Environmental requirements. 774.9 Section 774.9 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.9 Environmental requirements. The...

7 CFR 774.19 - Processing applications.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 7 2011-01-01 2011-01-01 false Processing applications. 774.19 Section 774.19 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.19 Processing applications...

7 CFR 774.19 - Processing applications.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 7 Agriculture 7 2012-01-01 2012-01-01 false Processing applications. 774.19 Section 774.19 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.19 Processing applications...

7 CFR 774.19 - Processing applications.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Processing applications. 774.19 Section 774.19 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.19 Processing applications...

7 CFR 774.19 - Processing applications.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 7 Agriculture 7 2014-01-01 2014-01-01 false Processing applications. 774.19 Section 774.19 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.19 Processing applications...

7 CFR 774.19 - Processing applications.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 7 Agriculture 7 2013-01-01 2013-01-01 false Processing applications. 774.19 Section 774.19 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.19 Processing applications...

7 CFR 774.20 - Funding applications.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 7 Agriculture 7 2013-01-01 2013-01-01 false Funding applications. 774.20 Section 774.20 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.20 Funding applications. Loan...

7 CFR 774.20 - Funding applications.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 7 Agriculture 7 2012-01-01 2012-01-01 false Funding applications. 774.20 Section 774.20 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.20 Funding applications. Loan...

7 CFR 774.20 - Funding applications.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 7 Agriculture 7 2014-01-01 2014-01-01 false Funding applications. 774.20 Section 774.20 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.20 Funding applications. Loan...

7 CFR 774.20 - Funding applications.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 7 2011-01-01 2011-01-01 false Funding applications. 774.20 Section 774.20 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.20 Funding applications. Loan...

7 CFR 774.20 - Funding applications.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Funding applications. 774.20 Section 774.20 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.20 Funding applications. Loan...

7 CFR 774.17 - Loan application.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Loan application. 774.17 Section 774.17 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.17 Loan application. A complete...

Evidence for apoptosis of murine macrophages by Actinobacillus actinomycetemcomitans infection.

PubMed

Kato, S; Muro, M; Akifusa, S; Hanada, N; Semba, I; Fujii, T; Kowashi, Y; Nishihara, T

1995-10-01

The gram-negative bacterium Actinobacillus actinomycetemcomitans is considered an important etiological agent in periodontal diseases. In this study, we show that A. actinomycetemcomitans strains are cytotoxic for the murine macrophage cell line J774.1. On the other hand, Porphyromonas gingivalis strains, other gram-negative oral species implicated in adult periodontitis, showed weak cytotoxic effects. For this to occur, A. actinomycetemcomitans had to gain entry into the macrophages, since cytotoxicity was prevented by cytochalasin D. We demonstrate that cell death induced by A. actinomycetemcomitans Y4 occurs through apoptosis, as shown by changes in nuclear morphology, an increase in the proportion of fragmented DNA, and the typical ladder pattern of DNA fragmentation indicative of apoptosis. We further sought to determine whether the cytotoxicity induced by A. actinomycetemcomitans Y4 could be modulated by the protein kinase inhibitors H7 and HA1004. Apoptotic cell death induced by A. actinomycetemcomitans Y4 was suppressed by H7 but was relatively unaffected by HA1004. These findings suggest that the signals of protein kinases may regulate apoptosis induced by A. actinomycetemcomitans Y4. The ability of A. actinomycetemcomitans to promote the apoptosis of macrophages may be important for the initiation of infection and the development of periodontal diseases.

Evidence for apoptosis of murine macrophages by Actinobacillus actinomycetemcomitans infection.

PubMed Central

Kato, S; Muro, M; Akifusa, S; Hanada, N; Semba, I; Fujii, T; Kowashi, Y; Nishihara, T

1995-01-01

The gram-negative bacterium Actinobacillus actinomycetemcomitans is considered an important etiological agent in periodontal diseases. In this study, we show that A. actinomycetemcomitans strains are cytotoxic for the murine macrophage cell line J774.1. On the other hand, Porphyromonas gingivalis strains, other gram-negative oral species implicated in adult periodontitis, showed weak cytotoxic effects. For this to occur, A. actinomycetemcomitans had to gain entry into the macrophages, since cytotoxicity was prevented by cytochalasin D. We demonstrate that cell death induced by A. actinomycetemcomitans Y4 occurs through apoptosis, as shown by changes in nuclear morphology, an increase in the proportion of fragmented DNA, and the typical ladder pattern of DNA fragmentation indicative of apoptosis. We further sought to determine whether the cytotoxicity induced by A. actinomycetemcomitans Y4 could be modulated by the protein kinase inhibitors H7 and HA1004. Apoptotic cell death induced by A. actinomycetemcomitans Y4 was suppressed by H7 but was relatively unaffected by HA1004. These findings suggest that the signals of protein kinases may regulate apoptosis induced by A. actinomycetemcomitans Y4. The ability of A. actinomycetemcomitans to promote the apoptosis of macrophages may be important for the initiation of infection and the development of periodontal diseases. PMID:7558299

Lysozyme activates Enterococcus faecium to induce necrotic cell death in macrophages.

PubMed

Gröbner, Sabine; Fritz, Evelyn; Schoch, Friederike; Schaller, Martin; Berger, Alexander C; Bitzer, Michael; Autenrieth, Ingo B

2010-10-01

Enterococci are commensal organisms in the alimentary tract. However, they can cause a variety of life-threatening infections, especially in nosocomial settings. We hypothesized that induction of cell death might enable these facultative pathogenic bacteria to evade the innate immune response and to cause infections of their host. We demonstrate that E. faecium when exposed to lysozyme induces cell death in macrophages in vitro and in vivo. Flow cytometric analyses of J774A.1 macrophages infected with E. faecium revealed loss of cell membrane integrity indicated by uptake of propidium iodide and decrease of the inner mitochondrial transmembrane potential DeltaPsi(m). Inhibition of caspases, treatment of macrophages with cytochalasin D, or rifampicin did not prevent cells from dying, suggesting cell death mechanisms that are independent of caspase activation, bacterial uptake, and intracellular bacterial replication. Characteristics of necrotic cell death were demonstrated by both lack of procaspase 3 activation and cell shrinkage, electron microscopy, and release of lactate dehydrogenase. Pretreatment of E. faecium with lysozyme and subsequently with broad spectrum protease considerably reduced cell death, suggesting that a bacterial surface protein is causative for cell death induction. Moreover, in a mouse peritonitis model we demonstrated that E. faecium induces cell death of peritoneal macrophages in vivo. Altogether, our results show that enterococci, under specific conditions such as exposure to lysozyme, induce necrotic cell death in macrophages, which might contribute to disseminated infections by these facultative pathogenic bacteria.

Modulating macrophage polarization with divalent cations in nanostructured titanium implant surfaces

NASA Astrophysics Data System (ADS)

Lee, Chung-Ho; Kim, Youn-Jeong; Jang, Je-Hee; Park, Jin-Woo

2016-02-01

Nanoscale topographical modification and surface chemistry alteration using bioactive ions are centrally important processes in the current design of the surface of titanium (Ti) bone implants with enhanced bone healing capacity. Macrophages play a central role in the early tissue healing stage and their activity in response to the implant surface is known to affect the subsequent healing outcome. Thus, the positive modulation of macrophage phenotype polarization (i.e. towards the regenerative M2 rather than the inflammatory M1 phenotype) with a modified surface is essential for the osteogenesis funtion of Ti bone implants. However, relatively few advances have been made in terms of modulating the macrophage-centered early healing capacity in the surface design of Ti bone implants for the two important surface properties of nanotopography and and bioactive ion chemistry. We investigated whether surface bioactive ion modification exerts a definite beneficial effect on inducing regenerative M2 macrophage polarization when combined with the surface nanotopography of Ti. Our results indicate that nanoscale topographical modification and surface bioactive ion chemistry can positively modulate the macrophage phenotype in a Ti implant surface. To the best of our knowledge, this is the first demonstration that chemical surface modification using divalent cations (Ca and Sr) dramatically induces the regenerative M2 macrophage phenotype of J774.A1 cells in nanostructured Ti surfaces. In this study, divalent cation chemistry regulated the cell shape of adherent macrophages and markedly up-regulated M2 macrophage phenotype expression when combined with the nanostructured Ti surface. These results provide insight into the surface engineering of future Ti bone implants that are harmonized between the macrophage-governed early wound healing process and subsequent mesenchymal stem cell-centered osteogenesis function.

45 CFR 77.4 - Remedial actions.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 45 Public Welfare 1 2011-10-01 2011-10-01 false Remedial actions. 77.4 Section 77.4 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION REMEDIAL ACTIONS APPLICABLE TO LETTER OF CREDIT ADMINISTRATION § 77.4 Remedial actions. If, after the conclusion of the procedures set forth in...

45 CFR 77.4 - Remedial actions.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 45 Public Welfare 1 2010-10-01 2010-10-01 false Remedial actions. 77.4 Section 77.4 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION REMEDIAL ACTIONS APPLICABLE TO LETTER OF CREDIT ADMINISTRATION § 77.4 Remedial actions. If, after the conclusion of the procedures set forth in...

7 CFR 774.22 - Loan closing.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Loan closing. 774.22 Section 774.22 Agriculture... SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.22 Loan closing. (a) Conditions. The... approval prior to closing. (b) Loan instruments and legal documents. The applicant will execute all loan...

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

PubMed

Kawakami, Tomoya; Koike, Atsushi; Amano, Fumio

2017-08-01

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

Use of Microsphere Technology for Targeted Delivery of Rifampin to Mycobacterium tuberculosis-Infected Macrophages

PubMed Central

Barrow, Esther L. W.; Winchester, Gary A.; Staas, Jay K.; Quenelle, Debra C.; Barrow, William W.

1998-01-01

Microsphere technology was used to develop formulations of rifampin for targeted delivery to host macrophages. These formulations were prepared by using biocompatible polymeric excipients of lactide and glycolide copolymers. Release characteristics were examined in vitro and also in two monocytic cell lines, the murine J774 and the human Mono Mac 6 cell lines. Bioassay assessment of cell culture supernatants from monocyte cell lines showed release of bioactive rifampin during a 7-day experimental period. Treatment of Mycobacterium tuberculosis H37Rv-infected monocyte cell lines with rifampin-loaded microspheres resulted in a significant decrease in numbers of CFU at 7 days following initial infection, even though only 8% of the microsphere-loaded rifampin was released. The levels of rifampin released from microsphere formulations within monocytes were more effective at reducing M. tuberculosis intracellular growth than equivalent doses of rifampin given as a free drug. These results demonstrate that rifampin-loaded microspheres can be formulated for effective sustained and targeted delivery to host macrophages. PMID:9756777

7 CFR 774.23 - Loan servicing.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Loan servicing. 774.23 Section 774.23 Agriculture... SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.23 Loan servicing. Loans will be serviced as a Non-program loan in accordance with 7 CFR part 766. If the loan is not repaid as agreed and...

YopJ-Induced Caspase-1 Activation in Yersinia-Infected Macrophages: Independent of Apoptosis, Linked to Necrosis, Dispensable for Innate Host Defense

PubMed Central

Zheng, Ying; Lilo, Sarit; Mena, Patricio; Bliska, James B.

2012-01-01

Yersinia outer protein J (YopJ) is a type III secretion system (T3SS) effector of pathogenic Yersinia (Yersinia pestis, Yersinia enterocolitica and Yersinia pseudotuberculosis) that is secreted into host cells. YopJ inhibits survival response pathways in macrophages, causing cell death. Allelic variation of YopJ is responsible for differential cytotoxicity in Yersinia strains. YopJ isoforms in Y. enterocolitica O:8 (YopP) and Y. pestis KIM (YopJKIM) strains have high cytotoxic activity. In addition, YopJKIM-induced macrophage death is associated with caspase-1 activation and interleukin-1β (IL-1β secretion. Here, the mechanism of YopJKIM-induced cell death, caspase-1 activation, and IL-1β secretion in primary murine macrophages was examined. Caspase-3/7 activity was low and the caspase-3 substrate poly (ADP-ribose) polymerase (PARP) was not cleaved in Y. pestis KIM5-infected macrophages. In addition, cytotoxicity and IL-1β secretion were not reduced in the presence of a caspase-8 inhibitor, or in B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax)/Bcl-2 homologous antagonist/killer (Bak) knockout macrophages, showing that YopJKIM-mediated cell death and caspase-1 activation occur independent of mitochondrial-directed apoptosis. KIM5-infected macrophages released high mobility group protein B1 (HMGB1), a marker of necrosis, and microscopic analysis revealed that necrotic cells contained active caspase-1, indicating that caspase-1 activation is associated with necrosis. Inhibitor studies showed that receptor interacting protein 1 (RIP1) kinase and reactive oxygen species (ROS) were not required for cytotoxicity or IL-β release in KIM5-infected macrophages. IL-1β secretion was reduced in the presence of cathepsin B inhibitors, suggesting that activation of caspase-1 requires cathepsin B activity. Ectopically-expressed YopP caused higher cytotoxicity and secretion of IL-1β in Y. pseudotuberculosis-infected macrophages than YopJKIM. Wild-type and congenic

Digital holographic microscopy as a technique to monitor macrophages infected by leishmania

NASA Astrophysics Data System (ADS)

Mendoza-Rodríguez, E.; Organista-Castelblanco, C.; Camacho, M.; Monroy-Ramírez, F.

2017-06-01

The Digital Holographic Microscopy in Transmission technique (DHM) is considered a useful tool in the noninvasive quantifying of transparent biological objects like living cells. In this work, we propose this technique to study and to monitor control macrophages infected by Leishmania (mouse lineJ774.A1). When the promastigotes enter in contact with healthy macrophages, they got phagocytosed and latterly confined in the formed parasitophorous vacuole. These processes change the morphology and density of the host macrophage. Both parameters can be measured in a label-free analysis of cells with the aid of the DHM technique. Our technique begins with the optical record of the holograms using a modified Mach-Zehnder interferometer and the reconstruction of the complex optical field transmitted by macrophages. In the latter point, we employ the angular spectrum algorithm. With the complex optical field reconstruction, we compute the field amplitude and the phase difference maps, which leads to describe one morphological characterization for the samples. Using phase difference maps is possible to measure internal variations for the integral refractive index, estimating the infection level of macrophages. Through the changes in the integral refractive index, it is also possible to describe and quantify in two different states the evolution of the infection. With these results some parameters of cells have been quantified, making the DHM technique a viable tool for diagnosis of biological samples under the presence of some pathogen.

Dragon (repulsive guidance molecule b) inhibits IL-6 expression in macrophages.

PubMed

Xia, Yin; Cortez-Retamozo, Virna; Niederkofler, Vera; Salie, Rishard; Chen, Shanzhuo; Samad, Tarek A; Hong, Charles C; Arber, Silvia; Vyas, Jatin M; Weissleder, Ralph; Pittet, Mikael J; Lin, Herbert Y

2011-02-01

Repulsive guidance molecule (RGM) family members RGMa, RGMb/Dragon, and RGMc/hemojuvelin were found recently to act as bone morphogenetic protein (BMP) coreceptors that enhance BMP signaling activity. Although our previous studies have shown that hemojuvelin regulates hepcidin expression and iron metabolism through the BMP pathway, the role of the BMP signaling mediated by Dragon remains largely unknown. We have shown previously that Dragon is expressed in neural cells, germ cells, and renal epithelial cells. In this study, we demonstrate that Dragon is highly expressed in macrophages. Studies with RAW264.7 and J774 macrophage cell lines reveal that Dragon negatively regulates IL-6 expression in a BMP ligand-dependent manner via the p38 MAPK and Erk1/2 pathways but not the Smad1/5/8 pathway. We also generated Dragon knockout mice and found that IL-6 is upregulated in macrophages and dendritic cells derived from whole lung tissue of these mice compared with that in respective cells derived from wild-type littermates. These results indicate that Dragon is an important negative regulator of IL-6 expression in immune cells and that Dragon-deficient mice may be a useful model for studying immune and inflammatory disorders.

Role of urease in megasome formation and Helicobacter pylori survival in macrophages

PubMed Central

Schwartz, Justin T.; Allen, Lee-Ann H.

2007-01-01

Previous studies have demonstrated that Helicobacter pylori (Hp) delays its entry into macrophages and persists inside megasomes, which are poorly acidified and accumulate early endosome autoantigen 1. Herein, we explored the role of Hp urease in bacterial survival in murine peritoneal macrophages and J774 cells. Plasmid-free mutagenesis was used to replace ureA and ureB with cat in Hp Strains 11637 and 11916. ureAB null Hp lacked detectable urease activity and did not express UreA or UreB as judged by immunoblotting. Deletion of ureAB had no effect on Hp binding to macrophages or the rate or extent of phagocytosis. However, intracellular survival of mutant organisms was impaired significantly. Immunofluorescence microscopy demonstrated that (in contrast to parental organisms) mutant Hp resided in single phagosomes, which were acidic and accumulated the lysosome marker lysosome-associated membrane protein-1 but not early endosome autoantigen 1. A similar phenotype was observed for spontaneous urease mutants derived from Hp Strain 60190. Treatment of macrophages with bafilomycin A1, NH4Cl, or chloroquine prevented acidification of phagosomes containing mutant Hp. However, only ammonium chloride enhanced bacterial viability significantly. Rescue of ureAB null organisms was also achieved by surface adsorption of active urease. Altogether, our data indicate a role for urease and urease-derived ammonia in megasome formation and Hp survival. PMID:16543403

Reduced expression of IL-12 p35 by SJL/J macrophages responding to Theiler's virus infection is associated with constitutive activation of IRF-3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dahlberg, Angela; Auble, Mark R.; Petro, Thomas M.

2006-09-30

Macrophages responding to viral infections may contribute to autoimmune demyelinating diseases (ADD). Macrophages from ADD-susceptible SJL/J mice responding to Theiler's Virus (TMEV) infection, the TLR7 agonist loxoribine, or the TLR4 agonist-LPS expressed less IL-12 p35 but more IL-12/23 p40 and IFN-{beta} than macrophages from ADD-resistant B10.S mice. While expression of IRF-1 and -7 was similar between B10.S and SJL/J TMEV-infected macrophages, SJL/J but not B10.S macrophages exhibited constitutively active IRF-3. In contrast to overexpressed IRF-1, IRF-5, and IRF-7, which stimulated p35 promoter reporter activity, overexpressed IRF-3 repressed p35 promoter activity in response to TMEV infection, loxoribine, IFN-{gamma}/LPS, but not IFN-{gamma}more » alone. IRF-3 lessened but did not eliminate IRF-1-stimulated p35 promoter activity. Repression by IRF-3 required bp -172 to -122 of the p35 promoter. The data suggest that pre-activated IRF-3 is a major factor in the differences in IL-12 production between B10.S and SJL/J macrophages responding to TMEV.« less

Rho is Required for the Initiation of Calcium Signaling and Phagocytosis by Fcγ Receptors in Macrophages

PubMed Central

Hackam, David J.; Rotstein, Ori D.; Schreiber, Alan; Zhang, Wei-jian; Grinstein, Sergio

1997-01-01

Phagocytosis of bacteria by macrophages and neutrophils is an essential component of host defense against infection. The mechanism whereby the interaction of opsonized particles with Fcγ receptors triggers the engulfment of opsonized particles remains incompletely understood, although activation of tyrosine kinases has been recognized as an early step. Recent studies in other systems have demonstrated that tyrosine kinases can in turn signal the activation of small GTPases of the ras superfamily. We therefore investigated the possible role of Rho in Fc receptor–mediated phagocytosis. To this end we microinjected J774 macrophages with C3 exotoxin from Clostridium botulinum, which ADP-ribosylates and inactivates Rho. C3 exotoxin induced the retraction of filopodia, the disappearance of focal complexes, and a global decrease in the F-actin content of J774 cells. In addition, these cells exhibited increased spreading and the formation of vacuolar structures. Importantly, inactivation of Rho resulted in the complete abrogation of phagocytosis. Inhibition of Fcγ receptor–mediated phagocytosis by C3 exotoxin was confirmed in COS cells, which become phagocytic upon transfection of the FcγRIIA receptor. Rho was found to be essential for the accumulation of phosphotyrosine and of F-actin around phagocytic cups and for Fcγ receptor–mediated Ca2+ signaling. The clustering of receptors in response to opsonin, an essential step in Fcγ-induced signaling, was the earliest event shown to be inhibited by C3 exotoxin. The effect of the toxin was specific, since clustering and internalization of transferrin receptors were unaffected by microinjection of C3. These data identify a role for small GTPases in Fcγ receptor–mediated phagocytosis by leukocytes. PMID:9294149

48 CFR 731.774 - Overseas recruitment incentive.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 5 2010-10-01 2010-10-01 false Overseas recruitment incentive. 731.774 Section 731.774 Federal Acquisition Regulations System AGENCY FOR INTERNATIONAL... Organizations 731.774 Overseas recruitment incentive. USAID's policies regarding overseas recruitment incentives...

Macrophage activation by heparanase is mediated by TLR-2 and TLR-4 and associates with plaque progression.

PubMed

Blich, Miry; Golan, Amnon; Arvatz, Gil; Sebbag, Anat; Shafat, Itay; Sabo, Edmond; Cohen-Kaplan, Victoria; Petcherski, Sirouch; Avniel-Polak, Shani; Eitan, Amnon; Hammerman, Haim; Aronson, Doron; Axelman, Elena; Ilan, Neta; Nussbaum, Gabriel; Vlodavsky, Israel

2013-02-01

Factors and mechanisms that activate macrophages in atherosclerotic plaques are incompletely understood. We examined the capacity of heparanase to activate macrophages. Highly purified heparanase was added to mouse peritoneal macrophages and macrophage-like J774 cells, and the levels of tumor necrosis factor-α, matrix metalloproteinase-9, interlukin-1, and monocyte chemotactic protein-1 were evaluated by ELISA. Gene expression was determined by RT-PCR. Cells collected from Toll-like receptor-2 and Toll-like receptor-4 knockout mice were evaluated similarly. Heparanase levels in the plasma of patients with acute myocardial infarction, stable angina, and healthy subjects were determined by ELISA. Immunohistochemistry was applied to detect the expression of heparanase in control specimens and specimens of patients with stable angina or acute myocardial infarction. Addition or overexpression of heparanase variants resulted in marked increase in tumor necrosis factor-α, matrix metalloproteinase-9, interlukin-1, and monocyte chemotactic protein-1 levels. Mouse peritoneal macrophages harvested from Toll-like receptor-2 or Toll-like receptor-4 knockout mice were not activated by heparanase. Plasma heparanase level was higher in patients with acute myocardial infarction, compared with patients with stable angina and healthy subjects. Pathologic coronary specimens obtained from vulnerable plaques showed increased heparanase staining compared with specimens of stable plaque and controls. Heparanase activates macrophages, resulting in marked induction of cytokine expression associated with plaque progression toward vulnerability.

Phagocytosis of Advanced Glycation End Products (AGEs) in Macrophages Induces Cell Apoptosis.

PubMed

Gao, Yuan; Wake, Hidenori; Morioka, Yuta; Liu, Keyue; Teshigawara, Kiyoshi; Shibuya, Megumi; Zhou, Jingxiu; Mori, Shuji; Takahashi, Hideo; Nishibori, Masahiro

2017-01-01

Advanced glycation end products (AGEs) are the products of a series of nonenzymatic modifications of proteins by reducing sugars. AGEs play a pivotal role in development of diabetic complications and atherosclerosis. Accumulation of AGEs in a vessel wall may contribute to the development of vascular lesions. Although AGEs have a diverse range of bioactivities, the clearance process of AGEs from the extracellular space, including the incorporation of AGEs into specific cells, subcellular localization, and the fate of AGEs, remains unclear. In the present study, we examined the kinetics of the uptake of AGEs by mouse macrophage J774.1 cells in vitro and characterized the process. We demonstrated that AGEs bound to the surface of the cells and were also incorporated into the cytoplasm. The temperature- and time-dependent uptake of AGEs was saturable with AGE concentration and was inhibited by cytochalasin D but not chlorpromazine. We also observed the granule-like appearance of AGE immunoreactivity in subcellular localizations in macrophages. Higher concentrations of AGEs induced intracellular ROS and 4-HNE, which were associated with activation of the NF- κ B pathway and caspase-3. These results suggest that incorporation of AGEs occurred actively by endocytosis in macrophages, leading to apoptosis of these cells through NF- κ B activation.

Phagocytosis of Advanced Glycation End Products (AGEs) in Macrophages Induces Cell Apoptosis

PubMed Central

Wake, Hidenori; Morioka, Yuta; Liu, Keyue; Shibuya, Megumi; Zhou, Jingxiu; Mori, Shuji; Takahashi, Hideo

2017-01-01

Advanced glycation end products (AGEs) are the products of a series of nonenzymatic modifications of proteins by reducing sugars. AGEs play a pivotal role in development of diabetic complications and atherosclerosis. Accumulation of AGEs in a vessel wall may contribute to the development of vascular lesions. Although AGEs have a diverse range of bioactivities, the clearance process of AGEs from the extracellular space, including the incorporation of AGEs into specific cells, subcellular localization, and the fate of AGEs, remains unclear. In the present study, we examined the kinetics of the uptake of AGEs by mouse macrophage J774.1 cells in vitro and characterized the process. We demonstrated that AGEs bound to the surface of the cells and were also incorporated into the cytoplasm. The temperature- and time-dependent uptake of AGEs was saturable with AGE concentration and was inhibited by cytochalasin D but not chlorpromazine. We also observed the granule-like appearance of AGE immunoreactivity in subcellular localizations in macrophages. Higher concentrations of AGEs induced intracellular ROS and 4-HNE, which were associated with activation of the NF-κB pathway and caspase-3. These results suggest that incorporation of AGEs occurred actively by endocytosis in macrophages, leading to apoptosis of these cells through NF-κB activation. PMID:29430285

Agglomeration, sedimentation, and cellular toxicity of alumina nanoparticles in cell culture medium

NASA Astrophysics Data System (ADS)

Yoon, Dokyung; Woo, Daekwang; Kim, Jung Heon; Kim, Moon Ki; Kim, Taesung; Hwang, Eung-Soo; Baik, Seunghyun

2011-06-01

The cytotoxicity of alumina nanoparticles (NPs) was investigated for a wide range of concentration (25-200 μg/mL) and incubation time (0-72 h) using floating cells (THP-1) and adherent cells (J774A.1, A549, and 293). Alumina NPs were gradually agglomerated over time although a significant portion of sedimentation occurred at the early stage within 6 h. A decrease of the viability was found in floating (THP-1) and adherent (J774A.1 and A549) cells in a dose-dependent manner. However, the time-dependent decrease in cell viability was observed only in adherent cells (J774A.1 and A549), which is predominantly related with the sedimentation of alumina NPs in cell culture medium. The uptake of alumina NPs in macrophages and an increased cell-to-cell adhesion in adherent cells were observed. There was no significant change in the viability of 293 cells. This in vitro test suggests that the agglomeration and sedimentation of alumina NPs affected cellular viability depending on cell types such as monocytes (THP-1), macrophages (J774A.1), lung carcinoma cells (A549), and embryonic kidney cells (293).

Contribution of macrophages in the contrast loss in iron oxide-based MRI cancer cell tracking studies

PubMed Central

Danhier, Pierre; Deumer, Gladys; Joudiou, Nicolas; Bouzin, Caroline; Levêque, Philippe; Haufroid, Vincent; Jordan, Bénédicte F.; Feron, Olivier; Sonveaux, Pierre; Gallez, Bernard

2017-01-01

Magnetic resonance imaging (MRI) cell tracking of cancer cells labeled with superparamagnetic iron oxides (SPIO) allows visualizing metastatic cells in preclinical models. However, previous works showed that the signal void induced by SPIO on T2(*)-weighted images decreased over time. Here, we aim at characterizing the fate of iron oxide nanoparticles used in cell tracking studies and the role of macrophages in SPIO metabolism. In vivo MRI cell tracking of SPIO positive 4T1 breast cancer cells revealed a quick loss of T2* contrast after injection. We next took advantage of electron paramagnetic resonance (EPR) spectroscopy and inductively coupled plasma mass spectroscopy (ICP-MS) for characterizing the evolution of superparamagnetic and non-superparamagnetic iron pools in 4T1 breast cancer cells and J774 macrophages after SPIO labeling. These in vitro experiments and histology studies performed on 4T1 tumors highlighted the quick degradation of iron oxides by macrophages in SPIO-based cell tracking experiments. In conclusion, the release of SPIO by dying cancer cells and the subsequent uptake of iron oxides by tumor macrophages are limiting factors in MRI cell tracking experiments that plead for the use of (MR) reporter-gene based imaging methods for the long-term tracking of metastatic cells. PMID:28467814

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

PubMed

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

2018-01-01

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

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

PubMed

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

2013-01-01

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

A Nitric Oxide Storage and Transport System That Protects Activated Macrophages from Endogenous Nitric Oxide Cytotoxicity*

PubMed Central

Lok, Hiu Chuen; Sahni, Sumit; Jansson, Patric J.; Kovacevic, Zaklina; Hawkins, Clare L.; Richardson, Des R.

2016-01-01

Nitric oxide (NO) is integral to macrophage cytotoxicity against tumors due to its ability to induce iron release from cancer cells. However, the mechanism for how activated macrophages protect themselves from endogenous NO remains unknown. We previously demonstrated by using tumor cells that glutathione S-transferase P1 (GSTP1) sequesters NO as dinitrosyl-dithiol iron complexes (DNICs) and inhibits NO-mediated iron release from cells via the transporter multidrug resistance protein 1 (MRP1/ABCC1). These prior studies also showed that MRP1 and GSTP1 protect tumor cells against NO cytotoxicity, which parallels their roles in defending cancer cells from cytotoxic drugs. Considering this, and because GSTP1 and MRP1 are up-regulated during macrophage activation, this investigation examined whether this NO storage/transport system protects macrophages against endogenous NO cytotoxicity in two well characterized macrophage cell types (J774 and RAW 264.7). MRP1 expression markedly increased upon macrophage activation, and the role of MRP1 in NO-induced 59Fe release was demonstrated by Mrp1 siRNA and the MRP1 inhibitor, MK571, which inhibited NO-mediated iron efflux. Furthermore, Mrp1 silencing increased DNIC accumulation in macrophages, indicating a role for MRP1 in transporting DNICs out of cells. In addition, macrophage 59Fe release was enhanced by silencing Gstp1, suggesting GSTP1 was responsible for DNIC binding/storage. Viability studies demonstrated that GSTP1 and MRP1 protect activated macrophages from NO cytotoxicity. This was confirmed by silencing nuclear factor-erythroid 2-related factor 2 (Nrf2), which decreased MRP1 and GSTP1 expression, concomitant with reduced 59Fe release and macrophage survival. Together, these results demonstrate a mechanism by which macrophages protect themselves against NO cytotoxicity. PMID:27866158

CCAAT/enhancer-binding protein beta inhibits proliferation in monocytic cells by affecting the retinoblastoma protein/E2F/cyclin E pathway but is not directly required for macrophage morphology.

PubMed

Gutsch, Romina; Kandemir, Judith D; Pietsch, Daniel; Cappello, Christian; Meyer, Johann; Simanowski, Kathrin; Huber, René; Brand, Korbinian

2011-07-01

Monocytic differentiation is orchestrated by complex networks that are not fully understood. This study further elucidates the involvement of transcription factor CCAAT/enhancer-binding protein β (C/EBPβ). Initially, we demonstrated a marked increase in nuclear C/EBPβ-liver-enriched activating protein* (LAP*)/liver-enriched activating protein (LAP) levels and LAP/liver-enriched inhibiting protein (LIP) ratios in phorbol 12-myristate 13-acetate (PMA)-treated differentiating THP-1 premonocytic cells accompanied by reduced proliferation. To directly study C/EBPβ effects on monocytic cells, we generated novel THP-1-derived (low endogenous C/EBPβ) cell lines stably overexpressing C/EBPβ isoforms. Most importantly, cells predominantly overexpressing LAP* (C/EBPβ-long), but not those overexpressing LIP (C/EBPβ-short), exhibited a reduced proliferation, with no effect on morphology. PMA-induced inhibition of proliferation was attenuated in C/EBPβ-short cells. In C/EBPβ(WT) macrophage-like cells (high endogenous C/EBPβ), we measured a reduced proliferation/cycling index compared with C/EBPβ(KO). The typical macrophage morphology was only observed in C/EBPβ(WT), whereas C/EBPβ(KO) stayed round. C/EBPα did not compensate for C/EBPβ effects on proliferation/morphology. Serum reduction, an independent approach known to inhibit proliferation, induced macrophage morphology in C/EBPβ(KO) macrophage-like cells but not THP-1. In PMA-treated THP-1 and C/EBPβ-long cells, a reduced phosphorylation of cell cycle repressor retinoblastoma was found. In addition, C/EBPβ-long cells showed reduced c-Myc expression accompanied by increased CDK inhibitor p27 and reduced cyclin D1 levels. Finally, C/EBPβ-long and C/EBPβ(WT) cells exhibited low E2F1 and cyclin E levels, and C/EBPβ overexpression was found to inhibit cyclin E1 promoter-dependent transcription. Our results suggest that C/EBPβ reduces monocytic proliferation by affecting the retinoblastoma/E2F/cyclin E

Testing Nucleoside Analogues as Inhibitors of Bacillus anthracis Spore Germination In Vitro and in Macrophage Cell Culture ▿

PubMed Central

Alvarez, Zadkiel; Lee, Kyungae; Abel-Santos, Ernesto

2010-01-01

Bacillus anthracis, the etiological agent of anthrax, has a dormant stage in its life cycle known as the endospore. When conditions become favorable, spores germinate and transform into vegetative bacteria. In inhalational anthrax, the most fatal manifestation of the disease, spores enter the organism through the respiratory tract and germinate in phagosomes of alveolar macrophages. Germinated cells can then produce toxins and establish infection. Thus, germination is a crucial step for the initiation of pathogenesis. B. anthracis spore germination is activated by a wide variety of amino acids and purine nucleosides. Inosine and l-alanine are the two most potent nutrient germinants in vitro. Recent studies have shown that germination can be hindered by isomers or structural analogues of germinants. 6-Thioguanosine (6-TG), a guanosine analogue, is able to inhibit germination and prevent B. anthracis toxin-mediated necrosis in murine macrophages. In this study, we screened 46 different nucleoside analogues as activators or inhibitors of B. anthracis spore germination in vitro. These compounds were also tested for their ability to protect the macrophage cell line J774a.1 from B. anthracis cytotoxicity. Structure-activity relationship analysis of activators and inhibitors clarified the binding mechanisms of nucleosides to B. anthracis spores. In contrast, no structure-activity relationships were apparent for compounds that protected macrophages from B. anthracis-mediated killing. However, multiple inhibitors additively protected macrophages from B. anthracis. PMID:20921305

49 CFR 236.774 - Movement, facing.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 49 Transportation 4 2010-10-01 2010-10-01 false Movement, facing. 236.774 Section 236.774 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... Movement, facing. The movement of a train over the points of a switch which face in a direction opposite to...

36 CFR 7.74 - Virgin Islands National Park.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Virgin Islands National Park. 7.74 Section 7.74 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR SPECIAL REGULATIONS, AREAS OF THE NATIONAL PARK SYSTEM § 7.74 Virgin Islands National Park. (a...

36 CFR 7.74 - Virgin Islands National Park.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Virgin Islands National Park. 7.74 Section 7.74 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR SPECIAL REGULATIONS, AREAS OF THE NATIONAL PARK SYSTEM § 7.74 Virgin Islands National Park. (a...

36 CFR 7.74 - Virgin Islands National Park.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Virgin Islands National Park. 7.74 Section 7.74 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR SPECIAL REGULATIONS, AREAS OF THE NATIONAL PARK SYSTEM § 7.74 Virgin Islands National Park. (a...

Regulation of adhesion behavior of murine macrophage using supported lipid membranes displaying tunable mannose domains

NASA Astrophysics Data System (ADS)

Kaindl, T.; Oelke, J.; Pasc, A.; Kaufmann, S.; Konovalov, O. V.; Funari, S. S.; Engel, U.; Wixforth, A.; Tanaka, M.

2010-07-01

Highly uniform, strongly correlated domains of synthetically designed lipids can be incorporated into supported lipid membranes. The systematic characterization of membranes displaying a variety of domains revealed that the equilibrium size of domains significantly depends on the length of fluorocarbon chains, which can be quantitatively interpreted within the framework of an equivalent dipole model. A mono-dispersive, narrow size distribution of the domains enables us to treat the inter-domain correlations as two-dimensional colloidal crystallization and calculate the potentials of mean force. The obtained results demonstrated that both size and inter-domain correlation can precisely be controlled by the molecular structures. By coupling α-D-mannose to lipid head groups, we studied the adhesion behavior of the murine macrophage (J774A.1) on supported membranes. Specific adhesion and spreading of macrophages showed a clear dependence on the density of functional lipids. The obtained results suggest that such synthetic lipid domains can be used as a defined platform to study how cells sense the size and distribution of functional molecules during adhesion and spreading.

Evidence for a gamma-interferon receptor that regulates macrophage tumoricidal activity

PubMed Central

1984-01-01

Gamma-interferon (IFN-gamma) is the macrophage-activating factor (MAF) produced by normal murine splenic cells and the murine T cell hybridoma 24/G1 that induces nonspecific tumoricidal activity in macrophages. Incubation of 24/G1 supernatants diluted to 8.3 IRU IFN-gamma/ml with 6 X 10(6) elicited peritoneal macrophages or bone marrow-derived macrophages for 4 h at 37 degrees C, resulted in removal of 80% of the MAF activity from the lymphokine preparation. Loss of activity appeared to result from absorption and not consumption because (a) 40% of the activity was removed after exposure to macrophage for 30 min at 4 degrees C, (b) no reduction of MAF activity was detected when the 24/G1 supernatant was incubated with macrophage culture supernatants, and (c) macrophage-treated supernatants showed a selective loss of MAF activity but not interleukin 2 (IL-2) activity. Absorption was dependent on the input of either IFN-gamma or macrophages and was time dependent at 37 degrees C but not at 4 degrees C. With four rodent species tested, absorption of murine IFN-gamma displayed species specificity. However, cultured human peripheral blood monocytes and the human histiocytic lymphoma cell line U937 were able to absorb the murine lymphokine. Although the majority of murine cell lines tested absorbed 24/G1 MAF activity, two murine macrophage cell lines, P388D1 and J774, were identified which absorbed significantly reduced amounts of natural IFN- gamma. Purified murine recombinant IFN-gamma was absorbed by elicited macrophages but not by P388D1. Normal macrophages but not P388D1 bound fluoresceinated microspheres coated with recombinant IFN-gamma and binding was inhibited by pretreatment of the normal cells with 24/G1 supernatants. Scatchard plot analysis showed that 12,000 molecules of soluble 125I-recombinant IFN-gamma bound per bone marrow macrophage with a Ka of 0.9 X 10(8) M-1. Binding was quantitatively inhibitable by natural IFN-gamma but not by murine IFN alpha. IFN

6-Mercaptopurine reduces macrophage activation and gut epithelium proliferation through inhibition of GTPase Rac1.

PubMed

Marinković, Goran; Hamers, Anouk A J; de Vries, Carlie J M; de Waard, Vivian

2014-09-01

Inflammatory bowel disease is characterized by chronic intestinal inflammation. Azathioprine and its metabolite 6-mercaptopurine (6-MP) are effective immunosuppressive drugs that are widely used in patients with inflammatory bowel disease. However, established understanding of their immunosuppressive mechanism is limited. Azathioprine and 6-MP have been shown to affect small GTPase Rac1 in T cells and endothelial cells, whereas the effect on macrophages and gut epithelial cells is unknown. Macrophages (RAW cells) and gut epithelial cells (Caco-2 cells) were activated by cytokines and the effect on Rac1 signaling was assessed in the presence or absence of 6-MP. Rac1 is activated in macrophages and epithelial cells, and treatment with 6-MP resulted in Rac1 inhibition. In macrophages, interferon-γ induced downstream signaling through c-Jun-N-terminal Kinase (JNK) resulting in inducible nitric oxide synthase (iNOS) expression. iNOS expression was reduced by 6-MP in a Rac1-dependent manner. In epithelial cells, 6-MP efficiently inhibited tumor necrosis factor-α-induced expression of the chemokines CCL2 and interleukin-8, although only interleukin-8 expression was inhibited in a Rac1-dependent manner. In addition, activation of the transcription factor STAT3 was suppressed in a Rac1-dependent fashion by 6-MP, resulting in reduced proliferation of the epithelial cells due to diminished cyclin D1 expression. These data demonstrate that 6-MP affects macrophages and gut epithelial cells beneficially, in addition to T cells and endothelial cells. Furthermore, mechanistic insight is provided to support development of Rac1-specific inhibitors for clinical use in inflammatory bowel disease.

Multi-walled carbon nanotubes injure the plasma membrane of macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hirano, Seishiro; Kanno, Sanae; Furuyama, Akiko

2008-10-15

Carbon nanotubes (CNTs) are emerging nanotechnology materials which are likely to be mass-produced in the near future. However, prior to mass-production, certain health-related concerns should first be addressed. For example, when inhaled, the thin-fibrous shape and the biopersistent characteristics of CNTs may cause pulmonary diseases, in a manner similar to asbestos. In the present study, mouse macrophages (J774.1) were exposed to highly-purified multi-walled CNTs (MWCNTs, 67 nm) or to UICC crocidolite in order to evaluate the toxicity of these nano-size fibers. The cytotoxicity of MWCNTs was found to be higher than that of crocidolite. The toxic effect of MWCNTs wasmore » not affected by N-acetylcysteine, an antioxidant, or buthionine sulfoximine, a glutathione synthesis inhibitor. cDNA microarray analyses suggested that the cytotoxicity of MWCNTs could not be explained satisfactorily by either an increase or decrease of gene expression, although mRNA levels of some cytokines were slightly increased by MWCNTs. Moreover, MWCNTs did not significantly activate either MAP kinases such as ERK, JNK and p38, nor common apoptosis pathways such as caspase 3 and PARP. Electron microscopic studies indicated that MWCNTs associate with the plasma membrane of macrophages and disrupt the integrity of the membrane. Several proteins were found to adsorb onto MWCNTs when MWCNT-exposed macrophages were gently lysed. One of these proteins was macrophage receptor with collagenous structure (MARCO). MARCO-transfected CHO-K1 cells associated with MWCNTs more rapidly than mock-transfected cells. These results indicate that MWCNTs probably trigger cytotoxic effects in phagocytotic cells by reacting with MARCO on the plasma membrane and rupturing the plasma membrane.« less

IL-4 directly signals tissue-resident macrophages to proliferate beyond homeostatic levels controlled by CSF-1

PubMed Central

Ruckerl, Dominik; Thomas, Graham D.; Hewitson, James P.; Duncan, Sheelagh; Brombacher, Frank; Maizels, Rick M.; Hume, David A.; Allen, Judith E.

2013-01-01

Macrophages (MΦs) colonize tissues during inflammation in two distinct ways: recruitment of monocyte precursors and proliferation of resident cells. We recently revealed a major role for IL-4 in the proliferative expansion of resident MΦs during a Th2-biased tissue nematode infection. We now show that proliferation of MΦs during intestinal as well as tissue nematode infection is restricted to sites of IL-4 production and requires MΦ-intrinsic IL-4R signaling. However, both IL-4Rα–dependent and –independent mechanisms contributed to MΦ proliferation during nematode infections. IL-4R–independent proliferation was controlled by a rise in local CSF-1 levels, but IL-4Rα expression conferred a competitive advantage with higher and more sustained proliferation and increased accumulation of IL-4Rα+ compared with IL-4Rα− cells. Mechanistically, this occurred by conversion of IL-4Rα+ MΦs from a CSF-1–dependent to –independent program of proliferation. Thus, IL-4 increases the relative density of tissue MΦs by overcoming the constraints mediated by the availability of CSF-1. Finally, although both elevated CSF1R and IL-4Rα signaling triggered proliferation above homeostatic levels, only CSF-1 led to the recruitment of monocytes and neutrophils. Thus, the IL-4 pathway of proliferation may have developed as an alternative to CSF-1 to increase resident MΦ numbers without coincident monocyte recruitment. PMID:24101381

IL-4 directly signals tissue-resident macrophages to proliferate beyond homeostatic levels controlled by CSF-1.

PubMed

Jenkins, Stephen J; Ruckerl, Dominik; Thomas, Graham D; Hewitson, James P; Duncan, Sheelagh; Brombacher, Frank; Maizels, Rick M; Hume, David A; Allen, Judith E

2013-10-21

Macrophages (MΦs) colonize tissues during inflammation in two distinct ways: recruitment of monocyte precursors and proliferation of resident cells. We recently revealed a major role for IL-4 in the proliferative expansion of resident MΦs during a Th2-biased tissue nematode infection. We now show that proliferation of MΦs during intestinal as well as tissue nematode infection is restricted to sites of IL-4 production and requires MΦ-intrinsic IL-4R signaling. However, both IL-4Rα-dependent and -independent mechanisms contributed to MΦ proliferation during nematode infections. IL-4R-independent proliferation was controlled by a rise in local CSF-1 levels, but IL-4Rα expression conferred a competitive advantage with higher and more sustained proliferation and increased accumulation of IL-4Rα(+) compared with IL-4Rα(-) cells. Mechanistically, this occurred by conversion of IL-4Rα(+) MΦs from a CSF-1-dependent to -independent program of proliferation. Thus, IL-4 increases the relative density of tissue MΦs by overcoming the constraints mediated by the availability of CSF-1. Finally, although both elevated CSF1R and IL-4Rα signaling triggered proliferation above homeostatic levels, only CSF-1 led to the recruitment of monocytes and neutrophils. Thus, the IL-4 pathway of proliferation may have developed as an alternative to CSF-1 to increase resident MΦ numbers without coincident monocyte recruitment.

A novel activity for substance P: stimulation of peroxisome proliferator-activated receptor-γ protein expression in human monocytes and macrophages

PubMed Central

Amoruso, A; Bardelli, C; Gunella, G; Ribichini, F; Brunelleschi, S

2008-01-01

Background and purpose: Substance P (SP) and peroxisome proliferator-activated receptor-γ (PPAR-γ) play important roles in different inflammatory conditions and are both expressed in human monocytes and macrophages. However, it is not known whether or not they interact. This study was undertaken to evaluate the effects of SP on PPAR-γ protein expression in monocytes and macrophages (MDMs: monocyte-derived macrophages) from healthy smokers and non-smokers. Experimental approach: PPAR-γ protein was detected by western blot and quantified by calculating the ratio between PPAR-γ and β-actin protein expression. Constitutive tachykinin NK1 receptor expression in monocytes and MDMs from healthy smokers and non-smokers was evaluated by western blot. Cytokine release was evaluated by ELISA. Key results: In the concentration range 10−10–10−6 M, SP stimulated PPAR-γ protein expression in monocytes and MDMs, being more effective in cells from healthy smokers. Moreover, in these cells there was a constitutively increased expression of NK1 receptors. SP-induced expression of the PPAR-γ protein was receptor-mediated, as it was reproduced by the NK1 selective agonist [Sar9Met(O2)11]SP and reversed by the competitive NK1 antagonist GR71251. SP-induced maximal effects were similar to those evoked by 15-deoxy-Δ12,14-prostaglandin J2; an endogenous PPAR-γ agonist, and were significantly reduced by a PPAR-γ antagonist. NK1 and PPAR-γ agonists exerted opposite effects on TNF-α release from monocytes and MDMs. Conclusions and implications: Enhancement of PPAR-γ protein expression represents a novel activity for SP, which could contribute to a range of chronic inflammatory disorders. PMID:18278062

PKC412 (CGP41251) modulates the proliferation and lipopolysaccharide-induced inflammatory responses of RAW 264.7 macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyatake, Katsutoshi; Institute for Genome Research, The University of Tokushima, Tokushima; Inoue, Hiroshi

2007-08-17

PKC412 (CGP41251) is a multitarget protein kinase inhibitor with anti-tumor activities. Here, we investigated the effects of PKC412 on macrophages. PKC412 inhibited the proliferation of murine RAW 264.7 macrophages through induction of G2/M cell cycle arrest and apoptosis. At non-toxic drug concentrations, PKC412 significantly suppressed the lipopolysaccharide (LPS)-induced release of TNF-{alpha} and nitric oxide, while instead enhancing IL-6 secretion. PKC412 attenuated LPS-induced phosphorylations of MKK4 and JNK, as well as AP-1 DNA binding activities. Furthermore, PKC412 suppressed LPS-induced Akt and GSK-3{beta} phosphorylations. These results suggest that the anti-proliferative and immunomodulatory effects of PKC412 are, at least in part, mediated throughmore » its interference with the MKK4/JNK/AP-1 and/or Akt/GSK-3{beta} pathways. Since macrophages contribute significantly to the development of both acute and chronic inflammation, PKC412 may have therapeutic potential and applications in treating inflammatory and/or autoimmune diseases.« less

48 CFR 731.774 - Overseas recruitment incentive.

Code of Federal Regulations, 2011 CFR

2011-10-01

... incentive. 731.774 Section 731.774 Federal Acquisition Regulations System AGENCY FOR INTERNATIONAL DEVELOPMENT GENERAL CONTRACTING REQUIREMENTS CONTRACT COST PRINCIPLES AND PROCEDURES Contracts With Nonprofit... are set forth in AIDAR 731.205-70. These policies are also applicable to contracts with a nonprofit...

Genetic and Phenotypic Characterization of a Salmonella enterica serovar Enteritidis Emerging Strain with Superior Intra-macrophage Replication Phenotype

PubMed Central

Shomer, Inna; Avisar, Alon; Desai, Prerak; Azriel, Shalhevet; Smollan, Gill; Belausov, Natasha; Keller, Nathan; Glikman, Daniel; Maor, Yasmin; Peretz, Avi; McClelland, Michael; Rahav, Galia; Gal-Mor, Ohad

2016-01-01

Salmonella enterica serovar Enteritidis (S. Enteritidis) is one of the ubiquitous Salmonella serovars worldwide and a major cause of food-born outbreaks, which are often associated with poultry and poultry derivatives. Here we report a nation-wide S. Enteritidis clonal outbreak that occurred in Israel during the last third of 2015. Pulsed field gel electrophoresis and whole genome sequencing identified genetically related strains that were circulating in Israel as early as 2008. Global comparison linked this outbreak strain to several clinical and marine environmental isolates that were previously isolated in California and Canada, indicating that similar strains are prevalent outside of Israel. Phenotypic comparison between the 2015 outbreak strain and other clinical and reference S. Enteritidis strains showed only limited intra-serovar phenotypic variation in growth in rich medium, invasion into Caco-2 cells, uptake by J774.1A macrophages, and host cell cytotoxicity. In contrast, significant phenotypic variation was shown among different S. Enteritidis isolates when biofilm-formation, motility, invasion into HeLa cells and uptake by THP-1 human macrophages were studied. Interestingly, the 2015 outbreak clone was found to possess superior intra-macrophage replication ability within both murine and human macrophages in comparison to the other S. Enteritidis strains studied. This phenotype is likely to play a role in the virulence and host-pathogen interactions of this emerging clone. PMID:27695450

Pancreatic Cancer Cell Exosome-Mediated Macrophage Reprogramming and the Role of MicroRNAs 155 and 125b2 Transfection using Nanoparticle Delivery Systems

PubMed Central

Su, Mei-Ju; Aldawsari, Hibah; Amiji, Mansoor

2016-01-01

Exosomes are nano-sized endosome-derived small intraluminal vesicles, which are important facilitators of intercellular communication by transporting contents, such as protein, mRNA, and microRNAs, between neighboring cells, such as in the tumor microenvironment. The purpose of this study was to understand the mechanisms of exosomes-mediated cellular communication between human pancreatic cancer (Panc-1) cells and macrophages (J771.A1) using a Transwell co-culture system. Following characterization of exosome-mediated cellular communication and pro-tumoral baseline M2 macrophage polarization, the Panc-1 cells were transfected with microRNA-155 (miR-155) and microRNA-125b-2 (miR-125b2) expressing plasmid DNA using hyaluronic acid-poly(ethylene imine)/hyaluronic acid-poly(ethylene glycol) (HA-PEI/HA-PEG) self-assembling nanoparticle-based non-viral vectors. Our results show that upon successful transfection of Panc-1 cells, the exosome content was altered leading to differential communication and reprogramming of the J774.A1 cells to an M1 phenotype. Based on these results, genetic therapies targeted towards selective manipulation of tumor cell-derived exosome content may be very promising for cancer therapy. PMID:27443190

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

PubMed

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

2016-02-01

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

Novel Keto-phospholipids Are Generated by Monocytes and Macrophages, Detected in Cystic Fibrosis, and Activate Peroxisome Proliferator-activated Receptor-γ*

PubMed Central

Hammond, Victoria J.; Morgan, Alwena H.; Lauder, Sarah; Thomas, Christopher P.; Brown, Sarah; Freeman, Bruce A.; Lloyd, Clare M.; Davies, Jane; Bush, Andrew; Levonen, Anna-Liisa; Kansanen, Emilia; Villacorta, Luis; Chen, Y. Eugene; Porter, Ned; Garcia-Diaz, Yoel M.; Schopfer, Francisco J.; O'Donnell, Valerie B.

2012-01-01

12/15-Lipoxygenases (LOXs) in monocytes and macrophages generate novel phospholipid-esterified eicosanoids. Here, we report the generation of two additional families of related lipids comprising 15-ketoeicosatetraenoic acid (KETE) attached to four phosphatidylethanolamines (PEs). The lipids are generated basally by 15-LOX in IL-4-stimulated monocytes, are elevated on calcium mobilization, and are detected at increased levels in bronchoalveolar lavage fluid from cystic fibrosis patients (3.6 ng/ml of lavage). Murine peritoneal macrophages generate 12-KETE-PEs, which are absent in 12/15-LOX-deficient mice. Inhibition of 15-prostaglandin dehydrogenase prevents their formation from exogenous 15-hydroxyeicosatetraenoic acid-PE in human monocytes. Both human and murine cells also generated analogous hydroperoxyeicosatetraenoic acid-PEs. The electrophilic reactivity of KETE-PEs is shown by their Michael addition to glutathione and cysteine. Lastly, both 15-hydroxyeicosatetraenoic acid-PE and 15-KETE-PE activated peroxisome proliferator-activated receptor-γ reporter activity in macrophages in a dose-dependent manner. In summary, we demonstrate novel peroxisome proliferator-activated receptor-γ-activating oxidized phospholipids generated enzymatically by LOX and 15-prostaglandin dehydrogenase in primary monocytic cells and in a human Th2-related lung disease. The lipids are a new family of bioactive mediators from the 12/15-LOX pathway that may contribute to its known anti-inflammatory actions in vivo. PMID:23060450

Novel keto-phospholipids are generated by monocytes and macrophages, detected in cystic fibrosis, and activate peroxisome proliferator-activated receptor-γ.

PubMed

Hammond, Victoria J; Morgan, Alwena H; Lauder, Sarah; Thomas, Christopher P; Brown, Sarah; Freeman, Bruce A; Lloyd, Clare M; Davies, Jane; Bush, Andrew; Levonen, Anna-Liisa; Kansanen, Emilia; Villacorta, Luis; Chen, Y Eugene; Porter, Ned; Garcia-Diaz, Yoel M; Schopfer, Francisco J; O'Donnell, Valerie B

2012-12-07

12/15-Lipoxygenases (LOXs) in monocytes and macrophages generate novel phospholipid-esterified eicosanoids. Here, we report the generation of two additional families of related lipids comprising 15-ketoeicosatetraenoic acid (KETE) attached to four phosphatidylethanolamines (PEs). The lipids are generated basally by 15-LOX in IL-4-stimulated monocytes, are elevated on calcium mobilization, and are detected at increased levels in bronchoalveolar lavage fluid from cystic fibrosis patients (3.6 ng/ml of lavage). Murine peritoneal macrophages generate 12-KETE-PEs, which are absent in 12/15-LOX-deficient mice. Inhibition of 15-prostaglandin dehydrogenase prevents their formation from exogenous 15-hydroxyeicosatetraenoic acid-PE in human monocytes. Both human and murine cells also generated analogous hydroperoxyeicosatetraenoic acid-PEs. The electrophilic reactivity of KETE-PEs is shown by their Michael addition to glutathione and cysteine. Lastly, both 15-hydroxyeicosatetraenoic acid-PE and 15-KETE-PE activated peroxisome proliferator-activated receptor-γ reporter activity in macrophages in a dose-dependent manner. In summary, we demonstrate novel peroxisome proliferator-activated receptor-γ-activating oxidized phospholipids generated enzymatically by LOX and 15-prostaglandin dehydrogenase in primary monocytic cells and in a human Th2-related lung disease. The lipids are a new family of bioactive mediators from the 12/15-LOX pathway that may contribute to its known anti-inflammatory actions in vivo.

Lentiviral infection of proliferating brain macrophages in HIV and simian immunodeficiency virus encephalitis despite sterile alpha motif and histidine-aspartate domain-containing protein 1 expression

PubMed Central

Lindgren, Allison A.; Filipowicz, Adam R.; Hattler, Julian B.; Kim, Soon Ok; Chung, Hye Kyung; Kuroda, Marcelo J.; Johnson, Edward M.; Kim, Woong-Ki

2018-01-01

Objective: HIV-1 infection of the brain and related cognitive impairment remain prevalent in HIV-1-infected individuals despite combination antiretroviral therapy. Sterile alpha motif and histidine-aspartate domain-containing protein 1 (SAMHD1) is a newly identified host restriction factor that blocks the replication of HIV-1 and other retroviruses in myeloid cells. Cell cycle-regulated phosphorylation at residue Thr592 and viral protein X (Vpx)-mediated degradation of SAMHD1 have been shown to bypass SAMHD1 restriction in vitro. Herein, we investigated expression and phosphorylation of SAMHD1 in vivo in relation to macrophage infection and proliferation during the neuropathogenesis of HIV-1 and simian immunodeficiency virus (SIV) encephalitis. Methods: Using brain and other tissues from uninfected and SIV-infected macaques with or without encephalitis, we performed immunohistochemistry, multilabel fluorescence microscopy and western blot to examine the expression, localization and phosphorylation of SAMHD1. Results: The number of SAMHD1+ nuclei increased in encephalitic brains despite the presence of Vpx. Many of these cells were perivascular macrophages, although subsets of SAMHD1+ microglia and endothelial cells were also observed. The SAMHD1+ macrophages were shown to be both infected and proliferating. Moreover, the presence of cycling SAMHD1+ brain macrophages was confirmed in the tissue of HIV-1-infected patients with encephalitis. Finally, western blot analysis of brain-protein extracts from SIV-infected macaques showed that SAMHD1 protein exists in the brain mainly as an inactive Thr592-phosphorylated form. Conclusion: The ability of SAMHD1 to act as a restriction factor for SIV/HIV in the brain is likely bypassed in proliferating brain macrophages through the phosphorylation-mediated inactivation, not Vpx-mediated degradation of SAMHD1. PMID:29698322

Immunostimulatory activity of snake fruit (Salacca edulis Reinw.) cultivar Pondoh Hitam extract on the activation of macrophages in vitro

NASA Astrophysics Data System (ADS)

Wijanarti, Sri; Putra, Agus Budiawan Naro; Nishi, Kosuke; Harmayani, Eni; Sugahara, Takuya

2017-05-01

Snake fruit (Salacca edulis Reinw) cultivar Pondoh Hitam is a tropical fruit produced in Indonesia. It is consumed freshly or processed and believed as the most delicious snake fruit cultivar. Snake fruit flesh contains high polisaccharides such as pectin and dietary fiber. Therefore, snake fruit is a potential immunostimulator candidates but the immunological effect of snake fruit flesh has not been reported. In the present study, immunostimulatory activity of snake fruit flesh extract (SFFE) on macrophages activation was evaluated. SFFE was prepared by extracting from snake fruit flesh with water, methanol 70%, and ethanol 70% for 15 h at 4°C. Then obtained SFFE was used to stimulated cytokine production in vitro using J774.1 cell line. The extract giving strongest stimulation was sellected for in vivo assay to stimulate cytokines production and gene expression using peritoneal macrophage (P-mac) of BALB/c mice. The results showed that SFFE exhibited immunostimulatory activities. Immunostimulatory activity could be indicated by macrophages activation characteristics such as cytokines production. Water extract of SFFE gave strongest stimulation on cytokines production in vitro and sellected for in vivo assay. In vivo assay showed that SFFE stimulated cytokines production as well as their gene expression levels. The optimum stimulation was demonstrated by SFFE 16.7 mg/g. Overall findings suggest that SFFE has a potent beneficial effects to promote the body health through activating macrophages.

Eicosapentaenoic acid membrane incorporation impairs ABCA1-dependent cholesterol efflux via a protein kinase A signaling pathway in primary human macrophages.

PubMed

Fournier, Natalie; Tardivel, Sylviane; Benoist, Jean-François; Vedie, Benoît; Rousseau-Ralliard, Delphine; Nowak, Maxime; Allaoui, Fatima; Paul, Jean-Louis

2016-04-01

A diet rich in n-3/n-6 polyunsaturated fatty acids (PUFAs) is cardioprotective. Dietary PUFAs affect the cellular phospholipids composition, which may influence the function of membrane proteins. We investigated the impact of the membrane incorporation of several PUFAs on ABCA1-mediated cholesterol efflux, a key antiatherogenic pathway. Arachidonic acid (AA) (C20:4 n-6) and docosahexaenoic acid (DHA) (C22:6 n-3) decreased or increased cholesterol efflux from J774 mouse macrophages, respectively, whereas they had no effect on efflux from human monocyte-derived macrophages (HMDM). Importantly, eicosapentaenoic acid (EPA) (C20:5 n-3) induced a dose-dependent reduction of ABCA1 functionality in both cellular models (-28% for 70μM of EPA in HMDM), without any alterations in ABCA1 expression. These results show that PUFA membrane incorporation does not have the same consequences on cholesterol efflux from mouse and human macrophages. The EPA-treated HMDM exhibited strong phospholipid composition changes, with high levels of both EPA and its elongation product docosapentaenoic acid (DPA) (C22:5 n-3), which is associated with a decreased level of AA. In HMDM, EPA reduced the ATPase activity of the membrane transporter. Moreover, the activation of adenylate cyclase by forskolin and the inhibition of cAMP phosphodiesterase by isobutylmethylxanthine restored ABCA1 cholesterol efflux in EPA-treated human macrophages. In conclusion, EPA membrane incorporation reduces ABCA1 functionality in mouse macrophages as well as in primary human macrophages and this effect seems to be PKA-dependent in human macrophages. Copyright © 2016 Elsevier B.V. All rights reserved.

Vitamin D binding protein-macrophage activating factor directly inhibits proliferation, migration, and uPAR expression of prostate cancer cells.

PubMed

Gregory, Kalvin J; Zhao, Bing; Bielenberg, Diane R; Dridi, Sami; Wu, Jason; Jiang, Weihua; Huang, Bin; Pirie-Shepherd, Steven; Fannon, Michael

2010-10-18

Vitamin D binding protein-macrophage activating factor (DBP-maf) is a potent inhibitor of tumor growth. Its activity, however, has been attributed to indirect mechanisms such as boosting the immune response by activating macrophages and inhibiting the blood vessel growth necessary for the growth of tumors. In this study we show for the first time that DBP-maf exhibits a direct and potent effect on prostate tumor cells in the absence of macrophages. DBP-maf demonstrated inhibitory activity in proliferation studies of both LNCaP and PC3 prostate cancer cell lines as well as metastatic clones of these cells. Flow cytometry studies with annexin V and propidium iodide showed that this inhibitory activity is not due to apoptosis or cell death. DBP-maf also had the ability to inhibit migration of prostate cancer cells in vitro. Finally, DBP-maf was shown to cause a reduction in urokinase plasminogen activator receptor (uPAR) expression in prostate tumor cells. There is evidence that activation of this receptor correlates with tumor metastasis. These studies show strong inhibitory activity of DBP-maf on prostate tumor cells independent of its macrophage activation.

Vitamin D Binding Protein-Macrophage Activating Factor Directly Inhibits Proliferation, Migration, and uPAR Expression of Prostate Cancer Cells

PubMed Central

Bielenberg, Diane R.; Dridi, Sami; Wu, Jason; Jiang, Weihua; Huang, Bin; Pirie-Shepherd, Steven; Fannon, Michael

2010-01-01

Background Vitamin D binding protein-macrophage activating factor (DBP-maf) is a potent inhibitor of tumor growth. Its activity, however, has been attributed to indirect mechanisms such as boosting the immune response by activating macrophages and inhibiting the blood vessel growth necessary for the growth of tumors. Methods and Findings In this study we show for the first time that DBP-maf exhibits a direct and potent effect on prostate tumor cells in the absence of macrophages. DBP-maf demonstrated inhibitory activity in proliferation studies of both LNCaP and PC3 prostate cancer cell lines as well as metastatic clones of these cells. Flow cytometry studies with annexin V and propidium iodide showed that this inhibitory activity is not due to apoptosis or cell death. DBP-maf also had the ability to inhibit migration of prostate cancer cells in vitro. Finally, DBP-maf was shown to cause a reduction in urokinase plasminogen activator receptor (uPAR) expression in prostate tumor cells. There is evidence that activation of this receptor correlates with tumor metastasis. Conclusions These studies show strong inhibitory activity of DBP-maf on prostate tumor cells independent of its macrophage activation. PMID:20976141

Flavonoid Apigenin Inhibits Lipopolysaccharide-Induced Inflammatory Response through Multiple Mechanisms in Macrophages

PubMed Central

Zhang, Xiaoxuan; Wang, Guangji; Gurley, Emily C.; Zhou, Huiping

2014-01-01

Background Apigenin is a non-toxic natural flavonoid that is abundantly present in common fruits and vegetables. It has been reported that apigenin has various beneficial health effects such as anti-inflammation and chemoprevention. Multiple studies have shown that inflammation is an important risk factor for atherosclerosis, diabetes, sepsis, various liver diseases, and other metabolic diseases. Although it has been long realized that apigenin has anti-inflammatory activities, the underlying functional mechanisms are still not fully understood. Methodology and Principal Findings In the present study, we examined the effect of apigenin on LPS-induced inflammatory response and further elucidated the potential underlying mechanisms in human THP-1-induced macrophages and mouse J774A.1 macrophages. By using the PrimePCR array, we were able to identify the major target genes regulated by apigenin in LPS-mediated immune response. The results indicated that apigenin significantly inhibited LPS-induced production of pro-inflammatory cytokines, such as IL-6, IL-1β, and TNF-α through modulating multiple intracellular signaling pathways in macrophages. Apigenin inhibited LPS-induced IL-1β production by inhibiting caspase-1 activation through the disruption of the NLRP3 inflammasome assembly. Apigenin also prevented LPS-induced IL-6 and IL-1β production by reducing the mRNA stability via inhibiting ERK1/2 activation. In addition, apigenin significantly inhibited TNF-α and IL-1β-induced activation of NF-κB. Conclusion and Significance Apigenin Inhibits LPS-induced Inflammatory Response through multiple mechanisms in macrophages. These results provided important scientific evidences for the potential application of apigenin as a therapeutic agent for inflammatory diseases. PMID:25192391

40 CFR 77.4 - Administrator's action on proposed offset plans.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 40 Protection of Environment 17 2013-07-01 2013-07-01 false Administrator's action on proposed offset plans. 77.4 Section 77.4 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS (CONTINUED) EXCESS EMISSIONS § 77.4 Administrator's action on proposed offset plans. (a...

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

PubMed Central

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

2013-01-01

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

7 CFR 774.10 - Other Federal, State, and local requirements.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 7 Agriculture 7 2013-01-01 2013-01-01 false Other Federal, State, and local requirements. 774.10 Section 774.10 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.10 Other Federal...

7 CFR 774.10 - Other Federal, State, and local requirements.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 7 Agriculture 7 2012-01-01 2012-01-01 false Other Federal, State, and local requirements. 774.10 Section 774.10 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.10 Other Federal...

7 CFR 774.10 - Other Federal, State, and local requirements.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 7 2011-01-01 2011-01-01 false Other Federal, State, and local requirements. 774.10 Section 774.10 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.10 Other Federal...

7 CFR 774.10 - Other Federal, State, and local requirements.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Other Federal, State, and local requirements. 774.10 Section 774.10 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.10 Other Federal...

7 CFR 774.10 - Other Federal, State, and local requirements.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 7 Agriculture 7 2014-01-01 2014-01-01 false Other Federal, State, and local requirements. 774.10 Section 774.10 Agriculture Regulations of the Department of Agriculture (Continued) FARM SERVICE AGENCY, DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.10 Other Federal...

CYP epoxygenase 2J2 prevents cardiac fibrosis by suppression of transmission of pro-inflammation from cardiomyocytes to macrophages

PubMed Central

Yang, Lei; Ni, Li; Duan, Quanlu; Wang, Xingxu; Chen, Chen; Chen, Song; Chaugai, Sandip; Zeldin, D.C.; Tang, Jia Rong; Wang, Dao Wen

2017-01-01

Cytochrome P450 epoxygenase (CYP450)-derived epoxyeicosatrienoic acids (EETs) are important regulators of cardiac remodeling; but the underlying mechanism remains unclear. The present study aimed to elucidate how EETs regulated cardiac fibrosis in response to isoprenaline (Iso) or angiotensin (Ang) II. Cardiac-specific human CYP2J2 transgenic mice (Tr) and wild-type (WT) C57BL/6 littermates were infused with Iso- or Ang II. Two weeks after infusion, Tr mice showed more alleviative cardiac fibrosis and inflammation compared with WT mice. In vitro, we found Iso or Ang II induced nuclear transfer of NF-κB p65 and inflammatory cytokines expression in cardiomyocytes. Furthermore, inflammation response emerged in macrophages cultured in cardiomyocytes-conditioned medium. When pretreatment with 14,15-EET in cardiomyocytes, the inflammatory response was markedly suppressed and the transmission of inflammation from cardiomyocytes to macrophages was reduced. In conclusion, CYP2J2 and EETs prevent cardiac fibrosis and cardiac dysfunction by suppressing transmission of pro-inflammation from cardiomyocytes to macrophages in heart, suggesting that elevation of EETs level could be a potential strategy to prevent cardiac fibrosis. PMID:25686540

Anti-leishmanial and toxicity activities of some selected Iranian medicinal plants.

PubMed

Kheiri Manjili, Hamidreza; Jafari, Hamidreza; Ramazani, Ali; Davoudi, Noushin

2012-11-01

Leishmaniasis is caused by protozoan parasites belonging to the genus Leishmania. Cutaneous leishmaniasis is the most common form of leishmaniasis in Iran. As there is not any vaccine for leishmaniasis, treatment is important to prevent the spreading of parasites. There is, therefore, a need to develop newer drugs from different sources. The aim of this study was to assess anti-leishmanial activity of the ethanolic extracts of 17 different medicinal plants against Leishmania major promastigotes and macrophage cell line J774. The selection of the hereby studied 17 plants was based on the existing information on their local ethnobotanic history. Plants were dried, powdered, and macerated in a hydroalcoholic solution. Resulting extracts have been assessed for in vitro anti-leishmanial and brine shrimp toxicity activities. Four plants, Caesalpinia gilliesii, Satureia hortensis, Carum copticum heirm, and Thymus migricus, displayed high anti-leishmanial activity (IC50, 9.76 ± 1.27, 15.625 ± 3.76, 15.625 ± 5.46, and 31.25 ± 15.44 μM, respectively) and were toxic against the J774 macrophage cell line at higher concentrations than those needed to inhibit the parasite cell growth (IC50, 45.13 ± 3.17, 100.44 ± 17.48, 43.76 ± 0.78, and 39.67 ± 3.29 μM, respectively). Glucantime as positive control inhibited the growth of L. major promastigotes with IC50 = 254 μg/ml on promastigotes (1 × 10(6)/100 μ/well) of a log phase culture, without affecting the growth of J774 macrophages. These data revealed that C. gilliesii, S. hortensis, C. copticum heirm, and T. migricus extracts contain active compounds, which could serve as alternative agents in the control of cutaneous leishmaniasis. The activity of these herbs against L. major promastigotes and macrophage cell line J774 was reported for the first time in our study.

Activation/proliferation and apoptosis of bystander goat lymphocytes induced by a macrophage-tropic chimeric caprine arthritis encephalitis virus expressing SIV Nef

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bouzar, Baya Amel; Rea, Angela; Hoc-Villet, Stephanie

Caprine arthritis encephalitis virus (CAEV) is the natural lentivirus of goats, well known for its tropism for macrophages and its inability to cause infection in lymphocytes. The viral genome lacks nef, tat, vpu and vpx coding sequences. To test the hypothesis that when nef is expressed by the viral genome, the virus became toxic for lymphocytes during replication in macrophages, we inserted the SIVsmm PBj14 nef coding sequences into the genome of CAEV thereby generating CAEV-nef. This recombinant virus is not infectious for lymphocytes but is fully replication competent in goat macrophages in which it constitutively expresses the SIV Nef.more » We found that goat lymphocytes cocultured with CAEV-nef-infected macrophages became activated, showing increased expression of the interleukin-2 receptor (IL-2R). Activation correlated with increased proliferation of the cells. Interestingly, a dual effect in terms of apoptosis regulation was observed in exposed goat lymphocytes. Nef was found first to induce a protection of lymphocytes from apoptosis during the first few days following exposure to infected macrophages, but later it induced increased apoptosis in the activated lymphocytes. This new recombinant virus provides a model to study the functions of Nef in the context of infection of macrophages, but in absence of infection of T lymphocytes and brings new insights into the biological effects of Nef on lymphocytes.« less

The alveolar macrophage.

PubMed

Bowden, D H

1984-04-01

The pulmonary macrophagic system is critical to the defense of the lung, keeping the alveoli clean and sterile and responding on demand with an adaptive outpouring of new cells into the air sacs. Under basal conditions alveolar macrophages, in common with other mononuclear phagocytes, are derived from the bone marrow. A population of macrophage precursors within the pulmonary interstitium provides a reserve pool capable of proliferation and delivery of phagocytes in response to unusually heavy loads of inhaled particles. This reserve system also produces macrophages when monocytic precursors in the bone marrow are depleted by diseases such as leukemia. The alveolar macrophage is destined to ingest particulate matter and to be eliminated along the mucociliary pathway; clearance by lymphatics is of minor importance and macrophages probably do not recross the alveolar epithelium to reach the pulmonary interstitial compartment. Although the protective role of the macrophage is dominant, this cell may participate, directly or indirectly, in the genesis of two major groups of chronic pulmonary disease, interstitial fibrosis and emphysema. Such inappropriate responses involve interactions with fibroblastic cells and tissue injury initiated by proteases secreted by the macrophage.

Acrolein increases macrophage atherogenicity in association with gut microbiota remodeling in atherosclerotic mice: protective role for the polyphenol-rich pomegranate juice.

PubMed

Rom, Oren; Korach-Rechtman, Hila; Hayek, Tony; Danin-Poleg, Yael; Bar, Haim; Kashi, Yechezkel; Aviram, Michael

2017-04-01

The unsaturated aldehyde acrolein is pro-atherogenic, and the polyphenol-rich pomegranate juice (PJ), known for its anti-oxidative/anti-atherogenic properties, inhibits macrophage foam cell formation, the hallmark feature of early atherosclerosis. This study aimed to investigate two unexplored areas of acrolein atherogenicity: macrophage lipid metabolism and the gut microbiota composition. The protective effects of PJ against acrolein atherogenicity were also evaluated. Atherosclerotic apolipoprotein E-deficient (apoE -/- ) mice that were fed acrolein (3 mg/kg/day) for 1 month showed significant increases in serum and aortic cholesterol, triglycerides, and lipid peroxides. In peritoneal macrophages isolated from the mice and in J774A.1 cultured macrophages, acrolein exposure increased intracellular oxidative stress and stimulated cholesterol and triglyceride accumulation via enhanced rates of their biosynthesis and over-expression of key regulators of cellular lipid biosynthesis: sterol regulatory element-binding proteins (SREBPs), 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCR), and diacylglycerol acyltransferase1 (DGAT1). Acrolein-fed mice demonstrated a major shift in the gut microbiota composition, including a significant phylum-level change in increased Firmicutes and decreased Bacteroidetes. At the family level, acrolein significantly increased the prevalence of Ruminococcaceae and Lachnospiraceae of which the Coprococcus genus was significantly and positively correlated with serum, aortic and macrophage lipid levels and peroxidation. The pro-atherogenic effects of acrolein on serum, aortas, macrophages, and the gut microbiota were substantially abolished by PJ. In conclusion, these findings provide novel mechanisms by which acrolein increases macrophage lipid accumulation and alters the gut microbiota composition in association with enhanced atherogenesis. Moreover, PJ was found as an effective strategy against acrolein atherogenicity.

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

PubMed

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

2015-10-01

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

Implication of matrix metalloproteinases 2 and 9 in ceramide 1-phosphate-stimulated macrophage migration.

PubMed

Ordoñez, Marta; Rivera, Io-Guané; Presa, Natalia; Gomez-Muñoz, Antonio

2016-08-01

Cell migration is a complex biological function involved in both physiologic and pathologic processes. Although this is a subject of intense investigation, the mechanisms by which cell migration is regulated are not completely understood. In this study we show that the bioactive sphingolipid ceramide 1-phosphate (C1P), which is involved in inflammatory responses, causes upregulation of metalloproteinases (MMP) -2 and -9 in J774A.1 macrophages. This effect was shown to be dependent on stimulation of phosphatidylinositol 3-kinase (PI3K) and extracellularly regulated kinases 1-2 (ERK1-2) as demonstrated by treating the cells with specific siRNA to knockdown the p85 regulatory subunit of PI3K, or ERK1-2. Inhibition of MMP-2 or MMP-9 pharmacologically or with specific siRNA to silence the genes encoding these MMPs abrogated C1P-stimulated macrophage migration. Also, C1P induced actin polymerization and potently increased phosphorylation of the focal adhesion protein paxillin, which are essential factors in the regulation of cell migration. As expected, blockade of paxillin activation with specific siRNA significantly reduced actin polymerization. In addition, inhibition of actin polymerization with cytochalasin D completely blocked C1P-induced MMP-2 and -9 expression as well as C1P-stimulated macrophage migration. It was also observed that pertussis toxin (Ptx) inhibited Akt, ERK1-2, and paxillin phosphorylation, and completely blocked cell migration. The latter findings support the notion that C1P-stimulated macrophage migration is a receptor mediated effect, and point to MMP-2 and -9 as possible therapeutic targets to control inflammation. Copyright © 2016 Elsevier Inc. All rights reserved.

Essential role of protein kinase C zeta in transducing a motility signal induced by superoxide and a chemotactic peptide, fMLP.

PubMed

Kuribayashi, Kageaki; Nakamura, Kiminori; Tanaka, Maki; Sato, Tsutomu; Kato, Junji; Sasaki, Katsunori; Takimoto, Rishu; Kogawa, Katsuhisa; Terui, Takeshi; Takayama, Tetsuji; Onuma, Takayuki; Matsunaga, Takuya; Niitsu, Yoshiro

2007-03-26

Under various pathological conditions, including infection, malignancy, and autoimmune diseases, tissues are incessantly exposed to reactive oxygen species produced by infiltrating inflammatory cells. We show augmentation of motility associated with morphological changes of human squamous carcinoma SASH1 cells, human peripheral monocytes (hPMs), and murine macrophage-like cell line J774.1 by superoxide stimulation. We also disclose that motility of hPMs and J774.1 induced by a chemotactic peptide (N-formyl-methionyl-leucyl-phenylalanine [fMLP]) was inhibited by superoxide dismutase or N-acetylcystein, indicating stimulation of motility by superoxide generated by fMLP stimulation. In these cells, protein kinase C (PKC) zeta was activated to phosphorylate RhoGDI-1, which liberated RhoGTPases, leading to their activation. These events were inhibited by dominant-negative PKCzeta in SASH1 cells, myristoylated PKCzeta peptides in hPMs and J774.1, or a specific inhibitor of RhoGTPase in SASH1, hPMs, and J774.1. These results suggest a new approach for manipulation of inflammation as well as tumor cell invasion by targeting this novel signaling pathway.

Sirtuin inhibition attenuates the production of inflammatory cytokines in lipopolysaccharide-stimulated macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fernandes, Claudia A.; Fievez, Laurence; Neyrinck, Audrey M.

Highlights: Black-Right-Pointing-Pointer Lipopolysaccharide-stimulated macrophages were treated with cambinol and sirtinol. Black-Right-Pointing-Pointer Cambinol and sirtinol decreased lipopolysaccharide-induced cytokines. Black-Right-Pointing-Pointer Cambinol decreased NF-{kappa}B activity but had no impact on p38 MAPK activation. Black-Right-Pointing-Pointer Sirtuins are an interesting target for the treatment of inflammatory diseases. -- Abstract: In several inflammatory conditions such as rheumatoid arthritis or sepsis, the regulatory mechanisms of inflammation are inefficient and the excessive inflammatory response leads to damage to the host. Sirtuins are class III histone deacetylases that modulate the activity of several transcription factors that are implicated in immune responses. In this study, we evaluated the impact ofmore » sirtuin inhibition on the activation of lipopolysaccharide (LPS)-stimulated J774 macrophages by assessing the production of inflammatory cytokines. The pharmacologic inhibition of sirtuins decreased the production of tumour necrosis factor-alpha (TNF-{alpha}) interleukin 6 (IL-6) and Rantes. The reduction of cytokine production was associated with decreased nuclear factor kappa B (NF-{kappa}B) activity and inhibitor kappa B alpha (I{kappa}B{alpha}) phosphorylation while no impact was observed on the phosphorylation status of p38 mitogen-activated kinase (p38 MAPK). This work shows that sirtuin pharmacologic inhibitors are a promising tool for the treatment of inflammatory conditions.« less

The Role of CD38 in Fcγ Receptor (FcγR)-mediated Phagocytosis in Murine Macrophages*

PubMed Central

Kang, John; Park, Kwang-Hyun; Kim, Jwa-Jin; Jo, Eun-Kyeong; Han, Myung-Kwan; Kim, Uh-Hyun

2012-01-01

Phagocytosis is a crucial event in the immune system that allows cells to engulf and eliminate pathogens. This is mediated through the action of immunoglobulin (IgG)-opsonized microbes acting on Fcγ receptors (FcγR) on macrophages, which results in sustained levels of intracellular Ca2+ through the mobilization of Ca2+ second messengers. It is known that the ADP-ribosyl cyclase is responsible for the rise in Ca2+ levels after FcγR activation. However, it is unclear whether and how CD38 is involved in FcγR-mediated phagocytosis. Here we show that CD38 is recruited to the forming phagosomes during phagocytosis of IgG-opsonized particles and produces cyclic-ADP-ribose, which acts on ER Ca2+ stores, thus allowing an increase in FcγR activation-mediated phagocytosis. Ca2+ data show that pretreatment of J774A.1 macrophages with 8-bromo-cADPR, ryanodine, blebbistatin, and various store-operated Ca2+ inhibitors prevented the long-lasting Ca2+ signal, which significantly reduced the number of ingested opsonized particles. Ex vivo data with macrophages extracted from CD38−/− mice also shows a reduced Ca2+ signaling and phagocytic index. Furthermore, a significantly reduced phagocytic index of Mycobacterium bovis BCG was shown in macrophages from CD38−/− mice in vivo. This study suggests a crucial role of CD38 in FcγR-mediated phagocytosis through its recruitment to the phagosome and mobilization of cADPR-induced intracellular Ca2+ and store-operated extracellular Ca2+ influx. PMID:22396532

42 CFR 423.774 - Eligibility determinations, redeterminations, and applications.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 42 Public Health 3 2010-10-01 2010-10-01 false Eligibility determinations, redeterminations, and applications. 423.774 Section 423.774 Public Health CENTERS FOR MEDICARE & MEDICAID SERVICES, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICARE PROGRAM VOLUNTARY MEDICARE PRESCRIPTION DRUG BENEFIT...

Antibacterial Activity of Ciprofloxacin-Encapsulated Cockle Shells Calcium Carbonate (Aragonite) Nanoparticles and Its Biocompatability in Macrophage J774A.1

PubMed Central

Isa, Tijani; Zakaria, Zuki Abu Bakar; Rukayadi, Yaya; Mohd Hezmee, Mohd Noor; Jaji, Alhaji Zubair; Imam, Mustapha Umar; Hammadi, Nahidah Ibrahim; Mahmood, Saffanah Khuder

2016-01-01

The use of nanoparticle delivery systems to enhance intracellular penetration of antibiotics and their retention time is becoming popular. The challenge, however, is that the interaction of nanoparticles with biological systems at the cellular level must be established prior to biomedical applications. Ciprofloxacin–cockle shells-derived calcium carbonate (aragonite) nanoparticles (C-CSCCAN) were developed and characterized. Antibacterial activity was determined using a modified disc diffusion protocol on Salmonella Typhimurium (S. Typhimurium). Biocompatibilittes with macrophage were evaluated using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-Bromo-2′-deoxyuridine (BrdU) assays. Transcriptional regulation of interleukin 1 beta (IL-1β) was determined using reverse transcriptase-polymerase chain reaction (RT-PCR). C-CSCCAN were spherical in shape, with particle sizes ranging from 11.93 to 22.12 nm. Encapsulation efficiency (EE) and loading content (LC) were 99.5% and 5.9%, respectively, with negative ζ potential. X-ray diffraction patterns revealed strong crystallizations and purity in the formulations. The mean diameter of inhibition zone was 18.6 ± 0.5 mm, which was better than ciprofloxacin alone (11.7 ± 0.9 mm). Study of biocompatability established the cytocompatability of the delivery system without upregulation of IL-1β. The results indicated that ciprofloxacin–nanoparticles enhanced the antibacterial efficacy of the antibiotic, and could act as a suitable delivery system against intracellular infections. PMID:27213349

Antibacterial Activity of Ciprofloxacin-Encapsulated Cockle Shells Calcium Carbonate (Aragonite) Nanoparticles and Its Biocompatability in Macrophage J774A.1.

PubMed

Isa, Tijani; Zakaria, Zuki Abu Bakar; Rukayadi, Yaya; Mohd Hezmee, Mohd Noor; Jaji, Alhaji Zubair; Imam, Mustapha Umar; Hammadi, Nahidah Ibrahim; Mahmood, Saffanah Khuder

2016-05-19

The use of nanoparticle delivery systems to enhance intracellular penetration of antibiotics and their retention time is becoming popular. The challenge, however, is that the interaction of nanoparticles with biological systems at the cellular level must be established prior to biomedical applications. Ciprofloxacin-cockle shells-derived calcium carbonate (aragonite) nanoparticles (C-CSCCAN) were developed and characterized. Antibacterial activity was determined using a modified disc diffusion protocol on Salmonella Typhimurium (S. Typhimurium). Biocompatibilittes with macrophage were evaluated using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-Bromo-2'-deoxyuridine (BrdU) assays. Transcriptional regulation of interleukin 1 beta (IL-1β) was determined using reverse transcriptase-polymerase chain reaction (RT-PCR). C-CSCCAN were spherical in shape, with particle sizes ranging from 11.93 to 22.12 nm. Encapsulation efficiency (EE) and loading content (LC) were 99.5% and 5.9%, respectively, with negative ζ potential. X-ray diffraction patterns revealed strong crystallizations and purity in the formulations. The mean diameter of inhibition zone was 18.6 ± 0.5 mm, which was better than ciprofloxacin alone (11.7 ± 0.9 mm). Study of biocompatability established the cytocompatability of the delivery system without upregulation of IL-1β. The results indicated that ciprofloxacin-nanoparticles enhanced the antibacterial efficacy of the antibiotic, and could act as a suitable delivery system against intracellular infections.

[Effects of thyroid hormone on macrophage dysfunction induced by oxidized low-density lipoprotein].

PubMed

Ning, Yu; Zhang, Ming; DU, Yun-Hui; Zhang, Hui-Na; Li, Lin-Yi; Qin, Yan-Wen; Wen, Wan-Wan; Zhao, Quan-Ming

2018-04-25

It has been recognized that patients with hypothyroidism have higher risks of atherosclerosis and coronary heart disease, however, the mechanisms are largely unknown. Considering that macrophage dysfunction plays an important role in the formation and development of atherosclerosis plaques, this study aimed to investigate the direct effects of thyroid hormone on macrophage functions and to provide new insight for the mechanism of hypothyroid atherosclerosis. RAW264.7 cells (mouse leukaemic monocyte macrophage cell line) were incubated with oxidized low-density lipoprotein (oxLDL) to establish macrophage foam cells model in vitro, and the protective effects of different concentration of thyroxine (T4) on the macrophage foam cells function were explored. The proliferation, migration and cell aging of macrophages were detected by MTT method, scratch test and β-galactosidase staining respectively. The ELISA method was used to detect the secretion of tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1), and interleukin-1β (IL-1β). Western blot analysis was applied to measure the phosphorylation of focal adhesion kinase (FAK), which was required for the process of proliferation and migration of macrophages. The results showed that oxLDL significantly inhibited the macrophage proliferation and migration, induced cell senescence, and promoted the secretion of TNF-α, MCP-1, and IL-1β; while T4 reversed those effects of oxLDL on macrophage in a concentration-dependent manner. Moreover, oxLDL increased the phosphorylation of FAK in macrophage, while T4 concentration-dependently reversed the effect. These results suggest that T4 modulates macrophage proliferation, migration, senescence, and secretion of inflammation factors in a concentration-dependent way.

The role of lipopolysaccharide in infectious bone resorption of periapical lesion.

PubMed

Hong, Chi-Yuan; Lin, Sze-Kwan; Kok, Sang-Heng; Cheng, Shih-Jung; Lee, Ming-Shu; Wang, Tong-Mei; Chen, Chuan-Shuo; Lin, Li-Deh; Wang, Juo-Song

2004-03-01

The role of lipopolysaccharide (LPS) in periapical lesion-induced bone resorption was investigated. Polymyxin B (PMB), a specific inhibitor of LPS, was evaluated to treat the apical lesion. Lipopolysaccharide isolated from two common endodontic pathogens, Fusobacterium nucleatum and Porphyromonas endodontalis, stimulated mouse macrophage (J774) to release interleukin-1alpha (IL-1 alpha) and tumor necrosis factor-alpha (TNF-alpha) in a time-dependent manner. Combination of LPS further enhanced the stimulation. PMB inhibited these effects significantly. LPS also stimulated matrix metalloproteinase-1 (MMP-1) gene expression in J774, whereas anti-IL-1 alpha and anti-TNF-alpha antibodies, as well as PMB, diminished this effect. A disease model of periapical lesion was established in Wistar rat. Administration of PMB reduced the extent of lesion-associated bone resorption by 76% to approximately 80%, and simultaneously reduced the numbers of MMP-1-producing macrophages. It is suggested that LPS released from the infected root canal triggers the synthesis of IL-1 alpha and TNF-alpha from macrophages. These pro-inflammatory cytokines up-regulate the production of MMP-1 by macrophages to promote periapical bone resorption.

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

PubMed

Falchi, Mario; Varricchio, Lilian; Martelli, Fabrizio; Masiello, Francesca; Federici, Giulia; Zingariello, Maria; Girelli, Gabriella; Whitsett, Carolyn; Petricoin, Emanuel F; Moestrup, Søren Kragh; Zeuner, Ann; Migliaccio, Anna Rita

2015-02-01

Cultures of human CD34(pos) cells stimulated with erythroid growth factors plus dexamethasone, a model for stress erythropoiesis, generate numerous erythroid cells plus a few macrophages (approx. 3%; 3:1 positive and negative for CD169). Interactions occurring between erythroblasts and macrophages in these cultures and the biological effects associated with these interactions were documented by live phase-contrast videomicroscopy. Macrophages expressed high motility interacting with hundreds/thousands of erythroblasts per hour. CD169(pos) macrophages established multiple rapid 'loose' interactions with proerythroblasts leading to formation of transient erythroblastic island-like structures. By contrast, CD169(neg) macrophages established 'tight' interactions with mature erythroblasts and phagocytosed these cells. 'Loose' interactions of CD169(pos) macrophages were associated with proerythroblast cytokinesis (the M phase of the cell cycle) suggesting that these interactions may promote proerythroblast duplication. This hypothesis was tested by experiments that showed that as few as 103 macrophages significantly increased levels of 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide incorporation frequency in S/G2/M and cytokinesis expressed by proerythroblasts over 24 h of culture. These effects were observed also when macrophages were co-cultured with dexamethasone directly conjugated to a macrophage-specific CD163 antibody. In conclusion, in addition to promoting proerythroblast proliferation directly, dexamethasone stimulates expansion of these cells indirectly by stimulating maturation and cytokinesis supporting activity of macrophages. Copyright© Ferrata Storti Foundation.

Proteomic analysis reveals different composition of extracellular vesicles released by two Trypanosoma cruzi strains associated with their distinct interaction with host cells.

PubMed

Ribeiro, Kleber Silva; Vasconcellos, Camilla Ioshida; Soares, Rodrigo Pedro; Mendes, Maria Tays; Ellis, Cameron C; Aguilera-Flores, Marcela; de Almeida, Igor Correia; Schenkman, Sergio; Iwai, Leo Kei; Torrecilhas, Ana Claudia

2018-01-01

Trypanosoma cruzi , the aetiologic agent of Chagas disease, releases vesicles containing a wide range of surface molecules known to affect the host immunological responses and the cellular infectivity. Here, we compared the secretome of two distinct strains (Y and YuYu) of T. cruzi , which were previously shown to differentially modulate host innate and acquired immune responses. Tissue culture-derived trypomastigotes of both strains secreted extracellular vesicles (EVs), as demonstrated by electron scanning microscopy. EVs were purified by exclusion chromatography or ultracentrifugation and quantitated using nanoparticle tracking analysis. Trypomastigotes from YuYu strain released higher number of EVs than those from Y strain, enriched with virulence factors trans -sialidase (TS) and cruzipain. Proteomic analysis confirmed the increased abundance of proteins coded by the TS gene family, mucin-like glycoproteins, and some typical exosomal proteins in the YuYu strain, which also showed considerable differences between purified EVs and vesicle-free fraction as compared to the Y strain. To evaluate whether such differences were related to parasite infectivity, J774 macrophages and LLC-MK2 kidney cells were preincubated with purified EVs from both strains and then infected with Y strain trypomastigotes. EVs released by YuYu strain caused a lower infection but higher intracellular proliferation in J774 macrophages than EVs from Y strain. In contrast, YuYu strain-derived EVs caused higher infection of LLC-MK2 cells than Y strain-derived EVs. In conclusion, quantitative and qualitative differences in EVs and secreted proteins from different T. cruzi strains may correlate with infectivity/virulence during the host-parasite interaction.

Proteomic analysis reveals different composition of extracellular vesicles released by two Trypanosoma cruzi strains associated with their distinct interaction with host cells

PubMed Central

Ribeiro, Kleber Silva; Vasconcellos, Camilla Ioshida; Soares, Rodrigo Pedro; Ellis, Cameron C.; Aguilera-Flores, Marcela; de Almeida, Igor Correia

2018-01-01

ABSTRACT Trypanosoma cruzi, the aetiologic agent of Chagas disease, releases vesicles containing a wide range of surface molecules known to affect the host immunological responses and the cellular infectivity. Here, we compared the secretome of two distinct strains (Y and YuYu) of T. cruzi, which were previously shown to differentially modulate host innate and acquired immune responses. Tissue culture-derived trypomastigotes of both strains secreted extracellular vesicles (EVs), as demonstrated by electron scanning microscopy. EVs were purified by exclusion chromatography or ultracentrifugation and quantitated using nanoparticle tracking analysis. Trypomastigotes from YuYu strain released higher number of EVs than those from Y strain, enriched with virulence factors trans-sialidase (TS) and cruzipain. Proteomic analysis confirmed the increased abundance of proteins coded by the TS gene family, mucin-like glycoproteins, and some typical exosomal proteins in the YuYu strain, which also showed considerable differences between purified EVs and vesicle-free fraction as compared to the Y strain. To evaluate whether such differences were related to parasite infectivity, J774 macrophages and LLC-MK2 kidney cells were preincubated with purified EVs from both strains and then infected with Y strain trypomastigotes. EVs released by YuYu strain caused a lower infection but higher intracellular proliferation in J774 macrophages than EVs from Y strain. In contrast, YuYu strain-derived EVs caused higher infection of LLC-MK2 cells than Y strain-derived EVs. In conclusion, quantitative and qualitative differences in EVs and secreted proteins from different T. cruzi strains may correlate with infectivity/virulence during the host–parasite interaction. PMID:29696081

Expression of the peroxisome proliferator-activated receptor γ (PPARγ) in human atherosclerosis and regulation in macrophages by colony stimulating factors and oxidized low density lipoprotein

PubMed Central

Ricote, Mercedes; Huang, Jannet; Fajas, Luis; Li, Andrew; Welch, John; Najib, Jamila; Witztum, Joseph L.; Auwerx, Johan; Palinski, Wulf; Glass, Christopher K.

1998-01-01

The peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-dependent transcription factor that has been demonstrated to regulate fat cell development and glucose homeostasis. PPARγ is also expressed in a subset of macrophages and negatively regulates the expression of several proinflammatory genes in response to natural and synthetic ligands. We here demonstrate that PPARγ is expressed in macrophage foam cells of human atherosclerotic lesions, in a pattern that is highly correlated with that of oxidation-specific epitopes. Oxidized low density lipoprotein (oxLDL) and macrophage colony-stimulating factor, which are known to be present in atherosclerotic lesions, stimulated PPARγ expression in primary macrophages and monocytic cell lines. PPARγ mRNA expression was also induced in primary macrophages and THP-1 monocytic leukemia cells by the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA). Inhibition of protein kinase C blocked the induction of PPARγ expression by TPA, but not by oxLDL, suggesting that more than one signaling pathway regulates PPARγ expression in macrophages. TPA induced the expression of PPARγ in RAW 264.7 macrophages by increasing transcription from the PPARγ1 and PPARγ3 promoters. In concert, these observations provide insights into the regulation of PPARγ expression in activated macrophages and raise the possibility that PPARγ ligands may influence the progression of atherosclerosis. PMID:9636198

Advanced glycation end products affect cholesterol homeostasis by impairing ABCA1 expression on macrophages.

PubMed

Kamtchueng Simo, Olivier; Ikhlef, Souade; Berrougui, Hicham; Khalil, Abdelouahed

2017-08-01

Reverse cholesterol transport (RCT), which is intimately linked to high-density lipoproteins (HDLs), plays a key role in cholesterol homeostasis and the prevention of atherosclerosis. The goal of the present study was to investigate the effect of aging and advanced glycation end products (AGEs) on RCT as well as on other factors that may affect the antiatherogenic property of HDLs. The transfer of macrophage-derived cholesterol to the plasma and liver and then to the feces for elimination was significantly lower in aged mice than in young mice. Chronic injection of d -galactose (D-gal) or AGEs also significantly reduced RCT (65.3% reduction in [ 3 H]cholesterol levels in the plasma of D-gal-treated mice after 48 h compared with control mice, P < 0.01). The injection of both D-gal and aminoguanidine hydrochloride increased [ 3 H]cholesterol levels in the plasma, although the levels were lower than those of control mice. The in vitro incubation of HDLs with dicarbonyl compounds increased the carbonyl and conjugated diene content of HDLs and significantly reduced PON1 paraoxonase activity (87.4% lower than control HDLs, P < 0.0001). Treating J774A.1 macrophages with glycated fetal bovine serum increased carbonyl formation (39.5% increase, P < 0.003) and reduced ABCA1 protein expression and the capacity of macrophages to liberate cholesterol (69.1% decrease, P < 0.0001). Our results showed, for the first time, that RCT is altered with aging and that AGEs contribute significantly to this alteration.

7 CFR 774.3 - Appeals.

Code of Federal Regulations, 2011 CFR

2011-01-01

... PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.3 Appeals. A loan applicant or borrower may request an appeal or review of an adverse decision made by the Agency in accordance with 7 CFR part 11. ...

7 CFR 774.3 - Appeals.

Code of Federal Regulations, 2010 CFR

2010-01-01

... PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.3 Appeals. A loan applicant or borrower may request an appeal or review of an adverse decision made by the Agency in accordance with 7 CFR part 11. ...

Inhibitory effect of trichodermanone C, a sorbicillinoid produced by Trichoderma citrinoviride associated to the green alga Cladophora sp., on nitrite production in LPS-stimulated macrophages.

PubMed

Marra, Roberta; Nicoletti, Rosario; Pagano, Ester; DellaGreca, Marina; Salvatore, Maria Michela; Borrelli, Francesca; Lombardi, Nadia; Vinale, Francesco; Woo, Sheridan L; Andolfi, Anna

2018-05-31

From the green alga Cladophora sp. collected in Italy, the marine fungal strain A12 of Trichoderma citrinoviride was isolated, identified and characterized. LC-MS qTOF analysis was applied to perform a metabolic profile of the fungal culture. Chromatographic techniques and spectroscopic methods were used to isolate and characterize the major secondary metabolites produced by this strain in liquid culture. In particular, four known sorbicillinoids (trichodermanone C, spirosorbicillinol A, vertinolide and sorbicillin) were purified and identified, together with 2-phenylethanol and tyrosol. Moreover, metabolomic analysis allowed to detect small amounts of trichodimerol, rezishanone A, 2',3'-dihydrosorbicillin and bisvertinol. For the first time a significant inhibitory effect on nitrite levels has been shown for trichodermanone C in lipopolysaccharide-stimulated J774A.1 macrophages.

Transcription factor NFAT5 promotes macrophage survival in rheumatoid arthritis

PubMed Central

Choi, Susanna; Choi, Soo Youn; Kwon, H. Moo; Hwang, Daehee; Park, Yune-Jung; Cho, Chul-Soo

2017-01-01

Defective apoptotic death of activated macrophages has been implicated in the pathogenesis of rheumatoid arthritis (RA). However, the molecular signatures defining apoptotic resistance of RA macrophages are not fully understood. Here, global transcriptome profiling of RA macrophages revealed that the osmoprotective transcription factor nuclear factor of activated T cells 5 (NFAT5) critically regulates diverse pathologic processes in synovial macrophages including the cell cycle, apoptosis, and proliferation. Transcriptomic analysis of NFAT5-deficient macrophages revealed the molecular networks defining cell survival and proliferation. Proinflammatory M1-polarizing stimuli and hypoxic conditions were responsible for enhanced NFAT5 expression in RA macrophages. An in vitro functional study demonstrated that NFAT5-deficient macrophages were more susceptible to apoptotic death. Specifically, CCL2 secretion in an NFAT5-dependent fashion bestowed apoptotic resistance to RA macrophages in vitro. Injection of recombinant CCL2 into one of the affected joints of Nfat5+/– mice increased joint destruction and macrophage infiltration, demonstrating the essential role of the NFAT5/CCL2 axis in arthritis progression in vivo. Moreover, after intra-articular injection, NFAT5-deficient macrophages were more susceptible to apoptosis and less efficient at promoting joint destruction than were NFAT5-sufficient macrophages. Thus, NFAT5 regulates macrophage survival by inducing CCL2 secretion. Our results provide evidence that NFAT5 expression in macrophages enhances chronic arthritis by conferring apoptotic resistance to activated macrophages. PMID:28192374

Macrosialin, a macrophage-restricted membrane sialoprotein differentially glycosylated in response to inflammatory stimuli [published erratum appears in J Exp Med 1992 Jan 1;175(1):309

PubMed Central

1991-01-01

Rat monoclonal antibody FA/11 has been used to identify macrosialin, a sialoglycoprotein confined to murine mononuclear phagocytes and related cells. Originally identified as a macrophage-associated glycoprotein predominantly localized in intracellular membranes (Smith, M.J., and G.L.E. Koch. 1987. J. Cell Sci. 87:113), the antigen is widely expressed on tissue macrophages, including those in lymphoid areas, and is expressed at low levels on isolated dendritic cells. Immuno- adsorption experiments reported here show that macrosialin is identical to the major 87-115-kD sialoglycoprotein previously identified by lectin blotting in exudate but not resident peritoneal macrophages (Rabinowitz, S., and S. Gordon. 1989. J. Cell Sci. 93:623). Resident peritoneal macrophages express low levels of macrosialin antigen in a glycoform that does not bind 125I wheat germ agglutinin or 125I peanut agglutinin; inflammatory stimuli upregulate expression of this antigen (up to 17-fold), in an alternative glycoform that is detected by these lectins. Pulse-chase experiments reveal a 44-kD core peptide that initially bears high-mannose chains (giving Mr 66 kD) and is subsequently processed to a mature protein of Mr 87-104 kD. Each glycoform contains N-linked glycan, as well as O-linked sugar structures that show alternative processing. Poly-N-acetyllactosamine structures are detected in the exudate cell glycoform only. This new marker for mononuclear phagocytes illustrates two strategies by which macrophages remodel their membranes in response to inflammatory stimuli. Its predominantly intracellular location and restricted cell distribution suggest a possible role in membrane fusion or antigen processing. PMID:1919437

Asian and Siberian ginseng as a potential modulator of immune function: an in vitro cytokine study using mouse macrophages.

PubMed

Wang, Huamin; Actor, Jeffrey K; Indrigo, Jessica; Olsen, Margaret; Dasgupta, Amitava

2003-01-01

Ginseng is a widely used herbal product in China, other Asian countries, and in the Unites States. There is a traditional belief that ginseng stimulates immune functions. In this study, the innate effects of Asian and Siberian ginsengs on cytokines and chemokines produced by cultured macrophages were examined. The effects of Asian and Siberian ginseng on cytokines and chemokines produced by cultured macrophages were examined. Mouse macrophages (J774A.1) were incubated with Asian or Siberian ginseng at varying concentrations (1, 10, 100, and 1000 microg/ml) for 24 h and then harvested for RNA isolation. The expression levels of IL-1beta, IL-12, TNF-alpha, MIP-1 alpha, and MIP-2 mRNA were measured by quantitative PCR. Our data showed that Asian ginseng induced a statistically significant increase in IL-12 expression at both mRNA and protein levels. However, the minor twofold increase is probably biologically insignificant. No significant increase of IL-12 by Siberian ginseng was observed at any dose level studied. No significant change in IL-1beta, IL-15, TNF-alpha, or MIP-1alpha mRNA was observed by either Asian or Siberian ginseng treatment. Our data showed statistically significant differential regulation of IL-12 by Asian ginseng. Siberian ginseng did not show a statistically significant increase. We conclude that both Asian ginseng and Siberian ginseng cannot significantly stimulate innate macrophage immune functions that influence cellular immune responses. Therefore, contrary to the popular belief, Asian and Siberian ginseng may not stimulate immune function.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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

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 cellsmore » 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.« less

30 CFR 937.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 937.774 Section 937.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE OREGON § 937.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit Rights...

30 CFR 921.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 921.774 Section 921.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE MASSACHUSETTS § 921.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit...

30 CFR 910.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 910.774 Section 910.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE GEORGIA § 910.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit Rights...

30 CFR 939.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 939.774 Section 939.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE RHODE ISLAND § 939.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit...

30 CFR 933.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 933.774 Section 933.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE NORTH CAROLINA § 933.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit...

30 CFR 912.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 912.774 Section 912.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE IDAHO § 912.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit Rights...

30 CFR 947.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 947.774 Section 947.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE WASHINGTON § 947.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit...

30 CFR 922.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 922.774 Section 922.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE MICHIGAN § 922.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit...

30 CFR 941.774 - Revision; renewal; and transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... sale of permit rights. 941.774 Section 941.774 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION... WITHIN EACH STATE SOUTH DAKOTA § 941.774 Revision; renewal; and transfer, assignment, or sale of permit rights. (a) Part 774 of this chapter, Revision; Renewal; and Transfer, Assignment, or Sale of Permit...

Antileishmanial and immunomodulatory activity of Xylopia discreta.

PubMed

López, R; Cuca, L E; Delgado, G

2009-10-01

This study aimed at determining the in vitro antileishmanial activity of the essential oil and eight extracts obtained from Xylopia discreta. J774 and U937 macrophages were exposed to the different substances to establish the median lethal concentration (LC(50)). The median effective concentration (EC(50)) was obtained by determining the reduction of Leishmania panamensis-infected cells. A selectivity index (SI) (LC(50)/EC(50)) >or= 20 defined a specific activity for one Xylopia discreta leaf extracts and for the essential oil, being these the two that showed the highest activity (SI = 64.8 and 110, respectively in J774 cells). To assess the substances' immunomodulatory activity, pro- and anti-inflammatory soluble mediators produced after treating infected macrophages were quantified by flow cytometry. The leaf methanol extract and the essential oil induced a differential production of monocyte chemoattractant protein-1, a chemokine associated with a Leishmania-resistant phenotype (Th1).

Ultrafine particles cause cytoskeletal dysfunctions in macrophages: role of intracellular calcium

PubMed Central

Möller, Winfried; Brown, David M; Kreyling, Wolfgang G; Stone, Vicki

2005-01-01

Background Particulate air pollution is reported to cause adverse health effects in susceptible individuals. Since most of these particles are derived form combustion processes, the primary composition product is carbon with a very small diameter (ultrafine, less than 100 nm in diameter). Besides the induction of reactive oxygen species and inflammation, ultrafine particles (UFP) can cause intracellular calcium transients and suppression of defense mechanisms of alveolar macrophages, such as impaired migration or phagocytosis. Methods In this study the role of intracellular calcium transients caused by UFP was studied on cytoskeleton related functions in J774A.1 macrophages. Different types of fine and ultrafine carbon black particles (CB and ufCB, respectively), such as elemental carbon (EC90), commercial carbon (Printex 90), diesel particulate matter (DEP) and urban dust (UD), were investigated. Phagosome transport mechanisms and mechanical cytoskeletal integrity were studied by cytomagnetometry and cell viability was studied by fluorescence microscopy. Macrophages were exposed in vitro with 100 and 320 μg UFP/ml/million cells for 4 hours in serum free medium. Calcium antagonists Verapamil, BAPTA-AM and W-7 were used to block calcium channels in the membrane, to chelate intracellular calcium or to inhibit the calmodulin signaling pathways, respectively. Results Impaired phagosome transport and increased cytoskeletal stiffness occurred at EC90 and P90 concentrations of 100 μg/ml/million cells and above, but not with DEP or UD. Verapamil and W-7, but not BAPTA-AM inhibited the cytoskeletal dysfunctions caused by EC90 or P90. Additionally the presence of 5% serum or 1% bovine serum albumin (BSA) suppressed the cytoskeletal dysfunctions. Cell viability showed similar results, where co-culture of ufCB together with Verapamil, W-7, FCS or BSA produced less cell dead compared to the particles only. PMID:16202162

Essential role of protein kinase C ζ in transducing a motility signal induced by superoxide and a chemotactic peptide, fMLP

PubMed Central

Kuribayashi, Kageaki; Nakamura, Kiminori; Tanaka, Maki; Sato, Tsutomu; Kato, Junji; Sasaki, Katsunori; Takimoto, Rishu; Kogawa, Katsuhisa; Terui, Takeshi; Takayama, Tetsuji; Onuma, Takayuki; Matsunaga, Takuya; Niitsu, Yoshiro

2007-01-01

Under various pathological conditions, including infection, malignancy, and autoimmune diseases, tissues are incessantly exposed to reactive oxygen species produced by infiltrating inflammatory cells. We show augmentation of motility associated with morphological changes of human squamous carcinoma SASH1 cells, human peripheral monocytes (hPMs), and murine macrophage-like cell line J774.1 by superoxide stimulation. We also disclose that motility of hPMs and J774.1 induced by a chemotactic peptide (N-formyl-methionyl-leucyl-phenylalanine [fMLP]) was inhibited by superoxide dismutase or N-acetylcystein, indicating stimulation of motility by superoxide generated by fMLP stimulation. In these cells, protein kinase C (PKC) ζ was activated to phosphorylate RhoGDI-1, which liberated RhoGTPases, leading to their activation. These events were inhibited by dominant-negative PKCζ in SASH1 cells, myristoylated PKCζ peptides in hPMs and J774.1, or a specific inhibitor of RhoGTPase in SASH1, hPMs, and J774.1. These results suggest a new approach for manipulation of inflammation as well as tumor cell invasion by targeting this novel signaling pathway. PMID:17389234

Selectively Reduced Intracellular Proliferation of Salmonella enterica Serovar Typhimurium within APCs Limits Antigen Presentation and Development of a Rapid CD8 T Cell Response1

PubMed Central

Albaghdadi, Homam; Robinson, Nirmal; Finlay, Brett; Krishnan, Lakshmi; Sad, Subash

2014-01-01

Ag presentation to CD8+ T cells commences immediately after infection, which facilitates their rapid expansion and control of pathogen. This paradigm is not followed during infection with virulent Salmonella enterica serovar Typhimurium (ST), an intracellular bacterium that causes mortality in susceptible C57BL/6J mice within 7 days and a chronic infection in resistant mice (129 × 1SvJ). Infection of mice with OVA-expressing ST results in the development of a CD8+ T cell response that is detectable only after the second week of infection despite the early detectable bacterial burden. The mechanism behind the delayed CD8+ T cell activation was evaluated, and it was found that dendritic cells/macrophages or mice infected with ST-OVA failed to present Ag to OVA-specific CD8+ T cells. Lack of early Ag presentation was not rescued when mice or dendritic cells/macrophages were infected with an attenuated aroA mutant of ST or with mutants having defective Salmonella pathogenicity island I/II genes. Although extracellular ST proliferated extensively, the replication of ST was highly muted once inside macrophages. This muted intracellular proliferation of ST resulted in the generation of poor levels of intracellular Ag and peptide-MHC complex on the surface of dendritic cells. Additional experiments revealed that ST did not actively inhibit Ag presentation, rather it inhibited the uptake of another intracellular pathogen, Listeria monocytogenes, thereby causing inhibition of Ag presentation against L. monocytogenes. Taken together, this study reveals a dichotomy in the proliferation of ST and indicates that selectively reduced intra-cellular proliferation of virulent pathogens may be an important mechanism of immune evasion. PMID:19692639

Generation of reactive oxygen species (ROS) is a key factor for stimulation of macrophage proliferation by ceramide 1-phosphate

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arana, Lide; Gangoiti, Patricia; Ouro, Alberto

2012-02-15

We previously demonstrated that ceramide 1-phosphate (C1P) is mitogenic for fibroblasts and macrophages. However, the mechanisms involved in this action were only partially described. Here, we demonstrate that C1P stimulates reactive oxygen species (ROS) formation in primary bone marrow-derived macrophages, and that ROS are required for the mitogenic effect of C1P. ROS production was dependent upon prior activation of NADPH oxidase by C1P, which was determined by measuring phosphorylation of the p40phox subunit and translocation of p47phox from the cytosol to the plasma membrane. In addition, C1P activated cytosolic calcium-dependent phospholipase A{sub 2} and protein kinase C-{alpha}, and NADPH oxidasemore » activation was blocked by selective inhibitors of these enzymes. These inhibitors, and inhibitors of ROS production, blocked the mitogenic effect of C1P. By using BHNB-C1P (a photolabile caged-C1P analog), we demonstrate that all of these C1P actions are caused by intracellular C1P. It can be concluded that the enzyme responsible for C1P-stimulated ROS generation in bone marrow-derived macrophages is NADPH oxidase, and that this enzyme is downstream of PKC-{alpha} and cPLA{sub 2}-{alpha} in this pathway. -- Highlights: Black-Right-Pointing-Pointer Ceramide 1-phosphate (C1P) stimulates reactive oxygen species (ROS) formation. Black-Right-Pointing-Pointer The enzyme responsible for ROS generation by C1P in macrophages is NADPH oxidase. Black-Right-Pointing-Pointer NADPH oxidase lies downstream of cPLA{sub 2}-{alpha} and PKC-{alpha} in this pathway. Black-Right-Pointing-Pointer ROS generation is essential for the stimulation of macrophage proliferation by C1P.« less

9 CFR 77.4 - Application for and retention of zones.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Application for and retention of zones. 77.4 Section 77.4 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS...

9 CFR 77.4 - Application for and retention of zones.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Application for and retention of zones. 77.4 Section 77.4 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS...

9 CFR 77.4 - Application for and retention of zones.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Application for and retention of zones. 77.4 Section 77.4 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS...

9 CFR 77.4 - Application for and retention of zones.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Application for and retention of zones. 77.4 Section 77.4 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS...

9 CFR 77.4 - Application for and retention of zones.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Application for and retention of zones. 77.4 Section 77.4 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS...

AMP-Activated Protein Kinase Interacts with the Peroxisome Proliferator-Activated Receptor Delta to Induce Genes Affecting Fatty Acid Oxidation in Human Macrophages.

PubMed

Kemmerer, Marina; Finkernagel, Florian; Cavalcante, Marcela Frota; Abdalla, Dulcineia Saes Parra; Müller, Rolf; Brüne, Bernhard; Namgaladze, Dmitry

2015-01-01

AMP-activated protein kinase (AMPK) maintains energy homeostasis by suppressing cellular ATP-consuming processes and activating catabolic, ATP-producing pathways such as fatty acid oxidation (FAO). The transcription factor peroxisome proliferator-activated receptor δ (PPARδ) also affects fatty acid metabolism, stimulating the expression of genes involved in FAO. To question the interplay of AMPK and PPARδ in human macrophages we transduced primary human macrophages with lentiviral particles encoding for the constitutively active AMPKα1 catalytic subunit, followed by microarray expression analysis after treatment with the PPARδ agonist GW501516. Microarray analysis showed that co-activation of AMPK and PPARδ increased expression of FAO genes, which were validated by quantitative PCR. Induction of these FAO-associated genes was also observed upon infecting macrophages with an adenovirus coding for AMPKγ1 regulatory subunit carrying an activating R70Q mutation. The pharmacological AMPK activator A-769662 increased expression of several FAO genes in a PPARδ- and AMPK-dependent manner. Although GW501516 significantly increased FAO and reduced the triglyceride amount in very low density lipoproteins (VLDL)-loaded foam cells, AMPK activation failed to potentiate this effect, suggesting that increased expression of fatty acid catabolic genes alone may be not sufficient to prevent macrophage lipid overload.

AMP-Activated Protein Kinase Interacts with the Peroxisome Proliferator-Activated Receptor Delta to Induce Genes Affecting Fatty Acid Oxidation in Human Macrophages

PubMed Central

Kemmerer, Marina; Finkernagel, Florian; Cavalcante, Marcela Frota; Abdalla, Dulcineia Saes Parra; Müller, Rolf; Brüne, Bernhard; Namgaladze, Dmitry

2015-01-01

AMP-activated protein kinase (AMPK) maintains energy homeostasis by suppressing cellular ATP-consuming processes and activating catabolic, ATP-producing pathways such as fatty acid oxidation (FAO). The transcription factor peroxisome proliferator-activated receptor δ (PPARδ) also affects fatty acid metabolism, stimulating the expression of genes involved in FAO. To question the interplay of AMPK and PPARδ in human macrophages we transduced primary human macrophages with lentiviral particles encoding for the constitutively active AMPKα1 catalytic subunit, followed by microarray expression analysis after treatment with the PPARδ agonist GW501516. Microarray analysis showed that co-activation of AMPK and PPARδ increased expression of FAO genes, which were validated by quantitative PCR. Induction of these FAO-associated genes was also observed upon infecting macrophages with an adenovirus coding for AMPKγ1 regulatory subunit carrying an activating R70Q mutation. The pharmacological AMPK activator A-769662 increased expression of several FAO genes in a PPARδ- and AMPK-dependent manner. Although GW501516 significantly increased FAO and reduced the triglyceride amount in very low density lipoproteins (VLDL)-loaded foam cells, AMPK activation failed to potentiate this effect, suggesting that increased expression of fatty acid catabolic genes alone may be not sufficient to prevent macrophage lipid overload. PMID:26098914

Bioelectric modulation of macrophage polarization

NASA Astrophysics Data System (ADS)

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

2016-02-01

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

Early Acidification of Phagosomes Containing Brucella suis Is Essential for Intracellular Survival in Murine Macrophages

PubMed Central

Porte, Françoise; Liautard, Jean-Pierre; Köhler, Stephan

1999-01-01

Brucella suis is a facultative intracellular pathogen of mammals, residing in macrophage vacuoles. In this work, we studied the phagosomal environment of these bacteria in order to better understand the mechanisms allowing survival and multiplication of B. suis. Intraphagosomal pH in murine J774 cells was determined by measuring the fluorescence intensity of opsonized, carboxyfluorescein-rhodamine- and Oregon Green 488-rhodamine-labeled bacteria. Compartments containing live B. suis acidified to a pH of about 4.0 to 4.5 within 60 min. Acidification of B. suis-containing phagosomes in the early phase of infection was abolished by treatment of host cells with 100 nM bafilomycin A1, a specific inhibitor of vacuolar proton-ATPases. This neutralization at 1 h postinfection resulted in a 2- to 34-fold reduction of opsonized and nonopsonized viable intracellular bacteria at 4 and 6 h postinfection, respectively. Ammonium chloride and monensin, other pH-neutralizing reagents, led to comparable loss of intracellular viability. Addition of ammonium chloride at 7 h after the beginning of infection, however, did not affect intracellular multiplication of B. suis, in contrast to treatment at 1 h postinfection, where bacteria were completely eradicated within 48 h. Thus, we conclude that phagosomes with B. suis acidify rapidly after infection, and that this early acidification is essential for replication of the bacteria within the macrophage. PMID:10417172

23 CFR 774.13 - Exceptions.

Code of Federal Regulations, 2010 CFR

2010-04-01

..., WILDLIFE AND WATERFOWL REFUGES, AND HISTORIC SITES (SECTION 4(F)) § 774.13 Exceptions. The Administration... result of the consultation under 36 CFR 800.5, that such work will not adversely affect the historic... recreation lands, wildlife and waterfowl refuges, and historic sites that are made, or determinations of...

15 CFR 774.2 - [Reserved

Code of Federal Regulations, 2010 CFR

2010-01-01

...” that “incorporate” commodities or software on the Commerce Control List (Supplement No. 1 to part 774... the practice of medicine (does not include medical research). (2) Commodities or software are considered “incorporated” if the commodity or software is: Essential to the functioning of the medical...

7 CFR 774.18 - Interest rate, terms and security requirements.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 7 2010-01-01 2010-01-01 false Interest rate, terms and security requirements. 774.18..., DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.18 Interest rate, terms and security requirements. (a) Interest rate. (1) The interest rate on the loan will be zero...

Medicinal potential from in vivo and acclimatized plants of Cleome rosea.

PubMed

Simões, Claudia; De Mattos, José Carlos P; Sabino, Kátia C C; Caldeira-de-Araújo, Adriano; Coelho, Marsen G P; Albarello, Norma; Figueiredo, Solange F L

2006-02-01

Methanolic extracts obtained from different organs of Cleome rosea, collected from its natural habitat and from in vitro-propagated plants, were submitted to in vitro biological assays. Inhibition of nitric oxide (NO) production by J774 macrophages and antioxidant effects by protecting the plasmid DNA from the SnCl(2)-induced damage were evaluated. Extracts from the stem of both origins and leaf of natural plants inhibited NO production. The plasmid DNA strand breaks induced by SnCl(2) were reduced by extracts from either leaf or stem of both sources. On the other hand, root extracts did not show any kind of effects on plasmid DNA, and presented significant toxic effects to J774 cells. The results showed that C. rosea presents medicinal potential and that the acclimatization process reduces the plant toxicity both to plasmid DNA and to J774 cells, suggesting the use of biotechnology tools to obtain elite plants as source of botanical material for pharmacological and phytochemical studies.

7 CFR 774.24 - Exception.

Code of Federal Regulations, 2010 CFR

2010-01-01

... SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.24 Exception. The Agency may grant an exception to any of the requirements of this section, if the proposed change is in the best financial interest of the Government and not inconsistent with the authorizing statute or other applicable law. ...

Genomewide effects of peroxisome proliferator-activated receptor gamma in macrophages and dendritic cells--revealing complexity through systems biology.

PubMed

Cuaranta-Monroy, Ixchelt; Kiss, Mate; Simandi, Zoltan; Nagy, Laszlo

2015-09-01

Systems biology approaches have become indispensable tools in biomedical and basic research. These data integrating bioinformatic methods gained prominence after high-throughput technologies became available to investigate complex cellular processes, such as transcriptional regulation and protein-protein interactions, on a scale that had not been studied before. Immunology is one of the medical fields that systems biology impacted profoundly due to the plasticity of cell types involved and the accessibility of a wide range of experimental models. In this review, we summarize the most important recent genomewide studies exploring the function of peroxisome proliferator-activated receptor γ in macrophages and dendritic cells. PPARγ ChIP-seq experiments were performed in adipocytes derived from embryonic stem cells to complement the existing data sets and to provide comparators to macrophage data. Finally, lists of regulated genes generated from such experiments were analysed with bioinformatics and system biology approaches. We show that genomewide studies utilizing high-throughput data acquisition methods made it possible to gain deeper insights into the role of PPARγ in these immune cell types. We also demonstrate that analysis and visualization of data using network-based approaches can be used to identify novel genes and functions regulated by the receptor. The example of PPARγ in macrophages and dendritic cells highlights the crucial importance of systems biology approaches in establishing novel cellular functions for long-known signaling pathways. © 2015 Stichting European Society for Clinical Investigation Journal Foundation.

The effect of space and parabolic flight on macrophage hematopoiesis and function

NASA Technical Reports Server (NTRS)

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

1995-01-01

We used weak electric fields to monitor macrophage spreading in microgravity. Using this technique, we demonstrated that bone marrow-derived macrophages responded to microgravity within 8 s. We also showed that microgravity differentially altered two processes associated with bone marrow-derived macrophage development. Spaceflight enhanced cellular proliferation and inhibited differentiation. These data indicate that the space/microgravity environment significantly affects macrophages.

Macrophages and cellular immunity in Drosophila melanogaster

PubMed Central

Gold, Katrina S.; Brückner, Katja

2016-01-01

The invertebrate Drosophila melanogaster has been a powerful model for understanding blood cell development and immunity. Drosophila is a holometabolous insect, which transitions through a series of life stages from embryo, larva and pupa to adulthood. In spite of this, remarkable parallels exist between Drosophila and vertebrate macrophages, both in terms of development and function. More than 90% of Drosophila blood cells (hemocytes) are macrophages (plasmatocytes), making this highly tractable genetic system attractive for studying a variety of questions in macrophage biology. In vertebrates, recent findings revealed that macrophages have two independent origins: self-renewing macrophages, which reside and proliferate in local microenvironments in a variety of tissues, and macrophages of the monocyte lineage, which derive from hematopoietic stem or progenitor cells. Like vertebrates, Drosophila possesses two macrophage lineages with a conserved dual ontogeny. These parallels allow us to take advantage of the Drosophila model when investigating macrophage lineage specification, maintenance and amplification, and the induction of macrophages and their progenitors by local microenvironments and systemic cues. Beyond macrophage development, Drosophila further serves as a paradigm for understanding the mechanisms underlying macrophage function and cellular immunity in infection, tissue homeostasis and cancer, throughout development and adult life. PMID:27117654

Protein arginine methyltransferase 1 modulates innate immune responses through regulation of peroxisome proliferator-activated receptor γ-dependent macrophage differentiation.

PubMed

Tikhanovich, Irina; Zhao, Jie; Olson, Jody; Adams, Abby; Taylor, Ryan; Bridges, Brian; Marshall, Laurie; Roberts, Benjamin; Weinman, Steven A

2017-04-28

Arginine methylation is a common posttranslational modification that has been shown to regulate both gene expression and extranuclear signaling events. We recently reported defects in protein arginine methyltransferase 1 (PRMT1) activity and arginine methylation in the livers of cirrhosis patients with a history of recurrent infections. To examine the role of PRMT1 in innate immune responses in vivo , we created a cell type-specific knock-out mouse model. We showed that myeloid-specific PRMT1 knock-out mice demonstrate higher proinflammatory cytokine production and a lower survival rate after cecal ligation and puncture. We found that this defect is because of defective peroxisome proliferator-activated receptor γ (PPARγ)-dependent M2 macrophage differentiation. PPARγ is one of the key transcription factors regulating macrophage polarization toward a more anti-inflammatory and pro-resolving phenotype. We found that PRMT1 knock-out macrophages failed to up-regulate PPARγ expression in response to IL4 treatment resulting in 4-fold lower PPARγ expression in knock-out cells than in wild-type cells. Detailed study of the mechanism revealed that PRMT1 regulates PPARγ gene expression through histone H4R3me2a methylation at the PPARγ promoter. Supplementing with PPARγ agonists rosiglitazone and GW1929 was sufficient to restore M2 differentiation in vivo and in vitro and abrogated the difference in survival between wild-type and PRMT1 knock-out mice. Taken together these data suggest that PRMT1-dependent regulation of macrophage PPARγ expression contributes to the infection susceptibility in PRMT1 knock-out mice. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The in vitro kinetics of the interactions between PEG-ylated magnetic-fluid-loaded liposomes and macrophages.

PubMed

Martina, Marie-Sophie; Nicolas, Valerie; Wilhelm, Claire; Ménager, Christine; Barratt, Gillian; Lesieur, Sylviane

2007-10-01

Binding and uptake kinetics of magnetic-fluid-loaded liposomes (MFL) by endocytotic cells were investigated in vitro on the model cell-line J774. MFL consisted of unilamellar phosphatidylcholine vesicles (mean hydrodynamic diameter close to 200nm) encapsulating 8-nm nanocrystals of maghemite (gamma-Fe(2)O(3)) and sterically stabilized by introducing 5mol% of distearylphosphatidylcholine poly(ethylene glycol)(2,000) (DSPE-PEG(2,000)) in the vesicle bilayer. The association processes with living macrophages were followed at two levels. On one hand, the lipid vesicles were imaged by confocal fluorescence microscopy. For this purpose 1mol% of rhodamine-marked phosphatidylethanolamine was added to the liposome composition. On the other hand, the iron oxide particles associated with cells were independently quantified by magnetophoresis. All the experiments were similarly performed with PEG-ylated or conventional MFL to point out the role of polymer coating. The results showed cell association with both types of liposomes resulting from binding followed by endocytosis. Steric stabilization by PEG chains reduced binding efficiency limiting the amount of MFL internalized by the macrophages. In contrast, PEG coating did not change the kinetics of endocytosis which exhibited the same first-order rate constant for both conventional and PEG-ylated liposomes. Moreover, lipids and iron oxide particle uptakes were perfectly correlated, indicating that MFL vesicle structure and encapsulation rate were preserved upon cell penetration.

Apoptosis of Trypanosoma musculi co-cultured with LPS activated macrophages: enhanced expression of nitric oxide synthase INF-gamma and caspase.

PubMed

Gugssa, A; Gebru, S; Lee, C M; Baccetti, B; Anderson, W

2005-08-01

Trypanosoma musculi-macrophage co-cultures were studied to investigate the biological role of lipopolysaccharide (LPS) induced cytokines in controlling the proliferation of parasites in vitro. Macrophages, isolated by peritoneal lavage, sustained the growth and proliferation of the parasites. Macrophages activated with LPS were characterized by up-regulation of nitric oxide synthase (iNOS) and phagocytosis of fluorescent latex spheres. Activated macrophages showed marked inhibition of the association and proliferation of the parasites. The LPS treated macrophages produced cytokines, especially interferon gamma (INF-gamma), which was detected by Western blot. Trypanosomes, inhibited from association with macrophages, did not proliferate and instead formed clusters held together by their flagella. Cells in these clusters were apoptotic, as demonstrated by the Apoptag reaction and gel fragmentation assay. In addition, high levels of caspase 8 and caspase 3 were shown in floating trypanosome clusters. The results would suggest that INF-gamma and other cytokines released by activated macrophages, possibly functioning through the INF-gammaR1, Fas ligand, CD95 or other death ligands in the trypanosome plasma membrane initiates the apoptosis cascade in trypanosomes.

Foxp3-positive macrophages display immunosuppressive properties and promote tumor growth

PubMed Central

Zorro Manrique, Soraya; Duque Correa, Maria Adelaida; Hoelzinger, Dominique B.; Dominguez, Ana Lucia; Mirza, Noweeda; Lin, Hsi-Hsien; Stein-Streilein, Joan; Gordon, Siamon

2011-01-01

Regulatory T cells (T reg cells) are characterized by the expression of the forkhead lineage-specific transcription factor Foxp3, and their main function is to suppress T cells. While evaluating T reg cells, we identified a population of Foxp3-positive cells that were CD11b+F4/80+CD68+, indicating macrophage origin. These cells were observed in spleen, lymph nodes, bone marrow, thymus, liver, and other tissues of naive animals. To characterize this subpopulation of macrophages, we devised a strategy to purify CD11b+F4/80+Foxp3+ macrophages using Foxp3-GFP mice. Analysis of CD11b+F4/80+Foxp3+ macrophage function indicated that these cells inhibited the proliferation of T cells, whereas Foxp3− macrophages did not. Suppression of T cell proliferation was mediated through soluble factors. Foxp3− macrophages acquired Foxp3 expression after activation, which conferred inhibitory properties that were indistinguishable from natural Foxp3+ macrophages. The cytokine and transcriptional profiles of Foxp3+ macrophages were distinct from those of Foxp3− macrophages, indicating that these cells have different biological functions. Functional in vivo analyses indicated that CD11b+F4/80+Foxp3+ macrophages are important in tumor promotion and the induction of T reg cell conversion. For the first time, these studies demonstrate the existence of a distinct subpopulation of naturally occurring macrophage regulatory cells in which expression of Foxp3 correlates with suppressive function. PMID:21670203

30 CFR 774.12 - Post-permit issuance information requirements for permittees.

Code of Federal Regulations, 2010 CFR

2010-07-01

... for permittees. 774.12 Section 774.12 Mineral Resources OFFICE OF SURFACE MINING RECLAMATION AND ENFORCEMENT, DEPARTMENT OF THE INTERIOR SURFACE COAL MINING AND RECLAMATION OPERATIONS PERMITS AND COAL... competent jurisdiction grants a stay of the cessation order and the stay remains in effect. (c) Within 60...

Biochemical Characterization, Action on Macrophages, and Superoxide Anion Production of Four Basic Phospholipases A2 from Panamanian Bothrops asper Snake Venom

PubMed Central

Rueda, Aristides Quintero; Rodríguez, Isela González; Arantes, Eliane C.; Setúbal, Sulamita S.; Calderon, Leonardo de A.; Zuliani, Juliana P.; Stábeli, Rodrigo G.; Soares, Andreimar M.

2013-01-01

Bothrops asper (Squamata: Viperidae) is the most important venomous snake in Central America, being responsible for the majority of snakebite accidents. Four basic PLA2s (pMTX-I to -IV) were purified from crude venom by a single-step chromatography using a CM-Sepharose ion-exchange column (1.5 × 15 cm). Analysis of the N-terminal sequence demonstrated that pMTX-I and III belong to the catalytically active Asp49 phospholipase A2 subclass, whereas pMTX-II and IV belong to the enzymatically inactive Lys49 PLA2s-like subclass. The PLA2s isolated from Panama Bothrops asper venom (pMTX-I, II, III, and IV) are able to induce myotoxic activity, inflammatory reaction mainly leukocyte migration to the muscle, and induce J774A.1 macrophages activation to start phagocytic activity and superoxide production. PMID:23509779

Administration of PDE4 Inhibitors Suppressed the Pannus-Like Inflammation by Inhibition of Cytokine Production by Macrophages and Synovial Fibroblast Proliferation

PubMed Central

Kobayashi, Katsuya; Suda, Toshio; Manabe, Haruhiko; Miki, Ichiro

2007-01-01

A marked proliferation of synovial fibroblasts in joints leads to pannus formation in rheumatoid arthritis (RA). Various kinds of cytokines are produced in the pannus. The purpose of this study is to elucidate the effects of phosphodiesterase 4 (PDE4) inhibitors in a new animal model for the evaluation of pannus formation and cytokine production in the pannus. Mice sensitized with methylated bovine serum albumin (mBSA) were challenged by subcutaneous implantation of a membrane filter soaked in mBSA solution in the back of the mice. Drugs were orally administered for 10 days. The granuloma formed around the filter was collected on day 11. It was chopped into pieces and cultured in vitro for 24 hr. The cytokines were measured in the supernatants. The type of cytokines produced in the granuloma was quite similar to those produced in pannus in RA. Both PDE4 inhibitors, KF66490 and SB207499, suppressed the production of IL-1β, TNF-α, and IL-12, and the increase in myeloperoxidase activity, a marker enzyme for neutrophils and hydroxyproline content. Compared to leflunomide, PDE4 inhibitors more strongly suppressed IL-12 production and the increase in myeloperoxidase activity. PDE4 inhibitors also inhibited lipopolysaccharide-induced TNF-α and IL-12 production from thioglycolate-induced murine peritoneal macrophages and the proliferation of rat synovial fibroblasts. These results indicate this model makes it easy to evaluate the effect of drugs on various cytokine productions in a granuloma without any purification step and may be a relevant model for evaluating novel antirheumatic drugs on pannus formation in RA. PDE4 inhibitors could have therapeutic effects on pannus formation in RA by inhibition of cytokine production by macrophages and synovial fibroblast proliferation. PMID:18274640

Administration of PDE4 inhibitors suppressed the pannus-like inflammation by inhibition of cytokine production by macrophages and synovial fibroblast proliferation.

PubMed

Kobayashi, Katsuya; Suda, Toshio; Manabe, Haruhiko; Miki, Ichiro

2007-01-01

A marked proliferation of synovial fibroblasts in joints leads to pannus formation in rheumatoid arthritis (RA). Various kinds of cytokines are produced in the pannus. The purpose of this study is to elucidate the effects of phosphodiesterase 4 (PDE4) inhibitors in a new animal model for the evaluation of pannus formation and cytokine production in the pannus. Mice sensitized with methylated bovine serum albumin (mBSA) were challenged by subcutaneous implantation of a membrane filter soaked in mBSA solution in the back of the mice. Drugs were orally administered for 10 days. The granuloma formed around the filter was collected on day 11. It was chopped into pieces and cultured in vitro for 24 hr. The cytokines were measured in the supernatants. The type of cytokines produced in the granuloma was quite similar to those produced in pannus in RA. Both PDE4 inhibitors, KF66490 and SB207499, suppressed the production of IL-1beta, TNF-alpha, and IL-12, and the increase in myeloperoxidase activity, a marker enzyme for neutrophils and hydroxyproline content. Compared to leflunomide, PDE4 inhibitors more strongly suppressed IL-12 production and the increase in myeloperoxidase activity. PDE4 inhibitors also inhibited lipopolysaccharide-induced TNF-alpha and IL-12 production from thioglycolate-induced murine peritoneal macrophages and the proliferation of rat synovial fibroblasts. These results indicate this model makes it easy to evaluate the effect of drugs on various cytokine productions in a granuloma without any purification step and may be a relevant model for evaluating novel antirheumatic drugs on pannus formation in RA. PDE4 inhibitors could have therapeutic effects on pannus formation in RA by inhibition of cytokine production by macrophages and synovial fibroblast proliferation.

Modelling the host-pathogen interactions of macrophages and Candida albicans using Game Theory and dynamic optimization.

PubMed

Dühring, Sybille; Ewald, Jan; Germerodt, Sebastian; Kaleta, Christoph; Dandekar, Thomas; Schuster, Stefan

2017-07-01

The release of fungal cells following macrophage phagocytosis, called non-lytic expulsion, is reported for several fungal pathogens. On one hand, non-lytic expulsion may benefit the fungus in escaping the microbicidal environment of the phagosome. On the other hand, the macrophage could profit in terms of avoiding its own lysis and being able to undergo proliferation. To analyse the causes of non-lytic expulsion and the relevance of macrophage proliferation in the macrophage- Candida albicans interaction, we employ Evolutionary Game Theory and dynamic optimization in a sequential manner. We establish a game-theoretical model describing the different strategies of the two players after phagocytosis. Depending on the parameter values, we find four different Nash equilibria and determine the influence of the systems state of the host upon the game. As our Nash equilibria are a direct consequence of the model parameterization, we can depict several biological scenarios. A parameter region, where the host response is robust against the fungal infection, is determined. We further apply dynamic optimization to analyse whether macrophage mitosis is relevant in the host-pathogen interaction of macrophages and C. albicans For this, we study the population dynamics of the macrophage- C. albicans interactions and the corresponding optimal controls for the macrophages, indicating the best macrophage strategy of switching from proliferation to attacking fungal cells. © 2017 The Author(s).

Macrophages and cellular immunity in Drosophila melanogaster.

PubMed

Gold, Katrina S; Brückner, Katja

2015-12-01

The invertebrate Drosophila melanogaster has been a powerful model for understanding blood cell development and immunity. Drosophila is a holometabolous insect, which transitions through a series of life stages from embryo, larva and pupa to adulthood. In spite of this, remarkable parallels exist between Drosophila and vertebrate macrophages, both in terms of development and function. More than 90% of Drosophila blood cells (hemocytes) are macrophages (plasmatocytes), making this highly tractable genetic system attractive for studying a variety of questions in macrophage biology. In vertebrates, recent findings revealed that macrophages have two independent origins: self-renewing macrophages, which reside and proliferate in local microenvironments in a variety of tissues, and macrophages of the monocyte lineage, which derive from hematopoietic stem or progenitor cells. Like vertebrates, Drosophila possesses two macrophage lineages with a conserved dual ontogeny. These parallels allow us to take advantage of the Drosophila model when investigating macrophage lineage specification, maintenance and amplification, and the induction of macrophages and their progenitors by local microenvironments and systemic cues. Beyond macrophage development, Drosophila further serves as a paradigm for understanding the mechanisms underlying macrophage function and cellular immunity in infection, tissue homeostasis and cancer, throughout development and adult life. Copyright © 2016. Published by Elsevier Ltd.

Embryonic and adult-derived resident cardiac macrophages are maintained through distinct mechanisms at steady state and during inflammation

PubMed Central

Epelman, Slava; Lavine, Kory J.; Beaudin, Anna E.; Sojka, Dorothy K.; Carrero, Javier A.; Calderon, Boris; Brija, Thaddeus; Gautier, Emmanuel L.; Ivanov, Stoyan; Satpathy, Ansuman T.; Schilling, Joel D.; Schwendener, Reto; Sergin, Ismail; Razani, Babak; Forsberg, E. Camilla; Yokoyama, Wayne; Unanue, Emil R.; Colonna, Marco; Randolph, Gwendalyn J.; Mann, Douglas L.

2014-01-01

Summary Cardiac macrophages are crucial for tissue repair after cardiac injury but have not been well characterized. Here we identify four populations of cardiac macrophages. At steady state, resident macrophages were primarily maintained through local proliferation. However, after macrophage depletion or during cardiac inflammation, Ly6chi monocytes contributed to all four macrophage populations, whereas resident macrophages also expanded numerically through proliferation. Genetic fate mapping revealed that yolk-sac and fetal monocyte progenitors gave rise to the majority of cardiac macrophages, and the heart was among a minority of organs in which substantial numbers of yolk-sac macrophages persisted in adulthood. CCR2 expression and dependence distinguished cardiac macrophages of adult monocyte versus embryonic origin. Transcriptional and functional data revealed that monocyte-derived macrophages coordinate cardiac inflammation, while playing redundant but lesser roles in antigen sampling and efferocytosis. These data highlight the presence of multiple cardiac macrophage subsets, with different functions, origins and strategies to regulate compartment. PMID:24439267

Evaluation of in vitro anti-inflammatory effects of crude ginger and rosemary extracts obtained through supercritical CO2 extraction on macrophage and tumor cell line: the influence of vehicle type.

PubMed

Justo, Oselys Rodriguez; Simioni, Patricia Ucelli; Gabriel, Dirce Lima; Tamashiro, Wirla Maria da Silva Cunha; Rosa, Paulo de Tarso Vieira; Moraes, Ângela Maria

2015-10-29

Numerous plants from have been investigated due to their anti-inflammatory activity and, among then, extracts or components of ginger (Zingiber officinale Roscoe) and rosemary (Rosmarinus officinalis L.), sources of polyphenolic compounds. 6-gingerol from ginger rhizome and carnosic acid and carnosol from rosemary leaves present anti-tumor, anti-inflammatory and antioxidant activities. However, the evaluation of the mechanisms of action of these and other plant extracts is limited due to their high hydrophobicity. Dimethylsulfoxide (DMSO) is commonly used as a vehicle of liposoluble materials to mammalian cells in vitro, presenting enhanced cell penetration. Liposomes are also able to efficiently deliver agents to mammalian cells, being capable to incorporate in their structure not only hydrophobic molecules, but also hydrophilic and amphiphilic compounds. Another strategy is based on the use of Pluronic F-68, a biocompatible low-foaming, non-ionic surfactant, to disperse hydrophobic components. Here, these three delivery approaches were compared to analyze their influence on the in vitro anti-inflammatory effects of ginger and rosemary extracts, at different concentrations, on primary mammalian cells and on a tumor cell line. Ginger and rosemary extracts free of organic solvents were obtained by supercritical fluid extraction and dispersed in DMSO, Pluronic F-68 or liposomes, in variable concentrations. Cell viability, production of inflammatory mediators and nitric oxide (NO) release were measured in vitro on J774 cell line and murine macrophages primary culture stimulated with bacterial lipopolysaccharide and interferon-γ after being exposed or not to these extracts. Ginger and rosemary extracts obtained by supercritical CO2 extraction inhibited the production of pro-inflammatory cytokines and the release of NO by peritoneal macrophages and J774 cells. The delivery vehicles influenced the anti-inflammatory effects. Comparatively, the ginger extract showed the

Differential impact of diabetes mellitus type II and arterial hypertension on collateral artery growth and concomitant macrophage accumulation.

PubMed

Ito, Wulf D; Lund, Natalie; Sager, Hendrik; Becker, Wiebke; Wenzel, Ulrich

2015-01-01

Diabetes mellitus type II and arterial hypertension are major risk factors for peripheral arterial disease and have been considered to reduce collateral growth (arteriogenesis). Collateral growth proceeds through different stages. Vascular proliferation and macrophage accumulation are hallmarks of early collateral growth. We here compare the impact of arterial hypertension and diabetes mellitus type II on collateral proliferation (Brdu incorporation) and macrophage accumulation (ED 2 staining) as well as collateral vessel function (collateral conductance) in a rat model of peripheral vascular disease (femoral artery occlusion), diabetes mellitus type II (Zucker fatty diabetic rats and Zucker lean rat controls) and arterial hypertension (induced via clip placement around the right renal arteriy). We furthermore tested the impact of monocyte chemoattractant protein-1 (MCP‑1) on collateral proliferation and macrophage accumulation in these models Diabetic animals showed reduced vascular proliferation and macrophage accumulation, which however did not translate into a change of collateral conductance. Hypertensive animals on the contrary had reduced collateral conductances without altered macrophage accumulation and only a marginal reduction in collateral proliferation. Infusion of MCP‑1 only enhanced vascular proliferation in diabetic animals. These findings illustrate that impaired monocyte/macrophage recruitment is responsible for reduced collateral growth under diabetic conditions but not in arterial hypertension suggesting that diabetes mellitus in particular affects early stages of collateral growth whereas hypertension has its impact on later remodeling stages. Successful pro-arteriogenic treatment strategies in a patient population that presents with diabetes mellitus and arterial hypertension need to address different stages of collateral growth and thus different molecular and cellular targets simultaneously.

Dextran loading protects macrophages from lipid peroxidation and induces a Keap1/Nrf2/ARE-dependent antioxidant response.

PubMed

Chechushkov, Anton; Zaitseva, Natalia; Vorontsova, Elena; Kozhin, Petr; Menshchikova, Elena; Shkurupiy, Vyacheslav

2016-12-01

Linear dextrans are often proposed as drug delivery systems with milder adverse effects and lower effective drug concentrations. Linear dextrans are polysaccharides that can potentially be used to load macrophages with drugs to transport them to a site of inflammation. Recently, it was reported that dextrans may exert a protective effect vis-à-vis drug cytotoxicity and during wound healing. The aim of the current work was to evaluate molecular mechanisms of action of dextrans that may be relevant to the cytoprotective effects. We determined the effect of treatment with 40- or 70-kDa dextran on production of reactive oxygen species, lipid peroxidation, and lysosomal pH in the J774 macrophage cell line. In addition, induction of Keap1/Nrf2/ARE and autophagic activity were evaluated. Dextrans of both molecular weights protected the cells from oxidative stress induced by cumene hydroperoxide and from lysosomal stress induced by ammonium chloride. The effect was associated with induction of the Keap1/Nrf2/ARE signaling pathway. Furthermore, dextran stimulated autophagy in a dose-dependent manner but inhibited the autophagosome-lysosome fusion in a time-dependent manner. This study shows possible cytoprotective effects of dextran under oxidative stress, and these findings may be used for the development of novel (dextran-based) drug delivery approaches. Copyright © 2016 Elsevier Inc. All rights reserved.

Effects of macrophage colony-stimulating factor on macrophages and their related cell populations in the osteopetrosis mouse defective in production of functional macrophage colony-stimulating factor protein.

PubMed Central

Umeda, S.; Takahashi, K.; Shultz, L. D.; Naito, M.; Takagi, K.

1996-01-01

The development of macrophage populations in osteopetrosis (op) mutant mice defective in production of functional macrophage colony-stimulating factor (M-CSF) and the response of these cell populations to exogenous M-CSF were used to classify macrophages into four groups: 1) monocytes, monocyte-derived macrophages, and osteoclasts, 2) MOMA-1-positive macrophages, 3) ER-TR9-positive macrophages, and 4) immature tissue macrophages. Monocytes, monocyte-derived macrophages, osteoclasts in bone, microglia in brain, synovial A cells, and MOMA-1- or ER-TR9-positive macrophages were deficient in op/op mice. The former three populations expanded to normal levels in op/op mice after daily M-CSF administration, indicating that they are developed and differentiated due to the effect of M-CSF supplied humorally. In contrast, the other cells did not respond or very slightly responded to M-CSF, and their development seems due to either M-CSF produced in situ or expression of receptor for M-CSF. Macrophages present in tissues of the mutant mice were immature and appear to be regulated by either granulocyte/macrophage colony-stimulating factor and/or interleukin-3 produced in situ or receptor expression. Northern blot analysis revealed different expressions of GM-CSF and IL-3 mRNA in various tissues of the op/op mice. However, granulocyte/macrophage colony-stimulating factor and interleukin-3 in serum were not detected by enzyme-linked immunosorbent assay. The immature macrophages differentiated and matured into resident macrophages after M-CSF administration, and some of these cells proliferated in response to M-CSF. Images Figure 4 Figure 6 Figure 8 Figure 10 Figure 11 PMID:8701995

AhR-dependent secretion of PDGF-BB by human classically activated macrophages exposed to DEP extracts stimulates lung fibroblast proliferation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaguin, Marie; Fardel, Olivier; Pôle Biologie, Centre Hospitalier Universitaire

2015-06-15

Lung diseases are aggravated by exposure to diesel exhaust particles (DEPs) found in air pollution. Macrophages are thought to play a crucial role in lung immune response to these pollutants, even if the mechanisms involved remain incompletely characterized. In the present study, we demonstrated that classically and alternative human macrophages (MΦ) exhibited increased secretion of PDGF-B in response to DEP extract (DEPe). This occurred via aryl hydrocarbon receptor (AhR)-activation because DEPe-induced PDGF-B overexpression was abrogated after AhR expression knock-down by RNA interference, in both M1 and M2 polarizing MΦ. In addition, TCDD and benzo(a)pyrene, two potent AhR ligands, also significantlymore » increased mRNA expression of PDGF-B in M1 MΦ, whereas some weak ligands of AhR did not. We next evaluated the impact of conditioned media (CM) from MΦ culture exposed to DEPe or of recombinant PDGF-B onto lung fibroblast proliferation. The tyrosine kinase inhibitor, AG-1295, prevents phosphorylations of PDGF-Rβ, AKT and ERK1/2 and the proliferation of MRC-5 fibroblasts induced by recombinant PDGF-B and by CM from M1 polarizing MΦ, strongly suggesting that the PDGF-BB secreted by DEPe-exposed MΦ is sufficient to activate the PDGF-Rβ pathway of human lung fibroblasts. In conclusion, we demonstrated that human MΦ, whatever their polarization status, secrete PDGF-B in response to DEPe and that PDGF-B is a target gene of AhR. Therefore, induction of PDGF-B by DEP may participate in the deleterious effects towards human health triggered by such environmental urban contaminants. - Highlights: • PDGF-B expression and secretion are increased by DEPe exposure in human M1 and M2 MΦ. • DEPe-induced PDGF-B expression is aryl-hydrocarbon-dependent. • DEPe-exposed M1 MΦ secrete sufficient PDGF-B to increase lung fibroblast proliferation.« less

Evaluation of the efficacy of photodynamic antimicrobial therapy using a phenothiazine compound and Laser (λ=660ηm) on the interface: macrophage vs S. aureus

NASA Astrophysics Data System (ADS)

de Oliveira, Susana C. P. S.; Monteiro, Juliana S. C.; Pires-Santos, Gustavo M.; Sampaio, Fernando José P.; Zanin, Fátima Antônia A.; Pinheiro, Antônio L. B.

2015-03-01

Nowadays photodynamic inactivation has been proposed as an alternative treatment for localized bacterial infections as a response to the problem of antibiotic resistance. Much is already known about the photodynamic inactivation of microorganisms: both antibiotic-sensitive and -resistant strains can be successfully photoinactivated and there is the additional advantage that repeated photosensitization of bacterial cells does not induce a selection of resistant strains. Staphylococcus spp. are opportunistic microorganisms known for their capacity to develop resistance against antimicrobial agents. The emergence of resistant strains of bacteria such as methicillin-resistant Staphylococcus aureus (MRSA) poses a major challenge to healthcare. MRSA is a major cause of hospital-acquired infection throughout the world and is now also prevalent in the community as well as nursing and residential homes. The aim of this study was to evaluate the phagocytic function of macrophages J774 against S. aureus in the presence and absence of AmPDT with phenothiazine compound (12.5 μg/mL) and low level laser (λ=660nm, 12 J/cm²). Experimental groups: Control group (L-P-), Phenothiazine group (L-P+) Laser group (L+P-), AmPDT group (L+P+).The tests presented in this study were performed in triplicate. This study showed that AmPDT induced bacterial death in about 80% as well as increasing phagocytic capacity of macrophages by approximately 20% and enhanced the antimicrobial activity by approximately 50% compared to the control group and enabling more intense oxidative burst.

Microparticles prepared with 50-190kDa chitosan as promising non-toxic carriers for pulmonary delivery of isoniazid.

PubMed

Oliveira, Paula M; Matos, Breno N; Pereira, Priscilla A T; Gratieri, Taís; Faccioli, Lucia H; Cunha-Filho, Marcílio S S; Gelfuso, Guilherme M

2017-10-15

Chitosan biocompatibility and mucoadhesiveness make it an ideal polymer for antituberculotic drugs microcapsulation for pulmonary delivery. Yet, previous study indicated toxicity problems to J-774.1-cells treated with some medium molecular weight (190-310kDa) chitosan microparticles. As polymer molecular weight is a crucial factor to be considered, this paper describes the preparation and characterization of chitosan (50-190kDa) microparticles containing isoniazid (INH). Cytotoxicity assays were also performed on murine peritoneal (J-774.1) and alveolar (AMJ2-C11) macrophages cell lines, followed by cytokines detection from AMJ2-C11 cells. Spray-drying process produced mucoadhesive microparticles from 3.2μm to 3.9μm, entrapping more than 89% of the drug and preserving their chemical stability. Drug release behavior could be controlled by the use of cross-linked or uncross-linked chitosan, the latter leading to a rapid drug release. Mucoadhesive potential of the microparticles was characterized following in vitro and ex vivo assays. Finally, a significant reduction on toxicity against peritoneal macrophages and no toxic effect on alveolar macrophages with use of such microparticles were observed. In conclusion, 50-190kDa chitosan microparticles may act as promising non-cytotoxic carriers for pulmonary delivery of INH showing marked alveoli macrophage activation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Macrophage Differentiation in Normal and Accelerated Wound Healing.

PubMed

Kotwal, Girish J; Chien, Sufan

2017-01-01

Chronic wounds pose considerable public health challenges and burden. Wound healing is known to require the participation of macrophages, but mechanisms remain unclear. The M1 phenotype macrophages have a known scavenger function, but they also play multiple roles in tissue repair and regeneration when they transition to an M2 phenotype. Macrophage precursors (mononuclear cells/monocytes) follow the influx of PMN neutrophils into a wound during the natural wound-healing process, to become the major cells in the wound. Natural wound-healing process is a four-phase progression consisting of hemostasis, inflammation, proliferation, and remodeling. A lag phase of 3-6 days precedes the remodeling phase, which is characterized by fibroblast activation and finally collagen production. This normal wound-healing process can be accelerated by the intracellular delivery of ATP to wound tissue. This novel ATP-mediated acceleration arises due to an alternative activation of the M1 to M2 transition (macrophage polarization), a central and critical feature of the wound-healing process. This response is also characterized by an early increased release of pro-inflammatory cytokines (TNF, IL-1 beta, IL-6), a chemokine (MCP-1), an activation of purinergic receptors (a family of plasma membrane receptors found in almost all mammalian cells), and an increased production of platelets and platelet microparticles. These factors trigger a massive influx of macrophages, as well as in situ proliferation of the resident macrophages and increased synthesis of VEGFs. These responses are followed, in turn, by rapid neovascularization and collagen production by the macrophages, resulting in wound covering with granulation tissue within 24 h.

Targeting Androgen Receptor to Suppress Macrophage-induced EMT and Benign Prostatic Hyperplasia (BPH) Development

PubMed Central

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

2012-01-01

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

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

PubMed

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

2012-10-01

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

Mitofusin 2 decreases intracellular lipids in macrophages by regulating peroxisome proliferator-activated receptor-γ

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Chun; Ge, Beihai; He, Chao

2014-07-18

Highlights: • Mfn2 decreases cellular lipid accumulation by activating cholesterol transporters. • PPARγ is involved in the Mfn2-mediated increase of cholesterol transporter expressions. • Inactivation of ERK1/2 and p38 is involved in Mfn2-induced PPARγ expression. - Abstract: Mitofusin 2 (Mfn2) inhibits atherosclerotic plaque formation, but the underlying mechanism remains elusive. This study aims to reveal how Mfn2 functions in the atherosclerosis. Mfn2 expression was found to be significantly reduced in arterial atherosclerotic lesions of both mice and human compared with healthy counterparts. Here, we observed that Mfn2 increased cellular cholesterol transporter expression in macrophages by upregulating peroxisome proliferator-activated receptor-γ, anmore » effect achieved at least partially by inhibiting extracellular signal-regulated kinase1/2 (ERK1/2) and p38 mitogen-activated protein kinases (MAPKs) pathway. These findings provide insights into potential mechanisms of Mfn2-mediated alterations in cholesterol transporter expression, which may have significant implications for the treatment of atherosclerotic heart disease.« less

An M1-like Macrophage Polarization in Decidual Tissue during Spontaneous Preterm Labor That Is Attenuated by Rosiglitazone Treatment.

PubMed

Xu, Yi; Romero, Roberto; Miller, Derek; Kadam, Leena; Mial, Tara N; Plazyo, Olesya; Garcia-Flores, Valeria; Hassan, Sonia S; Xu, Zhonghui; Tarca, Adi L; Drewlo, Sascha; Gomez-Lopez, Nardhy

2016-03-15

Decidual macrophages are implicated in the local inflammatory response that accompanies spontaneous preterm labor/birth; however, their role is poorly understood. We hypothesized that decidual macrophages undergo a proinflammatory (M1) polarization during spontaneous preterm labor and that PPARγ activation via rosiglitazone (RSG) would attenuate the macrophage-mediated inflammatory response, preventing preterm birth. In this study, we show that: 1) decidual macrophages undergo an M1-like polarization during spontaneous term and preterm labor; 2) anti-inflammatory (M2)-like macrophages are more abundant than M1-like macrophages in decidual tissue; 3) decidual M2-like macrophages are reduced in preterm pregnancies compared with term pregnancies, regardless of the presence of labor; 4) decidual macrophages express high levels of TNF and IL-12 but low levels of peroxisome proliferator-activated receptor γ (PPARγ) during spontaneous preterm labor; 5) decidual macrophages from women who underwent spontaneous preterm labor display plasticity by M1↔M2 polarization in vitro; 6) incubation with RSG reduces the expression of TNF and IL-12 in decidual macrophages from women who underwent spontaneous preterm labor; and 7) treatment with RSG reduces the rate of LPS-induced preterm birth and improves neonatal outcomes by reducing the systemic proinflammatory response and downregulating mRNA and protein expression of NF-κB, TNF, and IL-10 in decidual and myometrial macrophages in C57BL/6J mice. In summary, we demonstrated that decidual M1-like macrophages are associated with spontaneous preterm labor and that PPARγ activation via RSG can attenuate the macrophage-mediated proinflammatory response, preventing preterm birth and improving neonatal outcomes. These findings suggest that the PPARγ pathway is a new molecular target for future preventative strategies for spontaneous preterm labor/birth. Copyright © 2016 by The American Association of Immunologists, Inc.

In vivo responses of macrophages and perisinusoidal cells to cholestatic liver injury.

PubMed Central

Hines, J. E.; Johnson, S. J.; Burt, A. D.

1993-01-01

We investigated the response of macrophages and perisinusoidal (Ito) cells (PSCs) during the development of secondary biliary cirrhosis after ligation and division of the common bile duct. Liver tissue was obtained from three groups of male Wistar rats: 1) untreated controls (n = 3); 2) common bile duct-ligated (CBDL) animals (n = 15); and 3) sham-operated controls (n = 15). Material from animal groups 2 and 3 was obtained on days 3, 7, 14, 21, and 28 after operation; in all animals 5-bromo-2-deoxyuridine was administered intraperitoneally before death. Monocytes and macrophages were detected using the monoclonal antibody ED1 and tissue macrophages using the antibody ED2. Cell proliferation within the macrophage population was demonstrated by double labeling for ED2 and incorporated 5-bromo-2-deoxyuridine. PSCs were demonstrated in tissue sections by immunolocalization of desmin; proliferating PSCs were identified by double labeling for desmin and incorporated 5-bromo-2-deoxyuridine. Evidence of phenotypic modulation of PSCs was sought using anti-alpha-smooth muscle actin (alpha-SMA) antibody. Increased numbers of ED1- and ED2-positive cells were seen in CBDL animals at all time points. Local proliferation of macrophages could be identified and reached a peak at day 3, thereafter falling toward control values. Compared with those of controls, livers of CBDL animals showed increased numbers of desmin-positive PSCs in periportal zones from day 3 on, reaching a peak at day 14 (127.8 +/- 10.99 cells/0.635 mm2) and followed by a plateau. PSC proliferation peaked at days 3 and 7 (labeling indices 11.2% and 11.2%, respectively) and thereafter fell toward control values; no expansion of the PSC population was seen in sham-operated rats. Increased alpha-SMA-positive cells were also noted from day 3, with a peak at day 21 (231.1 +/- 11.52 cells/0.635 mm2) and followed by a plateau. En face labeling experiments in days 14, 21, and 28 CBDL animals showed cells co

In vitro activity of synthetic tetrahydroindeno[2,1-c]quinolines on Leishmania mexicana.

PubMed

Hernández-Chinea, Concepción; Carbajo, Erika; Sojo, Felipe; Arvelo, Francisco; Kouznetsov, Vladimir V; Romero-Bohórquez, Arnold R; Romero, Pedro J

2015-12-01

New synthetic compounds based on tetrahydroindenoquinoline structure were evaluated for their in vitro antileishmanial activities. The seven compounds assayed have antiproliferative activities against promastigotes of Leishmania mexicana. Compound 1 and 3 were the most active (IC50 1.0 μg/ml) and showed high selectivity towards the parasite. These compounds were selected to evaluate their effect on promastigote morphology and mitochondrial transmembrane potential as well as on the amastigote capability to survive into macrophages J774 cell line. Whereas compound 1 affected the promastigote cell cycle, compound 3 induced morphological changes and the total collapse of the mitochondrial transmembrane potential, a hallmark of apoptosis. Both compounds also affected the amastigote form of the parasite, decreasing their survival rate in J774 macrophages. Due to the greatest selectivity index, the apparent effect as apoptotic inducer and its sustained inhibition on intracellular amastigote replication, compound 3 is the best candidate to be tested in vivo. This compound is worth considering for the development of new antileishmanial drugs. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Uptake and intracellular activity of AM-1155 in phagocytic cells.

PubMed Central

Yamamoto, T; Kusajima, H; Hosaka, M; Fukuda, H; Oomori, Y; Shinoda, H

1996-01-01

The uptake and intracellular activity of AM-1155 in murine J774.1 macrophages and human polymorphonuclear leukocytes were investigated. AM-1155 penetrated phagocytic cells rapidly and reversibly, although the penetration process was not affected by metabolic inhibitors such as sodium fluoride, cyanide m-chlorophenylhydrazone, or ouabain or by nucleoside transport system inhibitors such as adenosine. The intracellular concentration-to-extracellular concentration ratio of AM-1155 in both cell types of phagocytes ranged from 5 to 7. These ratios were almost equal to those for sparfloxacin. The intracellular activity of AM-1155 in J774.1 macrophages, examined with Staphylococcus aureus 209P as a test bacterium, was dependent on the extracellular concentration. AM-1155 at a concentration of 1 microgram/ml reduced the number of viable cells of S. aureus ingested by more than 90%. The intracellular activity of AM-1155 was more potent than those of sparfloxacin, ofloxacin, ciprofloxacin, flomoxef, and erythromycin. These results suggest that the potent intracellular activity of AM-1155 might mainly be due to the high intracellular concentration and its potent in vitro activity. PMID:9124835

15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2010 CFR

2015-01-01

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15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2010 CFR

2010-01-01

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15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2013 CFR

2013-01-01

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15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2011 CFR

2011-01-01

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15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2012 CFR

2012-01-01

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15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2014 CFR

2014-01-01

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Modeling triple-negative breast cancer heterogeneity: effects of stromal macrophages, fibroblasts and tumor vasculature.

PubMed

Norton, Kerri-Ann; Jin, Kideok; Popel, Aleksander S

2018-05-08

A hallmark of breast tumors is its spatial heterogeneity that includes its distribution of cancer stem cells and progenitor cells, but also heterogeneity in the tumor microenvironment. In this study we focus on the contributions of stromal cells, specifically macrophages, fibroblasts, and endothelial cells on tumor progression. We develop a computational model of triple-negative breast cancer based on our previous work and expand it to include macrophage infiltration, fibroblasts, and angiogenesis. In vitro studies have shown that the secretomes of tumor-educated macrophages and fibroblasts increase both the migration and proliferation rates of triple-negative breast cancer cells. In vivo studies also demonstrated that blocking signaling of selected secreted factors inhibits tumor growth and metastasis in mouse xenograft models. We investigate the influences of increased migration and proliferation rates on tumor growth, the effect of the presence on fibroblasts or macrophages on growth and morphology, and the contributions of macrophage infiltration on tumor growth. We find that while the presence of macrophages increases overall tumor growth, the increase in macrophage infiltration does not substantially increase tumor growth and can even stifle tumor growth at excessive rates. Copyright © 2018. Published by Elsevier Ltd.

In vitro uptake of apoptotic body mimicking phosphatidylserine-quantum dot micelles by monocytic cell line

NASA Astrophysics Data System (ADS)

Maiseyeu, Andrei; Bagalkot, Vaishali

2014-04-01

A new quantum dot (QD) PEGylated micelle laced with phosphatidylserine (PS) for specific scavenger receptor-mediated uptake by macrophages is reported. The size and surface chemistry of PS-QD micelles were characterized by standard methods and the effects of their physicochemical properties on specific targeting and uptake were comprehensively studied in a monocytic cell line (J774A.1).

15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2010 CFR

2017-01-01

... 15 Commerce and Foreign Trade 2 2017-01-01 2017-01-01 false The Commerce Control List No. Supplement No. 1 to Part 774 Commerce and Foreign Trade Regulations Relating to Commerce and Foreign Trade (Continued) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS THE COMMERCE CONTROL LIST Pt. 774, Supp. No. 1...

15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2010 CFR

2016-01-01

... 15 Commerce and Foreign Trade 2 2016-01-01 2016-01-01 false The Commerce Control List No. Supplement No. 1 to Part 774 Commerce and Foreign Trade Regulations Relating to Commerce and Foreign Trade (Continued) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS THE COMMERCE CONTROL LIST Pt. 774, Supp. No. 1...

15 CFR Supplement No. 1 to Part 774 - The Commerce Control List

Code of Federal Regulations, 2010 CFR

2018-01-01

... 15 Commerce and Foreign Trade 2 2018-01-01 2018-01-01 false The Commerce Control List No. Supplement No. 1 to Part 774 Commerce and Foreign Trade Regulations Relating to Commerce and Foreign Trade (Continued) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS THE COMMERCE CONTROL LIST Pt. 774, Supp. No. 1...

Quinolactacins A, B and C: novel quinolone compounds from Penicillium sp. EPF-6. I. Taxonomy, production, isolation and biological properties.

PubMed

Kakinuma, N; Iwai, H; Takahashi, S; Hamano, K; Yanagisawa, T; Nagai, K; Tanaka, K; Suzuki, K; Kirikae, F; Kirikae, T; Nakagawa, A

2000-11-01

Quinolactacins A (1), B (2) and C (3), novel quinolone antibiotics have been found from the cultured broth of a fungal strain isolated from the larvae of the mulberry pyralid Margaronia pyloalis Welker). The fungal strain, EPF-6 was identified as Penicillium sp. from its morphological characteristics. Quinolactacins were obtained from the culture medium by solvent extraction and chromatographic purification. Compound 1 showed inhibitory activity against tumor necrosis factor (TNF) production induced by murine peritoneal macrophages and macrophage-like J774.1 cells stimulated with lipopolysaccharide (LPS).

Macrophage heterogeneity in tissues: phenotypic diversity and functions

PubMed Central

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

2014-01-01

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

Opsonization of Toxoplasma gondii tachyzoites with nonspecific immunoglobulins promotes their phagocytosis by macrophages and inhibits their proliferation in nonphagocytic cells in tissue culture.

PubMed

Vercammen, M; Scorza, T; El Bouhdidi, A; Van Beeck, K; Carlier, Y; Dubremetz, J F; Verschueren, H

1999-11-01

We have recently shown that Toxoplasma gondii tachyzoites grown in in vitro culture can bind unspecific immunoglobulin (Ig) through their Fc moiety. We show now that Fc receptors are also present on T. gondii within the host animal, and that intraperitoneal parasites in immunocompetent mice are saturated with unspecific Ig. We have also investigated the effect of the parasite's Fc receptor on the interaction of tachyzoites with mammalian cells, using the Vero cell line as a model for nonphagocytic host cells and murine peritoneal macrophages in primary culture as a model for phagocytic cells. Coating of tachyzoites with parasite-unrelated Ig did not enhance their invasive capacity in either target cell type, but slightly decreased the parasite proliferation. Moreover, phagocytosis by macrophages was increased by approximately 50% when parasites were coated with unspecific Ig. These results indicate that the Fc receptor on T. gondii affects the balance between invasion and phagocytosis in a way that is detrimental to the parasites.

Nuclear Proliferation: A Historical Overview

DTIC Science & Technology

2008-03-01

Talbert, “Nuclear Proliferation Technology Trends Analysis ,” Pacific Northwest National Laboratory, PNNL -14480 (September 2005), p. 92. 1973: Closed...L. Coles, and R. J. Talbert, “Nuclear Proliferation Technology Trends Analysis ,” Pacific Northwest National Laboratory, PNNL -14480 (September 2005...D. Zentner, G. L. Coles, and R. J. Talbert, “Nuclear Proliferation Technology Trends Analysis ,” Pacific Northwest National Laboratory, PNNL -14480

Apocynin suppresses the progression of atherosclerosis in apoE-deficient mice by inactivation of macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kinoshita, Hiroyuki; Matsumura, Takeshi, E-mail: takeshim@gpo.kumamoto-u.ac.jp; Ishii, Norio

Highlights: ► We examined the anti-athrogenic effect of apocynin in atherosclerotic model mice. ► Apocynin prevented atherosclerotic lesion formation. ► Apocynin suppressed ROS production in aorta and in macrophages. ► Apocynin suppressed cytokine expression and cell proliferation in macrophages. ► Apocynin may be beneficial compound for the prevention of atherosclerosis. -- Abstract: Production of reactive oxygen species (ROS) and other proinflammatory substances by macrophages plays an important role in atherogenesis. Apocynin (4-hydroxy-3-methoxy-acetophenone), which is well known as a NADPH oxidase inhibitor, has anti-inflammatory effects including suppression of the generation of ROS. However, the suppressive effects of apocynin on the progressionmore » of atherosclerosis are not clearly understood. Thus, we investigated anti-atherosclerotic effects of apocynin using apolipoprotein E-deficient (apoE{sup –/–}) mice in vivo and in mouse peritoneal macrophages in vitro. In atherosclerosis-prone apoE{sup –/–} mice, apocynin suppressed the progression of atherosclerosis, decreased 4-hydroxynonenal-positive area in atherosclerotic lesions, and mRNA expression of monocyte chemoattractant protein-1 (MCP-1) and interleukin-6 (IL-6) in aorta. In mouse peritoneal macrophages, apocynin suppressed the Ox-LDL-induced ROS generation, mRNA expression of MCP-1, IL-6 and granulocyte/macrophage colony-stimulating factor, and cell proliferation. Moreover, immunohistochemical studies revealed that apocynin decreased the number of proliferating cell nuclear antigen-positive macrophages in atherosclerotic lesions of apoE{sup –/–} mice. These results suggested that apocynin suppressed the formation of atherosclerotic lesions, at least in part, by inactivation of macrophages. Therefore, apocynin may be a potential therapeutic material to prevent the progression of atherosclerosis.« less

Inflammation and wound healing: The role of the macrophage

PubMed Central

Koh, Timothy J.; DiPietro, Luisa Ann

2013-01-01

The macrophage is a prominent inflammatory cell in wounds, but its role in healing remains incompletely understood. Macrophages have been described to have many functions in wounds, including host defense, the promotion and resolution of inflammation, the removal of apoptotic cells, and the support of cell proliferation and tissue restoration following injury. Recent studies suggest that macrophages exist in several different phenotypic states within the healing wound, and that the influence of these cells on each stage of repair varies with the specific phenotypes. While the macrophage is beneficial to the repair of normally healing wounds, this pleotropic cell type may promote excessive inflammation and/or fibrosis in certain circumstances. Emerging evidence suggests that macrophage dysfunction is a component of the pathogenesis of non-healing and poorly healing wounds. Due to advances in the understanding of this multi-functional cell, the macrophage continues to be an attractive therapeutic target both to reduce fibrosis and scarring, and to improve healing of chronic wounds. PMID:21740602

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Renard, C.; Vanderhaeghe, H.J.; Claes, P.J.

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

Pertussis toxin permeabilization enhances the traversal of Escherichia coli K1, macrophages, and monocytes in a cerebral endothelial barrier model in vitro.

PubMed

Seidel, Gabriela; Böcker, Kathrin; Schulte, Jessica; Wewer, Corinna; Greune, Lilo; Humberg, Verena; Schmidt, M Alexander

2011-03-01

The occasionally severe neurological complications following the human respiratory tract infection 'whooping cough' have been attributed to pertussis toxin (PT) expressed by the causative agent Bordetella pertussis. Disruption of the endothelial blood-brain barrier (BBB) by PT might facilitate the translocation of immune cells and of hematogenous microbial pathogens. To test this hypothesis, we investigated whether PT enhances the traversal of bacteria employing human brain microvascular endothelial cells (HBMEC) as an in vitro endothelial barrier model. PT incubation significantly increased the translocation of Escherichia coli K1 across the HBMEC barrier. Only intercellular E. coli K1 bacteria could be identified by electron microscopy suggesting paracellular translocation. In addition, the migration of differentiated HL60-derived macrophages and of human monocytic U937 cells through PT-treated HBMEC barriers was also enhanced. In comparison to E. coli C600, E. coli K1 showed prolonged survival in translocated HL60-derived and J774 macrophages as well as in U937 monocytes which suggested a contribution of the 'Trojan horse' mechanism. In summary, our findings demonstrate that the PT-induced permeabilization of endothelial barriers enhances the paracellular transmigration of microbes and immune cells. In vivo, this activity might lower the threshold of bacteremia facilitating secondary cerebral infections and the subsequent development of brain pathologies. Copyright © 2010 Elsevier GmbH. All rights reserved.

Peripheral blood "endothelial progenitor cells" are derived from monocyte/macrophages and secrete angiogenic growth factors.

PubMed

Rehman, Jalees; Li, Jingling; Orschell, Christie M; March, Keith L

2003-03-04

Endothelial progenitor cells (EPCs) have been isolated from peripheral blood and can enhance angiogenesis after infusion into host animals. It is not known whether the proangiogenic effects are a result of such events as endothelial differentiation and subsequent proliferation of EPCs or secondary to secretion of angiogenic growth factors. Human EPCs were isolated as previously described, and their phenotypes were confirmed by uptake of acetylated LDL and binding of ulex-lectin. EPC proliferation and surface marker expression were analyzed by flow cytometry, and conditioned medium was assayed for growth factors. The majority of EPCs expressed monocyte/macrophage markers such as CD14 (95.7+/-0.3%), Mac-1 (57.6+/-13.5%), and CD11c (90.8+/-4.9%). A much lower percentage of cells expressed the specific endothelial marker VE-cadherin (5.2+/-0.7%) or stem/progenitor-cell markers AC133 (0.16+/-0.05%) and c-kit (1.3+/-0.7%). Compared with circulating monocytes, cultured EPCs showed upregulation of monocyte activation and macrophage differentiation markers. EPCs did not demonstrate any significant proliferation but did secrete the angiogenic growth factors vascular endothelial growth factor, hepatocyte growth factor, granulocyte colony-stimulating factor, and granulocyte-macrophage colony-stimulating factor. Our findings suggest that acetylated LDL(+)ulex-lectin(+) cells, commonly referred to as EPCs, do not proliferate but release potent proangiogenic growth factors. The majority of acetylated LDL(+)ulex-lectin(+) cells are derived from monocyte/macrophages. The findings of low proliferation and endothelial differentiation suggest that their angiogenic effects are most likely mediated by growth factor secretion. These findings may allow for development of novel angiogenic therapies relying on secreted growth factors or on recruitment of endogenous monocytes/macrophages to sites of ischemia.

1, 25(OH){sub 2}D{sub 3}-induced interaction of vitamin D receptor with p50 subunit of NF-κB suppresses the interaction between KLF5 and p50, contributing to inhibition of LPS-induced macrophage proliferation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Dong; School of Public Health, North China University of Science and Technology, Tangshan, Hebei, 063000; Zhang, Ruo-nan

KLF5 and nuclear factor κB (NF-κB) regulate cell proliferation and inflammation. Vitamin D signaling through vitamin D receptor (VDR) exerts anti-proliferative and anti-inflammatory actions. However, an actual relationship between KLF5, NF-κB and VDR in the inflammation and proliferation of macrophages is still unclear. Here, we showed that LPS and proinflammatory cytokines stimulate KLF5 gene expression in macrophages, and that 1, 25(OH){sub 2}D{sub 3} suppresses LPS-induced KLF5 expression and cell proliferation via upregulation of VDR expression. Mechanistic studies suggested that KLF5 interacts with p50 subunit of NF-κB to cooperatively induce the expressions of positive cell cycle regulators cyclin B1 and Cdk1/Cdc2more » in LPS-treated macrophages. Further studies revealed that 1, 25(OH){sub 2}D{sub 3}-induced interaction of VDR with p50 decreases LPS-induced interaction of KLF5 with p50. Collectively, we identify a novel regulatory pathway in which 1, 25(OH){sub 2}D{sub 3} induces VDR expression and promotes VDR interaction with p50 subunit of NF-κB, which in turn attenuates the association of KLF5 with p50 subunit of NF-κB and thus exerts anti-inflammatory and anti-proliferative effects on macrophages. - Highlights: • 1, 25(OH){sub 2}D{sub 3} suppresses LPS-induced KLF5 expression via upregulation of VDR expression. • KLF5 interacts with NF-κB-p50 to cooperatively induce the expressions of positive cell cycle regulators cyclin B1 and Cdk1/Cdc2 in LPS-treated macrophages. • 1,25(OH){sub 2}D{sub 3} induces interaction of VDR with p50.« less

Autocrine IL-10 activation of the STAT3 pathway is required for pathological macrophage differentiation in polycystic kidney disease

PubMed Central

Peda, Jacqueline D.; Salah, Sally M.; Wallace, Darren P.; Fields, Patrick E.; Grantham, Connor J.; Fields, Timothy A.

2016-01-01

ABSTRACT Polycystic kidney disease (PKD) is characterized by slow expansion of fluid-filled cysts derived from tubules within the kidney. Cystic expansion results in injury to surrounding parenchyma and leads to inflammation, scarring and ultimately loss of renal function. Macrophages are a key element in this process, promoting cyst epithelial cell proliferation, cyst expansion and disease progression. Previously, we have shown that the microenvironment established by cystic epithelial cells can ‘program’ macrophages, inducing M2-like macrophage polarization that is characterized by expression of markers that include Arg1 and Il10. Here, we functionally characterize these macrophages, demonstrating that their differentiation enhances their ability to promote cyst cell proliferation. This observation indicates a model of reciprocal pathological interactions between cysts and the innate immune system: cyst epithelial cells promote macrophage polarization to a phenotype that, in turn, is especially efficient in promoting cyst cell proliferation and cyst growth. To better understand the genesis of this macrophage phenotype, we examined the role of IL-10, a regulatory cytokine shown to be important for macrophage-stimulated tissue repair in other settings. Herein, we show that the acquisition of the pathological macrophage phenotype requires IL-10 secretion by the macrophages. Further, we demonstrate a requirement for IL-10-dependent autocrine activation of the STAT3 pathway. These data suggest that the IL-10 pathway in macrophages plays an essential role in the pathological relationship between cysts and the innate immune system in PKD, and thus could be a potential therapeutic target. PMID:27491076

Transformation of Mouse Macrophages by Simian Virus 40

PubMed Central

Stone, Lawrence B.; Takemoto, Kenneth K.

1970-01-01

Studies were undertaken to prove that simian virus 40 (SV40) can transform the mouse macrophage, a cell type naturally restricted from deoxyribonucleic acid (DNA) replication. Balb/C macrophages infected with SV40 demonstrated T-antigen production and induced DNA synthesis simultaneously. In the absence of apparent division, these cells remained T antigen-positive for at least 45 days. SV40 could be rescued from nondividing, unaltered macrophages during the T antigen-producing period. Proliferating transformants appeared at an average of 66 days post-SV40 infection. Established cell lines were T antigen-positive and were negative for infectious virus, but yielded SV40 after fusion with African green monkey kidney cells. Their identity as transformed macrophages was substantiated by evaluation of cellular morphology, phagocytosis, acid phosphatase, β1c synthesis, and aminoacridine incorporation. Images PMID:4320698

Macrophage heterogeneity and cholesterol homeostasis: classically-activated macrophages are associated with reduced cholesterol accumulation following treatment with oxidized LDL.

PubMed

Chu, Eugene M; Tai, Daven C; Beer, Jennifer L; Hill, John S

2013-02-01

Macrophages are centrally involved during atherosclerosis development and are the predominant cell type that accumulates cholesterol in the plaque. Macrophages however, are heterogeneous in nature reflecting a variety of microenvironments and different phenotypes may be more prone to contribute towards atherosclerosis progression. Using primary human monocyte-derived macrophages, we sought to evaluate one aspect of atherogenic potential of different macrophage phenotypes by determining their propensity to associate with and accumulate oxidized low density lipoprotein (oxLDL). Classically-activated macrophages treated simultaneously with interferon γ (IFNγ) and tumor necrosis factor α (TNFα) associated with less oxLDL and accumulated less cholesterol compared to untreated controls. The combined treatment of IFNγ and TNFα reduced the mRNA expression of CD36 and the expression of both cell surface CD36 and macrophage scavenger receptor 1 (MSR1) protein. Under oxLDL loaded conditions, IFNγ and TNFα did not reduce macrophage protein expression of the transcription factor peroxisome proliferator-actived receptor γ (PPARγ) which is known to positively regulate CD36 expression. However, macrophages treated with IFNγ attenuated the ability of the PPARγ-specific agonist rosiglitazone from upregulating cell surface CD36 protein expression. Our results demonstrate that the observed reduction of cholesterol accumulation in macrophages treated with IFNγ and TNFα following oxLDL treatment was due at least in part to reduced cell surface CD36 and MSR1 protein expression. Copyright © 2012 Elsevier B.V. All rights reserved.

15 CFR Supplement No. 2 to Part 774 - General Technology and Software Notes

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false General Technology and Software Notes... REGULATIONS THE COMMERCE CONTROL LIST Pt. 774, Supp. 2 Supplement No. 2 to Part 774—General Technology and Software Notes 1. General Technology Note. The export of “technology” that is “required” for the...

Inhibition of Macrophage CD36 Expression and Cellular Oxidized Low Density Lipoprotein (oxLDL) Accumulation by Tamoxifen: A PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR (PPAR)γ-DEPENDENT MECHANISM.

PubMed

Yu, Miao; Jiang, Meixiu; Chen, Yuanli; Zhang, Shuang; Zhang, Wenwen; Yang, Xiaoxiao; Li, Xiaoju; Li, Yan; Duan, Shengzhong; Han, Jihong; Duan, Yajun

2016-08-12

Macrophage CD36 binds and internalizes oxidized low density lipoprotein (oxLDL) to facilitate foam cell formation. CD36 expression is activated by peroxisome proliferator-activated receptor γ (PPARγ). Tamoxifen, an anti-breast cancer medicine, has demonstrated pleiotropic functions including cardioprotection with unfully elucidated mechanisms. In this study, we determined that treatment of ApoE-deficient mice with tamoxifen reduced atherosclerosis, which was associated with decreased CD36 and PPARγ expression in lesion areas. At the cellular level, we observed that tamoxifen inhibited CD36 protein expression in human THP-1 monocytes, THP-1/PMA macrophages, and human blood monocyte-derived macrophages. Associated with decreased CD36 protein expression, tamoxifen reduced cellular oxLDL accumulation in a CD36-dependent manner. At the transcriptional level, tamoxifen decreased CD36 mRNA expression, promoter activity, and the binding of the PPARγ response element in CD36 promoter to PPARγ protein. Tamoxifen blocked ligand-induced PPARγ nuclear translocation and CD36 expression, but it increased PPARγ phosphorylation, which was due to that tamoxifen-activated ERK1/2. Furthermore, deficiency of PPARγ expression in macrophages abolished the inhibitory effect of tamoxifen on CD36 expression or cellular oxLDL accumulation both in vitro and in vivo Taken together, our study demonstrates that tamoxifen inhibits CD36 expression and cellular oxLDL accumulation by inactivating the PPARγ signaling pathway, and the inhibition of macrophage CD36 expression can be attributed to the anti-atherogenic properties of tamoxifen. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Redox Control of Inflammation in Macrophages

PubMed Central

Dehne, Nathalie; Grossmann, Nina; Jung, Michaela; Namgaladze, Dmitry; Schmid, Tobias; von Knethen, Andreas; Weigert, Andreas

2013-01-01

Abstract Macrophages are present throughout the human body, constitute important immune effector cells, and have variable roles in a great number of pathological, but also physiological, settings. It is apparent that macrophages need to adjust their activation profile toward a steadily changing environment that requires altering their phenotype, a process known as macrophage polarization. Formation of reactive oxygen species (ROS), derived from NADPH-oxidases, mitochondria, or NO-producing enzymes, are not necessarily toxic, but rather compose a network signaling system, known as redox regulation. Formation of redox signals in classically versus alternatively activated macrophages, their action and interaction at the level of key targets, and the resulting physiology still are insufficiently understood. We review the identity, source, and biological activities of ROS produced during macrophage activation, and discuss how they shape the key transcriptional responses evoked by hypoxia-inducible transcription factors, nuclear-erythroid 2-p45-related factor 2 (Nrf2), and peroxisome proliferator-activated receptor-γ. We summarize the mechanisms how redox signals add to the process of macrophage polarization and reprogramming, how this is controlled by the interaction of macrophages with their environment, and addresses the outcome of the polarization process in health and disease. Future studies need to tackle the option whether we can use the knowledge of redox biology in macrophages to shape their mediator profile in pathophysiology, to accelerate healing in injured tissue, to fight the invading pathogens, or to eliminate settings of altered self in tumors. Antioxid. Redox Signal. 19, 595–637. PMID:23311665

circRNA Mediates Silica-Induced Macrophage Activation Via HECTD1/ZC3H12A-Dependent Ubiquitination

PubMed Central

Zhou, Zewei; Jiang, Rong; Yang, Xiyue; Guo, Huifang; Fang, Shencun; Zhang, Yingming; Cheng, Yusi; Wang, Jing; Yao, Honghong; Chao, Jie

2018-01-01

Rationale: Phagocytosis of silicon dioxide (SiO2) into lung cells causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Circular RNAs (circRNAs) are a subclass of non-coding RNAs detected within mammalian cells; however, researchers have not determined whether circRNAs are involved in the pathophysiological process of silicosis. The upstream molecular mechanisms and functional effects on cell apoptosis, proliferation and migration were investigated to elucidate the role of circRNAs in SiO2-induced inflammation in pulmonary macrophages. Methods: Primary cultures of alveolar macrophages from healthy donors and patients as well as the RAW264.7 macrophage cell line were used to explore the functions of circHECTD1 (HECT domain E3 ubiquitin protein ligase 1) in macrophage activation. Results: The results of the experiments indicated that 1) SiO2 concomitantly decreased circHECTD1 levels and increased HECTD1 protein expression; 2) circHECTD1 and HECTD1 were involved in SiO2-induced macrophage activation via ubiquitination; and 3) SiO2-activated macrophages promoted fibroblast proliferation and migration via the circHECTD1/HECTD1 pathway. Tissue samples from silicosis patients confirmed the upregulation of HECTD1. Conclusions: Our study elucidated a link between SiO2-induced macrophage activation and the circHECTD1/HECTD1 pathway, thereby providing new insight into the potential use of HECTD1 in the development of novel therapeutic strategies for treating silicosis. PMID:29290828

circRNA Mediates Silica-Induced Macrophage Activation Via HECTD1/ZC3H12A-Dependent Ubiquitination.

PubMed

Zhou, Zewei; Jiang, Rong; Yang, Xiyue; Guo, Huifang; Fang, Shencun; Zhang, Yingming; Cheng, Yusi; Wang, Jing; Yao, Honghong; Chao, Jie

2018-01-01

Rationale: Phagocytosis of silicon dioxide (SiO 2 ) into lung cells causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Circular RNAs (circRNAs) are a subclass of non-coding RNAs detected within mammalian cells; however, researchers have not determined whether circRNAs are involved in the pathophysiological process of silicosis. The upstream molecular mechanisms and functional effects on cell apoptosis, proliferation and migration were investigated to elucidate the role of circRNAs in SiO 2 -induced inflammation in pulmonary macrophages. Methods: Primary cultures of alveolar macrophages from healthy donors and patients as well as the RAW264.7 macrophage cell line were used to explore the functions of circHECTD1 (HECT domain E3 ubiquitin protein ligase 1) in macrophage activation. Results: The results of the experiments indicated that 1) SiO 2 concomitantly decreased circHECTD1 levels and increased HECTD1 protein expression; 2) circHECTD1 and HECTD1 were involved in SiO 2 -induced macrophage activation via ubiquitination; and 3) SiO 2 -activated macrophages promoted fibroblast proliferation and migration via the circHECTD1/HECTD1 pathway. Tissue samples from silicosis patients confirmed the upregulation of HECTD1. Conclusions: Our study elucidated a link between SiO 2 -induced macrophage activation and the circHECTD1/HECTD1 pathway, thereby providing new insight into the potential use of HECTD1 in the development of novel therapeutic strategies for treating silicosis.

Monocyte to macrophage differentiation-associated (MMD) positively regulates ERK and Akt activation and TNF-α and NO production in macrophages.

PubMed

Liu, Qiang; Zheng, Jin; Yin, Dan-Dan; Xiang, Jie; He, Fei; Wang, Yao-Chun; Liang, Liang; Qin, Hong-Yan; Liu, Li; Liang, Ying-Min; Han, Hua

2012-05-01

Macrophage activation is modulated by both environmental cues and endogenous programs. In the present study, we investigated the role of a PAQR family protein, monocyte to macrophage differentiation-associated (MMD), in macrophage activation and unveiled its underlying molecular mechanism. Our results showed that while MMD expression could be detected in all tissues examined, its expression level is significantly up-regulated upon monocyte differentiation. Within cells, EGFP-MMD fusion protein could be co-localized to endoplasmic reticulum, mitochondria, Golgi apparatus, but not lysosomes and cytoplasm. MMD expression is up-regulated in macrophages after LPS stimulation, and this might be modulated by RBP-J, the critical transcription factor of Notch signaling. Overexpression of MMD in macrophages increased the production of TNF-α and NO upon LPS stimulation. We found that MMD overexpression enhanced ERK1/2 and Akt phosphorylation in macrophages after LPS stimulation. Blocking Erk or Akt by pharmacological agent reduced TNF-α or NO production in MMD-overexpressing macrophages, respectively. These results suggested that MMD modulates TNF-α and NO production in macrophages, and this process might involves Erk or Akt.

Cyclosporin A and tacrolimus (FK506) suppress expression of inducible nitric oxide synthase in vitro by different mechanisms

PubMed Central

Dusting, Gregory J; Akita, Kazuhiro; Hickey, Haruyo; Smith, Melanie; Gurevich, Vladimir

1999-01-01

The effects of the immunosuppressant drugs cyclosporin A and tacrolimus (FK506) on nitric oxide synthesis were examined in a murine macrophage cell line (J774) and rat vascular smooth muscle cells (VSMC) in culture for 24 and 48 h, respectively.Cyclosporin A (0.01–10 μM) inhibited by up to 90% accumulation of nitrite induced by lipopolysaccharide (LPS) in both cell lines, but FK506 (0.01–10 μM) had a weaker effect on nitrite accumulation in these cells. Cyclosporin A and FK506 (at 1 μM) also significantly inhibited nitrite production induced by recombinant murine interferon-γ (rIFNγ) and recombinant murine interleukin-1β (rIL-1β) in J774 and VSMC, respectively.In J774 cells, cyclosporin A (but not FK506) at 1 μM was inhibitory when co-incubated with the inducing agents but not when the cells were treated with the immunosuppressant before or after the inducer. In VSMC, nitrite production was inhibited by co-incubation of cyclosporin A or FK506 with the inducer, or when the immunosuppressants were pre-incubated with cells. In contrast, N-monomethyl L-arginine (NMMA) abolished nitrite production when incubated with either cell type during or after addition of inducing agent, but not if cells were preincubated with NMMA.RNA extracted from treated J774 and VSMC was subjected to reverse transcription–polymerase chain reaction (RT–PCR). Cyclosporin A, but not FK506, suppressed expression of mRNA for NOS2 in a concentration-dependent manner when co-incubated with LPS.The fact that the potency difference between cyclosporin A and FK506 for NO suppression is the opposite to that for inhibition of interleukin-2 generation suggests that the immunosuppressants act in J774 macrophages and VSMC through intracellular mechanisms that differ from those elucidated in T-cells. Cyclosporin A suppresses NOS2 gene transcription, but FK506 acts post-transcriptionally to suppress NO generation in VSMC.Taken together the present data suggest that therapeutic

Evaluation of the efficacy of photodynamic antimicrobial therapy using a phenothiazine compound and LED (red-orange) on the interface: macrophage vs S. aureus

NASA Astrophysics Data System (ADS)

Sampaio, Fernando José P.; de Oliveira, Susana C. P. S.; Monteiro, Juliana S. C.; Pires-Santos, Gustavo M.; Gesteira, Maria F. M.; Pinheiro, Antônio L. B.

2015-03-01

Antimicrobial Photodynamic therapy is a technique in which microorganisms are exposed to a photosensitizing drug and then irradiated with low-intensity visible light of the appropriate wavelength. The resulting photochemical reaction generates cytotoxic reactive oxygen species, such as singlet oxygen and free radicals, which are able to exert bactericidal effect. Much is already known about the photodynamic inactivation of microorganisms: both antibiotic-sensitive and - resistant strains can be successfully photo inactivated, and there is the additional advantage that repeated photosensitization of bacterial cells does not induce a selection of resistant strains. Recently, a series of studies have shown that it is possible to kill bacteria with a light source after the microorganisms have been sensitized with low concentration of dye, such as phenothiazines. The aim of this study was to evaluate the phagocytic function of macrophages J774 against S. aureus in the presence and absence of AmPDT with phenothiazine compound (12.5 μg/mL) and red-orange LED. Experimental groups: Control Group (L-F-), Phenothiazine group (L-F+) LED group (L+F-), Photodynamic therapy group (L+F+). The tests presented in this study were carried out in triplicate. This study demonstrated that AmPDT is able to increase about twice the phagocytic ability of macrophages; however, the bactericidal capacity of these cells did not show a substantial improvement, probably because the oxidative burst was less intense.

Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.

PubMed

Kim, Jong Hun; Lee, Eunjung; Friedline, Randall H; Suk, Sujin; Jung, Dae Young; Dagdeviren, Sezin; Hu, Xiaodi; Inashima, Kunikazu; Noh, Hye Lim; Kwon, Jung Yeon; Nambu, Aya; Huh, Jun R; Han, Myoung Sook; Davis, Roger J; Lee, Amy S; Lee, Ki Won; Kim, Jason K

2018-04-01

Obesity-mediated inflammation is a major cause of insulin resistance, and macrophages play an important role in this process. The 78-kDa glucose-regulated protein (GRP78) is a major endoplasmic reticulum chaperone that modulates unfolded protein response (UPR), and mice with GRP78 heterozygosity were resistant to diet-induced obesity. Here, we show that mice with macrophage-selective ablation of GRP78 (Lyz- GRP78 -/- ) are protected from skeletal muscle insulin resistance without changes in obesity compared with wild-type mice after 9 wk of high-fat diet. GRP78-deficient macrophages demonstrated adapted UPR with up-regulation of activating transcription factor (ATF)-4 and M2-polarization markers. Diet-induced adipose tissue inflammation was reduced, and bone marrow-derived macrophages from Lyz- GRP78 -/- mice demonstrated a selective increase in IL-6 expression. Serum IL-13 levels were elevated by >4-fold in Lyz- GRP78 -/- mice, and IL-6 stimulated the myocyte expression of IL-13 and IL-13 receptor. Lastly, recombinant IL-13 acutely increased glucose metabolism in Lyz- GRP78 -/- mice. Taken together, our data indicate that GRP78 deficiency activates UPR by increasing ATF-4, and promotes M2-polarization of macrophages with a selective increase in IL-6 secretion. Macrophage-derived IL-6 stimulates the myocyte expression of IL-13 and regulates muscle glucose metabolism in a paracrine manner. Thus, our findings identify a novel crosstalk between macrophages and skeletal muscle in the modulation of obesity-mediated insulin resistance.-Kim, J. H., Lee, E., Friedline, R. H., Suk, S., Jung, D. Y., Dagdeviren, S., Hu, X., Inashima, K., Noh, H. L., Kwon, J. Y., Nambu, A., Huh, J. R., Han, M. S., Davis, R. J., Lee, A. S., Lee, K. W., Kim, J. K. Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.

Blood vessel control of macrophage maturation promotes arteriogenesis in ischemia.

PubMed

Krishnasamy, Kashyap; Limbourg, Anne; Kapanadze, Tamar; Gamrekelashvili, Jaba; Beger, Christian; Häger, Christine; Lozanovski, Vladimir J; Falk, Christine S; Napp, L Christian; Bauersachs, Johann; Mack, Matthias; Haller, Hermann; Weber, Christian; Adams, Ralf H; Limbourg, Florian P

2017-10-16

Ischemia causes an inflammatory response that is intended to restore perfusion and homeostasis yet often aggravates damage. Here we show, using conditional genetic deletion strategies together with adoptive cell transfer experiments in a mouse model of hind limb ischemia, that blood vessels control macrophage differentiation and maturation from recruited monocytes via Notch signaling, which in turn promotes arteriogenesis and tissue repair. Macrophage maturation is controlled by Notch ligand Dll1 expressed in vascular endothelial cells of arteries and requires macrophage canonical Notch signaling via Rbpj, which simultaneously suppresses an inflammatory macrophage fate. Conversely, conditional mutant mice lacking Dll1 or Rbpj show proliferation and transient accumulation of inflammatory macrophages, which antagonizes arteriogenesis and tissue repair. Furthermore, the effects of Notch are sufficient to generate mature macrophages from monocytes ex vivo that display a stable anti-inflammatory phenotype when challenged with pro-inflammatory stimuli. Thus, angiocrine Notch signaling fosters macrophage maturation during ischemia.Molecular mechanisms of macrophage-mediated regulation of artery growth in response to ischemia are poorly understood. Here the authors show that vascular endothelium controls macrophage maturation and differentiation via Notch signaling, which in turn promotes arteriogenesis and ischemic tissue recovery.

Macrophage heterogeneity in liver injury and fibrosis.

PubMed

Tacke, Frank; Zimmermann, Henning W

2014-05-01

Hepatic macrophages are central in the pathogenesis of chronic liver injury and have been proposed as potential targets in combatting fibrosis. Recent experimental studies in animal models revealed that hepatic macrophages are a remarkably heterogeneous population of immune cells that fulfill diverse functions in homeostasis, disease progression, and regression from injury. These range from clearance of pathogens or cellular debris and maintenance of immunological tolerance in steady state conditions; central roles in initiating and perpetuating inflammation in response to injury; promoting liver fibrosis via activating hepatic stellate cells in chronic liver damage; and, finally, resolution of inflammation and fibrosis by degradation of extracellular matrix and release of anti-inflammatory cytokines. Cellular heterogeneity in the liver is partly explained by the origin of macrophages. Hepatic macrophages can either arise from circulating monocytes, which are recruited to the injured liver via chemokine signals, or from self-renewing embryo-derived local macrophages, termed Kupffer cells. Kupffer cells appear essential for sensing tissue injury and initiating inflammatory responses, while infiltrating Ly-6C(+) monocyte-derived macrophages are linked to chronic inflammation and fibrogenesis. In addition, proliferation of local or recruited macrophages may possibly further contribute to their accumulation in injured liver. During fibrosis regression, monocyte-derived cells differentiate into Ly-6C (Ly6C, Gr1) low expressing 'restorative' macrophages and promote resolution from injury. Understanding the mechanisms that regulate hepatic macrophage heterogeneity, either by monocyte subset recruitment, by promoting restorative macrophage polarization or by impacting distinctive macrophage effector functions, may help to develop novel macrophage subset-targeted therapies for liver injury and fibrosis. Copyright © 2014 European Association for the Study of the Liver

Dynamics of Salmonella infection of macrophages at the single cell level.

PubMed

Gog, Julia R; Murcia, Alicia; Osterman, Natan; Restif, Olivier; McKinley, Trevelyan J; Sheppard, Mark; Achouri, Sarra; Wei, Bin; Mastroeni, Pietro; Wood, James L N; Maskell, Duncan J; Cicuta, Pietro; Bryant, Clare E

2012-10-07

Salmonella enterica causes a range of diseases. Salmonellae are intracellular parasites of macrophages, and the control of bacteria within these cells is critical to surviving an infection. The dynamics of the bacteria invading, surviving, proliferating in and killing macrophages are central to disease pathogenesis. Fundamentally important parameters, however, such as the cellular infection rate, have not previously been calculated. We used two independent approaches to calculate the macrophage infection rate: mathematical modelling of Salmonella infection experiments, and analysis of real-time video microscopy of infection events. Cells repeatedly encounter salmonellae, with the bacteria often remain associated with the macrophage for more than ten seconds. Once Salmonella encounters a macrophage, the probability of that bacterium infecting the cell is remarkably low: less than 5%. The macrophage population is heterogeneous in terms of its susceptibility to the first infection event. Once infected, a macrophage can undergo further infection events, but these reinfection events occur at a lower rate than that of the primary infection.

Macrophages are required to coordinate mouse digit tip regeneration.

PubMed

Simkin, Jennifer; Sammarco, Mimi C; Marrero, Luis; Dawson, Lindsay A; Yan, Mingquan; Tucker, Catherine; Cammack, Alex; Muneoka, Ken

2017-11-01

In mammals, macrophages are known to play a major role in tissue regeneration. They contribute to inflammation, histolysis, re-epithelialization, revascularization and cell proliferation. Macrophages have been shown to be essential for regeneration in salamanders and fish, but their role has not been elucidated in mammalian epimorphic regeneration. Here, using the regenerating mouse digit tip as a mammalian model, we demonstrate that macrophages are essential for the regeneration process. Using cell-depletion strategies, we show that regeneration is completely inhibited; bone histolysis does not occur, wound re-epithelialization is inhibited and the blastema does not form. Although rescue of epidermal wound closure in the absence of macrophages promotes blastema accumulation, it does not rescue cell differentiation, indicating that macrophages play a key role in the redifferentiation of the blastema. We provide additional evidence that although bone degradation is a component, it is not essential to the overall regenerative process. These findings show that macrophages play an essential role in coordinating the epimorphic regenerative response in mammals. © 2017. Published by The Company of Biologists Ltd.

Differences in Intracellular Fate of Two Spotted Fever Group Rickettsia in Macrophage-Like Cells.

PubMed

Curto, Pedro; Simões, Isaura; Riley, Sean P; Martinez, Juan J

2016-01-01

Spotted fever group (SFG) rickettsiae are recognized as important agents of human tick-borne diseases worldwide, such as Mediterranean spotted fever (Rickettsia conorii) and Rocky Mountain spotted fever (Rickettsia rickettsii). Recent studies in several animal models have provided evidence of non-endothelial parasitism by pathogenic SFG Rickettsia species, suggesting that the interaction of rickettsiae with cells other than the endothelium may play an important role in pathogenesis of rickettsial diseases. These studies raise the hypothesis that the role of macrophages in rickettsial pathogenesis may have been underappreciated. Herein, we evaluated the ability of two SFG rickettsial species, R. conorii (a recognized human pathogen) and Rickettsia montanensis (a non-virulent member of SFG) to proliferate in THP-1 macrophage-like cells, or within non-phagocytic cell lines. Our results demonstrate that R. conorii was able to survive and proliferate in both phagocytic and epithelial cells in vitro. In contrast, R. montanensis was able to grow in non-phagocytic cells, but was drastically compromised in the ability to proliferate within both undifferentiated and PMA-differentiated THP-1 cells. Interestingly, association assays revealed that R. montanensis was defective in binding to THP-1-derived macrophages; however, the invasion of the bacteria that are able to adhere did not appear to be affected. We have also demonstrated that R. montanensis which entered into THP-1-derived macrophages were rapidly destroyed and partially co-localized with LAMP-2 and cathepsin D, two markers of lysosomal compartments. In contrast, R. conorii was present as intact bacteria and free in the cytoplasm in both cell types. These findings suggest that a phenotypic difference between a non-pathogenic and a pathogenic SFG member lies in their respective ability to proliferate in macrophage-like cells, and may provide an explanation as to why certain SFG rickettsial species are not associated

Differences in Intracellular Fate of Two Spotted Fever Group Rickettsia in Macrophage-Like Cells

PubMed Central

Curto, Pedro; Simões, Isaura; Riley, Sean P.; Martinez, Juan J.

2016-01-01

Spotted fever group (SFG) rickettsiae are recognized as important agents of human tick-borne diseases worldwide, such as Mediterranean spotted fever (Rickettsia conorii) and Rocky Mountain spotted fever (Rickettsia rickettsii). Recent studies in several animal models have provided evidence of non-endothelial parasitism by pathogenic SFG Rickettsia species, suggesting that the interaction of rickettsiae with cells other than the endothelium may play an important role in pathogenesis of rickettsial diseases. These studies raise the hypothesis that the role of macrophages in rickettsial pathogenesis may have been underappreciated. Herein, we evaluated the ability of two SFG rickettsial species, R. conorii (a recognized human pathogen) and Rickettsia montanensis (a non-virulent member of SFG) to proliferate in THP-1 macrophage-like cells, or within non-phagocytic cell lines. Our results demonstrate that R. conorii was able to survive and proliferate in both phagocytic and epithelial cells in vitro. In contrast, R. montanensis was able to grow in non-phagocytic cells, but was drastically compromised in the ability to proliferate within both undifferentiated and PMA-differentiated THP-1 cells. Interestingly, association assays revealed that R. montanensis was defective in binding to THP-1-derived macrophages; however, the invasion of the bacteria that are able to adhere did not appear to be affected. We have also demonstrated that R. montanensis which entered into THP-1-derived macrophages were rapidly destroyed and partially co-localized with LAMP-2 and cathepsin D, two markers of lysosomal compartments. In contrast, R. conorii was present as intact bacteria and free in the cytoplasm in both cell types. These findings suggest that a phenotypic difference between a non-pathogenic and a pathogenic SFG member lies in their respective ability to proliferate in macrophage-like cells, and may provide an explanation as to why certain SFG rickettsial species are not associated

Effect of sodium butyrate on cell proliferation and cell cycle in porcine intestinal epithelial (IPEC-J2) cells.

PubMed

Qiu, Yueqin; Ma, Xianyong; Yang, Xuefen; Wang, Li; Jiang, Zongyong

2017-04-01

Conflicting results have been reported that butyrate in normal piglets leads either to an increase or to a decrease of jejunal villus length, implying a possible effect on the proliferation of enterocytes. No definitive study was found for the biological effects of butyrate in porcine jejunal epithelial cells. The present study used IPEC-J2 cells, a non-transformed jejunal epithelial line to evaluate the direct effects of sodium butyrate on cell proliferation, cell cycle regulation, and apoptosis. Low concentrations (0.5 and 1 mM) of butyrate had no effect on cell proliferation. However, at 5 and 10 mM, sodium butyrate significantly decreased cell viability, accompanied by reduced levels of p-mTOR and PCNA protein. Sodium butyrate, in a dose-dependent manner, induced cell cycle arrest in G0/G1 phase and reduced the numbers of cells in S phase. In addition, relative expression of p21, p27, and pro-apoptosis bak genes, and protein levels of p21Waf1/Cip1, p27Kip1, cyclinD3, CDK4, and Cleave-caspase3 were increased by higher concentrations of sodium butyrate (1, 5, 10 mM), and the levels of cyclinD1 and CDK6 were reduced by 5 and 10 mM butyrate. Butyrate increased the phosphorylated form of the signaling molecule p38 and phosphorylated JNK. In conclusion, the present in vitro study indicated that sodium butyrate inhibited the proliferation of IPEC-J2 cells by inducing cell cycle arrest in the G0/G1 phase of cell cycles and by increasing apoptosis at high concentrations.

The G1/S Specific Cyclin D2 Is a Regulator of HIV-1 Restriction in Non-proliferating Cells

PubMed Central

Badia, Roger; Pujantell, Maria; Riveira-Muñoz, Eva; Puig, Teresa; Torres-Torronteras, Javier; Martí, Ramón; Clotet, Bonaventura; Ampudia, Rosa M.; Ballana, Ester

2016-01-01

Macrophages are a heterogeneous cell population strongly influenced by differentiation stimuli that become susceptible to HIV-1 infection after inactivation of the restriction factor SAMHD1 by cyclin-dependent kinases (CDK). Here, we have used primary human monocyte-derived macrophages differentiated through different stimuli to evaluate macrophage heterogeneity on cell activation and proliferation and susceptibility to HIV-1 infection. Stimulation of monocytes with GM-CSF induces a non-proliferating macrophage population highly restrictive to HIV-1 infection, characterized by the upregulation of the G1/S-specific cyclin D2, known to control early steps of cell cycle progression. Knockdown of cyclin D2, enhances HIV-1 replication in GM-CSF macrophages through inactivation of SAMHD1 restriction factor by phosphorylation. Co-immunoprecipitation experiments show that cyclin D2 forms a complex with CDK4 and p21, a factor known to restrict HIV-1 replication by affecting the function of the downstream cascade that leads to SAMHD1 deactivation. Thus, we demonstrate that cyclin D2 acts as regulator of cell cycle proteins affecting SAMHD1-mediated HIV-1 restriction in non-proliferating macrophages. PMID:27541004

Influence of particle geometry and PEGylation on phagocytosis of particulate carriers.

PubMed

Mathaes, Roman; Winter, Gerhard; Besheer, Ahmed; Engert, Julia

2014-04-25

Particle geometry of micro- and nanoparticles has been identified as an important design parameter to influence the interaction with cells such as macrophages. A head to head comparison of elongated, non-spherical and spherical micro- and nanoparticles with and without PEGylation was carried out to benchmark two phagocytosis inhibiting techniques. J774.A1 macrophages were incubated with fluorescently labeled PLGA micro- and nanoparticles and analyzed by confocal laser scanning microscope (CLSM) and flow cytometry (FACS). Particle uptake into macrophages was significantly reduced upon PEGylation or elongated particle geometry. A combination of both, an elongated shape and PEGylation, had the strongest phagocytosis inhibiting effect for nanoparticles. Copyright © 2014 Elsevier B.V. All rights reserved.

Reactive oxygen species (ROS) induced cytokine production and cytotoxicity of PAMAM dendrimers in J774A.1 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Naha, Pratap C., E-mail: pratap.naha@dit.i; NanoLab, Focas Research Institute, Dublin Institute of Technology, Kevin Street, Dublin 8; Davoren, Maria

2010-07-15

The immunotoxicity of three generations of polyamidoamine (PAMAM) dendrimers (G-4, G-5 and G-6) was evaluated in mouse macrophage cells in vitro. Using the Alamar blue and MTT assays, a generation dependent cytotoxicity of the PAMAM dendrimers was found whereby G-6 > G-5 > G-4. The toxic response of the PAMAM dendrimers correlated well with the number of surface primary amino groups, with increasing number resulting in an increase in toxic response. An assessment of intracellular ROS generation by the PAMAM dendrimers was performed by measuring the increased fluorescence as a result of intracellular oxidation of Carboxy H{sub 2}DCFDA to DCFmore » both quantitatively using plate reader and qualitatively by confocal laser scanning microscopy. The inflammatory mediators macrophage inflammatory protein-2 (MIP-2), tumour necrosis factor-{alpha} (TNF-{alpha}) and interleukin-6, (IL-6) were measured by the enzyme linked immunosorbant assay (ELISA) following exposure of mouse macrophage cells to PAMAM dendrimers. A generation dependent ROS and cytokine production was found, which correlated well with the cytotoxicological response and therefore number of surface amino groups. A clear time sequence of increased ROS generation (maximum at {approx} 4 h), TNF-{alpha} and IL-6 secretion (maximum at {approx} 24 h), MIP-2 levels and cell death ({approx} 72 h) was observed. The intracellular ROS generation and cytokine production induced cytotoxicity point towards the mechanistic pathway of cell death upon exposure to PAMAM dendrimers.« less

Effect of low power laser irradiation on macrophage phagocytic capacity

NASA Astrophysics Data System (ADS)

Lu, Cuixia; Song, Sheng; Tang, Yu; Zhou, Feifan

2011-03-01

Phagocytosis and subsequent degradation of pathogens by macrophages play a pivotal role in host innate immunity in mammals. Laser irradiation has been found to produce photobiological effects with evidence of interference with immunological functions. However, the effects of laser on the immune response have not been extensively characterized. In this study, we focused our attention on the effects of He-Ne laser on the phagocytic activity of macrophages by using flow cytometry (FCM). After irradiating at fluence of 0, 1, 2 J/cm2 with He-Ne laser (632.8 nm, 3mw), the cells were incubated with microsphere and then subjected to FACS analysis. The results showed that Low-power laser irradiation (LPLI) leads to an increase in phagocytosis on both mouse peritoneal macrophages and the murine macrophage-like cell line RAW264.7. In addition, we demonstrated that LPLI increased phagocytosis of microsphere in a dose-dependent manner, reaching a maximum at fluence of 2 J/cm2. Taken together, our results indicated that Low-power laser irradiation with appropriate dosage can enhance the phagocytosis of macrophage, and provided a theoretical base for the clinical use of the He-Ne laser.

The effect of aluminium and sodium impurities on the in vitro toxicity and pro-inflammatory potential of cristobalite

USGS Publications Warehouse

Nattrass, C.; Horwell, Claire J.; Damby, David; Brown, David; Stone, Vicki

2017-01-01

BackgroundExposure to crystalline silica (SiO2), in the form of quartz, tridymite or cristobalite, can cause respiratory diseases, such as silicosis. However, the observed toxicity and pathogenicity of crystalline silica is highly variable. This has been attributed to a number of inherent and external factors, including the presence of impurities. In cristobalite-rich dusts, substitutions of aluminium (Al) for silicon (Si) in the cristobalite structure, and impurities occluding the silica surface, have been hypothesised to decrease its toxicity. This hypothesis is tested here through the characterisation and in vitro toxicological study of synthesised cristobalite with incremental amounts of Al and sodium (Na) dopants. MethodsSamples of synthetic cristobalite with incremental amounts of Al and Na impurities, and tridymite, were produced through heating of a silica sol-gel. Samples were characterised for mineralogy, cristobalite purity and abundance, particle size, surface area and surface charge. In vitro assays assessed the ability of the samples to induce cytotoxicity and TNF-α production in J774 macrophages, and haemolysis of red blood cells. ResultsAl-only doped or Al+Na co-doped cristobalite contained between 1 and 4 oxide wt% Al and Na within its structure. Co-doped samples also contained Al- and Na-rich phases, such as albite. Doping reduced cytotoxicity to J774 macrophages and haemolytic capacity compared to non-doped samples. Al-only doping was more effective at decreasing cristobalite reactivity than Al+Na co-doping. The reduction in the reactivity of cristobalite is attributed to both structural impurities and a lower abundance of crystalline silica in doped samples. Neither non-doped nor doped crystalline silica induced production of the pro-inflammatory cytokine TNF-α in J774 macrophages. ConclusionsImpurities can reduce the toxic potential of cristobalite and may help explain the low reactivity of some cristobalite-rich dusts. Whilst further work

[Molecular mechanisms and relationship of M2-polarized macrophages with early response in multiple myeloma].

PubMed

Chen, X Y; Sun, R X; Zhang, W Y; Liu, T; Zheng, Y H; Wu, Y

2017-06-14

Objective: To investigate the relationship between M2-polarized macrophages and early response in multiple myeloma and its molecular mechanism. Methods: Two hundred and forty bone marrow biopsy tissue were collected and M2-polarized macrophages were stained by anti-CD163 monoclonal antibody. In vitro M2-polarized macrophages were derived from human peripheral blood mononuclear cell or THP-1 cells and identified by flow cytometry. Two myeloma cell lines RPMI 8226 and U266 were co-cultured with M2 macrophages using a transwell system. We measured myeloma cells proliferation through CCK-8 method and the pro-inflammatory cytokines expression (TNF-α and IL-6) by ELISA. Real time PCR was applied to measure chemokines (CCL2 and CCL3) , chemokine receptors (CCR2, CCR5) , VEGF and their receptors. In addition, flow cytometry was used to analyze the apoptosis of myeloma cells induced by dexamethasone. Results: ①Patients with high percentage of M2 macrophage involvement in bone marrow showed poorer response (23.9% versus 73.0%, χ (2)=60.31, P <0.001). ② In vitro the proliferation of RPMI 8226 cells ( P =0.005 at 24 h, P =0.020 at 36 h) or U266 myeloma cells ( P = 0.030 at 24h, P =0.020 at 36h) co-cultured with M2-polarized macrophages was higher than control group. ③In vitro the apoptotic rate of RPMI 8226 cells (29.0% versus 71.0%, t =4.97, P =0.008) or U266 myeloma cells (24.9% versus 67.7%, t =6.99, P =0.002) co-cultured with M2-polarized macrophages was lower than control group. ④ In vitro M2-polarized macrophages promoted myeloma cells secreting higher level of IL-6, TNF-α and higher expression of CCL2, CCL3, CCR2, CCR5, VEGFA, VEGFR-1,-2 compared with the non-macrophage co-culture system. Conclusion: M2-polarized macrophages promote myeloma cells proliferation and inhibit apoptosis through a very complex mechanism involving pro-inflammatory cytokines IL-6 and TNF-α, chemokines and related receptors such as CCL2, CCL3, CCR2, CCR3, and VEGF as well as related

Disruption of Serinc1, which facilitates serine-derived lipid synthesis, fails to alter macrophage function, lymphocyte proliferation or autoimmune disease susceptibility.

PubMed

Chu, Edward P F; Elso, Colleen M; Pollock, Abigail H; Alsayb, May A; Mackin, Leanne; Thomas, Helen E; Kay, Thomas W H; Silveira, Pablo A; Mansell, Ashley S; Gaus, Katharina; Brodnicki, Thomas C

2017-02-01

During immune cell activation, serine-derived lipids such as phosphatidylserine and sphingolipids contribute to the formation of protein signaling complexes within the plasma membrane. Altering lipid composition in the cell membrane can subsequently affect immune cell function and the development of autoimmune disease. Serine incorporator 1 (SERINC1) is a putative carrier protein that facilitates synthesis of serine-derived lipids. To determine if SERINC1 has a role in immune cell function and the development of autoimmunity, we characterized a mouse strain in which a retroviral insertion abolishes expression of the Serinc1 transcript. Expression analyses indicated that the Serinc1 transcript is readily detectable and expressed at relatively high levels in wildtype macrophages and lymphocytes. The ablation of Serinc1 expression in these immune cells, however, did not significantly alter serine-derived lipid composition or affect macrophage function and lymphocyte proliferation. Analyses of Serinc1-deficient mice also indicated that systemic ablation of Serinc1 expression did not affect viability, fertility or autoimmune disease susceptibility. These results suggest that Serinc1 is dispensable for certain immune cell functions and does not contribute to previously reported links between lipid composition in immune cells and autoimmunity. Copyright © 2016 Elsevier Ltd. All rights reserved.

Collaborating with the enemy: function of macrophages in the development of neoplastic disease.

PubMed

Eljaszewicz, Andrzej; Wiese, Małgorzata; Helmin-Basa, Anna; Jankowski, Michal; Gackowska, Lidia; Kubiszewska, Izabela; Kaszewski, Wojciech; Michalkiewicz, Jacek; Zegarski, Wojciech

2013-01-01

Due to the profile of released mediators (such as cytokines, chemokines, growth factors, etc.), neoplastic cells modulate the activity of immune system, directly affecting its components both locally and peripherally. This is reflected by the limited antineoplastic activity of the immune system (immunosuppressive effect), induction of tolerance to neoplastic antigens, and the promotion of processes associated with the proliferation of neoplastic tissue. Most of these responses are macrophages dependent, since these cells show proangiogenic properties, attenuate the adaptive response (anergization of naïve T lymphocytes, induction of Treg cell formation, polarization of immune response towards Th2, etc.), and support invasion and metastases formation. Tumor-associated macrophages (TAMs), a predominant component of leukocytic infiltrate, "cooperate" with the neoplastic tissue, leading to the intensified proliferation and the immune escape of the latter. This paper characterizes the function of macrophages in the development of neoplastic disease.

KRAS Mutation and Epithelial-Macrophage Interplay in Pancreatic Neoplastic Transformation.

PubMed

Bishehsari, Faraz; Zhang, Lijuan; Barlass, Usman; Preite, Nailliw; Turturro, Sanja; Najor, Matthew S; Shetuni, Brandon B; Zayas, Janet P; Mahdavinia, Mahboobeh; Abukhdeir, Abde M; Keshavarzian, Ali

2018-05-14

Pancreatic ductal adenocarcinoma (PDA) is characterized by epithelial mutations in KRAS and prominent tumor-associated inflammation, including macrophage infiltration. But knowledge of early interactions between neoplastic epithelium and macrophages in PDA carcinogenesis is limited. Using a pancreatic organoid model, we found that the expression of mutant KRAS in organoids increased i) ductal to acinar gene expression ratios, ii) epithelial cells proliferation, and iii) colony formation capacity in vitro, and endowed pancreatic cells with the ability to generate neoplastic tumors in vivo. KRAS mutations induced a pro-tumorigenic phenotype in macrophages. Altered macrophages decreased epithelial Pigment Epithelial Derived Factor (PEDF) expression and induced a cancerous phenotype. We validated our findings using annotated patient samples from The Cancer Genome Atlas (TCGA) as well as in our human PDA specimens. Epithelium-macrophage cross talk occurs early in pancreatic carcinogenesis where KRAS directly induces cancer-related phenotypes in epithelium, and also promotes a pro-tumorigenic phenotype in macrophages, in turn augmenting neoplastic growth. This article is protected by copyright. All rights reserved. © 2018 UICC.

Mineralocorticoid Receptor Deficiency in Macrophages Inhibits Neointimal Hyperplasia and Suppresses Macrophage Inflammation Through SGK1-AP1/NF-κB Pathways.

PubMed

Sun, Jian-Yong; Li, Chao; Shen, Zhu-Xia; Zhang, Wu-Chang; Ai, Tang-Jun; Du, Lin-Juan; Zhang, Yu-Yao; Yao, Gao-Feng; Liu, Yan; Sun, Shuyang; Naray-Fejes-Toth, Aniko; Fejes-Toth, Geza; Peng, Yong; Chen, Mao; Liu, Xiaojing; Tao, Jun; Zhou, Bin; Yu, Ying; Guo, Feifan; Du, Jie; Duan, Sheng-Zhong

2016-05-01

Restenosis after percutaneous coronary intervention remains to be a serious medical problem. Although mineralocorticoid receptor (MR) has been implicated as a potential target for treating restenosis, the cellular and molecular mechanisms are largely unknown. This study aims to explore the functions of macrophage MR in neointimal hyperplasia and to delineate the molecular mechanisms. Myeloid MR knockout (MMRKO) mice and controls were subjected to femoral artery injury. MMRKO reduced intima area and intima/media ratio, Ki67- and BrdU-positive vascular smooth muscle cells, expression of proinflammatory molecules, and macrophage accumulation in injured arteries. MMRKO macrophages migrated less in culture. MMRKO decreased Ki67- and BrdU-positive macrophages in injured arteries. MMRKO macrophages were less Ki67-positive in culture. Conditioned media from MMRKO macrophages induced less migration, Ki67 positivity, and proinflammatory gene expression of vascular smooth muscle cells. After lipopolysaccharide treatment, MMRKO macrophages had decreased p-cFos and p-cJun compared with control macrophages, suggesting suppressed activation of activator protein-1 (AP1). Nuclear factor-κB (NF-κB) pathway was also inhibited by MMRKO, manifested by decreased p-IκB kinase-β and p-IκBα, increased IκBα expression, decreased nuclear translocation of p65 and p50, as welll as decreased phosphorylation and expression of p65. Finally, overexpression of serum-and-glucocorticoid-inducible-kinase-1 (SGK1) attenuated the effects of MR deficiency in macrophages. Selective deletion of MR in myeloid cells limits macrophage accumulation and vascular inflammation and, therefore, inhibits neointimal hyperplasia and vascular remodeling. Mechanistically, MR deficiency suppresses migration and proliferation of macrophages and leads to less vascular smooth muscle cell activation. At the molecular level, MR deficiency suppresses macrophage inflammatory response via SGK1-AP1/NF-κB pathways. �

Macrophage Phenotypes Regulate Scar Formation and Chronic Wound Healing.

PubMed

Hesketh, Mark; Sahin, Katherine B; West, Zoe E; Murray, Rachael Z

2017-07-17

Macrophages and inflammation play a beneficial role during wound repair with macrophages regulating a wide range of processes, such as removal of dead cells, debris and pathogens, through to extracellular matrix deposition re-vascularisation and wound re-epithelialisation. To perform this range of functions, these cells develop distinct phenotypes over the course of wound healing. They can present with a pro-inflammatory M1 phenotype, more often found in the early stages of repair, through to anti-inflammatory M2 phenotypes that are pro-repair in the latter stages of wound healing. There is a continuum of phenotypes between these ranges with some cells sharing phenotypes of both M1 and M2 macrophages. One of the less pleasant consequences of quick closure, namely the replacement with scar tissue, is also regulated by macrophages, through their promotion of fibroblast proliferation, myofibroblast differentiation and collagen deposition. Alterations in macrophage number and phenotype disrupt this process and can dictate the level of scar formation. It is also clear that dysregulated inflammation and altered macrophage phenotypes are responsible for hindering closure of chronic wounds. The review will discuss our current knowledge of macrophage phenotype on the repair process and how alterations in the phenotypes might alter wound closure and the final repair quality.

Simplified Quantitative Assay System for Measuring Activities of Drugs against Intracellular Legionella pneumophila

PubMed Central

Higa, Futoshi; Kusano, Nobuchika; Tateyama, Masao; Shinzato, Takashi; Arakaki, Noriko; Kawakami, Kazuyoshi; Saito, Atsushi

1998-01-01

We developed a new simple assay for the quantitation of the activities of drugs against intracellular Legionella pneumophila. The cells of a murine macrophage-like cell line (J774.1 cells) allowed the intracellular growth and replication of the bacteria, which ultimately resulted in cell death. The infected J774.1 cell monolayers in 96-well microplates were first treated with antibiotics and were further cultured for 72 h. The number of viable J774.1 cells in each well was quantified by a colorimetric assay with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and an enzyme-linked immunosorbent assay reader. The number of growing bacteria in each well was also determined by counting the numbers of CFU on buffered charcoal yeast extract-α agar plates. Viable J774.1 cell counts, determined by the colorimetric assay, were inversely proportional to the number of intracellular replicating bacteria. The minimum extracellular concentrations (MIECs) of 24 antibiotics causing inhibition of intracellular growth of L. pneumophila were determined by the colorimetric assay system. The MIECs of beta-lactams and aminoglycosides were markedly higher than the MICs in buffered yeast extract-α broth. The MIECs of macrolides, fluoroquinolones, rifampin, and minocycline were similar to the respective MICs. According to their intracellular activities, clarithromycin and sparfloxacin were the most potent among the macrolides or fluoroquinolones tested in this study. Our results indicated that the MTT assay system allows comparative and quantitative evaluations of the intracellular activities of antibiotics and efficient processing of a large number of samples. PMID:9574712

Leishmania carbon metabolism in the macrophage phagolysosome- feast or famine?

PubMed

McConville, Malcolm J; Saunders, Eleanor C; Kloehn, Joachim; Dagley, Michael J

2015-01-01

A number of medically important microbial pathogens target and proliferate within macrophages and other phagocytic cells in their mammalian hosts. While the majority of these pathogens replicate within the host cell cytosol or non-hydrolytic vacuolar compartments, a few, including protists belonging to the genus Leishmania, proliferate long-term within mature lysosome compartments.  How these parasites achieve this feat remains poorly defined. In this review, we highlight recent studies that suggest that Leishmania virulence is intimately linked to programmed changes in the growth rate and carbon metabolism of the obligate intra-macrophage stages. We propose that activation of a slow growth and a stringent metabolic response confers resistance to multiple stresses (oxidative, temperature, pH), as well as both nutrient limitation and nutrient excess within this niche. These studies highlight the importance of metabolic processes as key virulence determinants in Leishmania.

M2 Polarization of Human Macrophages Favors Survival of the Intracellular Pathogen Chlamydia pneumoniae.

PubMed

Buchacher, Tanja; Ohradanova-Repic, Anna; Stockinger, Hannes; Fischer, Michael B; Weber, Viktoria

2015-01-01

Intracellular pathogens have developed various strategies to escape immunity to enable their survival in host cells, and many bacterial pathogens preferentially reside inside macrophages, using diverse mechanisms to penetrate their defenses and to exploit their high degree of metabolic diversity and plasticity. Here, we characterized the interactions of the intracellular pathogen Chlamydia pneumoniae with polarized human macrophages. Primary human monocytes were pre-differentiated with granulocyte macrophage colony-stimulating factor or macrophage colony-stimulating factor for 7 days to yield M1-like and M2-like macrophages, which were further treated with interferon-γ and lipopolysaccharide or with interleukin-4 for 48 h to obtain fully polarized M1 and M2 macrophages. M1 and M2 cells exhibited distinct morphology with round or spindle-shaped appearance for M1 and M2, respectively, distinct surface marker profiles, as well as different cytokine and chemokine secretion. Macrophage polarization did not influence uptake of C. pneumoniae, since comparable copy numbers of chlamydial DNA were detected in M1 and M2 at 6 h post infection, but an increase in chlamydial DNA over time indicating proliferation was only observed in M2. Accordingly, 72±5% of M2 vs. 48±7% of M1 stained positive for chlamydial lipopolysaccharide, with large perinuclear inclusions in M2 and less clearly bordered inclusions for M1. Viable C. pneumoniae was present in lysates from M2, but not from M1 macrophages. The ability of M1 to restrict chlamydial replication was not observed in M1-like macrophages, since chlamydial load showed an equal increase over time for M1-like and M2-like macrophages. Our findings support the importance of macrophage polarization for the control of intracellular infection, and show that M2 are the preferred survival niche for C. pneumoniae. M1 did not allow for chlamydial proliferation, but failed to completely eliminate chlamydial infection, giving further evidence

Tumor-associated macrophages as major source of APRIL in gastric MALT lymphoma.

PubMed

Munari, Fabio; Lonardi, Silvia; Cassatella, Marco A; Doglioni, Claudio; Cangi, Maria Giulia; Amedei, Amedeo; Facchetti, Fabio; Eishi, Yoshinobu; Rugge, Massimo; Fassan, Matteo; de Bernard, Marina; D'Elios, Mario M; Vermi, William

2011-06-16

Lymphoid hyperplasia of gastric mucosa associated with Helicobacter pylori (HP) infection represents a preneoplastic condition of the mucosa associated lymphoid tissue (MALT), which may evolve to a B-cell lymphoma. While it is well established that the initial neoplastic proliferation of B cells is antigen-driven and dependent on the helper activity of HP-specific T cells, it needs to be elucidated which cytokine or soluble factor(s) promote B-cell activation and lymphomagenesis. Herein, we originally report that gastric MALT lymphoma express high levels of a proliferation inducing ligand (APRIL), a novel cytokine crucial in sustaining B-cell proliferation. By immunohistochemistry, we demonstrate that APRIL is produced almost exclusively by gastric lymphoma-infiltrating macrophages located in close proximity to neoplastic B cells. We also show that macrophages produce APRIL on direct stimulation with both HP and HP-specific T cells. Collectively, our results represent the first evidence for an involvement of APRIL in gastric MALT lymphoma development in HP-infected patients.

Feedback inhibition of nitric oxide synthase activity by nitric oxide.

PubMed Central

Assreuy, J.; Cunha, F. Q.; Liew, F. Y.; Moncada, S.

1993-01-01

1. A murine macrophage cell line, J774, expressed nitric oxide (NO) synthase activity in response to interferon-gamma (IFN-gamma, 10 u ml-1) plus lipopolysaccharide (LPS, 10 ng ml-1). The enzyme activity was first detectable 6 h after incubation, peaked at 12 h and became undetectable after 48 h. 2. The decline in the NO synthase activity was not due to inhibition by stable substances secreted by the cells into the culture supernatant. 3. The decline in the NO synthase activity was significantly slowed down in cells cultured in a low L-arginine medium or with added haemoglobin, suggesting that NO may be involved in a feedback inhibitory mechanism. 4. The addition of NO generators, S-nitroso-acetyl-penicillamine (SNAP) or S-nitroso-glutathione (GSNO) markedly inhibited the NO synthase activity in a dose-dependent manner. The effect of NO on the enzyme was not due to the inhibition of de novo protein synthesis. 5. SNAP directly inhibited the inducible NO synthase extracted from activated J774 cells, as well as the constitutive NO synthase extracted from the rat brain. 6. The enzyme activity of J774 cells was not restored after the removal of SNAP by gel filtration, suggesting that NO inhibits NO synthase irreversibly. PMID:7682140

Inducible CYP2J2 and its product 11,12-EET promotes bacterial phagocytosis: a role for CYP2J2 deficiency in the pathogenesis of Crohn's disease?

PubMed

Bystrom, Jonas; Thomson, Scott J; Johansson, Jörgen; Edin, Matthew L; Zeldin, Darryl C; Gilroy, Derek W; Smith, Andrew M; Bishop-Bailey, David

2013-01-01

The epoxygenase CYP2J2 has an emerging role in inflammation and vascular biology. The role of CYP2J2 in phagocytosis is not known and its regulation in human inflammatory diseases is poorly understood. Here we investigated the role of CYP2J2 in bacterial phagocytosis and its expression in monocytes from healthy controls and Crohns disease patients. CYP2J2 is anti-inflammatory in human peripheral blood monocytes. Bacterial LPS induced CYP2J2 mRNA and protein. The CYP2J2 arachidonic acid products 11,12-EET and 14,15-EET inhibited LPS induced TNFα release. THP-1 monocytes were transformed into macrophages by 48h incubation with phorbol 12-myristate 13-acetate. Epoxygenase inhibition using a non-selective inhibitor SKF525A or a selective CYP2J2 inhibitor Compound 4, inhibited E. coli particle phagocytosis, which could be specifically reversed by 11,12-EET. Moreover, epoxygenase inhibition reduced the expression of phagocytosis receptors CD11b and CD68. CD11b also mediates L. monocytogenes phagocytosis. Similar, to E. coli bioparticle phagocytosis, epoxygenase inhibition also reduced intracellular levels of L. monocytogenes, which could be reversed by co-incubation with 11,12-EET. Disrupted bacterial clearance is a hallmark of Crohn's disease. Unlike macrophages from control donors, macrophages from Crohn's disease patients showed no induction of CYP2J2 in response to E. coli. These results demonstrate that CYP2J2 mediates bacterial phagocytosis in macrophages, and implicates a defect in the CYP2J2 pathway may regulate bacterial clearance in Crohn's disease.

Macrophage Phenotype and Function in Different Stages of Atherosclerosis

PubMed Central

Tabas, Ira; Bornfeldt, Karin E.

2016-01-01

The remarkable plasticity and plethora of biological functions performed by macrophages have enticed scientists to study these cells in relation to atherosclerosis for more than 50 years, and major discoveries continue to be made today. It is now understood that macrophages play important roles in all stages of atherosclerosis, from initiation of lesions and lesion expansion, to necrosis leading to rupture and the clinical manifestations of atherosclerosis, to resolution and regression of atherosclerotic lesions. Lesional macrophages are derived primarily from blood monocytes, although recent research has shown that lesional macrophage-like cells can also be derived from smooth muscle cells. Lesional macrophages take on different phenotypes depending on their environment and which intracellular signaling pathways are activated. Rather than a few distinct populations of macrophages, the phenotype of the lesional macrophage is more complex and likely changes during the different phases of atherosclerosis and with the extent of lipid and cholesterol loading, activation by a plethora of receptors, and metabolic state of the cells. These different phenotypes allow the macrophage to engulf lipids, dead cells, and other substances perceived as danger signals; efflux cholesterol to HDL; proliferate and migrate; undergo apoptosis and death; and secrete a large number of inflammatory and pro-resolving molecules. This review article, part of the Compendium on Atherosclerosis, discusses recent advances in our understanding of lesional macrophage phenotype and function in different stages of atherosclerosis. With the increasing understanding of the roles of lesional macrophages, new research areas and treatment strategies are beginning to emerge. PMID:26892964

FcγRI (CD64): an identity card for intestinal macrophages.

PubMed

De Calisto, Jaime; Villablanca, Eduardo J; Mora, J Rodrigo

2012-12-01

Macrophages are becoming increasingly recognized as key cellular players in intestinal immune homeostasis. However, differentiating between macrophages and dendritic cells (DCs) is often difficult, and finding a specific phenotypic signature for intestinal macrophage identification has remained elusive. In this issue of the European Journal of Immunology, Tamoutounour et al. [Eur. J. Immunol. 2012. 42: 3150-3166] identify CD64 as a specific macrophage marker that can be used to discriminate DCs from macrophages in the murine small and large intestine, under both steady-state and inflammatory conditions. The authors also propose a sequential 'monocyte-waterfall' model for intestinal macrophage differentiation, with implications for immune tolerance and inflammation at the gut mucosal interface. This Commentary will discuss the advantages and potential limitations of CD64 as a marker for intestinal macrophages. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Constant replenishment from circulating monocytes maintains the macrophage pool in adult intestine

PubMed Central

Scott, Charlotte L.; Perdiguero, Elisa Gomez; Geissmann, Frederic; Henri, Sandrine; Malissen, Bernard; Osborne, Lisa C.; Artis, David; Mowat, Allan McI.

2014-01-01

The paradigm that resident macrophages in steady-state tissues are derived from embryonic precursors has never been investigated in the intestine, which contains the largest pool of macrophages. Using fate mapping models and monocytopenic mice, together with bone marrow chimeric and parabiotic models, we show that embryonic precursors seeded the intestinal mucosa and demonstrated extensive in situ proliferation in the neonatal period. However these cells did not persist in adult intestine. Instead, they were replaced around the time of weaning by the CCR2-dependent influx of Ly6Chi monocytes that differentiated locally into mature, anti-inflammatory macrophages. This process was driven largely by the microbiota and had to be continued throughout adult life to maintain a normal intestinal macrophage pool. PMID:25151491

Infiltration of Macrophages Correlates with Severity of Allograft Rejection and Outcome in Human Kidney Transplantation.

PubMed

Bergler, Tobias; Jung, Bettina; Bourier, Felix; Kühne, Louisa; Banas, Miriam C; Rümmele, Petra; Wurm, Simone; Banas, Bernhard

2016-01-01

Despite substantial progress in recent years, graft survival beyond the first year still requires improvement. Since modern immunosuppression addresses mainly T-cell activation and proliferation, we studied macrophage infiltration into the allografts of 103 kidney transplant recipients during acute antibody and T-cell mediated rejection. Macrophage infiltration was correlated with both graft function and graft survival until month 36 after transplantation. Macrophage infiltration was significantly elevated in antibody-mediated and T-cell mediated rejection, but not in kidneys with established IFTA. Treatment of rejection with steroids was less successful in patients with more prominent macrophage infiltration into the allografts. Macrophage infiltration was accompanied by increased cell proliferation as well as antigen presentation. With regard to the compartmental distribution severity of T-cell-mediated rejection was correlated to the amount of CD68+ cells especially in the peritubular and perivascular compartment, whereas biopsies with ABMR showed mainly peritubular CD68 infiltration. Furthermore, severity of macrophage infiltration was a valid predictor of resulting creatinine values two weeks as well as two and three years after renal transplantation as illustrated by multivariate analysis. Additionally performed ROC curve analysis showed that magnitude of macrophage infiltration (below vs. above the median) was a valid predictor for the necessity to restart dialysis. Having additionally stratified biopsies in accordance to the magnitude of macrophage infiltration, differential CD68+ cell infiltration was reflected by striking differences in overall graft survival. The differences in acute allograft rejection have not only been reflected by different magnitudes of macrophage infiltration, but also by compartment-specific infiltration pattern and subsequent impact on resulting allograft function as well as need for dialysis initiation. There is a robust

Infiltration of Macrophages Correlates with Severity of Allograft Rejection and Outcome in Human Kidney Transplantation

PubMed Central

Bourier, Felix; Kühne, Louisa; Banas, Miriam C.; Rümmele, Petra; Wurm, Simone; Banas, Bernhard

2016-01-01

Objective Despite substantial progress in recent years, graft survival beyond the first year still requires improvement. Since modern immunosuppression addresses mainly T-cell activation and proliferation, we studied macrophage infiltration into the allografts of 103 kidney transplant recipients during acute antibody and T-cell mediated rejection. Macrophage infiltration was correlated with both graft function and graft survival until month 36 after transplantation. Results Macrophage infiltration was significantly elevated in antibody-mediated and T-cell mediated rejection, but not in kidneys with established IFTA. Treatment of rejection with steroids was less successful in patients with more prominent macrophage infiltration into the allografts. Macrophage infiltration was accompanied by increased cell proliferation as well as antigen presentation. With regard to the compartmental distribution severity of T-cell-mediated rejection was correlated to the amount of CD68+ cells especially in the peritubular and perivascular compartment, whereas biopsies with ABMR showed mainly peritubular CD68 infiltration. Furthermore, severity of macrophage infiltration was a valid predictor of resulting creatinine values two weeks as well as two and three years after renal transplantation as illustrated by multivariate analysis. Additionally performed ROC curve analysis showed that magnitude of macrophage infiltration (below vs. above the median) was a valid predictor for the necessity to restart dialysis. Having additionally stratified biopsies in accordance to the magnitude of macrophage infiltration, differential CD68+ cell infiltration was reflected by striking differences in overall graft survival. Conclusion The differences in acute allograft rejection have not only been reflected by different magnitudes of macrophage infiltration, but also by compartment-specific infiltration pattern and subsequent impact on resulting allograft function as well as need for dialysis

Swift obervation of PSN J10250739+1709146 in NGC 3239

NASA Astrophysics Data System (ADS)

Xu, D.

2012-01-01

The optical transient, PSN J10250739+1709146, was discovered by Moore, Newton and Puckett at 2012/01/07.387 (ref. http://www.cbat.eps.harvard.edu/unconf/followups/J10250739+1709146.html), and localised in NGC 3239 (z=0.002512). Spectroscopy at 2012/01/10.275 confirms this source as a young SN II (Cao et al., ATel #3855). A Swift-ToO was executed to observe the field of this SN starting from 2012/01/10.774 (exposure time: 2390s), with the primary aim to constrain any accompanying X-ray emission.

Toxicological Profiling of Metal Oxide Nanoparticles in Liver Context Reveals Pyroptosis in Kupffer Cells and Macrophages versus Apoptosis in Hepatocytes.

PubMed

Mirshafiee, Vahid; Sun, Bingbing; Chang, Chong Hyun; Liao, Yu-Pei; Jiang, Wen; Jiang, Jinhong; Liu, Xiangsheng; Wang, Xiang; Xia, Tian; Nel, André E

2018-04-24

The liver and the mononuclear phagocyte system are a frequent target for engineered nanomaterials, either as a result of particle uptake and spread from primary exposure sites or systemic administration of therapeutic and imaging nanoparticles. In this study, we performed a comparative analysis of the toxicological impact of 29 metal oxide nanoparticles (NPs), some commonly used in consumer products, in transformed or primary Kupffer cells (KCs) and hepatocytes. We not only observed differences between KCs and hepatocytes, but also differences in the toxicological profiles of transition-metal oxides (TMOs, e. g., Co 3 O 4 ) versus rare-earth oxide (REO) NPs ( e. g., Gd 2 O 3 ). While pro-oxidative TMOs induced the activation of caspases 3 and 7, resulting in apoptotic cell death in both cell types, REOs induced lysosomal damage, NLRP3 inflammasome activation, caspase 1 activation, and pyroptosis in KCs. Pyroptosis was accompanied by cell swelling, membrane blebbing, IL-1β release, and increased membrane permeability, which could be reversed by knockdown of the pore forming protein, gasdermin D. Though similar features were not seen in hepatocytes, the investigation of the cytotoxic effects of REO NPs could also be seen to affect macrophage cell lines such as J774A.1 and RAW 264.7 cells as well as bone marrow-derived macrophages. These phagocytic cell types also demonstrated features of pyroptosis and increased IL-1β production. Collectively, these findings demonstrate important mechanistic considerations that can be used for safety evaluation of metal oxides, including commercial products that are developed from these materials.

A solar super-flare as cause for the 14C variation in AD 774/5 ?

NASA Astrophysics Data System (ADS)

Neuhäuser, R.; Hambaryan, V. V.

2014-11-01

We present further considerations regarding the strong 14C variation in AD 774/5. For its cause, either a solar super-flare or a short gamma-ray burst were suggested. We show that all kinds of stellar or neutron star flares would be too weak for the observed energy input at Earth in AD 774/5. Even though Maehara et al. (2012) present two super-flares with {˜ 1035} erg of presumably solar-type stars, we would like to caution: These two stars are poorly studied and may well be close binaries, and/or having a M-type dwarf companion, and/or may be much younger and/or much more magnetic than the Sun - in any such case, they might not be true solar analog stars. From the frequency of large stellar flares averaged over all stellar activity phases (maybe obtained only during grand activity maxima), one can derive (a limit of) the probability for a large solar flare at a random time of normal activity: We find the probability for one flare within 3000 years to be possibly as low as 0.3 to 0.008 considering the full 1σ error range. Given the energy estimate in Miyake et al. (2012) for the AD 774/5 event, it would need to be {˜ 2000} stronger than the Carrington event as solar super-flare. If the AD 774/5 event as solar flare would be beamed (to an angle of only {˜ 24°}), 100 times lower energy would be needed. A new AD 774/5 energy estimate by Usoskin et al. (2013) with a different carbon cycle model, yielding 4 ot 6 time lower 14C production, predicts 4-6 times less energy. If both reductions are applied, the AD 774/5 event would need to be only ˜ 4 times stronger than the Carrington event in 1859 (if both had similar spectra). However, neither 14C nor 10Be peaks were found around AD 1859. Hence, the AD 774/5 event (as solar flare) either was not beamed that strongly, and/or it would have been much more than 4-6 times stronger than Carrington, and/or the lower energy estimate (Usoskin et al. 2013) is not correct, and/or such solar flares cannot form (enough) 14C and

Impairment of Macrophage Presenting Ability and Viability by Echinococcus granulosus Antigens.

PubMed

Mejri, Naceur; Hassen, Imed Eddine; Knapp, Jenny; Saidi, Mouldi

2017-03-01

Despite advances toward an improved understanding of the evasive mechanisms leading to the establishment of cystic echinococcosis, the discovery of specific immunosuppressive mechanisms and related factors are of great interest in the development of an immunotherapeutic approach. To elucidate immunosuppressive effects of bioactive factors contained in chromatographic fractions from hydatid cystic fluid (HCF) of Echinococcus granulosus. Hydatid cystic fluid was fractionated by reverse phase chromatography. Non-specific Concanavalin A-driven proliferation of spleen cells was used to determine specific inhibitory fractions. Trypan blue exclusion test and flowcytometry analysis were performed to check whether highly inhibitory fractions of HCF have apoptotic effect on peritoneal macrophages. Western blot analysis was used to determine proteolytic effects of parasitic antigens on major histocompatibility complex (MHC) class II (I-a) contained in membrane proteins extract from macrophages. High concentrations of HCF and few of chromatographic fractions suppressed spleen cells proliferation. Fractions 7 and 35 were the highest inhibitory fractions. Specifically fraction 35 and to a lesser extent HCF induced apoptosis in peritoneal naive macrophages. However, HCF and the fraction 7 proteolytically altered the expression of MHC class II molecules on peritoneal macrophages. The proteolytic molecule was identified to be a serine protease. Macrophages taken at the chronic and end phase from cystic echinococcosis-infected mice were able to uptake and process C-Ovalbumine-FITC. These cells expressed a drastically reduced level of (I-a) molecules. Our study present new aspects of immune suppression function of E. granulosus. Further molecular characterization of apoptotic and proteolytic factors might be useful to develop immunotherapeutic procedure to break down their inhibitory effects.

Inducible CYP2J2 and Its Product 11,12-EET Promotes Bacterial Phagocytosis: A Role for CYP2J2 Deficiency in the Pathogenesis of Crohn’s Disease?

PubMed Central

Bystrom, Jonas; Thomson, Scott J.; Johansson, Jörgen; Edin, Matthew L.; Zeldin, Darryl C.; Gilroy, Derek W.; Smith, Andrew M.; Bishop-Bailey, David

2013-01-01

The epoxygenase CYP2J2 has an emerging role in inflammation and vascular biology. The role of CYP2J2 in phagocytosis is not known and its regulation in human inflammatory diseases is poorly understood. Here we investigated the role of CYP2J2 in bacterial phagocytosis and its expression in monocytes from healthy controls and Crohns disease patients. CYP2J2 is anti-inflammatory in human peripheral blood monocytes. Bacterial LPS induced CYP2J2 mRNA and protein. The CYP2J2 arachidonic acid products 11,12-EET and 14,15-EET inhibited LPS induced TNFα release. THP-1 monocytes were transformed into macrophages by 48h incubation with phorbol 12-myristate 13-acetate. Epoxygenase inhibition using a non-selective inhibitor SKF525A or a selective CYP2J2 inhibitor Compound 4, inhibited E. coli particle phagocytosis, which could be specifically reversed by 11,12-EET. Moreover, epoxygenase inhibition reduced the expression of phagocytosis receptors CD11b and CD68. CD11b also mediates L. monocytogenes phagocytosis. Similar, to E. coli bioparticle phagocytosis, epoxygenase inhibition also reduced intracellular levels of L. monocytogenes, which could be reversed by co-incubation with 11,12-EET. Disrupted bacterial clearance is a hallmark of Crohn’s disease. Unlike macrophages from control donors, macrophages from Crohn’s disease patients showed no induction of CYP2J2 in response to E. coli. These results demonstrate that CYP2J2 mediates bacterial phagocytosis in macrophages, and implicates a defect in the CYP2J2 pathway may regulate bacterial clearance in Crohn’s disease. PMID:24058654

Expression of M2-polarized macrophages is associated with poor prognosis for advanced epithelial ovarian cancer.

PubMed

Lan, Chunyan; Huang, Xin; Lin, Suxia; Huang, Huiqiang; Cai, Qichun; Wan, Ting; Lu, Jiabin; Liu, Jihong

2013-06-01

Macrophages are polarized into two functionally distinct forms, M1 and M2, in response to different microenvironment. Tumor-associated macrophages (TAMs) generally have M2 phenotype and promote tumor progression. Few studies to date have described the infiltration of M2-polarized macrophages in ovarian cancer. We used two macrophages markers, CD68 and CD163, to analyze the expression of TAMs and to clarify the relationship between the M2 form and survival in advanced ovarian cancer. Clinical data of 110 patients with stages III-IV epithelial ovarian cancer at Sun Yat-sen University Cancer Center between 1999 and 2007 were retrospectively reviewed. Immunohistochemical staining of CD68 and CD163 was performed. Correlations between macrophage density and patient survival were analyzed. Our data showed that no significant difference was observed in survival between patients in the high- and the low-CD68 expression groups. In contrast, the progression-free survival (PFS) rates (p = 0.003) and overall survival (OS) rates (p = 0.004) were significantly higher in the low-CD163 expression group than in the high-CD163 expression group, respectively. Similarly, we also observed significantly improved 3-year PFS (49.8% vs. 11.0%, p < 0.001) and OS (77.4% vs. 45.0%, p < 0.001) rates in patients in the low-CD163/CD68 ratio group when compared with the high-CD163/CD68 ratio group. Multivariate analysis identified the density of CD163-positive cells as well as the ratio of CD163/CD68 as negative predictors for PFS and OS, respectively. Our results show that the infiltration of CD163-positive M2 macrophages as well as activation of macrophages towards the M2 phenotype may contribute to poor survival in advanced ovarian cancer.

Biomaterial Property Effects on Platelets and Macrophages: An in Vitro Study.

PubMed

Fernandes, Kelly R; Zhang, Yang; Magri, Angela M P; Renno, Ana C M; van den Beucken, Jeroen J J P

2017-12-11

The purpose of this study was to evaluate the effects of surface properties of bone implants coated with hydroxyapatite (HA) and β-tricalcium phosphate (β-TCP) on platelets and macrophages upon implant installation and compare them to grit-blasted Ti and Thermanox used as a control. Surface properties were characterized using scanning electron microscopy, profilometry, crystallography, Fourier transform infrared spectroscopy, and coating stability. For platelets, platelet adherence and morphology were assessed. For macrophages, morphology, proliferation, and polarization were evaluated. Surface characterization showed similar roughness of ∼2.5 μm for grit-blasted Ti discs, both with and without coating. Coating stability assessment showed substantial dissolution of HA and β-TCP coatings. Platelet adherence was significantly higher for grit-blasted Ti, Ti-HA, and Ti-β-TCP coatings compared to that of cell culture control Thermanox. Macrophage cultures revealed a decreased proliferation on both HA and β-TCP coated discs compared to both Thermanox and grit-blasted Ti. In contrast, secretion of pro-inflammatory cytokine TNF-α and anti-inflammatory cytokine TGF-β were marginal for grit-blasted Ti and Thermanox, while a coating-dependent increased secretion of pro- and anti-inflammatory cytokines was observed for HA and β-TCP coatings. The results demonstrated a significantly upregulated pro-inflammatory and anti-inflammatory cytokine secretion and marker gene expression of macrophages on HA and β-TCP coatings. Furthermore, HA induced an earlier M1 macrophage polarization but more M2 phenotype potency than β-TCP. In conclusion, our data showed that material surface affects the behaviors of first cell types attached to implants. Due to the demonstrated crucial roles of platelets and macrophages in bone healing and implant integration, this information will greatly aid the design of metallic implants for a higher rate of success in patients.

Does carbonation of steel slag particles reduce their toxicity? An in vitro approach.

PubMed

Ibouraadaten, Saloua; van den Brule, Sybille; Lison, Dominique

2015-06-01

Mineral carbonation can stabilize industrial residues and, in the steel industry, may contribute to simultaneously valorize CO2 emissions and slag. We hypothesized that, by restricting the leaching of metals of toxicological concern such as Cr and V, carbonation can suppress the toxicity of these materials. The cytotoxic activity (WST1 assay) of slag dusts collected from a stainless and a Linz-Donawitz (LD) steel plant, before and after carbonation, was examined in J774 macrophages. The release of Cr, V, Fe, Mn and Ni was measured after incubation in artificial lung fluids mimicking the extracellular and phagolysosomal milieu to which particles are confronted after inhalation. LD slag had the higher Fe, Mn and V content, and was more cytotoxic than stainless steel slag. The cytotoxic activity of LD but not of stainless dusts was reduced after carbonation. The cytotoxic activity of the dusts toward J774 macrophages necessitated a direct contact with the cells and was reduced in the presence of inhibitors of phagocytosis (cytochalasin D) or phagolysosome acidification (bafilomycin), pointing to a key role of metallic constituents released in phagolysosomes. This in vitro study supports a limited reduction of the cytotoxic activity of LD, but not of stainless, steel dusts upon carbonation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Potential Eye Drop Based on a Calix[4]arene Nanoassembly for Curcumin Delivery: Enhanced Drug Solubility, Stability, and Anti-Inflammatory Effect.

PubMed

Granata, Giuseppe; Paterniti, Irene; Geraci, Corrada; Cunsolo, Francesca; Esposito, Emanuela; Cordaro, Marika; Blanco, Anna Rita; Cuzzocrea, Salvatore; Consoli, Grazia M L

2017-05-01

Curcumin is an Indian spice with a wide spectrum of biological and pharmacological activities but poor aqueous solubility, rapid degradation, and low bioavailability that affect medical benefits. To overcome these limits in ophthalmic application, curcumin was entrapped in a polycationic calix[4]arene-based nanoaggregate by a simple and reproducible method. The calix[4]arene-curcumin supramolecular assembly (Calix-Cur) appeared as a clear colloidal solution consisting in micellar nanoaggregates with size, polydispersity index, surface potential, and drug loading percentage meeting the requirements for an ocular drug delivery system. The encapsulation in the calix[4]arene nanoassembly markedly enhanced the solubility, reduced the degradation, and improved the anti-inflammatory effects of curcumin compared to free curcumin in both in vitro and in vivo experiments. Calix-Cur did not compromise the viability of J774A.1 macrophages and suppressed pro-inflammatory marker expression in J774A.1 macrophages subjected to LPS-induced oxidative stress. Histological and immunohistochemical analyses showed that Calix-Cur reduced signs of inflammation in a rat model of LPS-induced uveitis when topically administrated in the eyes. Overall, the results supported the calix[4]arene nanoassembly as a promising nanocarrier for delivering curcumin to anterior ocular tissues.

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

PubMed

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

2015-11-01

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

Regulation of myeloproliferation and M2 macrophage programming in mice by Lyn/Hck, SHIP, and Stat5

PubMed Central

Xiao, Wenbin; Hong, Hong; Kawakami, Yuko; Lowell, Clifford A.; Kawakami, Toshiaki

2008-01-01

The proliferation and differentiation of hematopoietic stem cells (HSCs) is finely regulated by extrinsic and intrinsic factors via various signaling pathways. Here we have shown that, similar to mice deficient in the lipid phosphatase SHIP, loss of 2 Src family kinases, Lyn and Hck, profoundly affects HSC differentiation, producing hematopoietic progenitors with increased proliferation, reduced apoptosis, growth factor–independent survival, and skewed differentiation toward M2 macrophages. This phenotype culminates in a Stat5-dependent myeloproliferative disease that is accompanied by M2 macrophage infiltration of the lung. Expression of a membrane-bound form of SHIP in HSCs lacking both Lyn and Hck restored normal hematopoiesis and prevented myeloproliferation. In vitro and in vivo studies suggested the involvement of autocrine and/or paracrine production of IL-3 and GM-CSF in the increased proliferation and myeloid differentiation of HSCs. Thus, this study has defined a myeloproliferative transformation-sensitive signaling pathway, composed of Lyn/Hck, SHIP, autocrine/paracrine cytokines, and Stat5, that regulates HSC differentiation and M2 macrophage programming. PMID:18246197

Regulation of RAW 264.7 macrophages behavior on anodic TiO2 nanotubular arrays

NASA Astrophysics Data System (ADS)

Yao, Shenglian; Feng, Xujia; Li, Wenhao; Wang, Lu-Ning; Wang, Xiumei

2017-12-01

Titanium (Ti) implants with TiO2 nanotubular arrays on the surface could regulate cells adhesion, proliferation and differentiation to determine the bone integration. Additionally, the regulation of immune cells could improve osteogenesis or lead in appropriate immune reaction. Thus, we evaluate the behavior of RAW264.7 macrophages on TiO2 nanotubular arrays with a wide range diameter (from 20 to 120 nm) fabricated by an electrochemical anodization process. In this work, the proliferation, cell viability and cytokine/chemokine secretion were evaluated by CCK-8, live/dead staining and ELISA, respectively. SEM and confocal microscopy were used to observe the adhesion morphology. Results showed that the small size nanotube surface was benefit for the macrophages adhesion and proliferation, while larger size surface could reduce the inflammatory response. These findings contribute to the design of immune-regulating Ti implants surface that supports successful implantation.

β-elemene inhibits tumor-promoting effect of M2 macrophages in lung cancer.

PubMed

Yu, Xiaomu; Xu, Maoyi; Li, Na; Li, Zongjuan; Li, Hongye; Shao, Shujuan; Zou, Kun; Zou, Lijuan

2017-08-19

Macrophages in tumor are mostly M2-polarized and have been reported to promote tumorigenesis, which are also defined as tumor-associated macrophages (TAMs). β-elemene has therapeutic effects against several cancers, however, it remains unknown whether β-elemene could inhibit cancer by targeting TAMs. Herein, we examined the effect of β-elemene on macrophages to elucidate a novel mechanism of β-elemene in tumor therapy. We showed that the conditioned medium of M2 macrophages promoted lung cancer cells to migration, invasion and epithelial mesenchymal transition, which could be inhibited by β-elemene. Moreover, β-elemene regulated the polarization of macrophages from M2 to M1. β-elemene also inhibited the proliferation, migration, invasion of lung cancer cells and enhanced its radiosensitivity. These results indicate β-elemene suppresses lung cancer by regulating both macrophages and lung cancer cells, it is a promising drug for combination with chemotherapy or radiotherapy. Copyright © 2017 Elsevier Inc. All rights reserved.

Laboratory Aspects of Biological Warfare Agents

DTIC Science & Technology

2016-01-01

Embryonated chicken egg yolk sacs have typically been the method of choice for culture. They are inoculated when the embryos are 5-7 days old. The... chicken or mouse embryo fibroblasts, J774.16 mouse macrophages, L929 murine fibroblasts, HEL (human embryonic lung) or vero cells are more commonly...the family, Poxviridae, is a legacy of the original grouping of viruses associated with diseases that produced poxes in the skin, however, if

19 CFR 10.774 - Direct costs of processing operations.

Code of Federal Regulations, 2014 CFR

2014-04-01

... manufacture of the specific good, including fringe benefits, on-the-job training, and the costs of engineering..., design, engineering, and blueprint costs, to the extent that they are allocable to the specific good; (4... 19 Customs Duties 1 2014-04-01 2014-04-01 false Direct costs of processing operations. 10.774...

AMP-activated protein kinase enhances the phagocytic ability of macrophages and neutrophils

PubMed Central

Bae, Hong-Beom; Zmijewski, Jaroslaw W.; Deshane, Jessy S.; Tadie, Jean-Marc; Chaplin, David D.; Takashima, Seiji; Abraham, Edward

2011-01-01

Although AMPK plays well-established roles in the modulation of energy balance, recent studies have shown that AMPK activation has potent anti-inflammatory effects. In the present experiments, we examined the role of AMPK in phagocytosis. We found that ingestion of Escherichia coli or apoptotic cells by macrophages increased AMPK activity. AMPK activation increased the ability of neutrophils or macrophages to ingest bacteria (by 46±7.8 or 85±26%, respectively, compared to control, P<0.05) and the ability of macrophages to ingest apoptotic cells (by 21±1.4%, P<0.05 compared to control). AMPK activation resulted in cytoskeletal reorganization, including enhanced formation of actin and microtubule networks. Activation of PAK1/2 and WAVE2, which are downstream effectors of Rac1, accompanied AMPK activation. AMPK activation also induced phosphorylation of CLIP-170, a protein that participates in microtubule synthesis. The increase in phagocytosis was reversible by the specific AMPK inhibitor compound C, siRNA to AMPKα1, Rac1 inhibitors, or agents that disrupt actin or microtubule networks. In vivo, AMPK activation resulted in enhanced phagocytosis of bacteria in the lungs by 75 ± 5% vs. control (P<0.05). These results demonstrate a novel function for AMPK in enhancing the phagocytic activity of neutrophils and macrophages.—Bae, H. -B., Zmijewski, J. W., Deshane, J. S., Tadie, J. -M., Chaplin, D. D., Takashima, S., Abraham, E. AMP-activated protein kinase enhances the phagocytic ability of macrophages and neutrophils. PMID:21885655

The diverse functions of Src family kinases in macrophages

PubMed Central

Abram, Clare L.; Lowell, Clifford A.

2015-01-01

Macrophages are key components of the innate immune response. These cells possess a diverse repertoire of receptors that allow them to respond to a host of external stimuli including cytokines, chemokines, and pathogen-associated molecules. Signals resulting from these stimuli activate a number of macrophage functional responses such as adhesion, migration, phagocytosis, proliferation, survival, cytokine release and production of reactive oxygen and nitrogen species. The cytoplasmic tyrosine kinase Src and its family members (SFKs) have been implicated in many intracellular signaling pathways in macrophages, initiated by a diverse set of receptors ranging from integrins to Toll-like receptors. However, it has been difficult to implicate any given member of the family in any specific pathway. SFKs appear to have overlapping and complementary functions in many pathways. Perhaps the function of these enzymes is to modulate the overall intracellular signaling network in macrophages, rather than operating as exclusive signaling switches for defined pathways. In general, SFKs may function more like rheostats, influencing the amplitude of many pathways. PMID:18508521

Kinetics of tumor necrosis factor production by photodynamic-therapy-activated macrophages

NASA Astrophysics Data System (ADS)

Pass, Harvey I.; Evans, Steven; Perry, Roger; Matthews, Wilbert

1990-07-01

The ability of photodynamic therapy (PDT) to activate macrophages and produce cytokines, specifically tumor necrosis factor (TNF), is unknown. Three day thioglycolate elicited macrophages were incubated with 25 ug/mi Photofrin II (P11) for 2 hour, after which they were subjected to 630 nm light with fluences of 0-1800 J/m. The amount of TNF produced in the system as well as macrophage viability was measured 1, 3, 6, and 18 hours after POT. The level of TNF produced by the macrophages was significantly elevated over control levels 6 hours after POT and the absolute level of tumor necrosis factor production was influenced by the treatment energy and the resulting macrophage cytotoxicity. These data suggest that POT therapy induced cytotoxicity in vivo may be amplified by macrophage stimulation to secrete cytokines and these cytokines may also participate in other direct/indirect photodynamic therapy effects, i.e. immunosuppression, vascular effects.

Intratumoral Delivery of Interferonγ-Secreting Mesenchymal Stromal Cells Repolarizes Tumor-Associated Macrophages and Suppresses Neuroblastoma Proliferation In Vivo.

PubMed

Relation, Theresa; Yi, Tai; Guess, Adam J; La Perle, Krista; Otsuru, Satoru; Hasgur, Suheyla; Dominici, Massimo; Breuer, Christopher; Horwitz, Edwin M

2018-06-01

Neuroblastoma, the most common extracranial solid tumor in childhood, remains a therapeutic challenge. However, one promising patient treatment strategy is the delivery of anti-tumor therapeutic agents via mesenchymal stromal cell (MSC) therapy. MSCs have been safely used to treat genetic bone diseases such as osteogenesis imperfecta, cardiovascular diseases, autoimmune diseases, and cancer. The pro-inflammatory cytokine interferon-gamma (IFNγ) has been shown to decrease tumor proliferation by altering the tumor microenvironment (TME). Despite this, clinical trials of systemic IFNγ therapy have failed due to the high blood concentration required and associated systemic toxicities. Here, we developed an intra-adrenal model of neuroblastoma, characterized by liver and lung metastases. We then engineered MSCs to deliver IFNγ directly to the TME. In vitro, these MSCs polarized murine macrophages to the M1 phenotype. In vivo, we attained a therapeutically active TME concentration of IFNγ without increased systemic concentration or toxicity. The TME-specific IFNγ reduced tumor growth rate and increased survival in two models of T cell deficient athymic nude mice. Absence of this benefit in NOD SCID gamma (NSG) immunodeficient mouse model indicates a mechanism dependent on the innate immune system. IL-17 and IL-23p19, both uniquely M1 polarization markers, transiently increased in the tumor interstitial fluid. Finally, the MSC vehicle did not promote tumor growth. These findings reveal that MSCs can deliver effective cytokine therapy directly to the tumor while avoiding systemic toxicity. This method transiently induces inflammatory M1 macrophage polarization, which reduces tumor burden in our novel neuroblastoma murine model. Stem Cells 2018;36:915-924. © AlphaMed Press 2018.

Avian leukosis virus subgroup J promotes cell proliferation and cell cycle progression through miR-221 by targeting CDKN1B.

PubMed

Ren, Chaoqi; Yu, Mengmeng; Zhang, Yao; Fan, Minghui; Chang, Fangfang; Xing, Lixiao; Liu, Yongzhen; Wang, Yongqiang; Qi, Xiaole; Liu, Changjun; Zhang, Yanping; Cui, Hongyu; Li, Kai; Gao, Li; Pan, Qing; Wang, Xiaomei; Gao, Yulong

2018-06-01

Avian leukosis virus subgroup J (ALV-J), a highly oncogenic retrovirus, causes leukemia-like proliferative diseases in chickens. microRNAs post-transcriptionally suppress targets and are involved in the development of various tumors. We previously showed that miR-221 is upregulated in ALV-J-induced tumors. In this study, we analyzed the possible function of miR-221 in ALV-J tumorigenesis. The target validation system showed that CDKN1B is a target of miR-221 and is downregulated in ALV-J infection. As CDKN1B arrests the cell cycle and regulates its progression, we analyzed the proliferation of ALV-J-infected DF-1 cells. ALV-J-infection-induced DF1 cell derepression of G1/S transition and overproliferation required high miR-221 expression followed by CDKN1B downregulation. Cell cycle pathway analysis showed that ALV-J infection induced DF-1 cell overproliferation via the CDKN1B-CDK2/CDK6 pathway. Thus, miR-221 may play an important role in ALV-J-induced aggressive growth of DF-1 cells; these findings have expanded our insights into the mechanism underlying ALV-J infection and tumorigenesis. Copyright © 2018 Elsevier Inc. All rights reserved.

The role of HFE genotype in macrophage phenotype.

PubMed

Nixon, Anne M; Neely, Elizabeth; Simpson, Ian A; Connor, James R

2018-02-01

Iron regulation is essential for cellular energy production. Loss of cellular iron homeostasis has critical implications for both normal function and disease progression. The H63D variant of the HFE gene is the most common gene variant in Caucasians. The resulting mutant protein alters cellular iron homeostasis and is associated with a number of neurological diseases and cancer. In the brain, microglial and infiltrating macrophages are critical to maintaining iron homeostasis and modulating inflammation associated with the pathogenic process in multiple diseases. This study addresses whether HFE genotype affects macrophage function and the implications of these findings for disease processes. Bone marrow macrophages were isolated from wildtype and H67D HFE knock-in mice. The H67D gene variant in mice is the human equivalent of the H63D variant. Upon differentiation, the macrophages were used to analyze iron regulatory proteins, cellular iron release, migration, phagocytosis, and cytokine expression. The results of this study demonstrate that the H67D HFE genotype significantly impacts a number of critical macrophage functions. Specifically, fundamental activities such as proliferation in response to iron exposure, L-ferritin expression in response to iron loading, secretion of BMP6 and cytokines, and migration and phagocytic activity were all found to be impacted by genotype. Furthermore, we demonstrated that exposure to apo-Tf (iron-poor transferrin) can increase the release of iron from macrophages. In normal conditions, 70% of circulating transferrin is unsaturated. Therefore, the ability of apo-Tf to induce iron release could be a major regulatory mechanism for iron release from macrophages. These studies demonstrate that the HFE genotype impacts fundamental components of macrophage phenotype that could alter their role in degenerative and reparative processes in neurodegenerative disorders.

Toxicity and Carcinogenicity Mechanisms of Fibrous Antigorite

PubMed Central

Cardile, Venera; Lombardo, Laura; Belluso, Elena; Panico, Annamaria; Capella, Silvana; Balazy, Michael

2007-01-01

We studied the effects of fibrous antigorite on mesothelial MeT-5A and monocyte-macrophage J774 cell lines to further understand cellular mechanisms induced by asbestos fibers leading to lung damage and cancer. Antigorite is a mineral with asbestiform properties, which tends to associate with chrysotile or tremolite, and frequently occurs as the predominant mineral in the veins of several serpentinite rocks found abundantly in the Western Alps. Particles containing antigorite are more abundant in the breathing air of this region than those typically found in urban ambient air. Exposure of MeT-5A and J774 cells to fibrous antigorite at concentrations of 5–100 μg/ml for 72 hr induced dose-dependent cytotoxicity. Antigorite also stimulated the ROS production, induced the generation of nitrite and PGE2. MeT-5A cells were more sensitive to antigorite than J774 cells. The results of this study revealed that the fibrous antigorite stimulates cyclooxygenase and formation of hydroxyl and nitric oxide radicals. These changes represent early cellular responses to antigorite fibers, which lead to a host of pathological and neoplastic conditions because free radicals and PGE2 play important roles as mediators of tumor pathogenesis. Understanding the mechanisms of the cellular responses to antigorite and other asbestos particles should be helpful in designing rational prevention and treatment approaches. PMID:17431308

Acrolein increases 5-lipoxygenase expression in murine macrophages through activation of ERK pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Chae E.; Lee, Seung J.; Seo, Kyo W.

2010-05-15

Episodic exposure to acrolein-rich pollutants has been linked to acute myocardial infarction, and 5-lipoxygenase (5-LO) is involved in the production of matrix metalloproteinase-9 (MMP-9), which destabilizes atherosclerotic plaques. Thus, the present study determined the effect of acrolein on 5-LO/leukotriene B{sub 4} (LTB{sub 4}) production in murine macrophages. Stimulation of J774A.1 cells with acrolein led to increased LTB{sub 4} production in association with increased 5-LO expression. Acrolein-evoked 5-LO expression was blocked by pharmacological inhibition of the ERK pathway, but not by inhibitors for JNK and p38 MAPK pathways. In line with these results, acrolein exclusively increased the phosphorylation of ERK amongmore » these MAPK, suggesting a role for the ERK pathway in acrolein-induced 5-LO expression with subsequent production of LTB{sub 4}. Among the receptor tyrosine kinases including epidermal growth factor receptor (EGFR) and platelet derived growth factor receptor (PDGFR), acrolein-evoked ERK phosphorylation was attenuated by AG1478, an EGFR inhibitor, but not by AG1295, a PDGFR inhibitor. In addition, acrolein-evoked 5-LO expression was also inhibited by inhibition of EGFR pathway, but not by inhibition of PDGFR pathway. These observations suggest that acrolein has a profound effect on the 5-LO pathway via an EGFR-mediated activation of ERK pathway, leading to acute ischemic syndromes through the generation of LTB{sub 4}, subsequent MMP-9 production and plaque rupture.« less

Alveolar macrophage development in mice requires L-plastin for cellular localization in alveoli.

PubMed

Todd, Elizabeth M; Zhou, Julie Y; Szasz, Taylor P; Deady, Lauren E; D'Angelo, June A; Cheung, Matthew D; Kim, Alfred H J; Morley, Sharon Celeste

2016-12-15

Alveolar macrophages are lung-resident sentinel cells that develop perinatally and protect against pulmonary infection. Molecular mechanisms controlling alveolar macrophage generation have not been fully defined. Here, we show that the actin-bundling protein L-plastin (LPL) is required for the perinatal development of alveolar macrophages. Mice expressing a conditional allele of LPL (CD11c.Cre pos -LPL fl/fl ) exhibited significant reductions in alveolar macrophages and failed to effectively clear pulmonary pneumococcal infection, showing that immunodeficiency results from reduced alveolar macrophage numbers. We next identified the phase of alveolar macrophage development requiring LPL. In mice, fetal monocytes arrive in the lungs during a late fetal stage, maturing to alveolar macrophages through a prealveolar macrophage intermediate. LPL was required for the transition from prealveolar macrophages to mature alveolar macrophages. The transition from prealveolar macrophage to alveolar macrophage requires the upregulation of the transcription factor peroxisome proliferator-activated receptor-γ (PPAR-γ), which is induced by exposure to granulocyte-macrophage colony-stimulating factor (GM-CSF). Despite abundant lung GM-CSF and intact GM-CSF receptor signaling, PPAR-γ was not sufficiently upregulated in developing alveolar macrophages in LPL -/- pups, suggesting that precursor cells were not correctly localized to the alveoli, where GM-CSF is produced. We found that LPL supports 2 actin-based processes essential for correct localization of alveolar macrophage precursors: (1) transmigration into the alveoli, and (2) engraftment in the alveoli. We thus identify a molecular pathway governing neonatal alveolar macrophage development and show that genetic disruption of alveolar macrophage development results in immunodeficiency. © 2016 by The American Society of Hematology.

Capsaicin, a novel inhibitor of the NorA efflux pump, reduces the intracellular invasion of Staphylococcus aureus.

PubMed

Kalia, Nitin Pal; Mahajan, Priya; Mehra, Rukmankesh; Nargotra, Amit; Sharma, Jai Parkash; Koul, Surrinder; Khan, Inshad Ali

2012-10-01

To delineate the role of capsaicin (8-methyl-N-vanillyl-6-nonenamide) as an inhibitor of the NorA efflux pump and its impact on invasion of macrophages by Staphylococcus aureus. Capsaicin in combination with ciprofloxacin was tested for activity against S. aureus SA-1199B (NorA overproducing), SA-1199 (wild-type) and SA-K1758 (norA knockout). The role of NorA in the intracellular invasion of S. aureus and the ability of capsaicin to inhibit this invasion was established in J774 macrophage cell lines. The three-dimensional structure of NorA was predicted using an in silico approach and docking studies of capsaicin were performed. Capsaicin significantly reduced the MIC of ciprofloxacin for S. aureus SA-1199 and SA-1199B. Furthermore, capsaicin also extended the post-antibiotic effect of ciprofloxacin by 1.1 h at MIC concentration. There was a decrease in mutation prevention concentration of ciprofloxacin when combined with capsaicin. Inhibition of ethidium bromide efflux by NorA-overproducing S. aureus SA-1199B confirmed the role of capsaicin as a NorA efflux pump inhibitor (EPI). The most significant finding of this study was the ability of capsaicin to reduce the intracellular invasion of S. aureus SA-1199B (NorA overproducing) in J774 macrophage cell lines by 2 log(10). This study, for the first time, has shown that capsaicin, a novel EPI, not only inhibits the NorA efflux pump of S. aureus but also reduces the invasiveness of S. aureus, thereby reducing its virulence.

Brucella spp. of amphibians comprise genomically diverse motile strains competent for replication in macrophages and survival in mammalian hosts

PubMed Central

Al Dahouk, Sascha; Köhler, Stephan; Occhialini, Alessandra; Jiménez de Bagüés, María Pilar; Hammerl, Jens Andre; Eisenberg, Tobias; Vergnaud, Gilles; Cloeckaert, Axel; Zygmunt, Michel S.; Whatmore, Adrian M.; Melzer, Falk; Drees, Kevin P.; Foster, Jeffrey T.; Wattam, Alice R.; Scholz, Holger C.

2017-01-01

Twenty-one small Gram-negative motile coccobacilli were isolated from 15 systemically diseased African bullfrogs (Pyxicephalus edulis), and were initially identified as Ochrobactrum anthropi by standard microbiological identification systems. Phylogenetic reconstructions using combined molecular analyses and comparative whole genome analysis of the most diverse of the bullfrog strains verified affiliation with the genus Brucella and placed the isolates in a cluster containing B. inopinata and the other non-classical Brucella species but also revealed significant genetic differences within the group. Four representative but molecularly and phenotypically diverse strains were used for in vitro and in vivo infection experiments. All readily multiplied in macrophage-like murine J774-cells, and their overall intramacrophagic growth rate was comparable to that of B. inopinata BO1 and slightly higher than that of B. microti CCM 4915. In the BALB/c murine model of infection these strains replicated in both spleen and liver, but were less efficient than B. suis 1330. Some strains survived in the mammalian host for up to 12 weeks. The heterogeneity of these novel strains hampers a single species description but their phenotypic and genetic features suggest that they represent an evolutionary link between a soil-associated ancestor and the mammalian host-adapted pathogenic Brucella species. PMID:28300153

Direct recognition of superparamagnetic nanocrystals by macrophage scavenger receptor SR-AI.

PubMed

Chao, Ying; Karmali, Priya P; Mukthavaram, Rajesh; Kesari, Santosh; Kouznetsova, Valentina L; Tsigelny, Igor F; Simberg, Dmitri

2013-05-28

Scavenger receptors (SRs) are molecular pattern recognition receptors that have been shown to mediate opsonin-independent uptake of therapeutic and imaging nanoparticles, underlying the importance of SRs in nanomedicine. Unlike pathogens, engineered nanomaterials offer great flexibility in control of surface properties, allowing addressing specific questions regarding the molecular mechanisms of nanoparticle recognition. Recently, we showed that SR-type AI/II mediates opsonin-independent internalization of dextran superparamagnetic iron oxide (SPIO) nanoparticles via positively charged extracellular collagen-like domain. To understand the mechanism of opsonin-independent SPIO recognition, we tested the binding and uptake of nanoparticles with different surface coatings by SR-AI. SPIO coated with 10 kDa dextran was efficiently recognized and taken up by SR-AI transfected cells and J774 macrophages, while SPIO with 20 kDa dextran coating or cross-linked dextran hydrogel avoided the binding and uptake. Nanoparticle negative charge density and zeta-potential did not correlate with SR-AI binding/uptake efficiency. Additional experiments and computer modeling revealed that recognition of the iron oxide crystalline core by the positively charged collagen-like domain of SR-AI is sterically hindered by surface polymer coating. Importantly, the modeling revealed a strong complementarity between the surface Fe-OH groups of the magnetite crystal and the charged lysines of the collagen-like domain of SR-AI, suggesting a specific recognition of SPIO crystalline surface. These data provide an insight into the molecular recognition of nanocrystals by innate immunity receptors and the mechanisms whereby polymer coatings promote immune evasion.

Leishmania hijacking of the macrophage intracellular compartments.

PubMed

Liévin-Le Moal, Vanessa; Loiseau, Philippe M

2016-02-01

Leishmania spp., transmitted to humans by the bite of the sandfly vector, are responsible for the three major forms of leishmaniasis, cutaneous, diffuse mucocutaneous and visceral. Leishmania spp. interact with membrane receptors of neutrophils and macrophages. In macrophages, the parasite is internalized within a parasitophorous vacuole and engages in a particular intracellular lifestyle in which the flagellated, motile Leishmania promastigote metacyclic form differentiates into non-motile, metacyclic amastigote form. This phenomenon is induced by Leishmania-triggered events leading to the fusion of the parasitophorous vacuole with vesicular members of the host cell endocytic pathway including recycling endosomes, late endosomes and the endoplasmic reticulum. Maturation of the parasitophorous vacuole leads to the intracellular proliferation of the Leishmania amastigote forms by acquisition of host cell nutrients while escaping host defense responses. © 2015 FEBS.

Celastrol nanomicelles attenuate cytokine secretion in macrophages and inhibit macrophage-induced corneal neovascularization in rats.

PubMed

Li, Zhanrong; Li, Jingguo; Zhu, Lei; Zhang, Ying; Zhang, Junjie; Yao, Lin; Liang, Dan; Wang, Liya

The aim of the present study was to investigate the inhibitory effects of celastrol-loaded nanomicelles (CNMs) on activated macrophage-induced corneal neovascularization (CNV) in rats and cytokine secretion in macrophages. Using an angiogenesis assay in vitro, we detected the effects of CNMs on human umbilical vein endothelial cell (HUVEC) migration and invasion. In addition, the expression levels of cytokines secreted from hypoxia-induced macrophages were assessed through cytokine array analysis. The expression of hypoxia-inducible factors-1α (HIF-1α), nuclear factor-kappa B p65 (NF-κB p65), phospho-nuclear factor-kappa B p65 (phospho-NF-κB p65), p38 mitogen-activated protein kinase (p38 MAPK), phospho-p38 MAPK, extracellular signal-regulated kinase 1/2 (ERK1/2), and phospho-ERK1/2 was analyzed by western blotting. Activated macrophages were elicited through mineral oil lumbar injection, labeled with 1,19-dioctadecyl-3-3-39,39-tetramethylindocarbocyanine (DiI) and implanted into the corneal micro-pocket to induce CNV and to assess the antiangiogenic effect in rats. CNV was morphometrically analyzed using ImageJ software. Histopathological features were evaluated by immunofluorescence immunostaining for vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) on day 2 after surgery. In the present study, the results indicated that CNMs significantly inhibited the migration and invasion of HUVECs; remarkably attenuated the expression of VEGF, tumor necrosis factor-α, interleukin-1α, monocyte chemoattractant protein 1, cytokine-induced neutrophil chemoattractant 3, and MMP-9 protein; and downregulated ERK1/2, p38 MAPK, NF-κB activation, and HIF-1α expression in macrophages. The peritoneal cells elicited using mineral oil were highly purified macrophages, and the length and area of CNV were significantly decreased in the CNMs group compared with the control group. There was a significant reduction in the expression of VEGF and MMP-9 in

The Upregulation of Integrin αDβ2 (CD11d/CD18) on Inflammatory Macrophages Promotes Macrophage Retention in Vascular Lesions and Development of Atherosclerosis.

PubMed

Aziz, Moammir H; Cui, Kui; Das, Mitali; Brown, Kathleen E; Ardell, Christopher L; Febbraio, Maria; Pluskota, Elzbieta; Han, Juying; Wu, Huaizhu; Ballantyne, Christie M; Smith, Jonathan D; Cathcart, Martha K; Yakubenko, Valentin P

2017-06-15

Macrophage accumulation is a critical step during development of chronic inflammation, initiating progression of many devastating diseases. Leukocyte-specific integrin α D β 2 (CD11d/CD18) is dramatically upregulated on macrophages at inflammatory sites. Previously we found that CD11d overexpression on cell surfaces inhibits in vitro cell migration due to excessive adhesion. In this study, we have investigated how inflammation-mediated CD11d upregulation contributes to macrophage retention at inflammatory sites during atherogenesis. Atherosclerosis was evaluated in CD11d -/- /ApoE -/- mice after 16 wk on a Western diet. CD11d deficiency led to a marked reduction in lipid deposition in aortas and isolated macrophages. Macrophage numbers in aortic sinuses of CD11d -/- mice were reduced without affecting their apoptosis and proliferation. Adoptive transfer of fluorescently labeled wild-type and CD11d -/- monocytes into ApoE -/- mice demonstrated similar recruitment from circulation, but reduced accumulation of CD11d -/- macrophages within the aortas. Furthermore, CD11d expression was significantly upregulated on macrophages in atherosclerotic lesions and M1 macrophages in vitro. Interestingly, expression of the related ligand-sharing integrin CD11b was not altered. This difference defines their distinct roles in the regulation of macrophage migration. CD11d-deficient M1 macrophages demonstrated improved migration in a three-dimensional fibrin matrix and during resolution of peritoneal inflammation, whereas migration of CD11b -/- M1 macrophages was not affected. These results prove the contribution of high densities of CD11d to macrophage arrest during atherogenesis. Because high expression of CD11d was detected in several inflammation-dependent diseases, we suggest that CD11d/CD18 upregulation on proinflammatory macrophages may represent a common mechanism for macrophage retention at inflammatory sites, thereby promoting chronic inflammation and disease development

Immobilized heavy chain-hyaluronic acid polarizes lipopolysaccharide-activated macrophages toward M2 phenotype.

PubMed

He, Hua; Zhang, Suzhen; Tighe, Sean; Son, Ji; Tseng, Scheffer C G

2013-09-06

Despite the known anti-inflammatory effect of amniotic membrane, its action mechanism remains largely unknown. HC-HA complex (HC-HA) purified from human amniotic membrane consists of high molecular weight hyaluronic acid (HA) covalently linked to the heavy chain (HC) 1 of inter-α-trypsin inhibitor. In this study, we show that soluble HC-HA also contained pentraxin 3 and induced the apoptosis of both formyl-Met-Leu-Phe or LPS-activated neutrophils and LPS-activated macrophages while not affecting the resting cells. This enhanced apoptosis was caused by the inhibition of cell adhesion, spreading, and proliferation caused by HC-HA binding of LPS-activated macrophages and preventing adhesion to the plastic surface. Preferentially, soluble HC-HA promoted phagocytosis of apoptotic neutrophils in resting macrophages, whereas immobilized HC-HA promoted phagocytosis in LPS-activated macrophages. Upon concomitant LPS stimulation, immobilized HC-HA but not HA polarized macrophages toward the M2 phenotype by down-regulating IRF5 protein and preventing its nuclear localization and by down-regulating IL-12, TNF-α, and NO synthase 2. Additionally, IL-10, TGF-β1, peroxisome proliferator-activated receptor γ, LIGHT (TNF superfamily 14), and sphingosine kinase-1 were up-regulated, and such M2 polarization was dependent on TLR ligation. Collectively, these data suggest that HC-HA is a unique matrix component different from HA and uses multiple mechanisms to suppress M1 while promoting M2 phenotype. This anti-inflammatory action of HC-HA is highly desirable to promote wound healing in diseases heightened by unsuccessful transition from M1 to M2 phenotypes.

Immunoregulatory mechanisms of macrophage PPAR γ in mice with experimental inflammatory bowel disease

PubMed Central

Hontecillas, Raquel; Horne, William T.; Climent, Montse; Guri, Amir J.; Evans, C.; Zhang, Y.; Sobral, Bruno W.; Bassaganya-Riera, Josep

2010-01-01

Peroxisome proliferator-activated receptor γ (PPAR γ) is widely expressed in macrophages and has been identified as a putative target for the development of novel therapies against inflammatory bowel disease (IBD). Computational simulations identified macrophages as key targets for therapeutic interventions against IBD. This study aimed to characterize the mechanisms underlying the beneficial effects of macrophage PPAR γ in IBD. Macrophage-specific PPAR γ deletion significantly exacerbated clinical activity and colonic pathology, impaired the splenic and mesenteric lymph node regulatory T cell compartment, increased percentages of LP CD8+ T cells, increased surface expression of CD40, Ly6C, and TLR-4 in LP macrophages, and upregulated expression of colonic IFN-γ, CXCL9, CXCL10, IL-22, IL1RL1, CCR1, suppressor of cytokine signaling 3 and MCH class II in mice with IBD. Moreover, macrophage PPAR γ was required for accelerating pioglitazone-mediated recovery from DSS colitis, providing a cellular target for the anti-inflammatory effects of PPAR γ agonists in IBD. PMID:21068720

Immunoregulatory mechanisms of macrophage PPAR-γ in mice with experimental inflammatory bowel disease.

PubMed

Hontecillas, R; Horne, W T; Climent, M; Guri, A J; Evans, C; Zhang, Y; Sobral, B W; Bassaganya-Riera, J

2011-05-01

Peroxisome proliferator-activated receptor-γ (PPAR-γ) is widely expressed in macrophages and has been identified as a putative target for the development of novel therapies against inflammatory bowel disease (IBD). Computational simulations identified macrophages as key targets for therapeutic interventions against IBD. This study aimed to characterize the mechanisms underlying the beneficial effects of macrophage PPAR-γ in IBD. Macrophage-specific PPAR-γ deletion significantly exacerbated clinical activity and colonic pathology, impaired the splenic and mesenteric lymph node regulatory T-cell compartment, increased percentages of lamina propria (LP) CD8+ T cells, increased surface expression of CD40, Ly6C, and Toll-like receptor 4 (TLR-4) in LP macrophages, and upregulated expression of colonic IFN-γ, CXCL9, CXCL10, IL-22, IL1RL1, CCR1, suppressor of cytokine signaling 3, and MHC class II in mice with IBD. Moreover, macrophage PPAR-γ was required for accelerating pioglitazone-mediated recovery from dextran sodium sulfate (DSS) colitis, providing a cellular target for the anti-inflammatory effects of PPAR-γ agonists in IBD.

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

PubMed

Price, Christopher T D; Abu Kwaik, Yousef

2014-01-01

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

The DinJ/RelE Toxin-Antitoxin System Suppresses Bacterial Proliferation and Virulence of Xylella fastidiosa in Grapevine.

PubMed

Burbank, Lindsey P; Stenger, Drake C

2017-04-01

Xylella fastidiosa, the causal agent of Pierce's disease of grapes, is a slow-growing, xylem-limited, bacterial pathogen. Disease progression is characterized by systemic spread of the bacterium through xylem vessel networks, causing leaf-scorching symptoms, senescence, and vine decline. It appears to be advantageous to this pathogen to avoid excessive blockage of xylem vessels, because living bacterial cells are generally found in plant tissue with low bacterial cell density and minimal scorching symptoms. The DinJ/RelE toxin-antitoxin system is characterized here for a role in controlling bacterial proliferation and population size during plant colonization. The DinJ/RelE locus is transcribed from two separate promoters, allowing for coexpression of antitoxin DinJ with endoribonuclease toxin RelE, in addition to independent expression of RelE. The ratio of antitoxin/toxin expressed is dependent on bacterial growth conditions, with lower amounts of antitoxin present under conditions designed to mimic grapevine xylem sap. A knockout mutant of DinJ/RelE exhibits a hypervirulent phenotype, with higher bacterial populations and increased symptom development and plant decline. It is likely that DinJ/RelE acts to prevent excessive population growth, contributing to the ability of the pathogen to spread systemically without completely blocking the xylem vessels and increasing probability of acquisition by the insect vector.

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

PubMed

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

1993-01-01

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

Pharmacological Activation of Peroxisome Proliferator-Activated Receptor {Delta} Increases Sphingomyelin Synthase Activity in THP-1 Macrophage-Derived Foam Cell.

PubMed

Mou, Dongsheng; Yang, Hua; Qu, Changhua; Chen, Juan; Zhang, Chaogui

2016-08-01

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors, which mediate glucose and lipid homeostasis by regulating the expression of a large number of transcription factors. Sphingomyelin synthase (SMS) is a key enzyme in the synthesis of sphingomyelin (SM), and its expression and activity have been reported to be associated with atherosclerosis (AS). Although there have been many functional PPAR and SMS studies on atherosclerosis in recent years, few have investigated the correlation between the activation of PPARδ and the activity of SMS. In his study, macrophage-induced foam cells were utilized to model important pathological changes that occur in AS. The influence of PPARδ agonism by GW501516 on SMS and its product molecule SM were measured. Results indicated that the activation of PPARδ was correlated in a positive manner with the activity of SMS2, and the content of SM was dose dependently increased by GW501516. Together, this study represents the first to suggest that PPARδ activation may be a potential risk of AS through enhancing activity of SMS2.

In vitro biocorrosion of Ti-6Al-4V implant alloy by a mouse macrophage cell line.

PubMed

Lin, Hsin-Yi; Bumgardner, Joel D

2004-03-15

Corrosion of implant alloys releasing metal ions has the potential to cause adverse tissue reactions and implant failure. We hypothesized that macrophage cells and their released reactive chemical species (RCS) affect the alloy's corrosion properties. A custom cell culture corrosion box was used to evaluate how cell culture medium, macrophage cells and RCS altered the Ti-6Al-4V corrosion behaviors in 72 h and how corrosion products affected the cells. There was no difference in the charge transfer in the presence (75.2 +/- 17.7 mC) and absence (62.3 +/- 18.8 mC) of cells. The alloy had the lowest charge transfer (28.2 +/- 4.1 mC) and metal ion release (Ti < 10 ppb, V < 2 ppb) with activated cells (releasing RCS) compared with the other two conditions. This was attributed to an enhancement of the surface oxides by RCS. Metal ion release was very low (Ti < 20 ppb, V < 10 ppb) with nonactivated cells and did not change cell morphology, viability, and NO and ATP release compared with controls. However, IL-1beta released from the activated cells and the proliferation of nonactivated cells were greater on the alloy than the controls. In summary, macrophage cells and RCS reduced the corrosion of Ti-6Al-4V alloys as hypothesized. These data are important in understanding host tissue-material interactions. Copyright 2004 Wiley Periodicals, Inc. J Biomed Mater Res 68A: 717-724, 2004

Cavin1; a Regulator of Lung Function and Macrophage Phenotype

PubMed Central

Govender, Praveen; Romero, Freddy; Shah, Dilip; Paez, Jesus; Ding, Shi-Ying; Liu, Libin; Gower, Adam; Baez, Elizabeth; Aly, Sherif Shawky; Pilch, Paul; Summer, Ross

2013-01-01

Caveolae are cell membrane invaginations that are highly abundant in adipose tissue, endothelial cells and the lung. The formation of caveolae is dependent on the expression of various structural proteins that serve as scaffolding for these membrane invaginations. Cavin1 is a newly identified structural protein whose deficiency in mice leads to loss of caveolae formation and to development of a lipodystrophic phenotype. In this study, we sought to investigate the functional role of Cavin1 in the lung. Cavin1 deficient mice possessed dramatically altered distal lung morphology and exhibited significant physiological alterations, notably, increased lung elastance. The changes in distal lung architecture were associated with hypercellularity and the accumulation of lung macrophages. The increases in lung macrophages occurred without changes to circulating numbers of mononuclear cells and without evidence for increased proliferation. However, the increases in lung macrophages were associated with higher levels of macrophage chemotactic factors CXCL2 and CCL2 in BAL fluid from Cavin1−/− mice suggesting a possible mechanism by which these cells accumulate. In addition, lung macrophages from Cavin1−/− mice were larger and displayed measurable differences in gene expression when compared to macrophages from wild-type mice. Interestingly, macrophages were also increased in adipose tissue but not in liver, kidney or skeletal muscle from Cavin1−/− mice, and similar tissue specificity for macrophage accumulation was observed in lungs and adipose tissue from Caveolin1−/− mice. In conclusion, this study demonstrates an important role for Cavin1 in lung homeostasis and suggests that caveolae structural proteins are necessary for regulating macrophage number and phenotype in the lung. PMID:23634221

Mycoplasma fermentans-derived high-molecular-weight material induces interleukin-6 release in cultures of murine macrophages and human monocytes.

PubMed Central

Quentmeier, H; Schmitt, E; Kirchhoff, H; Grote, W; Mühlradt, P F

1990-01-01

A Mycoplasma fermentans-derived high-molecular-weight material (MDHM) is described which causes differentiation of concanavalin A-stimulated CBA/J or C57BL/6 mouse thymocytes to cytolytic effector T cells (CTLs). The effect of MDHM was inhibited by addition of monoclonal anti-interleukin-6 (IL-6) antibody. It could also be abolished after removal of adherent cells. However, adherent cell-depleted thymocytes could still form CTLs after addition of IL-6. The action of MDHM could thus be explained by the capacity of MDHM to stimulate IL-6 release from adherent cells. MDHM was active on macrophages from CBA/J and C3H/HeJ endotoxin nonresponder mice and was also capable of stimulating IL-6 release from human monocytes. On gel chromatography, MDHM had an apparent molecular size of 1.5 x 10(6) daltons. Treatment with RNase and DNase had no effect on either size or biological activity. Proteinase K did not abolish activity but reduced the apparent molecular size of MDHM. MDHM production by M. fermentans required either coculture with eucaryotic cell lines in RPMI 1640 medium with fetal calf serum or addition of eucaryotic cell sonic extracts to this medium. The biological activity of MDHM is not identical to that of a mitogen for murine spleen cells derived from M. arthritidis; MDHM caused only slight proliferation in this system compared with the mitogen from M. arthritidis, and the latter did not elicit IL-6 release from macrophages. The results are discussed in relation to mycoplasmas as putative etiological agents for rheumatoid arthritis, since high IL-6 titers were reported for synovial fluid from patients with this disease. PMID:2323816

7 CFR 774.18 - Interest rate, terms and security requirements.

Code of Federal Regulations, 2014 CFR

2014-01-01

..., DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.18 Interest rate... percent for 36 months or until the date of settlement of, completion of, or final distribution of assets... bankruptcy claim or 36 months from the date of the note. (2) However, any principal remaining thereafter will...

7 CFR 774.18 - Interest rate, terms and security requirements.

Code of Federal Regulations, 2011 CFR

2011-01-01

..., DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.18 Interest rate... percent for 36 months or until the date of settlement of, completion of, or final distribution of assets... bankruptcy claim or 36 months from the date of the note. (2) However, any principal remaining thereafter will...

7 CFR 774.18 - Interest rate, terms and security requirements.

Code of Federal Regulations, 2013 CFR

2013-01-01

..., DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.18 Interest rate... percent for 36 months or until the date of settlement of, completion of, or final distribution of assets... bankruptcy claim or 36 months from the date of the note. (2) However, any principal remaining thereafter will...

7 CFR 774.18 - Interest rate, terms and security requirements.

Code of Federal Regulations, 2012 CFR

2012-01-01

..., DEPARTMENT OF AGRICULTURE SPECIAL PROGRAMS EMERGENCY LOAN FOR SEED PRODUCERS PROGRAM § 774.18 Interest rate... percent for 36 months or until the date of settlement of, completion of, or final distribution of assets... bankruptcy claim or 36 months from the date of the note. (2) However, any principal remaining thereafter will...

SAA drives proinflammatory heterotypic macrophage differentiation in the lung via CSF-1R-dependent signaling

PubMed Central

Anthony, Desiree; McQualter, Jonathan L.; Bishara, Maria; Lim, Ee X.; Yatmaz, Selcuk; Seow, Huei Jiunn; Hansen, Michelle; Thompson, Michelle; Hamilton, John A.; Irving, Louis B.; Levy, Bruce D.; Vlahos, Ross; Anderson, Gary P.; Bozinovski, Steven

2014-01-01

Serum amyloid A (SAA) is expressed locally in chronic inflammatory conditions such as chronic obstructive pulmonary disease (COPD), where macrophages that do not accord with the classic M1/M2 paradigm also accumulate. In this study, the role of SAA in regulating macrophage differentiation was investigated in vitro using human blood monocytes from healthy subjects and patients with COPD and in vivo using an airway SAA challenge model in BALB/c mice. Differentiation of human monocytes with SAA stimulated the proinflammatory monokines IL-6 and IL-1β concurrently with the M2 markers CD163 and IL-10. Furthermore, SAA-differentiated macrophages stimulated with lipopolysaccharide (LPS) expressed markedly higher levels of IL-6 and IL-1β. The ALX/FPR2 antagonist WRW4 reduced IL-6 and IL-1β expression but did not significantly inhibit phagocytic and efferocytic activity. In vivo, SAA administration induced the development of a CD11chighCD11bhigh macrophage population that generated higher levels of IL-6, IL-1β, and G-CSF following ex vivo LPS challenge. Blocking CSF-1R signaling effectively reduced the number of CD11chighCD11bhigh macrophages by 71% and also markedly inhibited neutrophilic inflammation by 80%. In conclusion, our findings suggest that SAA can promote a distinct CD11chighCD11bhigh macrophage phenotype, and targeting this population may provide a novel approach to treating chronic inflammatory conditions associated with persistent SAA expression.—Anthony, D., McQualter, J. L., Bishara, M., Lim, E. X., Yatmaz, S., Seow, H. J., Hansen, M., Thompson, M., Hamilton, J. A., Irving, L. B., Levy, B. D., Vlahos, R., Anderson, G. P., Bozinovski, S. SAA drives proinflammatory heterotypic macrophage differentiation in the lung via CSF-1R-dependent signaling. PMID:24846388

Differential Kinetics of Aspergillus nidulans and Aspergillus fumigatus Phagocytosis.

PubMed

Gresnigt, Mark S; Becker, Katharina L; Leenders, Floris; Alonso, M Fernanda; Wang, Xiaowen; Meis, Jacques F; Bain, Judith M; Erwig, Lars P; van de Veerdonk, Frank L

2018-01-01

Invasive aspergillosis mainly occurs in immunocompromised patients and is commonly caused by Aspergillus fumigatus, while A.nidulans is rarely the causative agent. However, in chronic granulomatous disease (CGD) patients, A. nidulans is a frequent cause of invasive aspergillosis and is associated with higher mortality. Immune recognition of A. nidulans was compared to A. fumigatus to offer an insight into why A. nidulans infections are prevalent in CGD. Live cell imaging with J774A.1 macrophage-like cells and LC3-GFP-mCherry bone marrow-derived macrophages (BMDMs) revealed that phagocytosis of A. nidulans was slower compared to A. fumigatus. This difference could be attributed to slower migration of J774A.1 cells and a lower percentage of migrating BMDMs. In addition, delayed phagosome acidification and LC3-associated phagocytosis was observed with A. nidulans. Cytokine and oxidative burst measurements in human peripheral blood mononuclear cells revealed a lower oxidative burst upon challenge with A. nidulans. In contrast, A. nidulans induced significantly higher concentrations of cytokines. Collectively, our data demonstrate that A. nidulans is phagocytosed and processed at a slower rate compared to A. fumigatus, resulting in reduced fungal killing and increased germination of conidia. This slower rate of A. nidulans clearance may be permissive for overgrowth within certain immune settings. The Author(s). Published by S. Karger AG, Basel.

In vitro studies on the effect of particle size on macrophage responses to nanodiamond wear debris

PubMed Central

Thomas, Vinoy; Halloran, Brian A.; Ambalavanan, Namasivayam; Catledge, Shane A.; Vohra, Yogesh K.

2012-01-01

Nanostructured diamond coatings improve the smoothness and wear characteristics of the metallic component of total hip replacements and increase the longevity of these implants, but the effect of nanodiamond wear debris on macrophages needs to be determined to estimate the long-term inflammatory effects of wear debris. The objective was to investigate the effect of the size of synthetic nanodiamond particles on macrophage proliferation (BrdU incorporation), apoptosis (Annexin-V flow cytometry), metabolic activity (WST-1 assay) and inflammatory cytokine production (qPCR). RAW 264.7 macrophages were exposed to varying sizes (6, 60, 100, 250 and 500 nm) and concentrations (0, 10, 50, 100 and 200 μg ml−1) of synthetic nanodiamonds. We observed that cell proliferation but not metabolic activity was decreased with nanoparticle sizes of 6–100 nm at lower concentrations (50 μg ml−1), and both cell proliferation and metabolic activity were significantly reduced with nanodiamond concentrations of 200 μg ml−1. Flow cytometry indicated a significant reduction in cell viability due to necrosis irrespective of particle size. Nanodiamond exposure significantly reduced gene expression of tumor necrosis factor-α, interleukin-1β, chemokine Ccl2 and platelet-derived growth factor compared to serum-only controls or titanium oxide (anatase 8 nm) nanoparticles, with variable effects on chemokine Cxcl2 and vascular endothelial growth factor. In general, our study demonstrates a size and concentration dependence of macrophage responses in vitro to nanodiamond particles as possible wear debris from diamond-coated orthopedic joint implants. PMID:22342422

In vitro biocorrosion of Co-Cr-Mo implant alloy by macrophage cells.

PubMed

Lin, Hsin-Yi; Bumgardner, Joel D

2004-11-01

We hypothesized that macrophage cells and their released reactive chemical species (RCS) affect Co-Cr-Mo alloy's corrosion properties and that alloy corrosion products change macrophage cell behavior. A custom cell culture corrosion cell was used to evaluate how culture medium, cells, and RCS altered alloy corrosion in 3-day tests. Corrosion was evaluated by measuring total charge transfer at a constant potential using a potentiostat and metal ion release by atomic emission spectroscopy. Viability, proliferation, and NO (nitric oxide) and IL-1beta (interlukin-1beta) release were used to assess cellular response to alloy corrosion products. In the presence of activated cells, total charge transfers and Co ion release were the lowest (p < 0.05). This was attributed to an enhancement of the surface oxide by RCS. Cr and Mo release were not different between cells and activated cells. Low levels of metal ions did not affect cell viability, proliferation, or NO release, though IL-1beta released from the activated cells was higher on the alloy compared to the controls. These data support the hypothesis that macrophage cells and their RCS affect alloy corrosion. Changes in alloy corrosion by cells may be important to the development of host responses to the alloy and its corrosion products.

Synergy of anti-CD40, CpG and MPL in activation of mouse macrophages

PubMed Central

Shi, Yongyu; Felder, Mildred A.R.; Sondel, Paul M.; Rakhmilevich, Alexander L.

2015-01-01

Activation of macrophages is a prerequisite for their antitumor effects. Several reagents, including agonistic anti-CD40 monoclonal antibody (anti-CD40), CpG oligodeoxynucleotides (CpG) and monophosphoryl lipid A (MPL), can stimulate activation of macrophages. Our previous studies showed synergy between anti-CD40 and CpG and between anti-CD40 and MPL in macrophage activation and antitumor efficacy in mice. In the present study, we asked whether there was synergy among these three reagents. The activation of adherent peritoneal exudate cells (PEC) obtained from mice injected with anti-CD40 and then treated with CpG and/or MPL in vitro was determined by their ability to suppress proliferation of tumor cells and to produce various cytokines and chemokines in vitro. Cell sorting and histology followed by functional testing showed that macrophages were the main cell population in PEC activated by CD40 ligation in vivo. A combination of anti-CD40, CpG or MPL activated PEC to suppress proliferation of B16 cells and produce nitric oxide far greater than the single reagents or any of the double combinations of these reagents. In addition, the combination of all three reagents activated PEC to secrete IL-12, IFN-γ and MCP-1 to a greater degree than any single reagent or any two combined reagents. These results demonstrate that macrophages can be synergistically activated by anti-CD40, CpG and MPL, suggesting that this novel combined approach might be further investigated as potential cancer therapy. PMID:25829245

Synergy of anti-CD40, CpG and MPL in activation of mouse macrophages.

PubMed

Shi, Yongyu; Felder, Mildred A R; Sondel, Paul M; Rakhmilevich, Alexander L

2015-08-01

Activation of macrophages is a prerequisite for their antitumor effects. Several reagents, including agonistic anti-CD40 monoclonal antibody (anti-CD40), CpG oligodeoxynucleotides (CpG) and monophosphoryl lipid A (MPL), can stimulate activation of macrophages. Our previous studies showed synergy between anti-CD40 and CpG and between anti-CD40 and MPL in macrophage activation and antitumor efficacy in mice. In the present study, we asked whether there was synergy among these three reagents. The activation of adherent peritoneal exudate cells (PEC) obtained from mice injected with anti-CD40 and then treated with CpG and/or MPL in vitro was determined by their ability to suppress proliferation of tumor cells and to produce various cytokines and chemokines in vitro. Cell sorting and histology followed by functional testing showed that macrophages were the main cell population in PEC activated by CD40 ligation in vivo. A combination of anti-CD40, CpG or MPL activated PEC to suppress proliferation of B16 cells and produce nitric oxide far greater than the single reagents or any of the double combinations of these reagents. In addition, the combination of all three reagents activated PEC to secrete IL-12, IFN-γ and MCP-1 to a greater degree than any single reagent or any two combined reagents. These results demonstrate that macrophages can be synergistically activated by anti-CD40, CpG and MPL, suggesting that this novel combined approach might be further investigated as potential cancer therapy. Copyright © 2015 Elsevier Ltd. All rights reserved.

The Macrophage in the Development of Experimental Crescentic Glomerulonephritis

PubMed Central

Thomson, Napier M.; Holdsworth, Stephen R.; Glasgow, Eric F.; Atkins, Robert C.

1979-01-01

The role played by the macrophage in the development of injury in rabbit nephrotoxic nephritis (NTN) has been assessed by electron microscopy and glomerular culture of renal tissue obtained by several biopsies during the course of the disease. These observations have been correlated with the other immune, cellular, and biochemical events occurring in the glomerulus, ie, deposition of immunoglobulin and complement, proteinuria, polymorphonuclear leukocyte (PMN) exudation, fibrin deposition, crescent formation, and renal failure. A biphasic macrophage accumulation was detected, corresponding to the heterologous and autologous phases of the disease. In the autologous or crescentic phase, macrophages accumulated within the glomerular tuft from Day 5; their appearance coincided with the accumulation of PMN and development of proteinuria. Fibrin deposition in Bowman's space, which commenced on Days 6 and 7, was rapidly followed by the migration of macrophages from the glomeruli into Bowman's space. Within Bowman's space, macrophages were observed to phagocytose fibrin, transform into epithelioid and giant cells, and accumulate to form a substantial proportion of the cells forming the crescent. The inflammatory process of PMN exudation, macrophage accumulation, fibrin deposition, and crescent formation and the degree of renal failure reached a maximum by Days 12 to 14. Thereafter, resolution of the inflammatory process occurred so that by Day 40 macrophages had disappeared from the glomeruli. However, varying degrees of glomerular damage and renal failure persisted, occurring largely as a result of glomerulosclerosis and sclerosis of crescents. The tissue culture studies also demonstrated mesangial cell proliferation during the inflammatory process but did not show any abnormality of epithelial cell activity. This study demonstrates that the macrophages participate in NTN by accumulating in damaged glomeruli then migrating into Bowman's space (probably in response to

Rictor/mammalian target of rapamycin complex 2 promotes macrophage activation and kidney fibrosis.

PubMed

Ren, Jiafa; Li, Jianzhong; Feng, Ye; Shu, Bingyan; Gui, Yuan; Wei, Wei; He, Weichun; Yang, Junwei; Dai, Chunsun

2017-08-01

Mammalian target of rapamycin (mTOR) signalling controls many essential cellular functions. However, the role of Rictor/mTOR complex 2 (mTORC2) in regulating macrophage activation and kidney fibrosis remains largely unknown. We report here that Rictor/mTORC2 was activated in macrophages from the fibrotic kidneys of mice. Ablation of Rictor in macrophages reduced kidney fibrosis, inflammatory cell accumulation, macrophage proliferation and polarization after unilateral ureter obstruction or ischaemia/reperfusion injury. In bone marrow-derived macrophages (BMMs), deletion of Rictor or blockade of protein kinase Cα inhibited cell migration. Additionally, deletion of Rictor or blockade of Akt abolished interleukin-4-stimulated or transforming growth factor (TGF)-β1-stimulated macrophage M2 polarization. Furthermore, deletion of Rictor downregulated TGF-β1-stimulated upregulation of multiple profibrotic cytokines, including platelet-derived growth factor, vascular endothelial growth factor and connective tissue growth factor, in BMMs. Conditioned medium from TGF-β1-pretreated Rictor -/- macrophages stimulated fibroblast activation less efficiently than that from TGF-β1-pretreated Rictor +/+ macrophages. These results demonstrate that Rictor/mTORC2 signalling can promote macrophage activation and kidney fibrosis. Targeting this signalling pathway in macrophages may shine light on ways to protect against kidney fibrosis in patients with chronic kidney diseases. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Immunomodulatory effects of an acetylated Cyclocarya paliurus polysaccharide on murine macrophages RAW264.7.

PubMed

Liu, Xin; Xie, Jianhua; Jia, Shuo; Huang, Lixin; Wang, Zhijun; Li, Chang; Xie, Mingyong

2017-05-01

Polysaccharides (CP) extracted from the leaves of Cyclocarya paliurus (C. paliurus) have been shown to possess a variety of biological activities. In present study, CP was successfully modified to obtain its acetylated derivative Ac-CP. Its potential immunomodulatory activities on RAW264.7 macrophages were investigated. Results showed that the acetylated polysaccharide Ac-CP could significantly stimulate macrophage proliferation, its actions were significantly stronger than that of the corresponding unmodified polysaccharide, CP. Meanwhile, the NO production activities of macrophages were not significantly enhanced by Ac-CP compared to CP group. In addition, both the phagocytic activity and levels of cytokines TNF-a, IL-1β and IL-6 were enhanced in the RAW264.7 macrophages by stimulation of Ac-CP. These results indicated that the acetylated derivative Ac-CP could enhance the activation of peritoneal macrophages, and acetylation modification can enhance the immunomodulation function of CP, indicating the potential application of acetylated polysaccharide as an immunotherapeutic adjuvant. Copyright © 2017 Elsevier B.V. All rights reserved.

Effects of glutamine, taurine and their association on inflammatory pathway markers in macrophages.

PubMed

Sartori, Talita; Galvão Dos Santos, Guilherme; Nogueira-Pedro, Amanda; Makiyama, Edson; Rogero, Marcelo Macedo; Borelli, Primavera; Fock, Ricardo Ambrósio

2018-06-01

The immune system is essential for the control and elimination of infections, and macrophages are cells that act as important players in orchestrating the various parts of the inflammatory/immune response. Amino acids play important role in mediating functionality of the inflammatory response, especially mediating macrophages functions and cytokines production. We investigated the influence of glutamine, taurine and their association on the modulation of inflammatory pathway markers in macrophages. The RAW 264.7 macrophage cell line was cultivated in the presence of glutamine and taurine and proliferation rates, cell viability, cell cycle phases, IL-1α, IL-6, IL-10 and TNF-α as well as H 2 O 2 production and the expression of the transcription factor, NFκB, and its inhibitor, IκBα, were evaluated. Our results showed an increase in viable cells and increased proliferation rates of cells treated with glutamine concentrations over 2 mM, as well as cells treated with both glutamine and taurine. The cell cycle showed a higher percentage of cells in the phases S, G2 and M when they were treated with 2 or 10 mM glutamine, or with glutamine and taurine in cells stimulated with lipopolysaccharide. The pNFκB/NFκB showed reduced ratio expression when cells were treated with 10 mM of glutamine or with glutamine in association with taurine. These conditions also resulted in reduced TNF-α, IL-1α and H 2 O 2 production, and higher production of IL-10. These findings demonstrate that glutamine and taurine are able to modulate macrophages inflammatory pathways, and that taurine can potentiate the effects of glutamine, illustrating their immunomodulatory properties.

Long non-coding RNA cox-2 prevents immune evasion and metastasis of hepatocellular carcinoma by altering M1/M2 macrophage polarization.

PubMed

Ye, Yibiao; Xu, Yunxiuxiu; Lai, Yu; He, Wenguang; Li, Yanshan; Wang, Ruomei; Luo, Xinxi; Chen, Rufu; Chen, Tao

2018-03-01

Macrophages have been shown to demonstrate a high level of plasticity, with the ability to undergo dynamic transition between M1 and M2 polarized phenotypes. We investigate long non-coding RNA (lncRNA) cox-2 in macrophage polarization and the regulatory mechanism functions in hepatocellular carcinoma (HCC). Lipopolysaccharide (LPS) was used to induce RAW264.7 macrophages into M1 type, and IL-4 was to induce RAW264.7 macrophages into M2 type. We selected mouse hepatic cell line Hepal-6 and hepatoma cell line HepG2 for co-incubation with M1 or M2 macrophages. Quantitative real-time PCR was used to detect the expressions of lncRNA cox-2 and mRNAs. ELISA was conducted for testing IL-12 and IL-10 expressions; Western blotting for epithelial mesenchymal transition related factors (E-cadherin and Vimentin). An MTT, colony formation assay, flow cytometry, transwell assay, and stretch test were conducted to test cell abilities. The M1 macrophages had higher lncRNA cox-2 expression than that in the non-polarized macrophages and M2 macrophages. The lncRNA cox-2 siRNA decreased the expression levels of IL-12, iNOS, and TNF-α in M1 macrophages, increased the expression levels of IL-10, Arg-1, and Fizz-1 in M2 macrophages (all P < 0.05). The lncRNA cox-2 siRNA reduces the ability of M1 macrophages to inhibit HCC cell proliferation, invasion, migration, EMT, angiogenesis and facilitate apoptosis while strengthening the ability of M2 macrophages to promote proliferation HCC cell growth and inhibit apoptosis. These findings indicate that lncRNA cox-2 inhibits HCC immune evasion and tumor growth by inhibiting the polarization of M2 macrophages. © 2017 Wiley Periodicals, Inc.

Hepatocyte-specific PPARA expression exclusively promotes agonist-induced cell proliferation without influence from nonparenchymal cells

PubMed Central

Brocker, Chad N.; Yue, Jiang; Kim, Donghwan; Qu, Aijuan; Bonzo, Jessica A.

2017-01-01

Peroxisome proliferator-activated receptor-α (PPARA) is a nuclear transcription factor and key mediator of systemic lipid metabolism. Prolonged activation in rodents causes hepatocyte proliferation and hepatocellular carcinoma. Little is known about the contribution of nonparenchymal cells (NPCs) to PPARA-mediated cell proliferation. NPC contribution to PPARA agonist-induced hepatomegaly was assessed in hepatocyte (Ppara△Hep)- and macrophage (Ppara△Mac)-specific Ppara null mice. Mice were treated with the agonist Wy-14643 for 14 days, and response of conditional null mice was compared with conventional knockout mice (Ppara−/−). Wy-14643 treatment caused weight loss and severe hepatomegaly in wild-type and Ppara△Mac mice, and histological analysis revealed characteristic hepatocyte swelling; Ppara△Hep and Ppara−/− mice were protected from these effects. Ppara△Mac serum chemistries, as well as aspartate aminotransferase and alanine aminotransferase levels, matched wild-type mice. Agonist-treated Ppara△Hep mice had elevated serum cholesterol, phospholipids, and triglycerides when compared with Ppara−/− mice, indicating a possible role for extrahepatic PPARA in regulating circulating lipid levels. BrdU labeling confirmed increased cell proliferation only in wild-type and Ppara△Mac mice. Macrophage PPARA disruption did not impact agonist-induced upregulation of lipid metabolism, cell proliferation, or DNA damage and repair-related gene expression, whereas gene expression was repressed in Ppara△Hep mice. Interestingly, downregulation of inflammatory cytokines IL-15 and IL-18 was dependent on macrophage PPARA. Cell type-specific regulation of target genes was confirmed in primary hepatocytes and Kupffer cells. These studies conclusively show that cell proliferation is mediated exclusively by PPARA activation in hepatocytes and that Kupffer cell PPARA has an important role in mediating the anti-inflammatory effects of PPARA agonists. PMID

Brazilian propolis ethanol extract and its component kaempferol induce myeloid-derived suppressor cells from macrophages of mice in vivo and in vitro.

PubMed

Kitamura, Hiroshi; Saito, Natsuko; Fujimoto, Junpei; Nakashima, Ken-Ichi; Fujikura, Daisuke

2018-05-02

Brazilian green propolis is produced by mixing secretions from Africanized honey bees with exudate, mainly from Baccharis dracunculifolia. Brazilian propolis is especially rich in flavonoids and cinammic acid derivatives, and it has been widely used in folk medicine owing to its anti-inflammatory, anti-viral, tumoricidal, and analgesic effects. Moreover, it is applied to prevent metabolic disorders, such as type 2 diabetes and arteriosclerosis. Previously, we demonstrated that propolis ethanol extract ameliorated type 2 diabetes in a mouse model through the resolution of adipose tissue inflammation. The aims of this study were to identify the immunosuppressive cells directly elicited by propolis extract and to evaluate the flavonoids that induce such cells. Ethanol extract of Brazilian propolis (PEE; 100 mg/kg i.p., twice a week) was injected into lean or high fat-fed obese C57BL/6 mice or C57BL/6 ob/ob mice for one month. Subsequently, immune cells in visceral adipose tissue and the peritoneal cavity were monitored using FACS analysis. Isolated macrophages and the macrophage-like cell line J774.1 were treated with PEE and its constituent components, and the expression of immune suppressive myeloid markers were evaluated. Finally, we injected one of the identified compounds, kaempferol, into C57BL/6 mice and performed FACS analysis on the adipose tissue. Intraperitoneal treatment of PEE induces CD11b + , Gr-1 + myeloid-derived suppressor cells (MDSCs) in visceral adipose tissue and the peritoneal cavity of lean and obese mice. PEE directly stimulates cultured M1 macrophages to transdifferentiate into MDSCs. Among twelve compounds isolated from PEE, kaempferol has an exclusive effect on MDSCs induction in vitro. Accordingly, intraperitoneal injection of kaempferol causes accumulation of MDSCs in the visceral adipose tissue of mice. Brazilian PEE and its compound kaempferol strongly induce MDSCs in visceral adipose tissue at a relatively early phase of inflammation

Report: Over $774 Million of Puerto Rico State Revolving Funds at Risk

EPA Pesticide Factsheets

Report #17-P-0186, April 26, 2017. Over $774 million of state revolving funds is at risk due to Puerto Rico’s ongoing financial crisis. Restoration of the revolving funds in the near future is highly unlikely.

Evidence for a role of macrophage migration inhibitory factor in vascular disease.

PubMed

Chen, Zhiping; Sakuma, Masashi; Zago, Alexandre C; Zhang, Xiaobin; Shi, Can; Leng, Lin; Mizue, Yuka; Bucala, Richard; Simon, Daniel

2004-04-01

Inflammation plays an essential role in atherosclerosis and restenosis. Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that is widely expressed in vascular cells. However, there is no in vivo evidence that MIF participates directly in vascular injury and repair. Therefore, we investigated the effect of MIF blockade on the response to experimental angioplasty in atherosclerosis-susceptible mice. Carotid artery dilation (2.5 atm) and complete endothelial denudation were performed in male C57BL/6J LDL receptor-deficient mice treated with a neutralizing anti-MIF or isotype control monoclonal antibody. After 7 days and 28 days, intimal and medial sizes were measured and intima/media area ratio (I/M) was calculated. Intimal thickening and I/M were reduced significantly by anti-MIF compared with control antibody. Vascular injury was accompanied by progressive vessel enlargement or "positive remodeling" that was comparable in both treatment groups. MIF blockade was associated with reduced inflammation and cellular proliferation and increased apoptosis after injury. Neutralizing MIF bioactivity after experimental angioplasty in atherosclerosis-susceptible mice reduces vascular inflammation, cellular proliferation, and neointimal thickening. Although the molecular mechanisms responsible for these effects are not yet established, these data prompt further research directed at understanding the role of MIF in vascular disease and suggest novel therapeutic interventions for preventing atherosclerosis and restenosis.

Broad NE 8 lambda 774 emission from quasars in the HST-FOS snapshot survey (ABSNAP)

NASA Technical Reports Server (NTRS)

Hamann, Fred; Zuo, Lin; Tytler, David

1995-01-01

We discuss the strength and frequency of broad Ne VIII lambda 774 emission from quasars measured in the Hubble Space Telescope Faint Object Spectrograph (HST-FOS) snapshot survey (Absnap). Five sources in the survey have suitable redshifts (0.86 less than or equal to Z(sub em) less than or equal to 1.31), signal-to-noise ratios and no Lyman limit absorptions. Three of the five sources have a strong broad emission line near 774 A (rest), and the remaining two sources have a less securely measured line near this wavelength. We identify these lines with Ne VIII lambda 774 based on the measured wavelengths and theoretical estimates of various line fluxes (Hamann et al. 1995a). Secure Ne VIII detections occur in both radio-loud and radio-quiet sources. We tentatively conclude that broad Ne VIII lambda 774 emission is common in quasars, with typical strengths between approximately 25% and approximately 200% of O VI lambda 1034. These Ne VIII lambda 774 measurements imply that the broad emission line regions have a much hotter and more highly ionized component than previously recognized. They also suggest that quasar continua have substantial ionizing flux out to energies greater than 207 eV (greater than 15.2 ryd, lambda less than 60 A). Photoionization calculations using standard incident spectra indicate that the Ne VIII emission requires ionization parameters U greater than or = 5, total column densities N(sub H) greater than or = 10(sub 22)/sq cm and covering factors greater than or = 25%. The temperatures could be as high as approximately 10(exp 5) K. If the gas is instead collisionally ionized, strong Ne VIII would imply equilibrium temperatures in the range approximately 400,000 less than or approximately = T(sub e) less than or approximately = 10(exp 6) K. In either case, the highly ionized Ne VIII emission regions would appear as X-ray 'warm absorbers' if they lie along our line of sight to the X-ray continuum source.

Intracellular growth of Mycobacterium tuberculosis after macrophage cell death leads to serial killing of host cells

PubMed Central

Mahamed, Deeqa; Boulle, Mikael; Ganga, Yashica; Mc Arthur, Chanelle; Skroch, Steven; Oom, Lance; Catinas, Oana; Pillay, Kelly; Naicker, Myshnee; Rampersad, Sanisha; Mathonsi, Colisile; Hunter, Jessica; Wong, Emily B; Suleman, Moosa; Sreejit, Gopalkrishna; Pym, Alexander S; Lustig, Gila; Sigal, Alex

2017-01-01

A hallmark of pulmonary tuberculosis is the formation of macrophage-rich granulomas. These may restrict Mycobacterium tuberculosis (Mtb) growth, or progress to central necrosis and cavitation, facilitating pathogen growth. To determine factors leading to Mtb proliferation and host cell death, we used live cell imaging to track Mtb infection outcomes in individual primary human macrophages. Internalization of Mtb aggregates caused macrophage death, and phagocytosis of large aggregates was more cytotoxic than multiple small aggregates containing similar numbers of bacilli. Macrophage death did not result in clearance of Mtb. Rather, it led to accelerated intracellular Mtb growth regardless of prior activation or macrophage type. In contrast, bacillary replication was controlled in live phagocytes. Mtb grew as a clump in dead cells, and macrophages which internalized dead infected cells were very likely to die themselves, leading to a cell death cascade. This demonstrates how pathogen virulence can be achieved through numbers and aggregation states. DOI: http://dx.doi.org/10.7554/eLife.22028.001 PMID:28130921

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

PubMed Central

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

2016-01-01

Differentiated macrophages can self-renew in tissues and expand long-term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network controlling self-renewal. Single cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells. PMID:26797145

30 CFR 774.17 - Transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 30 Mineral Resources 3 2010-07-01 2010-07-01 false Transfer, assignment, or sale of permit rights... SYSTEMS UNDER REGULATORY PROGRAMS REVISION; RENEWAL; TRANSFER, ASSIGNMENT, OR SALE OF PERMIT RIGHTS; POST... § 774.17 Transfer, assignment, or sale of permit rights. (a) General. No transfer, assignment, or sale...

30 CFR 774.17 - Transfer, assignment, or sale of permit rights.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 30 Mineral Resources 3 2011-07-01 2011-07-01 false Transfer, assignment, or sale of permit rights... SYSTEMS UNDER REGULATORY PROGRAMS REVISION; RENEWAL; TRANSFER, ASSIGNMENT, OR SALE OF PERMIT RIGHTS; POST... § 774.17 Transfer, assignment, or sale of permit rights. (a) General. No transfer, assignment, or sale...

15 CFR Supplement No. 3 to Part 774 - Statements of Understanding

Code of Federal Regulations, 2011 CFR

2011-01-01

...” that “incorporate” commodities or software on the Commerce Control List (Supplement No. 1 to part 774... medical research). (2) Commodities or software are considered “incorporated” if the commodity or software... medical equipment; and exported or reexported with the medical equipment. (3) Except for such software...

[Distribution diversity of integrins and calcium channels on major human and mouse host cells of Leptospira species].

PubMed

Li, Cheng-xue; Zhao, Xin; Qian, Jing; Yan, Jie

2012-07-01

To determine the distribution of integrins and calcium channels on major human and mouse host cells of Leptospira species. The expression of β1, β2 and β3 integrins was detected with immunofluorescence assay on the surface of human monocyte line THP-1, mouse mononuclear-macrophage-like cell line J774A.1, human vascular endothelial cell line HUVEC, mouse vascular endothelial cell EOMA, human hepatocyte line L-02, mouse hepatocyte line Hepa1-6, human renal tubular epithelial cell line HEK-293, mouse glomerular membrane epithelial cell line SV40-MES13, mouse collagen blast line NIH/3T3, human and mouse platelets. The distribution of voltage gate control calcium channels Cav3.1, Cav3.2, Cav3.3 and Cav2.3, and receptor gate calcium channels P(2)X(1), P(2)2X(2), P(2)X(3), P(2)X(4), P(2)X(5), P(2)X(6) and P(2)X(7) were determined with Western blot assay. β1 integrin proteins were positively expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, L-02, Hepa1-6 and HEK-239 cells as well as human and mouse platelets. β2 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, and NIH/3T3 cells. β3 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, Hepa1-6, HEK-239 and NIH/3T3 cells as well as human and mouse platelets. P(2)X(1) receptor gate calcium channel was expressed on the membrane surface of human and mouse platelets, while P(2)X(5) receptor gate calcium channel was expressed on the membrane surface of J774A.1, THP-1, L-02, Hepa1-6, HEK-239 and HUVEC cells. However, the other calcium channels were not detected on the tested cell lines or platelets. There is a large distribution diversity of integrins and calcium channel proteins on the major human and mouse host cells of Leptospira species, which may be associated with the differences of leptospira-induced injury in different host cells.

The Increase in Mannose Receptor Recycling Favors Arginase Induction and Trypanosoma Cruzi Survival in Macrophages

PubMed Central

Garrido, Vanina V.; Dulgerian, Laura R.; Stempin, Cinthia C.; Cerbán, Fabio M.

2011-01-01

The macrophage mannose receptor (MR) is a pattern recognition receptor of the innate immune system that binds to microbial structures bearing mannose, fucose and N-acetylglucosamine on their surface. Trypanosoma cruzi antigen cruzipain (Cz) is found in the different developmental forms of the parasite. This glycoprotein has a highly mannosylated C-terminal domain that participates in the host-antigen contact. Our group previously demonstrated that Cz-macrophage (Mo) interaction could modulate the immune response against T. cruzi through the induction of a preferential metabolic pathway. In this work, we have studied in Mo the role of MR in arginase induction and in T. cruzi survival using different MR ligands. We have showed that pre-incubation of T. cruzi infected cells with mannose-Bovine Serum Albumin (Man-BSA, MR specific ligand) biased nitric oxide (NO)/urea balance towards urea production and increased intracellular amastigotes growth. The study of intracellular signals showed that pre-incubation with Man-BSA in T. cruzi J774 infected cells induced down-regulation of JNK and p44/p42 phosphorylation and increased of p38 MAPK phosphorylation. These results are coincident with previous data showing that Cz also modifies the MAPK phosphorylation profile induced by the parasite. In addition, we have showed by confocal microscopy that Cz and Man-BSA enhance MR recycling. Furthermore, we studied MR behavior during T. cruzi infection in vivo. MR was up-regulated in F4/80+ cells from T. cruzi infected mice at 13 and 15 days post infection. Besides, we investigated the effect of MR blocking antibody in T. cruzi infected peritoneal Mo. Arginase activity and parasite growth were decreased in infected cells pre-incubated with anti-MR antibody as compared with infected cells treated with control antibody. Therefore, we postulate that during T. cruzi infection, Cz may contact with MR, increasing MR recycling which leads to arginase activity up-regulation and intracellular

Inhibitory effect of Korean Red Ginseng on melanocyte proliferation and its possible implication in GM-CSF mediated signaling

PubMed Central

Oh, Chang Taek; Park, Jong Il; Jung, Yi Ra; Joo, Yeon Ah; Shin, Dong Ha; Cho, Hyoung Joo; Ahn, Soo Mi; Lim, Young-Ho; Park, Chae Kyu; Hwang, Jae Sung

2013-01-01

Korean Red Ginseng (KRG) has been reported to exert anticancer, anti-oxidant, and anti-inflammatory effects. However, there has been no report on the effect of KRG on skin pigmentation. In this study, we investigated the inhibitory effect of KRG on melanocyte proliferation. KRG extract (KRGE) at different concentrations had no effect on melanin synthesis in melan-A melanocytes. Saponin of KRG (SKRG) inhibited melanin content to 80% of the control at 100 ppm. Keratinocyte-derived factors induced by UV-irradiation were reported to stimulate melanogenesis, differentiation, proliferation, and dendrite formation. In this study, treatment of melan-A melanocytes with conditioned media from UV-irradiated SP-1 keratinocytes increased melanocyte proliferation. When UV-irradiated SP-1 keratinocytes were treated with KRGE or SKRG, the increase of melanocyte proliferation by the conditioned media was blocked. Granulocyte-macrophage colony-stimulating factor (GM-CSF) was produced and released from UV-irradiated keratinocytes. This factor has been reported to be involved in regulating the proliferation and differentiation of epidermal melanocytes. In this study, GM-CSF was significantly increased in SP-1 keratinocytes by UVB irradiation (30 mJ/cm2), and the proliferation of melan-A melanocytes increased significantly by GM-CSF treatment. In addition, the proliferative effect of keratinocyte-conditioned media on melan-A melanocytes was blocked by anti-GM-CSF treatment. KRGE or SKRG treatment decreased the expression of GM-CSF in SP-1 keratinocytes induced by UVB irradiation. These results demonstrate that UV irradiation induced GM-CSF expression in keratinocytes and KRGE or SKRG inhibited its expression. Therefore, KRG could be a good candidate for regulating UV-induced melanocyte proliferation. PMID:24235857

Optimization of cell receptor-specific targeting through multivalent surface decoration of polymeric nanocarriers

PubMed Central

D’Addio, Suzanne M.; Baldassano, Steven; Shi, Lei; Cheung, Lila; Adamson, Douglas H.; Bruzek, Matthew; Anthony, John E.; Laskin, Debra L.; Sinko, Patrick J.; Prud’homme, Robert K.

2013-01-01

Treatment of tuberculosis is impaired by poor drug bioavailability, systemic side effects, patient non-compliance, and pathogen resistance to existing therapies. The mannose receptor (MR) is known to be involved in the recognition and internalization of Mycobacterium tuberculosis. We present a new assembly process to produce nanocarriers with variable surface densities of mannose targeting ligands in a single step, using kinetically-controlled, block copolymer-directed assembly. Nanocarrier association with murine macrophage J774 cells expressing the MR is examined as a function of incubation time and temperature, nanocarrier size, dose, and PEG corona properties. Amphiphilic diblock copolymers are prepared with terminal hydroxyl, methoxy, or mannoside functionality and incorporated into nanocarrier formulations at specific ratios by Flash NanoPrecipitation. Association of nanocarriers protected by a hydroxyl-terminated PEG corona with J774 cells is size dependent, while nanocarriers with methoxy-terminated PEG coronas do not associate with cells, regardless of size. Specific targeting of the MR is investigated using nanocarriers having 0-75% mannoside-terminated PEG chains in the PEG corona. This is a wider range of mannose densities than has been previously studied. Maximum nanocarrier association is attained with 9% mannoside-terminated PEG chains, increasing uptake more than 3-fold compared to non-targeted nanocarriers with a 5 kg mol−1 methoxy-terminated PEG corona. While a 5 kg mol−1 methoxy-terminated PEG corona prevents non-specific uptake, a 1.8 kg mol−1 methoxy-terminated PEG corona does not sufficiently protect the nanocarriers from nonspecific association. There is continuous uptake of MR-targeted nanocarriers at 37°C, but a saturation of association at 4°C. The majority of targeted nanocarriers associate with J774E cells are internalized at 37°C and uptake is receptor-dependent, diminishing with competitive inhibition by dextran. This

Class A1 scavenger receptor modulates glioma progression by regulating M2-like tumor-associated macrophage polarization

PubMed Central

Zhang, Hanwen; Zhang, Wenbin; Sun, Xuan; Dang, Ruoyu; Zhou, Rongmei; Bai, Hui; Ben, Jingjing; Zhu, Xudong; Zhang, Yan; Yang, Qing; Xu, Yong; Chen, Qi

2016-01-01

Macrophages enhance glioma development and progression by shaping the tumor microenvironment. Class A1 scavenger receptor (SR-A1), a pattern recognition receptor primarily expressed in macrophages, is up-regulated in many human solid tumors. We found that SR-A1 expression in 136 human gliomas was positively correlated with tumor grade (P<0.01), but not prognosis or tumor recurrence. SR-A1-expressing macrophages originated primarily from circulating monocytes attracted to tumor tissue, and were almost twice as numerous as resident microglia in glioma tissues (P<0.001). The effects of SR-A1 on glioma proliferation and invasion were assessed in vivo using an SR-A1-deficient murine orthotopic glioma model. SR-A1 deletion promoted M2-like tumor-associated macrophage (TAM) polarization in mice by activating STAT3 and STAT6, which resulted in robust orthotopic glioma proliferation and angiogenesis. Finally, we found that HSP70 might be an endogenous ligand that activates SR-A1-dependent anti-tumorigenic pathways in gliomas, although its expression does not appear informative for diagnostic purposes. Our findings demonstrate a relationship between TAMs, SR-A1 expression and glioma growth and provide new insights into the pathogenic role of TAMs in glioma. PMID:27367025

Heterogeneity Within Macrophage Populations: A Possible Role for Colony Stimulating Factors

DTIC Science & Technology

1988-04-04

highest concentration ofriFN-yused (5.0 U/ml), a depression of T cell proliferation induced by the antigen-pulsed rGM-CSF-derived macrophages was...stimulation by rGM-CSF and nCSF-1 in bone marrow cells derived from normal mice and mice 3 and 7 days post-treatment with 5FU . Bone marrow cells

Polysaccharide of Dendrobium huoshanense activates macrophages via toll-like receptor 4-mediated signaling pathways.

PubMed

Xie, Song-Zi; Hao, Ran; Zha, Xue-Qiang; Pan, Li-Hua; Liu, Jian; Luo, Jian-Ping

2016-08-01

The present work aimed at investigating the pattern recognition receptor (PRR) and immunostimulatory mechanism of a purified Dendrobium huoshanense polysaccharide (DHP). We found that DHP could bind to the surface of macrophages and stimulate macrophages to secrete NO, TNF-α and IL-1β. To unravel the mechanism for the binding of DHP to macrophages, flow cytometry, confocal laser-scanning microscopy, affinity electrophoresis, SDS-PAGE and western blotting were employed to verify the type of PRR responsible for the recognition of DHP by RAW264.7 macrophages and peritoneal macrophages of C3H/HeN and C3H/HeJ macrophages. Results showed that toll-like receptor 4 (TLR4) was an essential receptor for macrophages to directly bind DHP. Further, the phosphorylation of ERK, JNK, Akt and p38 were observed to be time-dependently promoted by DHP, as well as the nuclear translocation of NF-κB p65. These results suggest that DHP activates macrophages via its direct binding to TLR4 to trigger TLR4 signaling pathways. Copyright © 2016 Elsevier Ltd. All rights reserved.

A critical role for macrophages in neovessel formation and the development of stenosis in tissue-engineered vascular grafts

PubMed Central

Hibino, Narutoshi; Yi, Tai; Duncan, Daniel R.; Rathore, Animesh; Dean, Ethan; Naito, Yuji; Dardik, Alan; Kyriakides, Themis; Madri, Joseph; Pober, Jordan S.; Shinoka, Toshiharu; Breuer, Christopher K.

2011-01-01

The primary graft-related complication during the first clinical trial evaluating the use of tissue-engineered vascular grafts (TEVGs) was stenosis. We investigated the role of macrophages in the formation of TEVG stenosis in a murine model. We analyzed the natural history of TEVG macrophage infiltration at critical time points and evaluated the role of cell seeding on neovessel formation. To assess the function of infiltrating macrophages, we implanted TEVGs into mice that had been macrophage depleted using clodronate liposomes. To confirm this, we used a CD11b-diphtheria toxin-receptor (DTR) transgenic mouse model. Monocytes infiltrated the scaffold within the first few days and initially transformed into M1 macrophages. As the scaffold degraded, the macrophage infiltrate disappeared. Cell seeding decreased the incidence of stenosis (32% seeded, 64% unseeded, P=0.024) and the degree of macrophage infiltration at 2 wk. Unseeded TEVGs demonstrated conversion from M1 to M2 phenotype, whereas seeded grafts did not. Clodronate and DTR inhibited macrophage infiltration and decreased stenosis but blocked formation of vascular neotissue, evidenced by the absence of endothelial and smooth muscle cells and collagen. These findings suggest that macrophage infiltration is critical for neovessel formation and provides a strategy for predicting, detecting, and inhibiting stenosis in TEVGs.—Hibino, N., Yi, T., Duncan, D. R., Rathore, A., Dean, E., Naito, Y., Dardik, A., Kyriakides, T., Madri, J., Pober, J. S., Shinoka, T., Breuer, C. K. A critical role for macrophages in neovessel formation and the development of stenosis in tissue-engineered vascular grafts. PMID:21865316

CD74-Downregulation of Placental Macrophage-Trophoblastic Interactions in Preeclampsia.

PubMed

Przybyl, Lukasz; Haase, Nadine; Golic, Michaela; Rugor, Julianna; Solano, Maria Emilia; Arck, Petra Clara; Gauster, Martin; Huppertz, Berthold; Emontzpohl, Christoph; Stoppe, Christian; Bernhagen, Jürgen; Leng, Lin; Bucala, Richard; Schulz, Herbert; Heuser, Arnd; Weedon-Fekjær, M Susanne; Johnsen, Guro M; Peetz, Dirk; Luft, Friedrich C; Staff, Anne Cathrine; Müller, Dominik N; Dechend, Ralf; Herse, Florian

2016-06-24

We hypothesized that cluster of differentiation 74 (CD74) downregulation on placental macrophages, leading to altered macrophage-trophoblast interaction, is involved in preeclampsia. Preeclamptic pregnancies feature hypertension, proteinuria, and placental anomalies. Feto-placental macrophages regulate villous trophoblast differentiation during placental development. Disturbance of this well-balanced regulation can lead to pathological pregnancies. We performed whole-genome expression analysis of placental tissue. CD74 was one of the most downregulated genes in placentas from preeclamptic women. By reverse transcriptase-polymerase chain reaction, we confirmed this finding in early-onset (<34 gestational week, n=26) and late-onset (≥34 gestational week, n=24) samples from preeclamptic women, compared with healthy pregnant controls (n=28). CD74 protein levels were analyzed by Western blot and flow cytometry. We identified placental macrophages to express CD74 by immunofluorescence, flow cytometry, and RT-PCR. CD74-positive macrophages were significantly reduced in preeclamptic placentas compared with controls. CD74-silenced macrophages showed that the adhesion molecules ALCAM, ICAM4, and Syndecan-2, as well as macrophage adhesion to trophoblasts were diminished. Naive and activated macrophages lacking CD74 showed a shift toward a proinflammatory signature with an increased secretion of tumor necrosis factor-α, chemokine (C-C motif) ligand 5, and monocyte chemotactic protein-1, when cocultured with trophoblasts compared with control macrophages. Trophoblasts stimulated by these factors express more CYP2J2, sFlt1, TNFα, and IL-8. CD74-knockout mice showed disturbed placental morphology, reduced junctional zone, smaller placentas, and impaired spiral artery remodeling with fetal growth restriction. CD74 downregulation in placental macrophages is present in preeclampsia. CD74 downregulation leads to altered macrophage activation toward a proinflammatory signature and

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

PubMed

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

2016-02-12

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

Macrophage-Specific Expression of IL-37 in Hyperlipidemic Mice Attenuates Atherosclerosis.

PubMed

McCurdy, Sara; Baumer, Yvonne; Toulmin, Emma; Lee, Bog-Hieu; Boisvert, William A

2017-11-15

Atherosclerosis, the progressive buildup of plaque within arterial blood vessels, can lead to fatal downstream events, such as heart attack or stroke. A key event contributing to the development of atherosclerosis is the infiltration of monocytes and its associated inflammation, as well as the formation of lipid-laden macrophage foam cells within the vessel wall. IL-37 is recognized as an important anti-inflammatory cytokine expressed especially by immune cells. This study was undertaken to elucidate the role of macrophage-expressed IL-37 in reducing the production and effects of proinflammatory cytokines, preventing foam cell formation, and reducing the development of atherosclerosis. Expression of human IL-37 was achieved with a macrophage-specific overexpression system, using the CD68 promoter in mouse primary bone marrow-derived macrophages via retroviral transduction. Macrophage IL-37 expression in vitro resulted in decreased mRNA (e.g., IL-1B, IL-6, and IL-12) and secreted protein production (e.g., IL-6, M-CSF, and ICAM-1) of key inflammatory mediators. IL-37 expression also inhibited macrophage proliferation, apoptosis, and transmigration, as well as reduced lipid uptake, compared with controls in vitro. The in vivo effects of macrophage-expressed IL-37 were investigated through bone marrow transplantation of transduced hematopoietic stem cells into irradiated atherosclerosis-prone Ldlr -/- mice. After 10 wk on a high-fat/high-cholesterol diet, mice with IL-37-expressing macrophages showed reduced disease pathogenesis, which was demonstrated by significantly less arterial plaque development and systemic inflammation compared with control mice. The athero-protective effect of macrophage-expressed IL-37 has implications for development of future therapies to treat atherosclerosis, as well as other chronic inflammatory diseases. Copyright © 2017 by The American Association of Immunologists, Inc.

Comparative Analysis of Transcriptomes of Macrophage Revealing the Mechanism of the Immunoregulatory Activities of a Novel Polysaccharide Isolated from Boletus speciosus Frost

PubMed Central

Ding, Xiang; Zhu, Hongqing; Hou, Yiling; Hou, Wanru; Zhang, Nan; Fu, Lei

2017-01-01

Background: The mechanism of the immunoregulatory activities of polysaccharide is still not clear. Materials and Methods: Here, we performed the B-cell, T-cell, and macrophage cell proliferation, the cell cycle analysis of macrophage cells, sequenced the transcriptomes of control group macrophages, and Boletus speciosus Frost polysaccharide (BSF-1) group macrophages using Illumina sequencing technology to identify differentially expressed genes (DEGs) to determine the molecular mechanisms of immunomodulatory activity of BSF-1 in macrophages. Results: These results suggested that BSF-1 could promote the proliferation of B-cell, T-cell, and macrophages, promote the proliferation of macrophage cells by abolishing cell cycle arrests in the G0/G1 phases, and promote cell cycle progression in S-phase and G2/M phase, which might induce cell division. A total of 12,498,414 and 11,840,624 bp paired-end reads were obtained for the control group and BSF-1 group, respectively, and they corresponded to a total size of 12.5 G bp and 11.8 G bp, respectively, after the low-quality reads and adapter sequences were removed. Approximately 81.83% of the total number of genes (8,257) were expressed reads per kilobase per million mapped reads (RPKM ≥1) and more than 1366 genes were highly expressed (RPKM >60) in the BSF-1 group. A gene ontology-enrichment analysis generated 13,042 assignments to cellular components, 13,094 assignments to biological processes, and 13,135 assignments to molecular functions. A Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the mitogen-activated protein kinase (MAPK) signaling pathways are significantly enriched for DEGs between the two cell groups. Conclusion: An analysis of transcriptome resources enabled us to examine gene expression profiles, verify differential gene expression, and select candidate signaling pathways as the mechanisms of the immunomodulatory activity of BSF-1. Based on the experimental data, we

Comparative Analysis of Transcriptomes of Macrophage Revealing the Mechanism of the Immunoregulatory Activities of a Novel Polysaccharide Isolated from Boletus speciosus Frost.

PubMed

Ding, Xiang; Zhu, Hongqing; Hou, Yiling; Hou, Wanru; Zhang, Nan; Fu, Lei

2017-01-01

The mechanism of the immunoregulatory activities of polysaccharide is still not clear. Here, we performed the B-cell, T-cell, and macrophage cell proliferation, the cell cycle analysis of macrophage cells, sequenced the transcriptomes of control group macrophages, and Boletus speciosus Frost polysaccharide (BSF-1) group macrophages using Illumina sequencing technology to identify differentially expressed genes (DEGs) to determine the molecular mechanisms of immunomodulatory activity of BSF-1 in macrophages. These results suggested that BSF-1 could promote the proliferation of B-cell, T-cell, and macrophages, promote the proliferation of macrophage cells by abolishing cell cycle arrests in the G0/G1 phases, and promote cell cycle progression in S-phase and G2/M phase, which might induce cell division. A total of 12,498,414 and 11,840,624 bp paired-end reads were obtained for the control group and BSF-1 group, respectively, and they corresponded to a total size of 12.5 G bp and 11.8 G bp, respectively, after the low-quality reads and adapter sequences were removed. Approximately 81.83% of the total number of genes (8,257) were expressed reads per kilobase per million mapped reads (RPKM ≥1) and more than 1366 genes were highly expressed (RPKM >60) in the BSF-1 group. A gene ontology-enrichment analysis generated 13,042 assignments to cellular components, 13,094 assignments to biological processes, and 13,135 assignments to molecular functions. A Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the mitogen-activated protein kinase (MAPK) signaling pathways are significantly enriched for DEGs between the two cell groups. An analysis of transcriptome resources enabled us to examine gene expression profiles, verify differential gene expression, and select candidate signaling pathways as the mechanisms of the immunomodulatory activity of BSF-1. Based on the experimental data, we believe that the significant antitumor activities of BSF-1 in

High salt reduces the activation of IL-4- and IL-13-stimulated macrophages.

PubMed

Binger, Katrina J; Gebhardt, Matthias; Heinig, Matthias; Rintisch, Carola; Schroeder, Agnes; Neuhofer, Wolfgang; Hilgers, Karl; Manzel, Arndt; Schwartz, Christian; Kleinewietfeld, Markus; Voelkl, Jakob; Schatz, Valentin; Linker, Ralf A; Lang, Florian; Voehringer, David; Wright, Mark D; Hubner, Norbert; Dechend, Ralf; Jantsch, Jonathan; Titze, Jens; Müller, Dominik N

2015-11-02

A high intake of dietary salt (NaCl) has been implicated in the development of hypertension, chronic inflammation, and autoimmune diseases. We have recently shown that salt has a proinflammatory effect and boosts the activation of Th17 cells and the activation of classical, LPS-induced macrophages (M1). Here, we examined how the activation of alternative (M2) macrophages is affected by salt. In stark contrast to Th17 cells and M1 macrophages, high salt blunted the alternative activation of BM-derived mouse macrophages stimulated with IL-4 and IL-13, M(IL-4+IL-13) macrophages. Salt-induced reduction of M(IL-4+IL-13) activation was not associated with increased polarization toward a proinflammatory M1 phenotype. In vitro, high salt decreased the ability of M(IL-4+IL-13) macrophages to suppress effector T cell proliferation. Moreover, mice fed a high salt diet exhibited reduced M2 activation following chitin injection and delayed wound healing compared with control animals. We further identified a high salt-induced reduction in glycolysis and mitochondrial metabolic output, coupled with blunted AKT and mTOR signaling, which indicates a mechanism by which NaCl inhibits full M2 macrophage activation. Collectively, this study provides evidence that high salt reduces noninflammatory innate immune cell activation and may thus lead to an overall imbalance in immune homeostasis.

Immunomodulatory effect of Parsley (Petroselinum crispum) essential oil on immune cells: mitogen-activated splenocytes and peritoneal macrophages.

PubMed

Yousofi, Alireza; Daneshmandi, Saeed; Soleimani, Neda; Bagheri, Kambiz; Karimi, Mohammad Hossein

2012-04-01

Parsley (Petroselinum crispum) has been traditionally used for the treatment of allergy, autoimmune and chronic inflammatory disorders. The present study aims to investigate the suppressive effects of parsley essential oil on mouse splenocytes and macrophages cells. Parsley essential oil was harvested. It was treated on splenocytes and phytohemagglutinin (PHA) (5 μg/mL) and lipopolysaccharide (LPS) (10 μg/mL) activated splenocytes in different concentrations (0.01-100 μg/mL); then, proliferation was assayed by methyl tetrazolium (MTT) method. Treatment was also performed on the macrophages and LPS-stimulated macrophages (10 μg/ml) and the nitrite levels were measured using the diazotization method based on the Griess reaction and MTT assay for evaluation of the viability of the macrophages. Proliferation of splenocytes in all the treated groups was suppressed. In PHA-stimulated splenocytes, the suppression was seen in all the examined concentrations (0.01-100 μg/mL), while in the unstimulated and LPS-stimulated groups suppression was relatively dose dependent and in high concentration (10 and100 μg/mL).The viability of the macrophages in all groups was the same and in the unstimulated groups; NO suppression was significant in all the concentrations but in LPS-stimulated groups, it was significant in the three higher concentrations (1, 10, and100 μg/mL). The results of this study indicate that parsley essential oil may be able to suppress the cellular and humoral immune response. It can also suppress both NO production and the functions of macrophages as the main innate immune cells. These results may suggest that parsley essential oil is a proper suppressant for different applications.

Neuroendocrine-like cells -derived CXCL10 and CXCL11 induce the infiltration of tumor-associated macrophage leading to the poor prognosis of colorectal cancer

PubMed Central

Liu, Lu; Xu, He-Yang; Wang, Jie; Chu, Zhong-Hua

2016-01-01

Our previous study revealed that neuroendocrine differentiation in colorectal cancer is one of the important factors leading to worse prognosis. In this study, we apply immunohistochemical staining, Western-blot, RT-PCR and ELISA to investigate the underlying mechanism that how the neuroendocrine differentiation to affect the prognosis of colorectal cancer. The interaction of colorectal cancer cells, neuroendocrine-like cells and tumor-associated macrophages in colorectal cancer progress is also investigated. By analyzing 82 cases of colorectal cancer patients treated in our institution, we found that colorectal adenocarcinoma with neuroendocrine differentiation had increasing number of tumor-associated macrophages and worse prognosis. Further evaluation of cytology showed that neuroendocrine cells have the ability to recruit tumor-associated macrophages to infiltrate the tumor tissue, and the tumor-associated macrophages enhance the proliferation and invasion abilities of the colon cancer cells. Moreover, we confirmed that CXCL10 and CXCL11 are the key chemokines in neuroendocrine-like cells and they promote the chemotaxis activity of tumor-associated macrophages. The secretion of CXCL10 and CXCL11 by neuroendocrine-like cells can recruit tumor-associated macrophages to infiltrate in tumor tissues. The latter enhances the proliferation and invasion of colorectal cancer cell and lead to poor prognosis. PMID:27034164

Mycelia extracts of fungal strains isolated from Cordyceps sinensis differently enhance the function of RAW 264.7 macrophages.

PubMed

Meng, Lan-Zhen; Lin, Bao-Qin; Wang, Bo; Feng, Kun; Hu, De-Jun; Wang, Lan-Ying; Cheong, Kit-Leong; Zhao, Jing; Li, Shao-Ping

2013-07-30

Cordyceps sinensis, an entomogenous fungus used in traditional Chinese medicine with multiple pharmacological activities. However, its usage has been limited due to the high price and short supply. Isolate of fungi strains from natural Cordyceps sinensis to achieve a large-scale production by fermentation is an alternative choice. The aim of this study was to investigate and compare the effects of mycelia extracts of different fungal stains isolated from natural Cordyceps sinensis on macrophage functions in vitro. Macrophages' proliferation, phagocytosis, nitric oxide (NO) production, cytokines secretion, iNOS, NF-κB p65 activation and translocation were investigated by the MTT assay, flow cytometry assay, Griess reagent method, ELISA, western blot and immunostaining assay, respectively. The results showed that the effects of cultured Cordyceps mycelia of different fungal strains isolated from natural Cordyceps sinensis on macrophages greatly variant. Among 17 Cordyceps aqueous extracts, only five extracts (UM01, QH11, BNQM, GNCC and DCXC) could significantly increase cell proliferation and NO production of RAW 264.7 mouse macrophages. Moreover, the five extracts, especially UM01 and QH11, significantly enhanced phagocytosis and promoted cytokines release of macrophages. Polysaccharides in cultured UM01 mycelia were found to be the main immune stimulating compounds. The variation of biological effects of fermented mycelia of different fungal strains from natural Cordyceps sinensis may be derived from their chemical diversity, especially polysaccharides, which need further study in future. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Mesenchymal Stem Cells Promote Diabetic Corneal Epithelial Wound Healing Through TSG-6-Dependent Stem Cell Activation and Macrophage Switch.

PubMed

Di, Guohu; Du, Xianli; Qi, Xia; Zhao, Xiaowen; Duan, Haoyun; Li, Suxia; Xie, Lixin; Zhou, Qingjun

2017-08-01

To explore the role and mechanism of bone marrow-derived mesenchymal stem cells (BM-MSCs) in corneal epithelial wound healing in type 1 diabetic mice. Diabetic mice were treated with subconjunctival injections of BM-MSCs or recombinant tumor necrosis factor-α-stimulated gene/protein-6 (TSG-6). The corneal epithelial wound healing rate was examined by fluorescein staining. The mRNA and protein expression levels of TSG-6 were measured by quantitative RT-PCR and Western blot. The infiltrations of leukocytes and macrophages were analyzed by flow cytometry and immunofluoresence staining. The effect of TSG-6 was further evaluated in cultured limbal epithelial stem/progenitor cells, macrophages, and diabetic mice by short hairpin RNA (shRNA) knockdown. Local MSC transplantation significantly promoted diabetic corneal epithelial wound healing, accompanied by elevated corneal TSG-6 expression, increased corneal epithelial cell proliferation, and attenuated inflammatory response. Moreover, in cultured human limbal epithelial stem/progenitor cells, TSG-6 enhanced the colony-forming efficiency, stimulated mitogenic proliferation, and upregulated the expression level of ΔNp63. Furthermore, in diabetic mouse cornea and in vitro macrophage culture, TSG-6 alleviated leukocyte infiltration and promoted the polarization of recruited macrophages to anti-inflammatory M2 phenotypes with increased phagocytotic capacity. In addition, the promotion of epithelial stem/progenitor cell activation and macrophage polarization by MSC transplantation was largely abrogated by shRNA knockdown of TSG-6. This study provided the first evidence of TSG-6 secreted by MSCs promoting corneal epithelial wound healing in diabetic mice through activating corneal epithelial stem/progenitor cells and accelerating M2 macrophage polarization.

Cell proliferation and differentiation in chemical leukemogenesis

NASA Technical Reports Server (NTRS)

Irons, R. D.; Stillman, W. S.; Clarkson, T. W. (Principal Investigator)

1993-01-01

In tissues such as bone marrow with normally high rates of cell division, proliferation is tightly coordinated with cell differentiation. Survival, proliferation and differentiation of early hematopoietic progenitor cells depend on the growth factors, interleukin 3 (IL-3) and/or granulocyte-macrophage colony stimulating factor (GM-CSF) and their synergism with other cytokines. We provide evidence that a characteristic shared by a diverse group of compounds with demonstrated leukemogenic potential is the ability to act synergistically with GM-CSF. This results in an increase in recruitment of a resting population of hematopoietic progenitor cells normally unresponsive to the cytokine and a twofold increase in the size of the proliferating cell population normally regarded to be at risk of transformation in leukemogenesis. These findings support the possibility that transient alterations in hematopoietic progenitor cell differentiation may be an important factor in the early stages of development of leukemia secondary to chemical or drug exposure.

Evaluation of in vitro macrophage differentiation during space flight

NASA Astrophysics Data System (ADS)

Ortega, M. Teresa; Lu, Nanyan; Chapes, Stephen K.

2012-05-01

We differentiated mouse bone marrow cells in the presence of recombinant macrophage colony stimulating (rM-CSF) factor for 14 days during the flight of space shuttle Space Transportation System (STS)-126. We tested the hypothesis that the receptor expression for M-CSF, c-Fms was reduced. We used flow cytometry to assess molecules on cells that were preserved during flight to define the differentiation state of the developing bone marrow macrophages; including CD11b, CD31, CD44, Ly6C, Ly6G, F4/80, Mac2, c-Fos as well as c-Fms. In addition, RNA was preserved during the flight and was used to perform a gene microarray. We found that there were significant differences in the number of macrophages that developed in space compared to controls maintained on Earth. We found that there were significant changes in the distribution of cells that expressed CD11b, CD31, F4/80, Mac2, Ly6C and c-Fos. However, there were no changes in c-Fms expression and no consistent pattern of advanced or retarded differentiation during space flight. We also found a pattern of transcript levels that would be consistent with a relatively normal differentiation outcome but increased proliferation by the bone marrow macrophages that were assayed after 14 days of space flight. There also was a surprising pattern of space flight influence on genes of the coagulation pathway. These data confirm that a space flight can have an impact on the in vitro development of macrophages from mouse bone marrow cells.

Evaluation of in vitro macrophage differentiation during space flight.

PubMed

Ortega, M Teresa; Lu, Nanyan; Chapes, Stephen K

2012-05-15

We differentiated mouse bone marrow cells in the presence of recombinant macrophage colony stimulating (rM-CSF) factor for 14 days during the flight of space shuttle Space Transportation System (STS)-126. We tested the hypothesis that the receptor expression for M-CSF, c-Fms was reduced. We used flow cytometry to assess molecules on cells that were preserved during flight to define the differentiation state of the developing bone marrow macrophages; including CD11b, CD31, CD44, Ly6C, Ly6G, F4/80, Mac2, c-Fos as well as c-Fms. In addition, RNA was preserved during the flight and was used to perform a gene microarray. We found that there were significant differences in the number of macrophages that developed in space compared to controls maintained on Earth. We found that there were significant changes in the distribution of cells that expressed CD11b, CD31, F4/80, Mac2, Ly6C and c-Fos. However, there were no changes in c-Fms expression and no consistent pattern of advanced or retarded differentiation during space flight. We also found a pattern of transcript levels that would be consistent with a relatively normal differentiation outcome but increased proliferation by the bone marrow macrophages that were assayed after 14 days of space flight. There also was a surprising pattern of space flight influence on genes of the coagulation pathway. These data confirm that a space flight can have an impact on the in vitro development of macrophages from mouse bone marrow cells.

Investigation of Dracocephalum kostchyi plant extract on the effective inflammatory transcription factors and mediators in activated macrophages.

PubMed

Kalantar, Kurosh; Gholijani, Nasser; Mousaei, Nashmin; Yazdani, Malihe; Amirghofran, Zahra

2018-06-07

Dracocephalum kotschyi is traditionally used for its anti-inflammatory effects. We aimed to investigate the effects of ethyl acetate extract of D. kotschyi on the expression of key inflammatory mediators and main signaling molecules involved in regulation of inflammation. Lipopolysaccharide (LPS)-stimulated J774.1 mouse macrophages were cultured in the presence of the plant extract and examined by the real time-PCR for gene expressions of interleukin (IL)-1β, tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2. Cytokine levels and phosphorylated forms of stress-activated protein kinases/c-Jun N-terminal kinase (SAPK/JNK), signal transducer and activator of transcription (STAT)-3, p38, IκB-α and nuclear factor (NF)-κB p65 were determined using ELISA. The extract significantly reduced the expression of key mediators of inflammation. iNOS expression level decreased from 138±8.5 fold in LPS-only treated cells to 6.5±2.6 fold after treatment with 25 ug/ml of the extract (p<0.001). Similarly, COX-2 expression decreased from 632 ±98.8 fold in control to 124 ±24.6 fold (p<0.01). Treatment of cells with the extract significantly reduced IL-1β and TNF-α cytokines at both gene and protein expression levels. The extract at 25 µg/ml caused significant decreases in phospho-SAPK/JNK and phospho-STAT3 levels in macrophages (p<0.01). Proteins of phospho-p38, NFκB-p65 and phospho-NF-κB p65 had a reduced levels in treated cells (p<0.05). No significant change in phospho-IĸB level was observed. These findings suggested that D. kotschyi with inhibition of NF-κB, SAPK/JNK, STAT-3 and p-38 might have reduced the expression levels of key inflammatory mediators and thus possibly have potential beneficial impact on inflammatory diseases. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

MiR-34b-5p Suppresses Melanoma Differentiation-Associated Gene 5 (MDA5) Signaling Pathway to Promote Avian Leukosis Virus Subgroup J (ALV-J)-Infected Cells Proliferaction and ALV-J Replication

PubMed Central

Li, Zhenhui; Luo, Qingbin; Xu, Haiping; Zheng, Ming; Abdalla, Bahareldin Ali; Feng, Min; Cai, Bolin; Zhang, Xiaocui; Nie, Qinghua; Zhang, Xiquan

2017-01-01

Avian leukosis virus subgroup J (ALV-J) is an oncogenic retrovirus that has a similar replication cycle to multiple viruses and therefore can be used as a model system for viral entry into host cells. However, there are few reports on the genes or microRNAs (miRNAs) that are responsible for the replication of ALV-J. Our previous miRNA and RNA sequencing data showed that the expression of miR-34b-5p was significantly upregulated in ALV-J-infected chicken spleens compared to non-infected chicken spleens, but melanoma differentiation-associated gene 5 (MDA5) had the opposite expression pattern. In this study, a dual-luciferase reporter assay showed that MDA5 is a direct target of miR-34b-5p. In vitro, overexpression of miR-34b-5p accelerated the proliferation of ALV-J-infected cells by inducing the progression from G2 to S phase and it promoted cell migration. Ectopic expression of MDA5 inhibited ALV-J-infected cell proliferation, the cell cycle and cell migration, and knockdown of MDA5 promoted proliferation, the cell cycle and migration. In addition, during ALV-J infections, MDA5 can detect virus invasion and it triggers the MDA5 signaling pathway. MDA5 overexpression can activate the MDA5 signaling pathway, and thus it can inhibit the mRNA and protein expression of the ALV-J env gene and it can suppress virion secretion. In contrast, in response to the knockdown of MDA5 by small interfering RNA (siRNA) or an miR-34b-5p mimic, genes in the MDA5 signaling pathway were significantly downregulated (P < 0.05), but the mRNA and protein expression of ALV-J env and the sample-to-positive ratio of virion in the supernatants were increased. This indicates that miR-34b-5p is able to trigger the MDA5 signaling pathway and affect ALV-J infections. Together, these results suggest that miR-34b-5p targets MDA5 to accelerate the proliferation and migration of ALV-J-infected cells, and it promotes ALV-J replication, via the MDA5 signaling pathway. PMID:28194372

MiR-34b-5p Suppresses Melanoma Differentiation-Associated Gene 5 (MDA5) Signaling Pathway to Promote Avian Leukosis Virus Subgroup J (ALV-J)-Infected Cells Proliferaction and ALV-J Replication.

PubMed

Li, Zhenhui; Luo, Qingbin; Xu, Haiping; Zheng, Ming; Abdalla, Bahareldin Ali; Feng, Min; Cai, Bolin; Zhang, Xiaocui; Nie, Qinghua; Zhang, Xiquan

2017-01-01

Avian leukosis virus subgroup J (ALV-J) is an oncogenic retrovirus that has a similar replication cycle to multiple viruses and therefore can be used as a model system for viral entry into host cells. However, there are few reports on the genes or microRNAs (miRNAs) that are responsible for the replication of ALV-J. Our previous miRNA and RNA sequencing data showed that the expression of miR-34b-5p was significantly upregulated in ALV-J-infected chicken spleens compared to non-infected chicken spleens, but melanoma differentiation-associated gene 5 ( MDA5 ) had the opposite expression pattern. In this study, a dual-luciferase reporter assay showed that MDA5 is a direct target of miR-34b-5p. In vitro , overexpression of miR-34b-5p accelerated the proliferation of ALV-J-infected cells by inducing the progression from G2 to S phase and it promoted cell migration. Ectopic expression of MDA5 inhibited ALV-J-infected cell proliferation, the cell cycle and cell migration, and knockdown of MDA5 promoted proliferation, the cell cycle and migration. In addition, during ALV-J infections, MDA5 can detect virus invasion and it triggers the MDA5 signaling pathway. MDA5 overexpression can activate the MDA5 signaling pathway, and thus it can inhibit the mRNA and protein expression of the ALV-J env gene and it can suppress virion secretion. In contrast, in response to the knockdown of MDA5 by small interfering RNA (siRNA) or an miR-34b-5p mimic, genes in the MDA5 signaling pathway were significantly downregulated ( P < 0.05), but the mRNA and protein expression of ALV-J env and the sample-to-positive ratio of virion in the supernatants were increased. This indicates that miR-34b-5p is able to trigger the MDA5 signaling pathway and affect ALV-J infections. Together, these results suggest that miR-34b-5p targets MDA5 to accelerate the proliferation and migration of ALV-J-infected cells, and it promotes ALV-J replication, via the MDA5 signaling pathway.

Differential Expression of Inward and Outward Potassium Currents in the Macrophage-like Cell Line J774.1

DTIC Science & Technology

1985-04-02

skeletal muscle (Adrian, Hodgkin & Chandler, 1970), and nerve (Goldman & Schauf , 1973; Thompson, 1977). The general mathematical formalism describes the...594-609. Goldman, L. & Schauf , C. L. (1973) Quantitative description of sodium and potassium currents and computed action potentials in Myxicola giant

The Role of Macrophages in Acute and Chronic Wound Healing and Interventions to Promote Pro-wound Healing Phenotypes

PubMed Central

Krzyszczyk, Paulina; Schloss, Rene; Palmer, Andre; Berthiaume, François

2018-01-01

Macrophages play key roles in all phases of adult wound healing, which are inflammation, proliferation, and remodeling. As wounds heal, the local macrophage population transitions from predominantly pro-inflammatory (M1-like phenotypes) to anti-inflammatory (M2-like phenotypes). Non-healing chronic wounds, such as pressure, arterial, venous, and diabetic ulcers indefinitely remain in inflammation—the first stage of wound healing. Thus, local macrophages retain pro-inflammatory characteristics. This review discusses the physiology of monocytes and macrophages in acute wound healing and the different phenotypes described in the literature for both in vitro and in vivo models. We also discuss aberrations that occur in macrophage populations in chronic wounds, and attempts to restore macrophage function by therapeutic approaches. These include endogenous M1 attenuation, exogenous M2 supplementation and endogenous macrophage modulation/M2 promotion via mesenchymal stem cells, growth factors, biomaterials, heme oxygenase-1 (HO-1) expression, and oxygen therapy. We recognize the challenges and controversies that exist in this field, such as standardization of macrophage phenotype nomenclature, definition of their distinct roles and understanding which phenotype is optimal in order to promote healing in chronic wounds. PMID:29765329

ALV-J infection induces chicken monocyte death accompanied with the production of IL-1β and IL-18.

PubMed

Dai, Manman; Feng, Min; Xie, Tingting; Li, Yuanfang; Ruan, Zhuohao; Shi, Meiqing; Liao, Ming; Zhang, Xiquan

2017-11-21

Immunosuppression induced by avian leukosis virus subgroup J (ALV-J) causes serious reproduction problems and secondary infections in chickens. Given that monocytes are important precursors of immune cells including macrophages and dendritic cells, we investigated the fate of chicken monocytes after ALV-J infection. Our results indicated that most monocytes infected with ALV-J including field or laboratory strains could not successfully differentiate into macrophages due to cells death. And cells death was dependent upon viral titer and accompanied with increased IL-1β and IL-18 mRNA levels. In addition, ALV-J infection up-regulated caspase-1 and caspase-3 activity in monocytes. Collectively, we found that ALV-J could cause cell death in chicken monocytes, especially pyroptosis, which may be a significant reason for ALV-J induced immunosuppression.

ALV-J infection induces chicken monocyte death accompanied with the production of IL-1β and IL-18

PubMed Central

Dai, Manman; Feng, Min; Xie, Tingting; Li, Yuanfang; Ruan, Zhuohao; Shi, Meiqing; Liao, Ming; Zhang, Xiquan

2017-01-01

Immunosuppression induced by avian leukosis virus subgroup J (ALV-J) causes serious reproduction problems and secondary infections in chickens. Given that monocytes are important precursors of immune cells including macrophages and dendritic cells, we investigated the fate of chicken monocytes after ALV-J infection. Our results indicated that most monocytes infected with ALV-J including field or laboratory strains could not successfully differentiate into macrophages due to cells death. And cells death was dependent upon viral titer and accompanied with increased IL-1β and IL-18 mRNA levels. In addition, ALV-J infection up-regulated caspase-1 and caspase-3 activity in monocytes. Collectively, we found that ALV-J could cause cell death in chicken monocytes, especially pyroptosis, which may be a significant reason for ALV-J induced immunosuppression. PMID:29245947

5-Lipoxygenase contributes to PPARγ activation in macrophages in response to apoptotic cells.

PubMed

von Knethen, Andreas; Sha, Lisa K; Kuchler, Laura; Heeg, Annika K; Fuhrmann, Dominik; Heide, Heinrich; Wittig, Ilka; Maier, Thorsten J; Steinhilber, Dieter; Brüne, Bernhard

2013-12-01

Macrophage polarization to an anti-inflammatory phenotype upon contact with apoptotic cells is a contributing hallmark to immune suppression during the late phase of sepsis. Although the peroxisome proliferator-activated receptor γ (PPARγ) supports this macrophage phenotype switch, it remains elusive how apoptotic cells activate PPARγ. Assuming that a molecule causing PPARγ activation in macrophages originates in the cell membrane of apoptotic cells we analyzed lipid rafts from apoptotic, necrotic, and living human Jurkat T cells which showed the presence of 5-lipoxygenase (5-LO) in lipid rafts of apoptotic cells only. Incubating macrophages with lipid rafts of apoptotic, but not necrotic or living cells, induced PPAR responsive element (PPRE)-driven mRuby reporter gene expression in RAW 264.7 macrophages stably transduced with a 4xPPRE containing vector. Experiments with lipid rafts of apoptotic murine EL4 T cells revealed similar results. To verify the involvement of 5-LO in activating PPARγ in macrophages, Jurkat T cells were incubated with the 5-LO inhibitor MK-866 prior to induction of apoptosis, which failed to induce mRuby expression. Similar results were obtained with lipid rafts of apoptotic EL4 T cells preexposed to the 5-LO inhibitors zileuton and CJ-13610. Interestingly, Jurkat T cells overexpressing 5-LO failed to activate PPARγ in macrophages, while their 5-LO overexpressing apoptotic counterparts did. Our results suggest that during apoptosis 5-LO gets associated with lipid rafts and synthesizes ligands that in turn stimulate PPARγ in macrophages. © 2013.

Regulatory T cells accumulate and proliferate in the ischemic hemisphere for up to 30 days after MCAO.

PubMed

Stubbe, Tobias; Ebner, Friederike; Richter, Daniel; Engel, Odilo; Randolf Engel, Odilo; Klehmet, Juliane; Royl, Georg; Meisel, Andreas; Nitsch, Robert; Meisel, Christian; Brandt, Christine

2013-01-01

Local and peripheral immune responses are activated after ischemic stroke. In our present study, we investigated the temporal distribution, location, induction, and function of regulatory T cells (Tregs) and the possible involvement of microglia, macrophages, and dendritic cells after middle cerebral artery occlusion (MCAO). C57BL/6J and Foxp3(EGFP) transgenic mice were subjected to 30 minutes MCAO. On days 7, 14, and 30 after MCAO, Tregs and antigen presenting cells were analyzed using fluorescence activated cell sorting multicolor staining and immunohistochemistry. A strong accumulation of Tregs was observed on days 14 and 30 in the ischemic hemisphere accompanied by the elevated presence and activation of microglia. Dendritic cells and macrophages were found on each analyzed day. About 60% of Foxp3(+) Tregs in ischemic hemispheres were positive for the proliferation marker Ki-67 on days 7 and 14 after MCAO. The transfer of naive CD4(+) cells depleted of Foxp3(+) Tregs into RAG1(-/-) mice 1 day before MCAO did not lead to a de novo generation of Tregs 14 days after surgery. After depletion of CD25(+) Tregs, no changes regarding neurologic outcome were detected. The sustained presence of Tregs in the brain after MCAO indicates a long-lasting immunological alteration and involvement of brain cells in immunoregulatory mechanisms.

Mononuclear-macrophages but not neutrophils act as major infiltrating anti-leptospiral phagocytes during leptospirosis.

PubMed

Chen, Xu; Li, Shi-Jun; Ojcius, David M; Sun, Ai-Hua; Hu, Wei-Lin; Lin, Xu'ai; Yan, Jie

2017-01-01

To identify the major infiltrating phagocytes during leptospirosis and examine the killing mechanism used by the host to eliminate Leptospira interrogans. Major infiltrating phagocytes in Leptospira-infected C3H/HeJ mice were detected by immunohistochemistry. Chemokines and vascular endothelial cell adhesion molecules (VECAMs) of Leptospira-infected mice and leptospirosis patients were detected by microarray and immunohistochemistry. Leptospira-phagocytosing and -killing abilities of human or mouse macrophages and neutrophils, and the roles of intracellular ROS, NO and [Ca2+]i in Leptospira-killing process were evaluated by confocal microscopy and spectrofluorimetry. Peripheral blood mononuclear-macrophages rather than neutrophils were the main infiltrating phagocytes in the lungs, liver and kidneys of infected mice. Levels of macrophage- but not neutrophil-specific chemokines and VECAMs were significantly increased in the samples from infected mice and patients. All macrophages tested had a higher ability than neutrophils to phagocytose and kill leptospires. Higher ROS and NO levels and [Ca2+]i in the macrophages were involved in killing leptospires. Human macrophages displayed more phagolysosome formation and a stronger leptospire-killing ability to than mouse macrophages. Mononuclear-macrophages but not neutrophils represent the main infiltrating and anti-leptospiral phagocytes during leptospirosis. A lower level of phagosome-lysosome fusion may be responsible for the lower Leptospira-killing ability of human macrophages.

Spliced XBP1 promotes macrophage survival and autophagy by interacting with Beclin-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tian, Ping-Ge; Jiang, Zhi-Xin; Li, Jian-Hua

Macrophage autophagy plays an important role in the development of atherosclerosis, but the precise mechanism mediating this process is unclear. The potential role of the X-box binding protein 1 (XBP1), a crucial transduction factor that is involved in endoplasmic reticulum stress and the unfolded protein response, in bone marrow-derived macrophage autophagy is unknown. This study mainly explores the roles of XBP1 mRNA splicing in bone marrow-derived macrophage autophagy. The present study shows that the transient overexpression of spliced XBP1 via adenovirus-mediated gene transfer induces autophagy and promotes proliferation in bone marrow-derived macrophages via the down-regulation of Beclin-1, but that themore » sustained overexpression of spliced XBP1 leads to apoptosis. When XBP1 is down-regulated in bone marrow-derived macrophages using siRNA, rapamycin-induced autophagosome formation is ablated. Furthermore, we have detected the overexpression of XBP1 in areas of atherosclerotic plaques in the arteries of ApoE−/− mice. These results demonstrate that XBP1 mRNA splicing plays an important role in maintaining the function of bone marrow-derived macrophages and provide new insight into the study and treatment of atherosclerosis. - Highlights: • XBP1 was up-regulated in atherosclerotic plaques of ApoE−/− mice. • Transient spliced XBP1 overexpression induced macrophages autophagy via Beclin-1. • Sustained spliced XBP1 overexpression triggered macrophages apoptosis. • Spliced XBP1 plays a key role in maintaining the macrophages survival.« less

Culture of Macrophage Colony-stimulating Factor Differentiated Human Monocyte-derived Macrophages.

PubMed

Jin, Xueting; Kruth, Howard S

2016-06-30

A protocol is presented for cell culture of macrophage colony-stimulating factor (M-CSF) differentiated human monocyte-derived macrophages. For initiation of experiments, fresh or frozen monocytes are cultured in flasks for 1 week with M-CSF to induce their differentiation into macrophages. Then, the macrophages can be harvested and seeded into culture wells at required cell densities for carrying out experiments. The use of defined numbers of macrophages rather than defined numbers of monocytes to initiate macrophage cultures for experiments yields macrophage cultures in which the desired cell density can be more consistently attained. Use of cryopreserved monocytes reduces dependency on donor availability and produces more homogeneous macrophage cultures.

Soluble factors from biofilm of Candida albicans and Staphylococcus aureus promote cell death and inflammatory response.

PubMed

de Carvalho Dias, Kassia; Barbugli, Paula Aboud; de Patto, Fernanda; Lordello, Virginia Barreto; de Aquino Penteado, Letícia; Medeiros, Alexandra Ivo; Vergani, Carlos Eduardo

2017-06-30

The objective of this study was to better understand the effects of soluble factors from biofilm of single- and mixed-species Candida albicans (C. albicans) and methicillin-sensitive Staphylococcus aureus (MSSA) cultures after 36 h in culture on keratinocytes (NOK-si and HaCaT) and macrophages (J774A.1). Soluble factors from biofilms of C. albicans and MSSA were collected and incubated with keratinocytes and macrophages, which were subsequently evaluated by cell viability assays (MTT). Lactate dehydrogenase (LDH) enzyme release was measured to assess cell membrane damage to keratinocytes. Cells were analysed by brightfield microscopy after 2 and 24 h of exposure to the soluble factors from biofilm. Cell death was detected by labelling apoptotic cells with annexin V and necrotic cells with propidium iodide (PI) and was visualized via fluorescence microscopy. Soluble factors from biofilm were incubated with J774A.1 cells for 24 h; the subsequent production of NO and the cytokines IL-6 and TNF-α was measured by ELISA. The cell viability assays showed that the soluble factors of single-species C. albicans cultures were as toxic as the soluble factors from biofilm of mixed cultures, whereas the soluble factors of MSSA cultures were less toxic than those of C. albicans or mixed cultures. The soluble factors from biofilm of mixed cultures were the most toxic to the NOK-si and HaCaT cells, as confirmed by analyses of PI labelling and cell morphology. Soluble factors from biofilm of single-species MSSA and mixed-species cultures induced the production of IL-6, NO and TNF-α by J744A.1 macrophages. The production of IL-6 and NO induced by the soluble factors from biofilm of mixed cultures was lower than that induced by the soluble factors from biofilm of single-species MSSA cultures, whereas the soluble factors from biofilm of C. albicans cultures induced only low levels of NO. Soluble factors from 36-h-old biofilm of C. albicans and MSSA cultures promoted cell death and

Exosomes derived from gastric cancer cells activate NF-κB pathway in macrophages to promote cancer progression.

PubMed

Wu, Lijun; Zhang, Xu; Zhang, Bin; Shi, Hui; Yuan, Xiao; Sun, Yaoxiang; Pan, Zhaoji; Qian, Hui; Xu, Wenrong

2016-09-01

Exosomes are nano-sized membrane vesicles secreted by both normal and cancer cells. Emerging evidence indicates that cancer cells derived exosomes contribute to cancer progression through the modulation of tumor microenvironment. However, the effects of exosomes derived from gastric cancer cells on macrophages are not well understood. In this study, we investigated the biological role of gastric cancer cells derived exosomes in the activation of macrophages. We demonstrated that gastric cancer cells derived exosomes activated macrophages to express increased levels of proinflammatory factors, which in turn promoted tumor cell proliferation and migration. In addition, gastric cancer cells derived exosomes remarkably upregulated the phosphorylation of NF-κB in macrophages. Inhibiting the activation of NF-κB reversed the upregulation of proinflammatory factors in macrophages and blocked their promoting effects on gastric cancer cells. Moreover, we found that gastric cancer cells derived exosomes could also activate macrophages from human peripheral blood monocytes through the activation of NF-κB. In conclusion, our results suggest that gastric cancer cells derived exosomes stimulate the activation of NF-κB pathway in macrophages to promote cancer progression, which provides a potential therapeutic approach for gastric cancer by interfering with the interaction between exosomes and macrophages in tumor microenvironment.